Sustained activation of p42/p44 mitogen-activated protein kinase during recovery from simulated ischaemia mediates adaptive cytoprotection in cardiomyocytes

Delayed cytoprotection (preconditioning) occurs 24 h after sublethal simula ted ischaemia and reperfusion (SI/R) in neonatal rat ventricular cardiomyoc ytes, SI/R was used to investigate the role of activation of mitogen-activa ted protein kinases (MAPKs), stress-activated protein kinases (SAPKs) and p hosphoinositide 3-kinase-dependent protein kinase B (PKB)/Akt in cytoprotec tion. SI resulted in transient dual (Thr/Tyr) phosphorylation of p42/p44-MA PK and p38-MAPK, weak phosphorylation of p46/p54-SAPK, but no phosphorylati on of PKB. 'Reperfusion' caused further transient phosphorylation of p38-MA PK, but sustained phosphorylation of p42/p44-MAPK (lasting 4 h) and of Ser( 473) of PKB (lasting 2 h). Furthermore, SI/R (24h) induced delayed protecti on against lethal SI, as determined by an increase in cell viability {biore duction of MTT [3-(4, 5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromid e]} and a decrease in cell injury (release of creatine kinase). Both protec tion and phosphorylation of p42/p44-MAPK were blocked by the MEK-1/2 (MAPK/ Erk kinase-1/2) inhibitor PD98059 (50 mu M) when given during SI/R, but not when given during SI alone. The p38-MAPK inhibitor SB203580 (10 mu M) bloc ked the p38-MAPK-dependent phosphorylation of activating transcription fact or 2 in vitro, and the phosphoinositide 3-kinase inhibitor wortmannin (100 nM) blocked PKB phosphorylation on Ser(473). However, neither SB203580 nor wortmannin had any effect on delayed protection. Therefore sustained activa tion of p42/p44-MAPK during simulated 'reperfusion' following sublethal SI mediates preconditioning in cardiomyocytes independently of transient activ ation of p38-MAPK or sustained activation of PKB.