ИСТИНА

Abstract: Objectives: Emerging evidence suggests that adult craniofacial tissues in vertebrates contain limited numbers of post-migratory neural crest-derived stem cells (NCSCs). We hypothesized that the ovine palate contains NCSCs equipped with a developmental potential equivalent to their rodent and human counterparts. Methods: Palatal tissue was extracted (ovineNCSCs) from female sheep (StStMU Animal Ethics Committee approval number 39, 16/04/2014) and human 3rd molars removed for medical reasons were used for periodontal ligamentum purification (hpdNCSC) if a written informed consent was obtained. The procedure of the NCSCs isolation is described in detail by Zeuner et al. (2017). In brief, both human pdNCSCs and ovine paldNCSCs were dissociated and cultured in serum-free media. Primary neurospheres were dissociated at days 8–10 and for adherent culture cultivated. Most of NCSCs populations can be isolated and enriched using magnetic cell separation (e.g. CliniMACS) based on their expression of the cell surface marker CD271. The yield and purity of the MACS-separation was assessed using flow cytometry. Results: Human ligamentum and ovine palatal tissues contained NCSCs capable for trilineage differentiation and ectodermal transdifferentiation. Human pdNCSCs had high proliferative activity (PDT- 24h). Cultivated ovineNCSCs were highly proliferative with an average population doubling time of 26h. Ovine paldNCSCs had higher osteogenic potential than human pdNCSCs. After isolation, both of investigated NCSCs formed neurospheres, express neural crest markers (e.g. HNK-1, Slug, CD271). We characterized the endogenous niche of the investigated NCSCs histologically and immunohistochemically and successfully cultivated the cells as adherent monolayer and as self-adherent neurosphere cultures. Conclusions: Our data suggest that ovineNCSCs are present in the ovine palate and human pdNCSCs in the periodontium. Magnetic separation is a feasible method to enrich human and ovine NCSCs for comparative research. Our study could pave the way allowing middle- and long-term studies of safety and efficacy of endogenous NCSCs transplantations. Disclosure Statement: The submitter must disclose the names of the organizations with which any author have a relationship, the nature of the relationship, and the clinical or research area involved. The following is submitted: None