Win Sanda, Than Tin Aung, Le Bao Han Allison, García-Ruiz Carmen, Fernandez-Checa Jose C, Kaplowitz Neil
University of Southern California Research Center for Liver Diseases, Division of Gastrointestinal and Liver Diseases, Keck School of Medicine, University of Southern California, Los Angeles, CA 90089-9121, USA.
Southern California Research Center for ALPD and Cirrhosis, Keck School of Medicine of the University of Southern California, Los Angeles, CA, USA; Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB), Consejo Superior Investigaciones Cientificas (CSIC) and Liver Unit-Hospital Clinic and CIBEREHD, Barcelona, Spain.
J Hepatol. 2015 Jun;62(6):1367-74. doi: 10.1016/j.jhep.2015.01.032. Epub 2015 Feb 7.
BACKGROUND & AIMS: Sustained c-Jun N-terminal kinase (JNK) activation by saturated fatty acids plays a role in lipotoxicity and the pathogenesis of non-alcoholic steatohepatitis (NASH). We have reported that the interaction of JNK with mitochondrial Sab leads to inhibition of respiration, increased reactive oxygen species (ROS), cell death and hepatotoxicity. We tested whether this pathway underlies palmitic acid (PA)-induced lipotoxicity in hepatocytes.
Primary mouse hepatocytes (PMH) from adeno-shlacZ or adeno-shSab treated mice and HuH7 cells were used.
In PMH, PA dose-dependently up to 1mM stimulated oxygen consumption rate (OCR) due to mitochondrial β-oxidation. At ⩾1.5mM, PA gradually reduced OCR, followed by cell death. Inhibition of JNK, caspases or treatment with antioxidant butylated hydroxyanisole (BHA) protected PMH against cell death. Sab knockdown or a membrane permeable Sab blocking peptide prevented PA-induced mitochondrial impairment, but inhibited only the late phase of both JNK activation (beyond 4h) and cell death. In PMH, PA increased p-PERK and its downstream target CHOP, but failed to activate the IRE-1α arm of the UPR. However, Sab silencing did not affect PA-induced PERK activation. Conversely, specific inhibition of PERK prevented JNK activation and cell death, indicating a major role upstream of JNK activation.
The effect of p-JNK on mitochondria plays a key role in PA-mediated lipotoxicity. The interplay of p-JNK with mitochondrial Sab leads to impaired respiration, ROS production, sustained JNK activation, and apoptosis.
饱和脂肪酸持续激活c-Jun氨基末端激酶（JNK）在脂毒性及非酒精性脂肪性肝炎（NASH）发病机制中起作用。我们曾报道JNK与线粒体Sab相互作用会导致呼吸抑制、活性氧（ROS）增加、细胞死亡及肝毒性。我们测试了该通路是否为棕榈酸（PA）诱导的肝细胞脂毒性的基础。
使用来自腺病毒-shlacZ或腺病毒-shSab处理小鼠的原代小鼠肝细胞（PMH）及HuH7细胞。
在PMH中，高达1mM的PA剂量依赖性刺激线粒体β-氧化导致的氧消耗率（OCR）。在⩾1.5mM时，PA逐渐降低OCR，随后细胞死亡。抑制JNK、半胱天冬酶或用抗氧化剂丁基羟基茴香醚（BHA）处理可保护PMH免受细胞死亡。敲低Sab或使用膜通透性Sab阻断肽可预防PA诱导的线粒体损伤，但仅抑制JNK激活（4小时后）和细胞死亡的晚期阶段。在PMH中，PA增加p-PERK及其下游靶点CHOP，但未能激活未折叠蛋白反应（UPR）的IRE-1α臂。然而，沉默Sab并不影响PA诱导的PERK激活。相反，特异性抑制PERK可预防JNK激活和细胞死亡，表明其在JNK激活上游起主要作用。
p-JNK对线粒体的作用在PA介导的脂毒性中起关键作用。p-JNK与线粒体Sab的相互作用导致呼吸受损、ROS产生、JNK持续激活及细胞凋亡。
