Effects Of Ca²⁺ Activated Transcription Factor Egr-1 On The Pancreatic ?-cells

Background:Type 2 diabetes is characterized by persistent hyperglycemia-induced ?-cell function decline and enhanced peripheral tissue insulin resistance.As the disease progresses,peripheral tissue insulin resistance increases,requiring more insulin to clear accumulated glucose in the plasma.Egr-1,a highly enriched Ca2+-dependent transcription factor in islets,can regulate various target gene expression in response to various stimuli.Egr-1 can now regulate cell proliferation,survival,cancer cell apoptosis,and negative regulation of insulin resistance.However,there are few studies on whether Egr-1 can regulate insulin synthesis and secretion.In this experiment,pancreatic ? cells were used as a model to investigate the effect of Egr-1 on islet ? cell function.Materials and Methods:INS-1 cells were treated with high glucose and high KCl for 5 min and 15 min,and the expression of Egr-1 was detected by QPCR.The expression of Egr-1 in INS-1 cells was silenced by siRNA,and the expression of INS1,INS2,MafA,Pdx-1,Cav1.2,Cav1.3,SNAP-25,Synataxin 1A and Vamp2 genes was detected by QPCR.Insulin cells with low expression of Egr-1 were stimulated with low-sugar,high-glucose and high-potassium SAB solutions and insulin secretion was detected.Egr-1 knockdown INS-1 cells were then stimulated with low sugar and high sugar SAB solutions and total ATP was detected.Finally,Egr-1 knockdown INS-1 cells were stimulated with low glucose,high glucose and high potassium Kreb's solution for 20s,2 min,3 min and 10 min to detect calcium images.Results:1)After QPCR detection,Egr-1 gene expression was significantly increased compared with the control group.2)Egr-1 silencing significantly down-regulated MafA,Pdx-1 and INS2 gene expression at mRNA levels,with no effect on other genes.3)Insulin secretion was reduced in the low glucose and high potassium treatment groups,but there was no change in the high glucose group.Insulin content was lower in the low-glucose and high-glucose groups,but not in the high-potassium group.Proinsulin were significantly reduced in all groups.4)There was no change in total ATP in the low sugar and high sugar treatment groups.5)There was no change in the[Ca2+]i peak in the high potassium group,and there was no change in the Ca2+ load curve integral(AUC)in the high potassium group and the low sugar group,and the Ca2+ load curve integral in the high sugar group was significantly increased.Conclusion:The Egr-1 gene regulates the expression of MafA and Pdx-1 genes in pancreatic ? cells,thereby regulating the expression and synthesis of the insulin INS gene.Egr-1 expression is increased when stimulated by acute high glucose and high potassium.Egr-1 gene silencing reduced insulin synthesis,decreased proinsulin in all groups,decreased insulin levels at low and high glucose levels,and decreased insulin secretion at low and high potassium levels.It is speculated that although the intracellular Ca2+ ion concentration is increased at high glucose,insulin secretion is unchanged because the insulin content is decreased.