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目的研究绞股蓝总苷(GPs)对胆固醇所致人脐静脉内皮细胞(hVECs)损伤的保护作用及机制。方法以体外培养的hVECs-12为研究对象,设正常对照组、溶媒组(二甲基亚砜)、胆固醇组(以50mg/L胆固醇处理48h)、3种浓度绞股蓝总苷(1、10、100μg/ml)组(按剂量在培养基中加入绞股蓝总苷预处理1h后再加入50mg/L的胆固醇处理48h)。分别用比色法、硝酸酶还原法及ELISA法检测细胞培液中乳酸脱氢酶(LDH)活性、一氧化氮(NO)浓度、单核细胞趋化蛋白-1(MCP-1)浓度;以DCFH-DA为荧光探针,流式细胞术检测细胞内活性氧水平;免疫细胞化学染色方法检测细胞内核因子-κBp65(NF-κBp65)核移位阳性细胞数。结果与正常对照组比较,胆固醇组各指标差异均有统计学意义(P<0.01);与胆固醇组比较,GPs可减少细胞培液中LDH活性、MCP-1浓度、并能增加NO浓度(P<0.01)、可降低细胞内ROS的水平(P<0.01);GPs能抑制NF-κB核移位(P<0.01),且随浓度升高有递减趋势。结论绞股蓝总苷对胆固醇所致的人脐静脉内皮细胞损伤具有保护作用,机制可能与减少细胞内ROS的生成,从而抑制NF-κB活化有关。
Objective To study the protective effect of gypenosides (GPs) on cholesterol-induced human umbilical vein endothelial cells (hVECs) injury and its mechanism. Methods The hVECs-12 cultured in vitro were divided into four groups: normal control group, DMSO group, cholesterol group (treated with 50 mg / L cholesterol for 48 h), three concentrations of Gypenosides (1, 100μg / ml) group (dose of Gynostemma glycosides in the medium pretreatment 1h and then added 50mg / L of cholesterol for 48h). The activities of lactate dehydrogenase (LDH), nitric oxide (NO) and monocyte chemoattractant protein-1 (MCP-1) in cell culture medium were detected by colorimetric method, nitrate reductase method and ELISA method, respectively. DCFH-DA was used as fluorescent probe to detect intracellular reactive oxygen species (ROS) levels by flow cytometry. The number of nuclear translocation-positive cells of nuclear factor-κBp65 (NF-κBp65) was detected by immunocytochemical staining. Results Compared with the normal control group, there were significant differences in each index of the cholesterol group (P <0.01). Compared with the cholesterol group, GPs could reduce the activity of LDH and MCP-1 and increase the concentration of NO in the culture medium <0.01). The GPs could inhibit the nuclear translocation of NF-κB (P <0.01), and decreased with increasing concentration of GPs. Conclusion Gypenosides has a protective effect on cholesterol-induced human umbilical vein endothelial cell injury, which may be related to the reduction of intracellular ROS production and the inhibition of NF-κB activation.