Inhibition of receptor-interacting protein kinase 1 improves experimental non-alcoholic fatty liver disease

Abstract

Background & Aims: In non-alcoholic fatty liver disease (NAFLD), hepatocytes can undergo necroptosis, a regulated form of necrotic cell death mediated by the receptorinteracting protein kinase (RIPK) 1. We herein assessed the potential of RIPK1 and its downstream effector mixed lineage kinase domain-like protein (MLKL), as therapeutic targets and markers of activity in NAFLD. Methods: C57/BL6J-mice were fed a normal chow diet (NCD) or high fat diet (HFD). The effect of RIPA-56, a highly specific inhibitor of RIPK1, was evaluated in either a prophylactic or a curative treatment of HFD-fed mice, and in primary human steatotic hepatocytes. RIPK1 and MLKL concentrations were measured in the serum of patients with NAFLD. Results: Both prophylactic and curative treatments of HFD-fed mice with RIPA-56, caused a down-regulation of MLKL and a reduction of liver injury, inflammation and fibrosis, characteristic of non-alcoholic steatohepatitis (NASH), as well as of steatosis. This latter effect was reproduced by treating primary human steatotic hepatocytes with RIPA-56 or necrosulfonamide (NSA), a specific inhibitor of human MLKL, and by knocking out (KO) MLKL in fat-loaded AML-12 mouse hepatocytes. MLKL KO in steatotic hepatocytes, caused an activation of the mitochondrial respiration, and an increase in b-oxidation. Along with MLKL decreased activation, RIPK3-KO mice exhibited increased activities of the liver mitochondrial respiratory chain complexes in experimental NASH. In patients with NAFLD, serum concentrations of RIPK1 and MLKL increased in correlation with the activity. Conclusion: The inhibition of RIPK1 improves NASH features in HFD-fed mice and reverses steatosis by an MLKL-dependent mechanism that involves at least partly an increase in mitochondrial respiration. RIPK1 and MLKL are potential serum markers of activity and promising therapeutic targets in NAFLD.

Necroptosis in Cholangiocarcinoma

“…Over the last years, several medications have been developed to modulate necroptosis, and numerous studies have provided the basis for moving some of these therapies toward clinical settings [2,18,31,54,55,59,72,111,112]. From what has been reported above, it is clear that RIPK1 and RIPK3 functions are essential for necroptotic cascade initiation and that it is possible to regulate necroptosis by acting on the functions of these molecules [4].…”

“…In the setting of non-alcoholic fatty liver disease (NAFLD)/non-alcoholic steatohepatitis (NASH), it was initially reported that RIPK3 might maintain white adipose tissue homeostasis and prevent glucose intolerance by regulating caspase-8 expression, thus reducing adipose tissue apoptosis and inflammation [2,57,59,[69][70][71]. In a high-fat diet mouse model, RIPK1 inhibition, and the subsequent downstream necrosome inactivation, improved all the histologic features of NASH, including liver inflammation and fibrosis [72]. Some studies indicated that the effects of RIPK3 and MLKL blockage are not consistent, with RIPK3 deletion that seemed to be harmful, while MLKL deletion that seemed to be beneficial in NAFLD [2,59,69,71].…”

“…In human NAFLD/NASH biopsy samples and serum, increased RIPK3 and MLKL expressions have been described, suggesting that necroptosis may also mediate liver injury, oxidative stress, and liver fibrosis in humans, through a positive feedback loop involving the activation of Jun N-terminal kinase [2,55,73,75]. Moreover, the serum concentrations of RIPK1 and MLKL increased with histological activity, and correlated with alanine transaminase (ALT) levels in patients with NAFLD, suggesting that a blood-based non-invasive assessment of necroptosis activity could be useful for identifying patients with NASH [72].…”

Necroptosis in Hepatosteatotic Ischaemia-Reperfusion Injury

“…In relation to liver injury, the RIPK1 protein level was upregulated in a bile duct ligation (BDL) model in mice [ 100 ]. Moreover, RIPK1 involvement has been suggested in the pathogenesis of NAFLD, as Majdi et al found an increased level of RIPK1 in the serum samples of NAFLD patients [ 101 ].…”

Role of necroptosis in chronic hepatic inflammation and liver disease in Cu/Zn superoxide dismutase deficient mice

“…DAMPs initiate and exacerbate the inflammatory process by binding to cell surface receptors of innate immune cells (13,14). Studies have shown that necroptosis is increased in NAFLD/NASH in mice, and inflammation associated with NAFLD/NASH with high fat feeding in mice is reduced by inhibiting necroptosis (15)(16)(17)(18)(19)(20)(21). In addition, studies with human subjects show that increased necroptosis is associated with NASH (16,17).…”