Haimerl Florian, Erhardt Annette, Sass Gabriele, Tiegs Gisa
Institute of Experimental and Clinical Pharmacology and Toxicology, University of Erlangen-Nuremberg, Erlangen D-91054 and the Division of Experimental Immunology and Hepatology, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, Hamburg D-20246, Germany.
Institute of Experimental and Clinical Pharmacology and Toxicology, University of Erlangen-Nuremberg, Erlangen D-91054 and the Division of Experimental Immunology and Hepatology, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, Hamburg D-20246, Germany; Institute of Experimental and Clinical Pharmacology and Toxicology, University of Erlangen-Nuremberg, Erlangen D-91054 and the Division of Experimental Immunology and Hepatology, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, Hamburg D-20246, Germany.
J Biol Chem. 2009 Jan 2;284(1):495-504. doi: 10.1074/jbc.M803533200. Epub 2008 Nov 10.
Tumor necrosis factor-alpha (TNFalpha) stimulation of hepatocytes induces either cell survival or apoptosis, which seems to be regulated by the ubiquitin-proteasome system. Here we investigated the role of TNFalpha-induced down-modulation of the de-ubiquitinating enzyme USP2 for hepatocyte survival. Inhibition of hepatocyte apoptosis by pre-treatment with TNFalpha (TNFalpha tolerance) was analyzed in the mouse model of galactosamine/TNFalpha-induced liver injury and in actinomycin D/TNFalpha-treated primary mouse hepatocytes. The role of USP2 for TNFalpha-induced hepatocyte survival was studied using small interference RNA or an expression clone. Injection of mice or preincubation of hepatocytes with TNFalpha caused a rapid down-regulation of hepatic USP2-41kD, the predominant USP2 isoform in the liver. In vitro an artificial knockdown of USP2 inhibited actinomycin D/TNFalpha-induced hepatocyte apoptosis, which was associated with elevated levels of the anti-apoptotic protein c-Flip(L/S) and a concomitant decrease of cellular levels of the ubiquitinligase Itch, a negative regulator of c-Flip. USP2-41kD overexpression abrogated TNFalpha tolerance in vitro, prevented accumulation of c-Flip(L/S) and resulted in elevated levels of Itch. Accordingly, c-Flip(L/S) protein levels were elevated in livers of TNFalpha-tolerant mice, which correlated to a switch from JNK and ERK to p38 signaling after galactosamine/TNF re-challenge. Our results indicate that TNFalpha-induced USP2 down-regulation is an effective cytoprotective mechanism in hepatocytes. Hence, USP2 could be a novel pharmacological target, and specific USP2 inhibitors might be potential candidates for the treatment of inflammation-related apoptotic liver damage.
肿瘤坏死因子-α（TNFα）刺激肝细胞可诱导细胞存活或凋亡，这似乎受泛素-蛋白酶体系统调控。在此，我们研究了TNFα诱导的去泛素化酶USP2下调对肝细胞存活的作用。在半乳糖胺/TNFα诱导的肝损伤小鼠模型以及放线菌素D/TNFα处理的原代小鼠肝细胞中，分析了用TNFα预处理（TNFα耐受）对肝细胞凋亡的抑制作用。使用小干扰RNA或表达克隆研究了USP2对TNFα诱导的肝细胞存活的作用。给小鼠注射TNFα或使肝细胞与TNFα预孵育会导致肝脏中USP2-41kD迅速下调，USP2-41kD是肝脏中主要的USP2亚型。在体外，人工敲低USP2可抑制放线菌素D/TNFα诱导的肝细胞凋亡，这与抗凋亡蛋白c-Flip（L/S）水平升高以及泛素连接酶Itch（c-Flip的负调节因子）细胞水平的相应降低有关。USP2-41kD过表达在体外消除了TNFα耐受，阻止了c-Flip（L/S）的积累，并导致Itch水平升高。因此，在TNFα耐受小鼠的肝脏中，c-Flip（L/S）蛋白水平升高，这与半乳糖胺/TNF再次攻击后从JNK和ERK信号转导切换到p38信号转导相关。我们的结果表明，TNFα诱导的USP2下调是肝细胞中一种有效的细胞保护机制。因此，USP2可能是一个新的药理学靶点，特异性USP2抑制剂可能是治疗炎症相关凋亡性肝损伤的潜在候选药物。
