Abstract 1359: Only the Catalytic p110alpha Isoform Mediates Class IA PI 3-Kinase-Induced Atherogenic Signals in Vascular Smooth Muscle Cells

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Marius Vantler ◽  
Lenard Mustafov ◽  
Evren Caglayan ◽  
Stephan Rosenkranz
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Dna Synthesis ◽  
Vascular Smooth Muscle Cells ◽  
Fold Increase ◽  
And Migration ◽  
Induced Apoptosis ◽  
Stimulation Of

Proliferation, migration, and apoptosis of vascular smooth muscle cells (VSMC) are pivotal determinants of the pathogenesis of vascular diseases, which are mainly controlled by growth factor dependent activation of PI 3-Kinase (PI3K). Growth factors like platelet-derived growth factor (PDGF) activate class IA PI3Ks containing one of three p110 catalytic subunits (p110alpha, p110beta, and p110delta). We investigated the specific function of these isoforms for PDGF-controlled proliferation, migration, and apoptosis of VSMC using novel isoform-specific inhibitors. PDGF-dependent proliferation and migration solely depended on p110alpha. Stimulation of VSMC with PDGF-BB (50 ng/ml) mediated a 2.5±0.4 increase ( p <0.05) of DNA-synthesis (BrdU incorporation assay) and induced a 3.4+/−0.7 fold increase ( p <0.05) of VSMC migration (modified Boyden-chamber). Inhibition of p110alpha with PIK075 (1 μ M, Ki=100 nM) completely abrogated PDGF-dependent DNA-synthesis and migration ( p <0,05), whereas inhibitors against p110beta (TGX 221, 1 μ M) or p110delta (IC87114 1 μ M) had no influence. Consistently, PDGF-induced DNA-synthesis and migration were suppressed by siRNA-dependent downregulation of p110alpha ( p <0,05) whereas p110beta or p110delta knockdown had no effect. Interestingly, stimulation of VSMC with PDGF-BB (50 ng/ml) induced anti- or proapoptotic effects depending on the duration of PDGFR activation. Incubation of VSMC with H 2 O 2 (50 μ M, 16h) led to a 2.8±0.7 fold increase ( p >0.05) of apoptosis (Cell Death Detection ELISA). Simultanous addition of PDGF-BB (50 ng/ml) significantly diminished the H 2 O 2 -induced apoptosis (52±7%, p >0.05). In contrast, prestimulation with PDGF-BB 24h prior to the addition of H 2 O 2 led to an increase of H 2 O 2 -induced apoptosis (7.8±1.3, p >0.05). The anti- as well as the proapoptotic effect depended strictly on p110alpha as PIK075 (1 μ M, p <0,05) or p110alpha specific siRNA completely abrogated PDGF-BB-mediated pro- as well as antiapoptotic effects. Our results demonstrate that only the catalytical PI3K subunit p110alpha mediates the growth factor-induced atherogenic responses. Therefore, p110alpha represents an interesting therapeutic target for prevention of atherosclerosis and restenosis formation.

Epidermal growth factor-mediated effects on equine vascular smooth muscle cells

1988 ◽  
Vol 255 (4) ◽  
pp. C447-C451 ◽  
Author(s):  
D. A. Grosenbaugh ◽  
M. S. Amoss ◽  
D. M. Hood ◽  
S. J. Morgan ◽  
J. D. Williams
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Dna Synthesis ◽  
Vascular Smooth Muscle Cells ◽  
Cellular Proliferation ◽  
Epidermal Growth ◽  
Dose Dependent

Epidermal growth factor (EGF) receptor binding kinetics and EGF-mediated stimulation of DNA synthesis and cellular proliferation were studied in cultured vascular smooth muscle cells (VSMC) from the equine thoracic aorta. Binding studies, using murine 125I-labeled EGF, indicate the presence of a single class of high-affinity binding sites (apparent KD = 2.8 X 10(-11) M), with an estimated maximal binding capacity of 5,800 sites/cell. EGF stimulated [3H]thymidine uptake in confluent quiescent monolayers in a dose-dependent fashion, half-maximal stimulation occurring at 7.5 X 10(-11) M. Likewise, EGF-mediated cellular proliferation was dose dependent (50% effective dose = 5 X 10(-11) M) under reduced serum concentrations. Equine VSMC contain specific receptors for EGF, and EGF can stimulate DNA synthesis and proliferation in these cultured cells, which suggests that EGF may participate in the proliferative changes observed in equine distal digital peripheral vascular disease.

Sphingolipids as mediators of effects of platelet-derived growth factor in vascular smooth muscle cells

1993 ◽  
Vol 265 (3) ◽  
pp. C740-C747 ◽  
Author(s):  
L. S. Jacobs ◽  
M. Kester
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Dna Synthesis ◽  
Vascular Smooth Muscle Cells ◽  
Platelet Derived Growth Factor ◽  
Sphingolipid Metabolism ◽  
A7r5 Cell ◽  
A7r5 Cells

The role of sphingolipids in mediating the action of platelet-derived growth factor (PDGF) has been investigated in the vascular smooth muscle-derived A7r5 cell line. L-Cycloserine (2 mM), an inhibitor of sphingolipid synthesis, caused time-dependent inhibition of [3H]serine incorporation into [3H]sphingomyelin in A7r5 cells. PDGF-AB (10 ng/ml), PDGF-BB (10 ng/ml), or sphingosine (10 microM) independently stimulated [3H]thymidine incorporation into DNA in A7r5 cells. L-Cycloserine (2 mM) inhibited stimulation of DNA synthesis by both PDGF-AB and PDGF-BB. L-Cycloserine (2 mM, 16 h) did not affect the ability of PDGF or sphingosine to increase intracellular free calcium ([Ca2+]i) in A7r5 cells loaded with the fluorescent indicator fura 2. Measurement of adenine nucleotide levels in A7r5 cell extracts by reverse-phase high-performance liquid chromatography indicated that treatment with L-cycloserine did not adversely affect cellular metabolism. To determine directly whether PDGF activates sphingolipid metabolism, A7r5 cells were labeled with [3H]serine for 48 h and then treated with PDGF-AB (10 ng/ml) for 1 h. Sphingolipids were separated by thin-layer chromatography and quantified by liquid scintillation counting. PDGF-AB stimulated an increase in [3H]sphingosine from 25.5 +/- 3.0 to 37.5 +/- 4.1 counts.min-1 (cpm).micrograms protein-1 and a concomitant decrease in [3H]ceramide from 24.3 +/- 3.2 to 18.5 +/- 2.9 cpm/micrograms protein. These data suggest that the PDGF-stimulated increase in [Ca2+]i is not sufficient for induction of DNA synthesis and that mitogenic effects of PDGF in vascular smooth muscle cells are mediated by sphingolipid metabolism.

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