Inhibition of extracellular signal-regulated kinase enhances ischemia/reoxygenation-induced apoptosis in cultured cardiac myocytes and exaggerates reperfusion injury in isolated perfused heart

Three major mammalian mitogen-activated protein kinases, extracellular sign al-regulated kinase (ERK), p38, and c-Jun NH2-terminal protein kinase (JNK) , have been identified in the cardiomyocyte, but their respective roles in the heart are not well understood. The present study explored their functio ns and cross talk in ischemia/reoxygenation (I/R)-induced cardiac apoptosis . Exposing rat neonatal cardiomyocytes to ischemia resulted in a rapid and transient activation of ERK, p38, and JNK, On reoxygenation, further activa tion of all 3 mitogen-activated protein kinases was noted; peak activities increased (fold) by 5.5, 5.2, and 6.2, respectively. Visual inspection of m yocytes exposed to I/R identified 18.6% of the cells as showing morphologic al features of apoptosis, which was further confirmed by DNA ladder and ter minal deoxyribonucleotide transferase-mediated dUTP nick end labeling (TUNE L), Myocytes treated with PD98059, a MAPK/ERK kinase (MEK1/MEK2) inhibitor, displayed a suppression of VR-induced ERK activation, whereas p38 and JNK activities were increased by 70.3% and 55.0%, respectively. In addition, th e number of apoptotic cells was increased to 33.4%. With pretreatment of ce lls with SB242719, a selective p38 inhibitor, or SB203580, a p38 and JNK2 i nhibitor, I/R+PD98059-induced apoptotic cells were reduced by 42.8% and 63. 3%, respectively. Hearts isolated from rats treated with PD98059 and subjec ted to global ischemia (30 minutes)/reoxygenation (1 hour) showed a diminis hed functional recovery compared with the vehicle group. Coadministration o f SB203580 attenuated the detrimental effects of PD98059 and significantly improved cardiac functional recovery. The data taken together suggest that ERK plays a protective role, whereas p38 and JNK mediate apoptosis in cardi omyocytes subjected to I/R, and the dynamic balance of their activities is critical in determining cardiomyocyte fate subsequent to reperfusional inju ry.