梓醇对模拟缺血再灌注损伤兔窦房结细胞凋亡及细胞骨架β-微管蛋白的影响

Effects of Catalpol on Apoptosis and β-Tubulin of Rabbit Sinoatrial Node Cells in Vitro Injured by Simulated Ischemia Reperfusion

目的观察地黄主要有效成分梓醇对模拟缺血再灌注损伤兔窦房结细胞凋亡及细胞骨架β-微管蛋白(β-tubulin)的影响,探讨其治疗病态窦房结综合征的机制。方法取新生乳兔窦房结细胞,分为正常组、模型组及梓醇高、中、低剂量组。以缺氧缺糖模拟缺血、恢复氧和糖的供应模拟再灌注造成窦房结细胞损伤模型。正常组与模型组给予等体积培养基,梓醇高、中、低剂量组给予终浓度为100、20、10μg/m L药物,应用酶标仪、流式细胞仪、激光共聚焦显微镜观察各组窦房结细胞凋亡率、β-tubulin的形态变化。结果模型组细胞凋亡率较正常组明显增加(P<0.01),β-tubulin裂解明显。与模型组比较,梓醇高、中、低剂量组细胞凋亡率明显降低(P<0.01),β-tubulin结构明显完整,荧光强度明显升高(P<0.01)。结论梓醇抑制模拟缺血再灌注引起的窦房结细胞凋亡,保护窦房结β-tubulin形态可能是地黄治疗病态窦房结综合征的机制之一。

Objective To observe the effects of catalpol, the effective component of RehmanniaeRadix, on atrionector cell apoptosis and β-tubulin in vitro rabbit injured by simulated ischemiareperfusion;To explore the mechanism of treating sick sinus syndrome. Methods Atrionector cellswere collected from newborn rabbits. Cells were divided into 5 groups：normal group, model group,catalpol high, medium and low dose groups. Anoxia and aglycaemia were established to simulateischemia. Atrionector cellular damage models were established by recovering the supply of oxygenand sugar. Normal control group and model group were given the same volume of culture medium,while catalpol high, medium and low dose groups were given medicine with relevantconcentrations （100, 20, 10 μg/mL, respectively）. ELISA, FCM, laser scanning confocal microscopywere used to observe apoptosis rate and β-tubulin of atrionector in each group. Results Apoptosisrate of the model group was obviously higher than the normal group （P 〈0.01）, and β-tubulincleavage was obvious. Apoptosis rate in catalpol high, medium and low dose groups weresignificantly lower than that of the model group （P 〈0.01）;β-tubulin structure were significantlymore complete compared with the model group;the fluorescence intensity was significantly higherthan that of model group （P 〈0.01）. Conclusion Catalpol can inhibit atrionector cellular apoptosiscaused by simulated ischemia reperfusion. Its protective effects on atrionector β-tubulin may bethe mechanism of the treatment of Rehmanniae Radix for sick sinus syndrome.