Extracellular ATP and adenosine induce cell apoptosis of human hepatoma Li‐7A cells via the A3 adenosine receptor

Extracellular ATP is a potent signaling molecule that modulates a myriad of cellular functions through the activation of P2 purinergic receptors and is cytotoxic to a variety of cells at higher concentrations. The mechanism of ATP‐elicited cytotoxicity is not fully understood. In this study, we investigated the effect of extracellular ATP on the human hepatoma Li‐7A cells. We observed a time‐ and dose‐dependent growth inhibition of Li‐7A cells by ATP, which is accompanied by an increase in the active form of caspase‐3 as well as increased cleavage of its substrate, poly (ADP‐ribose) polymerase. The cytotoxic effect of extracellular ATP was not mediated by the P2X 7 receptor, since (1) the effect was not abolished by the P2X 7 receptor antagonists oxidized ATP and KN‐62, and (2) extracellular ADP, AMP, and adenosine were also cytotoxic. We found that ATP and ADP were degraded to adenosine by Li‐7A cells and that treatment of Li‐7A cells by adenosine resulted in growth inhibition and caspase‐3 activation, indicating that adenosine is the apoptotic agent. Using adenosine receptor agonists and antagonists, as well as inhibitors of adenosine transport and deamination, we showed that the cytotoxic effect of adenosine is specifically mediated by the A3 receptor even though transcripts of A1, A2 A , A2 B , and a splice variant of the P2X 7 receptors were detected in Li‐7A cells by RT–PCR. Cytotoxicity caused by exogenous ATP and adenosine was completely abolished by the caspase‐3 inhibitor Z‐DEVD‐FMK, demonstrating the central role of caspase‐3 in apoptosis of Li‐7A cells.British Journal of Pharmacology (2003) 140 , 1009–1018. doi: 10.1038/sj.bjp.0705523