Abstract 4745: Exogenous aB-crystallin Improves Myocardial Function and Attenuates Apoptosis in Ischemia-Reperfusion Injury
Objective: Investigate the protective effect of exogenous αB-crystallin (heat-shock protein) (CryAB) on myocardial function after ischemia-reperfusion (I/R) injury through a potential anti-apoptotic mechanism.
Methods: Male C57BL/6 (H-2b) mice underwent temporary LAD artery occlusion (30 min). Intra-myocardial injections of 50 mcg CryAB (N=6) or PBS (control) (N=6) given 15 min prior to reperfusion. Intra-peritoneal injections administered every other day. MRI performed to assess LVEF (POD 40). To investigate the effect of CryAB on apoptosis after hypoxia/reoxygenation in vitro, murine atrial cardiomyocytes (HL-1) or human microvascular endothelial cells (HMEC-1) were incubated with CryAB (50 mcg) or PBS in a hypoxia chamber (1% O2, 5% CO2, 94% N2) for 6, 12, and 24 hours. Apoptosis detected by western blot (free bax, cleaved caspases-3, 9, PARP, Bcl-2) and ELISA analyses (cytoplasmic histone-associated DNA fragments and caspase-3 activity).
Results: In vivo: CryAB-treated mice had a 1.8-fold increase in LVEF vs. control mice on POD 40 (27±6% vs. 15±4%, p<0.005). In vitro:
HL-1: no difference in Bcl-2, free bax, cleaved caspases 3, 9, PARP protein expression and no significant difference in either cytoplasmic histone-associated DNA fragments or caspase-3 activity (2.12±0.92 vs. 2.22±0.63, p=0.72; 7168±874 vs. 7487±262, p=0.49, control vs. CryAB);
HMEC-1: significant increase in total Bcl-2 and decrease in free bax, cleaved caspases 3, 9, and PARP protein levels; 1.5-fold decrease in cytoplasmic histone-associated DNA fragments (0.1475±0.0187 vs. 0.0972±0.0231, p<0.01, control vs. CryAB) and 2.7-fold decrease in caspase-3 activity (27.43±7.60 vs. 10.42±2.19, p<0.005, control vs. CryAB).
Conclusion: Exogenous CryAB administration significantly improves cardiac function after I/R injury, in vivo. Interestingly, the protective anti-apoptotic affects of CryAB may target the endothelial cell, not cardiomyocytes, as previously hypothesized. Therefore, CryAB may be a novel therapeutic treatment for myocardial ischemia.