Hepatocyte-derived MANF mitigates ethanol-induced liver steatosis in mice via enhancing ASS1 activity and activating AMPK pathway

Han-yang Xu1,2, Yan-hong Jiao1,2, Shi-yu Li1,2, Xu Zhu1,2, Sheng Wang3, Yu-yang Zhang1,2, Yi-jun Wei1,2, Yu-jun Shen1,2, Wei Wang4, Yu-xian Shen1,2, Jun-tang Shao1,2
1 School of Basic Medical Sciences, Anhui Medical University, Hefei 230032, China
2 Biopharmaceutical Institute, Anhui Medical University, Hefei 230032, China
3 Center for Scientific Research of Anhui Medical University, Hefei 230032, China
4 Department of Hepatobiliary and Pancreatic Surgery, First Affiliated Hospital of Anhui Medical University, Hefei 230022, China
Correspondence to: Yu-xian Shen:, Jun-tang Shao:,
DOI: 10.1038/s41401-022-00920-8
Received: 2 March 2022
Accepted: 5 May 2022
Advance online: 2 June 2022


Hepatic steatosis plays a detrimental role in the onset and progression of alcohol-associated liver disease (ALD). Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an evolutionarily conserved protein related to the unfolded protein response. Recent studies have demonstrated that MANF plays an important role in liver diseases. In this study, we investigated the role of MANF in ethanol-induced steatosis and the underlying mechanisms. We showed that the hepatic MANF expression was markedly upregulated in mouse model of ALD by chronic-plus-single-binge ethanol feeding. Moreover, after chronic-plus-binge ethanol feeding, hepatocyte-specific MANF knockout (HKO) mice displayed more severe hepatic steatosis and liver injury than wild-type (WT) control mice. Immunoprecipitation-coupled MS proteomic analysis revealed that arginosuccinate synthase 1 (ASS1), a rate- limiting enzyme in the urea cycle, resided in the same immunoprecipitated complex with MANF. Hepatocyte-specific MANF knockout led to decreased ASS1 activity, whereas overexpression of MANF contributed to enhanced ASS1 activity in vitro. In addition, HKO mice displayed unique urea cycle metabolite patterns in the liver with elevated ammonia accumulation after ethanol feeding. ASS1 is known to activate AMPK by generating an intracellular pool of AMP from the urea cycle. We also found that MANF supplementation significantly ameliorated ethanol-induced steatosis in vivo and in vitro by activating the AMPK signaling pathway, which was partly ASS1 dependent. This study demonstrates a new mechanism in which MANF acts as a key molecule in maintaining hepatic lipid homeostasis by enhancing ASS1 activity and uncovers an interesting link between lipid metabolism and the hepatic urea cycle under excessive alcohol exposure.
Keywords: alcohol-associated liver diseases; mesencephalic astrocyte-derived neurotrophic factor; arginosuccinate synthase 1; liver steatosis; hepatic urea cycle

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