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Differential expression of nitric oxide synthases in porcine aortic endothelial cells during LPS-induced apoptosis
Background It is well known that nitric oxide (NO) is generated by a family of constitutively (nNOS and eNOS) or inducibly (iNOS) expressed enzymes and takes part in different aspects of the inflammatory response; nevertheless, its effective role in the pathogenesis of multiple organ dysfunction and septic shock is not fully understood. Methods To investigate the Nitric Oxide Synthases (NOSs) expression in endothelial cells during endotoxin exposure and the involvement of NO in lipopolysaccharide (LPS)-induced apoptosis, primary cultures of porcine Aortic Endothelial Cells (pAECs) were exposed to LPS for different time periods (1-24 h) and to LPS?+?L-NAME (15 h). Results Lipopolysaccharide induced an increase in mRNA and protein iNOS expression; on the contrary, the expression of eNOS was decreased. Furthermore, NOSs localisation was in part modified by LPS treatment. No alteration in the total level of Nitric Oxide was observed. L-NAME (5 mM) addition determined a slight decrease of LPS-induced apoptosis. Conclusions Endotoxin treatment strongly influenced NOS expression with an upregulation of iNOS and a simultaneous down regulation of eNOS. Moreover, in our model, the involvement of NO on LPS-induced apoptosis is very modest, suggesting that different pathways are involved in the regulation of this process.
Nitric oxide-scavenging compounds in Agrimonia pilosa Ledeb on LPS-induced RAW264.7 macrophages
The extract of Agrimonia pilosa Ledeb, with a high polyphenol content, inhibited nitrite accumulation as an indicator of nitric oxide (NO) in LPS-induced RAW264.7 macrophages. The NO inhibitory compounds in the extract were isolated using open column chromatography and HPLC, and five phenolic compounds, namely aromadendrin (AD), dihydrokaempferol 3-O-?-D-glucoside (DK3-O-glc), quercitrin (QC), aglimonolide-6-O-?-D-glucoside (AG6-O-glc) and loliolide (LL), were determined by
Junsei Taira; Hitoshi Nanbu; Katsuhiro Ueda
1,2,3,6-tetra-O-galloyl-beta-D-allopyranose gallotannin isolated, from Euphorbia jolkini, attenuates LPS-induced nitric oxide production in macrophages.
Nitric oxide (NO) is a pleiotropic regulator, critical to numerous biological processes, including vasodilatation and macrophage-mediated immunity. Macrophages express inducible NO synthase (iNOS) and produce NO after lipopolysaccharide (LPS) stimulation. Gallotannins are water-soluble polyphenols with wide-ranging biological activities. Various chemical structures of gallotannins occurring in medicinal and food plants that are used worldwide showed several remarkable biological and pharmacological activities. In the present study, we examined the inhibitory effects of gallotannin 1,2,3,6-tetra-O-galloyl-beta-D-allopyranose (GT24) isolated from Euphorbia jolkini on the LPS-induced NO production and underlying mechanisms of action. GT24 dose-dependently decreased LPS-induced NO production and iNOS expression in J774A.1 macrophages. In addition, GT24 inhibited LPS-induced activation of nuclear factor (NF)-kappaB as indicated by inhibition of degradation of I-kappaBalpha, nuclear translocation of NF-kappaB, and NF-kappaB dependent gene reporter assay. Our results suggest that GT24 possesses an inhibitory effect on the LPS-induced inflammatory reaction. PMID:20665470
Park, Seung-Bin; Kim, Mi-Sun; Lee, Hee Sang; Lee, Seung Ho; Kim, Sang-Hyun
Constituents of Limonia acidissima inhibit LPS-induced nitric oxide production in BV-2 microglia.
The ethyl acetate (EtOAc) soluble fraction of the 85% ethanol (EtOH) extract of the dried bark of Limonia acidissima potently inhibited nitric oxide (NO) production in lipopolysaccharide (LPS) activated BV-2 cells, a microglial cell line. Bioassay-guided column chromatography separation afforded a new stereoisomer of neolignan, (7'E)-(7R,8S)-4-hydroxy-3,5'-dimethoxy-4',7-epoxy-8,3'-neolig-7'-en-9,9'-diyil diacetate (1), together with two known lignans, (+)-yangambin (2) and (+)-syringaresinol (3), three known triterpenoids, hederatriol (4), basic acid methyl ester (5), and 3?-hydroxyolean-12-en-11-one (6), and four known fatty acid derivatives, cascarillic acid (7), (+)-?-dimorphecolic acid (8), 8(R)-hydroxylinoleic acid (9), and (6Z,9Z,12Z)-pentadecatrienoic acid (10). The structure of the new compound 1 was elucidated by detailed analysis of spectroscopic data and circular dichroism (CD) spectroscopy. Compounds 1, 3-6, and 8-10 isolated from L. acidissima significantly reduced NO production in LPS-stimulated BV-2 microglia cells. PMID:20578973
Kim, Ki Hyun; Ha, Sang Keun; Kim, Sun Yeou; Youn, Hyun Joo; Lee, Kang Ro
Geniposide suppresses LPS-induced nitric oxide, PGE2 and inflammatory cytokine by downregulating NF-?B, MAPK and AP-1 signaling pathways in macrophages.
Inflammatory responses are important to host immune reactions, but uncontrolled inflammatory mediators may aid in the pathogenesis of other inflammatory diseases. Geniposide, an iridoid glycoside found in the herb gardenia, is believed to have broad-spectrum anti-inflammatory effects in murine models but its mechanism of action is unclear. We investigated the action of this compound in murine macrophages stimulated by lipopolysaccharide (LPS), as the stimulation of macrophages by LPS is known to induce inflammatory reactions. We determined the effect of geniposide on LPS-induced production of the inflammatory mediators, nitric oxide (NO) and prostaglandin E2 (PGE2), the mRNA and protein expression of the NO and PGE2 synthases, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively, and the mRNA and protein expression of the inflammatory cytokine, tumor necrosis factor-? (TNF-?) and interleukin-6 (IL-6). Furthermore, nuclear factor (NF)-?B, mitogen-activated protein kinase (MAPK) and activator protein (AP)-1 activity were assayed. To understand the action of geniposide on the NF-?B and MAPK pathways, we studied the effect of NF-?B and MAPK inhibitors on the LPS-induced production of NO, PGE2 and TNF-?. Our findings clearly showed that geniposide mainly exerts its anti-inflammatory effects by inhibiting the LPS-induced NF-?B, MAPK and AP-1 signaling pathways in macrophages, which subsequently reduces overexpression of the inducible enzymes iNOS and COX-2 and suppresses the expression and release of the inflammatory factors, TNF-?, IL-6, NO and PGE2. Thus, geniposide shows promise as a therapeutic agent in inflammatory diseases. PMID:24735815
Shi, Qinghai; Cao, Jinjun; Fang, Li; Zhao, Hongyan; Liu, Zhengxiang; Ran, Jihua; Zheng, Xinchuan; Li, Xiaoling; Zhou, Yu; Ge, Di; Zhang, Hongming; Wang, Li; Ran, Ying; Fu, Jianfeng
Hypericum triquetrifolium-Derived Factors Downregulate the Production Levels of LPS-Induced Nitric Oxide and Tumor Necrosis Factor-? in THP-1 Cells.
Based on knowledge from traditional Arab herbal medicine, this in vitro study aims to examine the anti-inflammatory mechanism of Hypericum triquetrifolium by measuring the expression and release of pro-inflammatory cytokines, tumor necrosis factor-? (TNF-?) and interleukine-6 (IL-6), and inducible nitric oxide synthase (iNOS) in human monocytic cells, THP-1. The effects were assessed by measuring the levels of secretory proteins and mRNA of TNF-? and IL-6, the levels of nitric oxide (NO) secretion and the expression of iNOS in THP-1 cells. Cells were treated with 5??g lipopolysaccharide/ml (LPS) in the presence and absence of increasing concentrations of extracts from the aerial parts of H. triquetrifolium. During the entire experimental period, we used extract concentrations (up to 250??g?mL(-1)) that had no cytotoxic effects, as measured with MTT and LDH assays. Hypericum triquetrifolium extracts remarkably suppressed the LPS-induced NO release, significantly attenuated the LPS-induced transcription of iNOS and inhibited in a dose-dependent manner the expression and release of TNF-?. No significant effects were observed on the release of IL-6. Taken together, these results suggest that H. triquetrifolium probably exerts anti-inflammatory effects through the suppression of TNF-? and iNOS expressions. PMID:18955363
Saad, Bashar; Abouatta, Bernadette Soudah; Basha, Walid; Hmade, Alaa; Kmail, Abdalsalam; Khasib, Said; Said, Omar
Hypericum triquetrifolium--Derived Factors Downregulate the Production Levels of LPS-Induced Nitric Oxide and Tumor Necrosis Factor-? in THP-1 Cells
Based on knowledge from traditional Arab herbal medicine, this in vitro study aims to examine the anti-inflammatory mechanism of Hypericum triquetrifolium by measuring the expression and release of pro-inflammatory cytokines, tumor necrosis factor-? (TNF-?) and interleukine-6 (IL-6), and inducible nitric oxide synthase (iNOS) in human monocytic cells, THP-1. The effects were assessed by measuring the levels of secretory proteins and mRNA of TNF-? and IL-6, the levels of nitric oxide (NO) secretion and the expression of iNOS in THP-1 cells. Cells were treated with 5??g lipopolysaccharide/ml (LPS) in the presence and absence of increasing concentrations of extracts from the aerial parts of H. triquetrifolium. During the entire experimental period, we used extract concentrations (up to 250??g?mL?1) that had no cytotoxic effects, as measured with MTT and LDH assays. Hypericum triquetrifolium extracts remarkably suppressed the LPS-induced NO release, significantly attenuated the LPS-induced transcription of iNOS and inhibited in a dose-dependent manner the expression and release of TNF-?. No significant effects were observed on the release of IL-6. Taken together, these results suggest that H. triquetrifolium probably exerts anti-inflammatory effects through the suppression of TNF-? and iNOS expressions.
Saad, Bashar; AbouAtta, Bernadette Soudah; Basha, Walid; Hmade, Alaa; Kmail, Abdalsalam; Khasib, Said; Said, Omar
Rosmarinic Acid in Prunella vulgaris Ethanol Extract Inhibits LPS-induced Prostaglandin E2 and Nitric Oxide in RAW264.7 Mouse Macrophages
Prunella vulgaris has been used therapeutically for inflammation related conditions for centuries, but systematic studies of its anti-inflammatory activity are lacking and no specific active components have been identified. In this study, water and ethanol extracts of four P. vulgaris accessions were applied to RAW264.7 mouse macrophages and the ethanol extracts significantly inhibited lipopolysaccharide (LPS)-stimulated prostaglandin E2 (PGE2) and nitric oxide (NO) production at 30 ?g/mL without affecting cell viability. Extracts from different accessions of P. vulgaris were screened for anti-inflammatory activity to identify accessions with the greatest activity. The inhibition of PGE2 and NO production by selected extracts was dose-dependent, with significant effects seen at concentrations as low as 10 ?g/mL. Fractionation of ethanol extracts from the active accession, Ames 27664, suggested fractions 3 and 5 as possible major contributors to the overall activity. Rosmarinic acid (RA) content in P. vulgaris was found to independently inhibit inflammatory response, but it only partially explained the extracts' activity. LPS-induced cyclooxyginase-2 (COX-2) and nitric oxide synthase (iNOS) protein expression were both attenuated by P. vulgaris ethanol extracts, while RA only inhibited COX-2 expression.
Huang, Nan; Hauck, Cathy; Yum, Man-Yu; Rizshsky, Ludmila; Widrlechner, Mark P.; McCoy, Joe-Ann; Murphy, Patricia A.; Dixon, Philip M.; Nikolau, Basil J.; Birt, Diane F.
Withaferin A inhibits iNOS expression and nitric oxide production by Akt inactivation and down-regulating LPS-induced activity of NF-kappaB in RAW 264.7 cells.
Induction of inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production is thought to have beneficial immunomodulatory effects in acute and chronic inflammatory disorders. In Raw 264.7 cells stimulated with lipopolysaccharide (LPS) to mimic inflammation, withaferin A inhibited LPS-induced expression of both iNOS protein and mRNA in a dose-dependent manner. To investigate the mechanism by which withaferin A inhibits iNOS gene expression, we examined activation of mitogen-activated protein kinases (MAPKs) and Akt in Raw 264.7 cells. We did not observe any significant changes in the phosphorylation of p38 MAPK in cells treated with LPS alone or LPS plus withaferin A. However, LPS-induced Akt phosphorylation was markedly inhibited by withaferin A, while the phosphorylation of p42/p44 extracellular signal-regulated kinases (ERKs) was slightly inhibited by withaferin A treatment. Withaferin A prevented IkappaB phosphorylation, blocking the subsequent nuclear translocation of nuclear factor-kappaB (NF-kappaB) and inhibiting its DNA binding activity. LPS-induced p65 phosphorylation, which is mediated by extracellular signal-regulated kinase (ERK) and Akt pathways, was attenuated by withaferin A treatment. Moreover, LPS-induced NO production and NF-kappaB activation were inhibited by SH-6, a specific inhibitor of Akt. Taken together, these results suggest that withaferin A inhibits inflammation through inhibition of NO production and iNOS expression, at least in part, by blocking Akt and subsequently down-regulating NF-kappaB activity. PMID:18838070
Oh, Jung Hwa; Lee, Tae-Jin; Park, Jong-Wook; Kwon, Taeg Kyu
Inhibitory effects of (-)-?-bisabolol on LPS-induced inflammatory response in RAW264.7 macrophages.
Although (-)-?-bisabolol, a natural monocyclic sesquiterpene alcohol, is often used as a cosmetic soothing supplement, little is known about its mechanisms of anti-inflammatory effects. Therefore, this study was designed to investigate anti-inflammatory effects of (-)-?-bisabolol and its mechanisms of action. In this study, we found that (-)-?-bisabolol inhibited lipopolysaccharide (LPS)-induced production of nitric oxide (NO) and prostaglandin E(2) (PGE(2)) in RAW264.7 cells. In addition, expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) genes was reduced, as evidenced by Western blot and luciferase reporter assays for COX-2 and iNOS. To assess the mechanism of the anti-inflammatory property of (-)-?-bisabolol, its effects on the activity of AP-1 and NF-?B promoters were examined. LPS-induced activation of AP-1 and NF-?B promoters was significantly reduced by (-)-?-bisabolol. Consistently, (-)-?-bisabolol reduced LPS-induced phosphorylation of I?B?. In addition, while LPS-induced phosphorylation of ERK and p38 was attenuated by (-)-?-bisabolol, significant changes in the level of phosphorylated JNK were not observed. Our results indicate that (-)-?-bisabolol exerts anti-inflammatory effects by downregulating expression of iNOS and COX-2 genes through inhibition of NF-?B and AP-1 (ERK and p38) signaling. PMID:21771629
Kim, Seungbeom; Jung, Eunsun; Kim, Jang-Hyun; Park, Young-Ho; Lee, Jongsung; Park, Deokhoon
Requirement for STAT1 in LPS-induced gene expression in macrophages
This study examines the role of the sig- nal transducer and activator of transcription 1 (STAT1) in induction of lipopolysaccharide (LPS)- stimulated gene expression both in vitro and in vivo. LPS-induced expression of an interferon (IFN)-inducible 10-kDa protein (IP-10), IFN reg- ulatory factor-1 (IRF-1), and inducible nitric oxide synthase (iNOS) mRNAs was severely impaired in macrophages prepared from Stat12\\/2 mice,
Yoshihiro Ohmori; Thomas A. Hamilton
Achillea millefolium L. Essential Oil Inhibits LPS-Induced Oxidative Stress and Nitric Oxide Production in RAW 264.7 Macrophages
Achillea millefolium L. is a member of the Asteraceae family and has been used in folk medicine in many countries. In this study, 19 compounds in A. millefolium essential oil (AM-EO) have been identified; the major components are artemisia ketone (14.92%), camphor (11.64%), linalyl acetate (11.51%) and 1,8-cineole (10.15%). AM-EO can suppress the inflammatory responses of lipopolysaccharides (LPS)-stimulated RAW 264.7 macrophages, including decreased levels of cellular nitric oxide (NO) and superoxide anion production, lipid peroxidation and glutathione (GSH) concentration. This antioxidant activity is not a result of increased superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) activities, but rather occurs as a result of the down-regulation of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-? (TNF-?), interleukin-6 (IL-6) and heme oxygenase-1 (HO-1) expression, thus reducing the inflammatory response. Therefore, AM-EO can be utilized in many applications, including the treatment of inflammatory diseases in the future.
Chou, Su-Tze; Peng, Hsin-Yi; Hsu, Jaw-Cherng; Lin, Chih-Chien; Shih, Ying
Stevioside protects LPS-induced acute lung injury in mice.
Stevioside, a diterpene glycoside component of Stevia rebaudiana, has been known to exhibit anti-inflammatory properties. To evaluate the effect and the possible mechanism of stevioside in lipopolysaccharide (LPS)-induced acute lung injury, male BALB/c mice were pretreated with stevioside or dexamethasone 1 h before intranasal instillation of LPS. Seven hours later, tumor necrosis factor-?, interleukin-1?, and interleukin-6 in bronchoalveolar lavage fluid (BALF) were measured by using enzyme-linked immunosorbent assay. The number of total cells, neutrophils, and macrophages in the BALF were also determined. The right lung was excised for histological examination and analysis of myeloperoxidase activity and nitrate/nitrite content. Cyclooxygenase 2 (COX-2), inducible NO synthase (iNOS), nuclear factor-kappa B (NF-?B), inhibitory kappa B protein were detected by western blot. The results showed that stevioside markedly attenuated the LPS-induced histological alterations in the lung. Stevioside inhibited the production of pro-inflammatory cytokines and the expression of COX-2 and iNOS induced by LPS. In addition, not only was the wet-to-dry weight ratio of lung tissue significantly decreased, the number of total cells, neutrophils, and macrophages in the BALF were also significantly reduced after treatment with stevioside. Moreover, western blotting showed that stevioside inhibited the phosphorylation of I?B-? and NF-?B caused by LPS. Taken together, our results suggest that anti-inflammatory effect of stevioside against the LPS-induced acute lung injury may be due to its ability of inhibition of the NF-?B signaling pathway. Stevioside may be a promising potential therapeutic reagent for acute lung injury treatment. PMID:22968433
Yingkun, Nie; Zhenyu, Wang; Jing, Lin; Xiuyun, Lu; Huimin, Yu
Inhibition of IRAK-4 activity for rescuing endotoxin LPS-induced septic mortality in mice by lonicerae flos extract
Highlights: �Lonicerae flos extract (HS-23) is a clinical candidate, Phase I for sepsis treatment. �Here, HS-23 or its major constituents rescued LPS-induced septic mortality in mice. �As a mechanism, they directly inhibited IRAK-4-catalyzed kinase activity. �Thus, they suppressed LPS-induced expression of NF-?B/AP-1-target inflammatory genes. -- Abstract: Lonicerae flos extract (HS-23) is a clinical candidate currently undergoing Phase I trial in lipopolysaccharide (LPS)-injected healthy human volunteers, but its molecular basis remains to be defined. Here, we investigated protective effects of HS-23 or its major constituents on Escherichia coli LPS-induced septic mortality in mice. Intravenous treatment with HS-23 rescued LPS-intoxicated C57BL/6J mice under septic conditions, and decreased the levels of cytokines such as tumor necrosis factor ? (TNF-?), interleukin (IL)-1? and high-mobility group box-1 (HMGB-1) in the blood. Chlorogenic acid (CGA) and its isomers were assigned as major constituents of HS-23 in the protection against endotoxemia. As a molecular mechanism, HS-23 or CGA isomers inhibited endotoxin LPS-induced autophosphorylation of the IL-1 receptor-associated kinase 4 (IRAK-4) in mouse peritoneal macrophages as well as the kinase activity of IRAK-4 in cell-free reactions. HS-23 consequently suppressed downstream pathways critical for LPS-induced activation of nuclear factor (NF)-?B or activating protein 1 (AP-1) in the peritoneal macrophages. HS-23 also inhibited various toll-like receptor agonists-induced nitric oxide (NO) production, and down-regulated LPS-induced expression of NF-?B/AP-1-target inflammatory genes in the cells. Taken together, HS-23 or CGA isomers exhibited anti-inflammatory therapy against LPS-induced septic mortality in mice, at least in part, mediated through the inhibition of IRAK-4.
Park, Sun Hong; Roh, Eunmiri [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of)] [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of); Kim, Hyun Soo [Pharmaceutical R and D Center, Huons Co., Ltd., Anyang (Korea, Republic of)] [Pharmaceutical R and D Center, Huons Co., Ltd., Anyang (Korea, Republic of); Baek, Seung-Il [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of)] [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of); Choi, Nam Song [Pharmaceutical R and D Center, Huons Co., Ltd., Anyang (Korea, Republic of)] [Pharmaceutical R and D Center, Huons Co., Ltd., Anyang (Korea, Republic of); Kim, Narae; Hwang, Bang Yeon; Han, Sang-Bae [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of)] [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of); Kim, Youngsoo, E-mail: email@example.com [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of)] [College of Pharmacy, Chungbuk National University, Cheongju 361-763 (Korea, Republic of)
Carabrol suppresses LPS-induced nitric oxide synthase expression by inactivation of p38 and JNK via inhibition of I-{kappa}B{alpha} degradation in RAW 264.7 cells
Carabrol, isolated from Carpesium macrocephalum, showed anti-inflammatory potential in LPS-induced RAW 264.7 murine macrophages. In present study, carabrol demonstrated the inhibitory activity on pro-inflammatory cytokines such as IL-1{beta}, IL-6 and TNF-{alpha}. In addition, mRNA and protein levels of iNOS and COX-2 were reduced by carabrol. Molecular analysis revealed that these suppressive effects were correlated with the inactivation of p38 and JNK via inhibition of NF-{kappa}B activation. Immunoblotting showed that carabrol suppressed LPS-induced degradation of I-{kappa}B{alpha} and decreased nuclear translocation of p65. Taken together, these results suggest that carabrol can be a modulator of pro-inflammatory signal transduction pathway in RAW 264.7 cells.
Lee, Hwa Jin [Department of Life Science, Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of)] [Department of Life Science, Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of); Lim, Hyo Jin; Lee, Da Yeon [College of Pharmacy, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of)] [College of Pharmacy, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of); Jung, Hyeyoun [Department of Life Science, Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of)] [Department of Life Science, Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of); Kim, Mi-Ran; Moon, Dong-Cheul [College of Pharmacy, Chungbuk National University, Cheungju 361-763 (Korea, Republic of)] [College of Pharmacy, Chungbuk National University, Cheungju 361-763 (Korea, Republic of); Kim, Keun Il; Lee, Myeong-Sok [Department of Life Science, Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of)] [Department of Life Science, Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of); Ryu, Jae-Ha, E-mail: email@example.com [College of Pharmacy, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of)] [College of Pharmacy, Sookmyung Women's University, Seoul 140-742 (Korea, Republic of)
The Effect of the Aerial Part of Lindera akoensis on Lipopolysaccharides (LPS)-Induced Nitric Oxide Production in RAW264.7 Cells.
Four new secondary metabolites, 3?-((E)-Dodec-1-enyl)-4?-hydroxy-5?-methyldihydrofuran-2-one (1), linderinol (6), 4'-O-methylkaempferol 3-O-?-L-(4''-E-p-coumaroyl)rhamnoside (11) and kaempferol 3-O-?-L-(4''-Z-p-coumaroyl)rhamnoside (12) with eleven known compounds-3-epilistenolide D1 (2), 3-epilistenolide D2 (3), (3Z,4?,5?)-3-(dodec-11-ynylidene)-4-hydroxy-5-methylbutanolide (4), (3E,4?,5?)-3-(dodec-11-ynylidene)-4-hydroxy-5-methylbutanolide (5), matairesinol (7), syringaresinol (8), (+)-pinoresinol (9), salicifoliol (10), 4''-p-coumaroylafzelin (13), catechin (14) and epicatechin (15)-were first isolated from the aerial part of Lindera akoensis. Their structures were determined by detailed analysis of 1D- and 2D-NMR spectroscopic data. All of the compounds isolated from Lindera akoensis showed that in vitro anti-inflammatory activity decreases the LPS-stimulated production of nitric oxide (NO) in RAW 264.7 cell, with IC50 values of 4.1-413.8 �M. PMID:23624606
Yang, Chung-Ping; Huang, Guan-Jhong; Huang, Hui-Chi; Chen, Yu-Chang; Chang, Chi-I; Wang, Sheng-Yang; Chang, Hsun-Shuo; Tseng, Yen-Hsueh; Chien, Shih-Chang; Kuo, Yueh-Hsiung
The Effect of the Aerial Part of Lindera akoensis on Lipopolysaccharides (LPS)-Induced Nitric Oxide Production in RAW264.7 Cells
Four new secondary metabolites, 3?-((E)-Dodec-1-enyl)-4?-hydroxy-5?-methyldihydrofuran-2-one (1), linderinol (6), 4?-O-methylkaempferol 3-O-?-l-(4?-E-p-coumaroyl)rhamnoside (11) and kaempferol 3-O-?-l-(4?-Z-p-coumaroyl) rhamnoside (12) with eleven known compounds�3-epilistenolide D1 (2), 3-epilistenolide D2 (3), (3Z,4?,5?)-3-(dodec-11-ynylidene)-4-hydroxy-5-methylbutanolide (4), (3E,4?,5?)-3-(dodec-11-ynylidene)-4-hydroxy-5-methylbutanolide (5), matairesinol (7), syringaresinol (8), (+)-pinoresinol (9), salicifoliol (10), 4?-p-coumaroylafzelin (13), catechin (14) and epicatechin (15)�were first isolated from the aerial part of Lindera akoensis. Their structures were determined by detailed analysis of 1D- and 2D-NMR spectroscopic data. All of the compounds isolated from Lindera akoensis showed that in vitro anti-inflammatory activity decreases the LPS-stimulated production of nitric oxide (NO) in RAW 264.7 cell, with IC50 values of 4.1�413.8 ?M.
Yang, Chung-Ping; Huang, Guan-Jhong; Huang, Hui-Chi; Chen, Yu-Chang; Chang, Chi-I; Wang, Sheng-Yang; Chang, Hsun-Shuo; Tseng, Yen-Hsueh; Chien, Shih-Chang; Kuo, Yueh-Hsiung
Macelignan attenuates LPS-induced inflammation and reduces LPS-induced spatial learning impairments in rats.
Previous studies have shown that macelignan has anti-inflammatory and neuroprotective effects. Subsequently, in the current study, we demonstrate that oral administrations of macelignan reduce the hippocampal microglial activation induced by chronic infusions of lipopolysaccharide (LPS) into the fourth ventricle of Fisher-344 rat brains. A Morris water maze was used to evaluate the status of the hippocampal-dependent spatial learning in control rats with an artificial cerebrospinal fluid infusion, rats with chronic LPS infusions, and rats with chronic LPS infusions and oral administrations of macelignan. The rats with chronic LPS infusions showed spatial memory impairments relative to the control rats in the performance of the memory task. Daily administration of macelignan reduced the spatial memory impairments induced by the chronic LPS infusions. The results indicate that macelignan may possess therapeutic potential for the prevention of Alzheimer's disease. PMID:18940231
Cui, Chun-Ai; Jin, Da-Qing; Hwang, Yoo Kyeong; Lee, Im-Soon; Hwang, Jae Kwan; Ha, Ilho; Han, Jung-Soo
Macelignan attenuates LPS-induced inflammation and reduces LPS-induced spatial learning impairments in rats
Previous studies have shown that macelignan has anti-inflammatory and neuroprotective effects. Subsequently, in the current study, we demonstrate that oral administrations of macelignan reduce the hippocampal microglial activation induced by chronic infusions of lipopolysaccharide (LPS) into the fourth ventricle of Fisher-344 rat brains. A Morris water maze was used to evaluate the status of the hippocampal-dependent spatial learning in control
Chun-Ai Cui; Da-Qing Jin; Yoo Kyeong Hwang; Im-Soon Lee; Jae Kwan Hwang; Ilho Ha; Jung-Soo Han
Amelioration of LPS-Induced Inflammation Response in Microglia by AMPK Activation
Adenosine 5?-monophosphate-activated protein kinase (AMPK) is a key regulator of cellular energy homeostasis via modulating metabolism of glucose, lipid, and protein. In addition to energy modulation, AMPK has been demonstrated to associate with several important cellular events including inflammation. The results showed that ENERGI-F704 identified from bamboo shoot extract was nontoxic in concentrations up to 80??M and dose-dependently induced phosphorylation of AMPK (Thr-172) in microglia BV2 cells. Our findings also showed that the treatment of BV2 with ENERGI-F704 ameliorated the LPS-induced elevation of IL-6 and TNF-? production. In addition, ENERGI-F704 reduced increased production of nitric oxide (NO) and prostaglandin E2 (PGE2) via downregulating the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2), respectively. Moreover, ENERGI-F704 decreased activated nuclear translocation and protein level of NF-?B. Inhibition of AMPK with compound C restored decreased NF-?B translocation by ENERGI-F704. In conclusion, ENERGI-F704 exerts inhibitory activity on LPS-induced inflammation through manipulating AMPK signaling and exhibits a potential therapeutic agent for neuroinflammatory disease.
Chen, Chin-Chen; Lin, Jiun-Tsai; Cheng, Yi-Fang; Kuo, Cheng-Yi; Huang, Chun-Fang; Kao, Shao-Hsuan; Liang, Yao-Jen; Cheng, Ching-Yi; Chen, Han-Min
Intermedin Attenuates LPS-induced Inflammation in the Rat Testis
First reported as a vasoactive peptide in the cardiovascular system, intermedin (IMD), also known as adrenomedullin 2 (ADM2), is a hormone with multiple potent roles, including its antioxidant action on the pulmonary, central nervous, cardiovascular and renal systems. Though IMD may play certain roles in trophoblast cell invasion, early embryonic development and cumulus cell-oocyte interaction, the role of IMD in the male reproductive system has yet to be investigated. This paper reports our findings on the gene expression of IMD, its receptor components and its protein localization in the testes. In a rat model, bacterial lippolysaccharide (LPS) induced atypical orchitis, and LPS treatment upregulated the expression of IMD and one of its receptor component proteins, i.e. receptor activity modifying protein 2 (RAMP2). IMD decreased both plasma and testicular levels of reactive oxygen species (ROS) production, attenuated the increase in the gene expression of the proinflammatory cytokines tumor necrosis factor alpha (TNF?), interleukin 6 (IL6) and interleukin 1 beta (IL1?), rescued spermatogenesis, and prevented the decrease in plasma testosterone levels caused by LPS. The restorative effect of IMD on steroidogenesis was also observed in hydrogen peroxide-treated rat primary Leydig cells culture. Our results indicate IMD plays an important protective role in spermatogenesis and steroidogenesis, suggesting therapeutic potential for IMD in pathological conditions such as orchitis.
Liu, Yongjun; Huang, Chen; O, Wai-sum; Tang, Fai; Zhang, Jian V.
Role of reactive oxygen species in LPS-induced production of prostaglandin E2 in microglia.
We determined the roles of reactive oxygen species (ROS) in the expression of cyclooxygenase-2 (COX-2) and the production of prostaglandin E2 (PGE2) in lipopolysaccharide (LPS)-activated microglia. LPS treatment increased intracellular ROS in rat microglia dose-dependently. Pre-treatment with superoxide dismutase (SOD)/catalase, or SOD/catalase mimetics that can scavenge intracellular ROS, significantly attenuated LPS-induced release in PGE2. Diphenylene iodonium (DPI), a non-specific NADPH oxidase inhibitor, decreased LPS-induced PGE2 production. In addition, microglia from NADPH oxidase-deficient mice produced less PGE2 than those from wild-type mice following LPS treatment. Furthermore, LPS-stimulated expression of COX-2 (determined by RT-PCR analysis of COX-2 mRNA and western blot for its protein) was significantly reduced by pre-treatment with SOD/catalase or SOD/catalase mimetics. SOD/catalase mimetics were more potent than SOD/catalase in reducing COX-2 expression and PGE2 production. As a comparison, scavenging ROS had no effect on LPS-induced nitric oxide production in microglia. These results suggest that ROS play a regulatory role in the expression of COX-2 and the subsequent production of PGE2 during the activation process of microglia. Thus, inhibiting NADPH oxidase activity and subsequent ROS generation in microglia can reduce COX-2 expression and PGE2 production. These findings suggest a potential therapeutic intervention strategy for the treatment of inflammation-mediated neurodegenerative diseases. PMID:14756815
Wang, Tongguang; Qin, Liya; Liu, Bin; Liu, Yuxin; Wilson, Belinda; Eling, Thomas E; Langenbach, Robert; Taniura, Seijiro; Hong, Jau-Shyong
Ginger extract inhibits LPS induced macrophage activation and function
Background Macrophages play a dual role in host defence. They act as the first line of defence by mounting an inflammatory response to antigen exposure and also act as antigen presenting cells and initiate the adaptive immune response. They are also the primary infiltrating cells at the site of inflammation. Inhibition of macrophage activation is one of the possible approaches towards modulating inflammation. Both conventional and alternative approaches are being studied in this regard. Ginger, an herbal product with broad anti inflammatory actions, is used as an alternative medicine in a number of inflammatory conditions like rheumatic disorders. In the present study we examined the effect of ginger extract on macrophage activation in the presence of LPS stimulation. Methods Murine peritoneal macrophages were stimulated by LPS in presence or absence of ginger extract and production of proinflammatory cytokines and chemokines were observed. We also studied the effect of ginger extract on the LPS induced expression of MHC II, B7.1, B7.2 and CD40 molecules. We also studied the antigen presenting function of ginger extract treated macrophages by primary mixed lymphocyte reaction. Results We observed that ginger extract inhibited IL-12, TNF-?, IL-1? (pro inflammatory cytokines) and RANTES, MCP-1 (pro inflammatory chemokines) production in LPS stimulated macrophages. Ginger extract also down regulated the expression of B7.1, B7.2 and MHC class II molecules. In addition ginger extract negatively affected the antigen presenting function of macrophages and we observed a significant reduction in T cell proliferation in response to allostimulation, when ginger extract treated macrophages were used as APCs. A significant decrease in IFN-? and IL-2 production by T cells in response to allostimulation was also observed. Conclusion In conclusion ginger extract inhibits macrophage activation and APC function and indirectly inhibits T cell activation.
Inhibition of IRAK-4 activity for rescuing endotoxin LPS-induced septic mortality in mice by lonicerae flos extract.
Lonicerae flos extract (HS-23) is a clinical candidate currently undergoing Phase I trial in lipopolysaccharide (LPS)-injected healthy human volunteers, but its molecular basis remains to be defined. Here, we investigated protective effects of HS-23 or its major constituents on Escherichia coli LPS-induced septic mortality in mice. Intravenous treatment with HS-23 rescued LPS-intoxicated C57BL/6J mice under septic conditions, and decreased the levels of cytokines such as tumor necrosis factor ? (TNF-?), interleukin (IL)-1? and high-mobility group box-1 (HMGB-1) in the blood. Chlorogenic acid (CGA) and its isomers were assigned as major constituents of HS-23 in the protection against endotoxemia. As a molecular mechanism, HS-23 or CGA isomers inhibited endotoxin LPS-induced autophosphorylation of the IL-1 receptor-associated kinase 4 (IRAK-4) in mouse peritoneal macrophages as well as the kinase activity of IRAK-4 in cell-free reactions. HS-23 consequently suppressed downstream pathways critical for LPS-induced activation of nuclear factor (NF)-?B or activating protein 1 (AP-1) in the peritoneal macrophages. HS-23 also inhibited various toll-like receptor agonists-induced nitric oxide (NO) production, and down-regulated LPS-induced expression of NF-?B/AP-1-target inflammatory genes in the cells. Taken together, HS-23 or CGA isomers exhibited anti-inflammatory therapy against LPS-induced septic mortality in mice, at least in part, mediated through the inhibition of IRAK-4. PMID:24269819
Park, Sun Hong; Roh, Eunmiri; Kim, Hyun Soo; Baek, Seung-Il; Choi, Nam Song; Kim, Narae; Hwang, Bang Yeon; Han, Sang-Bae; Kim, Youngsoo
Effect of sesame antioxidants on LPS-induced NO production by BV2 microglial cells.
Sesame antioxidants have been shown to inhibit lipid peroxidation and regulate cytokine production. In this study, we focused on the effect of sesamin and sesamolin, on nitric oxide (NO) induction by lipopolysaccharides (LPS) in the murine microglial cell line BV-2 and rat primary microglia. The results showed that sesamin and sesamolin significantly inhibited NO production, iNOS mRNA and protein expression in LPS-stimulated BV-2 cells. Sesamin or sesamolin significantly reduced LPS-activated p38 MAPK of BV-2 cells. Furthermore, SB203580, a specific inhibitor of p38 MAP kinase, dose-dependently inhibited NO production in LPS-stimulated BV-2 cells. Taken together, the inhibition of NO production might be due to the reduction of LPS-induced p38 MAPK signal pathway by sesamin and sesamolin. PMID:14534426
Hou, Rolis Chien-Wei; Chen, Huan-Lian; Tzen, Jason T C; Jeng, Kee-Ching G
Silibinin Inhibits LPS-Induced Macrophage Activation by Blocking p38 MAPK in RAW 264.7 Cells.
We demonstrate herein that silibinin, a polyphenolic flavonoid compound isolated from milk thistle (Silybum marianum), inhibits LPS-induced activation of macrophages and production of nitric oxide (NO) in RAW 264.7 cells. Western blot analysis showed silibinin inhibits iNOS gene expression. RT-PCR showed that silibinin inhibits iNOS, TNF-?, and IL1?. We also showed that silibinin strongly inhibits p38 MAPK phosphorylation, whereas the ERK1/2 and JNK pathways are not inhibited. The p38 MAPK inhibitor abrogated the LPS-induced nitrite production, whereas the MEK-1 inhibitor did not affect the nitrite production. A molecular modeling study proposed a binding pose for silibinin targeting the ATP binding site of p38 MAPK (1OUK). Collectively, this series of experiments indicates that silibinin inhibits macrophage activation by blocking p38 MAPK signaling. PMID:24244809
Youn, Cha Kyung; Park, Seon Joo; Lee, Min Young; Cha, Man Jin; Kim, Ok Hyeun; You, Ho Jin; Chang, In Youp; Yoon, Sang Pil; Jeon, Young Jin
Attenuation of lipopolysaccharide (LPS)-induced cytotoxicity by tocopherols and tocotrienols?
Lipopolysaccharide (LPS) induces host inflammatory responses and tissue injury and has been implicated in the pathogenesis of various age-related diseases such as acute respiratory distress syndrome, vascular diseases, and periodontal disease. Antioxidants, particularly vitamin E, have been shown to suppress oxidative stress induced by LPS, but the previous studies with different vitamin E isoforms gave inconsistent results. In the present study, the protective effects of ?- and ?-tocopherols and ?- and ?-tocotrienols on the oxidative stress induced by LPS against human lung carcinoma A549 cells were studied. They suppressed intracellular reactive oxygen formation, lipid peroxidation, induction of inflammatory mediator cytokines, and cell death. Tocopherols were incorporated into cultured cells much slower than tocotrienols but could suppress LPS-induced oxidative stress at much lower intracellular concentration than tocotrienols. Considering the bioavailability, it was concluded that ?-tocopherol may exhibit the highest protective capacity among the vitamin E isoforms against LPS-induced oxidative stress.
Nishio, Keiko; Horie, Masanori; Akazawa, Yoko; Shichiri, Mototada; Iwahashi, Hitoshi; Hagihara, Yoshihisa; Yoshida, Yasukazu; Niki, Etsuo
LPS induces melanogenesis through p38 MAPK activation in human melanocytes.
We have observed that lipopolysaccharide (LPS) induces pigmentation in melanocytes and in this study have examined whether these responses are mediated by the p38 mitogen-activated protein kinase (MAPK) signaling pathway. LPS appears to stimulate the pigmentation of melanocytes and cultured skin. LPS was found to induce the expression of microphthalmia-associated transcription factor (MITF) and tyrosinase protein in cells. Stimulation of melanocytes with LPS led to time dependent phosphorylation of p38 MAPK. Furthermore, p38 MAPK functionally regulated the LPS-induced melanin formation in melanocytes; a p38 MAPK inhibitor, SB203580, almost completely attenuated the LPS-mediated up-regulation of melanin synthesis and induction of MITF and tyrosinase expression. These findings indicate that activation of p38 MAPK plays an important role in LPS-induced melanogenesis by up-regulating MITF and tyrosinase expression. PMID:18478240
Ahn, Joo Hee; Jin, Sun Hee; Kang, Hee Young
Pyranocoumarins from Glehnia littoralis inhibit the LPS-induced NO production in macrophage RAW 264.7 cells.
A new dihydropyranocoumarin, (+)-cis-(3'S,4'S)-diisobutyrylkhellactone (1), together with five known compounds, 3'-senecioyl-4'-acetylkhellactone (2), 3'-isovaleryl-4'-acetylkhellactone (3), 3',4'-disenecioylkhellactone (4), 3'-isovaleryl-4'-senecioylkhellactone (5), and 3',4'-diisovalerylkhellactone (6), was isolated from Glehnia littoralis. Their chemical structures were elucidated based on the spectroscopic data interpretation, particularly 1D and 2D NMR data including HMQC and HMBC. All the isolated compounds showed the potential to inhibit LPS-induced nitric oxide production in RAW 264.7 cells with IC50 values ranging from 7.4 to 44.3?M. PMID:24813739
Lee, Jin Woo; Lee, Chul; Jin, Qinghao; Yeon, Eung Tae; Lee, Dongho; Kim, Soo-Young; Han, Sang Bae; Hong, Jin Tae; Lee, Mi Kyeong; Hwang, Bang Yeon
Myrrh Inhibits LPS-Induced Inflammatory Response and Protects from Cecal Ligation and Puncture-Induced Sepsis
Myrrh has been used as an antibacterial and anti-inflammatory agent. However, effect of myrrh on peritoneal macrophages and clinically relevant models of septic shock, such as cecal ligation and puncture (CLP), is not well understood. Here, we investigated the inhibitory effect and mechanism(s) of myrrh on inflammatory responses. Myrrh inhibited LPS-induced productions of inflammatory mediators such as nitric oxide, prostaglandin E2, and tumor necrosis factor-? but not of interleukin (IL)-1? and IL-6 in peritoneal macrophages. In addition, Myrrh inhibited LPS-induced activation of c-jun NH2-terminal kinase (JNK) but not of extracellular signal-regulated kinase (ERK), p38, and nuclear factor-?B. Administration of Myrrh reduced the CLP-induced mortality and bacterial counts and inhibited inflammatory mediators. Furthermore, administration of Myrrh attenuated CLP-induced liver damages, which were mainly evidenced by decreased infiltration of leukocytes and aspartate aminotransferase/alanine aminotransferase level. Taken together, these results provide the evidence for the anti-inflammatory and antibacterial potential of Myrrh in sepsis.
Kim, Min-Sun; Bae, Gi-Sang; Park, Kyoung-Chel; Koo, Bon Soon; Kim, Byung-Jin; Lee, Hye-Jin; Seo, Sang-Wan; Shin, Yong Kook; Jung, Won-Seok; Cho, Jung-Hee; Kim, Youn-Chul; Kim, Tae-Hyeon; Song, Ho-Joon; Park, Sung-Joo
Kuromoji (Lindera umbellata) essential oil inhibits LPS-induced inflammation in RAW 264.7 cells.
Kuromoji (Lindera umbellata) essential oil (KEO) has long been used in Japan as a traditional medicine. It contains linalool (C10H18O), a naturally occurring small terpenoid. For this study, we investigated the anti-inflammatory effect of KEO in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Mouse macrophage-like RAW 264.7 cells were stimulated with LPS. Then they were treated with 25 or 50 �g/mL of KEO for 24 h. KEO suppressed LPS-induced pro-inflammatory cytokine production such as that of nitric oxide (NO), interleukin-6 (IL-6), and tumor necrosis factor-? (TNF-?) in a dose-dependent manner. In addition, inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) mRNA expression and protein levels were suppressed by treatment with KEO cells. In addition, by treatment with 25 or 50 �g/mL of linalool showed the same anti-inflammatory effect. The results suggest that KEO and linalool can be regarded as a natural resource for use in anti-inflammatory therapeutic products. PMID:23470753
Maeda, Hayato; Yamazaki, Mao; Katagata, Yohtaro
ETD-02-045 (GAVETT) GPRA # 10108 Neutrophils Play a Critical Role in the Development of LPS-Induced Airway Disease. Jordan D. Savov, Stephen H. Gavett*, David M. Brass, Daniel L. Costa*, and David A. Schwartz ABSTRACT We investigated the role of neutrophils...
Emodin inhibits LPS-induced inflammatory response by activating PPAR-? in mouse mammary epithelial cells.
Emodin, an anthraquinone derivative isolated from the rhizomes of Rheum palmatum, has been reported to have a protective effect against lipopolysaccharide (LPS)-induced mastitis. However, the underlying molecular mechanisms are not well understood. The aim of this study was to investigate the molecular mechanisms of emodin in modifying lipopolysaccharide (LPS)-induced signaling pathways in mouse mammary epithelial cells (MEC). The pro-inflammatory cytokines were determined by ELISA. Nuclear factor-?B (NF-?B), inhibitory kappa B (I?B?) protein, p38, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and PPAR-? were determined by Western blotting. The results showed that emodin suppressed tumor necrosis factor-alpha (TNF-?), interleukin-6 (IL-6), iNOS and COX-2 expression. We also found that emodin inhibited LPS-induced NF-?B activation, I?B? degradation, phosphorylation of ERK, JNK and P38. Furthermore, emodin could activate PPAR-? and the anti-inflammatory effects of emodin can be reversed by GW9662, a specific antagonist for PPAR-?. In conclusion, our results demonstrate that emodin activates PPAR-?, thereby attenuating LPS-induced inflammatory response. PMID:24874440
Yang, Zhengtao; Zhou, Ershun; Wei, Dong; Li, Depeng; Wei, Zhengkai; Zhang, Wen; Zhang, Xichen
Effects of voluntary wheel running on LPS-induced sickness behavior in aged mice.
Peripheral stimulation of the innate immune system with LPS causes exaggerated neuroinflammation and prolonged sickness behavior in aged mice. Regular moderate intensity exercise has been shown to exert anti-inflammatory effects that may protect against inappropriate neuroinflammation and sickness in aged mice. The purpose of this study was to test the hypothesis that voluntary wheel running would attenuate LPS-induced sickness behavior and proinflammatory cytokine gene expression in ~22-month-old C57BL/6J mice. Mice were housed with a running wheel (VWR), locked-wheel (Locked), or no wheel (Standard) for 10 weeks, after which they were intraperitoneally injected with LPS across a range of doses (0.02, 0.08, 0.16, 0.33 mg/kg). VWR mice ran on average 3.5 km/day and lost significantly more body weight and body fat, and increased their forced exercise tolerance compared to Locked and Shoebox mice. VWR had no effect on LPS-induced anorexia, adipsia, weight-loss, or reductions in locomotor activity at any LPS dose when compared to Locked and Shoebox groups. LPS induced sickness behavior in a dose-dependent fashion (0.33>0.02 mg/kg). Twenty-four hours post-injection (0.33 mg/kg LPS or Saline) we found a LPS-induced upregulation of whole brain TNF?, IL-1?, and IL-10 mRNA, and increased IL-1? and IL-6 in the spleen and liver; these effects were not attenuated by VWR. We conclude that VWR does not reduce LPS-induced exaggerated or prolonged sickness behavior in aged animals, or 24h post-injection (0.33 mg/kg LPS or Saline) brain and peripheral proinflammatory cytokine gene expression. The necessity of the sickness response is critical for survival and may outweigh the subtle benefits of exercise training in aged animals. PMID:23277090
Martin, Stephen A; Pence, Brandt D; Greene, Ryan M; Johnson, Stephanie J; Dantzer, Robert; Kelley, Keith W; Woods, Jeffrey A
Oroxylin-A Rescues LPS-Induced Acute Lung Injury via Regulation of NF-?B Signaling Pathway in Rodents
Background and Purpose Successful drug treatment for sepsis-related acute lung injury (ALI) remains a major clinical problem. This study was designed to assess the beneficial effects of post-treatment of oroxylin A (OroA), a flavonoid, in ameliorating lipopolysaccharides (LPS)-induced lung inflammation and fatality. Experimental Approach Rats were injected with LPS (10 mg/kg, iv) to induce ALI, and OroA was given (15 mg/kg, iv) 1 hr or 6 hrs after LPS challenge. Twenty four hrs after LPS challenge, biochemical changes in the blood and lung tissues, and morphological/histological alterations in the lung associated with inflammation and injury were examined. Therapeutic effect of OroA was assessed by measuring the survival rate in endotoxemic mice. Key Results LPS (10 mg/kg, iv) significantly altered WBC counts, elevated plasma tumor necrosis factor (TNF)-? and nitric oxide (NO), increased pulmonary edema, thickened alveolar septa, and decreased survival rate. These changes were ameliorated by OroA (15 mg/kg, iv) administered 1 hr or 6 hrs after LPS challenge. This post-treatment also significantly attenuated LPS-induced activation of nuclear factor-?B (NF-?B) and the release of high mobility group box 1 (HMGB1) in lung tissues. Furthermore, post-treatment with OroA (60 mg/kg, ip) administered 1 hr or 6 hrs after LPS challenge in mice significantly increased survival rate. Conclusion and Implication OroA administered after induction of ALI by LPS significantly prevent and revere lung tissues injuries with increased survival rate. Positive post-treatment effects of OroA suggest that OroA is a potentially useful candidate for managing lung inflammation in LPS-induced endotoxemia and septic shock.
Tseng, Tzu-Ling; Chen, Mei-Fang; Tsai, Ming-Jen; Hsu, Yung-Hsiang; Chen, Chin-Piao; Lee, Tony J. F.
NOS2 regulation of LPS-induced airway inflammation via S-nitrosylation of NF-?B p65
Inducible nitric oxide synthase (NOS2) expression is increased in the airway epithelium in acute inflammatory disorders although the physiological impact remains unclear. We have previously shown that NOS2 inhibits NF-?B (p50-p65) activation in respiratory epithelial cells by inducing S-nitrosylation of the p65 monomer (SNO-p65). In addition, we have demonstrated that mouse lung SNO-p65 levels are acutely depleted in a lipopolysaccharide (LPS) model of lung injury and that augmenting SNO-p65 levels before LPS treatment results in decreased airway epithelial NF-?B activation, airway inflammation, and lung injury. We now show that aerosolized LPS induces NOS2 expression in the respiratory epithelium concomitant with an increase in lung SNO-p65 levels and a decrease in airway NF-?B activity. Genetic deletion of NOS2 results in an absence of SNO-p65 formation, persistent NF-?B activity in the respiratory epithelium, and prolonged airway inflammation. These results indicate that a primary function of LPS-induced NOS2 expression in the respiratory epithelium is to modulate the inflammatory response through deactivation of NF-?B via S-nitrosylation of p65, thereby counteracting the initial stimulus-coupled denitrosylation.
Kelleher, Zachary T.; Potts, Erin N.; Brahmajothi, Mulugu V.; Foster, Matthew W.; Auten, Richard L.; Foster, W. Michael
Effects of cannabinoid receptor ligands on LPS-induced pulmonary inflammation in mice
The effects of cannabinoid receptor agonists WIN 55,212-2, ?9-tetrahydrocannabinol (?9-THC), arachidonoylethanolamide (anandamide) and palmitoylethanolamide on lipopolysaccharide (LPS) -induced bronchopulmonary inflammation in mice were investigated. WIN 55,212-2 and ?9-THC induced a concentration-dependent decrease in TNF-? level in the bronchoalveolar lavage fluid (BALF) (maximum inhibition 52.7% and 36.9% for intranasal doses of 750 nmol.kg?1 and 2.65 mmol.kg?1, respectively). This effect was accompanied
E. Berdyshev; E. Boichot; M. Corbel; N. Germain; V Lagente
Attenuation of LPS-induced lung inflammation by glucosamine in rats.
Acute inflammation is often observed during acute lung injury (ALI) and acute respiratory distress syndrome. Glucosamine is known to act as an anti-inflammatory molecule. The effects of glucosamine on acute lung inflammation and its associated mechanisms remain unclear. The present study sought to address how glucosamine plays an anti-inflammatory role in acute lung inflammation in vivo and in vitro. Using the LPS intratracheal instillation-elicited rat lung inflammation model, we found that glucosamine attenuated pulmonary edema and polymorphonuclear leukocyte infiltration, as well as the production of TNF-?, IL-1?, cytokine-induced neutrophil chemoattractant (CINC)-1, macrophage inflammatory protein (MIP)-2, and nitric oxide (NO) in the bronchoalveolar lavage fluid (BALF) and in the cultured medium of BALF cells. The expression of TNF-?, IL-1?, IFN-?, CINC-1, MIP-2, monocyte chemotactic protein-1, and inducible NO synthase (iNOS) in LPS-inflamed lung tissue was also suppressed by glucosamine. Using the rat alveolar epithelial cell line L2, we noted that the cytokine mixture (cytomix)-regulated production and mRNA expression of CINC-1 and MIP-2, NO production, the protein and mRNA expression of iNOS, iNOS mRNA stability, and iNOS promoter activity were all inhibited by glucosamine. Furthermore, glucosamine reduced LPS-mediated NF-?B signaling by decreasing I?B phosphorylation, p65 nuclear translocation, and NF-?B reporter activity. Overexpression of the p65 subunit restored the inhibitory action of glucosamine on cytomix-regulated NO production and iNOS expression. In conclusion, glucosamine appears to act as an anti-inflammatory molecule in LPS-induced lung inflammation, at least in part by targeting the NF-?B signaling pathway. PMID:23898954
Chuang, Kun-Han; Peng, Yen-Chun; Chien, Han-Yun; Lu, Meng-Lun; Du, Hsin-I; Wu, Yuh-Lin
Effects of ginsenoside Re on LPS-induced inflammatory mediators in BV2 microglial cells
Background Microglial activation plays an important role in neurodegenerative diseases by producing several pro-inflammatory enzymes and pro-inflammatory cytokines. Lipopolysaccharide (LPS)-induced inflammation leads to the activation of microglial cells in the central nervous system (CNS) and is associated with the pathological mechanisms of neurodegenerative diseases, including PD, AD, and ALS. Ginseng is a natural antioxidant used in herbal medicine and contains ginsenosides (Rb1, Rg1, Rg3, Re, and Rd), which have anti-neoplastic and anti-stress properties. This study demonstrates the involvement of the anti-inflammatory signaling pathway, ginsenoside-Re (G-Re), which is one of the ginsenosides mediated by LPS-induced neuroinflammation in BV2 microglial cells. Methods BV2 microglial cells were pretreated with 2 ?g/ml G-Re and stimulated with 1 ?g/ml LPS to induce neuroinflammation. To investigate the effect of G-Re on LPS-induced cell signaling, we performed western blotting and immunofluorescence using specific antibodies, such as phospho-p38, COX2, and iNOS. Results Pretreatment with 2 ?g/ml G-Re was neuroprotective against 1 ?g/ml LPS-treated microglial cells. The neuroprotective events induced by G-Re treatment in neuroinflammation occurred via the phospho-p38, iNOS, and COX2 signaling pathways in BV2 cells. Conclusion Taken together, we suggest that G-Re exerts a beneficial effect on neuroinflammatory events in neurodegenerative diseases.
Protective effect of resveratrol against LPS-induced extracellular lipoperoxidation in AR42J cells partly via a Myd88-dependent signaling pathway
Lipopolysaccharides (LPS) are major components of the cell wall of Gram negative bacteria implicated in the pathogenesis of bacterial infection. Resveratrol is a polyphenolic phytoalexin exhibiting antioxidant and anti-inflammatory properties. We investigated the protective effects of this natural compound on LPS-induced proinflammatory effect using non-myeloid AR42J pancreatic cells. We found that LPS dose-dependently increased extracellular malondialdehyde (MDA) and nitric oxide
Hichem Sebai; Elodie Ristorcelli; Veronique Sbarra; Sonia Hovsepian; Guy Fayet; Ezzedine Aouani; Dominique Lombardo
The role of macrophages in LPS-induced lethality and tissue injury.
In the present study we investigated the role of mononuclear phagocytes in the pathogenesis of lipopolysaccharide (LPS)-induced lethality and tissue injury. Since hepatic and splenic macrophages are the primary sites of localization of i.v.-injected LPS, we selectively eliminated these macrophages using liposome-encapsulated dichloromethylene diphosphonate (DMDP). After double DMDP-liposome treatment the phagocytic cells in the liver and spleen were completely eliminated, except for the macrophages in the white pulp of the spleen which were affected to a lesser extent by this treatment. An i.v. injection of LPS into DMDP- and saline-pretreated mice showed that the latter animals exhibited febrile-associated symptoms such as lethargy and ruffled fur, but that macrophage elimination abrogated these symptoms. Although after double saline- or DMDP-pretreatment the LD50 appears to be 1 mg and 630 micrograms, respectively, the differences in lethality between both groups of mice were not statistically significant. Therefore, we concluded that hepatic and splenic macrophages are not necessary for LPS-induced lethality. The role of macrophages in LPS-induced local tissue damage was studied by comparing the histopathological changes in hepatic and splenic tissue between DMDP- and saline-pretreated mice. A sublethal dose of LPS induced similar hepatic lesions in macrophage-depleted and saline-pretreated mice, whereas the histopathological changes in the spleen were much more pronounced after DMDP-pretreatment. Particularly in the inner periarteriolar lymphocyte sheath (PALS) of these mice, the number of T cells was considerably reduced and extensive cellular necrosis could be found. These data strongly suggest that the local tissue damage resulting from LPS injection may not be due to its localization in mononuclear phagocytes but rather to interaction with other cell types. Images Figure 1 Figure 2 Figure 3
Groeneveld, P H; Claassen, E; Kuper, C F; Van Rooijen, N
Fasting Exacerbates and Feeding Diminishes LPS-Induced Liver Injury in the Rat
Introduction Enteral nutrition improves clinical outcomes. The effects of feeding on LPS induced liver injury are unknown. We hypothesized\\u000a that feeding would attenuate liver injury from LPS. Methods Fasted or fed rats were given LPS (20�mg\\/kg ip) or saline for 5 h and sacrificed. Serum aminotransferases and cytokines (immunoassay)\\u000a were measured. Oxidative stress protein (iNOS, COX2, and HO1) assessments (Western
Sasha D. Adams; Benjamin A. Delano; Kenneth S. Helmer; David W. Mercer
Minocycline attenuates lipopolysaccharide (LPS)-induced neuroinflammation, sickness behavior, and anhedonia
BACKGROUND: Activation of the peripheral innate immune system stimulates the secretion of CNS cytokines that modulate the behavioral symptoms of sickness. Excessive production of cytokines by microglia, however, may cause long-lasting behavioral and cognitive complications. The purpose of this study was to determine if minocycline, an anti-inflammatory agent and purported microglial inhibitor, attenuates lipopolysaccharide (LPS)-induced neuroinflammation, sickness behavior, and anhedonia.
Christopher J Henry; Yan Huang; Angela Wynne; Mark Hanke; Justin Himler; Michael T Bailey; John F Sheridan; Jonathan P Godbout
Endothelial P2Y 2 receptor regulates LPS-induced neutrophil transendothelial migration in vitro
Previous studies showed that P2 receptors are involved in neutrophil migration via stimulation of chemokine release and by facilitating chemoattractant gradient sensing. Here, we have investigated whether these receptors are involved in LPS-induced neutrophil transendothelial migration (TEM) using a Boyden chamber where neutrophils migrated through a layer of lipopolysaccharide (LPS)-stimulated human umbilical vein endothelial cells (HUVECs). In line with a
Filip Kukulski; Fethia Ben Yebdri; Fariborz Bahrami; Michel Fausther; Alain Tremblay; Jean S�vigny
Extracellular nucleotides mediate LPS-induced neutrophil migration in vitro and in vivo
Extracellular nucleotides are emerging as important inflammatory mediators. Here, we demonstrate that these molecules mediate LPS- induced neutrophil migration in vitro and in vivo. Apyrase, a nucleotide scavenger, reduced the abil- ity of LPS-stimulated monocytes to recruit neutro- phils, as assayed using a modified Boyden cham- ber. This effect resulted from the inhibition of IL-8 release from monocytes. Furthermore, LPS-in-
Filip Kukulski; Fethia Ben Yebdri; Julie Lefebvre; Michel Warny; Philippe A. Tessier; Jean Sevigny
Sphingomyelin synthase 2 (SMS2) deficiency attenuates LPS-induced lung injury
Sphingomyelin synthase (SMS) catalyzes the synthesis of sphingomyelin (SM) and is required for maintenance of plasma membrane microdomain fluidity. Of the two isoforms of mammalian SMS, SMS1 is mostly present in the trans-Golgi apparatus, whereas SMS2 is predominantly found at the plasma membrane. SMS2 has a role in receptor mediated response to inflammation in macrophages, however, the role of SMS2 in vascular permeability, pulmonary edema, and lung injury have not been investigated. To define the role of SMS activation in lung injury, we utilized a lipopolysaccharide (LPS)-induced lung edema model. SMS activity was measured and correlated with the severity of lung injury. Within 4 h of LPS treatment, SMS activity was increased significantly and remained upregulated up to 24 h. Comparison of LPS-induced lung injury in SMS2 knockout (SMS2?/?) and wild-type littermate control mice showed that inflammation, cytokine induction, and lung injury were significantly inhibited in SMS2?/? mice. Our results suggest that a deficiency of SMS2 can diminish the extent of pulmonary edema and lung injury. Furthermore, we show that depletion of SMS2 was sufficient to decrease MAP kinase-JNK activation, severity of LPS-induced pulmonary neutrophil influx, and inflammation, suggesting a novel role of SMS2 activation in lung injury.
Gowda, Satish; Yeang, Calvin; Wadgaonkar, Sunil; Anjum, Fatima; Grinkina, Natalia; Cutaia, Michael; Jiang, Xian-Chen
Alcohol exacerbates LPS-induced fibrosis in subclinical acute pancreatitis.
The role of pancreatic acinar cells in initiating fibrogenic responses during the early stages of alcoholic acute pancreatitis has not been evaluated. We investigated the ability of injured acinar cells to generate pancreatic fibrosis in acute pancreatitis. Rats were fed either an ethanol-containing or control diet over 14 weeks and euthanized 3 or 24 hours after a single lipopolysaccharide injection. Profibrotic transforming growth factor-? of acinar cells and pancreatic fibrosis were assessed by immunofluorescence, histological characteristics, and electron microscopy. Human pancreatic tissues were also evaluated. Periacinar cell fibrosis and collagen were exacerbated 24 hours after endotoxemia in alcohol-fed rats. Alcohol exposure exacerbated acinar cell-specific production of transforming growth factor ? in response to lipopolysaccharide in vivo and in acinar cell-like AR42J cells in vitro. Although a morphological examination showed no visible signs of necrosis, early pancreatic fibrosis can be initiated by little or no pancreatic necrosis. Transforming growth factor ? was also significantly increased in human acinar cells from patients with acute/recurrent pancreatitis compared with chronic pancreatitis tissue. Alcohol exacerbates lipopolysaccharide-induced pancreatic fibrosis during the early onset of mild, subclinical, acute pancreatitis. We suggest that multiple, subclinical, acute pancreatitis episodes can accumulate in fibrosis during the development of chronic pancreatitis, even if there is no history of acute pancreatitis. PMID:24091223
Gu, Haitao; Fortunato, Franco; Bergmann, Frank; B�chler, Markus W; Whitcomb, David C; Werner, Jens
Anti-inflammatory effects of chicanine on murine macrophage by down-regulating LPS-induced inflammatory cytokines in I?B?/MAPK/ERK signaling pathways.
Schisandra chinensis Baill is a Chinese traditional medicine with multiple pharmacological activities. In this study, chicanine, one of the major lignan compounds of S. chinesis, was investigated for suppressive effects on lipopolysaccharide (LPS)-induced inflammatory responses in murine macrophages (RAW 264.7 cells). Chicanine was found to have anti-inflammatory properties with the inhibition of nitric oxide (NO) and Prostaglandin E (2) (PGE2) production and nuclear factor-?B (NF-?B) signaling in LPS-stimulated RAW 264.7 cells with no cytotoxic effects. Treatment of RAW 264.7 cells with chicanine down-regulated LPS-induced expression of pro-inflammatory cytokines including TNF?, IL-1?, MCP-1, G-CSF, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS). These inhibitory effects were found with the blockage of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinases 1 and 2 (ERK 1/2), and also I?B-? phosphorylation. These results indicated that anti-inflammatory actions of chicanine in macrophages involved inhibition of LPS-induced TLR4-I?B?/MAPK/ERK signaling pathways. PMID:24361309
Chen, Haixia; Sohn, Johann; Zhang, Likang; Tian, Jingge; Chen, Shuhan; Bjeldanes, Leonard F
Adenosine receptor A3 is a critical mediator in LPS-induced pulmonary inflammation.
Adenosine receptor A(3) (A(3)) regulates directed movement of polymorphonuclear cells (PMNs) to sites of inflammation and has been implicated as a relevant mediator in models of inflammatory diseases. Here, we sought to characterize the role of A(3) in a murine model of lung inflammation. Initial studies revealed that pulmonary A(3) transcript levels were elevated following LPS exposure in vivo. In addition, inhalation of LPS increased the accumulation of PMNs in wild-type and A(3)(-/-) mice in all lung compartments. Pretreatment with the specific A(3)-agonist Cl-IB-MECA significantly decreased migration of PMNs into lung interstitium and alveolar air space of wild-type mice but not of A(3)(-/-) mice. Lower PMN counts were associated with reduced levels of TNF-? and IL-6 in the alveolar space of wild-type mice that received Cl-IB-MECA. In addition, Cl-IB-MECA attenuated LPS-induced microvascular permeability in wild-type mice as assessed by the extravasation of Evans blue. In pulmonary microvascular endothelial cells, Cl-IB-MECA reduced LPS-induced cytoskeletal remodeling and cell retraction, consistent with a specific role of A(3) for maintaining endothelial integrity. Migratory activity of human PMNs across an endothelial or epithelial monolayer was reduced when A(3) was activated on PMNs. Studies in chimeric mice, however, revealed that Cl-IB-MECA required A(3) on both hematopoietic and nonhematopoietic cells to reduce transmigration in vivo. Together, our results shed new light on the role of A(3) in LPS-induced PMN trafficking in the lung and suggest pharmacological modulation of A(3)-dependent pathways as a promising approach in lung inflammation. PMID:20639349
Wagner, Rosalyn; Ngamsri, Kristian-Christos; Stark, Stefanie; Vollmer, Irene; Reutershan, J�rg
Molecular hydrogen reduces LPS-induced neuroinflammation and promotes recovery from sickness behaviour in mice.
Molecular hydrogen has been shown to have neuroprotective effects in mouse models of acute neurodegeneration. The effect was suggested to be mediated by its free-radical scavenger properties. However, it has been shown recently that molecular hydrogen alters gene expression and protein phosphorylation. The aim of this study was to test whether chronic ad libitum consumption of molecular hydrogen-enriched electrochemically reduced water (H-ERW) improves the outcome of lipopolysaccharide (LPS)-induced neuroinflammation. Seven days after the initiation of H-ERW treatment, C57Bl/6 mice received a single injection of LPS (0.33 mg/kg i.p.) or an equivalent volume of vehicle. The LPS-induced sickness behaviour was assessed 2 h after the injection, and recovery was assessed by monitoring the spontaneous locomotor activity in the homecage for 72 h after the administration of LPS. The mice were killed in the acute or recovery phase, and the expression of pro- and antiinflammatory cytokines in the hippocampus was assessed by real-time PCR. We found that molecular hydrogen reduces the LPS-induced sickness behaviour and promotes recovery. These effects are associated with a shift towards anti-inflammatory gene expression profile at baseline (downregulation of TNF- ? and upregulation of IL-10). In addition, molecular hydrogen increases the amplitude, but shortens the duration and promotes the extinction of neuroinflammation. Consistently, molecular hydrogen modulates the activation and gene expression in a similar fashion in immortalized murine microglia (BV-2 cell line), suggesting that the effects observed in vivo may involve the modulation of microglial activation. Taken together, our data point to the regulation of cytokine expression being an additional critical mechanism underlying the beneficial effects of molecular hydrogen. PMID:22860058
Spulber, Stefan; Edoff, Karin; Hong, Lie; Morisawa, Shinkatsu; Shirahata, Sanetaka; Ceccatelli, Sandra
Molecular Hydrogen Reduces LPS-Induced Neuroinflammation and Promotes Recovery from Sickness Behaviour in Mice
Molecular hydrogen has been shown to have neuroprotective effects in mouse models of acute neurodegeneration. The effect was suggested to be mediated by its free-radical scavenger properties. However, it has been shown recently that molecular hydrogen alters gene expression and protein phosphorylation. The aim of this study was to test whether chronic ad libitum consumption of molecular hydrogen-enriched electrochemically reduced water (H-ERW) improves the outcome of lipopolysaccharide (LPS)-induced neuroinflammation. Seven days after the initiation of H-ERW treatment, C57Bl/6 mice received a single injection of LPS (0.33 mg/kg i.p.) or an equivalent volume of vehicle. The LPS-induced sickness behaviour was assessed 2 h after the injection, and recovery was assessed by monitoring the spontaneous locomotor activity in the homecage for 72 h after the administration of LPS. The mice were killed in the acute or recovery phase, and the expression of pro- and antiinflammatory cytokines in the hippocampus was assessed by real-time PCR. We found that molecular hydrogen reduces the LPS-induced sickness behaviour and promotes recovery. These effects are associated with a shift towards anti-inflammatory gene expression profile at baseline (downregulation of TNF- ? and upregulation of IL-10). In addition, molecular hydrogen increases the amplitude, but shortens the duration and promotes the extinction of neuroinflammation. Consistently, molecular hydrogen modulates the activation and gene expression in a similar fashion in immortalized murine microglia (BV-2 cell line), suggesting that the effects observed in vivo may involve the modulation of microglial activation. Taken together, our data point to the regulation of cytokine expression being an additional critical mechanism underlying the beneficial effects of molecular hydrogen.
Spulber, Stefan; Edoff, Karin; Hong, Lie; Morisawa, Shinkatsu; Shirahata, Sanetaka; Ceccatelli, Sandra
LPS-induced cytokine production in human dendritic cells is regulated by sialidase activity
Removal of sialic acid from glycoconjugates on the surface of monocytes enhances their response to bacterial LPS. We tested the hypothesis that endogenous sialidase activity creates a permissive state for LPS-induced cytokine production in human monocyte-derived DCs. Of the four genetically distinct sialidases (Neu1�4), Neu1, Neu3, and Neu4 are expressed in human monocytes, but only Neu1 and Neu3 are up-regulated as cells differentiate into DCs. Neu1 and Neu3 are present on the surface of monocytes and DCs and are also present intracellularly. DCs contain a greater amount of sialic acid than monocytes, but the amount of sialic acid/mg total protein declines during differentiation to DCs. This relative hyposialylation of cells does not occur in mature DCs grown in the presence of zanamivir, a pharmacologic inhibitor of Neu3 but not Neu1, or DANA, an inhibitor of Neu1 and Neu3. Inhibition of sialidase activity during differentiation to DCs causes no detectable change in cell viability or expression of DC surface markers. Differentiation of monocytes into DCs in the presence of zanamivir results in reduced LPS- induced expression of IL-6, IL-12p40, and TNF-? by mature DCs, demonstrating a role for Neu3 in cytokine production. A role for Neu3 is supported by inhibition of cytokine production by DANA in DCs from Neu1�/� and WT mice. We conclude that sialidase-mediated change in sialic acid content of specific cell surface glycoconjugates in DCs regulates LPS-induced cytokine production, thereby contributing to development of adaptive immune responses.
Stamatos, Nicholas M.; Carubelli, Ivan; van de Vlekkert, Diantha; Bonten, Erik J.; Papini, Nadia; Feng, Chiguang; Venerando, Bruno; d'Azzo, Alessandra; Cross, Alan S.; Wang, Lai-Xi; Gomatos, Peter J.
Endothelial P2Y2 receptor regulates LPS-induced neutrophil transendothelial migration in vitro.
Previous studies showed that P2 receptors are involved in neutrophil migration via stimulation of chemokine release and by facilitating chemoattractant gradient sensing. Here, we have investigated whether these receptors are involved in LPS-induced neutrophil transendothelial migration (TEM) using a Boyden chamber where neutrophils migrated through a layer of lipopolysaccharide (LPS)-stimulated human umbilical vein endothelial cells (HUVECs). In line with a role of P2 receptors, neutrophil TEM was inhibited by the P2 receptor antagonists suramin and reactive blue 2 (RB-2) acting on the basolateral, but not luminal, HUVECs' P2 receptors. HUVECs express P2Y(1), P2Y(2), P2Y(4), P2Y(6) and P2Y(11). The involvement of P2Y(4) was unlikely as this receptor is insensitive to suramin while P2Y(1), P2Y(6) and P2Y(11) were excluded with available selective antagonists, leaving P2Y(2) as the only candidate. Indeed, the P2Y(2) knockdown in HUVECs inhibited neutrophil TEM compared to control HUVECs transfected with scrambled siRNA. Moreover, UTP, a P2Y(2) ligand, markedly potentiated LPS-induced TEM. Interestingly, IL-8 and ICAM-1 had a modest effect on neutrophil TEM in this 3h assay which was significantly diminished by the inhibition of Rho kinase in HUVECs with Y27632. In summary, endothelial P2Y(2) receptors control the early LPS-induced neutrophil TEM in vitro via Rho kinase activation. PMID:20022380
Kukulski, Filip; Ben Yebdri, Fethia; Bahrami, Fariborz; Fausther, Michel; Tremblay, Alain; S�vigny, Jean
Minocycline attenuates lipopolysaccharide (LPS)-induced neuroinflammation, sickness behavior, and anhedonia
Background Activation of the peripheral innate immune system stimulates the secretion of CNS cytokines that modulate the behavioral symptoms of sickness. Excessive production of cytokines by microglia, however, may cause long-lasting behavioral and cognitive complications. The purpose of this study was to determine if minocycline, an anti-inflammatory agent and purported microglial inhibitor, attenuates lipopolysaccharide (LPS)-induced neuroinflammation, sickness behavior, and anhedonia. Methods In the first set of experiments the effect of minocycline pretreatment on LPS-induced microglia activation was assessed in BV-2 microglia cell cultures. In the second study, adult (3�6 m) BALB/c mice received an intraperitoneal (i.p.) injection of vehicle or minocycline (50 mg/kg) for three consecutive days. On the third day, mice were also injected (i.p.) with saline or Escherichia coli LPS (0.33 mg/kg) and behavior (i.e., sickness and anhedonia) and markers of neuroinflammation (i.e., microglia activation and inflammatory cytokines) were determined. In the final study, adult and aged BALB/c mice were treated with the same minocycline and LPS injection regimen and markers of neuroinflammation were determined. All data were analyzed using Statistical Analysis Systems General Linear Model procedures and were subjected to one-, two-, or three-way ANOVA to determine significant main effects and interactions. Results Minocycline blocked LPS-stimulated inflammatory cytokine secretion in the BV-2 microglia-derived cell line and reduced LPS-induced Toll-like-receptor-2 (TLR2) surface expression on brain microglia. Moreover, minocycline facilitated the recovery from sickness behavior (i.e., anorexia, weight loss, and social withdrawal) and prevented anhedonia in adult mice challenged with LPS. Furthermore, the minocycline associated recovery from LPS-induced sickness behavior was paralleled by reduced mRNA levels of Interleukin (IL)-1?, IL-6, and indoleamine 2, 3 dioxygenase (IDO) in the cortex and hippocampus. Finally, in aged mice, where exaggerated neuroinflammation was elicited by LPS, minocycline pretreatment was still effective in markedly reducing mRNA levels of IL-1?, TLR2 and IDO in the hippocampus. Conclusion These data indicate that minocycline mitigates neuroinflammation in the adult and aged brain and modulates the cytokine-associated changes in motivation and behavior.
Henry, Christopher J; Huang, Yan; Wynne, Angela; Hanke, Mark; Himler, Justin; Bailey, Michael T; Sheridan, John F; Godbout, Jonathan P
Iloprost improves endothelial barrier function in LPS-induced lung injury
RATIONALE Protective effects of prostacyclin and its stable analog Iloprost are mediated by elevation of intracellular cAMP leading to enhancement of peripheral actin cytoskeleton and cell-cell adhesive structures. This study tested hypothesis that iloprost may exhibit protective effects against lung injury and endothelial barrier dysfunction induced by bacterial wall lypopolysacharide (LPS). METHODS Endothelial barrier dysfunction was assessed by measurements of transendothelial permeability, morphologically, and analysis of LPS-activated inflammatory signaling. In vivo, C57BL/6J mice were challenged with LPS with or without iloprost or 8-bromoadenosine-3?,5?-cyclic monophosphate (Br-cAMP) treatment. Lung injury was monitored by measurements of bronchoalveolar lavage protein content, cell count, and Evans blue extravasation. RESULTS Iloprost and Br-cAMP attenuated disruption of endothelial monolayer and suppressed activation of p38 mitogen activated protein (MAP) kinase, NF?B pathway, Rho signaling, ICAM1 expression, and neutrophil migration after LPS challenge. In vivo, iloprost was effective against LPS-induced protein and neutrophil accumulation in bronchoalveolar lavage fluid and reduced myeloperoxidase activation, ICAM-1 expression, and Evans blue extravasation in the lungs. Inhibition of Rac activity abolished barrier protective and anti-inflammatory effects of iloprost and Br-cAMP. CONCLUSION Iloprost-induced elevation of intracellular cAMP triggers Rac signaling, which attenuates LPS-induced NF?B and p38 MAPK inflammatory pathways and Rho-dependent mechanism of endothelial permeability.
Birukova, Anna A.; Wu, Tinghuai; Tian, Yufeng; Meliton, Angelo; Sarich, Nicolene; Tian, Xinyong; Leff, Alan; Birukov, Konstantin G.
Propylene-Glycol Aggravates LPS-Induced Sepsis through Production of TNF-? and IL-6.
Background: Propylene glycol (1,2-propanediol, PG) is a commonly used solvent for oral, intravenous, as well as topical pharmaceutical preparations. While PG is generally considered to be safe, it has been known that large intravenous doses given over a short period of time can be toxic. Objective: To evaluate the effect of PG in sepsis induced by the bacterial endotoxin lipopolysaccharide (LPS). Methods: Balb/c mice were treated with LPS (1 mg/kg b.w., i.p.) with or without PG (5 g/kg b.w. i.v.). The survival rate and the production of inflammatory cytokines were measured. In RAW264.7 mouse macrophages encoding NF-kB-luc reporter gene, the nuclear transcription factor kappa-B (NF-kB) activation was measured. Results: We found that intravenous PG increased the mortality rate in sepsis induced by the bacterial endotoxin lipopolysaccharide (LPS) in mice. In accordance with that, PG enhanced LPS-induced production of inflammatory cytokines, including tumor necrosis factor-? (TNF-?) and interleukin-6 (IL-6) in vivo. PG also increased the LPS-induced macrophage activation in vitro as detected by measuring NF-kB activation. Conclusion: Our results indicate that drugs containing high doses of PG can pose a risk when administered to patients suffering from or prone to Gram negative bacterial infection. PMID:24975968
Marton, Annamaria; Kolozsi, Csongor; Kusz, Erzsebet; Olah, Zoltan; Letoha, Tamas; Vizler, Csaba; Pecze, Laszlo
Anti-inflammatory Effects of Triptolide in LPS-Induced Acute Lung Injury in Mice.
Triptolide is one of the main active components of Chinese herb Tripterygium wilfordii Hook F, which has been demonstrated to have anti-inflammatory properties. The aim of this study was to investigate the effects of triptolide on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice and to clarify the possible mechanisms. Mice were administered intranasally with LPS to induce lung injury. Triptolide was administered intraperitoneally 1�h before LPS challenge. Triptolide-treated mice exhibited significantly reduced leukocyte, myeloperoxidase (MPO) activity, edema of the lung, as well as TNF-?, IL-1?, and IL-6 production in the bronchoalveolar lavage fluid compared with LPS-treated mice. Additionally, Western blot analysis showed that triptolide inhibited the phosphorylation of inhibitor-kappa B kinase-alpha (I?B-?), p65, nuclear factor kappa B (NF-?B), p38, extracellular receptor kinase (ERK), and Jun N-terminal kinase (JNK) and the expression of Toll-like receptor 4 (TLR4) caused by LPS. In conclusion, our results suggested that the promising anti-inflammatory mechanism of triptolide may be that triptolide activates peroxisome proliferation-activated receptor gamma (PPAR-?), thereby attenuating an LPS-induced inflammatory response. Triptolide may be a promising potential therapeutic reagent for ALI treatment. PMID:24706025
Wei, Dong; Huang, Zhihong
Vascular barrier protective effects of orientin and isoorientin in LPS-induced inflammation in vitro and in vivo.
Endothelial cell protein C receptor (EPCR) can be shed from the cell surface, and this process is mediated by tumor necrosis factor-? converting enzyme (TACE), and high levels of soluble EPCR are involved in vascular inflammation. Orientin, one of the C-glycosyl flavonoids, has been known to have anxiolytic and antioxidative activities. However, the effect of orientin on lipopolysaccharide (LPS)-induced inflammatory response has not been studied. Here we investigated the barrier protective effects of orientin against pro-inflammatory responses induced by LPS and the associated signaling pathways. We found that orientin inhibited LPS-induced barrier disruption, expression of cell adhesion molecules (CAMs), and adhesion/transendothelial migration of monocytes to human endothelial cells. Orientin induced potent inhibition of phorbol-12-myristate 13-acetate (PMA) and LPS-induced EPCR shedding. Orientin also suppressed LPS-induced hyperpermeability and leukocyte migration in vivo. Furthermore, orientin suppressed the production of tumor necrosis factor-? (TNF-?) or Interleukin (IL)-6 and the activation of nuclear factor-?B (NF-?B) or extracellular regulated kinases (ERK) 1/2 by LPS. Moreover, treatment with orientin resulted in reduced LPS-induced lethal endotoxemia. These results suggest that orientin protects vascular barrier integrity by inhibiting hyperpermeability, expression of CAMs, and adhesion and migration of leukocytes, thereby endorsing its usefulness as a therapy for vascular inflammatory diseases. PMID:24792192
Lee, Wonhwa; Ku, Sae-Kwang; Bae, Jong-Sup
BHBA Suppresses LPS-Induced Inflammation in BV-2 Cells by Inhibiting NF-?B Activation
?-Hydroxybutyric acid (BHBA) has neuroprotective effects, but the underlying molecular mechanisms are unclear. Microglial activation plays an important role in neurodegenerative diseases by producing several proinflammatory enzymes and proinflammatory cytokines. The current study investigates the potential mechanisms whereby BHBA affects the expression of potentially proinflammatory proteins by cultured murine microglial BV-2 cells stimulated with lipopolysaccharide (LPS). The results showed that BHBA significantly reduced LPS-induced protein and mRNA expression levels of iNOS, COX-2, TNF-?, IL-1?, and IL-6. Blocking of GPR109A by PTX resulted in a loss of this anti-inflammatory effect in BV-2 cells. Western blot analysis showed that BHBA reduced LPS-induced degradation of I?B-? and translocation of NF-?B, while no effect was observed on MAPKs phosphorylation. All results imply that BHBA significantly reduces levels of proinflammatory enzymes and proinflammatory cytokines by inhibition of the NF-?B signaling pathway but not MAPKs pathways, and GPR109A is essential to this function. Overall, these data suggest that BHBA has a potential as neuroprotective drug candidate in neurodegenerative diseases.
Fu, Shou-Peng; Li, Su-Nan; Wang, Jian-Fa; Li, Yang; Xie, Shan-Shan; Xue, Wen-Jing; Liu, Hong-Mei; Huang, Bing-Xu; Lv, Qing-Kang; Lei, Lian-Cheng; Liu, Guo-Wen; Wang, Wei; Liu, Ju-Xiong
Stiffness-Activated GEF-H1 Expression Exacerbates LPS-Induced Lung Inflammation
Acute lung injury (ALI) is accompanied by decreased lung compliance. However, a role of tissue mechanics in modulation of inflammation remains unclear. We hypothesized that bacterial lipopolysacharide (LPS) stimulates extracellular matrix (ECM) production and vascular stiffening leading to stiffness-dependent exacerbation of endothelial cell (EC) inflammatory activation and lung barrier dysfunction. Expression of GEF-H1, ICAM-1, VCAM-1, ECM proteins fibronectin and collagen, lysyl oxidase (LOX) activity, interleukin-8 and activation of Rho signaling were analyzed in lung samples and pulmonary EC grown on soft (1.5 or 2.8 kPa) and stiff (40 kPa) substrates. LPS induced EC inflammatory activation accompanied by expression of ECM proteins, increase in LOX activity, and activation of Rho signaling. These effects were augmented in EC grown on stiff substrate. Stiffness-dependent enhancement of inflammation was associated with increased expression of Rho activator, GEF-H1. Inhibition of ECM crosslinking and stiffening by LOX suppression reduced EC inflammatory activation and GEF-H1 expression in response to LPS. In vivo, LOX inhibition attenuated LPS-induced expression of GEF-H1 and lung dysfunction. These findings present a novel mechanism of stiffness-dependent exacerbation of vascular inflammation and escalation of ALI via stimulation of GEF-H1 - Rho pathway. This pathway represents a fundamental mechanism of positive feedback regulation of inflammation.
Mambetsariev, Isa; Tian, Yufeng; Wu, Tinghuai; Lavoie, Tera; Solway, Julian; Birukov, Konstantin G.; Birukova, Anna A.
Cannabidiol (CBD) Enhances Lipopolysaccharide (LPS)-Induced Pulmonary Inflammation in C57BL/6 Mice
Cannabidiol (CBD) is a plant-derived cannabinoid that has been predominantly characterized as anti-inflammatory. However, it is clear that immune effects of cannabinoids can vary with cannabinoid concentration, or type or magnitude of immune stimulus. The present studies demonstrate that oral administration of CBD enhanced lipopolysaccharide (LPS)-induced pulmonary inflammation in C57BL/6 mice. The enhanced inflammatory cell infiltrate as observed in bronchoalveolar lavage fluid (BALF) was comprised mainly of neutrophils, with some monocytes. Concomitantly, CBD enhanced pro-inflammatory cytokine mRNA production, including tumor necrosis factor-? (Tnfa), interleukins (IL) 6 and 23 (Il6, Il23), and granulocyte colony stimulating factor (Gcsf). These results demonstrate that the CBD-mediated enhancement of LPS-induced pulmonary inflammation is mediated at the level of transcription of a variety of pro-inflammatory genes. The significance of these studies is that CBD is part of a therapeutic currently in use for spasticity and pain in multiple sclerosis patients, and therefore it is important to further understand mechanisms by which CBD alters immune function.
Karmaus, Peer W. F.; Wagner, James G.; Harkema, Jack R.; Kaminski, Norbert E.; Kaplan, Barbara L.F.
Calorie restriction attenuates lipopolysaccharide (LPS)-induced microglial activation in discrete regions of the hypothalamus and the subfornical organ.
Calorie restriction (CR) has been shown to increase longevity and elicit many health promoting benefits including delaying immunosenescence and attenuating neurodegeneration in animal models of Alzheimer's disease and Parkinson's disease. CR also suppresses microglial activation following cortical injury and aging. We previously demonstrated that CR attenuates lipopolysaccharide (LPS)-induced fever and shifts hypothalamic signaling pathways to an anti-inflammatory bias; however, the effects of CR on LPS-induced microglial activation remain largely unexplored. The current study investigated regional changes in LPS-induced microglial activation in mice exposed to 50% CR for 28days. Immunohistochemistry was conducted to examine changes in ionized calcium-binding adapter molecule-1 (Iba1), a protein constitutively expressed by microglia, in a total of 27 brain regions involved in immunity, stress, and/or thermoregulation. Exposure to CR attenuated LPS-induced fever, and LPS-induced microglial activation in a subset of regions: the arcuate nucleus (ARC) and ventromedial nucleus of the hypothalamus (VMH) and the subfornical organ (SFO). Microglial activation in the ARC and VMH was positively correlated with body temperature. These data suggest that CR exerts effects on regionally specific populations of microglia; particularly, in appetite-sensing regions of the hypothalamus, and/or regions lacking a complete blood brain barrier, possibly through altered pro- and anti-inflammatory signaling in these regions. PMID:24291211
Radler, Morgan E; Hale, Matthew W; Kent, Stephen
Punicalagin Inhibits Inflammation in LPS-Induced RAW264.7 Macrophages via the Suppression of TLR4-Mediated MAPKs and NF-?B Activation.
Punicalagin (2,3,hexahydroxydiphenoyl-gallagyl-D-glucose and referred to as PUN) is a bioactive ellagitannin isolated from pomegranate, which is widely used for the treatment of inflammatory bowel disease (IBD), diarrhea, and ulcers in Chinese traditional medicine. In this study, we detected the anti-inflammation potentials of PUN in lipopolysaccharide (LPS)-induced macrophages and tried to uncover the underlying mechanism. Results demonstrated that PUN (25, 50, or 100�?M) treatment could significantly decrease the LPS-induced production of nitric oxide), prostaglandin E2 (PGE2), interleukin (IL)-1?, IL-6, and tumor necrosis factor (TNF)-? in RAW264.7 cells. Molecular research showed that PUN inhibited the activation of upstream mediator nuclear factor-?B by suppressing the phosphorylation of I?B? and p65. Results also indicated that PUN could suppress the phosphorylation of mitogen-activated protein kinase including p38, c-Jun N-terminal kinase, and extracellular signal-regulated kinase. In conclusion, we observed that PUN could inhibit LPS-induced inflammation, and it may be a potential choice for the treatment of inflammation diseases. PMID:24473904
Xu, Xiaolong; Yin, Peng; Wan, Changrong; Chong, Xinlu; Liu, Mingjiang; Cheng, Peng; Chen, Jiajia; Liu, Fenghua; Xu, Jianqin
Yohimbine promotes cardiac NE release and prevents LPS-induced cardiac dysfunction via blockade of presynaptic ?2A-adrenergic receptor.
Myocardial depression is an important contributor to mortality in sepsis. We have recently demonstrated that ?2-adrenoceptor (AR) antagonist, yohimbine (YHB), attenuates lipopolysaccharide (LPS)-induced myocardial depression. However, the mechanisms for this action of YHB are unclear. Here, we demonstrated that YHB decreased nitric oxide (NO) and tumor necrosis factor-alpha (TNF-?) levels in the myocardium and plasma, attenuated cardiac and hepatic dysfunction, but not kidney and lung injuries in endotoxemic mice. Immunohistochemical analysis revealed that cardiac ?2A-AR was mostly located in sympathetic nerve presynaptic membrane; YHB decreased cardiac ?2A-AR level and promoted cardiac norepinephrine (NE) release in endotoxemic mice. Reserpine that exhausted cardiac NE without markedly decreasing plasma NE level abrogated the inhibitory effects of YHB on cardiac TNF-? and iNOS expression as well as cardiac dysfunction, but not the suppressive effects of YHB on plasma TNF-? and NO elevation in LPS-challenged mice. Furthermore, both reserpine and YHB significantly inhibited LPS-induced myocardial apoptosis. ?1-AR, ?2-AR, but not ?1-AR antagonists reversed the inhibitory effect of YHB on LPS-stimulated myocardial apoptosis. However, ?1-AR antagonist attenuated LPS-caused cardiomyocyte apoptosis, partly abolished the protective effect of YHB on the left ventricular ejection fraction in endotoxemic mice. Altogether, these findings indicate that YHB attenuates LPS-induced cardiac dysfunction, at least in part, through blocking presynaptic ?2A-AR and thus increasing cardiac NE release. YHB-elevated cardiac NE improves cardiac function via suppressing cardiac iNOS and TNF-? expression, activating ?1-AR and inhibiting cardiomyocyte apoptosis through ?1- and ?2-AR in endotoxemic mice. However, cardiac ?1-AR activation promotes LPS-induced cardiomyocyte apoptosis. PMID:23691077
Wang, Yiyang; Yu, Xiaohui; Wang, Faqiang; Wang, Yuan; Wang, Yanping; Li, Hongmei; Lv, Xiuxiu; Lu, Daxiang; Wang, Huadong
Transiently enhanced LPS-induced fever following hyperthermic stress in rabbits
NASA Astrophysics Data System (ADS)
Hyperthermia has been shown to induce an enhanced febrile response to the bacterial-derived endotoxin lipopolysaccharide (LPS). The aim of the present study was to test the hypothesis that the enhanced LPS-induced fever seen in heat stressed (HS) animals is caused by leakage of intestinal bacterial LPS into the circulation. Male rabbits were rendered transiently hyperthermic (a maximum rectal temperature of 43�C) and divided into three groups. They were then allowed to recover in a room at 24�C for 1, 2 or 3 days post-HS. One day after injection with LPS, the post-HS rabbits exhibited significantly higher fevers than the controls, though this was not seen in rabbits at either 2 or 3 days post-HS. The plasma levels of endogenous LPS were significantly increased during the HS as compared to those seen in normothermic rabbits prior to HS. LPS fevers were not induced in these animals. One day post-HS, rabbits that had been pretreated with oral antibiotics exhibited significantly attenuated LPS levels. When challenged with human recombinant interleukin-1? instead of LPS, the 1-day post-HS rabbits did not respond with enhanced fevers. The plasma levels of TNF? increased similarly during LPS-induced fevers in both the control and 1-day post-HS rabbits, while the plasma levels of corticosterone and the osmolality of the 1-day post-HS rabbits showed no significant differences to those seen prior to the HS. These results suggest that the enhanced fever in the 1-day post-HS rabbits is LPS specific, and may be caused by increased leakage of intestinal endotoxin into blood circulation.
Shibata, Masaaki; Uno, Tadashi; Riedel, Walter; Nishimaki, Michiyo; Watanabe, Kaori
Stabilization of Nrf2 by tBHQ prevents LPS-induced apoptosis in differentiated PC12 cells
The inflammatory reaction plays an important role in the pathogenesis of the neurodegenerative disorders. tert-butylhydroquinone\\u000a (tBHQ) exhibits a wide range of pharmacological activities including anti-oxidative and anti-inflammatory action. In this\\u000a study, we tried to elucidate possible effects of tBHQ on lipopolysaccharide (LPS)-induced inflammatory reaction and its underlying\\u000a mechanism in neuron-like PC12 cells. tBHQ inhibited LPS-induced generation of reactive oxygen species
Fariba Khodagholi; Solaleh Khoramian Tusi
Andrographolide Protects against LPS-Induced Acute Lung Injury by Inactivation of NF-?B
Background Nuclear factor-?B (NF-?B) is a central transcriptional factor and a pleiotropic regulator of many genes involved in acute lung injury. Andrographolide is found in the plant of Andrographis paniculata and widely used in Traditional Chinese Medicine, exhibiting potently anti-inflammatory property by inhibiting NF-?B activity. The purpose of our investigation was designed to reveal the effect of andrographolide on various aspects of LPS induced inflammation in vivo and in vitro. Methods and Results In vivo, BALB/C mice were subjected to LPS injection with or without andrographolide treatments to induce ALI model. In vitro, MLE-12 cells were stimulated with LPS in the presence and absence of andrographolide. In vivo, pulmonary inflammation, pulmonary edema, ultrastructure changes of type II alveolar epithelial cells, MPO activity, total cells, neutrophils, macrophages, TNF-?, IL-6 and IL-1? in BALF, along with the expression of VCAM-1 and VEGF were dose-dependently attenuated by andrographolide. Meanwhile, in vitro, the expression of VCAM-1 and VEGF was also reduced by andrographolide. Moreover, our data showed that andrographolide significantly inhibited the ratios of phospho-IKK?/total IKK?, phospho-I?B?/total I?B? and phospho-NF-?B p65/total NF-?B p65, and NF-?B p65 DNA binding activities, both in vivo and in vitro. Conclusions These results indicate that andrographolide dose-dependently suppressed the severity of LPS-induced ALI, more likely by virtue of andrographolide-mediated NF-?B inhibition at the level of IKK? activation. These results suggest andrographolide may be considered as an effective and safe drug for the potential treatment of ALI.
Zhu, Tao; Wang, Dao-xin; Zhang, Wei; Liao, Xiu-qing; Guan, Xian; Bo, Hong; Sun, Jia-yang; Huang, Ni-wen; He, Jing; Zhang, Yun-kun; Tong, Jing; Li, Chang-yi
Phenolic glycosides from Alangium salviifolium leaves with inhibitory activity on LPS-induced NO, PGE(2), and TNF-alpha production.
Three new phenolic glycosides, salviifosides A-C (13), and three known compounds salicin (4), kaempferol (5), and kaempferol 3-O-beta-d-glucopyranoside (6) were isolated from the leaves of Alangium salviifolium (L.f.) Wangerin (Alangiaceae). The structures of the new metabolites were determined on the basic of spectroscopic analyses including two dimensional NMR. The anti-inflammatory activities of new compounds (1-3) were investigated on lipopolysaccharide (LPS)-induced murine macrophage cells line, RAW 264.7. Salviifoside B (2) potentially inhibits the productions of nitric oxide (NO), prostaglandin E(2) (PGE(2)), and tumor necrosis factor-alpha (TNF-alpha). PMID:19500975
Tran, Manh Hung; Nguyen, Hai Dang; Kim, Jin Cheol; Choi, Jae Sue; Lee, Hyeong Kyu; Min, Byung-Sun
Persistence of LPS-induced lung inflammation in surfactant protein-C-deficient mice.
Pulmonary surfactant protein-C (SP-C) gene-targeted mice (Sftpc(-/-)) develop progressive lung inflammation and remodeling. We hypothesized that SP-C deficiency reduces the ability to suppress repetitive inflammatory injury. Sftpc(+/+) and Sftpc(-/-) mice given three doses of bacterial LPS developed airway and airspace inflammation, which was more intense in the Sftpc(-/-) mice at 3 and 5 days after the final dose. Compared with Sftpc(+/+)mice, inflammatory injury persisted in the lungs of Sftpc(-/-) mice 30 days after the final LPS challenge. Sftpc(-/-) mice showed LPS-induced airway goblet cell hyperplasia with increased detection of Sam pointed Ets domain and FoxA3 transcription factors. Sftpc(-/-) type II alveolar epithelial cells had increased cytokine expression after LPS exposure relative to Sftpc(+/+) cells, indicating that type II cell dysfunction contributes to inflammatory sensitivity. Microarray analyses of isolated type II cells identified a pattern of enhanced expression of inflammatory genes consistent with an intrinsic low-level inflammation resulting from SP-C deficiency. SP-C-containing clinical surfactant extract (Survanta) or SP-C/phospholipid vesicles blocked LPS signaling through the LPS receptor (Toll-like receptor [TLR] 4/CD14/MD2) in human embryonic kidney 293T cells, indicating that SP-C blocks LPS-induced cytokine production by a TLR4-dependent mechanism. Phospholipid vesicles alone did not modify the TLR4 response. In vivo deficiency of SP-C leads to inflammation, increased cytokine production by type II cells, and persistent inflammation after repetitive LPS stimulation. PMID:23795648
Glasser, Stephan W; Maxfield, Melissa D; Ruetschilling, Teah L; Akinbi, Henry T; Baatz, John E; Kitzmiller, Joseph A; Page, Kristen; Xu, Yan; Bao, Erik L; Korfhagen, Thomas R
LPS-induced IL8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
BACKGROUND: The release of LPS by bacteria stimulates both immune and specific epithelial cell types to release inflammatory mediators. It is known that LPS induces the release of IL-8 by intestinal mucosal cells. Because it is now emerging that bacteria may induce alteration of epigenetic patterns in host cells, we have investigated whether LPS-induced IL-8 activation in human intestinal epithelial
Tiziana Angrisano; Raffaela Pero; Silvia Peluso; Simona Keller; Silvana Sacchetti; Carmelo B Bruni; Lorenzo Chiariotti; Francesca Lembo
Anti-inflammatory effect of spilanthol from Spilanthes acmella on murine macrophage by down-regulating LPS-induced inflammatory mediators.
Spilanthes acmella (Paracress), a common spice, has been administered as a traditional folk medicine for years to cure toothaches, stammering, and stomatitis. Previous studies have demonstrated its diuretic, antibacterial, and anti-inflammatory activities. However, the active compounds contributing to the anti-inflammatory effect have seldom been addressed. This study isolates the active compound, spilanthol, by a bioactivity-guided approach and indicates significant anti-inflammatory activity on lipopolysaccharide-activated murine macrophage model, RAW 264.7. The anti-inflammatory mechanism of paracress is also investigated. Extracts of S. acmella are obtained by extraction with 85% ethanol, followed by liquid partition against hexane, chloroform, ethyl acetate, and butanol. The ethyl acetate extract exhibits a stronger free radical scavenging capacity than other fractions do, as determined by DPPH and ABTS radical scavenging assays. The chloroform extract significantly inhibits nitric oxide production ( p < 0.01) and is selected for further fractionation to yield the active compound, spilanthol. The diminished levels of LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX-2) mRNA and protein expression support the postulation that spilanthol inhibits proinflammatory mediator production at the transcriptional and translational levels. Additionally, the LPS-stimulated IL-1beta, IL-6, and TNF-alpha productions are dose-dependently reduced by spilanthol. The LPS-induced phosphorylation of cytoplasmic inhibitor-kappaB and the nuclear NF-kappaB DNA binding activity are both restrained by spilanthol. Results of this study suggest that spilanthol, isolated from S. acmella, attenuates the LPS-induced inflammatory responses in murine RAW 264.7 macrophages partly due to the inactivation of NF-kappaB, which negatively regulates the production of proinflammatory mediators. PMID:18321049
Wu, Li-Chen; Fan, Nien-Chu; Lin, Ming-Hui; Chu, Inn-Ray; Huang, Shu-Jung; Hu, Ching-Yuan; Han, Shang-Yu
Chitosan oligosaccharides block LPS-induced O-GlcNAcylation of NF-?B and endothelial inflammatory response.
It is known that chitosan oligosaccharides (COS) suppress LPS-induced vascular endothelial inflammatory response by mechanism involving NF-?B blockade. It remains unknown how COS inhibit NF-?B. We provided evidence both in cultured endothelial cells and mouse model supporting a new mechanism. Regardless of the endothelial cell types, the LPS-induced NF-?B-dependent inflammatory gene expression was suppressed by COS, which was associated with reduced NF-?B nucleus translocation. LPS enhanced O-GlcNAc modification of NF-?B/p65 and activated NF-?B pathway, which could be prevented either by siRNA knockdown of O-GlcNAc transferase (OGT) or pretreatment with COS. Inhibition of either mitogen-activated protein kinase or superoxide generation abolishes LPS-induced NF-?B O-GlcNAcylation. Consistently, aortic tissues from LPS-treated mice presented enhanced NF-?B/p65 O-GlcNAcylation in association with upregulated gene expression of inflammatory cytokines in vascular tissues; however, pre-administration of COS prevented these responses. In conclusion, COS decreased OGT-dependent O-GlcNAcylation of NF-?B and thereby attenuated LPS-induced vascular endothelial inflammatory response. PMID:24274545
Li, Yu; Liu, Hongtao; Xu, Qing-Song; Du, Yu-Guang; Xu, Jian
Morphine preconditioning protects against LPS-induced neuroinflammation and memory deficit.
Recent studies show that morphine possesses protective preconditioning effects in different ischemia/reperfusion models. However, there is very little information about the antineuroinflammatory role of morphine and its protective effect against memory deficit. In the present study, we evaluated the role of morphine preconditioning in a model of mild neuroinflammation induced by intraperitoneal lipopolysaccharide (LPS) injection (1 mg/kg). Rats were trained on passive avoidance apparatus and challenged with LPS 20 h later. Four hours after LPS, rats were subjected to passive avoidance testing and then for the assessments of inflammatory and apoptotic cell death mediators in the hippocampus. LPS significantly increased the nuclear NF-?B and expression of COX-2, IL-1?, and TNF-?, augmented the activity of caspase-3 and PARP cleavage, and in parallel shortened the latencies to enter the dark compartment. Although morphine injection in a noninflammatory context was able to induce a neuroinflammatory response and memory loss, morphine preconditioning at the dose of 4 mg/kg significantly prevented the LPS-induced neuroinflammation and memory deficit. Morphine preconditioning was abolished by naloxone and, therefore, is dependent on opioid receptors. These results suggest that acute morphine injection, in spite of the induction of a neuroinflammatory response and amnesia per se, exerts an antineuroinflammatory role and protects from cell death and memory deficit in an inflammatory context. PMID:22388653
Rostami, Farzaneh; Oryan, Shahrbanoo; Ahmadiani, Abolhassan; Dargahi, Leila
Granzyme K synergistically potentiates LPS-induced cytokine responses in human monocytes.
Granzymes are serine proteases released by cytotoxic lymphocytes to induce apoptosis in virus-infected cells and tumor cells. Evidence is emerging that granzymes also play a role in controlling inflammation. Granzyme serum levels are elevated in patients with autoimmune diseases and infections, including sepsis. However, the function of extracellular granzymes in inflammation largely remains unknown. Here, we show that granzyme K (GrK) binds to Gram-negative bacteria and their cell-wall component lipopolysaccharide (LPS). GrK synergistically enhances LPS-induced cytokine release in vitro from primary human monocytes and in vivo in a mouse model of LPS challenge. Intriguingly, these extracellular effects are independent of GrK catalytic activity. GrK disaggregates LPS from micelles and augments LPS-CD14 complex formation, thereby likely boosting monocyte activation by LPS. We conclude that extracellular GrK is an unexpected direct modulator of LPS-TLR4 signaling during the antimicrobial innate immune response. PMID:24711407
Wensink, Annette C; Kemp, Vera; Fermie, Job; Garc�a Laorden, M Isabel; van der Poll, Tom; Hack, C Erik; Bovenschen, Niels
p52-independent nuclear translocation of RelB promotes LPS-induced attachment
The NF-{kappa}B signaling pathways have a critical role in the development and progression of various cancers. In this study, we demonstrated that the small cell lung cancer cell line (SCLC) H69 expressed a unique NF-{kappa}B profile as compared to other cancer cell lines. The p105/p50, p100/p52, c-Rel, and RelB protein and mRNA transcripts were absent in H69 cells but these cells expressed RelA/p65. The activation of H69 cells by lipopolysaccharide (LPS) resulted in the induction of RelB and p100 expression. The treatment also induced the nuclear translocation of RelB without the processing of p100 to p52. Furthermore, LPS-induced {beta}1 integrin expression and cellular attachment through an NF-{kappa}B-dependent mechanism. Blocking RelB expression prevented the increase in the expression of {beta}1 integrin and the attachment of H69. Taken together, the results suggest that RelB was responsible for the LPS-mediated attachment and may play an important role in the progression of some cancers.
Saito, T. [Laboratory of Immunology, National Institute on Aging, NIH Biomedical Research Center, Baltimore, MD 21224 (United States)] [Laboratory of Immunology, National Institute on Aging, NIH Biomedical Research Center, Baltimore, MD 21224 (United States); Sasaki, C.Y., E-mail: email@example.com [Laboratory of Immunology, National Institute on Aging, NIH Biomedical Research Center, Baltimore, MD 21224 (United States); Rezanka, L.J.; Ghosh, P.; Longo, D.L. [Laboratory of Immunology, National Institute on Aging, NIH Biomedical Research Center, Baltimore, MD 21224 (United States)] [Laboratory of Immunology, National Institute on Aging, NIH Biomedical Research Center, Baltimore, MD 21224 (United States)
Polyphenols from blueberries modulate inflammation cytokines in LPS-induced RAW264.7 macrophages.
Polyphenols including 3-glucoside/arabinoside/galactoside-based polymers of delphinidins, petunidins, peonidins, malvidins and cyanidins are one type of biological macromolecules, which are extraordinarily rich in blueberries. Anti-inflammatory activity of blueberry polyphenols (BPPs) was investigated by using lipopolysaccharide (LPS) induced RAW264.7 macrophages. The results showed that BPPs suppressed the gene expression of IL-1? (interleukin-1?), IL-6 and IL-12p35. The inhibition effect on IL-1? and IL-6 mRNA was most obvious at the concentration of 10-200?g/mL BPPs. But the inhibition effect on IL-12p35 mRNA was increased with the increasing concentration of BPPs. When fixed at 100?g/mL BPPs, the most significant inhibition on IL-1?, IL-6 and IL-12p35 mRNA expression was detected at 12-48h. In conclusion, BPPs exhibit anti-inflammation activity by mediating and modulating the balances in pro-inflammatory cytokines of IL-1?, IL-6, and IL-12. PMID:24905959
Cheng, Anwei; Yan, Haiqing; Han, Caijing; Wang, Wenliang; Tian, Yaoqi; Chen, Xiangyan
Fresh organically grown ginger (Zingiber officinale): composition and effects on LPS-induced PGE2 production.
Gas chromatography in conjunction with mass spectrometry, a technique previously employed to analyze non-volatile pungent components of ginger extracts modified to trimethylsilyl derivatives, was applied successfully for the first time to analyze unmodified partially purified fractions from the dichloromethane extracts of organically grown samples of fresh Chinese white and Japanese yellow varieties of ginger, Zingiber officinale Roscoe (Zingiberaceae). This analysis resulted in the detection of 20 hitherto unknown natural products and 31 compounds previously reported as ginger constituents. These include paradols, dihydroparadols, gingerols, acetyl derivatives of gingerols, shogaols, 3-dihydroshogaols, gingerdiols, mono- and diacetyl derivatives of gingerdiols, 1-dehydrogingerdiones, diarylheptanoids, and methyl ether derivatives of some of these compounds. The thermal degradation of gingerols to gingerone, shogaols, and related compounds was demonstrated. The major constituent in the two varieties was [6]-gingerol, a chemical marker for Z. officinale. Mass spectral fragmentation patterns for all the compounds are described and interpreted. Anti-inflammatory activities of silica gel chromatography fractions were tested using an in vitro PGE2 assay. Most of the fractions containing gingerols and/or gingerol derivatives showed excellent inhibition of LPS-induced PGE2 production. PMID:15280001
Jolad, Shivanand D; Lantz, R Clark; Solyom, Aniko M; Chen, Guan Jie; Bates, Robert B; Timmermann, Barbara N
Phospholipid Scramblase Expression in the Pregnant Mouse Uterus in LPS-Induced Preterm Delivery
Phospholipid scramblases (PLSCR), stimulated by proinflammatory cytokines, are thought to mediate the loss of lipid asymmetry in cell membranes, allowing for specific reactions in the coagulation cascade. The PLSCR may therefore provide a link between inflammation, coagulation, and, because thrombin is a uterotonic, preterm birth (PTB). To explore the relationship between PLSCR expression and inflammation-related PTB, we utilized reverse transcriptase�polymerase chain reaction and Western blot studies to quantify messenger RNA (mRNA) and protein expression for the 4 PLSCR homologues (PLSCR 1-4). Uteri from day 15 pregnant mice were harvested at several time points after intrauterine lipopolysaccharide (LPS) injection (or normal saline, for controls). Expression of mRNA in all 4 Plscr isoforms was demonstrated. Lipopolysaccharide treatment resulted in increased expression of PLSCR-1 and a decrease in Plscr4 mRNA, thereby demonstrating modulation of PLSCR-1 and PLSCR-4 in LPS-induced PTB. Additionally, protein expression was confirmed for all except PLSCR-4, with increased expression of PLSCR-1 after LPS treatment.
Oppenheimer, Karen H.; Sweet, Leigh M.; Phillippe, Mark
CKBM stimulates MAPKs but inhibits LPS-induced IFN-gamma in lymphocytes.
CKBM is an herbal formula composed of five Chinese medicinal herbs (Panax ginseng, Schisandra chinensis, Fructus crataegi, Ziziphus jujuba and Glycine max) supplemented with processed Saccharomyces cerevisiae. It has been demonstrated that CKBM is capable of triggering the release of IL-6 and TNFalpha from human peripheral blood mononuclear cells. In this report, T-lymphocytic Sup-T1 cells and B-lymphocytic Ramos cells were utilized as cellular models to investigate how CKBM regulates intracellular signaling as well as the production of cytokines. CKBM stimulated the three major subgroups of mitogen-activated protein kinase (i.e. ERK, JNK and p38) in Sup-T1 cells, but only triggered the activation of ERK and p38 in Ramos cells. The induction of mitogen-activated protein kinases (MAPK) activations varied with the duration of treatment, as well as with the dosage of CKBM. In terms of cytokine production, treatment of CKBM alone did not trigger the release of IL-1beta and IFNgamma, but it suppressed the LPS-induced IFNgamma production from both Sup-T1 cells and Ramos cells. In view of the therapeutic effects of traditional Chinese medicines in inflammatory and autoimmune disorders, the results suggest that CKBM may exhibit its immuno-modulatory effects by regulating intracellular signaling as well as cytokine production in different lymphocytic cell types. PMID:16775808
Chan, Anthony S L; Yip, Eric C H; Yung, Lisa Y; Pang, Haihong; Luk, Sharon C W; Pang, Shiu F; Wong, Yung H
Effect of Pluchea lanceolata bioactives in LPS-induced neuroinflammation in C6 rat glial cells.
Neuroinflammation plays a significant role in various chronic and acute pathological conditions of the central nervous system. In the Indian system of medicine, Pluchea lanceolata is used to treat the neurological disorders. We investigated the effect of major pentacyclic triterpene and its naturally occurring acetate derivative isolated from P. lanceolata on lipopolysaccharide (LPS)-stimulated neuroinflammatory condition associated to inflammatory cytokine production in rat astrocytoma cell line (C6). The log concentration dependence of Pluchea bioactive taraxasterol (Tx) significantly (p?0.05) attenuates the release of pro-inflammatory cytokines, such as TNF-?, IFN-?, and IL-6, while its in situ produced acetyl derivative, i.e., taraxasterol acetate (TxAc), did not inhibit the LPS-induced IL-6 production at lower concentration (p?>?0.05). Surflex-Dock molecular modeling study was performed to simulate the binding capacity of compounds into the active site of the TNF-? (2AZ5), tumor protein P53 (2VUK), and NF-kappa-B (1RAM). The differential inhibition of cytokines by Tx and TxAc was further confirmed by high docking scores showing the high affinity to target proteins. Findings of the study demonstrated the comparatively greater role of Pluchea triterpene than its in situ produced acetate derivate in neuroinflammation-associated disorders. PMID:24101125
Srivastava, Pooja; Mohanti, Shilpa; Bawankule, Dnyaneshwar Umrao; Khan, Feroz; Shanker, Karuna
IL8 is an essential mediator of the increased delayed-phase vascular permeability in LPS-induced rabbit pleurisy
We investigated the involvement of IL-8 in the delayed vascular permeability (VP) in rabbit lipopolysaccharide (LPS)-pleurisy. Maximal level of interleukin-8 (IL-8) was detected in pleural fluid at 2 h after LPS injection and anti-IL-8 inhibited the delayed VP by 90%. Injection of homologous IL-8 induced VP, the time-course of which pre- ceded that of LPS-induced delayed VP. Production of IL-8
Takumi Fukumoto; Akihiro Matsukawa; Teizo Yoshimura; Sunao Edamitsu; Susumu Ohkawara; Katsumasa Takagi; Masaru Yoshinaga
Involvement of mitogen-activated protein kinases and NF{kappa}B in LPS-induced CD40 expression on human monocytic cells
CD40 is a costimulatory molecule linking innate and adaptive immune responses to bacterial stimuli, as well as a critical regulator of functions of other costimulatory molecules. The mechanisms regulating lipopolysaccharide (LPS)-induced CD40 expression have not been adequately characterized in human monocytic cells. In this study we used a human monocytic cell line, THP-1, to investigate the possible mechanisms of CD40 expression following LPS exposure. Exposure to LPS resulted in a dose- and time-dependent increase in CD40 expression. Further studies using immunoblotting and pharmacological inhibitors revealed that mitogen-activated protein kinases (MAPKs) and NF{kappa}B were activated by LPS exposure and involved in LPS-induced CD40 expression. Activation of MAPKs was not responsible for LPS-induced NF{kappa}B activation. TLR4 was expressed on THP-1 cells and pretreatment of cells with a Toll-like receptor 4 (TLR4) neutralizing antibody (HTA125) significantly blunted LPS-induced MAPK and NF{kappa}B activation and ensuing CD40 expression. Additional studies with murine macrophages expressing wild type and mutated TLR4 showed that TLR4 was implicated in LPS-induced ERK and NF{kappa}B activation, and CD40 expression. Moreover, blockage of MAPK and NF{kappa}B activation inhibited LPS-induced TLR4 expression. In summary, LPS-induced CD40 expression in monocytic cells involves MAPKs and NF{kappa}B.
Wu Weidong [Center for Environmental Medicine, Asthma, and Lung Biology, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)]|[Department of Pediatrics, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)], E-mail: firstname.lastname@example.com; Alexis, Neil E. [Center for Environmental Medicine, Asthma, and Lung Biology, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)]|[Department of Pediatrics, University of North Carolina, Chapel Hill, North Carolina 27599 (United States); Chen Xian [Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)]|[Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina 27599 (United States); Bromberg, Philip A. [Center for Environmental Medicine, Asthma, and Lung Biology, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)]|[Department of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599 (United States); Peden, David B. [Center for Environmental Medicine, Asthma, and Lung Biology, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)]|[Department of Pediatrics, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)]|[Department of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)
Inhibition of LPS-induced airway neutrophilic inflammation in healthy volunteers with an oral CXCR2 antagonist
Background Inhaled lipopolysaccharide (LPS) induces a dose-dependent, acute neutrophilic response in the airways of healthy volunteers that can be quantified in induced sputum. Chemokines, such as CXCL1 and CXCL8, play an important role in neutrophilic inflammation in the lung through the activation of CXCR2 and small molecule antagonists of these receptors have now been developed. We investigated the effect of AZD8309, a CXCR2 antagonist, compared with placebo on LPS-induced inflammation measured in sputum of healthy volunteers. Methods Twenty healthy subjects were randomized in a double-blind placebo-controlled, cross-over study. AZD8309 (300�mg) or placebo was dosed twice daily orally for 3�days prior to challenge with inhaled LPS and induced sputum was collected 6�h later. Results Treatment with AZD8309 showed a mean 77% reduction in total sputum cells (p?0.001) and 79% reduction in sputum neutrophils (p?0.05) compared with placebo after LPS challenge. There was also a reduction in neutrophil elastase activity (p?0.05) and CXCL1 (p?0.05) and trends for reductions in sputum macrophages (47%), leukotriene B4 (39%) and CXCL8 (52%). Conclusions AZD8309 inhibited LPS-induced inflammation measured in induced sputum of normal volunteers, indicating that this treatment may be useful in the treatment of neutrophilic diseases of the airways, such as COPD, severe asthma and cystic fibrosis. Trial registration NCT00860821.
Protective effects and mechanisms of mogroside V on LPS-induced acute lung injury in mice.
Abstract Context: Mogroside V, a compound isolated from Momordica grosvenori Swingle, which belongs to the Cucurbitaceae, is a traditional Chinese medicine reported to have anti-inflammatory potential in murine macrophages and a murine ear edema model. Objective: To investigate the effects and mechanisms of action of this compound in a model of acute lung injury (ALI) induced by lipopolysaccharides (LPS). Materials and methods: Female BALB/c mice were treated with commercial mogroside V (2.5, 5 and 10?mg/kg) for 1?h prior to intranasal injection of LPS (10??g in 50??l). After 12?h, airway inflammation in the ALI model was determined by the wet/dry weight (W/D) ratio, myeloperoxidase (MPO) activity of lung tissue, leukocyte recruitment and cytokine levels in the bronchoalveolar lavage fluid (BALF). Additionally, lung tissue was examined by histology and western blotting to investigate the changes in pathology and the signalling in the presence and absence of mogroside V. Results: Mogroside V at 5 and 10?mg/kg inhibited airway inflammation induced by LPS as measured by the decrease in the histological changes (44 and 67.3% reduction in lung injury score, respectively), a 28.9 and 55.3% reduction in lung MPO activity, and inflammatory cell counts, interleukin-1? (IL-1?, 382 and 280?pg/ml, respectively), IL-6 (378 and 232?pg/ml, respectively) and tumor necrosis factor-? (TNF-?, 12.5 and 7.8?ng/ml, respectively) levels in the BALF. Additionally, mogroside V treatment reduced the activation of cyclooxygenase 2 (COX-2), inducible NO synthase (iNOS), and the nuclear factor (NF)-?B. Discussions and conclusions: Together, these data suggest that mogroside V has the potential to protect against LPS-induced airway inflammation in a model of ALI. PMID:24621273
Shi, Dongfang; Zheng, Meizhu; Wang, Yumeng; Liu, Chunming; Chen, Shan
LPS-Induced Delayed Preconditioning Is Mediated by Hsp90 and Involves the Heat Shock Response in Mouse Kidney
Introduction We and others demonstrated previously that preconditioning with endotoxin (LPS) protected from a subsequent lethal LPS challenge or from renal ischemia-reperfusion injury (IRI). LPS is effective in evoking the heat shock response, an ancient and essential cellular defense mechanism, which plays a role in resistance to, and recovery from diseases. Here, by using the pharmacological Hsp90 inhibitor novobiocin (NB), we investigated the role of Hsp90 and the heat shock response in LPS-induced delayed renal preconditioning. Methods Male C57BL/6 mice were treated with preconditioning (P: 2 mg/kg, ip.) and subsequent lethal (L: 10 mg/kg, ip.) doses of LPS alone or in combination with NB (100 mg/kg, ip.). Controls received saline (C) or NB. Results Preconditioning LPS conferred protection from a subsequent lethal LPS treatment. Importantly, the protective effect of LPS preconditioning was completely abolished by a concomitant treatment with NB. LPS induced a marked heat shock protein increase as demonstrated by Western blots of Hsp70 and Hsp90. NB alone also stimulated Hsp70 and Hsp90 mRNA but not protein expression. However, Hsp70 and Hsp90 protein induction in LPS-treated mice was abolished by a concomitant NB treatment, demonstrating a NB-induced impairment of the heat shock response to LPS preconditioning. Conclusion LPS-induced heat shock protein induction and tolerance to a subsequent lethal LPS treatment was prevented by the Hsp90 inhibitor, novobiocin. Our findings demonstrate a critical role of Hsp90 in LPS signaling, and a potential involvement of the heat shock response in LPS-induced preconditioning.
Kaucsar, Tamas; Bodor, Csaba; Godo, Maria; Szalay, Csaba; Revesz, Csaba; Nemeth, Zalan; Mozes, Miklos; Szenasi, Gabor; Rosivall, Laszlo; Soti, Csaba; Hamar, Peter
The LPS-induced neutrophil recruitment into rat air pouches is mediated by TNFalpha: likely macrophage origin.
The role of resident cells during the lipopolysaccharide (LPS)-induced neutrophil recruitment into rat air pouches was investigated. In this model, LPS (Escherichia coli, O55: B5 strain; 2-2000 ng) induced a dose- and time-dependent neutrophil recruitment accompanied by the generation of a tumour necrosis factor-alpha (TNFalpha)-like activity. Dexamethasone (0.05-5 mug) and cycloheximide (6 ng), injected 2 h before LPS into the pouches, inhibited the neutrophil recruitment and the generation of the TNFalpha-like activity, while the H1-receptor antagonist mepyramine (1 and 4 mg/kg, i.p., 0.5 h before LPS) and the PAF-receptor antagonist WEB 2170 (0.05 and 1 mg/kg, i.p., 0.5 h before LPS) had no effect. Purified alveolar macrophages (AM) were used to replenish the pouches of cycloheximide-treated recipient rats. AM provided by PBS-treated animals led to the recovery of the LPS-induced neutrophil recruitment and of the TNFalpha-like formation contrasting with those from cycloheximide-treated animals (1 mg/kg, i.p.). When delivered in situ, liposome-encapsulated clodronate, a macrophage depletor, significantly impaired both the LPSinduced neutrophil recruitment and the TNFalpha-like activity. An anti-murine TNFalpha polyclonal antibody (0.5 h before LPS) was also effective. These results emphasize the pivotal role of macrophages for LPS-induced neutrophil recruitment via the formation of TNFalpha. PMID:18472868
Arreto, C D; Dumarey, C; Nahori, M A; Vargaftig, B B
Wogonin inhibits LPS-induced tumor angiogenesis via suppressing PI3K/Akt/NF-?B signaling.
Wogonin has been shown to have anti-angiogenesis and anti-tumor effects. However, whether wogonin inhibits LPS-induced tumor angiogenesis is not well known. In this study, we investigated the effect of wogonin on inhibiting LPS-induced tumor angiogenesis and further probed the underlying mechanisms. ELISA results revealed that wogonin could suppress LPS-induced VEGF secretion from tumor cells. Transwell assay, tube formation assay, rat aortic ring assay and CAM model were used to evaluate the effect of wogonin on angiogenesis induced by MCF-7 cell (treated with LPS) in vitro and in vivo. The inhibitory effect of wogonin on angiogenesis in LPS-treated MCF-7 cells was then confirmed by the above in vitro and in vivo assays. The study of the molecular mechanism showed that wogonin could suppress PI3K/Akt signaling activation. Moreover, wogonin inhibited nuclear translocation of NF-?B and its binding to DNA. The result of real-time PCR and luciferase reporter assay suggested that VEGF expression was down-regulated by wogonin primarily at the transcriptional level. IGF-1 and p65 expression plasmid were used to activate PI3K/Akt and NF-?B pathways, and to observe the effect of wogonin on the simualtion of PI3K/Akt/NF-?B signaling. Taken together, the result suggested that wogonin was a potent inhibitor of tumor angiogenesis and provided a new insight into the mechanisms of wogonin against cancer. PMID:24858369
Zhao, Kai; Song, Xiuming; Huang, Yujie; Yao, Jing; Zhou, Mi; Li, Zhiyu; You, Qidong; Guo, Qinglong; Lu, Na
The LPS-induced neutrophil recruitment into rat air pouches is mediated by TNF?: likely macrophage origin
The role of resident cells during the lipopolysaccharide (LPS)-induced neutrophil recruitment into rat air pouches was investigated. In this model, LPS (Escherichia coli, O55: B5 strain; 2�2000 ng) induced a dose� and time-dependent neutrophil recruitment accompanied by the generation of a tumour necrosis factor-? (TNF?)-like activity. Dexamethasone (0.05�5 mug) and cycloheximide (6 ng), injected 2 h before LPS into the pouches, inhibited the neutrophil recruitment and the generation of the TNF?-like activity, while the H1-receptor antagonist mepyramine (1 and 4 mg/kg, i.p., 0.5 h before LPS) and the PAF-receptor antagonist WEB 2170 (0.05 and 1 mg/kg, i.p., 0.5 h before LPS) had no effect. Purified alveolar macrophages (AM) were used to replenish the pouches of cycloheximide-treated recipient rats. AM provided by PBS-treated animals led to the recovery of the LPS-induced neutrophil recruitment and of the TNF?-like formation contrasting with those from cycloheximide-treated animals (1 mg/kg, i.p.). When delivered in situ, liposome-encapsulated clodronate, a macrophage depletor, significantly impaired both the LPSinduced neutrophil recruitment and the TNF?-like activity. An anti-murine TNF? polyclonal antibody (0.5 h before LPS) was also effective. These results emphasize the pivotal role of macrophages for LPS-induced neutrophil recruitment via the formation of TNF?.
Arreto, C-D.; Dumarey, C.; Nahori, M-A.; Vargaftig, B. B.
The Fusarium toxin deoxynivalenol (DON) modulates the LPS induced acute phase reaction in pigs.
The systemic effects of the Fusarium toxin deoxynivalenol (DON) and of bacterial lipopolysaccharides (LPS) were studied in male castrated pigs (40.4 � 3.7 kg) infused intravenously with either DON or LPS alone (100 ?g DON/kg/h, 7.5 ?g/LPS/kg/h), or together (100 ?g DON plus 7.5 ?g/LPS/kg/h). The Control group received a saline infusion (n=6/treatment, 24h observation period). An additional DON infusion did not exacerbate the clinical signs observed in LPS-infused pigs. For example, rectal temperature climaxed after 4h (40.4 � 0.2�C) and 5h (40.1 � 0.3�C), in the LPS and LPS+DON group, respectively. Saline and DON alone did not induce an acute phase reaction as indicated by unaltered plasma levels of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) while LPS caused a significant rise of both cytokines. TNF-alpha plasma peak concentrations were significantly higher in the LPS compared to the DON+LPS group (94.3 � 17.2 ng/mL vs. 79.2 � 15.7 ng/mL) while IL-6 climaxed earlier in the latter group (3h p.i. vs. 2h p.i.). From the tested clinical-chemical plasma characteristics the total bilirubin concentration and the ASAT activity were strongly elevated by the LPS infusion and additionally increased and decreased by DON, respectively. In conclusion, the LPS-induced effects were only marginally modified by DON. PMID:23603058
D�nicke, Sven; Brosig, Bianca; Kersten, Susanne; Kluess, Jeannette; Kahlert, Stefan; Panther, Patricia; Diesing, Anne-Kathrin; Rothk�tter, Hermann-Josef
Disparate roles of marrow- and parenchymal cell-derived TLR4 signaling in murine LPS-induced systemic inflammation
Systemic inflammatory response syndrome (SIRS) occurs in a range of infectious and non-infectious disease processes. Toll-like receptors (TLRs) initiate such responses. We have shown that parenchymal cell TLR4 activation drives LPS-induced systemic inflammation; SIRS does not develop in mice lacking TLR4 expression on parenchymal cells. The parenchymal cell types whose TLR4 activation directs this process have not been identified. Employing a bone marrow transplant model to compartmentalize TLR4 signaling, we characterized blood neutrophil and cytokine responses, NF-?B1 activation, and Tnf-?, Il6, and Ccl2 induction in several organs (spleen, aorta, liver, lung) near the time of LPS-induced symptom onset. Aorta, liver, and lung gene responses corresponded with both LPS-induced symptom onset patterns and plasma cytokine/chemokine levels. Parenchymal cells in aorta, liver, and lung bearing TLR4 responded to LPS with chemokine generation and were associated with increased plasma chemokine levels. We propose that parenchymal cells direct SIRS in response to LPS.
Juskewitch, Justin E.; Platt, Jeffrey L.; Knudsen, Bruce E.; Knutson, Keith L.; Brunn, Gregory J.; Grande, Joseph P.
Antibacterial cathelicidin peptide CAP11 inhibits the lipopolysaccharide (LPS)-induced suppression of neutrophil apoptosis by blocking the binding of LPS to target cells
Objective: The action of antibacterial cathelicidin CAP11 (cationic antibacterial polypeptide of 11�kDa) on the lipopolysaccharide (LPS)-induced suppression of neutrophil apoptosis was evaluated in vitro.
I. Nagaoka; S. Yomogida; H. Tamura; M. Hirata
Icariin attenuates LPS-induced acute inflammatory responses: Involvement of PI3K\\/Akt and NF-?B signaling pathway
This study aimed to investigate the mechanism underlying the attenuation of LPS-induced lung inflammation by icariin in vivo and in vitro. The anti-inflammatory effects of icariin on LPS-induced acute inflammatory and the molecular mechanism were investigated. Pretreatment with icarrin (20mg\\/kg) could attenuate acute lung inflammation by inhibiting mRNA expressions of tumor necrosis factor alpha (TNF-?), interleukin-6 (IL-6), metalloproteinase cycloxygenase-2 (COX-2),
Chang-Qing Xu; Bao-Jun Liu; Jin-Feng Wu; Yan-Chun Xu; Xiao-Hong Duan; Yu-Xue Cao; Jing-Cheng Dong
2-phenylethynesulfonamide Prevents Induction of Pro-inflammatory Factors and Attenuates LPS-induced Liver Injury by Targeting NHE1-Hsp70 Complex in Mice
The endotoxin-mediated production of pro-inflammatory cytokines plays an important role in the pathogenesis of liver disorders. Heat shock protein (Hsp70) overexpression has established functions in lipopolysaccharide (LPS)-mediated inflammatory response. However, little is known about the role of Hsp70 activity in LPS signaling. We hypothesized that inhibition of Hsp70 substrate binding activity can ameliorate LPS-induced liver injury by decreasing induction of pro-inflammatory factors. In this study, C57/BL6 mice were injected intraperitoneally with LPS and 2-phenylethynesulfonamide (PES), an inhibitor of Hsp70 substrate binding activity. We found that i. PES prevented LPS-induced increase in serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity, infiltration of inflammatory cells, and liver cell apoptosis; ii. PES reduced inducible nitric oxide synthase (iNOS) protein expression as well as serum nitric oxide (NO), tumor necrosis factor-? (TNF-?), and interleukin-6 (IL-6) content in LPS-stimulated mice; iii. PES reduced the mRNA level of iNOS, TNF-?, and IL-6 in LPS-stimulated liver. iiii. PES attenuated the degradation of inhibitor of ?B-? (I?B-?) as well as the phosphorylation and nuclear translocation of nuclear factor-?B (NF-?B) in LPS-stimulated liver. Similar changes in the protein expression of inflammatory markers, I?B-? degradation, and NF-?B phosphorylation and nuclear translocation were observed in RAW 264.7 cells. Further mechanistic studies revealed that PES remarkably reduced the elevation of [Ca2+]i and intracellular pH value (pHi) in LPS-stimulated RAW 264.7 cells. Furthermore, PES significantly reduced the increase in Na+/H+ exchanger 1 (NHE1) association to Hsp70 in LPS-stimulated macrophages and liver, suggesting that NHE1-Hsp70 interaction is required for the involvement of NHE1 in the inflammation response. In conclusion, inhibition of Hsp70 substrate binding activity in vivo reduces the induction of pro-inflammatory factors and prevents LPS-induced liver injury likely by disrupting NHE1-Hsp70 interaction which consequently reduces the activation of I?B-?-NF-?B pathway in liver.
Huang, Chao; Wang, Jia; Chen, Zhuo; Wang, Yuzhe; Zhang, Wei
Open-field and LPS-induced sickness behavior in young chickens: effects of embryonic cocaine and/or ritanserin.
Exposure to drugs of abuse during embryogenesis may adversely affect nervous, immune, and endocrine systems development. We compared exposure on embryonic day 18 (E18) by single or multiple cocaine (COC) injections (56.25 mg/kg total dose for both) or saline on hatching and activity measures. In saline-exposed controls, repeated testing, age, and gender affected activity levels. A single or multiple COC injections increased the median latency to explore and multiple COC injections decreased the median number of lines crossed by female chicks in the open field. We also determined if pretreatment with the serotonin2 (5-HT2) receptor antagonist ritanserin could attenuate COC's effects on open-field behavior as well as behaviors sensitive to immune system stimulation (lipopolysaccharide (LPS)-induced sickness behavior). Eggs containing embryos were pretreated on E17 with 0.4 mg ritanserin/kg or its vehicle followed by multiple COC injections or saline on E18. E18 COC treatment decreased the median number of lines crossed and distress vocalizations in females. Ritanserin pretreatment mitigated the COC induced effects. E18 COC exposure also suppressed LPS-induced sickness behaviors in both males and females, increasing food consumption and the time spent awake and active, as well as decreasing the time spent sleeping. Ritanserin alone had no effect on the food consumed or time spent active, nor did this dose affect COC-induced alterations in sickness behavior. Ritanserin alone decreased time spent sleeping and also failed to affect the COC-induced suppression. Thus, embryonic COC exposure can suppress open field and LPS-induced sickness behavior in the young chick, and ritanserin pretreatment can block the former, but not the latter effects at the dose chosen for these experiments. PMID:9715802
Schrott, L M; Getty, M E; Wacnik, P W; Sparber, S B
The Transcription Factor C/EBP-? Mediates Constitutive and LPS-Inducible Transcription of Murine SerpinB2
SerpinB2 or plasminogen activator inhibitor type 2 (PAI-2) is highly induced in macrophages in response to inflammatory stimuli and is linked to the modulation of innate immunity, macrophage survival, and inhibition of plasminogen activators. Lipopolysaccharide (LPS), a potent bacterial endotoxin, can induce SerpinB2 expression via the toll-like receptor 4 (TLR4) by ?1000-fold over a period of 24 hrs in murine macrophages. To map the LPS-regulated SerpinB2 promoter regions, we transfected reporter constructs driven by the ?5 kb 5'-flanking region of the murine SerpinB2 gene and several deletion mutants into murine macrophages. In addition, we compared the DNA sequence of the murine 5? flanking sequence with the sequence of the human gene for homologous functional regulatory elements and identified several regulatory cis-acting elements in the human SERPINB2 promoter conserved in the mouse. Mutation analyses revealed that a CCAAT enhancer binding (C/EBP) element, a cyclic AMP response element (CRE) and two activator protein 1 (AP-1) response elements in the murine SerpinB2 proximal promoter are essential for optimal LPS-inducibility. Electrophoretic mobility shift (EMSA) and chromatin immunoprecipitation (ChIP) assays demonstrated that LPS induces the formation of C/EBP-? containing complexes with the SerpinB2 promoter. Importantly, both constitutive and LPS-induced SerpinB2 expression was severely abrogated in C/EBP-?-null mouse embryonic fibroblasts (MEFs) and primary C/EBP-?-deficient peritoneal macrophages. Together, these data provide new insight into C/EBP-?-dependent regulation of inflammation-associated SerpinB2 expression.
Gade, Padmaja; Mahony, Donna; Buzza, Marguerite S.; Kalvakolanu, Dhananjaya V.; Antalis, Toni M.
Cobalt Protoporphyrin Accelerates TFEB Activation and Lysosome Reformation during LPS-Induced Septic Insults in the Rat Heart
Lipopolysaccharide (LPS)-induced myocardial dysfunction is caused, at least in part, by mitochondrial dysfunction. Mitochondrial dysfunction and the oxidative damage associated with it are scavenged through various cellular defense systems such as autophagy to prevent harmful effects. Our recent study has demonstrated that cobalt protoporphyrin IX (CoPPIX), a potent inducer of heme oxygenase-1 (HO-1), ameliorates septic liver injuries by enhancing mitochondrial autophagy in rats. In our current study, we show that CoPPIX (5 mg/kg s.c.) not only accelerates the autophagic response but also promotes lysosome reformation in the rat heart treated with LPS (15 mg/kg i.p.). Lysosomal membrane-associated protein-2 (LAMP2), which is essential to the maintenance of lysosomal functions in the heart, is depleted transiently but restored rapidly during LPS administration in the rat. Activation of transcription factor EB (TFEB), a master regulator of lysosomal biogenesis and autophagy, was also observed, indicating a hyper consumption and subsequent reformation of the lysosome to meet the increased demand for autophagosome cleaning. CoPPIX was found to promote these processes and tended to restore the LPS-induced suppression of cardiac performances whilst chloroquine (CQ; 20 mg/kg i.p.), an inhibitor of lysosomes and autophagic protein degradation, abrogates these beneficial effects. The cardioprotective effect of CoPPIX against LPS toxicity was also observed via decreased levels of cardiac releasing enzymes in the plasma. Taken together, our current data indicate that lysosome reformation mediated by TFEB may be involved in cardioprotection against LPS-induced septic insults, and serve as a novel mechanism by which CoPPIX protects the heart against oxidative stress.
Unuma, Kana; Aki, Toshihiko; Funakoshi, Takeshi; Yoshida, Ken-ichi; Uemura, Koichi
Inhibitory effects of cyclic AMP elevating agents on lipopolysaccharide (LPS)-induced microvascular permeability change in mouse skin
Anti-inflammatory effects of cyclic AMP elevating agents were examined in a mouse model of lipopolysaccharide (LPS)-induced microvascular permeability change. Vascular permeability on the back skin was measured by the local accumulation of Pontamine sky blue (PSB) after subcutaneous injection of LPS (400??g site?1) from Salmonella typhimurium. Dye leakage in the skin was significantly increased 2?h after injection of LPS. This LPS-induced dye leakage was suppressed by phosphodiesterase inhibitors, including pentoxifylline (160?mg?kg?1), milrinone (5?�?10?mg?kg?1), rolipram (0.5?�?10?mg?kg?1) and zaprinast (5?�?10?mg?kg?1). The dye leakage was also inhibited by ?-adrenoceptor agonists, including isoproterenol (0.5?�?5?mg?kg?1) and salbutamol (0.05?�?5?mg?kg?1), an adenylate cyclase activator, forskolin (5?mg?kg?1), and a cell permeable cyclic AMP analogue, 8-bromo-cyclic AMP (8-Br-cAMP, 10?mg?kg?1). LPS caused a transient increase in serum TNF-? level peaking at 1?h after the injection. This increase in serum TNF-? was completely blocked by a pretreatment with pentoxifylline (160?mg?kg?1), milrinone (5?mg?kg?1), rolipram (1?mg?kg?1), zaprinast (10?mg?kg?1), salbutamol (0.5?mg?kg?1), forskolin (1?mg?kg?1) and 8-Br-cAMP (10?mg?kg?1). LPS caused an increase in serum IL-1? level peaking at 3?h after injection. This increase in serum IL-1? was not significantly suppressed by the cyclic AMP elevating agents. Our study suggests that cyclic AMP elevating agents attenuate LPS-induced microvascular permeability change by suppressing TNF-? up regulation.
Irie, Kaoru; Fujii, Emiko; Ishida, Hiroyasu; Wada, Keiji; Suganuma, Taiyo; Nishikori, Tomohiro; Yoshioka, Toshimasa; Muraki, Takamura
Anti-inflammatory activity of cinnamon water extract in vivo and in vitro LPS-induced models
Background Cinnamon bark is one of the most popular herbal ingredients in traditional oriental medicine and possesses diverse pharmacological activities including anti-bacterial, anti-viral, and anti-cancer properties. The goal of this study is to investigate the in vivo and in vitro inhibitory effect of cinnamon water extract (CWE) on lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-? and its underlying intracellular mechanisms. Methods CWE was orally administrated to mice for 6 days prior to intraperitoneal injection of LPS. Serum levels of TNF-? and interleukin (IL)-6 were determined 1 hour after LPS stimulation. Peritoneal macrophages from thioglycollate-injected mice were isolated and assayed for viability, cytokine expression and signaling molecules upon LPS stimulation. CWE was further fractioned according to molecular size, and the levels of total polyphenols and biological activities of each fraction were measured. Results The oral administration of CWE to mice significantly decreased the serum levels of TNF-? and IL-6. CWE treatment in vitro decreased the mRNA expression of TNF-?. CWE blocked the LPS-induced degradation of I?B? as well as the activation of JNK, p38 and ERK1/2. Furthermore, size-based fractionation of CWE showed that the observed inhibitory effect of CWE in vitro occurred in the fraction containing the highest level of total polyphenols. Conclusions Treatment with CWE decreased LPS-induced TNF-? in serum. In vitro inhibition of TNF-? gene by CWE may occur via the modulation of I?B? degradation and JNK, p38, and ERK1/2 activation. Our results also indicate that the observed anti-inflammatory action of CWE may originate from the presence of polyphenols.
Low level laser therapy reduces acute lung inflammation in a model of pulmonary and extrapulmonary LPS-induced ARDS.
The present study aimed to investigate the effects low level laser therapy (LLLT) in a LPS-induced pulmonary and extrapulmonary acute respiratory distress syndrome (ARDS) in BALB/c mice. Laser (830nm laser, 9J/cm(2), 35mW, 80s per point, 3 points per application) was applied in direct contact with skin, 1h after LPS administration. Mice were distributed in control (n=6; PBS), ARDS IT (n=7; LPS orotracheally 10?g/mouse), ARDS IP (n=7; LPS intra-peritoneally 100?g/mouse), ARDS IT+Laser (n=9; LPS intra-tracheally 10?g/mouse), ARDS IP+Laser (n=9; LPS intra-peritoneally 100?g/mouse). Twenty-four hours after last LPS administration, mice were studied for pulmonary inflammation by total and differential cell count in bronchoalveolar lavage (BAL), cytokines (IL-1beta, IL-6, KC and TNF-alpha) levels in BAL fluid and also by quantitative analysis of neutrophils number in the lung parenchyma. LLLT significantly reduced pulmonary and extrapulmonary inflammation in LPS-induced ARDS, as demonstrated by reduced number of total cells (p<0.001) and neutrophils (p<0.001) in BAL, reduced levels of IL-1beta, IL-6, KC and TNF-alpha in BAL fluid and in serum (p<0.001), as well as the number of neutrophils in lung parenchyma (p<0.001). LLLT is effective to reduce pulmonary inflammation in both pulmonary and extrapulmonary model of LPS-induced ARDS. PMID:24792475
Oliveira, Manoel Carneiro; Greiffo, Fl�via Regina; Rigonato-Oliveira, Nicole Cristine; Cust�dio, Ricardo Wesley Alberca; Silva, Vanessa Roza; Damaceno-Rodrigues, Nilsa Regina; Almeida, Francine Maria; Albertini, Regiane; Lopes-Martins, Rodrigo �lvaro B; de Oliveira, Luis Vicente Franco; de Carvalho, Paulo de Tarso Camillo; Ligeiro de Oliveira, Ana Paula; Leal, Ernesto C�sar P; Vieira, Rodolfo P
Walnut extract inhibits LPS-induced activation of BV-2 microglia via internalization of TLR4: possible involvement of phospholipase D2.
Walnuts are a rich source of essential fatty acids, including the polyunsaturated fatty acids alpha-linolenic acid and linoleic acid. Essential fatty acids have been shown to modulate a number of cellular processes in the brain, including the activation state of microglia. Microglial activation can result in the generation of cytotoxic intermediates and is associated with a variety of age-related and neurodegenerative conditions. In vitro, microglial activation can be induced with the bacterial cell wall component lipopolysaccharide (LPS). In the present study, we generated a methanolic extract of English walnuts (Juglans regia) and examined the effects of walnut extract exposure on LPS-induced activation in BV-2 microglial cells. When cells were treated with walnut extract prior to LPS stimulation, production of nitric oxide and expression of inducible nitric oxide synthase were attenuated. Walnut extract also induced a decrease in tumor necrosis-alpha (TNFalpha) production. We further found that walnut extract induced internalization of the LPS receptor, toll-like receptor 4, and that the anti-inflammatory effects of walnut were dependent on functional activation of phospholipase D2. These studies represent the first to describe the anti-inflammatory effects of walnuts in microglia, which could lead to nutritional interventions in the prevention and treatment of neurodegeneration. PMID:20213499
Willis, Lauren M; Bielinski, Donna F; Fisher, Derek R; Matthan, Nirupa R; Joseph, James A
Microbial transformation of deoxyandrographolide and their inhibitory activity on LPS-induced NO production in RAW 264.7 macrophages.
A series of analogues of deoxyandrographolide (1) transformed by Cunninghamella blakesleana AS 3.2004 were isolated and identified by spectral methods including 2D NMR. Among them, 3-oxo-17,19-dihydroxy-7,13-ent-labdadien-15,16-olide (9), 3-oxo-19-hydroxy-1,13-ent-labdadien-15,16-olide (16), 3-oxo-1?-hydroxy-14-deoxy-andrographolide (17) and 3-oxo-2?-hydroxy-14-deoxyandrographolide (18) are new compounds. And their structure-activity relationships (SAR) of inhibitory activity on LPS-induced NO production in RAW 264.7 macrophage cells were also discussed. PMID:22264489
Deng, Sa; Zhang, Bao Jing; Wang, Chang Yuan; Tian, Yan; Yao, Ji Hong; An, Lei; Huang, Shan Shan; Peng, Jin Yong; Liu, Ke Xin; Ma, Xiao Chi
Artemisia fukudo essential oil attenuates LPS-induced inflammation by suppressing NF-kappaB and MAPK activation in RAW 264.7 macrophages.
In the present study, the chemical constituents of Artemisia fukudo essential oil (AFE) were investigated using GC-MS. The major constituents were alpha-thujone (48.28%), beta-thujone (12.69%), camphor (6.95%) and caryophyllene (6.01%). We also examined the effects of AFE on the production of nitric oxide (NO), prostaglandin E(2) (PGE(2)), tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and IL-6, in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Western blotting and RT-PCR tests indicated that AFE has potent dose-dependent inhibitory effects on pro-inflammatory cytokines and mediators. We investigated the mechanism by which AFE inhibits NO and PGE(2) by examining the level of nuclear factor-kappaB (NF-kappaB) activation within the mitogen-activated protein kinase (MAPK) pathway, which is an inflammation-induced signal pathway in RAW 264.7 cells. AFE inhibited LPS-induced ERK, JNK, and p38 phosphorylation. Furthermore, AFE inhibited the LPS-induced phosphorylation and degradation of Ikappa-B-alpha, which is required for the nuclear translocations of the p50 and p65 NF-kappaB subunits in RAW 264.7 cells. Our results suggest that AFE might exert an anti-inflammatory effect by inhibiting the expression of pro-inflammatory cytokines. Such an effect is mediated by a blocking of NF-kappaB activation which consequently inhibits the generation of inflammatory mediators in RAW264.7 cells. AFE may be useful for treating inflammatory diseases. PMID:20156520
Yoon, W J; Moon, J Y; Song, G; Lee, Y K; Han, M S; Lee, J S; Ihm, B S; Lee, W J; Lee, N H; Hyun, C G
Chloroform fraction of Solanum tuberosum L. cv Jayoung epidermis suppresses LPS-induced inflammatory responses in macrophages and DSS-induced colitis in mice.
In this study, the authors investigated the molecular mechanism underlying the antiinflammatory effects of the chloroform fraction of the peel of 'Jayoung' (CFPJ), a color-fleshed potato, on lipopolysaccharide (LPS)-induced RAW 264.7 macrophages and in mice with dextran sulfate sodium (DSS)-induced colitis. CFPJ inhibited the expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at the transcription level, and attenuated the transcriptional activity of nuclear factor-?B (NF-?B) by reducing the translocation of NF-?B depending on degradation of inhibitory ?B-? (I?B-?). Furthermore, CFPJ attenuated the phosphorylations of mitogen-activated protein kinase kinases3/6 (MKK3/6) and of p38. In colitis model, CFPJ significantly reduced the severity of colitis and the productions and protein levels of pro-inflammatory mediators in colonic tissue. These results suggest that the anti-inflammatory effects of CFPJ are associated with the suppression of NF-?B and p38 activation in macrophages, and support its possible therapeutic role for the treatment of colitis. PMID:24184733
Lee, Seung-Jun; Shin, Ji-Sun; Choi, Hye-Eun; Lee, Kyoung-Goo; Cho, Young-Wuk; An, Hyo-Jin; Jang, Dae Sik; Jeong, Jin-Cheol; Kwon, Oh-Keun; Nam, Jung-Hwan; Lee, Kyung-Tae
Salidroside Reduces Cell Mobility via NF-?B and MAPK Signaling in LPS-Induced BV2 Microglial Cells
The unregulated activation of microglia following stroke results in the production of toxic factors that propagate secondary neuronal injury. Salidroside has been shown to exhibit protective effects against neuronal death induced by different insults. However, the molecular mechanisms responsible for the anti-inflammatory activity of salidroside have not been elucidated clearly in microglia. In the present study, we investigated the molecular mechanism underlying inhibiting LPS-stimulated BV2 microglial cell mobility of salidroside. The protective effect of salidroside was investigated in microglial BV2 cell, subjected to stretch injury. Moreover, transwell migration assay demonstrated that salidroside significantly reduced cell motility. Our results also indicated that salidroside suppressed LPS-induced chemokines production in a dose-dependent manner, without causing cytotoxicity in BV2 microglial cells. Moreover, salidroside suppressed LPS-induced activation of nuclear factor kappa B (NF-?B) by blocking degradation of I?B? and phosphorylation of MAPK (p38, JNK, ERK1/2), which resulted in inhibition of chemokine expression. These results suggest that salidroside possesses a potent suppressive effect on cell migration of BV2 microglia and this compound may offer substantial therapeutic potential for treatment of ischemic strokes that are accompanied by microglial activation.
Hu, Haixia; Li, Zuanfang; Zhu, Xiaoqin; Lin, Ruhui; Chen, Lidian
Newly synthesized 'hidabeni' chalcone derivatives potently suppress LPS-induced NO production via inhibition of STAT1, but not NF-?B, JNK, and p38, pathways in microglia.
Chalcones are open-chain flavonoids that are biosynthesized in various plants. Some of them possess anti-inflammatory activity. We previously found that chalcone glycosides from Brassica rapa L. 'hidabeni' suppress lipopolysaccharide (LPS)-induced nitric oxide (NO) production in rat microglia highly aggressively proliferating immortalized (HAPI) cells. In this study, to explore chalcone derivatives with potent NO inhibitory activity, we synthesized ten compounds based on 'hidabeni' chalcone and examined their effects on LPS-triggered inducible NO synthase (iNOS) expression and NO production. Compounds C4 and C10 potently inhibited NO production (IC50: 4.19, 2.88?�M, respectively). C4 and C10 suppressed LPS-induced iNOS expression via the inhibition of the signal transduction and activator of transcription 1 (STAT1), but not nuclear factor-kappa B (NF-?B), c-Jun N terminal kinase (JNK), and p38, pathways. C10, but not C4, inhibited activation of the MEK/extracellular signal-regulated kinase (ERK) pathway. C4 and C10 also suppressed LPS-induced expression of interferon regulatory factor 1 (IRF-1), which is an important transcription factor involved in iNOS expression. Our findings indicate that these chalcone derivatives are candidate compounds for preventing microglia-mediated neuroinflammation. PMID:24882415
Hara, Hirokazu; Ikeda, Ryoko; Ninomiya, Masayuki; Kamiya, Tetsuro; Koketsu, Mamoru; Adachi, Tetsuo
Antinociceptive Effect of Tetrandrine on LPS-Induced Hyperalgesia via the Inhibition of IKK? Phosphorylation and the COX-2/PGE2 Pathway in Mice
Tetrandrine (TET) is a bisbenzylisoquinoline alkaloid that is isolated from the Stephania Tetrandra. It is known to possess anti-inflammatory and immunomodulatory effects. We have shown that TET can effectively suppress the production of bacterial lipopolysaccharide (LPS)-induced inflammatory mediators, including cyclooxygenases (COXs), in macrophages. However, whether TET has an antinociceptive effect on LPS-induced hyperalgesia is unknown. In the present study, we investigated the potential antinociceptive effects of TET and the mechanisms by which it elicits its effects on LPS-induced hyperalgesia. LPS effectively evoked hyperalgesia and induced the production of PGE2 in the sera, brain tissues, and cultured astroglia. TET pretreatment attenuated all of these effects. LPS also activated inhibitor of ?B (I?B) kinase ? (IKK?) and its downstream components in the I?B/nuclear factor (NF)-?B signaling pathway, including COX-2; the increase in expression levels of these components was significantly abolished by TET. Furthermore, in primary astroglia, knockdown of IKK?, but not IKK?, reversed the effects of TET on the LPS-induced increase in I?B phosphorylation, P65 phosphorylation, and COX-2. Our results suggest that TET can effectively exert antinociceptive effects on LPS-induced hyperalgesia in mice by inhibiting IKK? phosphorylation, which leads to the reduction in the production of important pain mediators, such as PGE2 and COX-2, via the IKK?/I?B/NF-?B pathway.
Zhao, Hengguang; Luo, Fuling; Li, Hongzhong; Zhang, Li; Yi, Yongfen; Wan, Jingyuan
Apigenin Protects Endothelial Cells from Lipopolysaccharide (LPS)-Induced Inflammation by Decreasing Caspase-3 Activation and Modulating Mitochondrial Function
Acute and chronic inflammation is characterized by increased reactive oxygen species (ROS) production, dysregulation of mitochondrial metabolism and abnormal immune function contributing to cardiovascular diseases and sepsis. Clinical and epidemiological studies suggest potential beneficial effects of dietary interventions in inflammatory diseases but understanding of how nutrients work remains insufficient. In the present study, we evaluated the effects of apigenin, an anti-inflammatory flavonoid abundantly found in our diet, in endothelial cells during inflammation. Here, we show that apigenin reduced lipopolysaccharide (LPS)-induced apoptosis by decreasing ROS production and the activity of caspase-3 in endothelial cells. Apigenin conferred protection against LPS-induced mitochondrial dysfunction and reestablished normal mitochondrial complex I activity, a major site of electron leakage and superoxide production, suggesting its ability to modulate endothelial cell metabolic function during inflammation. Collectively, these findings indicate that the dietary compound apigenin stabilizes mitochondrial function during inflammation preventing endothelial cell damage and thus provide new translational opportunities for the use of dietary components in the prevention and treatment of inflammatory diseases.
Duarte, Silvia; Arango, Daniel; Parihar, Arti; Hamel, Patrice; Yasmeen, Rumana; Doseff, Andrea I.
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) prevents lipopolysaccharide (LPS)-induced, sepsis-related severe acute lung injury in mice
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an energy metabolism-related enzyme in the glycolytic pathway. Recently, it has been reported that GAPDH has other physiological functions, such as apoptosis, DNA repair and autophagy. Some in vitro studies have indicated immunological aspects of GAPDH function, although there is no definite study discussing the advantage of GAPDH as a therapeutic target. Here, we show that GAPDH has an anti-inflammatory function by using a lipopolysaccharide (LPS)-induced, sepsis-related severe acute lung injury (ALI) mouse model, which is referred to as acute respiratory distress syndrome (ARDS) in humans. GAPDH pre-injected mice were protected from septic death, and their serum levels of proinflammatory cytokines were significantly suppressed. In lung tissue, LPS-induced acute injury and neutrophil accumulation were strongly inhibited by GAPDH pre-injection. Pulmonary, proinflammatory cytokine gene expression and serum chemokine expression in GAPDH pre-injected mice were also reduced. These data suggest the therapeutic potential of GAPDH for sepsis-related ALI/ARDS.
Takaoka, Yuki; Goto, Shigeru; Nakano, Toshiaki; Tseng, Hui-Peng; Yang, Shih-Ming; Kawamoto, Seiji; Ono, Kazuhisa; Chen, Chao-Long
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) prevents lipopolysaccharide (LPS)-induced, sepsis-related severe acute lung injury in mice.
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an energy metabolism-related enzyme in the glycolytic pathway. Recently, it has been reported that GAPDH has other physiological functions, such as apoptosis, DNA repair and autophagy. Some in vitro studies have indicated immunological aspects of GAPDH function, although there is no definite study discussing the advantage of GAPDH as a therapeutic target. Here, we show that GAPDH has an anti-inflammatory function by using a lipopolysaccharide (LPS)-induced, sepsis-related severe acute lung injury (ALI) mouse model, which is referred to as acute respiratory distress syndrome (ARDS) in humans. GAPDH pre-injected mice were protected from septic death, and their serum levels of proinflammatory cytokines were significantly suppressed. In lung tissue, LPS-induced acute injury and neutrophil accumulation were strongly inhibited by GAPDH pre-injection. Pulmonary, proinflammatory cytokine gene expression and serum chemokine expression in GAPDH pre-injected mice were also reduced. These data suggest the therapeutic potential of GAPDH for sepsis-related ALI/ARDS. PMID:24902773
Takaoka, Yuki; Goto, Shigeru; Nakano, Toshiaki; Tseng, Hui-Peng; Yang, Shih-Ming; Kawamoto, Seiji; Ono, Kazuhisa; Chen, Chao-Long
Acute Hypoxia Decreases E. coli LPS-Induced Cytokine Production and NF-?B Activation in Alveolar Macrophages*
Reductions in alveolar oxygenation during lung hypoxia/reoxygenation (H/R) injury are common after gram-negative endotoxemia. However, the effects of H/R on endotoxin-stimulated cytokine production by alveolar macrophages are unclear and may depend upon thresholds for hypoxic oxyradical generation in situ. Here TNF-? and IL-? production were determined in rat alveolar macrophages stimulated with E. coli lipopolysaccharide (LPS, serotype O55:B5) while exposed to either normoxia for up to 24 h, to brief normocarbic hypoxia (1.5 h at an atmospheric PO2 = 10 � 2 mm Hg), or to combined H/R. LPS-induced TNF-? and IL-? were reduced at the peak of hypoxia and by reoxygenation in LPS + H/R cells (P < 0.01) compared with normoxic controls despite no changes in reduced glutathione (GSH) or in PGE2 production. Both TNF-? mRNA and NF-?B activation were reduced by hypoxia that suppressed superoxide anion generation. Thus, dynamic reductions in the ambient PO2 of alveolar macrophages that do not deplete GSH suppress LPS-induced TNF-? expression, IL-? production, and NF-?B activation even as oxyradical production is decreased.
Matuschak, George M.; Nayak, Ravi; Doyle, Timothy M.; Lechner, Andrew J.
Expression of Coagulation-Related Protein Genes During LPS-Induced Preterm Delivery in the Pregnant Mouse
Preterm delivery (PTD) has been associated with inflammation along with activation of the coagulation pathway. These studies sought to characterize the expression of several coagulation pathway genes including plasminogen activator inhibitor 1 (PAI-1), tissue factor (TF), protease-activated receptor 1 (Par1), protease-activated receptor 2 (Par2), fibrinogen-like protein 2 (Fgl2), and thrombomodulin (TM) during lipopolysaccharide (LPS)-induced PTD in day 15 pregnant CD-1 mice. Western blot studies confirmed protein expression for PAI-1, Par1, Par2, Fgl2, and TM in the mouse uterus. Quantitative reverse transcriptase polymerase chain reaction (RT-PCR) confirmed increased PAI-1 messenger RNA (mRNA) in the uteri, lung, kidney, and liver tissues at 2 to 6 hours after LPS injection. In contrast, TF expression significantly decreased by 12 hours in uterine tissue; whereas, its expression was unchanged in the other maternal tissues. The uterine mRNA for Par1, Par2, Fgl2, and TM remained stable. In summary, these studies have confirmed expression of coagulation pathway genes within the pregnant uterus; some of which are modulated during LPS-induced PTD.
Diamond, Allaire K.; Sweet, Leigh M.; Oppenheimer, Karen H.; Bradley, Diana F.
Resveratrol Inhibits LPS-Induced MAPKs Activation via Activation of the Phosphatidylinositol 3-Kinase Pathway in Murine RAW 264.7 Macrophage Cells
Background Resveratrol is a natural polyphenolic compound that has cardioprotective, anticancer and anti-inflammatory properties. We investigated the capacity of resveratrol to protect RAW 264.7 cells from inflammatory insults and explored mechanisms underlying inhibitory effects of resveratrol on RAW 264.7 cells. Methodology/Principal Findings Murine RAW 264.7 cells were treated with resveratrol (1, 5, and 10 �M) and/or LPS (5 �g/ml). Nitric oxide (NO) and prostaglandin E2 (PGE2) were measured by Griess reagent and ELISA. The mRNA and protein levels of proinflammatory proteins and cytokines were analysed by ELISA, RT-PCR and double immunofluorescence labeling, respectively. Phosphorylation levels of Akt, cyclic AMP-responsive element-binding protein (CREB), mitogen-activated protein kinases (MAPKs) cascades, AMP-activated protein kinase (AMPK) and expression of SIRT1(Silent information regulator T1) were measured by western blot. Wortmannin (1 �M), a specific phosphatidylinositol 3-kinase (PI3-K) inhibitor, was used to determine if PI3-K/Akt signaling pathway might be involved in resveratrol�s action on RAW 264.7 cells. Resveratrol significantly attenuated the LPS-induced expression of nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-? (TNF-?) and interleukin-1? (IL-1?) in RAW 264.7 cells. Resveratrol increased Akt phosphorylation in a time-dependent manner. Wortmannin, a specific phosphatidylinositol 3-kinase (PI3-K) inhibitor, blocked the effects of resveratrol on LPS-induced RAW 264.7 cells activation. In addition, PI3-K inhibition partially abolished the inhibitory effect of resveratrol on the phosphorylation of cyclic AMP-responsive element-binding protein (CREB) and mitogen-activated protein kinases (MAPKs) cascades. Meanwhile, PI3-K is essential for resveratrol-mediated phosphorylation of AMPK and expression of SIRT1. Conclusion and Implications This investigation demonstrates that PI3-K/Akt activation is an important signaling in resveratrol-mediated activation of AMPK phosphorylation and SIRT1 expression, and inhibition of phosphorylation of CREB and MAPKs activation, proinflammatory mediators and cytokines production in response to LPS in RAW 264.7 cells.
Liu, Bin; Deng, Yi-Shu; Zhan, Dong; Chen, Yuan-Li; He, Ying; Liu, Jing; Zhang, Zong-Ji; Sun, Jun; Lu, Di
LPS induces pp60c-src-mediated tyrosine phosphorylation of Hsp90 in lung vascular endothelial cells and mouse lung
Heat shock protein 90 (Hsp90) inhibitors were initially developed as anticancer agents; however, it is becoming increasing clear that they also possess potent anti-inflammatory properties. Posttranslational modifications of Hsp90 have been reported in tumors and have been hypothesized to affect client protein- and inhibitor-binding activities. In the present study we investigated the posttranslational modification of Hsp90 in inflammation. LPS, a prototypical inflammatory agent, induced concentration- and time-dependent tyrosine (Y) phosphorylation of Hsp90? and Hsp90? in bovine pulmonary arterial and human lung microvascular endothelial cells (HLMVEC). Mass spectrometry identified Y309 as a major site of Y phosphorylation on Hsp90? (Y300 of Hsp90?). LPS-induced Hsp90 phosphorylation was prevented by the Hsp90 inhibitor 17-allyl-amino-demethoxy-geldanamycin (17-AAG) in vitro as well as in lungs from LPS-treated mice, in vivo. Furthermore, 17-AAG prevented LPS-induced pp60src activation. LPS-induced Hsp90 phosphorylation was also prevented by the pp60src inhibitor PP2. Additionally, Hsp90 phosphorylation was induced by infecting cells with a constitutively active pp60src adenovirus, whereas either a dominant-negative pp60src adenovirus or reduced expression of pp60src by a specific siRNA prevented the LPS-induced Y phosphorylation of Hsp90. Transfection of HLMVEC with the nonphosphorylatable Hsp90? Y300F mutant prevented LPS-induced Hsp90? tyrosine phosphorylation but not pp60src activation. Furthermore, the Hsp90? Y300F mutant showed a reduced ability to bind the Hsp90 client proteins eNOS and pp60src and HLMVEC transfected with the mutant exhibited reduced LPS-induced barrier dysfunction. We conclude that inflammatory stimuli cause posttranslational modifications of Hsp90 that are Hsp90-inhibitor sensitive and may be important to the proinflammatory actions of Hsp90.
Barabutis, Nektarios; Handa, Vaishali; Dimitropoulou, Christiana; Rafikov, Ruslan; Snead, Connie; Kumar, Sanjiv; Joshi, Atul; Thangjam, Gagan; Fulton, David; Black, Stephen M.; Patel, Vijay
In vivo Angiotensin II AT1 receptor blockade selectively inhibits LPS-induced innate immune response and ACTH release in rat pituitary gland.
Systemic lipopolysaccharide (LPS) administration induces an innate immune response and stimulates the hypothalamic-pituitary-adrenal axis. We studied Angiotensin II AT(1) receptor participation in the LPS effects with focus on the pituitary gland. LPS (50 microg/kg, i.p.) enhanced, 3h after administration, gene expression of pituitary CD14 and that of Angiotensin II AT(1A) receptors in pituitary and hypothalamic paraventricular nucleus (PVN); stimulated ACTH and corticosterone release; decreased pituitary CRF(1) receptor mRNA and increased all plasma and pituitary pro-inflammatory factors studied. The AT(1) receptor blocker (ARB) candesartan (1mg/kg/day, s.c. daily for 3 days before LPS) blocked pituitary and PVN AT(1) receptors, inhibited LPS-induced ACTH but not corticosterone secretion and decreased LPS-induced release of TNF-alpha, IL-1beta and IL-6 to the circulation. The ARB reduced LPS-induced pituitary gene expression of IL-6, LIF, iNOS, COX-2 and IkappaB-alpha; and prevented LPS-induced increase of nNOS/eNOS activity. The ARB did not affect LPS-induced TNF-alpha and IL-1beta gene expression, IL-6 or IL-1beta protein content or LPS-induced decrease of CRF(1) receptors. When administered alone, the ARB increased basal plasma corticosterone levels and basal PGE(2) mRNA in pituitary. Our results demonstrate that the pituitary gland is a target for systemically administered LPS. AT(1) receptor activity is necessary for the complete pituitary response to LPS and is limited to specific pro-inflammatory pathways. There is a complementary and complex influence of the PVN and circulating cytokines on the initial pituitary response to LPS. Our findings support the proposal that ARBs may be considered for the treatment of inflammatory conditions. PMID:19427376
S�nchez-Lemus, Enrique; Benicky, Julius; Pavel, Jaroslav; Saavedra, Juan M
Enhanced PDE4B expression augments LPS-inducible TNF expression in ethanol-primed monocytes: relevance to alcoholic liver disease
Increased plasma and hepatic TNF-? expression is well documented in patients with alcoholic hepatitis and is implicated in the pathogenesis of alcoholic liver disease. We have previously shown that monocytes from patients with alcoholic hepatitis show increased constitutive and LPS-induced NF-?B activation and TNF-? production. Our recent studies showed that chronic ethanol exposure significantly decreased cellular cAMP levels in both LPS-stimulated and unstimulated monocytes and Kupffer cells, leading to an increase in LPS-inducible TNF-? production by affecting NF-?B activation and induction of TNF mRNA expression. Accordingly, the mechanisms underlying this ethanol-induced decrease in cellular cAMP leading to an increase in TNF expression were examined in monocytes/macrophages. In this study, chronic ethanol exposure was observed to significantly increase LPS-inducible expression of cAMP-specific phosphodiesterase (PDE)4B that degrades cellular cAMP. Increased PDE4B expression was associated with enhanced NF-?B activation and transcriptional activity and subsequent priming of monocytes/macrophages leading to enhanced LPS-inducible TNF-? production. Selective inhibition of PDE4 by rolipram abrogated LPS-mediated TNF-? expression at both protein and mRNA levels in control and ethanol-treated cells. Notably, PDE4 inhibition did not affect LPS-inducible NF-?B activation but significantly decreased NF-?B transcriptional activity. These findings strongly support the pathogenic role of PDE4B in the ethanol-mediated priming of monocytes/macrophages and increased LPS-inducible TNF production and the subsequent development of alcoholic liver disease (ALD). Since enhanced TNF expression plays a significant role in the evolution of clinical and experimental ALD, its downregulation via selective PDE4B inhibitors could constitute a novel therapeutic approach in the treatment of ALD.
Gobejishvili, Leila; Barve, Shirish; Joshi-Barve, Swati; McClain, Craig
Sequential release of TNF? and phospholipase A2 in a rat model of LPS-induced pleurisy
The levels of extracellular phospholipase A2 (sPLA2) and TNF?, and cell accumulation were measured in the pleural washings obtained at different times following the induction of Escherichia coli lipopolysaccharide (LPS, 100 ?g/cavity) pleurisy in rats. TNF? peaked at 2 hours (3036 � 160.3 units/ml) and decreased thereafter. Conversely, levels of sPLA2 peaked at 48 hours (1.97 � 0.64 ng/ml) and were increased further (14.02 � 4.16 ng/ml) by pretreatment with anti-TNF? antibody. Cell accumulation was not affected by antibody pretreatment. These data indicate that the sPLA2 enzyme is involved in LPS-induced pleurisy. The enzyme seems not to be stimulated by TNF? which may be involved in the downregulation of sPLA2 in this model of inflammation.
Bucci, M.; D?Acquisto, F.; Parente, L.; Cirino, G.
LPS-Induced Lung Inflammation in Marmoset Monkeys - An Acute Model for Anti-Inflammatory Drug Testing
Increasing incidence and substantial morbidity and mortality of respiratory diseases requires the development of new human-specific anti-inflammatory and disease-modifying therapeutics. Therefore, new predictive animal models that closely reflect human lung pathology are needed. In the current study, a tiered acute lipopolysaccharide (LPS)-induced inflammation model was established in marmoset monkeys (Callithrix jacchus) to reflect crucial features of inflammatory lung diseases. Firstly, in an ex vivo approach marmoset and, for the purposes of comparison, human precision-cut lung slices (PCLS) were stimulated with LPS in the presence or absence of the phosphodiesterase-4 (PDE4) inhibitor roflumilast. Pro-inflammatory cytokines including tumor necrosis factor-alpha (TNF-?) and macrophage inflammatory protein-1 beta (MIP-1?) were measured. The corticosteroid dexamethasone was used as treatment control. Secondly, in an in vivo approach marmosets were pre-treated with roflumilast or dexamethasone and unilaterally challenged with LPS. Ipsilateral bronchoalveolar lavage (BAL) was conducted 18 hours after LPS challenge. BAL fluid was processed and analyzed for neutrophils, TNF-?, and MIP-1?. TNF-? release in marmoset PCLS correlated significantly with human PCLS. Roflumilast treatment significantly reduced TNF-? secretion ex vivo in both species, with comparable half maximal inhibitory concentration (IC50). LPS instillation into marmoset lungs caused a profound inflammation as shown by neutrophilic influx and increased TNF-? and MIP-1? levels in BAL fluid. This inflammatory response was significantly suppressed by roflumilast and dexamethasone. The close similarity of marmoset and human lungs regarding LPS-induced inflammation and the significant anti-inflammatory effect of approved pharmaceuticals assess the suitability of marmoset monkeys to serve as a promising model for studying anti-inflammatory drugs.
Seehase, Sophie; Lauenstein, Hans-Dieter; Schlumbohm, Christina; Switalla, Simone; Neuhaus, Vanessa; Forster, Christine; Fieguth, Hans-Gerd; Pfennig, Olaf; Fuchs, Eberhard; Kaup, Franz-Josef; Bleyer, Martina; Hohlfeld, Jens M.; Braun, Armin
The regulation of cytochrome P450 2E1 during LPS-induced inflammation in the rat
It is well known that inflammatory and infectious conditions differentially regulate cytochrome P450 (P450)-mediated drug metabolism in the liver. We have previously outlined a potential pathway for the downregulation in hepatic cytochrome P450 following LPS-mediated inflammation in the CNS (Abdulla, D., Goralski, K.B., Garcia Del Busto Cano, E., Renton, K.W., 2005. The signal transduction pathways involved in hepatic cytochrome P450 regulation in the rat during an LPS-induced model of CNS inflammation. Drug Metab. Dispos). The purpose of this study was to outline the effects of LPS-induced peripheral and central nervous system inflammation on hepatic cytochrome P450 2E1 (CYP2E1) in vivo, an enzyme that plays an important role in various physiological and pathological states. We report an increase in hepatic mRNA expression of CYP2E1 that occurred as early as 2-3 h following either the intraperitoneal (i.p.) injection of 5 mg/kg LPS or i.c.v. administration of 25 {mu}g of LPS. This increase in CYP2E1 mRNA expression was sustained for 24 h. In sharp contrast to the increase in hepatic CYP2E1 mRNA, we observed a significant reduction in the catalytic activity of this enzyme 24 h following either the i.c.v. or i.p. administration of LPS. Cycloheximide or actinomycin-D did not change the LPS-mediated downregulation in hepatic CYP2E1 catalytic activity. Our results support the idea that LPS acts at two different levels to regulate hepatic CYP2E1: a transcriptional level to increase CYP2E1 mRNA expression and a post-transcriptional level to regulate CYP2E1 protein and activity.
Abdulla, Dalya [Department of Pharmacology, Sir Charles Tupper Medical Bldg., Dalhousie University, Halifax, Nova Scotia, B3H 4H7 (Canada); Goralski, Kerry B. [Department of Pharmacology, Sir Charles Tupper Medical Bldg., Dalhousie University, Halifax, Nova Scotia, B3H 4H7 (Canada); College of Pharmacy, Burbidge Building, Dalhousie University, Halifax, Nova Scotia, B3H 3J5 (Canada); Renton, Kenneth W. [Department of Pharmacology, Sir Charles Tupper Medical Bldg., Dalhousie University, Halifax, Nova Scotia, B3H 4H7 (Canada)]. E-mail: email@example.com
Gigantol isolated from the whole plants of Cymbidium goeringii inhibits the LPS-induced iNOS and COX-2 expression via NF-kappaB inactivation in RAW 264.7 macrophages cells.
During our efforts to find bioactive natural products with anti-inflammatory activity, we isolated gigantol from the whole plants of Cymbidium goeringii (Orchidaceae) by activity-guided chromatographic fractionation. Gigantol was found to have potent inhibitory effects on LPS-induced nitric oxide (NO) and prostaglandin E (2) (PGE (2)) production in RAW 264.7 cells. Consistent with these findings, gigantol suppressed the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at the protein and mRNA levels in RAW 264.7 cells in a concentration-dependent manner. Our data also indicate that gigantol is a potent inhibitor of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) release and influenced the mRNA expression levels of these cytokines in a dose-dependent manner. Furthermore, a reporter gene assay for nuclear factor kappa B (NF-kappaB) and an electromobility shift assay (EMSA) demonstrated that gigantol effectively inhibited the activation of NF-kappaB, which is necessary for the expression of iNOS, COX-2, TNF-alpha, IL-1beta and IL-6. Thus, our studies suggest that gigantol inhibits LPS-induced iNOS and COX-2 expression by blocking NF- kappaB activation. PMID:16924582
Won, Jong-Heon; Kim, Ji-Yeon; Yun, Kyung-Jin; Lee, Jin-Hee; Back, Nam-In; Chung, Hae-Gon; Chung, Sun A; Jeong, Tae-Sook; Choi, Myung-Sook; Lee, Kyung-Tae
Santamarin, a sesquiterpene lactone isolated from Saussurea lappa, represses LPS-induced inflammatory responses via expression of heme oxygenase-1 in murine macrophage cells.
Saussurea lappa C.B. Clarke (Compositae) is indigenous to India and Pakistan. The dried root of S. lappa has been traditionally used for alleviating pain in abdominal distention and tenesmus, indigestion with anorexia, dysentery, nausea, and vomiting. Santamarin is a sesquiterpene lactone isolated from S. lappa. In the present study, santamarin inhibited inducible nitric oxide synthase (iNOS) protein, reduced iNOS-derived nitric oxide (NO), suppressed COX-2 protein and reduced COX-derived PGE(2) production in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and murine peritoneal macrophages. Similarly, santamarin reduced tumor necrosis factor-? (TNF-?) and interleukin-1? (IL-1?) production. In addition, santamarin suppressed the phosphorylation and degradation of I?B-? as well as the nuclear translocation of p65 in response to LPS in RAW264.7 cells. Furthermore, santamarin induced heme oxygenase (HO)-1 expression mRNA and protein level that plays a cytoprotective role against inflammation. The induction of HO-1 is primarily regulated at the transcriptional level, and its induction by various agents is mediated by the nuclear transcription factor E2-related factor 2 (Nrf2), master regulator of antioxidant responses. Unbound Nrf2 translocates into the nucleus and binds to the antioxidant response element (ARE) in the upstream promoter region of many antioxidative genes, where it initiates their transcription. The effects of santamarin on LPS-induced NO, PGE(2), TNF-?, and IL-1? production were partially reversed by the HO-1 inhibitor, tin protoporphyrin (SnPP). Therefore, our data suggest that the anti-inflammatory effect of santamarin in macrophages may be exerted through a novel mechanism that involves HO-1 expression. PMID:22564506
Choi, Hyun-Gyu; Lee, Dong-Sung; Li, Bin; Choi, Yeon Ho; Lee, Seung-Ho; Kim, Youn-Chul
Pharmacological inhibition of GSK-3 in a guinea pig model of LPS-induced pulmonary inflammation: I. Effects on lung remodeling and pathology
Background Glycogen synthase kinase-3 (GSK-3) is a constitutively active kinase that regulates multiple signalling proteins and transcription factors involved in a myriad of cellular processes. The kinase acts as a negative regulator in ?-catenin signalling and is critically involved in the smad pathway. Activation of both pathways may contribute to pulmonary features of chronic obstructive pulmonary disease (COPD). Methods In the present study, we investigated the effect of the selective GSK-3 inhibitor SB216763 on pulmonary pathology in a guinea pig model of lipopolysaccharide (LPS)-induced COPD. Guinea pigs were instilled intranasally with LPS or saline twice weekly for 12�weeks and pre-treated with either intranasally instilled SB216763 or corresponding vehicle 30�min prior to each LPS/saline challenge. Results Repeated LPS exposures activated ?-catenin signalling, primarily in the airway epithelium and submucosa. LPS also induced pulmonary inflammation and tissue remodelling as indicated by inflammatory cell influx, increased pulmonary fibronectin expression and enhanced small airway collagen content. Inhibition of GSK-3 by SB216763 did not affect LPS-induced inflammatory cell influx, but prevented the small airway remodelling and, unexpectedly, inhibited the activation of ?-catenin in vivo. LPS or SB216763 treatment had no effect on the airway smooth muscle content and alveolar airspace size. However, GSK-3 inhibition prevented LPS-induced right ventricle hypertrophy. Conclusions Our findings indicate that GSK-3 inhibition prevents LPS-induced pulmonary pathology in guinea pigs, and that locally reduced LPS-induced ?-catenin activation appears in part to underlie this effect.
Protective effect of resveratrol against LPS-induced extracellular lipoperoxidation in AR42J cells partly via a Myd88-dependent signaling pathway.
Lipopolysaccharides (LPS) are major components of the cell wall of Gram negative bacteria implicated in the pathogenesis of bacterial infection. Resveratrol is a polyphenolic phytoalexin exhibiting antioxidant and anti-inflammatory properties. We investigated the protective effects of this natural compound on LPS-induced proinflammatory effect using non-myeloid AR42J pancreatic cells. We found that LPS dose-dependently increased extracellular malondialdehyde (MDA) and nitric oxide without affecting their intracellular level whereas resveratrol abolished all these deleterious effects. LPS increased CD14 expression; IRAK1 and a phosphorylated form of p38 MAPK protein. Resveratrol counteracted LPS effect by decreasing CD14 and IRAK1 expression but unexpectedly increased the p38 MAPK protein phosphorylation. Altogether, our data highlighted the functionality of the TLR4-Myd88 signaling pathway in LPS pro-oxidant effect using non-myeloid cells. They further suggested that resveratrol exerted antioxidant properties either by a Myd88-dependent way not involving IRAK1 or by a TRIF dependent pathway. PMID:20035708
Sebai, Hichem; Ristorcelli, Elodie; Sbarra, Veronique; Hovsepian, Sonia; Fayet, Guy; Aouani, Ezzedine; Lombardo, Dominique
Neolignans from the fruits of Magnolia obovata and their inhibition effect on NO production in LPS-induced RAW 264.7 cells.
Three new neolignans, named 9-methoxyobovatol (6), magnobovatol (7), and 2-hydroxyobovaaldehyde (9), along with six known ones, magnolol (1), honokiol (2), isomagnolol (3), obovatol (4), obovatal (5), and obovaaldehyde (8), were isolated from the fruits of Magnolia obovata using silica gel and ODS column chromatography. From the results of spectroscopic data including EIMS, IR, 1H- and 13C-NMR, DEPT, and 2D-NMR (gCOSY, gHSQC, gHMBC), the chemical structures were determined. All isolated compounds were evaluated for inhibition activity on nitric oxide production in LPS-induced RAW 264.7 cells, and compounds 1-4, 6, 7, and 9 showed significant activity with IC50 values of 15.8 � 0.3, 3.3 � 1.2, 14.1 � 0.9, 6.2 � 1.2, 14.8 � 2.3, 14.2 � 1.2, and 14.8 � 3.2 �M, respectively, without any visible toxic effect. PMID:23970426
Seo, Kyeong-Hwa; Lee, Dae-Young; Lee, Dong-Sung; Park, Ji-Hae; Jeong, Rak-Hun; Jung, Ye-Jin; Shrestha, Sabina; Chung, In-Sik; Kim, Geum-Soog; Kim, Youn-Chul; Baek, Nam-In
Ketamine\\/Xylazine attenuates LPS-induced iNOS expression in various rat tissues 1,2 1 Presented at the annual meeting of the Association of Academic Surgery, Boston, MA, November 7�9, 2002. 2 Supported by NIGMS grant GM38529 and GM08792
Ketamine and xylazine (K\\/X) are commonly used in combination as an anesthetic agent in experimental animal models. We previously noted that K\\/X attenuated lipopolysaccharide (LPS)-induced liver injury, gastric stasis, and reduced symptoms of endotoxemia. Because ketamine attenuates expression of several proinflammatory genes, we examined the effects of K\\/X on inducible nitric oxide synthase (iNOS), which has been implicated in endotoxin-induced
Kenneth S Helmer; Yan Cui; Ashvin Dewan; David W Mercer
Riluzole partially rescues age-associated, but not LPS-induced, loss of glutamate transporters and spatial memory.
Impaired memory may result from synaptic glutamatergic dysregulation related to chronic neuroinflammation. GLT1 is the primary excitatory amino acid transporter responsible for regulating extracellular glutamate levels in the hippocampus. We tested the hypothesis that if impaired spatial memory results from increased extracellular glutamate due to age or experimentally induced chronic neuroinflammation in the hippocampus, then pharmacological augmentation of the glutamate transporter GLT1 will attenuate deficits in a hippocampal-dependent spatial memory task. The profile of inflammation-related genes and proteins associated with normal aging, or chronic neuroinflammation experimentally-induced via a four-week LPS infusion into the IV(th) ventricle, were correlated with performance in the Morris water maze following treatment with Riluzole, a drug that can enhance glutamate clearance by increasing GLT1 expression. Age-associated inflammation was qualitatively different from LPS-induced neuro-inflammation in young rats. LPS produced a pro-inflammatory phenotype characterized by increased IL-1� expression in the hippocampus, whereas aging was not associated with a strong central pro-inflammatory response but with a mixed peripheral immune phenotype. Riluzole attenuated the spatial memory impairment, the elevation of serum cytokines and the decrease in GLT1 gene expression in Aged rats, but had no effect on young rats infused with LPS. Our findings highlight the therapeutic potential of reducing glutamatergic function upon memory impairment in neurodegenerative diseases associated with aging. PMID:23709339
Brothers, Holly M; Bardou, Isabelle; Hopp, Sarah C; Kaercher, Roxanne M; Corona, Angela W; Fenn, Ashley M; Godbout, Jonathan P; Wenk, Gary L
EP2 and EP4 receptors on muscularis resident macrophages mediate LPS-induced intestinal dysmotility via iNOS upregulation through cAMP/ERK signals
Intestinal resident macrophages play an important role in gastrointestinal dysmotility by producing prostaglandins (PGs) and nitric oxide (NO) in inflammatory conditions. The causal correlation between PGs and NO in gastrointestinal inflammation has not been elucidated. In this study, we examined the possible role of PGE2 in the LPS-inducible inducible NO synthase (iNOS) gene expression in murine distal ileal tissue and macrophages. Treatment of ileal tissue with LPS increased the iNOS and cyclooxygenase (COX)-2 gene expression, which lead to intestinal dysmotility. However, LPS did not induce the expression of iNOS and COX-2 in tissue from macrophage colony-stimulating factor-deficient op/op mice, indicating that these genes are expressed in intestinal resident macrophages. iNOS and COX-2 protein were also expressed in dextran-phagocytized macrophages in the muscle layer. CAY10404, a COX-2 inhibitor, diminished LPS-dependent iNOS gene upregulation in wild-type mouse ileal tissue and also in RAW264.7 macrophages, indicating that PGs upregulate iNOS gene expression. EP2 and EP4 agonists upregulated iNOS gene expression in ileal tissue and isolated resident macrophages. iNOS mRNA induction mediated by LPS was decreased in the ileum isolated from EP2 or EP4 knockout mice. In addition, LPS failed to decrease the motility of EP2 and EP4 knockout mice ileum. EP2- or EP4-mediated iNOS expression was attenuated by KT-5720, a PKA inhibitor and PD-98059, an ERK inhibitor. Forskolin or dibutyryl-cAMP mimics upregulation of iNOS gene expression in macrophages. In conclusion, COX-2-derived PGE2 induces iNOS expression through cAMP/ERK pathways by activating EP2 and EP4 receptors in muscularis macrophages. NO produced in muscularis macrophages induces dysmotility during gastrointestinal inflammation.
Tajima, Tsuyoshi; Murata, Takahisa; Aritake, Kosuke; Urade, Yoshihiro; Michishita, Masaki; Matsuoka, Toshiyuki; Narumiya, Shuh; Ozaki, Hiroshi
Suppressive Effect of CORM-2 on LPS-Induced Platelet Activation by Glycoprotein Mediated HS1 Phosphorylation Interference
In recent years, it has been discovered that septic patients display coagulation abnormalities. Platelets play a major role in the coagulation system. Studies have confirmed that carbon monoxide (CO) has important cytoprotective and anti-inflammatory function. However, whether CO could alter abnormal activation of platelets and coagulation and thereby reduce the incidence of mortality during sepsis has not been defined. In this report, we have used CO-releasing molecules (CORM-2) to determine whether CO inhibits LPS-induced abnormal activation of platelets and have explored the potential mechanisms. LPS was used to induce activation of platelets in vitro, which were purified from the peripheral venous blood of healthy adult donors. CORM-2 was applied as a potential therapeutic agent. CORM-2 preconditioning and delayed treatment were also studied. We found that in the LPS groups, the function of platelets such as spreading, aggregation, and release were enhanced abnormally. By contrast, the platelets in the CORM-2 group were gently activated. Further studies showed that the expression of platelet membrane glycoproteins increased in the LPS group. Coincidently, both hematopoietic lineage cell-specific protein 1 and its phosphorylated form also increased dramatically. These phenomena were less dramatically seen in the CORM-2 groups. Taken together, we conclude that during LPS stimulation, platelets were abnormally activated, and this functional state may be associated with the signal that is transmitted between membrane glycoproteins and HS1. CORM-released CO suppresses the abnormal activation of platelets by interfering with glycoprotein-mediated HS1 phosphorylation.
Liu, Dadong; Liang, Feng; Wang, Xu; Cao, Jie; Qin, Weiting; Sun, Bingwei
Effects of Supplemental Glutamine on Growth Performance, Plasma Parameters and LPS-induced Immune Response of Weaned Barrows after Castration
Two experiments were conducted to investigate the effects of supplemental glutamine on growth performance, plasma parameters and LPS-induced immune response of weaned barrows after castration. In experiment 1, forty-eight weaned male piglets were used and fed maize and soybean meal diets supplemented with 0 (Control) or 2% L-Gln (Gln+) for 25 days. The results indicated that the Gln+ group tended to increase average daily gain compared to control in stages of days 7 to 14 and 0 to 25. The Gln+ had significantly better feed efficiency than the control group did during days 14 to 25 and 0 to 25. The plasma blood urea nitrogen and alkaline phosphatase contents of Gln+ group were higher than those of the control group on day 14 post-weaning. In experiment 2, sixteen weaned male piglets were injected with E. coli K88+ lipopolysaccharide (LPS) on day 14 post-weaning. The results showed that the Gln+ group had lower concentrations of plasma adrenocorticotrophic hormone and cortisol than the control group on day 14 pre-LPS challenge. In addition, Gln+ group had higher plasma IgG concentration than the control group for pre- or post-LPS challenged on day 14 post-weaning. In summary, dietary supplementation of Gln was able to alleviate the stressful condition and inflammation associated with castration in weaned barrows, and to improve their immunity and growth performance in the early starter stage.
Hsu, C. B.; Lee, J. W.; Huang, H. J.; Wang, C. H.; Lee, T. T.; Yen, H. T.; Yu, B.
Microbial transformation of acetyl-11-keto-?-boswellic acid and their inhibitory activity on LPS-induced NO production.
The capabilities of 20 strains of fungi to transform acetyl-11-keto-?-boswellic (AKBA) were screened. And biotransformation of AKBA by Cunninghamella blakesleana AS 3.970 afforded five metabolites (1-5), while two metabolites (6, 7) were isolated from biotransformation of Cunninghamella elegans AS 3.1207. The chemical structures of these metabolites were identified by spectral methods including 2D NMR and their structures were elucidated as 7?-hydroxy-3-acety-11-keto-?-boswellic acid (1), 21?-dihydroxy-3-acety-11-keto-?-boswellic acid (2), 7?,22?-dihydroxy-3-acety-11-keto-?-boswellic acid (3), 7?,16?-dihydroxy-3-acety-11-keto-?-boswellic acid (4), 7?,15?-dihydroxy-3-acety-11-keto-?-boswellic acid (5); 7?,15?,21?-trihydroxy-3-acety-11-keto-?-boswellic acid (6) and 15?,21?-dihydroxy-3-acety-11-keto-?-boswellic acid (7). All these products are previously unknown. Their primary structure-activity relationships (SAR) of inhibition activity on LPS-induced NO production in RAW 264.7 macrophage cells were evaluated. PMID:23391590
Sun, Yan; Liu, Dan; Xi, Ronggang; Wang, Xiaobo; Wang, Yan; Hou, Jie; Zhang, Baojing; Wang, Changyuan; Liu, Kexin; Ma, Xiaochi
S-Form Lipopolysaccharide (LPS), but Not Lipid A or R-Chemo-type LPS, Induces Interleukin6 Production in Vitamin D 3-Differentiated THP1 Cells
Bacterial lipopolysaccharide (LPS) induces the production of various inflammatory cytokines and the inducibility is considered attributable to the glycolipid part of LPS called lipid A. We report anin vitromodel in which lipid A is not necessarily a minimal structure for the LPS activity. Vitamin D3-differentiated THP-1 cells, cultured human monocytic leukemia cells, produced a high level of interleukin-6 (IL-6) by
Yasuo Suda; Kazue Aoyama; Kazumi Arimoto; Toshihide Tamura; Shoichi Kusumoto
Abstract The immune system is believed to be a sensitive indicator for adverse polychlorinated biphenyl (PCB)-induced health effects. Four commercial PCB mixtures (Aroclors) or six individual PCB congeners were evaluated for their effect on splenocyte viability and lip...
CaMKI? regulates AMPK-dependent, TORC-1 independent autophagy during LPS-induced acute lung neutrophilic inflammation1,2
Autophagy is an evolutionarily conserved cytoplasmic process regulated by the energy rheostats mTOR and AMPK that recycles damaged or unused proteins and organelles. It has been described as an important effector arm of immune cells. We have shown that the cytoplasmically oriented calcium/calmodulin-dependent protein kinase I ? (CaMKI?) regulates the inflammatory phenotype of the macrophage (M?). Here, we hypothesize that CaMKI? mediates M? autophagy. LPS induced autophagy in RAW 264.7 cells and murine peritoneal M? that was attenuated with biochemical CaMK inhibition or CaMKI? siRNA. Inhibition of CaMKI? reduced LPS-induced p-Thr172AMPK and TORC1 activity, and expression of a constitutively active CaMKI? but not a kinase-deficient mutant induced p-Thr172AMPK and autophagy that was attenuated by the AMPK inhibitor Compound C. Co-immunoprecipitation and in vitro kinase assays demonstrated that CaMKI? activates AMPK, thereby inducing ATG7, which also localizes to this CaMKI?-AMPK complex. During LPS-induced lung inflammation, C57Bl/6 mice receiving CaMKI?siRNA displayed reduced lung and bronchoalveolar immune cell autophagy that correlated with reduced neutrophil recruitment, myeloperoxidase activity, and air space cytokine concentration. Independently inhibiting autophagy, using siRNA targeting the PI3 kinase VPS34, yielded similar reductions in lung autophagy and neutrophil recruitment. Thus, a novel CaMKI?-AMPK pathway is rapidly activated in M? exposed to LPS and regulates an early autophagic response, independent of TORC1 inhibition. These mechanisms appear to be operant in vivo in orchestrating LPS-induced lung neutrophil recruitment and inflammation.
Guo, Lanping; Stripay, Jennifer L.; Zhang, Xianghong; Collage, Richard D.; Hulver, Mei; Carchman, Evie H.; Howell, Gina M.; Zuckerbraun, Brian S.; Lee, Janet S.; Rosengart, Matthew R.
Bis-N-norgliovictin, a small-molecule compound from marine fungus, inhibits LPS-induced inflammation in macrophages and improves survival in sepsis.
Sepsis is a highly lethal disorder characterized by systemic inflammation, and Toll-like receptor 4 (TLR4) in macrophages plays a crucial role in modulating innate immune response and outcome of sepsis. During the screening of natural products against inflammation, we identified bis-N-norgliovictin, a small-molecule compound isolated from marine-derived fungus, significantly inhibited lipopolysaccharide (LPS, ligand of TLR4)-induced tumor necrosis factor-? (TNF-?) production in RAW264.7 cells. In this study, we evaluated the effect of bis-N-norgliovictin on TLR4-mediated inflammation in mouse macrophages and LPS-induced sepsis model. In RAW264.7 and mouse peritoneal macrophages, bis-N-norgliovictin dose-dependently inhibited LPS-induced production of TNF-?, interleukin-6 (IL-6), interferon-? (IFN-?) and monocyte chemoattractant protein (MCP-1), but without suppressing cell viability. The anti-inflammatory effect was attributed to the down-regulation of TLR4-triggered myeloid differentiation primary response protein 88 (MyD88)-dependent and TIR-containing adapter inducing interferon-? (TRIF)-dependent signaling pathways, including p38 and c-Jun N-terminal kinase (JNK) of mitogen-activated protein kinases (MAPKs), nuclear factor-?B (NF-?B) and interferon regulatory factor 3 (IRF3) cascades. Importantly, bis-N-norgliovictin also protected mice against LPS-induced endotoxic shock. Intravenous injection of bis-N-norgliovictin 1h before LPS challenge dose-dependently inhibited LPS-induced increases in serum levels of TNF-?, IL-6, MCP-1 and IL-10, attenuated liver and lung injury and diminished M1 macrophage polarization in liver. Our results demonstrate that bis-N-norgliovictin exhibit potent anti-inflammatory effect both in vitro and in vivo. These findings suggest that bis-N-norgliovictin can be a useful therapeutic candidate for the treatment of sepsis and other inflammatory diseases. PMID:23438875
Song, Yuxian; Dou, Huan; Gong, Wei; Liu, Xianqin; Yu, Zhiguo; Li, Erguang; Tan, Renxiang; Hou, Yayi
Chronic Morphine Treatment Inhibits LPS Induced Angiogenesis: Implications in Wound healing
Delayed wound healing is a chronic problem in opioid drug abusers. We investigated the role chronic morphine plays on later stages of wound healing events using an angiogenesis model. Our results show that morphine treatment resulted in a significant decrease in inflammation induced angiogenesis. To delineate the mechanisms involved we investigate the role of hypoxia inducible factor 1 alpha (HIF-1 alpha), a potent inducer of angiogenic growth factor. Morphine treatment resulted in a significant decrease in the expression and nuclear translocation of HIF-1 alpha with a concurrent suppression in vascular endothelial growth factor (VEGF) synthesis. Cells of the innate immune system play a dominant role in the angiogenic process. Morphine treatment inhibited early recruitment of both neutrophils and monocytes towards an inflammatory signal with a significant decrease in the monocyte chemoattractant MCP-1. Taken together, our studies show that morphine regulates the wound repair process on multiple levels. Morphine acts both directly and indirectly in suppressing angiogenesis.
Martin, Josephine L.; Charboneau, Richard; Barke, Roderick A.; Roy, Sabita
Protection against LPS-Induced Pulmonary Edema through the Attenuation of Protein Tyrosine Phosphatase-1B Oxidation
One hallmark of acute lung injury is the disruption of the pulmonary endothelial barrier. Such disruption correlates with increased endothelial permeability, partly through the disruption of cell�cell contacts. Protein tyrosine phosphatases (PTPs) are known to affect the stability of both cell�extracellular matrix adhesions and intercellular adherens junctions (AJs). However, evidence for the role of select PTPs in regulating endothelial permeability is limited. Our investigations noted that the inhibition of PTP1B in cultured pulmonary endothelial cells (ECs), as well as in the vasculature of intact murine lungs via the transient overexpression of a catalytically inactive PTP1B, decreased the baseline resistance of cultured EC monolayers and increased the formation of edema in murine lungs, respectively. In addition, we observed that the overexpression of wild-type PTP1B enhanced basal barrier function in vitro. Immunohistochemical analyses of pulmonary ECs and the coimmunoprecipitation of murine lung homogenates demonstrated the association of PTP1B with the AJ proteins ?-catenin, p120-catenin, and VE-cadherin both in vitro and ex vivo. Using LPS in a model of sepsis-induced acute lung injury, we showed that reactive oxygen species were generated in response to LPS, which correlated with enhanced PTP1B oxidation, inhibited phosphatase activity, and attenuation of the interactions between PTP1B and ?-catenin, as well as enhanced ?-catenin tyrosine phosphorylation. Finally, the overexpression of a cytosolic PTP1B fragment, shown to be resistant to nicotinamide adenine dinucleotide phosphate�reduced oxidase�4 (Nox4)-mediated oxidation, significantly attenuated LPS-induced endothelial barrier dysfunction and the formation of lung edema, and preserved the associations of PTP1B with AJ protein components, independent of PTP1B phosphatase activity. We conclude that PTP1B plays an important role in maintaining the pulmonary endothelial barrier, and PTP1B oxidation appears to contribute to sepsis-induced pulmonary vascular dysfunction, possibly through the disruption of AJs.
Grinnell, Katie L.; Chichger, Havovi; Braza, Julie; Duong, Huetran
Therapeutic Effect of Intravenous Infusion of Perfluorocarbon Emulsion on LPS-Induced Acute Lung Injury in Rats
Acute lung injury (ALI) and its more severe form, acute respiratory distress syndrome (ARDS) are the leading causes of death in critical care. Despite extensive efforts in research and clinical medicine, mortality remains high in these diseases. Perfluorocarbon (PFC), a chemical compound known as liquid ventilation medium, is capable of dissolving large amounts of physiologically important gases (mainly oxygen and carbon dioxide). In this study we aimed to investigate the effect of intravenous infusion of PFC emulsion on lipopolysaccharide (LPS) induced ALI in rats and elucidate its mechanism of action. Forty two Wistar rats were randomly divided into three groups: 6 rats were treated with saline solution by intratracheal instillation (control group), 18 rats were treated with LPS by intratracheal instillation (LPS group) and the other 18 rats received PFC through femoral vein prior to LPS instillation (LPS+PFC group). The rats in the control group were sacrificed 6 hours later after saline instillation. At 2, 4 and 6 hours of exposure to LPS, 6 rats in the LPS group and 6 rats in LPS+PFC group were sacrificed at each time point. By analyzing pulmonary pathology, partial pressure of oxygen in the blood (PaO2) and lung wet-dry weight ratio (W/D) of each rat, we found that intravenous infusion of PFC significantly alleviated acute lung injury induced by LPS. Moreover, we showed that the expression of pulmonary myeloperoxidase (MPO), intercellular adhesion molecule-1 (ICAM-1) of endothelial cells and CD11b of polymorphonuclear neutrophils (PMN) induced by LPS were significantly decreased by PFC treatment in vivo. Our results indicate that intravenous infusion of PFC inhibits the infiltration of PMNs into lung tissue, which has been shown as the core pathogenesis of ALI/ARDS. Thus, our study provides a theoretical foundation for using intravenous infusion of PFC to prevent and treat ALI/ARDS in clinical practice.
Lv, Qi; Yin, Xiaofeng; Song, Jianqi; Landen, Ning Xu; Fan, Haojun
Therapeutic effect of intravenous infusion of perfluorocarbon emulsion on LPS-induced acute lung injury in rats.
Acute lung injury (ALI) and its more severe form, acute respiratory distress syndrome (ARDS) are the leading causes of death in critical care. Despite extensive efforts in research and clinical medicine, mortality remains high in these diseases. Perfluorocarbon (PFC), a chemical compound known as liquid ventilation medium, is capable of dissolving large amounts of physiologically important gases (mainly oxygen and carbon dioxide). In this study we aimed to investigate the effect of intravenous infusion of PFC emulsion on lipopolysaccharide (LPS) induced ALI in rats and elucidate its mechanism of action. Forty two Wistar rats were randomly divided into three groups: 6 rats were treated with saline solution by intratracheal instillation (control group), 18 rats were treated with LPS by intratracheal instillation (LPS group) and the other 18 rats received PFC through femoral vein prior to LPS instillation (LPS+PFC group). The rats in the control group were sacrificed 6 hours later after saline instillation. At 2, 4 and 6 hours of exposure to LPS, 6 rats in the LPS group and 6 rats in LPS+PFC group were sacrificed at each time point. By analyzing pulmonary pathology, partial pressure of oxygen in the blood (PaO2) and lung wet-dry weight ratio (W/D) of each rat, we found that intravenous infusion of PFC significantly alleviated acute lung injury induced by LPS. Moreover, we showed that the expression of pulmonary myeloperoxidase (MPO), intercellular adhesion molecule-1 (ICAM-1) of endothelial cells and CD11b of polymorphonuclear neutrophils (PMN) induced by LPS were significantly decreased by PFC treatment in vivo. Our results indicate that intravenous infusion of PFC inhibits the infiltration of PMNs into lung tissue, which has been shown as the core pathogenesis of ALI/ARDS. Thus, our study provides a theoretical foundation for using intravenous infusion of PFC to prevent and treat ALI/ARDS in clinical practice. PMID:24489970
Hou, Shike; Ding, Hui; Lv, Qi; Yin, Xiaofeng; Song, Jianqi; Land�n, Ning Xu; Fan, Haojun
B and T lymphocyte attenuator inhibits LPS-induced endotoxic shock by suppressing Toll-like receptor 4 signaling in innate immune cells
Although innate immune responses are necessary for the initiation of acquired immune responses and the subsequent successful elimination of pathogens, excessive responses occasionally result in lethal endotoxic shock accompanied by a cytokine storm. B and T lymphocyte attenuator (BTLA), a coinhibitory receptor with similarities to cytotoxic T-lymphocyte antigen (CTLA)-4 and programmed death (PD)-1, is expressed in not only B and T cells but also dendritic cells (DCs) and macrophages (M?s). Recently, several studies have reported that BTLA-deficient (BTLA?/?) mice show enhanced pathogen clearance compared with WT mice in early phase of infections. However, the roles of BTLA expressed on innate cells in overwhelming and uncontrolled immune responses remain unclear. Here, we found that BTLA?/? mice were highly susceptible to LPS-induced endotoxic shock. LPS-induced TNF-? and IL-12 production in DCs and M?s was significantly enhanced in BTLA?/? mice. BTLA?/? DCs also produced high levels of TNF-? on stimulation with Pam3CSK4 but not poly(I:C) or CpG, suggesting that BTLA functions as an inhibitory molecule on Toll-like receptor signaling at cell surface but not endosome. Moreover, BTLA?/? DCs showed enhanced MyD88- and toll/IL-1R domain-containing adaptor inducing IFN (TRIF)-dependent signaling on LPS stimulation, which is associated with impaired accumulation of Src homology 2-containing protein tyrosine phosphatase in lipid rafts. Finally, we found that an agonistic anti-BTLA antibody rescued mice from LPS-induced endotoxic shock, even if the antibody was given to mice that had developed a sign of endotoxic shock. These results suggest that BTLA directly inhibits LPS responses in DCs and M?s and that agonistic agents for BTLA might have therapeutic potential for LPS-induced endotoxic shock.
Kobayashi, Yoshihisa; Iwata, Arifumi; Suzuki, Kotaro; Suto, Akira; Kawashima, Saki; Saito, Yukari; Owada, Takayoshi; Kobayashi, Midori; Watanabe, Norihiko; Nakajima, Hiroshi
Treatment of experimental mouse bladder tumour by LPS-induced epithelial cell shedding.
The purpose of the present study was to explore the therapeutic potential of serial administration of shedding-inducing endotoxin in a mouse tumour bladder model. The studies were conducted with two variants derived from the MBT-2 tumour namely, T5 and T50, the latter being far more aggressive than the former. It was found that T5 tumours responded to intravesical lipopolysaccharides (LPS) instillation by a considerable reduction in their pace of growth (P < 0.0001) when treatment was initiated 3 days after tumour implantation, but not when started after 7 days. The T50 variant did not respond to LPS when treated 3 days after implantation, but a considerable reduction in rate of growth occurred when treatment was started after 1-2 days. Shedding induced by intravesically instilled LPS was found to retard considerably the progression rate of experimental bladder tumour.
Nativ, O.; Medalia, O.; Mor, Y.; Shajrawi, I.; Sabo, E.; Aronson, M.
Sulforaphane suppressed LPS-induced inflammation in mouse peritoneal macrophages through Nrf2 dependent pathway
Sulforaphane (SFN) is a natural isothiocyanate that is present in cruciferous vegetables such as broccoli and cabbage. Previous studies have shown that SFN is effective in preventing carcinogenesis induced by carcinogens in rodents, which is related in part to its potent anti-inflammation properties. In the present study, we compared the anti-inflammatory effect of SFN on LPS-stimulated inflammation in primary peritoneal
Wen Lin; Rachel T. Wu; Tienyuan Wu; Tin-Oo Khor; Hu Wang; Ah-Ng Kong
Anti?inflammatory effects of triptolide by inhibiting the NF??B signalling pathway in LPS?induced acute lung injury in a murine model.
Triptolide is one of the main active components in the Chinese herb Tripterygium wilfordii Hook�F, which has been demonstrated to possess anti?inflammatory properties. The aim of this study was to investigate the effects of triptolide on lipopolysaccharide (LPS)?induced acute lung injury (ALI) in mice and to explore the possible mechanisms. Mice were administered LPS intranasally to induce lung injury, and triptolide was administered intraperitoneally 1�h prior to the LPS challenge. Triptolide?treated mice exhibited significantly reduced levels of leukocytes, myeloperoxidase activity and edema of the lung, as well as tumour necrosis factor??, interleukin (IL)?1? and IL?6 production in the bronchoalveolar lavage fluid compared with LPS?treated mice. Additionally, western blot analysis showed that triptolide inhibited the LPS?induced phosphorylation of nuclear factor of ? light polypeptide gene enhancer in B cells inhibitor?? and nuclear factor ??light?chain?enhancer of activated B�cells?p65 (NF??B�p65) and the expression of Toll?like receptor�4 (TLR4). In conclusion, the results from the present study suggest that the anti?inflammatory effect of triptolide against LPS?induced ALI may be due to its ability to inhibit the TLR4?mediated NF??B signalling pathway. Triptolide may therefore be a promising potential therapeutic agent for ALI treatment, which may ultimately aid the clinical therapy for patients with ALI. PMID:24789089
Wang, Xian; Zhang, Lei; Duan, Wei; Liu, Bin; Gong, Ping; Ding, Yusong; Wu, Xiongwen
Upregulation of miR-146a contributes to the suppression of inflammatory responses in LPS-induced acute lung injury.
Despite the critical role of microRNA in inflammatory response, little is known about its function in inflammation-induced Acute Lung Injury (ALI)/Acute Respiratory Distress Syndrome (ARDS). To investigate the potential role of microRNA146a (miR-146a) in ALI, we used lipopolysaccharide (LPS)-induced ALI rat model. Our data revealed that LPS-induced lung injury in rats resulted in significant upregulation of proinflammatory cytokine tumor necrosis factor-alpha (TNF-?), IL-6, IL-1?, and miR-146a expression. LPS treatment also leads to higher expression of miR-146a as well as increase in secretion of TNF-?, IL-6, and IL-1? in alveolar macrophage (AM) NR8383 cells in a time-dependent manner. Manipulation with miR146a mimic significantly suppressed LPS-mediated TNF-?, IL-6, and IL-1? induction in NR8383 cells by repressing expression of IRAK-1 and TRAF-6. These data clearly indicate that the upregulation of miR146a suppresses inflammatory mediators in LPS induced-ALI model. Therefore, miR-146a may be therapeutically targeted as a mean to repress inflammatory response following ALI. PMID:23848342
Zeng, Zhenguo; Gong, Honghan; Li, Yong; Jie, Kemin; Ding, Chengzhi; Shao, Qiang; Liu, Fen; Zhan, Yian; Nie, Cheng; Zhu, Weifeng; Qian, Kejian
Redox Regulation of Lipopolysaccharide(LPS)Induced Interleukin8 (IL8) Gene Expression Mediated by NF?B and AP1 in Human Astrocytoma U373 Cells
LPS-induced expression of the IL-8 gene was markedly enhanced by H2O2or by deprivation of the cellular antioxidant glutathione by L-buthionine-(S,R)-sulfoximine (BSO) in human astrocytoma U373 cells. In contrast, it was markedly suppressed by the reductant N-acetyl-L-cysteine (NAC) and other antioxidants. Transient expression analysis using the chloramphenicol acetyltransferase assay revealed that activation of the IL-8 promoter by LPS was stimulated by
Chihiro Tanaka; Hideaki Kamata; Hidenori Takeshita; Hitoshi Yagisawa; Hajime Hirata
Inhibition of LPS-induced apoptosis in differentiated-PC12 cells by new triazine derivatives through NF-?B-mediated suppression of COX2
Anti-inflammatory therapy approaches have been in the focus of attention in the treatment of neurodegenerative diseases, such as Alzheimer's disease (AD). In this study, we examined the role of new 1,2,4-triazine derivatives against cytotoxicity exerted by lipopolysaccharide (LPS) in differentiated rat pheochromocytoma (PC12) cell line. Our results indicated that LPS-induced cell death can be inhibited in the presence of some
Niloufar Ansari; Fariba Khodagholi; Mahmoudreza Ramin; Mohsen Amini; Hamid Irannejad; Leila Dargahi; Azim Dehghani Amirabad
Lipoxin A4 and platelet activating factor are involved in E. coli or LPS-induced lung inflammation in CFTR-deficient mice.
CFTR (cystic fibrosis transmembrane conductance regulator) is expressed by both neutrophils and platelets. Lack of functional CFTR could lead to severe lung infection and inflammation. Here, we found that mutation of CFTR (F508del) or inhibition of CFTR in mice led to more severe thrombocytopenia, alveolar neutrocytosis and bacteriosis, and lower lipoxin A4/MIP-2 (macrophage inhibitory protein-2) or lipoxin A4/neutrophil ratios in the BAL (bronchoalveolar lavage) during acute E. coli pneumonia. In vitro, inhibition of CFTR promotes MIP-2 production in LPS-stimulated neutrophils; however, lipoxin A4 could dose-dependently suppress this effect. In LPS-induced acute lung inflammation, blockade of PSGL-1 (P-selectin glycoprotein ligand-1) or P-selectin, antagonism of PAF by WEB2086, or correction of mutated CFTR trafficking by KM11060 could significantly increase plasma lipoxin A4 levels in F508del relevant to wildtype mice. Concurrently, F508del mice had higher plasma platelet activating factor (PAF) levels and PAF-AH activity compared to wildtype under LPS challenge. Inhibiting hydrolysis of PAF by a specific PAF-AH (PAF-acetylhydrolase) inhibitor, MAFP, could worsen LPS-induced lung inflammation in F508del mice compared to vehicle treated F508del group. Particularly, depletion of platelets in F508del mice could significantly decrease plasma lipoxin A4 and PAF-AH activity and deteriorate LPS-induced lung inflammation compared to control F508del mice. Taken together, lipoxin A4 and PAF are involved in E. coli or LPS-induced lung inflammation in CFTR-deficient mice, suggesting that lipoxin A4 and PAF might be therapeutic targets for ameliorating CFTR-deficiency deteriorated lung inflammation. PMID:24671173
Wu, Haiya; Yang, Jun; Su, Emily M; Li, Ling; Zhao, Caiqi; Yang, Xi; Gao, Zhaowei; Pan, Mengyao; Sun, Peiyu; Sun, Wei; Jiang, Yiyi; Su, Xiao
Effect of a novel leukotoriene synthesis inhibitor, BAY x1005, on the antigen- and LPS-induced airway hyperresponsiveness in guinea pigs
Due to the inhibition of 5-lipoxygenase-activating protein (FLAP), BAY x1005 is a new selective inhibitor of leukotriene synthesis. The effects of BAY x1005 on the antigen- and bacterial lipopolysaccharide (LPS)-induced airway hyperresponsiveness in guinea pigs were investigated. Six times provocation of aeroantigen caused biphasic increases in airway resistance which peaked at 1 hr (immediate phase reaction) and 4 hrs (late
Hiroichi Nagai; Hiroshi Takeda; Takashi Uno; Hiroyuki Tanaka; Akihiko Matsuo
Toona sinensis Inhibits LPS-Induced Inflammation and Migration in Vascular Smooth Muscle Cells via Suppression of Reactive Oxygen Species and NF-?B Signaling Pathway
Toona sinensis is one of the most popular vegetarian cuisines in Taiwan and it has been shown to possess antioxidant, antiangiogenic, and anticancer properties. In this study, we investigated the antiatherosclerotic potential of aqueous leaf extracts from Toona sinensis (TS; 25�100??g/mL) and its major bioactive compound, gallic acid (GA; 5??g/mL), in LPS-treated rat aortic smooth muscle (A7r5) cells. We found that pretreatment with noncytotoxic concentrations of TS and GA significantly inhibited inflammatory NO and PGE2 production by downregulating their precursors, iNOS and COX-2, respectively, in LPS-treated A7r5 cells. Furthermore, TS and GA inhibited LPS-induced intracellular ROS and their corresponding mediator, p47phox. Notably, TS and GA pretreatment significantly inhibited LPS-induced migration in transwell assays. Gelatin zymography and western blotting demonstrated that treatment with TS and GA suppressed the activity or expression of MMP-9, MMP-2, and t-PA. Additionally, TS and GA significantly inhibited LPS-induced VEGF, PDGF, and VCAM-1 expression. Further investigation revealed that the inhibition of iNOS/COX-2, MMPs, growth factors, and adhesion molecules was associated with the suppression of NF-?B activation and MAPK (ERK1/2, JNK1/2, and p38) phosphorylation. Thus, Toona sinensis may be useful for the prevention of atherosclerosis.
Yang, Hsin-Ling; Huang, Pei-Jane; Liu, Yi-Ru; Kumar, K. J. Senthil; Hsu, Li-Sung; Lu, Te-Ling; Chia, Yi-Chen; Takajo, Tokuko; Kazunori, Anzai; Hseu, You-Cheng
Genistein Suppresses LPS-Induced Inflammatory Response through Inhibiting NF-?B following AMP Kinase Activation in RAW 264.7 Macrophages
Genistein, the major isoflavone in soybean, was recently reported to exert beneficial effects in metabolic disorders and inflammatory diseases. In the present study, we investigated the effects and mechanisms of a dietary concentration of genistein on the inflammatory response in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages. Our results demonstrated that genistein effectively inhibited the LPS-induced overproduction of tumor necrosis factor-alpha (TNF-?) and interleukin 6 (IL-6), as well as LPS-induced nuclear factor kappa B (NF-?B) activation. In addition, the data also showed that genistein prevented LPS-induced decrease in adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. These effects were obviously attenuated by an AMPK inhibitor. Taken together, our results suggest that the dietary concentration of genistein is able to attenuate inflammatory responses via inhibition of NF-?B activation following AMPK stimulation. The data provide direct evidence for the potential application of low concentrations of genistein in the prevention and treatment of inflammatory diseases.
Ji, Guiyuan; Zhang, Yupei; Yang, Qinhe; Cheng, Shaobin; Hao, Jing; Zhao, Xihong; Jiang, Zhuoqin
Rhizoma Coptidis Inhibits LPS-Induced MCP-1/CCL2 Production in Murine Macrophages via an AP-1 and NF?B-Dependent Pathway
Introduction. The Chinese extract Rhizoma coptidis is well known for its anti-inflammatory, antioxidative, antiviral, and antimicrobial activity. The exact mechanisms of action are not fully understood. Methods. We examined the effect of the extract and its main compound, berberine, on LPS-induced inflammatory activity in a murine macrophage cell line. RAW 264.7 cells were stimulated with LPS and incubated with either Rhizoma coptidis extract or berberine. Activation of AP-1 and NF?B was analyzed in nuclear extracts, secretion of MCP-1/CCL2 was measured in supernatants. Results. Incubation with Rhizoma coptidis and berberine strongly inhibited LPS-induced monocyte chemoattractant protein (MCP)-1 production in RAW cells. Activation of the transcription factors AP-1 and NF?B was inhibited by Rhizoma coptidis in a dose- and time-dependent fashion. Conclusions. Rhizoma coptidis extract inhibits LPS-induced MCP-1/CCL2 production in vitro via an AP-1 and NF?B-dependent pathway. Anti-inflammatory action of the extract is mediated mainly by its alkaloid compound berberine.
Remppis, Andrew; Bea, Florian; Greten, Henry Johannes; Buttler, Annette; Wang, Hongjie; Zhou, Qianxing; Preusch, Michael R.; Enk, Ronny; Ehehalt, Robert; Katus, Hugo; Blessing, Erwin
Toona sinensis inhibits LPS-induced inflammation and migration in vascular smooth muscle cells via suppression of reactive oxygen species and NF-?B signaling pathway.
Toona sinensis is one of the most popular vegetarian cuisines in Taiwan and it has been shown to possess antioxidant, antiangiogenic, and anticancer properties. In this study, we investigated the antiatherosclerotic potential of aqueous leaf extracts from Toona sinensis (TS; 25-100 ?g/mL) and its major bioactive compound, gallic acid (GA; 5 ?g/mL), in LPS-treated rat aortic smooth muscle (A7r5) cells. We found that pretreatment with noncytotoxic concentrations of TS and GA significantly inhibited inflammatory NO and PGE2 production by downregulating their precursors, iNOS and COX-2, respectively, in LPS-treated A7r5 cells. Furthermore, TS and GA inhibited LPS-induced intracellular ROS and their corresponding mediator, p47(phox). Notably, TS and GA pretreatment significantly inhibited LPS-induced migration in transwell assays. Gelatin zymography and western blotting demonstrated that treatment with TS and GA suppressed the activity or expression of MMP-9, MMP-2, and t-PA. Additionally, TS and GA significantly inhibited LPS-induced VEGF, PDGF, and VCAM-1 expression. Further investigation revealed that the inhibition of iNOS/COX-2, MMPs, growth factors, and adhesion molecules was associated with the suppression of NF-?B activation and MAPK (ERK1/2, JNK1/2, and p38) phosphorylation. Thus, Toona sinensis may be useful for the prevention of atherosclerosis. PMID:24723997
Yang, Hsin-Ling; Huang, Pei-Jane; Liu, Yi-Ru; Kumar, K J Senthil; Hsu, Li-Sung; Lu, Te-Ling; Chia, Yi-Chen; Takajo, Tokuko; Kazunori, Anzai; Hseu, You-Cheng
Genomic Instability in Liver Cells Caused by an LPS-Induced Bystander-Like Effect
Bacterial infection has been linked to carcinogenesis, however, there is lack of knowledge of molecular mechanisms that associate infection with the development of cancer. We analyzed possible effects of the consumption of heat-killed E. coli O157:H7 cells or its cellular components, DNA, RNA, protein or lipopolysaccharides (LPS) on gene expression in na�ve liver cells. Four week old mice were provided water supplemented with whole heat-killed bacteria or bacterial components for a two week period. One group of animals was sacrificed immediately, whereas another group was allowed to consume uncontaminated tap water for an additional two weeks, and liver samples were collected, post mortem. Liver cells responded to exposure of whole heat-killed bacteria and LPS with alteration in ?H2AX levels and levels of proteins involved in proliferation, DNA methylation (MeCP2, DNMT1, DNMT3A and 3B) or DNA repair (APE1 and KU70) as well as with changes in the expression of genes involved in stress response, cell cycle control and bile acid biosynthesis. Other bacterial components analysed in this study did not lead to any significant changes in the tested molecular parameters. This study suggests that lipopolysaccharides are a major component of Gram-negative bacteria that induce molecular changes within na�ve cells of the host.
Kovalchuk, Igor; Walz, Paul; Thomas, James; Kovalchuk, Olga
Epidural analgesia with morphine or buprenorphine in ponies with lipopolysaccharide (LPS)-induced carpal synovitis
This study evaluated the analgesia effects of the epidural administration of 0.1 mg/kg bodyweight (BW) of morphine or 5 ?g/kg BW of buprenorphine in ponies with radiocarpal joint synovitis. Six ponies were submitted to 3 epidural treatments: the control group (C) received 0.15 mL/kg BW of a 0.9% sodium chloride (NaCl) solution; group M was administered 0.1 mg/kg BW of morphine; and group B was administered 5 ?g/kg BW of buprenorphine, both diluted in 0.9% NaCl to a total volume of 0.15 mL/kg BW administered epidurally at 10 s/mL. The synovitis model was induced by injecting 0.5 ng of lipopolysaccharide (LPS) in the left or right radiocarpal joint. An epidural catheter was later introduced in the lumbosacral space and advanced up to the thoracolumbar level. The treatment started 6 h after synovitis induction. Lameness, maximum angle of carpal flexion, heart rate, systolic arterial pressure, respiratory rate, temperature, and intestinal motility were evaluated before LPS injection (baseline), 6 h after LPS injection (time 0), and 0.5, 1, 2, 4, 6, 8, 10, 12, 16, 20, and 24 h after treatments. Although the model of synovitis produced clear clinical signs of inflammation, the lameness scores in group C were different from the baseline for only up to 12 h. Both morphine and buprenorphine showed a reduction in the degree of lameness starting at 0.5 and 6 h, respectively. Reduced intestinal motility was observed at 0.5 h in group M and at 0.5 to 1 h in group B. Epidural morphine was a more effective analgesic that lasted for more than 12 h and without side effects. It was concluded that morphine would be a valuable analgesic option to alleviate joint pain in the thoracic limbs in ponies.
Freitas, Gabrielle C.; Carregaro, Adriano B.; Gehrcke, Martielo I.; De La Corte, Flavio D.; Lara, Valeria M.; Pozzobon, Ricardo; Brass, Karin E.
Cloning and analysis of gene regulation of a novel LPS-inducible cDNA
The expression of many genes is altered upon the activation of macrophages by bacterial LPS. These genes play a crucial role in the orchestration of various responses to protect the host against infection. A novel 2.3 kilobase (kb) cDNA, designated IRG1, was obtained from a cDNA library prepared with RNA isolated from RAW 264.7 following lipopolysaccharide stimulation. Sequence analysis of the clone revealed no identity to any known genes but showed the presence of many potential phosphorylation sites suggesting that IRG1 protein product may be regulated at this level. Furthermore, IRG1 contains the motif for glycosaminoglycan attachment site, implying that IRG1 may be a proteoglycan. By interspecific backcross analysis, IRG1 was mapped to mouse chromosome 14 linked to Tyrp2 and Rap2a. The IRG1 message appears 1.5 h following LPS exposure and its induction was not dependent on new protein synthesis. In fact, cycloheximide induced the expression of IRG1, suggesting that a protein repressor prevents the expression of IRG1 when uninduced. The role of the protein kinase A pathway in regulating the induction of IRG1 by LPS is questionable, because although forskolin inhibited its induction, neither dibutyrl-cAMP nor 8-(4-chlorophenylthio)-cAMP had much effect on its expression. In contrast, activation of protein kinase C potentiated the LPS response. Chelation of extracellular calcium inhibited IRG1 4 h after LPS induction, while increasing intracellular calcium had little effect on the levels of the IRG1 transcript. Inhibiting tyrosine phosphorylation abrogated the induction of IRG1 by LPS. Hence, the induction of IRG1 by LPS is mediated by tyrosine kinase and protein kinase C pathway. 80 refs., 5 figs.
Lee, C.G.L.; O`Brien, W.E. [Baylor College of Medicine, Houston, TX (United States); Jenkins, N.A.; Gilbert, D.J.; Copeland, N.G. [Frederick Cancer Research and Development Center, MD (United States)
LPS-Induced Galectin-3 Oligomerization Results in Enhancement of Neutrophil Activation
Galectin-3 (Gal 3) is a glycan-binding protein that can be secreted by activated macrophages and mast cells at inflammation sites and plays an important role in inflammatory diseases caused by Bacteria and their products, such as lipopolysaccharides (LPS). Although it is well established that Gal 3 can interact with LPS, the pathophysiological importance of LPS/Gal 3 interactions is not fully understood. Data presented herein demonstrate for the first time that the interaction of Gal 3, either via its carbohydrate binding C-terminal domain or via its N-terminal part, with LPS from different bacterial strains, enhances the LPS-mediated neutrophil activation in vitro. Gal 3 allowed low LPS concentrations (1 �g/mL without serum, 1 ng/mL with serum) to upregulate CD11b expression and reactive oxygen species (ROS) generation on human neutrophils in vitro and drastically enhanced the binding efficiency of LPS to the neutrophil surface. These effects required LPS preincubation with Gal 3, before neutrophil stimulation and involved specific Gal 3/LPS interaction. A C-terminal Gal-3 fragment, which retains the lectin domain but lacks the N-terminal part, was still able to bind both to Escherichia coli LPS and to neutrophils, but had lost the ability to enhance neutrophil response to LPS. This result emphasizes the importance of an N-terminus-mediated Gal 3 oligomerization induced by its interaction with LPS. Finally we demonstrated that Balb/C mice were more susceptible to LPS-mediated shock when LPS was pretreated with Gal 3. Altogether, these results suggest that multimeric interactions between Gal 3 oligomers and LPS potentiate its pro-inflammatory effects on neutrophils.
Fermino, Marise Lopes; Polli, Claudia Danella; Toledo, Karina Alves; Liu, Fu-Tong; Hsu, Dan K.; Roque-Barreira, Maria Cristina; Pereira-da-Silva, Gabriela
Distinct LPS-induced signals regulate LPS uptake and morphological changes in medfly hemocytes.
Recently we demonstrated that lipopolysaccharide (LPS) promotes activation of the Ras/ERK cascade in medfly hemocytes and that phagocytosis of Escherichia coli by insect hemocytes is mediated by an integrin-dependent process via the activation of FAK/Src complex (J Biol Chem 273 (1998) 14813; FEBS Letters 496 (2001) 55). In the current study we wanted to further elucidate the effects of LPS on medfly hemocytes, in order to better understand the regulation of the evolutionary conserved signaling mechanisms between insects and mammals. We initially observed that different stimuli, including LPS, E. coli, RGD, fibronectin and heat shock activate hemocyte ERK. The response of hemocytes to these stimuli denoted that hemocyte ERK is evidently stimulated by at least an LPS receptor and via an integrin-mediated process. The medfly hemocytes respond to LPS by changing their morphology, inducing the activation of several signaling pathways, including Ras/MEK/ERK, PI-3K/ERK and Rho pathways and contributing to LPS uptake. Experiments based on inhibitors of specific signaling pathways, such as manumycin A, toxin A, U0126, PD98059 and wortmannin revealed that Ras, MEK and PI-3K are involved in the activation of ERK. Whether PI-3K is an intermediate of Ras/MEK/ERK pathway or activates ERK via other signaling pathway it remains to be elucidated. ERK is not activated via Rho pathway, denoting that Rho may not be an upstream effector molecule of ERK pathway. Regarding the role(s) that these kinases play in hemocytes, it can be suggested that PI-3K and Rho GTPases can modulate hemocyte shape changes, whereas ERK, Ras and MEK cannot. In addition, PI-3K as well as Ras and MEK through ERK activation participate in LPS endocytosis. Therefore, PI-3K shares a dual role; it is involved both in cell shape changes and in LPS endocytosis. Since ERK activation appears to be independent of the integrity of actin filaments, as cytochalasin D and latrunculin A did not block ERK activation, it can be concluded that LPS endocytosis is independent of actin cytoskeleton remodeling as is the case in mammalian systems. PMID:14563359
Soldatos, Anastasios N; Metheniti, Aristea; Mamali, Irene; Lambropoulou, Maria; Marmaras, Vassilis J
Pulmonary Permeability Assessed by Fluorescent-Labeled Dextran Instilled Intranasally into Mice with LPS-Induced Acute Lung Injury
Background Several different methods have been used to assess pulmonary permeability in response to acute lung injury (ALI). However, these methods often involve complicated procedures and algorithms that are difficult to precisely control. The purpose of the current study is to establish a feasible method to evaluate alterations in lung permeability by instilling fluorescently labeled dextran (FITC-Dextran) intranasally. Methods/Principal Findings For the mouse model of direct ALI, lipopolysaccharide (LPS) was administered intranasally. FITC-Dextran was instilled intranasally one hour before the mice were euthanized. Plasma fluorescence intensities from the LPS group were significantly higher than in the control group. To determine the reliability and reproducibility of the procedure, we also measured the lung wet-to-dry weight ratio, the protein concentration of the bronchoalveolar lavage fluid, tight and adherens junction markers and pathological changes. Consistent results were observed when the LPS group was compared with the control group. Simultaneously, we found that the concentration of plasma FITC-Dextran was LPS dose-dependent. The concentration of plasma FITC-Dextran also increased with initial intranasal FITC-Dextran doses. Furthermore, increased fluorescence intensity of plasma FITC-Dextran was found in the intraperitoneally LPS-induced ALI model. Conclusion/Significance In conclusion, the measurement of FITC-Dextran in plasma after intranasal instillation is a simple, reliable, and reproducible method to evaluate lung permeability alterations in vivo. The concentration of FITC-Dextran in the plasma may be useful as a potential peripheral biomarker of ALI in experimental clinical studies.
Chen, Honglei; Wu, Shaoping; Lu, Rong; Zhang, Yong-guo; Zheng, Yuanyuan; Sun, Jun
CD11c+ Alveolar Macrophages are a Source of IL-23 during LPS-induced Acute Lung Injury
Acute lung injury (ALI) is a severe pulmonary disease causing high numbers of fatalities worldwide. Innate immune responses are an integral part of the pathophysiologic events during ALI. Interleukin-23 (IL-23) is a proinflammatory mediator known to direct the inflammatory responses in various settings of infection, autoimmunity and cancer. IL-23 has been associated with proliferation and effector functions in Th17 cells. Surprisingly, little is known about production of IL-23 during ALI. In this study we found expression of mRNA for IL-23p19 to be 10-fold elevated in lung homogenates of C57BL/6 mice after lipopolysaccharide (LPS)-induced ALI. Likewise, concentrations of IL-23, protein significantly increased in bronchoalveolar lavage fluids. Experiments with IL-23 deficient mice showed that endogenous IL-23 was required for production of IL-17A during LPS-ALI. CD11c-diphtheria toxin receptor transgenic mice were used to selectively deplete CD11c+ cells, the data suggesting that IL-23 production is dependent at least in part on CD11c+ cells during ALI. No alterations of IL-23 levels were observed in Rag-1 deficient mice as compared to wild type C57BL/6 mice following ALI. The mouse alveolar macrophage cell line, MH-S, as well as primary alveolar macrophages displayed abundant surface expression of CD11c. Activation of these macrophages by LPS resulted in release of IL-23 in vitro. Our findings identify CD11c+ macrophages in the lung are likely an important source of IL-23 during ALI, which may be helpful for better understanding of this disease.
Bosmann, Markus; Grailer, Jamison J.; Russkamp, Norman F.; Ruemmler, Robert; Zetoune, Firas S.; Sarma, J. Vidya; Ward, Peter A.
Polygonum cuspidatum, compared with baicalin and berberine, inhibits inducible nitric oxide synthase and cyclooxygenase-2 gene expressions in RAW 264.7 macrophages
Polygonum cuspidatum water extract (PCWE) was shown to be a potent inhibitor of lipopolysaccharide (LPS)-induced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). PCWE was compared to baicalin isolated from Scutellaria baicalensis Georgi and berberine of Coptidis rhizoma and Phellodendri cortex, for their effects on LPS-induced nitric oxide (NO) production and iNOS and COX-2 gene expressions in RAW
Kyung-Woon Kim; Ki-Tai Ha; Cheol-Soo Park; Un-Ho Jin; Hyen Wook Chang; In-Seon Lee; Cheorl-Ho Kim
IL1 ?- and LPS-induced serotonin secretion is increased in EC cells derived from Crohn's disease
Gut mucosal enterochromaffin (EC) cells are regarded as key regulators of intestinal motility and fluid secretion via secretion of serotonin (5HT), are increased in numbers in mucosal inflammation and located in close proximity to immune cells. We examined whether interleukin (IL)1? and Escherichia coli lipopolysaccharide (LPS) induced EC cell 5HT release through Toll-like/IL-1 (TIL) receptor activation, nuclear factor kappa B (NF?B) and mitogen-activated protein kinase (MAPK) phosphorylation and evaluated whether somatostatin could inhibit this phenomenon. Pure (>98%) human intestinal EC cells were isolated by fluorescent activated cell sorting from preparations of normal (n = 5) and Crohn's colitis (n = 6) mucosa. 5HT release was measured (ELISA), and NF?B and ERK phosphorylation quantitated (ELISA) in response to IL1? and LPS. 5HT secretion was increased by both E. coli LPS (EC50 = 5 ng mL?1) and IL1? (EC50 = 0.05 pmol L?1) >2-fold (P < 0.05) in Crohn's EC cells compared with normal EC cells. Secretion was reversible by the TLR4 antagonist, E. coli K12 LPS (IC50 = 12 ng mL?1) and the IL1? receptor antagonist (ILRA; IC50 = 3.4 ng mL?1). IL1? caused significant (P < 0.05) NF?B and MAPK phosphorylation (40�55%). The somatostatin analogue, lanreotide inhibited IL1b-stimulated secretion in Crohn's (IC50 = 0.61 nmol L?1) and normal EC cells (IC50 = 1.8 nmol L?1). Inter-leukins (IL1?) and bacterial products (E. coli LPS) stimulated 5HT secretion from Crohn's EC cells via TIL receptor activation (TLR4 and IL1?). Immune-mediated alterations in EC cell secretion of 5HT may represent a component of the pathogenesis of abnormal bowel function in Crohn's disease. Inhibition of EC cell-mediated 5HT secretion may be an alternative therapeutic strategy in the amelioration of inflammatory bowel disease symptomatology.
Baicalein inhibits nuclear factor-?B and apoptosis via c-FLIP and MAPK in d-GalN\\/LPS induced acute liver failure in murine models
The hepatoprotective effects and molecular mechanisms of baicalein on acute liver failure induced by d-galactosamine (d-GalN)\\/lipopolysaccharides (LPS) were investigated in vivo. Mice were administered with different doses of baicalein (50, 100 or 150mg\\/kg, p.o.) 1h before injection of d-GalN (700mg\\/kg)\\/LPS (10?g\\/kg) and then sacrificed 6h after treatment with d-GalN\\/LPS. Pretreatment with baicalein prevented d-GalN\\/LPS-induced liver damage by preventing associated increases
Yan-Ling Wu; Li-Hua Lian; Ying Wan; Ji-Xing Nan
Phillyrin attenuates LPS-induced pulmonary inflammation via suppression of MAPK and NF-?B activation in acute lung injury mice.
Phillyrin (Phil) is one of the main chemical constituents of Forsythia suspensa (Thunb.), which has shown to be an important traditional Chinese medicine. We tested the hypothesis that Phil modulates pulmonary inflammation in an ALI model induced by LPS. Male BALB/c mice were pretreated with or without Phil before respiratory administration with LPS, and pretreated with dexamethasone as a control. Cytokine release (TNF-?, IL-1?, and IL-6) and amounts of inflammatory cell in bronchoalveolar lavage fluid (BALF) were detected by ELISA and cell counting separately. Pathologic changes, including neutrophil infiltration, interstitial edema, hemorrhage, hyaline membrane formation, necrosis, and congestion during acute lung injury in mice were evaluated via pathological section with HE staining. To further investigate the mechanism of Phil anti-inflammatory effects, activation of MAPK and NF-?B pathways was tested by western blot assay. Phil pretreatment significantly attenuated LPS-induced pulmonary histopathologic changes, alveolar hemorrhage, and neutrophil infiltration. The lung wet-to-dry weight ratios, as the index of pulmonary edema, were markedly decreased by Phil pretreatment. In addition, Phil decreased the production of the proinflammatory cytokines including (TNF-?, IL-1?, and IL-6) and the concentration of myeloperoxidase (MPO) in lung tissues. Phil pretreatment also significantly suppressed LPS-induced activation of MAPK and NF-?B pathways in lung tissues. Taken together, the results suggest that Phil may have a protective effect on LPS-induced ALI, and it potentially contributes to the suppression of the activation of MAPK and NF-?B pathways. Phil may be a new preventive agent of ALI in the clinical setting. PMID:23751215
Zhong, Wei-ting; Wu, Yi-chun; Xie, Xian-xing; Zhou, Xuan; Wei, Miao-miao; Soromou, Lanan-Wassy; Ci, Xin-xin; Wang, Da-cheng
The monoacylglycerol lipase inhibitor JZL184 attenuates LPS-induced increases in cytokine expression in the rat frontal cortex and plasma: differential mechanisms of action
Background and purpose JZL184 is a selective inhibitor of monoacylglycerol lipase (MAGL), the enzyme that preferentially catabolizes the endocannabinoid 2-arachidonoyl glycerol (2-AG). Here, we have studied the effects of JZL184 on inflammatory cytokines in the brain and plasma following an acute immune challenge and the underlying receptor and molecular mechanisms involved. Experimental approach JZL184 and/or the CB1 receptor antagonist, AM251 or the CB2receptor antagonist, AM630 were administered to rats 30 min before lipopolysaccharide (LPS). 2 h later cytokine expression and levels, MAGL activity, 2-AG, arachidonic acid and prostaglandin levels were measured in the frontal cortex, plasma and spleen. Key results JZL184 attenuated LPS-induced increases in IL-1?, IL-6, TNF-? and IL-10 but not the expression of the inhibitor of NFkB (I?B?) in rat frontal cortex. AM251 attenuated JZL184-induced decreases in frontal cortical IL-1? expression. Although arachidonic acid levels in the frontal cortex were reduced in JZL184-treated rats, MAGL activity, 2-AG, PGE2 and PGD2 were unchanged. In comparison, MAGL activity was inhibited and 2-AG levels enhanced in the spleen following JZL184. In plasma, LPS-induced increases in TNF-? and IL-10 levels were attenuated by JZL184, an effect partially blocked by AM251. In addition, AM630 blocked LPS-induced increases in plasma IL-1? in the presence, but not absence, of JZL184. Conclusion and implications Inhibition of peripheral MAGL in rats by JZL184 suppressed LPS-induced circulating cytokines that in turn may modulate central cytokine expression. The data provide further evidence for the endocannabinoid system as a therapeutic target in treatment of central and peripheral inflammatory disorders. Linked Articles This article is part of a themed section on Cannabinoids. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2013.169.issue-4 & http://dx.doi.org/10.1111/bph.2012.167.issue-8
Kerr, DM; Harhen, B; Okine, BN; Egan, LJ; Finn, DP; Roche, M
Effect of Rabbit Epididymal Antimicrobial Peptide, REHb?P, on LPS-Induced Proinflammatory Cytokine Responses in Human Vaginal Cells In Vitro
Antimicrobial peptides (AMP's) protect epithelial surfaces including epididymis against pathogens and play a key role in orchestrating various defensive responses. Recently, we have identified one such AMP, rabbit epididymal hemoglobin-? subuit (REHb?P) from the epididymal fluid of rabbit, Oryctologus cuniculus. The demonstration of a protective role of REHb?P in epididymal epithelial cells (EPEC's) led us to investigate: (1) the identification of LPS interactive domain in REHb?P, and (2) whether the REHb?P of rabbit origin mediates vaginal cellular immune responses of another species (human). HeLa-S3, human vaginal epithelial cells (hVECs) were exposed to LPS or the LPS-stimulated cells treated with REHb?P or neutral peptide, nREHb?P. Effect of LPS and cytokines (IL-6 and IL-1?) and chemokines (IL-8, MCP-1) levels was determined in the culture supernatants. In response to the LPS, hVECs synthesized these mediators and the levels were significantly higher than controls. This enhancing effect was ameliorated when the LPS-induced hVECs were treated with REHb?P. Similar results were obtained on NF-?B protein and hBD-1 mRNA expression. Confocal microscopy studies revealed that REHb?P attenuated the LPS-induced internalization of E. coli by macrophages. The chemotaxis studies performed using Boyden chamber Transwell assay, which showed elevated migration of U937 cells when the supernatants of LPS-induced hVECs were used, and the effect was inhibited by REHb?P. REHb?P was found to be localized on the acrosome of rabbit spermatozoa, suggesting its role in sperm protection beside sperm function. In conclusion, REHb?P may have the potential to develop as a therapeutic agent for reproductive tract infections (RTI's).
Reddy, K. V. R.; Sukanya, D.; Patgaonkar, M. S.; Selvaakumar, C.
Etazolate abrogates the lipopolysaccharide (LPS)-induced downregulation of the cAMP/pCREB/BDNF signaling, neuroinflammatory response and depressive-like behavior in mice.
Increasing evidence has indicated that immune challenge by bacterial lipopolysaccharide (LPS) induces depressive-like behavior, neuroinflammatory response and upregulates phosphodiesterase-4 (PDE4), an enzyme that specifically hydrolyzes cyclic adenosine monophosphate (cAMP). However, whether the potential PDE4 inhibitor etazolate prevents the LPS-induced depressive-like behavior remains unclear. Here using a model of depression induced by the repeated administration of LPS during 16days, and then investigated the influence of LPS on the expression of PDE4, interleukin-1? (IL-1?) and antidepressant action of etazolate in mice through forced swimming, novelty suppressed feeding, sucrose preference and open-field tests. Our results showed that etazolate pretreatment facilitated the recovery from weight loss and prevented the depressive-like behavior induced by repeated LPS administration. Moreover, the antidepressant action of etazolate was paralleled by significantly reducing the expression levels of PDE4A, PDE4B, PDE4D and IL-1? and up-regulating the cAMP/phosphorylated cAMP response-element binding protein (pCREB)/brain-derived neurotrophic factor (BDNF) signaling in the hippocampus and prefrontal cortex of mice. These results indicate that the effects of etazolate on the depressive-like behavior induced by repeated LPS treatment may partially depend on the inhibition of PDE4 subtypes, the activation of the cAMP/pCREB/BDNF signaling and the anti-inflammatory responses in the hippocampus and prefrontal cortex. PMID:24434771
Guo, J; Lin, P; Zhao, X; Zhang, J; Wei, X; Wang, Q; Wang, C
Aqueous Extract of Gracilaria tenuistipitata Suppresses LPS-Induced NF-?B and MAPK Activation in RAW 264.7 and Rat Peritoneal Macrophages and Exerts Hepatoprotective Effects on Carbon Tetrachloride-Treated Rat
In addition to the previous investigations of bioactivity of aqueous extract of the edible Gracilaria tenuistipitata (AEGT) against H2O2-induced DNA damage and hepatitis C virus replication, the purpose of this study is to evaluate the potential therapeutic properties of AEGT against inflammation and hepatotoxicity using lipopolysaccharide (LPS)-stimulated mouse RAW 264.7 cells, primary rat peritoneal macrophages and carbon tetrachloride (CCl4)-induced acute hepatitis model in rats. AEGT concentration-dependently inhibited the elevated RNA and protein levels of inducible nitric oxide synthase and cyclooxygenase-2, thereby reducing nitric oxide and prostaglandin E2 levels, respectively. Moreover, AEGT significantly suppressed the production of LPS-induced proinflammatory cytokines, including interleukin (IL)-1?, IL-6 and tumor necrosis factor-?. These inhibitory effects were associated with the suppression of nuclear factor-kappa B activation and mitogen-activated protein kinase phosphorylation by AEGT in LPS-stimulated cells. In addition, we highlighted the hepatoprotective and curative effects of AEGT in a rat model of CCl4-intoxicated acute liver injury, which was evident from reduction in the elevated serum aspartate aminotransferase and alanine aminotransferase levels as well as amelioration of histological damage by pre-treatment or post-treatment of AEGT. In conclusion, the results demonstrate that AEGT may serve as a potential supplement in the prevention or amelioration of inflammatory diseases.
Tseng, Chin-Kai; Lin, Chun-Kuang; Chang, Hsueh-Wei; Wu, Yu-Hsuan; Yen, Feng-Lin; Chang, Fang-Rong; Chen, Wei-Chun; Yeh, Chi-Chen; Lee, Jin-Ching
Aqueous extract of Gracilaria tenuistipitata suppresses LPS-induced NF-?B and MAPK activation in RAW 264.7 and rat peritoneal macrophages and exerts hepatoprotective effects on carbon tetrachloride-treated rat.
In addition to the previous investigations of bioactivity of aqueous extract of the edible Gracilaria tenuistipitata (AEGT) against H2O2-induced DNA damage and hepatitis C virus replication, the purpose of this study is to evaluate the potential therapeutic properties of AEGT against inflammation and hepatotoxicity using lipopolysaccharide (LPS)-stimulated mouse RAW 264.7 cells, primary rat peritoneal macrophages and carbon tetrachloride (CCl4)-induced acute hepatitis model in rats. AEGT concentration-dependently inhibited the elevated RNA and protein levels of inducible nitric oxide synthase and cyclooxygenase-2, thereby reducing nitric oxide and prostaglandin E2 levels, respectively. Moreover, AEGT significantly suppressed the production of LPS-induced proinflammatory cytokines, including interleukin (IL)-1?, IL-6 and tumor necrosis factor-?. These inhibitory effects were associated with the suppression of nuclear factor-kappa B activation and mitogen-activated protein kinase phosphorylation by AEGT in LPS-stimulated cells. In addition, we highlighted the hepatoprotective and curative effects of AEGT in a rat model of CCl4-intoxicated acute liver injury, which was evident from reduction in the elevated serum aspartate aminotransferase and alanine aminotransferase levels as well as amelioration of histological damage by pre-treatment or post-treatment of AEGT. In conclusion, the results demonstrate that AEGT may serve as a potential supplement in the prevention or amelioration of inflammatory diseases. PMID:24475143
Tseng, Chin-Kai; Lin, Chun-Kuang; Chang, Hsueh-Wei; Wu, Yu-Hsuan; Yen, Feng-Lin; Chang, Fang-Rong; Chen, Wei-Chun; Yeh, Chi-Chen; Lee, Jin-Ching
Modulation of B lymphocyte differentiation by calcitonin gene-related peptide (CGRP). II. Inhibition of LPS-induced kappa light chain expression by CGRP.
The presence of calcitonin gene-related peptide (CGRP) in nerve endings in lymphoid organs, around blood vessels, and in bone marrow suggests that it might influence the function and differentiation of lymphoid cells. Previous studies identified specific CGRP receptors on mature T and B lymphocytes and on 70Z/3 pre-B cells. In these studies, it was found that CGRP stimulated a rapid, prolonged elevation of cAMP with in 70Z/3 cells with an ED50 of approximately 20 fM. Following CGRP treatment, cAMP levels peaked within 5 min and were still elevated after 60 min. The effect of CGRP on surface immunoglobulin (sIg) expression was examined by treating 70Z/3 cells with CGRP, or combinations of CGRP and LPS, and then measuring sIg expression by FACS. When 70Z/3 cells were treated with LPS, CGRP, or calcitonin for 48 hr, only LPS induced the expression of sIg, increasing the percentage of cells expressing sIg from less than 10% positive in untreated cells to 80-98% positive. Subsequent experiments examined the effect of CGRP on LPS-induced sIg. CGRP inhibited LPS-induced sIg expression at concentrations ranging from 10(-15) to 10(-7) M. The maximal inhibition was observed at CGRP concentrations ranging from 10(-10) to 10(-8) M, and the maximal reduction of sIg expression was about 40%. The inhibitory effect of CGRP was specific in that it could not be mimicked by calcitonin and could be blocked by the CGRP receptor antagonist CGRP8-37. A similar dose-dependent inhibitory effect on LPS induction was observed in 70Z/3 cells treated with LPS and dibutyryl cAMP, suggesting that the inhibitory effect of CGRP on sIg expression is mediated by stimulation of intracellular cAMP. The inhibitory effect of CGRP on LPS induction of sIg appears to be mediated by a reduction in the expression of both mu and kappa mRNA. When mu and kappa expression were examined by Northern blot analysis, it was found that CGRP caused a 50% reduction in the amount of mu and kappa mRNA induced by LPS. The ability of CGRP to inhibit differentiation of 70Z/3 cells and sIg expression provides evidence that CGRP can influence the differentiation of lymphopoietic precursors. PMID:8396499
McGillis, J P; Humphreys, S; Rangnekar, V; Ciallella, J
Sulfated derivatives of 20(S)-ginsenoside Rh2 and their inhibitory effects on LPS-induced inflammatory cytokines and mediators.
Ginsenoside Rh2 is one of the most important ginsenosides in ginseng with antitumor, antidiabetic, antiallergic, and anti-inflammatory effects. However, the extremely poor oral bioavailability induced by its low water solubility greatly limits the potency of Rh2 in clinical use. Therefore, in this study we sulfated 20(S)-ginsenoside Rh2 with chlorosulfonic acid and pyridine method, and got two new sulfated derivatives, Rh2-B1 and Rh2-B2, with higher water solubility. Their chemical structures were characterized by spectroscopic methods (IR, MS and NMR). Additionally, Rh2-B1 and Rh2-B2 had the greater anti-inflammatory effects than Rh2 through inhibiting inflammatory cytokines and mediators in LPS-induced mouse RAW264.7 macrophages cells. These results suggested that the sulfated modification of Rh2 improved its water solubility and the sulfated derivatives could be more potential candidates for developing as anti-inflammatory agents. PMID:23266729
Fu, Ben-Dong; Bi, Wen-Yan; He, Chang-Liang; Zhu, Wei; Shen, Hai-Qing; Yi, Peng-Fei; Wang, Lu; Wang, Da-Cheng; Wei, Xu-Bin
Calcineurin Inactivation Leads to Decreased Responsiveness to LPS in Macrophages and Dendritic Cells and Protects Against LPS-Induced Toxicity In Vivo
Microbial components such as lipopolysaccharide (LPS) bind to Toll-like receptors (TLR) and activate innate and inflammatory responses. Responses to LPS and other microbial components are limited by the activation of negative feedback mechanisms that reduce responsiveness to subsequent LPS exposure, often termed LPS tolerance. Our laboratory has previously shown that calcineurin, a phosphatase known for its activation of T cells via NFAT, negatively regulates the TLR pathway in macrophages; consequently, calcineurin inhibitors (FK506 and Cyclosporin A) mimic TLR ligands in activating the TLR pathway, NF-?B, and associated innate and inflammatory responses. This study investigated the physiological consequences of calcineurin inactivation for LPS-induced inflammatory responses in vitro and in vivo using two models: calcineurin inhibition by FK506 (tacrolimus) and myeloid cell-specific calcineurin deletion. Activation of dendritic cells and macrophages with FK506 in vitro was shown to induce a state of reduced responsiveness to LPS (i.e., a form of LPS tolerance). Similarly, macrophages from FK506-treated mice or from mice in which the calcineurin B1 (CnB1) subunit was conditionally knocked out in myeloid cells were found to have diminished LPS-induced inflammatory responses. In addition, mice with CnB1-deficient myeloid cells and mice undergoing FK506 treatment showed improved survival and recovery when challenged with high doses of systemic LPS compared to controls. These results demonstrate that inactivation of calcineurin in macrophages and other myeloid cells by inhibition or deletion can induce a form of LPS tolerance and protect the host from LPS toxicity in vivo.
Jennings, Charay; Kusler, Brenda; Jones, Patricia P.
Caffeic acid regulates LPS-induced NF-?B activation through NIK/IKK and c-Src/ERK signaling pathways in endothelial cells.
The redox sensitive, proinflammatory nuclear transcription factor NF-?B plays a key role in inflammation. In a redox state disrupted by oxidative stress, pro-inflammatory genes are upregulated by the activation of NF-?B via diverse kinases. Thus, the search and characterization of new substances that modulate NF-?B are topics of considerable research interest. Caffeic acid is a component of garlic, some fruits, and coffee, and is widely used as a phenolic agent in beverages. In the present study, caffeic acid was examined with respect to the modulation of inflammatory NF-?B activation via the redox-related c-Src/ERK and NIK/IKK pathways via the reduction of oxidative stress. YPEN-1 cells (an endothelial cell line) were used to explore the molecular mechanism underlying the anti-inflammatory effect of caffeic acid by examining its modulation of NF-?B signaling pathway by LPS. Our results show that LPS-induced oxidative stress-related NF-?B activation upregulated pro-inflammatory COX-2, NF-?B targeting gene which were all inhibited effectively by caffeic acid. Our study shows that caffeic acid inhibits the activation of NF-?B via the c-Src/ERK and NIK/IKK signal transduction pathways. Our results indicate that antioxidative effect of caffeic acid and its restoration of redox balance are responsible for its anti-inflammatory action. Thus, the study provides new information regarding the anti-inflammatory properties of caffeic acid and the roles in the regulation of LPS-induced oxidative stress induces alterations in signal transduction pathways. PMID:23888332
Kim, So Ra; Jung, Yu Ri; Kim, Dae Hyun; An, Hye Jin; Kim, Mi Kyung; Kim, Nam Deuk; Chung, Hae Young
Prepared and screened a modified TNF-alpha molecule as TNF-alpha autovaccine to treat LPS induced endotoxic shock and TNF-alpha induced cachexia in mouse.
Overexpression of TNF-alpha in the body is critically involved in many diseases. A strategy to construct TNF-alpha autovaccine by introducing a T cell helper epitope to the protein has been developed and may be an alternative because it is cheaper and highly efficient. However, the induction of high level anti-TNF-alpha neutralizing autoantibodies by TNF-alpha autovaccine is depend on a proper T cell help epitope. In order to evaluate the effect of different T helper cell epitopes on the immunogenicity of mouse TNF-alpha (mTNF-alpha), three T helper cell epitopes, TT (QYIKANSKFIGITEL), HEL (NTDGSTDYGILQINSR), and PADRE (AKFVAAWTLKA), were chosen for this study. The sequence (amino acids 126-140) of mTNF-alpha was replaced with those of the T cell help epitopes, respectively. The three fusion proteins (mTNF-TT, mTNF-HEL, mTNF-PADRE) were expressed in Escherichia coli and purified with a simple strategy. The abilities of the proteins elicited TNF-alpha autoantibodies in BALB/c mice were investigated. The results showed that mTNF-PADRE is the most effective among the three modified TNF-alpha molecules. In the absence of adjuvant, the therapeutic effect of TNF-PADRE on LPS induced endotoxic shock mice and mTNF-alpha induced cachexia mice was observed. This study suggests that mTNF-PADRE may be a better candidate of mTNF-alpha autovaccine. PMID:17592730
Wan, Yi; Xue, Xiaochang; Li, Meng; Zhang, Xiaoyong; Qin, Xin; Zhang, Cun; You, Yanjie; Wang, Weihua; Jiang, Changli; Wu, Shouzhen; Liu, Yan; Zhu, Wenhua; Ran, Yonggang; Zhang, Zhen; Han, Wei; Zhang, Yingqi
Hericium erinaceus suppresses LPS-induced pro-inflammation gene activation in RAW264.7 macrophages.
The aim of this study was to investigate the anti-inflammatory properties of each fraction of Hericium erinaceus (HE). The ethanol extract from HE was partitioned with different solvents in the order of increasing polarity. The treatment with 10-100 ?g/mL of each fraction did not reduce RAW 264.7 cell viability except ethyl acetate fraction. Among the various extracts, the chloroform fraction showed the most potent activity against nitric oxide (NO), prostaglandin E(2) (PGE(2)) and reactive oxygen species (ROS). The western blotting and reverse transcriptase polymerase chain reaction (RT-PCR) analyses revealed that chloroform fraction from HE (CHE) significantly reduced the protein level of iNOS and cyclooxygenase-2 (COX-2) or mRNA levels of iNOS in lipopolysaccharide-induced macrophages. Furthermore, CHE inhibited the translocation of nuclear factor (NF)-?B p65 subunit, phsophorylation of I-?B, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK) in a dose-dependent manner. Furthermore, the activation of both activator protein-1 (AP-1) and NF ?B in the nucleus were abrogated by CHE with luciferase assay. In conclusion, these results indicate that CHE may provide an anti-inflammatory effect by attenuating the generation of excessive NO, PGE(2), and ROS and by suppressing the expression of pro-inflammatory genes through the inhibition of NF-?B and JNK activity. PMID:22126451
Kim, Young-Ock; Lee, Sang-Won; Oh, Chung-Hun; Rhee, Yun-Hee
Memantine protects against LPS-induced neuroinflammation, restores behaviorally-induced gene expression and spatial learning in the rat
Neuroinflammation is reliably associated with the pathogenesis of a number of neurodegenerative diseases, and can be detected by the presence of activated microglia. Neuroinflammation can be induced by chronic lipopolysaccharide (LPS) infusion into the 4th ventricle of the rat resulting in region-selective microglia activation and impaired hippocampal-dependent memory. Furthermore, this treatment results in altered behaviorally-induced expression of the immediate early
S. Rosi; A. Vazdarjanova; V. Ramirez-Amaya; P. F. Worley; G. L. Wenk
Lipopolysaccharide-induced Diaphragmatic Contractile Dysfunction in Mice Lacking the Inducible Nitric Oxide Synthase
The goal of this study was to evaluate the importance of the inducible nitric oxide synthase (iNOS) in lipopolysaccharide (LPS)-induced diaphragmatic contractile dysfunction. Many investigators have proposed that iNOS induction in the ventilatory and limb muscles of animals injected with Escherichia coli LPS leads to impaired muscle contractility and increased fatigability. We tested this proposal by examining wild-type mice and
The fatty acid amide hydrolase (FAAH) inhibitor PF-3845 acts in the nervous system to reverse LPS-induced tactile allodynia in mice
BACKGROUND AND PURPOSE Inflammatory pain presents a problem of clinical relevance and often elicits allodynia, a condition in which non-noxious stimuli are perceived as painful. One potential target to treat inflammatory pain is the endogenous cannabinoid (endocannabinoid) system, which is comprised of CB1 and CB2 cannabinoid receptors and several endogenous ligands, including anandamide (AEA). Blockade of the catabolic enzyme fatty acid amide hydrolase (FAAH) elevates AEA levels and elicits antinociceptive effects, without the psychomimetic side effects associated with ?9-tetrahydrocannabinol (THC). EXPERIMENTAL APPROACH Allodynia was induced by intraplantar injection of LPS. Complementary genetic and pharmacological approaches were used to determine the strategy of blocking FAAH to reverse LPS-induced allodynia. Endocannabinoid levels were quantified using mass spectroscopy analyses. KEY RESULTS FAAH (?/?) mice or wild-type mice treated with FAAH inhibitors (URB597, OL-135 and PF-3845) displayed an anti-allodynic phenotype. Furthermore, i.p. PF-3845 increased AEA levels in the brain and spinal cord. Additionally, intraplantar PF-3845 produced a partial reduction in allodynia. However, the anti-allodynic phenotype was absent in mice expressing FAAH exclusively in the nervous system under a neural specific enolase promoter, implicating the involvement of neuronal fatty acid amides (FAAs). The anti-allodynic effects of FAAH-compromised mice required activation of both CB1 and CB2 receptors, but other potential targets of FAA substrates (i.e. �-opioid, TRPV1 and PPAR? receptors) had no apparent role. CONCLUSIONS AND IMPLICATIONS AEA is the primary FAAH substrate reducing LPS-induced tactile allodynia. Blockade of neuronal FAAH reverses allodynia through the activation of both cannabinoid receptors and represents a promising target to treat inflammatory pain. LINKED ARTICLES This article is part of a themed section on Cannabinoids in Biology and Medicine. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.165.issue-8. To view Part I of Cannabinoids in Biology and Medicine visit http://dx.doi.org/10.1111/bph.2011.163.issue-7
Booker, Lamont; Kinsey, Steven G; Abdullah, Rehab A; Blankman, Jacqueline L; Long, Jonathan Z; Ezzili, Cyrine; Boger, Dale L; Cravatt, Benjamin F; Lichtman, Aron H
Monocyte B7 and Sialyl Lewis X modulates the efficacy of IL-10 down-regulation of LPS-induced monocyte tissue factor in whole blood.
Recent studies have investigated the use of anti-inflammatory cytokine, interleukin 10 (IL-10) to control the development of disseminated intravascular coagulation (DIC) in sepsis by down-regulation of monocyte tissue factor (MTF) induced by lipopolysaccharide (LPS) in the initial phase of the disease. In vitro and in vivo human studies have shown that a minimal (<1 h) delay in IL-10 treatment significantly reduces the cytokines ability to inhibit LPS-induced MTF expression and the end products of coagulation. In this whole blood in vitro study we investigated the role of lymphocyte and platelet interactions with monocytes to up-regulate MTF expression in the presence of IL-10 in the initial phase of exposure to LPS. Individual blockade of monocyte B7 or platelet P-selectin significantly (35%) reduced MTF expression (P<0.05). IL-10 showed a dose-dependent inhibition of LPS (0.1 microg/ml) induced MTF expression, with 56% inhibition at 1 ng/ml, maximizing at 5 ng/ml IL-10 (75%; P<0.05). Simultaneous exposure to LPS and IL-10 (1 ng/ml) or addition of IL-10 1 h after LPS, with individual B7 and P-selectin blockade significantly enhanced the inhibition of MTF expression by IL-10 (P<0.05). We conclude that the efficacy of IL-10 to control DIC could be enhanced by a simultaneous B7 and P-selectin blockade. PMID:9695978
Warnes, G; Biggerstaff, J P; Francis, J L
SOCS3 Expression Correlates with Severity of Inflammation, Expression of Proinflammatory Cytokines, and Activation of STAT3 and p38 MAPK in LPS-Induced Inflammation In Vivo
SOCS3 is an inducible endogenous negative regulator of JAK/STAT pathway, which is relevant in inflammatory conditions. We used a model of LPS-induced periodontal disease in rats to correlate SOCS3 expression with the inflammatory status. In vitro we used a murine macrophage cell line to assess the physical interaction between SOCS3 and STAT3 by coimmunoprecipitation. 30?ug of LPS from Escherichia coli were injected in the gingival tissues on the palatal aspect of first molars of the animals 3x/week for up to 4 weeks. Control animals were injected with the vehicle (PBS). The rats were sacrificed at 7, 15, and 30 days. Inflammation and gene expression were assessed by stereometric analysis, immunohistochemistry, RT-qPCR, and western blot. LPS injections increased inflammation, paralleled by an upregulation of SOCS3, of the proinflammatory cytokines IL-1?, IL-6, and TNF-? and increased phosphorylation of STAT3 and p38 MAPK. SOCS3 expression accompanied the severity of inflammation and the expression of proinflammatory cytokines, as well as the activation status of STAT3 and p38 MAPK. LPS stimulation in a macrophage cell line in vitro induced transient STAT3 activation, which was inversely correlated with a dynamic physical interaction with SOCS3, suggesting that this may be a mechanism for SOCS3 regulatory function.
Chaves de Souza, Joao Antonio; Nogueira, Andressa Vilas Boas; Chaves de Souza, Pedro Paulo; Kim, Yeon Jung; Silva Lobo, Caroline; Pimentel Lopes de Oliveira, Guilherme Jose; Cirelli, Joni Augusto; Garlet, Gustavo Pompermaier; Rossa, Carlos
Ginsenoside Rh2 Downregulates LPS-Induced NF-?B Activation through Inhibition of TAK1 Phosphorylation in RAW 264.7 Murine Macrophage
The present study was carried out to evaluate the inhibitory effects of ginsenoside Rh2 on nuclear-factor- (NF-) ?B in lipopolysaccharide- (LPS-) activated RAW 264.7 murine macrophages. RAW 264.7 cells were pretreated with indicated concentrations of ginsenoside Rh2 for 1?h prior to the incubation of LPS (1??g/mL) for indicated time period. Ginsenoside Rh2 reduced CD14 and Toll-like receptor 4 (TLR4) expressions 24?h after LPS stimulation. Furthermore, ginsenoside Rh2 significantly inhibited TGF-beta-activated kinase 1 (TAK1) phosphorylation 30?min after LPS stimulation. Ginsenoside Rh2 was further shown to inhibit NF-?B p65 translocation into the nucleus by suppressing I?B-? degradation. Also, LPS increased mRNA expression of TNF-? and IL-1? time-dependently, while TQ reduced TNF-? within 3?h and IL-1? within 1?h. And we firstly found that pretreatment of ginsenoside Rh2 successively inhibited hypoxia-inducible factor- (HIF-) 1? expression increased by LPS. In conclusion, ginsenoside Rh2 may inhibit LPS-induced NF-?B activation and reduce HIF-1? accumulation, suggesting that ginsenoside Rh2 may be considered as a potential therapeutic candidate for chronic inflammatory diseases.
Lian, Li-Hua; Jin, Quan; Song, Shun-Zong; Wu, Yan-Ling; Bai, Ting; Jiang, Shuang; Li, Qian; Yang, Ning; Nan, Ji-Xing
High glucose increases LPS-induced DC apoptosis through modulation of ERK1/2, AKT and Bax/Bcl-2
Background This study investigates the effect of glucose on the LPS-induced apoptosis of dendritic cells in the intestinal tract of mice and the dendritic cell line DC2.4. Methods Flow cytometry was used to detect dendritic cell apoptosis both in vivo and in vitro. Hoechst 33258 staining was used to detect the morphological changes characteristic of apoptotic nuclei. Expression of apoptosis related proteins was investigated by western blot analysis and immunohistochemistry. Results Pretreatment with a high concentration of glucose increased apoptosis of LPS-treated dendritic cells both in vivo and in vitro at 24�h. No effect was evident at the earlier time points of 15�min and 6�h in vitro. Furthermore, at 24�hours the expression of the survival proteins AKT, ERK and Bcl-2 was decreased, while the expression of the proapoptotic protein Bax was increased. AKT, ERK, Bcl-2 and Bax were mainly located in the cytoplasm by immunohistochemistry. Conclusions These results suggest that high glucose concentrations might prime dendritic cells for apoptosis induced by LPS in the intestinal tract through upregulating the expression of Bax and downregulating the expression of AKT, ERK and Bcl-2. Therefore, this study may give clues to understanding the immunological mechanism behind gastrointestinal complications in diabetes mellitus.
Modulation of LPS-induced CD4+ T-cell activation and apoptosis by antioxidants in untreated asymptomatic HIV infected participants: an in vitro study.
Persistent immune activation characterises HIV infection and is associated with depletion of CD4+ T-cells and increased risk of disease progression. Early loss of gut mucosal integrity results in the translocation of microbial products such as lipopolysaccharide (LPS) into the systemic circulation. This is an important source of on-going immune stimulation. The purpose of this study was to determine levels of CD4+ T-cell activation (%CD25 expression) and apoptosis (% annexin V/7-AAD) in asymptomatic, untreated HIV infection at baseline and after stimulation with LPS and incubation with or without vitamin C and N-acetylcysteine. LPS induced a significant (P < 0.03) increase in %CD25 expression, annexin V, and 7-AAD in HIV positive individuals. NAC in combination with vitamin C, significantly (P = 0.0018) reduced activation and early apoptosis of CD4+ T-cells to a greater degree than with either antioxidant alone. Certain combinations of antioxidants could be important in reducing the harmful effects of chronic immune activation and thereby limit CD4+ T-cell depletion. Importantly, we showed that CD4+ T-cells of the HIV positive group responded better to a combination of the antioxidants at this stage than those of the controls. Therefore, appropriate intervention at this asymptomatic stage could rescue the cells before repetitive activation results in the death of CD4+ T-cells. PMID:24348678
Mburu, S; Marnewick, J L; Abayomi, A; Ipp, H
Anti-inflammatory effects of hydrophilic and lipophilic statins with hyaluronic acid against LPS-induced inflammation in porcine articular chondrocytes.
The objective of the study is to understand the therapeutic effects of lipophilic (simvastatin) and hydrophilic statins (pravastatin) combined with/without hyaluronic acid for osteoarthritis by an in vitro LPS-induced inflammatory model of articular chondrocytes. HA in combination with different doses of simvastatin or pravastatin were used. Beside cytotoxicity, the influence of statins on NO production, pro-inflammatory cytokine, inflammatory mediators, and NF-?B p50 protein were analyzed. Finally, TUNEL assay was performed to detect DNA strand breakage. Two statins were less able to lower NF-?B activity when they were administrated along without HA. The gene expression demonstrates that simvastatin and pravastatin had the ability to decrease pro-inflammatory and inflammatory mediator levels. High dose simvastatin with or without HA down regulated inflammatory cytokines, but resulted in higher cytotoxicity. TUNEL assay confirms the regulatory effect of statins with or without HA over the apoptosis of chondrocytes, especially in hydrophilic statins. The significant down-regulation of inflammatory mediators suggests that intra-articular injection of HA in combination with statins might feasibly slow the progress of osteoarthritis. Administration of simvastatin or pravastatin with hyaluronic acid may produce beneficial effects for OA treatment, but with better results when hydrophilic statin was used. PMID:24302463
Chang, Chih-Hung; Hsu, Yuan-Ming; Chen, Yu-Chun; Lin, Feng-Huei; Sadhasivam, Subramaniam; Loo, Siow-Tung; Savitha, Sivasubramanian
Inhibition of sPLA2-IIA Prevents LPS-Induced Neuroinflammation by Suppressing ERK1/2-cPLA2? Pathway in Mice Cerebral Cortex
Neuroinflammation is involved in various central nervous system (CNS) disorders, including brain infections, ischemia, trauma, stroke, and degenerative CNS diseases. In the CNS inflammation, secretory phospholipase A2-IIA (sPLA2-IIA) acts as a mediator, resulting in the generation of the precursors of pro-inflammatory lipid mediators, such as prostaglandins (PGs) and leukotrienes (LTs). However, the role of sPLA2-IIA in neuroinflammation is more complicated and remains unclear yet. In the present study, we investigated the effect of sPLA2-IIA inhibition by specific inhibitor SC-215 on the inflammation in LPS-induced mice cerebral cortex and primary astrocytes. Our results showed that the inhibition of sPLA2-IIA alleviated the release of PGE2 by suppressing the activation of ERK1/2, cPLA2?, COX-2 and mPGES-1. These findings demonstrated that sPLA2-IIA showed the potential to regulate the neuroinflammation in vivo and in vitro, indicating that sPLA2-IIA might be a novel target for the treatment of acute neuroinflammation.
Xiang, Yanxiao; Chen, Lin; Liu, Huiqing; Liu, Xiaoqian; Wei, Xinbing; Sun, Baozhu; Wang, Tian; Zhang, Xiumei
Protective Effect of the Fruit Hull of Gleditsia sinensis on LPS-Induced Acute Lung Injury Is Associated with Nrf2 Activation
The fruit hull of Gleditsia sinensis (FGS) has been prescribed as a traditional eastern Asian medicinal remedy for the treatment of various respiratory diseases, but the efficacy and underlying mechanisms remain poorly characterized. Here, we explored a potential usage of FGS for the treatment of acute lung injury (ALI), a highly fatal inflammatory lung disease that urgently needs effective therapeutics, and investigated a mechanism for the anti-inflammatory activity of FGS. Pretreatment of C57BL/6 mice with FGS significantly attenuated LPS-induced neutrophilic lung inflammation compared to sham-treated, inflamed mice. Reporter assays, semiquantitative RT-PCR, and Western blot analyses show that while not affecting NF-?B, FGS activated Nrf2 and expressed Nrf2-regulated genes including GCLC, NQO-1, and HO-1 in RAW 264.7 cells. Furthermore, pretreatment of mice with FGS enhanced the expression of GCLC and HO-1 but suppressed that of proinflammatory cytokines in including TNF-? and IL-1? in the inflamed lungs. These results suggest that FGS effectively suppresses neutrophilic lung inflammation, which can be associated with, at least in part, FGS-activating anti-inflammatory factor Nrf2. Our results suggest that FGS can be developed as a therapeutic option for the treatment of ALI.
Choi, Jun-Young; Kwun, Min Jung; Kim, Kyun Ha; Lyu, Ji Hyo; Han, Chang Woo; Jeong, Han-Sol; Ha, Ki-Tae; Jung, Hee-Jae; Lee, Beom-Joon; Sadikot, Ruxana T.; Christman, John W.; Jung, Sung-Ki; Joo, Myungsoo
Critical Roles of the WASP N-Terminal Domain and Btk in LPS-Induced Inflammatory Response in Macrophages
While Wiskott-Aldrich syndrome protein (WASP) plays critical roles in TCR signaling as an adaptor molecule, how it transduces innate immune signals remains to be elucidated. To investigate the roles of WASP in innate immune cells, we established bone marrow-derived macrophage (BMDM) cell lines from WASP15 transgenic (Tg) mice overexpressing the WASP N-terminal region (exons 1�5). Upon LPS stimulation, WASP15 Tg BMDM cell lines produce lower levels of inflammatory cytokines, such as TNF-?, IL-6, and IL-12p40 than the wild-type BMDM cell line. In addition, the production of nitric oxide by WASP15 Tg BMDM cells in response to LPS and IFN-? was significantly impaired. Furthermore, we uncovered that the WASP N-terminal domain associates with the Src homology (SH) 3 domain of Bruton's tyrosine kinase (Btk). Overexpression of the WASP N-terminal domain diminishes the extent of tyrosine phosphorylation of endogenous WASP in WASP15 Tg BMDM cells, possibly by interfering with the specific binding between endogenous WASP and Btk during LPS signaling. These observations strongly suggest that the interaction between WASP N-terminal domain and Btk plays important roles in the LPS signaling cascade in innate immunity.
Sakuma, Chisato; Sato, Mitsuru; Takenouchi, Takato; Chiba, Joe; Kitani, Hiroshi
CD40 is a co-stimulatory surface molecule actively expressed on mature dendritic cells (DC). Recent studies suggest that endotoxin (LPS) inhalation induces DC maturation in the airways of healthy volunteers. To characterize the effect of LPS on CD40 expression and underlying mech...
Anti-inflammatory effect of anemarsaponin B isolated from the rhizomes of Anemarrhena asphodeloides in LPS-induced RAW 264.7 macrophages is mediated by negative regulation of the nuclear factor-kappaB and p38 pathways.
Anemarrhena asphodeloides is widely used in traditional Chinese medicine, and is known to have anti-diabetic and diuretic effects. In this study, we evaluated the anti-inflammatory effects of anemarsaponin B (ASB), a steroidal saponin isolated from the rhizomes of A. asphodeloides (Liliaceae), in LPS-stimulated RAW 264.7 macrophage cell line. ASB significantly and dose-dependently decreased the protein and mRNA levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). ASB also reduced the expressions and productions of pro-inflammatory cytokines, including those of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6). Electrophoretic mobility shift assay (EMSA) and reporter gene assays revealed that ASB attenuated the LPS-induced DNA binding and transcriptional activity of nuclear factor-kappa B (NF-kappaB). In addition, it was found that pretreatment with ASB significantly inhibited the nuclear translocation of the p65 subunit of NF-kappaB by blocking the phosphorylation of inhibitory kappa B-alpha (IkappaB-alpha). On the other hand, ASB inhibited the phosphorylation of MAP kinase kinases 3/6 (MKK3/6) and mixed lineage kinase 3 (MLK3), which are both involved in the p38 pathway. Taken together, these results suggest that anti-inflammatory effect of ASB in LPS-treated RAW 264.7 macrophages is associated with the inhibition of NF-kappaB transcriptional activity, possibly via the p38 MAP kinase pathway. PMID:19375480
Kim, Ji-Yeon; Shin, Ji-Sun; Ryu, Jong Hoon; Kim, Sun Yeou; Cho, Young-Wuk; Choi, Jung-Hye; Lee, Kyung-Tae
c-Jun N-terminal kinase inhibition and alpha-tocopherol protect midbrain dopaminergic neurons from interferon-gamma/lipopolysaccharide-induced injury without affecting nitric oxide production.
Interferon-gamma (IFN-gamma)/lipopolysaccharide (LPS) induces delayed dopaminergic neuron loss in midbrain slice cultures, because of nitric oxide production resulting from p38 mitogen-activated protein kinase (p38 MAPK)-dependent induction of inducible nitric oxide synthase (iNOS). In this study, we show that inhibition of c-Jun N-terminal kinase (JNK), but not of extracellular signal-regulated kinase, protects dopaminergic neurons from IFN-gamma/LPS-induced degeneration. In contrast to a p38 MAPK inhibitor, SB203580, however, a JNK inhibitor, anthra[1,9-cd]pyrazol-6(2H)-one (SP600125), did not suppress IFN-gamma/LPS-induced iNOS expression and nitric oxide production. Involvement of NADPH oxidase-derived superoxide production in dopaminergic neurodegeneration was not obvious, in that superoxide dismutase/catalase or manganese 3-methoxy-N,N'-bis(salicylidene)ethylenediamine chloride (EUK-134), a superoxide dismutase/catalase mimetic, did not afford neuroprotection. Moreover, the NADPH oxidase inhibitors apocynin and diphenylene iodonium were protective against IFN-gamma/LPS cytotoxicity only at concentrations that suppressed nitric oxide production. Notably, alpha-tocopherol effectively prevented IFN-gamma/LPS-induced dopaminergic neuron degeneration, without affecting iNOS induction and nitric oxide production. These results underscore the neuroprotective potential of JNK inhibitor and alpha-tocopherol, in the sense that both agents could rescue dopaminergic neurons under inflammatory conditions associated with robust increases in nitric oxide production. PMID:16307444
Shibata, Haruki; Katsuki, Hiroshi; Okawara, Mitsugi; Kume, Toshiaki; Akaike, Akinori
Role of p38 MAPK in LPS induced pro-inflammatory cytokine and chemokine gene expression in equine leukocytes.
Endotoxemia occurs when bacterial lipopolysaccharide (LPS) in the blood induces a dysregulated inflammatory response, resulting in circulatory shock and multi-organ failure. Laminitis is a common complication in endotoxemic horses and is frequently the reason for humane euthanasia of these cases. Blood leukocytes are a principal target of LPS in endotoxemia leading to activation of multiple signal transduction pathways involved in the induction of a number of pro-inflammatory genes. In other animal models, the p38 mitogen activated protein kinase (MAPK) pathway has been associated with induced expression of tumor necrosis factor-alpha (TNFalpha), interleukin (IL)-1beta, IL-6 and IL-8. The goal of this study was to determine the role of the p38 MAPK pathway in the induction of these pro-inflammatory cytokine and chemokine genes in LPS-stimulated equine leukocytes. Stimulation of equine peripheral blood leukocytes resulted in an increase in TNFalpha, IL-1beta, IL-6 and IL-8 mRNA levels. Pharmacological inhibition of p38 MAPK activity with SB203580 or SB202190 reduced the ability of LPS stimulation to increase mRNA concentrations for all four genes. However, only SB203580 pretreatment significantly reduced LPS-stimulated IL-1beta and IL-8 mRNA expression and only pretreatment with SB202190 significantly reduced LPS-stimulated TNFalpha and IL-6 mRNA expression. From this study we conclude TNFalpha, IL-1beta, IL-6 and IL-8 are induced upon LPS stimulation of equine leukocytes and that this induction of gene expression is dependent on the p38 MAPK pathway. However, there are differences in the efficacy of the p38 inhibitors tested here that may be explained by differences in specificity or potency. This study provides evidence for the use of selective p38 MAPK inhibitors as potential therapeutics for the treatment of equine endotoxemia. PMID:19070370
Neuder, Laura E; Keener, Jamie M; Eckert, Rachael E; Trujillo, Jennifer C; Jones, Samuel L
The binding site for the transcription factor, NF-?B, on the cystathionine ?-lyase promoter is critical for LPS?induced cystathionine ?-lyase expression.
Hydrogen sulfide (H2S) is regarded as the third endogenous gaseous signaling molecule. Cystathioine ?-lyase (CSE), one of the three enzymes in the transsulfuration pathway, is responsible for the production of endogenous H2S. The H2S/CSE signaling pathway is involved in the inflammation induced by lipopolysaccharides�(LPS). Therefore, in this study, we investigated the effects of the binding site (on the CSE promoter) for the transcription factor, nuclear factor (NF)-?B, on the transcriptional regulation of the CSE gene in mammalian cells treated with LPS. For this purpose, HEK-293 and COS-7 cells were transfected with 5��g pGL4.12-KM1478 or 5��g pGL4.12-KM1478m (mutant) together with the pRL-CMV control vector (0.032��g for the HEK-293 cells, 0.0032��g for the COS-7�cells). Subsequently, the cells were treated with LPS for 6�h. The expression of CSE was measured by RT-qPCR. cDNA pooled from J774.1A and RAW264.7 cells treated with LPS for 6�h was used to estimate the quantity of the transcripts. Our results revealed that LPS markedly increased the mRNA and protein expression levels of the CSE gene in the J774.1A and RAW264.7 cells following treatment with LPS for 6�h. In addition, we found that the GGGACATTCC DNA sequence on the promoter of the CSE gene was closely associated with the transcriptional regulation of the CSE gene in the HEK-293 and COS-7 cells treated with LPS. Taken together, our data suggest that the NF-?B binding site on CSE promoter is critical for LPS-induced CSE expression in mammalian cells. PMID:24866963
Wang, Maoxian; Guo, Zhanyun; Wang, Shilong
Translation control of TAK1 mRNA by hnRNP K modulates LPS-induced macrophage activation.
Macrophage activation by bacterial lipopolysaccharides (LPS) is induced through Toll-like receptor 4 (TLR4). The synthesis and activity of TLR4 downstream signaling molecules modulates the expression of pro- and anti-inflammatory cytokines. To address the impact of post-transcriptional regulation on that process, we performed RIP-Chip analysis. Differential association of mRNAs with heterogeneous nuclear ribonucleoprotein K (hnRNP K), an mRNA-specific translational regulator in differentiating hematopoietic cells, was studied in noninduced and LPS-activated macrophages. Analysis of interactions affected by LPS revealed several mRNAs encoding TLR4 downstream kinases and their modulators. We focused on transforming growth factor-?-activated kinase 1 (TAK1) a central player in TLR4 signaling. HnRNP K interacts specifically with a sequence in the TAK1 mRNA 3' UTR in vitro. Silencing of hnRNP K does not affect TAK1 mRNA synthesis or stability but enhances TAK1 mRNA translation, resulting in elevated TNF-?, IL-1?, and IL-10 mRNA expression. Our data suggest that the hnRNP K-3' UTR complex inhibits TAK1 mRNA translation in noninduced macrophages. LPS-dependent TLR4 activation abrogates translational repression and newly synthesized TAK1 boosts macrophage inflammatory response. PMID:24751651
Liepelt, Anke; Mossanen, Jana C; Denecke, Bernd; Heymann, Felix; De Santis, Rebecca; Tacke, Frank; Marx, Gernot; Ostareck, Dirk H; Ostareck-Lederer, Antje
p120 Modulates LPS-Induced NF-?B Activation Partially through RhoA in Bronchial Epithelial Cells
p120-Catenin (p120) is an adherens junction protein recognized to regulate cell-cell adhesion. Emerging evidence indicates that p120 may also play an important role in inflammatory responses, and the regulatory mechanisms are still unknown. In the present study, we showed that p120 was associated with airway inflammation. p120 downregulation induced nuclear factor-?B (NF-?B) activation, accompanied with I?B? degradation, p65 nuclear translocation, and increased expression of interleukin-8 (IL-8) in lipopolysaccharide (LPS)- treated C57BL mice and human bronchial epithelial cells (BECs). Moreover, we first found that p120 directly coprecipitated with RhoA in BECs. After LPS stimulation, although total RhoA and p120-bound RhoA were unchanged, RhoA activity was increased. Y27632, a ROCK inhibitor, could partially inhibit nuclear translocation of p65. Overexpression of p120 inactivated RhoA and NF-?B in BECs, whereas p120 loss significantly increased RhoA activity, p65 nuclear translocation, and IL-8 expression. Taken together, our study supports the regulatory role of p120 in airway inflammation and reveals that p120 may modulate NF-?B signaling partially through RhoA.
Qin, Shenghui; Zhang, Yanli; Liu, Liwei; Wu, Renliang
Organ-specific protection against LPS-induced vascular leak is dependent on the endothelial protein C receptor (EPCR)
Objective To study the role of the endothelial protein C receptor (EPCR) in the modulation of susceptibility to inflammation-induced vascular leak in vivo. Approach/Results Genetically modified mice with low, <10%-EPCR expression (EPCRlow) and control mice were challenged with lipopolysaccharides (LPS) in a mouse model of endotoxemia. Infrared fluorescence and quantification of albumin-bound Evans Blue (EB) in tissues and intravascular plasma volumes were used to assess plasma extravasation. Pair wise analysis of EPCRlow and control mice matched for gender, age, and weight allowed determination of EPCR-dependent vascular leak. Kidney, lung, and brain were the organs with highest discriminative increased EB accumulation in EPCRlow versus control mice in response to LPS. Histology of kidney and lung confirmed the EPCR-specific pathology. In addition to severe kidney injury in response to LPS, EPCRlow and anti-EPCR-treated wild-type mice suffered from enhanced albuminuria and profound renal hemorrhage versus controls. Intravascular volume loss at the same extent of weight loss in EPCRlow mice compared to control mice provided proof that plasma leak was the predominant cause of EB tissue accumulation Conclusions This study demonstrates an important protective role for EPCR in vivo against vascular leakage during inflammation and suggests that EPCR-dependent vascular protection is organ specific.
von Drygalski, Annette; Furlan-Freguia, Christian; Ruf, Wolfram; Griffin, John H.; Mosnier, Laurent O.
Ron receptor deficient alveolar myeloid cells exacerbate LPS-induced acute lung injury in the murine lung
Previous studies have shown that the Ron receptor tyrosine kinase is an important regulator of the acute lung inflammatory response induced by intranasal administration of bacterial lipopolysaccharide (LPS). Compared to wild type mice, complete loss of the Ron receptor in all cell types in vivo was associated with increased lung damage as determined by histological analyses and several markers of lung injury including increases in pro-inflammatory cytokines such as TNF?. TNF? is a multifunctional cytokine secreted by macrophages, which plays a major role in inflammation and is a central mediator of several disease states including rheumatoid arthritis and sepsis. Based on increased TNF? production observed in the Ron-deficient mice, we hypothesized that Ron receptor function in the inflammatory cell compartment is essential for the regulating lung injury in vivo. To test this hypothesis we generated myeloid lineage-specific Ron deficient mice. In this study, we report that loss of Ron signaling selectively in myeloid cells results in increased lung injury following intranasal administration of LPS as measured by increases in TNF? production, ensuing neutrophil accumulation and increased lung histopathology. These findings corroborate the role of Ron receptor tyrosine kinase as a negative regulator of inflammation and further demonstrate the in vivo significance of Ron signaling selectively in myeloid cells as a major regulator of this response in vivo. These data authenticate Ron as a potential target in innate immunity and TNF?-mediated pathologies.
Nikolaidis, Nikolaos M.; Kulkarni, Rishikesh M.; Gray, Jerilyn K.; Collins, Margaret H.; Waltz, Susan E.
Furanodien-6-one from Commiphora erythraea inhibits the NF-?B signalling and attenuates LPS-induced neuroinflammation.
We investigated the in vitro anti-inflammatory activity of 1(10),4-furanodien-6-one, one the most active compounds of the hexane extract of Commiphora erythraea (Ehrenb.) Engl., by exposing microglial BV-2 cells to lipopolysaccharide. We showed that furanodien-6-one pre-treatment restored cell viability and ROS to control levels while halving NO generation. Production of pro-inflammatory IL-6, IL-23, IL-17, TGF-?, and INF-?, significantly induced by LPS, was also markedly reduced by furanodien-6-one treatment. We further showed that furanodien-6-one protects primary neuronal cultures against the inflammatory/toxic insults of LPS-treated BV-2 conditioned media, indicating that furanodien-6-one exerts anti-inflammatory/cytoprotective effects in neuronal cells. We then investigated whether furanodien-6-one exerts anti-inflammatory properties in an in vivo model of microglial activation. In adult mice ip-injected with LPS we found that furanodien-6-one had strong cerebral anti-inflammatory properties by inhibiting liver and brain TNF? as well as IL-1? expression. Results were not unexpected since FTIR-metabolomic analyses showed that furanodien-6-one-treated mice had a reduced dissimilarity to control animals and that the response to LPS treatment was markedly modified by furanodien-6-one. In conclusion our data provide strong evidence of the anti-inflammatory properties of furanodien-6-one that could be exploited to counteract degenerative pathologies based on neuroinflammation. PMID:23357788
Bellezza, Ilaria; Mierla, Annalisa; Grottelli, Silvia; Marcotullio, Maria Carla; Messina, Federica; Roscini, Luca; Cardinali, Gianluigi; Curini, Massimo; Minelli, Alba
Intraperitoneal injection of lactoferrin ameliorates severe albumin extravasation and neutrophilia in LPS-induced inflammation in neonatal rats.
Lactoferrin (LF) plays various anti-inflammatory roles in inflammation experimentally induced by lipopolysaccharides (LPS). But the effects of LF on albumin extravasation and neutrophilia have not been elucidated. We aimed to study the effects of LF on albumin extravasation, neutrophilia and/or on other symptoms in inflammation caused by LPS in rats. Human lactoferrin (hLF) was injected (10 mg/100 mL in PBS) 18 h, or 15 min prior to, or 60 min after intraperitoneal injection of LPS in 13 days old Sprague Dawley rats. Prophylactic injection of hLF significantly ameliorated albumin extravasation in ascitic fluid at 5 h and neutrophilia in the blood at 24 h after LPS injection, but the after-injection of hLF did not. Interestingly, an injection of rat anti-TNFalpha IgG 15 min prior to LPS injection did not ameliorate albumin extravasation. Prophylactic injection of hLF significantly ameliorated other symptoms like mortality, and the decrease of phagocytotic activity of peritoneal polymorpho-nuclear leukocytes (PMNL), but did not ameliorate the decrease of platelets in the plasma. These findings suggest that hLF may be available as a medical treatment prior to surgery for prophylaxis of side effects like albumin extravasation or neutrophilia. PMID:16415506
Yajima, Masako; Yajima, Takaji; Kuwata, Tamotsu
Role of TLR signaling in Francisella tularensis-LPS-induced, antibody-mediated protection against Francisella tularensis challenge.
Immunization with Ft-LPS provokes an antigen-specific, B-1a cell-derived antibody response that protects WT mice against an otherwise lethal challenge with Ft LVS. However, this same regimen offers limited protection to TLR2(-/-) mice, despite production of WT levels of anti-Ft-LPS antibodies. As Ft-LPS exhibits no TLR2 agonist activity, and macrophage-induced cytokine production in response to Ft LVS is overwhelmingly TLR2-dependent, we hypothesized that treatment of TLR2(-/-) mice with an alternative, MyD88-dependent TLR agonist would compensate for reduced recognition of Ft LVS in TLR2(-/-) mice and thereby, restore Ft-LPS-mediated protection. Administration of the nontoxic TLR4 agonist, synthetic Escherichia coli MPL, at the time of Ft-LPS immunization or Ft LVS challenge, fully protected TLR2(-/-) mice, whereas treatment of WT or TLR2(-/-) mice with MPL alone conferred partial protection. The TLR5 agonist, flagellin, also synergized with Ft-LPS to protect TLR2(-/-) mice from lethal Ft LVS challenge. In contrast to Ft LVS, Ft-LPS pretreatment failed to protect mice against i.n. challenge with Ft Schu S4, whereas MPL, administered in the absence or presence of Ft-LPS, conferred significant, albeit partial, protection. MPL treatment of macrophages increased the uptake of Ft LVS and decreased intracellular bacterial survival while shifting the macrophage-differentiation phenotype from "alternatively activated" to "classically activated". Collectively, our data suggest that optimal, Ft-LPS-mediated protection against Ft LVS infection requires two discrete events, i.e., production of Ft-LPS-specific antibody, as well as TLR-mediated macrophage activation, to fully control Francisella infection. PMID:21750122
Cole, Leah E; Mann, Barbara J; Shirey, Kari Ann; Richard, Katharina; Yang, Yang; Gearhart, Patricia J; Chesko, Kirsty L; Viscardi, Rose M; Vogel, Stefanie N
Role of TLR signaling in Francisella tularensis-LPS-induced, antibody-mediated protection against Francisella tularensis challenge
Immunization with Ft-LPS provokes an antigen-specific, B-1a cell-derived antibody response that protects WT mice against an otherwise lethal challenge with Ft LVS. However, this same regimen offers limited protection to TLR2?/? mice, despite production of WT levels of anti-Ft-LPS antibodies. As Ft-LPS exhibits no TLR2 agonist activity, and macrophage-induced cytokine production in response to Ft LVS is overwhelmingly TLR2-dependent, we hypothesized that treatment of TLR2?/? mice with an alternative, MyD88-dependent TLR agonist would compensate for reduced recognition of Ft LVS in TLR2?/? mice and thereby, restore Ft-LPS-mediated protection. Administration of the nontoxic TLR4 agonist, synthetic Escherichia coli MPL, at the time of Ft-LPS immunization or Ft LVS challenge, fully protected TLR2?/? mice, whereas treatment of WT or TLR2?/? mice with MPL alone conferred partial protection. The TLR5 agonist, flagellin, also synergized with Ft-LPS to protect TLR2?/? mice from lethal Ft LVS challenge. In contrast to Ft LVS, Ft-LPS pretreatment failed to protect mice against i.n. challenge with Ft Schu S4, whereas MPL, administered in the absence or presence of Ft-LPS, conferred significant, albeit partial, protection. MPL treatment of macrophages increased the uptake of Ft LVS and decreased intracellular bacterial survival while shifting the macrophage-differentiation phenotype from �alternatively activated� to �classically activated�. Collectively, our data suggest that optimal, Ft-LPS-mediated protection against Ft LVS infection requires two discrete events, i.e., production of Ft-LPS-specific antibody, as well as TLR-mediated macrophage activation, to fully control Francisella infection.
Cole, Leah E.; Mann, Barbara J.; Shirey, Kari Ann; Richard, Katharina; Yang, Yang; Gearhart, Patricia J.; Chesko, Kirsty L.; Viscardi, Rose M.; Vogel, Stefanie N.
Tumor Necrosis Factor-? from Macrophages Enhances LPS-Induced Clara Cell Expression of Keratinocyte-Derived Chemokine
Tumor necrosis factor (TNF)-? is a cytokine produced by alveolar macrophages in response to LPS in the lung. Clara cells are bronchiolar epithelial cells that produce a variety of proinflammatory cytokines in response to LPS but not to TNF-?. In this study, we examined whether TNF-? affects Clara cell cytokine production in the setting of LPS stimulation. Using a transformed murine Clara cell line (C22), we observed that both LPS and TNF-? induced production of keratinocyte-derived chemokine (KC) and monocyte chemoattractant protein (MCP)-1. We also found that simultaneous LPS and TNF-? stimulation is synergistic for KC production, but additive for MCP-1 production. By using a Transwell coculture system of RAW264.7 macrophages and Clara cells isolated from C57Bl/6 mice, we found that macrophages produce a soluble factor that enhances Clara cell KC production in response to LPS. Cocultures of Clara cells from mice deficient in TNF-? receptors with RAW264.7 macrophages demonstrated that the effect of macrophages on Clara cells is mediated primarily via TNF-?. To determine whether these findings occur in vivo, we treated wild-type and TNF receptor�deficient mice intratracheally with LPS and examined the expression of KC. LPS-treated, TNF receptor�deficient mice showed much less KC mRNA in airway epithelial cells compared with wild-type mice. In contrast, a similar number of KC-expressing cells was seen in the lung periphery. Thus, upregulation of KC by Clara cells in the setting of LPS stimulation is largely dependent on TNF-? originating from alveolar macrophages. These findings shed light on macrophage�Clara cell interactions in regulating the pulmonary inflammatory response to LPS.
Elizur, Arnon; Adair-Kirk, Tracy L.; Kelley, Diane G.; Griffin, Gail L.; deMello, Daphne E.; Senior, Robert M.
Investigation of sanguinarine and chelerythrine effects on LPS-induced inflammatory gene expression in THP-1 cell line.
Quaternary benzo[c]phenanthridine alkaloids sanguinarine and chelerythrine have been used in folk medicine for their wide range of useful properties. One of their major effect is also anti-inflammatory activity, that is not clarified in detail. This study focused on the ability of these alkaloids to modulate the gene expression of pro-inflammatory tumour necrosis factor ? (TNF-?), monocyte chemoattractant protein 1 (MCP-1, also known as CCL-2), interleukin (IL)-6, IL-1? and anti-inflammatory cytokines IL-1 receptor antagonist (IL-1RA) and IL-10. The effect of these alkaloids was compared with that of conventional drug prednisone. Human monocyte-derived macrophages were pre-treated with alkaloids or prednisone and inflammatory reaction was induced by lipopolysaccharide. Changes of gene expression at the transcriptional level of mentioned cytokines were measured. In our study mainly affected pro-inflammatory cytokines were CCL-2 and IL-6. Two hours after LPS stimulation, cells influenced by sanguinarine and chelerythrine significantly declined the CCL-2 expression by a factors of 3.5 (p<0.001) and 1.9 (p<0.01); for those treated with prednisone the factor was 5.3 (p<0.001). Eight hours after LPS induction, both alkaloids significantly diminished the CCL-2 expression. The lower expression was found for sanguinarine--lower by a factor of 4.3 than for cells treated with the vehicle (p<0.001). Two hours after LPS stimulation, cells treated with sanguinarine decreased the IL-6 mRNA level by a factor of 3.9 (p<0.001) compared with cells treated with the vehicle. Chelerythrine decreased the level of IL-6 mRNA by a factor of 1.6 (p<0.001). Sanguinarine decreased gene expression of CCL-2 and IL-6 more than chelerythrine and its effect was quite similar to prednisone. Four hours after LPS stimulation, cells pre-treated with sanguinarine exhibited significantly higher expression (a factor of 1.7, p<0.001) of IL-1RA than cells without sanguinarine treatment. Our results help to clarify possible mechanisms of action of these alkaloids in the course of inflammation. PMID:22592163
P?n?�kov�, K; Koll�r, P; M�ller Z�valov�, V; T�borsk�, E; Urbanov�, J; Ho�ek, J
LPS-induced TLR4 signaling in human colorectal cancer cells increases beta1 integrin-mediated cell adhesion and liver metastasis.
Infectious complications resulting from resection of colorectal cancer (CRC) elevates the risk of cancer recurrence and metastasis, but the reason for this risk relationship is unknown. Defining the mechanisms responsible may offer opportunities to improve outcomes in a majority of patients whose tumors are resected as part of their therapy. The complex formed between Toll receptor TLR4 and myeloid differentiation factor MD2 defines a major cell surface receptor for lipopolysaccharide (LPS), a gram-negative bacterial antigen that has been implicated in infectious complications after CRC resection. As the TLR4/MD2 complex is expressed on CRC cells, we hypothesized that LPS may promote liver metastasis in CRC by stimulating TLR4 signaling. In support of this hypothesis, we report here that LPS enhances liver metastasis of human CRC cells that express TLR4/MD2 after intrasplenic graft of immunocompromised nude mice. Compared with TLR4 nonexpressing, nonmetastatic CRC cells, we observed increased in vitro adherence to different extracellular matrices and human umbilical vein endothelial cells (HUVEC). Furthermore, we observed an increased likelihood of in vivo capture within hepatic sinusoids after LPS treatment. No differences were apparent in phosphorylation of p38 and MAPK isoforms, but in metastatic CRC cells expressing surface TLR4 treatment with LPS increased Ser473 phosphorylation of AKT kinase. We showed that enhanced adherence elicited by LPS in these cells could be blocked at three different levels, using Eritoran (TLR4 small molecule antagonist), PI-103 (PI3K inhibitor), or anti-?1 integrin blocking antibodies. Taken together, the results indicate that stimulation of the TLR4/MD2 complex by LPS activates PI3K/AKT signaling and promotes downstream ?1 integrin function, thereby increasing the adhesiveness and metastatic capacity of CRC cells. Our findings suggest that inhibiting LPS-induced TLR4 signaling could improve therapeutic outcomes by preventing cancer metastasis during the perioperative period of CRC resection. PMID:21363926
Hsu, Rich Y C; Chan, Carlos H F; Spicer, Jonathan D; Rousseau, Mathieu C; Giannias, Betty; Rousseau, Simon; Ferri, Lorenzo E
Polygonum cuspidatum, compared with baicalin and berberine, inhibits inducible nitric oxide synthase and cyclooxygenase-2 gene expressions in RAW 264.7 macrophages.
Polygonum cuspidatum water extract (PCWE) was shown to be a potent inhibitor of lipopolysaccharide (LPS)-induced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). PCWE was compared to baicalin isolated from Scutellaria baicalensis Georgi and berberine of Coptidis rhizoma and Phellodendri cortex, for their effects on LPS-induced nitric oxide (NO) production and iNOS and COX-2 gene expressions in RAW 264.7 macrophages. Both PCWE and the compounds inhibited LPS-induced NO production in a concentration-dependent manner without a cytotoxicity. The decrease in NO production was in parallel with the inhibition of LPS-induced iNOS gene expression by PCWE and the compounds. In contrast, iNOS enzyme activity was not inhibited by PCWE and two agents. In addition, only PCWE inhibited LPS-induced prostaglandin E2 (PGE2) production and COX-2 gene expression without affecting COX-2 enzyme activity, while baicalin or berberine did not. Furthermore, N-nitro-L-arginine (NLA) and N-nitro-L-arginine methyl ester (L-NAME) pretreatment enhanced LPS-induced iNOS protein expression, which was inhibited by these PCWE and two agents, although LPS-induced COX-2 protein expression was not affected by NLA and L-NAME. PCWE inhibited PGE2 production and COX-2 protein expression in NLA/LPS or L-NAME/LPS-co-treated RAW 264.7 cell, however, baicalin or berberine did not. From the results, it was concluded that co-treatment with NOS inhibitors and PCWE effectively blocks acute production of NO and inhibits expression of iNOS and COX-2 genes. PMID:17553752
Kim, Kyung-Woon; Ha, Ki-Tai; Park, Cheol-Soo; Jin, Un-Ho; Chang, Hyen Wook; Lee, In-Seon; Kim, Cheorl-Ho
Regulation of ENaC-mediated alveolar fluid clearance by insulin via PI3K/Akt pathway in LPS-induced acute lung injury
Background Stimulation of epithelial sodium channel (ENaC) increases Na+ transport, a driving force of alveolar fluid clearance (AFC) to keep alveolar spaces free of edema fluid that is beneficial for acute lung injury (ALI). It is well recognized that regulation of ENaC by insulin via PI3K pathway, but the mechanism of this signaling pathway to regulate AFC and ENaC in ALI remains unclear. The aim of this study was to investigate the effect of insulin on AFC in ALI and clarify the pathway in which insulin regulates the expression of ENaC in vitro and in vivo. Methods A model of ALI (LPS at a dose of 5.0 mg/kg) with non-hyperglycemia was established in Sprague-Dawley rats receiving continuous exogenous insulin by micro-osmotic pumps and wortmannin. The lungs were isolated for measurement of bronchoalveolar lavage fluid(BALF), total lung water content(TLW), and AFC after ALI for 8 hours. Alveolar epithelial type II cells were pre-incubated with LY294002, Akt inhibitor and SGK1 inhibitor 30 minutes before insulin treatment for 2 hours. The expressions of ?-,?-, and ?-ENaC were detected by immunocytochemistry, reverse transcriptase polymerase chain reaction (RT-PCR) and western blotting. Results In vivo, insulin decreased TLW, enchanced AFC, increased the expressions of ?-,?-, and ?-ENaC and the level of phosphorylated Akt, attenuated lung injury and improved the survival rate in LPS-induced ALI, the effects of which were blocked by wortmannin. Amiloride, a sodium channel inhibitor, significantly reduced insulin-induced increase in AFC. In vitro, insulin increased the expressions of ?-,?-, and ?-ENaC as well as the level of phosphorylated Akt but LY294002 and Akt inhibitor significantly prevented insulin-induced increase in the expression of ENaC and the level of phosphorylated Akt respectively. Immunoprecipitation studies showed that levels of Nedd4-2 binding to ENaC were decreased by insulin via PI3K/Akt pathway. Conclusions Our study demonstrated that insulin alleviated pulmonary edema and enhanced AFC by increasing the expression of ENaC that dependent upon PI3K/Akt pathway by inhibition of Nedd4-2.
1,25-Dihydroxyvitamin D3 Promotes a Sustained LPS-Induced NF-?B-Dependent Expression of CD55 in Human Monocytic THP-1 Cells
The vitamin D3 system imposes immunosuppressive effects on monocytic cells, in part, by inhibiting NF-?B-dependent expression of proinflammatory mediators. CD55, a cell surface complement regulatory protein that promotes protective and anti-inflammatory properties, is reportedly an NF-?B target gene transiently induced in monocytic cells by the bacterial endotoxin LPS. CD55 is elevated on white cells in women experiencing preterm labor (a pathophysiology commonly associated with bacterial infection) and failure to maintain CD55 was associated with subsequent preterm delivery. We examined the influence of vitamin D3 signaling on LPS-induced expression of CD55 in human monocytic THP-1 cells using quantitative PCR, immunoblot, immunohistochemistry, and NF-?B activation pathway inhibitors. Non-NF-?B targets CD14 and CD11b, which modulate bacterial surveillance and eradication, respectively, were also examined. LPS produced a rapid transient 1.6-fold increase in CD55 mRNA. 1,25-D3 alone did not affect CD55 mRNA expression within the first 48 h. However, in 1,25-D3 pretreated cells, LPS produced a >4-fold immediate and sustained increase in CD55 mRNA and protein expression, which was blocked by NF-?B inhibitors. Our results unexpectedly suggest that vitamin D3 signaling may promote an anti-inflammatory response through an NF-?B-dependent increase in CD55 expression. As expected, LPS or 1,25-D3 alone led to sustained increases in CD14 and CD11b expression. In 1,25-D3 pretreated cells, LPS differentially regulated protein expression - CD14 (21-fold increase) and CD11b (a transient 2-fold decrease) - principally at the posttranscriptional level. The coordinated temporal expression of CD55, CD14 and CD11b would contribute to an anti-inflammatory response by providing protection against complement-mediated cell lysis during pathogen recognition and eradication. Overall, the vitamin D3 system may play a role coordinating an anti-inflammatory response pattern of the host complement immune system. This may be particularly important when considering the high rates of preterm births in blacks, a population that exhibits reduced circulating vitamin D3 levels.
Izban, Michael G.; Nowicki, Bogdan J.; Nowicki, Stella
Lipopolysaccharide-induced alveolar epithelial permeability: the role of nitric oxide.
Intratracheal instillation of lipopolysaccharide (LPS) in the rat has been used as a model of acute lung inflammation. Among the early events in this process is a transient increase in airspace epithelial permeability which peaks 4 h after intratracheal instillation of LPS. The increased epithelial permeability is concomitant with the influx of neutrophils into the airspaces, peaking 8 h postinstillation. We have investigated the mechanism of this LPS-induced increase in epithelial permeability. The role of the neutrophil in LPS-induced epithelial permeability was assessed by pretreatment with neutrophil antibody to abolish neutrophil influx, which did not affect the increase in epithelial permeability. Because LPS instillation also induced increased tumor necrosis factor alpha (TNF-alpha) activity in bronchoalveolar lavage (BAL) fluid, and its release by cultured BAL leukocytes from treated animals, TNF-alpha antibody was coinstilled intratracheally with LPS in rats. TNF-alpha antibody eliminated TNF-alpha activity in BAL fluid, but had no effect on LPS-induced increased epithelial permeability. Increased levels of nitric oxide (NO), measured as nitrite, were also present in BAL fluid from LPS-treated rat lungs and LPS-elicited BAL leukocytes produced increased NO in culture. Treatment of rats with the specific NO synthase inhibitor L-NMMA significantly diminished the LPS-induced increased epithelial permeability. These data suggest that NO is involved in LPS-induced changes in epithelial integrity. However, other mechanisms should be evoked in addition to NO to explain completely the increased epithelial permeability produced by LPS. PMID:9563715
Li, X Y; Donaldson, K; MacNee, W
Differential sensitivity to LPS-induced myocardial dysfunction in the isolated Brown Norway and Dahl S rat hearts: roles of mitochondrial function, NF?B activation and TNF-? production
Recently we reported that BN rats were more resistant to lipopolysaccharide (LPS)-induced myocardial dysfunction than SS rats. This differential sensitivity was exemplified by reduced production of proinflammatory cytokines and diminished NF?B pathway activation. To further clarify the mechanisms of different susceptibility of these two strains to endotoxin, this study was designed to examine the alterations of cardiac and mitochondrial bioenergetics, proinflammatory cytokines, and signaling pathways after hearts were isolated and exposed to LPS ex vivo. Isolated BN and SS hearts were perfused with LPS (4 ?g/ml) for 30 min in the Langendorff preparation. LPS depressed cardiac function as evident by reduced left ventricular developed pressure as well as decreased peak rate of contraction and relaxation in SS hearts, but not in BN heart. These findings are consistent with our previous in vivo data. Under complex I substrates a higher O2 consumption and H2O2 production were observed in mitochondria from SS hearts than that from BN hearts. LPS significantly increased H2O2 levels in both SS and BN heart mitochondria; however the increase in O2 consumption and H2O2 production in BN heart mitochondria was much lower than that in SS heart mitochondria. Additionally LPS significantly decreased complex I activity in SS hearts but not in BN hearts. Furthermore, LPS induced higher levels of TNF-? and increased phosphorylation of I?B and p65 more in SS hearts than BN hearts. Our results clearly demonstrate that less mitochondrial dysfunction combined with a reduced production of TNF-? and diminished activation of NF?B are involved in the mechanisms by which isolated BN hearts were more resistant to LPS-induced myocardial dysfunction.
An, Jianzhong; Du, Jianhai; Wei, Na; Guan, Tongju; Camara, Amadou K.S.; Shi, Yang
Synthesis and effects of new caffeic acid derivatives on nitric oxide production in lipopolysaccharide-induced RAW 264.7 macrophages
In this study, 20 new derivatives of caffeic acid esters were synthesized and their inhibitory activities against the lipopolysaccharide (LPS)-induced nitric oxide (NO) production in RAW264.7 macrophages were determined. Compounds 3l, 3r, 3s and 3t were found to decrease nitrite levels in a dose-dependent manner in LPS-induced cells and showed potent inhibitory activities against the NO production in RAW264.7 macrophages with IC50 values of 7.4, 5.9, 3.3 and 2.2 ?M, respectively. They could be selected as compromising compounds for the later pharmacological study.
Zhang, Jie; Xu, Liu-Xin; Xu, Xu-Sheng; Li, Bo-Wei; Wang, Rui; Fu, Jian-Jun
Plumbagin inhibits LPS-induced inflammation through the inactivation of the nuclear factor-kappa B and mitogen activated protein kinase signaling pathways in RAW 264.7 cells.
Plumbagin (PL) has been reported to exhibit anti-carcinogenic, anti-inflammatory and analgesic activities, but little is known about its mechanism. In this study, we investigated the anti-inflammatory property of PL and its mechanism of action. Although no significant cytotoxicity of PL was observed over the concentration range tested, PL (2.5-7.5 ?M) significantly and dose-dependently suppressed the secretion of pro-inflammatory mediators and inhibited the expression of TNF-?, IL-1?, IL-6 and iNOS in LPS-stimulated RAW 264.7 cells. Furthermore, PL consistently suppressed the activity of iNOS in LPS-induced RAW 264.7 cells. To elucidate the mechanism underlying the anti-inflammatory activity of PL, we assessed the effects of PL on the MAPK pathway and the activity and expression of NF-?B. These experiments demonstrated that PL significantly reduced the luciferase activity of an NF-?B promoter reporter and p65 nuclear translocation. The LPS-induced phosphorylation of MAP kinases was also attenuated by PL; significant changes were observed in the levels of phosphorylated ERK1/2, JNK and p38 MAPK. Additionally, MAPK inhibitors confirmed the inhibitory effect of PL on the MAPK pathway. Taken together, these data suggest that PL exerts its anti-inflammatory effects by down-regulating the expression of pro-inflammatory mediators through inhibition of NF-?B and MAPK signaling in LPS-stimulated RAW 264.7 cells. PMID:24296134
Wang, Tingyu; Wu, Feihua; Jin, Zhigui; Zhai, Zanjing; Wang, Yugang; Tu, Bing; Yan, Wei; Tang, Tingting
Gomisin A decreases the LPS-induced expression of iNOS and COX-2 and activation of RIP2/NF-?B in mouse peritoneal macrophages.
Abstract Gomisin A (GA), a lignan component contained in the fruit of Schisandra chinensis Baillon, improves hepatic cell degeneration, vasodilatory activity and insulin sensitivity. These effects also impact the immune system, including various inflammatory mediators and cytokines. In this study, the anti-inflammatory effect of GA on lipopolysaccharide-stimulated mouse peritoneal macrophages was studied. Pretreatment with GA attenuated the expression of receptor-interacting protein 2 (RIP2) and I?B kinase-? (IKK-?) as well as IKK-? phosphorylation. The activation of nuclear factor-kappa B (NF-?B) in the nucleus, the phosphorylation of I?B? and degradation of I?B? in the cytosol were suppressed by GA. GA decreased the production and mRNA expression of the inflammatory cytokines tumor necrosis factor-alpha (TNF-?) and interleukin (IL)-6. In addition, expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and production of nitric oxide were decreased by pretreatment with GA. In conclusion, these results show that the anti-inflammatory properties of GA potentially result from the inhibition of COX-2, iNOS, IL-6, TNF-? and NO through the down-regulation of RIP2 and NF-?B activation. These results impact the development of potential health products for preventing and treating inflammatory diseases. PMID:24749675
Jeong, Hyun-Ja; Han, Na-Ra; Kim, Kyu-Yeob; Choi, Il-Sook; Kim, Hyung-Min
Evaluation of natural products on inhibition of inducible cyclooxygenase (COX2) and nitric oxide synthase (iNOS) in cultured mouse macrophage cells
The inhibitors of prostaglandin biosynthesis and nitric oxide production have been considered as potential anti-inflammatory and cancer chemopreventive agents. In this study, we evaluated approximately 170 methanol extracts of natural products including Korean herbal medicines for the inhibition of prostaglandin E2 production (for COX-2 inhibitors) and nitric oxide formation (for iNOS inhibitors) in lipopolysaccharide (LPS)-induced mouse macrophages RAW264.7 cells. As
Chae Hee Hong; Sun Kyung Hur; O-Jin Oh; Sun Sook Kim; Kyung Ae Nam; Sang Kook Lee
Dietary Blue Pigments Derived from Genipin, Attenuate Inflammation by Inhibiting LPS-Induced iNOS and COX-2 Expression via the NF-?B Inactivation
Background and Purpose The edible blue pigments produced by gardenia fruits have been used as value-added colorants for foods in East Asia for 20 years. However, the biological activity of the blue pigments derived from genipin has not been reported. Methodology/Principal Findings The anti-inflammatory effect of blue pigments was studied in lipopolysaccharide (LPS) stimulated RAW 264.7 macrophage in vitro. The secretions of nitric oxide (NO) and prostaglandin E2 (PGE2) were inhibited in concentration-dependent manner by blue pigments. Real-time reverse-transcription polymerase chain reaction (Real-time RT-PCR) analyses demonstrated that the mRNA expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin (IL)-6, and tumor necrosis factor alpha (TNF-?) was inhibited, moreover, ELISA results showed that the productions of IL-6 and TNF-? were inhibited. Cell-based ELISA revealed the COX-2 protein expression was inhibited. The proteome profiler array showed that 12 cytokines and chemokines involved in the inflammatory process were down-regulated by blue pigments. Blue pigments inhibited the nuclear transcription factor kappa-B (NF-?B) activation induced by LPS, and this was associated with decreasing the DNA-binding activity of p65 and p50. Furthermore, blue pigments suppressed the degradation of inhibitor of ?B (I?B) ?, Inhibitor of NF-?B Kinase (IKK) ?, IKK-?, and phosphorylation of I?B-?. The anti-inflammatory effect of blue pigments in vivo was studied in carrageenan-induced paw edema and LPS-injecting ICR mice. Finally, blue pigments significantly inhibited paw swelling and reduced plasma TNF-? and IL-6 production in vivo. Conclusions and Implications These results suggest that the anti-inflammatory properties of blue pigments might be the results from the inhibition of iNOS, COX-2, IL-6, IL-1?, and TNF-? expression through the down-regulation of NF-?B activation, which will provide strong scientific evidence for the edible blue pigments to be developed as a new health-enhancing nutritional food for the prevention and treatment of inflammatory diseases.
Wang, Qiang-Song; Xiang, Yaozu; Cui, Yuan-Lu; Lin, Ke-Ming; Zhang, Xin-Fang
The protein kinase PKR is critical for LPS-induced iNOS production but dispensable for inflammasome activation in macrophages.
Inflammasomes are multi-protein platforms that drive the activation of caspase-1 leading to the processing and secretion of biologically active IL-1? and IL-18. Different inflammasomes including NOD-like receptor (NLR) family pyrin domain-containing 3 (NLRP3), NLR caspase-recruitment domain-containing 4 (NLRC4) and absent in melanoma 2 (AIM2) are activated and assembled in response to distinct microbial or endogenous stimuli. However, the mechanisms by which upstream stimuli trigger inflammasome activation remain poorly understood. Double-stranded RNA-activated protein kinase (PKR), a protein kinase activated by viral infection, has been recently shown to be required for the activation of the inflammasomes. Using macrophages from two different mouse strains deficient in PKR, we found that PKR is important for the induction of the inducible nitric oxide synthase (iNOS). However, PKR was dispensable for caspase-1 activation, processing of pro-IL-1?/IL-18 and secretion of IL-1? induced by stimuli that trigger the activation of NLRP3, NLRC4 and AIM2. These results indicate that PKR is not required for inflammasome activation in macrophages. PMID:23401008
He, Yuan; Franchi, Luigi; N��ez, Gabriel
Anti-inflammatory effects of anthocyanins-rich extract from bilberry (Vaccinium myrtillus L.) on croton oil-induced ear edema and Propionibacterium acnes plus LPS-induced liver damage in mice.
Abstract Bilberry (Vaccinium myrtillus L.) has been known to play a protective role in human health due to its high anthocyanin content. This study investigated the anti-inflammatory effects of bilberry extract (BE, containing 42.04% anthocyanin) on Propionibacterium acnes (P. acnes) plus lipopolysaccharide (LPS) induced liver injury and croton oil-induced ear edema in mice. Results showed that BE could effectively inhibit croton oil-induced ear edema and liver inflammation provoked by P. acnes plus LPS, as reflected by the reduced plasma alanine aminotransferase and aspartate aminotransferase activities. These findings were confirmed by hepatic pathological examination. Moreover, BE administration markedly suppressed the increase of liver mRNA levels of iNOS, TNF-?, IL-1? and IL-6, and the protein levels of iNOS, TNF-? and NF-?B. In addition, liver malondialdehyde and NO contents were significantly reduced by BE treatment. These results indicated that BE has potent protective effects on acute and immunological inflammation, which might contribute to the study of the anti-inflammatory effects of natural products and healthy food. PMID:24548119
Luo, Hui; Lv, Xiao-Dan; Wang, Guo-En; Li, Yi-Fang; Kurihara, Hiroshi; He, Rong-Rong
Helminth excreted/secreted antigens repress expression of LPS-induced Let-7i but not miR-146a and miR-155 in human dendritic cells.
MicroRNAs have emerged as key regulators of immune responses. They influence immune cells' function and probably the outcome of several infections. Currently, it is largely unknown if helminth parasites and their antigens modify host microRNAs expression. The aim of this study was to explore if excreted/secreted antigens of Taenia crassiceps regulate LPS-induced miRNAs expression in human dendritic cells. We found that these antigens repressed LPS-let-7i induction but not mir-146a or mir-155 and this correlates with a diminished inflammatory response. This let-7i downregulation in dendritic cells constitutes a novel feature of the modulatory activity that helminth-derived antigens exert on their host. PMID:23509825
Terrazas, Luis I; S�nchez-Mu�oz, Fausto; P�rez-Miranda, Magaly; Mej�a-Dom�nguez, Ana M; Ledesma-Soto, Yadira; Bojalil, Rafael; G�mez-Garc�a, Lorena
Aldose Reductase Mediates Endotoxin-Induced Production of Nitric oxide and Cytotoxicity in Murine Macrophages.
Aldose reductase (AR) is a ubiquitously expressed protein with pleiotrophic roles as an efficient catalyst for the reduction of toxic lipid aldehydes and mediator of hyperglycemia, cytokine and growth factor �induced redox sensitive signals that cause secondary diabetic complications. Although AR inhibition has been shown to be protective against oxidative stress signals, the role of AR in regulating nitric oxide (NO) synthesis and NO-mediated apoptosis has not been elucidated to date. We therefore investigated the role of AR in regulating lipopolysaccharide (LPS)-induced NO synthesis and apoptosis in RAW 264.7 macrophages. Inhibition or RNA interference ablation of AR suppressed LPS-stimulated production of NO and over-expression of iNOS mRNA. Inhibition or ablation of AR also prevented the LPS-induced apoptosis, cell cycle arrest, activation of caspase-3, p38-MAPK, JNK, NF-?B and AP1. In addition, AR inhibition prevented the LPS-induced down-regulation of Bcl-xl and up-regulation of Bax and Bak in macrophages. L-arginine increased and L-NAME decreased the severity of cell death caused by LPS and AR inhibitors prevented it. Furthermore, inhibition of AR prevents cell death caused by HNE and GS-HNE, but not GS-DHN. Our findings for the first time suggest that AR catalyzed lipid aldehyde-glutathione conjugates regulates the LPS-induced production of inflammatory marker NO and cytotoxicity in RAW 264.7 cells. Inhibition or ablation of AR activity may be potential therapeutic target in endotoximia and other inflammatory diseases.
Ramana, Kota V; Reddy, Aramati BM.; Tammali, Ravinder; Srivastava, Satish K.
Protein Kinase C ? (PKC?)-Extracellular Signal-regulated Kinase 1/2 (ERK1/2) Signaling Cascade Regulates Glycogen Synthase Kinase-3 (GSK-3) Inhibition-mediated Interleukin-10 (IL-10) Expression in Lipopolysaccharide (LPS)-induced Endotoxemia*
Glycogen synthase kinase-3 (GSK-3) modulates a wide array of cellular processes, including embryonic development, cell differentiation, survival, and apoptosis. Recently, it was reported that a GSK-3 inhibitor attenuates lipopolysaccharide (LPS)-induced septic shock and regulates the mortality of endotoxemic mice. However, the detailed mechanism of reduced mortality via GSK-3 inhibition is not well defined. Herein, we showed that GSK-3 inhibition induces extracellular signal-regulated kinase 1/2 (ERK1/2) activation under LPS-stressed conditions via protein kinase C ? (PKC?) activation. Furthermore, PKC?-induced ERK1/2 activation by the inhibition of GSK-3 provoked the production of interleukin (IL)-10, playing a crucial role in regulating endotoxemia. Using a mitogen-activated protein kinase kinase-1 (MEK-1) and PKC? inhibitor, we confirmed that GSK-3 inhibition induces PKC? and subsequent ERK1/2 activation, resulting in increased IL-10 expression under LPS-treated conditions. We verified that septic shock caused by LPS is attenuated by GSK-3 inhibition using a GSK-3 inhibitor. This relieved endotoxemia induced by GSK-3 inhibition was restored in an ERK1/2-dependent manner. Taken together, IL-10 expression produced by GSK-3 inhibition-induced ERK1/2 activation via PKC? relieved LPS-mediated endotoxemia. This finding suggests that IL-10 hyperexpression resulting from GSK-3 inhibition-induced ERK activation could be a new therapeutic pathway for endotoxemia.
Noh, Kyung Tae; Son, Kwang Hee; Jung, In Duk; Kang, Hyun Kyu; Hwang, Sun Ae; Lee, Won Suk; You, Ji Chang; Park, Yeong-Min
Intraislet release of interleukin 1 inhibits beta cell function by inducing beta cell expression of inducible nitric oxide synthase
Cytokines, released in and around pancreatic islets during insulitis, have been proposed to participate in beta-cell destruction associated with autoimmune diabetes. In this study we have evaluated the hypothesis that local release of the cytokine interleukin 1 (IL-1) by nonendocrine cells of the islet induce the expression of inducible nitric oxide synthase (iNOS) by beta cells which results in the inhibition of beta cell function. Treatment of rat islets with a combination of tumor necrosis factor (TNF) and lipopolysaccharide (LPS), conditions known to activate macrophages, stimulate the expression of iNOS and the formation of nitrite. Although TNF+LPS induce iNOS expression and inhibit insulin secretion by intact islets, this combination does not induce the expression of iNOS by beta or alpha cells purified by fluorescence activated cell sorting (Facs). In contrast, IL-1 beta induces the expression of iNOS and also inhibits insulin secretion by both intact islets and Facs-purified beta cells, whereas TNF+LPS have no inhibitory effects on insulin secretion by purified beta cells. Evidence suggests that TNF+LPS inhibit insulin secretion from islets by stimulating the release of IL-1 which subsequently induces the expression of iNOS by beta cells. The IL-1 receptor antagonist protein completely prevents TNF+LPS-induced inhibition of insulin secretion and attenuates nitrite formation from islets, and neutralization of IL-1 with antisera specific for IL-1 alpha and IL-1 beta attenuates TNF+LPS-induced nitrite formation by islets. Immunohistochemical localization of iNOS and insulin confirm that TNF+LPS induce the expression of iNOS by islet beta cells, and that a small percentage of noninsulin-containing cells also express iNOS. Local release of IL-1 within islets appears to be required for TNF+LPS-induced inhibition of insulin secretion because TNF+LPS do not stimulate nitrite formation from islets physically separated into individual cells. These findings provide the first evidence that a limited number of nonendocrine cells can release sufficient quantities of IL-1 in islets to induce iNOS expression and inhibit the function of the beta cell, which is selectively destroyed during the development of autoimmune diabetes.
The nitric oxide hypothesis of aging
�Nitric oxide (NO), generated by endothelial (e) NO synthase (NOS) and neuronal (n) NOS, plays a ubiquitous role in the body in controlling the function of almost every, if not every, organ system. Bacterial and viral products, such as bacterial lipopolysaccharide (LPS), induce inducible (i) NOS synthesis that produces massive amounts of NO toxic to the invading viruses and bacteria,
S. M McCann; J Licinio; M.-L Wong; W. H Yu; S Karanth; V Rettorri
Dietary Blue Pigments Derived from Genipin, Attenuate Inflammation by Inhibiting LPS-Induced iNOS and COX2 Expression via the NF-?B Inactivation
Background and PurposeThe edible blue pigments produced by gardenia fruits have been used as value-added colorants for foods in East Asia for 20 years. However, the biological activity of the blue pigments derived from genipin has not been reported.Methodology\\/Principal FindingsThe anti-inflammatory effect of blue pigments was studied in lipopolysaccharide (LPS) stimulated RAW 264.7 macrophage in vitro. The secretions of nitric
Qiang-Song Wang; Yaozu Xiang; Yuan-Lu Cui; Ke-Ming Lin; Xin-Fang Zhang
Pepsin-pancreatin protein hydrolysates from extruded amaranth inhibit markers of atherosclerosis in LPS-induced THP-1 macrophages-like human cells by reducing expression of proteins in LOX-1 signaling pathway
Background Atherosclerosis is considered a progressive disease that affects arteries that bring blood to the heart, to the brain and to the lower end. It derives from endothelial dysfunction and inflammation, which play an important role in the thrombotic complications of atherosclerosis. Cardiovascular disease is the leading cause of death around the world and one factor that can contribute to its progression and prevention is diet. Our previous study found that amaranth hydrolysates inhibited LPS-induced inflammation in human and mouse macrophages by preventing activation of NF-?B signaling. Furthermore, extrusion improved the anti-inflammatory effect of amaranth protein hydrolysates in both cell lines, probably attributed to the production of bioactive peptides during processing. Therefore, the objective of this study was to compare the anti-atherosclerotic potential of pepsin-pancreatin hydrolysates from unprocessed and extruded amaranth in THP-1 lipopolysaccharide-induced human macrophages and suggest the mechanism of action. Results Unprocessed amaranth hydrolysate (UAH) and extruded amaranth hydrolysate (EAH) showed a significant reduction in the expression of interleukin-4 (IL-4) (69% and 100%, respectively), interleukin-6 (IL-6) (64% and 52%, respectively), interleukin-22 (IL-22) (55% and 70%, respectively). Likewise, UAH and EAH showed a reduction in the expression of monocyte-chemo attractant protein-1 (MCP-1) (35% and 42%, respectively), transferrin receptor-1 (TfR-1) (48% and 61%, respectively), granulocyte-macrophage colony-stimulating factor (GM-CSF) (59% and 63%, respectively), and tumor necrosis factor-? (TNF-?) (60% and 63%, respectively). Also, EAH reduced the expression of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) (27%), intracellular adhesion molecule-1 (ICAM-1) (28%) and matrix metalloproteinase-9 (MMP-9) (19%), important molecular markers in the atherosclerosis pathway. EAH, led to a reduction of 58, 52 and 79% for LOX-1, ICAM-1 and MMP-9, respectively, by confocal microscopy. Conclusions Extruded amaranth hydrolysate showed potential anti-atherosclerotic effect in LPS-induced THP-1 human macrophage-like cells by reducing the expression of proteins associated with LOX-1 signaling pathway.
Lucidone inhibits iNOS and COX-2 expression in LPS-induced RAW 264.7 murine macrophage cells via NF-kappaB and MAPKs signaling pathways.
The anti-inflammatory mechanism of lucidone isolated from the fruits of Lindera erythrocarpa Makino was investigated. Our data indicate that lucidone significantly inhibits the production of NO and PGE(2) autacoids in LPS-induced RAW 264.7 murine macrophage cells. Moreover, it also notably decreased the secretion of tumor necrosis factor-alpha (TNF-alpha). Consistent with these observations, the mRNA and protein expression levels of iNOS and COX-2 were also inhibited by lucidone in a dose-dependent manner. Lucidone also reduced the translocation of NF-kappaB induced by LPS, which is associated with the prevention of the degradation of I-kappaB, and subsequently decreased p65/p50 protein levels in the nucleus. Lucidone also inhibited NF-kappaB activation by impairing the binding of NF-kappaB to its cis-acting element. In addition, lucidone inhibited JNK and p38MAPKs signals, which are the most significant signals involved in NO, PGE(2) and TNF-alpha production; NF-kappaB/AP-1 activation was also inhibited by lucidone. Taken together, the anti-inflammatory activity of lucidone might be caused by the inhibition of iNOS and COX-2 expressions through downregulation of NF-kappaB and AP-1 binding. PMID:19194838
Senthil Kumar, K J; Wang, Sheng-Yang
Differential migration, LPS-induced cytokine, chemokine and NO expression in immortalized BV-2 and HAPI cell lines and primary microglial cultures
Microglial cells are hematopoietically derived monocytes of the CNS and serve important neuromodulatory, neurotrophic and neuroimmune roles. Following insult to the CNS, microglia develop a reactive phenotype, migrate to the site of injury, proliferate, and release a range of proinflammatory, anti-inflammatory and neurotrophic factors. Isolation of primary microglial cell cultures has been an integral step in elucidating the many roles of these cells. In addition to primary microglial cells, several immortalized cell lines have been created to model primary microglia in vitro, including murine derived BV-2 cells and rat derived HAPI cells. Here we compare rat primary microglial, BV-2 and HAPI cells in experiments assessing migration, expression of activation markers and production and release of NO (nitric oxide), cytokines and chemokines. BV-2 and HAPI cells responded similarly to primary microglia in experiments assessing migration, Iba1 expression, and NO release. However, BV-2 and HAPI cells did not model primary microglia in experiments assessing TNF?, IL-1?, IL-6 and MCP-1 expression and release and pERK 44/42 (extracellular receptor kinase) expression following LPS treatment. These results indicate that BV-2 and HAPI cell cultures only partially model primary microglia and that their use should therefore be carefully considered.
Horvath, Ryan J.; Nutile-McMenemy, Nancy; Alkaitis, Matthew S.; De Leo, Joyce A.
Rhynchophylline attenuates LPS-induced pro-inflammatory responses through down-regulation of MAPK/NF-?B signaling pathways in primary microglia.
Excessive activation of microglial cells has been implicated in various types of neuroinflammation. Suppression of microglial activation would have therapeutic benefits, leading to the alleviation of the progression of neurodegeneration. In this study, the inhibitory effects of rhynchophylline (RIN), a tetracyclic oxindole alkaloid component isolated from Uncaria rhynchophylla (Miq.) Jacks., on the production of pro-inflammatory mediators were investigated in lipopolysaccharide (LPS)-stimulated microglia. The results showed that RIN markedly reduced the production of nitric oxide (NO), prostaglandins E(2) (PGE(2) ), monocyte chemoattractant protein (MCP-1), tumor necrosis factor-? (TNF-?) and interleukin-1? (IL-1?) in LPS-activated microglia. The mRNA expression levels of iNOS and COX-2 were also depressed by RIN in a concentration-dependent manner. Further studies revealed that RIN blocked I?B? phosphorylation and degradation, inhibited the phosphorylation of mitogen-activated protein kinases (MAPKs). In summary, these data suggest that RIN suppresses inflammatory responses of microglia and may act as a potential therapeutic agent for various neurodegenerative diseases involving neuroinflammation. PMID:22322985
Song, Yu; Qu, Rong; Zhu, Shenghua; Zhang, Ruiguo; Ma, Shiping
SGLT-1-mediated glucose uptake protects intestinal epithelial cells against LPS-induced apoptosis and barrier defects: a novel cellular rescue mechanism?
Excessive apoptosis induced by enteric microbes leads to epithelial barrier defects. This mech- anism has been implicated in the pathogenesis of inflammatory bowel diseases (IBD) and bacterial enter- itis. The sodium-dependent glucose cotransporter (SGLT-1) is responsible for active glucose uptake in enterocytes. The aim was to investigate the effects of SGLT-1 glucose uptake on enterocyte apoptosis and barrier defects induced
Linda C. H. Yu; Andrew N. Flynn; Jerrold R. Turner; Andre G. Buret
A Role of Cell Apoptosis in Lipopolysaccharide (LPS)-induced Nonlethal Liver Injury in d -galactosamine ( d -GalN)-sensitized Rats
Lipopolysaccharide (LPS) is implicated in the pathology of acute liver injury and can induce lethal liver failure when simultaneously\\u000a administered with d-galactosamine (d-GalN). At the present time, nonlethal liver failure, the liver injury of clinical implication, is incompletely understood\\u000a following challenge by low-dose LPS\\/d-GalN. We report here our investigation of the effects of liver injury following a nonlethal dose LPS\\/d-GalN
Liang-Ming Liu; Ji-Xiang Zhang; Jie Luo; Hong-Xing Guo; Huan Deng; Jian-Yong Chen; Sui-Lin Sun
Differential Inhibition by Nimesulide of the Early and Late Phases of Intravenous and Intracerebroventricular-LPS-Induced Fever in Guinea Pigs
Objectives: The findings that inducible cyclooxygenase (COX)-2, but not constitutive COX-1, is upregulated in the brain of conscious rats ?1.5 h after intraperitoneal pyrogen administration, that the systemic administration of COX-2 inhibitors abolishes fever, and that COX-2-deficient mice do not develop fever in response to intraperitoneal lipopolysaccharide (LPS) have strongly implicated COX-2 in the mediation of the febrile response. However,
Alexandre A. Steiner; Shuxin Li; Clark M. Blatteis
Novel Mechanism of Attenuation of LPS-Induced NF-?B Activation by the Heat Shock Protein 90 Inhibitor, 17-N-allylamino-17-demethoxygeldanamycin, in Human Lung Microvascular Endothelial Cells.
Heat shock protein (hsp) 90 inhibition attenuates NF-?B activation and blocks inflammation. However, the precise mechanism of NF-?B regulation by hsp90 in the endothelium is not clear. We investigated the mechanisms of hsp90 inhibition by 17-N-allylamino-17-demethoxygeldanamycin (17-AAG) on NF-?B activation by LPS in primary human lung microvascular endothelial cells. Transcriptional activation of NF-?B was measured by luciferase reporter assay, gene expression by real-time RT-PCR, DNA binding of transcription factors by chromatin immunoprecipitation assay, protein-protein interaction by coimmunoprecipitation/immunoblotting, histone deacetylase (HDAC)/histone acetyltransferase enzyme activity by fluorometry, and nucleosome eviction by partial microccocal DNase digestion. In human lung microvascular endothelial cells, 17-AAG-induced degradation of IKB? was accomplished regardless of the phosphorylation/ubiquitination state of the protein. Hence, 17-AAG did not block LPS-induced NF-?B nuclear translocation and DNA binding activity. Instead, 17-AAG blocked the recruitment of the coactivator, cAMP response element binding protein binding protein, and prevented the assembly of a transcriptionally competent RNA polymerase II complex at the ?B elements of the IKB? (an NF-?B-responsive gene) promoter. The effect of LPS on IKB? mRNA expression was associated with rapid deacetylation of histone-H3(Lys9) and a dramatic down-regulation of core histone H3 binding. Even though treatment with an HDAC inhibitor produced the same effect as hsp90 inhibition, the effect of 17-AAG was independent of HDAC. We conclude that hsp90 inhibition attenuates NF-?B transcriptional activation by preventing coactivator recruitment and nucleosome eviction from the target promoter in human lung endothelial cells. PMID:24303801
Thangjam, Gagan S; Dimitropoulou, Chistiana; Joshi, Atul D; Barabutis, Nektarios; Shaw, Mary C; Kovalenkov, Yevgeniy; Wallace, Chistopher M; Fulton, David J; Patel, Vijay; Catravas, John D
Quince (Cydonia oblonga Miller) peel polyphenols modulate LPS-induced inflammation in human THP-1-derived macrophages through NF-?B, p38MAPK and Akt inhibition.
Chronic inflammation is a hallmark of several pathologies, such as rheumatoid arthritis, gastritis, inflammatory bowel disease, atherosclerosis and cancer. A wide range of anti-inflammatory chemicals have been used to treat such diseases while presenting high toxicity and numerous side effects. Here, we report the anti-inflammatory effect of a non-toxic, cost-effective natural agent, polyphenolic extract from the Tunisian quince Cydonia oblonga Miller. Lipopolysaccharide (LPS) treatment of human THP-1-derived macrophages induced the secretion of high levels of the pro-inflammatory cytokine TNF-? and the chemokine IL-8, which was inhibited by quince peel polyphenolic extract in a dose-dependent manner. Concomitantly, quince polyphenols enhanced the level of the anti-inflammatory cytokine IL-10 secreted by LPS-treated macrophages. We further demonstrated that the unexpected increase in IL-6 secretion that occurred when quince polyphenols were associated with LPS treatment was partially responsible for the polyphenols-mediated inhibition of TNF-? secretion. Biochemical analysis showed that quince polyphenols extract inhibited the LPS-mediated activation of three major cellular pro-inflammatory effectors, nuclear factor-kappa B (NF-?B), p38MAPK and Akt. Overall, our data indicate that quince peel polyphenolic extract induces a potent anti-inflammatory effect that may prove useful for the treatment of inflammatory diseases and that a quince-rich regimen may help to prevent and improve the treatment of such diseases. PMID:22252293
Essafi-Benkhadir, Khadija; Refai, Amira; Riahi, Ichrak; Fattouch, Sami; Karoui, Habib; Essafi, Makram
Inhibitory effects of the beta-adrenergic receptor agonist zilpaterol on the LPS-induced production of TNF-alpha in vitro and in vivo.
In this study the anti-inflammatory properties of zilpaterol, a beta2-adrenergic receptor (AR) agonist specifically developed as a growth promoter in cattle were investigated. Although zilpaterol has a different structure compared with the beta2-AR agonists known to date, it was noted that it was able to bind to both the beta2-AR (Ki = 1.1 x 10(-6)) and the beta1-AR (Ki = 1.0 x 10(-5)). Using lipopolysaccharide (LPS)-exposed U937 macrophages, the production of cyclic adenosine-3',5'-cyclic monophosphate (cAMP) and tumor necrosis factor alpha (TNF-alpha) were investigated. Zilpaterol inhibited TNF-alpha release and induced intracellular cAMP levels in a dose-dependent manner. The inhibition of TNF-alpha release and induction of cAMP production was mainly mediated via the beta2-AR, as indicated by addition of beta1- and beta2-specific antagonists. The effects of zilpaterol were investigated in LPS-treated male Wistar rats after pretreatment with zilpaterol. Zilpaterol dosed at 500 microg/kg body weight reduced the TNF-alpha plasma levels. In conclusion, zilpaterol is a beta2-adrenergic agonist and an inhibitor of TNF-alpha production induced by LPS both in vivo and in vitro. PMID:16343285
Verhoeckx, K C M; Doornbos, R P; van der Greef, J; Witkamp, R F; Rodenburg, R J T
New phenylpropanoid and other compounds from Illicium lanceolatum with inhibitory activities against LPS-induced NO production in RAW 264.7 macrophages.
Illicium lanceolatum is a traditional Chinese medicine (TCM) for treating inflammatory diseases. Anti-inflammatory activities of I. lanceolatum stems and leaves were tested using ear edema models induced by dimethyl benzene in mice. Bioassay-guided fractionation of the ethanol extract of I. lanceolatum leaves and stems revealed that the ethyl acetate fraction exhibited inhibitory potency to dimethyl benzene-induced edema in the mouse ear. Phytochemical investigation on the active fraction led to the isolation of a new phenylpropanoid (1), together with fifteen known compounds. This is the first report of the isolation of 2-16 from I. lanceolatum. Of these compounds, compounds 1, 2 and 3 showed inhibitory activity on LPS-stimulated NO production in RAW 264.7 macrophages with IC50 values of 27.58, 26.59 and 34.35 ?g/mL, respectively. I. lanceolatum stems and leaves can be exploited to alleviate inflammatory diseases, which makes the rare medicinal plant resources sustainable. PMID:24613803
Gui, Xuan; Wang, Guowei; Zhang, Naidan; Huang, Baokang
Differential protection among fractionated blueberry polyphenolic families against DA-, A?42 and LPS-induced decrements in Ca2+ buffering in primary hippocampal cells
It has been postulated that at least part of the loss of cognitive function in aging may be the result of deficits in Ca2+ recovery (CAR) and increased oxidative/inflammatory (OX/INF) stress signaling. However, previous research showed that aged animals supplemented with blueberry (BB) extract, showed fewer deficits in CAR, as well as motor and cognitive functional deficits. A recent subsequent experiment has shown that DA- or A?42-induced deficits in CAR in primary hippocampal neuronal cells (HNC) were antagonized by BB extract, and (OX/INF) signaling was reduced. Present experiments assessed the most effective BB polyphenol fraction that could protect against OX/INF-induced deficits in CAR, ROS generation, or viability. HNCs treated with BB extract, BB fractions (e.g., proanthocyanidin, PAC), or control medium were exposed to dopamine (DA, 0.1mM), amyloid beta (A?42, 25 �M) or lipopolysaccharide (LPS, 1�g/ml). Results indicated that the degree of protection against deficits in CAR varied as a function of the stressor and was generally greater against A?42 and LPS than DA. The whole BB, anthocyanin (ANTH) and pre-C18 fractions offered the greatest protection, while chlorogenic acid offered the lowest protection. Protective capabilities of the various fractions against ROS depended upon the stressor, where the BB extract and the combined PAC (high and low m.w.) fraction offered the best protection against LPS and A?42 but were less effective against DA-induced ROS. The high and low m.w. PACs and the ANTH fractions enhanced ROS production regardless of the stressor used and this reflected increased activation of stress signals (e.g., P38 MAPK). The viability data indicated that the whole BB and combined PAC fraction showed greater protective effects against the stressors than the more fractionated polyphenolic components. Thus, these results suggest that, except for a few instances, the lesser the polyphenolic fractionation the greater the effects, especially with respect to prevention of ROS and stress signal generation, and viability.
Joseph, James A.; Shukitt-Hale, Barbara; Brewer, Gregory J.; Weikel, Karen A.; Kalt, Wilhelmina; Fisher, Derek R.
MR imaging and targeting of a specific alveolar macrophage subpopulation in LPS-induced COPD animal model using antibody-conjugated magnetic nanoparticles
Purpose Targeting and noninvasive imaging of a specific alveolar macrophage subpopulation in the lung has revealed the importance for early and better diagnosis and therapy of chronic obstructive pulmonary disease (COPD). In this study, the in vivo effect of pulmonary administration of iron oxide nanoparticles on the polarization profile of macrophages was assessed, and a noninvasive free-breathing magnetic resonance imaging (MRI) protocol coupled with the use of biocompatible antibody-conjugated superparamagnetic iron oxide (SPIO) nanoparticles was developed to enable specific targeting and imaging of a particular macrophage subpopulation in lipopolysaccharide-induced COPD mice model. Materials and methods Enzyme-linked immunosorbent assay, Real-time polymerase chain reaction, and flow cytometry analysis were performed to assess the biocompatibility of PEGylated dextran-coated SPIO nanoparticles. Specific biomarkers for M1 and M2 macrophages subsets were selected for conjugation with magnetic nanoparticles. MRI protocol using ultra-short echo time sequence was optimized to enable simultaneous detection of inflammation progress in the lung and detection of macrophages subsets. Flow cytometry and immunohistochemistry analysis were finally performed to confirm MRI readouts and to characterize the polarization profile of targeted macrophages. Results The tested SPIO nanoparticles, under the current experimental conditions, were found to be biocompatible for lung administration in preclinical settings. Cluster of differentiation (CD)86- and CD206-conjugated magnetic nanoparticles enabled successful noninvasive detection of M1 and M2 macrophage subpopulations, respectively, and were found to co-localize with inflammatory regions induced by lipopolysaccharide challenge. No variation in the polarization profile of targeted macrophages was observed, even though a continuum switch in their polarization might occur. However, further confirmatory studies are required to conclusively establish this observation. Conclusion Coupling of magnetic iron oxide nanoparticles with a specific antibody targeted to a particular macrophage subpopulation could offer a promising strategy for an early and better diagnosis of pulmonary inflammatory diseases using noninvasive MRI.
Al Faraj, Achraf; Shaik, Asma Sultana; Afzal, Sibtain; Al Sayed, Baraa; Halwani, Rabih
Effect of 635 nm irradiation on high glucose-boosted inflammatory responses in LPS-induced MC3T3-E1 cells.
Hyperglycemia occurs in patients with poorly controlled diabetes mellitus and contributes to bone resorption and increased susceptibility to bacterial infections. Hyperglycemia can incite low-grade inflammation that can contribute to the resorption of bone, especially the periodontal bone. The increased susceptibility to periodontal infections can contribute to bone resorption through the activation of osteoclasts. In this study, the osteoblastic, clonal cell line, MC3T3-E1, was used in an in vitro model of hyperglycemia and lipopolysaccharide-induced reactive oxygen species generation to determine the potential anti-inflammatory effect of 635 nm light-emitting diode (LED) irradiation or whether 635 nm LED irradiation can be a potential anti-inflammatory treatment. LED irradiation of MC3T3-E1 cells stimulated with lipopolysaccharide in a high glucose-containing medium decreased the level of cyclooxygenase gene and protein expression and reduced the level of prostaglandin E2 expression by decreasing the amount of reactive oxygen species generation. LED irradiation also inhibited the osteoclastogenesis in MC3T3-E1 cells by regulating the receptor activator of nuclear factor kappa-B ligand and osteoprotegerin. These findings reveal the mechanisms which are important in the pathogenesis of diabetic periodontitis and highlight the beneficial effects of 635 nm LED irradiation in reducing the adverse effects of diabetic periodontitis. PMID:22699799
Kwon, HyukIl; Lim, WonBong; Kim, JiSun; Jeon, SangMi; Kim, SangWoo; Karna, Sandeep; Cha, HyunRok; Kim, OkJoon; Choi, HongRan
Analysis of LPS-Induced, NF?B-Dependent Interleukin-8 Transcription in Kidney Embryonic Cell Line Expressing TLR4 Using Luciferase Assay.
Gene expression is orchestrated by a complex network of signal transduction pathways that typically originate on cell surface receptors and culminate in DNA-binding transcription factors, which translocate to the nucleus and bind cis-regulatory elements in promoter regions of genes, thereby inducing de novo synthesis of the nascent RNA transcripts and their splicing. Gene expression arrays monitor abundance of the matured, spliced cDNA, which undergoes additional posttranscriptional modifications that greatly affect the half-life of the cDNA. Thus, the relative abundance of cDNA is not necessarily commensurable with the activity of promoters of the corresponding genes. In contrast, reporter gene assays provide valuable insight into the regulation of gene expression at the level of transcription and allow for discerning the contribution of individual transcription factors into changes in gene expression. Here, we describe a robust reporter gene assay method that is useful for exploration of transcription regulatory network, which regulates gene expression in response to inflammation. The method is exemplified by using the promoter region of the prototypic pro-inflammatory chemokine interleukin-8 (IL-8, CXCL8), which plays an important role in immune response as well as carcinogenesis. Using the luciferase reporter gene assay, we analyze the activation status of the IL-8 promoter in lipopolysaccharide (LPS)-stimulated human embryonic kidney cells. PMID:24908317
Yunusova, Tamara; Akhtar, Mumtaz; Poltoratsky, Vladimir
Fluoxetine up-regulates expression of cellular FLICE-inhibitory protein and inhibits LPS-induced apoptosis in hippocampus-derived neural stem cell
Fluoxetine is a widely used antidepressant compound which inhibits the reuptake of serotonin in the central nervous system. Recent studies have shown that fluoxetine can promote neurogenesis and improve the survival rate of neurons. However, whether fluoxetine modulates the proliferation or neuroprotection effects of neural stem cells (NSCs) needs to be elucidated. In this study, we demonstrated that 20 {mu}M fluoxetine can increase the cell proliferation of NSCs derived from the hippocampus of adult rats by MTT test. The up-regulated expression of Bcl-2, Bcl-xL and the cellular FLICE-inhibitory protein (c-FLIP) in fluoxetine-treated NSCs was detected by real-time RT-PCR. Our results further showed that fluoxetine protects the lipopolysaccharide-induced apoptosis in NSCs, in part, by activating the expression of c-FLIP. Moreover, c-FLIP induction by fluoxetine requires the activation of the c-FLIP promoter region spanning nucleotides -414 to -133, including CREB and SP1 sites. This effect appeared to involve the phosphatidylinositol-3-kinase-dependent pathway. Furthermore, fluoxetine treatment significantly inhibited the induction of proinflammatory factor IL-1{beta}, IL-6, and TNF-{alpha} in the culture medium of LPS-treated NSCs (p < 0.01). The results of high performance liquid chromatography coupled to electrochemical detection further confirmed that fluoxentine increased the functional production of serotonin in NSCs. Together, these data demonstrate the specific activation of c-FLIP by fluoxetine and indicate the novel role of fluoxetine for neuroprotection in the treatment of depression.
Chiou, S.-H. [Department of Medical Research and Education, Taipei Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan (China) and Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan (China)]. E-mail: firstname.lastname@example.com; Chen, S.-J. [Department of Ophthalmology, Taipei Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan (China)]. E-mail: email@example.com; Peng, C-H. [Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan (China); Department of Ophthalmology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Chang, Y.-L. [Department of Pharmacy, Taipei Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan (China); Ku, H.-H. [Institute of Anatomy and Cell Biology, National Yang-Ming University, Taipei, Taiwan (China); Hsu, W.-M. [Department of Ophthalmology, Taipei Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan (China); Ho, Larry L.-T. [Department of Medical Research and Education, Taipei Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan (China); Lee, C.-H. [Department of Medical Research and Education, Taipei Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan (China)
Suppression of NF-?B by dieckol extracted from Ecklonia cava negatively regulates LPS induction of inducible nitric oxide synthase gene.
Dieckol, extracted from brown algae, Ecklonia cava, is suggested to elicit anti-inflammatory or anti-tumorigenic activities. However, dieckol-mediated regulatory mechanism for inflammatory response still remains elusive. Here, we show that dieckol suppressed lipopolysaccharide (LPS)-induced inducible nitric oxide synthase (iNOS) expression in mouse leukemic macrophage Raw264.7 cells. Also, dieckol decreased LPS-induced both nitric oxide (NO) production and iNOS promoter-driven transcriptional activity in a dose-dependent manner. On the other hand, LPS-mediated NF-?B activity was inhibited by dieckol treatment. Moreover, results revealed that dieckol diminished LPS-mediated p65 nuclear translocation or I?B? phosphorylation dose-dependently, and reduced LPS-induced phosphorylation of mitogen-activated protein kinases (MAPKs), significantly p38MAPK. Collectively, these findings suggest that dieckol acts as a negative regulator of LPS-mediated iNOS induction through suppression of NF-?B activity, implying a mechanistic role of dieckol in regulation of inflammatory response. PMID:24744158
Choi, Hye-Jin; Park, Jung-Hwan; Lee, Bong Ho; Chee, Hee Youn; Lee, Kyung Bok; Oh, Sang-Muk
Microarray and Pathway Analysis Reveal Distinct Mechanisms Underlying Cannabinoid-Mediated Modulation of LPS-Induced Activation of BV-2 Microglial Cells
Cannabinoids are known to exert immunosuppressive activities. However, the mechanisms which contribute to these effects are unknown. Using lipopolysaccharide (LPS) to activate BV-2 microglial cells, we examined how ?9-tetrahydrocannabinol (THC), the major psychoactive component of marijuana, and cannabidiol (CBD) the non-psychoactive component, modulate the inflammatory response. Microarray analysis of genome-wide mRNA levels was performed using Illumina platform and the resulting expression patterns analyzed using the Ingenuity Pathway Analysis to identify functional subsets of genes, and the Ingenuity System Database to denote the gene networks regulated by CBD and THC. From the 5338 transcripts that were differentially expressed across treatments, 400 transcripts were found to be upregulated by LPS, 502 by CBD+LPS and 424 by THC+LPS, while 145 were downregulated by LPS, 297 by CBD+LPS and 149 by THC+LPS, by 2-fold or more (p?0.005). Results clearly link the effects of CBD and THC to inflammatory signaling pathways and identify new cannabinoid targets in the MAPK pathway (Dusp1, Dusp8, Dusp2), cell cycle related (Cdkn2b, Gadd45a) as well as JAK/STAT regulatory molecules (Socs3, Cish, Stat1). The impact of CBD on LPS-stimulated gene expression was greater than that of THC. We attribute this difference to the fact that CBD highly upregulated several genes encoding negative regulators of both NF?B and AP-1 transcriptional activities, such as Trib3 and Dusp1 known to be modulated through Nrf2 activation. The CBD-specific expression profile reflected changes associated with oxidative stress and glutathione depletion via Trib3 and expression of ATF4 target genes. Furthermore, the CBD affected genes were shown to be controlled by nuclear factors usually involved in regulation of stress response and inflammation, mainly via Nrf2/Hmox1 axis and the Nrf2/ATF4-Trib3 pathway. These observations indicate that CBD, and less so THC, induce a cellular stress response and that this response underlies their high immunosuppressant activities.
Juknat, Ana; Kozela, Ewa; Rimmerman, Neta; Levy, Rivka; Gao, Fuying; Coppola, Giovanni; Geschwind, Daniel; Vogel, Zvi
Comparative analysis of the acute response of the trout, O. mykiss, head kidney to in vivo challenge with virulent and attenuated infectious hematopoietic necrosis virus and LPS-induced inflammation
Background The response of the trout, O. mykiss, head kidney to bacterial lipopolysaccharide (LPS) or active and attenuated infectious hematopoietic necrosis virus (IHNV and attINHV respectively) intraperitoneal challenge, 24 and 72 hours post-injection, was investigated using a salmonid-specific cDNA microarray. Results The head kidney response to i.p. LPS-induced inflammation in the first instance displays an initial stress reaction involving suppression of major cellular processes, including immune function, followed by a proliferative hematopoietic-type/biogenesis response 3 days after administration. The viral response at the early stage of infection highlights a suppression of hematopoietic and protein biosynthetic function and a stimulation of immune response. In fish infected with IHNV a loss of cellular function including signal transduction, cell cycle and transcriptional activity 72 hours after infection reflects the tissue-specific pathology of IHNV infection. attIHNV treatment on the other hand shows a similar pattern to native IHNV infection at 24 hours however at 72 hours a divergence from the viral response is seen and replace with a recovery response more similar to that observed for LPS is observed. Conclusion In conclusion we have been able to identify and characterise by transcriptomic analysis two different types of responses to two distinct immune agents, a virus, IHNV and a bacterial cell wall component, LPS and a 'mixed' response to an attenuated IHNV. This type of analysis will lead to a greater understanding of the physiological response and the development of effective immune responses in salmonid fish to different pathogenic and pro-inflammatory agents.
MacKenzie, Simon; Balasch, Joan C; Novoa, Beatriz; Ribas, Laia; Roher, Nerea; Krasnov, Aleksei; Figueras, Antonio
Lipopolysaccharide-induced murine embryonic resorption involves nitric oxide-mediated inhibition of the NAD+-dependent 15-hydroxyprostaglandin dehydrogenase.
The initial inactivation of prostaglandins (PGs) is mediated by 15-hydroxyprostaglandin dehydrogenase (15-PGDH). PGs are potent mediators of several biological processes, including inflammation and reproduction. In uterus, PGs play a key role in infection-induced pregnancy loss, in which concentration of this mediator increased. This process is accompanied with the induction of nitric oxide synthase expression and a marked increase in uterine levels of nitric oxide. There is no information concerning nitric oxide contribution to potential changes in PG catabolism, but experimental evidence suggests that nitric oxide modulates PG pathways. The specific objectives of the study were to evaluate the protein expression of HPGD (15-PGDH) and to characterize the nitric oxide-dependent regulation of this enzyme in a model of lipopolysaccharide (LPS)-induced embryonic resorption. Results show that LPS decreased HPGD protein expression and augmented PGE synthase activity; therefore, PGE? levels increased in uterus in this inflammatory condition. Just as LPS, the treatment with a nitric oxide donor diminished HPGD protein expression in uterine tissue. In contrast, the inhibition of nitric oxide synthesis both in control and in LPS-treated mice increased 15-PGDH levels. Also, we have found that this enzyme and PGE? levels are not modulated by peroxynitrite, an oxidant agent derived from nitric oxide. This study suggests that LPS and nitric oxide promote a decrease in the ability of the uterus for PG catabolism during bacterially triggered pregnancy loss in mice. PMID:22843771
Aisemberg, Julieta; Bariani, Mar�a V; Vercelli, Claudia A; Wolfson, Manuel L; Franchi, Ana M
Inhibition of lipopolysaccharide-inducible nitric oxide synthase and IL1? through suppression of NF-?B activation by 3-(1?-1?-dimethyl-allyl)-6-hydroxy-7-methoxy-coumarin isolated from Ruta graveolens L
The Ruta graveolens L. plant is used in traditional medicine to treat a large number of diseases. The methanol (50%) extract of the whole plant was observed to inhibit the expression of inducible nitric oxide synthase (iNOS) and the cycloxygenase-2 (COX-2) gene in lipopolysaccharide (LPS)-induced macrophage cells (J774A.1, [Raghav, S.K., Gupta, B., Agrawal, C., Goswami, K., Das, H.R., 2006b. Anti-inflammatory
Sunil Kumar Raghav; Bhawna Gupta; Anju Shrivastava; Hasi Rani Das
LPS-induced inflammation in the chicken is associated with CCAAT/enhancer binding protein beta-mediated fat mass and obesity associated gene down-regulation in the liver but not hypothalamus
Background The fat mass and obesity associated gene (FTO) is widely investigated in humans regarding its important roles in obesity and type 2 diabetes. Studies in mammals demonstrate that FTO is also associated with inflammation markers. However, the association of FTO with inflammation in chickens remains unclear. In this study, male chickens on day 28 posthatching were injected intraperitoneally with lipopolysaccharide (LPS) or saline to investigate whether the FTO gene is involved in LPS-induced inflammation. Results We detected significant down-regulation of FTO mRNA in the liver (P?0.01), but not in the hypothalamus, 2 and 24�h after LPS challenge. Toll-like receptor (TLR) 2 (P?0.01) and TLR4 (P?0.01) followed the same pattern as FTO, being suppressed significantly in liver but not in hypothalamus. IL-1? was dramatically up-regulated (P?0.01) in both liver and hypothalamus 2�h after LPS challenge, while activation of IL-6 was observed in the liver (P?0.01), but not in hypothalamus. The 5?-flanking sequence of the chicken FTO gene contains nine predicted binding sites for CCAAT/enhancer binding protein beta (C/EBP beta) and one for signal transducer and activator of transcription 3 (STAT3). Significant elevation of C/EBP beta was detected in the liver (P?0.01), but not in the hypothalamus, 2�h after LPS challenge. Lipopolysaccharide challenge increased the C/EBP beta binding to FTO promoter in the liver (P?0.01 for fragment 1, P < 0.05 for fragment 2), although the protein content of C/EBP beta was not altered. Moreover, injection of LPS resulted in enhanced phosphorylation of liver STAT3, a downstream transcription factor in IL-6 signaling. Although phosphorylated STAT3 was not detected to directly bind to FTO promoter, it was found to interact with C/EBP beta. Conclusion Our results reveal that FTO expression in liver, but not in hypothalamus, is affected by the i.p. injection of LPS, which may be mediated through tissue-specific FTO transcriptional regulation by C/EBP beta and STAT3 interaction.
Bifunctional effects of fucoidan on the expression of inducible nitric oxide synthase
Algal fucoidan is a marine sulfated polysaccharide with a wide variety of biological activities including anti-thrombotic and anti-inflammatory effects. This study evaluated the effect of fucoidan on the expression of inducible nitric oxide synthase (iNOS) in a macrophage cell line, RAW264.7. Low concentration range of fucoidan (10 {mu}g/ml) increased the basal expression level of iNOS in quiescent macrophages. However, we found for the first time that fucoidan inhibited the release of nitric oxide (NO) in RAW264.7 cells stimulated with lipopolysaccharide (LPS). Western blot analysis revealed that fucoidan suppressed the LPS-induced expression of the inducible nitric oxide synthase (iNOS) gene. Moreover, the activation of both nuclear factor-{kappa}B (NF-{kappa}B) and activator protein 1 (AP-1) are key steps in the transcriptional activation of the iNOS gene. Here, it was revealed that fucoidan selectively suppressed AP-1 activation, and that the activation of AP-1 appears to be essential for the induction of iNOS in activated macrophages. This inhibitory effect on AP-1 activation by fucoidan might be associated with its NO blocking and anti-inflammatory effects.
Yang, Jin Won [College of Pharmacy, Chosun University, Gwangju (Korea, Republic of); Yoon, Se Young [College of Pharmacy, Chosun University, Gwangju (Korea, Republic of); LG Household and Healthcare Ltd., Research Park, Daejeon (Korea, Republic of); Oh, Soo Jin [College of Pharmacy and Research Center for Transgenic Cloned Pigs, Chungnam National University, Daejeon (Korea, Republic of); Kim, Sang Kyum [College of Pharmacy and Research Center for Transgenic Cloned Pigs, Chungnam National University, Daejeon (Korea, Republic of); Kang, Keon Wook [College of Pharmacy, Chosun University, Gwangju (Korea, Republic of)]. E-mail: email@example.com
Alachlor and carbaryl suppress lipopolysaccharide-induced iNOS expression by differentially inhibiting NF-{kappa}B activation
Nitric oxide (NO) produced by macrophages plays an important role in host defense and inflammation. We found that two agrochemicals, alachlor and carbaryl, inhibit lipopolysaccharide (LPS)-induced NO production by macrophages. In the present study, we investigated this inhibitory mechanism in RAW 264 cells. Both chemicals inhibited LPS-induced iNOS protein and mRNA expression as well as murine iNOS promoter activity. When treating these chemicals with reducing agents, the inhibition by carbaryl was reversed, but not the inhibition by alachlor. These chemicals also inhibited LPS-induced interferon-{beta} (IFN-{beta}) expression, an indispensable factor for LPS-induced iNOS expression. The inhibited iNOS expression, however, was not restored by exogenous IFN-{beta} supplementation. LPS-induced nuclear translocation of NF-{kappa}B, which is necessary for the expression of IFN-{beta} and iNOS, was inhibited by these chemicals: however, the LPS-induced degradation of I{kappa}B-{alpha} and I{kappa}B-{beta} was inhibited only by alachlor. These results indicate that alachlor and carbaryl differentially impair the LPS-induced NF-{kappa}B activation, leading to the inhibition of NO production.
Shimomura-Shimizu, Mifumi [Division of Microbiology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501 (Japan); Sugiyama, Kei-ichi [Division of Microbiology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501 (Japan); Muroi, Masashi [Division of Microbiology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501 (Japan); Tanamoto, Ken-ichi [Division of Microbiology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501 (Japan)]. E-mail: firstname.lastname@example.com
Enhancement of lipopolysaccharide-induced nitric oxide and interleukin-6 production by PEGylated gold nanoparticles in RAW264.7 cells
NASA Astrophysics Data System (ADS)
While the immunogenicity and cytotoxicity of gold nanoparticles (AuNPs) are noted by many researchers, the mechanisms by which AuNPs exert these effects are poorly understood. In this study, we investigated the effects of polyethylene glycolylated AuNPs (PEG@AuNPs) on lipopolysaccharide (LPS)-induced nitric oxide (NO) and interleukin-6 (IL-6) production and the associated molecular mechanism in RAW264.7 cells. The results showed that PEG@AuNPs were internalized more quickly by LPS-activated RAW264.7 cells than unstimulated cells, and they reached saturation within 24 hours. PEG@AuNPs enhanced LPS-induced production of NO and IL-6 and inducible nitric oxide synthase (iNOS) expression in RAW264.7 cells, partially by activating p38 mitogen-activated protein kinases (p38 MAPK) and nuclear factor-kappaB pathways. In addition, the p38 MAPK inhibitor SB203580 attenuated PEG@AuNP-enhanced LPS-induced NO production and iNOS expression. Overproduction of NO and IL-6 is known to be closely correlated with the pathology of many diseases and inflammations. Thus, it is speculated that the highly biocompatible gold nanoparticles can induce immunotoxicity due to their potency to stimulate macrophages to release aberrant or excessive pro-inflammatory mediators.While the immunogenicity and cytotoxicity of gold nanoparticles (AuNPs) are noted by many researchers, the mechanisms by which AuNPs exert these effects are poorly understood. In this study, we investigated the effects of polyethylene glycolylated AuNPs (PEG@AuNPs) on lipopolysaccharide (LPS)-induced nitric oxide (NO) and interleukin-6 (IL-6) production and the associated molecular mechanism in RAW264.7 cells. The results showed that PEG@AuNPs were internalized more quickly by LPS-activated RAW264.7 cells than unstimulated cells, and they reached saturation within 24 hours. PEG@AuNPs enhanced LPS-induced production of NO and IL-6 and inducible nitric oxide synthase (iNOS) expression in RAW264.7 cells, partially by activating p38 mitogen-activated protein kinases (p38 MAPK) and nuclear factor-kappaB pathways. In addition, the p38 MAPK inhibitor SB203580 attenuated PEG@AuNP-enhanced LPS-induced NO production and iNOS expression. Overproduction of NO and IL-6 is known to be closely correlated with the pathology of many diseases and inflammations. Thus, it is speculated that the highly biocompatible gold nanoparticles can induce immunotoxicity due to their potency to stimulate macrophages to release aberrant or excessive pro-inflammatory mediators. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr31355c
Liu, Zhimin; Li, Wenqing; Wang, Feng; Sun, Chunyang; Wang, Lu; Wang, Jun; Sun, Fei
Ethyl acetate extract from Angelica Dahuricae Radix inhibits lipopolysaccharide-induced production of nitric oxide, prostaglandin E2 and tumor necrosis factor-alphavia mitogen-activated protein kinases and nuclear factor-kappaB in macrophages.
Angelica dahurica (Umbelliferae) has been used to treat headache of common cold, supraorbital neuralgia, painful swelling on the body, nasal stuffiness, leukorrhea and arthralgia due to wind-dampness in Korean traditional medicine. It is also claimed to be effective in the treatment of acne, erythema, headache, toothache, sinusitis, colds and flu. The present study focused whether the ethyl acetate extract from Angelica Dahuricae Radix (EAAD) inhibits production of nitric oxide (NO), prostaglandin E(2) (PGE(2)) and tumor necrosis factor (TNF)-alpha, as well as expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), nuclear factor-kappaB (NF-kappaB) and mitogen-activated protein kinases (MAPKs) in lipopolysaccharide (LPS)-stimulated macrophages. EAAD inhibited LPS-induced NO, PGE(2) and TNF-alpha production as well as expression of iNOS and COX-2 in RAW 264.7 cells. EAAD inhibited LPS-induced TNF-alpha production in THP-1 cells. Furthermore, EAAD suppressed LPS-induced phosphorylation of p38 MAPK and extracellular-signal regulated kinases 1/2 (ERK1/2), I-kappaBalpha degradation, and NF-kappaB activation in RAW 264.7 cells. These results suggest that EAAD has the inhibitory effects on LPS-induced TNF-alpha, NO and PGE(2) production, and expression of iNOS and COX-2 in macrophage through blockade in the phosphorylation of MAPKs, following I-kappaBalpha degradation and NF-kappaB activation. PMID:17229575
Kang, Ok-Hwa; Lee, Go-Hoon; Choi, Hyuk Joon; Park, Pil Sang; Chae, Hee-Sung; Jeong, Seung-Il; Kim, Youn-Chul; Sohn, Dong Hwan; Park, Hyun; Lee, John Hwa; Kwon, Dong-Yeul
Disruption of renal peritubular blood flow in lipopolysaccharide-induced renal failure: role of nitric oxide and caspases.
Acute renal failure (ARF) is a frequent and serious complication of endotoxemia caused by lipopolysaccharide (LPS) and contributes significantly to mortality. The present studies were undertaken to examine the roles of nitric oxide (NO) and caspase activation on renal peritubular blood flow and apoptosis in a murine model of LPS-induced ARF. Male C57BL/6 mice treated with LPS (Escherichia coli) at a dose of 10 mg/kg developed ARF at 18 h. Renal failure was associated with a significant decrease in peritubular capillary perfusion. Vessels with no flow increased from 7 +/- 3% in the saline group to 30 +/- 4% in the LPS group (P < 0.01). Both the inducible NO synthase inhibitor L-N(6)-1-iminoethyl-lysine (L-NIL) and the nonselective caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp fluoromethylketone (Z-VAD) prevented renal failure and reversed perfusion deficits. Renal failure was also associated with an increase in renal caspase-3 activity and an increase in renal apoptosis. Both L-NIL and Z-VAD prevented these changes. LPS caused an increase in NO production that was blocked by L-NIL but not by Z-VAD. Taken together, these data suggest NO-mediated activation of renal caspases and the resulting disruption in peritubular blood flow are an important mechanism of LPS-induced ARF. PMID:15998845
Tiwari, Manish M; Brock, Robert W; Megyesi, Judit K; Kaushal, Gur P; Mayeux, Philip R
Suppressive effect of novel aromatic diamine compound on nuclear factor-kappaB-dependent expression of inducible nitric oxide synthase in macrophages.
N1-benzyl-4-methylbenzene-1,2-diamine (BMD) is a novel synthetic compound. In the present study, BMD compound was discovered to inhibit nitric oxide (NO) production in macrophages RAW 264.7. BMD compound attenuated lipopolysaccharide (LPS)-induced synthesis of both mRNA and protein of inducible nitric oxide synthase (iNOS), and inhibited LPS-induced iNOS promoter activity, indicating that the aromatic diamine compound could down-regulate iNOS expression at the transcription level. As a mechanism of the anti-inflammatory action, suppression of BMD compound on nuclear factor (NF)-kappaB activation has been documented. BMD compound exhibited dose-dependent inhibitory effect on LPS-mediated NF-kappaB transcriptional activity in the macrophages. Further, the compound inhibited LPS-mediated nuclear translocation of NF-kappaB p65 and DNA binding activity of NF-kappaB complex, in parallel, but did not affect LPS-mediated degradation of inhibitory kappaBalpha protein (IkappaBalpha). These results indicate that BMD compound could inhibit nuclear localization step of NF-kappaB p65 without affecting IkappaBalpha degradation. Finally, BMD compound could provide an invaluable tool to investigate NF-kappaB-dependent iNOS expression, in addition to its therapeutic potential in NO-associated inflammatory diseases. PMID:16183054
Shin, Hyun-Mo; Byung Hak Kim; Eun Yong Chung; Jung, Sang-Hun; Yeong Shik Kim; Kyung Rak Min; Kim, Youngsoo
Inhibition of inducible nitric oxide synthase expression by novel nonsteroidal anti-inflammatory derivatives with gastrointestinal-sparing properties.
1. The effects of novel nitric oxide-releasing nonsteroidal anti-inflammatory compounds (NO-NSAIDs) on induction of nitric oxide (NO) synthase by bacterial lipopolysaccharide (LPS) were examined in a murine cultured macrophage cell line, J774. 2. LPS-induced nitrite production was markedly attenuated by the nitroxybutylester derivatives of flurbiprofen (FNBE), aspirin, ketoprofen, naproxen, diclofenac and ketorolac, with each compound reducing accumulated nitrite levels by > 40% at the maximum concentrations (100 micrograms ml-1) used. 3. Further examination revealed that nitrite production was inhibited in a concentration-dependent (1-100 micrograms ml-1) manner by FNBE which at 100 micrograms ml-1 decreased LPS-stimulated levels by 63.3 +/- 8.6% (n = 7). The parent compound flurbiprofen was relatively ineffective over the same concentration-range, inhibiting nitrite accumulation by 24 +/- 0.9% (n = 3) at the maximum concentration used (100 micrograms ml-1). 4. FNBE reduced LPS-induced nitrite production when added to cells up to 4 h after LPS. Thereafter, FNBE caused very little or no reduction in nitrite levels. Furthermore NO-NSAIDs (100 micrograms ml-1) did not inhibit the metabolism of L-[3H]-arginine to citrulline by NO synthase isolated from LPS-activated macrophages. 5. Western blot analysis demonstrated that NO synthase expression was markedly attenuated following co-incubation of J774 cell with LPS (1 microgram ml-1; 24 h) and FNBE (100 micrograms ml-1; 24 h). Thus taken together, these findings indicate that NO-NSAIDs inhibit induction of NO synthase without directly affecting enzyme activity. 6. In conclusion our results indicate that NO-NSAIDs can inhibit the inducible L-arginine-NO pathway, and are capable of suppressing NO synthesis by inhibiting expression of NO synthase. The clinical implications of these findings remain to be established. PMID:8730734
Cirino, G; Wheeler-Jones, C P; Wallace, J L; Del Soldato, P; Baydoun, A R
Inhibition of inducible nitric oxide synthase expression by novel nonsteroidal anti-inflammatory derivatives with gastrointestinal-sparing properties.
1. The effects of novel nitric oxide-releasing nonsteroidal anti-inflammatory compounds (NO-NSAIDs) on induction of nitric oxide (NO) synthase by bacterial lipopolysaccharide (LPS) were examined in a murine cultured macrophage cell line, J774. 2. LPS-induced nitrite production was markedly attenuated by the nitroxybutylester derivatives of flurbiprofen (FNBE), aspirin, ketoprofen, naproxen, diclofenac and ketorolac, with each compound reducing accumulated nitrite levels by > 40% at the maximum concentrations (100 micrograms ml-1) used. 3. Further examination revealed that nitrite production was inhibited in a concentration-dependent (1-100 micrograms ml-1) manner by FNBE which at 100 micrograms ml-1 decreased LPS-stimulated levels by 63.3 +/- 8.6% (n = 7). The parent compound flurbiprofen was relatively ineffective over the same concentration-range, inhibiting nitrite accumulation by 24 +/- 0.9% (n = 3) at the maximum concentration used (100 micrograms ml-1). 4. FNBE reduced LPS-induced nitrite production when added to cells up to 4 h after LPS. Thereafter, FNBE caused very little or no reduction in nitrite levels. Furthermore NO-NSAIDs (100 micrograms ml-1) did not inhibit the metabolism of L-[3H]-arginine to citrulline by NO synthase isolated from LPS-activated macrophages. 5. Western blot analysis demonstrated that NO synthase expression was markedly attenuated following co-incubation of J774 cell with LPS (1 microgram ml-1; 24 h) and FNBE (100 micrograms ml-1; 24 h). Thus taken together, these findings indicate that NO-NSAIDs inhibit induction of NO synthase without directly affecting enzyme activity. 6. In conclusion our results indicate that NO-NSAIDs can inhibit the inducible L-arginine-NO pathway, and are capable of suppressing NO synthesis by inhibiting expression of NO synthase. The clinical implications of these findings remain to be established. Images Figure 4
Cirino, G.; Wheeler-Jones, C. P.; Wallace, J. L.; Del Soldato, P.; Baydoun, A. R.
?-Dihydroxychalcone-glycoside (?-DHC) isolated from the heartwood of Pterocarpus marsupium inhibits LPS induced MAPK activation and up regulates HO-1 expression in murine RAW 264.7 macrophage.
Three phenolic glycosides isolated from the heartwood of Pterocarpus marsupium showed significant free radical and superoxide ion scavenging activity and antioxidant potential that were comparable to, or several folds higher than those of standard antioxidants, trolox and ascorbic acid. The effective concentrations of these compounds were far below their cytotoxic levels. Compound 3, which was characterized to be ?-dihydroxychalcone-glycoside (?-DHC), was the most potent one. Subsequent studies demonstrated that ?-DHC effectively reduced nitric oxide and cytokine production by the LPS stimulated RAW 264.7 mouse macrophage cell line. The compound effectively attenuated the expression of inflammation-mediating enzymes COX-2 and iNOS at the mRNA as well as protein levels in a concentration dependent manner. It prevented phosphorylation of all the three MAPKs (JNK, ERK, p38) and eventually blocked the activation of downstream elements contributing to inflammation. Phosphorylation of I?B-? and subsequent translocation of NF-?B into the nucleus were restricted, while the expression of stress responsive gene HO-1 was up-regulated. ?-DHC targeted Keap-1 by modifying its cysteine thiols, dissociating it from Nrf-2 and facilitating nuclear entry of the latter; and this in turn induced HO-1 expression. Thus ?-DHC exerts its anti-inflammatory activity in a dual manner: by down regulating MAPKs and restricting nuclear stabilization of NF-?B at one end, and by disrupting Nrf-2-Keap-1 complex on the other. In conclusion, the anti-inflammatory potential together with its high therapeutic index envisages ?-DHC as a prospective candidate molecule for the development of therapeutic strategy against inflammatory disorders. PMID:24675710
Chakraborty, Prarthana; Saraswat, Ghungroo; Kabir, Syed N
Puerarin suppresses production of nitric oxide and inducible nitric oxide synthase in lipopolysaccharide-induced N9 microglial cells through regulating MAPK phosphorylation, O-GlcNAcylation and NF-?B translocation.
Microglial cells play a critical role in mediating central nervous system in?ammatory processes. Activated microglial cells induced by proinflammatory factor, such as lipopolysaccharide (LPS), release many kinds of neurotoxic cytokines including reactive oxygen species (ROS) which contributes to the pathogenesis of neurodegenerative diseases. Puerarin, extracted from kudzu root, possesses the characteristic of neuroprotection, antioxidation and anticancer. In the present study, we observed that LPS induced over-production of nitric oxide (NO) and increased the level of intracellular ROS in N9 microglial cells, but it was inhibited by puerarin. Furthermore, treatment with puerarin on N9 cells suppressed the over-expression of inducible nitric oxide synthase (iNOS) induced by LPS which is implicated in intracellular O-linked ?-N-acetylglucosamine (O-GlcNAc) level, phosphorylation of mitogen-activated protein kinase (MAPK) and nuclear factor ?B (NF-?B) signaling pathway. We also observed that the enhanced phosphorylation of p38, JNK and ERK1/2 in N9 cells induced by LPS were inhibited by puerarin, otherwise the down-regulation of O-GlcNAcylation level of protein in N9 cell induced by LPS was up-regulated by pretreatment with puerarin. These results indicate that puerarin effectively inhibits microglia activation induced by LPS through inhibiting expression of iNOS, production of NO and ROS which was mediated via regulating O-GlcNAcylation, phosphorylation of MAPK and NF-?B translocation. PMID:22246431
Zheng, Gao-Ming; Yu, Chao; Yang, Zhu
A molecular pharmacology study into the anti-inflammatory actions of Euphorbia hirta L. on the LPS-induced RAW 264.7�cells through selective iNOS protein inhibition
Euphorbia hirta L. has been widely used in India and Chinese society. The molecular pharmacology basis of its anti-inflammatory effect is\\u000a revealed in this work. The ethanol extract of Euphorbia hirta L. (Eh) and its active component were studied in lipopolysaccharide (LPS)-activated macrophage cells (RAW 264.7) as an established\\u000a inflammation model. After activation, nitric oxide (NO) production and expression of
Mei-Fen Shih; Yih-Dih Cheng; Chia-Rui Shen; Jong-Yuh Cherng
Dual Specificity Phosphatase 1 Regulates Human Inducible Nitric Oxide Synthase Expression by p38 MAP Kinase
The role of dual specificity phosphatase 1 (DUSP1) in inducible nitric oxide synthase (iNOS) expression in A549 human pulmonary epithelial cells, J774 mouse macrophages and primary mouse bone marrow-derived macrophages (BMMs) was investigated. iNOS expression was induced by a cytokine mixture (TNF, IFN? and IL-1?) in A549 cells and by LPS in J774 cells, and it was inhibited by p38 MAPK inhibitors SB202190 and BIRB 796. Stimulation with cytokine mixture or LPS enhanced also DUSP1 expression. Down-regulation of DUSP1 by siRNA increased p38 MAPK phosphorylation and iNOS expression in A549 and J774 cells. In addition, LPS-induced iNOS expression was enhanced in BMMs from DUSP1(?/?) mice as compared to that in BMMs from wild-type mice. The results indicate that DUSP1 suppresses iNOS expression by limiting p38 MAPK activity in human and mouse cells. Compounds that enhance DUSP1 expression or modulate its function may be beneficial in diseases complicated with increased iNOS-mediated NO production.
Turpeinen, Tuija; Nieminen, Riina; Taimi, Ville; Heittola, Taina; Sareila, Outi; Clark, Andrew R.; Moilanen, Eeva; Korhonen, Riku
Altered Regulation of Renal Nitric Oxide and Atrial Natriuretic Peptide Systems in Lipopolysaccharide-induced Kidney Injury
Nitric oxide (NO) and atrial natriuretic peptide (ANP) may induce vascular relaxation by increasing the production of cyclic guanosine monophosphate (cGMP), an important mediator of vascular tone during sepsis. This study aimed to determine whether regulation of NO and the ANP system is altered in lipopolysaccharide (LPS)-induced kidney injury. LPS (10 mg.kg-1) was injected in the tail veins of male Sprague-Dawley rats; 12 hours later, the kidneys were removed. Protein expression of NO synthase (NOS) and neutral endopeptidase (NEP) was determined by semiquantitative immunoblotting. As an index of synthesis of NO, its stable metabolites (nitrite/nitrate, NOx) were measured using colorimetric assays. mRNA expression of the ANP system was determined by real-time polymerase chain reaction. To determine the activity of guanylyl cyclase (GC), the amount of cGMP generated in response to sodium nitroprusside (SNP) and ANP was calculated. Creatinine clearance decreased and fractional excretion of sodium increased in LPS-treated rats compared with the controls. Inducible NOS protein expression increased in LPS-treated rats, while that of endothelial NOS, neuronal NOS, and NEP remained unchanged. Additionally, urinary and plasma NOx levels increased in LPS-treated rats. SNP-stimulated GC activity remained unchanged in the glomerulus and papilla in the LPS-treated rats. mRNA expression of natriuretic peptide receptor (NPR)-C decreased in LPS-treated rats, while that of ANP and NPR-A did not change. ANP-stimulated GC activity reduced in the glomerulus and papilla. In conclusion, enhancement of the NO/cGMP pathway and decrease in ANP clearance were found play a role in the pathogenesis of LPS-induced kidney injury.
Bae, Eun Hui; Kim, In Jin; Ma, Seong Kwon; Lee, Jong Un
The NADPH oxidase inhibitor apocynin induces nitric oxide synthesis via oxidative stress
We have recently shown that apocynin elicits an oxidative stress in N11 mouse glial cells and other cell types. Here we report that apocynin increased the accumulation of nitrite, the stable derivative of nitric oxide (NO), in the extracellular medium of N11 cell cultures, and the NO synthase (NOS) activity in cell lysates. The increased synthesis of NO was associated with increased expression of inducible NOS (iNOS) mRNA, increased nuclear translocation of the redox-sensitive transcription factor NF-{kappa}B and decreased intracellular level of its inhibitor IkB{alpha}. These effects, accompanied by increased production of H{sub 2}O{sub 2}, were very similar to those observed after incubation with bacterial lipopolysaccharide (LPS) and were inhibited by catalase. These results suggest that apocynin, similarly to LPS, induces increased NO synthesis by eliciting a generation of reactive oxygen species (ROS), which in turn causes NF-{kappa}B activation and increased expression of iNOS. Therefore, the increased bioavailability of NO reported in the literature after in vivo or in vitro treatments with apocynin might depend, at least partly, on the drug-elicited induction of iNOS, and not only on the inhibition of NADPH oxidase and the subsequent decreased scavenging of NO by oxidase-derived ROS, as it is often supposed.
Riganti, Chiara [Dipartimento di Genetica, Biologia e Biochimica, Universita di Torino, Via Santena 5/bis, 10126 Torino (Italy)], E-mail: email@example.com; Costamagna, Costanzo; Doublier, Sophie; Miraglia, Erica; Polimeni, Manuela; Bosia, Amalia; Ghigo, Dario [Dipartimento di Genetica, Biologia e Biochimica, Universita di Torino, Via Santena 5/bis, 10126 Torino (Italy)
Hexane fraction from Sargassum fulvellum inhibits lipopolysaccharide-induced inducible nitric oxide synthase expression in RAW 264.7 cells via NF-?B pathways.
Sargassum fulvellum (Turner) C. Agardh has been used to treat various inflammatory diseases, including lump, dropsy, swollen and painful scrotum, and urination problems for several centuries with no side effects. This study aims to investigate the anti-inflammatory effect of the hexane fraction of S. fulvellum (HFS) in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells and phorbol 12-myristate 13-acetate (PMA)-induced mouse-ear edema. The anti-inflammatory activity of HFS in LPS-stimulated RAW 264.7 cells was investigated by assessing the inhibition of nitric oxide (NO) and pro-inflammatory cytokine production during Griess reaction and enzyme-linked immunosorbent assay (ELISA), respectively. The molecular mechanisms that underlie the anti-inflammatory action of HFS were investigated by analyzing the activation of transcription factor and its upstream signaling proteins. Additionally, an in vivo study of the anti-inflammatory effect of HFS was carried out using PMA-induced mouse-ear edema. HFS inhibited LPS-induced NO production in a dose-dependent manner and suppressed the expression of inducible NO synthase (iNOS) in the RAW 264.7 cells. Further, HFS reduced the production of pro-inflammatory cytokines in the LPS-stimulated RAW 264.7 cells. HFS significantly inhibited LPS-induced nuclear factor kappa B (NF-?B) transcriptional activity and NF-?B translocation into the nucleus by preventing degradation of inhibitor ?B-?. Moreover, HFS inhibited the activation of Akt and mitogen-activated protein kinases (MAPKs) in the LPS-stimulated RAW 264.7 cells. Furthermore, HFS suppressed PMA-induced mouse-ear edema. The above data indicate that the anti-inflammatory effects of HFS on LPS-stimulated cells are associated with the suppression of NF-?B through the inhibition of MAPKs and Akt phosphorylation. PMID:23711142
Gwon, Wi-Gyeong; Lee, Min-Sup; Kim, Jong-Soon; Kim, Jae-Il; Lim, Chi-Won; Kim, Nam-Gil; Kim, Hyeung-Rak
CHP1002, a novel andrographolide derivative, inhibits pro-inflammatory inducible nitric oxide synthase and cyclooxygenase-2 expressions in RAW264.7 macrophages via up-regulation of heme oxygenase-1 expression.
Andrographolides, a type of diterpene lactone, are widely known to have anti-inflammatory and anti-oxidative properties. CHP1002, a synthetic derivative of andrographolide, has similar anti-inflammatory action in mouse ear swelling test and rat paw edema test. In the present study, the mechanism of anti-inflammatory effects of CHP1002 was investigated in RAW264.7 macrophages. CHP1002 potently suppressed inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expressions in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. CHP1002 reduced the production of iNOS-derived nitric oxide (NO) and COX-2-derived prostaglandin E2 (PGE2). CHP1002 induced heme oxygenase-1 (HO-1) expression via activation of extracellular signal-regulated kinase (ERK) and NF-E2 related factor 2 transcription factor (Nrf2). Down-regulation of LPS-induced iNOS and COX-2 expressions was partially reversed by the HO-1 inhibitor zinc protoporphyrin (ZnPP). In addition, CHP1002 significantly attenuated LPS-induced TNF-?, IL-1? and IL-6 production. CHP1002 effectively induced HO-1 and was capable of inhibiting some macrophage-derived pro-inflammatory mediators, which may be closely correlated with its anti-inflammatory action. PMID:23261362
Zhang, Bo; Yan, Lingdi; Zhou, Peilan; Dong, Zhaoqi; Feng, Siliang; Liu, Keliang; Gong, Zehui
Bromelain improves decrease in defecation in postoperative rats: modulation of colonic gene expression of inducible nitric oxide synthase.
Ileus continues to be a common consequence of abdominal surgery, causing significant patient discomfort and often leading to more serious problems. The therapy available is limited, hence, ileus remains an important clinical problem. Activation of inducible nitric oxide synthase (iNOS) directly modulates intestinal dysmotility after bowel manipulation and plays an essential role in initiating intestinal inflammation. Nuclear factor (NF)-kappaB is known to be a critical component of iNOS gene transcriptional activation in response to inflammatory stimuli. Bromelain is a crude extract from the pineapple stem, which is sold as a nutritional supplement to "promote digestive health" and as an anti-inflammatory medication in some developed countries. Here, we have found that oral administration of bromelain improves decrease in defecation in abdominal postoperative rats. Results showed that bromelain increased the wet weight, dry weight, water content and number of fecal pellets in laparotomized plus mechanically manipulated rats, suggesting improvement of postoperative ileus. Furthermore, bromelain treatment inhibited overexpressed iNOS mRNA and restored down-regulated inhibitor kappaBalpha mRNA in the colon of the postoperative rats. From the in vitro experiments, bromelain inhibits lipopolysaccharide (LPS)-induced nitrite overproduction in macrophage cell lines and LPS-induced NF-kappaB luciferase reporter gene expression in RAW264.7 macrophages transfected with NF-kappaB luciferase reporter gene. Thus, our findings suggest that bromelain improves decrease in defecation in postoperative rats, at least in part, by inhibiting colonic iNOS overexpression via NF-kappaB pathway. Our data indicates that bromelain may benefit patients with postoperative ileus. PMID:16137711
Wen, Suping; Huang, Tom H W; Li, George Q; Yamahara, Johji; Roufogalis, Basil D; Li, Yuhao
Anti-angiogenic and inhibitory activity on inducible nitric oxide production of the mushroom Ganoderma lucidum
Fresh fruit bodies of Ganoderma lucidum were extracted with 70% ethanol at room temperature. The extract (GL) showed significant anti-angiogenic activity, which was detected using a chick embryo chorioallantoic membrane assay. GL significantly inhibited LPS-induced NO production in RAW 264.7 macrophages. These results support the anti-tumor effect of Ganoderma lucidum.
Yun Seon Song; Sun-Hyoung Kim; Jae-Hoon Sa; Changbae Jin; Chang-Jin Lim; Eun-Hee Park
Bauer Ketones 23 and 24 from Echinacea paradoxa var. paradoxa Inhibit Lipopolysaccharide-induced Nitric Oxide, Prostaglandin E2 and Cytokines in RAW 264.7 Mouse Macrophages
Among the nine Echinacea species, E. purpurea, E. angustifolia and E. pallida, have been widely used to treat the common cold, flu and other infections. In our study, ethanol extracts of these three Echinacea species and E. paradoxa, including its typical variety, E. paradoxa var. paradoxa, were screened in lipopolysaccharide (LPS)-stimulated macrophage cells to assess potential anti-inflammatory activity. Echinacea paradoxa var. paradoxa, rich in polyenes/polyacetylenes, was an especially efficient inhibitor of LPS-induced production of nitric oxide (NO), prostaglandin E2 (PGE2), interleukin-1 beta (IL-1?) and interleukin-6 (IL-6) by 46%, 32%, 53% and 26%, respectively, when tested at 20 ?g/ml in comparison to DMSO control. By bioactivity-guided fractionation, pentadeca-8Z-ene-11, 13-diyn-2-one (Bauer ketones 23, compound 1) and pentadeca-8Z, 13Z-dien-11-yn-2-one (Bauer ketone 24, compound 2) from E. paradoxa var. paradoxa were found primarily responsible for inhibitory effects on NO and PGE2 production. Moreover, Bauer ketone 24 (compound 2) was the major contributor to inhibition of inflammatory cytokine production in LPS-induced mouse macrophage cells. These results provide a rationale for exploring the medicinal effects of the Bauer ketone-rich taxon, E. paradoxa var. paradoxa, and confirm the anti-inflammatory properties of Bauer ketones 23 and 24.
Zhang, Xiaozhu; Rizshsky, Ludmila; Hauck, Catherine; Qu, Luping; Widrlechner, Mark P.; Nikolau, Basil J.; Murphy, Patricia A.; Birt, Diane F.
Inhibition of the induction of the inducible nitric oxide synthase in murine brain microglial cells by sodium salicylate.
The induction of the inducible nitric oxide synthase (iNOS) has been proposed to play a role in a variety of inflammatory diseases. Sodium salicylate (NaSal) is the most commonly used anti-inflammatory agent. We investigated whether NaSal can diminish the induction of iNOS in murine brain microglial cells. In primary cultures, interferon-gamma (IFN-gamma) or lipopolysaccharide (LPS) separately did not stimulate nitric oxide (NO) production, whereas IFN-gamma combined with LPS synergistically induced iNOS. NaSal inhibited both the production of NO and expression of iNOS in microglial cells. Synergy between IFN-gamma and LPS was mainly dependent on tumour necrosis factor-alpha (TNF-alpha) secretion as the increase of the induction of the iNOS by IFN-gamma plus LPS was associated with the increase of TNF-alpha secretion and IFN-gamma plus LPS-induced TNF-alpha secretion by microglial cells was decreased by the treatment with NaSal. These results suggest a possible use of NaSal in managing inflammation of the central nervous system through inhibition of the iNOS induction. Images Figure 2 Figure 3
Kim, H; Lee, E; Shin, T; Chung, C; An, N
Nitric oxide-induced calcium release
Ryanodine receptors (RyRs), located in the sarcoplasmic/endoplasmic reticulum (SR/ER) membrane, are required for intracellular Ca2+ release that is involved in a wide range of cellular functions. In addition to Ca2+-induced Ca2+ release in cardiac cells and voltage-induced Ca2+ release in skeletal muscle cells, we recently identified another mode of intracellular Ca2+ mobilization mediated by RyR, i.e., nitric oxide-induced Ca2+ release (NICR), in cerebellar Purkinje cells. NICR is evoked by neuronal activity, is dependent on S-nitrosylation of type 1 RyR (RyR1) and is involved in the induction of long-term potentiation (LTP) of cerebellar synapses. In this addendum, we examined whether peroxynitrite, which is produced by the reaction of nitric oxide with superoxide, may also have an effect on the Ca2+ release via RyR1 and the cerebellar LTP. We found that scavengers of peroxynitrite have no significant effect either on the Ca2+ release via RyR1 or on the cerebellar LTP. We also found that an application of a high concentration of peroxynitrite does not reproduce neuronal activity-dependent Ca2+ release in Purkinje cells. These results support that NICR is induced by endogenous nitric oxide produced by neuronal activity through S-nitrosylation of RyR1.
Kakizawa, Sho; Yamazawa, Toshiko; Iino, Masamitsu
Aminoguanidine selectively inhibits inducible nitric oxide synthase.
1. Endotoxin induces nitric oxide synthase in vascular tissue, including rat main pulmonary artery. Currently available agents that cause inhibition of nitric oxide synthase are relatively non-selective between the constitutive and inducible forms of the enzyme. 2. Aminoguanidine caused a dose-dependent increase in phenylephrine-induced tension in intact and endothelium-denuded pulmonary artery rings from endotoxin-treated rats, but had no effect on sham-treated controls. 3. Contraction caused by aminoguanidine in endothelium-denuded vessels from endotoxin-treated rats was unaffected by indomethacin (10 microM), and by cimetidine and mepyramine (both 10 microM), excluding an effect of aminoguanidine mediated by arachidonic acid metabolites or histamine. 4. Contraction caused by aminoguanidine in endothelium-denuded vessels from endotoxin-treated rats was abolished by L-arginine (2 mM) and L-NG-monomethyl arginine (300 microM), but unaffected by D-arginine and D-NG-monomethyl arginine, suggesting that its action is mediated by the L-arginine/nitric oxide pathway.(ABSTRACT TRUNCATED AT 250 WORDS)
Griffiths, M. J.; Messent, M.; MacAllister, R. J.; Evans, T. W.
Anti-inflammatory effect of anemarsaponin B isolated from the rhizomes of Anemarrhena asphodeloides in LPS-induced RAW 264.7 macrophages is mediated by negative regulation of the nuclear factor-?B and p38 pathways
Anemarrhena asphodeloides is widely used in traditional Chinese medicine, and is known to have anti-diabetic and diuretic effects. In this study, we evaluated the anti-inflammatory effects of anemarsaponin B (ASB), a steroidal saponin isolated from the rhizomes of A. asphodeloides (Liliaceae), in LPS-stimulated RAW 264.7 macrophage cell line. ASB significantly and dose-dependently decreased the protein and mRNA levels of inducible
Ji-Yeon Kim; Ji-Sun Shin; Jong Hoon Ryu; Sun Yeou Kim; Young-Wuk Cho; Jung-Hye Choi; Kyung-Tae Lee
Mechanism for Prenatal LPS-Induced DA Neuron Loss.
National Technical Information Service (NTIS)
A small percentage of patients with Parkinson's disease (PD) are caused by genetic defects (familial Parkinson's disease). The etiologies of the majority of patients are still unknown. Recent advance in our laboratories suggests that prenatal exposure to ...
P. M. Carvey
Mechanism for Prenatal LPS-Induced DA Neuron Loss.
National Technical Information Service (NTIS)
In nonfamilial Parkinson's Disease (PD) the etiologies of the majority of patients are still unknown. However, recent advances by the authors suggest that prenatal exposure to the bacterial toxin lipopolysaccharide (LPS) could be an important etiology for...
P. M. Carvey
Taurine Chloramine Inhibits LPS-Induced Glucose Uptake
Inflammatory cells use glucose as a primary source of metabolic energy, and thus increased uptake of glucose and high rates\\u000a of glycolysis are characteristics of inflamed cells. Taurine chloramine (TauCl) is the product of a reaction between cellular\\u000a taurine and hypochlorous acid (HOCl\\/OCl-), the latter produced by the halide-dependent myeloperoxidase (MPO) system in inflammatory cells. Taurine, a major metabolite\\u000a of
Chaekyun Kim; Seongtag Kim
Ginger extract inhibits LPS induced macrophage activation and function
BACKGROUND: Macrophages play a dual role in host defence. They act as the first line of defence by mounting an inflammatory response to antigen exposure and also act as antigen presenting cells and initiate the adaptive immune response. They are also the primary infiltrating cells at the site of inflammation. Inhibition of macrophage activation is one of the possible approaches
Sudipta Tripathi; David Bruch; Dilip S Kittur
Nitric oxide inhibits tissue factor synthesis, expression and activity in human monocytes by prior formation of peroxynitrite
Objective: Nitric oxide (NO) has antithrombotic properties by regulating platelet function, whereas direct effects on plasmatic coagulation\\u000a are rarely described. In sepsis and inflammation, when synthesis of NO, oxygen radicals and toxic metabolites is crucial,\\u000a the expression of tissue factor (TF) on monocytes stimulated by lipopolysaccharides (LPS) induces intravascular coagulation.\\u000a This study was performed to examine the influence of NO
M. Gerlach; D. Keh; G. Bezold; S. Spielmann; I. K�rer; R. U. Peter; K. J. Falke; H. Gerlach
Functional effects of econazole on inducible nitric oxide synthase: production of a calmodulin-dependent enzyme.
1. We performed experiments to examine the effects of an anti-fungal imidazole compound, econazole, on the regulation and effects of lipopolysaccharide-inducible nitric oxide synthase (iNOS) activity in rat aortic rings and cultured J774 murine macrophage cells. 2. In endothelium-intact rings of thoracic aorta, phenylephrine caused a concentration-dependent contraction with EC50 of 1.9 +/- 0.15 x 10(-8) M (n = 5). Following incubation with lipopolysaccharide (LPS, 5 micrograms ml-1) for 8 h there was a right-shift in the concentration-response curve (EC50 3.1 +/- 0.28 x 10(-7) M, P < 0.05) with a depression in the maximum contraction from 1.44 +/- 0.25 g to 0.86 +/- 0.26 g (n = 4). Co-incubation of rings with econazole (1 x 10(-5) M) partially inhibited the LPS-induced loss of reactivity to phenylephrine (EC50 6.5 +/- 0.72 x 10(-8) M) and fully inhibited the reduction in maximum tension (1.49 +/- 0.19 g; n = 5). 3. In J774 cells, incubation with LPS (10 micrograms ml-1, 24 h) resulted in significant nitrite production that was inhibited by co-incubation with econazole (IC50 5.0 +/- 0.9 x 10(-6) M; n = 5). In cells stimulated with LPS, production of L-[3H]-citrulline from L-[3H]-arginine was 6.41 +/- 0.22 pmol mg-1 protein min-1 (n = 3). This was inhibited by 92 +/- 6% by addition of NG-monomethyl-L-arginine (L-NMMA, 1 x 10(-3) M; n = 3) to the homogenate but not by econazole (1 x 10(-5) M; n = 3). In contrast pretreatment of cells with econazole (1 x 10(-5) M) markedly reduced the LPS-induced [3H]-citrulline production (0.86 +/- 0.053 pmol mg-1 protein min-1; P < 0.01; n = 3). 4. In cells treated with LPS and econazole, L-[3H]-citrulline production was restored in a concentration-dependent manner by addition of calmodulin (1 x 10(-8)-3 x 10(-7) M) with an IC50 of 4.2 +/- 0.9 x 10(-8) M. 5. We have shown that econazole inhibits the functional and biochemical activity of iNOS in rat aortic rings and cultured J774 cells. Treatment of cells with econazole renders the NO synthase functionally inactive. In econazole-treated cells enzyme activity is restored by calmodulin suggesting that econazole may inhibit the binding of this essential co-factor to the enzyme following its production. These studies may have implications for the design of novel anti-inflammatory agents working through the L-arginine-nitric oxide pathway.
Bogle, R. G.; Vallance, P.
Lycopene suppresses proinflammatory response in lipopolysaccharide-stimulated macrophages by inhibiting ROS-induced trafficking of TLR4 to lipid raft-like domains.
We recently showed that lycopene inhibited lipopolysaccharide (LPS)-induced productions of nitric oxide (NO) and interleukin-6 (IL-6) in murine RAW264.7 macrophages by mechanisms related to inhibition of ERK and nuclear factor-?B. Since the assembly of Toll-like receptor 4 (TLR4) in lipid rafts is a key element in LPS induced signaling, we investigated whether this process would be influenced by lycopene. We found that pretreatment of RAW264.7 cells with lycopene inhibited LPS-induced recruitment of TLR4 into fractions - enriched with lipid raft marker. By the methods of immunoprecipitation and immunoblotting, we also found that lycopene inhibited the subsequent formation of the complex of TLR4 with its adaptors including myeloid differentiation primary-response protein 88 and TIR domain-containing adaptor-inducing IFN-?. We also found that the lycopene induced inhibition was associated with reduced formation of reactive oxygen species (ROS), which was an upstream mechanism for the effects of lycopene, because treating the cells with the antioxidant N-acetyl-l-cysteine and NADPH oxidase inhibitor diphenyleneiodonium chloride significantly inhibited LPS-induced recruitment of TLR4 into lipid raft-like domains as well as the production of proinflammatory molecule NO and IL-6. Thus, our findings suggest that lycopene may prevent LPS-induced TLR4 assembly into lipid rafts through reducing intracellular ROS level. PMID:23246157
Zou, Jun; Feng, Dan; Ling, Wen-Hua; Duan, Rui-Dong
Inhibitory action of novel aromatic diamine compound on lipopolysaccharide-induced nuclear translocation of NF-kappaB without affecting IkappaB degradation.
4-Methyl-N1-(3-phenyl-propyl)-benzene-1,2-diamine (JSH-23) is a novel chemically synthetic compound. The aromatic diamine JSH-23 compound exhibited inhibitory effect with an IC(50) value of 7.1 microM on nuclear factor (NF)-kappaB transcriptional activity in lipopolysaccharide (LPS)-stimulated macrophages RAW 264.7, and interfered LPS-induced nuclear translocation of NF-kappaB without affecting IkappaB degradation. This mechanism of action is very rare for controlling NF-kappaB activation. Furthermore, the compound inhibited not only LPS-induced expressions of tumor necrosis factor-alpha, interleukin (IL)-1beta, IL-6 and inducible nitric oxide synthase and cyclooxygenase-2 but also LPS-induced apoptosis of the RAW 264.7 cells. PMID:15280016
Shin, Hyun-Mo; Kim, Min-Hee; Kim, Byung Hak; Jung, Sang-Hun; Kim, Yeong Shik; Park, Hye Ji; Hong, Jin Tae; Min, Kyung Rak; Kim, Youngsoo
Recognition of Betaine as an Inhibitor of Lipopolysaccharide-Induced Nitric Oxide Production in Activated Microglial Cells
Background: Neuroinflammation, as a major outcome of microglia activation, is an important factor for progression of neurodegenerative disorders including Alzheimer's disease and Parkinson's disease. Microglial cells, as the first-line defense in the central nervous system, act as a source of neurotoxic factors such as nitric oxide (NO), a free radical which is involved in neuronal cell death. The aim of this study was to inhibit production of NO in activated microglial cells in order to decrease neurological damages that threat the central nervous system. Methods: An in vitro model of a newborn rat brain cell culture was used to examine the effect of betaine on the release of NO induced by lipopolysaccharide (LPS). Briefly, primary microglial cells were stimulated by LPS and after 2 minutes, they were treated by different concentrations of betaine. The production of NO was assessed by the Griess assay while cell viability was determined by the MTT assay. Results: Our investigations indicated that LPS-induced NO release was attenuated by betaine, suggesting that this compound might inhibit NO release. The effects of betaine on NO production in activated microglial cells after 24 h were "dose-dependent". It means that microglial cells which were treated with higher concentrations of betaine, released lower amounts of NO. Also our observations showed that betaine compound has no toxic effect on microglial cells. Conclusion: Betaine has an inhibitory effect on NO release in activated microglial cells and may be an effective therapeutic component to control neurological disorders.
Amiraslani, Banafsheh; Sabouni, Farzaneh; Abbasi, Shahsanam; Nazem, Habiballah; Sabet, Mohammadsadegh
Preparation and biological evaluation of enzyme-assisted extracts from edible seaweed (Enteromorpha prolifera) as antioxidant, anti-acetylcholinesterase and inhibition of lipopolysaccharide-induced nitric oxide production in murine macrophages.
The multifunctional bioactive materials were prepared from Enteromorpha prolifera by enzyme-assisted extraction using four proteases and seven carbohydrases, and the biological activities of the enzyme-assisted extracts were evaluated as antioxidant, anti-acetylcholinesterase (AChE) and anti-inflammatory effect as the measures of inhibition of lipopolysaccharide (LPS)-induced nitric oxide (NO) production in RAW264.7 cells. The enzyme-assisted extracts were rich in polyphenols in the range 124 � 4.2 to 844 � 9.1 mg/100 g and flavonoids in the range 453 � 6.0 to 675 � 5.2 mg/100 g, and Protamex and Viscozyme extracts, which were rich in polyphenols and flavonoids, showed the highest 2,2-diphenyl-1-picrylhydrazyl scavenging, hydrogen peroxide scavenging, ferrous ion chelating and reducing power. Flavourzyme extract (89.92%) and Promozyme extract (93.64%) showed the highest AChE inhibitory activities at the concentration of 1.0 mg/ml. All enzyme-assisted extracts showed no cytotoxic effect on RAW264.7 cells at the tested concentration and significantly inhibited the LPS-induced NO production in RAW264.7 cells. PMID:21913802
Ahn, Chang-Bum; Park, Pyo-Jam; Je, Jae-Young
Chronic diabetic nephropathy: role of inducible nitric oxide synthase
.�� Nitric oxide (NO) is a multifunctional mediator that has been implicated in the short-term hemodynamic alterations that occur\\u000a in acute streptozocin (STZ)-induced diabetes. We investigated the role of NO produced by inducible nitric oxide synthase (iNOS)\\u000a in chronic STZ diabetic nephropathy. Diabetes was induced in C57BL\\/6 and iNOS knockout (KO) mice with two intraperitoneal\\u000a injections of STZ, 100 mg\\/kg.
Howard Trachtman; Stephen Futterweit; Elyse Pine; Jared Mann; Elsa Valderrama
Protective effects of agmatine against D-galactosamine and lipopolysaccharide-induced fulminant hepatic failure in mice.
Fulminant hepatic failure (FHF) is a life-threatening syndrome characterized by massive hepatic necrosis and high mortality. There is no effective therapy for the disease other than liver transplantation. This study aimed to investigate the effect of agmatine, inducible nitric oxide synthase (iNOS) inhibitor, on D-galactosamine and lipopolysaccharide (GalN/LPS)-induced FHF in mice and explore its possible mechanism(s). Male Swiss albino mice were injected with a single dose agmatine (14 mg/kg, IP) 8 h prior to challenge with a single intraperitoneal injection of both GalN (800 mg/kg) and LPS (50 ?g/kg). Agmatine significantly attenuated all GalN/LPS-induced biochemical and pathological changes in liver. It prevented the increase of serum transaminases and alkaline phosphatase (ALP). In addition, agmatine markedly attenuated GalN/LPS-induced necrosis and inflammation. Agmatine significantly reduced oxidative stress and enhanced antioxidant enzymes. Importantly, agmatine decreased total nitric oxide (NO) and pro-inflammatory cytokine, tumor necrosis factor-alpha (TNF-?). These findings reveal that agmatine has hepatoprotective effects against GalN/LPS-induced FHF in mice that may be related to its ability to suppress oxidative stress, NO synthesis and TNF-? production. Therefore, agmatine may serve as a novel therapeutic strategy for hepatic inflammatory diseases. PMID:23989863
El-Agamy, Dina S; Makled, Mirhan N; Gamil, Nareman M
Zinc protoporphyrin inhibition of lipopolysaccharide-, lipoteichoic acid-, and peptidoglycan-induced nitric oxide production through stimulating iNOS protein ubiquitination
In the present study, zinc protoporphyrin (ZnPP), but not ferric protoporphyrin (FePP), tin protoporphyrin (SnPP), or zinc chloride (ZnCl{sub 2}), at the doses of 0.5, 1, and 2 {mu}M, dose-dependently inhibited lipopolysaccharide- (LPS), lipoteichoic acid (LTA), and peptidoglycan (PGN)-induced inducible nitric oxide (iNOS) and nitric oxide (NO) production with an increase in heme oxygenase 1 (HO-1) protein in RAW264.7 macrophages in a serum-free condition. NO inhibition and HO-1 induction by ZnPP were blocked by the separate addition of fetal bovine serum (FBS) and bovine serum albumin (BSA). A decrease in the iNOS/NO ratio and an increase in HO-1 protein by ZnPP were identified in three different conditions including ZnPP pretreatment, ZnPP co-treatment, and ZnPP post-treatment with LPS and LTA. Activation of c-Jun N-terminal kinases (JNKs) and extracellular regulated kinases (ERKs) were detected in LPS-, LTA-, and PGN-treated RAW264.7 cells, and iNOS/NO production was blocked by adding the JNK inhibitor, SP600125, but not the ERK inhibitor, PD98059. However, ZnPP addition potentiated ERK and JNK protein phosphorylation stimulated by LPS, LTA, and PGN. Increases in total protein ubiquitination and ubiquitinated iNOS proteins were detected in ZnPP-treated macrophages elicited by LPS according to Western and immunoprecipitation/Western blotting assays, respectively. The decrease in LPS-induced iNOS protein by ZnPP was reversed by adding the proteasome inhibitors MG132 and lactacystin. The reduction in HO-1 protein induced by ZnPP via transfection of HO-1 small interfering RNA did not affect the inhibitory effect of ZnPP against LPS-induced iNOS/NO production and protein ubiquitination induced by ZnPP in macrophages. Data of the present study provide the first evidence to support ZnPP effectively inhibiting inflammatory iNOS/NO production through activation of protein ubiquitination in a HO-1-independent manner in macrophages.
Chow, J.-M. [Section of Hematology-Oncology, Department of Internal Medicine, Taipei Municipal Wan Fang Hospital, Taipei Medical University, Taipei 110, Taiwan (China); Lin, H.-Y.; Shen, S.-C. [Graduate Institute of Medical Sciences, Taipei Medical University, Taipei 110, Taiwan (China); Wu, M.-S. [Department of Gastroenterology, Taipei Medical University Wan Fang Hospital, Taiwan (China); Lin, C.-W. [Graduate Institute of Pharmacy, School of Pharmacy, Taipei Medical University, Taipei 110, Taiwan (China); Chiu, W.-T. [Department of Neurosurgery, Taipei Medical University-Wan Fang Hospital, Taipei, Taiwan (China); Lin, C.-H. [Graduate Institute of Medical Sciences, Taipei Medical University, Taipei 110, Taiwan (China)], E-mail: firstname.lastname@example.com; Chen, Y.-C. [Graduate Institute of Medical Sciences, Taipei Medical University, Taipei 110, Taiwan (China); Cancer Research Center and Orthopedics Research Center, Taipei Medical University Hospital, Taipei, Taiwan (China)], E-mail: email@example.com
Role of Inducible Nitric Oxide Synthase-Derived Nitric Oxide in Silica-Induced Pulmonary Inflammation and Fibrosis
Inhalation of crystalline silica can produce lung inflammation and fibrosis. Inducible nitric oxide synthase (iNOS)-derived nitric oxide (NO) is believed to be involved in silica-induced lung disease. To investigate the role of iNOS-derived NO in this disease, the responses of iNOS knockout (KO) versus C57Bl\\/6J wild-type (WT) mice to silica were compared. Male mice (8�10 wk old, mean body weight
Patti C. Zeidler; Ann Hubbs; Lori Battelli; Vincent Castranova
Antifibrotic Role of Inducible Nitric Oxide Synthase
Long-term treatment in rats with l-NAME, an isoform-non-specific inhibitor of nitric oxide synthase (NOS), leads to fibrosis of the heart and kidney, suggesting that nitric oxide (NO) may play a role in preventing tissue fibrosis. In this process, a likely target of NO is the quenching of reactive oxygen species (ROS) through peroxynitrite formation, and one possible source for this
M. G. Ferrini; D. Vernet; T. R. Magee; A. Shahed; A. Qian; J. Rajfer; N. F. Gonzalez-Cadavid
Ganoderma lucidum inhibits inducible nitric oxide synthase expression in macrophages
Nitric oxide (NO) is a principal mediator in many physiological and pathological processes. Overproduction of NO via the inducible nitric oxide synthase (iNOS) has cytotoxic effect through the formation of peroxynitrite with superoxide anion. The iNOS is mainly expressed in macrophages and is able to produce large amount of NO. The expression of iNOS is mainly regulated at the transcriptional
Connie W. H. Woo; Ricky Y. K. Man; Yaw L. Siow; Patrick C. Choy; Eric W. Y. Wan; Chak S. Lau; Karmin O
Lipopolysaccharide-induced fever in Pekin ducks is mediated by prostaglandins and nitric oxide and modulated by adrenocortical hormones.
Information on avian fever is limited, and, in particular, very little is known about the mediators and modulators of the febrile response in birds. Therefore, in this study, the possible mediatory roles of nitric oxide (NO) and prostaglandins (PGs), together with a potential modulatory role for adrenocortical hormones in the generation of fever was investigated in conscious Pekin ducks. Their body temperatures were continuously measured by abdominally implanted temperature-sensitive data loggers. The febrile response induced by intramuscular injection of LPS at a dose of 100 microg/kg was compared with and without inhibition of NO production by N-nitro-L-arginine methyl ester (L-NAME), inhibition of PG synthesis (by diclofenac), and elevation of circulating concentrations of dexamethasone and corticosterone (by exogenous administration). LPS administration induced a marked, monophasic fever with a rise in temperature of more than 1 degrees C after 3-4 h. In the presence of L-NAME, diclofenac, and adrenocorticoids at doses that had no effect upon normal body temperature in afebrile ducks, there was a significant inhibition of the LPS-induced fever. In addition, during the febrile response, the blood concentration of corticosterone was significantly elevated (from a basal level of 73.6 +/- 9.8 ng/ml to a peak level of 132.6 +/- 16.5 ng/ml). The results strongly suggest that the synthesis of both NO and PGs is a vital step in the generation of fever in birds and that the magnitude of the response is subject to modulation by adrenocorticoids. PMID:16037125
Gray, David A; Maloney, Shane K; Kamerman, Peter R
U-Bang-Haequi Tang: A Herbal Prescription that Prevents Acute Inflammation through Inhibition of NF-?B-Mediated Inducible Nitric Oxide Synthase
Since antiquity, medical herbs have been prescribed for both treatment and preventative purposes. Herbal formulas are used to reduce toxicity as well as increase efficacy in traditional Korean medicine. U-bang-haequi tang (UBT) is a herbal prescription containing Arctii fructus and Forsythia suspensa as its main components and has treated many human diseases in traditional Korean medicine. This research investigated the effects of UBT against an acute phase of inflammation. For this, we measured induction of nitric oxide (NO) and related proteins in macrophage cell line stimulated by lipopolysaccharide (LPS). Further, paw swelling was measured in carrageenan-treated rats. Carrageenan significantly induced activation of inflammatory cells and increases in paw volume, whereas oral administration of 0.3 or 1?g/kg/day of UBT inhibited the acute inflammatory response. In RAW264.7 cells, UBT inhibited mRNA and protein expression levels of iNOS. UBT treatment also blocked elevation of NO production, nuclear translocation of NF-?B, phosphorylation of I?-B? induced by LPS. Moreover, UBT treatment significantly blocked the phosphorylation of p38 and c-Jun NH2-terminal kinases by LPS. In conclusion, UBT prevented both acute inflammation in rats as well as LPS-induced NO and iNOS gene expression through inhibition of NF-?B in RAW264.7 cells.
Hwangbo, Min; Jung, Ji Yun; Ki, Sung Hwan; Park, Sang Mi; Jegal, Kyung Hwan; Lee, Ju-Hee; Kang, Seung Ho; Park, Sun-Dong; Ku, Sae Kwang; Zhao, Rong Jie; Jee, Seon Young
U-Bang-Haequi Tang: A Herbal Prescription that Prevents Acute Inflammation through Inhibition of NF-?B-Mediated Inducible Nitric Oxide Synthase.
Since antiquity, medical herbs have been prescribed for both treatment and preventative purposes. Herbal formulas are used to reduce toxicity as well as increase efficacy in traditional Korean medicine. U-bang-haequi tang (UBT) is a herbal prescription containing Arctii fructus and Forsythia suspensa as its main components and has treated many human diseases in traditional Korean medicine. This research investigated the effects of UBT against an acute phase of inflammation. For this, we measured induction of nitric oxide (NO) and related proteins in macrophage cell line stimulated by lipopolysaccharide (LPS). Further, paw swelling was measured in carrageenan-treated rats. Carrageenan significantly induced activation of inflammatory cells and increases in paw volume, whereas oral administration of 0.3 or 1?g/kg/day of UBT inhibited the acute inflammatory response. In RAW264.7 cells, UBT inhibited mRNA and protein expression levels of iNOS. UBT treatment also blocked elevation of NO production, nuclear translocation of NF-?B, phosphorylation of I?-B? induced by LPS. Moreover, UBT treatment significantly blocked the phosphorylation of p38 and c-Jun NH2-terminal kinases by LPS. In conclusion, UBT prevented both acute inflammation in rats as well as LPS-induced NO and iNOS gene expression through inhibition of NF-?B in RAW264.7 cells. PMID:24959187
Hwangbo, Min; Jung, Ji Yun; Ki, Sung Hwan; Park, Sang Mi; Jegal, Kyung Hwan; Cho, Il Je; Lee, Ju-Hee; Kang, Seung Ho; Park, Sun-Dong; Ku, Sae Kwang; Kim, Sang Chan; Zhao, Rong Jie; Jee, Seon Young; Kim, Young Woo
Soybean glyceollins mitigate inducible nitric oxide synthase and cyclooxygenase-2 expression levels via suppression of the NF-?B signaling pathway in RAW 264.7 cells
Glyceollins, produced to induce disease resistance responses against specific species, such as an incompatible pathogen Phytophthora sojae in soybeans, have the potential to exhibit anti-inflammatory activity in RAW 264.7 cells. To investigate the anti-inflammatory effects of elicited glyceollins via a signaling pathway, we studied the glyceollin signaling pathway using several assays including RNA and protein expression levels. We found that soybean glyceollins significantly reduced LPS-induced nitric oxide (NO) and prostaglandin E2 (PGE2) production, as well as the expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) via the suppression of NF-?B activation. Glyceollins also inhibited the phosphorylation of I?B? kinase (IKK), the degradation of I?B?, and the formation of NF-?B-DNA binding complex in a dose-dependent manner. Furthermore, they inhibited pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-?, interleukin (IL)-1? and IL-18, but increased the generation of the anti-inflammatory cytokine IL-10. Collectively, the present data show that glyceollins elicit potential anti-inflammatory effects by suppressing the NF-?B signaling pathway in RAW 264.7 cells.
Inhibitory effect of resveratrol dimerized derivatives on nitric oxide production in lipopolysaccharide-induced RAW 264.7 cells.
Four types of resveratrol dimerized analogues were synthesized and evaluated in vitro on LPS-induced NO production in RAW 264.7 cells. The results showed that several compounds, especially those containing 1,2-diphenyl-2,3-dihydro-1H-indene core (type I), exhibited good inhibitory activities. Among 25 analogues, 12b showed a significant inhibitory activity (49% NO production at 10 ?M, IC50=3.38 ?M). Further study revealed that compound 12b could suppress LPS-induced iNOS expression, NO production, and IL-1? release in a concentration-dependently manner. The mechanism of action (MOA) involved for its anti-inflammatory responses was through signaling pathways of p38 MAPK and JNK1/2, but not ERK1/2. PMID:23791078
Zhong, Chen; Liu, Xin-Hua; Chang, Jun; Yu, Jian-Ming; Sun, Xun
Supercritical extract of Seabuckthorn Leaves (SCE200ET) inhibited endotoxemia by reducing inflammatory cytokines and nitric oxide synthase 2 expression.
Endotoxins from infectious organisms lead to sepsis, a systemic inflammatory response, and a major cause of death. Numerous studies have shown the potential role of plants and plant-derived compounds in the suppression of LPS induced endotoxemia in vivo. In the present study, we have identified a plant namely Seabuckthorn (Hippophae rhamnoides L.) as a potent agent for the treatment of endotoxemia. The objective of the study was to investigate the influence of Supercritical Extract of Seabuckthorn Leaves (SCE200ET) and its active component Isorhamnetin (IR) on the LPS induced endotoxemia in Balb/c mice by measuring the level of nitric oxide (NO), TNF-? and IL-6. Expression of COX-2 and iNOS was measured to understand the involvement of various pathways in the mechanism of action of SCE200ET and IR. The results indicated that SCE200ET and IR inhibited LPS induced NO production by peritoneal macrophages. Cytokines mediated effector functions were influenced by the reduction of IL-6 and TNF-? production and CD40 expression was also markedly diminished in the extract or IR treated groups. In addition, the anti-inflammatory properties were further characterized by decreased expression of COX-2 and iNOS proteins. Fractionation and phytochemical analysis of the extract by RP-HPLC led to identification of isorhamnetin, as bioactive component. Thus, SCE200ET extract and its active component Isorhamnetin could be potential therapeutic agents for the treatment of endotoxin induced sepsis. PMID:24594274
Jayashankar, Bindhya; Mishra, K P; Ganju, L; Singh, S B
Nitric oxide-induced calcium release: activation of type 1 ryanodine receptor by endogenous nitric oxide.
Ryanodine receptors (RyRs), located in the sarcoplasmic/endoplasmic reticulum (SR/ER) membrane, are required for intracellular Ca2+ release that is involved in a wide range of cellular functions. In addition to Ca2+-induced Ca2+ release in cardiac cells and voltage-induced Ca2+ release in skeletal muscle cells, we recently identified another mode of intracellular Ca2+ mobilization mediated by RyR, i.e., nitric oxide-induced Ca2+ release (NICR), in cerebellar Purkinje cells. NICR is evoked by neuronal activity, is dependent on S-nitrosylation of type 1 RyR (RyR1) and is involved in the induction of long-term potentiation (LTP) of cerebellar synapses. In this addendum, we examined whether peroxynitrite, which is produced by the reaction of nitric oxide with superoxide, may also have an effect on the Ca2+ release via RyR1 and the cerebellar LTP. We found that scavengers of peroxynitrite have no significant effect either on the Ca2+ release via RyR1 or on the cerebellar LTP. We also found that an application of a high concentration of peroxynitrite does not reproduce neuronal activity-dependent Ca2+ release in Purkinje cells. These results support that NICR is induced by endogenous nitric oxide produced by neuronal activity through S-nitrosylation of RyR1. PMID:23247505
Kakizawa, Sho; Yamazawa, Toshiko; Iino, Masamitsu
Ethanol extract of Magnolia officinalis prevents lipopolysaccharide-induced memory deficiency via its antineuroinflammatory and antiamyloidogenic effects.
Magnolia bark contains several compounds such as magnolol, honokiol, 4-O-methylhonokiol, obovatol, and other neolignan compounds. These compounds have been reported to have various beneficial effects in various diseases. There is sufficient possibility that ethanol extract of Magnolia officinalis is more effective in amyloidogenesis via synergism of these ingredients. Neuroinflammation has been known to play a critical role in the pathogenesis of Alzheimer's disease (AD). We investigated whether the ethanol extract of M.?officinalis (10?mg/?kg in 0.05% ethanol) prevents memory dysfunction and amyloidogenesis in AD mouse model by intraperitoneal lipopolysaccharide (LPS, 250?�g/?kg/day for seven times) injection. We found that ethanol extract of M.?officinalis prevented LPS-induced memory deficiency as well as inhibited the LPS-induced elevation of inflammatory proteins, such as inducible nitric oxide synthase and cyclooxygenase 2, and activation of astrocytes and microglia. In particular, administration of M.?officinalis ethanol extract inhibited LPS-induced amyloidogenesis, which resulted in the inhibition of amyloid precursor protein, beta-site amyloid-precursor-protein-cleaving enzyme 1 and C99. Thus, this study shows that ethanol extract of M.?officinalis prevents LPS-induced memory impairment as well as amyloidogenesis via inhibition of neuroinflammation and suggests that ethanol extract of M.?officinalis might be a useful intervention for neuroinflammation-associated diseases such as AD. PMID:22628265
Lee, Young-Jung; Choi, Dong-Young; Yun, Yeo-Pyo; Han, Sang Bae; Kim, Hwan Mook; Lee, Kiho; Choi, Seok Hwa; Yang, Mhan-Pyo; Jeon, Hyun Soo; Jeong, Jea-Hwang; Oh, Ki-Wan; Hong, Jin Tae
Arginase Activity Mediates Retinal Inflammation in Endotoxin-Induced Uveitis
Arginase has been reported to reduce nitric oxide bioavailability in cardiovascular disease. However, its specific role in retinopathy has not been studied. In this study, we assessed the role of arginase in a mouse model of endotoxin-induced uveitis induced by lipopolysaccharide (LPS) treatment. Measurement of arginase expression and activity in the retina revealed a significant increase in arginase activity that was associated with increases in both mRNA and protein levels of arginase (Arg)1 but not Arg2. Immunofluorescence and flow cytometry confirmed this increase in Arg1, which was localized to glia and microglia. Arg1 expression and activity were also increased in cultured Muller cells and microglia treated with LPS. To test whether arginase has a role in the development of retinal inflammation, experiments were performed in mice deficient in one copy of the Arg1 gene and both copies of the Arg2 gene or in mice treated with a selective arginase inhibitor. These studies showed that LPS-induced increases in inflammatory protein production, leukostasis, retinal damage, signs of anterior uveitis, and uncoupling of nitric oxide synthase were blocked by either knockdown or inhibition of arginase. Furthermore, the LPS-induced increase in Arg1 expression was abrogated by blocking NADPH oxidase. In conclusion, these studies suggest that LPS-induced retinal inflammation in endotoxin-induced uveitis is mediated by NADPH oxidase-dependent increases in arginase activity.
Zhang, Wenbo; Baban, Babak; Rojas, Modesto; Tofigh, Sohrab; Virmani, Suvika K.; Patel, Chintan; Behzadian, M. Ali; Romero, Maritza J.; Caldwell, Robert W.; Caldwell, Ruth B.
Malabaricone C suppresses lipopolysaccharide-induced inflammatory responses via inhibiting ROS-mediated Akt/IKK/NF-?B signaling in murine macrophages.
Malabaricone C (MLB-C), isolated from nutmeg, is a phenolic diarylnonanoid that is known to exert a variety of pharmacological activities. In the present study, we investigated the molecular actions of MLB-C against lipopolysaccharide (LPS)-induced inflammatory responses in RAW264.7 cells and murine peritoneal macrophages. MLB-C inhibited the production of nitric oxide (NO), prostaglandin E(2) (PGE(2)), interleukin-6 (IL-6), and interferon-? (INF-?) in a dose-dependent manner. Consistent with NO and PGE(2) inhibition, MLB-C suppressed LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression as well as the promoter activities of COX-2 and iNOS. MLB-C pretreatment prevented LPS-induced nuclear factor-kappa B (NF-?B) activation through the inhibition of phosphorylation of I?B kinase (IKK), phosphorylation and degradation of I?B?, and nuclear translocation of NF-?B. In addition, MLB-C blocked LPS-induced serine 536 phosphorylation and transcriptional activity of RelA/p65 subunit of NF-?B. Further study demonstrated that MLB-C inhibited LPS-induced Akt phosphorylation, which is an upstream activator of NF-?B, by reducing reactive oxygen species (ROS) accumulation, without affecting phosphorylation of mitogen-activated protein kinases (MAPKs). These findings indicate that MLB-C exerts an anti-inflammatory effect through the inhibition of NF-?B activation by inhibiting interconnected ROS/Akt/IKK/NF-?B signaling pathways. PMID:22917708
Kang, Jungwon; Tae, Nara; Min, Byung Sun; Choe, Jongseon; Lee, Jeong-Hyung
The leaf extract of Spondias mombin L. displays an anti-inflammatory effect and suppresses inducible formation of tumor necrosis factor-? and nitric oxide (NO).
Extracts of Spondias mombin L. (Anacardiacea) is used in the traditional medicine of Africa and Latin America to treat many inflammatory conditions, with repeated claims of efficacy. However, there are no scientific data yet to support these claims and the mechanism through which the extract may be acting is still unknown. This study was undertaken to investigate the effects of the methanolic extract of the leaf of S. mombin (SM) on inflammation and to uncover some of the possible mechanisms that could explain any observed changes. The anti-inflammatory activity of the extract was investigated in Wistar rats using intraplantar injection of carrageenan as an in vivo model of inflammation. The effect of oral supplementation of the SM extract on tumor necrosis factor (TNF)-? levels after an intraperitoneal lipopolysaccharide (LPS; 1 mg/kg) challenge was investigated in mice. The effect of SM on TNF-? and inducible nitric oxide (iNO) production by LPS-stimulated bone marrow-derived macrophages (BM-M�) was also investigated in vitro. BM-M� were preincubated for 2 h with SM (0-100 ?g/ml), activated with LPS, and then TNF-? and NO production measured in the cell-free conditioned culture supernatant after 24 h of incubation. The study showed that pre-treatment of rats with the SM extract (at 100, 200, and 400 mg/kg, per os) caused a significant dose-related inhibition of carrageenan-induced paw edema over a 4-h period. In treated mice, LPS-inducible (systemic) TNF-? levels were found to be significantly lower as a result of their receiving the SM extract. In vitro, SM treatment caused a dose-dependent decrease in LPS-inducible TNF-? and NO production by BM-M� compared to the effects of treatment of the cells with LPS alone. Taken together, the results of these studies suggest that supplementation with SM extract can alleviate inflammatory responses and that this could possibly be via a suppression of the production of pro-inflammatory mediators and cytokines such as TNF-? and iNO. PMID:21261441
Nworu, Chukwuemeka S; Akah, Peter A; Okoye, Festus B C; Toukam, Donatien Kamdem; Udeh, Judith; Esimone, Charles O
Inducible nitric oxide synthase expression in human cerebral infarcts
The inducible or �immunological� isoform of nitric oxide synthase (iNOS) is induced in many cell types by inflammatory stimuli\\u000a and synthesizes toxic amounts of NO. In rodent models of focal cerebral ischemia, iNOS is expressed in neutrophils invading\\u000a the injured brain and in local blood vessels. Studies with iNOS inhibitors and iNOS null mice indicate that NO produced by\\u000a iNOS
Colleen Forster; H. Brent Clark; M. Elizabeth Ross; Constantino Iadecola
Acanthopanax koreanum fruit waste inhibits lipopolysaccharide-induced production of nitric oxide and prostaglandin E2 in RAW 264.7 macrophages.
The Acanthopanax koreanum fruit is a popular fruit in Jeju Island, but the byproducts of the alcoholic beverage prepared using this fruit are major agricultural wastes. The fermentability of this waste causes many economic and environmental problems. Therefore, we investigated the suitability of using A. koreanum fruit waste (AFW) as a source of antiinflammatory agents. AFWs were extracted with 80% EtOH. The ethanolic extract was then successively partitioned with hexane, CH(2)Cl(2), EtOAc, BuOH, and water. The results indicate that the CH(2)Cl(2) fraction (100 microg/mL) of AFW inhibited the LPS-induced nitric oxide (NO) and prostaglandin E(2) (PGE(2)) production in RAW 264.7 cells by 79.6% and 39.7%, respectively. These inhibitory effects of the CH(2)Cl(2) fraction of AFWs were accompanied by decreases in the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins and iNOS and COX-2 mRNA in a dose-dependent pattern. The CH(2)Cl(2) fraction of AFWs also prevented degradation of IkappaB-alpha in a dose-dependent manner. Ursolic acid was identified as major compound present in AFW, and CH(2)Cl(2) extracts by high performance liquid chromatography (HPLC). Furthermore using pure ursolic acid as standard and by HPLC, AFW and CH(2)Cl(2) extracts was found to contain 1.58 mg/g and 1.75 mg/g, respectively. Moreover, we tested the potential application of AFW extracts as a cosmetic material by performing human skin primary irritation tests. In these tests, AFW extracts did not induce any adverse reactions. Based on these results, we suggest that AFW extracts be considered possible anti-inflammatory candidates for topical application. PMID:20368786
Yang, Eun-Jin; Moon, Ji-Young; Lee, Jung-Soon; Koh, Jaesook; Lee, Nam Ho; Hyun, Chang-Gu
Acanthopanax koreanum Fruit Waste Inhibits Lipopolysaccharide-Induced Production of Nitric Oxide and Prostaglandin E2 in RAW 264.7 Macrophages
The Acanthopanax koreanum fruit is a popular fruit in Jeju Island, but the byproducts of the alcoholic beverage prepared using this fruit are major agricultural wastes. The fermentability of this waste causes many economic and environmental problems. Therefore, we investigated the suitability of using A. koreanum fruit waste (AFW) as a source of antiinflammatory agents. AFWs were extracted with 80% EtOH. The ethanolic extract was then successively partitioned with hexane, CH2Cl2, EtOAc, BuOH, and water. The results indicate that the CH2Cl2 fraction (100 ?g/mL) of AFW inhibited the LPS-induced nitric oxide (NO) and prostaglandin E2 (PGE2) production in RAW 264.7 cells by 79.6% and 39.7%, respectively. These inhibitory effects of the CH2Cl2 fraction of AFWs were accompanied by decreases in the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins and iNOS and COX-2 mRNA in a dose-dependent pattern. The CH2Cl2 fraction of AFWs also prevented degradation of I?B-? in a dose-dependent manner. Ursolic acid was identified as major compound present in AFW, and CH2Cl2 extracts by high performance liquid chromatography (HPLC). Furthermore using pure ursolic acid as standard and by HPLC, AFW and CH2Cl2 extracts was found to contain 1.58?mg/g and 1.75?mg/g, respectively. Moreover, we tested the potential application of AFW extracts as a cosmetic material by performing human skin primary irritation tests. In these tests, AFW extracts did not induce any adverse reactions. Based on these results, we suggest that AFW extracts be considered possible anti-inflammatory candidates for topical application.
Yang, Eun-Jin; Moon, Ji-Young; Lee, Jung-Soon; Koh, Jaesook; Lee, Nam Ho; Hyun, Chang-Gu
UV Induced Oxidation of Nitric Oxide
NASA Technical Reports Server (NTRS)
Nitric oxide in a gaseous stream is converted to nitrogen dioxide using oxidizing species generated at least in part using in situ UV radiation sources. The sources of the oxidizing species include oxygen and/or hydrogen peroxide. The oxygen may be a component of the gaseous stream or added to the gaseous stream, preferably near a UV radiation source, and is converted to ozone by the UV irradiation. The hydrogen peroxide is decomposed through a combination of vaporization and UV irradiation. The hydrogen peroxide is preferably stored at stable concentration levels, i.e., approximately 50% by volume and increased in concentration in a continuous process preceding vaporization within the flow channel of the gaseous stream and in the presence of the UV radiation sources.
Parrish, Clyde, F. (Inventor); Luecke, Dale E. (Inventor)
Plumbagin, a vitamin K3 analogue, abrogates lipopolysaccharide-induced oxidative stress, inflammation and endotoxic shock via NF-?B suppression.
Plumbagin has been reported to modulate cellular redox status and suppress NF-?B. In the present study, we investigated the effect of plumbagin on lipopolysaccharide (LPS)-induced endotoxic shock, oxidative stress and inflammatory parameters in vitro and in vivo. Plumbagin inhibited LPS-induced nitric oxide, TNF-?, IL-6 and prostaglandin-E2 production in a concentration-dependent manner in RAW 264.7 cells without inducing any cell death. Plumbagin modulated cellular redox status in RAW cells. Plumbagin treatment significantly reduced MAPkinase and NF-?B activation in macrophages. Plumbagin prevented mice from endotoxic shock-associated mortality and decreased serum levels of pro-inflammatory markers. Plumbagin administration ameliorated LPS-induced oxidative stress in peritoneal macrophages and splenocytes. Plumbagin also attenuated endotoxic shock-associated changes in liver and lung histopathology and decreased the activation of ERK and NF-?B in liver. These findings demonstrate the efficacy of plumbagin in preventing LPS-induced endotoxemia and also provide mechanistic insights into the anti-inflammatory effects of plumbagin. PMID:24234154
Checker, Rahul; Patwardhan, Raghavendra S; Sharma, Deepak; Menon, Jisha; Thoh, Maikho; Sandur, Santosh K; Sainis, Krishna B; Poduval, T B
Protective effect of pretreatment with low-dose lipopolysaccharide on D-galactosamine-induced acute liver failure
Background and aims: The effect of low-dose lipopolysaccharide (LPS) induced nitric oxide (NO) on liver damage and survival in rats with acute liver failure caused by a lethal dose of D-galactosamine (D-gal) was studied. Results: Ninety percent of control animals died within 4 days after D-gal injection, but pretreatment with low-dose LPS significantly decreased mortality to 5%. There was marked
Toru Kono; Hiromi Kotani; Toshiyuki Asama; Noriaki Mamiya; Kei Ohara; Masashi Yoneda; Jun Iwamoto; Shinichi Kasai
Nitric oxide mediates relative airway hyporesponsiveness to lipopolysaccharide in surfactant protein A-deficient mice.
Surfactant protein A (SP-A) mediates innate immune cell responses to LPS, a cell wall component of gram-negative bacteria that is found ubiquitously in the environment and is associated with adverse health effects. Inhaled LPS induces lung inflammation and increases airway responsiveness (AR). However, the role of SP-A in mediating LPS-induced AR is not well-defined. Nitric oxide (NO) is described as a potent bronchodilator, and previous studies showed that SP-A modulates the LPS-induced production of NO. Hence, we tested the hypothesis that increased AR, observed in response to aerosolized LPS exposure, would be significantly reduced in an SP-A-deficient condition. Wild-type (WT) and SP-A null (SP-A(-/-)) mice were challenged with aerosolized LPS. Results indicate that despite similar inflammatory indices, LPS-treated SP-A(-/-) mice had attenuated AR after methacholine challenge, compared with WT mice. The attenuated AR could not be attributed to inherent differences in SP-D concentrations or airway smooth muscle contractile and relaxation properties, because these measures were similar between WT and SP-A(-/-) mice. LPS-treated SP-A(-/-) mice, however, had elevated nitrite concentrations, inducible nitric oxide synthase (iNOS) expression, and NOS activity in their lungs. Moreover, the administration of the iNOS-specific inhibitor 1400W completely abrogated the attenuated AR. Thus, when exposed to aerosolized LPS, SP-A(-/-) mice demonstrate a relative airway hyporesponsiveness that appears to be mediated at least partly via an iNOS-dependent mechanism. These findings may have clinical significance, because recent studies reported associations between surfactant protein polymorphisms and a variety of lung diseases. PMID:20348208
Pastva, Amy M; Walker, Julia K L; Maddox, Lee A; Mukherjee, Sambuddho; Giamberardino, Charles; Hsia, Bethany; Potts, Erin; Zhu, Hongmei; Degan, Simone; Sunday, Mary E; Lawson, Barbara L; Korfhagen, Thomas R; Schwartz, David A; Eu, Jerry P; Foster, William M; McMahon, Timothy J; Que, Loretta; Wright, Jo Rae
Effect of nitric oxide on megakaryocyte growth induced by thrombopoietin
The present study investigated the effect of nitric oxide (NO) on megakaryocyte (Mk) proliferation induced by thrombopoietin (TPO). Low-density mononuclear cells (MNCs) and CD34+ cells from human bone marrow (BM) were cultured in liquid medium in the presence of sodium nitroprusside (SNP) or (Z)-1-[2-(aminoethyl)-N-(2-ammonioethyl) amino] diazen-1-ium-1, 2-diolate (DETA\\/NO) and then stimulated with TPO. Mk number decreased in both NO donors,
M Schattner; R. G Pozner; I Engelberger; A Gorostizaga; N Maugeri; R Gomez; A Pasqualini; O Torres; M. A Lazzari
Cardiac Weight in Hypertension Induced by Nitric Oxide Synthase Blockade
Wistar rats given a nitric oxide synthase inhibitor, iVc-nitro-L-arginine-methyl ester (L-NAME), for 4 weeks develop time- and dose-dependent hypertension without cardiac hypertrophy. This initial study of the relation between left ventricular weight and L-NAME-induced hypertension has now been extended by giving 50 mg\\/kg per day L-NAME to Wistar rats (n=30) for 8 weeks and comparing results with those from control
Jean-Francois Arnal; Abdel-Ilah El Amrani; Gilles Chatellier; Joel Menard; Jean-Baptiste Michel
Radiation-induced decomposition of the tributyl phosphate-nitric acid system: Role of nitric acid
Radiation-induced decomposition of tributyl phosphate-nitric acid as a two-component system has been studied. Degradation products, dibutylphosphoric acid (DBP) and monobutylphosphoric acid (MBP), were determined by separation-extraction method. 0.59, 0.78 and 1.38 are the G (DBP) values and 0.15, 0.17 and 0.13 are the G (MBP) values obtained for pure TBP, TBP-3M HNO3 extract and TBP-5M HNO3 extract, respectively. G (�HNO3)
M. V. Krishnamurthy; R. Sampathkumar
Antidepressant-like effect of nitric oxide synthase inhibitors and sildenafil against lipopolysaccharide-induced depressive-like behavior in mice.
Inflammation, oxidative and nitrosative stress underlie depression being assessed in rodents by the systemic administration of lipopolysacharide (LPS). There is an increasing body of evidence of an involvement of nitric oxide (NO) pathway in depression, but this issue was not investigated in LPS-induced model. Thus, herein we evaluated the effects of NO-pathway-modulating drugs, named aminoguanidine, l-NAME, sildenafil and l-arginine, on the behavioral (forced swimming test [FST], sucrose preference [SPT] and prepulse inhibition [PPI] of the startle) and neurochemical (glutathione [GSH], lipid peroxidation, IL-1?) alterations in the prefrontal cortex, hippocampus and striatum as well as in BDNF levels in the hippocampus 24h after LPS (0.5mg/kg, i.p.) administration, a time-point related to depressive-like behavior. Twenty-four hours post LPS there was an increase in immobility time in the FST, decrease in sucrose preference and PPI levels accompanied by a decrease in GSH levels and an increase in lipid peroxidation, IL-1? and hippocampal BDNF levels suggestive of a depressive-like state. The pretreatment with the NOS inhibitors, l-NAME and aminoguanidine as well as sildenafil prevented the behavioral and neurochemical alterations induced by LPS, although sildenafil and l-NAME were not able to prevent the increase in hippocampal BDNF levels induced by LPS. The iNOS inhibitor, aminoguanidine, and imipramine prevented all behavioral and neurochemical alterations induced by LPS. l-arginine did not prevent the alterations in immobility time, sucrose preference and GSH induced by LPS. Taken together our results show that the NO-cGMP pathway is important in the modulation of the depressive-like alterations induced by LPS. PMID:24662848
Tomaz, V S; Cordeiro, R C; Costa, A M N; de Lucena, D F; Nobre J�nior, H V; de Sousa, F C F; Vasconcelos, S M M; Vale, M L; Quevedo, J; Mac�do, D
Nonsteroidal Anti-inflammatory Drugs Inhibit Expression of the Inducible Nitric Oxide Synthase Gene
Increased nitric oxide production is associated with acute and chronic inflammatory processes. Accordingly, we tested the hypothesis that the therapeutic action of nonsteroidal anti-inflammatory drugs could be attributed at least in part to inhibition of excess nitric oxide production. We report here that sodium salicylate, aspirin, ibuprofen, and indomethacin markedly inhibited the appearance of the inducible inflammatory nitric oxide synthase
E. E. Aeberhard; S. A. Henderson; N. S. Arabolos; J. M. Griscavage; F. E. Castro; C. T. Barrett; L. J. Ignarro
Expression, activity and functional significance of inducible nitric oxide synthase in the failing human heart
Objectives. The study was designed to evaluate the functional impact of nitric oxide (NO) generation within the myocardium on cardiac contraction in the failing human heart.Background. Heart failure is associated with activation of cytokines and expression of inducible nitric oxide synthase (NOS II), which generates NO from L-arginine. Nitric oxide has been shown to modulate myocardial performance, raising the possibility
Helmut Drexler; Stephanie K�stner; Armin Strobel; Roland Studer; Otto E Brodde; Gerd Hasenfu�
Antioxidant properties of lutein contribute to the protection against lipopolysaccharide-induced uveitis in mice
Background Lutein is an important eye-protective nutrient. This study investigates the protective effects and mechanisms of lutein on lipopolysaccharides (LPS)-induced uveitis in mice. Methods Lutein, suspended in drinking water at a final concentration of 12.5 and 25 mg/mL, was administered to mice at 0.1 mL/10 g body weight for five consecutive days. Control and model group received drinking water only. Uveitis was induced by injecting LPS (100 mg per mouse) into the footpad in the model and lutein groups on day 5 after the last drug administration. Eyes of the mice were collected 24 hours after the LPS injection for the detection of indicators using commercial kits and reverse transcription-polymerase chain reaction. Results LPS-induced uveitis was confirmed by significant pathological damage and increased the nitric oxide level in eye tissue of BALB/C mice 24 hours after the footpad injection. The elevated nitric oxide level was significantly reduced by oral administration of lutein (125 and 500 mg/kg/d for five days) before LPS injection. Moreover, lutein decreased the malondialdehyde content, increased the oxygen radical absorbance capacity level, glutathione, the vitamin C contents and total superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities. Lutein further increased expressions of copper-zinc SOD, manganese SOD and GPx mRNA. Conclusion The antioxidant properties of lutein contribute to the protection against LPS-induced uveitis, partially through the intervention of inflammation process.
?-Lipoic acid protects mitochondrial enzymes and attenuates lipopolysaccharide-induced hypothermia in mice.
Hypothermia is a key symptom of sepsis, but the mechanism(s) leading to hypothermia during sepsis is largely unknown and thus no effective therapy is available for hypothermia. Therefore, it is important to investigate the mechanism and develop effective therapeutic methods. Lipopolysaccharide (LPS)-induced hypothermia accompanied by excess nitric oxide (NO) production leads to a reduction in energy production in wild-type mice. However, mice lacking inducible nitric oxide synthase did not suffer from LPS-induced hypothermia, suggesting that hypothermia is associated with excess NO production during sepsis. This observation is supported by the treatment of wild-type mice with ?-lipoic acid (LA) in that it effectively attenuates LPS-induced hypothermia with decreased NO production. We also found that LA partially restored ATP production, and activities of the mitochondrial enzymes involved in energy metabolism, which were inhibited during sepsis. These data suggest that hypothermia is related to mitochondrial dysfunction, which is probably compromised by excess NO production and that LA administration attenuates hypothermia mainly by protecting mitochondrial enzymes from NO damage. PMID:24675228
Hiller, Sylvia; DeKroon, Robert; Xu, Longquan; Robinette, Jennifer; Winnik, Witold; Alzate, Oscar; Simington, Stephen; Maeda, Nobuyo; Yi, Xianwen
Yu Ping Feng San, an Ancient Chinese Herbal Decoction, Regulates the Expression of Inducible Nitric Oxide Synthase and Cyclooxygenase-2 and the Activity of Intestinal Alkaline Phosphatase in Cultures
Yu Ping Feng San (YPFS), a Chinese herbal decoction comprising Astragali Radix (AR; Huangqi), Atractylodis Macrocephalae Rhizoma (AMR; Baizhu), and Saposhnikoviae Radix (SR; Fangfeng), has been used clinically to treat inflammatory bowel diseases (IBD). Previously, we demonstrated a dual role of YPFS in regulating cytokine release in cultured macrophages. In this study, we elucidated the anti-inflammatory effect of YPFS that is mediated through modulating the expression of three key enzymes involved in IBD: inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and intestinal alkaline phosphatase (IALP). In a lipopolysaccharide (LPS)-induced chronic-inflammation model of cultured murine macrophages, YPFS treatment suppressed the activation of iNOS and COX-2 expression in a dose-dependent manner. Conversely, application of YPFS in cultured small intestinal enterocytes markedly induced the expression of IALP in a time-dependent manner, which might strengthen the intestinal detoxification system. A duality of YPFS in modulating the expression of iNOS and COX-2 was determined here. The expression of iNOS and COX-2 in macrophages was induced by YPFS, and this activation was partially blocked by the NF-?B-specific inhibitor BAY 11-7082, indicating a role of NF-?B signaling. These YPFS-induced changes in gene regulation strongly suggest that the anti-inflammatory effects of YPFS are mediated through the regulation of inflammatory enzymes.
Du, Crystal Y. Q.; Choi, Roy C. Y.; Dong, Tina T. X.; Lau, David T. W.; Tsim, Karl W. K.
The spin trap 5,5-dimethyl-1-pyrroline N-oxide inhibits lipopolysaccharide-induced inflammatory response in RAW 264.7 cells
Aim Exposure of macrophages to lipopolysaccharide (LPS) induces oxidative and inflammatory stresses, which cause cell damage. Antioxidant and anti-inflammatory properties have been attributed to the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO), commonly used in free radical analysis, but these aspects of DMPO have been little explored. In this study, we sought to establish the anti-inflammatory activity of DMPO, presumably by removing free radicals which otherwise help activate inflammatory response and damage cells. Main methods RAW 264.7 macrophages were treated with LPS and/or DMPO for different time points, cell damage, production of inflammatory mediators, inducible nitric oxide synthase (iNOS) expression, NF-?B p65 activation, phosphorylation of MAPKs and Akt, and intracellular reactive oxygen species (ROS) were determined. Key findings After cells were treated with LPS and/or DMPO for 24 h, DMPO reduced the LPS-induced inflammatory response as indicated by downregulated iNOS expression and production of inflammatory mediators. Accordingly, DMPO protected cells from LPS-induced cytotoxicity. In order to understand the mechanistic basis of these DMPO effects, the NF-?B p65 activation and the phosphorylation of MAPKs and Akt were examined. We found, by assaying cells treated with LPS and/or DMPO for 15-60 min, that DMPO inhibited the phosphorylation of MAPKs, Akt, and I?B?, and reduced the NF-?B p65 translocation. Furthermore, we demonstrated that DMPO inhibited LPS-induced ROS production. Significance DMPO showed the anti-inflammatory activity and attenuated LPS-induced cell damage, most likely by reducing ROS production and thus preventing the subsequent inflammatory activation and damage.
Zhai, Zili; Gomez-Mejiba, Sandra E.; Zhu, Hua; Lupu, Florea; Ramirez, Dario C.
PPAR? agonists inhibit nitric oxide production by enhancing iNOS degradation in LPS-treated macrophages
Background and purpose: Nitric oxide (NO) production through the inducible nitric oxide synthase (iNOS) pathway is increased in response to pro-inflammatory cytokines and bacterial products. In inflammation, NO has pro-inflammatory and regulatory effects. Peroxisome proliferator-activated receptors (PPARs), members of the nuclear steroid receptor superfamily, regulate not only metabolic but also inflammatory processes. The aim of the present study was to investigate the role of PPAR? in the regulation of NO production and iNOS expression in activated macrophages. Experimental approach: The effects of PPAR? agonists were investigated on iNOS mRNA and protein expression, on NO production and on the activation of transcription factors NF-?B and STAT1 in J774 murine macrophages exposed to bacterial lipopolysaccharide (LPS). Key results: PPAR? agonists GW7647 and WY14643 reduced LPS-induced NO production in a dose-dependent manner as measured by the accumulation of nitrite into the culture medium. However, PPAR? agonists did not alter LPS-induced iNOS mRNA expression or activation of NF-?B or STAT1 which are important transcription factors for iNOS. Nevertheless, iNOS protein levels were reduced by PPAR? agonists in a time-dependent manner. The reduction was markedly greater after 24?h incubation than after 8?h incubation. Treatment with the proteasome inhibitors, lactacystin or MG132, reversed the decrease in iNOS protein levels caused by PPAR? agonists. Conclusions and implications: The results suggest that PPAR? agonists reduce LPS-induced iNOS expression and NO production in macrophages by enhancing iNOS protein degradation through the proteasome pathway. The results offer an additional mechanism underlying the anti-inflammatory effects of PPAR? agonists.
Paukkeri, E-L; Leppanen, T; Sareila, O; Vuolteenaho, K; Kankaanranta, H; Moilanen, E
Synthesis of new heterocyclic lupeol derivatives as nitric oxide and pro-inflammatory cytokine inhibitors.
A series of heterocyclic derivatives including indoles, pyrazines along with oximes and esters were synthesized from lupeol and evaluated for anti-inflammatory activity through inhibition of lipopolysaccharide (LPS) induced nitric oxide (NO) production in RAW 264.7 and J774A.1 cells. All the synthesized molecules of lupeol were found to be more active in inhibiting NO production with an IC50 of 18.4-48.7?M in both the cell lines when compared to the specific nitric oxide synthase (NOS) inhibitor, L-NAME (IC50=69.21 and 73.18?M on RAW 264.7 and J774A.1 cells, respectively). The halogen substitution at phenyl ring of indole moiety leads to potent inhibition of NO production with half maximal concentration ranging from 18.4 to 41.7?M. Furthermore, alkyl (11, 12) and p-bromo/iodo (15, 16) substituted compounds at a concentration of 20?g/mL exhibited mild inhibition (29-42%) of LPS-induced tumor necrosis factor alpha (TNF-?) and weak inhibition (10-22%) towards interleukin 1-beta (IL-1?) production in both the cell lines. All the derivatives were found to be non-cytotoxic when tested at their IC50 (?M). These findings suggest that the derivatives of lupeol could be a lead to potent inhibitors of NO. PMID:24909081
Bhandari, Pamita; Patel, Neeraj Kumar; Bhutani, Kamlesh Kumar
Expression and activity of nitric oxide synthase isoforms in methamphetamine-induced striatal dopamine toxicity.
Nitric oxide is implicated in methamphetamine (METH)-induced neurotoxicity; however, the source of the nitric oxide has not been identified. Previous work has also revealed that animals with partial dopamine loss induced by a neurotoxic regimen of methamphetamine fail to exhibit further decreases in striatal dopamine when re-exposed to methamphetamine 7-30 days later. The current study examined nitric oxide synthase expression and activity and protein nitration in striata of animals administered saline or neurotoxic regimens of methamphetamine at postnatal days 60 and/or 90, resulting in four treatment groups: Saline:Saline, METH:Saline, Saline:METH, and METH:METH. Acute administration of methamphetamine on postnatal day 90 (Saline:METH and METH:METH) increased nitric oxide production, as evidenced by increased protein nitration. Methamphetamine did not, however, change the expression of endothelial or inducible isoforms of nitric oxide synthase, nor did it change the number of cells positive for neuronal nitric oxide synthase mRNA expression or the amount of neuronal nitric oxide synthase mRNA per cell. However, nitric oxide synthase activity in striatal interneurons was increased in the Saline:METH and METH:METH animals. These data suggest that increased nitric oxide production after a neurotoxic regimen of methamphetamine results from increased nitric oxide synthase activity, rather than an induction of mRNA, and that constitutively expressed neuronal nitric oxide synthase is the most likely source of nitric oxide after methamphetamine administration. Of interest, animals rendered resistant to further methamphetamine-induced dopamine depletions still show equivalent degrees of methamphetamine-induced nitric oxide production, suggesting that nitric oxide production alone in response to methamphetamine is not sufficient to induce acute neurotoxic injury. PMID:23230214
Friend, Danielle M; Son, Jong H; Keefe, Kristen A; Fricks-Gleason, Ashley N
Nitric oxide synthases and cyclophosphamide-induced cystitis in rats.
The role of inducible (iNOS) and neuronal nitric oxide (nNOS) synthases and of tachykinin NK1 receptors on the pathogenesis of cyclophosphamide (CYP)-induced cystitis was investigated, in rats. CYP-induced cystitis was characterized by large increases in bladder-protein plasma extravasation (PPE), increases in the urinary excretion of nitric oxide (NO) metabolites and histological evidences of urothelial damage, edema, extensive white blood cell infiltrates and vascular congestion of the bladder. The specific iNOS inhibitor, S-methylthiourea (MITU), produced marked inhibition (>90%) of CYP-induced increases in PPE associated with amelioration of tissue inflammatory changes. Treatment with 7-nitroindazole (7-NI; 20, 40 and 80 mg/kg), a selective nNOS inhibitor, did not significantly reduce CYP-induced increases in PPE and failed to produce histological improvement. In addition, treatment with MITU, but not with 7-NI, inhibited the increases in the urinary excretion of NO metabolites induced by CYP treatment. WIN 51,708 (17-beta-hydroxy-17-alpha-ethynyl-androstano[3,2-b]pyrimido[1,2-a]benzimidazole; WIN), a selective NK1-receptor antagonist, reduced the increases in EPP and ameliorated the inflammatory changes in the bladder induced by CYP. However, the maximal degree of protection achieved with WIN was significantly less than that produced by MITU. Combined treatment with the iNOS inhibitor and the NK1 antagonist produced no greater effect than that produced by the iNOS inhibitor alone. Our results suggest that NO plays a fundamental role in the production of the cystitis associated with CYP treatment. The iNOS, and not nNOS, seems responsible for the inflammatory changes. Part of the increases in NO may due to activation of NK1 receptors by neuropeptides such as substance P possibly released from primary afferent fibers. PMID:11284451
Alfieri, A B; Malave, A; Cubeddu, L X
Effect of l-arginine�nitric oxide system on chemical-induced diabetes mellitus
Several in vitro studies have suggested that nitric oxide may be the mediator of cytokine-induced beta-cell destruction. On the other hand, in vivo studies have given conflicting results: some studies suggesting that nitric oxide synthase inhibitors do not suppress streptozotocin-induced diabetes in mice, while others revealed that nitric oxide synthase inhibitors can reduce the incidence of insulin-dependent diabetes mellitus in
I. Krishna Mohan; U. N. Das
Phenylpropanoid ester from Zingiber officinale and their inhibitory effects on the production of nitric oxide.
A new phenylpropanoid ester mixture, (E)-geranylferulic acid (1a) and (Z)-geranylferulic acid (1b), along with 13 known compounds, [6]-gingerol (2), [8]-gingerol (3), [10]-gingerdione (4), 1-dehydro-[6]-gingerdione (5), 1-dehydro-[8]-gingerdione (6), [6]-paradol (7), [8]-paradol (8), [6]-gingeroldiacetate (9), 6-hydroxy-[6]-shogaol (10), galanolactone (11), trans-�-sesquiphellandrol (12), trans-sesquipiperitol (13), and 4?,5?-dihydroxybisabola-2,10-diene (14) were isolated from ethanol extract of Zingiber officinale. Their structures were determined based on the spectroscopic (1D, 2D-NMR and MS) and chemical evidence. All of the isolates were evaluated for their potential to inhibit LPS-induced production of nitric oxide in murine macrophage RAW264.7 cells. Compounds 1-12 were found to inhibit nitric oxide production with IC(50) values ranging from 5.5 to 28.5 ?M. PMID:22370785
Hong, Seong Su; Oh, Joa Sub
Nitric Oxide Signaling in Hypergravity-Induced Neuronal Plasticity
NASA Technical Reports Server (NTRS)
The goal of this research project was to identify the neurons and circuits in the vestibular nuclei and nucleus prepositus hypoglossi that utilize nitric oxide (NO) for intercellular signaling during gravity-induced plasticity. This objective was pursued using histochemical and immunocytochemical approaches to localize NO-producing neurons and characterize the fine morphology of the cells in ground-based studies of normal rats, rats adapted to hypergravity, and rats adapted to hypergravity and then re-adapted to the 1G environment. NO-producing neurons were identified and studied using four methodologies: i) immunocytochemistry employing polyclonal antibodies directed against neuronal nitric oxide synthase (nNOS), to provide an indication of the capacity of a cell for NO production; ii) immunocytochemistry employing a monoclonal antibody directed against L-citrulline, to provide an indirect index of the enzyme's activity; iii) histochemistry based on the NADPH-diaphorase reaction, for fuI1 cytological visualization of neurons; and iv) double immunofluorescence to co-localize nNOS and L-citrulline in individual vestibular nuclei (VN) and neurons.
Holstein, Gay R.
Mechanism of Inducible Nitric Oxide Synthase Exclusion from Mycobacterial Phagosomes
Mycobacterium tuberculosis is sensitive to nitric oxide generated by inducible nitric oxide synthase (iNOS). Consequently, to ensure its survival in macrophages, M. tuberculosis inhibits iNOS recruitment to its phagosome by an unknown mechanism. Here we report the mechanism underlying this process, whereby mycobacteria affect the scaffolding protein EBP50, which normally binds to iNOS and links it to the actin cytoskeleton. Phagosomes harboring live mycobacteria showed reduced capacity to retain EBP50, consistent with lower iNOS recruitment. EBP50 was found on purified phagosomes, and its expression increased upon macrophage activation, paralleling expression changes seen with iNOS. Overexpression of EBP50 increased while EBP50 knockdown decreased iNOS recruitment to phagosomes. Knockdown of EBP50 enhanced mycobacterial survival in activated macrophages. We tested another actin organizer, coronin-1, implicated in mycobacterium-macrophage interaction for contribution to iNOS exclusion. A knockdown of coronin-1 resulted in increased iNOS recruitment to model latex bead phagosomes but did not increase iNOS recruitment to phagosomes with live mycobacteria and did not affect mycobacterial survival. Our findings are consistent with a model for the block in iNOS association with mycobacterial phagosomes as a mechanism dependent primarly on reduced EBP50 recruitment.
Davis, Alexander S; Vergne, Isabelle; Master, Sharon S; Kyei, George B; Chua, Jennifer; Deretic, Vojo
Regulation of the expression of inducible nitric oxide synthase.
Nitric oxide (NO) generated by the inducible isoform of nitric oxide synthase (iNOS) is involved in complex immunomodulatory and antitumoral mechanisms and has been described to have multiple beneficial microbicidal, antiviral and antiparasital effects. However, dysfunctional induction of iNOS expression seems to be involved in the pathophysiology of several human diseases. Therefore iNOS has to be regulated very tightly. Modulation of expression, on both the transcriptional and post-transcriptional level, is the major regulation mechanism for iNOS. Pathways resulting in the induction of iNOS expression vary in different cells or species. Activation of the transcription factors NF-kappaB and STAT-1alpha and thereby activation of the iNOS promoter seems to be an essential step for the iNOS induction in most human cells. However, at least in the human system, also post-transcriptional mechanisms involving a complex network of RNA-binding proteins build up by AUF1, HuR, KSRP, PTB and TTP is critically involved in the regulation of iNOS expression. Recent data also implicate regulation of iNOS expression by non-coding RNAs (ncRNAs). PMID:20438856
Pautz, Andrea; Art, Julia; Hahn, Susanne; Nowag, Sebastian; Voss, Cornelia; Kleinert, Hartmut
Down-regulatory effect of usnic acid on nuclear factor-kappaB-dependent tumor necrosis factor-alpha and inducible nitric oxide synthase expression in lipopolysaccharide-stimulated macrophages RAW 264.7.
The purpose of this study was to investigate the molecular mechanisms that are responsible for the antiinflammatory effect of usnic acid (UA). UA is one of the most common and abundant lichen metabolites. The present study examined the effects of UA on the tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) production induced by lipopolysaccharide (LPS) in RAW264.7 macrophages and the underlying molecular mechanisms. UA decreased the TNF-alpha level in LPS-stimulated RAW264.7 macrophages in dose-dependent manner, the IC(50) value was 12.8 microM. RT-PCR analysis indicated that it inhibited TNF-alpha mRNA expression. Furthermore, it inhibited NO production in LPS-activated RAW264.7 macrophages, the IC(50) value was 4.7 microM. Western blot analysis showed that UA attenuated LPS-induced synthesis of iNOS protein and nuclear translocation of NF-kappaB p65 in the macrophages, in parallel. UA also inhibited LPS-mediated I-kappaBalpha degradation. Taken together, this suggests that UA has an antiinflammatory effect by inhibiting TNF-alpha and iNOS expression, possibly through suppression of nuclear translocation of NF-kappaB p65 and I-kappaBalpha degradation. PMID:19003951
Jin, Ju-qing; Li, Cui-qin; He, Lang-chong
Nonsteroidal anti-inflammatory drugs inhibit expression of the inducible nitric oxide synthase gene.
Increased nitric oxide production is associated with acute and chronic inflammatory processes. Accordingly, we tested the hypothesis that the therapeutic action of nonsteroidal anti-inflammatory drugs could be attributed at least in part to inhibition of excess nitric oxide production. We report here that sodium salicylate, aspirin, ibuprofen, and indomethacin markedly inhibited the appearance of the inducible inflammatory nitric oxide synthase in rat alveolar macrophages activated with lipopolysaccharide and interferon gamma. We attribute the mechanism of nitric oxide synthase inhibition by nonsteroidal anti-inflammatory drugs to pretranslational control of enzyme expression and not to direct inhibition of enzymatic activity. These observations indicate that the chronic anti-inflammatory action of nonsteroidal anti-inflammatory drugs may be due not only to inhibition of prostaglandin synthesis but also to inhibition of inducible nitric oxide synthase gene expression and nitric oxide synthesis. PMID:7535524
Aeberhard, E E; Henderson, S A; Arabolos, N S; Griscavage, J M; Castro, F E; Barrett, C T; Ignarro, L J
In vivo Expression of Inducible Nitric Oxide Synthase in Experimentally Induced Neurologic Diseases
The purpose of this study was to investigate the induction of inducible nitric oxide synthase (iNOS) mRNA in the brain tissue of rats and mice under the following experimental conditions: in rats infected with borna disease virus and rabies virus, in mice infected with herpes simplex virus, and in rats after the induction of experimental allergic encephalitis. The results showed
Hilary Koprowski; Yong Mu Zheng; Ellen Heber-Katz; Nigel Fraser; Lucy Rorke; Zhen Fang Fu; Cathleen Hanlon; Bernhard Dietzschold
Methanol Extract of Antrodia camphorata Protects against Lipopolysaccharide-Induced Acute Lung Injury by Suppressing NF-?B and MAPK Pathways in Mice.
Antrodia camphorata (AC) has been used as a herbal medicine for drug intoxication for the treatment of inflammation syndromes and liver-related diseases in Taiwan. This study demonstrates the protective effect of the methanol extract of AC (MAC) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. Mice were treated with MAC 1 h before the intratracheal (I.T.) instillation of LPS challenge model. Lung injury was evaluated 6 h after LPS induction. Pretreatment with MAC markedly improved LPS-induced histological alterations and edema in lung tissues. Moreover, MAC also inhibited the release of pro-inflammatory mediators such as nitric oxide (NO), tumor necrosis factor alpha (TNF-?), interleukin-1 beta (IL-1?), and IL-6 at 6 h in the bronchoalveolar lavage fluid (BALF) during LPS-induced lung injury. Furthermore, MAC reduced total cell number and protein concentrations in the BALF the pulmonary wet/dry weight (W/D) ratio, and myeloperoxidase activity and enhanced superoxide dismutase (SOD) activity in lung tissues. MAC also efficiently blocked protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and phosphorylation of mitogen-activated protein kinases (MAPKs) and inhibited the degradation of nuclear factor-kappa B (NF-?B) and I?B?. This is the first investigation in which MAC inhibited acute lung edema effectively, which may provide a potential target for treating ALI. MAC may utilize the NF-?B and MAPKs pathways and the regulation of SOD activity to attenuate LPS-induced nonspecific pulmonary inflammation. PMID:24849405
Huang, Guan-Jhong; Deng, Jeng-Shyan; Chen, Chin-Chu; Huang, Ching-Jang; Sung, Ping-Jyun; Huang, Shyh-Shyun; Kuo, Yueh-Hsiung
Potential chemoprevention of LPS-stimulated nitric oxide and prostaglandin E? production by ?-L-rhamnopyranosyl-(1?6)-?-D-glucopyranosyl-3-indolecarbonate in BV2 microglial cells through suppression of the ROS/PI3K/Akt/NF-?B pathway.
?-l-Rhamnopyranosyl-(1?6)-?-d-glucopyranosyl-3-indolecarbonate (RG3I) is a chemical constituent isolated from the commonly used Asian traditional medicinal plant, Clematis mandshurica; however, no studies have been reported on its anti-inflammatory properties. In the present study, we found that RG3I attenuates the lipopolysaccharide (LPS)-induced DNA-binding activity of nuclear factor-?B (NF-?B) via the dephosphorylation of PI3K/Akt in BV2 microglial cells, leading to a suppression of nitric oxide (NO) and prostaglandin E2 (PGE2) production, along with that of their regulatory genes, inducible NO synthase (iNOS) and cyclooxygenase-2 (Cox-2). Further, the PI3K/Akt inhibitor, LY294002 diminished the expression of LPS-stimulated iNOS and COX-2 genes by suppressing NF-?B activity. Moreover, RG3I significantly inhibited LPS-induced reactive oxygen species (ROS) generation similar to the ROS inhibitors, N-acetylcysteine (NAC) and glutathione (GSH). Notably, NAC and GSH abolished the LPS-induced expression of iNOS and Cox-2 in BV2 microglial cells by inhibiting NF-?B activity. Taken together, our data indicate that RG3I suppresses the production of proinflammatory mediators such as NO and PGE2 as well as their regulatory genes in LPS-stimulated BV2 microglial cells by inhibiting the PI3K/Akt- and ROS-dependent NF-?B signaling pathway, suggesting that RG3I may be a good candidate to regulate LPS-induced inflammatory response. PMID:24486459
Dilshara, Matharage Gayani; Lee, Kyoung-Tae; Choi, Yung Hyun; Moon, Dong-Oh; Lee, Hak-Ju; Yun, Sung Gyu; Kim, Gi-Young
TIPE2 Negatively Regulates Inflammation by Switching Arginine Metabolism from Nitric Oxide Synthase to Arginase
TIPE2, the tumor necrosis factor (TNF)-alpha-induced protein 8-like 2 (TNFAIP8L2), plays an essential role in maintaining immune homeostasis. It is highly expressed in macrophages and negatively regulates inflammation through inhibiting Toll-like receptor signaling. In this paper, we utilized RAW264.7 cells stably transfected with a TIPE2 expression plasmid, as well as TIPE2-deficient macrophages to study the roles of TIPE2 in LPS-induced nitric oxide (NO) and urea production. The results showed that TIPE2-deficiency significantly upregulated the levels of iNOS expression and NO production in LPS-stimulated macrophages, but decreased mRNA levels of arginase I and urea production. However, TIPE2 overexpression in macrophages was capable of downregulating protein levels of LPS-induced iNOS and NO, but generated greater levels of arginase I and urea production. Furthermore, TIPE2?/? mice had higher iNOS protein levels in lung and liver and higher plasma NO concentrations, but lower levels of liver arginase I compared to LPS-treated WT controls. Interestingly, significant increases in I?B degradation and phosphorylation of JNK, p38, and I?B were observed in TIPE2-deficient macrophages following LPS challenge. These results strongly suggest that TIPE2 plays an important role in shifting L-arginase metabolism from production of NO to urea, during host inflammatory response.
Geng, Minghong; Zhang, Wenqian; Cui, Jian; Liu, Suxia
Opposite effect of oxidative stress on inducible nitric oxide synthase and haem oxygenase-1 expression in intestinal inflammation: anti-inflammatory effect of carbon monoxide.
Inducible nitric oxide synthase (iNOS) is expressed in intestinal epithelial cells (IEC) of patients with active inflammatory bowel disease (IBD) and in IEC of endotoxaemic rats. The induction of iNOS in IEC is an element of the NF-kappaB-mediated survival pathway. Haem oxygenase-1 (HO-1) is an AP-1-regulated gene that is induced by oxidative stress. The enzyme produces carbon monoxide (CO), which may attenuate the inflammatory response. The aim of this study was to investigate the regulation and interaction of iNOS and HO-1 in response to inflammation and oxidative stress. Male Wistar rats were treated with the thiol-modifying agent diethylmaleate (DEM) to induce oxidative stress and rendered endotoxaemic by LPS injection. Human colonic biopsies and the human colon carcinoma cell line DLD-1 were treated with DEM and the lipid peroxidation end-product 4-hydroxynonenal to induce oxidative stress and exposed to cytokine mix (CM) to mimic inflammation. In some experiments, cells were incubated with 250-400 ppm CO prior to and during stimulation with CM. HO-1 and iNOS expression was evaluated by RT-PCR, western blotting, and immunohistology. NF-kappaB activation was evaluated by EMSA. LPS induced iNOS but not HO-1 in epithelial cells of the ileum and colon. Oxidative stress strongly induced HO-1 in epithelial and inflammatory cells. Combined oxidative stress and endotoxaemia decreased iNOS expression but strongly induced HO-1 expression. Similarly, CM induced iNOS but not HO-1 in colonic biopsies and DLD-1 cells. Oxidative stress prevented iNOS induction in an NF-kappaB-dependent manner but increased HO-1 expression in CM-exposed DLD-1 cells. CO inhibited iNOS mRNA induction in CM-stimulated DLD-1 cells. These data demonstrate opposite regulation of iNOS and HO-1 in intestinal epithelial cells in response to cytokine exposure and oxidative stress. These findings suggest that iNOS (NF-kappaB driven) and HO-1 (AP-1 driven) represent mutually exclusive survival mechanisms in intestinal epithelial cells. PMID:15476266
Dijkstra, Gerard; Blokzijl, Hans; Bok, Lisette; Homan, Manon; van Goor, Harry; Faber, Klaas Nico; Jansen, Peter L M; Moshage, Han
Effect of the inhalation of nitric oxide on 5-hydroxytryptamine-induced pulmonary hypertension in calves.
In healthy anaesthetized Friesian-Holstein calves, pulmonary hypertension was induced by means of a continuous intravenous administration of serotonin (0.025 mg/kg body/weight/min). Afterwards, the anaesthetized calves inhaled 40 and 80 ppm of nitric oxide using an open system. The influences of the administration of serotonin and the inhalation of nitric oxide on the haemodynamic and blood gas parameters were investigated. The inhalation of 40 and 80 ppm of nitric oxide during serotonin-induced pulmonary hypertension in calves resulted in a significant fall of the mean pulmonary artery pressure. The inhalation of nitric oxide also induced an amelioration of intrapulmonary oxygen transport. The intravenous administration of serotonin in calves resulted in severe systemic hypotension. Hence, the influence of the inhalation of nitric oxide on the systemic arterial pressure could not be evaluated. PMID:8968160
Sustronck, B; Van Loon, G; Gasthuys, F; Foubert, L; Deprez, P; Muylle, E
Methylmalonate-induced seizures are attenuated in inducible nitric oxide synthase knockout mice
Methylmalonic acidemias consist of a group of inherited neurometabolic disorders caused by deficiency of methylmalonyl-CoA mutase activity clinically and biochemically characterized by neurological dysfunction, methylmalonic acid (MMA) accumulation, mitochondrial failure and increased reactive species production. Although previous studies have suggested that nitric oxide (NO) plays a role in the neurotoxicity of MMA, the involvement of NO-induced nitrosative damage from inducible
Leandro Rodrigo Ribeiro; Michele Rechia Fighera; Mauro Schneider Oliveira; Ana Fl�via Furian; Leonardo Magno Rambo; Ana Paula de Oliveira Ferreira; Andr� Luiz Lopes Saraiva; Mauren Assis Souza; Frederico Diniz Lima; Danieli Valnes Magni; Renata Dezengrini; Eduardo Furtado Flores; D. Allan Butterfield; Juliano Ferreira; Adair Roberto Soares dos Santos; Carlos Fernando Mello; Luiz Fernando Freire Royes
Role of inducible nitric oxide synthase in trinitrobenzene sulphonic acid induced colitis in mice
BACKGROUNDStudies using inhibitors of nitric oxide synthase (NOS) to date are inconclusive regarding the role of inducible NOS (iNOS) in intestinal inflammation.AIMS(1) To examine the role of iNOS in the development of chronic intestinal inflammation; (2) to identify the cellular source(s) of iNOS.METHODSColitis was induced by an intrarectal instillation of trinitrobenzene sulphonic acid (TNBS, 60 mg\\/ml, 30% ethanol), in wild
D-M McCafferty; M Miampamba; E Sihota; K A Sharkey; P Kubes
Inhibition of inducible nitric oxide synthase and cyclooxygenase-2 in lipopolysaccharide-stimulated RAW264.7 cells by carboxybutyrylated glucosamine takes place via down-regulation of mitogen-activated protein kinase-mediated nuclear factor-?B signaling
Glucosamine (GlcN) has been reported to possess several biomedical properties, and currently a great deal of attention has been focused on improving the functional properties of GlcN for different applications. Therefore, this study was conducted to introduce a carboxybutyryl functional group to GlcN and to find out the inhibitory mechanism of a novel GlcN derivative, carboxybutyrylated GlcN (CGlcN), on the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in bacterial lipopolysaccharide (LPS)-induced mouse macrophages (RAW264.7 cells). In the initial experiments, the production of NO and prostaglandin E2 (PGE2) was inhibited by CGlcN pretreatment and suggested the possibility of down-regulating their respective genes, iNOS and COX-2. Reverse transcription-polymerase chain reaction and Western blot analysis revealed that CGlcN can affect both transcriptional and translational levels of iNOS and COX-2 expression. The data from the nuclear factor-?B (NF-?B) promoter gene transfection experiment supported the idea that inhibition of iNOS and COX-2 is caused by the down-regulation of their transcription factor, NF-?B. Following stimulation with LPS, p38 mitogen-activated protein kinase (p38 MAPK) and c-Jun N-terminal kinase (JNK) present upstream of NF-?B signaling were also inhibited by CGlcN treatment. However, the protein level of another MAPK, extracellular signal-regulated kinase (ERK), remained unaffected. Moreover, following treatment with CGlcN, the protein expression of I-?B kinase (IKK) clearly confirmed that its down-regulation directly inhibited the degradation of I?B and release of NF-?B. Therefore, it can be concluded that CGlcN is capable of inhibiting iNOS and COX-2 expression in LPS-induced RAW264.7 cells via attenuation of NF-?B signaling by p38 MAPK and JNK, but not by ERK.
Rajapakse, Niranjan; Kim, Moon-Moo; Mendis, Eresha; Kim, Se-Kwon
Nitric oxide stress in sporadic inclusion body myositis muscle fibres: inhibition of inducible nitric oxide synthase prevents interleukin-1?-induced accumulation of ?-amyloid and cell death.
Sporadic inclusion body myositis is a severely disabling myopathy. The design of effective treatment strategies is hampered by insufficient understanding of the complex disease pathology. Particularly, the nature of interrelationships between inflammatory and degenerative pathomechanisms in sporadic inclusion body myositis has remained elusive. In Alzheimer's dementia, accumulation of ?-amyloid has been shown to be associated with upregulation of nitric oxide. Using quantitative polymerase chain reaction, an overexpression of inducible nitric oxide synthase was observed in five out of ten patients with sporadic inclusion body myositis, two of eleven with dermatomyositis, three of eight with polymyositis, two of nine with muscular dystrophy and two of ten non-myopathic controls. Immunohistochemistry confirmed protein expression of inducible nitric oxide synthase and demonstrated intracellular nitration of tyrosine, an indicator for intra-fibre production of nitric oxide, in sporadic inclusion body myositis muscle samples, but much less in dermatomyositis or polymyositis, hardly in dystrophic muscle and not in non-myopathic controls. Using fluorescent double-labelling immunohistochemistry, a significant co-localization was observed in sporadic inclusion body myositis muscle between ?-amyloid, thioflavine-S and nitrotyrosine. In primary cultures of human myotubes and in myoblasts, exposure to interleukin-1? in combination with interferon-? induced a robust upregulation of inducible nitric oxide synthase messenger RNA. Using fluorescent detectors of reactive oxygen species and nitric oxide, dichlorofluorescein and diaminofluorescein, respectively, flow cytometry revealed that interleukin-1? combined with interferon-? induced intracellular production of nitric oxide, which was associated with necrotic cell death in muscle cells. Intracellular nitration of tyrosine was noted, which partly co-localized with amyloid precursor protein, but not with desmin. Pharmacological inhibition of inducible nitric oxide synthase by 1400W reduced intracellular production of nitric oxide and prevented accumulation of ?-amyloid, nitration of tyrosine as well as cell death inflicted by interleukin-1? combined with interferon-?. Collectively, these data suggest that, in skeletal muscle, inducible nitric oxide synthase is a central component of interactions between interleukin-1? and ?-amyloid, two of the most relevant molecules in sporadic inclusion body myositis. The data further our understanding of the pathology of sporadic inclusion body myositis and may point to novel treatment strategies. PMID:22436237
Schmidt, Jens; Barthel, Konstanze; Zsch�ntzsch, Jana; Muth, Ingrid E; Swindle, Emily J; Hombach, Anja; Sehmisch, Stephan; Wrede, Arne; L�hder, Fred; Gold, Ralf; Dalakas, Marinos C
Inhibition of inducible nitric oxide synthase by Acanthopanax senticosus extract in RAW264.7 macrophages
Aim of the studyThe herb Acanthopanax senticosus (Siberian ginseng) has long been used as a traditional medicine. However, little is known about anti-inflammatory effects and its mechanisms of action. Excess production of nitric oxide (NO) is one of the characteristics of inflammation. In this study we examined the effects of A. senticosus extract (ASE) on NO production and inducible nitric
Qiu-Ye Lin; Li-Ji Jin; Zhen-Hui Cao; Yong-Ping Xu
Nitric oxide and its modulators in chronic constriction injury-induced neuropathic pain in rats
This study was conducted to examine the role of nitric oxide (NO) in peripheral neuropathy induced by chronic constriction injury of sciatic nerve of rats by using NO precursor, NO donors and nitric oxide synthase (NOS) inhibitors. Chronic constriction injury of sciatic nerve of rats resulted in peripheral neuropathy as confirmed by nociceptive behavioural tests using mechanical, thermal and cold
Ajit K. Naik; Surendra K. Tandan; Dinesh Kumar; Shailesh P. Dudhgaonkar
Thymoquinone suppresses expression of inducible nitric oxide synthase in rat macrophages
The objective of the present study was to determine the immunomodulatory role of thymoquinone (TQ) regarding its effect on the production of nitric oxide (NO) by rat peritoneal macrophages. Under certain conditions, macrophages and certain other cells can produce high concentrations of NO from its precursor l-arginine via inducible nitric oxide synthase (iNOS) pathway. TQ has been established as the
A El-Mahmoudy; H Matsuyama; M. A Borgan; Y Shimizu; M. G El-Sayed; N Minamoto; T Takewaki
Effect of Nitric Oxide on the Anticancer Activity of the Topoisomerase-Active Drugs Etoposide and Adriamycin in Human Melanoma Cells
Nitric oxide (?NO) was originally identified as an innate cytotoxin. However, in tumors it can enhance resistance to chemotherapy and exacerbate cancer progression. Our previous studies indicated that ?NO/?NO-derived species react with etoposide (VP-16) in vitro and form products that show significantly reduced activity toward HL60 cells and lipopolysaccharide (LPS)-induced macrophages. Here, we further confirm the hypothesis that �NO generation contributes to VP-16 resistance by examining interactions of ?NO with VP-16 in inducible nitric-oxide synthase (iNOS)�expressing human melanoma A375 cells. Inhibition of iNOS catalysis by N6-(1-iminoethyl)-l-lysine dihydrochloride (L-NIL) in human melanoma A375 cells reversed VP-16 resistance, leading to increased DNA damage and apoptosis. Furthermore, we found that coculturing A375 melanoma cells with LPS-induced macrophage RAW cells also significantly reduced VP-16 cytotoxicity and DNA damage in A375 cells. We also examined the interactions of ?NO with another topoisomerase active drug, Adriamycin, in A375 cells. In contrast, to VP-16, ?NO caused no significant modulation of cytotoxicity or Adriamycin-dependent apoptosis, suggesting that ?NO does not interact with Adriamycin. Our studies support the hypothesis that ?NO oxidative chemistry can detoxify VP-16 through direct nitrogen oxide radical attack. Our results provide insights into the pharmacology and anticancer mechanisms of VP-16 that may ultimately contribute to increased resistance, treatment failure, and induction of secondary leukemia in VP-16�treated patients.
Kumar, Ashutosh; Bhattacharjee, Suchandra; Espey, Michael G.; Mason, Ronald P.
Effect of nitric oxide on the anticancer activity of the topoisomerase-active drugs etoposide and adriamycin in human melanoma cells.
Nitric oxide (�NO) was originally identified as an innate cytotoxin. However, in tumors it can enhance resistance to chemotherapy and exacerbate cancer progression. Our previous studies indicated that (�NO/�NO-derived species react with etoposide (VP-16) in vitro and form products that show significantly reduced activity toward HL60 cells and lipopolysaccharide (LPS)-induced macrophages. Here, we further confirm the hypothesis that (�)NO generation contributes to VP-16 resistance by examining interactions of �NO with VP-16 in inducible nitric-oxide synthase (iNOS)-expressing human melanoma A375 cells. Inhibition of iNOS catalysis by N(6)-(1-iminoethyl)-L-lysine dihydrochloride (L-NIL) in human melanoma A375 cells reversed VP-16 resistance, leading to increased DNA damage and apoptosis. Furthermore, we found that coculturing A375 melanoma cells with LPS-induced macrophage RAW cells also significantly reduced VP-16 cytotoxicity and DNA damage in A375 cells. We also examined the interactions of (�)NO with another topoisomerase active drug, Adriamycin, in A375 cells. In contrast, to VP-16, (�)NO caused no significant modulation of cytotoxicity or Adriamycin-dependent apoptosis, suggesting that (?)NO does not interact with Adriamycin. Our studies support the hypothesis that (�)NO oxidative chemistry can detoxify VP-16 through direct nitrogen oxide radical attack. Our results provide insights into the pharmacology and anticancer mechanisms of VP-16 that may ultimately contribute to increased resistance, treatment failure, and induction of secondary leukemia in VP-16-treated patients. PMID:24049059
Sinha, Birandra K; Kumar, Ashutosh; Bhattacharjee, Suchandra; Espey, Michael G; Mason, Ronald P
Phlorofucofuroeckol A suppresses expression of inducible nitric oxide synthase, cyclooxygenase-2, and pro-inflammatory cytokines via inhibition of nuclear factor-?B, c-Jun NH2-terminal kinases, and Akt in microglial cells.
Microglial activation has been implicated in many neurological disorders for its inflammatory and neurotrophic effects. In this study, we investigated the effects of phlorofucofuroeckol A isolated from Ecklonia stolonifera Okamura on the production of inflammatory mediators in lipopolysaccharide (LPS)-stimulated microglia. Pre-treatment of phlorofucofuroeckol A attenuated the productions of nitric oxide, prostaglandin E2, and pro-inflammatory cytokines in LPS-stimulated microglia. Profoundly, phlorofucofuroeckol A treatment showed inactivation of nuclear factor-?B (NF-?B) by preventing the degradation of inhibitor ?B-? and the nuclear translocation of p65 NF-?B subunit. Moreover, phlorofucofuroeckol A inhibited the activation of c-Jun NH2-terminal kinases (JNKs), p38 mitogen-activated protein kinase (MAPK), and Akt, but not that of extracellular signal-regulated kinase. These results indicate that phlorofucofuroeckol A inhibits the LPS-induced expression of inflammatory mediators through inactivation of NF-?B, JNKs, p38 MAPK, and Akt pathways. These findings suggest that phlorofucofuroeckol A can be considered as a nutraceutical candidate for the treatment of neuroinflammation in neurodegenerative diseases. PMID:22993079
Kim, A-Reum; Lee, Min-Sup; Choi, Ji-Woong; Utsuki, Tadanobu; Kim, Jae-Il; Jang, Byeong-Churl; Kim, Hyeung-Rak
Suppression of lipopolysaccharide-induced of inducible nitric oxide synthase and cyclooxygenase-2 by Sanguis Draconis, a dragon's blood resin, in RAW 264.7 cells.
Sanguis Draconis (SD) is a kind of dragon's blood resin that is obtained from Daemomorops draco (Palmae). It is used in traditional medicine and has shown anti-inflammatory activity in some diseases. In this study, we examined the effects of Sanguis Dranonis ethanol extract (SDEE) on LPS-induced inflammation using RAW 264.7 cells. Our data indicated that SDEE inhibits LPS-stimulated NO, PGE2, IL-1 beta and TNF-alpha release, and iNOS and COX-2 expression. Furthermore, SDEE suppressed the LPS-induced p65 expression of NF-kappa B, which was associated with the inhibition of I kappa B-alpha degradation. We also found that the expression of HO-1 was significantly increased in RAW 264.7 cells by SDEE. These results suggest among possibilities of anti-inflammation that SDEE inhibits the production of NO and PGE2 by the down-regulation of iNOS and COX-2 gene expression via the suppression of NF-kappaB (p65) activation. SDEE can induce HO-1 over-expression in macrophage cells, which indicates that it may possess antioxidant properties. This result means that SEDD its anti-inflammatory effects in macrophages may be through a novel mechanism that involves the action of HO-1. Thus, SD could provide a potential therapeutic approach for inflammation-associated disorders. PMID:18060707
Choy, Cheuk-Sing; Hu, Chien-Ming; Chiu, Wen-Ta; Lam, Carlos-Shu Kei; Ting, Yih; Tsai, Shin-Han; Wang, Tzu-Chien
Modified apple polysaccharides suppress the migration and invasion of colorectal cancer cells induced by lipopolysaccharide.
Metastasis is the major cause of death in colorectal cancer (CRC). In colitis-associated carcinogenesis, the activation of nuclear factor-?B (NF-?B) occurs via lipopolysaccharide (LPS) binding to the toll-like receptor 4 (TLR4). The LPS/TLR4/NF-?B pathway contributes to the development and metastasis of colitis-associated colon cancer. In the present study, we hypothesized that an extracted modified Fuji apple polysaccharide (MAP) would alter the LPS/TLR4/NF-?B pathway. Thus, we evaluated the effect of MAP in vitro on the LPS/TLR4/NF-?B pathway in CRC cells (HT-29 and SW620 cells). The results suggest that (i) MAP competed with LPS for binding to TLR4 to reduce LPS-induced NF-?B expression and (ii) MAP suppressed the nuclear translocation of NF-?B p65. MAP significantly decreased LPS-induced expression of TLR4, cyclooxygenase-2, matrix metallopeptidase 9 (MMP9), matrix metallopeptidase 2, inducible nitric oxide synthase, and prostaglandin E2, and it increased the protein expression of the inhibitor of ?B? and NF-?B p65 in cytoplasm when it was given in combination with LPS. These results indicate that MAP suppressed LPS-induced migration and invasiveness of CRC cells by targeting the LPS/TLR4/NF-?B pathway. Therefore, we propose that MAP has potential for the clinical prevention of CRC cell metastasis. PMID:24074742
Zhang, Dian; Li, Yu-hua; Mi, Man; Jiang, Feng-Liang; Yue, Zheng-gang; Sun, Yang; Fan, Lei; Meng, Jin; Zhang, Xin; Liu, Li; Mei, Qi-Bing
Potential involvement of nitric oxide synthase but not inducible nitric oxide synthase in the development of experimental corneal neovascularization
AIM To investigate the effect of nitric oxide and its synthetase on experimental corneal neovascularization (CRNV). METHODS CRNV was induced by alkali injury in mice, nitric oxide synthetase (NOS) was inhibited by NG-nitro-L-arginine (L-NAME) and inducible nitric oxide synthetase (iNOS) was inhibited by aminoguanidine hemisulfate salt (AG). The inhibitory effect was detected at day 2 and 4 after corneal alkali injury by reverse transcription polymerase chain reaction (RT-PCR). CRNV was compared between the control and the treated mice by microscopic observation and corneal whole mount CD31 immunostaining. RESULTS The inhibition of L-NAME to NOS and AG to iNOS after corneal injury was confirmed by RT-PCR (P<0.05). Compared with control mice, L-NAME treated mice exhibited significantly decreased CRNV areas (P<0.05). In contrast, AG treatment failed to attenuate alkali induced CRNV (P>0.05). CONCLUSION Our findings suggest that NOS but not iNOS plays a critical role in alkali injury induced CRNV.
Chen, Yuan; Liu, Gao-Qin; Lu, Pei-Rong
Prostacyclin prevents nitric oxide-induced megakaryocyte apoptosis
We have previously demonstrated that nitric oxide (NO) triggers CD34+-derived megakaryocyte apoptosis. We here show that prostacyclin (PGI2) inhibits PAPA/NO-induced megakaryocyte death detected by fluorescent microscopy and flow cytometry. The cAMP-specific phosphodiesterase inhibitor, Ro 20-1724, and the permeable analog dibutyryl-cAMP also delayed apoptosis. PGI2 effect was fully prevented when adenylyl cyclase activity was suppressed by SQ 22536, and partially reversed by the permeable protein kinase A inhibitor PKI 14-22 amide. ELISA showed that while both PGI2 and NO alone or synergistically raised cAMP, only NO was able to increase intracellular cGMP levels. Treatment of megakaryocytes with PGI2 abolished both basal and NO-raised cGMP levels. Addition of 8-pCPT-cGMP or activation of soluble guanylyl cyclase by BAY 41-2272 induced cell death in a concentration-dependent manner, and ODQ, an inhibitor of guanylyl cyclase, prevented both PAPA/NO- or BAY 41-2272-induced apoptosis. Specific cGMP phosphodiesterase inhibition by Zaprinast or suppression of adenylyl cyclase by SQ 22536 enhanced the PAPA/NO proapoptotic effect. PGI2 completely inhibited NO-mediated generation and the increased activity of the cleaved form of caspase-3. In conclusion, our results demonstrate that contrary to their well-known direct and synergistic inhibitory effects on platelets, PGI2 and NO regulate opposite megakaryocyte survival responses through a delicate balance between intracellular cyclic nucleotide levels and caspase-3 activity control.
Gabriel Pozner, Roberto; Negrotto, Soledad; D'Atri, Lina Paola; Lidia Kotler, Monica; Angela Lazzari, Maria; Martin Gomez, Ricardo; Schattner, Mirta
Induction of inducible nitric oxide synthase gene expression by Pokeweed mitogen
The present study has characterized the expression of iNOS gene in Pokeweed mitogen (PWM)-driven murine macrophage RAW 264.7 cells. PWM significantly induced nitric oxide production in a dose-dependent manner. Quantitative reverse transcription-polymerase chain reaction analysis demonstrated that the inducible nitric oxide synthase gene expression is increased by PWM treatment. Since iNOS transcription has recently been shown to be under the
Young Jin Jeon; Jung Sup Lee; Hye Gwang Jeong
Therapeutic effect of C-phycocyanin extracted from blue green algae in a rat model of acute lung injury induced by lipopolysaccharide.
C-Phycocyanin (CPC), extracted from blue green algae, is a dietary nutritional supplement due to its several beneficial pharmacological effects. This study was conducted to evaluate whether CPC protects against lipopolysaccharide- (LPS-) induced acute lung injury (ALI) in rats. Rats were challenged with LPS (5?mg/kg body weight) intratracheally to induce ALI. After 3?h LPS instillation, rats were administrated with CPC (50?mg/kg body weight, i.p.) for another 3?h. Our results showed that posttreatment with CPC significantly inhibited LPS-induced elevation of protein concentration, nitrite/nitrate level, release of proinflammatory cytokines, the number of total polymorphonuclear cells in bronchoalveolar lavage fluid, and lung edema evidenced by decrease of lung wet/dry weight ratio accompanied by a remarkable improvement of lung histopathological alterations. Furthermore, CPC significantly attenuated LPS-induced myeloperoxidase activity, O2 (-) formation, expression of inducible nitric oxide synthase, and cyclooxygenase-2 as well as nuclear factor-kappa B (NF- ? B) activation in lungs. Additionally, CPC significantly downregulated proapoptotic proteins such as caspase-3 and Bax, but upregulated antiapoptotic proteins such as Bcl-2 and Bcl-XL in lungs exposed to LPS. These findings indicate that CPC could be potentially useful for treatment of LPS-related ALI by inhibiting inflammatory responses and apoptosis in lung tissues. PMID:23573157
Leung, Pak-On; Lee, Hao-Hsien; Kung, Yu-Chien; Tsai, Ming-Fan; Chou, Tz-Chong
Therapeutic Effect of C-Phycocyanin Extracted from Blue Green Algae in a Rat Model of Acute Lung Injury Induced by Lipopolysaccharide
C-Phycocyanin (CPC), extracted from blue green algae, is a dietary nutritional supplement due to its several beneficial pharmacological effects. This study was conducted to evaluate whether CPC protects against lipopolysaccharide- (LPS-) induced acute lung injury (ALI) in rats. Rats were challenged with LPS (5?mg/kg body weight) intratracheally to induce ALI. After 3?h LPS instillation, rats were administrated with CPC (50?mg/kg body weight, i.p.) for another 3?h. Our results showed that posttreatment with CPC significantly inhibited LPS-induced elevation of protein concentration, nitrite/nitrate level, release of proinflammatory cytokines, the number of total polymorphonuclear cells in bronchoalveolar lavage fluid, and lung edema evidenced by decrease of lung wet/dry weight ratio accompanied by a remarkable improvement of lung histopathological alterations. Furthermore, CPC significantly attenuated LPS-induced myeloperoxidase activity, O2? formation, expression of inducible nitric oxide synthase, and cyclooxygenase-2 as well as nuclear factor-kappa B (NF-?B) activation in lungs. Additionally, CPC significantly downregulated proapoptotic proteins such as caspase-3 and Bax, but upregulated antiapoptotic proteins such as Bcl-2 and Bcl-XL in lungs exposed to LPS. These findings indicate that CPC could be potentially useful for treatment of LPS-related ALI by inhibiting inflammatory responses and apoptosis in lung tissues.
Leung, Pak-on; Lee, Hao-Hsien; Kung, Yu-Chien; Tsai, Ming-Fan; Chou, Tz-Chong
GAPDH regulates cellular heme insertion into inducible nitric oxide synthase
Heme proteins play essential roles in biology, but little is known about heme transport inside mammalian cells or how heme is inserted into soluble proteins. We recently found that nitric oxide (NO) blocks cells from inserting heme into several proteins, including cytochrome P450s, hemoglobin, NO synthases, and catalase. This finding led us to explore the basis for NO inhibition and to identify cytosolic proteins that may be involved, using inducible NO synthase (iNOS) as a model target. Surprisingly, we found that GAPDH plays a key role. GAPDH was associated with iNOS in cells. Pure GAPDH bound tightly to heme or to iNOS in an NO-sensitive manner. GAPDH knockdown inhibited heme insertion into iNOS and a GAPDH mutant with defective heme binding acted as a dominant negative inhibitor of iNOS heme insertion. Exposing cells to NO either from a chemical donor or by iNOS induction caused GAPDH to become S-nitrosylated at Cys152. Expressing a GAPDH C152S mutant in cells or providing a drug to selectively block GAPDH S-nitrosylation both made heme insertion into iNOS resistant to the NO inhibition. We propose that GAPDH delivers heme to iNOS through a process that is regulated by its S-nitrosylation. Our findings may uncover a fundamental step in intracellular heme trafficking, and reveal a mechanism whereby NO can govern the process.
Chakravarti, Ritu; Aulak, Kulwant S.; Fox, Paul L.; Stuehr, Dennis J.
Inducible nitric oxide synthase as a possible target in hypertension.
Nitric oxide (NO) is an important vasodilator produced by vascular endothelium. Its enzymatic formation is derived from three different synthases: neuronal (nNOS), endothelial (eNOS) and inducible (iNOS) synthases. While relatively small amounts of NO produced by eNOS are important to cardiovascular homeostasis, high NO levels produced associated with iNOS activity may have detrimental consequences to the cardiovascular system and contribute to hypertension. In this article, we reviewed current literature and found mounting evidence indicating that increased iNOS expression and activity contribute to the pathogenesis of hypertension and its complications. Excessive amounts of NO produced by iNOS up-regulation can react with superoxide anions forming peroxynitrite, thereby promoting nitrosative stress and endothelial dysfunction. In addition, abnormal iNOS activity can up-regulate arginase activity, allowing it to compete with eNOS for L-arginine, thereby resulting in reduced NO bioavailability. This may also lead to eNOS uncoupling with enhanced production of superoxide anions instead of NO. All these alterations mediated by iNOS apparently contribute to hypertension and its complications. We also reviewed current evidence showing the effects of iNOS inhibitors on different animal models of hypertension. iNOS inhibition apparently exerts antihypertensive effects, decreases oxidative and nitrosative stress, and improves vascular function. Together, these studies highlight the possibility that iNOS is a potential pharmacological target in hypertension. PMID:24102471
Oliveira-Paula, Gustavo H; Lacchini, Riccardo; Tanus-Santos, Jose E
Inducible Nitric Oxide Synthase Mediates Hypoxia-Induced Hypoxia-Inducible Factor1? Activation and Vascular Endothelial Growth Factor Expression in Oxygen-Induced Retinopathy
Objective: Previous studies provided evidence that many factors contribute to retinal angiogenesis, including inducible nitric oxide synthase (iNOS), hypoxia-inducible factor-1? (HIF-1?) and vascular endothelial growth factor (VEGF). But the role of nitric oxide generated by iNOS in the regulation of expression of hypoxia-inducible genes in retinopathy of prematurity remains unclear. So we sought to better define the molecular basis of
Tao He; Ming Ai; Xiao-Hui Zhao; Yi-Qiao Xing
t10,c12 conjugated linoleic acid induces compensatory growth after immune challenge
Previous work demonstrated that feeding commercial preparations of conjugated linoleic acid (CLA) [a 50:50 mixture of c9,t11 and t10,c12 CLA (cCLA)] partially overcame lipopolysaccharide (LPS)-induced growth depression. The objective of this study was to determine which CLA isomer was responsible for the reduction of LPS-induced growth depression. Dietary cCLA supplementation for 3 weeks protected mice from LPS-induced weight loss 24
Daniel E. Butz; Guangming Li; Mark E. Cook
PKC412 (CGP41251) modulates the proliferation and lipopolysaccharide-induced inflammatory responses of RAW 264.7 macrophages
PKC412 (CGP41251) is a multitarget protein kinase inhibitor with anti-tumor activities. Here, we investigated the effects of PKC412 on macrophages. PKC412 inhibited the proliferation of murine RAW 264.7 macrophages through induction of G2/M cell cycle arrest and apoptosis. At non-toxic drug concentrations, PKC412 significantly suppressed the lipopolysaccharide (LPS)-induced release of TNF-{alpha} and nitric oxide, while instead enhancing IL-6 secretion. PKC412 attenuated LPS-induced phosphorylations of MKK4 and JNK, as well as AP-1 DNA binding activities. Furthermore, PKC412 suppressed LPS-induced Akt and GSK-3{beta} phosphorylations. These results suggest that the anti-proliferative and immunomodulatory effects of PKC412 are, at least in part, mediated through its interference with the MKK4/JNK/AP-1 and/or Akt/GSK-3{beta} pathways. Since macrophages contribute significantly to the development of both acute and chronic inflammation, PKC412 may have therapeutic potential and applications in treating inflammatory and/or autoimmune diseases.
Miyatake, Katsutoshi [Department of Orthopedics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima (Japan); Institute for Genome Research, The University of Tokushima, Tokushima (Japan); Inoue, Hiroshi [Institute for Genome Research, The University of Tokushima, Tokushima (Japan)]. E-mail: firstname.lastname@example.com; Hashimoto, Kahoko [Department of Life and Environmental Sciences, Faculty of Engineering, Chiba Institute of Technology, Chiba (Japan); Takaku, Hiroshi [Department of Life and Environmental Sciences, Faculty of Engineering, Chiba Institute of Technology, Chiba (Japan); Takata, Yoichiro [Department of Orthopedics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima (Japan); Nakano, Shunji [Department of Orthopedics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima (Japan); Yasui, Natsuo [Department of Orthopedics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima (Japan); Itakura, Mitsuo [Institute for Genome Research, The University of Tokushima, Tokushima (Japan)
Role of nitric oxide in hematosuppression and benzene-induced toxicity
It is becoming increasingly apparent that nitric oxide plays a multifunctional role in regulating inflammatory processes in the body. Although nitric oxide and its oxidation products are cytotoxic toward certain pathogens, they can also cause tissue injury and suppress proliferation. Cytokines and growth factors released at sites of inflammation or injury stimulate both immune and nonimmune cells to produce nitric oxide. Nowhere in the body is this more detrimental than in the bone marrow, for the continuous production of hematopoietic precursors is essential for normal blood cell maturation. Our laboratories have discovered that, in response to inflammatory mediators, bone marrow cells readily produce nitric oxide. Nitric oxide production is enhanced by hematopoietic growth factors including interleukin-3, macrophage colony stimulating factor, and granulocyte-macrophage colony-stimulating factor. When bone marrow cells produce nitric oxide, hematopoiesis is impaired, an effect that is potentiated by colony-stimulating factors. Treatment of mice with benzene, which suppresses bone marrow cell development, was found to markedly enhance the ability of bone marrow cells to produce nitric oxide in response to inflammatory mediators alone and in combination with hematopoietic growth factors. Taken together, these data suggest that nitric oxide may be an important mediator of benzene-induced bone marrow suppression. 38 refs., 3 figs.
Laskin, D.L.; Heck, D.E.; Punjabi, C.J.; Laskin J.D. [Rutgers Univ., Piscataway, NJ (United States)]|[UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ (United States)
Inhibition of Inducible Nitric Oxide Synthase in Murine Visceral Larva Migrans: Effects on Lung and Liver Damage
The roles of nitric oxide production and oxidative process were studied in mice infected with Toxocara canis and treated with aminoguanidine which is a specific inhibitor of inducible nitric oxide synthase (iNOS). Relations of nitric oxide synthase inhibition and tissue pathology were assessed by biochemical, histological and immunohistochemical methods. In experiments, Balb\\/c albino mice were inoculated with T. canis eggs
Cihan Demirci; Aysen Gargili; Handan Cetinkaya; Ilhan Uyaner; Basak Boynuegri; M. Koray Gumustas
Role of Nitric Oxide and Reactive Oxygen Species in Platelet-Activating Factor-Induced Microvascular Leakage
Platelet-activating factor (PAF), released during inflammatory responses, increases microvascular permeability to fluid and macromolecules. Previous studies in the hamster cheek pouch microcirculation have shown that PAF-induced increases in permeability can be diminished by pretreatment with a nitric oxide synthase inhibitor indicating that nitric oxide is required for PAF to cause leakage, although nitric oxide itself does not cause leakage. We
Richard E. Klabunde; Denise E. Anderson
The costimulatory immunogen LPS induces the B-Cell clones that infiltrate transplanted human kidneys
The mechanism of chronic rejection of transplanted human kidneys is unknown. An understanding of this process is important because, chronic rejection ultimately leads to loss of the kidney allograft in most transplants. One feature of chronic rejection is the infiltration of ectopic B-cell clusters that are clonal into the transplanted kidney. We now show that the antibodies produced by these B-cells react strongly with the core carbohydrate region of LPS. Since LPS is a costimulatory immunogen that can react with both the B-cell receptor (BCR) and the Toll-like receptor 4 (TLR4), these results suggest a mechanism for the selective pressure that leads to clonality of these B-cell clusters and opens the possibility that infection and the attendant exposure to LPS plays a role in the chronic rejection of human kidney transplants. If confirmed by clinical studies, these results suggest that treating patients with signs of chronic rejection with antibiotics may improve kidney allograft survival.
Grover, Rajesh K.; Cheng, Julong; Peng, Yingjie; Jones, Teresa M.; Ruiz, Diana I.; Ulevitch, Richard J.; Glass, John I.; Dennis, Edward A.; Salomon, Daniel R.; Lerner, Richard A.
Roquefort cheese proteins inhibit Chlamydia pneumoniae propagation and LPS-induced leukocyte migration.
Inflammation in atherosclerosis, which could be associated with some subclinical infections such as C. pneumoniae, is one of the key factors responsible for the development of clinical complications of this disease. We report that a proprietary protein extract isolated from Roquefort cheese inhibits the propagation of C. pneumoniae in a human HL cell line in a dose-dependent manner, as revealed by the immunofluorescence analysis. These changes were accompanied by a significant reduction in the infective progeny formation over the protein extract range of 0.12-0.5 ?g/mL. Moreover, short term feeding of mice with Roquefort cheese (twice, 10 mg per mouse with an interval of 24 hours) led to the inhibition of the migration of peritoneal leukocytes caused by intraperitoneal injection of E. coli lipopolysaccharide. These changes were complemented by a reduction in neutrophil count and a relative increase in peritoneal macrophages, suggesting that ingestion of Roquefort could promote regenerative processes at the site of inflammation. The ability of this protein to inhibit propagation of Chlamydia infection, as well as the anti-inflammatory and proregenerative effects of Roquefort itself, may contribute to the low prevalence of cardiovascular mortality in France where consumption of fungal fermented cheeses is the highest in the world. PMID:23737705
Petyaev, Ivan M; Zigangirova, Naylia A; Kobets, Natalie V; Tsibezov, Valery; Kapotina, Lydia N; Fedina, Elena D; Bashmakov, Yuriy K
LPS-Induced Genes in Intestinal Tissue of the Sea Cucumber Holothuria glaberrima
Metazoan immunity is mainly associated with specialized cells that are directly involved with the immune response. Nevertheless, both in vertebrates and invertebrates other organs might respond to immune activation and participate either directly or indirectly in the ongoing immune process. However, most of what is known about invertebrate immunity has been restricted to immune effector cells and little information is available on the immune responses of other tissues or organs. We now focus on the immune reactions of the intestinal tissue of an echinoderm. Our study employs a non-conventional model, the echinoderm Holothuria glaberrima, to identify intestinal molecules expressed after an immune challenge presented by an intra-coelomic injection of lipopolysaccharides (LPS). The expression profiles of intestinal genes expressed differentially between LPS-injected animals and control sea water-injected animals were determined using a custom-made Agilent microarray with 7209 sea cucumber intestinal ESTs. Fifty (50) unique sequences were found to be differentially expressed in the intestine of LPS-treated sea cucumbers. Seven (7) of these sequences represented homologues of known proteins, while the remaining (43) had no significant similarity with any protein, EST or RNA database. The known sequences corresponded to cytoskeletal proteins (Actin and alpha-actinin), metabolic enzymes (GAPDH, Ahcy and Gnmt), metal ion transport/metabolism (major yolk protein) and defense/recognition (fibrinogen-like protein). The expression pattern of 11 genes was validated using semi-quantitative RT-PCR. Nine of these corroborated the microarray results and the remaining two showed a similar trend but without statistical significance. Our results show some of the molecular events by which the holothurian intestine responds to an immune challenge and provide important information to the study of the evolution of the immune response.
Ramirez-Gomez, Francisco; Ortiz-Pineda, Pablo A.; Rivera-Cardona, Gabriela; Garcia-Arraras, Jose E.
Prophylactic effect of liposomal N-acetylcysteine against LPS-induced liver injuries
The aim of this study was to evaluate and compare the effectiveness of N-acetylcysteine (NAC) and liposomally-encapsulated NAC (L-NAC) in ameliorating the hepatotoxic effects of lipopolysaccharide (LPS). LPS, a major cell wall molecule of Gram-negative bacteria and the principal initiator of septic shock, causes liver injury in vivo that is dependent on neutrophils, platelets, and several inflammatory mediators, including tumour
Misagh Alipour; Abdelwahab Omri; Milton G. Smith; Zacharias E. Suntres
Lack of effect of oral administration of resveratrol in LPS-induced systemic inflammation
Purpose��The high mortality index due to sepsis and the lack of an effective treatment requires the search for new compounds that can\\u000a serve as therapy for this disease. Resveratrol, a well-known anti-inflammatory natural compound, might be a good candidate\\u000a for the treatment of sepsis. The aim of this work was to study the effects of oral administration of resveratrol, before
M. Larrosa; M. Azor�n-Ortu�o; M. J. Ya�ez-Gasc�n; M. T. Garc�a-Conesa; F. Tom�s-Barber�n; J. C. Esp�n
Low-level diode laser therapy reduces lipopolysaccharide (LPS)-induced bone cell inflammation.
In this study, the aim is to investigate the cytologic effects of inflammatory bone cells after in vitro low-level laser therapy (LLLT). A human osteosarcoma cell line (MG63) was cultured, infected with lipopolysaccharide (LPS) and exposed to low-level laser treatment at 5 or 10�J/cm(2) using a 920�nm diode laser. MG63 cell attachment was observed under a microscope, and cell viability was quantified by mitochondrial colorimetric assay (MTT). LPS-treated MG63 cells were irradiated with LLLT, and the inflammatory markers iNOS, TNF-? and IL-1, were analyzed by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. The data were collected and analyzed by one-way analysis of variance (ANOVA); p?0.05 indicated a statistically significant difference. Low-level laser treatment on MG63 cells increased their ability to attach and survive. After irradiation, the expression levels of iNOS, TNF-? and IL-1 in LPS-infected MG63 cells decreased over time (p?0.05). Conclusions: low-level diode laser treatment increased the MG63 cell proliferative ability and decreased the expression of inflammatory mediators in MG63 cells. PMID:22002329
Huang, Tsui Hsien; Lu, Yu Chuan; Kao, Chia Tze
The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...
Fresh organically grown ginger ( Zingiber officinale): composition and effects on LPS-induced PGE 2 production
Gas chromatography in conjunction with mass spectrometry, a technique previously employed to analyze non-volatile pungent components of ginger extracts modified to trimethylsilyl derivatives, was applied successfully for the first time to analyze unmodified partially purified fractions from the dichloromethane extracts of organically grown samples of fresh Chinese white and Japanese yellow varieties of ginger, Zingiber officinale Roscoe (Zingiberaceae). This analysis
Shivanand D. Jolad; R. Clark Lantz; Aniko M. Solyom; Guan Jie Chen; Robert B. Bates; Barbara N. Timmermann
Treatment of LPS-Induced Tissue Injury: Role of Liposomal Antioxidants.
National Technical Information Service (NTIS)
Tissue injury is a common occurrence in multiple organ failure, a possible clinical complication of Gram-negative bacterial sepsis. Gram-negative bacteria, in part through lipopolysaccharide (LPS), tumor necrosis factor, and other cytokines, activate neut...
Z. E. Suntres P. N. Shek
Up-regulation of podoplanin involves in neuronal apoptosis in LPS-induced neuroinflammation.
Podoplanin (PDPN) is a mucin-type transmembrane sialoglycoprotein expressed in multiple tissues in adult animals, including the brain, lungs, kidney, and lymphoid organs. Studies of this molecule have demonstrated its great importance in tumor metastasis, platelet aggregation, and lymphatic vessel formation. However, information regarding its regulation and possible function in the central nervous system is still limited. In this study, we performed a neuroinflammatory model by lipopolysaccharide (LPS) lateral ventral injection in adult rats and detected increased expression of PDPN in the brain cortex. Immunofluorescence indicated that PDPN was located in the neurons, but not astrocytes. Moreover, there was a concomitant up-regulation of active caspase-3, cyclin D1, and CDK4 in vivo and vitro studies. In addition, the expression of these three proteins in cortical primary neurons was decreased after knocking down PDPN by siRNA. Collectively, all these results suggested that the up-regulation of PDPN might be involved in neuronal apoptosis in neuroinflammation after LPS injection. PMID:24821010
Song, Yan; Shen, Jianhong; Lin, Yuchang; Shen, Jiabing; Wu, Xinming; Yan, Yaohua; Zhou, Li; Zhang, Haiyan; Zhou, Ying; Cao, Maohong; Liu, Yonghua
We have previously demonstrated in CD-1 mice that pre-administration of N-acetyl cysteine (NAC) or the p38 MAP kinase inhibitor (SB203580) reduces acute lung injury and inflammation following pulmonary exposures to diesel exhaust particles (DEP) or lipopolysaccharide (LPS). Here ...
Soluble and immobilized anti-Ig antibodies in the regulation of LPS-induced lymphoblasts.
Anti-immunoglobulin (anti-Ig) causes suppression of secretion of immunoglobulin by LPS-activated pig B lymphoblasts. The cellular level at which anti-Ig influences immunoglobulin secretion has been investigated, using soluble anti-Ig which enters B cells, and anti-Ig immobilized onto acrylamide bead or plastic surfaces which can act only at the B-cell surface. Suppression of secretion only occurred with soluble anti-Ig, indicating that the intracellular processing of antibody after its complexing on the cell surface was necessary for suppression to occur. Immobilized anti-Ig acted as effectively as soluble antibody in activation of resting B cells into mitosis, showing that the activating signal of anti-Ig is received at the cell surface. Electron microscopy has shown that the block to secretion after soluble anti-Ig resulted from the accumulation of smooth membrane-bounded intracellular vesicles which, by immunofluorescence, contained immunoglobulin. The formation of vesicles was intimately associated with the intracellular localization of 125I-labelled anti-Ig which was used to follow cellular processing of anti-Ig. Images Figure 2 Figure 3 Figure 4
Symons, D B; Clarkson, C A; Hall, F J
Mast cells are necessary for the hypothermic response to LPS-induced sepsis
As central nervous system residents, mast cells contain many cytokines and are localized primarily near large blood vessels in the diencephalon and within the leptomeninges, making them candidates for immune to neural �cross talk.� Using mast cell-deficient KitW-sh/W-sh mice, we assessed the role of these cells in the thermoregulatory component of the immune response to lipopolysaccharide (LPS). KitW-sh/W-sh and wild-type (WT) mice differed in several respects in response to injection of a high dose of LPS (1 mg/kg ip). Core temperature (Tc) of WT mice decreased by ?3�C, whereas KitW-sh/W-sh mice did not become hypothermic but instead exhibited pronounced low-frequency Tc oscillations around their baseline temperature. In addition, KitW-sh/W-sh mice had lower levels of whole brain TNF-? but no differences in IL-1?, IL-6, IFN-?, or histamine compared with WT mice following injection of the high dose of LPS, consistent with the role of TNF-? in sepsis. KitW-sh/W-sh mice had increased resistance to LPS, and some survived a dose of LPS that was lethal in littermate controls. In contrast, KitW-sh/W-sh and WT mice were similar in other aspects, namely, in the hyperthermia following injection of TNF-? (1.5 ?g icv), reduced nighttime Tc and locomotor activity (to 1 mg/kg LPS), response to a low dose of LPS (10 ?g/kg ip), and response to subcutaneous turpentine injection. These results indicate that mast cells play a role in the regulation of thermoregulatory responses and survival following sepsis induction and suggest a brain site of action.
Nautiyal, Katherine M.; McKellar, Heather; Silverman, Ann-Judith; Silver, Rae
VIP Inhibits P. gingivalis LPS-induced IL18 and IL18BPa in Monocytes
IL-18 is a pro-inflammatory cytokine that is important in the regulation of T-cells and is elevated in inflammatory disorders such as periodontal disease. Vasoactive intestinal peptide (VIP) modulates immune responses to the periodontal pathogen Porphyromonas gingivalis (Pg). Our objective was to investigate the effect of Pg LPS on IL-18 and its natural inhibitor, IL-18 binding protein (IL-18BPa), in human monocytes,
N. Foster; K. Andreadou; L. Jamieson; P. M. Preshaw; J. J. Taylor
CpG DNA and LPS induce distinct patterns of activation in human monocytes
A specific set of immune functions is switched on in response to DNA containing unmethylated CpG dinucleotides in particular base contexts (�CpG motifs�). Plasmids, viral vectors and antisense oligodeoxynucleotides used for DNA vaccination, gene replacement or gene blockade contain immunostimulatory CpG motifs which may have independent biological activity. Although the immune stimulatory effects of CpG motifs on murine cells are
G Hartmann; A M Krieg
Molecular determinants of LPS-induced acute renal inflammation: Implication of the kinin B1 receptor.
Acute renal inflammation represents a complex disease and its molecular basis remains incompletely defined. We examined changes of global renal gene expression in lipopolysacharide-treated wild-type and kinin B(1) receptor-knockout mice to better comprehend molecular mechanisms of acute renal inflammation and possible implications of the kinin B(1) receptor in early (inflammatory) stages of renal disease. Microarray data revealed that LPS-mediated renal inflammation is associated with strong induction of gene families that are mostly involved in inflammatory and immune response and cell adhesion, as well as genes associated with metabolism, signal transduction and transport. Downregulated by the LPS challenge were genes and pathways that are necessary for normal renal function, including those implicated in metabolism, transport, protein biosynthesis and, cytoskeleton organization, regulation of transcription and signal transduction. Moreover, we show that B(1) receptor ablation could be protective against inflammation-related kidney injuries. PMID:19538936
Bascands, Jean-Loup; Bachvarova, Magdalena; Neau, Eric; Schanstra, Joost P; Bachvarov, Dimcho
Transcriptional profiling of the LPS induced NF-?B response in macrophages
BACKGROUND: Exposure of macrophages to bacterial products such as lipopolysaccharide (LPS) results in activation of the NF-?B transcription factor, which orchestrates a gene expression programme that underpins the macrophage-dependent immune response. These changes include the induction or repression of a wide range of genes that regulate inflammation, cell proliferation, migration and cell survival. This process is tightly regulated and loss
Omar Sharif; Viacheslav N Bolshakov; Stephanie Raines; Peter Newham; Neil D Perkins
Plexin-A1 is required for Toll-like receptor-mediated microglial activation in the development of lipopolysaccharide-induced encephalopathy
Recent investigations have suggested that semaphorins, which are known repulsive axon guidance molecules, may play a crucial role in maintaining brain homeostasis by regulating microglial activity. Sema3A, secreted in higher amounts from injured neurons, is considered to suppress excessive inflammatory responses by inducing microglial apoptosis through its binding to Plexin-A1 receptors on activated microglia. To clarify the in vivo role of Plexin-A1-mediated signaling in lipopolysaccharide (LPS)-induced injury in mouse brain, we examined the neuroinflammatory changes initiated by LPS administration to the cerebral ventricles of wild-type (WT) and Plexin-A1-deficient (?/?) mice. WT mice administered LPS exhibited a significantly higher expression of COX-2, iNOS, IL-1? and TNF-? in the hippocampus, and a significantly greater ventricular enlargement and intracerebral infiltration of leukocytes, as compared with the saline-treated group. By contrast, Plexin-A1?/? mice administered LPS did not exhibit a significantly increased expression of COX-2, iNOS, IL-1? or TNF-? in the hippocampus as compared with the saline-treated group. Plexin-A1?/? mice administered LPS did not show significant increases in ventricle size or infiltration of leukocytes into the brain, as compared with the saline-treated group. In WT, but not in the Plexin-A1?/? primary microglia treated with LPS, Sema3A induced significantly more nitric oxide production than in the immunoglobulin G control. These results revealed the crucial role of the Sema3A-Plexin-A1 interaction in the Toll-like receptor 4-mediated signaling of the LPS-induced activation of microglia. Thus, results of the present study revealed the essential role of Plexin-A1 in the development of LPS-induced neuroinflammation in mice, suggesting the possible application of microglial control of the semaphorin-plexin signaling system to the treatment of LPS-induced encephalopathy and other psychiatric diseases associated with neuroinflammation.
The immunomodulation of inducible nitric oxide in scallop Chlamys farreri.
Nitric oxide (NO) is an important signalling molecule which plays an indispensable role in immunity of all vertebrates and invertebrates. In the present study, the immunomodulation of inducible NO in scallop Chlamys farreri was examined by monitoring the alterations of haemocyte behaviours and related immune molecules in response to the stimulations of LPS and/or with S-Methylisothiourea Sulphate (SMT), an inhibitor of inducible NO synthase (NOS). The total activity of NOS and NO concentration in the haemolymph of scallop C. farreri increased significantly at 3, 6 and 12 h after LPS stimulation respectively, whereas their increases were fully repressed when scallops were treated in the collaborating of LPS and SMT. Meanwhile, some cellular and humoral immune parameters were determined after the stimulation of LPS and SMT to investigate the role of inducible NO in innate immunity of scallop. After LPS stimulation, the highest levels of haemocytes apoptosis and phagocytosis were observed at 24 h (38.5 � 2.5%, P < 0.01) and 12 h (38.6 � 0.2%, P < 0.01), respectively, and the reactive oxygen species (ROS) level (5.88 � 0.90%, P < 0.01) of haemocytes and anti-bacterial activity of haemolymph (10.0 � 2.2%, P < 0.01) all elevated dramatically at 12 h. Although the activity of lysozyme and phenoloxidase (PO) in haemolymph both declined at 48 h (93.0 � 6.3 U mgprot(-1), 0.40 � 0.06 U mgprot(-1), P < 0.01), superoxide dismutase (SOD) activity and GSH concentration both increased to the highest level at 24 h post treatment (99.2 � 8.1 U mgprot(-1), 93.0 � 6.3 nmol mgprot(-1), P < 0.01). After the collaborating treatment of LPS and SMT, the apoptosis index increased much higher from 48 h, while the increase of haemocytes phagocytosis, ROS level and haemolymph anti-bacteria activities were suppressed completely at 12 h. The declines of lysozyme and PO activity in haemolymph were reversed at 48 h, and the rise of SOD activity and GSH concentration started earlier from 3 h. These results indicated clearly that NO could participate in the scallop immunity and play a crucial role in the modulation of immune response including haemocytes apoptosis and phagocytosis, anti-bacterial activity and redox homeostasis in the haemolymph of scallop. PMID:23073205
Jiang, Qiufen; Zhou, Zhi; Wang, Lingling; Shi, Xiaowei; Wang, Jingjing; Yue, Feng; Yi, Qilin; Yang, Chuanyan; Song, Linsheng
Inducible nitric oxide synthase haplotype associated with migraine and aura.
Migraine is a complex neurological disorder with a clear neurogenic inflammatory component apparently including enhanced nitric oxide (NO) formation. Excessive NO amounts possibly contributing to migraine are derived from increased expression and activity of inducible NO synthase (iNOS). We tested the hypothesis that two functional, clinically relevant iNOS genetic polymorphisms (C(-1026)A-rs2779249 and G2087A-rs2297518) are associated with migraine with or without aura. We studied 142 healthy women without migraine (control group) and 200 women with migraine divided into two groups: 148 with migraine without aura (MWA) and 52 with aura (MA). Genotypes were determined by real-time polymerase chain reaction using the Taqman(�) allele discrimination assays. The PHASE 2.1 software was used to estimate the haplotypes. The A allele for the G2087A polymorphism was more commonly found in the MA group than in the MWA group (28 vs. 18%; P < 0.05). No other significant differences in the alleles or genotypes distributions were found (P > 0.05). The haplotype combining both A alleles for the two polymorphisms was more commonly found in the MA group than in the control group or in the MWA group (19 vs. 10 or 8%; P = 0.0245 or 0.0027, respectively). Our findings indicate that the G2087A and the C(-