论文部分内容阅读
目的 探讨多房棘球蚴感染宿主CD4~+T淋巴细胞缺失的机制,多房棘球蚴感染与宿主T淋巴细胞亚群凋亡相互关系。方法 利用尼龙柱和补体法从多房棘球蚴感染12wk,25wk和正常对照组BALB/c 小鼠脾脏分离出纯CD4~+、CDS8~+T细胞,在体外分别经多房棘球蚴抗原(EmAg)、抗CD3抗原(anti-CD3)、白细胞介素-2(IL-2)、肿瘤坏死因子α(TNFα)及商陆丝裂原(PWM)刺激培养16h,用DNA凝胶电泳分析;透射电镜观察形态学变化;原位末端标记法(Tunel)和碘化丙锭(PI)双染后,流式细胞仪(FCM)分析凋亡细胞数和凋亡发生的细胞周期。结果 感染25wk小鼠,CD4~+T细胞DNA电泳图均出现典型“梯形”条带,透射电镜见染色质浓染、胞质出泡、凋亡小体形成;CD8~+T细胞DNA电泳国无梯形带,透射电镜见染色质浓染,电子密度增强。FCM分析,感染12wk小鼠,CD4~+、CD8~+T细胞凋亡数与正常对照组差异无显著性意义(P>0.05);感染25wk小鼠,CD4~+T细胞凋亡数较正常对照组显著增高(P<0.01),也显著高于同组CD8~+T细胞(P<0.01)。凋亡主要发生在细胞周期S期。结论 多房棘球蚴寄生宿主后期,可诱导宿主成熟CD4~+T细胞发生活化诱导性死亡(AICD),使宿主处于免疫抑制状态。
Objective To investigate the mechanism of the absence of CD4 ~ + T lymphocytes in the host of Echinococcus multilocularis infection and the relationship between the infection of Echinococcosis and the apoptosis of host T lymphocyte subsets. Methods Pure CD4 + and CD8 + T cells were isolated from BALB / c mice spleen of Echinococcus multilocularis infection at 12wk, 25wk and normal control groups by nylon column and complement method, respectively. Echinococcus multilocularis antigen The cells were cultured for 16h with EmAg, anti-CD3, IL-2, TNFα and PWM. DNA gel electrophoresis analysis The morphological changes were observed by transmission electron microscopy. After double staining with Tunel and PI, the number of apoptotic cells and the cell cycle of apoptosis were analyzed by flow cytometry (FCM). Results The DNA ladder of CD4 ~ + T cells infected by 25wk mice showed typical “trapezoidal” bands. Transmission electron microscopy showed chromatin staining, cytoplasmic effusion and apoptotic bodies formation. DNA electrophoresis of CD8 ~ + T cells No trapezoidal band, transmission electron microscopy of chromatin staining, enhanced electron density. FCM analysis showed that the number of apoptosis of CD4 ~ +, CD8 ~ + T cells in 12wk infected mice had no significant difference compared with that of the normal control group (P> 0.05). The number of apoptotic CD4 ~ + T cells in infected 25wk mice was more than normal Control group was significantly higher (P <0.01), also significantly higher than the same group of CD8 ~ T cells (P <0.01). Apoptosis occurs mainly in the cell cycle S phase. Conclusions Echinococcus multilocularis can induce host-induced CD4 ~ + T-cell activation-induced death (AICD) at the late stage of host parasitism, and make the host immunosuppressed.