KLF9沉默对高糖刺激诱导的内皮细胞损伤的影响

KLF9 silencing reduces endothelial cell injury induced by high glucose

目的:探讨Krüppel样因子9(KLF9)对高糖刺激下内皮细胞损伤的影响。方法:人脐静脉内皮细胞(HUVEC)分为4组,空白对照组不处理,其他3组分别给予高糖(33.5 mmol/L葡萄糖)刺激12、24、48 h,采用qRT-PCR检测细胞中KLF9 mRNA表达水平。另取HUVEC分为3组,ScRNA组(转染空载体ScRNA12 h后用5.5 mmol/L葡萄糖处理48 h)、HG-ScRNA组(转染空载体12 h后用33.5 mmol/L葡萄糖处理48 h)和HG-KLF9 siRNA组(转染KLF9 siRNA 12 h后用33.5 mmol/L葡萄糖处理48 h),采用CCK-8法检测各组细胞活力,TUNEL染色检测细胞凋亡,ELISA法检测细胞中炎症因子水平,氧化应激因子检测试剂盒检测细胞氧化应激水平,免疫荧光染色检测NF-E2相关因子2(NRF2)表达及核转位水平。结果:与空白对照组相比,高糖刺激24、48 h后细胞中KLF9 mRNA表达水平升高(P<0.05)。与ScRNA组相比,HG-ScRNA组细胞中炎症因子(TNF-α、IL-1和IL-6)水平增高,细胞活性氧和丙二醛含量升高,超氧化物歧化酶2以及谷胱甘肽还原酶活性降低,细胞增殖能力降低,凋亡增多,NRF2表达以及核转位水平降低(P<0.05);与HG-ScRNA组相比,HG-KLF9 siRNA组以上指标均有改善(P<0.05)。结论:沉默KLF9可减轻高糖诱导的内皮细胞炎症、氧化应激水平,恢复NRF2的核表达,从而减轻内皮细胞损伤。

Aim:To explore the effects of KLF9 on endothelial cell injury under high glucose.Methods:Human umbilical vein endothelial cells(HUVEC)were divided into 4 groups,the blank control group was not treated,and the other 3 groups were stimulated with high glucose(33.5 mmol/L glucose)for 12,24 and 48 hours,respectively.The expression of KLF9 mRNA was detected by qRT-PCR.HUVEC were divided into 3 groups,ScRNA group(ScRNA transfection for 12 hours and 5.5 mmol/L glucose treatment for 48 hours),HG-ScRNA group(ScRNA transfection for 12 hours and 33.5 mmol/L glucose treatment for 48 hours)and HG-KLF9 siRNA group(KLF9 siRNA transfection for 12 hours and 33.5 mmol/L glucose treatment for 48 hours).CCK-8 assay was used to detect cell viability,TUNEL staining was used to detect cell apoptosis,ELISA was used to detect the levels of inflammatory factors,oxidative stress level was detected by testing kits,and immunofluorescence staining was used to detect the expression and nuclear translocation of NRF2.Results:KLF9 mRNA in endothelial cells increased after 24 and 48 hours high glucose stimulation(P<0.05).Compared with ScRNA group,the levels of inflammatory factors(TNF-α,IL-1 and IL-6)in HG-ScRNA group were increased significantly,the levels of ROS and MDA were increased,the activities of SOD2 and Gpx were decreased,the proliferation of cells was reduced,the apoptosis was increased,and the expression and nuclear translocation of NRF2 were decreased(P<0.05);compared with HG-ScRNA group,the above indexes in HG-KLF9 siRNA group were all improved(P<0.05).Conclusion:KLF9 silencing can alleviate the inflammation and oxidative stress induced by high glucose,and restore the expression of NRF2,thus alleviate the injury of endothelial cells.