【摘要】 目的确定α-毒素诱导脐静脉内皮细胞(HUV-EC-C)的凋亡在金葡菌L型垂直感染中的作用。方法在培养的HUV-EC-C细胞中加入不同浓度(0、10、30、90、270 ng/ml)的金葡菌α-毒素,处理不同时间(0、2、4、6、8 h)后经Annexin V-PI染色,流式细胞仪检测HUV-ECC细胞的凋亡率。通过ELISA检测α肿瘤坏死因子(TNF-α)、caspase-3与caspase-8的表达量,并观察加入TNF-α中和抗体、caspases-3抑制剂zDEVD-FMK、caspase-8抑制剂zIETD-fmk对α-毒素诱导HUV-EC-C细胞凋亡的影响。结果α-毒素能够诱导HUV-EC-C细胞凋亡并表现为时间、剂量依赖性,明显增加HUV-EC-C细胞中TNF-α、caspase-3与caspase-8的表达;在加入TNF-α中和抗体、zDEVD-FMK、zIETD-fmk后可部分阻断α-毒素能够诱导的HUV-EC-C细胞凋亡。结论α-毒素通过TNF-α及caspase-8介导的外源性死亡途径诱导HUV-EC-C细胞凋亡,表明α-毒素诱导内皮细胞凋亡是金葡菌L型垂直感染影响胎儿发育的主要机制之一。更多还原

【Abstract】 Objective To investigateα-toxin-induced apoptosis of umbilical vein endothelial cells and explore its role in vertical infection of Staphylococcus aureus L-form.Methods HUV-EC-C cells exposed to different concentrations(0,10,30,90,and 270 ng/ml) ofα-toxin for different time lengths(0,2,4,6,and 8 h) were examined for apoptosis using flow cytometry with Annexin V-PI staining.The levels of tumor necrosis factor-α(TNF-α) and the activities of,caspase-3 and caspase-8 in the cell culture were detected by ELISA and colorimetric method,respectively.α-Toxin-induced cell apoptosis was also analyzed in HUV-EC-C cells treated with a neutralizing antibody of TNF-αor with the inhibitory peptides of caspase-3(zDEVD-FMK) and caspase-8(zIETD-fmk).Resultsα-Toxin induced apoptosis of HUV-EC-C cells in a dose- and time-dependent manner and caused significantly enhanced expression of TNF-αand the activation of both caspase-3 and caspase-8.Inhibition of TNF-αwith its neutralizing antibody and the inhibitory peptides of caspase-3 or -8 all significantly decreasedα-toxin-induced cell apoptosis,and the caspase-3 inhibitor completely blockedα-toxin-induced cell apoptosis.Conclusionα-Toxin-induced apoptosis is partially mediated by the extrinsic cell death pathway of TNF-αand caspase-8 and plays an important role in the vertical infection of S.aureus L-form to affect fetal growth and development.更多还原