Sturgeons are among the most ancient linages of actinopterygians. At present, many sturgeon species are critically endangered. Surrogate production could be used as an affordable and a time-efficient method for endangered sturgeons. Our study established a method for identifying and isolating type A spermatogonia from different developmental stages of testes using flow cytometric cell sorting (FCM). Flow cytometric analysis of a whole testicular cell suspension showed several well-distinguished cell populations formed according to different values of light scatter parameters. FCM of these different cell populations was performed directly on glass slides for further immunocytochemistry to identify germ cells. Results showed that the cell population in gate P1 on a flow cytometry plot (with high forward scatter and high side scatter parameter values) contains the highest amount of type A spermatogonia. The sorted cell populations were characterized by expression profiles of 10 germ cell specific genes. The result confirmed that setting up for the P1 gate could precisely sort type A spermatogonia in all tested testicular developmental stages. The P2 gate, which was with lower forward scatter and side scatter values mostly, contained type B spermatogonia at a later maturing stage. Moreover, expressions of plzf, dnd, boule, and kitr were significantly higher in type A spermatogonia than in later developed germ cells. In addition, plzf was firstly found as a reliable marker to identify type A spermatogonia, which filled the gap of identification of spermatogonial stem cells in sterlet. It is expected to increase the efficiency of germ stem cell culture and transplantation with plzf identification. Our study thus first addressed a phenotypic characterization of a pure type A spermatogonia population in sterlet. FCM strategy can improve the production of sturgeons with surrogate broodstock and further the analysis of the cellular and molecular mechanisms of sturgeon germ cell development.

• Keywords
• PLZF, fluorescence-activated cell sorting, germ stem cell, gonad, spermatogonia, sturgeon,
• Publication type
• Journal Article MeSH

Spermatogenesis is a continuous and dynamic developmental process, in which a single diploid spermatogonial stem cell (SSC) proliferates and differentiates to form a mature spermatozoon. Herein, we summarize the accumulated knowledge of SSCs and their distribution in the testes of teleosts. We also reviewed the primary endocrine and paracrine influence on spermatogonium self-renewal vs. differentiation in fish. To provide insight into techniques and research related to SSCs, we review available protocols and advances in enriching undifferentiated spermatogonia based on their unique physiochemical and biochemical properties, such as size, density, and differential expression of specific surface markers. We summarize in vitro germ cell culture conditions developed to maintain proliferation and survival of spermatogonia in selected fish species. In traditional culture systems, sera and feeder cells were considered to be essential for SSC self-renewal, in contrast to recently developed systems with well-defined media and growth factors to induce either SSC self-renewal or differentiation in long-term cultures. The establishment of a germ cell culture contributes to efficient SSC propagation in rare, endangered, or commercially cultured fish species for use in biotechnological manipulation, such as cryopreservation and transplantation. Finally, we discuss organ culture and three-dimensional models for in vitro investigation of fish spermatogenesis.

• Keywords
• fish, florescence-activated cell sorting (FACS), germ cell culture, magnetic-activated cell sorting (MACS), spermatogenesis, spermatogonial stem cell (SSC),
• MeSH
• Cell Culture Techniques * MeSH
• Stem Cells cytology   ultrastructure   MeSH
• Fishes metabolism   MeSH
• Cell Separation * MeSH
• Spermatogenesis MeSH
• Spermatogonia cytology   ultrastructure   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH