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Recent experimental work has revealed unusual features of the effect of certain drugs on cardiac inwardly rectifying potassium currents, including the constitutively active and acetylcholine-induced components of acetylcholine-sensitive current (IKAch). These unusual features have included alternating susceptibility of the current components to activation and inhibition induced by ethanol or nicotine applied at various concentrations, and significant correlation between the drug effect and the current magnitude measured under drug-free conditions. To explain these complex drug effects, we have developed a new type of quantitative model to offer a possible interpretation of the effect of ethanol and nicotine on the IKAch channels. The model is based on a description of IKAch as a sum of particular currents related to the populations of channels formed by identical assemblies of different α-subunits. Assuming two different channel populations in agreement with the two reported functional IKAch-channels (GIRK1/4 and GIRK4), the model was able to simulate all the above-mentioned characteristic features of drug-channel interactions and also the dispersion of the current measured in different cells. The formulation of our model equations allows the model to be incorporated easily into the existing integrative models of electrical activity of cardiac cells involving quantitative description of IKAch. We suppose that the model could also help make sense of certain observations related to the channels that do not show inward rectification. This new ionic channel model, based on a concept we call population type, may allow for the interpretation of complex interactions of drugs with ionic channels of various types, which cannot be done using the ionic channel models available so far. [ABSTRACT FROM AUTHOR]

Streptococcus suis is an important porcine bacterial pathogen and a zoonotic agent responsible for sudden death, septic shock and meningitis, of which serotype 2 is the most widespread, with serotype 14 also causing infections in humans in South-East Asia. Knowledge of its pathogenesis and virulence are almost exclusively based on these two serotypes. Though serotype 9 is responsible for the greatest number of porcine cases in Spain, the Netherlands and Germany, very little information is currently available regarding this serotype. Of the different virulence factors, the capsular polysaccharide (CPS) is required for S. suis virulence as it promotes resistance to phagocytosis and killing and masks surface components responsible for host cell activation. However, these roles have been described for serotypes 2 and 14, whose CPSs are structurally and compositionally similar, both containing sialic acid. Consequently, we evaluated herein the interactions of serotype 9 with host cells and the role of its CPS, which greatly differs from those of serotypes 2 and 14. Results demonstrated that serotype 9 adhesion to but not invasion of respiratory epithelial cells was greater than that of serotypes 2 and 14. Furthermore serotype 9 was more internalized by macrophages but equally resistant to whole blood killing. Though recognition of serotypes 2, 9 and 14 by DCs required MyD88-dependent signaling, in vitro pro-inflammatory mediator production induced by serotype 9 was much lower. In vivo, however, serotype 9 causes an exacerbated inflammatory response, which combined with persistent bacterial presence, is probably responsible for host death during the systemic infection. Though presence of the serotype 9 CPS masks surface components less efficiently than those of serotypes 2 and 14, the serotype 9 CPS remains critical for virulence as it is required for survival in blood and development of clinical disease, and this regardless of its unique composition and structure. [ABSTRACT FROM AUTHOR]

The developmental speed of new antimicrobials does not meet the emergence of multidrug-resistant bacteria sufficiently. A potential shortcut is assessing the antimicrobial activity of already approved drugs. Intrudingly, the antibacterial action of glatiramer acetate (GA) has recently been discovered. GA is a well-known and safe immunomodulatory drug particular effective against Gram-negative bacteria, which disrupts biological membranes by resembling the activity of antimicrobial peptides. Thus, GA can potentially be included in treatment strategies used to combat infections caused by multidrug-resistant Gram-negatives. One potential application is chronic respiratory infections caused by Pseudomonas aeruginosa, however the safety of GA inhalation has never been assessed. Here, the safety of inhaling nebulized GA is evaluated in a preclinical pig model. The potential side effects, i.e., bronchoconstriction, respiratory tract symptoms and systemic- and local inflammation were assessed by ventilator monitoring, clinical observation, biochemistry, flowcytometry, and histopathology. No signs of bronchoconstriction assessed by increased airway peak pressure, Ppeak, or decreased oxygen pressure were observed. Also, there were no signs of local inflammation in the final histopathology examination of the pulmonary tissue. As we did not observe any potential pulmonary side effects of inhaled GA, our preliminary results suggest that GA inhalation is safe and potentially can be a part of the treatment strategy targeting chronic lung infections caused by multidrug-resistant Gram-negative bacteria. [ABSTRACT FROM AUTHOR]

The study of protein N-glycosylation is essential in biological and biopharmaceutical research as N-glycans have been reported to regulate a wide range of physiological and pathological processes. Monitoring glycosylation in diagnosis, prognosis, as well as biopharmaceutical development and quality control are important research areas. A number of techniques for the analysis of protein N-glycosylation are currently available. Here we examine three methodologies routinely used for the release of N-glycans, in the effort to establish and standardize glycoproteomics technologies for quantitative glycan analysis from cultured cell lines. N-glycans from human gamma immunoglobulins (IgG), plasma and a pool of four cancer cell lines were released following three approaches and the performance of each method was evaluated. [ABSTRACT FROM AUTHOR]

Background: This study investigated the incidence and predictors of retreatment after discontinuation of either entecavir (ETV) or tenofovir disoproxil fumarate (TDF) treatment in Taiwan. Methods: A total of 535 non-cirrhotic chronic hepatitis B (CHB) patients undergoing either ETV (n = 358) or TDF (n = 177) treatment were enrolled. Patients were followed for at least 12 months after stopping ETV or TDF treatment. Most patients (86.3%) fulfilled the retreatment criteria of Taiwan's National Health Plan. Results: The 5-year cumulative rates of clinical relapse and retreatment were 52.1% and 47%, respectively, in 160 hepatitis B e antigen (HBeAg)-positive patients, and were 62% and 54.8%, respectively, in 375 HBeAg-negative patients. The median duration from the end of treatment until clinical relapse and retreatment was 40 and 57 weeks, respectively, for all patients. Multivariate Cox regression analysis revealed that discontinuing TDF treatment, old age, male gender, and higher baseline HBsAg levels were independent factors of retreatment in HBeAg-positive patients; old age, HBV genotype B, and higher baseline and end-of-treatment HBsAg levels were independent factors in HBeAg-negative patients. A total of 18.8% of retreated patients satisfied the retreatment criteria of hepatic decompensation according to Taiwan's National Health Plan. Of the 64 patients who had clinical relapse without retreatment, 17 achieved sustained virological remission and 26 did not experience clinical relapse until their last visit after clinical relapse. Four patients developed HBsAg loss. Conclusions: The 5-year retreatment rate was about 50% in HBeAg-positive and HBeAg-negative patients. Discontinuing TDF treatment was an independent factor of retreatment in HBeAg-positive patients. [ABSTRACT FROM AUTHOR]

The nicotinic acetylcholine receptor (nAChR) is an excitatory pentameric ligand-gated ion channel (pLGIC), homologous to the inhibitory γ-aminobutyric acid (GABA) type A receptor targeted by pharmaceuticals and endogenous sedatives. Activation of the GABAA receptor by the neurosteroid allopregnanolone can be inhibited competitively by thyroid hormone (L-3,3’,5-triiodothyronine, or T3), but modulation of nAChR by T3 or neurosteroids has not been investigated. Here we show that allopregnanolone inhibits the nAChR from Torpedo californica at micromolar concentrations, as do T3 and the anionic neurosteroid pregnenolone sulfate (PS). We test for the role of protein and ligand charge in mediated receptor inhibition by varying pH in a narrow range around physiological pH. We find that both T3 and PS become less potent with increasing pH, with remarkably similar trends in IC50 when T3 is neutral at pH < 7.3. After deprotonation of T3 (but no additional deprotonation of PS) at pH 7.3, T3 loses potency more slowly with increasing pH than PS. We interpret this result as indicating the negative charge is not required for inhibition but does increase activity. Finally, we show that both T3 and PS affect nAChR channel desensitization, which may implicate a binding site homologous to one that was recently indicated for accelerated desensitization of the GABAA receptor by PS. [ABSTRACT FROM AUTHOR]

Concentrations of free amino acids and [K+] in human sweat can be many times higher than in plasma. Conversely, [Na+] and [Cl-] in sweat are hypotonic to plasma. It was hypothesised that the amino acids and K+ were directly or indirectly associated with the resorption of Na+ and Cl- in the sweat duct. The implication would be that, as resources of these components became limiting during prolonged exercise then the capacity to resorb [Na+] and [Cl-] would diminish, resulting in progressively higher levels in sweat. If this were the case, then [Na+] and [Cl-] in sweat would have inverse relationships with [K+] and the amino acids during exercise. Forearm sweat was collected from 11 recreational athletes at regular intervals during a prolonged period of cycling exercise after 15, 25, 35, 45, 55 and 65 minutes. The subjects also provided passive sweat samples via 15 minutes of thermal stimulation. The sweat samples were analysed for concentrations of amino acids, Na+, Cl-, K+, Mg2+ and Ca2+. The exercise sweat had a total amino acid concentration of 6.4 ± 1.2mM after 15 minutes which was lower than the passive sweat concentration at 11.6 ± 0.8mM (p<0.05) and showed an altered array of electrolytes, indicating that exercise stimulated a change in sweat composition. During the exercise period, [Na+] in sweat increased from 23.3 ± 3.0mM to 34.6 ± 2.4mM (p<0.01) over 65 minutes whilst the total concentrations of amino acids in sweat decreased from 6.4 ± 1.2mM to 3.6 ± 0.5mM. [Na+] showed significant negative correlations with the concentrations of total amino acids (r = -0.97, p<0.05), K+ (r = -0.93, p<0.05) and Ca2+ (r = -0.83, p<0.05) in sweat. The results supported the hypothesis that amino acids and K+, as well as Ca2+, were associated with resorption of Na+ and Cl-. [ABSTRACT FROM AUTHOR]

Raman chemometric urinalysis (Rametrix™) was used to analyze 235 urine specimens from healthy individuals. The purpose of this study was to establish the “range of normal” for Raman spectra of urine specimens from healthy individuals. Ultimately, spectra falling outside of this range will be correlated with kidney and urinary tract disease. Rametrix™ analysis includes direct comparisons of Raman spectra but also principal component analysis (PCA), discriminant analysis of principal components (DAPC) models, multivariate statistics, and it is available through GitHub as the Rametrix™ LITE Toolbox for MATLAB®. Results showed consistently overlapping Raman spectra of urine specimens with significantly larger variances in Raman shifts, found by PCA, corresponding to urea, creatinine, and glucose concentrations. A 2-way ANOVA test found that age of the urine specimen donor was statistically significant (p < 0.001) and donor sex (female or male identification) was less so (p = 0.0526). With DAPC models and blind leave-one-out build/test routines using the Rametrix™ PRO Toolbox (also available through GitHub), an accuracy of 71% (sensitivity = 72%; specificity = 70%) was obtained when predicting whether a urine specimen from a healthy unknown individual was from a female or male donor. Finally, from female and male donors (n = 4) who contributed first morning void urine specimens each day for 30 days, the co-occurrence of menstruation was found statistically insignificant to Rametrix™ results (p = 0.695). In addition, Rametrix™ PRO was able to link urine specimens with the individual donor with an average of 78% accuracy. Taken together, this study established the range of Raman spectra that could be expected when obtaining urine specimens from healthy individuals and analyzed by Rametrix™ and provides the methodology for linking results with donor characteristics. [ABSTRACT FROM AUTHOR]

We identified a mouse strain, HLB444, carrying an N-ethyl-N-nitrosourea (ENU)-induced mutation in a highly conserved C2H2 zinc-finger DNA binding motif of the transcriptional regulator KLF15 that exhibits resistance to diet-induced obesity. Characterization of the HLB444 mutant model on high-fat and chow diets revealed a number of phenotypic differences compared to wild-type controls. When fed a high fat diet, HLB444 had lower body fat, resistance to hepatosteatosis, lower circulating glucose and improved insulin sensitivity compared to C57BL/6J controls. Gut microbial profiles in HLB444 generated from 16S rRNA sequencing of fecal samples differed from controls under both chow and high fat diets. HLB444 shares similar phenotypic traits with engineered full- and adipose-specific Klf15 knockout strains; however, some phenotypic differences between this mutant and the other models suggest that the Klf15 mutation in HLB444 is a hypomorphic variant. The HLB444 model will inform further annotation of transcriptional functions of KLF15, especially with respect to the role of the first zinc-finger domain. [ABSTRACT FROM AUTHOR]

We previously reported that allogeneic, intraperitoneally administered “Neo-Islets,” composed of cultured pancreatic islet cells co-aggregated with high numbers of immunoprotective and cytoprotective Adipose-derived Stem Cells, reestablished, through omental engraftment, redifferentiation and splenic and omental up-regulation of regulatory T-cells, normoglycemia in autoimmune Type-1 Diabetic Non-Obese Diabetic (NOD) mice without the use of immunosuppressive agents or encapsulation devices. Based on these observations, we are currently testing this Neo-Islet technology in an FDA guided pilot study (INAD 012–776) in insulin-dependent, spontaneously diabetic pet dogs by ultrasound-guided, intraperitoneal administration of 2x10e5 Neo-Islets/kilogram body weight to metabolically controlled (blood glucose, triglycerides, thyroid and adrenal functions) and sedated animals. We report here interim observations on the first 4 canine Neo-Islet-treated, insulin-dependent pet dogs that are now in the early to intermediate-term follow-up phase of the planned 3 year study (> 6 months post treatment). Current results from this translational study indicate that in dogs, Neo-Islets appear to engraft, redifferentiate and physiologically produce insulin, and are rejected by neither auto- nor allo-immune responses, as evidenced by (a) an absent IgG response to the allogeneic cells contained in the administered Neo-Islets, and (b) progressively improved glycemic control that achieves up to a 50% reduction in daily insulin needs paralleled by a statistically significant decrease in serum glucose concentrations. This is accomplished without the use of anti-rejection drugs or encapsulation devices. No adverse or serious adverse events related to the Neo-Islet administration have been observed to date. We conclude that this minimally invasive therapy has significant translational relevance to veterinary and clinical Type 1 diabetes mellitus by achieving complete and at this point partial glycemic control in two species, i.e., diabetic mice and dogs, respectively. [ABSTRACT FROM AUTHOR]

In the course of the Maillard reaction, which occurs during heating of food but also under physiological condition, a broad spectrum of reaction products is formed. Among them, the advanced glycation endproducts (AGEs) Nε-carboxymethyllysine (CML), pyrraline (Pyr), methylglyoxal-derived hydroimidazolone 1 (MG-H1) and Nε-carboxyethyllysine (CEL) are the quantitatively dominating compounds during later reaction stages. Those dietary glycation compounds are under discussion as to be associated with chronic inflammation and the pathophysiological consequences of diseases such as diabetes. In the present study, the concentration of individual glycation compounds in saliva was monitored for the first time and related to their dietary uptake. Fasting saliva of 33 metabolically healthy subjects was analyzed with HPLC-MS/MS. The observed levels of individual glycation compounds ranged from 0.5 to 55.2 ng/ml and differed both intra- and interindividually. Patterns did not correlate with subject-related features such as vegetarianism or sports activities, indicating that dietary intake may play an important role. Therefore, six volunteers were asked to eat a raw food diet free of glycation compounds for two days. Within two days, salivary Pyr was lowered from median 1.7 ng/ml to a minimum level below the limit of detection, and MG-H1 decreased from 3.6 to 1.7 ng/ml in in a time-dependent manner after two days. Salivary CML and CEL concentrations were not affected. Therefore, measuring Pyr and MG-H1 in saliva is a suitable diagnostic tool to monitor the dietary intake and metabolic transit of glycation compounds present in heated foods. [ABSTRACT FROM AUTHOR]

Comparing the efficacy of inhaled antibiotics can be difficult in small groups of patients with cystic fibrosis and mild lung disease. In a feasibility study we compared Aztreonam lysine for inhalation solution (AZLI; Cayston®) to standard inhaled antibiotic therapy in patients with cystic fibrosis and near normal spirometry. To detect treatment responses we used both lung clearance index (LCI) and forced expiratory volume in one second (FEV1). At baseline, median FEV1 was 87% pred. and median LCI was 8.6 (upper limit of normal: 7.0). After 4 weeks, LCI improved by -0.36 after AZLI and deteriorated by +0.12 after tobramycin treatment (p = 0.039). No significant differences between treatments (p = 0.195) were observed using FEV1. These results suggest that lung clearance index can be used to detect treatment induced changes in subjects with mild lung disease. [ABSTRACT FROM AUTHOR]

Objective: The mechanism by which G-protein-coupled receptor 40 (GPR40) signaling amplifies glucose-stimulated insulin secretion through activation of protein kinase C (PKC) is unknown. We examined whether a GPR40 agonist, GW9508, could stimulate conventional and novel isoforms of PKC at two glucose concentrations (3 mM and 20 mM) in INS-1D cells. Methods: Using epifluorescence microscopy, we monitored relative changes in the cytosolic fluorescence intensity of Fura2 as a marker of change in intracellular Ca2+ ([Ca2+]i) and relative increases in green fluorescent protein (GFP)-tagged myristoylated alanine-rich C kinase substrate (MARCKS-GFP) as a marker of PKC activation in response to GW9508 at 3 mM and 20 mM glucose. To assess the activation of the two PKC isoforms, relative increases in membrane fluorescence intensity of PKCα-GFP and PKCε-GFP were measured by total internal reflection fluorescence microscopy. Specific inhibitors of each PKC isotype were constructed and synthesized as peptide fusions with the third α-helix of the homeodomain of Antennapedia. Results: At 3 mM glucose, GW9508 induced sustained MARCKS-GFP translocation to the cytosol, irrespective of changes in [Ca2+]i. At 20 mM glucose, GW9508 induced sustained MARCKS-GFP translocation but also transient translocation that followed sharp increases in [Ca2+]i. Although PKCα translocation was rarely observed, PKCε translocation to the plasma membrane was sustained by GW9508 at 3 mM glucose. At 20 mM glucose, GW9508 induced transient translocation of PKCα and sustained translocation as well as transient translocation of PKCε. While the inhibitors (75 μM) of each PKC isotype reduced GW9508-potentiated, glucose-stimulated insulin secretion in INS-1D cells, the PKCε inhibitor had a more potent effect. Conclusion: GW9508 activated PKCε but not PKCα at a substimulatory concentration of glucose. Both PKC isotypes were activated at a stimulatory concentration of glucose and contributed to glucose-stimulated insulin secretion in insulin-producing cells. [ABSTRACT FROM AUTHOR]

Functional variability among boys with Duchenne muscular dystrophy (DMD) is well recognised and complicates interpretation of clinical studies. We hypothesised that boys with DMD could be clustered into groups sharing similar trajectories of ambulatory function over time, as measured by the North Star Ambulatory Assessment (NSAA) total score. We also explored associations with other variables such as age, functional abilities, and genotype. Using the NorthStar Clinical Network database, 395 patients with >1 NSAA assessment were identified. We utilised latent class trajectory analysis of longitudinal NSAA scores, which produced evidence for at least four clusters of boys sharing similar trajectories versus age in decreasing order of clinical severity: 25% of the boys were in cluster 1 (NSAA falling to ≤ 5 at age ~10y), 35% were in cluster 2 (NSAA ≤ 5 ~12y), 21% in were cluster 3 (NSAA≤ 5 ~14y), and 19% in cluster 4 (NSAA > 5 up to 15y). Mean ages at diagnosis of DMD were similar across clusters (4.2, 3.9, 4.3, and 4.8y, respectively). However, at the first NSAA assessment, a significant (p<0.05) association was observed between earlier declining clusters and younger age, worse NSAA, slower rise from supine, slower 10 metre walk/run times, and younger age of steroid initiation. In order to assess the probability of observing complete loss of function for individual NSAA items, we examined the proportion of patients who shifted from a score of 1 or 2 at baseline to a score of 0. We also assessed the probability of gain of function using the inverse assessment and stratified the probability of deterioration, improvement–or static behavior–by age ranges and using baseline functional status. Using this tool, our study provides a comprehensive assessment of the NSAA in a large population of patients with DMD and, for the first time, describes discrete clusters of disease progression; this will be invaluable for future DMD clinical trial design and interpretation of findings. [ABSTRACT FROM AUTHOR]

Background: In Fetal Growth Restriction ‘fetal programming’ may take place via DNA methylation, which has implications for short-term and long-term health outcomes. Small-for-gestational age fetuses are considered fetal growth restricted, characterized by brain-sparing when fetal Doppler hemodynamics are abnormal, expressed as a cerebroplacental ratio (CPR) <1. We aimed to determine whether brain-sparing is associated with altered DNA methylation of selected genes. Methods: We compared DNA methylation of six genes in 41 small-for-gestational age placentas with a normal or abnormal CPR. We selected EPO, HIF1A, VEGFA, LEP, PHLDA2, and DHCR24 for their role in angiogenesis, immunomodulation, and placental and fetal growth. DNA methylation was analyzed by pyrosequencing. Results: Growth restricted fetuses with an abnormal CPR showed hypermethylation of the VEGFA gene at one CpG (VEGFA-309, p = .001) and an overall hypomethylation of the LEP gene, being significant at two CpGs (LEP-123, p = .049; LEP-51, p = .020). No differences in methylation were observed for the other genes. Conclusions: VEGFA and LEP genes are differentially methylated in placentas of small-for-gestational age fetuses with brain-sparing. Hypermethylation of VEGFA-309 in abnormal CPR-placentas could indicate successful compensatory mechanisms. Methylation of LEP-51 is known to suppress LEP expression. Hypomethylation in small-for-gestational age placentas with abnormal CPR may result in hyperleptinemia and predispose to leptin-resistance later in life. [ABSTRACT FROM AUTHOR]

The gastric pathogen Helicobacter pylori requires a noncanonical cytosolic chemoreceptor transducer-like protein D (TlpD) for efficient colonization of the mammalian stomach. Here, we reconstituted a complete chemotransduction signaling complex in vitro with TlpD and the chemotaxis (Che) proteins CheW and CheA, enabling quantitative assays for potential chemotaxis ligands. We found that TlpD is selectively sensitive at micromolar concentrations to bleach (hypochlorous acid, HOCl), a potent antimicrobial produced by neutrophil myeloperoxidase during inflammation. HOCl acts as a chemoattractant by reversibly oxidizing a conserved cysteine within a 3His/1Cys Zn-binding motif in TlpD that inactivates the chemotransduction signaling complex. We found that H. pylori is resistant to killing by millimolar concentrations of HOCl and responds to HOCl in the micromolar range by increasing its smooth-swimming behavior, leading to chemoattraction to HOCl sources. We show related protein domains from Salmonella enterica and Escherichia coli possess similar reactivity toward HOCl. We propose that this family of proteins enables host-associated bacteria to sense sites of tissue inflammation, a strategy that H. pylori uses to aid in colonizing and persisting in inflamed gastric tissue. [ABSTRACT FROM AUTHOR]

A persistent obstacle for constructing kinetic models of metabolism is uncertainty in the kinetic properties of enzymes. Currently, available methods for building kinetic models can cope indirectly with uncertainties by integrating data from different biological levels and origins into models. In this study, we use the recently proposed computational approach iSCHRUNK (n ilico Approach to aracterization and eduction of certainty in the inetic Models), which combines Monte Carlo parameter sampling methods and machine learning techniques, in the context of Bayesian inference. Monte Carlo parameter sampling methods allow us to exploit synergies between different data sources and generate a population of kinetic models that are consistent with the available data and physicochemical laws. The machine learning allows us to data-mine the a priori generated kinetic parameters together with the integrated datasets and derive posterior distributions of kinetic parameters consistent with the observed physiology. In this work, we used iSCHRUNK to address a design question: can we identify which are the kinetic parameters and what are their values that give rise to a desired metabolic behavior? Such information is important for a wide variety of studies ranging from biotechnology to medicine. To illustrate the proposed methodology, we performed Metabolic Control Analysis, computed the flux control coefficients of the xylose uptake (XTR), and identified parameters that ensure a rate improvement of XTR in a glucose-xylose co-utilizing S. cerevisiae strain. Our results indicate that only three kinetic parameters need to be accurately characterized to describe the studied physiology, and ultimately to design and control the desired responses of the metabolism. This framework paves the way for a new generation of methods that will systematically integrate the wealth of available omics data and efficiently extract the information necessary for metabolic engineering and synthetic biology decisions. [ABSTRACT FROM AUTHOR]

L-5-methyltetrahydrofolate is the predominant folate form in human milk but is currently not approved as a folate source for infant and follow-on formula. We aimed to assess the suitability of L-5-methyltetrahydrofolate as a folate source for infants. Growth and tolerance in healthy term infants fed formulae containing equimolar doses of L-5-methyltetrahydrofolate (10.4 μg/ 100 ml, n = 120, intervention group) or folic acid (10.0 μg/ 100 ml, n = 120, control group) was assessed in a randomized, double-blind, parallel, controlled trial. A reference group of breastfed infants was followed. Both formulae were well accepted without differences in tolerance or occurrence of adverse events. The most common adverse events were common cold, poor weight gain or growth, rash, eczema, or dry skin and respiratory tract infection. Weight gain (the primary outcome) was equivalent in the two groups (95% CI -2.11; 1.68 g/d). In line with this, there was only a small difference in absolute body weight adjusted for birth weight and sex at visit 4 (95% CI -235; 135 g). Equivalence was also shown for gain in head circumference but not for recumbent length gain and increase in calorie intake. Given the nature of the test, this does not indicate an actual difference, and adjusted means at visit 4 were not significantly different for any of these parameters. Infants receiving formula containing L-5-methyltetrahydrofolate had lower mean plasma levels of unmetabolized folic acid (intervention: 0.73 nmol/L, control: 1.15 nmol/L, p<0.0001) and higher levels of red cell folate (intervention: 907.0 ±192.8 nmol/L, control: 839.4 ±142.4 nmol/L, p = 0.0095). We conclude that L-5-methyltetrahydrofolate is suitable for use in infant and follow-on formula, and there are no indications of untoward effects. Trial registration: This trial was registered at ClinicalTrials.gov (). [ABSTRACT FROM AUTHOR]

Background: Accurate and accessible diagnosis is key for the control of visceral leishmaniasis (VL). Yet, current diagnostic tests for VL have severe limitations: they are invasive or not suitable as point of care (POC) test or their performance is suboptimal in East Africa. We analysed the antigens in the VL serodiagnostics development pipeline to identify shortcomings and to propose strategies in the development of an alternative POC test for VL in East Africa. Objectives: The objective of this study was to identify and to analyse all antigens for VL serodiagnosis that have been published before 2018 in order to identify candidates and gaps in the pipeline for a new POC test in East Africa. Methods: A systematic literature search was performed on PubMed for original research articles on Leishmania-specific antigens for antibody detection of VL in humans. From each article, the following information was extracted: the antigen name, test format and characteristics, its reported sensitivity and specificity and study cohort specifications. Results: One hundred and seven articles containing information about 96 tests based on 89 different antigens were included in this study. Eighty six of these tests, comprising 80 antigens, were evaluated in phase I and II studies only. Only 20 antigens, all of which are native, contain a carbohydrate and/or lipid moiety. Twenty-four antigens, of which 7 are non-native, are composed of antigen mixtures. Nineteen tests, comprising 18 antigens, have been evaluated on East African specimens, of which only 2 (rK28 based immunochromatographic test and intact promastigote based indirect fluorescent antibody technique) consistently showed sensitivities above 94 and specificities above 97% in a phase III study and one in a phase II study (dot blot with SLA). Only rK28 is a non-native mixture of antigens which we consider suitable for further evaluation and implementation. Conclusions: The development pipeline for an alternative serodiagnostic test for VL is almost empty. Most antigens are not sufficiently evaluated. Non-protein antigens and antigen mixtures are being neglected. We propose to expand the evaluation of existing antigen candidates and to investigate the diagnostic potential of defined non-native carbohydrate and lipid antigens for VL serodiagnosis in East Africa. [ABSTRACT FROM AUTHOR]

Human fumarylacetoacetate hydrolase (FAH) domain containing protein 1 (FAHD1) is a mitochondrial oxalocatate decarboxylase, the first of its kind identified in eukaryotes. The physiological role of FAHD1 in other eukaryotes is still poorly understood. In C. elegans loss of the FAHD1 ortholog FAHD-1 was reported to impair mitochondrial function, locomotion and egg-laying behavior, yet the underlying mechanisms remained unclear. Using tissue-specific rescue of fahd-1(-) worms, we find that these phenotypic abnormalities are at least in part due to fahd-1’s function in neurons. Moreover, we show that egg-laying defects in fahd-1(-) worms can be fully rescued by external dopamine administration and that depletion of fahd-1 expression induces expression of several enzymes involved in serotonin biosynthesis. Together, our results support a role for fahd-1 in modulating serotonin levels and suggest this protein as a novel link between metabolism and neurotransmitter signaling in the nervous system. Finally, we propose a model to explain how a metabolic defect could ultimately lead to marked changes in neuronal signaling. [ABSTRACT FROM AUTHOR]

The energy expenditure and substrate utilisation were measured in 5 men pre- and post- a 67 day, 1750km unassisted Antarctic traverse from the Hercules Inlet to the Ross Sea Ice via the South pole pulling sledges weighing 120kg whilst experiencing temperatures as low as -57°C. A 36-hours protocol in a whole body calorimeter was employed to measure periods of rest, sleep and three periods of standardised stepping exercises at 80, 100 and 120 steps min-1; participants were fed isocalorically. Unlike previous expeditions where large weight loss was reported, only a modest loss of body weight (7%, P = 0.03) was found; fat tissue was reduced by 53% (P = 0.03) together with a small, but not statistically significant, increase in lean tissue weight (P = 0.18). This loss occurred despite a high-energy intake (6500 kcal/day) designed to match energy expenditure. An energy balance analysis suggested the loss in body weight could be due to the energy requirements of thermoregulation. Differences in energy expenditure [4.9 (0.1) vs 4.5 (0.1) kcal/min. P = 0.03], carbohydrate utilisation [450 (180) vs 569 (195) g/day; P = 0.03] and lipid utilisation [450 (61) vs 388 (127) g/day, P = 0.03] at low levels of exertion were different from pre-expedition values. Only carbohydrate utilisation remained statistically significant when normalised to body weight. The differences in energy expenditure and substrate utilisation between the pre- and post-expedition for other physiological states (sleeping, resting, higher levels of exercise, etc) were small and not statistically significant. Whilst inter-subject variability was large, there was a tendency for increased carbohydrate utilisation, post-expedition, when fasted that decreased upon feeding. [ABSTRACT FROM AUTHOR]

During the lactation period, rat pups are fed by the dam, and the patterns of mother-pup interaction change during this period. Additionally, there are changes in feeding; first, mother´s milk is the only food needed for sustenance, and later, it is combined with solid food and water. GH serum concentrations depend on both maternal-pup interaction and energy metabolism. In the artificial rearing (AR) procedure, pups are deprived of mother-pup interaction, and the feeding pattern is controlled. This rearing paradigm has been used in rats to analyze the effects of maternal deprivation on social behavior. In the present study, we analyzed the variation in GH, acylated ghrelin and IGF-1 serum concentrations throughout the lactation period in AR pups. At pnd7, the maternal rearing (MR) pups responded to a 4 h fast with a drop in GH serum concentration, which is a well-known response to maternal deprivation. GH serum levels in the AR pups did not change, suggesting an adaptation phenomenon. A dopamine inhibitory effect of GH secretion was observed in pnd7 cultured somatotropes, suggesting dopamine regulation of GH secretion at this age. Acylated ghrelin serum levels in the AR pups showed an inverted pattern compared to that in the MR pups, which was related to the artificial feeding pattern. IGF-1 serum levels were lower in the AR pups than in MR pups, which was associated with hepatic GH resistance and with low Igf1 mRNA expression at pnd7. Interestingly, at pnd14, both pup groups showed high hepatic Igf1 mRNA expression but low IGF-1 serum levels, and this was inverted at pnd21. However, serum glucose levels were lower in the AR pups at pnd14 but reached the same levels as the MR pups at pnd21. Moreover, hepatomegaly and higher hepatic GH-receptor levels were observed in the AR pups at pnd21, which was in agreement with an absence of a solid food meal. During AR, the pups lost the maternal interaction-stimulated GH secretion, which correlated with lower IGF-1 serum levels during the first week of postnatal development. Later, the AR pups exhibited hepatic responses, in order to satisfy the metabolic demand for the normal weaning, with low carbohydrates levels in their meal. [ABSTRACT FROM AUTHOR]

Klotho is a soluble or membrane-bound anti-aging protein, whose protective actions are important for a prudential function of many organs. Because Klotho and cerebral small vessel disease (SVD) are associated with ageing process and endothelial dysfunction, it is possible that Klotho has an association with cerebral SVD. We aimed to investigate the association of plasma Klotho concentration with the presence, burden and progression of cerebral SVD. We prospectively enrolled 262 patients with first-ever acute cerebral infarction, performed brain MRI and collected their blood samples within 24 hours of admission. An enzyme-linked immunosorbent assay was used for evaluating plasma Klotho concentration. We estimated the total SVD score of each patient after determining the presence and burden of high-grade white matter hyperintensities (HWMHs), cerebral microbleeds (CMBs), high-grade perivascular spaces (HPVSs) and asymptomatic lacunar infarctions (ALIs). Univariate and multivariate analyses were conducted to investigate association of Klotho with cerebral SVD and the total SVD score. Of the 262 patients, 152 (58.0%) were men. The mean age of these patients was 64.7 years. The mean ± standard deviation of plasma Klotho concentration was 329.8 ± 194.1 pg/mL. In multivariate analysis, plasma Klotho concentration was negatively associated with the presence of HWMHs [Odds ratio (OR): 0.13, p = 0.047], HPVSs (OR: 0.22, p = 0.024) and ALIs (OR: 0.53, p = 0.021) but not associated with the presence of CMBs (OR: 0.39, p = 0.404). Plasma Klotho concentration was also negatively related to the total SVD score (unstandardized coefficients beta: −0.895, standard error = 0.317, p = 0.005, R2 = 0.239). Furthermore, plasma Klotho concentration was negatively related to the presence (OR: 0.75, 95% CI: 0.59–0.96, p = 0.025) and severity of cerebral SVD progression (OR: 0.72, 95% CI: 0.56–0.92, p = 0.009). In conclusion, plasma Klotho concentration was negatively associated with the presence, burden and progression of cerebral SVD. [ABSTRACT FROM AUTHOR]

Aberrant DNA methylation patterns are common in cancers and environmental pollutant exposed subjects. Up to date, few studies have examined the aberrant DNA methylation patterns in benzene exposed workers. We recruited 141 benzene-exposed workers, including 83 benzene-exposed workers from a shoe factory in Wenzhou and 58 workers from a painting workshop in Wuhu, 35 workers in Wuhu were followed from 2009 to 2013, and 48 indoor workers as controls from Wenzhou. We used high-resolution melting (HRM) to quantitate human samples of DNA methylation in long interspersed nuclear element-1 (LINE-1), (6)-methylguanine-DNA methyltransferase (MGMT), and DNA mismatch repair gene human mutator L homologue 1 (hMLH1). AML-5 cells were treated with benzoquinone (BQ) and hydroquinone (HQ), and the promoter methylation of MGMT and hMLH1 was detected using the bisulfite sequencing PCR method. The degree of LINE-1 methylation in benzene-exposed workers was significantly lower than that of the controls (p<0.001), and the degree of MGMT (p<0.001) and hMLH1 (p = 0.01) methylation was significantly higher than that of the controls. The in vitro study validated the aberrant hypermethylation of hMLH1 after treatment with BQ. Among the cohort workers who were followed from 2009 to 2013, the LINE1 methylation elevated in 2013 than 2009 (p = 0.004), and premotor methylation in hMLH1 reduced in 2013 than 2009 (p = 0.045) with the reduction of the benzene exposure. This study provides evidence that benzene exposure can induce LINE-1 hypomethylation and DNA repair gene hypermethylation. [ABSTRACT FROM AUTHOR]

Although the literature on the Maillard reaction in infant formulas is extensive, most studies have focused on model systems, and in only a few cases on real food systems. Therefore, the objective of the present study was to determine the status of the Maillard reaction, both the early and advanced phases, in a variety of commercial infant formulas available on the Swedish market. Ten powder and liquid milk-based infant formulas from three manufacturers were selected to determine available lysine and CML contents, the two established indicators of the reaction. The products were also characterized with respect to protein content, carbohydrates composition, water content and water activity. In order to be able to compare the impact of different processing steps applied on powder and liquid formulas, the solid formulas contained similar ingredients as their corresponding liquid ones. Our findings showed that powder and liquid formulas contained similar available lysine concentrations regardless of the manufacturer, showing 27.14–36.57% decrease in the available lysine, compared to the reference skim milk powder in this study. The CML concentrations were in a broad range of 68.77–507.99 mg / kg protein. In the case of one manufacturer, liquid infant formulas had significantly higher CML content, compared to the powder products (p < 0.05). The results from this study are a step taken towards better understanding of the extent of the Maillard reaction in real complex systems of infant formulas. [ABSTRACT FROM AUTHOR]

Glycoconjugation to target the Warburg effect provides the potential to enhance selective uptake of anticancer or imaging agents by cancer cells. A Warburg effect targeting group, rationally designed to facilitate uptake by glucose transporters and promote cellular accumulation due to phosphorylation by hexokinase (HK), has been synthesised. This targeting group, the C2 modified glucose analogue 2-(2-[2-(2-aminoethoxy)ethoxy]ethoxy)-D-glucose, has been conjugated to the fluorophore nitrobenzoxadiazole to evaluate its effect on uptake and accumulation in cancer cells. The targeting vector has demonstrated inhibition of glucose phosphorylation by HK, indicating its interaction with the enzyme and thereby confirming the potential to facilitate an intracellular trapping mechanism for compounds it is conjugated with. The cellular uptake of the fluorescent analogue is dependent on the glucose concentration and is so to a greater extent than is that of the widely used fluorescent glucose analogue, 2-NBDG. It also demonstrates selective uptake in the hypoxic regions of 3D spheroid tumour models whereas 2-NBDG is distributed primarily through the normoxic regions of the spheroid. The increased selectivity is consistent with the blocking of alternative uptake pathways. [ABSTRACT FROM AUTHOR]

The biopharmaceutical industry is growing at a fast pace, making nowadays 20% of the pharma market. Within this market, therapeutic monoclonal antibodies (mAbs) are the dominant product class. With the patent expirations, biosimilars and, perhaps more relevant, biobetters, are in fast development. Thus, a comprehensive characterization at the molecular level of antibodies heterogeneity such as glycoforms, post-translational modifications (PTMs) and sequence variations is of utmost importance. Mass spectrometry (MS)-based approaches are undoubtedly the most powerful analytical strategies to monitor and define an array of critical quality attributes on mAbs. In this work, we demonstrate the analytical power of the Q-TOF MS platform for comprehensive and detailed analysis at molecular levels of an in-house produced mAb. This methodology involves minimal sample preparation procedures and provides an extensive collection of valuable data in a short period of time. [ABSTRACT FROM AUTHOR]

IgA nephropathy, a common chronic kidney disease, has various possible outcomes. Therefore, the identification of novel prognostic biomarkers is needed. To this purpose, we used gas chromatography mass spectrometry to search for metabolites capable of predicting the phenotypes of IgA nephropathy in hyper IgA (HIGA) mice, an established model mice for IgA nephropathy. We measured the plasma metabolite levels in 12- and 22-week-old mice, prior to the manifestation of IgA nephropathy phenotypes, and statistically investigated the associations between these metabolites and the phenotypes of IgA nephropathy, such as the urine protein levels and histological phenotypes of the kidney at 32 weeks. We observed that in plasma samples collected from 12- and 22-week-old HIGA mice, the urinary protein levels were significantly associated with 8 and 10 metabolites, the glomerular cellular component levels were significantly associated with 8 and 7 metabolites, and the mesangial substrate levels were significantly associated with 8 and 8 metabolites, respectively. Among the candidate metabolites associated with the phenotypes of IgA nephropathy, coniferyl alcohol levels were significantly higher in HIGA mice at all of the 12, 22, and 32 weeks of age. Since this study was an observational study, we could not elucidate the underlying mechanisms; however, we were able to identify new candidate metabolites, such as coniferyl alcohol, as being potentially involved in the pathogenesis of IgA nephropathy. These results might help to develop novel laboratory tests and therapeutic reagents for IgA nephropathy in the future. [ABSTRACT FROM AUTHOR]

Despite having been sequenced over a decade ago, the functional significance of much of the mammalian genome remains unknown. The mouse has become the preeminent mammalian model for identifying endogenous gene function in vivo. Here we characterize the phenotype of a loss-of function allele for the evolutionarily conserved transcription factor, Elongation Factor Homolog 1 (Elof1). Recent work utilizing the yeast homolog, Elf1, has demonstrated that Elf1 associates with the RNA polymerase II complex to promote elongation by relieving the association of the template DNA strand with bound histones. Loss of Elof1 results in developmental delay and morphological defects during early mouse development resulting in peri-gastrulation lethality. Although Elof1 is highly conserved we observe tissue specific expression during gastrulation and in adult murine tissues, suggesting there may be other genes with similar function in diverse tissues or that mElof1 has adopted lineage specific functions. To better understand its function in mammalian transcription, we examined splice variants and find that Elof1 regulates mutually exclusive exon use in vivo. Distinct from what has been demonstrated in yeast, we demonstrate that Elof1 is essential for viability during mammalian gastrulation which may be due to a role mediating tissue specific exclusive exon use, a regulatory function unique to higher eukaryotes. [ABSTRACT FROM AUTHOR]

In insects, the gustatory system has a critical function not only in selecting food and feeding behaviours but also in growth and metabolism. Gustatory receptors play an irreplaceable role in insect gustatory signalling. Trichogramma chilonis is an effective biocontrol agent against agricultural insect pests. However, the molecular mechanism of gustation in T. chilonis remains elusive. In this study, we found that T. chilonis adults had a preference for D-fructose and that D-fructose contributed to prolong longevity and improve fecundity. Then, We also isolated the full-length cDNA encoding candidate gustatory receptor (TchiGR43a) based on the transcriptome data of T. chilonis, and observed that the candidate gustatory receptor gene was expressed from the larval to adult stages. The expression levels of TchiGR43a were similar between female and male. A Xenopus oocyte expression system and two-electrode voltage-clamp recording further verified the function analysis of TchiGR43a. Electrophysiological results showed that TchiGR43a was exclusively tuned to D-fructose. By the studies of behaviour, molecular biology and electrophysiology in T. chilonis, our results lay a basic fundation of further study on the molecular mechanisms of gustatory reception and provide theoretical basis for the nutritional requirement of T. chilonis in biocontrol. [ABSTRACT FROM AUTHOR]

Background: Systematic reviews of randomised controlled trials (RCTs) have suggested that maternal vitamin D (25[OH]D) and calcium supplementation increase birth weight. However, limitations of many trials were highlighted in the reviews. Our aim was to combine genetic and RCT data to estimate causal effects of these two maternal traits on offspring birth weight.Methods and Findings: We performed two-sample mendelian randomisation (MR) using genetic instrumental variables associated with 25(OH)D and calcium that had been identified in genome-wide association studies (GWAS; sample 1; N = 122,123 for 25[OH]D and N = 61,275 for calcium). Associations between these maternal genetic variants and offspring birth weight were calculated in the UK Biobank (UKB) (sample 2; N = 190,406). We used data on mother-child pairs from two United Kingdom birth cohorts (combined N = 5,223) in sensitivity analyses to check whether results were influenced by fetal genotype, which is correlated with the maternal genotype (r ≈ 0.5). Further sensitivity analyses to test the reliability of the results included MR-Egger, weighted-median estimator, 'leave-one-out', and multivariable MR analyses. We triangulated MR results with those from RCTs, in which we used randomisation to supplementation with vitamin D (24 RCTs, combined N = 5,276) and calcium (6 RCTs, combined N = 543) as an instrumental variable to determine the effects of 25(OH)D and calcium on birth weight. In the main MR analysis, there was no strong evidence of an effect of maternal 25(OH)D on birth weight (difference in mean birth weight -0.03 g [95% CI -2.48 to 2.42 g, p = 0.981] per 10% higher maternal 25[OH]D). The effect estimate was consistent across our MR sensitivity analyses. Instrumental variable analyses applied to RCTs suggested a weak positive causal effect (5.94 g [95% CI 2.15-9.73, p = 0.002] per 10% higher maternal 25[OH]D), but this result may be exaggerated because of risk of bias in the included RCTs. The main MR analysis for maternal calcium also suggested no strong evidence of an effect on birth weight (-20 g [95% CI -44 to 5 g, p = 0.116] per 1 SD higher maternal calcium level). Some sensitivity analyses suggested that the genetic instrument for calcium was associated with birth weight via exposures that are independent of calcium levels (horizontal pleiotropy). Application of instrumental variable analyses to RCTs suggested that calcium has a substantial effect on birth weight (178 g [95% CI 121-236 g, p = 1.43 × 10-9] per 1 SD higher maternal calcium level) that was not consistent with any of the MR results. However, the RCT instrumental variable estimate may have been exaggerated because of risk of bias in the included RCTs. Other study limitations include the low response rate of UK Biobank, which may bias MR estimates, and the lack of suitable data to test whether the effects of genetic instruments on maternal calcium levels during pregnancy were the same as those outside of pregnancy.Conclusions: Our results suggest that maternal circulating 25(OH)D does not influence birth weight in otherwise healthy newborns. However, the effect of maternal circulating calcium on birth weight is unclear and requires further exploration with more research including RCT and/or MR analyses with more valid instruments. [ABSTRACT FROM AUTHOR]

The signaling lymphocytic activation molecule (SLAM) family is comprised of nine distinct receptors that are expressed exclusively on hematopoietic cells. Most of these transmembrane receptors are homotypic by nature and downstream signaling occurs when cells that express the same SLAM receptor interact. Previous studies have determined that anti-SLAMF6 antibodies can have a therapeutic effect in autoimmunity and cancer. However, little is known about the role of SLAMF6 in the adaptive immune responses and in order to utilize SLAMF6 interventional approaches, a better understanding of the biology of this receptor in T cell is warranted. Accordingly, the objective of our study was to investigate both functionally and structurally the role of SLAMF6 in T cell receptor (TCR) mediated responses. Biochemical and genetic experiments revealed that SLAMF6 was required for productive TCR downstream signaling. Interestingly, SLAMF6 ectodomain was required for its function, but not for its recruitment to the immunological synapse. Flow-cytometry analysis demonstrated that tyrosine 308 of the tail of SLAMF6 was crucial for its ability to enhance T cell function. Imaging studies revealed that SLAMF6 clustering, specifically with the TCR, resulted in dramatic increase in downstream signaling. Mechanistically, we showed that SLAMF6 enhanced T cell function by increasing T cell adhesiveness through activation of the small GTPase Rap1. Taken together SLAMF6 is an important regulator of T cell activation where both its ectodomain and its endodomain contribute differentially to T cell functions. Additional studies are underway to better evaluate the role of anti-SLAMF6 approaches in specific human diseases. [ABSTRACT FROM AUTHOR]

Protein tyrosine phosphatase receptor type Z (PTPRZ) is preferentially expressed in the central nervous system as two transmembrane receptor isoforms PTPRZ-A/B and one secretory isoform PTPRZ-S. Ptprz-knockout mice lacking the expression of all three isoforms show behavioral, learning, and neurological abnormalities, including increased exploratory activities to novelty, deficits in spatial and contextual learning, and reduced responses to methamphetamine, relative to wild-type mice. To investigate whether PTPRZ isoforms play distinct physiological roles, we herein performed behavioral studies on two knock-in mouse lines: One expresses the catalytically inactive Cys-1930 to Ser (CS) mutants of PTPRZ-A/B, while the other generated in the present study expresses catalytically active mutants of PTPRZ-A/B lacking the negative regulatory PTP-D2 domain and C-terminal PDZ-binding motif (ΔD2) instead of wild-type PTPRZ-A/-B. In contrast to Ptprz-knockout mice, neither increased responses to novelty in the open field nor memory impairments in the inhibitory-avoidance task were observed in Ptprz-CS or Ptprz-ΔD2 mice. However, the effects of methamphetamine on locomotor activity were significantly weaker in Ptprz-KO mice and CS mutant mice than in wild-type mice, but were normal in ΔD2 mutant mice. Furthermore, microdialysis experiments revealed that methamphetamine-evoked dopamine release in the nucleus accumbens was reduced in Ptprz-KO mice and CS mutant mice. These results suggest that the extracellular region of PTPRZ, including the secretory isoform, is crucial for behavioral responses to novelty and the formation of aversive memories, whereas the PTPase activities of PTPRZ receptor isoforms are involved in regulating the dopaminergic system. [ABSTRACT FROM AUTHOR]

Poly(N,N-dimethylaminoethyl methacrylate) (PDMAEMA) is a promising quite new polymer with very interesting properties. The thermal degradation process of PDMAEMA was investigated. The polymer was heated at specific time intervals, then heating was stopped, and infrared analysis was performed to obtain information on the structure of the solid residue. The thermal degradation process has a two-stage character. The limit temperature for the first decomposition step was about 390°C, after which the second stage of sample decomposition began. The order of disintegration of the macromolecules was determined. Activation energy values for the thermal decomposition process have been calculated; they are 89.8 kJ/mol for the first stage and 17.7 kJ/mol for the second stage of the degradation process. [ABSTRACT FROM AUTHOR]

Mannitol, a sugar alcohol used in commercial food products, has been previously shown to induce sex-biased mortality in female Drosophila melanogaster when ingested at a single concentration (1 M). We hypothesized that sex differences in energy needs, related to reproductive costs, contributed to the increased mortality we observed in females compared to males. To test this, we compared the longevity of actively mating and non-mating flies fed increasing concentrations of mannitol. We also asked whether mannitol-induced mortality was concentration-dependent for both males and females, and if mannitol’s sex-biased effects were consistent across concentrations. Females and males both showed concentration-dependent increases in mortality, but female mortality was consistently higher at concentrations of 0.75 M and above. Additionally, fly longevity decreased further for both sexes when housed in mixed sex vials as compared to single sex vials. This suggests that the increased energetic demands of mating and reproduction for both sexes increased the ingestion of mannitol. Finally, larvae raised on mannitol produced expected adult sex ratios, suggesting that sex-biased mortality due to the ingestion of mannitol occurs only in adults. We conclude that sex and reproductive status differences in mannitol ingestion drive sex-biased differences in adult fly mortality. [ABSTRACT FROM AUTHOR]

Background: Cancer patients undergoing Doxorubicin (DOX) treatment are susceptible to acute and chronic cardiac anomalies, including aberrant arrhythmias, ventricular dysfunction, and heart failure. To stratify patients at high risk for DOX -related heart failure (CHF), diagnostic techniques have been sought. While echocardiography is used for monitoring LVEF and LV volumes due to its wide-availability and cost-efficiency, it may not identify early stages of the initiation of DOX-induced systolic heart failure. To address these limitations, PET tracers could also provide noninvasive assessment of early and reversible metabolic changes of the myocardium. Objective: Herein, we report a preliminary investigation of 68Ga-Galmydar potential to monitor Dox-induced cardiomyopathy in vivo, ex vivo, and in cellulo employing both nuclear- and optical imaging. Methods and results: To assess 68Ga-Galmydar ability for monitoring DOX-induced cardiomyopathy, microPET imaging was performed 5 d post treatment of rats either with a single dose of DOX (15 mg/kg) or vehicle as a control (saline) and images were co-registered for anatomical reference using CT. Following tail-vein injection of the radiotracer in rats at 60 min, micro-PET/CT static scan (10 min acquisition), 68Ga-Galmydar demonstrated 1.91-fold lower uptake in hearts of DOX-treated (standard uptake value; SUV: 0.92, n = 3) rats compared with their vehicle treated (SUV: 1.76, n = 3) control counterparts. For correlation of PET imaging data, post-imaging quantitative biodistribution studies were also performed, wherein excised organs were counted for γ activity, and normalized to injected dose. The post imaging pharmacokinetic data also demonstrated heart uptake values of 2.0 fold lower for DOX treated rats(%ID/g; DOX: 0.44 ± 0.1, n = 3) compared to their vehicle-treated controls (%ID/g; Control: 0.89 ± 0.03, n = 3, p = 0.04). Employing the fluorescent traits of Galmydar, live cell fluorescence imaging indicated a gradual decrease in uptake and retention of Galmydar within mitochondria of H9c2 cells following DOX-treatment, while indicating dose-dependent and time-dependent uptake profiles. Following depolarization of electronegative transmembrane gradients at the mitochondrial membrane, the uptake of the probe was decreased in H9c2 cells, and the uptake profiles were found to be identical, using both fluorescence and radiotracer bioassays. Finally, the decreased uptake of the metalloprobe in H9c2 cells also correlated with caspase-3 expression resulting from DOX-induced cardiotoxicity and cell death. Conclusions: 68Ga-Galmydar could provide a noninvasive assessment of DOX-related and likely reversible metabolic changes at earliest stages. Further studies with other chemotherapeutics (potentially capable of inducing cardiomyopathy) are underway. [ABSTRACT FROM AUTHOR]

Background: The incidence of ocular candidiasis (OC) in patients with candidemia varies across different reports, and the issue of whether routine ophthalmoscopy improves outcomes has been raised. This study investigated the incidence of OC and evaluate whether the extent of OC impacts the clinical outcomes. Methods: This retrospective study included non-neutropenic patients with candidemia who underwent treatment at one of 15 medical centers between 2010 and 2016. Chorioretinitis without other possible causes for the ocular lesions and endophthalmitis was classified as a probable OC. If signs of chorioretinitis were observed in patients with a systemic disease that causes similar ocular lesions, they were classified as a possible OC. Results: In total, 781 of 1089 patients with candidemia underwent an ophthalmic examination. The prevalence of OC was 19.5%. The time from the collection of a positive blood culture to the initial ophthalmic examination was 5.0 ± 3.9 days in patients with OC. The leading isolate was Candida albicans (77.9%). Possible OC was associated with unsuccessful treatments (resolution of ocular findings) (odds ratio: 0.354, 95% confidence interval: 0.141–0.887), indicating an overdiagnosis in patients with a possible OC. If these patients were excluded, the incidence fell to 12.8%. Endophthalmitis and/or macular involvement, both of which require aggressive therapy, were detected in 43.1% of patients; a significantly higher incidence of visual symptoms was observed in these patients. Conclusion: Even when early routine ophthalmic examinations were performed, a high incidence of advanced ocular lesions was observed. These results suggest that routine ophthalmic examinations are still warranted in patients with candidemia. [ABSTRACT FROM AUTHOR]

Over consumption of fructose may lead to obesity and dyslipidemia and cause fructosylation-induced alterations in the structure and function of proteins. The aim of this study was to investigate the role of fructosylated-HSA-AGE in the pathogenesis of fatty liver (NAFLD and NASH) by biochemical, immunological and histological studies. Immunogenicity of fructosylated-HSA-AGE was probed by inducing antibodies in rabbits. Fructosylated-HSA-AGE was found to be highly immunogenic. Furthermore, fructosylated-HSA-AGE caused mild fibrosis with steatosis and portal inflammation of hepatocytes in experimental animals. Liver function test and dyslipidemic parameters in immunized animals were also found to be raised. Ultrasonography, which should form part of the assessment of chronically raised transaminases, shows fatty infiltration. Interestingly, alanine aminotransferase (ALT), aspartate aminotransferase (AST), bilirubin, total cholesterol (TC) and triglyceride (TG) profiles confirms USG images of overweight, obese patients. Thus, present study demonstrates that fructosylated-HSA-AGE is hepatotoxic, immunologically active and may cause dyslipidemia. [ABSTRACT FROM AUTHOR]

Staphylococcal enterotoxins (SEs) produced by Staphylococcus aureus are known as causative agents of emetic food poisoning. We previously demonstrated that SEA binds with submucosal mast cells and evokes mast cell degranulation in a small emetic house musk shrew model. Notably, primates have been recognized as the standard model for emetic assays and analysis of SE emetic activity. However, the mechanism involved in SEA-induced vomiting in primates has not yet been elucidated. In the present study, we established common marmosets as an emetic animal model. Common marmosets were administered classical SEs, including SEA, SEB and SEC, and exhibited multiple vomiting responses. However, a non-emetic staphylococcal superantigen, toxic shock syndrome toxin-1, did not induce emesis in these monkeys. These results indicated that the common marmoset is a useful animal model for assessing the emesis-inducing activity of SEs. Furthermore, histological analysis uncovered that SEA bound with submucosal mast cells and induced mast cell degranulation. Additionally, ex vivo and in vivo pharmacological results showed that SEA-induced histamine release plays a critical role in the vomiting response in common marmosets. The present results suggested that 5-hydroxytryptamine also plays an important role in the transmission of emetic stimulation on the afferent vagus nerve or central nervous system. We conclude that SEA induces histamine release from submucosal mast cells in the gastrointestinal tract and that histamine contributes to the SEA-induced vomiting reflex via the serotonergic nerve and/or other vagus nerve. [ABSTRACT FROM AUTHOR]

Sorghum is used as a substitute for rice in dog food, owing to its nutritional similarity and low cost. However, its use has been associated with negative effects, like a reduction in palatability, digestibility, and enzyme activity, which can decrease nutrient absorption. The presence of condensed tannins (CT) in sorghum may cause these effects. Another tannin group, the hydrolysable tannins (HT), is known for its antioxidant properties. Research has shown the nutritional effects of sorghum on dogs, but the effect of HT on dogs remains unknown. We evaluated the effects of substituting rice with sorghum containing CT and inclusion of commercial extract of HT on digestibility, fecal and urinary characteristics, and postprandial blood glucose levels in adult dogs. Eight adult Beagle were randomly subjected to 4 treatments: (R) 50% rice; (RS) 25% rice + 25% sorghum; (RHT) 50% rice + 0.10% HT; (RSHT) 25% rice + 25% sorghum + 0,10% HT. Tannins did not affect food intake. The digestibility of dry matter, organic matter, crude protein (CP), acid hydrolyzed fat, gross energy, and metabolizable energy (ME) decreased with sorghum inclusion (P < 0.05). Greater fecal dry matter was observed with the RHT diet. HT associated with sorghum, RSHT diet, reduced ME (P < 0.05). Sorghum inclusion enhanced fecal output, without altering fecal score (P > 0.05). No alterations in urinary characteristics were observed. Sorghum and HT did not affect the postprandial blood glucose response measured by the area under the curve (P > 0.05). The substitution of rice by sorghum decreased CP digestibility and ME of the diets. Sorghum can be considered as a source of carbohydrates with lower digestibility of protein and energy than rice. HT may potentiate the effect of CT, but more research is needed to evaluate its potential use in dog nutrition. [ABSTRACT FROM AUTHOR]

Background: Vancomycin is frequently used in hemodialysis (HD) and in hemodiafiltration (HDF) patients and is usually administered in the last 30 or 60 minutes of a dialysis session. Vancomycin pharmacokinetics are not well described in HDF patients. The aim of this study is to develop a population pharmacokinetic (PPK) model and dosing regimen for vancomycin in HDF patients and to evaluate its applicability in low-flux (LF-HD) patients. Methods: Two-compartment PPK models were developed using data from HDF patients (n = 17), and was parameterized as follows: non-renal clearance (CLm), renal clearance as a fraction of creatinine clearance (fr), central volume of distribution (V1), intercompartmental clearance (CL12), peripheral volume of distribution (V2) and extracorporeal extraction ratio (Eec). We evaluated the final model in a cohort of LF-HD patients (n = 21). Dosing schemes were developed for a vancomycin 24-h AUC of 400 mg*h/L. Results: Model parameters (± SD) were: CLm = 0.473 (0.271) L/h, fr = 0.1 (fixed value), V1 = 0.278 (0.092) L/kgLBMc, CL12 = 9.96 L/h (fixed value), V2 = 0.686 (0.335) L/kgLBMc and Eec = 0.212 (0.069). The model reliably predicted serum levels of vancomycin in both HDF and LF-HD patients during and between dialysis sessions. The median of the prediction error (MDPE) as a measure of bias is -0.7% (95% CI: -3.4%-1.7%) and the median of the absolute values of the prediction errors (MDAPE) as a measure of precision is 7.9% (95% CI: 6.0%-9.8%). In both HDF and LF-HD, the optimal vancomycin loading dose for a typical patient weighing 70 kg is 1700 mg when administered during the last 60 minutes of the hemodialysis session. Maintenance dose is 700 mg if administered during the last 30 or 60 minutes of the hemodialysis session. Conclusion: The developed PPK model for HDF is also capable of predicting serum levels of vancomycin in patients on LF-HD. A dosing regimen was developed for the use of vancomycin in HDF and LF-HD. [ABSTRACT FROM AUTHOR]

Hyperandrogenism is a risk factor of cerebrovascular diseases as androgens can alter markedly the regulation of cerebrovascular tone. We examined the combined impact of androgen excess and vitamin D deficiency (VDD), a common co-morbidity in hyperandrogenic disorders, on remodeling and testosterone-induced vascular responses of anterior cerebral arteries (ACA) in order to evaluate the interplay between androgens and VDD in the cerebral vasculature. Male and female Wistar rats were either fed with vitamin D deficient or vitamin D supplemented diet. Half of the female animals from both groups received transdermal testosterone treatment. After 8 weeks, vessel lumen, wall thickness and testosterone-induced vascular tone of isolated ACA were determined using pressure microangiometry and histological examination. Androgen receptor protein expression in the wall of cerebral arteries was examined using immunohistochemistry. In female rats only combined VDD and testosterone treatment decreased the lumen and increased the wall thickness of ACA. In males, however VDD by itself was able to decrease the lumen and increase the wall thickness. Vascular reactivity showed similar alterations: in females, testosterone constricted the ACA only after combined VDD and hyperandrogenism, whereas in males VDD resulted in increased testosterone-induced contractions in spite of decreased androgen receptor expression. In conclusion, a marked interplay between hyperandrogenism and VDD results in inward remodeling and enhanced testosterone-induced constrictions of cerebral arteries, which might compromise the cerebral circulation and thus, increase the risk of stroke in the long term. In addition, the early cerebrovascular manifestation of VDD appears to require androgen excess and thus, depends on gender. [ABSTRACT FROM AUTHOR]

Theoretical models of time perception suggest a simple bottom-up relationship between physiological arousal and perceived duration. Increases in physiological arousal lengthen the perceived duration of events whereas decreases in physiological arousal reduce them. Whilst this relationship has been demonstrated for highly arousing negatively valenced stimuli, it has not been demonstrated for other classes of distorting stimuli (e.g. positively valenced or low arousal stimuli). The current study tested the effect of valence (positive and negative) and arousal level (high and low) on the relationship between physiological arousal and perceived duration. Sympathetic nervous system (SNS) and parasympathetic nervous system (PSNS) activity was measured during a verbal estimation task in which participants judged the duration of high and low arousal, positive, negative and neutrally valenced IAPS images. SNS and PSNS activity were indexed by measuring Pre-Ejection Period (PEP) and High Frequency Heart-rate Variability (HF-HRV) respectively. SNS reactivity was predicative of perceived duration, but only for high arousal negatively valenced stimuli, with decreases in PEP being associated with longer duration estimates. SNS and PSNS activity was not predictive of perceived duration for the low arousal negative stimuli or the low and high arousal positive stimuli. We therefore propose a new model suggesting that emotional distortions to time result from a combination of bottom-up (physiological arousal) and top-down (threat detection) factors. [ABSTRACT FROM AUTHOR]

Chronic fatigue, in its various manifestations, frequently co-occur with pain, sleep disturbances and depression and is a non-communicable condition which is rapidly becoming endemic worldwide. However, it is handicapped by a lack of objective definitions and diagnostic measures. This has prompted the World Health Organization to develop an international instrument whose intended purpose is to improve quality of life (QOL), with energy and fatigue as one domain of focus. To complement this objective, the interface between detoxification, the exposome, and xenobiotic-sensing by nuclear receptors that mediate induction of biotransformation-linked genes, is stimulating renewed attention to a rational development of strategies to identify the metabolic profiles in complex multifactorial conditions like fatigue. Here we present results from a seven-year study of a cohort of 576 female patients suffering from low to high levels of chronic fatigue, in which phase I and phase II biotransformation was assessed. The biotransformation profiles used were based on hepatic detoxification challenge tests through oral caffeine, acetaminophen and acetylsalicylic acid ingestion coupled with oxidative stress analyses. The interventions indicated normal phase I but increased phase II glucuronidation and glycination conjugation. Complementarity was indicated between a fatigue scale, medical symptoms and associated energy-related parameters by application of Chi-square Automatic Interaction Detector (CHAID) analysis. The presented study provides a cluster of data from which we propose that multidisciplinary inputs from the combination of a fatigue scale, medical symptoms and biotransformation profiles provide the rationale for the development of a comprehensive laboratory instrument for improved diagnostics and personalized interventions in patients with chronic fatigue with a view to improving their QOL. [ABSTRACT FROM AUTHOR]

Regulatory agencies establish that a broad physicochemical and biological characterization is necessary for the evaluation of comparability between a biosimilar candidate product and a reference commercial drug. Between them, conformational characterization of proteins is of vital importance to determine its folding and biological functions. In this work, the conformational features of a novel monoclonal antibody (called 5G4) were evaluated by means of circular dichroism spectroscopy and fluorescence. Secondary structure and thermal stability of mAbs were determined by circular dichroism in the far ultraviolet, while three-dimensional folding of proteins was analyzed by both circular dichroism in the near ultraviolet and intrinsic tryptophan fluorescence. In all experiments, Herceptin (Roche) was used as control. Both antibodies showed a composition of secondary structure predominantly of β-sheets (55–56%) and thermal stability of ~ 75°C, suggesting structural similarity. The three-dimensional folding of proteins was also similar due to the absorption spectra of the aromatic residues and the emission wavelength maxima by fluorescence were comparable. The values of the fluorescence attenuation constant (Stern-Volmer constant) for increasing concentrations of acrylamide were also similar, suggesting a degree of exposure of tryptophan residues similar, although it was slightly decreased for Herceptin. Our data permit to consider that 5G4 monoclonal antibody showed similar conformational characteristics when compared with Herceptin. [ABSTRACT FROM AUTHOR]

Glycine receptors (GlyRs) are ligand-gated chloride channels mediating inhibitory neurotransmission in the brain stem and spinal cord. They function as pentamers composed of alpha and beta subunits for which 5 genes have been identified in human (GLRA1, GLRA2, GLRA3, GLRA4, GLRB). Several in vitro studies showed that the pentameric subtype composition as well as its stoichiometry influence the distribution and the molecular function of the receptor. Moreover, mutations in some of these genes are involved in different human conditions ranging from tinnitus to epilepsy and hyperekplexia, suggesting distinct functions of the different subunits. Although the beta subunit is essential for synaptic clustering of the receptor, the specific role of each alpha subtype is still puzzling in vivo. The zebrafish genome encodes for five glycine receptor alpha subunits (glra1, glra2, glra3, glra4a, glra4b) thus offering a model of choice to investigate the respective role of each subtype on general motor behaviour. After establishing a phylogeny of GlyR subunit evolution between human and zebrafish, we checked the temporal expression pattern of these transcripts during embryo development. Interestingly, we found that glra1 is the only maternally transmitted alpha subunit. We also showed that the expression of the different GlyR subunits starts at different time points during development. Lastly, in order to decipher the role of each alpha subunit on the general motor behaviour of the fish, we knocked out individually each alpha subunit by CRISPR/Cas9-targeted mutagenesis. Surprisingly, we found that knocking out any of the alpha2, 3, a4a or a4b subunit did not lead to any obvious developmental or motor phenotype. However, glra1-/- (hitch) embryos depicted a strong motor dysfunction from 3 days, making them incapable to swim and thus leading to their premature death. Our results infer a strong functional redundancy between alpha subunits and confirm the central role played by glra1 for proper inhibitory neurotransmission controlling locomotion. The genetic tools we developed here will be of general interest for further studies aiming at dissecting the role of GlyRs in glycinergic transmission in vivo and the hitch mutant (hic) is of specific relevance as a new model of hyperekplexia. [ABSTRACT FROM AUTHOR]

Background: Vitamin D may play a role in skeletal muscle because of the discovery of VDR in skeletal muscle. However, vitamin D deficiency is a global problem, including athletes. Studies examining the effect of vitamin D supplementation on muscle function in athletes have inconsistent results. Therefore, we aimed to quantitatively summarize the evidence for the effect of vitamin D supplementation on skeletal muscle strength and explosive power of athletes using a meta-analysis. Methods: PubMed, EMBASE, Cochrane Library, and Web of Science were searched for studies to identify randomized controlled trials or controlled trials meeting the inclusion criteria. By a meta-analysis, effect sizes (standardized mean differences, SMD) with 95% confidence intervals (CI) was calculated to compare reported outcomes across studies, I2 index was used to assessing heterogeneity, and heterogeneity factors were identified by regression analysis. The potential publication and sensitivity analyses were also assessed. Results: Eight RCTs involving 284 athletes were included. The protocols used to evaluate the muscle strength of athletes were inconsistent across the included studies, and muscle explosive power was assessed via vertical jump tests. The results indicated that vitamin D supplementation had no impact on overall muscle strength outcomes (SMD 0.05, 95% CI: -0.39 to 0.48, p = 0.84). In subgroup analysis, vitamin D supplementation had an effect on lower-limb muscle strength (SMD 0.55, 95% CI:0.12 to 0.98, p = 0.01) but not upper-limb muscle strength (SMD -0.19, 95% CI:-0.73 to 0.36, p = 0.50) or muscle explosive power (SMD 0.05, 95% CI:-0.24 to 0.34, p = 0.73). Vitamin D supplementation was more effective for athletes trained indoors (SMD 0.48, 95% CI:0.06 to 0.90, p = 0.02). Conclusions: Vitamin D supplementation positively affected lower limb muscle strength in athletes, but not upper limb muscle strength or muscle power. Different muscle groups and functions may respond differently to vitamin D supplementation. Additional studies should focus on determining the appropriate vitamin D supplementation methods and optimal serum 25(OH)D levels for athletes. Registration: The protocol for our study is registered in the international prospective register of systematic reviews (PROSPERO registration number ). [ABSTRACT FROM AUTHOR]

Background: Assessing Mycobacterium tuberculosis (TB) viability by fluorescein diacetate (FDA) microscopy can predict TB culture results, treatment response and infectiousness. However, diverse methods have been published. We aimed to optimise FDA microscopy, minimising sputum processing, biohazard and complexity for use in resource-constrained settings. Methods and results: Optimization: Patients with smear-positive pulmonary TB before treatment and healthy control participants provided sputa. These were divided into equal aliquots that were tested directly or after NaOH centrifuge-decontamination. Each aliquot was cultured and used to prepare slides (n = 80). FDA microscopy used: 1 or 3 drops of sputum; with/out acid-alcohol wash; with/out phenol sterilization; with 0/30/60 seconds KMnO4 quenching. Control samples all had negative culture and microscopy results. FDA microscopy had higher sensitivity when performed directly (without centrifuge-decontamination) on 1 drop of sputum (P<0.001), because 3 drops obscured microscopy. Acid-alcohol wash and KMnO4 quenching made bacilli easier to identity (P = 0.005). Phenol sterilization did not impair microscopy (P>0.1). Validation: The 2 protocols that performed best in the optimization experiments were reassessed operationally by comparing duplicate slides (n = 412) stained with KMnO4 quenching for 30 versus 60 seconds. FDA microscopy results were similar (P = 0.4) and highly reproducible, with 97% of counts agreeing within +/-1 logarithm. Storage: Smear microscopy slides and aliquots of the sputum from which they were made were stored for 4 weeks. Twice-weekly, paired slides (n = 80) were stained with freshly prepared versus stored FDA and read quantitatively. Storing sputum, microscopy slides or FDA solution at 4°C or room temperature had no effect on FDA microscopy results (all P>0.2). Cost: Material costs for each slide tested by FDA microscopy using reagents purchased locally were USD $0.05 and required the same equipment, time and skills as auramine acid-fast microscopy. Conclusions: We recommend a simple, bio-secure protocol for FDA microscopy that provides sensitive and repeatable results without requiring centrifugation. [ABSTRACT FROM AUTHOR]

Background: Regional anticoagulation with citrate during renal replacement therapy (RRT) reduces the risk of bleeding, extends dialyzer lifespan and is cost-effective. Therefore, current guidelines recommend its use if patients are not anticoagulated for another reason and if there are no contraindications against citrate. RRT with regional citrate anticoagulation has been established in critically ill patients as continuous veno-venous hemodialysis (CVVHD) to reduce citrate load. However, CVVHD is inferior regarding middle molecule clearance compared to continuous veno-venous hemofiltration (CVVH). The use of a high cut-off dialyzer in CVVHD may thus present an option for middle molecule clearance similar to CVVH. This may allow combining the advantages of both techniques. Methods: In this prospective, randomized, single-blinded single-center-trial, sixty patients with acute renal failure and established indication for renal replacement therapy were randomized 1:1 into two groups. The control group was put on CVVHD using regional citrate anticoagulation and a high-flux dialyzer, while the intervention group was on CVVHD using regional citrate anticoagulation and a high-cut-off dialyzer. The concentrations of urea, creatinine, β2-microglobulin, myoglobin, interleukin 6 and albumin were measured pre- and post-dialyzer 1, 6, 12, 24 and 48 hours after initiating CVVHD. Results: Mean plasma clearance for β2-microglobulin was 19.6±5.8 ml/min in the intervention group vs. 12.2±3.6 ml/min in the control group (p<0.001). For myoglobin (8.0±4.5 ml/min vs. 0.2±3.6 ml/min, p<0.001) and IL-6 (1.5±4.3 vs. -2.5±3.5 ml/min, p = 0.002) a higher mean plasma clearance using high-cut-off dialyzer could be detected too, but no difference for urea, creatinine and albumin could be observed concerning this parameter between the two groups. Conclusion: CVVHD using a high cut-off dialyzer results in more effective middle molecule clearance than that with high-flux dialyzer. Trial registration: German Clinical Trials Register (, registered 26th November 2013) [ABSTRACT FROM AUTHOR]

Progesterone membrane receptor component 1 (Pgrmc1) is a cytochrome b5-related protein with wide-ranging functions studied most extensively in non-neural tissues. We previously demonstrated that Pgrmc1 is widely distributed in the brain with highest expression in the limbic system. To determine Pgrmc1 functions in cells of these regions, we compared transcriptomes of control siRNA-treated and Pgrmc1 siRNA-treated N42 hypothalamic cells using whole genome microarrays. Our bioinformatics analyses suggested that Pgrmc1 plays a role in immune functions and likely regulates proinflammatory cytokine signaling. In follow-up studies, we showed that one of these cytokines, TNFα, increased expression of rtp4, ifit3 and gbp4, genes found on microarrays to be among the most highly upregulated by Pgrmc1 depletion. Moreover, either Pgrmc1 depletion or treatment with the Pgrmc1 antagonist, AG-205, increased both basal and TNFα-induced expression of these genes in N42 cells. TNFα had no effect on levels of Rtp4, Ifit3 or Gbp4 mRNAs in mHippoE-18 hippocampal control cells, but Pgrmc1 knock-down dramatically increased basal and TNFα-stimulated expression of these genes. P4 had no effect on gbp4, ifit3 or rtp4 expression or on the ability of Pgrmc1 to inhibit TNFα induction of these genes. However, a majority of the top upstream regulators of Pgrmc1 target genes were related to synthesis or activity of steroids, including P4, that exert neuroprotective effects. In addition, one of the identified Pgrmc1 targets was Nr4a1, an orphan receptor important for the synthesis of most steroidogenic molecules. Our findings indicate that Pgrmc1 may exert neuroprotective effects by suppressing TNFα-induced neuroinflammation and by regulating neurosteroid synthesis. [ABSTRACT FROM AUTHOR]