In Vitro Ubiquitination: Self-Ubiquitination, Chain Formation, and Substrate Ubiquitination Assays.
Ubiquitination of proteins in vitro has evolved as an indispensable tool for the functional analysis of this posttranslational modification. In vitro ubiquitination is particularly helpful to study conjugation mechanisms. The efficiency of the ubiquitination reaction depends in part on the quality of the enzymes utilized. Here we introduce the assay developed in our lab to study HECT E3 ligases. It involves bacterially expressed E1, His-tagged Ube2D3 (also called UbcH5c, the best E2 for Nedd4), untagged Nedd4, and untagged ubiquitin (Ub). As tags may impair specific activity of the enzymes or even interfere with the enzymatic reaction, they should be avoided, removed, or kept to a minimal size whenever possible, unless proven to be without consequence. The protocol described here is suitable for other E3 ligases capable of forming Ub chains as pseudo-product of the enzyme reaction. It is also adapted to include substrates. In this case, substrates should be tagged and purified after the reaction is completed to allow the detection of the ubiquitinated products.


Beclin1 overexpression inhibitis proliferation, invasion and migration of CaSki cervical cancer cells.
The influence of the autophagy-related gene Beclin1 on proliferation, invasion and metastasis of the cervical cancer CaSki cells and its possible mechanism in vitro were here targeted. After the overexpression vector pcDNA3.1-Beclin1 and RNA interference vector pSUPER-Beclin1 were transfected into CaSki cells in vitro, stable expression cell lines demonstration Beclin1 expression was upregulated, and VEGF and MMP-9 expression were decreased, leading to cell arrest in the G0/G1 phase of the cell cycle. MTT assays further revealed proliferation of cells was significantly inhibited in Beclin1-overexpressing transfectant cells, with invasion and metastasis also being inhibited in Transwell chamber assays. The present results suggest that Beclin1 inhibits invasion and metastasis of cervical cancer CaSki cells in vitro. Mechanisms probably involve Beclin1 inhibition of cell proliferation, and decreased expression of VEGF and MMP-9 proteins.


Retroperitoneoscopic distal pancreatectomy: a new surgical approach.
The traditional laparoscopic surgery is difficult to deal with the deep lesions of the body and tail of the pancreas, which may damage the visceral organs of the abdominal cavity and cause abdominal adhesion and other related complications. This paper introduces the operation procedure of retroperitoneoscopy in pancreatic surgery, and evaluates its feasibility in clinical application. Retrospective analysis was performed on patients with retroperitoneal pancreatectomy in our hospital. The anatomical features of the fascia, surgical plane composition and surgical pathway of the fascia of the retroperitoneoscopic pancreatectomy were observed during the operation, and the surgical safety and feasibility were analyzed. The following parameters were evaluated: operation time, blood loss, pancreatic fistula, postoperative gastro-intestinal recovery, hospital stay. All 3 patients had a smooth operation and no serious complications occurred. During retroperitoneal laparoscopic pancreatectomy, there is a vascularized plane between the posterior fascia of the pancreas and the prerenal fascia, which can avoid injury of the visceral organs and retroperitoneal vessels. The anterior renal fascia should be used as the posterior boundary of the safe separation plane. The surgical plane based on the anatomy of the fascia and interstitial dissection is the theoretical basis of modern surgery, which is safe, fast and effective. The inter-prerenal fascia plane is the correct and safe anatomical plane of posterior laparoscopic surgery.


[Halitosis in 1999].
Bad breath is a major concern to the general population and a source of an important profit industry world wide. Between 50 and 60 per cent of the population suffer from chronic halitosis. This can have consequences for private or professional life. Reasons can imply many specialities: ENT, gastro-enterology, pneumology, hepatology, genetics and psychiatry (a high percentage of the patients who come to the clinic with a primary complaint of halitosis do not have a detectable problem). Nevertheless, 85% are stomatological and are explained by the release of volatile sulfur compounds. These substances have a very offensive odor in very low concentrations. A specialized device called a halimeter is available to measure the volatile sulphur compounds in mouth air but in practice the objective assessment of malodor is still best performed by the organoleptic method. A careful examination can determine the patient's problem in most cases. Initial treatment strategy should focus on the exact cause and on oral hygiene. In addition to the usual recommendations, the daily use of tongue cleaners is very beneficial. Other than etiologic therapy, masking can be achieved by oral care products (mouth rinses, toothpastes, chewing gums) which contain metal ions, especially zinc, that inhibit odor formation because of its affinity to sulphur compounds.


Targeting bone metastases starting from the preneoplastic niche: home sweet home.
The metastatic process is a multistep coordinated event with a high degree of efficiency. Specific subpopulations of cancer stem cells, with tumor-initiating and migratory capacity, can selectively migrate towards sites that are able to promote survival and/or proliferation of metastatic tumor cells through a microenvironment modification. Cross-talk between the bone microenvironment and cancer cells can facilitate bone tropism of cancer cells. Fully understanding this complexity represents a major challenge in anti-cancer research and a mandatory step towards the development of new drugs potentially able not only to reduce the consequences of bone lesions but also to target the metastatic process in visceral sites.


A commercial ELISA for detection of interferon gamma in white rhinoceros.
Bovine tuberculosis (bTB), caused by Mycobacterium bovis, is endemic in Kruger National Park, South Africa, home to the largest population of white rhinoceros (Ceratotherium simum) in the world. In 2016, the first cases of naturally occurring bTB were reported in white rhinoceros; however, there is a lack of understanding of infection and disease process in this species. Prevention and control of transmission depends on the availability of accurate tools to detect M. bovis infection. Interferon gamma (IFN-γ) assays are a reliable detection method for TB in other animal species, and studies have indicated that these tests can be used in white rhinoceros. We sought to screen and optimize a commercial IFN-γ enzyme-linked immunosorbent assay (ELISA) to detect endogenous white rhinoceros IFN-γ in mitogen-stimulated whole blood as a basis for developing a test for M. bovis infection. Optimizations included identifying ELISA antibodies and determining the effect of sample matrix, ELISA plate incubation temperature, ELISA linearity, assay reproducibility, and the assay's limit of quantification. The optimized assay employed an equine IFN-γ antibody pair that was used to create a commercial ELISA kit. This ELISA had a linear response to recombinant equine and endogenous rhinoceros IFN-γ (range: 7.8-125 pg/mL). When incubated at 37°C, the ELISA was highly reproducible, with an optimal recovery and a low limit of quantification, indicating that the Mabtech equine IFN-γ ELISAPRO kit is a robust assay for measuring white rhinoceros IFN-γ.


Theoretical study on rotation of pyrrole rings in porphyrin and N-confused porphyrin.
Rotation of pyrrole rings in regular porphyrins and N-confused porphyrins is theoretically investigated by DFT calculations. While the inversion of the pyrrole rings in the regular porphyrins requires high activation energies (36.5-49.1 kcal/mol), the inversion of the confused pyrrole rings in the N-confused porphyrins requires much lower activation energies (18.1-24.5 kcal/mol). This marked difference can be explained by the intramolecular hydrogen bondings and aromatic stabilization due to the [18]annulenic substructures, where confusion and NH tautomerism play an important role. In both of the macrocycles, 360 degrees rotation of their pyrrole rings would be difficult possibly due to the small cavity. Alternatively, a reaction pathway for the production of N-fused porphyrin from N-confused porphyrin is obtained, which is consistent with the experimental observation.


Pigmentation and sporulation are alternative cell fates in Bacillus pumilus SF214.
Bacillus pumilus SF214 is a spore forming bacterium, isolated from a marine sample, able to produce a matrix and a orange-red, water soluble pigment. Pigmentation is strictly regulated and high pigment production was observed during the late stationary growth phase in a minimal medium and at growth temperatures lower than the optimum. Only a subpopulation of stationary phase cells produced the pigment, indicating that the stationary culture contains a heterogeneous cell population and that pigment synthesis is a bimodal phenomenon. The fraction of cells producing the pigment varied in the different growth conditions and occurred only in cells not devoted to sporulation. Only some of the pigmented cells were also able to produce a matrix. Pigment and matrix production in SF214 appear then as two developmental fates both alternative to sporulation. Since the pigment had an essential role in the cell resistance to oxidative stress conditions, we propose that within the heterogeneous population different survival strategies can be followed by the different cells.


χ2-BidLSTM: A Feature Driven Intrusion Detection System Based on χ2 Statistical Model and Bidirectional LSTM.
In a network architecture, an intrusion detection system (IDS) is one of the most commonly used approaches to secure the integrity and availability of critical assets in protected systems. Many existing network intrusion detection systems (NIDS) utilize stand-alone classifier models to classify network traffic as an attack or as normal. Due to the vast data volume, these stand-alone models struggle to reach higher intrusion detection rates with low false alarm rates( FAR). Additionally, irrelevant features in datasets can also increase the running time required to develop a model. However, data can be reduced effectively to an optimal feature set without information loss by employing a dimensionality reduction method, which a classification model then uses for accurate predictions of the various network intrusions. In this study, we propose a novel feature-driven intrusion detection system, namely χ2-BidLSTM, that integrates a χ2 statistical model and bidirectional long short-term memory (BidLSTM). The NSL-KDD dataset is used to train and evaluate the proposed approach. In the first phase, the χ2-BidLSTM system uses a χ2 model to rank all the features, then searches an optimal subset using a forward best search algorithm. In next phase, the optimal set is fed to the BidLSTM model for classification purposes. The experimental results indicate that our proposed χ2-BidLSTM approach achieves a detection accuracy of 95.62% and an F-score of 95.65%, with a low FAR of 2.11% on NSL-KDDTest+. Furthermore, our model obtains an accuracy of 89.55%, an F-score of 89.77%, and an FAR of 2.71% on NSL-KDDTest-21, indicating the superiority of the proposed approach over the standard LSTM method and other existing feature-selection-based NIDS methods.


A descriptive study on wrist and hand sensori-motor impairment and function following distal radius fracture intervention.
Descriptive cross-sectional design. Wrist and hand sensori-motor impairment have been observed after distal radius fracture (DRF) treatment. This impairment and its relationship to function lack research. The primary aim of this exploratory study was to determine the magnitude of wrist and hand sensori-motor impairment following surgical and non-surgical treatment among older patients following DRF. Secondary aims were to determine the relationship between wrist and hand sensori-motor impairment with function and pain as well as the relationships among wrist and hand sensori-motor impairment and function and age following DRF. Ten Test (TT), active joint position sense (JPS), electromyography (EMG), computerized hand-grip dynamometer (CHD), and the Patient-Rated Wrist Evaluation (PRWE) were used to assess twenty-four female participants 8 weeks following DRF treatment and their 24 matched-control healthy counterparts on wrist and hand sensibility, proprioception, muscle recruitment, grip force, muscle fatigue, and functional status. Participants following DRF demonstrated significantly (p < .05) greater sensory (i.e., JPS, TT), and motor (i.e., EMG, CHD) deficits than their control counterparts. A significantly higher functional deficit (i.e., PRWE) also existed among participants following DRF than the control group. Participants following surgical and non-surgical DRF treatment were found to be statistically different only on total grip force. Group differences on JPS and total grip force revealed the strongest effect size with the highest correlations to PRWE. EMG and muscle fatigue ratio group differences revealed a weaker effect size with a fair degree of correlation to PRWE. Pain significantly correlated with sensori-motor function. Age did not correlate with any measured variable. Significant wrist and hand sensori-motor impairment and functional deficits among older females 8 weeks following DRF surgical and non-surgical interventions were revealed. JPS and total grip force were the most clinically meaningful tests for assessing the sensori-motor status as well as explaining functional disability and pain levels for these patients. 2c.


The efficacy of excimer laser (308 nm) for vitiligo at different body sites.
The treatment with XeCl-excimer laser generated 308-nm UVB radiation has shown promising results in patients with vitiligo. In this controlled, prospective trial we studied the primary efficacy (start and grade of repigmentation) and patient's satisfaction of XeCl-excimer laser for treatment of vitiligo patches at different body sites and re-evaluated the achieved repigmentation 12 months after the end of therapy. Twenty-five patients with generalized or localized vitiligo with a total of 85 lesions at different body sites were enrolled in this study. Vitiligo patches were treated with 308-nm XeCl-excimer laser 3 times a week for 6 to 10 weeks. The overall repigmentation grade of each treated lesion was evaluated once a week on a 5 point scale rating from 0 (no repigmentation), 1 (1-5%), 2 (6-25%), 3 (26-50%), 4 (51-75%), to 5 (76-100%). Twenty-four patients completed the study. Within 6 to 10 weeks of treatment 67% of the patients (16/24) developed follicular repigmentation of at least one of their vitiligo lesions. Lesion repigmentation started after a mean of 13 treatments in lesions located on the face, trunk, arm, and/or leg (high-responder location), and after a mean of 22 treatments in lesions located on the elbow, wrist, dorsum of the hand, knee, and/or dorsum of the foot (low-responder location). Untreated control lesions and lesions located on the fingers did not achieve any repigmentation. After 10 weeks of treatment repigmentation of more than 75% was found in 25% (7/28) of lesions of the high-responder location group versus 2% (1/43) of lesions of the low-responder location group. In most cases, laser-induced repigmentation was persistent, as determined 12 months after the end of treatment. 308-nm excimer laser is an effective modality for the treatment of vitiligo. However, similar to other non-surgical treatment modalities, the therapeutic effect is mainly dependent on the location of vitiligo lesions.


Analysis of recombinant soluble mouse D-factor/LIF receptor.
The recombinant soluble mouse D-factor/LIF receptor (sD-factor-R) was expressed in COS-7 cells. Scatchard analysis of the bindings of mouse 125I-D-factor and human 125I-D-factor to the sD-factor-R indicated dissociation constants (Kd) of 12 and 0.267 nM, respectively, which were comparable to those of the binding protein in mouse serum. The apparent molecular masses of the sD-factor-R and human D-factor observed by gel filtration chromatography were 150 and 50 kDa, respectively. The size of the sD-factor-R.human D-factor complex was approximately 200 kDa, indicating that D-factor forms a 1:1 complex with the sD-factor-R. The sD-factor-R inhibited the induction of differentiation of mouse myeloid leukemic M1 cells by mouse D-factor by blocking the binding of this factor to the cells.


Creatine and Nicotinamide Prevent Oxidant-Induced Senescence in Human Fibroblasts.
Dermal fibroblasts provide structural support by producing collagen and other structural/support proteins beneath the epidermis. Fibroblasts also produce insulin-like growth factor-1 (IGF-1), which binds to the IGF-1 receptors (IGF-1Rs) on keratinocytes to activate signaling pathways that regulate cell proliferation and cellular responses to genotoxic stressors like ultraviolet B radiation. Our group has determined that the lack of IGF-1 expression due to fibroblast senescence in the dermis of geriatric individuals is correlated with an increased incidence of skin cancer. The present studies tested the hypothesis that pro-energetics creatine monohydrate (Cr) and nicotinamide (NAM) can protect normal dermal human fibroblasts (DHF) against experimentally induced senescence. To that end, we used an experimental model of senescence in which primary DHF are treated with hydrogen peroxide (H2O2) in vitro, with senescence measured by staining for beta-galactosidase activity, p21 protein expression, and senescence associated secretory phenotype cytokine mRNA levels. We also determined the effect of H2O2 on IGF-1 mRNA and protein expression. Our studies indicate that pretreatment with Cr or NAM protects DHF from the H2O2-induced cell senescence. Treatment with pro-energetics post-H2O2 had no effect. Moreover, these agents also inhibited reactive oxygen species generation from H2O2 treatment. These studies suggest a potential strategy for protecting fibroblasts in geriatric skin from undergoing stress-induced senescence, which may maintain IGF-1 levels and therefore limit carcinogenesis in epidermal keratinocytes.


Adult onset glutaric aciduria type I presenting with a leukoencephalopathy.
Glutaric aciduria type I usually presents with an acute metabolic crisis during infancy. The authors report a previously healthy 19-year-old woman who presented with recurrent headaches, oculomotor symptoms, and a severe leukoencephalopathy on MRI. The diagnosis was made by urinary organic acid analysis and confirmed by enzyme studies. Genetic analysis revealed compound heterozygosity with a deletion c.219delC in exon 3 and a novel missense mutation R132G in exon 5 of the glutaryl CoA dehydrogenase (GCDH) gene.


Isocitrate Dehydrogenase Mutations in Myelodysplastic Syndromes and in Acute Myeloid Leukemias.
Acute myeloid leukemia (AML) is a heterogeneous disease generated by the acquisition of multiple genetic and epigenetic aberrations which impair the proliferation and differentiation of hematopoietic progenitors and precursors. In the last years, there has been a dramatic improvement in the understanding of the molecular alterations driving cellular signaling and biochemical changes determining the survival advantage, stimulation of proliferation, and impairment of cellular differentiation of leukemic cells. These molecular alterations influence clinical outcomes and provide potential targets for drug development. Among these alterations, an important role is played by two mutant enzymes of the citric acid cycle, isocitrate dehydrogenase (IDH), IDH1 and IDH2, occurring in about 20% of AMLs, which leads to the production of an oncogenic metabolite R-2-hydroxy-glutarate (R-2-HG); this causes a DNA hypermethylation and an inhibition of hematopoietic stem cell differentiation. IDH mutations differentially affect prognosis of AML patients following the location of the mutation and other co-occurring genomic abnormalities. Recently, the development of novel therapies based on the specific targeting of mutant IDH may contribute to new effective treatments of these patients. In this review, we will provide a detailed analysis of the biological, clinical, and therapeutic implications of IDH mutations.


[Relationship of fatty liver with HBV infection, hyperlipidemia and abnormal alanine aminotransferase].
To investigate the prevalence of fatty liver in Guangzhou and its relation to hepatitis B virus (HBV) infection, hyperlipidemia and abnormal alanine aminotransferase (ALT). A retrospective analysis was made on clinical data of 4365 participants who received health check-up in our hospital. The prevalence of fatty liver was 18.2%, 19.2% in males and 17.1% in females, respectively. There was no significant difference between males and females (P = 0.07). Among 793 subjects with fatty liver, 440 were males and 353 were females. The prevalence of fatty liver was 16.7% in HBV infection group and 18.3% in the group without HBV infection, no significant difference was seen between these two groups (P = 0.45). The prevalence of fatty liver was 42.1% in hyperlipidemia group, and 11.6% in the group with normal serum triglyceride (TG) and total cholesterol (TC), respectively, there was a significant difference between these two groups (P < 0.05). The abnormal ALT was seen in 32.5% of the fattty liver group, which was significantly higher than that (8.6%) in the group without fatty liver (P < 0.05). The prevalence of fatty liver was not significantly different between males and females in Guangzhou. Fatty liver was not related with HBV infection but closely related with age and hyperlipidemia. Fatty liver was a common cause of abnormal ALT.


A reversed-phase high-performance liquid chromatography method for the determination of cotrimoxazole (trimethoprim/ sulphamethoxazole) in children treated for malaria.
A high-performance liquid chromatography (HPLC) method was developed for the simultaneous analysis of trimethoprim (TMP), sulphamethoxazole (SMX), and acetylsulphamethoxazole (AcSMX) in small amounts of blood. The method involved precipitation with 50 microL trichloracetic acid (1M) to 125 microL plasma or serum sample. 60 microL supernatant was added to 60 microL mobile phase, modified with 50microL 1 M sodium hydroxide/mL. The mobile phase consisted of 20% acetonitrile and 80% phosphate buffer adjusted to pH 6.15. Using 125 microL of the sample, limits of quantitation were 0.1 microg/mL for TMP, 1.0 microg/mL for SMX, and 1.0 microg/mL for AcSMX. The precision of the method was 2% to 11% over the range of concentrations tested, 0.5-30 microg/mL for TMP, 5-300 microg/mL for SMX, and 2.5-150 microg/mL for AcSMX, respectively. No interference with other commonly used drugs was observed. The method is rapid, simple, specific, and sensitive enough for pharmacokinetic studies. The small amount of blood required makes it suitable for pediatric patients. The method was used to analyze samples from Tanzanian children aged 6-59 months participating in a cotrimoxazole (TMP/SMX)/chloroquine randomized trial for the treatment of uncomplicated malaria. Venous blood samples from 68 children were collected 2 hours after the first dose of TMP/SMX (4 mg/kg TMP/20 mg/kg SMX at two divided doses for 5 days) and again at treatment day 4. Individual variations in plasma concentrations of TMP, SMX, and AcSMX were considerable. The mean and SEM plasma concentrations (g/mL) of TMP, SMX, and AcSMX 2 hours after the first treatment dose were 2.0 +/- 1.0 (range 0.5-6), 53 +/- 22 (range 24-146), and 13.5 +/- 12 (range 0-65), respectively. On the fourth day the attained plasma concentrations were not significantly different from samples collected after the first dose.


Steroidogenic enzymes in skin.
The gonadal synthesis of testosterone from cholesterol involves four enzymes, namely, cytochrome P-450 side-chain cleavage enzyme, cytochrome P-450 17a-hydroxylase/lyase, 3b-hydroxysteroid dehydrogenase, and 17b-hydroxysteroid dehydrogenase. A significant part of the plasma-borne testosterone is converted in androgen target tissues, such as the skin, to the more potent androgen dihydrotestosterone by the steroid 5a-reductase type 1 and type 2 isoenzymes. Dihydrotestosterone, which binds to the nuclear androgen receptor with much greater affinity than testosterone, is the androgen responsible for a process leading to androgenetic alopecia. Consequently, the 5a-reductase inhibitor finasteride was developed and has proven efficacious in promoting hair growth as a consequence of lowering scalp and plasma dihydrotestosterone levels. In contrast to the direct synthesis of dihydrotestosterone from testosterone, biologically inactive C19-steroids produced by glandular and peripheral tissues may also feed into the scalp skin production of dihydrotestosterone by the local expression of reductive 17b-hydroxysteroid dehydrogenase, oxidative 3a-hydroxysteroid dehydrogenase, and 3b-hydroxysteroid dehydrogenase enzymes. Aberrant expression of one or more of these enzymes, could conceivably result in increased scalp dihydrotestosterone levels, and possibly, acceleration of the balding process in genetically predisposed men and women.


Human bronchial epithelial cells differentiate to 3D glandular acini on basement membrane matrix.
To create a model system that investigates mechanisms resulting in hyperplasia and hypertrophy of respiratory tract submucosal glands, we developed an in vitro three-dimensional (3D) system wherein normal human bronchial epithelial (HBE) cells differentiated into glandular acini when grown on a basement membrane matrix. The differentiation of primary HBE cells into glandular acini was monitored temporally by light microscopy. Apoptosis-induced lumen formation was observed by immunofluorescence analysis. The acinar cells expressed and secreted MUC5B mucin (marker for glandular mucous cells) and lysozyme, lactoferrin, and zinc-α2-glycoprotein (markers for glandular serous cells) at Day 22. β-Tubulin IV, a marker for ciliated cells, was not detected. Expression of mucous and serous cell markers in HBE glandular acini demonstrated that HBE cells grown on a basement membrane matrix differentiated into acini that exhibit molecular characteristics of respiratory tract glandular acinar cells. Inhibition studies with neutralizing antibodies resulted in a marked decrease in size of the spheroids at Day 7, demonstrating that laminin (a major component of the basement membrane matrix), the cell surface receptor integrin α6, and the cell junction marker E-cadherin have functional roles in HBE acinar morphogenesis. No significant variability was detected in the average size of glandular acini formed by HBE cells from two normal individuals. These results demonstrated that this in vitro model system is reproducible, stable, and potentially useful for studies of glandular differentiation and hyperplasia.


PI3Kδ inhibitor, GS-1101 (CAL-101), attenuates pathway signaling, induces apoptosis, and overcomes signals from the microenvironment in cellular models of Hodgkin lymphoma.
GS-1101 (CAL-101) is an oral PI3Kδ-specific inhibitor that has shown preclinical and clinical activity in non-Hodgkin lymphoma and chronic lymphocytic leukemia. To investigate the potential role of PI3Kδ in Hodgkin lymphoma (HL), we screened 5 HL cell lines and primary samples from patients with HL for PI3Kδ isoform expression and constitutive PI3K pathway activation. Inhibition of PI3Kδ by GS-1101 resulted in the inhibition of Akt phosphorylation. Cocultures with stroma cells induced Akt activation in HL cells, and this effect was blocked by GS-1101. Conversely, production of the stroma-stimulating chemokine, CCL5, by HL cells was reduced by GS-1101. GS-1101 also induced dose-dependent apoptosis of HL cells at 48 hours. Reductions in cell viability and apoptosis were enhanced when combining GS-1101 with the mTOR inhibitor everolimus. Our findings suggest that excessive PI3Kδ activity is characteristic in HL and support clinical evaluation of GS-1101, alone and in combination, as targeted therapy for HL.


Peptide Self-Assembled Nanostructures with Distinct Morphologies and Properties Fabricated by Molecular Design.
Six surfactant-like peptides with the same amino acid composition but different primary sequences are designed, including G3A3V3I3K3, K3I3V3A3G3, I3V3A3G3K3, K3G3A3V3I3, V3G3I3A3K3, and K3A3I3G3V3. These peptides form antiparallel β-sheets during self-assembly. Because the constituent residues have different side chain size and hydrophobicity, sequence changes adjust group distribution and hydrophobicity on the two sides of a given β-sheet. This consequently tunes the binding energy of the side-to-side pairing conformations and leads to different self-assembled structures. G3A3V3I3K3 and K3I3V3A3G3 form short nanorods with diameters of 8.5 ± 1.0 nm and lengths <150 nm. I3V3A3G3K3 and K3G3A3V3I3 form nanosheets with heights of 4.0 ± 0.5 nm and limited lengths and widths. V3G3I3A3K3 and K3A3I3G3V3 form long fibrils with diameters of 7.0 ± 1.0 nm and lengths of micrometer scale. These nanostructures exhibit different capacity in encapsulating insoluble hydrophobic drug molecules and delivering them into the cells. The nanosheets of I3V3A3G3K3 and K3G3A3V3I3 can encapsulate both nile red and doxorubicin molecules to an extent of up to 17-23% in mole ratio. Moreover, the shape and size of the nanostructures affect the drug delivery into cells greatly, with the nanosheets and short rods exhibiting higher efficiency than the long fibrils. The study provides new insights into programmed peptide self-assembly toward specific functionalities.


Functional and health-promoting properties of probiotics' exopolysaccharides; isolation, characterization, and applications in the food industry.
Exopolysaccharides (EPS) are extracellular sugar metabolites/polymers of some slim microorganisms and, a wide variety of probiotics have been broadly investigated for their ability to produce EPS. EPS originated from probiotics have potential applications in food, pharmaceutical, cosmetology, wastewater treatment, and textiles industries, nevertheless slight is recognized about their function. The present review purposes to comprehensively discuss the structure, classification, biosynthesis, extraction, purification, sources, health-promoting properties, techno-functional benefits, application in the food industry, safety, toxicology, analysis, and characterization methods of EPS originated from probiotic microorganisms. Various studies have shown that probiotic EPS used as stabilizers, emulsifiers, gelling agents, viscosifiers, and prebiotics can alter the nutritional, texture, and rheological characteristics of food and beverages and play a major role in improving the quality of these products. Numerous studies have also proven the beneficial health effects of probiotic EPS, including antioxidant, antimicrobial, anti-inflammatory, immunomodulatory, anticancer, antidiabetic, antibiofilm, antiulcer, and antitoxin activities. Although the use of probiotic EPS has health effects and improves the organoleptic and textural properties of food and pharmaceutical products and there is a high tendency for their use in related industries, the production yield of these products is low and requires basic studies to support their products in large scale.


The oriental armyworm ( Mythimna separata) feeding induces systemic defence responses within and between maize leaves.
Maize ( Zea mays) is a staple cereal crop cultivated all over the world but that is threatened by various insects. Feeding of the lepidopteran insect Mythimna separata triggers defence signalling and increases anti-herbivore benzoxazinoids (Bxs) in the insect-damaged maize leaves. However, the herbivory-elicited within-leaf and leaf-to-leaf systemic signalling in maize remains largely unexplored. Here, we show that simulated M. separata herbivory and mechanical wounding elicited increased levels of jasmonic acid (JA), JA-Ile (JA-isoleucine conjugate) and Bxs in the damaged areas and in specific systemic regions within a leaf. Importantly, increased contents of Bxs were detected in a systemic leaf, and consistently, this leaf exhibited increased defence against M. separata. Increased JA/JA-Ile and altered transcriptome, including Bx biosynthesis genes, were detected in systemic leaves after wounding or simulated herbivory treatments, although only simulated herbivory induced increase of the contents of Bxs systemically. Promoter and co-expression analysis revealed that transcription factors bHLH57 and WRKY34 may regulate Bx biosynthesis genes in systemic leaves. Moreover, leaf ablation experiment indicated that the systemic signal rapidly exited the local leaves within 30 min after elicitation. This study provides new insight into the temporal and spatial regulation of defence responses of maize against lepidopteran insects. This article is part of the theme issue 'Biotic signalling sheds light on smart pest management'.


The activation of Rac1 by M3 muscarinic acetylcholine receptors involves the translocation of Rac1 and IQGAP1 to cell junctions and changes in the composition of protein complexes containing Rac1, IQGAP1, and actin.
The abilities of the M(3) muscarinic acetylcholine receptor (mAChR) and Rac1 to regulate similar cellular responses, including cadherin-mediated adhesion, prompted us to investigate Rac1 regulation by M(3) mAChR. We characterized changes in Rac1 induced by stimulating transfected M(3) mAChR in Chinese hamster ovary cells stably expressing hemagglutinin (HA)-tagged wild-type or mutant Rac1. mAChR activation converts endogenous Rac1 to the GTP-bound form in cells expressing HA-Rac1 but not in cells expressing dominant negative HA-Rac1(Asn-17) or constitutively active HA-Rac1(Val-12). The competitive binding of endogenous IQGAP1 by HA-Rac1(Val-12) may diminish the mAChR-mediated activation of endogenous Rac1. HA-Rac1 and HA-Rac1(Val-12), but not HA-Rac1(Asn-17), accumulate with IQGAP1 at cell junctions during mAChR-induced cell-cell compaction. Co-localization studies suggest that Rac1 can accumulate at junctions without IQGAP1, but IQGAP1 cannot accumulate at junctions without Rac1. mAChR activation also induces GTP-independent changes in Rac1 because mAChR activation redistributes HA-Rac1(Asn-17), which does not bind GTP. Actin associates with complexes containing HA-Rac1 or HA-Rac1(Val-12) after prolonged mAChR activation. We also demonstrate that Rac1 participates in mAChR-induced cell-cell compaction and c-Jun phosphorylation. These results indicate that M(3) mAChR activation converts Rac1 to the GTP-bound form, alters interactions between Rac1, IQGAP1, and actin, and causes the junctional accumulation of Rac1 and IQGAP1.


The presence and severity of chronic kidney disease predicts all-cause mortality in type 1 diabetes.
This study aimed to identify clinical features associated with premature mortality in a large contemporary cohort of adults with type 1 diabetes. The Finnish Diabetic Nephropathy (FinnDiane) study is a national multicenter prospective follow-up study of 4,201 adults with type 1 diabetes from 21 university and central hospitals, 33 district hospitals, and 26 primary health care centers across Finland. During a median 7 years of follow-up, there were 291 deaths (7%), 3.6-fold (95% CI 3.2-4.0) more than that observed in the age- and sex-matched general population. Excess mortality was only observed in individuals with chronic kidney disease. Individuals with normoalbuminuria showed no excess mortality beyond the general population (standardized mortality ratio [SMR] 0.8, 95% CI 0.5-1.1), independent of the duration of diabetes. The presence of microalbuminuria, macroalbuminuria, and end-stage kidney disease was associated with 2.8, 9.2, and 18.3 times higher SMR, respectively. The increase in mortality across each stage of albuminuria was equivalent to the risk conferred by preexisting macrovascular disease. In addition, the glomerular filtration rate was independently associated with mortality, such that individuals with impaired kidney function, as well as those demonstrating hyperfiltration, had an increased risk of death. An independent graded association was observed between the presence and severity of kidney disease and mortality in a large contemporary cohort of individuals with type 1 diabetes. These findings highlight the clinical and public health importance of chronic kidney disease and its prevention in the management of type 1 diabetes.


Early Activation by Ethylene of the Tonoplast H-Pumping ATPase in the Latex from Hevea brasiliensis.
The treatment of Hevea brasiliensis (rubber tree) bark by chloro-2-ethyl phosphonic acid (ethrel), an ethylene-producing compound, induces a significant increase in the tonoplast H(+)-translocating ATPase activity in the latex during the first 24 hours after the application of the stimulating agent. Moreover, the tonoplast-bound ATPase is highly activated when vacuoles (lutoids) are resuspended in ultrafiltrated cytosol. This effect is amplified during ethrel stimulation. Preliminary assays to characterize the endogenous effector(s) suggest that the activator(s) could be a heat-resistant compound with a low molecular weight, most likely an anion. The activation of the tonoplast-bound ATPase and the associated activation of the protons translocation across the lutoid membrane, could explain the cytosolic alkalinization observed in latex following the ethrel treatment of Hevea bark, which results in an enhanced rubber production.


Cardiorespiratory function, resting metabolic rate and heart rate variability in coal miners exposed to hypobaric hypoxia in highland workplace.
Owing to intermittent/acute exposure to hypobaric hypoxia, highland miners may often suffer, the physiological characteristics between highland and lowland miners, however, are rarely reported. The objective of this study was to compare the physiological characteristics of coal miners working at disparate altitudes. Twenty-three male coal mining workers acclimating to high altitude for 30 ± 6 days in Tibet (highland group; approx. 4500 m above sea level; 628.39 millibar), and 22 male coal mining workers in Hebei (lowland group; less than 100 m above sea level; 1021.82 millibar) were recruited. Tests were conducted to compare ventilatory parameters, circulation parameters, resting metabolic rate (RMR), and heart rate variability (HRV) indices between the two groups in resting state. Ventilation volume per minute (VE) of the highland group was markedly raised compared to that of the lowland group (11.70 ± 1.57 vs. 8.94 ± 1.97 L/min, p = 0.000). In the meanwhile, O2 intake per heart beat (VO2/HR) was strikingly decreased (3.54 ± 0.54 vs. 4.36 ± 0.69 ml/beat, p = 0.000). Resting metabolic rate relevant to body surface area (RMR/BSA) was found no significant difference between the two groups. Evident reduction in standard deviation of NN intervals (SDNN) and remarkable increase in ratio of low- and high- frequency bands (LF/HF) were manifest in highland miners compared to that of lowland ones (110.82 ± 33.34 vs. 141.44 ± 40.38, p = 0.008 and 858.86 ± 699.24 vs. 371.33 ± 171.46, p = 0.003; respectively). These results implicate that long-term intermittent exposure to high altitude can lead miners to an intensified respiration, a compromised circulation and a profound sympathetic-parasympathetic imbalance, whereas the RMR in highland miners does not distinctly decline.


LPA6-RhoA signals regulate junctional complexes for polarity and morphology establishment of maturation stage ameloblasts.
Lysophosphatidic acid (LPA) is a potent bioactive phospholipid that exerts various functions upon binding to six known G protein-coupled receptors (LPA1-6); however; its role in a tooth remains unclear. This study aimed to explore the impact of the LPA/LPA receptor 6 (LPA6)/RhoA signaling axis on maturation stage ameloblasts (M-ABs), which are responsible for enamel mineralization. The expression of LPA6 and LPA-producing synthetic enzymes during ameloblast differentiation was explored through immunobiological analysis of mouse incisors and molars. To elucidate the role of LPA6 in ameloblasts, incisors of LPA6 KO mice were analyzed. In vitro experiments using ameloblast cell lines were performed to validate the function of LPA-LPA6-RhoA signaling in ameloblasts. LPA6 and LPA-producing enzymes were strongly expressed in M-ABs. In LPA6 knockout mice, M-ABs exhibited abnormal morphology with the loss of cell polarity, and an abnormal enamel epithelium containing cyst-like structures was formed. Moreover, the expression of E-cadherin and zonula occludens-1 (ZO-1) significantly decreased in M-ABs. In vitro experiments demonstrated that LPA upregulated the expression of E-cadherin, ZO-1, and filamentous actin (F-actin) at the cellular membrane, whereas LPA6 knockdown decreased their expression and changed cell morphology. Furthermore, we showed that RhoA signaling mediates LPA-LPA6-induced junctional complexes. This study demonstrated that LPA-LPA6-RhoA signaling is essential for establishing proper cell morphology and polarity, via cell-cell junction and actin cytoskeleton expression and stability, of M-ABs. These results highlight the biological significance of bioactive lipids in a tooth, providing a novel molecular regulatory mechanism of ameloblasts.


Lymphatic mapping in endometrial cancer: comparison of hysteroscopic and subserosal injection and the distribution of sentinel lymph nodes.
Endometrial cancer incidence increases over the age of 65 and lymphadenectomy in these women is a morbid procedure. Sentinel lymph node (SLN) should avoid extensive lymphadenectomy in node negative patients. The aim of this prospective study is to determine the feasibility and usefulness of lymphatic mapping and SLN identification in the management of endometrial cancer. From January 2004 to December 2007 101 women with endometrial cancer participated in the study. We injected Tc hysteroscopically, peritumorally 2 hours before laparotomy in 24 women. We applied Tc and blue dye subserously after laparotomy and before adhesiolysis in 67 women. Ten patients with metastatic disease in ovary, omentum, peritoneum, and bulky nodes were excluded from analysis. We detected SLN in 12 women (50%) in hysteroscopic group and in 49 women (73.1%) in subserous group. We identified 133 SLNs in 61 women. We found 20 SLNs (15.0%) in supraobturator region, 78 (58.6%) in external iliac area, 11 (8.3%) in paraaortal area, 13 (9.8%) on common iliac artery, 8 (6.0%) in medial part of lateral parametrium, and 3 (2.3%) in presacral area. Sentinel lymph node identification is a new strategy that can be used to examine nodal status with a high successful rate in breast, cervical, and vulvar cancer. Results in endometrial cancer are not as successful, however. In the future, it will be necessary to find optimal timing, the best route of application, and the "right" size of the Tc particles. Subserous application seems to be superior to hysteroscopic application.


Recognition of affective prosody in brain-damaged patients and healthy controls: a neurophysiological study using EEG and whole-head MEG.
A passive oddball experiment was used in which stimuli were emotional exclamations differing in their affective tone. In both electroencephalography (EEG) and magnetoencephalography (MEG), deviants elicited an N300 component, sometimes accompanied by a slow wave. Both components had a symmetrical distribution, but the former was more posterior than the latter. The same responses to prosodic stimuli were significant in 6 of 27 patients with severe disorders of consciousness (persistent vegetative state and minimally conscious state) and in all 3 of the examined locked-in patients, indicating that the procedure can be applied for testing neurological patients. The occurrence of significant responses depended on the presence or absence of a lesion to the right temporal lobe. Obviously, the N300 depends on the activity of the right temporal cortex but does not originate there. We suggest that the component is related not to the recognition of affective prosody as such, but to the following detection of affective mismatch due to violations of emotional context of stimulation.


Nicotine-Derived Compounds as Therapeutic Tools Against Post-Traumatic Stress Disorder.
Post-traumatic stress disorder (PTSD) is an anxiety disorder that develops after experiencing trauma. Actual therapies do not help majority of patients with PTSD. Moreover, extinguished fear memories usually reappear in the individuals when exposed to trauma cues. New drugs to reduce the impact of conditioned cues in eliciting abnormal fear responses are urgently required. Cotinine, the main metabolite of nicotine, decreased anxiety and depressive-like behavior, and enhanced fear extinction in mouse models of PTSD. Cotinine, considered a positive modulator of the α7 nicotinic acetylcholine receptor (α7nAChR), enhances fear extinction in rodents in a manner dependent on the activity of the αnAChRs. Cotinine stimulates signaling pathways downstream of α7nAChR including the protein kinase B (Akt)/glycogen synthase kinase 3β (GSK3β) pathway and the extracellular signal-regulated kinases (ERKs). The stimulation of these factors promotes synaptic plasticity and the extinction of fear. In this review, we discuss the hypothesis that cotinine relieves PTSD symptoms and facilitates fear memory extinction by promoting brain plasticity through the positive modulation of presynaptic nAChRs and its effectors in the brain.


Small-molecule proteostasis regulators for protein conformational diseases.
Protein homeostasis (proteostasis) is essential for cellular and organismal health. Stress, aging and the chronic expression of misfolded proteins, however, challenge the proteostasis machinery and the vitality of the cell. Enhanced expression of molecular chaperones, regulated by heat shock transcription factor-1 (HSF-1), has been shown to restore proteostasis in a variety of conformational disease models, suggesting this mechanism as a promising therapeutic approach. We describe the results of a screen comprised of ∼900,000 small molecules that identified new classes of small-molecule proteostasis regulators that induce HSF-1-dependent chaperone expression and restore protein folding in multiple conformational disease models. These beneficial effects to proteome stability are mediated by HSF-1, FOXO, Nrf-2 and the chaperone machinery through mechanisms that are distinct from current known small-molecule activators of the heat shock response. We suggest that modulation of the proteostasis network by proteostasis regulators may be a promising therapeutic approach for the treatment of a variety of protein conformational diseases.


Treatment of familial male precocious puberty with spironolactone, testolactone, and deslorelin.
Combined antiandrogen (spironolactone) and aromatase inhibitor (testolactone) are effective for the short term treatment of familial male precocious puberty. During this therapy, plasma testosterone levels remain in the adult range, since spironolactone blocks the testosterone receptor without significantly affecting plasma testosterone levels. After our initial 18-month pilot study, we continued to treat eight boys with the combined therapy for 2.0-4.2 yr. During this time all boys exhibited a pubertal rise in gonadotropin secretion and a diminishing response to treatment, which was manifested by the recurrence of clinical features of puberty and an increase in the bone maturation rate (P < 0.05). Addition of the LHRH agonist deslorelin (4 micrograms/kg.day, sc) to the combined therapy decreased peak LH, plasma testosterone, bone maturation rate, and growth velocity (P < 0.05) over the next year. We conclude that the rise in gonadotropin levels during central activation of hypothalamic LHRH secretion in boys with familial male precocious puberty causes a partial escape from the combined effect of spironolactone and testolactone. The addition of deslorelin to the combined therapy appears to restore the control of puberty in this setting.


Tegaserod, a 5-hydroxytryptamine type 4 receptor partial agonist, is devoid of electrocardiographic effects.
Certain GI prokinetic agents have been shown to affect cardiac repolarization, which may be associated with life-threatening arrhythmias. The selective 5-hydroxytryptamine type 4 receptor partial agonist tegaserod is a novel promotile agent developed for the treatment of functional motility disorders such as irritable bowel syndrome (IBS). The aim of the study was to investigate the cardiac safety profile of tegaserod through analysis of electrocardiographic data from clinical studies conducted in patients with IBS and a study conducted in healthy male subjects. In three randomized, double blind, placebo-controlled, parallel group clinical studies, 2516 IBS patients with symptoms of abdominal pain and constipation received tegaserod 2 or 6 mg b.i.d. (n = 1679) or placebo (n = 837) for 12 wk. In an additional study, 36 healthy male subjects received iv. single doses of tegaserod (0.8 mg to 20 mg) or placebo. Standard 12-lead electrocardiograms were recorded at baseline and during treatment. Baseline values were compared with data collected during the treatment period. The proportion of patients with prolongation of the QTc interval was the same for placebo and tegaserod, as was the frequency of overall electrocardiographic abnormalities. No ventricular or supraventricular tachycardia was observed. Comparable electrocardiographic results were obtained during placebo and tegaserod treatment. In healthy volunteers, tegaserod at i.v. doses resulting in plasma concentrations up to 100 times those measured after therapeutic doses (6 mg b.i.d.) did not influence electrocardiographic parameters. Tegaserod is devoid of electrocardiographic effects and is not expected to adversely influence cardiac function. These data confirm preclinical findings.


Site-directed mutagenesis of the phosphorylation sites in the mouse glucocorticoid receptor.
The functional significance of receptor phosphorylation in mediating the actions of glucocorticoids remains undefined. The identification of seven phosphorylation sites in the mouse glucocorticoid receptor (Bodwell, J. E., Orti, E., Coull, J. M., Pappin, D. J. C., Smith, L. I., and Swift, F. (1991) J. Biol. Chem. 266, 7549-7555) permits a direct examination of the potential regulatory role of glucocorticoid receptor phosphorylation in transactivation. Using oligonucleotide-directed mutagenesis of the mouse glucocorticoid receptor cDNA, we have substituted alanine or aspartate for the residues phosphorylated in this ligand-dependent transcription factor. COS-1 cells were cotransfected with mutant receptor cDNA expression vectors and a reporter plasmid containing the glucocorticoid-inducible mouse mammary tumor virus promoter linked to chloramphenicol acetyltransferase in order to characterize the effect of these substitutions on receptor-mediated gene expression. Substitution of alanine or aspartate at single phosphorylation sites does not prevent receptor transactivation. Receptors containing multiple substitutions of alanine or aspartate at the major phosphorylation sites in the acidic domain elicit levels of hormone-induced reporter gene expression that are comparable to wild-type receptors. Mutant receptors substituted with alanine at the five phosphorylation sites conserved among the rat, human, and mouse receptors exhibit a 22% decrease in transcriptional activity. Receptors mutated at all seven sites display a similar modest reduction. These results demonstrate that receptor phosphorylation at these seven identified residues is not a major determinant in glucocorticoid receptor transcriptional activity at the mouse mammary tumor virus promoter.


Management of early postoperative coronary artery bypass graft failure.
Perioperative graft failure following coronary artery bypass grafting (CABG) may result in acute myocardial ischaemia. Whether acute percutaneous coronary intervention, emergency reoperation or conservative intensive care treatment should be used is currently unknown. Between 2003 and 2009, 39 of the 5598 patients who underwent isolated CABG surgery underwent early postoperative coronary angiography for suspected myocardial ischaemia. Following angiography, two groups were identified: patients who underwent immediately reintervention (group 1); and those treated conservatively (group 2). Primary study endpoints were mortality and postoperative myocardial infarct size. Postoperative coronary angiography revealed early perioperative bypass graft failure in 32 of 39 patients. Acute percutaneous coronary intervention was performed in 15 patients, redo-CABG in 4 patients and conservative treatment in 13 patients. The number of failing bypass grafts were significantly higher in group 1 compared with group 2 (P = 0.0251). A trend toward lower post-procedural peak cardiac troponin T and creatinine phosphokinase serum levels in group 1 was observed (163.0 vs. 206.0 and 4.35 vs. 5.53, respectively) (P = 0.0662 and 0.1648). Early reintervention may limit the extent of myocardial cellular damage compared with conservative medical strategy in patients with myocardial ischaemia due to early graft failure.


Likelihood based approaches to handling data below the quantification limit using NONMEM VI.
To evaluate the likelihood-based methods for handling data below the quantification limit (BQL) using new features in NONMEM VI. A two-compartment pharmacokinetic model with first-order absorption was chosen for investigation. Methods evaluated were: discarding BQL observations (M1), discarding BQL observations but adjusting the likelihood for the remaining data (M2), maximizing the likelihood for the data above the limit of quantification (LOQ) and treating BQL data as censored (M3), and like M3 but conditioning on the observation being greater than zero (M4). These four methods were compared using data simulated with a proportional error model. M2, M3, and M4 were also compared using data simulated from a positively truncated normal distribution. Successful terminations and bias and precision of parameter estimates were assessed. For the data simulated with a proportional error model, the overall performance was best for M3 followed by M2 and M1. M3 and M4 resulted in similar estimates in analyses without log transformation. For data simulated with the truncated normal distribution, M4 performed better than M3. Analyses that maximized the likelihood of the data above the LOQ and treated BQL data as censored provided the most accurate and precise parameter estimates.


Essential role of the B23/NPM core domain in regulating ARF binding and B23 stability.
How cells coordinate inhibition of growth and division during genotoxic events is fundamental to our understanding of the origin of cancer. Despite increasing interest and extensive study, the mechanisms that link regulation of DNA synthesis and ribosomal biogenesis remain elusive. Recently, the tumor suppressor p14(ARF) (ARF) has been shown to interact functionally with the nucleolar protein B23/NPM (B23) and inhibit rRNA biogenesis. However, the molecular basis of the ARF-B23 interaction is hitherto unclear. Here we show that a highly conserved motif in the B23 oligomerization domain is essential for mediating ARF binding in vivo. Mutagenesis of conserved B23 core residues (L102A, G105A, G107A) prevented B23 from interacting with ARF. Modeling of the B23 core indicated that substitutions in the GSGP loop motif could trigger conformational changes in B23 thereby obstructing ARF binding. Interestingly, the GSGP loop mutants were unstable, defective for oligomerization, and delocalized from the nucleolus to the nucleoplasm. B23 core mutants displayed increased ubiquitination and proteasomal degradation. We conclude that the functional integrity of the B23 core motif is required for stability, efficient nucleolar localization as well as ARF binding.


Deubiquitinases: Novel Therapeutic Targets in Immune Surveillance?
Inflammation is a protective response of the organism to tissue injury or infection. It occurs when the immune system recognizes Pathogen-Associated Molecular Patterns (PAMPs) or Damage-Associated Molecular Pattern (DAMPs) through the activation of Pattern Recognition Receptors. This initiates a variety of signalling events that conclude in the upregulation of proinflammatory molecules, which initiate an appropriate immune response. This response is tightly regulated since any aberrant activation of immune responses would have severe pathological consequences such as sepsis or chronic inflammatory and autoimmune diseases. Accumulative evidence shows that the ubiquitin system, and in particular ubiquitin-specific isopeptidases also known as deubiquitinases (DUBs), plays crucial roles in the control of these immune pathways. In this review we will give an up-to-date overview on the role of DUBs in the NF-κB pathway and inflammasome activation, two intrinsically related events triggered by activation of the membrane TLRs as well as the cytosolic NOD and NLR receptors. Modulation of DUB activity by small molecules has been proposed as a way to control dysregulation or overactivation of these key players of the inflammatory response. We will also discuss the advances and challenges of a potential use of DUBs as therapeutic targets in inflammatory pathologies.


Ornithine decarboxylase, polyamines, and pyrrolizidine alkaloids in senecio and crotalaria.
When tested for ornithine and arginine decarboxylases, pyrrolizidine alkaloid-bearing Senecio riddellii, S. longilobus (Compositae), and Crotalaria retusa (Leguminosae) plants exhibited only ornithine decarboxylase activity. This contrasts with previous studies of four species of pyrrolizidine alkaloid-bearing Heliotropium (Boraginaceae) in which arginine decarboxylase activity was very high relative to that of ornithine decarboxylase. Unlike Heliotropium angiospermum and Heliotropium indicum, in which endogenous arginine was the only detectable precursor of putrescine channeled into pyrrolizidines, in the species studied here-using difluoromethylornithine and difluoromethylarginine as the enzyme inhibitors-endogenous ornithine was the main if not the only precursor of putrescine converted into the alkaloid aminoalcohol moiety. In S. riddellii and C. retusa at flowering, ornithine decarboxylase activity was present mainly in leaves, especially the young ones. However, other very young organs such as inflorescence and growing roots exhibited much lower or very low activities; the enzyme activity in stems was negligible. There was no correlation between the enzyme activity and polyamine or alkaloid content in either species. In both species only free polyamines were detected except for C. retusa roots and inflorescence-with relatively very high levels of these compounds-in which conjugated putrescine, spermidine, and spermine were also found; agmatine was not identified by HPLC in any plant organ except for C. retusa roots with rhizobial nodules. Organ- or age-dependent differences in the polyamine levels were small or insignificant. The highest alkaloid contents were found in young leaves and inflorescence.


The Roles of Mitochondrial Folate Metabolism in Supporting Mitochondrial DNA Synthesis, Oxidative Phosphorylation, and Cellular Function.
Folate-mediated one-carbon metabolism (FOCM) is compartmentalized within human cells to the cytosol, nucleus, and mitochondria. The recent identifications of mitochondria-specific, folate-dependent thymidylate [deoxythymidine monophosphate (dTMP)] synthesis together with discoveries indicating the critical role of mitochondrial FOCM in cancer progression have renewed interest in understanding this metabolic pathway. The goal of this narrative review is to summarize recent advances in the field of one-carbon metabolism, with an emphasis on the biological importance of mitochondrial FOCM in maintaining mitochondrial DNA integrity and mitochondrial function, as well as the reprogramming of mitochondrial FOCM in cancer. Elucidation of the roles and regulation of mitochondrial FOCM will contribute to a better understanding of the mechanisms underlying folate-associated pathologies.


[Decrease of paraoxonase activity in the first day after abdominal operations in women].
Paraoxonase (PON1) is one of enzymes participating in antioxidative defence mechanism, which protects the organism from the results of action of reactive oxygen species. PON1 is an esterase contained in plasma high density lipoproteins (HDL) which protects HDL from peroxidation and plasma membranes from free radical injury. PON1 serum activity is diminished in some diseases (atherosclerosis, diabetes, coronary artery disease) and after cardiosurgery operations. But there is no information about changes in PON1 activity after abdominal operations. Of this study was the assessment of PON1 serum activity after the abdominal operations in women. 40 women operated in the 1st Department of Gynaecology of Lublin Medical University were studied, including 34 patients after total abdominal hysterectomy with salpingo-oophorectomy and 6 patients after myomectomy or cyst enucleation. The blood to experiments was collected before operation, and 8 and 24 hours after. PON1 serum activity was assessed spectrophotometrically toward two synthetic substrates: paraoxon and phenyl acetate. PON1 activity toward paraoxon before operation was 146.8 +/- 17.3 U/ml, and after 8 and 24 hours after 107.6 +/- 11.9 U/ml and 119.9 +/- 16.9 U/ml, respectively (results statistically significant). PON1 activity toward phenyl acetate (arylesterase activity) before operation was 182.7 +/- 16.4 U/ml and did not significantly changed after the surgery. It is observed decreased PON1 serum activity in the early postoperative period after abdominal operations in women, what can be connected with the utilisation of PON1 in antioxidative defence mechanism.


The Role of a Physician-Staffed Helicopter in Emergency Care of Patients on Isolated Danish Islands.
Emergency calls may lead to the dispatch of either ground ambulances or helicopter emergency medical services (HEMS). For residents on isolated islands, the HEMS can reduce the time to hospital admission and lead to improved outcomes. This study investigated the emergency care for residents on isolated islands with a focus on the role of a physician-staffed helicopter. The data were obtained from Danish national registries and databases. We included data on emergency calls from isolated islands from the time of emergency call to discharge. We identified 1130 emergency calls from which 775 patients were registered with a hospital admission. Of these, 41% were transported by the HEMS and 36% by a ground ambulance. The median time to admission was 83 min (IQR 66-104) and 90 min (IQR 45-144) for the HEMS and ground ambulance, respectively (p = 0.26). The overall 30-day mortality was 6.2% (95% CI: 4.6-8.1%), and 37% of all the patients were admitted to the hospital with an unspecified diagnosis. The emergency calls from isolated islands led to the dispatch of the HEMS in 41% of the cases. The use of the HEMS did not significantly reduce the time to admission but was used in a greater proportion of patients with an acute cardiac disease (66%) or stroke (67%).


Raised levels of Il-6, Il-17a, and Il-22 in fatal leptospirosis.
Clinical manifestations of leptospirosis range from mild, common cold-like illness, to a life-threatening condition. The host immune response has been hypothesized to play a major role in leptospirosis outcome. Increased levels of inflammatory mediators, such as cytokines, may promote tissue damage that lead to increased disease severity. The question is whether cytokines levels may predict the outcome of leptospirosis and guide patient management. This study aimed to assess the association between Th1-, Th2-, and Th17-related cytokines with the clinical outcome of patients with leptospirosis. Different cytokine levels were measured in fifty-two plasma samples of hospitalized patients diagnosed with leptospirosis in Malaysia (January 2016-December 2017). Patients were divided into two separate categories: survived (n = 40) and fatal outcome (n = 12). Nineteen plasma samples from healthy individuals were obtained as controls. Cytokine quantification was performed using Simple Plex™ assays from ProteinSimple (San Jose, CA, USA). Measurements were done in triplicate and statistical analysis was performed using GraphPad software and SPSS v20. IL-6 (p = 0.033), IL-17A (p = 0.022), and IL-22 (p = 0.046) were significantly elevated in fatal cases. IL-17A concentration (OR 1.115; 95% CI 1.010-1.231) appeared to be an independent predictor of fatality of leptospirosis. Significantly higher levels of TNF-α (p ≤ 0.0001), IL-6 (p ≤ 0.0001), IL-10 (p ≤ 0.0001), IL-12 (p ≤ 0.0001), IL17A (p ≤ 0.0001), and IL-18 (p ≤ 0.0001) were observed among leptospirosis patients in comparison with healthy controls. Our study shows that certain cytokine levels may serve as possible prognostic biomarkers in leptospirosis patients.


[Preparation of rabbit antibody against human Id3 and identification of intracellular localization of Id3 in tumor cells].
To express and purify the recombinant human inhibitor of differentiation 3 (Id3) in E.coli and prepare rabbit polyclonal antibody against Id3. The expression vector Id3/pET32a was transformed into E.coli BL21(DE3) and expression of histidine (His)-tagged Id3 fusion protein was induced with IPTG and confirmed by SDS-PAGE and Western blot techniques. Id3 fusion protein was purified through immolbilized Ni(2+); absorption chromatographic column.The purified protein was used to immunize rabbits.The titer and specificity of rabbit antisera were evaluated by immunodouble gel diffusion, ELISA and Western blot techniques. The collected rabbit antisera were purified with polypeptide affinity column. The intracellular distributions of Id3 expression in human human breast cancer cells (MCF7), prostate cancer cells (PC-3M) and lung adenocarcinoma cells (A549) were observed by indirect immunofluorescence assay(IFA). Id3 fusion protein with 6xhis-tag was successfully expressed in E.coli BL 21. The expressed Id3 with the relative molecular mass size of 34 kD was purified specifically through affinity chromatographic column and confirmed by Western blot. The titers of the rabbit anti-Id3 polyclonal antibodies were 1:8 by immunodouble gel diffusion and 1:8 000 by ELISA. Western blot revealed that the prepared antisera reacted specifically with Id3. IFA showed that in A549 , MCF7 and PC-3M cells, Id3 was mainly distributed in nucleoplasm and the Id3 expression level was significantly higher in MCF7 and PC-3M than in A549 cells. The expression and purification of human Id3 protein and preparation of polyclonal antibody against Id3 provide useful tools for the further study of Id3.


Differential transactivation by orphan nuclear receptor NOR1 and its fusion gene product EWS/NOR1: possible involvement of poly(ADP-ribose) polymerase I, PARP-1.
In extraskeletal myxoid chondrosarcoma, a chromosomal translocation creates a gene fusion between EWS and an orphan nuclear receptor, NOR1. The resulting fusion protein EWS/NOR1 has been believed to lead to malignant transformation by functioning as a transactivator for NOR1-target genes. By comparing the gene expression profiles of NOR1- and EWS/NOR1-overexpressing cells, we found that they largely shared up-regulated genes, but no significant correlation was observed with respect to the transactivation levels of each gene. In addition, the proteins associated with NOR1 and EWS/NOR1 were mostly the same in these cells. The results suggest that these proteins differentially transactivate overlapping target genes through a similar transcriptional machinery. To clarify the mechanisms underlying the transcriptional divergence between NOR1 and EWS/NOR1, we searched for alternatively associated proteins, and identified poly(ADP-ribose) polymerase I (PARP-1) as an NOR1-specific binding protein. Consistent with its binding properties, PARP-1 acted as a transcriptional repressor of NOR1, but not EWS/NOR1, in a luciferase reporter assay employing PARP-1(-/-) fibroblasts. Interestingly, suppressive activity of PARP-1 was observed in a DNA response element-specific manner, and in a subtype-specific manner toward the NR4A family (Nur77, Nurr1, and NOR1), suggesting that PARP-1 plays a role in the diversity of transcriptional regulation mediated by the NR4A family in normal cells. Altogether, our findings suggest that NOR1 and EWS/NOR1 regulate overlapping target genes differently by utilizing associated proteins, including PARP-1; and that EWS/NOR1 may acquire oncogenic activities by avoiding (or gaining) transcription factor-specific modulation by the associated proteins.


Valvular strands and cerebral ischemia. Effect of demographics and strand characteristics.
Valvular strands, thin filamentous material attached to the mitral or aortic valve, are seen during transesophageal echocardiography and have been associated with stroke. Little is known about this association in different age, sex, and race-ethnic subgroups and the effect of various strand characteristics on this association. From patients referred for transesophageal echocardiography, 73 patients with recent ischemic stroke (68) or transient ischemic attack (5) were age matched to 73 stroke- and transient ischemic attack-free control subjects. The association between valvular strands and cerebral ischemia was evaluated for the overall group and demographic subgroups. The effect of strand location, length, number, and valve thickness was also determined. An association between cerebral ischemia and valvular strands was observed (odds ratio [OR] = 4.4; 95% confidence interval [CI] = 2.0 to 9.6). The association was found for both men and women and among all three race-ethnic groups. The OR was greater in those who were younger (12.5 [95% CI = 2.4 to 64.5] for age < 60, 4.8 [95% CI = 1.3 to 18.2] for age 60 to 69, and 1.8 [95% CI = 0.5 to 6.4] for age > or = 70 years). Strands on both the mitral (OR = 3.5; 95% CI = 1.5 to 7.9) and aortic (OR = 3.7; 95% CI = 1.1 to 11.9) valve were associated with cerebral ischemia, whereas the number and length of strands were not. The effect of strands was independent of mitral or aortic valve thickness. Valvular strands, whether mitral or aortic, are associated with ischemic stroke, especially among younger persons.


The Role of Feeding Practices as a Determinant of the pufa Index in Children with Early Childhood Caries.
To evaluate the prevalence and severity of untreated dental caries among young children (aged <6 years) with early childhood caries (ECC) using the pufa index and to examine the relationship between caries and feeding practices. A cross-sectional study of children with ECC was undertaken in two stages: a questionnaire survey based on interviews and an intraoral examination using the defs and pufa indices. The overall prevalence of untreated dental caries in our sample of 238 ECC-affected children was 72.3%. The abscess component (code a) was the most common condition. There was an increase in the ECC patterns and the severity of untreated dental caries with increasing age for all teeth groups. ECC and untreated caries lesions for the various teeth groups and study sites varied inversely with current feeding status. The majority of the children had posterior ECC, and the fewest were affected by ulceration (code u) as a consequence of untreated dental caries. Except for the posterior caries pattern, all other caries patterns and pufa components (code u alone, p<0.05) had a significantly higher prevalence among children with nocturnal feeding practices (p<0.001). Binary logistic regression analyses revealed that older age (p<0.05) and nocturnal feeding practices (p<0.001) were significantly associated with pufa scores>0. These data provide valuable evidence of the effect of nocturnal feeding as an important determinant of the increased prevalence of untreated caries lesions among children with ECC.


Microbially Synthesized Repeats of Mussel Foot Protein Display Enhanced Underwater Adhesion.
Mussels strongly adhere to a variety of surfaces by secreting byssal threads that contain mussel foot proteins (Mfps). Recombinant production of Mfps presents an attractive route for preparing advanced adhesive materials. Using synthetic biology strategies, we synthesized Mfp5 together with Mfp5 oligomers containing two or three consecutive, covalently-linked Mfp5 sequences named Mfp5(2) and Mfp5(3). The force and work of adhesion of these proteins were measured underwater with a colloidal probe mounted on an atomic force microscope and the adsorption was measured with a quartz crystal microbalance. We found positive correlations between Mfp5 molecular weight and underwater adhesive properties, including force of adhesion, work of adhesion, protein layer thickness, and recovery distance. DOPA-modified Mfp5(3) displayed a high force of adhesion (201 ± 36 nN μm-1) and a high work of adhesion (68 ± 21 fJ μm-1) for a cure time of 200 s, which are higher than those of previously reported Mfp-mimetic adhesives. Results presented in this study highlight the power of synthetic biology in producing biocompatible and highly adhesive Mfp-based materials.


The role of expectations in patient satisfaction with medical care.
The purpose of this article is to review research about the role of expectations in patients or consumer satisfaction and to identify theories to account for the process by which patient satisfaction is achieved. Studies are examined to suggest how expectations and satisfaction may be defined or measured and to describe the empirical support for an expectation and satisfaction relationship. The application of an expectation and satisfaction model to health care marketing is discussed.


Clinical status of anti-cancer agents derived from marine sources.
The chemical, biological and ecological diversity of the marine ecosystem has contributed immensely in the discovery of extremely potent compounds that have shown potent activities in antitumor, analgesia, antiinflammatory, immunomodulation, allergy, anti-viral etc. The compounds of marine origin are diverse in structural class from simple linear peptides to complex macrocyclic polyethers. The recent advances in the sophisticated instruments for the isolation and characterization of marine natural products and development of high-throughput screening, have substantially increased the rate of discovery of various compounds of biomedical application. Didemnin was the first marine peptide that entered in human clinical trials in US for the treatment of cancer and other compounds such as dolastatin-10, soblidotin, didemnin B, ecteinascidin 743, girolline, aplidine, cryptophycins (also arenastatin A), bryostatin 1, ILX 651, kahalalide F, E7389, discodermolide, ES-285 (spisulosine), HTI-286 (hemiasterlin derivative), squalamine, KRN-7000, vitilevuamide, Laulimalide, Curacin A, diazonamide, peloruside A, eleutherobin, sarcodictyin, thiocoraline, salicylihalimides A, ascididemnin, CGX-1160, CGX-1007dictyodendrins, GTS-21 (aka DMBX), manoalide, IPL-576,092 (aka HMR-4011A) have entered in the clinical trials. This article summarize clinical status and synthetic advances of some of these compounds.


Serial changes in the biophysical profile in patients undergoing cervical ripening with a controlled release PGE2 vaginal pessary.
The purpose of this study was to evaluate the effects of exogenous administration of PGE2 upon the components of the biophysical profile. The study group included 17 nulliparas at > or = 38 weeks gestation, with a Bishop score of < or = 4, requiring induction of labor. A controlled release vaginal pessary containing 10 mg of PGE2, designed to release hormones at approximately 0.8 mg per hour in vitro, was used for 12 hours of cervical ripening. The BPP was performed by the same sonographer at three intervals: prior to pessary insertion, at 6 hours, and 12 hours. None of these patients had membrane rupture or went into spontaneous labor during the ripening process. All patients subsequently required amniotomy and oxytocin. The proportion of patients scoring 2 points for fetal breathing movements decreased from 59% at baseline to 0% at 12 hours, P < 0.0005, and the proportion of patients with fetal body movements decreased from 100% at baseline to 25% at 12 hours, P < 0.0005. However, the other components of the biophysical profile were not affected. The mean maternal plasma PGE2 metabolite concentrations were 235 pg/ml, 475 pg/ml, and 466 pg/ml at 0, 6 and 12 hours, respectively, P < 0.005. In term patients, vaginal administration of the PGE2 pessary was associated with improved Bishop score over 12 hours and significant increases in maternal plasma PGEM levels at 6 hours and 12 hours. These changes were inversely related to fetal breathing and body movements.


Improvement of the Bonding Properties of Mineral Trioxide Aggregate by Elastin-Like Polypeptide Supplementation.
Elastin-like polypeptide (ELP) supplementation was previously reported to enhance the physical properties of mineral trioxide aggregate (MTA). The aim of this study was to investigate the effect of ELP supplementation on the bonding properties of MTA to dentin. Two types of ELPs were synthesized and mixed with MTA in a 0.3 liquid/powder ratio. The push-out bond strength test and interfacial observation with scanning electron microscopy were performed for ELP-supplemented MTA. The porosity of MTA fillings in the cavity was observed with microcomputed tomography. The stickiness, flow rate, and contact angle were additionally measured for potential increased bonding properties. ELP supplementation improved the bond strength of MTA to dentin. MTA supplemented by a specific ELP exhibited a less porous structure, higher stickiness, and higher flow rate. ELPs also decreased the contact angle to dentin. This research data verifies that ELP improves the bonding properties of MTA to a tooth structure. The sticky and highly flowable characteristics of ELP-supplemented MTA may provide intimate contact with dentin and supply a less porous cement structure, which might improve the bonding properties of MTA.


Endoscopic treatment of tracheocele in pediatric patients.
Acquired tracheal pouch known also as tracheocele is a rare air-filled diverticulum of tracheal pars membranacea. This disease may be due to esophageal or tracheal surgery, orotracheal intubation, or increased intralunimal pressure through a weak area of tracheal wall. When symptomatic and medical therapy is insufficient, this disorder must be treated surgically. Several surgical methods ranging from open neck or thoracic surgery to endoscopic managements have been reported. We report the case history of a 7-year-old boy affected by recurrent pneumonia due to tracheal pouch. The patient underwent successful brushing of tracheocele plus fibrin glue application by rigid tracheobroncoscopy. Furthermore, particular emphasis on endoscopic treatments of tracheal diverticulum is also presented.


[Plant potyvirus evolution: the survey of the genetic structure of populations].
The Potyvirus is the largest genus of the largest family of plant RNA viruses, the Potyviridae. The potyviruses infect not only dicotyledonous but also monocotyledonous plants. The potyvirus phylogeny shows that the genus probably originated from a virus of monocotyledonous plants and that it first diverged approximately 7250 years ago in Southwest Eurasia or North Africa. Turnip mosaic virus (TuMV) belongs to the genus Potyvirus and infects a wide range of plant species, most from the family Brassicaceae. TuMV is most studied a potyvirus species for molecular evolution and the genetic structure of populations. The use of computer programs for better understanding of the evolution and the genetic structures of populations of potyviruses and TuMV are illustrated.


Characterization of lignin in situ by photoacoustic spectroscopy.
Photoacoustic spectroscopy is a recently developed nondestructive analytical technique that provides ultraviolet, visible, and infrared absorption spectra from intensely light scattering, solid, and/or optically opaque materials not suitable for conventional spectrophotometric analysis. In wood and other lignocellulosics, the principal ultraviolet absorption bands, in the absence of photosynthetic pigments, arise from the aromatic lignin component of the cell walls. Photoacoustic spectra of extracted lignin fragments (milled wood lignin) and synthetic lignin-like polymers contain a single major absorption band at 280 nanometers with an absorption tail extending beyond 400 nanometers. Photoacoustic spectra of pine, maple, and oak lignin in situ contain a broad primary absorption band at 300 nanometers and a longer wavelength shoulder around 370 nanometers. Wheat lignin in situ, on the other hand, exhibits two principle absorption peaks, at 280 nanometers and 320 nanometers. The presence of absorption bands at wavelengths greater than 300 nanometers in intact lignin could result from (a) interacting, nonconjugated chromophores, or (b) the presence of more highly conjugated structural components formed as the result of oxidation of the polymer. Evidence for the latter comes from the observation that, on the outer surface of senescent, field-dried wheat culms (stems), new absorption bands in the 350 to 400 nanometer region predominate. These new bands are less apparent on the outer surface of presenescent wheat culms and are virtually absent on the inner surface of either senescent or presenescent culms, suggesting that the appearance of longer wavelength absorption bands in senescent wheat is the result of accumulated photochemical modifications of the ligin polymer. These studies also demonstrate photoacoustic spectroscopy to be an important new tool for the investigation of insoluble plant components.


Mechanistic Insights into Rhenium-Catalyzed Regioselective C-Alkenylation of Phenols with Internal Alkynes.
A (μ-aryloxo)rhenium complex was isolated and confirmed as a key precatalyst for rhenium-catalyzed ortho-alkenylation (C-alkenylation) of unprotected phenols with alkynes. The reaction exclusively provided ortho-alkenylphenols; the formation of para or multiply alkenylated phenols and hydrophenoxylation (O-alkenylation) products was not observed. Several mechanistic experiments excluded a classical Friedel-Crafts-type mechanism, leading to the proposed phenolic hydroxyl group assisted electrophilic alkenylation as the most plausible reaction mechanism. For this purpose, the use of rhenium, a metal between the early and late transition metals in the periodic table, was key for the activation of both the soft carbon-carbon triple bond of the alkyne and the hard oxygen atom of the phenol, at the same time. ortho-Selective alkenylation with allenes also provided the corresponding adducts with a substitution pattern different from that obtained by the addition reaction with alkynes.


Chiral Pyridinium Phosphoramide as a Dual Brønsted Acid Catalyst for Enantioselective Diels-Alder Reaction.
Chiral pyridinium phosphoramide 1·HX was designed to be a new class of chiral Brønsted acid catalyst in which both the pyridinium proton and the adjacent imide-like proton activated by the electron-withdrawing pyridinium moiety could work cooperatively as strong dual proton donors. The potential of 1·HX was shown in the enantioselective Diels-Alder reactions of 1-amino dienes with various dienophiles including N-unsubstituted maleimide, which has yet to be successfully used in an asymmetric Diels-Alder reaction.


Regulated exopolysaccharide production in Myxococcus xanthus.
Myxococcus xanthus fibrils are cell surface-associated structures composed of roughly equal amounts of polysaccharide and protein. The level of M. xanthus polysaccharide production under different conditions in the wild type and in several mutants known to have alterations in fibril production was investigated. Wild-type exopolysaccharide increased significantly as cells entered the stationary phase of growth or upon addition of Ca2+ to growing cells, and the polysaccharide-induced cells exhibited an enhanced capacity for cell-cell agglutination. The activity of the key gluconeogenic pathway enzyme phosphoenolpyruvate carboxykinase (Pck) also increased under these conditions. Most fibril-deficient mutants failed to produce polysaccharide in a stationary-phase- or Ca2+-dependent fashion. However, regulation of Pck activity was generally unimpaired in these mutant strains. In an stk mutant, which overproduces fibrils, polysaccharide production and Pck activity were constitutively high under the conditions tested. Polysaccharide production increased in most fibril-deficient strains when an stk mutant allele was present, indicating that these fibril-deficient mutants retained the basic cellular components required for fibril polysaccharide production. In contrast to other divalent cations tested, Sr2+ effectively replaced Ca2+ in stimulating polysaccharide production, and either Ca2+ or Sr2+ was required for fruiting-body formation by wild-type cells. By using transmission electron microscopy of freeze-substituted log-phase wild-type cells, fibril material was observed as a cell surface-associated layer of uniform thickness composed of filaments with an ordered structure.


Cost-effectiveness of a helpline for suicide prevention.
We evaluated the cost-effectiveness and budget impact of a suicide helpline in Belgium, consisting of a telephone- and a chat service. An age- and gender-dependent Markov model with a ten-year time horizon and a one-year cycle length was developed, assuming a societal perspective, to predict cumulative costs and quality-adjusted life-years (QALYs) in the helpline users. The model included six transition states: the initial state (at risk), first attempt, re-attempt, follow-up, suicide and death from other causes. Data on the effect of the helpline and costs associated with model states were obtained from the literature. One-way and probabilistic sensitivity analyses were performed to capture uncertainty. In addition, the budget impact of the helpline was analysed. Over ten years, the telephone- as well as the chat service could avoid about 36% of suicides and attempts in this high-risk population. In males, 0.063 QALYs (95% confidence interval, CI 0.030-0.097) and 0.035 QALYs (95%CI -0.026-0.096) were gained by users of the telephone- and chat service respectively. The corresponding values for females were 0.019 QALYs (95%CI -0.015-0.052) and a QALY-neutral result of -0.005 (95%CI -0.071-0.062). There were net societal savings of respectively €2382 (95%CI 1953-2859) and €2282 (95%CI 1855-2758) in male users; €2171 (95%CI 1735-2664) and €2458 (95%CI 1945-3025) in female users. At the population level, an investment of €218,899 saved €1,452,022 for the public health service (national health insurance), mainly due to the telephone service. The analysis predicted that both means of telemedicine for suicide prevention in Flanders are cost-saving, and have a modest effect on QALYs.


Formulation of technetium-99m-aerosol colloid with improved delivery efficiency for lung ventilation imaging.
Static imaging of lung ventilation in multiple views or with SPECT requires a 99mTc aerosol which does not wash out during imaging. Commercially available 99mTc colloids have suitable physical properties but are inefficiently delivered from jet nebulizers. An in vitro system was used to compare the delivery of various 99mTc radiopharmaceuticals from nebulizer solutions. Delivery of noncolloidal agents (pertechnetate, pentetate, imidodiphosphate, albumin) was much greater than that of colloids. However, addition of phosphate buffer to an in-house 99mTc-tin colloid produced an agent that was delivered as efficiently as the noncolloidal tracers and which shows promise for clinical use.


On using a normal approximation for the noncentral t-distribution in determining upper limits for future sample relative repeatability and reproducibility standard deviations.
Formulas, based on a normal approximation for the noncentral t-distribution, were developed to compute 100p% one-tailed upper limits for future sample relative repeatability and relative reproducibility standard deviations (RSDr,% and RSDR,%) collaboratively obtained under a completely randomized model. The accuracy of the formulas for obtaining a one-tailed upper limit for the future sample RSDr,% was assessed by comparing the computed noncentral t-distribution-based upper limits with the one-tailed upper limits based on a normal approximation for the noncentral t-distribution. The accuracy of the normal approximation formula for obtaining a one-tailed upper limit for a future sample RSDR,% was assessed by comparing the formula-based one-tailed upper limits with those obtained in a Monte Carlo simulation study.


Description of the topographical changes associated to the different stages of the DsbA catalytic cycle.
This paper provides a description of the surface topography of DsbA, the bacterial disulfide-bond forming enzyme, in the different phases of its catalytic cycle. Three representative states, that is, oxidized and reduced protein and a covalent complex mimicking the DsbA-substrate disulfide intermediate, have been investigated by a combination of limited proteolysis experiments and mass spectrometry methodologies. Protease-accessible sites are largely distributed in the oxidized form with a small predominance inside the thioredoxin domain. Proteolysis occurs even in secondary structure elements, revealing a significant mobility of the protein. Many cleavage sites disappear in the reduced form and most of the remaining ones appear with strongly reduced kinetics. The protein within the complex shows an intermediate behavior. This variation of flexibility in DsbA is probably the determining factor for the course of its catalytic cycle. In particular, the great mobility of the oxidized protein might facilitate the accommodation of its various substrates, whereas the increasing rigidity from the complexed to the reduced form could help the release of oxidized products. The formation of the complex between PID peptide and DsbA does not significantly protect the enzyme against proteolysis, reinforcing the results previously obtained by calorimetry concerning the weakness of their interaction. The few cleavage sites observed, however, are in favor of the presence of the peptide in the binding site postulated from crystallographic studies. As for the peptide itself, the proteolytic pattern and the protection effect exerted by DsbA could be explained by a preferential orientation within the binding site.


The ethics of human stem cell research.
The medical and clinical promise of stem cell research is widely heralded, but moral judgments about it collide. This article takes general stock of such judgments and offers one specific resolution. It canvasses a spectrum of value judgments on sources, complicity, adult stem cells, and public and private contexts. It then examines how debates about abortion and stem cell research converge and diverge. Finally, it proposes to extend the principle of "nothing is lost" to current debates. This extension links historical discussions of the ethics of direct killing with unprecedented possibilities that in vitro fertilization procedures yield. A definite normative region to inhabit is located, within a larger range of rival value judgments. The creation of embryos for research purposes only should be resisted, yet research on "excess' embryos is permissible by virtue of an appeal to the "nothing is lost" principle.


Prevalence of hypertension in the rural adult population of Osun State, southwestern Nigeria.
The purpose of this study was to determine the prevalence of hypertension in two rural communities of Osun State, Nigeria. A consenting adult population of the Alajue and Obokun rural communities in southwestern Nigeria that presented for the screening exercise participated in this community-based cross-sectional descriptive study. Two hundred and fifty-nine respondents aged older than 18 years completed a standardized, pretested, structured questionnaire as part of activities celebrating World Kidney Day and World Glaucoma Day in 2011. Anthropometric data and blood pressure were recorded, and the data were analyzed using the Statistical Package for Social Sciences version 17. The mean age of the respondents was 49.7 ± 1.6 years, 100 (38.6%) were males, 84 (32.4%) were farmers, and 111 (42.9%) were traders. The prevalence of hypertension was 13.16% (present in 34 respondents). Seventeen (6.6%) had isolated systolic hypertension, while 11 (4.2%) had isolated diastolic hypertension. Two hundred and thirty-six (91.1%) undertook daily exercise lasting at least 30 minutes and 48 (18.5%) had ever taken antihypertensive drugs on a regular basis. Four respondents (1.6%) claimed a family history of hypertension. The average body mass index (BMI) among respondents was 23.4 ± 4.9 kg/m(2), 51 (19.6%) had a BMI of 25.0-29.9, and 30 (11.5%) had a BMI ≥ 30. A significant association existed between age older than 40 years and having hypertension (P < 0.05), while no relationship existed between age and BMI or between gender and hypertension (P > 0.05). Rates of older age and high BMI were significantly higher among hypertensives than among normotensives. Respondents with BMI ≥ 25 had at least a three times greater likelihood of developing hypertension than those with BMI < 25 (odds ratio 2.9, 95% confidence interval 0.007-0.056, P = 0.011). The prevalence of hypertension is high in this study population and we recommend scaling up primary prevention efforts to reduce this in Nigerian communities.


[Prevalence of metabolic health in Mallorca obese patients].
Aims: to assess the prevalence of metabolic health in Mallorca obese patients. Methods: participants were classified in metabolically healthy obese (MHO) and metabolically non-healthy obese (MNHO). Food, toxic and lifestyle habits, time of obesity evolution, breastfeeding, obesity in childhood and family history of obesity and diabetes mellitus, as well as glycemia, total cholesterol, HDL-cholesterol and triglyceridemia were evaluated in 457 obese patients. Results: prevalence of MHO was 49.2% and that of MNHO was 50.8%. MHO phenotype decreased with age. All patients showed inadequate habits. Consumption of fruits, salads and vegetables, tobacco and physical activity were similar between both groups; 37.4% of patients consumed sugary sweet drinks, and 52.9% consumed alcohol, higher in MNHO (4.3%) than in MHO (0.4%). MNHO showed higher values of BMI, abdominal circumference, fat percentage and visceral fatty index, as well as all metabolically studied outcomes. Conclusions: more than half of assessed obese population showed metabolic complications, but all obese population showed similar inadequate food and lifestyle habits. Increase of age, low educational level, years of obesity evolution, and visceral localization of fat are associated with a metabolically non-healthy status. Criteria to define and typify the metabolic state of obese subjects should be unified.


Corneal epithelium following intraepikeratophakia.
The corneal epithelium in eight patients was surveyed by biomicroscopy and specular microscopy before and after the intraepikeratophakia procedure. Autografting of the epithelium was also examined by electron microscopy. Although the superficial layer of the epithelium was damaged, the wing cell layer remained intact, contributing to the epithelialization of the cornea. Epithelialization of the cornea after surgery was completed in three days in all cases. The epithelium was stained with fluorescein, revealing abnormalities in the superficial layer. This stained superficial epithelium was replaced with new epithelium within seven days, but the new, unstained epithelium exhibited an abnormal pattern when observed under the specular microscope. Abnormally shaped cells, cells with nuclei, and white cells were observed, and normal maturation of the epithelial cells was disrupted. These abnormalities lasted more than eight months in three patients. Patient selection and careful follow-up is advised because of the abnormalities of the epithelium.


Differential contribution of protein phosphatase 1α to cell transformation of different cell types.
Protein phosphorylation plays roles in cell transformation. Numerous protein kinase enzymes actively participate in the formation of various types of cancer by phosphorylating downstream substrates. Aurora‑A is a widely known Serine/Threonine (Ser/Thr) oncogenic kinase, which is upregulated in more than twenty types of human cancer. This enzyme phosphorylates a wide range of substrates. For example, Aurora‑A induces cell transformation by phosphorylating hepatoma upregulated protein (HURP) at four serine residues, which in turn decreases the phosphorylated levels of cell‑growth suppressive Jun N‑terminal kinase (p‑JNK). Various protein phosphatase enzymes are considered tumor suppressors by the dephosphorylation and consequent inactivation of their oncogenic substrates. Protein phosphatase 1α (PP1α), for instance, acts on Aurora‑A by dephosphorylating its substrates. However, the role of PP1α in cancer progression remains ambiguous. PP1α is overexpressed in several cancer tissues, and induces cell apoptosis and differentiation or it inhibits tumor formation in other types of cells. In addition, positive and negative correlations between PP1α expression and lung cancer development have been documented. These observations suggest the differential regulation of PP1α in various cancer tissues, or propose an ambiguous contribution of PP1α to lung cancer development. In order to investigate these contradictory conclusions, it was reported that the chromosomal region covering the PP1α locus was subjected to DNA alterations, such as gain or loss in various human cancer types by a study based on literature search. Upregulation of PP1α was noted in a collection of lung cancer tissues, and was required for the cell transformation of the lung cancer cell line A549. In contrast to this finding, overexpression of ectopic PP1α inhibited cell proliferation in 293T cells. Mechanistic studies revealed that PP1α activated AKT in A549 cells, whereas it further inactivated AKT and disrupted the HURP/JNK signaling cascade in 293T cells. Collectively, the data indicated that PP1α exerted an oncogenic function in lung cancer, while exhibiting various effects on cell transformation in different types of cells via distinct or opposite mechanisms.