Signaling Pathways Transduced through the Elastin Receptor Facilitate Proliferation of Arterial Smooth Muscle Cells

Abstract

In this report we demonstrate that soluble peptides, elastin degradation products stimulate proliferation of arterial smooth muscle cells. We show that these effects are due to generation of intracellular signals transduced through the cell surface elastin receptor, which consists of peripheral 67-kDa elastin-binding protein (EBP) (spliced variant of ␤-galactosidase), immobilized to the transmembrane sialidase and the protective protein. We found that elastin receptor-transduced signaling triggers activation of G proteins, opening of L-type calcium channels, and a sequential activation of tyrosine kinases: FAK, c-Src, platelet-derived growth factorreceptor kinase and then Ras-Raf-MEK1/2-ERK1/2 phosphorylation cascade. This, in turn, causes an increase in expression of cyclins and cyclin-dependent kinases, and a consequent increase in cellular proliferation. The EBP-transduced signals also induce tyrosine kinase-dependent phosphorylation of ␤-tubulin, LC3, microtubule-associated protein 1, and ␣-actin and troponin-T, which could be linked to reorganization of cytoskeleton. We have also disclosed that induction of these signals can be abolished by anti-EBP antibody or by galactosugars, which cause shedding of EBP from the cell surface. Moreover, elastin-derived peptides did not induce proliferation of EBP-deficient cells derived from patients bearing a nonsense mutation of the ␤-galactosidase gene or sialidase-deficient cells from patients with congenital sialidosis.It has been well established that formation of neointima in vascular diseases is associated with impaired assembly of tropoelastin into insoluble elastin (1-6) and with extensive degradation of the elastin-rich extracellular matrix by numerous proteinases leaking from the serum and secreted from the infiltrating platelets, leukocytes, and activated vascular cells (7-10). It has also been suggested that local accumulation of non-assembled tropoelastin and small elastin-derived peptides may constitute an important factor in the activation of the normally quiescent medial SMC 1 into the proliferative and migratory phenotype, which participates in the formation of the occlusive neointima in vascular diseases (2,3,(11)(12)(13)(14)(15) Elastin does not contain the RGD sequence and does not interact with cell surface integrins. Our previous studies demonstrated that numerous cell types, including vascular myocytes, express the cell surface elastin receptor complex, which consists of three subunits (25,26), and that the average cell contains ϳ2 ϫ 10 6 elastin binding sites with the binding affinity (K d ) of 8 nM (27). We found that two of those subunits (55-and 61-kDa) are anchored to the plasma membrane, whereas the third, a peripheral 67-kDa protein, actually binds elastin (25). This major functional component of the receptor complex was named the elastin binding protein (EBP). The repeat hexapeptide in tropoelastin, VGVAPG, has been identified as a chief ligand for high affinity binding to this cell surface receptor (25-27). It has been later established t...

Elastic fiber homeostasis requires lysyl oxidase–like 1 protein

“…The following antibodies to elastin, fibrillin-1 and fibrillin-2 were from Elastin Products Company: PR385 (rabbit antibody to mouse elastin, exons 6-17); PR387 (rabbit antibody to mouse elastin, exons 31-36); RT675 (goat antibody to rat elastin) used for double labeling of elastin with LOXL1, LOX or fibulin-5; PR210AP (rabbit antibody to fibrillin-1); PR225 (rabbit antibody to fibrillin-2). EBP antibody (EBP2112) 5 was a gift from A. Hinek (The Hospital for Sick Children, Toronto, Canada). We carried out immunoblotting and immunofluorescence staining of unfixed cryosections as described 27 .…”

“…Once laid down, they are thought to remain stable 2 , except in the uterine tract where cycles of active remodeling occur 3 . Loss of elastic fibers underlies connective tissue aging and important diseases including emphysema [4][5][6][7] . Failure to maintain elastic fibers is explained by a theory of antielastase-elastase imbalance 8 , but little is known about the role of renewal.…”

“…Our data also show an unexpected, causal link between elastic fiber homeostasis and pelvic prolapse, a clinical condition strongly correlated with both childbirth and advanced age [21][22][23] . Elastin polymer and soluble elastin-derived peptides also have signaling roles in cell adhesion, migration and proliferation 5,24 . Thus, the combination of loss of elastin polymer and accumulation of tropoelastin may have additionally contributed to the histopathologic changes seen in the LOXL1 mutant mice.…”

Interaction between the Elastin Peptide VGVAPG and Human Elastin Binding Protein

“…Elastin peptide concentration in the blood flow is usually low, but it can reach high values in aged subjects, notably in those developing pathologies where elastin is massively degraded such as aneurysms (15,16). Unlike insoluble fibrous elastin, elastin peptides promote cell proliferation, cell chemotaxis, angiogenesis, and proteases release in a wide variety of normal cells (13,(17)(18)(19). The effects promoted by VGVAPG, a sequence repeated six times in human tropoelastin, are the most documented.…”

Elastin Based Constructs

“…The most well documented of these receptors is EBP, which binds to multiple sites including the VGVAPG sequence on exon 24 of tropoelastin (Rodgers and Weiss 2005). Upon binding elastin, this receptor activates intracellular signaling pathways involved in cell proliferation, chemotaxis, migration and cell morphology for a range of cell types including SMCs, endothelial cells, fibroblasts, monocytes, leukocytes and mesenchymal cells (Senior, Griffin et al 1980;Indik, Abrams et al 1990;Faury, Ristori et al 1994;Faury, Ristori et al 1995;Kamisato, Uemura et al 1997;Jung, Rutka et al 1998;Mochizuki, Brassart et al 2002). Other cell receptors, including a less documented glycoprotein termed elastonectin and G protein-coupled receptor can bind elastin through the VGVAPG sequence (Hornebeck, Tixier et al 1986).…”