Objective-Vascular smooth muscle cell (VSMC) proliferation is an important component of atherosclerosis, restenosis after angioplasty and stent placement, and vein graft failure. Outside-in signaling from the cadherin:␤-catenin complex can increase transcription of the cell-cycle gene cyclin D1; however, its role in VSMC proliferation has only recently been considered. Methods and Results-We examined the involvement of R-cadherin and ␤-catenin in VSMC proliferation in ballooninjured carotid arteries in vivo and aortic rings in vitro. The number of medial VSMCs positive for R-cadherin was significantly reduced by 32%Ϯ5%, 52%Ϯ10%, and 23%Ϯ2% at 0.25, 24, and 48 hours after injury in vivo, respectively. These changes in cadherin expression coincided with the detection of nuclear ␤-catenin and elevated cyclin D1 expression. Furthermore, loss of R-cadherin expression was associated with medial VSMC proliferation. Inhibition of classical cadherin function with a HAV peptide and R-cadherin neutralizing antibodies significantly increased proliferation by 4.3Ϯ1.0-fold and 4.1Ϯ0.98-fold, and increased the number of cells with ␤-catenin in the nucleus and expressing cyclin D1 in aortic rings. Conclusions-These results suggest that R-cadherin expression and ␤-catenin signaling may be associated with increased cyclin D1 expression and VSMC proliferation and may therefore play an important role in vascular disease. Key Words: smooth muscle Ⅲ cadherin Ⅲ proliferation Ⅲ intimal thickening V ascular smooth muscle cell (VSMC) proliferation plays a key role in pathological processes characterized by neointimal thickening, such as atherosclerosis, vascular rejection, and restenosis after angioplasty and stent placement. 1,2 Balloon injury of arteries causes quiescent, medial cells to lose their differentiated, contractile phenotype and proliferate for up to 3 days. [3][4][5] After this first wave of medial VSMC proliferation, medial VSMCs migrate and appear in the intima 4 days after injury. A new proliferative wave is then observed in the intima, which peaks at Ϸ7 days. Increased synthesis of extracellular matrix components continues for at least 4 weeks after injury. 6 The cadherins are a family of transmembrane glycoproteins that mediate calcium-dependent homophilic cell-cell interaction. 7 One cell type can express multiple cadherins, and the expression pattern is cell type-specific. 8 The extracellular domain of cadherins promotes the cell-cell adhesion through a binding site, which contains a HAV motif in the classical type I cadherins. 9 The cytoplasmic region connects cadherins to the cytoskeletal components through the catenin proteins. In addition to serving a structural function by linking to the actin cytoskeleton, the classical cadherins act as signaling receptors that affect cell behavior, including cell proliferation, migration, and differentiation. 10,11 In the absence of Wnt signals, ␤-catenin is either coupled to cadherins and the cytoskeleton at the plasma membrane or targeted for proteosomal degradation by the a...

ObjectivesInhibition of vascular smooth muscle cell (VSMC) migration is a potential strategy for reducing intimal thickening during in-stent restenosis and vein graft failure. In this study, we examined the effect of disrupting the function of the VSMC adhesion molecule, N-cadherin, using antagonists, neutralizing antibodies, and a dominant negative, on VSMC migration and intimal thickening. Migration was assessed by the scratch-wound assay of human saphenous vein VSMCs and in a human saphenous vein ex vivo organ culture model of intimal thickening.ResultsInhibition of cadherin function using a pan-cadherin antagonist, significantly reduced migration by 53% ± 8% compared with the control peptide (n = 3; P < .05). Furthermore, inhibition of N-cadherin function with an N-cadherin antagonist, neutralizing antibodies, and adenoviral expression of dominant negative N-cadherin (RAd dn-N-cadherin), significantly reduced migration by 31% ± 2%, 23% ± 1% and 32% ± 7% compared with controls, respectively (n = 3; P < .05). Inhibition of cadherin function significantly increased apoptosis by between 1.5- and 3.3-fold at the wound edge. In an ex vivo model of intimal thickening, inhibition of N-cadherin function by infection of human saphenous vein segments with RAd dn-N-cadherin significantly reduced VSMC migration by 55% and increased VSMC apoptosis by 2.7-fold. As a result, intimal thickening was significantly suppressed by 54% ± 14%. Importantly, there was no detrimental effect of dn-N-cadherin on endothelial coverage; in fact, it was significantly increased, as was survival of cultured human saphenous vein endothelial cells.ConclusionsUnder the condition of this study, cell-cell adhesion mediated by N-cadherin regulates VSMC migration via modulation of viability. Interestingly, inhibition of N-cadherin function significantly retards intimal thickening via inhibition of VSMC migration and promotion of endothelial cell survival. We suggest that disruption of N-cadherin-mediated cell-cell contacts is a potential strategy for reducing VSMC migration and intimal thickening.

Objective-Vascular smooth muscle cell (VSMC) apoptosis is thought to contribute to atherosclerotic plaque instability.Cadherin mediates calcium-dependent homophilic cell-cell contact. We studied the role of N-cadherin in VSMC apoptosis. Methods and Results-Human saphenous vein VSMCs were grown in agarose-coated wells to allow cadherin-mediated aggregate formation. Cell death and apoptosis were determined after disruption of cadherins using several approaches (nՆ3 per approach). Calcium removal from culture medium or addition of nonspecific cadherin antagonist peptides significantly decreased aggregate formation and increased cell death by apoptosis (34Ϯ6% versus 75Ϯ1% and 19Ϯ1% versus 40Ϯ5%, respectively; PϽ0.05). Specific inhibition of N-cadherin using antagonists and neutralizing antibodies similarly increased apoptosis. Supporting this, overexpression of full-length N-cadherin significantly reduced VSMC apoptosis from 44Ϯ10% to 20Ϯ3% (PϽ0.05), whereas abolishing N-cadherin expression by overexpression of a dominant-negative N-cadherin significantly, even in the presence of cell-matrix contacts, increased apoptosis from 9Ϯ2% to 50Ϯ1% (PϽ0.05). Interestingly, cell-cell contacts provided a similar degree of protection from apoptosis to cell-matrix contacts. Finally, N-cadherin-mediated cell-cell contacts initiated anti-apoptotic signaling by increasing Akt and Bad phosphorylation. Key Words: apoptosis Ⅲ atherosclerosis Ⅲ N-cadherin Ⅲ smooth muscle A therosclerotic plaques that have ruptured have thin fibrous caps that lack vascular smooth muscle cells (VSMCs). Stable plaques, by contrast, are rich in fibrous tissue and VSMCs. Thinning of the fibrous cap and loss of VSMCs in the late stages of atherosclerosis could therefore contribute to plaque instability. 1 VSMCs are thought to contribute to plaque stability by producing the extracellular matrix that in conjunction with the VSMCs provides the fibrous cap tensile strength and encapsulates the lipid core. 2 VSMC apoptosis is considered an important underlying mechanism that leads to reduced VSMC numbers and extracellular matrix in advanced atherosclerotic plaques, hence the thinning of the fibrous cap and plaque instability. 3 In fact, a recent study in which induction of VSMC apoptosis in the fibrous cap of mouse lesions caused plaque rupture and thrombosis provides direct evidence for the involvement of VSMC apoptosis and plaque instability. 4 Although abundant evidence indicates that VSMC apoptosis occurs in atherosclerosis, the mechanisms that initiate and regulate apoptosis are not fully understood. The presence of inflammatory cells, cytokines, modified low-density lipoprotein cholesterol, reactive oxygen species, and the systemic effects of altered blood pressure and flow are thought to promote apoptosis in atherosclerotic plaques. 5 However, apoptosis of VSMCs appears to be a counterbalance of pro-apoptotic factors such as fas or tumor necrosis factor, and anti-apoptotic factors, known as survival factors. It has been proposed that all cells are pro...