Yang Jun, Gallagher Scott F, Haines Krista, Epling-Burnette P K, Bai Fenqi, Gower William R, Mastorides Stephen, Norman James G, Murr Michel M
Department of Surgery, James A Haley Veterans Hospital, University of South Florida, Tampa, Florida 33601, USA.
J Gastrointest Surg. 2004 Feb;8(2):166-74. doi: 10.1016/j.gassur.2003.10.016.
Liver injury is an important prognostic indicator during acute pancreatitis. The aim of this study was to determine the role of Fas ligand (FasL) in hepatocyte injury. Liver parenchymal enzymes were measured in cocultures of hepatocytes and Kupffer cells treated with elastase. FasL and FasL mRNA were measured in elastase-treated Kupffer cells. Hepatocytes were treated with FasL and their viability was assessed by monotetrazolium (MTT), apoptosis by flow cytometry, as well as caspase-3 and p38-mitogen-activated protein kinase (MAPK) by immunoblotting. Elastase increased aspartate aminotransferase and lactate dehydrogenase in cocultures of hepatocyte and Kupffer cells (P<0.040). Elastase increased FasL production from Kupffer cells (P=0.02) and upregulated FasL mRNA (FasL/beta-2 microglobulin (BMG): 0.23+/-0.03 vs. 0.11+/-0.003; P=0.04). FasL increased alanine aminotransferase and lactate dehydrogenase (P<0.03) and reduced hepatocyte viability by 45% (P=0.01). FasL increased the number of dually labeled cells with AnnexinV/7AAD (P=0.03) while upregulating cleavage of caspase-3 and the phosphorylation of p38-MAPK. FasL antibody attenuated the FasL-related increase in dually labeled cells (P=0.02), the cleavage of caspase-3, and phosphorylation of p38-MAPK. Pancreatic elastase upregulates FasL within Kupffer cells. FasL induces hepatocyte injury and death and upregulates p38-MAPK and caspase-3 within hepatocytes. The ability to manipulate interactions between Kupffer cells and hepatocytes may have important therapeutic implications.
肝损伤是急性胰腺炎期间一个重要的预后指标。本研究的目的是确定Fas配体（FasL）在肝细胞损伤中的作用。在用弹性蛋白酶处理的肝细胞和库普弗细胞共培养物中测量肝实质酶。在弹性蛋白酶处理的库普弗细胞中测量FasL和FasL mRNA。用FasL处理肝细胞，并通过单四氮唑蓝（MTT）评估其活力，通过流式细胞术评估细胞凋亡，通过免疫印迹评估半胱天冬酶-3和p38丝裂原活化蛋白激酶（MAPK）。弹性蛋白酶增加了肝细胞和库普弗细胞共培养物中的天冬氨酸转氨酶和乳酸脱氢酶（P<0.040）。弹性蛋白酶增加了库普弗细胞中FasL的产生（P=0.02）并上调了FasL mRNA（FasL/β2微球蛋白（BMG）：0.23±0.03对0.11±0.003；P=0.04）。FasL增加了丙氨酸转氨酶和乳酸脱氢酶（P<0.03）并使肝细胞活力降低了45%（P=0.01）。FasL增加了膜联蛋白V/7氨基放线菌素D双标记细胞的数量（P=0.03），同时上调了半胱天冬酶-3的裂解和p38-MAPK的磷酸化。FasL抗体减弱了双标记细胞中与FasL相关的增加（P=0.02）、半胱天冬酶-3的裂解和p38-MAPK的磷酸化。胰腺弹性蛋白酶上调库普弗细胞内的FasL。FasL诱导肝细胞损伤和死亡，并上调肝细胞内的p38-MAPK和半胱天冬酶-3。操纵库普弗细胞与肝细胞之间相互作用的能力可能具有重要的治疗意义。
