缺氧复氧对内皮细胞间隙连接通讯功能的影响初探

Effect of Hypoxia/Reoxygenation on Gap Junctional Intercellular Communication in Cultured Human Endothelial Cells

目的 观察缺氧复氧刺激对内皮细胞间隙连接通讯功能的影响 ,初步探讨缺氧复氧引起内皮细胞损伤的机制。方法 将人脐静脉内皮细胞株 ECV30 4分为两组 ,常氧对照组在常氧状态下培养 12～ 18小时 ;缺氧复氧组先在氧浓度低于 1%状态下培养 12～ 14小时 ,再置于常氧状态复氧 0～ 6小时。应用荧光漂白回复技术对细胞间隙连接通讯功能进行分析 ,结果用荧光回复率表示。结果 常氧对照组内皮细胞荧光回复率为 34 .4%±16 .5 %;缺氧复氧组经缺氧处理以及复氧 4小时和复氧 6小时处理后 ,其内皮细胞荧光回复率分别为 42 .7%±2 2 .7%、37.8%± 13.2 %和 43.0 %± 16 .2 %,与常氧对照组相比差异无显著性 (P>0 .0 5 ) ;经复氧 2小时处理后 ,内皮细胞的荧光回复率为 2 5 .0± 7.4%,与常氧对照组和经缺氧、复氧 4小时、复氧 6小时处理组相比 ,差异十分显著(P<0 .0 1)。结论 在本实验条件下 ,单独缺氧 12～ 14小时不影响内皮细胞间隙连接通讯功能 ,而复氧 2小时则能显著抑制间隙连接通讯功能。由此提示 ,间隙连接通讯功能改变参与了内皮细胞缺氧复氧损伤的早期反应。

Objective This study was conducted to investigate the effect of hypoxia/reoxygenation on endothelial gap junctional intercellular communication (GJIC) and make an approach to the possible mechanism therein involved. Methods After human umbilical vein endothelial cells (ECs) had grown to confluence, they were divided into two groups and were exposed to different conditions. ECs in group one were incubated in normoxia for 12-18 hours. ECs in group two were exposed to hypoxia (pO 2<1%) for 12-14 hours and then were reoxygenated in normoxia for 0-6 hours. GJIC was detected by means of a fluorescence recovery after photobleaching (FRAP) and was represented by the fluorescence recovery rate. Results In normoxia group, the fluorescence recovery rate of ECs was 34.4%±16.5%. There was no difference in fluorescence recovery rate between the normoxia group, the hypoxia sub-group, and the 4-hour and 6-hour reoxygenation sub-groups (P>0 05). However, after 2 hours of reoxygenation treatment, the fluorescence recovery rate (25.0%±7.4%) was significantly lower than that of the normoxia group, the hypoxia sub-group, and the 4-hour and 6-hour reoxygenation sub-groups (P<0.01). Conclusion Under the condition of this experiment, hypoxia treatment alone did not influence endothelial GJIC, but GJIC was significantly reduced after 2 hours of reoxygenation. These suggest that the functional change of GJIC has been involved in the early reaction to the adverse effects of hypoxia/reoxygenation on endothelial cells.