Spermatogonial stem cell in zebrafish, Danio rerio: functional characterization and its regulation by FSH (follicule stimulating hormone)

Abstract

Spermatogonial stem cells (SSCs) are the male germline stem cells responsible for maintaining the spermatogenic process along the entire reproductive life throughout cycles of self-renewal and/or differentiation. Although important for the maintenance of the spermatogenesis, the molecular characterization of SSCs remains unknown for several vertebrates, including teleost fish. In this context, the previous work, developed by the undergrad student (FAPESP 2018/17111-3), has characterized some molecular markers of SSCs in zebrafish using specific antibodies for this species through Western Blot (WB) and immunofluorescence (IF). Our results revealed that pluripotency markers Pou5f3, Nanog and Nanos3 are preferentially expressed in undifferentiated spermatogonia. As the cells differentiate, the expression of these markers decreased until undetectable levels, such as in spermatocytes, spermatids and spermatozoa. Based on that and to continue the previous work, the aim of the current project is to examine whether these markers co-localize and if they are associated with the cell cycle of the SSCs by BrdU (Bromodeoxyuridine). Finally, we aim to investigate the regulation of the pluripotent markers in response to follicle stimulating hormone (FSH), which is considered an important endocrine factor involved on the spermatogonial activity in zebrafish. To achieve these goals, techniques such as, IF, WB and testis tissue culture, will be employed. This methodology is already standardize by the Reproductive and Molecular Biology group at UNESP/Botucatu. The obtained results will contribute for a better understanding about the SSC biology in teleosts, as well as, to elucidate the spermatogonial differentiation processes and its integration with the endocrine and paracrine signals from the stem cell niche. Moreover, this project will provide means to understand the SSC regulation under physiological conditions, as well as, for future studies on ecotoxicology. (AU)