In liver ischemic preconditioning (IP), stimulation of
adenosine A2a receptors (A2aR) prevents
ischemia/reperfusion injury by promoting
diacylglycerol-mediated activation of
protein kinase C (PKC). By concerting
diacylglycerol to
phosphatidic acid,
diacylglycerol kinases (DGKs) act as terminator of
diacylglycerol signalling. This study investigates the role of DGK in the development of hepatocyte IP. DGK activity and cell viability were evaluated in isolated rat hepatocytes preconditioned by 10 min
hypoxia followed by 10 min re-oxygenation or by the treatment with the A2aR agonist,
CGS21680, and subsequently exposed to prolonged
hypoxia. We observed that after IP or A2aR activation, a decrease in DGK activity was associated with the onset of hepatocyte tolerance to
hypoxia. CGS21680-induced stimulation of A2aR specifically inhibited DGK
isoform theta by activating RhoA-
GTPase. Consistently, both
siRNA-mediated downregulation of DGK theta and hepatocyte pretreatment with the DGK inhibitor
R59949 induced cell tolerance to
hypoxia. The pharmacological inhibition of DGK was associated with the
diacylglycerol-dependent activation of PKC delta and epsilon and of their downstream target
p38 MAPK. In conclusion, we unveil a novel signalling pathway contributing to the onset of hepatocyte preconditioning, which through RhoA-
GTPase, couples A2aR to the downregulation of DGK. Such an inhibition is essential for the sustained accumulation of
diacylglycerol required for triggering PKC-mediated survival signals.