问号钩端螺旋体跨血管内皮细胞转运分子机制研究  

Molecular mechanism of transcytosis of Leptospira interrogans across vascular endothelial cells

作　　者：李丹[1] 王运中[2] 计雪强[2] 邵雪君[2] 朱宏[2] 严杰[3] 李阳 Li Dan;Wang Yunzhong;Ji Xueqiang;Shao Xuejun;Zhu Hong;Yan Jie;Li Yang(Department of Clinical Laboratory,the Second Affiliated Hospital of SoochowUniversity,Suzhou 215004,China;Department of Clinical Laboratory,Children's Hospital of Soochow University,Suzhou 215025,China;Department of Medical Microbiology and Parasitology,Zhejiang University School of Medicine,Hangzhou 310058,China)

机构地区：[1]苏州大学附属第二医院检验科,215004 [2]苏州大学附属儿童医院检验科,215025 [3]浙江大学医学院病原生物学系,杭州310058

出　　处：《中华微生物学和免疫学杂志》2020年第5期344-349,共6页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金(81671974)。

摘　　要：目的探讨问号钩端螺旋体(简称钩体)跨血管内皮细胞转运分子机制。方法采用Transwell法了解问号钩体赖株穿越人脐静脉血管内皮细胞(HUVEC)单层能力。采用透射电镜和激光共聚焦显微镜法检测HUVEC内吞问号钩体后形成内吞泡情况。采用细胞内吞抑制试验确定HUVEC内吞问号钩体方式。采用激光共聚焦显微镜法检测HUVEC胞内问号钩体与溶酶体标志分子LAMP1共定位情况。采用暗视野显微镜Petroff-Hausser计数法检测HUVEC胞内问号钩体出胞情况。结果问号钩体赖株能迅速穿越HUVEC细胞单层。问号钩体通过PI3K-微丝依赖的细胞内吞方式进入HUVEC并形成内吞泡。HUVEC胞内问号钩体不与LAMP1共定位,提示问号钩体内吞泡不与溶酶体融合。HUVEC胞内问号钩体通过FAK-微丝/微管途径出胞。结论问号钩体通过跨细胞转运方式穿越人血管内皮细胞导致体内播散并加重病情。Objective To investigate the molecular mechanism of transcytosis of Leptospira interrogans(L.interrogans)across vascular endothelial cells.Methods Transwell assay was performed to observe the ability of L.interrogans strain Lai across the monolayer of human vascular endothelial cells(HUVEC).Transmission electron microscopy and laser confocal microscopy were used to detect the endocytic vesicles containing L.interrogans strain Lai in HUVEC.The leptospiral endocytic pathway was determined by endocytic inhibition test.Laser confocal microscopy was also used detect the co-localization of L.interrogans with lysosomal marker LAMP1 in HUVEC.The exocytosis of L.interrogans from HUVEC was detected using Petroff-Hausser counting chamber and darkfield microscopy.Results L.interrogans strain Lai could rapidly transmigrate through HUVEC monolayers and be internalized into HUVEC by PI3K-microfilament-dependent endocytosis to form leptospiral endocytic vesicles.The internalized L.interrogans did not co-localize with LAMP1,indicating the leptospiral endocytic vesicles did not fuse with lysosomes.The exocytosis of internalized L.interrogans was through FAK-microfilament/microtubule pathway.Conclusions L.interrogans strain Lai could transmigrate through HUVEC by transcytosis to diffuse in vivo and cause disease aggravation.

关 键 词：问号钩端螺旋体 败血症 血管内皮细胞 跨细胞转运 体内播散 

分 类 号：R377[医药卫生—病原生物学]