感染细胞内问号钩端螺旋体吞噬泡形成及其与溶酶体的融合

Formation of the Leptospira-contained phagosome and fusion of the phagosome and lysosome in Leptospira interrogans infected cells

目的了解问号钩端螺旋体(简称钩体)感染宿主细胞后吞噬泡形成、吞噬泡与溶酶体融合及感染细胞超微结构改变。方法用问号钩体黄疸出血群赖型56601株感染小鼠单核巨噬样细胞J774A.1和猴肾成纤维细胞Cos-7。采用透射电镜观察J774A.1细胞和Cos-7细胞胞内钩体吞噬泡的形成及感染细胞超微结构的改变。采用免疫双荧光染色法和激光共聚焦显微镜,观察含钩体吞噬泡与溶酶体融合情况。结果问号钩体56601株感染J774A.1细胞30 min、感染Cos-7细胞15 min即可在胞浆中发现膜包绕的含钩体吞噬泡,吞噬泡内钩体均保持原有生理弯曲。56601株问号钩体感染Cos-7细胞2h后,钩体吞噬泡膜开始消失,但未发现J774A.1细胞内钩体吞噬泡膜消失的现象。J774A.1细胞内钩体吞噬泡与溶酶体发生共区域化,表明吞噬泡与溶酶体发生融合。J774A.1细胞感染钩体后出现染色质浓缩形成的凋亡小体样结构、细胞空泡变性、线粒体肿胀等超微结构病变,但Cos-7细胞感染钩体后其超微结构基本正常。结论问号钩体感染J774A.1细胞和Cos-7细胞后可迅速形成吞噬泡并可能与钩体Ⅲ型分泌系统产物有关。J774A.1细胞内钩体吞噬泡可与溶酶体发生融合。不同细胞的钩体吞噬泡膜消失及超微结构改变有明显差异。

Objective To explore the formation of cellular phagosome, fusion of phagosome and lysosome, and uhrastructure changes in Leptospira-infected cells. Methyls Mouse mononuclear-macrophage like cell J774A. 1 and monkey kidney fibroblast Cos-7 were infected with Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai strain 56601. The formation of cellular phagosome and uhrastructure changes in the Leptospirainfection J774A. 1 cells and Cos-7 cells were observed in EM. By using double immunofluorescence method and laser scanning confocal microscopy, the fusion of cellular Leptospira-contained phagosome with lysosome was observed. Results The cytoplasmic Leptospira-contained phagosomes were enclosed by membrane in J774A. 1 cells at the 30 min and in Cos-7 cells at the 15 min after infection of L. interrogans strain 56601. Leptospira in the phagosomes maintained their original physiological curves. The membrane of Leptospira-contained phagosomes in Cos- 7 cells in 2 h after infection of L. interrogans strain 56601 became disappeared whereas no similar phenomenon in Leptospira-infected J774A. 1 cells could be found. The co-localiztion of Leptospira-contained phagosomes and lysosomes in J774A. 1 cells occurred, indicating the fusion of phagosome and lysosome. The Leptospira-infected J774A. 1 cells showed ultrastructural pathological changes such as chromatin condensation to form nucleic apoptotic body-hke structure, vacuolar degeneration and swelling mitochondrion, whereas no ultrastructural pathological change in the Leptospira-infected Cos-7 cells was presented. Conclusion The cellular phagosomes can quickly form in J774A. 1 and Cos-7 cells infected with Leptospira, which may be related to the expression products of leptospiral typeⅢ secretary system. The Leptospira-contained phagosomes in J774A. 1 cell can fuse with lysosome. There are remarkably different disappearance of the Leptospira-contained phagosome membrane and the ultrastructural changes in different host cells.