The Role Of Pancreatic Long Non-coding RNA Meg3 In The Mechanism Regulation Of Insulin Synthesis And Secretion

Long non-coding RNAs(lncRNAs)generally are greater in size than 200 nucleotides,without function of the coding protein.In recent years,a growing number of studies have demonstrated that lncRNAs express with specificity of time and space in tissues and cells,are involved in proliferation,apoptosis,migration,differentiation and many other important process of cells,their abnormal expressions are closely related to the occurrence of a variety of human diseases.Recent studies show that lncRNAs can influence diabetes susceptibility gene expression,which increases the risk of both type 1 and 2 diabetes,suggest lncRNAs may be closely related with the occurrence of diabetes.Maternally expressed gene 3(MEG3),the locus located at human chromosome 14q32.3,encodes an lncRNA MEG3.Be homologous Meg3 gene in mice with human.Existing research shows that lncRNA MEG3 expressed abnormally in islets of patients with type 2 diabetes,but its role in the pancreatic beta cells function has not yet been reported.We used real-time quantitative PCR(qRT-PCR,qPCR)technology to detect lncRNA Meg3 in different tissues and cells in normal mice,and in islets of non-obese diabetic(NOD)mice and db/db mice.Detection of lncRNA Meg3 expression in MIN6 cells and mouse islet cells,which treated with different concentration of sugar.Specific knockdown of lncRNA Meg3 expression in vitro and in vivo by specificity small RNA interference,and detect MIN6 cell proliferation,apoptosis,as well as synthesis and secretion of insulin through the thiazole blue(MTT),flow cytometry,radioimmunoassay(RIA),western blotting.Detection influence of lncRNA Meg3 on the synthesis and secretion of insulin in vivo by intraperitoneal glucose tolerancetest(IPGTT),enzyme-linked immunosorbent assay(ELISA),immunohistochemical.Using RNA Binding Protein Immunoprecipitation(RIP),Chromatin Immunoprecipitation(ChIP)to investigate the molecular mechanism of lncRNA Meg3 in insulin synthesis regulation.The results showed that lncRNA Meg3,regulated by the sugar concentration,is abundantly expressed in mouse islet cells.And it expressed in islet of diabetes mice significantly lower than in normal.These suggest that lncRNA Meg3 contact islet function may be important.By specific knockdown of Meg3 in the islet cell,cells vitality decline,apoptosis lncreased,insulin synthesis and secretion reduced,the transcription factors v-Maf musculoaponeurotic fibrosarcoma oncogene family,protein A(MafA)and Pancreatic and duodenal homeobox1(Pdx1)expressions downregulated.Immunohistochemical results show reducing the area of insulin positive cells.Through bioinformatics analysis,RIP and ChIP,we found lncRNA Meg3 inhibited MafA negative transcription factors Rad21/Smc3/Sin3α expression by combining EZH2 in mice pancreas,affected the synthesis of insulin.Our finding confirmed the lncRNA Meg3 maintain essential islet function in mice,the loss of expression leads to reduce of the insulin synthesis and secretion.Its expression reduced in islet of diabetes mice also suggested the lncRNA Meg3 may be involved in occur of the diabetes.This study aims to discuss the role of lncRNA Meg3 in mice islet function,further enrich and perfect the mechanism regulation network of islet cell function.