title: Molecular markers of putative spermatogonial stem cells in the domestic cat.
abstract: Spermatogonial stem cells (SSCs) are an important tool for fertility preservation and species conservation. The ability to expand SSCs by in vitro culture is a crucial premise for their use in assisted reproduction. Because SSCs represent a small proportion of the germ cells in the adult testis, culture success is aided by pre-enrichment through sorting techniques based on cell surface-specific markers. Given the importance of the domestic cat as a model for conservation of endangered wild felids, herein we sought to examine culture conditions as well as molecular markers for cat SSCs. Using a cell culture medium for mouse SSCs supplemented with glial cell-derived neurotrophic factor (GDNF), germ cells from prepuberal cat testes remained viable in culture for up to 43 days. Immunohistochemistry for promyelocytic leukaemia zinc finger (PLZF) protein on foetal, prepuberal and adult testis sections revealed a pattern of expression consistent with the labelling of undifferentiated spermatogonia. Fluorescence-activated cell sorting (FACS) with an antibody against epithelial cell adhesion molecule (EPCAM) was used to sort live cells. Then, the gene expression profile of EPCAM-sorted cells was investigated through RT-qPCR. Notably, EPCAM (+) cells expressed relatively high levels of CKIT (CD117), a surface protein typically expressed in differentiating germ cells but not SSCs. Conversely, EPCAM (-) cells expressed relatively high levels of POU domain class 5 transcription factor 1 (POU1F5 or OCT4), clearly a germ line stem cell marker. These results suggest that cat SSCs would probably be found within the population of EPCAM (-) cells. Future studies should identify additional surface markers that alone or in combination can be used to further enrich SSCs from cat germ cells.
authors.affiliation: Center for Regenerative Medicine (CMRB), Barcelona, Spain.|Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Madrid, Spain.
authors.forename: S J
authors.initials: SJ
authors.lastname: Bedford-Guaus
authors.affiliation: Department of Chemistry, Korea University, Seoul, Korea.
authors.forename: S
authors.initials: S
authors.lastname: Kim
authors.affiliation: Center for Regenerative Medicine (CMRB), Barcelona, Spain.
authors.forename: L
authors.initials: L
authors.lastname: Mulero
authors.affiliation: Center for Regenerative Medicine (CMRB), Barcelona, Spain.
authors.forename: J M
authors.initials: JM
authors.lastname: Vaquero
authors.affiliation: Center for Regenerative Medicine (CMRB), Barcelona, Spain.
authors.forename: C
authors.initials: C
authors.lastname: Morera
authors.affiliation: Centre Veterinari La Sagrera, Barcelona, Spain.
authors.forename: R
authors.initials: R
authors.lastname: Adan-Milanès
authors.affiliation: Center for Regenerative Medicine (CMRB), Barcelona, Spain.|Reproductive Medicine Service, Department of Obstetrics & Gynecology, Hospital Universitari Quirón-Dexeus, Barcelona, Spain.
authors.forename: A
authors.initials: A
authors.lastname: Veiga
authors.affiliation: Center for Regenerative Medicine (CMRB), Barcelona, Spain.|Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Madrid, Spain.|Institució Catalana de Recerca i Estudis Avançats, Barcelona, Spain.
authors.forename: Á
authors.initials: Á
authors.lastname: Raya
mesh_terms: D000072956:Adult Germline Stem Cells / Q000737:chemistry*; D000818:Animals; D015415:Biomarkers / Q000032:analysis*; D002415:Cats; D002469:Cell Separation / Q000379:methods / Q000662:veterinary; D002478:Cells, Cultured; D003247:Conservation of Natural Resources; D056727:Endangered Species; D000071858:Epithelial Cell Adhesion Molecule; D005434:Flow Cytometry / Q000662:veterinary; D007150:Immunohistochemistry / Q000662:veterinary; D051741:Kruppel-Like Transcription Factors / Q000032:analysis; D008297:Male; D023421:Models, Animal; D012741:Sexual Maturation; D013093:Spermatogonia / Q000737:chemistry; D013737:Testis / Q000166:cytology; D059467:Transcriptome
chemical_list: D015415:Biomarkers; D000071858:Epithelial Cell Adhesion Molecule; D051741:Kruppel-Like Transcription Factors
keywords: Felis catus; epithelial cell adhesion molecule; promyelocytic leukaemia zinc finger protein; spermatogonial stem cells