Eipel C, Bordel R, Nickels R M, Menger M D, Vollmar B
Dept. of Experimental Surgery, Univ. of Rostock, D-18055 Rostock, Germany.
Am J Physiol Gastrointest Liver Physiol. 2004 May;286(5):G769-76. doi: 10.1152/ajpgi.00275.2003. Epub 2004 Jan 8.
Apoptotic hepatocytes have been demonstrated to represent an important signal for transmigration of leukocytes sequestered in sinusoids during endotoxemia in vivo. Beside leukocytes, platelets and their adhesion to endothelial cells and leukocytes have been implicated in inflammatory liver injury. Using in vivo multifluorescence microscopy, we examined the possibility that hepatocellular apoptosis causes both leukocytes and platelets to colocalize within the sinusoidal microvasculature of endotoxemic livers. We further addressed the issue whether cellular colocalization with apoptotic hepatocytes is cause or consequence of apoptosis. Intraperitoneal exposure of rats with LPS (5 mg/kg) induced liver injury after 6 and 16 h, as given by nutritive perfusion failure (20 +/- 2 and 21 +/- 2%), intrahepatic leukocyte (60 +/- 10 and 121 +/- 48 cells/mm(2)), and platelet (12 +/- 4 and 34 +/- 4 cells/mm(2)) accumulation as well as parenchymal cell apoptosis (4 +/- 1 and 11 +/- 2 cells/mm(2)) and caspase cleavage (4.7 +/- 2.4- and 7.0 +/- 3.0-fold increase; P < 0.05 vs. saline-exposed controls). Higher doses of LPS (10 mg/kg ip) further increased intrahepatic leukocyte and platelet accumulation but not the extent of parenchymal apoptosis. Detailed spatial analysis revealed colocalization of leukocytes (range 12-24%) but barely of platelets (<6%) with apoptotic hepatocytes in all endotoxemic groups studied. It is of interest, however, that platelets were found at increasing rates in colocalization with leukocytes at 6 and 16 h after LPS exposure (5 mg/kg LPS: 7 +/- 3 and 25 +/- 6%; 10 mg/kg LPS: 11 +/- 4 and 14 +/- 1%). Platelet-leukocyte events significantly correlated with the extent of caspase cleavage as an indicator of tissue apoptosis (P < 0.05; r = 0.82). Blockade of apoptosis by a pan-caspase inhibitor caused a significant reduction of leukocyte adherence and platelet-leukocyte colocalization on LPS exposure. On the other hand, leukocytopenic animals revealed reduced hepatocyte apoptosis, although values still exceeded those of controls, and in leuko- and thrombocytopenic animals, hepatocyte apoptosis was found reduced to control values. Taken together, LPS-associated hepatocyte apoptosis seems to be initiated by circulating blood cells that become adherent within the liver but might also contribute to further sustain the inflammatory cell-cell response.
凋亡的肝细胞已被证明是体内内毒素血症期间滞留于肝血窦内的白细胞迁移的重要信号。除白细胞外，血小板及其与内皮细胞和白细胞的黏附也与炎症性肝损伤有关。利用体内多荧光显微镜，我们研究了肝细胞凋亡是否会导致白细胞和血小板在内毒素血症肝脏的肝血窦微血管内共定位。我们还进一步探讨了与凋亡肝细胞的细胞共定位是凋亡的原因还是结果这一问题。腹腔注射脂多糖（LPS，5mg/kg）6小时和16小时后可诱导大鼠肝损伤，表现为营养性灌注衰竭（20±2%和21±2%）、肝内白细胞（60±10和121±48个细胞/mm²）和血小板（12±4和34±4个细胞/mm²）积聚，以及实质细胞凋亡（4±1和11±2个细胞/mm²）和半胱天冬酶裂解（增加4.7±2.4倍和7.0±3.0倍；与生理盐水处理的对照组相比，P<0.05）。更高剂量的LPS（10mg/kg腹腔注射）进一步增加了肝内白细胞和血小板的积聚，但实质细胞凋亡程度未增加。详细的空间分析显示，在所有研究的内毒素血症组中，白细胞与凋亡肝细胞共定位（范围为12 - 24%），而血小板与凋亡肝细胞共定位的比例极低（<6%）。然而，有趣的是，在LPS暴露后6小时和16小时，发现血小板与白细胞共定位的比例增加（5mg/kg LPS：7±3%和25±6%；10mg/kg LPS：11±4%和14±1%）。血小板 - 白细胞事件与作为组织凋亡指标的半胱天冬酶裂解程度显著相关（P<0.05；r = 0.82）。泛半胱天冬酶抑制剂阻断凋亡可显著降低LPS暴露时白细胞的黏附和血小板 - 白细胞的共定位。另一方面，白细胞减少的动物肝细胞凋亡减少，尽管其值仍超过对照组，而白细胞和血小板均减少的动物，肝细胞凋亡减少至对照值。综上所述，LPS相关的肝细胞凋亡似乎由循环血细胞引发，这些血细胞在肝脏内黏附，但也可能有助于进一步维持炎症性细胞间反应。
