摘要
目的:探讨沉默生长分化因子15(GDF15)对高糖诱导的人肾小管上皮细胞HK-2凋亡和氧化应激的影响及作用机制。方法:采用Western blot法检测16例糖尿病肾病(DN)患者和正常肾组织中GDF15蛋白表达水平。转染GDF15小干扰RNA(si-GDF15)、无义阴性序列(si-NC)至HK-2细胞,Western blot法检测细胞中GDF15蛋白水平(验证转染效果)。HK-2细胞分为对照组、高糖组、si-GDF15+高糖组、si-NC+高糖组,流式细胞仪检测各组细胞凋亡,2′,7′-二氯二氢荧光素二乙酸酯(DCFH-DA)法检测各组细胞中活性氧簇(ROS)含量,酶联免疫吸附法检测各组细胞中丙二醛(MDA)、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)水平,Western blot法检测各组细胞中Bcl-2、Bax、细胞核因子E2相关因子2(Nrf2)和血红素加氧酶-1(HO-1)蛋白表达水平。结果:与正常肾组织比较,DN患者肾组织中GDF15蛋白表达水平升高(P<0.001)。与对照组比较,高糖组HK-2细胞凋亡率、细胞中Bax、HO-1和细胞核Nrf2蛋白、MDA以及ROS水平升高(P<0.001),细胞中Bcl-2和细胞质Nrf2蛋白水平、SOD和GSH-Px水平降低(P<0.001)。与si-NC+高糖组比较,si-GDF15+高糖组HK-2细胞凋亡率、细胞中Bax和细胞质Nrf2蛋白、MDA及ROS水平降低(P<0.001),细胞中Bcl-2、HO-1和细胞核Nrf2蛋白水平、SOD和GSH-Px水平升高(P<0.001)。结论:沉默GDF15基因表达可抑制高糖诱导的肾小管上皮细胞HK-2凋亡和氧化应激,其作用机制与激活Nrf2/HO-1信号通路有关。
Aim:To investigate the effects of silencing growth differentiation factor 15(GDF15)on high glucose-induced apoptosis and oxidative stress in human renal tubular epithelial cells HK-2 and its mechanism.Methods:Western blot method was used to detect the expression levels of GDF15 protein in 16 patients with diabetic nephropathy(DN)and normal kidney tissues.si-GDF15 and si-NC were transfected into HK-2 cells,and Western blot was used to detect the GDF15 protein level in cells to verify transfection efficiency.HK-2 cells were divided into control group,high glucose group,si-GDF15+high glucose group,and si-NC+high glucose group.Flow cytometry was used to detect the apoptosis of cells.DCFH-DA method was used to detect the content of reactive oxygen species(ROS)in cells.ELISA was used to detect the levels of MDA,SOD and GSH-Px in cells,and Western blot was used to detect the levels of Bcl-2,Bax,Nrf2 and HO-1 protein.Results:Compared with normal kidney tissues,the expression of GDF15 protein in renal tissues of DN patients was significantly increased(P<0.05).Compared with the control group,the apoptosis rate of HK-2 cells,the levels of Bax,HO-1 and nuclear Nrf2 protein,and the levels of MDA and ROS in the high glucose group were significantly increased(P<0.001),while the levels of Bcl-2 and cytoplasmic Nrf2 protein and the levels of SOD and GSH-Px were significantly decreased(P<0.001).Compared with si-NC+high glucose group,the apoptosis rate of HK-2 cells,the levels of Bax and cytoplasmic Nrf2 protein,and the levels of MDA and ROS in si-GDF15+high glucose group were significantly decreased(P<0.001),while the levels of Bcl-2,HO-1 and nuclear Nrf2 protein and the levels of SOD and GSH-Px were significantly increased(P<0.001).Conclusion:Silencing GDF15 gene can inhibit high glucose-induced apoptosis and oxidative stress in renal tubular epithelial cells,and its mechanism is related to the activation of Nrf2/HO-1 signaling pathway.
作者
郭海燕
邢志华
王丽丽
解菊芬
GUO Haiyan;XING Zhihua;WANG Lili;XIE Jufen(Department of Internal Medicine, Clinical Department, Fenyang College, Shanxi Medical University, Fenyang,Shanxi 032200;Department of Endocrinology, Fenyang Hospital, Shanxi Province, Fenyang,Shanxi 032200;Department of Hematology, Fenyang Hospital, Shanxi Province, Fenyang,Shanxi 032200)
出处
《郑州大学学报（医学版）》
CAS
北大核心
2020年第3期368-373,共6页
Journal of Zhengzhou University(Medical Sciences)
基金
山西省自然科学基金项目(2017016050)。
