Emerging mechanistic insights of selective autophagy in hepatic diseases

Macroautophagy (hereafter referred to as autophagy), a highly conserved metabolic process, regulates cellular homeostasis by degrading dysfunctional cytosolic constituents and invading pathogens via the lysosomal system. In addition, autophagy selectively recycles specific organelles such as damaged mitochondria (via mitophagy), and lipid droplets (LDs; via lipophagy) or eliminates specialized intracellular pathogenic microorganisms such as hepatitis B virus (HBV) and coronaviruses (via virophagy). Selective autophagy, particularly mitophagy, plays a key role in the preservation of healthy liver physiology, and its dysfunction is connected to the pathogenesis of a wide variety of liver diseases. For example, lipophagy has emerged as a defensive mechanism against chronic liver diseases. There is a prominent role for mitophagy and lipophagy in hepatic pathologies including non-alcoholic fatty liver disease (NAFLD), hepatocellular carcinoma (HCC), and drug-induced liver injury. Moreover, these selective autophagy pathways including virophagy are being investigated in the context of viral hepatitis and, more recently, the coronavirus disease 2019 (COVID-19)-associated hepatic pathologies. The interplay between diverse types of selective autophagy and its impact on liver diseases is briefly addressed. Thus, modulating selective autophagy (e.g., mitophagy) would seem to be effective in improving liver diseases. Considering the prominence of selective autophagy in liver physiology, this review summarizes the current understanding of the molecular mechanisms and functions of selective autophagy (mainly mitophagy and lipophagy) in liver physiology and pathophysiology. This may help in finding therapeutic interventions targeting hepatic diseases via manipulation of selective autophagy.

Mice fed with HFD or MCD; Huh7 cells treated with PA [Gonzalez-Rodriguez et al., 2014] Activation of IBTKα, a member of UPR results in induction of autophagy and activation of NFKB triggering hepatocyte lipotoxicity SFA (PA) treated HepG2 cells [ Willy et al., 2017] Hepatic steatosis induces defective autophagy in the liver and inhibits autophagic proteolysis In human NAFLD, chronic hepatitis B and chronic hepatitis C patient samples [ Fukuo et al., 2014] Defective autophagy in AFLD mTOR activation and a decrease levels of TFEB-mediated lysosomal gene expression lead to defective autophagy Chronic feeding plus acute binge alcohol ("Gao-binge") in mice [Chao et al., 2018] Activation of ACAC/ACC activity and increased malonyl CoA content in liver tissues suppress autophagy.

Activities of LAL decreases in hepatocytes
Chronic ethanol exposure in rats [Kharbanda et al., 1996;Schulze et al., 2017] Defective autophagy in HCC Higher expressions of SQSTM1 and GPC3 (glypican 3; a tumor marker for HCC) indicate defective autophagy liver cancer. [Bao et al., 2014] Impaired autophagy by increasing oxidative stress leads to start hepatocarcinogenesis.

Patients with HCC cells with HCV infection
Liver-specific atg5 -/mice [Tian et al., 2015] HuR increased autophagy mRNA expressions leads to impaired autophagy in HCC cells.
Targeted deletion of BECN1 in embryonic stem (ES) cells or mice [ Yue et al., 2003]

Actions of mitophagy in liver injury: steatosis and fatty liver diseases
Mitochondrial dysfunction is associated with ALFD and NAFLD Liver specimens with ALFD and NAFLD patients [ Gordon, 1973; Ke 2020] Defense against fatty liver by PINK1-PRKN-dependent mitophagy Rat liver specimens (ethanol treatment) [Eid et at. 2013;Wiliam et al.2015;Eid et al. 2016a,b] Mitochondrial dysfunction is associated with NAFLD Liver specimens from patients with NAFLD [ Caldwell et al., 1999;Ke et al.2020] Defense against NAFLD by PRKN-dependent mitophagy Liver tissues from alcat1 KO mice fed with a HFD

Suppression of NAFLD development by TH-induced mitophagy
HepG2 cells and mouse liver tissues fed with a MCD [ Sinha and Yen, 2016] Initiation of DRAM-mediated mitophagy in the development of NAFLD HepG2 cells treated with OA [Pang et al., 2018] Megamitochondria by defective PRKN-independent mitophagy in fatty liver Liver tissues from liver-specific dnm1l -/-, opa1 -/mice and dnm1l -/-opa1 -/mice with MCD feeding [Yamada et al., 2018] Increased mitochondrial protein degradation by activated mitophagy Liver tissues from LDLR KO with a WD feeding  Inflammasome activation by inhibition of mitophagy Mouse liver tissues fed a HFCD and rat primary hepatocyte treated with PA

Actions of mitophagy in liver cancer
Suppression of hepatoma cell growth and liver tumor by ConAactivated BNIP3-dependent mitophagy BALB/c hepatoma cell line ML-1; liver specimens from NOD/SCID mice treated with ConA Lei and Chang, 2007] Activation of Dox-activated cell death of hepatoma cells

HepG2 cells treated with Dox (adriamycin) [Qian and Yang 2009] Improvement of Dox-activated cell death of HepG2 cells by curcumin
HepG2 cells treated with Dox and curcumin [Qian et 2011] Cytotoxicity of sorafenib is amplified in hepatoma cells by melatonin-activated mitophagy

HepG2, Hep3B and Huh7 cells treated with melatonin and sorafenib [ Prieto-Domínguez et al.2016] Cell death of hepatoma cells is triggered by inducing DRAMdependent mitophagy
HepG2, Hep3B and Huh7 cells [ Liu et al., 2014] Inhibition of initiation of HCC by FUNDC1-activated mitophagy through suppression of inflammasome Liver tissue specimens from HCC patients and liver tissues from liver-specific fundc1 KO mice  Elevation of HCC cell viability by upregulated DNM1L and decreased MFN1 levels Liver tissue specimens of HCC patients and mouse xenograft models, Bel7402 and SMMC7721 cell lines [ Huang et al., 2016] Preservation of the stemness of CSCs by activating NANOG and initiation of mitophagy HepG2, Hep3B and Huh7 cells [ Liu et al., 2017]

Actions of mitophagy in other liver diseases
Melatonin-induced mitophagy protects against CCl4-induced liver fibrosis CCl4 treated rat liver tissue specimens [ Kang et al., 2016] CCl4 activates of PINK1-PRKN-dependent mitophagy in Kupffer cells but TIMD4/TIM-4 suppresses it CCl4 treated mouse liver tissue specimens [Wu et al 2020] Fine particulate matter (PM2.5) activates HSCs and causes liver fibrosis; and inhibition of mitophagy alleviates the fibrosis LX-2 cells and primary HSCs [Qiu et al., 2019] Defective mitophagy promotes inflammasome activation in the HSC model Liver specimens from patients with acute liver failure and mice treated with LPS and LX-2 cells treated with H2O2, LPS, NAC or FCCP [ Tian et al., 2018] NR4A1-PRKDC-TP53 axis acts as a signaling pathway for AFLD pathogenesis Hepatocytes from nr4a1 KO mice, and liver-specific prkdc KO mice (ethanol treatment)  Defective mitophagy increases lipogenesis via upregulation of lipogenic enzymes Liver tissues from bnip3 KO mice and PMH from bnip3null mice [Glick et al., 2012] TH-activated mitophagy increases FA β-oxidation through inducing CPT1α expression Liver specimens from thr KO mice and HepG2 cells [ Singh et al., 2018] HFD-fed REDD1 KO mice increase CPT1A, BNIP3 and PRKN expression in the livers Liver specimens from ddit4/redd1 KO mice treated with HFD [ Dumas et al., 2020] Insulin resistance (IR) inhibits mitophagy Liver specimens from B6 mice with HFD and PMH [ Liu et al., 2009] Any defect in PRKN-dependent mitophagy does not change in obesity and IR Liver specimens from prkn KO mice and PMH treated with HFD [Costa et al., 2016;Edmunds et al., 2019] Loss of FUNDC1-mediated mitochondrial turnover induce adipose tissue-associated macrophage infiltration Supplementary Figure 1 Figure S1 legend. Molecular mechanisms of various stages of autophagy. Autophagy is activated in response to various cellular stresses and is triggered by a decrease in rapamycin complex 1 (mTORC1) activity due to the activation of AMP-activated protein kinase (AMPK) or p53 signaling. mTORC1 suppresses the activity of Unc-51-like autophagy activating kinase 1 (ULK1) complex. Therefore, inhibition of mTORC1 causes the initialization of the ULK1-mediated formation of the isolation (autophagosomal) membrane (IM) in association with the class III phosphatidylinositide 3-kinase (PI3K) complex (PI3KC3). The IM expands into an autophagosome (AP) with a double-layer membrane, which can engulf any cellular component, including proteins, damaged organelles, and lipid droplets. The AP merges with the lysosome (via LAMP-1, 2), forming autophagolysosome (APL) or autolysosome (AL), and resulting in the degradation of the cargo by cathepsins and the autophagic lysosome reformation (ALR). The nucleation, elongation and maturation of the IM are dependent on two ubiquitin-like conjugation systems (ATG12 and ATG8), which involve multiple autophagy proteins, including Beclin1, ATG5, ATG16 and MT-associated protein 1 light chain 3 (LC3). The AL provides an acidic milieu for hydrolytic enzymes to digest the engulfed components. Nuclear localization of transcription factor EB (TFEB) is critical to the formation of lysosomes and to the enhanced expression of autophagy proteins. Importantly, autophagy could be selective of mitochondria (mitophagy) or ER (ER-phagy). (Al-Bari et al 2021, reprinted with permission from IJMS).