论文部分内容阅读
本文旨在探讨细胞膜微结构域中胆固醇在小鼠巨噬细胞摄取土拉弗朗西斯菌LVS株中的作用。用电穿孔法将穿梭质粒pFNLTP6gro-gfp导入土拉弗朗西斯菌LVS株;细胞胆固醇用菲律平Ⅲ染色,结合单克隆抗体的小窝蛋白-1用键合了Alexa594的羊抗鼠二抗显色;用Z轴电动聚焦的荧光显微镜分析土拉弗朗西斯菌LVS株感染;用甲基-β-环糊精干扰富含脂质的胞膜,损耗胆固醇,以评估膜微结构域的损伤对土拉弗朗西斯菌入侵的影响,菲律平Ⅲ用于检测胆固醇损耗的效果。结果显示,细胞胆固醇在早期与摄取的土拉弗朗西斯菌疫苗株共定位,膜微结构域相关成分小窝蛋白-1与两者接触密切;土拉弗朗西斯菌入侵巨噬细胞需要胆固醇丰富的膜结构域。结果提示,胆固醇丰富的膜微结构域和小窝蛋白-1在土拉弗朗西斯菌早期进入巨噬细胞过程中发挥重要作用。
This article aims to investigate the role of cholesterol in the cell membrane micro-domain in the ingestion of the macrophages of F. tularensis LVS by mouse macrophages. The shuttle plasmid pFNLTP6gro-gfp was introduced by electroporation into the F. tularensis strain LVS. Cell cholesterol was stained with phenylemphine III, and the caveolin-1 conjugated to the monoclonal antibody was visualized with a goat anti-mouse secondary antibody conjugated to Alexa594 ; Analysis of the infection of L. tularensis strain LVS with a z-axis electrofocusing fluorescent microscope; interference of the lipid-rich plasma membrane with methyl-beta-cyclodextrin to loss of cholesterol to assess the effects of membrane microdomain damage on Tura The impact of Francisella invasion, Philippine Ping Ⅲ for the detection of cholesterol depletion effect. The results showed that the cellular cholesterol co-localized with the inoculum of Toraxella fastidiosa vaccine strain in the early stage and the caveolin-1, a component of the membrane micro-domain, was in close contact with both of them; the invasion of Taraxaci inoculated with macrophages required a cholesterol-rich membrane structure area. The results suggest that the cholesterol-rich membrane micro-domain and caveolin-1 play an important role in the early entry of F. tularensis into macrophages.