Jaeschke H, Fisher M A, Lawson J A, Simmons C A, Farhood A, Jones D A
Department of Pharmacology, Pharmacia & Upjohn, Inc., Kalamazoo, MI 49007, USA.
J Immunol. 1998 Apr 1;160(7):3480-6.
Endotoxin (ET)-induced liver failure is characterized by parenchymal cell apoptosis and inflammation leading to liver cell necrosis. Members of the caspase family have been implicated in the signal transduction pathway of apoptosis. The aim of this study was to characterize ET-induced hepatic caspase activation and apoptosis and to investigate their effect on neutrophil-mediated liver injury. Treatment of C3Heb/FeJ mice with 700 mg/kg galactosamine (Gal) and 100 microg/kg Salmonella abortus equi ET increased caspase 3-like protease activity (Asp-Val-Glu-Asp-substrate) by 1730 +/- 140% at 6 h. There was a parallel enhancement of apoptosis (assessed by DNA fragmentation ELISA and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay). In contrast, activity of caspase 1 (IL-1beta-converting enzyme)-like proteases (Tyr-Val-Ala-Asp-substrate) did not change throughout the experiment. Caspase 3-like protease activity and apoptosis was not induced by Gal/ET in ET-resistant mice (C3H/HeJ). Furthermore, only murine TNF-alpha but not IL-1alphabeta increased caspase activity and apoptosis. Gal/ET caused neutrophil-dependent hepatocellular necrosis at 7 h (area of necrosis, 45 +/- 3%). Delayed treatment with the caspase 3-like protease inhibitor Z-Val-Ala-Asp-CH2F (Z-VAD) (10 mg/kg at 3 h) attenuated apoptosis by 81 to 88% and prevented liver cell necrosis (< or = 5%). Z-VAD had no effect on the initial inflammatory response, including the sequestration of neutrophils in sinusoids. However, Z-VAD prevented neutrophil transmigration and necrosis. Our data indicate that activation of the caspase 3 subfamily of cysteine proteases is critical for the development of parenchymal cell apoptosis. In addition, excessive hepatocellular apoptosis can be an important signal for transmigration of primed neutrophils sequestered in sinusoids.
内毒素（ET）诱导的肝衰竭的特征是实质细胞凋亡和炎症，进而导致肝细胞坏死。半胱天冬酶家族成员参与了凋亡信号转导途径。本研究的目的是明确ET诱导的肝脏半胱天冬酶激活和凋亡的特征，并研究它们对中性粒细胞介导的肝损伤的影响。用700mg/kg氨基半乳糖（Gal）和100μg/kg马流产沙门氏菌ET处理C3Heb/FeJ小鼠，6小时时半胱天冬酶3样蛋白酶活性（Asp-Val-Glu-Asp底物）增加了1730±140%。凋亡也有平行增加（通过DNA片段化ELISA和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法评估）。相比之下，在整个实验过程中，半胱天冬酶1（IL-1β转化酶）样蛋白酶（Tyr-Val-Ala-Asp底物）的活性没有变化。在ET抗性小鼠（C3H/HeJ）中，Gal/ET未诱导半胱天冬酶3样蛋白酶活性和凋亡。此外，只有小鼠TNF-α而非IL-1αβ增加了半胱天冬酶活性和凋亡。Gal/ET在7小时时导致中性粒细胞依赖性肝细胞坏死（坏死面积为45±3%）。用半胱天冬酶3样蛋白酶抑制剂Z-Val-Ala-Asp-CH2F（Z-VAD，3小时时10mg/kg）延迟处理可使凋亡减少81%至88%，并预防肝细胞坏死（≤5%）。Z-VAD对初始炎症反应没有影响，包括中性粒细胞在肝血窦中的滞留。然而，Z-VAD可预防中性粒细胞迁移和坏死。我们的数据表明，半胱氨酸蛋白酶半胱天冬酶3亚家族的激活对实质细胞凋亡的发生至关重要。此外，过度的肝细胞凋亡可能是肝血窦中预激活的中性粒细胞迁移的重要信号。
