title: Smooth muscle cell rigidity and extracellular matrix organization influence endothelial cell spreading and adhesion formation in coculture.
abstract: Efforts to develop functional tissue-engineered blood vessels have focused on improving the strength and mechanical properties of the vessel wall, while the functional status of the endothelium within these vessels has received less attention. Endothelial cell (EC) function is influenced by interactions between its basal surface and the underlying extracellular matrix. In this study, we utilized a coculture model of a tissue-engineered blood vessel to evaluate EC attachment, spreading, and adhesion formation to the extracellular matrix on the surface of quiescent smooth muscle cells (SMCs). ECs attached to and spread on SMCs primarily through the alpha(5)beta(1)-integrin complex, whereas ECs used either alpha(5)beta(1)- or alpha(v)beta(3)-integrin to spread on fibronectin (FN) adsorbed to plastic. ECs in coculture lacked focal adhesions, but EC alpha(5)beta(1)-integrin bound to fibrillar FN on the SMC surface, promoting rapid fibrillar adhesion formation. As assessed by both Western blot analysis and quantitative real-time RT-PCR, coculture suppressed the expression of focal adhesion proteins and mRNA, whereas tensin protein and mRNA expression were elevated. When attached to polyacrylamide gels with similar elastic moduli as SMCs, focal adhesion formation and the rate of cell spreading increased relative to ECs in coculture. Thus, the elastic properties are only one factor contributing to EC spreading and focal adhesion formation in coculture. The results suggest that the softness of the SMCs and the fibrillar organization of FN inhibit focal adhesions and reduce cell spreading while promoting fibrillar adhesion formation. These changes in the type of adhesions may alter EC signaling pathways in tissue-engineered blood vessels.
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708-0281, USA.
authors.forename: Charles S
authors.initials: CS
authors.lastname: Wallace
authors.forename: Sophie A
authors.initials: SA
authors.lastname: Strike
authors.forename: George A
authors.initials: GA
authors.lastname: Truskey
mesh_terms: D000906:Antibodies / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology*; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D048430:Cell Shape / Q000502:physiology; D018920:Coculture Techniques; D004730:Endothelium, Vascular / Q000166:cytology* / Q000502:physiology; D005109:Extracellular Matrix / Q000502:physiology*; D005353:Fibronectins / Q000502:physiology; D006801:Humans; D039081:Integrin alpha5beta1 / Q000037:antagonists & inhibitors / Q000276:immunology / Q000502:physiology; D039302:Integrin alphaVbeta3 / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000502:physiology; D023822:Tissue Engineering
chemical_list: D000906:Antibodies; D005353:Fibronectins; D039081:Integrin alpha5beta1; D039302:Integrin alphaVbeta3