Effects of angiotensin II and of phorbol ester on protein kinase C activity and on prostacyclin production in cultured rat aortic smooth-muscle cells

Abstract

The role of the Ca2+-sensitive phospholipid-dependent protein kinase C (PKC) was studied in cultured rat aortic smooth-muscle cells, known to respond to angiotensin II (Ang II) by producing prostacyclin, determined by the release of 6-oxo-prostaglandin F1 alpha. PKC activity was measured in the cytosol and the solubilized membrane fraction after DEAE-cellulose chromatography using a linear NaCl gradient. Ang II stimulated the activity of PKC in the cytosolic and in the membrane fractions of aortic smooth-muscle cells. These increases in PKC activity were concentration-dependent and occurred rapidly, reaching a plateau within 10 min. In contrast, phorbol 12-myristate 13-acetate (PMA) rapidly decreased cytosolic PKC activity and at the same time increased membrane PKC activity to reach a plateau after 20 min. Cytosolic PKC activity from control and Ang II-stimulated cells was found to be less dependent on [Ca2+] than was the highly [Ca2+]-dependent membrane PKC activity from the same cells. In contrast, membrane PKC activity from PMA-treated cells was largely [Ca2+]-independent. In the presence of 10 nM-PMA, the sensitivity of cultured smooth-muscle cells towards Ang II was increased, and maximal values of Ang II-induced prostacyclin production were enhanced by about 60%. In cells incubated with both Ang II and PMA, an additive effect on membrane PKC activity was observed, whereas cytosolic PKC activity was suppressed as in cells treated with PMA alone. These results suggest that an increase of the membrane, but not the cytosolic, PKC activity represents a positive signal in the prostacyclin production induced by Ang II stimulation of aortic smooth-muscle cells. PMA seems to induce a state of activation of membrane PKC which does not need increased intracellular [Ca2+] to be fully expressed, whereas Ang II-stimulated membrane PKC activity requires higher Ca2+ concentrations. The possibility exists that the addition of both signals leads to the augmentation of Ang II-stimulated prostacyclin production.

Identification of an Essential Signaling Cascade for Mitogen-activated Protein Kinase Activation by Angiotensin II in Cultured Rat Vascular Smooth Muscle Cells

“…PKC is known to activate MAPK presumably by directly phosphorylating Raf-1 (60). Since the present study showed that Ang II-induced MAPK activation was PI-PLC-dependent in VSMC and Ang II has been shown to stimulate PKC activity in VSMC through PI-PLC activation (14,61), Ang II could activate the Raf-1-MAPK system by activated PKC. Indeed, it has been reported that prolonged phorbol ester treatment significantly suppressed Ang II-induced Raf-1 and MAPK activation in cultured rat VSMC (20).…”

Increased renal and vascular cytosolic phospholipase A2activity in rats with cirrhosis and ascites

“…As in mesangial cells, vasoconstrictor factors stimulate PG synthesis in cultured vascular smooth muscle cells, mainly prostaglandin E 2 and prostaglandin I 2 , which induce cell relaxation. 36,37 Only a few studies have been published assessing PG production in vascular tissue in cirrhosis. The urinary excretion of 2,3-dinor-6-keto-prostaglandin F 1␣ , a prostaglandin I 2 metabolite used to estimate the extrarenal production of prostaglandin I 2 , is increased in patients with cirrhosis, particularly in those with ascites, and correlates inversely with arterial pressure.…”

Effects of phorbol ester‐sensitive PKC (c/nPKC) activation on the production of adiponectin in 3T3‐L1 adipocytes

“…Our previous study (13) showed that activation of atypical PKC (aPKC) led to NF-jB activation and a decrease of PPARc expression. Considering the fact that various hormones, including insulin, angiotensin II, glucagons, parathyroid hormone, endothelin-1, thrombin, and vitamin D3, and high glucose concentration, activate c/nPKC (14)(15)(16)(17)(18)(19), evaluating the role of c/nPKC on gene expression in adipocyte may provide a new understanding of the regulation of insulin sensitivity.…”