Angiogenic response of endothelial cells seeded dispersed versus on beads in fibrin gels

Abstract

Induction of an inter-connected microvessel network in a tissue-engineered construct prior to implantation may be an alternative to improve the success rate of cell/tissue survival and wound integration. Conditions of endothelial cell-seeding density and distribution were investigated in two 3-D angiogenesis culture systems. Endothelial cells were either seeded dispersed in a fibrin gel, or subconfluent on micro-beads (Cytodex) prior to being embedded in fibrin. Human fibroblasts and growth factors were introduced to optimize angiogenesis. A density higher than 4 x 10(4) cells/ml of fibrin was necessary to induce angiogenic-like structures (i.e., sprouting, cord-, lumen-like structures) by 14 days in the dispersed cell model. Endothelial cells on micro-beads also exhibited angiogenic-like structures that were inter-connected to those on neighboring beads. The sizes of the angiogenic-like structures were larger on beads compared to those found in the dispersed cell model. High cell density was needed in angiogenesis when cells were seeded separately, whereas the association of endothelial cells on bead surfaces significantly reduced the cell density used. Moreover, increasing bead density was not necessary to facilitate further angiogenic formation. Micro-spheres may represent a potential support for endothelial cells in microvessel networking, with subsequent applications in the pre-vascularization of bio-implants.