Abstract
Calcium-independent phospholipase A(2) (iPLA(2)) is the predominant phospholipase A(2) present in myocardium, and its pathophysiological role in acute myocardial infarction has been suggested by the rapid increase in membrane-associated iPLA(2) activity during myocardial ischaemia and reperfusion (I/R). We therefore examined iPLA(2) in mitochondrial fractions prepared from Langendorff-perfused adult rabbit hearts. Our studies indicate that iPLA(2)beta is present in rabbit heart mitochondrial inner membranes with no apparent translocation during ischaemia, I/R or preconditioning. Mitochondrion-associated iPLA(2) was catalytically competent and exhibited 2-, 3- and 2.5-fold increases in measured iPLA(2) activity following ischaemia, I/R and preconditioning, respectively, when compared with the activity of iPLA(2) measured in mitochondria from control hearts. Mitochondrial phospholipids are essential for maintaining the ordered structure and function of the organelle. I/R resulted in a rapid overall decrease in phosphatidylcholine and phosphatidylethanolamine glycerophospholipid species, as determined by electrospray ionization MS, that was partially alleviated by pretreatment of hearts with the iPLA(2)-specific inhibitor, bromoenol lactone (BEL). Pretreatment of I/R hearts with 10 microM BEL significantly reduced the infarct size almost to that of continuously perfused hearts and was cardioprotective only when administered prior to ischaemia. Cardioprotection by BEL was reversed by the simultaneous perfusion of 100 microM 5-hydroxydecanoate, implicating the mitochondrial K(ATP) channel in BEL-mediated protection from I/R. Preconditioning also significantly reduced the infarct size in response to I/R but protection was lost by concurrent perfusion of 10 microM arachidonic acid. Taken together, these data strongly implicate mitochondria-associated iPLA(2) in the signal transduction of myocardial I/R injury.