LncRNA LEGLTBC Functions As A CeRNA To Antagonize The Effects Of MiR-199a-3p On The Downregulation Of FoxM1 In Glucolipotoxicity-Induced INS-1 Beta Cell Proliferation Of Damage

Objective : In this study,we intended to explore the role of long non-coding RNA-LEGLTBC(Lnc RNA-LEGLTBC)in the proliferation and damage of rat INS-1cells induced by glucolipotoxicity.Methods:(1)LEGLTBC expression was determined by q RT-PCR in INS-1 cells(rat islet β cell lines)incubated with high glucose and palmitate(HG/PA)at different time points.After LEGLTBC was inhibited or overexpressed,the proliferation of INS-1 cells incubated with HG/PA was detected by EDU staining and CCK-8.(2)Bioinformatics software RNA22 V2 and Reg RNA 2.0 predicted the binding of LEGLTBC to mi R-199a-3p.Binding ability of both was detected by double luciferase report.The expression of mi R-199a-3p in INS-1 cells at different time points after HG/PA incubation was determined by q RT-PCR.The expression of mi R-199a-3p was determined by q RT-PCR after LEGLTBC was overexpressed or knocked out.After the expression of mi R-199a-3p was reduced,the proliferation of INS-1 cells incubated with HG/PA was detected by EDU staining and CCK-8.(3)Bioinformatics software Target Scan predicted the binding of mi R-199a-3p to Fox M1.The combination of them was proved by luciferase experiment.The expression of Fox M1 protein in INS-1 cells after HG/PA incubation at different time points and mi R-199a-3p knockdown was determined by Western blotting.After Fox M1 silencing,the expression of cyclin Cyclin D1 in INS-1 cells incubated with Hg /PA was determined,and the proliferation ability of INS-1 cells was determined by EDU staining and CCK-8.(4)The expression of Fox M1 in INS-1 cells incubated with HG/PA was determined by Western blotting after LEGLTBC overexpression or silenced.The expression of Fox M1 and Cyclin D1 in INS-1 cells incubated with HG/PA was determined by Western blotting after LEGLTBC overexpression or LEGLTBC + silenced Fox M1.Results:(1)The results of q RT-PCR showed that the expression of LEGLTBC induced by Hg /PA was decreased in a time-dependent manner compared with that of INS-1 cells cultured under normal conditions.Inhibition of LEGLTBC expression by si RNA aggravated HG/PA induced impaired INS-1 cell proliferation.In contrast,adenovirus overexpression of LEGLTBC significantly recovered Ins-1 cell proliferation.(2)The level of mi R-199a-3p in INS-1 cells treated with HG /PA gradually increased with the prolonging of incubation time of HG /PA,which was contrary to the trend of LEGLTBC expression.The luciferase assay showed that mi R-199a-3p simulant and LEGLTBC-WT significantly reduced the luciferase activity in INS-1 cells,but had no significant effect on the luciferase activity in cells co-transfected with mi R-199a-3p simulant and LEGLTBC-MUT.In addition,the overexpression of LEGLTBC significantly down-regulated the expression of mi R-199a-3p,and silencing LEGLTBC promoted the expression of mi R-199a-3p.Reducing the expression of mi R-199a-3p reduced the proliferation slowdown of INS-1cells induced by HG/PA incubation.(3)The expression of Fox M1 protein in INS-1cells incubated with HG/PA was decreased in a time dependent manner.The luciferase assay showed that the luciferase activity in INS-1 cells was significantly reduced by co-transfection of mi R-199a-3p simulant and Fox M1-WT,while the luciferase activity in cells co-transfected with mi R-199a-3p simulant and Fox M1-MUT had no significant effect.Deduction of mi R-199a-3p increased Fox M1 expression.The expression of Fox M1 was silenced,and the expression of Cyclin D1 was significantly decreased in INS-1 cells incubated with HG/PA.EDU cell proliferation assay and CCK-8 assay also showed that inhibition of Fox M1 expression could aggravate the proliferation and cell viability injury of INS-1 cells induced by glucolipotoxicity.(4)Fox M1 expression increased when LEGLTBC was overexpressed,but decreased when LEGLTBC was inhibited.Overexpression of LEGLTBC or LEGLTBC + silenced Fox M1 resulted in increased FOXM1 expression in HG /PA treated INS-1 cells,while si-Fox M1 transfection decreased LEGLTBC-induced Fox M1 expression enhancement.In addition,overexpression of LEGLTBC increased the expression of c Cyclin D1 in INS-1 cells treated with HG /PA,while the protective effect of overexpression of LEGLTBC was inhibited after Fox M1 silenced.Conclusion: As a competitive endogenous RNA,LEGLTBC interacts with mi R-199a-3p and affects the expression of target gene Fox M1,thereby regulating HG/PA induced proliferation and injury of islet β cells.In this study,we investigated the role of Lnc RNA in the glucolipotoxicity injury of pancreatic beta cells,and revealed a new mechanism of glucolipotoxicity leading to the damage of pancreatic beta cells,thus providing a new idea for the protection of pancreatic beta cells.