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内毒素诱导的肝细胞及枯否细胞凋亡与肝脏损伤的关系

更新时间:2012-10-14 13:58:39  
袁建成 周立新 秦孝健 肖光夏 黎鳌



  【摘要】 目的 探讨内毒素、肿瘤坏死因子(TNF)在诱导枯否细胞及肝细胞凋亡过程中的作用及其与肝脏损伤的关系。方法 采用细胞死亡ELISA法分别检测枯否细胞,以及和肝脏细胞联合培养时,对不同浓度的内毒素,在不同时相点两种细胞的凋零密度,同时观察培养上清中丙氨酸转氨酶(ALT),乳酸脱氢酶(LDH)含量变化。结果 枯否细胞的凋亡数量随着培养时间的延长(3~24小时)和内毒素浓度的增加(0~10 μg/ml)明显增加。而枯否细胞与肝细胞联合培养时,在内毒素浓度大于1 μg/ml时,细胞凋亡数显著上升。TNF单克隆抗体可阻断内毒素诱导的枯否细胞及肝细胞凋亡。LDH及ALT含量在内毒素浓度高于1 μg/ml,且培养6小时后,有明显升高,晚于凋亡出现。结论 TNF介导了内毒素所致的枯否细胞及肝细胞的凋零。而且细胞凋亡早于细胞损伤的出现。大量凋零细胞的出现,使枯否细胞对内毒素的灭活作用降低进而可能导致肝细胞的凋零及损伤。
  【关键词】 脱噬作用  枯否细胞  肝细胞  肿瘤  坏死因子

The relationship between LPS-induced apoptosis in hepatocyte, Kupffer cell and hepatocytic damage Yuan Jiancheng, Zhou Lixin, Qin Xiaojian, et al. Institute of Burn Research, South Western Hospital, Third Military Medical University, Chongqing, 400038
  【Abstract】 Objective To study the role of tumor necrosis factor (TNFα) in the mechanisms of liver injury and the relationship between LPS-induced apoptosis in hepatocyte (HC), kupffer cell (KC), and hepatic damage. Methods In vitro, kupffer cells (1×10/ml) were stimulated with different concentrations of endotoxin (1~10 μg/ml) for 1-24hr; the kupffer cells, which had been cultured with endotoxin for 24hr, were isolated to coculture with hepatocytes (1∶1) for 1~24hr. Two kinds of cells of apoptosis and the block effect of TNFα antibody were detected by cell death ELISA, and the supernatant was performed to determine the levels of alanine aminotransferase (ALT), lactate dehydrogenase (LDH) at different time points. Results Following stimulation with LPS, the number of positive apoptotic KC increased in a concentration and time (3h~24h) dependent manner. In contrast, coincubation of HC with LPS-stimulated KC resulted in a marked increase in positive apoptotic hepatocyte when LPS concentration was greater than 1 μg/ml. TNFα antibody blocked apoptosis in both KC and HC. Only when the LPS concentration was higher than 1 μg/ml and cultured for 6hr, the release

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