Lipotoxicity in NASH

Exposure to a sedentary lifestyle, fat-rich and fiber-poor diets, positive caloric imbalance and an extended life disrupt the metabolic homeostasis causing non-alcoholic fatty liver disease. A subset of patients with this emerging public health problem may develop non-alcoholic steatohepatitis (NASH) and progress to cirrhosis and liver cancer [[1]Fuchs M. Sanyal A.J. Non-alcoholic fatty liver disease: a pathophysiological perspective.in: Arias I.M. Alter H.J. Boyer J.L. Cohen D.E. Fausto F. Shafritz D.A. Wolkoff A.W. The liver. Biology and pathobiology. 5th ed. Wiley & Sons, 2010: 719-741Google Scholar]. Understanding the molecular mechanisms promoting liver injury in NASH is not only of biomedical and public health interest, but also key to develop new avenues for specific treatment interventions. In this Snapshot article, we will outline current evolving concepts of signaling cascades that may be linked to liver injury in NASH. It is important to acknowledge that a significant amount of information has been obtained from in vitro and animal studies that may not entirely reflect the situation in the liver of patients with NASH. In addition, multiple pathways leading to apoptosis may be operational in NASH patients at the same time, but the relative contribution of each one is unknown. Nevertheless, targeting apoptotic pathways in NASH may represent a viable therapeutic strategy particularly in the context of acute liver injury augmented by fatty liver or in the setting of transplantation using highly steatotic donor livers. Major sources of hepatic saturated fatty acids (SFAs) are adipose tissue, diet, and de novo lipogenesis from glucose. Under physiological conditions, SFA are transported to mitochondria for β-oxidation or esterified for either excretion in very low density lipoproteins or storage as lipid droplets. SFA can be released from lipid droplets via macrolipophagy (Fig. 1A). Multiple mechanisms are concurrently operative to produce liver injury in hepatocytes overwhelmed by SFA, primarily from adipocyte lipolysis, and free cholesterol from de novo synthesis [1Fuchs M. Sanyal A.J. Non-alcoholic fatty liver disease: a pathophysiological perspective.in: Arias I.M. Alter H.J. Boyer J.L. Cohen D.E. Fausto F. Shafritz D.A. Wolkoff A.W. The liver. Biology and pathobiology. 5th ed. Wiley & Sons, 2010: 719-741Google Scholar, 2Simonen P. Kotronen A. Hallikainen M. Sevastianova K. Makkonen J. Hakkarainen A. et al.Cholesterol synthesis is increased and absorption decreased in non-alcoholic fatty liver disease independent of obesity.J Hepatol. 2011; 54: 153-159Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar]. These lipids, and in particular SFA, can activate a variety of intracellular responses resulting in lipotoxic stress in the endoplasmic reticulum (ER) and mitochondria, respectively. As a consequence, apoptosis occurs which represents a key pathogenic feature of NASH. Metabolic stress from lipids has also been linked to macrolipophagy dysfunction, presumably at the level of autophagosome-lysosome fusion [[3]Singh R. Kaushik S. Wang Y. Xiang Y. Novak I. Komatsu MTanaka K.E. et al.Autophagy regulates lipid metabolism.Nature. 2009; 458: 1131-1135Crossref PubMed Scopus (2522) Google Scholar]. Whether this promotes additional lipid accumulation or increased lipolysis in NASH remains to be addressed. Continuous excessive availability of SFA likely generates lipid intermediates that will shift normal triacylglyceride formation towards induction of ER stress with accumulation of unfolded or misfolded proteins in the ER [[4]Mantzaris M.D. Tsianos E.V. Galaris D. Interruption of triacylglycerol synthesis in the endoplasmic reticulum is the initiating event for saturated fatty acid-induced lipotoxicity in liver cells.FEBS J. 2011; 278: 519-530Crossref PubMed Scopus (61) Google Scholar]. This perturbation promotes a state of oxidative stress and triggers an evolutionary conserved adaptive response activating signaling pathways that result in translational arrest and degradation of proteins and production of antioxidants to allow recovery and cell survival (Fig. 1B). This adaptive response is activated by at least three ER stress sensors, activating transcription factor 6 (ATF6), inositol requiring enzyme 1 (IRE1), and PKR-like ER kinase (PERK), respectively. Failure to upregulate activating transcription factor 4 (ATF4) and efficiently degrade proteins in response to activating X-box protein 1 (XBP1) despite translational arrest of proteins synthesis via eukaryotic initiation factor 2α (eIF2α) may identify NASH patients at particular risk to progress to cirrhosis due to insufficient degradation of unfolded proteins [[1]Fuchs M. Sanyal A.J. Non-alcoholic fatty liver disease: a pathophysiological perspective.in: Arias I.M. Alter H.J. Boyer J.L. Cohen D.E. Fausto F. Shafritz D.A. Wolkoff A.W. The liver. Biology and pathobiology. 5th ed. Wiley & Sons, 2010: 719-741Google Scholar]. It remains to be determined whether the antioxidant response via combined activation of ATF4 and the transcription factor NRF2 is impaired in patients with NASH [[5]Hardwick R.N. Fisher C.D. Canet M.J. Lake A.D. Cherrington N.J. Diversity in antioxidant response enzymes in progressive stages of human nonalcoholic fatty liver disease.Drug Metab Dispos. 2010; 38: 2293-2301Crossref PubMed Scopus (143) Google Scholar]. Once the adaptive response machinery fails and the ER stress continues, then the alarm response is activated and promotes apoptosis (Fig. 1C). IRE1, PERK, and ATF6 all converge at the level of C/EBP homologous protein (CHOP) which forms a heteromeric complex with c-Jun to positively regulate p53 upregulated modulator of apoptosis (PUMA) expression with subsequent B-cell lymphoma 2-associated X protein (Bax) activation [[6]Cazanave S.C. Elmi N.A. Akazawa Y. Bronk S.F. Mott J.L. Gores G.J. CHOP and AP-1 cooperatively mediate PUMA expression during lipoapoptosis.Am J Physiol Gastrointest Liver Physiol. 2010; 299: G236-G243Crossref PubMed Scopus (148) Google Scholar]. As a consequence, mitochondrial membrane channel formation results in the release of cytochrome c into the cytosol, promoting activation of downstream effector caspase-3 and -7, proteases that dismantle the cell and cause cell death by apoptosis. IRE1 can also bind tumor necrosis factor receptor-associated factor 2 (TRAF2) to activate apoptosis signal-regulating kinase 1 (ASK1) and downstream Jun N-terminal kinase (JNK) to facilitate formation of the c-Jun/CHOP heteromeric complex [[7]Kim I. Xu W. Reed J.C. Cell death and endoplasmic reticulum stress: disease relevance and therapeutic opportunities.Nat Rev Drug Discov. 2008; 7: 1013-1030Crossref PubMed Scopus (1446) Google Scholar]. CHOP in addition upregulates death receptors (e.g. DR5) that sensitize hepatocytes to circulating death ligands. Mitochondrial dysfunction may be further augmented by stress-induced release of calcium by the ER. The toll-like receptor 4 (TLR4) is a pattern-recognition receptor that activates a proinflammatory-signaling pathway in response to SFAs. This pathway (Fig. 1D) is initiated by recruiting adaptor molecules Toll/IL-1 receptor domain containing adaptor protein (TIRAP) and myeloid differentiation factor 88 (MyD88) that ultimately leads to activation of nuclear factor κB with production of tumor necrosis factors α (TNFα) [[8]Takeda K. Akira S. Toll-like receptors in innate immunity.Int Immunol. 2005; 17: 1-14Crossref PubMed Scopus (2656) Google Scholar]. Binding of TNFα to the TNF receptor forms a complex consisting of TNF receptor associated death domain protein (TRADD), TNF receptor associated factor 2 (TRAF2), and receptor interacting protein (RIP). This complex activates a pro-apoptotic ASK1/JNK pathway that ultimately results in Bim activation and mitochondrial dysfunction. Activation of forkhead box-containing protein, class 0, member 3a (FoxO3a) mediated by SFA-stimulated protein phosphatase 2a activity may work in concert with JNK to activate Bim [[9]Barreyro F.J. Kobayashi S. Bronk S.F. Werneburg N.W. Mahli H. Gores G.J. Transcriptional regulation of Bim by FoxO3A mediates hepatocyte lipoapoptosis.J Biol Chem. 2007; 282: 27141-27154Crossref PubMed Scopus (163) Google Scholar]. The TRADD/TRAF2/RIP complex may be internalized and after recruiting Fas-associated protein with death domain (FADD), the mitochondrial amplification loop is activated by caspase 8. Lysosomal involvement in apoptosis has also been described in patients with NASH (Fig. 1D). Upon cytosol-to-lysosome translocation of Bax and lysosomal membrane permeabilization, cathepsin B is released into cytosol with subsequent increased production of TNFα via activation of the NF-κB pathway [[10]Werneburg N.W. Guicciardi M.E. Bronk S.F. Gores G.J. Tumor necrosis factor-alpha associated lysosomal permeabilization is cathepsin B dependent.Am J Physiol Gastrointest Liver Physiol. 2002; 283: G947-G956Crossref PubMed Scopus (162) Google Scholar]. Under physiological conditions, hepatocytes are relatively resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and Fas cytotoxicity. However, in patients with NASH hepatocytes appear to be sensitized and hepatic upregulation of the TRAIL receptor DR5 was observed (Fig. 1E). The authors declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.