Platelet-derived Growth Factor Stimulates Protein Kinase A through a Mitogen-activated Protein Kinase-dependent Pathway in Human Arterial Smooth Muscle Cells

Abstract

The abilities of platelet-derived growth factor (PDGF) and insulin-like growth factor (IGF-I) to regulate cAMP metabolism and mitogen-activated protein kinase (MAP kinase) activity were compared in human arterial smooth muscle cells (hSMC). PDGF-BB stimulated cAMP accumulation up to 150-fold in a concentration-dependent manner (EC 50 Ϸ 0.7 nM). The peak of cAMP formation and cAMP-dependent protein kinase (PKA) activity occurred approximately 5 min after the addition of PDGF and rapidly declined thereafter. Incubating cells with PDGF and 3-isobutyl-1-methylxanthine (IBMX, a phosphodiesterase inhibitor) enhanced the accumulation of cAMP and PKA activity by an additional 2.5-3-fold, whereas IBMX alone was essentially without effect. The PDGF-stimulated increase in cAMP was prevented by addition of the cyclooxygenase inhibitor indomethacin, consistent with release of prostaglandins stimulating cAMP. PDGF, but not IGF-I, stimulated MAPK activity, cytosolic phospholipase A 2 (cPLA 2) phosphorylation, and cAMP synthesis which indicated a key role for MAP kinase in the activation of cPLA 2. Further, PDGF stimulated the rapid release of arachidonic acid and synthesis of prostaglandin E 2 (PGE 2) which could be inhibited by a cPLA 2 inhibitor (AACOCF 3). Calcium mobilization was required for PDGF-induced arachidonic acid release and PGE 2 synthesis but not for MAPK activation, whereas PKC was required for PGE 2-mediated activation of PKA. In summary, these results demonstrate that PDGF increases cAMP formation and PKA activity through a MAP kinase-mediated activation of cPLA 2 , arachidonic acid release, and PGE 2 synthesis in human arterial smooth muscle cells.

The mitogen-activated protein kinase pathway can mediate growth inhibition and proliferation in smooth muscle cells. Dependence on the availability of downstream targets.

“…PGE 2 is one of the principal growth-inhibitory prostaglandins secreted by human SMCs (23). PGE 2 secretion into the culture medium was determined as previously described (24). For immunoblotting experiments, lysates from cells incubated with PDGF-BB for 5 min with or without a 30-min preincubation with 10-30 M PD 098059 or 10 M indomethacin were used for immunoblotting and detection of cPLA 2 band shift due to phosphorylation as described (24).…”

Specifying the cellular responses to IGF signals: roles of IGF-binding proteins

“…Are the signal transduction pathways activated by the IGF-IR that lead to directed migration different from those that lead to proliferation? Early studies using human SMCs suggested that IGF-I stimulation did not activate MAPK in this cell type (Takagi et al 1995, Graves et al 1996, Koyama et al 1996. However, others reported an increase in MAPK activity after IGF-I treatment in rat SMCs (Thommes et al 1996).…”

Kinases: positive and negative regulators of apoptosis

“…44,45,[62][63][64][65][66] Ras is capable of activating both Raf [67][68][69] and Mekk; 70 however, activation of the Erk pathway can also occur in the absence of Ras activation. 71 The pathways leading to Erk activation are reported to be triggered by receptor tyrosine kinases, 72,73 G-protein linked receptors, [74][75][76] PKC, 71,77,78 and calcium. 46,[79][80][81][82] Several lines of evidence suggest that the Erk pathway has an important role in modulating the survival of hematopoietic cells.…”