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Endothelial and smooth muscle-derived neuropilin-like protein regulates platelet-derived growth factor signaling in human vascular smooth muscle cells by modulating receptor ubiquitination.

ABSTRACT: Endothelial and smooth muscle cell-derived neuropilin-like protein (ESDN) is up-regulated in the neointima of remodeling arteries and modulates vascular smooth muscle cell (VSMC) growth. Platelet-derived growth factor (PDGF) is the prototypic growth factor for VSMCs and plays a key role in vascular remodeling. Here, we sought to further define ESDN function in primary human VSMCs. ESDN down-regulation by RNA interference significantly enhanced PDGF-induced VSMC DNA synthesis and migration. This was associated with increased ERK1/2, Src, and PDGF receptor (PDGFR)beta phosphorylation, without altering total PDGFRbeta expression levels. In binding assays, ESDN down-regulation significantly increased (125)I-PDGF maximum binding (B(max)) to PDGF receptors on VSMCs without altering the binding constant (K(d)), raising the possibility that ESDN regulates PDGFR processing. ESDN down-regulation significantly reduced ligand-induced PDGFRbeta ubiquitination. This was associated with a significant reduction in the expression level of c-Cbl, an E3 ubiquitin ligase that ubiquitinylates PDGFRbeta. Thus, ESDN modulates PDGF signaling in VSMCs via regulation of PDGFR surface levels. The ESDN effect is mediated, at least in part, through effects on PDGFRbeta ubiquitination. ESDN may serve as a target for regulating PDGFRbeta signaling in VSMCs.

SUBMITTER: Guo X

PROVIDER: S-EPMC2785569 | biostudies-other | 2009 Oct

REPOSITORIES: biostudies-other

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Endothelial and smooth muscle-derived neuropilin-like protein regulates platelet-derived growth factor signaling in human vascular smooth muscle cells by modulating receptor ubiquitination.

Guo Xiaojia X Nie Lei L Esmailzadeh Leila L Zhang Jiasheng J Bender Jeffrey R JR Sadeghi Mehran M MM

The Journal of biological chemistry 20090819 43

Endothelial and smooth muscle cell-derived neuropilin-like protein (ESDN) is up-regulated in the neointima of remodeling arteries and modulates vascular smooth muscle cell (VSMC) growth. Platelet-derived growth factor (PDGF) is the prototypic growth factor for VSMCs and plays a key role in vascular remodeling. Here, we sought to further define ESDN function in primary human VSMCs. ESDN down-regulation by RNA interference significantly enhanced PDGF-induced VSMC DNA synthesis and migration. This ...[more]

PMID: 19696027

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Project description:Platelet-derived growth factor-BB activates platelet-derived growth factor receptor-? and promotes vascular smooth muscle cell phenotypic transformation. Elevated levels of non-muscle myosin IIB (SMemb) are found in secretory smooth muscle cells along with inflammatory mediators, such as intercellular adhesion molecule-1, which can amplify neutrophil infiltration into the brain. In the present study, we investigated the role of platelet-derived growth factor-BB/platelet-derived growth factor receptor-? following intracerebral hemorrhage-induced brain injury in mice, with emphasis on its ability to promote vascular smooth muscle cell phenotypic transformation followed by increased intercellular adhesion molecule-1 expression and elevated neutrophil infiltration in the vicinity of the hematoma. We also determined the extent to which plasmin from the hematoma influences the platelet-derived growth factor-BB/platelet-derived growth factor receptor-? system subsequent to intracerebral hemorrhage.Controlled in vivo laboratory study.Animal research laboratory.One hundred and fifty six eight-week-old male CD1 mice.Brain injury was induced by autologous arterial blood or plasmin injection into mouse brains. Small interfering RNA targeting platelet-derived growth factor receptor-? was administered 24 hours before intracerebral hemorrhage. A platelet-derived growth factor receptor antagonist, Gleevec, was administered following intracerebral hemorrhage. A mitogen-activated protein kinase-activated protein kinase 2 inhibitor (KKKALNRQLGVAA) was delivered with platelet-derived growth factor-BB in naïve animals. Platelet-derived growth factor-BB was injected with a plasmin inhibitor (?-aminocaproic acid) in intracerebral hemorrhage mice. Plasmin-injected mice were given platelet-derived growth factor receptor-? small interfering RNA 24 hours before the operation. Neurological deficits, brain edema, western blots, and immunofluorescence were evaluated.Platelet-derived growth factor receptor-? small interfering RNA attenuated SMemb and intercellular adhesion molecule-1 expression and neutrophil infiltration at 24 hours post injury and reduced neurological deficits and brain edema at 24 and 72 hours following intracerebral hemorrhage. The platelet-derived growth factor receptor antagonist, Gleevec, reduced SMemb and intercellular adhesion molecule-1 expression. Platelet-derived growth factor receptor-? activation led to increased expression of intercellular adhesion molecule-1 and was reversed by KKKALNRQLGVAA in naïve mice. Plasmin inhibition suppressed platelet-derived growth factor receptor-? activation and neutrophil infiltration, whereas exogenous platelet-derived growth factor-BB increased platelet-derived growth factor receptor-? activation, regardless of plasmin inhibition. Platelet-derived growth factor receptor-? small interfering RNA decreased the expression of intercellular adhesion molecule-1 by plasmin injection.The platelet-derived growth factor-BB/platelet-derived growth factor receptor-? system contributes to neuroinflammation through vascular smooth muscle cell phenotypic transformation near the hematoma via the p38 mitogen-activated protein kinase/mitogen-activated protein kinase-activated protein kinase 2 pathway following intracerebral hemorrhage. Plasmin is hypothesized to be upstream of the proposed neuroinflammatory system. The therapeutic intervention targeting the platelet-derived growth factor-BB/platelet-derived growth factor receptor-? is a novel strategy to prevent plasmin-induced brain injury following intracerebral hemorrhage.

Project description:ObjectiveWe examined the role of syndecan-1 in modulating the phenotype of vascular smooth muscle cells in the context of endogenous inflammatory factors and altered microenvironments that occur in disease or injury-induced vascular remodeling.Methods and resultsVascular smooth muscle cells (vSMCs) display a continuum of phenotypes that can be altered during vascular remodeling. While the syndecans have emerged as powerful and complex regulators of cell function, their role in controlling vSMC phenotype is unknown. Here, we isolated vSMCs from wild type (WT) and syndecan-1 knockout (S1KO) mice. Gene expression and western blotting studies indicated decreased levels of α-smooth muscle actin (α-SMA), calponin, and other vSMC-specific differentiation markers in S1KO relative to WT cells. The spread area of the S1KO cells was found to be greater than WT cells, with a corresponding increase in focal adhesion formation, Src phosphorylation, and alterations in actin cytoskeletal arrangement. In addition, S1KO led to increased S6RP phosphorylation and decreased AKT and PKC-α phosphorylation. To examine whether these changes were present in vivo, isolated aortae from aged WT and S1KO mice were stained for calponin. Consistent with our in-vitro findings, the WT mice aortae stained higher for calponin relative to S1KO. When exposed to the inflammatory cytokine TNF-α, WT vSMCs had an 80% reduction in syndecan-1 expression. Further, with TNF-α, S1KO vSMCs produced increased pro-inflammatory cytokines relative to WT. Finally, inhibition of interactions between syndecan-1 and integrins αvβ3 and αvβ5 using the inhibitory peptide synstatin appeared to have similar effects on vSMCs as knocking out syndecan-1, with decreased expression of vSMC differentiation markers and increased expression of inflammatory cytokines, receptors, and osteopontin.ConclusionsTaken together, our results support that syndecan-1 promotes vSMC differentiation and quiescence. Thus, the presence of syndecan-1 would have a protective effect against vSMC dedifferentiation and this activity is linked to interactions with integrins αvβ3 and αvβ5.

Dataset Information

Endothelial and smooth muscle-derived neuropilin-like protein regulates platelet-derived growth factor signaling in human vascular smooth muscle cells by modulating receptor ubiquitination.

Publications

Endothelial and smooth muscle-derived neuropilin-like protein regulates platelet-derived growth factor signaling in human vascular smooth muscle cells by modulating receptor ubiquitination.

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