Necroptosis is a key pathogenic event in human and experimental murine models of non-alcoholic steatohepatitis

Clin Sci (Lond). 2015 Oct 1;129(8):721-39. doi: 10.1042/CS20140732. Epub 2015 Jun 15.


Hepatocyte cell death, inflammation and oxidative stress constitute key pathogenic mechanisms underlying non-alcoholic fatty liver disease (NAFLD). We aimed to investigate the role of necroptosis in human and experimental NAFLD and its association with tumour necrosis factor α (TNF-α) and oxidative stress. Serum markers of necrosis, liver receptor-interacting protein 3 (RIP3) and phosphorylated mixed lineage kinase domain-like (MLKL) were evaluated in control individuals and patients with NAFLD. C57BL/6 wild-type (WT) or RIP3-deficient (RIP3(-/-)) mice were fed a high-fat choline-deficient (HFCD) or methionine and choline-deficient (MCD) diet, with subsequent histological and biochemical analysis of hepatic damage. In primary murine hepatocytes, necroptosis and oxidative stress were also assessed after necrostatin-1 (Nec-1) treatment or RIP3 silencing. We show that circulating markers of necrosis and TNF-α, as well as liver RIP3 and MLKL phosphorylation were increased in NAFLD. Likewise, RIP3 and MLKL protein levels and TNF-α expression were increased in the liver of HFCD and MCD diet-fed mice. Moreover, RIP3 and MLKL sequestration in the insoluble protein fraction of NASH (non-alcoholic steatohepatitis) mice liver lysates represented an early event during stetatohepatitis progression. Functional studies in primary murine hepatocytes established the association between TNF-α-induced RIP3 expression, activation of necroptosis and oxidative stress. Strikingly, RIP3 deficiency attenuated MCD diet-induced liver injury, steatosis, inflammation, fibrosis and oxidative stress. In conclusion, necroptosis is increased in the liver of NAFLD patients and in experimental models of NASH. Further, TNF-α triggers RIP3-dependent oxidative stress during hepatocyte necroptosis. As such, targeting necroptosis appears to arrest or at least impair NAFLD progression.

Keywords: chronic liver diseases; mixed lineage kinase domain-like (MLKL); necroptosis; non-alcoholic steatohepatitis (NASH); oxidative stress; receptor-interacting protein 3 (RIP3).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Case-Control Studies
  • Cell Death
  • Choline Deficiency
  • Diet, High-Fat
  • Disease Models, Animal
  • Hepatocytes / metabolism*
  • Humans
  • Liver / metabolism
  • Liver / pathology
  • Male
  • Methionine / deficiency
  • Mice
  • Mice, Inbred C57BL
  • Non-alcoholic Fatty Liver Disease / etiology*
  • Non-alcoholic Fatty Liver Disease / metabolism
  • Non-alcoholic Fatty Liver Disease / pathology
  • Oxidative Stress
  • Palmitic Acid
  • Rats
  • Reactive Oxygen Species / metabolism
  • Receptor-Interacting Protein Serine-Threonine Kinases / metabolism
  • Tumor Necrosis Factor-alpha / metabolism


  • Reactive Oxygen Species
  • Tumor Necrosis Factor-alpha
  • Palmitic Acid
  • Methionine
  • RIPK3 protein, human
  • Receptor-Interacting Protein Serine-Threonine Kinases