Involvement of Cpi-17 and zipper-interacting protein kinase in the regulation of protein kinase C-alpha, protein kinase C-epsilon on vascular calcium sensitivity after hemorrhagic shock.

Our previous studies have found that protein kinase C (PKC) alpha and epsilon take part in the regulation of calcium sensitization and vascular reactivity after hemorrhagic shock. However, the regulatory mechanism is still unclear. The aim of this study was to investigate the role of PKC-potentiated inhibitory protein for protein phosphatase 1 of 17 kDa (CPI-17) and zipper-interacting protein kinase (ZIPK) in the regulation of PKC-alpha and PKC-epsilon on vascular calcium sensitivity after hemorrhagic shock. Using superior mesenteric artery from hemorrhagic shock rats (at 40 mmHg for 2 h), the effects of PKC-alpha and PKC-epsilon agonists on calcium sensitivity and the phosphorylation of 20-kDa myosin light chain (MLC20), and the modulating effects of CPI-17 and ZIPK in this process were observed. In hypoxia-treated vascular smooth muscle cells (VSMCs), the effects of PKC-alpha and PKC-epsilon agonists on protein expression and activity of CPI-17 and ZIPK and the modulating effects of CPI-17 and ZIPK on the regulation of PKC-alpha and PKC-epsilon on the activity of myosin light chain phosphatase (MLCP) were observed. The results indicated that PKC-alpha and PKC-epsilon agonists thymelea toxin and carbachol seemed to increase the calcium sensitivity and MLC20 phosphorylation of superior mesenteric artery after hemorrhagic shock and antagonize the increase of MLCP activity in VSMC after hypoxia. The effects of PKC-alpha and PKC-epsilon agonists were abolished by CPI-17 and ZIPK neutralizing antibody. The protein expression and activity of CPI-17 and ZIPK in VSMC were suppressed after hypoxia but increased by PKC-alpha and PKC-epsilon agonists. Immunoprecipitation experiment showed that PKC-alpha and PKC-epsilon, especially PKC-epsilon, may directly bind with ZIPK but not CPI-17 in VSMC, and ZIPK may directly bind with CPI-17. These results suggest that CPI-17 and ZIPK participate in the regulation of PKC-alpha and PKC-epsilon on vascular calcium sensitivity after hemorrhagic shock. Protein kinase C alpha and epsilon regulation of MLCP-MLC20 phosphorylation pathway in the process of VSMC contraction is mainly through ZIPK; CPI-17 may play an indirect modulating role.