alpha-6 Integrin Expression in Bovine Spermatogonial Cells Purified by Discontinuous Percoll Density Gradient

Autor(es)

de Barros, F. R. O. ^[1] ; Worst, R. A. ; Saurin, G. C. P. ; Mendes, C. M. ; Assumpcao, M. E. O. A. ; Visintin, J. A.

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Dept Anim Reprod, VRA, Sch Vet Med & Anim Sci, Lab Vitro Fertilizat Cloni, BR-05508270 Sao Paulo - Brazil

• Tipo de documento: Artigo Científico
• Fonte: REPRODUCTION IN DOMESTIC ANIMALS; v. 47, n. 6, p. 887-890, DEC 2012.
• Citações Web of Science: 9

Resumo

The study of spermatogonial stem cells (SSCs) provides a model to better understand adult stem cell biology. Besides the biomedical potential to perform studies of infertility in many species, SSCs hold a promising application at animal transgenesis. Because stem cells are thought to be associated with basement membranes, expression of alpha-6 integrin has been investigated as a marker of type A spermatogonial cells, which are considered SSCs because of their undifferentiated status and self-renewal ability. In this manner, the aim of this study was to isolate type A SSCs from adult bulls by a two-step enzymatic procedure followed by a discontinuous Percoll density gradient purification and verify the expression of alpha-6 integrin by flow cytometry and real-time RT-PCR before and after Percoll purification. Spermatogonial cells were successfully obtained using the two-step enzymatic digestion. An average of 1 x 10(5) viable cells per gram of testis was isolated. However, the discontinuous Percoll did not purify isolated cells regarding alpha-6 integrin expression. Flow cytometry analysis demonstrated no differences in the alpha-6 integrin expression between cell samples before and after Percoll purification (p = 0.5636). The same was observed in the real-time PCR analysis (p > 0.05). In addition to alpha-6 integrin, the expression of GFR alpha-1 and PGP9.5, known bovine SSCs markers, was detected in all samples studied. Considering that Percoll can reduce cell viability, it is possible to conclude that Percoll density gradient is not suitable to purify bovine SSC, according to alpha-6 integrin expression. (AU)

• Processo FAPESP: 09/52241-6 - Modificacao de celulas-tronco espermatogoniais para producao de bovinos transgenicos.
• Beneficiário: Flavia Regina Oliveira de Barros
• Modalidade de apoio: Bolsas no Brasil - Doutorado
• Processo FAPESP: 10/51654-2 - Aplicação de células-tronco espermatogoniais na produção de animais transgênicos
• Beneficiário: José Antonio Visintin
• Modalidade de apoio: Auxílio à Pesquisa - Regular