Obtenção e caracterização de populações de células-tronco CD200 e CD34 positivas da pele na espécie canina

• Ano de defesa: 2016
• Autor(a) principal: Castro, Raquel Vasconcelos Guimarães de [UNESP]
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Estadual Paulista (Unesp)
• Palavras-chave em Português: Flow cytometry; Cell culture; Hair follicle; Immunofluorescence; Bulge; Citometria de fluxo; Cultivo celular; Folículo piloso; Imunofluorescência
• Link de acesso: http://hdl.handle.net/11449/139466

Skin is an extensive and easily accessible organ, possessing various cell types which are in constant renovation. Some stem cell types found in the skin have the potential of self-renewal, proliferation and differentiation, processes which are responsible for the organ’s maintenance of homeostasis and the healing of wounds. Previous studies suggested the presence of a stem cell niche at the bulge region of the hair follicle, which contains cell populations positive for CD200 and CD34. Thus, this work sought to identify these cell populations in canine cultures using the following methods: 1. 1- Collecting samples of adult and fetus canine skin, isolating and culturing these cells in vitro using a method of simple enzymatic digestion; 2- Testing the cell cultures for CD200 and CD34 in vitro, comparing them with analyzed tissue material. Hematoxylin and eosin staining were conducted for the biopsies and analysis of fetal and adult canine skin, with the extracted cell cultures being characterized for the presence of the proteins CD200 and CD34 through immunofluorescence, immunocytochemistry and flow cytometry. In both adult and fetal tissue samples, CD200 and CD34 immunofluorescence results were negative. In immunocytochemistry, both fetal and adult cultures cells tested positive for CD34 and CD200. The pluripotency marker OCT4 was also tested, and was positive for fetal culture cells. Flow cytometer results showed that, for samples with a double staining of CD200 and CD34 the average percentage of marked cells was 3.1% in adults and 0.33% in fetal cells (n=3). For the CD200 marker alone, positive cells were found only in adult cultures, representing 2.8% of the total population (n=3). In conclusion, the results suggest that obtaining bulge stem cells from both fetuses and adults, with use of CD200 and CD34 markers, is validated through the simple enzymatic digestion and cell culture methods utilized in this study. The present work is the first step towards developing new research involving this niche in dogs.