Jornal de pesquisa e terapia com células-tronco

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Guidelines to Optimize Survival and Migration Capacities of Equine Mesenchymal Stem Cells

Sarah Broeckx, Cynthia de Vries, Marc Suls, Deborah J Guest and Jan H Spaas

Background: Although mesenchymal stem cell (MSC) therapy is commonly being used in veterinary medicine, no specific guidelines are described to guarantee an optimal cell survival and migration. However, this is indispensable in order to assure an optimal clinical use. Methods: In the present study, equine peripheral blood (PB)-derived MSCs were isolated and characterized. In order to determine the optimal parameters for long-term cryopreservation, the effects of different dimethyl sulfoxide (DMSO) concentrations (5-20%) and thawing methods (temperature-controlled or hand-thawed) on MSC viability were evaluated by means of trypan blue staining. Afterwards, MSC storage was evaluated for 12 months using only 10% of DMSO as a cryoprotectant without rate-controlled freezing. Then, the influence of frequently used anaesthetics and injectable gels on MSC viability was assessed. Finally, the migration capacity of MSCs through hyaluronic acid at different concentrations was assessed. Results: Firstly, we demonstrated that the DMSO concentration and the thawing method had no considerable influence on MSC viability within the 4 hour evaluation after thawing. Secondly, anaesthetics were highly cytotoxic and only 10% of the MSCs survived 3 hours of their presence. Hyaluronic acid- and glycosaminoglycan-based gels were compatible with MSCs and allowed a high cell survival (more than 90%) within 48 hours of follow-up. However, in a polyacrylamide gel, approximately 80% of the MSCs died within 48 hours. Finally, the concentration of the gel influenced the migration capacity of the MSCs. Indeed, only 60% of the MSCs in 20mg/ml of hyaluronic acid attached to the underlying surface within 48 hours, whereas almost 100% of the MSCs in 10mg/ml of the same substance were able to migrate through the gel. Conclusion: The present study reports practical guidelines for the clinical application of equine PB-derived MSCs.