A new technique for measuring the cell growth and metabolism of endothelial cells seeded on vascular prostheses.
For the improvement of vascular graft patency, an endothelial cell (EC) lining is desirable. It is essential that the EC remains viable after being seeded onto the prosthetic graft. The aim of this study was to adapt an Alamar redox assay (ABRA) as a technique to monitor the viability of ECs seeded on prosthetic grafts. To test the graft types, we seeded human umbilical vein ECs on compliant polyurethane (CPU), expanded polytetrafluoroethylene, and Dacron at a density of 2 x 10(5) cell/cm(2). After 24 h of incubation, ABRA was added, and the absorbance was measured at 4, 8, and 24 h. To assess seeded cell concentrations on grafts, we seeded CPU at densities ranging from 1 x 10(5) to 8 x 10(5) cell/cm(2). The validity of the test was assessed with sodium azide and mitomycin C, known physiological perturbators. ABRA reduction demonstrated that ECs were viable and functional postseeding on the prosthetic grafts. A significant correlation was observed with ABRA reduction and cell concentrations (p < 0.001). The acid phosphatase assay demonstrated enzyme activity in the cells, but they were not maintained under normal physiological conditions. The ABRA bioreduced product was soluble, stable, and noncytotoxic over 24 h. The assay is independent of the geometry or physiochemistry of the graft type. The technique allows the continuous assessment of the metabolism and viability of seeded cells, is simple to perform, and does not destroy the cells or graft materials.
Seifalian
A M
AM
Vascular Haemodynamic Laboratory, University Department of Surgery, Royal Free and University College Medical School, University College London and the Royal Free Hospital, London, England. a.seifalian@rfc.ucl.ac.uk
Salacinski
H J
HJ
Punshon
G
G
Krijgsman
B
B
Hamilton
G
G
D001672:Biocompatible Materials; D001705:Bioprosthesis; D001808:Blood Vessels; D002455:Cell Division; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D006801:Humans; D010084:Oxidation-Reduction; D011108:Polymers
D001672:Biocompatible Materials; D011108:Polymers