Use of Human Gingival Fibroblasts for Pre-Vascularization Strategies in Oral Tissue Engineering.

Abstract

Cocultures of human gingival fibrobasts (hGF) and endothelial cells could enhance regeneration and repair models as well as improve vascularization limitations in tissue engineering. The aim of this study was to assess if hGF could support formation of stable vessel-like networks. Explant primary hGF were isolated from gum surgical wastes collected from healthy patients with no history of periodontitis. Human umbilical vein endothelial cells (HUVEC) were two-dimensional (2D) and three-dimensional (3D) cocultured in vitro with hGF at a cell ratio of 1:1 and medium of 1:1 of their respective media during at least 31days. Vessel quantification of HUVEC networks was performed. In order to investigate the pericyte-like properties of hGF, the expression of perivascular markers α-SMA, NG2, CD146 and PDGFR-β was studied using immunocytochemistry and flow cytometry on 2D cultures. hGF were able to support a long-lasting HUVEC network at least 31days, even in the absence of a bioreactor with flow. As observed, HUVEC started to communicate with each other from day 7, constructing a network. Their interconnection increased significantly between day 2 and day 21 and lasted beyond the 31days of observation. Moreover, we tried to explain the stability of the networks obtained and showed that a small population of hGF in close vicinity of HUVEC networks expressed perivascular markers. These findings highlight a new interesting property concerning hGF, accentuating their relevance in tissue engineering and periodontal regeneration. These promising results need to be confirmed using more 3D applications and in vivo testing.