基于NFAT5介导的EMT研究益糖康改善高血糖对肾脏早期损伤的作用机制  

Research on Mechanism of Yitangkang(益糖康)Improving Early Renal Injury Induced by Hyperglycemia Based on NFAT5 Mediated EMT

作　　者：郭隽馥[1,2] 李乡南 杨宇峰[1] 石岩 GUO Junfu;LI Xiangnan;YANG Yufeng;SHI Yan(Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning,China;Shenyang Key Laboratory for TCM Emotional Disorder,Shenyang 110847,Liaoning,China)

机构地区：[1]辽宁中医药大学,辽宁沈阳110847 [2]沈阳市中医神志病学重点实验室,辽宁沈阳110847

出　　处：《中华中医药学刊》2024年第1期35-40,I0007,共7页Chinese Archives of Traditional Chinese Medicine

基　　金：中国博士后科学基金面上项目(2019M65141);辽宁省“兴辽英才计划”青年拔尖人才项目(XLYC2203180);辽宁省“百千万人才工程”项目(2020921039)。

摘　　要：目的探讨活化T细胞核因子5(nuclear factor of activated T cells 5,NFAT5)对高糖诱导人肾小管上皮HK-2细胞上皮间质转化(epithelial-mesenchymal transition,EMT)的影响以及益糖康改善高血糖对肾脏早期损伤的作用机制。方法SD雄性大鼠20只,随机分成生理盐水组和益糖康组,每组10只。生理盐水组给予1 mL/100 g生理盐水灌胃,益糖康组给予2.10 g/100 g益糖康煎剂灌胃,连续5 d,腹主动脉取血,离心分离上清。通过高糖诱导HK-2细胞构建早期糖尿病肾病体外模型,并加入相应的药物进行干预。分别采用qRT-PCR和Western blot实验检测转染NC-siRNA(NC-siRNA组)/NFAT5-siRNA(NFAT5-siRNA组)或生理盐水含药血清处理(NS组)/益糖康含药血清处理(YTK组)以及益糖康含药血清处理同时转染GV492-Empty(YTK+GV492-Empty组)/GV492-NFAT5(YTK+GV492-NFAT5组)细胞中NFAT5、E-cadherin以及Vimentin mRNA和蛋白表达水平的改变。再分别采用划痕和Transwell实验检测各组细胞迁移力和侵袭力的改变。结果转染NFAT5-siRNA能够明显抑制NFAT5 mRNA(P<0.05)和蛋白(P<0.05)的表达;与NC-siRNA组相比较,NFAT5-siRNA组细胞中E-cadherin mRNA(P<0.05)和蛋白(P<0.01)的表达水平显著升高,Vimentin mRNA(P<0.05)和蛋白(P<0.01)的表达水平显著降低,细胞的迁移力(P<0.05)和侵袭力(P<0.01)均明显减弱。与NS组相比较,YTK组细胞中NFAT5 mRNA(P<0.05)和蛋白(P<0.05)的表达水平明显降低。转染GV492-NFAT5能够明显促进NFAT5 mRNA(P<0.05)和蛋白(P<0.01)的表达;与YTK+GV492-Empty组相比较,YTK+GV492-NFAT5组细胞中E-cadherin mRNA(P<0.01)和蛋白(P<0.01)的表达水平显著降低,Vimentin mRNA(P<0.05)和蛋白(P<0.01)的表达水平显著升高,细胞的迁移力(P<0.05)和侵袭力(P<0.05)均明显增强。结论在高糖环境下,NFAT5会促进人肾小管上皮细胞发生EMT;益糖康能够通过下调NFAT5表达抑制肾小管上皮细胞发生EMT从而改善高血糖对肾脏的早期损伤。Objective To investigate the effect of nuclear factor of activated T cells 5(NFAT5)on epithelial-mesenchymal transition(EMT)of human renal tubular epithelial HK-2 cells induced by high glucose and the mechanism of Yitangkang(益糖康)improving early renal injury induced by hyperglycemia.Methods Twenty male SD rats were randomly divided into normal saline group and Yitangkang group,with 10 rats in each group.Normal saline group was given normal saline by intragastric administration(1 mL/100 g)and Yitangkang group was given Yitangkang decoction by intragastric administration(2.10 g/100 g),for 5 consecutive days.The abdominal aorta blood was taken out and the supernatant was separated by centrifugation.HK-2 cells were induced by high glucose to construct an in vitro model of early diabetic nephropathy and the corresponding drugs were added to intervene the cells.The qRT-PCR and Western Blot assays were performed to detect the expression levels of NFAT5,E-cadherin and Vimentin mRNA and protein in the cells transfected with NC-siRNA(NC-siRNA group)/NFAT5-siRNA(NFAT5-siRNA group)or the cells treated with normal saline drug-containing serum(NS group)/Yitangkang drug-containing serum(YTK group)or the cells treated with Yitangkang drug-containing serum and transfected with GV492-Empty(YTK+GV492-Empty group)/treated with Yitangkang drug-containing serum and transfected with GV492-NFAT5(YTK+GV492-NFAT5 group),respectively.The wound healing and Transwell assay were used to test the migration and invasion abilities of cells in different groups.Results Transfection of NFAT5-siRNA could significantly inhibit the expressions of NFAT5 mRNA and protein(P<0.01,P<0.05).Compared with those of the NC-siRNA group,the expression levels of E-cadherin mRNA and protein(P<0.05,P<0.01)in NFAT5-siRNA group were significantly increased,the expression levels of Vimentin mRNA and protein(P<0.05,P<0.01)were significantly decreased.The migration and invasion abilities of cells were significantly decreased(P<0.05,P<0.01).Compared with those of the

关 键 词：T细胞核因子5 细胞上皮间质转化 益糖康 糖尿病肾病 

分 类 号：R289.5[医药卫生—方剂学]