摘要
目的探讨四跨膜蛋白8(Tspan 8)对糖尿病肾小管上皮细胞自噬活性的影响及机制。方法将HK-2细胞分为正常组(NG,5.5mmol/L D-葡萄糖)和高糖组(HG,30mmol/L D-葡萄糖)。高糖培养的HK-2细胞分别转染Tspan 8基因的随机无靶点阴性对照(shCtrl)和特异性基因敲减序列(siRNA)。Western blot和qPCR检测Tspan 8表达;Western blot检测蛋白激酶B(Akt)及其磷酸化水平(p-Akt),哺乳动物雷帕霉素靶蛋白(mTOR)及其磷酸化水平(Ⅱ/Ⅰ)。结果高糖刺激后,Tspan 8表达升高约1.5倍(P<0.01);与shCtrl组比较,Tspan 8siRNA组LC3Ⅱ/LC3Ⅰ比值升高3倍,p-Akt表达降低约1.5倍,p-mTOR表达降低约2倍(P<0.01)。结论 Tspan8可能通过激活mTOR信号通路,负向调节糖尿病肾小管上皮细胞的自噬活性。
Objective To analyze the effects and molecular mechanism of Tetraspanin 8（Tspan 8）on autophagy activity of diabetic renal tubular epithelial cells. Methods The cultured HK-2 cells were divided into 2 groups：normal group（NG,5.5 mmol/L D-glucose）,high glucose group（HG,30 mmol/L D-glucose）.In HG group,HK-2 cells were transfected with random non-target negative control（shCtrl）and Tspan 8 specific knockdown sequences（siRNA）,respectively.The expression of Tspan 8 was measured by western blot and real-time fluorescence quantitative PCR.The expression of Akt,p-Akt,mTOR,p-mTOR,LC3Ⅰ and LC3Ⅱ was detected by western blot. Results After high glucose stimulation,the expression of Tspan 8 increased by approximately 1.5 folds（P〈0.01）.Compared with shCtrl group,the Tspan 8 siRNA group had a 3-fold increase in LC3Ⅱ/LC3Ⅰratio,a 1.5-fold decrease in p-Akt expression and a2-fold decrease in p-mTOR expression（P〈0.01）. Conclusion Tspan 8 may negatively regulate the autophagy activity of diabetic renal tubular epithelial cells by activating the mTOR signaling pathway.
作者
庄兰艮
杨青青
金国玺
时照明
ZHUANG Langen;YANG Qingqing;J IN Guoxi(Department of Endocrinology,The First Affiliated Hospital of Bengbu Medical College,Bengbu 233000,China)
出处
《中国糖尿病杂志》
CAS
CSCD
北大核心
2018年第10期860-863,共4页
Chinese Journal of Diabetes
基金
蚌埠医学院科技发展基金(BYKF1734)
关键词
四跨膜蛋白
糖尿病
肾小管
自噬
Tetraspanin
Diabetes mellitus
Renal tubule
Autophagy
