Vascular smooth muscle cell responses to 3D polyurethane scaffolds for tissue engineering applications.

摘要

One promising strategy in vascular tissue engineering is the design of hybrid vascular substitutes where vascular cells infiltrate biostable porous scaffolds, which provide a favourable environment for guided cell repopulation and act as a mechanically supporting layer after the tissue regeneration process. The aim of the present work was to characterize interactions of human coronary artery smooth muscle cells (HCASMC) with 3D porous biostable polyurethane scaffolds. To initially assess the feasibility of polyurethanes as potential materials for vascular tissue engineering applications, HCASMC and human coronary artery endothelial cells were cultured onto films, and results showed that polyurethanes supported the adhesion of vascular cells, the formation of endothelial monolayers and the maintenance of the differentiated smooth muscle cell phenotype. We therefore used a pressure differential/particulate leaching technique to fabricate 3D polyurethane scaffolds with pores generated from either paraffin or NR4Cl porogens. SEM and microCT studies of the fabricated scaffolds showed that the current scaffolds had highly open and interconnected pore structures, with an average porosity of 84%. In comparison to scaffolds fabricated from NR4Cl porogens, the use of paraffin porogens resulted in scaffolds with highly spherical pores, thinner struts, micropores between macropores, and higher surface area to volume ratio. Following uniaxial compression and extension tests, scaffolds fabricated from NR4Cl porogens had superior mechanical properties, which are suitable for vascular tissue engineering. HCASMC interactions on scaffolds revealed that cells adhered to the scaffolds and displayed an elongated morphology with parallel alignment. Surface modification with Matrigel or fibronectin promoted further cellular infiltration and proliferation. Deep in the scaffold, cells were encountered that formed actin-rich lamellipodial extensions spreading along the struts and lacked stress fibers, suggesting active cell migration. HCASMC started producing collagen as judged by histochemical analysis but appeared to lack elastin production. Western blot analyses following successful cell recovery from the scaffolds indicated that HCASMC, after culturing for 4 and 7 days, expressed similar amounts of smalpha-actin and calponin regardless of ECM coating. Taken together, our data showed that biostable polyurethanes can be used to fabricate porous and highly interconnected 3D scaffolds that promote HCASMC attachment, proliferation, migration, differentiation and collagen production.;Keywords: tissue engineering, polyurethane scaffolds, vascular grafts, mechanical properties, vascular smooth muscle cell, cell adhesion, cell proliferation, cell infiltration, marker protein expression, extracellular matrix production