缺氧-复氧对培养的大鼠心肌细胞损伤与细胞凋亡的影响.docVIP

???? 缺氧-复氧对培养的大鼠心肌细胞损伤与细胞凋亡的影响 ???? ????【摘要】　目的　探讨培养的大鼠心肌细胞在缺氧(缺血)和缺氧-复氧(再灌注)状态下心肌细胞损伤和细胞凋亡情况。方法　采用1～3天新生SD大鼠，常规方式的培养心肌细胞，给予模拟缺氧(缺血)液、模拟复氧(再灌注)液以及干预因素(终浓度为200 U/mL的SOD)处理，分为缺氧-复氧损伤组(H/R组)、缺氧-复氧损伤组+SOD(H/R+SOD组)、缺氧预处理组(HP组)和正常对照组(Control组)，进行以下指标观察：①台盼蓝排斥法计算细胞存活率；②乳酸脱氢酶(LDH)活性测定；③心肌细胞内丙二醛(MDA)含量；④心肌细胞内游离钙测定；⑤流式细胞仪分析细胞凋亡；⑥透射电镜观察细胞超微结构。结果　H/R组的细胞存活率为(42.5±5.5)%，明显低于H/R+SOD组和HP组的细胞存活率(分别为64.5%±5.0%和66.4%±5.4%，P＜0.05)；而H/R组心肌细胞内的脂质过氧化产物丙二醛(MDA)含量则明显增高(3.53±1.51)nmol/L，与HP组和H/R+SOD组的心肌细胞内MDA含量(1.52±1.03)nmol/L和(1.93±0.69)nmol/L比较有显著差异(P＜0.05)；培养液中H/R组心肌酶活性增高，均提示了缺氧-复氧过程中心肌细胞损伤严重；同时，伴有心肌细胞内游离钙增加，也显著高于HP组和H/R+SOD组(P＜0.01)；而且细胞内钙的增加与细胞凋亡的数量呈正相关(γ=0.92,P＜0.01)。H/R组心肌细胞凋亡率为(18.52±6.15)%，也明显高于HP组和H/R+SOD组(分别为4.62%±2.57%和6.94%±2.71%，P＜0.01)。H/R组细胞超微结构可见多数细胞呈胞浆浓缩、核固缩深染等改变，而其他各组则少见。结论　缺氧-复氧可加重心肌细胞损伤，伴着心肌细胞凋亡的增加；细胞内钙离子过多可能是促发心肌凋亡的因素。SOD和或缺氧预处理具有抗心肌细胞损伤和减轻心肌细胞凋亡的作用，而且可能是通过降低细胞内钙离子浓度而起作用的。　　【关键词】　培养心肌细胞　缺氧-复氧　缺氧预处理　SOD　细胞损伤与凋亡 Effect of hypoxia-reoxygenation on cell injury and apoptosis in cultured neonatal rat myocytes Yao Zhen, Feng Jianzhang, Fu Yonghen, et al.（Guangdong Provincial Cardiovascular Research Institute, Guangzhou 510100） 　　【Abstract】　Objective　To study the effects of hypoxia-reoxygenation on cell injury and apoptosis in cultured neonatal rat myocytes. Methods　Cultured myocytes of neonatal SD rats have been divided into four groups-Hypoxia/Reoxygenation(H/R), Hypoxia/Reoxygenation+SOD(H/R+SOD), Hypoxia Preconditioning(HP) and Control. Myocyte viability, the activity of LDH in culture, the contents of intracellular MDA and free Cal-cium （［Ca2+］i） in myocytes were evaluated in each group. Also, apoptotic rate in each group was assayed by flow cytometer with PI staining method. The ultrastructure of cardiomyocytes was observed by a Philips CM10 transmission electron microscope. Results　Myocyte viability in H/R group was much lower than that in control, H/R+SOD and HP groups(P＜0.01 or 0.05). The cultural contents of LDH, the myocyte contents of MDA and ［Ca2+］i were a