恩格列净及达格列净对棕榈酸诱导的心肌细胞损伤的保护作用及其机制

Protective Effect of Empagliflozin and Dapagliflozin on Palmitic Acid-induced Cardiomyocytes Injury and Its Underlying Mechanism

关键词:  恩格列净  达格列净  棕榈酸  心肌细胞

Key words:  empagliflozin  dapagliflozin  palmitic acid  cardiomyocytes

摘要:

目的 探讨钠-葡萄糖共转运蛋白-2抑制剂恩格列净（empagliflozin，EMPA）及达格列净（dapagliflozin，DAPA）对棕榈酸（palmitic acid，PA）诱导H9c2心肌细胞损伤的保护作用及其可能机制。方法 采用PA诱导H9c2心肌细胞损伤，CCK-8法筛选PA、EMPA及DAPA的最佳给药浓度；Western blotting检测TLR4、p-AKT、p-mTOR、Nrf2和HO-1的蛋白表达水平；ELISA法检测IL-1β、IL-6和TNF-α的含量；荧光显微镜和流式细胞术检测PA诱导H9c2心肌细胞中ROS水平。结果 PA 100 μmol·L^–1刺激24 h可诱导H9c2心肌细胞存活率明显下降（P<0.01），IL-1β、IL-6、TNF-α、TLR4蛋白表达、p-mTOR蛋白表达及ROS水平明显增加（P<0.01），p-AKT、Nrf2和HO-1蛋白表达显著降低（P<0.01）；与模型组比较，经EMPA及DAPA处理后，细胞存活率明显增加（P<0.01），IL-1β、IL-6、TNF-α、TLR4蛋白表达、p-mTOR蛋白表达及ROS水平明显降低（P<0.05或P<0.01），p-AKT、Nrf2和HO-1蛋白表达显著上调（P<0.05或P<0.01）。结论 EMPA及DAPA对PA诱导的心肌细胞损伤均具有保护作用，其机制可能与下调TLR4、p-mTOR蛋白表达，增强AKT蛋白磷酸化，激活Nrf2/HO-1通路，抑制ROS的生成有关。

Abstract:

OBJECTIVE To investigate the protective effect of sodium-glucose cotransporter 2 inhibitors empagliflozin (EMPA) and dapagliflozin(DAPA) on palmitic acid(PA)-induced injury of rat H9c2 cardiomyocytes and its possible mechanism. METHODS H9c2 myocardial cell injury was induced by PA. CCK-8 method was used to screen the optimal dosage of PA, EMPA and DAPA. The protein expression levels of TLR4, p-AKT, p-mTOR, Nrf2 and HO-1 were detected by Western blotting. The expressions of IL-1β, IL-6, TNF-α were detected by ELISA. ROS levels in PA-induced H9c2 cardiomyocytes were determined by fluorescence microscopy and flow cytometry. RESULTS The survival rate of H9c2 cardiomyocytes decreased significantly after treatment of PA 100 μmol·L^-1 for 24 h(P<0.01), the expression of IL-1β, IL-6, TNF-α, TLR4, p-mTOR and ROS were significantly increased(P<0.01), and the protein expression of p-AKT, Nrf2 and HO-1 was significantly decreased(P<0.01). Compared with the model group, the cell survival rate was significantly increased after EMPA and DAPA treatment(P<0.01), the expression of IL-1β, IL-6, TNF-α, TLR4, p-mTOR and ROS levels were significantly decreased(P<0.05 or P<0.01), and the protein expression of p-AKT, Nrf2 and HO-1 was significantly up-regulated(P<0.05 or P<0.01). CONCLUSION EMPA and DAPA have protective effects on PA-induced cardiomyocyte injury, and the mechanism may be related to down-regulation of TLR4 and p-mTOR protein expression, enhancement of AKT protein phosphorylation, activation of Nrf2/HO-1 pathway, and inhibition of ROS generation.