Abstract 1780: Caspase-independent Apoptosis Is Activated In The Late Stage Of Heart Failure In Mice Overexpressing Crma In Vivo

Abstract

Caspase inhibition is thought to be an effective strategy for inhibiting apoptosis in heart. Recent data, however, suggest that caspase-independent apoptosis might be activated when caspases are inhibited. Here, we examined whether caspase-independent apoptosis involving apoptosis inducing factor (AIF) is important in the development of heart failure in the setting of caspase inhibition in vivo. Method: We created transgenic mice with cardiac specific overexpression of cytokine response modifier A (CrmA), a known inhibitor of caspases, using αMHC promoter (CrmA Tg mice). A single dose of doxorubicin (DOX; 20 mg/kg) was injected i.p. into male mice to generate acute cardiomyopathy. A quantitative analysis of cardiac apoptosis was performed using TUNEL assay. Results: Caspase activity was completely inhibited in CrmA Tg mice hearts 6 days after DOX injection, whereas the WT hearts showed significant caspase activation. We also observed significantly improved survival in CrmA Tg compared to WT mice (81% vs 38%, p<0.05) 6 days after DOX injection. However, at 12 days, CrmA Tg and WT mice had a similar rate of cardiac apoptosis, and the survival benefit did not persist. Further analysis revealed that CrmA Tg but not WT mice hearts were characterized by a significant release of AIF from the mitochondria at 12 days. This evidence of AIF-induced apoptosis suggests a late activation of AIF in the setting of caspase inhibition. CrmA Tg mice were also pre-treated with 4-AN (4-amino-1,8-napthalimide, 3 mg/kg), a potent inhibitor of PARP, and examined 12 days after DOX injection. PARP is an important regulator of AIF-induced apoptosis, and PARP inhibition significantly improved DOX-impaired cardiac function (+91%, p<0.01) and survival compared to CrmA Tg mice without 4-AN (93% vs. 13%, p<0.05). In addition 4-AN pretreatment resulted in a significant inhibition of PARP activity, blockade of AIF released from mitochondria, and less apoptosis compared to CrmA Tg without 4-AN. Conclusion: In the setting of caspase inhibition, apoptosis can be induced through caspase-independent mechanisms, predominately via AIF, in a model of heart failure in vivo . The concurrent inhibition of AIF activation provides a more effective and complete inhibition of apoptosis.