Evidence for Protein Kinase C-Mediated Activation of Rho- Kinase in a Porcine Model of Coronary Artery Spasm

OBJECTIVE—We have recently demonstrated that protein kinase C (PKC) and Rho-kinase play important roles in coronary vasospasm in a porcine model. However, it remains to be examined whether there is an interaction between the two molecules to cause the spasm. METHODS AND RESULTS—A segment of left porcine coronary artery was chronically treated with IL-1β–bound microbeads in vivo. Two weeks after the operation, phorbol ester caused coronary spasm in vivo and coronary hypercontractions in vitro at the IL-1β–treated segment; both were significantly inhibited by hydroxyfasudil, a specific Rho-kinase inhibitor. Guanosine 5′-[γ-thio]triphosphate (GTPγS), which activates Rho with a resultant activation of Rho-kinase, enhanced Ca sensitization of permeabilized vascular smooth muscle cells, which were resistant to the blockade of PKC by calphostin C. The GTPγS-induced Ca sensitization was greater in the spastic segment than in the control segment. Western blot analysis revealed that only PKCδ isoform was activated during the hypercontraction. CONCLUSIONS—These results demonstrate that PKC and Rho-kinase coexist on the same intracellular signaling pathway, with PKC located upstream on Rho-kinase, and that among the PKC isoforms, only PKCδ may be involved. Thus, the strategy to inhibit Rho-kinase rather than PKC may be a more specific and useful treatment for coronary spasm.