高糖抑制PI3K/AKT通路诱导人冠状动脉内皮细胞凋亡的研究

目的探讨高糖对人冠状动脉内皮细胞凋亡及微颗粒释放的影响及机制。方法根据细胞培养基中葡萄糖浓度不同分为3组：正常对照组（5．5mmol/L），中糖组（17．6mmo/L），高糖组（33．3mmol/L），另设甘露醇对照组（20．0mmol/L）。通过Hoechst 33258荧光染色观察凋亡形态学变化，应用流式细胞术检测内皮微颗粒细胞凋亡率及微颗粒，ELISA法检测人冠状动脉内皮细胞的凋亡相关蛋白Bad、Bax的表达以及PBK、p-Akt蛋白的表达。结果随着葡萄糖浓度增加，人冠状动脉内皮细胞微颗粒的释放及细胞凋亡率逐渐增加，高糖组最高（P〈0．01）；人冠状动脉内皮细胞分泌Bad、Bax逐渐增多，高糖组最高（P〈0．01）；人冠状动脉内皮细胞分泌PBK、p-Akt逐渐减少，高糖组减少最明显（P〈0．01）。与甘露醇对照组比较，正常对照组微颗粒的释放，细胞凋亡率，分泌Bad、Bax、P13K、p-Akt差异均无统计学意义（P〉0．05）。结论高糖可以促进人冠状动脉内皮细胞凋亡及微颗粒的释放．其机制可能同PDK／AKT途径相关。

High glucose induces human endothelial cell apoptosis through inhibiting PDK/AKT pathway

Objective To investigate the effect of high glucose on human coronary endothelial cell （HCAEC） apoptosis and its mechanism. Methods Cultured HCAECs were treated with normal （5.5 mmol/L）, medium （17.6 mmol/L） and high concentration of glucose （33.3 mmol/L）, or mannitol （20.0 mmol/L）, respectively. Hoechst 33258 fluorescence staining was used to observe the morphological changes of apoptosis. The absolute number of endothelial microparticles and the apoptotic cells percentage were identified by flow cytometer; the expression of Bad, Bax, PI3K and p- Akt in cell lysates were detected by ELISA. Results HCAECs microparticles and apoptotic cells were increased with the increasing glucose concentrations and those in high glucose group increased most （ P 〈 0.01 ）. Bad and Bax secreted by HCAECs increased with the increasing of glucose concentrationsand those in high glucose group increased most （P 〈 0. 01 ）. PI3K and p-Akt secreted by HCAECs secreted were decreased and those in high glucose group decreased most （P 〈0. 01）. There were no significant differences in HCAECs microparticles, apoptotic cells, Bad, Bax, PI3K and p-Akt expressions between control group and mannitol group （P 〉 0.05 ）. Conclusion High glucose induces HCAECs apoptosis and EMPs release, in which PI3K/AKT pathway may be involved.