Abstract:
Objectives: In order to develop the technology for porous albumin stents to serve as blood vessels we tested the ability of albumin substratum to support the adhesion and growth of vascular endothelial cells (VEC). Our specific aims were to find the appropriate albumin source and type and further improve the albumin substratum by enriching it with various plasma and extracellular components.
Methods: The albumin substratum was prepared by applying 0.2 ml of a sterile albumin solution at a concentration of 27 mg/ml into a polystyrene well (16 mm in diameter). The albumin layer was air dried for 48 h at room temperature and then heated (850C for 60 min). VEC were plated (1X105 cells/well) on the albumin surface and the rate of adhesion was measured after 24 h by washing the non-adherent cells followed by typinization and counting of the adhered cells. The growth of the cells was determined by further culturing of the adhered cells for 48 h followed by tripsinization and cell counting.
Results: Adhesion and growth of VEC on six different albumin types from two sources and various degrees of purity were studied. Our results demonstrated that a fatty acid and ?-globulin free albumin preparation (Sigma Cat. # A7030) was up to 48% better than the other preparations in supporting VEC adhesion and growth. In order to further improve the VEC adhesion and growth on albumin surfaces, we added to the albumin substratum various extracellular and plasma components, i.e. fetal calf serum, bovine pepsinized type I collagen, fibronectin and high density lipoprotein (HDL). The fetal calf serum at high concentrations (more than 50%) was toxic to VEC but at low concentrations (1.5%- 2.5%) slightly supported both cell attachment and growth (up to 18% increase). Fibronectin at the concentration range of 5-50 µg/ml did not improved cell adhesion and growth. HDL (50-500 µg/ml) slightly improved cell adhesion and growth (up to 17%), but the most significant increase in cell number (up to 70%) was achieved by collagen at the concentration of 25 µg/ml.
Conclusions: Albumin was found to be a compatible substratum to VEC adhesion and growth. The best albumin preparation supporting VEC adhesion and growth was found to be a fatty acid and ?-globulin free albumin preparation (Sigma Cat. # A7030). The adhesion and growth of VEC was further improved by including bovine pepsinized type I collagen (25 ?g/ml) in the albumin substratum. Further in-vivo studies in a wound healing model to test the capacity of the specific albumin and collagen composition proposed in this study are in progress.
