title: Phenotypic characterization and in vitro propagation and transplantation of the Nile tilapia (Oreochromis niloticus) spermatogonial stem cells.
abstract: In association with in vitro culture and transplantation, isolation of spermatogonial stem cells (SSCs) is an excellent approach for investigating spermatogonial physiology in vertebrates. However, in fish, the lack of SSC molecular markers represents a great limitation to identify/purify these cells, rendering it difficult to apply several valuable biotechnologies in fish-farming. Herein, we describe potential molecular markers, which served to phenotypically characterize, cultivate and transplant Nile tilapia SSCs. Immunolocalization revealed that Gfra1 is expressed exclusively in single type A undifferentiated spermatogonia (Aund, presumptive SSCs). Likewise, the expression of Nanos2 protein was observed in Aund cells. However, Nanos2-positive spermatogonia have also been identified in cysts with two to eight germ cells that encompass type A differentiated spermatogonia (Adiff). Moreover, we also established effective primary culture conditions that allowed the Nile tilapia spermatogonia to expand their population for at least one month while conserving their original undifferentiated (stemness) characteristics. The maintenance of Aund spermatogonial phenotype was demonstrated by the expression of early germ cell specific markers and, more convincingly, by their ability to colonize and develop in the busulfan-treated adult Nile tilapia recipient testes after germ cell transplantation. In addition to advancing our knowledge on the identity and physiology of fish SSCs, these findings provide the first step in establishing a system that will allow fish SSCs expansion in vitro, representing an important progress towards the development of new biotechnologies in aquaculture, including the possibility of producing transgenic fish.
authors.affiliation: Laboratory of Cellular Biology, Department of Morphology, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais 31270-901, Brazil.
authors.forename: Samyra Maria
authors.initials: SM
authors.lastname: Santos Nassif Lacerda
authors.forename: Guilherme Mattos Jardim
authors.initials: GM
authors.lastname: Costa
authors.forename: Mariana de Araújo
authors.initials: Mde A
authors.lastname: da Silva
authors.forename: Paulo Henrique Almeida
authors.initials: PH
authors.lastname: Campos-Junior
authors.forename: Tânia Mara
authors.initials: TM
authors.lastname: Segatelli
authors.forename: Marco Túlio Diniz
authors.initials: MT
authors.lastname: Peixoto
authors.forename: Rodrigo Ribeiro
authors.initials: RR
authors.lastname: Resende
authors.forename: Luiz Renato
authors.initials: LR
authors.lastname: de França
mesh_terms: D000818:Animals; D023681:Cichlids / Q000378:metabolism*; D029941:Fish Proteins / Q000378:metabolism; D008297:Male; D013093:Spermatogonia / Q000166:cytology*; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000166:cytology*; D013737:Testis / Q000166:cytology
chemical_list: D029941:Fish Proteins
keywords: Gfra1; Nanos2; Nile tilapia (Oreochromis niloticus); Spermatogonial stem cells; Spermatogonial transplantation; Stem cell markers