Downregulation of N-cadherin in the neointima stimulates migration of smooth muscle cells by RhoA deactivation.
OBJECTIVE
The aim of the study was to analyze whether cadherin- and Rho-family GTPases-mediated dynamic rearrangement of cell-cell adhesion play an important role during human arterial smooth muscle cell (haSMC) migration.


METHODS
Expression patterns of N-cadherin and beta-catenin were analyzed in a domestic pig restenosis model after 14, 28, and 90 days as well as in quiescent and migratory haSMCs in vitro. N-cadherin expression was upregulated by transient sense; downregulation was induced by antisense transfection. For functional inhibition, antibody GC-4 was used. Cell migration was quantified using Boyden chamber assays. Regulation of RhoA GTPase was tested by assessment of RhoA activity.


RESULTS
In vivo analysis of N-cadherin expression in a porcine restenosis model revealed downregulation in the neointima after 14 days. After 28 days, N-cadherin expression was slightly restored, while after 90 days, no difference between medial and neointimal expression was detectable. beta-Catenin levels remained unchanged during the whole period. According to the in vivo situation, N-cadherin was significantly downregulated in migratory haSMCs compared to quiescent cells in vitro. After N-cadherin overexpression, haSMC migration was reduced by 87% (P<0.001). By contrast, inhibition of N-cadherin in quiescent haSMCs by GC-4 increased the migratory potential by 87% (P<0.01). In haSMCs overexpressing N-cadherin, a significant upregulation of RhoA activity was demonstrated, while RhoA activity was blocked by GC-4.


CONCLUSIONS
These results indicate that the regulation of haSMC attachment by N-cadherins is essential for haSMC migration. Modification of N-cadherin expression and activity induces RhoA signaling with relevance for the reorganization of the actin cytoskeleton.
Blindt
Rüdiger
R
Medical Clinic I, University Hospital Aachen, Pauwelsstr 30, 52074 Aachen, Germany. ruediger.blindt@post.rwth-aachen.de
Bosserhoff
Anja-Katrin
AK
Dammers
Julia
J
Krott
Nicole
N
Demircan
Lütfü
L
Hoffmann
Rainer
R
Hanrath
Peter
P
Weber
Christian
C
Vogt
Felix
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015820:Cadherins / Q000032:analysis* / Q000235:genetics; D002465:Cell Movement; D023903:Coronary Restenosis / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D023421:Models, Animal; D009130:Muscle, Smooth / Q000378:metabolism* / Q000473:pathology; D009389:Neovascularization, Pathologic; D016376:Oligonucleotides, Antisense / Q000235:genetics; D013552:Swine; D015534:Trans-Activators / Q000378:metabolism; D014162:Transfection / Q000379:methods; D017539:Tunica Intima / Q000378:metabolism*; D051176:beta Catenin; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D000911:Antibodies, Monoclonal; C495270:CTNNB1 protein, human; D015820:Cadherins; D003598:Cytoskeletal Proteins; D016376:Oligonucleotides, Antisense; D015534:Trans-Activators; D051176:beta Catenin; D020742:rhoA GTP-Binding Protein