Abstract P271: Genetically Induced Moderate Inhibition of the Proteasome in Cardiomyocytes Exacerbates Myocardial Ischemia-Reperfusion Injury in Mice
Proteasome dysfunction is implicated human ischemic heart disease and observed in experimental myocardial ischemia-reperfusion (I/R) injury. Altered proteasome activities in I/R hearts were reported but it is unclear whether proteasome function in I/R hearts is adequate. Moreover, effects of pharmacological inhibition of the proteasome on I/R injury remain controversial. Hence we sought to determine the adequacy of proteasomal function in I/R hearts and the impact of moderate cardiomyocyte-restricted proteasome inhibition (CR-PSMI) on I/R injury in intact animals. First, myocardial I/R was created by ligation (30min) and subsequent release of the left anterior descending artery in the transgenic (tg) mice overexpressing GFPdgn, a previously validated surrogate proteasome substrate. Compared with the sham controls, myocardial GFPdgn protein levels in the remote area, the board zone, and the area at risk (AAR) of the ventricle 24hrs after reperfusion were significantly increased, indicative of proteasome functional insufficiency (PFI) in the I/R heart. The most important proteasome peptidase resides in β5 subunits of the 20S proteasome. The clinically used proteasome inhibitor bortezomib targets specially the β5 subunit. To achieve CR-PSMI in intact animals, we engineered and tested a catalytically inactive mouse β5 subunit mutant (T60A-β5) and created multiple tg mouse lines in which T60A-β5 was overexpressed under the control of an attenuated mouse mhc6 promoter. Baseline analyses of the tg lines confirm the ability of T60A-β5 to replace endogenous β5 and inhibit proteasome chymotrypsin-like activities in the heart in a dose-dependent manner. A stable tg line with ∼40% replacement was used in this study. The same I/R procedure caused a significantly greater infarct size in T60A-β5 tg mice (66.2% of AAR) than in the littermate Ntg mice (53.4% of AAR). Under either the sham surgery or the I/R condition, a decrease in phosphorylated AKT and an increase in PKCδ proteins were evident in the tg hearts, compared with the respective Ntg groups. These results show that I/R causes PFI in the heart and PFI plays a pathogenic role in I/R injury. A potential mechanism by which PFI contributes to I/R injury is to suppress AKT activation and exacerbate PKCδ signaling.
- © 2011 by American Heart Association, Inc.