Abstract 501: The Platelet-Derived Growth Factor β Receptor Mediates Anti- as well as Pro-Apoptotic Signals in Vascular Smooth Muscle Cells via PI 3 Kinase Dependent Pathways
VSMC apoptosis plays a critical role in the development of atherosclerosis. Intimal VSMC apoptosis leads to plaque destabilization, whereas VSMC survival promotes neointimal growth. The PDGFβR activates cytoplasmic signaling molecules such as Src, PI3K, RasGAP, SHP-2 and PLCγ and mediates survival signals in many cell types. We characterized the role of each signaling molecule for VSMC apoptosis. PDGF-BB induced anti- as well as proapoptotic effects in VSMC depending on the stimulation time. Incubation of VSMC with H2O2 (100 μM, 12 h) increased apoptosis to 5.3±1.1-fold (p<<med>0.05). PDGF-BB (50 ng/ml) diminished H2O2-induced apoptosis (52±7%, p<<med>0.05). In contrast, chronic stimulation (96 h) with PDGF-BB augmented apoptosis to 3.1±0.4-fold (p<<med>0.05). To characterize the anti- and pro-apoptotic pathways, we used mutated βPDGFR in which the binding sites for the signaling molecules were individually deleted. To bypass endogenous PDGFR in VSMC we used chimeric receptors containing the extracellular ligand binding domain of the M-CSF receptor. Short term activation of the chimeric wild type receptor (WT) with M-CSF (50 ng/ml) reduced H2O2-induced apoptosis to the same extend as the endogenous PDGFR. Deletion of the PI3K binding site abolished the anti-apoptotic effect, whereas deletion of the binding sites for Src, RasGAP, SHP-2 or PLCγ had no influence. Exploration of downstream signaling events revealed that PDGF-BB transiently activates the antiapoptotic Akt signaling pathway in a PI3K-dependent manner, leading to the phosphorylation of BAD and Forkhead transcription factors. Chronic stimulation of WT-expressing VSMC with M-CSF (50 ng/ml, 96 h) increased apoptosis to 2.8±0.6-fold (p<<med>0.05), correlating with a decreased expression of the antiapoptotic protein Bcl-2 (Real Time PCR and Westernblot analysis). Deletion of the PI3K binding site abolished the pro-apoptotic effect, whereas a mutant which only binds PI3K was able to mediate the proapoptotic effect as WT. In summary, short-time activation of the βPDGFR induces cell survival by transient activation of the Akt signaling pathway, whereas chronic stimulation leads to increased cell death by the regulation of genes such as Bcl-2. Both effects are mediated by PI3K-dependent signaling pathways.