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Pancreatic β cell dysfunction and reduction due to glucose toxicity play a crucial role in the development of type 2 diabetes mellitus (T2DM). Irisin, a novel exercise-induced myokine, reduces obesity, improves insulin resistance and lowers blood glucose by promoting the browning of white adipose tissue, thereby enhancing thermogenesis and increasing energy expenditure. Recent studies have reported that irisin promotes cell proliferation and protects cells from apoptosis. However, the effects of irisin on pancreatic β cells are unknown. Thus, the aim of this study was to investigate the effects and the potential underlying mechanisms of irisin on pancreatic β cell proliferation and apoptosis induced by high glucose. Both in vitro (INS-1 cells) and in vivo (a T2DM rat model) experiments were conducted. Irisin significantly increased the proliferation of INS-1 cells, with the most significant effect observed at 24 h with 100 ng/ml irisin. Irisin also promoted INS-1 cell proliferation via the ERK and p38 MAPK signaling pathways, protected the cells from high-glucose-induced apoptosis by regulating the expression of caspases, Bad, Bax, Bcl-2 and Bcl-xl, and improved pancreatic β cell function. Irisin significantly reduced the body weight and blood glucose values and increased the serum insulin levels of the diabetic rats. An oral glucose tolerance test (OGTT) indicated that irisin also improved the glucose tolerance of T2DM rats. Together, these findings suggest that irisin may have applications in the prevention and treatment of T2DM because of its protective effect on the secretion of pancreatic β cells.

Vaspin (visceral adipose tissue-derived serine protease inhibitor) is a recently discovered adipokine that has been implicated in diabetes mellitus and other metabolic disorders. However, the effects of vaspin on pancreatic β cell function and related mechanisms are not fully understood. Thus, the present study was performed to investigate the effects of vaspin on pancreatic β cell function and the potential underlying mechanisms. Both in vitro (rat insulinoma cells, INS-1) and in vivo (high fat diet fed rats) experiments were conducted. The results showed that vaspin significantly increased INS-1 cell secretory function. Potential mechanisms were explored using inhibitors, western blot and real-time PCR techniques. We found that vaspin increased the levels of IRS-2 mRNA and IRS-2 total protein, while decreased the serine phosphorylation level of IRS-2 protein. Moreover, vaspin increased the Akt phosphorylation protein level which was reversed by PI3K inhibitor ly294002. In addition, vaspin increased the phosphorylation levels of mTOR and p70S6K, which was inhibited by rapamycin. Meanwhile, we found that the NF-κB mRNA and protein levels were reduced after vaspin treatment, similar to the effect of NF-κB inhibitor TPCK. Furthermore, vaspin increased the glucose stimulated insulin secretion (GSIS) level, lowered blood glucose level and improved the glucose tolerance and insulin sensitivity of high fat diet fed rats. Hyperglycemic clamp test manifested that vaspin improved islet β cell function. Together, these findings provide a new understanding of the function of vaspin on pancreatic β cell and suggest that it may serve as a potential agent for the prevention and treatment of type 2 diabetes.

This study investigated the effects of long-term simulated weightlessness on liver morphology, enzymes, glycogen, and apoptosis related proteins by using two-month rat-tail suspension model (TS), and liver injury improvement by rat-tail suspension with resistance training model (TS&RT). Microscopically the livers of TS rats showed massive granular degeneration, chronic inflammation, and portal fibrosis. Mitochondrial and endoplasmic reticulum swelling and loss of membrane integrity were observed by transmission electron microscopy (TEM). The similar, but milder, morphological changes were observed in the livers of TS&RT rats. Serum biochemistry analysis revealed that the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were significantly higher (p

Promiscuous inhibition of the human ether-à-go-go-related gene (hERG) potassium channel by drugs poses a major risk for life threatening arrhythmia and costly drug withdrawals. Current knowledge of this phenomenon is derived from a limited number of known drugs and tool compounds. However, in a diverse, naïve chemical library, it remains unclear which and to what degree chemical motifs or scaffolds might be enriched for hERG inhibition. Here we report electrophysiology measurements of hERG inhibition and computational analyses of >300,000 diverse small molecules. We identify chemical 'communities' with high hERG liability, containing both canonical scaffolds and structurally distinctive molecules. These data enable the development of more effective classifiers to computationally assess hERG risk. The resultant predictive models now accurately classify naïve compound libraries for tendency of hERG inhibition. Together these results provide a more complete reference map of characteristic chemical motifs for hERG liability and advance a systematic approach to rank chemical collections for cardiotoxicity risk.

ObjectiveThis study was performed to investigate the therapeutic effects of iguratimod in a lupus mouse model.MethodsFemale MRL/lpr mice were treated with iguratimod, vehicle solution or cyclophosphamide. Proteinuria was monitored and kidney injury was blindly scored by a renal pathologist. Serum anti-double-stranded DNA antibodies were monitored by radioimmunoassay. Kidney IgG and CD20 were stained by immunohistochemistry. Splenic lymphocyte phenotypes were analyzed by flow cytometry. BAFF, IL-17A, IL-6, and IL-21 levels in serum and splenic lymphocytes were detected by ELISA or quantitative PCR.ResultsCompared with the vehicle-treated controls, MRL/lpr mice treated with iguratimod showed less protenuria, less acute pathological lesions and no chronic changes in the kidneys. There were significant differences in glomerular injury and vasculitis scores, as well as in the semi-quantitative analysis of immune complex deposition between the two groups. Disease activity markers in sera (anti-dsDNA antibodies and immunoglobulin levels) were reduced and hypocomplementemia was attenuated. Lymphocyte expression of BAFF, IL-6, IL-17A and IL-21 was decreased. The abnormal splenic B220+ T cell and plasma cell populations in MRL/lpr mice were reduced by iguratimod treatment, with recovery of the total B cell population and inhibition of B cell infiltration of the kidney tissue. The dosage of iguratimod used in this study showed no significant cytotoxic effects in vivo and no overt side-effects were observed.ConclusionIguratimod ameliorates immune nephritis in MRL/lpr mice via a non-antiproliferative mechanism. Our data suggest a potential therapeutic role of iguratimod in lupus.

Avian metapneumovirus (aMPV) emerged as an important respiratory pathogen causing acute respiratory tract infection in avian species. Here we used a chicken aMPV subgroup C (aMPV/C) isolate to inoculate experimentally BALB/c mice and found that the aMPV/C can efficiently replicate and persist in the lungs of mice for at least 21 days with a peak viral load at day 6 postinoculation. Lung pathological changes were characterized by increased inflammatory cells. Immunochemical assay showed the presence of viral antigens in the lungs and significant upregulation of pulmonary inflammatory cytokines and chemokines including MCP-1, MIP-1α, RANTES, IL-1β, IFN-γ, and TNF-α were detected following inoculation. These results indicate for the first time that chicken aMPV/C may replicate in the lung of mice. Whether aMPV/C has potential as zoonotic pathogen, further investigation will be required.

Growing evidence suggests that drugs interact with diverse molecular targets mediating both therapeutic and toxic effects. Prediction of these complex interactions from chemical structures alone remains challenging, as compounds with different structures may possess similar toxicity profiles. In contrast, predictions based on systems-level measurements of drug effect may reveal pharmacologic similarities not evident from structure or known therapeutic indications. Here we utilized drug-induced transcriptional responses in the Connectivity Map (CMap) to discover such similarities among diverse antagonists of the human ether-à-go-go related (hERG) potassium channel, a common target of promiscuous inhibition by small molecules. Analysis of transcriptional profiles generated in three independent cell lines revealed clusters enriched for hERG inhibitors annotated using a database of experimental measurements (hERGcentral) and clinical indications. As a validation, we experimentally identified novel hERG inhibitors among the unannotated drugs in these enriched clusters, suggesting transcriptional responses may serve as predictive surrogates of cardiotoxicity complementing existing functional assays.

Methylene blue (MB) has unique energy-enhancing and antioxidant properties and is FDA-approved drug to treat methemoglobinemia and cyanide poisoning. This study evaluated the efficacy of MB to treat ischemic stroke in rats using longitudinal MRI and behavioral measures. Rats were subjected to 60-minute middle-cerebral-artery occlusion. In a randomized double-blinded design, vehicle or MB was administered after reperfusion. The initial lesion volumes at 30 minutes post-ischemia were not significantly different between the two groups (P = 0.92). The final infarct volumes two days after stroke increased in the vehicle group but decreased in the MB group, yielding a 30% difference in infarct volume (P = 0.03). Tracking tissue fate on a pixel-by-pixel basis showed that MB salvaged more initial core pixels compared to controls (22±3% versus 11±3%, P = 0.03), and more mismatch pixels compared to controls (83±3% versus 61±8%, P = 0.02). This study demonstrates MB treatment minimizes ischemic brain injury and improves functional outcomes.

By restoring mitochondrial function, methylene blue (MB) is an effective neuroprotectant in many neurological disorders (e.g., Parkinson's and Alzheimer's diseases). MB has also been proposed as a brain metabolic enhancer because of its action on mitochondrial cytochrome c oxidase. We used in vitro and in vivo approaches to determine how MB affects brain metabolism and hemodynamics. For in vitro, we evaluated the effect of MB on brain mitochondrial function, oxygen consumption, and glucose uptake. For in vivo, we applied neuroimaging and intravenous measurements to determine MB's effect on glucose uptake, cerebral blood flow (CBF), and cerebral metabolic rate of oxygen (CMRO(2)) under normoxic and hypoxic conditions in rats. MB significantly increases mitochondrial complex I-III activity in isolated mitochondria and enhances oxygen consumption and glucose uptake in HT-22 cells. Using positron emission tomography and magnetic resonance imaging (MRI), we observed significant increases in brain glucose uptake, CBF, and CMRO(2) under both normoxic and hypoxic conditions. Further, MRI revealed that MB dramatically increased CBF in the hippocampus and in the cingulate, motor, and frontoparietal cortices, areas of the brain affected by Alzheimer's and Parkinson's diseases. Our results suggest that MB can enhance brain metabolism and hemodynamics, and multimetric neuroimaging systems offer a noninvasive, nondestructive way to evaluate treatment efficacy.

BackgroundWhole exome sequencing provides a labor-saving and direct means of genetic diagnosis of hereditary disorders in which the pathogenic gene harbors a large cohort of exons. We set out to demonstrate a suitable example of genetic diagnosis of MEN 2A/FMTC (multiple endocrine neoplasia type 2/familial medullary thyroid carcinoma) using this approach.Methodology/principal findingsWe sequenced the whole exome of six individuals from a large Chinese MEN2A/FMTC pedigree to identify the variants of the RET (REarranged during Transfection) protooncogene and followed this by validation. Then prophylactic or surgical thyroidectomy with modified or level VI lymph node dissection and adrenalectomy were performed for the carriers. The cases were closely followed up. Massively parallel sequencing revealed four missense mutations of RET. We unexpectedly discovered that the proband's daughter with MEN 2A-related MTC presented a novel p.C634Y/V292M/R67H/R982C compound mutation, due to the involvement of p.C634Y in the proband with MEN 2A and p.V292M/R67H/R982C in the proband's husband with FMTC. In the maternal origin, p.C634Y caused bilateral MTC in all 5 cases and bilateral pheochromocytoma in 2 of the 5; the earliest onset age was 28 years. In the paternal origin, one of the six p.V292M/R67H/R982C carriers presented bilateral MTC (70 years old), one only had bilateral C-cell hyperplasia (44 years), two had bilateral multi-nodules (46 and 48 years) and two showed no abnormality (22 and 19 years).Conclusions/significanceThe results confirmed the successful clinical utility of whole exome sequencing, and our data suggested that the p.C634Y/V292M/R67H/R982C mutation of RET exhibited a more aggressive clinical phenotype than p.C634Y or p.V292M/R67H/R982C, while p.V292M/R67H/R982C presented a relatively milder pathogenicity of MTC and likely predisposed to FMTC.

BACKGROUND: A collection of in vitro evidence has demonstrated that Notch signaling plays a key role in the growth of neurites in differentiated neurons. However, the effects of Notch signaling on axon outgrowth in an in vivo condition remain largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: In this study, the neural tubes of HH10-11 chick embryos were in ovo electroporated with various Notch transgenes of activating or inhibiting Notch signaling, and then their effects on commissural axon outgrowth across the floor plate midline in the chick developing central nerve system were investigated. Our results showed that forced expression of Notch intracellular domain, constitutively active form of RBPJ, or full-length Hes1 in the rostral hindbrain, diencephalon and spinal cord at stage HH10-11 significantly inhibited commissural axon outgrowth. On the other hand, inhibition of Notch signaling by ectopically expressing a dominant-negative form of RBPJ promoted commissural axonal growth along the circumferential axis. Further results revealed that these Notch signaling-mediated axon outgrowth defects may be not due to the alteration of axon guidance since commissural axon marker TAG1 was present in the axons in floor plate midline, and also not result from the changes in cell fate determination of commissural neurons since the expression of postmitotic neuron marker Tuj1 and specific commissural markers TAG1 and Pax7 was unchanged. CONCLUSIONS/SIGNIFICANCE: We first used an in vivo system to provide evidence that forced Notch signaling negatively regulates commissural axon outgrowth.

To explore the effects of long-term weightlessness on the renal tissue, we used the two months tail suspension model to simulate microgravity and investigated the simulated microgravity on the renal morphological damages and related molecular mechanisms. The microscopic examination of tissue structure and ultrastructure was carried out for histopathological changes of renal tissue morphology. The immunohistochemistry, real-time PCR and Western blot were performed to explore the molecular mechanisms associated the observations. Hematoxylin and eosin (HE) staining showed severe pathological kidney lesions including glomerular atrophy, degeneration and necrosis of renal tubular epithelial cells in two months tail-suspended rats. Ultrastructural studies of the renal tubular epithelial cells demonstrated that basal laminas of renal tubules were rough and incrassate with mitochondria swelling and vacuolation. Cell apoptosis in kidney monitored by the expression of Bax/Bcl-2 and caspase-3 accompanied these pathological damages caused by long-term microgravity. Analysis of the HSP70 protein expression illustrated that overexpression of HSP70 might play a crucial role in inducing those pathological damages. Glucose regulated protein 78 (GRP78), one of the endoplasmic reticulum (ER) chaperones, was up-regulated significantly in the kidney of tail suspension rat, which implied that ER-stress was associated with apoptosis. Furthermore, CHOP and caspase-12 pathways were activated in ER-stress induced apoptosis. Resistance training not only reduced kidney cell apoptosis and expression of HSP70 protein, it also can attenuate the kidney impairment imposed by weightlessness. The appropriate optimization might be needed for the long term application for space exploration.

BACKGROUND: The mechanisms underlying neurotoxicity caused by L-DOPA are not yet completely known. Based on recent findings, we speculated that the increased expression of divalent metal transporter 1 without iron-response element (DMT1-IRE) induced by L-DOPA might play a critical role in the development of L-DOPA neurotoxicity. To test this hypothesis, we investigated the effects of astrocyte-conditioned medium (ACM) and siRNA DMT-IRE on L-DOPA neurotoxicity in cortical neurons. METHODS AND FINDINGS: We demonstrated that neurons treated with L-DOPA have a significant dose-dependent decrease in neuronal viability (MTT Assay) and increase in iron content (using a graphite furnace atomic absorption spectrophotometer), DMT1-IRE expression (Western blot analysis) and ferrous iron (55Fe(II)) uptake. Neurons incubated in ACM with or without L-DOPA had no significant differences in their morphology, Hoechst-33342 staining or viability. Also, ACM significantly inhibited the effects of L-DOPA on neuronal iron content as well as DMT1-IRE expression. In addition, we demonstrated that infection of neurons with siRNA DMT-IRE led to a significant decrease in DMT1-IRE expression as well as L-DOPA neurotoxicity. CONCLUSION: The up-regulation of DMT1-IRE and the increase in DMT1-IRE-mediated iron influx play a key role in L-DOPA neurotoxicity in cortical neurons.