Spermatogonial stem cells (SSCs) are distinct in their ability to self-renew, transmit genetic information, and persist throughout the life of an individual. These characteristics make SSCs a useful tool for addressing diverse challenges such as efficient transgenic production in non-rodent, biomedical animal models or preservation of the male genome for species in which survival of frozen-thawed sperm is low. A requisite first step to accessing this technology in felids is the establishment of molecular markers. This study was designed to evaluate, in the domestic cat (Felis catus), expression both in situ and following enrichment in vitro of six genes (GFRA1, GPR125, ZBTB16, POU5F1, THY1, UCHL1) that had been previously identified as SSC markers in other species. Antibodies for surface markers GFRA1, GPR125, and THY1 could not be validated, while western blot analysis of prepubertal, peripubertal, and adult cat testis confirmed protein expression for the intracellular markers UCHL1, ZBTB16, and POU5F1. Co-localization of the markers by immunohistochemistry revealed that several cells within the subpopulation adjacent to the basement membrane of the seminiferous tubules and identified morphologically as spermatogonia, expressed all three intracellular markers. Studies performed on cheetah (Acinonyx jubatus) and Amur leopard (Panthera pardus orientalis) testis exhibited a conserved expression pattern in protein molecular weights, relative abundance, and localization of positive cells within the testis. The expression of the three intracellular SSC marker proteins in domestic and wild cat testes confirms conservation of these markers in felids. Enrichment of marker transcripts following differential plating was also observed. These markers will facilitate further studies in cell enrichment and in vitro culture of felid SSCs enabling both production of transgenic domestic cats and preservation of the male genome from rare and endangered felids.
