白藜芦醇抑制Gly-LDL诱导人脐静脉内皮细胞损伤的作用机制研究

Study on the inhibitory effect of resveratrol on injury of human umbilical vein endothelial cells

目的探究白藜芦醇(RSV)对糖化低密度脂蛋白(Gly-LDL)致人脐静脉内皮细胞(HUVECs)损伤的影响及作用机制。方法体外培养人脐静脉内皮细胞,Gly-LDL诱导其损伤,设置为模型组;用不同浓度(5、10、20μmol/L)白藜芦醇干预内皮细胞24 h,分别设置为白藜芦醇低浓度组、白藜芦醇中浓度组和白藜芦醇高浓度组;另设置空白组。通过CCK-8法检测细胞增殖能力,划痕实验观察细胞迁移能力,流式细胞术测定细胞凋亡水平,ELISA法检测内皮细胞中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)的含量,Western blot法检测Toll样受体4(TLR4)、缺氧诱导因子-1α(HIF-1α)、IL-6和TNF-α的蛋白表达,实时荧光定量PCR法检测TLR4、HIF-1α、IL-6和TNF-α的mRNA表达。结果与空白组比较,模型组细胞增殖和迁移能力下降(P<0.05,P<0.01),细胞凋亡率增加(P<0.01),血清IL-6和TNF-α含量增加(P<0.01),内皮细胞中TLR4、HIF-1α、IL-6和TNF-α的蛋白和mRNA表达均增加(P<0.05,P<0.01)。与模型组相比,白藜芦醇干预后,Gly-LDL损伤人脐静脉内皮细胞情况减轻,白藜芦醇中浓度组和白藜芦醇高浓度组的细胞增殖、迁移能力增强(P<0.05),细胞凋亡率下降(P<0.05),血清IL-6和TNF-α含量下降(P<0.05),内皮细胞中TLR4、HIF-1α、IL-6和TNF-α的蛋白表达和mRNA表达均减少(P<0.05)。结论白藜芦醇能有效减轻Gly-LDL对人脐静脉内皮细胞的炎症损伤,增强细胞增殖、迁移能力,抑制细胞凋亡,其机制可能与白藜芦醇下调TLR4/HIF-1α信号通路有关。

Objective To explore the effect and mechanism of resveratrol on human umbilical vein endothelial cells injury induced by glycosylated low density lipoprotein(Gly-LDL).Methods Human umbilical vein endothelial cells were cultured in vitro.The model group was induced by Gly-LDL.The low-concentration,medium-concentration and high-concentration RSV groups were established by treating with 5μmol/L,10μmol/L and 20μmol/L resveratrol respectively for 24 hours.The cell proliferative capacity was detected by Cell Counting Kit-8(CCK-8),the ability of cell migration was observed by scratch essay,the level of apoptosis was measured by flow cytometry,the contents of interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in endothelial cells were detected by ELISA,and the protein expressions of Toll-like receptor 4(TLR4),hypoxia-inducible factor-1(HIF-1α),IL-6 and TNF-αwere detected by Western-blot.The mRNA expressions of TLR4,HIF-1α,IL-6 and TNF-αwere detected by real-time fluorescence quantitative PCR.Results Compared with the blank group,the cell proliferative capacity and migration ability decreased(P<0.05,P<0.01),the rate of apoptosis increased(P<0.01),the contents of IL-6 and TNF-αin serum increased(P<0.01),and the protein expression and mRNA expression of TLR4,HIF-1α,IL-6 and TNF-αin endothelial cells increased in the model group(P<0.05,P<0.01).Compared with the model group,the injury of human umbilical vein endothelial cells induced by Gly-LDL was alleviated after the intervention of resveratrol.The ability of cell proliferation and migration was enhanced(P<0.05),the apoptosis rate was decreased(P<0.05),the contents of serum IL-6 and TNF-αwere decreased(P<0.05),and the protein expression and mRNA expression of TLR4,HIF-1α,IL-6 and TNF-αin endothelial cells were decreased in the medium-concentration and high-concentration RSV groups(P<0.05).Conclusion Resveratrol seems to reduce the inflammatory injury of human umbilical vein endothelial cells induced by Gly-LDL,enhance the ability of cell proliferation and migration,and inhibit cellapoptosis,which may be related to the down-regulation of TLR4/HIF-1αsignal pathway by resveratrol.