BREAKDOWN AND RELEASE OF MYOFILAMENT PROTEINS DURING ISCHEMIA AND ISCHEMIA REPERFUSION IN RAT HEARTS - IDENTIFICATION OF DEGRADATION PRODUCTS AND EFFECTS ON THE PCA-FORCE RELATION/

Our objective in experiments reported here was to identify myofilament proteins of rat hearts either lost or degraded by cardiac ischemia (1 5- or 60-minute duration) with and without 45 minutes of reperfusion. We correlated these changes with alterations in myofilament sensitivit y to Ca2+ and maximum force generation. Protein degradation and loss w ere assessed by high-performance liquid chromatography, SDS-PAGE, West ern blotting analysis, and amino acid sequencing. Compared with nonisc hemic control hearts, bundles of skinned fibers from hearts subjected to ischemia alone demonstrated a decrease in maximum force generation and an increase in sensitivity to Ca2+. These changes in function were increased with the duration of the ischemia and with reperfusion. Wit h increasing duration of ischemia, there was an increased loss and deg radation of myofibrillar alpha-actinin and troponin I (TnI) at its C-t erminus. alpha-Actinin and TnI were most susceptible to ischemia, but with 60 minutes of ischemia/reperfusion, there was also degradation of myosin light chain-1 (MLC1) involving a clip of residues 1 to 19. The MLC1 degradation product was detected in the reperfusion effluent (al ong with troponin T, tropomyosin, and alpha-actinin) but not in the ti ssue with 60 minutes of ischemia with no reperfusion. Moreover, with i schemia the following proteins became associated with the myofibrils: GAPDH and proteins of the mitochondrial ATP synthase complex. Our resu lts provide new evidence regarding the mechanism by which ischemia/rep erfusion causes myocardial injury and support the hypothesis that an i mportant element in the injury is altered activity and structure of th e myofilaments.