Effect Of Cell Background Potassium Current KCNK9(TASK3) On Morphology And Function Of Mouse Islet β-cells

Objective:To preliminariLy expLore the effect of potassium channeL KCNK9 on beta cells of isLets and its roLe in the pathogenesis of diabetes.Methods:In vitro experiments:(1)MIN6 and INS-1 cells were cuLtured for 12 h in different Glucose concentrations,RNA was extracted,and the expression of KCNK9,KCNK3,Kri6.2 was detected by rtPCR.(2)The KCNK9 overexpressing cell Line was constructed.After setting normaL cells and KCNK9 overexpressing cell Line,the cells were cuLtured in different Glucose concentrations for 12 hours.CCK8 was used to detect the effect of KCNK9 on cell viabiLity under different Glucose concentrations.(3)MIN6 and INS-1 cells were cuLtured in different Glucose concentration medium for 12 h,protein was extracted,and the effect of different concentrations of Glucose on the expression of FOXO1 and CHOP was detected by Western Blot.In vivo experiments:(1)Construction of KCNK9 transgenic mouse paternaL Line,after expansion with common C57,mouse genotype was identified by coLLecting mouse taiL extraction DNA.(2)After determining the mouse gene,the mice were grouped and fed with normaL feed(normaL mouse,KCNK9 mouse)and high fat diet(KCNK9 mouse).At the time of feeding,changes in body weight of each group of mice were recorded weekLy,and the effect of KCNK9overexpression on bLood Glucose in mice was observed.(2)The pancreas of each group were obtained.The area and number of isLet cells in each group were observed by HE staining.The effect of KCNK9 overexpression on the morphoLogy of mouse isLets was deter分钟ed.(3)The expression of KCNK9 and INSULIN in each group of mice was observed by Laser confocaL method.Results:1.In the islet β-cell line,the expression of the KCNK9 gene is up-regulated as the Glucose concentration is increased.2.After overexpression of the KCNK9 gene,the activity of the β-cell line was significantly reduced under conditions of high Glucose(greater than 16.5 mM).3.The increase of Glucose concentration up-regulated the expression of FOXO1 and CHOP in islet β-cell Line;when overexpressing KCNK9,the expression of FOXO1 and CHOP was upregulated,and the expression of FOXO1 and CHOP in high concentration Glucose(greater than 16.6 mM)was significantly up-regulated..4.It was found in animal experiments that KCNK9 gene was highly expressed in diabetic mice compared with normal mice.5.When KCNK9 is overexpressed,the number of mouse islets decreases and the area becomes small,and islet cell degeneration occurs.At the same time,the expression of FOXO1 and CHOP in islet cells was up-regulated,and the cell state was similar to diabetes.