Identification of paraxanthine as the most potent caffeine-derived inhibitor of connective tissue growth factor expression in liver parenchymal cells.

Recently, we identified hepatocytes as the major cellular source of profibrogenic connective tissue growth factor (CTGF/CCN2) in the liver. Based on reports of a hepatoprotective effect of coffee consumption, we were the first to provide evidence that caffeine suppresses transforming growth factor (TGF)-beta dependent and -independent CTGF expression in hepatocytes in vitro and in vivo, thus suggesting this xanthine-alkaloid as a potential therapeutic agent. This study aims at comparing the inhibitory capacities of caffeine and its three demethylated derivates paraxanthine, theophylline and theobromine on CTGF expression in hepatocytes and hepatic stellate cells (HSC). Our data suggest paraxanthine as the most important pharmacological repressor of hepatocellular CTGF expression among the caffeine-derived metabolic methylxanthines with an inhibitory dosage (ID)50 of 1.15 mM, i.e. 3.84-fold lower than what is observed for caffeine. In addition, paraxanthine displayed the least cell toxicity as proven by the water-soluble tetrazolium-1 cell vitality assay. However, caffeine or any of the metabolites did not inhibit CTGF expression in HSC. At the toxicological threshold concentration of 1 mM for paraxanthine, we observed an inhibition of hepatocellular CTGF synthesis by 44%, which was strongly reverted in the presence of the specific competitive cyclic adenosine monophosphate inhibitor Rp-adenosine 3',5-cyclic monophosphorothioate triethylammonium salt. Furthermore, CTGF protein expression induced by various concentrations of TGF-beta (0.13-1 ng/ml) is still reduced by, on average, 27%/45% in the presence of paraxanthine (1.25 mM/2.5 mM). Our data provide an evidence-based suggestion of the caffeine-derived primary metabolite paraxanthine as a potentially powerful antifibrotic drug by its inhibitory effect on (hepatocellular) CTGF synthesis.