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Spermatogonial stem cells identified by molecular expression of PLZF, integrin β1 and reactivity to Dolichos biflorus agglutinin in alpaca adult testes
Author(s) -
Valdivia Martha,
CastañedaZegarra Sergio,
Lévano Gloria,
Lazo Jorge,
Reyes Jhakelin,
Bravo Zezé,
Santiani Alexei,
Mujica Fidel,
Ruíz Jaime,
Gonzales Gustavo F.
Publication year - 2019
Publication title -
andrologia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.633
H-Index - 59
eISSN - 1439-0272
pISSN - 0303-4569
DOI - 10.1111/and.13283
Subject(s) - biology , andrology , trypan blue , flow cytometry , agglutinin , microbiology and biotechnology , sperm , homeobox protein nanog , gene , lectin , cell , genetics , embryonic stem cell , medicine , induced pluripotent stem cell
The identification system of spermatogonial stem cell (SSC) was established in alpaca using the molecular expression as well as the reactivity pattern to Dolichos biflorus agglutinin (DBA) by flow cytometry. Twenty‐four testicles with their epididymis were recovered from adult alpacas at the slaughterhouse of Huancavelica—Perú. Samples were transported to the Laboratory of Reproductive Physiology at Universidad Nacional Mayor de San Marcos. Testes were selected for our study when the progressive motility of epididymal spermatozoa (ESPM) was above 30%. Isolation of SSC was performed with two enzymatic digestions. Finally, sperm viability was evaluated by means of the trypan blue vital stain in spermatogonial round cells. Samples with more than 80% viability were selected. Isolated cells cultured for 2 days were used for identifying the presence of SSCs by the expression of integrin β1 (116 bp) and PLZF (206 bp) genes. Spermatogonia were classified according to the DBA reactivity. Spermatogonia with a strong positive to DBA (sDBA + ) were classified as SSC (Mean ± SEM =4.44 ± 0.68%). Spermatogonia in early differentiation stages stained weakly positive with DBA (wDBA + ) (Mean ± SEM =37.44 ± 3.07%) and differentiated round cells as DBA negative (Mean ± SEM =54.12 ± 3.18%). With the use of molecular and DBA markers, it is possible to identify easily the spermatogonial stem cells in alpaca.