Adenosine A _2a receptor stimulation prevents hepatocyte lipotoxicity and non-alcoholic steatohepatitis (NASH) in rats

NEFA (non-esterified 'free' fatty acid)-mediated lipotoxicity plays a critical role in the pathogenesis of NASH (non-alcoholic steatohepatitis). In the light of the growing need for new therapeutic options for NASH, we investigated the action of A _2aR (adenosine A _2a receptor) stimulation against lipotoxicity. The effects of the A _2aR agonist CGS21680 [2-p-(2-carboxyethyl)phenethylamino-5′-Nethylcarboxyamidoadenosine] were evaluated 'in vitro' in liver cells exposed to SA (stearic acid) and 'in vivo' in rats with NASH induced by 8 weeks of feeding with an MCD diet (methionine/cholinedeficient diet). In cultured hepatocytes, SA promoted apoptosis by inducing MKK4 (mitogenactivated protein kinase kinase 4)/SEK1 (stress-activated protein kinase/extracellular-signalregulated kinase kinase-1) and JNK-1/2 (c-Jun N-terminal kinase-1/2) activation. CGS21680 addition prevented JNK-1/2 activation and reduced apoptosis without interfering with lipid accumulation. CGS21680 action required PI3K (phosphoinositide 3-kinase)/Akt-mediated block of MKK4/SEK1. Consistently, PI3K inhibition with wortmannin abolished the cytoprotective action of CGS21680 and reverted MKK4 inhibition. SA lipotoxicity was also prevented by transfecting HTC cells with a specific MKK4/SEK1 siRNA (small interfering RNA). In rats receiving the MCD diet, the development of NASH was associated with MKK4/SEK1 and JNK-1/2 activation. CGS21680 (0.5 mg/kg of body weight, intraperitoneal) administration to MCD-fed rats prevented JNK-1/2 activation by acting on MKK4/SEK1. CGS21680 also effectively reduced NASH-associated ALT (alanine aminotransferase) release, hepatocyte apoptosis, liver inflammation and fibrosis without affecting hepatic steatosis. Taken together, these results demonstrate that, by inhibiting JNK-1/2, A _2aR stimulation reduces lipotoxicity and ameliorates NASH, giving a rationale to investigate A _2aR agonists as possible new therapeutic agents in preventing fatty liver progression to NASH.