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TGF-?/Smad3 inhibit vascular smooth muscle cell apoptosis through an autocrine signaling mechanism involving VEGF-A.

ABSTRACT: We have previously shown that in the presence of elevated Smad3, transforming growth factor-? (TGF-?) transforms from an inhibitor to a stimulant of vascular smooth muscle cell (SMC) proliferation and intimal hyperplasia (IH). Here we identify a novel mechanism through which TGF-?/Smad3 also exacerbates IH by inhibiting SMC apoptosis. We found that TGF-? treatment led to inhibition of apoptosis in rat SMCs following viral expression of Smad3. Conditioned media from these cells when applied to naive SMCs recapitulated this effect, suggesting an autocrine pathway through a secreted factor. Gene array of TGF-?/Smad3-treated cells revealed enhanced expression of vascular endothelial growth factor (VEGF), a known inhibitor of endothelial cell apoptosis. We then evaluated whether VEGF is the secreted mediator responsible for TGF-?/Smad3 inhibition of SMC apoptosis. In TGF-?/Smad3-treated cells, VEGF mRNA and protein as well as VEGF secretion were increased. Moreover, recombinant VEGF-A inhibited SMC apoptosis and a VEGF-A-neutralizing antibody reversed the inhibitory effect of conditioned media on SMC apoptosis. Stimulation of SMCs with TGF-? led to the formation of a complex of Smad3 and hypoxia-inducible factor-1? (HIF-1?) that in turn activated the VEGF-A promoter and transcription. In rat carotid arteries following arterial injury, Smad3 and VEGF-A expression were upregulated. Moreover, Smad3 gene transfer further enhanced VEGF expression as well as inhibited SMC apoptosis. Finally, blocking either the VEGF receptor or Smad3 signaling in injured carotid arteries abrogated the inhibitory effect of Smad3 on vascular SMC apoptosis. Taken together, our study reveals that following angioplasty, elevation of both TGF-? and Smad3 leads to SMC secretion of VEGF-A that functions as an autocrine inhibitor of SMC apoptosis. This novel pathway provides further insights into the role of TGF-? in the development of IH.

SUBMITTER: Shi X

PROVIDER: S-EPMC4123076 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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TGF-β/Smad3 inhibit vascular smooth muscle cell apoptosis through an autocrine signaling mechanism involving VEGF-A.

Shi X X Guo L-W LW Seedial S M SM Si Y Y Wang B B Takayama T T Suwanabol P A PA Ghosh S S DiRenzo D D Liu B B Kent K C KC

Cell death & disease 20140710

We have previously shown that in the presence of elevated Smad3, transforming growth factor-β (TGF-β) transforms from an inhibitor to a stimulant of vascular smooth muscle cell (SMC) proliferation and intimal hyperplasia (IH). Here we identify a novel mechanism through which TGF-β/Smad3 also exacerbates IH by inhibiting SMC apoptosis. We found that TGF-β treatment led to inhibition of apoptosis in rat SMCs following viral expression of Smad3. Conditioned media from these cells when applied to na ...[more]

PMID: 25010983

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Project description:Elastin is predominantly comprised of crosslinked tropoelastin. For many years elastin was considered to serve a solely structural role but is now being increasingly identified as causal in cell signaling, development and repair. We introduced tropoelastin into an in vitro model in which airway smooth muscle cells (ASMCs) were stimulated with transforming growth factor (TGF)-?1 to examine the modulatory effect of this modular elastin sequence on release of angiogenic factors and matrix metalloproteinases (MMPs). Human ASMCs were presented to surfaces coated with tropoelastin or collagen and controls, then stimulated with TGF-?1. Transcript levels of vascular endothelial growth factor (VEGF) and connective tissue growth factor (CTGF) were quantified 4 and 24 h after TGF-?1 stimulation. Protein VEGF release from cells and CTGF sequestered at cell surfaces were measured by ELISA at 24 and 48 h. TGF-?1 increased VEGF mRNA 2.4 fold at 4 h and 5 fold at 24 h, accompanied by elevated cognate protein release 3 fold at 24 h and 2.5 fold at 48 h. TGF-?1 stimulation increased CTGF mRNA 6.9 fold at 4 h and 11.8 fold at 24 h, accompanied by increased sequestering of its protein counterpart 1.2 fold at 24 h and 1.4 fold at 48 h. Pre-incubation of cells with tropoelastin did not modulate VEGF or CTGF mRNA expression, but combined with TGF-?1 stimulation it led to enhanced VEGF release 5.1-fold at 24h and 4.4-fold at 48 h. Pre-incubation with tropoelastin decreased CTGF sequestering 0.6-fold at 24 and 48 h, and increased MMP-2 production. Collagen pre-incubation under the same conditions displayed no effect on TGF-?1 stimulation apart from a slightly decreased (0.9 fold) sequestered CTGF at 48 h. As CTGF is known to anchor VEGF to the matrix and inhibit its angiogenic activity, a process which can be reversed by digestion with MMP-2, these findings reveal that elastin sequences can disrupt the balance of angiogenic factors, with implications for aberrant angiogenesis. The results suggest a model of molecular crosstalk and support an active role for elastin in vascular remodeling.

Dataset Information

TGF-?/Smad3 inhibit vascular smooth muscle cell apoptosis through an autocrine signaling mechanism involving VEGF-A.

Publications

TGF-β/Smad3 inhibit vascular smooth muscle cell apoptosis through an autocrine signaling mechanism involving VEGF-A.

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