Lipid-induced ASK1 activation in hepatocytes and Kupffer cells mediates the increased vulnerability of fatty liver to ischemia/reperfusion injury in mice

Lipid-induced ASK1 activation in hepatocytes and Kupffer cells mediates the increased vulnerability of fatty liver to ischemia/reperfusion injury in mice Chiara Imarisio1*, Elisa Alchera1*, Bangalore R Chandrashekar1*, Guido Valente2, Antonia Follenzi1, Elena Trisolini1, Renzo Boldorini1 and Rita Carini1 [email protected], [email protected], [email protected], [email protected], [email protected] [email protected], [email protected], [email protected] 1Department of Health Science, University of Piemonte Orientale, 28100 Novara, Italy. 2Department of Translational Medicine, University of Piemonte Orientale, 28100 Novara, Italy. * These Authors equally contributed to the study ABSTRACT Background & Aims: Steatosis enhances parenchymal injury and inflammation in liver exposed to ischemia/reperfusion (I/R). Several alterations, such as ER stress and increased ROS production are associated to such effects but a final and targetable pathogenic mechanism is still undetermined. This study investigates “in vitro” and “in vivo” a molecular mediator of the interplay among ER stress, ROS and cytotoxic/survival pathways and its role in the susceptibility of steatotic hepatocytes (HP) and Kupffer cells (KC) to hypoxia/reoxygenation (H/R) and of fatty liver to I/R injury and inflammation. Methods: Control or steatotic (treated with palmitic acid, PA) primary mouse HP and KC were exposed to H/R to “in vitro” simulate I/R exposure. C57BL/6 mice fed 9 weeks with control or High Fat diet underwent to a non-lethal partial hepatic I/R. Results: In HP, PA increases H/R damage, induces ROS production and enhances the stimulation of the ASK1/JNK cytotoxic axis activated by the ER stress mediator TRAF2 during H/R. Prevention of ROS production nullifies the increased susceptibility of HP to H/R and the enhanced ASK1/JNK activation. ASK1 inhibition also completely protects JNK activation and HP damage. In KC, PA alone induces TRAF2 and a consequent ASK1 and p38 MAPK activation. PA also increases KC damage induced by H/R, but oppositely to HP, ASK1 inhibition enhances H/R damage by preventing the stimulation of the survival mediator p38 MAPK. In mice liver, steatosis induces the expression of activated ASK1 in KC, whereas upon I/R exposure, activated ASK1 expression is evident in both in KC and HP. “In vivo”, ASK1 inhibition prevents ASK1, JNK and p38 MAPK activation and protects fatty mice liver from I/R-induced transaminases release and from the increase of TNF-alpha and iNOS. Conclusions: Our results show that: 1) Lipids increase ASK1/JNK activation induced by ER in HP by rising cellular ROS; 2) Lipids activate ASK1/p38 MAPK in KC by promoting ER stress; 3) ASK1 is cytoxic for HP and protective for KC; 4) ASK1 inhibition protects I/R injury and inflammation of fatty liver. These observations indicate that steatosis, by stimulating ASK1, contextually promotes I/R induced liver injury and inflammation by increasing HP damage and protecting the resident hepatic macrophages (KC) and evidence the potentiality of ASK1 inhibitors as novel therapeutic agents to prevent hepatic damage and reduce inflammatory reactions consequent to fatty liver surgery.