CAJ | 학술논문

目的探讨问号钩端螺旋体(简称钩体)侵入人或鼠单核-巨噬细胞方式及其吞噬泡形成差异性.方法采用透射电镜观察问号钩体黄疸出血群赖型赖株侵入小鼠单核-巨噬样细胞J774A.1和佛波酯(PMA)激活的人单核细胞THP-1后吞噬泡形成情况.采用免疫荧光联合激光共聚焦显微镜及荧光分光光度仪等方法,观察细胞内吞抑制剂单丹磺酰尸胺(MDC)、氧化酚砷(PAO)阻断及内吞相关网格蛋白抗体封闭前后,J774A.1细胞和PMA激活的THP-1细胞内问号钩体赖株数量的变化.结果 J774A.1细胞内问号钩体存在于吞噬泡内,THP-1细胞内问号钩体无吞噬泡膜包绕.MDC和PAO能以剂量依赖方式抑制J774A.1和THP-1细胞内吞问号钩体,其中10 μmol/L以上MDC和1 μmol/L以上PAO阻断的J774A.1和THP-1细胞内问号钩体数量明显少于未阻断细胞(P＜0.05).网格蛋白抗体封闭后,J774A.1和THP-1细胞内问号钩体数量也明显减少(P＜0.05).结论问号钩体以网格蛋白依赖性内吞途径侵入人或鼠单核-巨噬细胞.人或鼠单核-巨噬细胞内问号钩体吞噬泡形成有明显差异,这可能是人或鼠感染问号钩体后发病情况不同的原因之一.
목적 탐토문호구단라선체(간칭구체)침입인혹서단핵-거서세포방식급기탄서포형성차이성.방법 채용투사전경관찰문호구체황달출혈군뢰형뢰주침입소서단핵-거서양세포J774A.1화불파지(PMA)격활적인단핵세포THP-1후탄서포형성정황.채용면역형광연합격광공취초현미경급형광분광광도의등방법,관찰세포내탄억제제단단광선시알(MDC)、양화분신(PAO)조단급내탄상관망격단백항체봉폐전후,J774A.1세포화PMA격활적THP-1세포내문호구체뢰주수량적변화.결과 J774A.1세포내문호구체존재우탄서포내,THP-1세포내문호구체무탄서포막포요.MDC화PAO능이제량의뢰방식억제J774A.1화THP-1세포내탄문호구체,기중10 μmol/L이상MDC화1 μmol/L이상PAO조단적J774A.1화THP-1세포내문호구체수량명현소우미조단세포(P＜0.05).망격단백항체봉폐후,J774A.1화THP-1세포내문호구체수량야명현감소(P＜0.05).결론 문호구체이망격단백의뢰성내탄도경침입인혹서단핵-거서세포.인혹서단핵-거서세포내문호구체탄서포형성유명현차이,저가능시인혹서감염문호구체후발병정황불동적원인지일.
Objective To determine the modality of Leptospira interrogans invading human and murine mononuclear-macrophages and diversity of leptospiral phagocytotic vesicle formation. Methods Transmission electron microscopy was applied to observe the invasion of L. interrogans serogroup Icterohaemorrhagiae serovar Lai strain Lai into murine mononuclear-macrophage-like cell line J774A. 1 and PMA-activated human monocyte line THP-1 and the formation of leptospiral phagocytotic vesicles. By using immunofluorescence plus either laser confocal microscopy or fluorescence spectrophotometry, the changes of intracellular leptospiral numbers in J774A. 1 and PMA-activated THP-1 cells before and after block with endocytosis inhibitors monodansylcadaverin (MDC), phenylarsine oxide (PAO) and clathrin antibody were investigated. Results The leptospires in J774A. 1 cells were located in phagocytotic vesicles while the leptospires in THP-1 cells had no package with phagocytotic vesicle membrane. Both MDC and PAO presented the effect inhibiting endocytosis of L. interrogans into J774A. 1 and THP-1 cells in dose-dependent manner. The numbers of leptospires in J774A. 1 and THP-1 cells that pre-blocked with 10 μmol/L or above MDC and 1 μmol/L or above PAO were significantly less than that in the two cells untreated with MDC and PAO (P＜0.05＝. After J774A. 1 and THP-1 cells were blocked with clathrin antibody, the numbers of intracellular leptospires were also remarkbly decreased ( P<0.05 ).Conclusion Leptospira interrogans can invade into both human and murine mononuclear-macrophages through the way of clathrin-dependent endocytosis. There is an opposite diversity of leptospiral phagocytotic vesicle formations in human and murine mononuclear-macrophages, which may result in the difference of pathogenesis in human and mice after infected with L. interrogans.