Targeted disruption of PDE3B, but not PDE3A, protects murine heart from ischemia/reperfusion injury.

Although inhibition of cyclic nucleotide phosphodiesterase type 3 (PDE3) has been reported to protect rodent heart against ischemia/reperfusion (I/R) injury, neither the specific PDE3 isoform involved nor the underlying mechanisms have been identified. Targeted disruption of PDE3 subfamily B (PDE3B), but not of PDE3 subfamily A (PDE3A), protected mouse heart from I/R injury in vivo and in vitro, with reduced infarct size and improved cardiac function. The cardioprotective effect in PDE3B(-/-) heart was reversed by blocking cAMP-dependent PKA and by paxilline, an inhibitor of mitochondrial calcium-activated K channels, the opening of which is potentiated by cAMP/PKA signaling. Compared with WT mitochondria, PDE3B(-/-) mitochondria were enriched in antiapoptotic Bcl-2, produced less reactive oxygen species, and more frequently contacted transverse tubules where PDE3B was localized with caveolin-3. Moreover, a PDE3B(-/-) mitochondrial fraction containing connexin-43 and caveolin-3 was more resistant to Ca(2+)-induced opening of the mitochondrial permeability transition pore. Proteomics analyses indicated that PDE3B(-/-) heart mitochondria fractions were enriched in buoyant ischemia-induced caveolin-3-enriched fractions (ICEFs) containing cardioprotective proteins. Accumulation of proteins into ICEFs was PKA dependent and was achieved by ischemic preconditioning or treatment of WT heart with the PDE3 inhibitor cilostamide. Taken together, these findings indicate that PDE3B deletion confers cardioprotective effects because of cAMP/PKA-induced preconditioning, which is associated with the accumulation of proteins with cardioprotective function in ICEFs. To our knowledge, our study is the first to define a role for PDE3B in cardioprotection against I/R injury and suggests PDE3B as a target for cardiovascular therapies.

Disruption of caveolae blocks ischemic preconditioning-mediated S-nitrosylation of mitochondrial proteins.

AIMS: Nitric oxide (NO) and protein S-nitrosylation (SNO) play important roles in ischemic preconditioning (IPC)-induced cardioprotection. Mitochondria are key regulators of preconditioning, and most proteins showing an increase in SNO with IPC are mitochondrial. The aim of this study was to address how IPC transduces NO/SNO signaling to mitochondria in the heart. RESULTS: In this study using Langendorff perfused mouse hearts, we found that IPC-induced cardioprotection was blocked by treatment with either N-nitro-L-arginine methyl ester (L-NAME, a constitutive NO synthase inhibitor), ascorbic acid (a reducing agent to decompose SNO), or methyl-?-cyclodextrin (M?CD, a cholesterol sequestering agent to disrupt caveolae). IPC not only activated AKT/eNOS signaling but also led to translocation of eNOS to mitochondria. M?CD treatment disrupted caveolar structure, leading to dissociation of eNOS from caveolin-3 and blockade of IPC-induced activation of the AKT/eNOS signaling pathway. A significant increase in mitochondrial SNO was found in IPC hearts compared to perfusion control, and the disruption of caveolae by M?CD treatment not only abolished IPC-induced cardioprotection, but also blocked the IPC-induced increase in SNO. INNOVATION: These results provide mechanistic insight into how caveolae/eNOS/NO/SNO signaling mediates cardioprotection induced by IPC. CONCLUSION: Altogether these results suggest that caveolae transduce eNOS/NO/SNO cardioprotective signaling in the heart.