CCN1/Integrin α₅β₁ Instigates Free Fatty Acid-Induced Hepatocyte Lipid Accumulation and Pyroptosis through NLRP3 Inflammasome Activation

Hyperlipidemia with high blood levels of free fatty acids (FFA) is the leading cause of non-alcoholic steatohepatitis. CCN1 is a secreted matricellular protein that drives various cellular functions, including proliferation, migration, and differentiation. However, its role in mediating FFA-induced pro-inflammatory cell death and its underlying molecular mechanisms have not been characterized. In this study, we demonstrated that CCN1 was upregulated in the livers of obese mice. The increase in FFA-induced CCN1 was evaluated in vitro by treating hepatocytes with a combination of oleic acid and palmitic acid (2:1). Gene silencing using specific small interfering RNAs (siRNA) revealed that CCN1 participated in FFA-induced intracellular lipid accumulation, caspase-1 activation, and hepatocyte pyroptosis. Next, we identified integrin α₅β₁ as a potential receptor of CCN1. Co-immunoprecipitation demonstrated that the binding between CCN1 and integrin α₅β₁ increased in hepatocytes upon FFA stimulation in the livers of obese mice. Similarly, the protein levels of integrin α₅ and β₁ were increased in vitro and in vivo. Experiments with specific siRNAs confirmed that integrin α₅β₁ played a part in FFA-induced intracellular lipid accumulation, NLRP3 inflammasome activation, and pyroptosis in hepatocytes. In conclusion, these results provide novel evidence that the CCN1/integrin α₅β₁ is a novel mediator that drives hepatic lipotoxicity via NLRP3-dependent pyroptosis.