Hepatocyte-specific MET deletion exacerbates acetaminophen-induced hepatotoxicity in mice

Despite the well-known role of MET in liver regeneration following partial-hepatectomy (PHx), its role in the clinically-relevant acetaminophen (APAP)-induced liver injury (AILI) model remains unexplored. AILI markedly differs from PHx because it is associated with massive liver necrosis. This study aims to delineate the role of MET specifically in AILI. Hepatocyte-specific MET-KO mice were given a toxic-dose of APAP and assessed for hepatotoxicity/regeneration parameters. MET deletion strikingly exacerbated initial hepatotoxicity and impaired subsequent proliferative response, culminating in significant mortality. Mechanistically, MET deletion enhanced JNK-activation and its mitochondrial translocation, resulting in excessive mitochondrial oxidative-damage, releasing cell-death inducer AIF into cytosol. Excess JNK-activation was attributed to reduced inhibitory activity of AKT on JNK in the absence of MET-signaling. Pharmacological activation of AKT reduced JNK-activation and hepatotoxicity in MET-KO mice. RNA-sequencing/immunoblotting not only showed repression of proliferative/survival signaling, but also activation of cell death/senescence pathways along with impaired unfolded-protein-response in MET-KO mice. Analysis of published single-nucleus RNA-sequencing data showed proliferation in livers from APAP-induced ALF patients was associated with strong activation of HGF/MET signaling in hepatocytes, with spatial-transcriptomics showing striking induction of HGF surrounding the necrotic-zones. Interestingly, 35% of the genes altered in human-ALF were regulated by MET in the mouse AILI-model. In conclusion, our study demonstrates that MET is crucial for restraining hepatotoxicity following APAP overdose via inhibiting mitochondrial cell-death signaling pathway. Wild-type and hepatocyte-specific MET knockout mice were given 300 mg/kg of Acetaminophen (APAP). Liver tissues were harvested at 0, 6, and 24hr post-APAP overdose.

尽管已知间质上皮转化因子（MET）在部分肝切除术（PHx）后的肝脏再生中发挥关键作用，但其在临床相关的对乙酰氨基酚（APAP）诱导性肝损伤（AILI）模型中的作用仍未被阐明。AILI与PHx存在显著差异，该模型伴随大规模肝脏坏死。本研究旨在明确MET在AILI中的特异性作用。研究人员给予肝细胞特异性MET敲除（MET-KO）小鼠毒性剂量的APAP，并对其肝毒性与再生相关参数进行评估。MET缺失显著加重了初始肝损伤，并损害了后续的增殖反应，最终导致小鼠死亡率显著升高。机制层面分析显示，MET缺失增强了c-Jun氨基末端激酶（JNK）的激活及其线粒体易位，引发过度的线粒体氧化损伤，并将细胞死亡诱导因子凋亡诱导因子（AIF）释放至胞质中。过度的JNK激活归因于MET信号缺失时，蛋白激酶B（AKT）对JNK的抑制活性减弱。在MET-KO小鼠中，通过药理学手段激活AKT可降低JNK的激活水平与肝毒性。RNA测序（RNA-sequencing）与免疫印迹（immunoblotting）分析结果显示，MET-KO小鼠不仅存在增殖/存活信号通路的抑制，还出现了细胞死亡与衰老通路的激活，同时未折叠蛋白反应（UPR）受损。对已发表的单细胞核RNA测序（single-nucleus RNA-sequencing）数据的分析表明，对乙酰氨基酚诱导的急性肝衰竭（ALF）患者肝脏中的肝细胞增殖，与肝细胞内肝细胞生长因子（HGF）/MET信号通路的强激活密切相关；空间转录组学（spatial-transcriptomics）分析则显示，坏死区域周围的HGF表达显著上调。值得注意的是，在人类ALF中发生表达异常的基因中，有35%在小鼠AILI模型中受到MET的调控。综上所述，本研究证实MET通过抑制线粒体细胞死亡信号通路，在APAP过量诱导的肝损伤中发挥关键的肝毒性抑制作用。本实验中，研究人员向野生型与肝细胞特异性MET敲除小鼠给予300 mg/kg剂量的对乙酰氨基酚（APAP），并于APAP给药后0、6及24小时采集肝脏组织。