金黄色葡萄球菌诱导人脐静脉内皮细胞凋亡的观察

Observation on the apoptosis of human umbilical vein endothelial cells induced by Staphylococcus aureus

目的观察人脐静脉内皮细胞株ECV-304对金黄色葡萄球菌(S.aureus)感染的反应。方法常规方法制备金黄色葡萄球菌悬液,以1∶1、1∶10和1∶100的感染复数感染ECV-304细胞1、3、6、12h和24h后,估算细胞吞噬细菌数,采用Hoechst33258染色,DNAladder及细胞凋亡相关因子RT-PCR检测细菌感染后细胞凋亡情况。结果金黄色葡萄球菌感染复数越高,细胞内吞噬的活细菌数越多。金黄色葡萄球菌感染导致ECV-304细胞发生凋亡,凋亡具有时间依赖性,随着感染时间的延长,凋亡小体增多,DNA片段也随之增多。抗凋亡细胞因子Bcl2mRNA表达量随着感染时间的延长下降,而促凋亡细胞因子Bax和Caspase-3表达量增多。结论金黄色葡萄球菌感染ECV-304细胞后,通过下调抗凋亡细胞因子Bcl2、上调促凋亡细胞因子Bax和Caspase-3诱导细胞凋亡。金黄色葡萄球菌感染内皮细胞的可能致病机制为临床治疗提供了实验依据。

Objective To study the response of human umbilical vein endothelial cells （ECV-304） to Staphylococcus aureus infection. Methods S.aureus suspension was collected routinely and used to infect ECV-304 at a multiplicity of infections （MOI） of 1：1,1：10 and 1：100 for 1 to 24 h. The number of living ceils was estimated by the colony forming assay.Apoptotic nuclei were stained with Hoechst33258.Apoptotic DNA fragments were analyzed using gel electrophoresis and the expression of apoptotic factors was analyzed using RT-PCR. Results The number of intracellular bacteria was increased with an increasing in MOI. S.aureus induced apoptosis in EC^-304 in a time-dependent manner. Apoptotic bodies and DNA fragments were time-dependently increased. The level of Bcl2 mRNA Bcl2 was decreased, and the expression level of Bax and Caspase-3 was increased. Conclusion S.aureus was found to induce apoptosis in ECV-304 through the down-regulation of Bcl2 and the up-regulation of Bax and Caspase-3.