Prostacyclin and PPARα Agonists Control Vascular Smooth Muscle Cell Apoptosis and Phenotypic Switch through Distinct 14-3-3 Isoforms

We hypothesized that prostacyclin (PGI(2)) protects vascular smooth muscle cell (VSMC) against apoptosis and phenotypic switch through peroxisome proliferator-activated receptor-alpha (PPAR alpha) activation and 14-3-3 upregulation. Here we showed that transfection of rat aortic VSMC, A-10, with PGI(2)-producing vectors, Ad-COPI, resulted in attenuated H2O2-induced apoptosis accompanied by a selective increase in 14-3-3 beta and 14-3-3 theta expression. Carbaprostacyclin (cPGI(2)) and Wy14,643 exerted a similar effect. The effects of PGI(2) were abrogated by MK886, a PPAR alpha antagonist, but not GSK3787, a PPAR delta antagonist. PPAR alpha transfection upregulated 14-3-3 beta and theta expression and attenuated H2O2-induced apoptosis. H2O2-induced 14-3-3 beta but not 14-3-3 theta degradation was blocked by a caspase 3 inhibitor. Furthermore, 14-3-3 beta but not 14-3-3 theta overexpression reduced, while 14-3-3 beta siRNA aggravated apoptosis. VSMC contractile proteins and serum response factor (SRF) were reduced in H2O2-treated A-10 cells which were concurrently prevented by caspase 3 inhibitor. By contrast, PGI(2) prevented H2O2-induced SM22 alpha and Calponin-1 degradation without influencing SRF. cPGI(2) and Wy14,643 also effectively blocked VSMC phenotypic switch induced by growth factors (GFs). GFs suppressed 14-3-3 beta, theta, epsilon and eta isoforms and cPGI(2) prevented the decline of beta, theta and eta, but not epsilon. 14-3-3 theta siRNA abrogated the protective effect of cPGI(2) on SM22 alpha and Calponin-1 while 14-3-3 theta or 14-3-3 beta overexpression partially restored SM22 alpha. These results indicated that PGI(2) protects VSMCs via PPAR alpha by upregulating 14-3-3 beta and 14-3-3 theta. 14-3-3 beta upregulation confers resistance to apoptosis whereas 14-3-3 theta and beta upregulation protects SM22 alpha and Calponin-1 from degradation.