Meagre <i>Argyrosomus regius</i> (Asso, 1801) Stem Spermatogonia: Histological Characterization, Immunostaining, In Vitro Proliferation, and Cryopreservation

• Main Authors: Rosa Zupa, Nicola A. Martino, Giuseppina Marzano, Maria E. Dell’Aquila, Aldo Corriero
• Format: Article
• Language: English
• Published: MDPI AG 2020-05-01
• Series: Animals
• Subjects: stem spermatogonia; fish reproduction; germ cell characterization; gem cell culture
• Online Access: https://www.mdpi.com/2076-2615/10/5/851

The meagre, <i>Argyrosomus regius</i>, is a valued fish species of which aquaculture production might be supported by the development of a stem germ cell xenotransplantation technology. Meagre males were sampled at a fish farm in the Ionian Sea (Italy) at the beginning and end of the reproductive season. Small and large Type A undifferentiated spermatogonia were histologically identified in the germinal epithelium. Among the tested stemness markers, anti-oct4 and anti-vasa antibodies labeled cells likely corresponding to the small single Type A spermatogonia; no labeling was obtained with anti-GFRA1 and anti-Nanos2 antibodies. Two types of single A spermatogonia were purified via density gradient centrifugation of enzymatically digested testes. Testes from fish in active spermatogenesis resulted in a more efficient spermatogonial stem cell (SSC) yield. After cell seeding, meagre SSCs showed active proliferation from Day 7 to Day 21 and were cultured up to Day 41. After cryopreservation in dimethyl-sulfoxide-based medium, cell viability was 28.5%. In conclusion, these results indicated that meagre SSCs could be isolated, characterized, cultured in vitro, successfully cryopreserved, and used after thawing. This is a first step towards the development of a xenotransplantation technology that might facilitate the reproduction of this valuable species in captivity.