Vascular calcification is an active cell-mediated process that reduces elasticity of blood vessels and increases blood pressure. Until now, the molecular basis of vascular calcification has not been fully understood. We previously reported that microtubule disturbances mediate vascular calcification. Here, we found that protein kinase C (PKC) signaling acted as a novel coordinator between cytoskeletal changes and hyperphosphatemia-induced vascular calcification. Phosphorylation and expression of both PKC alpha and PKC delta decreased during inorganic phosphate (Pi)-induced vascular smooth muscle cell (VSMC) calcification. Knockdown of PKC isoforms by short interfering RNA as well as PKC inactivation by Go6976 or rottlerin treatment revealed that specific inhibition of PKC alpha. and PKC delta accelerated Pi-induced calcification both in VSMCs and ex vivo aorta culture through upregulation of osteogenic signaling. Additionally, inhibition of PKC alpha and PKC delta induced disassembly of microtubule and actin, respectively. In summary, our results indicate that cytoskeleton perturbation via PKC alpha, and PKC delta inactivation potentiates vascular calcification through osteogenic signal induction. (C) 2014 Elsevier Inc. All rights reserved.