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The leucine-rich and immunoglobulin-like domains (LRIG) gene family contains LRIG1, 2 and 3. LRIG1 is a negative regulator of EGFR, but little is known about the function of LRIG2. To determine the role of LRIG2 in the progression of glioma, we performed RNA interference-mediated knockdown of LRIG2 in a human glioma cell line (GL15). Downregulation of LRIG2 expression resulted in: rapid EGF-mediated loss of EGFR; decreased proliferation; G(0)/G(1) arrest; increased spontaneous apoptosis; enhanced cell adhesion and increased invasion capability of GL15 cells in vitro. These findings indicate that LRIG2 possesses distinct functions compared with LRIG1 and validate the attractiveness of LRIG2 as a target in glioma therapy.

Protocatechuic acid (PCA, 3,4-dihydroxybenzoic acid), the main metabolite of anthocyanins, is widely distributed in fruits and vegetables and has been reported to possess a strong antioxidant activity. Herein, we aimed to investigate the protective effect of PCA against high palmitic-acid (PA)-induced oxidative damage and the underling molecular mechanisms in human umbilical vein endothelial cells (HUVECs). PCA reduced the levels of intracellular reactive oxygen species and malondialdehyde and increased the activities of endogenous antioxidant enzymes, including superoxide dismutase, glutathione peroxidase 1, and heme oxygenase 1 (HO-1). Metabolomic analysis showed that PCA affected numerous metabolites, especially some of which were related with energy metabolism. PCA also upregulated the phosphorylation of adenosine-monophosphate-activated protein kinase (AMPK) at Thr through activating liver kinase B1 and then promoted the expression of p-Nrf2 and HO-1. Moreover, PCA reversed the decreased expression of peroxisome proliferator-activated receptor γ coactivator 1α and significantly increased the mitochondrial density. Collectively, these results demonstrated that PCA attenuated PA-induced oxidative damage in HUVECs via an AMPK-dependent pathway.

Phloretin, a dihydrochalcone structural flavonoid compound, possesses antioxidant activity. In this study, we conducted studies to explore the function of phloretin on high palmitic acid-induced oxidative stress in human umbilical vein endothelial cells and investigated the potential mechanism using ribonucleic acid sequencing (RNA-Seq). Our findings reveal that phloretin significantly decreased the levels of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA), increased superoxide dismutase (SOD) and glutathione peroxidase-1 (Gpx-1) activity, and restored the loss of mitochondrial membrane potential (MMP). Next, whole transcriptome analysis was performed using RNA-Seq The results indicated more than 3000 differentially expressed genes (DEGs). Gene Ontology analysis revealed that the DEGs were categorized functionally, mainly by the biological processes, cell metabolism, and cellular response to chemical stimulus. The Kyoto Encyclopedia of Genes and Genomes indicated that they were mainly enriched in cAMP, apoptosis, and cytoskeletal regulation signaling pathways. Furthermore, on the basis of the results of RNA-Seq and Western blotting, our study verified that phloretin upregulated the expression of p-Nrf2 and HO-1 by promoting the phosphorylation of AMPK at Thr 172 through activation of liver kinase B1. In conclusion, phloretin attenuates PA-induced oxidative stress in HUVECs via the AMPK/Nrf2 antioxidative pathway.

ObjectivesGenome-wide association studies (GWASs) have discovered associations of numerous SNPs and genes with obesity. However, the underlying molecular mechanisms through which these SNPs and genes affect the predisposition to obesity remain not fully understood. Aims of our study are to comprehensively characterize obesity GWAS SNPs and genes through computational approaches.MethodsFor obesity GWAS identified SNPs, functional annotation, effects on miRNAs binding and impact on protein phosphorylation were performed via RegulomeDB and 3DSNP, miRNASNP, and the PhosSNP 1.0 database, respectively. For obesity associated genes, protein-protein interaction network construction, gene ontology and pathway enrichment analyses were performed by STRING, PANTHER and STRING, respectively.ResultsA total of 445 SNPs are significantly associated with obesity related phenotypes at threshold P < 5×10−8. A number of SNPs were eQTLs for obesity associated genes, some SNPs located at binding sites of obesity related transcription factors. SNPs that might affect miRNAs binding and protein phosphorylation were identified. Protein-protein interaction network analysis identified the highly-interconnected “hub” genes. Obesity associated genes mainly involved in metabolic process and catalytic activity, and significantly enriched in 15 signal pathways.ConclusionsOur results provided the targets for follow-up experimental testing and further shed new light on obesity pathophysiology.

The aim of the present study was to investigate the effects of P2X7R short hairpin (sh)RNA on the proliferation and apoptosis of MCF-7 cells, and to detect the expression of P2X7R in breast cancer and MCF-7 cells. In order to detect the expression of P2X7R in normal breast and breast cancer tissues, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blot analysis and immunohistochemistry were performed. A P2X7-targeted shRNA sequence and a scrambled sequence were inserted into the pLKO.1 expression vector, and MCF-7 cells with stable transfection of P2X7R-shRNA and P2X7R-scrambled shRNA (control) were selected. qRT-PCR was used to detect the mRNA expression levels of P2X7R in the MCF-7 cells transfected with P2X7R-shRNA and scrambled shRNA. In addition, protein expression levels of P2X7R in the fresh tumor tissues were detected by western blot analysis. An MTT assay was used to detect the proliferation rate at different time points, while flow cytometry was used to detect the growth inhibition and apoptosis rate of the stably transfected MCF-7 cells. P2X7R expression levels in the breast cancer tissues were higher when compared with the normal breast tissue, and a positive correlation was observed with the estrogen receptor (ER), as shown by qRT-PCR, western blot analysis and immunohistochemistry. Plasmids expressing P2X7 gene-specific shRNA and scrambled shRNA were constructed and transfected into MCF-7 cells. The qRT-PCR results revealed lower mRNA expression levels of P2X7 in the P2X7R-shRNA cells when compared with the scrambled shRNA cells. Furthermore, western blot analysis demonstrated that P2X7 protein was highly expressed in the MCF-7 cells transfected with scrambled shRNA, while low expression was observed in the P2X7R-shRNA-transfected cells. Following transfection of the recombinant plasmids into the MCF-7 cells, the proliferation rate in each group was analyzed. The P2X7R-shRNA and KN-62 groups were shown to have significantly reduced rates of proliferation when compared with the normal control group. In addition, flow cytometry revealed that the P2X7R-shRNA and KN-62 groups exhibited a reduced level of cell proliferation and a higher rate of apoptosis. In conclusion, P2X7R was shown to be overexpressed in breast cancer tissues and positively associated with ER expression. A P2X7R-shRNA expression vector effectively inhibited P2X7R expression in MCF-7 breast cancer cells, which subsequently induced cell apoptosis and reduced the levels of cell proliferation. These results indicated that P2X7R may serve as a potential target for breast cancer treatment and prevention.

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d-chiro-Inositol (DCI) is a biologically active component found in tartary buckwheat, which can reduce hyperglycemia and ameliorate insulin resistance. However, the mechanism underlying the antidiabetic effects of DCI remains largely unclear. This study investigated the effects and underlying molecular mechanisms of DCI on hepatic gluconeogenesis in mice fed a high fat diet and saturated palmitic acid-treated hepatocytes. DCI attenuated free fatty acid uptake by the liver via lipid trafficking inhibition, reduced diacylglycerol deposition, and hepatic PKCε translocation. Thus, DCI could improve insulin sensitivity by suppressing hepatic gluconeogenesis. Subsequent analyses revealed that DCI decreased hepatic glucose output and the expression levels of PEPCK and G6 Pase in insulin resistant mice through PKCε-IRS/PI3K/AKT signaling pathway. Likewise, such effects of DCI were confirmed in HepG2 cells with palmitate-induced insulin resistance. These findings indicate a novel pathway by which DCI prevents hepatic gluconeogenesis, reduces lipid deposition, and ameliorates insulin resistance via regulation of PKCε-PI3K/AKT axis.