OXIDIZED PHOSPHATIDYLCHOLINES ARE LOCALIZED WITHIN MITOCHONDRIA DURING MYOCARDIAL ISCHEMIA REPERFUSION INJURY

Abstract

Myocardial Ischemia-Reperfusion (I/R) injury occurs due to rapid reperfusion of ischemic myocardium, after an ischemic episode. Oxidized phosphatidylcholines (OxPCs) are considered to play an important role in IR injury. The purpose of present study was to determine the production and cellular localization of OxPCs during ischemia reperfusion injury in mice. IR in adult male mice was stimulated by ligation of the left anterior descending coronary artery for 60 min and then reperfusion for 24 hours. Myocardial infarction was confirmed by echocardiography and TTC staining. OxPCs in myocardial tissue were quantified using reverse phase liquid chromatography coupled to 4000 QTrap mass spectrometer. In addition, cardiomyocytes were also isolated from adult rats using the Langendorff apparatus and cells were perfused with deoxygenated buffer followed by oxygenated buffer. The cells were fractionated and the nuclear, mitochondrial and cytosolic fractions were analyzed to identify OxPCs. The OxPC level in sham control group was 3.78±0.62ng/mg tissue which significantly increased to 13.54±5.06ng/mg tissue in IR group. This 3.6 fold increase (p-value = 0.03) was seen in both the fragmented and non-fragmented OxPCs. The isolated ischemic cardiomyocytes showed a 1.6 times increase in OxPC levels as compared to control. Subcellular fractionation of cardiomyocytes showed a preferential localization of OxPCs within the mitochondrial fraction, with 56% of total OxPCs present in the mitochondria as compared to other cell fractions. In addition, there was 9.37ng/mg of fragmented OxPCs in the mitochondrial fraction, whereas 5.22ng/mg was in the rest of the cell. The result of the study shows that myocardial I/R leads to a significant increase in OxPC production within myocardial tissue. We are reporting for the first time, that OxPCs are preferentially generated within the mitochondria fraction of cardiomyocytes during I/R. The increase of apoptotic fragmented OxPCs, suggests that the mitochondria might be the epicenter of OxPC mediated apoptosis during I/R.