摘要
目的:研究14-3-3蛋白在心肌细胞缺氧预处理中的表达及意义。方法:实验用SD新生大鼠(1～3d龄),雌雄不拘,无菌取出乳鼠心脏,经分离附着组织,胰蛋白酶消化,纯化制成心肌细胞。在培养的第4天随机分为3组:正常对照组、缺氧/复氧(A/R)组、缺氧预处理(APC)组。各组分别进行以下指标观察:①心肌细胞搏动频率;②细胞存活率(MTT法);③培养液中乳酸脱氢酶(LDH)活性;④透射电镜观察细胞超微结构;⑤Western Blotting法检测14-3-3蛋白的表达变化。RT-PCR法检测14-3-3蛋白η、σmRNA的表达变化。结果:A/R组和APC组的14-3-3蛋白表达均上调,分别是正常对照组的(3.61±0.37)倍和(5.52±0.49)倍,A/R组和APC组与正常对照组比较、A/R组与APC组比较,均P<0.01。A/R组、APC组心肌细胞14-3-3η亚型mRNA分别为正常对照组的(1.82±0.30)倍、(2.93±0.52)倍,差异均有统计学意义(P<0.01);APC组与A/R组比较,差异亦有统计学意义(P<0.01)。A/R组、APC组心肌细胞14-3-3σ亚型mRNA表达量分别为正常对照组的(1.13±0.18)倍、(1.21±0.14倍),A/R组、APC组与正常对照组比较,A/R组与APC组比较,差异均无统计学意义(P>0.05)。结论:14-3-3η亚型mRNA可能参与心肌细胞APC后的早期保护作用。
Objective：To investigate the expression of 14-3-3 protein in the acute myocardial cells A/R injury.Method：Cultured myocardial cells of neonatal SD rats were randomly divided into three groups：control group; anoxia reoxygenation （A/R） group;anoxia preconditioning group （APC）.In each group,the myocardial cells were subjected to anoxia （3 h） followed by reoxygenation （1 h） 24 h after anoxic preconditioning on the model of the cultured myocardial cells. At the end of experiment,myocardial cells beating rate,cell viability and the activity of LDH in culture were measured. Furthermore,the morphology or ultrastructure assessment of myocardial cells was observed by inverted microscope or transmission electron microscope,the expression of 14-3-3 protein was measured by western blotting techniques,and the expression of 14-3-3η,σmRNA was measured by RT-PCR. Result：①It is verified that cardiomyocytes after APC are offered more capacity to tolerate A/R damage from the level of cells.②14-3-3 protein and 14-3-3η,σmRNA showed a basic expression in myocardial cells under normal condition,14-3-3 protein and 14-3-3ηmRNA was enlarged after anoxia-reoxygenation injury.Conclusion：It is suggested that 14-3-3ηmRNA may be involved in the process of protection induced by anoxia preconditioning.
出处
《临床心血管病杂志》
CAS
CSCD
北大核心
2010年第2期144-148,共5页
Journal of Clinical Cardiology
基金
国家重点基础研究发展计划资助课题(No:2009CB526405)
国家自然科学基金资助课题(No:30460048)
