title: Vascular smooth muscle cells orchestrate the assembly of type I collagen via alpha2beta1 integrin, RhoA, and fibronectin polymerization.
abstract: Assembly of collagen into fibrils is widely studied as a spontaneous and entropy-driven process. To determine whether vascular smooth muscle cells (SMCs) impact the formation of collagen fibrils, we microscopically tracked the conversion of soluble to insoluble collagen in human SMC cultures, using fluorescent type I collagen at concentrations less than that which supported self-assembly. Collagen microaggregates were found to form on the cell surface, initially as punctate collections and then as an increasingly intricate network of fibrils. These fibrils displayed 67-nm periodicity and were found in membrane-delimited cellular invaginations. Fibril assembly was inhibited by an anti-alpha2beta1 integrin antibody and accelerated by an alpha2beta1 integrin antibody that stimulates a high-affinity binding state. Newly assembled collagen fibrils were also found to co-localize with newly assembled fibronectin fibrils. Moreover, inhibition of fibronectin assembly with an anti-alpha5beta1 integrin antibody completely inhibited collagen assembly. Collagen fibril formation was also linked to the cytoskeleton. Fibrils formed on the stretched tails of SMCs, ran parallel to actin microfilament bundles, and formed poorly on SMCs transduced with retrovirus containing cDNA for dominant-negative RhoA and robustly on SMCs expressing constitutively active RhoA. Lysophosphatidic acid, which activates RhoA and stimulates fibronectin assembly, stimulated collagen fibril formation, establishing for the first time that collagen polymerization can be regulated by soluble agonists of cell function. Thus, collagen fibril formation is under close cellular control and is dynamically integrated with fibronectin assembly, opening new possibilities for modifying collagen deposition.
authors.affiliation: Robarts Research Institute (Vascular Biology Group) and London Health Science Centre, 339 Windermere Road, London, Ontario, Canada N6A 5A5.
authors.forename: Shaohua
authors.initials: S
authors.lastname: Li
authors.forename: Caroline
authors.initials: C
authors.lastname: Van Den Diepstraten
authors.forename: Sudhir J
authors.initials: SJ
authors.lastname: D'Souza
authors.forename: Bosco M C
authors.initials: BM
authors.lastname: Chan
authors.forename: J Geoffrey
authors.initials: JG
authors.lastname: Pickering
mesh_terms: D000199:Actins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D024042:Collagen Type I / Q000096:biosynthesis* / Q000737:chemistry; D003599:Cytoskeleton / Q000502:physiology; D024022:Fibrillar Collagens / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D038982:Integrin alpha2beta1 / Q000378:metabolism*; D008246:Lysophospholipids / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000502:physiology*; D011108:Polymers / Q000378:metabolism; D012995:Solubility; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
chemical_list: D000199:Actins; D024042:Collagen Type I; D024022:Fibrillar Collagens; D005353:Fibronectins; D038982:Integrin alpha2beta1; D008246:Lysophospholipids; D011108:Polymers; D020742:rhoA GTP-Binding Protein