Alpha 8 integrin expression is required for maintenance of the smooth muscle cell differentiated phenotype.
OBJECTIVE
Vascular smooth muscle cell (VSMC) de-differentiation is a prerequisite for migration from the tunica media to the intima after vascular injury. Integrin cell adhesion molecules participate in VSMC phenotype modulation. Alpha 8 beta 1 integrin is a differentiation marker of VSMCs and its knockdown heightens migration. In the present study, we examined whether or not alpha 8 integrin is required for the maintenance of VSMC differentiated phenotype.


METHODS
Alpha 8 integrin in rat VSMC was knocked down by short interference RNA (siRNA) targeting alpha 8 integrin in comparison to a non-silencing siRNA. Cytoskeletal and morphological changes in VSMC were examined by immunofluorescence staining. The expression of phenotype-dependent markers was analyzed by immunoblotting.


RESULTS
Alpha 8 integrin gene silencing evoked drastic changes in characteristics of the VSMC differentiated phenotype, including VSMC morphology, actin fibre organization, focal adhesion assembly and the expression of phenotype-dependent markers in favor of de-differentiation. Then, we investigated whether or not phenotype modulation induced by alpha 8 integrin gene silencing could be reversed by an inducer of VSMC differentiation. Transforming growth factor-beta (TGF-beta) failed to upregulate smooth muscle-myosin heavy chain as well as the assembly of parallel actin fibres in VSMCs transfected by siRNA-alpha 8. In addition, TGF-beta-induced vinculin localization at the tip of the cells was impaired by alpha 8 integrin gene silencing.


CONCLUSIONS
These data suggest that alpha 8 integrin expression is required for maintenance of the VSMC differentiated phenotype, a state that is crucial for non-motile VSMCs.
Zargham
Ramin
R
Institut de recherches cliniques de Montréal, Université de Montréal and Department of Experimental Medicine, McGill University, 110, avenue des Pins ouest, Montréal, Québec, Canada H2W 1R7.
Thibault
Gaétan
G
D000199:Actins / Q000032:analysis / Q000378:metabolism; D000818:Animals; D015415:Biomarkers / Q000032:analysis; D015153:Blotting, Western / Q000379:methods; D002339:Carotid Arteries; D002454:Cell Differentiation; D002465:Cell Movement; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D018390:Gene Targeting; D039001:Integrin alpha Chains / Q000235:genetics / Q000378:metabolism*; D008297:Male; D018613:Microscopy, Confocal; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000032:analysis / Q000378:metabolism; D010641:Phenotype; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism / Q000494:pharmacology; D017539:Tunica Intima / Q000166:cytology / Q000378:metabolism; D016596:Vinculin / Q000032:analysis / Q000378:metabolism
D000199:Actins; D015415:Biomarkers; D039001:Integrin alpha Chains; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; C093856:integrin alpha8; D016596:Vinculin; D018995:Myosin Heavy Chains