Unknown

Dataset Information

0

Simplified isolation and enrichment of spermatogonial stem-like cells from pubertal domestic cats (Felis catus).


ABSTRACT: The efficiency of spermatogonial stem cell (SSC) isolation and culture from pubertal donors is currently poor primarily, because of contamination with other testicular cells. This study aimed to purify SSC-like cells using different extracellular matrixes and a discontinuous gradient density. In experiment 1, testes (n=6) were analyzed for histology and SSC-related protein expressions (laminin, SSEA-4, DDX-4 and GFR?-1). After enzymatic digestion, the cell suspension was plated onto either a laminin- or gelatin-coated dish. The number of SSC-like cells was determined at 15, 30 and 60 min of culture (experiment 2). Experiment 3 was performed to test whether or not the additional step of Percoll gradient density centrifugation could really improve purification of SSC-like cells. Testicular histology revealed complete spermatogenesis with laminin expression essentially at the basal lamina of the seminiferous tubules. SSEA-4 and GFR?-1 co-localized with DDX-4 in the spermatogonia. The relative percentage of SSC-like cells, as determined by cells expressing SSEA-4 (59.42 ± 2.18%) and GFR?-1 (42.70 ± 1.28%), revealed that the highest SSC-like cell purity was obtained with the 15-min laminin-coated dish compared with other incubation times and gelatin treatment (P<0.05). Percoll treatment prior to laminin selection (15 min) significantly improved SSC-like cell recovery (91.33 ± 0.14%, P<0.001) and purity (83.82 ± 2.05% for SSEA-4 and 64.39 ± 1.51% for GFR?-1, P<0.05). These attached cells demonstrated a typical SSC-like cell morphology and also expressed POU5F1, RET and ZBTB16 mRNA. In conclusion, double enrichment with Percoll gradient density centrifugation and laminin plating highly enriched the SSC-like cells population.

SUBMITTER: Tiptanavattana N 

PROVIDER: S-EPMC4667649 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature

altmetric image

Publications

Simplified isolation and enrichment of spermatogonial stem-like cells from pubertal domestic cats (Felis catus).

Tiptanavattana Narong N   Techakumphu Mongkol M   Tharasanit Theerawat T  

The Journal of veterinary medical science 20150614 11


The efficiency of spermatogonial stem cell (SSC) isolation and culture from pubertal donors is currently poor primarily, because of contamination with other testicular cells. This study aimed to purify SSC-like cells using different extracellular matrixes and a discontinuous gradient density. In experiment 1, testes (n=6) were analyzed for histology and SSC-related protein expressions (laminin, SSEA-4, DDX-4 and GFRα-1). After enzymatic digestion, the cell suspension was plated onto either a lam  ...[more]

Similar Datasets

2016-05-01 | GSE73263 | GEO
| S-EPMC6449508 | biostudies-literature
| S-EPMC4619199 | biostudies-literature
| S-EPMC8366510 | biostudies-literature
| S-EPMC4858209 | biostudies-literature
| S-EPMC7584099 | biostudies-literature
| S-EPMC4976283 | biostudies-literature
| S-EPMC4558093 | biostudies-literature
| S-EPMC4138527 | biostudies-literature
| S-EPMC10029910 | biostudies-literature