DNA damage and methylation induced by glyphosate in human peripheral blood mononuclear cells (in vitro study).
Glyphosate is a very important herbicide that is widely used in the agriculture, and thus the exposure of humans to this substance and its metabolites has been noted. The purpose of this study was to assess DNA damage (determination of single and double strand-breaks by the comet assay) as well as to evaluate DNA methylation (global DNA methylation and methylation of p16 (CDKN2A) and p53 (TP53) promoter regions) in human peripheral blood mononuclear cells (PBMCs) exposed to glyphosate. PBMCs were incubated with the compound studied at concentrations ranging from 0.1 to 10 mM for 24 h. The study has shown that glyphosate induced DNA lesions, which were effectively repaired. However, PBMCs were unable to repair completely DNA damage induced by glyphosate. We also observed a decrease in global DNA methylation level at 0.25 mM of glyphosate. Glyphosate at 0.25 mM and 0.5 mM increased p53 promoter methylation, while it did not induce statistically significant changes in methylation of p16 promoter. To sum up, we have shown for the first time that glyphosate (at high concentrations from 0.5 to 10 mM) may induce DNA damage in leucocytes such as PBMCs and cause DNA methylation in human cells.


Do oestrogens regulate placental growth in the mouse?
Some effects of litter size, ovariectomy and oestradiol (E2) on placental growth were determined. In order to increase variation in litter size, some mice had one oviduct removed and the remainder were unilaterally ovariectomized 3-4 weeks prior to mating. Thus after mating mice became unilaterally pregnant. Growth was examined qualitatively by measuring changes in placental DNA, protein, RNA : DNA ratios and in vivo incorporation of [5-3H]uridine into RNA. Spontaneous placenta hyperplasia due to low litter size occurred to an equal extent in intact mice and mice which were ovariectomized on day 10 of pregnancy and given 1.0 mg progesterone (P) and 10 ng E2 daily. Ovariectomy on day 10 increased the incorporation of [5-3H]uridine into placenta RNA on day 16 in mice with six or more foetuses. When mice were ovariectomized on day 5 and given 1.0 mg P and 10 or 60 ng E2 daily to maintain gestation, rates of placental growth remained similar to the rate in control, intact mice. The results suggest that, unlike the case in the rat, oestrogens do not regulate placental growth in the mouse.


Fear is not critical to classically conditioned analgesia: the effects of periaqueductal gray lesions and administration of chlordiazepoxide.
A high correlation between fear and analgesia classically conditioned to footshock in rats has been reported in the literature. However, it has never been directly tested whether or not fear is in fact causal to the production of conditioned analgesia. We therefore tested whether conditioned analgesia could be elicited in the absence of fear by employing two independent methods of fear suppression. First, areas of the periaqueductal gray (PAG) previously implicated in fear were selectively lesioned. Lesions of the dorsolateral PAG significantly attenuated conditioned analgesia and markedly decreased fear responses. Second, fear was attenuated via administration of chlordiazepoxide (CDP). Rats which had been conditioned while in the presence of CDP showed no reduction in conditioned pain inhibition. These results demonstrate that: (1) fear is not causal to classically conditioned analgesia and (2) the anatomical substrates for fear and conditioned analgesia are distinct but partially overlapping. The fact that fear is not a critical antecedent for classically conditioned analgesia suggests that classical conditioning techniques may be applied clinically to increase the effectiveness of some analgesic manipulations.


Islet oxygen consumption and insulin secretion tightly coupled to calcium derived from L-type calcium channels but not from the endoplasmic reticulum.
The aim of the study was to test whether the source of intracellular calcium (Ca2+) is a determinant of beta cell function. We hypothesized that elevations in cytosolic Ca2+ caused by the release of Ca2+ from the endoplasmic reticulum (ER) have little physiologic impact on oxygen consumption and insulin secretion. Ca2+ release from the ER was induced in isolated rat islets by acetylcholine and response of oxygen consumption rate (OCR), NAD(P)H, cytosolic Ca2+, and insulin secretory rate (ISR) were measured. Glucose increased all four parameters, and thereafter acetylcholine further increased cytosolic Ca2+, OCR, and ISR. To assess the contribution of Ca2+ release from the ER in mediating the effects of acetylcholine, ER Ca2+ stores were first emptied by inhibiting the sarcoendoplasmic reticulum Ca2+-ATPase, which subsequently reduced the effect of acetylcholine on cytosolic Ca2+ but not its effects on OCR or ISR. As predicted, OCR and ISR were acutely sensitive to changes in L-type Ca2+ channel activity; nimodipine completely inhibited glucose-stimulated ISR and suppressed OCR by 36%, despite only inhibiting cytosolic Ca2+ by 46%. Moreover, in the presence of nimodipine and high glucose, acetylcholine still elevated cytosolic Ca2+ levels above those observed in the presence of high glucose alone but did not significantly stimulate ISR. In conclusion, Ca2+ flux through L-type Ca2+ channels was tightly coupled to changes in OCR and ISR. In contrast, the results obtained support the notion that Ca2+ release from the ER has little or no access to the intracellular machinery that regulates OCR and ISR.


A novel extracorporeal CO(2) removal system: results of a pilot study of hypercapnic respiratory failure in patients with COPD.
Hypercapnic respiratory failure in patients with COPD frequently requires mechanical ventilatory support. Extracorporeal CO2 removal (ECCO2R) techniques have not been systematically evaluated in these patients. This is a pilot study of a novel ECCO2R device that utilizes a single venous catheter with high CO2 removal rates at low blood flows. Twenty hypercapnic patients with COPD received ECCO2R. Group 1 (n = 7) consisted of patients receiving noninvasive ventilation with a high likelihood of requiring invasive ventilation, group 2 (n = 2) consisted of patients who could not be weaned from noninvasive ventilation, and group 3 (n = 11) consisted of patients on invasive ventilation who had failed attempts to wean. The device was well tolerated, with complications and rates similar to those seen with central venous catheterization. Blood flow through the system was 430.5 ± 73.7 mL/min, and ECCO2R was 82.5 ± 15.6 mL/min and did not change significantly with time. Invasive ventilation was avoided in all patients in group 1 and both patients in group 2 were weaned; PaCO2 decreased significantly (P < .003) with application of the device from 78.9 ± 16.8 mm Hg to 65.9 ± 11.5 mm Hg. In group 3, three patients were weaned, while the level of invasive ventilatory support was reduced in three patients. One patient in group 3 died due to a retroperitoneal bleed following catheterization. This single-catheter, low-flow ECCO2R system provided clinically useful levels of CO2 removal in these patients with COPD. The system appears to be a potentially valuable additional modality for the treatment of hypercapnic respiratory failure.


Ionic-radius-driven selection of the main-group-metal cage for intermetalloid clusters [Ln@Pbx Bi14-x](q-) and [Ln@Pby Bi13-y](q-) (x/q=7/4, 6/3; y/q=4/4, 3/3).
Reactions of the binary, pseudo-homoatomic Zintl anion (Pb2 Bi2 )(2-) with Ln(C5 Me4 H)3 (Ln=La, Ce, Nd, Gd, Sm, Tb) in the presence of [2.2.2]crypt in ethane-1,2-diamine/toluene yielded ten [K([2.2.2]crypt)](+) salts of lanthanide-doped semimetal clusters with 13 or 14 surface atoms. Single-crystal X-ray diffraction and energy-dispersive X-ray spectroscopy indicated the presence of the anions [Ln@Pb6 Bi8 ](3-), [Ln@Pb3 Bi10 ](3-), [Ln@Pb7 Bi7 ](4-), or [Ln@Pb4 Bi9 ](4-) in single or double salts; the latter showed various ratios of the components in the solid state. The anions are the first ternary intermetalloid clusters comprising only elements of the sixth period of the periodic table, namely, Pb, Bi and lanthanides. This study, which was complemented by ESI mass spectrometry and (139) La NMR spectroscopy in solution, rationalizes a continuous development of the ratio of 13:14-atom cages with the ionic radius of the embedded Ln(3+) ion, which seems to select the most suitable cage type. Quantum chemical investigations helped to analyze this situation in more detail and to explain the observed subtle influence of the atomic radii. Magnetic measurements confirmed that the embedded Ln(3+) ions keep their expected paramagnetic or diamagnetic nature.


Mass spectrometry imaging and LC/MS reveal decreased cerebellar phosphoinositides in Niemann-Pick type C1-null mice.
Niemann-Pick disease type C1 (NPC1) is a lipid storage disorder in which cholesterol and glycosphingolipids accumulate in late endosomal/lysosomal compartments because of mutations in the NPC1 gene. A hallmark of NPC1 is progressive neurodegeneration of the cerebellum as well as visceral organ damage; however, the mechanisms driving this disease pathology are not fully understood. Phosphoinositides are phospholipids that play distinct roles in signal transduction and vesicle trafficking. Here, we utilized a consensus spectra analysis of MS imaging data sets and orthogonal LC/MS analyses to evaluate the spatial distribution of phosphoinositides and quantify them in cerebellar tissue from Npc1-null mice. Our results suggest significant depletion of multiple phosphoinositide species, including PI, PIP, and PIP2, in the cerebellum of the Npc1-null mice in both whole-tissue lysates and myelin-enriched fractions. Additionally, we observed altered levels of the regulatory enzyme phosphatidylinositol 4-kinase type 2α in Npc1-null mice. In contrast, the levels of related kinases, phosphatases, and transfer proteins were unaltered in the Npc1-null mouse model, as observed by Western blot analysis. Our discovery of phosphoinositide lipid biomarkers for NPC1 opens new perspectives on the pathophysiology underlying this fatal neurodegenerative disease.


Pressure ulcer prevention is everyone's business: the PUPS project.
Prevention of pressure ulcers is one of the greatest healthcare challenges in terms of reducing patient harm. The literature shows that although numerous reports and policy documents have been published, pressure ulcer prevention remains an ongoing challenge. A number of innovations have been published offering practising nurses and managers ideas for raising awareness of skin care and preventing pressure ulcers. The majority of these have focused on patients in hospital settings with very little in the literature related to care-home and community initiatives. This article reports on an innovative approach to education for pressure ulcer prevention through collaboration between patients, carers and health and social care professionals.


Posterodorsal septal lesions impair performance on both shift and stay working memory tasks.
Two groups of rats were trained preoperatively on either a shift or a stay problem in a T-maze. Training trials consisted of two runs, an "information run" in which a subject was forced to go down one of the two arms of the T-maze, followed immediately by a "choice run" in which the subject could choose either arm. In the shift condition, rats were rewarded with wet mash only for choosing the arm opposite the one they entered on the information run. In the stay condition, rats were rewarded for entering the arm that was entered on the information run. In both conditions, rats ultimately learned to perform with median accuracy of 100%, but the shift group reached this level of performance after fewer trials than the stay group. In a subsequent phase, the delay between information runs and choice runs was increased from 0 to 30, 60, 90, 210 and then decreased back to 0 s. Choice accuracy in both groups declined as the delay increased and returned to 100% at the 0-s delay. Half of the subjects in each condition then received either lesions of the posterodorsal septum-aimed at disconnecting the septum and hippocampus-or control surgery. Postoperative retention deficits resulted from posterodorsal septal lesions in both shift and stay conditions. There was some recovery of performance but no indication of "savings" during postoperative training. These results indicate that deficits in maze performance by rats with septo-hippocampal damage are not restricted to tasks that require alternation of spatial locations. This finding falsifies the notion that maze deficits reflect a spontaneous alternation deficit or changed "spatial strategy," but it supports the hypothesis of a working memory deficit in these animals.


Expression of PTCH gene in ameloblastoma and odontogenic keratocyst: A comparative study.
This is a cross-sectional comparative study, aimed to quantify the expression of patched (PTCH) gene in ameloblastoma, odontogenic keratocyst (OKC) and also the comparison of both the expressions. Genomic deoxyribonucleic acid (DNA) was extracted and quantified, and the expression of the PTCH gene was done in 17 cases of ameloblastoma and 17 cases of OKC by quantitative real-time polymerase chain reaction (RT-qPCR). It was observed that there was an overexpression of the PTCH gene in both ameloblastoma and OKC with a good mean cycle threshold (CT) value of 32.71 ± 2.432 and 34.69 ± 1.875, respectively. When comparing the PTCH expression between the two, ameloblastoma showed higher expression than the OKC and the difference is statistically significant with P value of 0.025. Our findings suggest that there is overexpression of PTCH in ameloblastoma and OKC, but it is highly expressed in ameloblastoma when compared to OKC. Overexpression of PTCH may constitute the activation of the Sonic Hedgehog pathway and may suggest the mechanism for the development of ameloblastoma and OKC. Hence it can be used as a valuable marker for early diagnosis and in the identification of therapeutic targets.


Kin-Driver: a database of driver mutations in protein kinases.
Somatic mutations in protein kinases (PKs) are frequent driver events in many human tumors, while germ-line mutations are associated with hereditary diseases. Here we present Kin-driver, the first database that compiles driver mutations in PKs with experimental evidence demonstrating their functional role. Kin-driver is a manual expert-curated database that pays special attention to activating mutations (AMs) and can serve as a validation set to develop new generation tools focused on the prediction of gain-of-function driver mutations. It also offers an easy and intuitive environment to facilitate the visualization and analysis of mutations in PKs. Because all mutations are mapped onto a multiple sequence alignment, analogue positions between kinases can be identified and tentative new mutations can be proposed for studying by transferring annotation. Finally, our database can also be of use to clinical and translational laboratories, helping them to identify uncommon AMs that can correlate with response to new antitumor drugs. The website was developed using PHP and JavaScript, which are supported by all major browsers; the database was built using MySQL server. Kin-driver is available at: http://kin-driver.leloir.org.ar/


Large globin RNA molecules and their processing.
RNA containing beta-globin message sequences larger than 2000 nucleotides could be detected in nuclei of murine erythroid cells using cloned beta-globin cDNA. Under steady-state conditions, when nuclear RNA was separated on denaturing agarose gels and covalently bound to diazobenzyloxymethyl-paper, a 4200-nucleotide and a approximately equal to 3500-nucleotide band could be seen. The presence of these large molecules could also be visualized under the electron microscope after hybridization to a beta-globin genomic DNA fragment. We suggest that these molecules are precursors to mature mRNAs. In addition to these large molecules, a series of molecules smaller than 2000 nucleotides were seen. These are postulated to be processing intermediates in the maturation of beta-globin mRNA.


Comparative metabolism of four limonoids in human liver microsomes using ultra-high-performance liquid chromatography coupled with high-resolution LTQ-Orbitrap mass spectrometry.
Limonoids, characterized by a triterpenoid skeleton with a furan ring, are unique secondary metabolites widely distributed in the families of Rutaceae, particularly in Citrus species and Meliaceae. Studies on health benefits have demonstrated that limonoids have a range of biological activities. Dietary intake of citrus limonoids may provide a protective effect against the onset of various cancers and other xenobiotic related diseases. However, few studies about the metabolic profiles of limonoids have been carried out. The objectives of this study were to investigate the metabolic profiles of four limonoids (limonin, obacunone, nominin and gedunin) in human liver microsomes (HLMs) using ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC/HRMS) and to identify the cytochrome P450 (CYP) enzymes involved in the formation of their metabolites by recombinant human CYP enzymes. Based on the accurate HR-MS/MS spectra and the proposed MS/MS fragmentation pathways, four metabolites of limonin (M1-1, M1-2, M1-3 and M1-4), eight metabolites ofobacunone (M2-1, M2-2, M2-3, M2-4, M2-5, M2-6, M2-7 and M2-8), six metabolites of nominin (M3-1, M3-2, M3-3, M3-4, M3-5 and M3-6) and three metabolites of gedunin (M4-1, M4-2 and M4-3) in HLMs were tentatively identified and the involved CYPs were investigated. The results demonstrated that reduction at C-7 and C-16, hydroxylation and reaction of glycine with reduction limonoids were the major metabolic pathways of limonoids in HLMs. Among them, glycination with reduction was the unique metabolic process of limonoids observed for the first time. CYP2D6 and CYP3A4 played an important role in the isomerization and glycination of limonoids in HLMs, whereas other CYP isoforms were considerably less active. The results might help to understand the metabolic process of limonoids in vitro such as the unidentified metabolites of limonin glucoside observed in the medium of microbes and the biotransformation of limonin in juices. Moreover, it would be beneficial for us to further study the pharmacokinetic behavior of limonoids in vivo systematically.


Genome-wide identification and expression analysis of calcium-dependent protein kinase and its closely related kinase genes in Capsicum annuum.
As Ca2+ sensors and effectors, calcium-dependent protein kinases (CDPKs) play important roles in plant growth, development, and response to environmental cues. However, no CDPKs have been characterized in Capsicum annuum thus far. Herein, a genome wide comprehensive analysis of genes encoding CDPKs and CDPK-related protein kinases (CRKs) was performed in pepper, a total of 31 CDPK genes and five closely related kinase genes were identified, which were phylogenetically divided into four distinct subfamilies and unevenly distributed across nine chromosomes. Conserved sequence and exon-intron structures were found to be shared by pepper CDPKs within the same subfamily, and the expansion of the CDPK family in pepper was found to be due to segmental duplication events. Five CDPKs in the C. annuum variety CM334 were found to be mutated in the Chiltepin variety, and one CDPK present in CM334 was lost in Chiltepin. The majority of CDPK and CRK genes were expressed in different pepper tissues and developmental stages, and 10, 12, and 8 CDPK genes were transcriptionally modified by salt, heat, and Ralstonia solanacearum stresses, respectively. Furthermore, these genes were found to respond specifically to one stress as well as respond synergistically to two stresses or three stresses, suggesting that these CDPK genes might be involved in the specific or synergistic response of pepper to salt, heat, and R. solanacearum. Our results lay the foundation for future functional characterization of pepper CDPK and its closely related gene families.


Caller satisfaction with the Cannabis Information and Helpline.
We assessed caller satisfaction with an illicit drug helpline in Australia (the Cannabis Information and Helpline, CIH). A 10-min telephone interview was conducted with 200 volunteers who called the service during 2009 (121 called regarding themselves and 79 called regarding another person's cannabis use). Callers were a mean of 43 years of age and typically female (59%). Callers showed high levels of satisfaction with the service: the mean CSQ-8 score was 28.2 (SD = 4.3). Participants who found the service easy to access and those who felt their needs were met reported the highest satisfaction. A linear regression analysis was conducted to identify the significant predictors of participants' satisfaction with the CIH. The ease with which the participant was able to get through to a counsellor (standardized beta = 2.37, P < 0.02) and whether the participant felt that all their needs were met (standardized beta = -4.26, P < 0.001) were the only significant predictors of total satisfaction with the call. Despite the recognition that telephone services are possibly the easiest health-care service to access, ensuring consistent availability and accessibility remains paramount although not easy.


Selective Manipulation of Biomolecules with Insulator-Based Dielectrophoretic Tweezers.
Insulator-based dielectrophoretic (iDEP) trapping, separating, and concentrating nanoscale objects is carried out using a non-metal, unbiased, mobile tip acing as a tweezers. The spatial control and manipulation of fluorescently-labeled polystyrene particles and DNA were performed to demonstrate the feasibility of the iDEP tweezers. Frequency-dependent iDEP tweezers' strength and polarity were quantitatively determined using two theoretical approaches to DNA, which resulted in a factor of 2 ~ 40 differences between them. In either approach, the strength of iDEP was at least 4-order of magnitude stronger than the thermal force, indicating iDEP was a dominant force for trapping, holding, and separating DNA. The trapping strength and volume of the iDEP tweezers were also determined, which further supports direct capture and manipulation of DNA at the tip end.


Assessing the Relationship of Mammographic Breast Density and Proliferative Breast Disease.
Increased breast density and a history of benign breast biopsy are both considered risk factors for developing breast cancer. Understanding the specifics of these risk factors and their relationship to each other can lead to a better understanding of a patient's propensity for breast cancer development and improved surveillance strategies. We included 245 women who underwent a benign breast biopsy without atypia between October 2011 and June 2013. Biopsies were performed for suspicious calcifications as well as masses and architectural distortion. Lesions biopsied were divided into two groups: calcified and noncalcified lesions. The patient's breast density was assessed on most recent mammogram and was classified using the American College of Radiology BI-RADS density categories. Based on histologic diagnosis, each case was classified as proliferative or nonproliferative breast disease. The median age of the cohort (n = 245) was 55 years (range, 40-84 years). There were 162 (66%) postmenopausal women in the study. A core biopsy was performed for calcifications in 33.5% cases and for noncalcified lesions in 58% cases. In patients with dense breast tissue, an underlying proliferative histology was found significantly more frequently with calcifications (66.7%) as opposed to noncalcified lesions (35.9%) (RR = 2.3 (1.3-4.0); χ(2) = 8.7; p = 0.003). In nondense breast patients, there was no significant difference (RR = 1.1 (0.7-1.8); χ(2) = 0.1; p = 0.738). In the postmenopausal group, women with dense breasts had proliferative histology significantly more frequently than women with nondense breasts (55.3% versus 38.3%; p < 0.05), regardless of the underlying lesion type. Postmenopausal women with dense breasts who underwent a breast biopsy with benign histology had a significantly higher likelihood of having proliferative breast disease, regardless of underlying lesion type. Women with dense breasts also showed proliferative histology significantly more often for calcifications as opposed to noncalcified lesions.


Biotransformation of agallochaexcoerin A by Aspergillus flavus.
Agallochaexcoerin A (1), a seco-manoyl oxide diterpenoid was metabolised by pathogenic fungus, Aspergillus flavus, in growth media to yield a new metabolite, termed agallochaexcoerin G (2). It was confirmed by using IR, UV, (1)H NMR and HR-ESI-MS techniques. This microbial bioconversion was achieved by unusual dehydration at C-4 position.


Educational card games for understanding gastrointestinal physiology.
In the last few years, there has been an emphasis on the development of creative educational materials that supplement the traditional lecture format. The new materials should engage students in interactive learning and enhance critical thinking, small group discussion, and problem-solving skills. To help students understand and apply basic science concepts in a challenging, interactive format, we developed two card games. Although the principles of the games can be adapted to many scientific disciplines, these specific games provide a unique opportunity to integrate, analyze, and interpret basic concepts of gastrointestinal (GI) physiology. Go GI and GI Rummy were developed to assist students in the understanding of GI physiology and were designed to function as a tool for learning lecture material. Both games were evaluated by medical, graduate, and high school students. Student evaluation of the educational material showed that the games were successful in promoting the learning of GI physiology and engaging students in the discussion of GI concepts. Through this new approach, the students' level of understanding and ability to apply and synthesize materials were enhanced.


Biosynthesis and intracellular transport of enzymes of peroxisomal beta-oxidation.
Three peroxisomal enzymes of beta-oxidation from rat liver were synthesized in a cell-free protein-synthesizing system derived from a lysate of rabbit reticulocytes. The in vitro products of acyl-CoA oxidase (EC 220.127.116.11) and a bifunctional protein containing enoyl-CoA hydratase (EC 18.104.22.168) and 3-hydroxyacyl-CoA dehydrogenase (EC 188.8.131.52) activities were apparently the same in size and charge as the subunit of the respective mature enzymes; that of 3-ketoacyl-CoA thiolase (EC 126.96.36.199) was about 3,000 Da larger and more basic than its mature subunit. The free polysome fraction of rat liver was 3.1-5.7 times more active than the membrane-bound polysome fraction in the synthesis of the three peroxisomal enzymes; these values were similar to those for cytosolic enzymes and differed from that for serum albumin. In isolated rat hepatocytes, radiolabeled acyl-CoA oxidase and bifunctional protein increased with time with no appreciable change in the subunit size. On the other hand, the labeled putative precursor of 3-ketoacyl-CoA thiolase, as well as the mature form of the enzyme, was detected in the hepatocytes. The radioactivity of the putative precursor reached a plateau in 30 min; that of the mature subunit appeared after a lag time of about 5 min and increased with time up to 90 min. In pulse-chase experiments, the putative precursor disappeared with an apparent half-life of several minutes. When the hepatocytes were fractionated into the cytosolic and the particulate fractions, one half of labeled acyl-CoA oxidase and 60% of the bifunctional protein were recovered in the cytosolic fraction after 10 min of labeling, whereas 70-80% of the labeled enzymes were recovered in the particulate fraction after 40-60 min of labeling. These results indicate that the three enzymes of peroxisomal beta-oxidation are synthesized on free polysomes, released into the cytosol, and then transported into peroxisomes. Our findings also indicate that 3-ketoacyl-CoA thiolase undergoes proteolytic processing during maturation. The temporal sequence of the proteolytic cleavage and intracellular transport of the thiolase remains to be determined.


Insomnia in Cancer--Associations and Implications.
Insomnia is a common condition in cancer patients. In spite of the high prevalence its associations have not been well studied. Existing data suggests that insomnia is related to depression and pain. However, the impact of ongoing chemotherapy on sleep is not investigated. To study the relationship between insomnia and chemotherapy after analysing confounding variables. Consecutive patients who visited New England Oncology Clinic in Tamworth were recruited. Insomnia was assessed with the Bergen insomnia scale. The Montgomery Asberg Depression rating scale was used to measure depression. Pain was assessed with the Brief Pain inventory. Chronic medical conditions, type of cancer, side effects to chemotherapy, role of steroids and other drugs were studied as confounders. A total of 56 patients participated in the study. Age ranged from 33 to 83 years (mean: 63.6, SD=10.97). There were 29 men and 27 women. 42 patients received at least one form of chemotherapy and 15 were receiving radiotherapy at the time of assessment. Mean insomnia score was significantly higher in those receiving chemotherapy than in those without chemotherapy (8.92 vs 17.2, two tailed p=0.005, 95% CI=2.63-13.71). There was no significant variation in insomnia scores in terms of chronic medical condition, type of cancer, psychiatric history, use of steroids or adverse effects of chemotherapy. However, total insomnia score was correlated with depression rating score (Pearson correlation, r=0.39, p=0.003) and magnitude of pain (r=0.37, p=0.006). On regression analysis only pain was found to be predictive of insomnia. Insomnia in patients with cancer is found to be associated with concurrent chemotherapy and correlated with degree of depression and pain. Identifying factors related to insomnia in cancer population has implications in its management and patient education.


Disruption of the Francisella noatunensis subsp. orientalis pdpA Gene Results in Virulence Attenuation and Protection in Zebrafish.
Several Francisella spp., including Francisella noatunensis, are regarded as important emerging pathogens of wild and farmed fish. However, very few studies have investigated the virulence factors that allow these bacterial species to be pathogenic in fish. The Francisella pathogenicity island (FPI) is a well-described, gene-dense region encoding major virulence factors for the genus Francisella. pdpA is a member of the pathogenicity-determining protein genes carried by the FPI that are implicated in the ability of the mammalian pathogen Francisella tularensis to escape and replicate in infected host cells. Using a sacB suicide approach, we generated pdpA knockouts to address the role of PdpA as a virulence factor for F. noatunensis. Because polarity can be an issue in gene-dense regions, we generated two different marker-based mutants in opposing polarity (the F. noatunensis subsp. orientalis ΔpdpA1 and ΔpdpA2 strains). Both mutants were attenuated (P < 0.0001) in zebrafish challenges and displayed impaired intracellular replication (P < 0.05) and cytotoxicity (P < 0.05), all of which could be restored to wild-type (WT) levels by complementation for the ΔpdpA1 mutant. Importantly, differences were found for bacterial burden and induction of acute-phase and proinflammatory genes for the F. noatunensis subsp. orientalis ΔpdpA1 and ΔpdpA2 mutants compared to the WT during acute infection. In addition, neither mutant resulted in significant histopathological changes. Finally, immunization with the F. noatunensis subsp. orientalis ΔpdpA1 mutant led to protection (P < 0.012) against an acute 40% lethal dose (LD40) challenge with WT F. noatunensis in the zebrafish model of infection. Taken together, the results from this study further demonstrate physiological similarities within the genus Francisella relative to their phylogenetic relationships and the utility of zebrafish for addressing virulence factors for the genus.


Predictive value of serum ALT and T-cell receptor beta variable chain for HBeAg seroconversion in chronic hepatitis B patients during tenofovir treatment.
Effective antiviral therapy plays a key role in slowing the progression of chronic hepatitis B (CHB). Identification of serum indices, including hepatitis B e antigen (HBeAg) expression and seroconversion, will facilitate evaluation of the efficacy of antiviral therapy in HBeAg-positive CHB patients. The biochemical, serological, virological parameters, and the frequency of circulating CD4CD25 regulatory T cell (Treg) in 32 patients were measured at baseline and every 12 weeks during 96 weeks of tenofovir disoproxil fumarate (TDF) treatment. The relationship between the hepatitis B virus (HBV) deoxyribonucleic acid (DNA) and Treg and alanine aminotransferase (ALT) levels was analyzed, respectively. The molecular profiles of T-cell receptor beta variable chain (TRBV) were determined using gene melting spectral pattern. For the seroconverted 12 patients, ALT declined to normal levels by week 24 and remained at this level in subsequent treatment; moreover, the predictive cutoff value of ALT for HBeAg seroconversion (SC) was 41.5 U/L at week 24. The positive correlation between HBV DNA and Treg and ALT was significant in SC patients, but not in non-SC patients. Six TRBV families (BV3, BV11, BV12, BV14, BV20, and BV24) were predominantly expressed in SC patients at baseline. The decline of ALT could be used to predict HBeAg seroconversion for CHB patients during TDF treatment. In addition, the profile of Tregs and TRBVs may be associated with HBeAg seroconversion and could also be a potential indicator for predicting HBeAg SC and treatment outcome for CHB patients.


Prenatal and perinatal risk factors and testicular cancer: a hospital-based case-control study.
Some evidence exists to support the hypothesis that elevated levels of circulating maternal estrogens during early pregnancy may increase risk of testicular germ cell cancer. However, the results from studies evaluating maternal factors have been mixed. We evaluated maternal factors, particularly those associated with excess estrogen levels, as risk factors for testicular cancer. We conducted a hospital-based case-control study at The University of Texas M. D. Anderson Cancer Center in Houston, Texas of 144 testicular cancer patients diagnosed between 1990 and 1996 and 86 friend controls matched to cases on age, race, and state of residence. Risk factor data about the mother, the son, and the pregnancy were obtained from the mothers by telephone interviews and from the sons by self-administered questionnaires. Extreme nausea during the first trimester of pregnancy was associated with an elevated risk of testicular cancer [odds ratio (OR) = 2.0; 95% confidence interval (CI) = 1.0-3.9]. Adjustment for potential confounders slightly lowered this risk (OR = 1.8; 95% CI = 0.9-3.8). Risks were modestly increased for other factors that are proxy measures for maternal estrogens, including preterm delivery (OR = 2.2; 95% CI = 0.4-12.9), birth weight <3000 g (OR = 2.4: 95% CI = 0.7-8.1), and birth weight >4000 g (OR = 1.7; 95% CI = 0.9-3.2), albeit nonsignificantly so. Our finding that severe nausea was associated with increased testicular cancer risk adds evidence to support the in utero estrogen exposure hypothesis because nausea early in pregnancy is related to rising levels of circulating estrogens. For other factors, which are less direct measures of maternal estrogens, the modest associations found indicate a suggestive pattern in support of the excess estrogen hypothesis.


Violent video games affecting our children.
Exposure to media violence is associated with increased aggression and its sequelae. Unfortunately, the majority of entertainment video games contain violence. Moreover, children of both genders prefer games with violent content. As there is no compulsory legislative standards to limit the type and amount of violence in video games, concerned adults must assume an oversight role.


Effects of different fat sources (saturated and unsaturated) on reproductive performance and biological indices of ewes during flushing period.
This study was conducted to evaluate the effect of feeding different sources of fat during flushing period on the reproductive performance, lambing percent, and twin numbers of Afshari ewes. A total of 84 ewes (mean weight 48 ± 3 kg; age: 3-4 years) were divided into seven groups of 12 animals and received flushing-specific rations for 5 weeks. The control group just received a basic ration (non-flushing). Lipid sources were calcium salt of palm oil (CaP), pure palm oil (PO), calcium salt of flaxseed (CaFL), calcium salt of sunflower oil (CaSF), flaxseed oil (FLO), and sunflower oil (SFO). Estrous cycles were synchronized in all ewes using 14-day CIDRs followed by 400-IU PMSG injection at the time of CIDR removal. Fertility and lambing percent were higher in ewes fed with diets containing calcium salts of flaxseed and SFO, as compared to other treatments. Total number of lambs in flushing treatments was significantly higher than that in the control group (p < 0.01). Serum cholesterol and progesterone levels were significantly greater in omega-3 (CaFL) and omega-6 (CaSF) treatments relative to other treatments (p < 0.01). It was concluded that supplementing the flushing diet with calcium salts of fatty acids (CSFA) increased blood metabolites and hormones related to reproductive performance; and improved fertility, lambing rate and ewes of CaFL treatment have the highest number of lambs (16 lambs) between different groups. Using saturated and unsaturated fatty acids, especially in their CSFA forms during flushing period, could improve the reproduction problems induced by progesterone deficiency, lack of durability of the fetus due to hormonal instability, and abortion control factors.


Therapeutic Role of MiR-140-5p for the Treatment of Non-small Cell Lung Cancer.
Lung cancer is the second most common cancer in both men and women, after prostate and breast cancer, respectively. There are two main types of primary lung cancer, non-small cell lung cancer (NSCLC), which accounts for approximately 85-90% of all lung cancer cases, and small cell lung cancer (SCLC), which accounts for the other 10-15% of lung cancers. MiRNAs are small molecules that post-transcriptionally regulate many genes and contribute to many disease aetiologies, including tumours. In lung cancer, the down-regulation of miR-140-5p leads to disease progression. In this study a miR-140-5p-only treatment and miR-140-5p combined with other chemotherapeutics have been studied in vitro. When transfected into NSCLC, the overexpression of miR-140-5p reduced the migration and invasion properties of malignant cells and, also improved their adhesion onto the artificial extracellular matrix. When miRNA-140-5p replacement treatment was combined with other drugs commonly used in clinical practice, such as gefinitib, DMH1 and cisplatin, it enhanced their efficacy by reducing the migration and invasion ability of cancer cells, thus suggesting that it acts synergistically with known compounds for the treatment of NSCLC. Additionally, some endothelial mesenchymal transition (EMT) markers appeared to be regulated by miR-140-5p. Novel direct targets of miR-140-5p have not been investigated in this study, but our results indicate the involvement of miR-140-5p in lung cancer invasion. The preliminary data from this study imply that when miR-140-5p levels are restored; maybe synergistically support current therapies for NSCLC though further validation, especially in vivo is required.


Effect of dilauryl succinate on reproduction of the cock and hen and preventive effect of vitamin E.
Fertility of White Leghorn cocks fed a diet containing 12% of dilauryl succinate (LS) for 16 weeks was significantly lower than that of cocks fed the control diet. Little difference in fertility and hatchability of fertile eggs was observed between White Leghorn hens fed the control or LS diets, but the percentage of chicks at hatch, which showed signs of encephalomalacia with lesions on the cerebellum, and the percentage of the hens and offspring having fragile erythrocytes were much higher when LS was fed. Total tocopherol in the plasma of the cocks and hens fed LS and in the eggs laid by the hens fed LS was lower than that in birds fed the control diet, though that of the hens was within the range of plasma tocopherol of normal hens. Offspring of the cocks and hens fed LS died from encephalomalacia earlier than those fed the control diet. These observations with LS feeding were all prevented by oral administration of dl-alpha-tocopheryl acetate. The responses of the cocks and hens to LS were compared with those to linoleic acid in the literature. The direct effect of LS, or more likely of monolauryl succinate, independent of peroxides from unsaturated fatty acids was discussed.


Chemical carcinogens: implications for cancer treatment.
Carcinogen exposure has been associated with enhanced cancer immunogenicity that is often attributed to neoantigen generation. However, the broader, neoantigen-independent impact of carcinogens on immune responses to cancer cells remains underexplored. In this issue of the JCI, Huang et al. uncover a mechanism wherein carcinogen-treated cancer cells exhibit an inability to establish an immunosuppressive tumor microenvironment (TME) due to reduced M-CSF expression. Intriguingly, the so-called carcinogen-induced tumor-associated macrophages (TAMs) within this TME exhibited anti-tumor properties instead of the conventional immunosuppressive phenotype. This phenomenon extended to human lung cancers, as evidenced by TAM reprogramming in smokers versus nonsmokers. This study substantially advances our understanding of carcinogen-mediated effects on cancer immunogenicity, potentially redirecting approaches to cancer immunotherapy.


[Determination of prostaglandinum E1 by fluorescence extinction method].
In this paper, we have studied the fluorescence extinction effect of methyl alcohol by prostaglandinum E1 (PG E1) and found the linear relation between the fluorescence extinction intensity and prostaglandinum E1 concentration. Its linear equation is deltaF = 0.222c + 9.33 x 10(-2)(deltaF: fluorescence extinction value; c: PG E1 concentration microg/mL), correlation coeficient r = 0.99930. The results derived from fluorescence extinction method correspond to the results from violet spectrophotometric method.


CORTICOSTEROID TREATMENT IN THE SETTING OF DECOMPENSATED LIVER CIRRHOSIS WITH RELATIVE ADRENAL INSUFFICIENCY: A CASE REPORT AND A BRIEF REVIEW OF THE LITERATURE.
Relative adrenal insufficiency (RAI) is the term used to describe inadequate production or action of glucocorticoids with respect to the severity of the illness. RAI is frequently found in critically ill patients particularly with septic complications and it is also present in both critically ill and stable patients with liver cirrhosis. In the following study a case report of a patient with decompensated cirrhosis and RAI is presented followed by a brief review of the literature. A 65-year-old male with liver cirrhosis of alcoholic etiology was admitted to hospital with bilateral leg edema, ascites, and marked weakness. At admission, his blood pressure was 82/52 mmHg and he had sinus tachycardia of 130/min. Laboratory analysis revealed hyponatremia (122 mmol/L), while ascites fluid analysis showed no infection. During the first 48 hours of hospitalization the patient remained persistently hypotensive despite adequate vascular filling and the addition of noradrenaline. A standard-dose short synacthen test was performed which revealed a poor cortisol response, which is a compatible criterion for the diagnosis of RAI. Intravenous hydrocortisone therapy was initiated, which resulted in a rapid improvement in patient's general condition, and increase in blood pressure. As the patient became hemodynamically stable without the need of noradrenaline, the hydrocortisone dose was weaned progressively, and he was discharged after 18 days of hospitalization in a stable condition.


Effects of expiratory muscle strength training on swallowing function in acute stroke patients with dysphagia.
[Purpose] This study was conducted to identify the effects of expiratory muscle strength training on swallowing function in acute stroke patients with dysphagia. [Subjects and Methods] A total of 18 stroke patients with dysphagia were enrolled in the study. All participants were randomly assigned to either an experimental group (n=9) or a control group (n=9). All participants performed traditional-swallowing rehabilitation therapy in 30-minute sessions five times a week for four weeks; however, only the experimental group received expiratory muscle strength training. [Results] Both groups showed significant improvements after mediation. When compared with the control group, the functional dysphagia scale, vallecular residue, and penetration-aspiration scale were significantly improved in the experimental group. [Conclusion] Expiratory muscle strength training is an effective intervention for impaired swallowing function in acute strike patients with dysphagia.


Kale (Brassica oleracea var. acephala) as a superfood: Review of the scientific evidence behind the statement.
Kale (Brassica oleracea var. acephala) is a cruciferous vegetable, characterized by leaves along the stem, which, in recent years, have gained a great popularity as a ´superfood´. Consequently, in a popular culture it is listed in many ´lists of the healthiest vegetables´. Without the doubt, a scientific evidences support the fact that cruciferous vegetables included in human diet can positively affect health and well-being, but remains unclear why kale is declared superior in comparison with other cruciferous. It is questionable if this statement about kale is triggered by scientific evidence or by some other factors. Our review aims to bring an overview of kale's botanical characteristics, agronomic requirements, contemporary and traditional use, macronutrient and phytochemical content and biological activity, in order to point out the reasons for tremendous kale popularity.


Uptake and release of 3-H-metaraminol by rat lung. Basic aspects, ionic and energy requirements.
The accumulation of 3H-metaraminol in incubated lung tissue of the rat was examined in vitro. Lung tissue concentrated tritium after incubation with 3H-metaraminol. This uptake proceeds against a concentration gradient and shows saturation kinetics (Km 12.5 X 10(-7) M and Vmax 2.105 nM/g/15 min.) with great affinity for 3H-metaraminol. The accumulation of 3H-metaraminol is dependent of temperature and energy supply, and requires the presence of sodium ions. The loss of accumulated radioactivity showed the possibility that a small fraction of metaraminol is apparently firmly boune (e.g. neuronal sites), while a greater portion is taken up by extraneuronal structures (e.g. capillary endothelial cells). The first localization could represent uptake-1 and the second uptake-2. The results obtained also confirm the important role of the lungs as an organ which participates in the fate of endogenous substances and drugs.


Comparison of Renal Complications between Endografts with Suprarenal and Infrarenal Fixation.
Surgeons have multiple grafts options available for the endovascular treatment of abdominal aortic aneurysm (EVAR), and some hypothesize that suprarenal fixation endografts may result in higher rates of renal complications than infrarenal endografts. This study aimed to compare the outcomes of contemporary suprarenal and infrarenal endografts. The Targeted Vascular Module of the National Surgical Quality Improvement Project was utilised to identify patients undergoing EVAR for infrarenal aneurysm from 2011 to 2013. Pre-operative and operative variables and 30 day outcomes were compared among suprarenal (Zenith and Endurant) and infrarenal fixation devices (Excluder). Renal complications included creatinine increase > 2 mg/dL or new dialysis, as defined by NSQIP. Multivariate regression was completed to account for patient demographics, comorbidities, and operative characteristics. A total of 3587 patients were evaluated including 2273 (63%) with suprarenal grafts and 1314 (37%) with infrarenal grafts. Patients with suprarenal grafts were less commonly white (84% vs. 88%, p < .01) and more commonly male (83% vs. 80%, p = .03). There were no differences in age or comorbidities. Renal complications (1.1% vs. 0.1%, p < .01) and length of stay more than 2 days (34% vs. 25%, p < .01) occurred more commonly after suprarenal fixation. After adjustment, suprarenal grafts had significantly higher rates of renal complications (OR, 12.0; 95% CI, 1.6-91) and length of stay more than 2 days (OR, 1.4; 95% CI, 1.2-1.7). Overall rates of renal complications following EVAR are low. Patients selected for suprarenal stent grafts are at increased risk of renal complications and prolonged length of stay, which may be due to selection bias, deployment techniques, or the presence of a bare stent overlying the renal arteries. Further studies are necessary to evaluate the mechanism and duration of renal dysfunction and important long-term outcomes of interest.


Microglial depletion aggravates the severity of acute and chronic seizures in mice.
Microglia are the resident immune cells of the center nervous system and participate in various neurological diseases. Here we determined the function of microglia in epileptogenesis using microglial ablation approaches. Three different microglia-specific genetic tools were used, CX3CR1CreER/+:R26iDTA/+, CX3CR1CreER/+:R26iDTR/+, and CX3CR1CreER/+:Csf1rFlox/Flox mice. We found that microglial depletion led to worse kainic acid (KA)-induced status epilepticus, higher mortality rate, and increased neuronal degeneration in the hippocampus. In KA-induced chronic spontaneous recurrent seizures, microglial depletion increased seizure frequency, interictal spiking, and seizure duration. Therefore, microglial depletion aggravates the severity of KA-induced acute and chronic seizures. Interestingly, microglial repopulation reversed the effects of depletion upon KA-induced status epilepticus. Our results demonstrate a beneficial role of microglia in suppressing both acute and chronic seizures, suggesting that microglia are a potential therapeutic target for the management of epilepsy.


M34 actin regulatory protein is a sensitive diagnostic marker for early- and late-stage mammary carcinomas.
At present, there is no available molecular marker that reliably detects the earliest stages of epithelial transformation in the majority of patients affected with incipient breast carcinoma. Here we introduce M34 protein, a mammalian actin filament regulatory protein, as a highly sensitive and easily detected positive cellular marker for both early and late stages of breast carcinoma. In this study, 24 human lactation duct neoplasms from postmenopausal women, including fibroadenoma, ductal carcinoma in situ, intraductal lobular papilloma, and metastatic adenocarcinoma, were analyzed for the presence of M34 protein by histochemical staining of paraffin and fresh-frozen sections. All 24 neoplasias tested positive for M34, whereas none of the 4 normal breast tissues stained for the protein. M34 identification was strongly positive for transformed epithelium in all tumor types tested. Twelve precancerous lesions of fibroadenoma (n = 4), intraductal papilloma (n = 4), and incipient ductal carcinoma in situ (n = 4) all showed high levels of M34 staining, suggesting that precancerous tumors, as well as the earliest stages of mammary carcinoma, can be sensitively detected. Furthermore, anti-M34 antibody selectively stained all 12 advanced-stage metastatic adenocarcinoma cell masses and micrometastases in axillary lymph nodes tested. Single-cell micrometastases embedded in connective tissue or lymph node parenchyma could be clearly resolved by M34 with horseradish peroxidase staining. Lymphocytes, normal ductal endothelium, and vascular endothelial cells were M34-negative, as were muscle, nerve, and adipose tissues. Low-level M34 staining was detected in connective tissue fibroblasts, macrophages, and neutrophils. To our knowledge, no previously reported markers have shown high sensitivity of detection for both the earliest and most advanced stages of breast carcinoma. Consequently, M34 appears uniquely suited for diagnosis of the earliest stages of lactation duct transformation as well as for advanced-stage mammary carcinoma metastases in surgical margins and axillary lymph nodes.


[Progress of biological air filter (BAF) development in manned spacecraft cabin].
The contaminants originating from human metabolism, material off-gassing and waste processing, may influence human health and the growth and development of higher plants when they accumulate at some degree in the spacecraft cabin. So the contaminants concentrations must be controlled below the spacecraft maximum allowable concentration (SMAC). For the long manned space missions and planetary habitation, biological technique is available for the removal of the contaminants. The biological air filter, BAF, is a system that degrades the contaminants into carbon dioxide, water and salts. It holds many advantages such as small weight and volume, low power consumption, easy maintenance and good working performance under the condition of microgravity. Its wide application will be seen in the space field in near future.


Application of Dexmedetomidine as an Opioid Substitute in Opioid-Free Anesthesia: A Systematic Review and Meta-analysis.
Opioid-based general anesthesia was previously used to alleviate perioperative pain; however, several complications associated with using anesthesia have raised several concerns. Various studies have investigated the application prospect of using opioid-free general anesthesia, such as dexmedetomidine, as an opioid substitute. We performed a systematic review and meta-analysis to explore and highlight the safety and effectiveness of dexmedetomidine as an opioid substitute for opioid-free anesthesia. A systematic review and meta-analysis. We screened for suitable clinical trials from electronic databases, including "PubMed," "Cochrane Library," "EMBASE," and "Web of Science." Eligible trials were included in this meta-analysis. The quality of the screened randomized controlled trials (RCTs) was determined using the risk of bias assessment criteria by the Cochrane Collaboration tool. We used the "Review Manager 5.3" and "Stata 10.0" software to perform the meta-analysis. We evaluated the quality of evidence using the "Grading of Recommendations Assessment, Development, and Evaluation" approach. For the analysis, we included 32 RCTs encompassing 2,509 patients. In the opioid-free group, the 2-hour postoperative pain score of patients (mean difference = -0.53, 95% CI: -1.00, -0.07; P = 0.02, I2=78%) was significantly lower compared to those in the opioid-based group. In addition, several patients required rescue analgesia (risk ratio = 0.70, 95% CI: 0.58, 0.84, P < 0.05, I2 = 71%) and opioids postsurgery. However, the duration of extubation and postanesthesia care unit, as well as the incidences of bradycardia, were high in patients receiving dexmedetomidine as opioid-free general anesthesia. Subgroup analysis for different anesthesia-maintaining drugs had not been conducted. The heterogeneity did not reduce after subgroup analysis. Different doses of dexmedetomidine had not been evaluated. These findings indicate that opioid-free general anesthesia based on dexmedetomidine could be effective; however, prolonged extubation time and cardiovascular complications are a few risks associated with dexmedetomidine.


[Malignancy in gastric carcinoma, with special reference to the DNA ploidy and proliferative activity].
Analysis of DNA ploidy patterns was performed on 129 primary gastric cancers and the results correlated with histologic findings and in vivo bromodeoxyuridine (BrdU) labeling. Forty-nine cases were diploid (38%) and 80 cases were aneuploid. There was no correlation between DNA ploidy and histologic type. In aneuploid cancers, incidence of lymphatic invasion, lymph node metastasis and rate of advanced cases were significantly higher than those in diploid tumors. During the follow-up period of 5-10 years, 23 of the 40 patients (55%) with aneuploid tumors died of disease within 3-120 months. Only 13 of the 36 patients (36%) with diploid tumors died of disease. The BrdU labeling indices (BrdU LI) were from 2.8% to 26.7%, with a mean of 10.4%. There was no correlation between BrdU LI and histologic type or stage. The mean BrdU LI of early cancers was 8.1%, of advanced cancers, 11.9%. BrdU LI of cancers with lymphatic invasion or lymph node metastasis was higher than those without them. The mean BrdU LI of diploid cancer was 6.0%, of aneuploid cancers, 11.9%. There was a good correlation between BrdU labeling indices and DNA ploidy patterns. These results indicate that determination of DNA ploidy patterns and growth fractions by BrdU labeling may be useful in the conjecture of prognosis of the patient and in the selection of patients from various modalities.


Genetic evaluation of inherited muscle diseases.
Inherited muscle diseases are common and can present to the neurologist as anything from an incidentally found high creatine kinase level with muscle soreness in an adult to severe weakness at birth. Although a vast number of gene mutations can cause muscle degeneration, a solid understanding of the clinical presentation and gene diagnostic methods of the three most common forms of inherited muscle disease (Duchenne, myotonic, and facioscapulohumeral muscular dystrophy) will allow for the accurate diagnosis in most cases. Additionally, a general understanding of the cellular pathways involved in maintaining proper muscle fiber integrity helps provide a road map for evaluating less common disorders that are more of a diagnostic challenge, including limb-girdle muscular dystrophy, distal myopathy, congenital myopathy, and congenital muscular dystrophy.


Mechanisms and clinical significance of multidrug resistance.
Tumor cells often become refractory to diverse drugs with different mechanisms of cytotoxic action. This paper reviews the current state of our knowledge of multidrug resistance, the limitations of present concepts to fully explain the diversity of the MDR phenotypes, and the clinical relevance of these studies derived largely from cell culture systems. The authors discuss the use of markers associated with the multi-drug resistance phenotype to identify potentially drug-resistant tumors and outline strategies that might be used to overcome the resistance phenomenon.


Fourier analysis to measure diffusion coefficients and resolve mixtures on a continuous electrophoresis chip.
We report a method to measure diffusion coefficients of fluorescent solutes in the 10(2)-10(6) Da molecular mass range in a glass-PDMS chip. Upon applying a permanent electric field, the solute is introduced through a narrow channel into a wide analysis chamber where it migrates along the injection axis and diffuses in two dimensions. The diffusion coefficient is extracted after 1D Fourier transform of the resulting stationary concentration pattern. Analysis is straightforward, requiring no numerical integration or velocity field simulation. The diffusion coefficients measured for fluorescein, rhodamine green-labeled oligonucleotides, and YOYO-1-stained dsDNA fragments agree with the literature values and with our own fluorescence correlation spectroscopy measurements. As shown for 151 and 1257 base pair dsDNA mixtures, the present method allows us to rely on diffusion to quantitatively characterize the nature and the composition of binary mixtures. In particular, we implement a DNA hybridization assay to illustrate the efficiency of the proposed protocol for library screening.


MicroRNA-374b inhibits liver cancer progression via down regulating programmed cell death-1 expression on cytokine-induced killer cells.
Programmed cell death-1 (PD-1) is an oncogene associated with suppressing proliferation and cytokine production of T cells in the progression of liver cancer. microRNAs (miRs) regulate gene expression via specific binding to the target 3'untranslated region of mRNA. In the present study, miR-374b was indicated to interact with PD-1 and affect the tumor-targeting capacity of cytokine-induced killer (CIK) cells. miR-374b inhibitor significantly increased PD-1 expression in CIK cells. A synthetic small interfering (si)RNA targeting PD-1 was employed to silence the expression level of PD-1 in CIK cells. Then, the antitumor effect of siPD-1 in CIK cells was investigated. In vitro study demonstrated that IFN-γ secretion and the concentration of lactate dehydrogenase were significantly increased in the PD-1 knockdown group; however, the viability of HepG2 cells in the PD-1 knockdown group had significantly decreased, compared with the HepG2 cells in the negative control group. In vivo study indicated that mice inoculated with HepG2 cells and CIK cells with PD-1 knocked down had a significantly smaller tumor volume, compared with the control group. To conclude, human CIK cells transfected with siPD-1 can target liver cancer cells and enhance immunotherapy efficacy, and therefore they have potential in the immunotherapy of liver cancer.


A genetic switch controls the production of flagella and toxins in Clostridium difficile.
In the human intestinal pathogen Clostridium difficile, flagella promote adherence to intestinal epithelial cells. Flagellar gene expression also indirectly impacts production of the glucosylating toxins, which are essential to diarrheal disease development. Thus, factors that regulate the expression of the flgB operon will likely impact toxin production in addition to flagellar motility. Here, we report the identification a "flagellar switch" that controls the phase variable production of flagella and glucosylating toxins. The flagellar switch, located upstream of the flgB operon containing the early stage flagellar genes, is a 154 bp invertible sequence flanked by 21 bp inverted repeats. Bacteria with the sequence in one orientation expressed flagellum and toxin genes, produced flagella, and secreted the toxins ("flg phase ON"). Bacteria with the sequence in the inverse orientation were attenuated for flagellar and toxin gene expression, were aflagellate, and showed decreased toxin secretion ("flg phase OFF"). The orientation of the flagellar switch is reversible during growth in vitro. We provide evidence that gene regulation via the flagellar switch occurs post-transcription initiation and requires a C. difficile-specific regulatory factor to destabilize or degrade the early flagellar gene mRNA when the flagellar switch is in the OFF orientation. Lastly, through mutagenesis and characterization of flagellar phase locked isolates, we determined that the tyrosine recombinase RecV, which catalyzes inversion at the cwpV switch, is also responsible for inversion at the flagellar switch in both directions. Phase variable flagellar motility and toxin production suggests that these important virulence factors have both advantageous and detrimental effects during the course of infection.


CRY1/2 Selectively Repress PPARδ and Limit Exercise Capacity.
Cellular metabolite balance and mitochondrial function are under circadian control, but the pathways connecting the molecular clock to these functions are unclear. Peroxisome proliferator-activated receptor delta (PPARδ) enables preferential utilization of lipids as fuel during exercise and is a major driver of exercise endurance. We show here that the circadian repressors CRY1 and CRY2 function as co-repressors for PPARδ. Cry1-/-;Cry2-/- myotubes and muscles exhibit elevated expression of PPARδ target genes, particularly in the context of exercise. Notably, CRY1/2 seem to repress a distinct subset of PPARδ target genes in muscle compared to the co-repressor NCOR1. In vivo, genetic disruption of Cry1 and Cry2 enhances sprint exercise performance in mice. Collectively, our data demonstrate that CRY1 and CRY2 modulate exercise physiology by altering the activity of several transcription factors, including CLOCK/BMAL1 and PPARδ, and thereby alter energy storage and substrate selection for energy production.


Medical libraries from Ankara to London.
Thirteen European libraries were visited and are described with special attention given to the Turkish and Hungarian library systems. Information and personal impressions give insight into European ideas of librarianship.


Identification of a common cyanobacterial symbiont associated with Azolla spp. through molecular and morphological characterization of free-living and symbiotic cyanobacteria.
Symbiotically associated cyanobacteria from Azolla mexicana and Azolla pinnata were isolated and cultured in a free-living state. Morphological analyses revealed differences between the free-living isolates and their symbiotic counterparts, as did restriction fragment length polymorphism (RFLP) analyses with both single-copy glnA and rbcS gene probes and a multicopy psbA gene probe. RFLP analyses with Anabaena sp. strain PCC 7120 nifD excision element probes, including an xisA gene probe, detected homologous sequences in DNA extracted from the free-living isolates. Sequences homologous to these probes were not detected in DNA from the symbiotically associated cyanobacteria. These analyses indicated that the isolates were not identical to the major cyanobacterial symbiont species residing in leaf cavities of Azolla spp. Nevertheless, striking similarities between several free-living isolates were observed. In every instance, the isolate from A. pinnata displayed banding patterns virtually identical to those of free-living cultures previously isolated from Azolla caroliniana and Azolla filiculoides. These results suggest the ubiquitous presence of a culturable minor cyanobacterial symbiont in at least three species of Azolla.


Methods for assessing the extent of exposure and effects of air pollution.
In many places in Europe, the ambient air pollution exceeds the levels considered to be safe for health. The objective of the paper is to review and summarise the methods of assessment of its impact on health, and to indicate the contributions of various research disciplines, particularly environmental epidemiology. The framework for assessment of impact is based on a four stage model: assessment of release of pollutant; assessment of exposure; assessment of the consequence; and risk estimation. Epidemiology is crucial in providing the data for the assessment of consequence. The criteria that determine the use of epidemiological studies for this task include lack of bias, correct control of confounding, and measured estimates of exposure. At present, those criteria are easier to satisfy for studies of short term effects on health than for the delayed consequences of exposure, or exposure accumulated over a prolonged period. Combinations of results from various populations through meta-analysis of existing studies or conducting multicentre studies is often necessary to increase the reliability of the consequence assessment stage. To assess the impact on health systematically helps to focus on actions to limit air pollutants with the greatest impacts on human health and on the most affected populations. This method allows identification of the most pertinent questions which have to be answered by studies on relations between pollution and health and on exposure of populations to air pollutants. Epidemiology has considerable potential to contribute to this research.


Subclinical ketosis in dairy cows.
Subclinical ketosis is defined as a preclinical stage of ketosis. The peak prevalence of subclinical ketosis occurs during the fourth week of lactation. Herd-related factors, breed, parity, and season are other important determinants. Subclinical ketosis can be revealed by determining levels of plasma glucose, plasma NEFA and blood, and milk or urine ketone body concentration. There are theoretical and practical advantages of using milk ketone bodies. Most authors are agreed on approximate lower and upper borderlines for subclinical ketosis. The risk of an outbreak of clinical symptoms has been evaluated by some authors. Most authors have found significant negative relationships between energy balance and ketone body concentration. Some disagreement may be attributable to the fact that the diets used in different experiments can have different glucogenic potential, even if the energy content is the same. This affects the relationship between energy balance and ketone body concentration, as the ketone body level is influenced by both the energy balance and plasma glucose. Feeding silage with high butyric acid content increases the risk of subclinical ketosis. There are indications that cows with the highest milk yield directly after calving are at greatest risk for developing ketosis. Increased ketone body level secondarily reduces milk production, a decrease that has been quantified by some authors. Subclinical ketosis causes delayed reproductive functions return to normal after calving, increased intervals from calving to first and last service, and an increased frequency of ovarian cysts. The routine determination of milk acetone levels in control programs can be used to evaluate the status of individual cows, to indicate the energy feeding in early lactation at a herd level, and to evaluate sires for breeding. The heritability and the tendency toward a positive genetic correlation between milk acetone and milk yield have also been discussed, as have aspects of nutritional prevention. Factors such as energy- and protein-rich roughage, tasty high-energy concentrates, suitable feeding during the dry period, and division of the concentrates into at least four meals are considered to be important.


Cadmium affects focal adhesion kinase (FAK) in mesangial cells: involvement of CaMK-II and the actin cytoskeleton.
The toxic metal ion cadmium (Cd(2+)) induces pleiotropic effects on cell death and survival, in part through effects on cell signaling mechanisms and cytoskeletal dynamics. Linking these phenomena appears to be calmodulin-dependent activation of the Ca(2+)/calmodulin-dependent protein kinase II (CaMK-II). Here we show that interference with the dynamics of the filamentous actin cytoskeleton, either by stabilization or destabilization, results in disruption of focal adhesions at the ends of organized actin structures, and in particular the loss of vinculin and focal adhesion kinase (FAK) from the contacts is a result. Low-level exposure of renal mesangial cells to CdCl2 disrupts the actin cytoskeleton and recapitulates the effects of manipulation of cytoskeletal dynamics with biological agents. Specifically, Cd(2+) treatment causes loss of vinculin and FAK from focal contacts, concomitant with cytoskeletal disruption, and preservation of cytoskeletal integrity with either a calmodulin antagonist or a CaMK-II inhibitor abrogates these effects of Cd(2+). Notably, inhibition of CaMK-II decreases the migration of FAK-phosphoTyr925 to a membrane-associated compartment where it is otherwise sequestered from focal adhesions in a Cd(2+)-dependent manner. These results add further insight into the mechanism of the CaMK-II-dependent effects of Cd(2+) on cellular function.


Schwann cell-derived Apolipoprotein D controls the dynamics of post-injury myelin recognition and degradation.
Management of lipids, particularly signaling lipids that control neuroinflammation, is crucial for the regeneration capability of a damaged nervous system. Knowledge of pro- and anti-inflammatory signals after nervous system injury is extensive, most of them being proteins acting through well-known receptors and intracellular cascades. However, the role of lipid binding extracellular proteins able to modify the fate of lipids released after injury is not well understood. Apolipoprotein D (ApoD) is an extracellular lipid binding protein of the Lipocalin family induced upon nervous system injury. Our previous study shows that axon regeneration is delayed without ApoD, and suggests its participation in early events during Wallerian degeneration. Here we demonstrate that ApoD is expressed by myelinating and non-myelinating Schwann cells and is induced early upon nerve injury. We show that ApoD, known to bind arachidonic acid (AA), also interacts with lysophosphatidylcholine (LPC) in vitro. We use an in vivo model of nerve crush injury, a nerve explant injury model, and cultured macrophages exposed to purified myelin, to uncover that: (i) ApoD regulates denervated Schwann cell-macrophage signaling, dampening MCP1- and Tnf-dependent macrophage recruitment and activation upon injury; (ii) ApoD controls the over-expression of the phagocytosis activator Galectin-3 by infiltrated macrophages; (iii) ApoD controls the basal and injury-triggered levels of LPC and AA; (iv) ApoD modifies the dynamics of myelin-macrophage interaction, favoring the initiation of phagocytosis and promoting myelin degradation. Regulation of macrophage behavior by Schwann-derived ApoD is therefore a key mechanism conditioning nerve injury resolution. These results place ApoD as a lipid binding protein controlling the signals exchanged between glia, neurons and blood-borne cells during nerve recovery after injury, and open the possibility for a therapeutic use of ApoD as a regeneration-promoting agent.


Concise review: Current understanding of extracellular vesicles to treat neuropathic pain.
Extracellular vesicles (EVs) including exosomes are vesicular vesicles with phospholipid bilayer implicated in many cellular interactions and have the ability to transfer multiple types of cargo to cells. It has been found that EVs can package various molecules including proteins and nucleic acids (DNA, mRNA, and noncoding RNA). The discovery of EVs as carriers of proteins and various forms of RNA, such as microRNAs (miRNA) and long noncoding RNAs (lncRNA), has raised great interest in the field of drug delivery. Despite the underlying mechanisms of neuropathic pain being unclear, it has been shown that uncontrolled glial cell activation and the neuroinflammation response to noxious stimulation are important in the emergence and maintenance of neuropathic pain. Many studies have demonstrated a role for noncoding RNAs in the pathogenesis of neuropathic pain and EVs may offer possibilities as carriers of noncoding RNAs for potential in neuropathic pain treatment. In this article, the origins and clinical application of EVs and the mechanism of neuropathic pain development are briefly introduced. Furthermore, we demonstrate the therapeutic roles of EVs in neuropathic pain and that this involve vesicular regulation of glial cell activation and neuroinflammation.


Changes in predictive cuing modulate the hemispheric distribution of the P1 inhibitory response to attentional targets.
Brain activity related to orienting of attention with spatial cues and brain responses to attentional targets are influenced the probabilistic contingency between cues and targets. Compared to predictive cues, cues predicting at chance the location of targets reduce the filtering out of uncued locations and the costs in reorienting attention to targets presented at these locations. Slagter et al. (2016) have recently suggested that the larger target related P1 component that is found in the hemisphere ipsilateral to validly cued targets reflects stimulus-driven inhibition in the processing of the unstimulated side of space contralateral to the same hemisphere. Here we verified whether the strength of this inhibition and the amplitude of the corresponding P1 wave are modulated by the probabilistic link between cues and targets. Healthy participants performed a task of endogenous orienting once with predictive and once with non-predictive directional cues. In the non-predictive condition we observed a drop in the amplitude of the P1 ipsilateral to the target and in the costs of reorienting. No change in the inter-hemispheric latencies of the P1 was found between the two predictive conditions. The N1 facilitatory component was unaffected by predictive cuing. These results show that the predictive context modulates the strength of the inhibitory P1 response and that this modulation is not matched with changes in the inter-hemispheric interaction between the P1 generators of the two hemispheres.


Phonological phrase boundaries constrain the online syntactic analysis of spoken sentences.
Two experiments tested whether phonological phrase boundaries constrain online syntactic analysis in French. Pairs of homophones belonging to different syntactic categories (verb and adjective) were used to create sentences with a local syntactic ambiguity (e.g., [le petit chien mort], in English, the dead little dog, vs. [le petit chien] [mord], in English, the little dog bites, where brackets indicate phonological phrase boundaries). An expert speaker recorded the sentences with either a maximally informative prosody or a minimally informative one. Participants correctly assigned the appropriate syntactic category to the target word, even without any access to the lexical disambiguating information, in both a completion task (Experiment 1) and an abstract word detection task (Experiment 2). The size of the experimental effect was modulated by the prosodic manipulation (maximally vs. minimally informative), guaranteeing that prosody played a crucial role in disambiguation. The authors discuss the implications of these results for models of online speech perception and language acquisition.


Diagnostic testing holds the key to NTD elimination.
"Fit-for-purpose" diagnostic tests have emerged as a prerequisite to achieving global targets for the prevention, control, elimination, and eradication of neglected tropical diseases (NTDs), as highlighted by the World Health Organization's (WHO) new roadmap. There is an urgent need for the development of new tools for those diseases for which no diagnostics currently exist and for improvement of existing diagnostics for the remaining diseases. Yet, efforts to achieve this, and other crosscutting ambitions, are fragmented, and the burden of these 20 debilitating diseases immense. Compounded by the Coronavirus Disease 2019 (COVID-19) pandemic, programmatic interruptions, systemic weaknesses, limited investment, and poor commercial viability undermine global efforts-with a lack of coordination between partners, leading to the duplication and potential waste of scant resources. Recognizing the pivotal role of diagnostic testing and the ambition of WHO, to move forward, we must create an ecosystem that prioritizes country-level action, collaboration, creativity, and commitment to new levels of visibility. Only then can we start to accelerate progress and make new gains that move the world closer to the end of NTDs.


Crude dietary polysaccharide fraction isolated from jackfruit enhances immune system activity in mice.
Crude polysaccharides (PSs) were isolated from the fruit pulp of jackfruit, and their chemical composition determined and evaluated for an immune regulatory activity in mice. The PSs were isolated from water extracts of jackfruit pulp (JFP) using the ethanol precipitation method. The resulting precipitates were further purified by dialysis and protein depletion by the Sevage method. The phenol-sulfuric method was used to determine the content of the PSs. The composition of PSs was determined by the Sephadex-G200 column chromatography and high-performance liquid chromatography methods. The thymus index and macrophage phagocytic function methods in mice were used to evaluate the immune regulatory activity of JFP-PSs. The JFP-PSs content in jackfruit was about 21% (w/w) and the yield of crude PSs was 3.91%. The single molecular mass weight PS was the main constituent of JFP-PSs. The major monosaccharide residues were rhamnose, glucose, galactose, and arabinose. The JFP-PSs enhanced the thymus weight index and the phagocytic rate after 30 days of subchronic p.o. administration to mice at 4.5 mg/kg. The JFP contains single molecular PS and JFP-PS has immune-stimulating activities in mice. These data suggest that at least some of the traditional uses of JFP can be ascribed to its immunomodulatory effects.


Expression and distribution of osteopontin and matrix metalloproteinase (MMP)-3 and -7 in mouse salivary glands.
We investigated the expression and distribution of osteopontin in mouse salivary glands. Western blot analysis showed intense positive bands at the predicted molecular mass (about 60 kDa) in mouse parotid and sublingual glands. However, a cross-reacted band around 30 kDa was strongly detected in submandibular glands. Indirect immunofluorescent analysis showed that osteopontin was localized at the luminal (apical) membranes of the acinar cells in parotid and sublingual glands. However, it was not detected in acinar cells of submandibular glands. No expression was found in ductal cells of any glands. We also examined the expression of matrix metalloproteinase (MMP)-3 and -7. In parotid gland, MMP-3 was observed at 57 kDa, indicating a latent form, but MMP-7 was not detected. In contrast, MMP-7 definitely was observed at 28 kDa area in submandibular gland, whereas MMP-3 was not detected. These results suggest that osteopontin localizes at luminal sites of acinar cells and may be associated with saliva secretion in mouse salivary gland. It is also suggested that osteopontin may be cleaved by MMP-7 in mouse submandibular gland.


Vonoprazan fumarate, a novel potassium-competitive acid blocker, in the management of gastroesophageal reflux disease: safety and clinical evidence to date.
Potassium-competitive acid blocker (P-CAB) is a class of drug that competitively blocks the potassium-binding site of H+, K+-adenosine triphosphate (ATP)ase. Although the history of this class of drugs started over 30 years ago, clinical use of two P-CABs, revaprazan and vonoprazan, were only recently approved in Korea and Japan, respectively. Among them, vonoprazan has several advantages over conventional proton-pump inhibitors (PPIs), including rapid onset of action, long duration of acid suppression, fewer interindividual variations in terms of acid suppression, and minimum dietary influence on its action. These advantages of vonoprazan have been proved in clinical trials conducted for license approvals for several acid-related diseases. In this review article, current evidence of vonoprazan in the management of gastroesophageal reflux disease (GERD) will be summarized. Since the clinical trial data, as well as postmarketed clinical data, have consistently demonstrated superiority of vonoprazan over conventional PPIs in terms of achieving healing of mucosal breaks and maintaining the healing, it may provide an excellent, if not complete, option for fulfilling some of the unmet needs for current GERD therapy. The safety problem of vonoprazan is also discussed, as more pronounced hypergastrinemia inevitably ensues with its use.


Multiple self-care behaviors and associated factors in community-dwelling patients with hypertension in Myanmar.
This study aimed to identify the prevalence of self-care behaviors and the associated factors among hypertensive patients in primary care in Myanmar. This cross-sectional study was conducted from April to May 2019 among 410 hypertensive patients in Myitkyina Township, Kachin State, Myanmar. Hypertensive patients aged 30-70 years old and being registered at the community health centers in Myitkyina Township were selected using multi-stage cluster random sampling. Self-care behaviors were measured by Hypertensive Self-Care Activity Level Effect (H-SCALE). Chi-square test and multiple logistic regression analysis were used to explore the associated factors. Prevalence of adherence to multiple self-care behaviors were low: avoidance of tobacco use at 50.2%, followed by physical activity at 24.9%, medication at 24.1%, weight management at 9.5%, and healthy diet at 2.7%, while abstinence from harmful alcohol drinking was high at 97.8%. Multiple logistic regression analysis indicated that younger patients, low family income, inadequate knowledge, and no comorbidity were associated with non-adherence to medication. Living in rural area and having poor self-efficacy were associated with non-adherence to weight management, while being younger, female and having poor self-efficacy were also associated with non-adherence to physical activity. Compared with Kachin, other ethnics were more likely to be non-adherent to avoidance of tobacco use. Although a majority of respondents were not harmful drinkers, adherence to medication, healthy diet, physical activity, weight management and avoidance of tobacco use were very low. Health practitioners should provide education programs for hypertensive patients to direct them towards practical techniques in managing their blood pressure.


Exopolysaccharide production and ropy phenotype are determined by two gene clusters in putative probiotic strain Lactobacillus paraplantarum BGCG11.
Lactobacillus paraplantarum BGCG11, a putative probiotic strain isolated from a soft, white, artisanal cheese, produces a high molecular-weight heteropolysaccharide, exopolysaccharide (EPS)-CG11, responsible for the ropy phenotype and immunomodulatory activity of the strain. In this study, a 26.4-kb region originating from the pCG1 plasmid, previously shown to be responsible for the production of EPS-CG11 and a ropy phenotype, was cloned, sequenced, and functionally characterized. In this region 16 putative open reading frames (ORFs), encoding enzymes for the production of EPS-CG11, were organized in specific loci involved in the biosynthesis of the repeat unit, polymerization, export, regulation, and chain length determination. Interestingly, downstream of the eps gene cluster, a putative transposase gene was identified, followed by an additional rfb gene cluster containing the rfbACBD genes, the ones most probably responsible for dTDP-L-rhamnose biosynthesis. The functional analysis showed that the production of the high-molecular-weight fraction of EPS-CG11 was absent in two knockout mutants, one in the eps and the other in the rfb gene cluster, as confirmed by size exclusion chromatography analysis. Therefore, both eps and rfb genes clusters are prerequisites for the production of high-molecular-weight EPS-CG11 and for the ropy phenotype of strain L. paraplantarum BGCG11.


Effect of glycogen depletion on the oxygen uptake slow component in humans.
Previous studies have indicated that the (.-)VO(2) slow component is related to the recruitment of type II muscle fibres. We therefore hypothesised that an exercise and dietary regimen designed to deplete type I muscle fibres of glycogen would result in a greater contribution of type II muscle fibres to the exercise power output and therefore a larger amplitude of the (.-)VO(2) slow component. Eight male subjects took part in this study. On day 1, the subjects reported to the laboratory at 8 a.m., and completed a 9 min constant-load cycling test at a work rate equivalent to 85 % (.-)VO(2) peak. On day 2 at 12 p.m., the subjects were fed a 4200 kJ meal (60 % protein, 40 % fat); at 6 p.m. they completed a 2 h cycling test at 60 % (.-)VO(2) peak. On day 3 at 8 a.m., the subjects performed an exercise test identical to that of day 1. Metabolic and respiratory measurements indicated that our experimental design was effective in reducing the muscle glycogen content. (.-)VO(2) was significantly higher (by approximately 140 ml x min (-1)) throughout exercise following glycogen depletion but no appreciable changes in (.-)VO(2) kinetics were found: neither the time constant of the primary response (from 35.4 +/- 2.5 to 33.2 +/- 4.4 s) nor the amplitude of the slow component (from 404 +/- 95 to 376 +/- 81 ml x min (-1)) was significantly altered. Therefore, we suggest that the increased (.-)VO(2) throughout exercise and the unaltered (.-)VO(2) slow component following glycogen depletion might be explained by a shift towards a greater reliance on fat metabolism in type I muscle fibres with no appreciable change in fibre type recruitment patterns.


Glycan modulation and sulfoengineering of anti-HIV-1 monoclonal antibody PG9 in plants.
Broadly neutralizing anti-HIV-1 monoclonal antibodies, such as PG9, and its derivative RSH hold great promise in AIDS therapy and prevention. An important feature related to the exceptional efficacy of PG9 and RSH is the presence of sulfated tyrosine residues in their antigen-binding regions. To maximize antibody functionalities, we have now produced glycan-optimized, fucose-free versions of PG9 and RSH in Nicotiana benthamiana. Both antibodies were efficiently sulfated in planta on coexpression of an engineered human tyrosylprotein sulfotransferase, resulting in antigen-binding and virus neutralization activities equivalent to PG9 synthesized by mammalian cells ((CHO)PG9). Based on the controlled production of both sulfated and nonsulfated variants in plants, we could unequivocally prove that tyrosine sulfation is critical for the potency of PG9 and RSH. Moreover, the fucose-free antibodies generated in N. benthamiana are capable of inducing antibody-dependent cellular cytotoxicity, an activity not observed for (CHO)PG9. Thus, tailoring of the antigen-binding site combined with glycan modulation and sulfoengineering yielded plant-produced anti-HIV-1 antibodies with effector functions superior to PG9 made in CHO cells.


The pathogen Neisseria meningitidis requires oxygen, but supplements growth by denitrification. Nitrite, nitric oxide and oxygen control respiratory flux at genetic and metabolic levels.
The human pathogen Neisseria meningitidis is the major causative agent of bacterial meningitis. The organism is usually treated as a strict aerobe and is cultured under fully aerobic conditions in the laboratory. We demonstrate here that although N. meningitidis fails to grow under strictly anaerobic conditions, under oxygen limitation the bacterium expresses a denitrification pathway (reduction of nitrite to nitrous oxide via nitric oxide) and that this pathway supplements growth. The expression of the gene aniA, which encodes nitrite reductase, is regulated by oxygen depletion and nitrite availability via transcriptional regulator FNR and two-component sensor-regulator NarQ/NarP respectively. Completion of the two-step denitrification pathway requires nitric oxide (NO) reduction, which proceeds after NO has accumulated during batch growth under oxygen-limited conditions. During periods of NO accumulation both nitrite and NO reduction are observed aerobically, indicating N. meningitidis can act as an aerobic denitrifier. However, under steady-state conditions in which NO is maintained at a low concentration, oxygen respiration is favoured over denitrification. NO inhibits oxidase activity in N. meningitidis with an apparent Ki NO = 380 nM measured in intact cells. The high respiratory flux to nitrite after microaerobic growth and the finding that accumulation of the denitrification intermediate NO inhibits oxygen respiration support the view that denitrification is a pathway of major importance in N. meningitidis.


The microbiome and its relevance in complex wounds.
Chronic wounds include arterial, venous, diabetic and pressure ulcers. The term "chronic wound" defines wounds that have not healed within three months. The possible causes that transform a simple wound into a chronic one are the object of study, and research has focused on infection as one of the crucial factors in producing and maintaining chronic wounds. In fact, 60% of chronic wounds are colonized by bacteria living in a biofilm. A biofilm is a complex aggregation of microorganisms characterized by the secretion of an adhesive and protective matrix. The two most common biofilm generators are Staphylococcus aureus and Pseudomonas aeruginosa. Several conditions constitute risk factors for the formation of biofilms, such as diabetes mellitus, venous insufficiency, malnutrition, cancer, oedema, and repetitive trauma. There are no standard diagnostic tests to determine the presence of biofilms, but there are several clinical indications which can help, and performing a tissue biopsy is better than a swab. Wound bed preparation is a crucial part of biofilm treatment and surgical or conservative sharp wound debridement are the preferred treatments that are effective in removing or reducing biofilms. We report three cases of complex wounds and their treatment.


Attachment, self-worth, and peer-group functioning in middle childhood.
We evaluated links between peer-group functioning and indicators of attachment security in relation to both mother and father in middle childhood, among 73 10-year-olds (37 girls). Children's perceptions of security with both parents, coping styles with mother, and self-worth were assessed. Classmates, teachers, and mothers evaluated the participants' peer-related behavioral characteristics. Children's perceptions of security to both parents were related to others' appraisals of their social competence; perceptions of security to father were related to lower aggression. We did not find child gender effects, but children had higher security scores in relation to mother than to father. Self-worth perceptions mediated the relation between attachment and social competence.


Progress toward improved therapies for inborn errors of metabolism.
Because of their prevalence, severity and lack of effective treatments, inborn errors of metabolism need novel and more effective therapeutic approaches. The opportunity for an early treatment coming from expanded newborn screening has made this need even more urgent. To meet this demand, a growing number of novel treatments are entering in the phase of clinical development. Strategies to overcome the detrimental consequences of the enzyme deficiencies responsible for inborn errors of metabolism have been focused on multiple fronts at the levels of the gene, RNA, protein and whole cell. These strategies have been accomplished using a wide spectrum of approaches ranging from small molecules to enzyme replacement therapy, cell and gene therapy. The applications of new technologies in the field of inborn errors of metabolism, such as genome editing, RNA interference and cell reprogramming, along with progress in pre-existing strategies, such as gene therapy or cell transplantation, have tremendous potential for clinical translation.


Editorial Commentary: Tunnel Widening and Loss of Reduction May Be Inevitable After Acromioclavicular Stabilization With Cortical Suspensory Devices Whether Inset (Top Hats) or Onlay (Buttons).
Despite widely varying techniques, coracoclavicular (CC) stabilization after traumatic, unstable acromioclavicular (AC) joint dislocations shows notable rates of perioperative complications, radiographic loosening, and, ultimately, loss of anatomic reduction. Loss of reduction can occur in upward of 40% of cases after CC repair or reconstruction. Complications and unplanned reoperations can approach 30% and 1.2% to 5.4%, respectively. While we posit that AC joint congruity confers greater shoulder function, anatomic reduction does not always correlate with patient satisfaction or favorable outcomes, which is estimated in up to 88% of surgically treated cases. AC joint repair or reconstruction often uses cortical suspensory button devices to maintain reduction until sufficient soft tissue healing of the CC and AC ligaments. Over time, the so-called windshield wiper phenomenon of the suture-button construct may lead to tunnel osteolysis and, in severe cases, secondary fracture of the coracoid or clavicle. Current efforts to characterize the extent of tunnel widening and ascertain its implications for radiographic outcomes are critical. Whether using inset, low-profile top hat devices or onlay buttons with adjustable loop fixation, loss of AC joint congruity remains unacceptably high. We should strive to more consistently trade a "bump for a scar," not a bump and a scar!


Commitment profiles among nurses: combinations of organizational commitment forms and work engagement, psychological distress, and turnover intention.
To clarify the combined effect of the sub-factors of organizational commitment, this study examined the relationships between organizational commitment profiles and work engagement, psychological distress, and turnover intention among nurses. A cross-sectional survey was conducted; 455 nurses (38 men and 417 women) were included in the statistical analysis. We extracted six clusters through k-means cluster analysis and applied a one-way analysis of variance and χ2 test for work engagement, psychological distress, and turnover intention. Consequently, significant differences were found in work engagement and turnover intention (both p<0.05), and no significant difference was found in psychological distress. These results indicate the formation of affective and normative commitment among nurses in working energetically or preventing turnover. Additionally, no negative effects related to increases in continuance commitment were identified in this study.


The synergistic effect of low-dose cyclosporine and fluocinolone acetonide on the survival of rat allogeneic skin grafts.
Both CsA and topical FA can prolong the survival of skin allografts under the proper conditions. This study was performed to determine if there is a synergistic effect between these two compounds. Buf (RT1b) rat split-thickness skin grafts were transplanted onto the backs of Lew (RT1l) rats. The MST for the control group was 9.89 +/- 0.35 days. In rats given oral CsA, 2.5 or 5 mg/kg, daily from the second day of grafting, the MST was 16.0 +/- 1.9 and 13.6 +/- 0.4 days (blood CsA levels were 166 +/- 20 and 640 +/- 32 ng/ml at the time of rejection, respectively. Topical FA applied daily beginning 72 hr after grafting resulted in a MST of 24.1 +/- 3.6 days. When both topical FA and 5 mg/kg oral CsA were used, the allograft survival time was more than 100 days in 4 of 7 animals. When oral CsA 2.5 mg/kg was combined with topical CsA and FA, the allograft rejection was delayed until 50 days postgrafting in four of six animals. The synergistic effects of oral CsA and topical FA is significant, and thus allows for the use of a subtherapeutic dosage of each compound and provides a potentially safe means for prolonging skin allograft survival.