Role of extracellular vesicles in hypoxia-induced hepatic injury in non-alcoholic fatty liver disease

Transition from isolated steatosis to non-alcoholic steatohepatitis is a key issue in non-alcoholic fatty liver disease (NAFLD). Recent observations in patients with obstructive sleep apnea syndrome (OSAS), suggest that intermittent hypoxia (IH) may contribute to disease progression. Among other mechanisms, release of extracellular vesicles (EV) by injured hepatocytes has been implicated in NAFLD progression. We investigated the effects of hypoxia on liver injury in experimental models of NAFLD though EV. Methods: Hepatocytes treated with free fatty acids were subjected to chemically induced hypoxia (CH) using cobalt chloride (CoCl2). Triglyceride (TG) content, lipotoxicity, pro-inflammatory and pro-fibrotic cytokines and cleaved caspase-1 were assessed. Also, Kupffer cells (KC) and human stellate cell (LX-2) were treated with conditioned medium (CM) and EVs from hypoxic fat-laden hepatocytes. The choline deficient L-amino acid diet (CDAA)-fed mice model used to assess the effects of IH on experimental NAFLD. Results: Both in vitro and in vivo models of NAFLD exhibited increased TG content, lipotoxicity and pro-inflammatory and pro-fibrotic cytokines compared to non-treated hepatocytes or control mice. Cleaved caspase-1 increased in 1) CH-exposed steatotic hepatocytes, 2) CDAA-fed mice exposed to IH, and 3) in EV (from steatotic hepatocytes and CDAA-fed mice). CM and EVs from hypoxic fat-laden hepatic cells evoked a pro-inflammatory and pro-fibrotic phenotype in KC and LX-2 respectively. Conclusion: Our findings in experimental models of NAFLD indicate that hypoxia may increase liver injury and promote disease progression. These results suggest EV and their content (caspase-1) as a potential novel biomarker in NAFLD and OSAS.