MiR-126-3p Inhibits IRS-2 And Plays A Key Role In The Regulation Of β Cell Proliferation

Objective:mi R-126 has many target genes, including VEGF, PI3 K, Spred1, IRS1 for sugar metabolism. There is decreased content of mi R-126 in the plasma of patients with type 2 diabetes. Our preliminary findings, IRS-1 and mi R-126 are PIK3R2 target, so mi R-126 may be IRS-1 / PI3 K / Akt pathway that are involved in β cell metabolic regulation. However bioinformatics data analysis shows that mi R-126 IRS2 is a target gene, and IRS1 and IRS2 functions are inseparable, and therefore, it is necessary to adjust the predicted verified IRS2 mi R-126, and physiological significance of the metabolic regulation in β cells.Methods:A. IRS2 study regulation mi R-126(1) Bioinformatics method to predict micro RNAs-125-3p "seed" on the target gene sequence and IRS2Use mi Rbase, TargetScan, Bibiserve, Pictar microRNA target gene prediction and other Web sites and software, the secondary structure of the target gene and micro RNA-126-3p were predictive analysis, and find the "seed" sequence IRS2 mi R-126 function.(2) Dual luciferase reporter gene assay whether the regulation of gene expression of mi R-126 IRS2.(3) Western blot and Real-Time PCR were used to detect whether the regulation of gene expression of micro RNA-126-3p IRS2.Second, through the Real-Time PCR method, to detect high glucose INS1 cultured cells stimulated by high glucose detection of endogenous mi R-126-3p and IRS1, IRS2 expression.Third, the high glucose INS1 cells, CCK8 assay cell proliferation rate, study micro RNA-126 for βcell proliferation.Results:(1) Dual-luciferase reporter gene experiments confirmed, mi R-126-3p by seed sequence 3’UTR IRS-2 in the 6 bases can inhibit the expression of the luciferase reporter gene in the seed sequence Mutation of two bases can reverse this inhibitory prove mi R-126 can inhibit the expression of IRS2.(2) Western blot and Real-Time PCR method were proved, mi R-126-3P IRS2 can inhibit the expression of m RNA and protein levels, and significant changes in the proportion of IRS1 / IRS2.(3) High glucose can cause rapid increase of endogenous mi R-126-3p, 4h that reached its peak, IRS2 and IRS1 also a slight change, but change in IRS2 is more intense than the IRS1. Insulin can accelerates the endogenous mi R-126-3p.(4) mi R-126-3p can significantly inhibit the proliferation of INS-1 cells, high glucose increases micro RNA-126 can inhibit cell proliferation.Conclusion: mi R-126-3p inhibition of IRS2, can significantly alter the proportion of IRS1 / IRS2, inhibit cell proliferation.