柚皮苷对高糖诱导的血管内皮细胞损伤及PI3K/AKT/eNOS信号通路的影响

Naringin protects human umbilical vein endothelial cells against injury induced by high glucose through PI3K/AKT/eNOS pathway

目的:研究柚皮苷(naringin,Nar)对高糖(high glucose,HG)诱导人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)损伤的作用及对PI3K/AKT/e NOS信号通路的影响。方法:利用HG(33 mmol/L glucose)培养基孵育HUVECs不同时间(6、12、24、48、72 h)后,用胰岛素(5 m IU/L)刺激细胞15 min,建立内皮细胞损伤模型,分别测定各组上清液中NO水平、细胞中e NOS和磷酸化e NOS(p-e NOS)的水平。对损伤的HUVECs用不同浓度的Nar(5、10、25、50、100 mg/L)孵育不同时间(6、12、24、48和72 h),用胰岛素(5 m IU/L)刺激细胞15min,检测细胞上清中的NO水平,观察Nar对HG诱导HUVECs损伤的作用。对损伤细胞先分别用PI3K抑制剂LY294002(10μmol/L)和AKT抑制剂AKT inhibitorⅣ(0.5μmol/L)预处理24 h,再用50 mg/L的Nar处理24 h,用胰岛素(5 m IU/L)刺激细胞15 min,检测细胞上清中NO的水平,以及e NOS、p-e NOS、PI3K、AKT及p-AKT的蛋白水平,探讨Nar减轻HG对HUVECs损伤作用的机制。结果:Nar的量效和时效结果显示,50 mg/L Nar预处理HUVECs 24 h后,其减轻HG对HUVECs损伤的作用最为显著,表现为NO和p-e NOS的水平升高(P<0.05)。加入PI3K和AKT抑制剂后,Nar减轻HG致内皮细胞的损伤及上调p-e NOS和NO水平的作用完全取消(P<0.05)。结论:Nar能减轻HG诱导的血管内皮细胞损伤,其作用机制可能是通过PI3K/AKT/e NOS信号通路介导的。

AIM：To investigate the protective effect of naringin （ Nar） on the injury of human umbilical vein endothelial cells （ HUVECs） induced by 33 mmol/L high glucose （ HG） and to explore its possible mechanisms.METH-ODS：The injury model was established by treating HUVECs with HG medium for the indicated time （6, 12, 24, 48 and 72 h） , and then the levels of NO, eNOS and p-eNOS were detected, respectively.The effects of Nar on high glucose-in-duced endothelial cell injury were observed.HUVECs were treated with Nar at concentrations of 5, 10, 25, 50 and 100 mg/L for 6 h, 12 h, 24 h, 36 h and 48 h.The levels of NO in the supernatants were measured.The effects of Nar on HG-injured HUVECs were explored by treating the cells with 10 μmol/L of LY294002, a PI3K inhibitor, or 0.5 μmol/L of AKT inhibitorⅣ, an AKT inhibitor, and then the levels of NO, PI3K, AKT, eNOS and their phosphorylated proteins were determined by Western blot.RESULTS：Nar at concentration of 50 mg/L significantly attenuated the injury of endothelial＆amp;nbsp;cells induced by high glucose （ P〈0.01） , and the protective effects of Nar were abolished by pretreating with the inhibitor of PI3K or AKT （P〈0.01）.CONCLUSION： Nar protects endothelial cells against the injury induced by high glucose through PI3K/AKT/eNOS pathway.