膜胆固醇对重组创伤弧菌溶细胞素活性的影响  

Membrane Cholesterol Influences the Activity of Recombinant Vibrio Vulnificus Hemolysin

作　　者：丁卉[1] 潘娌妮[1] 郭秋[1] 楼永良[1] 严杰[2] 伍越[1] 王福乐[1] 

机构地区：[1]温州医学院检验医学院浙江省医学遗传学重点实验室,温州325035 [2]浙江大学医学院病原生物学教研室,杭州310031

出　　处：《中国细胞生物学学报》2010年第5期696-701,共6页Chinese Journal of Cell Biology

基　　金：浙江省自然科学资金(No.Y2090468);浙江省科技计划(No.84008012)资助项目~~

摘　　要：研究膜胆固醇对重组创伤弧菌溶细胞素(recombinant Vibrio vulnificus hemolysin,rVvhA)生物学活性的影响,以进一步明确rVvhA的分子致病机制。利用甲基-β-环糊精(MβCD)可以去除细胞膜上胆固醇的能力,rVvhA作用细胞之前,细胞预先经5 mmol/L MβCD处理。测定MβCD预处理组和未处理组(即rVvhA直接作用组)细胞的存活率、细胞内钙离子浓度、培养上清中钾离子浓度及细胞凋亡与坏死情况。结果显示MβCD预处理组的细胞存活率较rVvhA直接作用组高、rVvhA引起的钙离子内流和钾离子释放受到抑制、细胞凋亡率也明显降低。结果表明,在rVvhA活性发挥中细胞膜上的胆固醇是必须存在的,胆固醇的降低会抑制rVvhA的活性。To study the effect of membrane cholesterol on the activity of recombinant Vibrio vulnificus hemolysin（rVvhA）.Before treated with rVvhA,J774A.1 cells were incubated with 5 mmol/L MβCD for 30 min to remove cholesterol from cells.After rVvhA treatment,the cell viability was determined by MTT assay.The intracellular [Ca^（2＋）]was determined using Fluo-3 AM fluorescent probe combined with Laser scanning confocal microscope, the release of cellular K~＋into reaction medium and the cell apoptosis were detected according to detect kit instructions. The results showed cell pretreated with MβCD increased the cell viability,blocked rVvhA induced Ca^（2＋）influx and K~＋ release,and inhibited the rVvhA induced-apoptosis.In conclusion,membrane cholesterol is required for activity of rVvhA,the depletion of cholesterol can inhibited the activity of rVvhA.

关 键 词：创伤弧菌 溶细胞素 甲基-β-环糊精 胆固醇 

分 类 号：R378[医药卫生—病原生物学]