高糖环境对人脐静脉血管内皮细胞凋亡及Caspase-3表达的影响

Effects of high glucose on apoptosis of human umbilical vein endothelial cells and expression of Caspase-3

目的探讨高糖环境对人脐静脉血管内皮细胞(HUVEC)凋亡的影响,及其与Caspase-3表达的相关性。方法将培养成功的HUVEC以不同浓度的葡萄糖培养48 h,TUNEL法观察细胞凋亡情况;流式细胞仪(Annexin V-FITC及PI双荧光)检测细胞存活、凋亡和坏死情况;Western blotting法观察细胞中活性Caspase-3的表达。结果光学显微镜观察高糖组(33 mmol/L)内凋亡细胞明显多于正常糖浓度组(5.5 mmol/L);随机选择3个视野,高糖组TUNEL阳性的细胞数多于正常糖浓度组。流式细胞仪结果显示:正常糖浓度组细胞凋亡率为(4.10±2.16)%,细胞坏死率为(0.47±0.05)%,存活细胞率为(94.44±2.13)%;高糖组细胞凋亡率为(16.13±1.51)%,细胞坏死率为(1.09±0.27)%,存活细胞率为(81.99±1.13)%,凋亡细胞百分率与正常糖浓度组相比差异有统计学意义(P<0.05);Western blotting法显示正常糖浓度组Caspase-3/β-actin为0.062±0.005,高糖组Caspase-3/β-actin为0.125±0.05,差异有统计学意义(P<0.05),高糖组活性Caspase-3表达量增强。结论高糖环境培养下的HUVEC凋亡增加,可能与活性Caspase-3表达增强有关。

Objective To investigate the effects of high glucose on the apoptosis of human umbilical vein endothelial cells（HUVECs） and its correlation with Caspase-3 expression.Methods HUVECs were cultured by glucose solution of different concentrations for 48 h.The apoptosis of cells was observed by the TUNEL staining.The survival,apoptosis,and necrosis of cells were detected by the flow cytometry（Annexin V-FITC and PI dual fluorescence）.The expression of cleaved Caspase-3 of cells was observed by the Western Blot analysis.Results The apoptotic cells of the high glucose group（33 mmol/L） were significantly more than those of the normal glucose group（5.5 mmol/L） under the optic microscope.The TUNEL-positive nuclei of the high glucose group were more than those of the normal glucose group in 3 randomly selected fields.The results of flow cytometry showed that for the normal glucose group;the apoptosis rate was（4.10 ±2.16）%,the necrosis rate was（0.47 ± 0.05）%;and the survival rate was（94.44 ±2.13）%.While for the high glucose group,the apoptosis rate was（16.13 ± 1.51）%;the necrosis rate was（1.09 ± 0.27）%;and the survival rate was（81.99 ± 1.13）%.The difference of apoptosis rate of two groups was statistically significant（P 〈 0.05）.The results of Western blotting method showed that the Caspase-3/β-actin levels of the normal glucose group and high glucose group were 0.062 ±0.005 and 0.125 ±0.05,respectively.The difference was statistically significant（P 〈0.05）.The expression of cleaved Caspase-3 of the high glucose group increased.Conclusion The apoptosis of HUVECs under high glucose increases,which may be relevant to the increase of expression of cleaved Caspase-3.