DAB2 promotes pulmonary fibrosis and may act as an intermediate between IGF­1R and PI3K/AKT signaling pathways.

Idiopathic pulmonary fibrosis (IPF) is a heterogeneous lung disease associated with high mortality. Disabled-2 (DAB2), an adapter protein, regulates cell-fibrinogen adhesion and fibrinogen uptake. DAB2 is differentially expressed in mouse fibrotic lungs induced by bleomycin according to a genome microarray analysis based on Gene Expression Omnibus database. However, the role of DAB2 in IPF has not been revealed. A bleomycin-induced mouse model of pulmonary fibrosis was constructed in the present study. It found that the expression of DAB2 was upregulated in bleomycin-induced fibrotic lung tissue with collagen fiber deposition and pulmonary interstitium thickening. Colocalization of DAB2 with α-smooth muscle actin (SMA) was observed in lung tissue sections. In vitro, human lung fibroblast MRC-5 cells were treated with TGF-ß1 and the expression of DAB2 was increased. Knockdown of DAB2 suppressed cell proliferation and the expression of α-SMA, collagen I, collagen IV and fibronectin in TGF-ß1-treated MRC-5 cells. The phosphorylation levels of PI3K and AKT were suppressed in DAB2-knockdown cells. IGF-1/IGF-1R has been reported to promote pulmonary fibrosis and activate the PI3K/Akt signaling. In the present study, the activation of IGF-1/IGF-1R signaling pathways in bleomycin-induced fibrotic lung tissues were positively associated with DAB2 expression. The phosphorylation level of IGF-1R was increased in MRC-5 cells with TGF-ß1 treatment, and DAB2 expression was decreased by silencing of IGF-1R. This suggested that DAB2 might be a downstream target of the IGF-1R pathway and thus induced PI3K/AKT signaling activation and fibrogenesis. The current study demonstrated the importance of DAB2 in pulmonary fibrosis and suggested the potential of IGF-1R/DAB2/PI3K in the pathogenesis of IPF.

Overexpression of steroid receptor coactivators alleviates hyperglycemia-induced endothelial cell injury in rats through activating the PI3K/Akt pathway.

Hyperglycemia is a major factor in vascular endothelial injury that finally leads to a cardiovascular event. Steroid receptor coactivators (SRCs) are a group of non-DNA binding proteins that induce structural changes in steroid receptors (nuclear receptors) critical for transcriptional activation. SRCs, namely, SRC-1, SRC-2, and SRC-3, are implicated in the regulation of vascular homeostasis. In this study we investigate the role of SRCs in hyperglycemia-induced endothelial injury. Aortic endothelial cells were prepared from normal and diabetic rats, respectively. Diabetic rats were prepared by injection of streptozotocin (50 mg/kg, i.p.). The expression levels of SRC-1 and SRC-3 were significantly decreased in endothelial cells from the diabetic rats. Similar phenomenon was also observed in aortic endothelial cells from the normal rats treated with a high glucose (25 mM) for 4 h or 8 h. The expression levels of SRC-2 were little affected by hyperglycemia. Overexpression of SRC-1 and SRC-3 in high glucose-treated endothelial cells significantly increased the cell viability, suspended cell senescence, and inhibited cell apoptosis compared with the control cells. We further showed that overexpression of SRC-1 and SRC-3 markedly suppressed endothelial injury through restoring nitric oxide production, upregulating the expression of antioxidant enzymes (SOD, GPX, and CAT), and activating the PI3K/Akt pathway. The beneficial effects of SRC-1 and SRC-3 overexpression were blocked by treatment with the PI3K inhibitor LY294002 (10 mM) or with the Akt inhibitor MK-2206 (100 nM). In conclusion, hyperglycemia decreased SRC-1 and SRC-3 expression levels in rat aortic endothelial cells. SRC-1 and SRC-3 overexpression might protect against endothelial injury via inhibition of oxidative stress and activation of PI3K/Akt pathway.

Differential neurometabolite alterations in brains of medication-free individuals with bipolar disorder and those with unipolar depression: a two-dimensional proton magnetic resonance spectroscopy study.

OBJECTIVES: Bipolar disorder (BD) is a mental disorder characterized by periods of elevated mood and depression. Many individuals with BD are initially misdiagnosed and treated for unipolar depression (UD). In this study, we report direct comparisons between medication-free individuals with BD and those with UD in terms of the neurometabolites in the anterior cingulate cortex (ACC), medial prefrontal cortex (mPFC), parietal cortex (PC), and posterior cingulate cortex (PCC) of the brain. METHODS: Participants included medication-free patients with BD or UD, and matched healthy controls. All patients were in the depressive state and had similar symptoms. All subjects were subjected to a multi-voxel proton magnetic resonance spectroscopy procedure with a 3.0 T GE Signa MR scanner. After post-processing, the absolute concentrations of glycerophosphocholine + phosphocholine (GPC + PC), phosphocreatine + creatine (PCr + Cr), Glx (glutamate + glutamine), myo-inositol (MI), and N-acetyl aspartate (NAA) from the above brain regions were compared across the three groups. RESULTS: Patients with BD showed significantly higher levels of Glx in their ACC, lower GPC + PC, PCr + Cr, MI, and NAA in their PC, and lower NAA in their mPFC, compared to healthy controls; patients with UD presented significantly lower levels of GPC + PC, PCr + Cr, and NAA in their PCC, and lower Glx in their mPFC. All analyzed brain metabolites, except Glx, were significantly lower in the PC of patients with BD, whereas levels of GPC + PC, PCr + Cr, and NAA were significantly reduced in the PCC of patients with UD. CONCLUSIONS: These results add to the evidence of brain metabolite differences in brains of patients with UD and BD which may be of help in differentiating these two mood disorders.

Treatment with PPARδ agonist alleviates non-alcoholic fatty liver disease by modulating glucose and fatty acid metabolic enzymes in a rat model.

Non-alcoholic fatty liver disease (NAFLD) is an increasingly common condition which is associated with certain features of metabolic syndrome and insulin resistance. Peroxisome proliferator­activated receptor (PPAR)δ is an important regulator of energy metabolism and insulin resistance in diabetes. However, the function of PPARδ in NAFLD has not yet been fully elucidated. In the present study, in order to explore the function of PPARδ in NAFLD, we created a rat model of NALFD induced by a high-fat diet (HFD) and treated the rats with GW501516, a PPARδ agonist. We found that the lipid levels decreased, and hepatocellular ballooning and inflammatory cell infiltration were also significantly decreased following treatment of the rats with GW501516 compared to the untreated rats. Treatment with GW501516 also significantly decreased the homeostasis model assessment of insulin resistance (HOMA-IR) index, as well as the low­density lipoprotein (LDL) levels. In addition, treatment with GW501516 increased the levels of insulin­like growth factor­1 (IGF-1) and high­density lipoprotein (HDL) compared to the HFD group. Furthermore, the elevated levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma­glutamyl transpeptidase (GGT) and alkaline phosphatase (ALP) in the HFD group were all restored to the normal control levels following treatment with GW501516. RT­qPCR and immunohistochemical staining revealed that the expression levels of sterol regulatory element binding protein­1c (SREBP­1c) and glucose transporter 2 (GLUT­2) were both restored to normal control levels following treatment with GW501516. Also, the levels of enzymes related to lipid metabolism were increased following treatment with GW501516. In conclusion, our findings demonstrate that treatment with GW501516 alleviates NAFLD by modulating glucose and fatty acid metabolism.

The anti-fibrotic effects of microRNA-153 by targeting TGFBR-2 in pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial fibrotic lung disease with an undefined etiology and no effective treatments. By binding to cell surface receptors, transforming growth factor-ß (TGF-ß) plays a pivotal role in lung fibrosis. Therefore, the screening of microRNAs (miRNAs), especially those interrupting the effects of TGF-ß, may provide information not only on the pathomechanism, but also on the treatment of this disease. In the present study, we found that miR-153 expression was dysregulated in the lungs of mice with experimental pulmonary fibrosis and TGF-ß1 decreased miR-153 expression in pulmonary fibroblasts. Moreover, increased miR-153 levels attenuated, whereas the knock down of miR-153 promoted the pro-fibrogenic activity of TGF-ß1, and miR-153 reduced the contractile and migratory activities of fibroblasts. In addition, TGFBR2, a transmembrane serine/threonine kinase receptor for TGF-ß, was identified as a direct target of miR-153. Furthermore, by post-transcriptional regulation of the expression of TGFBR2, phosphorylation of SMAD2/3 was also influenced by miR-153. These data suggest that miR-153 disturbs TGF-ß1 signal transduction and its effects on fibroblast activation, acting as an anti-fibrotic element in the development of pulmonary fibrosis.

miR-29c regulates NAV3 protein expression in a transgenic mouse model of Alzheimer's disease.

The microRNA-29 family (miRNA-29s) has three mature members, miR-29a, miR-29b and miR-29c, which have been implicated in the regulation of the pathogenesis of Alzheimer's disease (AD). The miR-29 family members exhibit differential regulation in various diseases and different subcellular distribution. In the present study, we initially investigated differential expression of miR-29c in the hippocampus and the frontal cortex of the young APPswe/PSΔE9 mouse brain, accompanied by inverse expression of neurone navigator 3 (NAV3), a regulator of axon guidance. We observed that miR-29c directly mediated downregulation of NAV3 protein expression in vitro. The mouse NAV3 mRNA has a functional miR-29c binding site in the 3' UTR, which localized in the position between 830-836 bp of 3'UTR region, slightly different from human NAV3 mRNA binding site. These observations suggest that miR-29c may be involved in neurodegenerative processes by regulating NAV3 expression in the young AD mouse.

Melatonin inhibits tunicamycin-induced endoplasmic reticulum stress and insulin resistance in skeletal muscle cells.

The prevalence of type 2 diabetes mellitus (T2D) is increasing worldwide. Melatonin possesses various beneficial metabolic actions, decreased levels of which may accelerate T2D. Endoplasmic reticulum stress (ERS) has been linked to insulin resistance in multiple tissues, but the role of melatonin on ERS and insulin resistance in skeletal muscle has not yet been investigated. In this study, the results showed that tunicamycin decreased insulin-stimulated Akt phosphorylation, but promoted the phosphorylation of protein kinase R-like ER protein kinase (PERK) time-dependently in C2C12 cells. Consistently, ERS gene markers, including binding immunoglobulin protein (BIP)/glucose regulated protein 78 (GRP78) expression and the splicing of X box binding protein 1 (XBP-1), were activated by tunicamycin time-dependently. Interestingly, melatonin pretreatment reversed the elevated PERK phosphorylation, as well as the activation of Bip expression and XBP-1 splicing, and prevented the inhibitory effect of tunicamycin on Akt phosphorylation. In addition, the insulin-provoked glucose transport was reduced by tunicamycin, and then promoted by melatonin pretreatment. A strong phosphorylation of inositol-requiring enzyme 1 (IRE-1), c-JUN NH2-terminal kinase (JNK), and insulin receptor substrate 1 (IRS-1) serine, and simultaneously, a dramatic decrease of IRS-1 tyrosine phosphorylation were observed in the presence of tunicamycin, leading to a blockade of insulin signaling, which was reversed by melatonin pretreatment. Furthermore, luzindole pretreatment acted inversely with melatonin action on glucose uptake and insulin signaling. Therefore, these results demonstrated that melatonin pretreatment inhibited the activated role of tunicamycin on ERS and insulin resistance through melatonin receptor-mediated IRE-1/JNK/IRS-1 insulin signaling in skeletal muscle cells.

TCF2 attenuates FFA-induced damage in islet ß-cells by regulating production of insulin and ROS.

Free fatty acids (FFAs) are cytotoxic to pancreatic islet ß-cells and play a crucial role in the diabetes disease process. A recent study revealed a down-regulation of transcription factor 2 (TCF2) levels during FFA-mediated cytotoxicity in pancreatic ß-cells. However, its function during this process and the underlying mechanism remains unclear. In this study, treatment with palmitic acid (PA) at high levels (400 and 800 µM) decreased ß-cell viability and TCF2 protein expression, along with the glucose-stimulated insulin secretion (GSIS). Western and RT-PCR analysis confirmed the positive regulatory effect of TCF2 on GSIS through promotion of the key regulators pancreatic duodenal homeobox-1 (PDX1) and glucose transporter 2 (GLUT2) in ß-cells. In addition, both PI3K/AKT and MEK/ERK showed decreased expression in PA (800 µM)-treated ß-cells. Overexpression of TCF2 could effectively restore the inhibitory effect of PA on the activation of PI3K/AKT and MEK/ERK as well as ß-cell viability, simultaneously, inhibited PA-induced reactive oxygen species (ROS) generation. After blocking the PI3K/AKT and MAPK/ERK signals with their specific inhibitor, the effect of overexpressed TCF2 on ß-cell viability and ROS production was obviously attenuated. Furthermore, a protective effect of TCF2 on GSIS by positive modulation of JNK-PDX1/GLUT2 signaling was also confirmed. Accordingly, our study has confirmed that TCF2 positively modulates insulin secretion and further inhibits ROS generation via the PI3K/AKT and MEK/ERK signaling pathways. Our work may provide a new therapeutic target to achieve prevention and treatment of diabetes.

CCN5 overexpression inhibits profibrotic phenotypes via the PI3K/Akt signaling pathway in lung fibroblasts isolated from patients with idiopathic pulmonary fibrosis and in an in vivo model of lung fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive interstitial lung disease with unknown etiology and undefined treatment modality. Fibroblasts are regarded as the major cell type that mediates the onset and progression of lung fibrosis by secreting large amounts of extracellular matrix (ECM) proteins, such as connective tissue growth factor (CTGF/CCN2). Current knowledge confers a crucial role of CCN2 in lung fibrosis. CCN5, another member of the CCN family, has been suggested to play an inhibitory role in some fibrotic diseases, such as cardiac fibrosis. However, the role of CCN5 in the process of IPF remains unknown. In the present study, using western blot analysis, we demonstrate that CCN2 is highly expressed in fibroblasts derived from IPF tissue, but is only slightly expressed in normal human lung fibroblasts. However, CCN5 was weakly expressed in all the above cells. qRT-PCR revealed that transforming growth factor (TGF)-ß1 stimulation increased CCN2 expression in the IPF-derived cultures of primary human lung fibroblasts (PIFs) in a time- and concentration-dependent manner, but only slightly affected the expression of CCN5. The overexpression of CCN5 induced by the transfection of PIFs with recombinant plasmid did not affect cell viability, proliferation and apoptosis; however, it significantly suppressed the expression of CCN2, α-smooth muscle actin (α-SMA) and collagen type I. The TGF-ß1-induced upregulation of the phosphorylation of Akt was reversed by CCN5 overexpression. Our results also demonstrated that adenovirus-mediated CCN5 overexpression in a mouse model of bleomycin-induced IPF significantly decreased the hydroxyproline content in the lungs, as well as TGF-ß1 expression in bronchoalveolar lavage fluid. Taken together, our data demonstrate that CCN5 exerts an inhibitory effect on the fibrotic phenotypes of pulmonary fibroblasts in vitro and in vivo, and as such may be a promising target for the treatment of IPF.

Upregulation of adenosine A2A receptors induced by atypical antipsychotics and its correlation with sensory gating in schizophrenia patients.

Sensory gating deficits have been found in patients with schizophrenia and their unaffected relatives. However, the underlying neurobiological mechanism of this deficit remains unclear. Pre-clinical studies have implicated adenosine in sensory gating deficits in schizophrenia. Therefore, the current study investigated a possible relationship between peripheral adenosine A2A receptor (ADORA2A) and sensory gating indices (P50 measures) in medication-free schizophrenia (n=31) and healthy (n=21) groups. The effects of six-week antipsychotic treatment were examined. At baseline, schizophrenia patients showed impaired sensory gating compared to healthy controls. However, there was no significant difference in ADORA2A gene expression among groups. In addition, ADORA2A expression was not correlated with sensory gating at any time point. Following treatment, we found a significant upregulation of ADORA2A expression. Intriguingly, we observed a significant positive association between ADORA2A upregulation and baseline P50 amplitudes in the schizophrenia group. A main finding of the current pilot study is the upregulation of ADORA2A expression following treatment with antipsychotics. In addition, this upregulation was predicted by baseline P50 amplitude, an observation that awaits replication in an expanded sample.

MicroRNA-153 negatively regulates the expression of amyloid precursor protein and amyloid precursor-like protein 2.

Increased expression of the amyloid precursor protein (APP) is a crucial risk factor of Alzheimer's disease (AD). Amyloid precursor-like protein 2 (APLP2), a homologue of APP, is also suggested to participate in AD pathogenesis. Accumulating evidence suggest the regulatory role of microRNA on AD-related genes. Here we showed that the levels of miR-153 were significantly decreased at early- and late-stage of AD in APPswe/PSΔE9 murine model. Moreover, a binding site of miR-153 on APP and APLP2-3'UTR was identified, respectively, by luciferase assay. Gain and loss of function experiments demonstrated that miR-153 suppressed the expression of APP and APLP2. Using miR-153 transgenic mouse model, we testified that miR-153 downregulated the expression of APP and APLP2 protein in vivo. Furthermore, closely related expression patterns of miR-153 and APP/APLP2 during brain development indicated a physiological regulation role of miR-153 on the two genes. In a neuronal cell line treated with Aß(42) peptides and H(2)O(2,) the levels of miR-153 varied during time-course leading to corresponding changes of APLP2 protein, indicating Aß peptides and oxidative stress influence the expression of miR-153. Thus, miR-153 contributes to post-transcriptional regulation of APP/APLP2, suggesting a possible role for miR-153 in neuro-pathological conditions.

[Significance and effect of Glucosidorum Tripterygii tororum on expressions of Foxp3 in regulatory T cells in type 1 diabetic rat model].

AIM: To investigate the significance and effect of Glucosidorum Tripterygii tororum (GTT) on the expressions of Foxp3 in CD4(+)CD25(+) regulatory T cells (Tr) in type 1 diabetic rat model. METHODS: 48 rats were divided evenly into 3 groups by random. Diabetic model was developed by multiple low dose intraperitoneal injection of streptozotocin in group II, III, and group I was treated as normal control. After the establishment of the model, administration of GTT was conducted in group III, which was performed once a day, lasting three monthes. Lymphocyte proliferation was detected by lymphocyte transformation test (MTT assay). The expression of Foxp3 mRNA was measured in peripheral blood and spleen by using real-time PCR. The expression of Foxp3 protein in lymphoid node and spleen was detected by immunohistochemistry. RESULTS: The blood glucose level in group II, III was obviously higher than that of control group (P<0.01), followed with lymphocyte infiltrating in pancreatic islets. The infiltrating degree in group III was decreased than that of group II. The proliferation of splenic lymphocyte in group II, III was evidently increased comparing with the control group (P<0.01). After administration of GTT, the proliferation in group III was lower than that in group II. In group II, III, the expressions of Foxp3 mRNA and protein increased markedly compared with control group (P<0.05). Moreover, the level in group III was especially enhanced, but compared with group II, there was no significantly difference (P>0.05). CONCLUSION: Foxp3(+) Tr may involve in the pathogenesy of type 1 diabetes indused by STZ. GTT did have therapeutical effect on type 1 diabetes, possibly through upregulating the expressions of Foxp3 in Tr.