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Cell transplantation is a useful therapy for treating peripheral nerve injuries. The clinical use of Schwann cells (SCs), however, is limited because of their limited availability. An emerging solution to promote nerve regeneration is to apply injured nerves with stem cells derived from various tissues. In this study, different types of allogeneic cells including SCs, adipose-derived adult stem cells (ASCs), dental pulp stem cells (DPSCs), and the combination of SCs with ASCs or DPSCs were seeded on nerve conduits to test their efficacy in repairing a 15-mm-long critical gap defect of rat sciatic nerve. The regeneration capacity and functional recovery were evaluated by the histological staining, electrophysiology, walking track, and functional gait analysis after 8 weeks of implantation. An in vitro study was also performed to verify if the combination of cells led to syner-gistic neurotrophic effects (NGF, BDNF, and GDNF). Experimental rats receiving conduits seeded with a combination of SCs and ASCs had the greatest functional recovery, as evaluated by the walking track, functional gait, nerve conduction velocity (NCV), and histological analysis. Conduits seeded with cells were always superior to the blank conduits without cells. Regarding NCV and the number of blood vessels, conduits seeded with SCs and DPSCs exhibited better values than those seeded with DPSCs only. Results from the in vitro study confirmed the synergistic NGF production from the coculture of SCs and ASCs. It was concluded that coculture of SCs with ASCs or DPSCs in a conduit promoted peripheral nerve regeneration over a critical gap defect.

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Human gingival fibroblasts (HGF) were recently found to be a source of mesenchymal stem cells. Their behavior on a biomaterial has not been reported so far. The effect of culturing HGF on chitosan membranes on their chondrogenic differentiation was investigated in this study. HGF were first cultured on chitosan membranes and spheroid formation of HGF was observed. Next, HGF on chitosan were induced with chondrogenesis induction medium and their chondrogenic differentiation potential was expressed by assessing the expression of chondrogenesis related genes at both mRNA and protein levels by reverse transcription-polymerase chain reaction (RT-PCR) and immunostaining, respectively. We discovered that the chondrogenic differentiation potential of HGF could be enhanced simply by culturing HGF on chitosan membranes. Expression of neural crest and stemness genes were also analyzed by RT-PCR to evaluate the stemness and self-renewal of HGF spheroids. We found that spheroid formation helped to increase and maintain the expression of stemness genes in HGF. To understand the aspects of the chitosan membranes that induced spheroid formation of HGF, mechanical and physical properties of the chitosan membranes were examined. The migration of HGF on chitosan membranes was also monitored to speculate the process of spheroid formation. In addition, the roles of the Rho/Rho-associated kinase (ROCK) pathway and connexin 43 (Cx43) in spheroid formation were explored. Treatment of HGF cultured on chitosan with the ROCK-activity inhibitor Y27632 clearly inhibited spheroid formation, suggesting that the Rho/ROCK pathway was involved in spheroid formation. The increased Cx43 activity of HGF spheroids on chitosan indicated that the gap junction intercellular communication was regulated by spheroid formation. It was concluded that culturing HGF on chitosan may activate the Rho/ROCK pathway, which led to spheroid formation and gap junction regulation. These changes may contribute to the enhanced chondrogenic differentiation potential of HGF on chitosan.