Regulation of Inositol Phospholipid Hydrolysis by Extended Treatment with Angiotensin II in Human Aortic Smooth Muscle Cells

approved: Theresa M. Filtz Long-term stimuli of many systems leads to decreased cellular responsiveness, or desensitization. We characterized the desensitization of angiotensin II (Ang 11)-mediated inositol phospholipid (IP) hydrolysis in cultured human aortic smooth muscle cells (HASMC). Although it has been suggested that the desensitization induced by long-term Mg II exposure may result partially from down-regulation of Ang II receptor, this is not sufficient to explain fully desensitization in many systems. Post-receptor desensitization of IP hydrolysis may also result from phosphorylation or changes in protein levels of the effector enzyme, PLC-. We identified the major PLC-f isoenzymes expressed by HASMC as PLC-Fl and PLC-3. Mg II pretreatment reduced IP accumulation induced by Ang II (lp.M) in a time-dependent manner. Phorbol ester-12myristrate13-acetate (PMA), a protein kinase C (PKC) activator, also reduced Ang Il-stimulated IP accumulation. These results suggest that PKC activation may negatively regulate Ang Il-stimulated IP signaling in HASMC, similar to rat cells. In addition, PKC also reduced IP accumulation stimulated by A1F4, directly activating the G protein. It suggests that the majority of PKC-induced desensitization of Mg TI-stimulated IP signaling occurs downstream of the Mg II receptor in HASMC. However, both PLC-p 1 and PLC-3, expected candidates for PKC phosphorylation, were phosphorylated independently of PKC activation or inhibition, indicating that PKC might not be involved in direct phosphorylation of PLC-1 and PLC-3. Furthermore, PLC-1, but not PLC-3, was highly phosphorylated under basal conditions, suggesting that PLC-1 and PLC-3 may play different roles in IP signaling in HASMC. Redacted for Privacy