缺血缺氧损伤对肠上皮细胞肌动蛋白及其结合蛋白的影响

目的 观察大鼠肠上皮细胞(IEC)缺血缺氧损伤后肌动蛋白及其结合蛋白的变化,探讨其与肠上皮细胞凋亡的关系及钙通道阻断剂对其的影响.方法 建立大鼠IEC分离与原代培养和体外模拟缺血缺氧环境,设立对照组(A组),缺血组(B组),缺氧组(C组),缺血缺氧组(D组)及加用维拉珀米2 mg/L的相应对照组.利用流式细胞仪(FCM)计算凋亡细胞率,激光共聚焦显微镜技术(LSCM)观察肌动蛋白(F-actin)、踝蛋白(talin)和α-辅肌动蛋白(α-actin)的形态及荧光强度的变化.结果 B、C、D组IEC凋亡较A组明显增加(P<0.05);加用钙通道阻断剂后的B1、C1、D1组较相应对照组减少(P<0.05);缺血缺氧损伤导致F-actin、talin和α-actin形态的改变,以D组最显著,同时B、C、D组的荧光强度较A组明显减弱(P<0.01),加用钙通道阻断剂后的B1、C1、D1组F-actin、talin和α-actin形态有所恢复,荧光强度增强,与损伤组比较,差异有统计学意义(P<0.05).结论 缺血缺氧损伤导致的肌动蛋白及其结合蛋白的改变与IEC脱落、凋亡相关;钙通道阻断剂可以减轻这种改变.

The change of F-actin and actin binding protein in intstinal epithelial cells under the ischemia and anoxia injury

Objective To investigate the relationship between the apoptosis and the change of F-actin and actin binding protein in intstinal epithelial cells (IEC) under the ischemia and anoxia injury, and the effect of calcium channel antagonist. Methods Primary culture of rat IECs and ischemia and an-oxia injury model in vitro were established. IECs were divided into 8 groups : group A ( control group), group B (ischemia group) ,group C (anoxia group) ,group D (ischemia and anoxia group) ,group A1 ( A +2 mg/L Verapamil) ,group B1 (B +2 mg/L Verapamil) ,group C1 (C +2 mg/L Verapamil) ,group D1 ( D + 2 mg/L Verapamil). Apoptosis rate (AR) was determined by flow cytometry. F-actin, talin and α-ac-tin were observed by LSCM. Results The AR in groups B, C and D was significantly higher than in group A (P < 0.05). The AR in groups B1, C1 and D1 was lower than in groups B. C and D ( P < 0.05 ). The form and arragement of F-actin, talin and α-actin in groups B, C and D were changed with the decrease of fluorescence intensity compared with group A (P < 0.01 ), the most significant in group D. The change in groups B1, C1 and D1 was reduced and the fluorescence intensity was increased as compared with the cor-responding injury groups (P < 0.05). Conclusion The ischemia and anoxia injury results in the break-down of F-actin and actin binding protein, which is related to the apotosis of IEC. Calcium channel antago-nist can decrease AR and reduce the change of these cytoskeleton.