Resumption of spermatogenesis in senescent goldfish Carassius auratus (Linnaeus, 1758) through spermatogonial cell therapy

Abstract

In recent times, stem cell research has gained considerable prominence because of its applications in assisted reproductive technology and the treatment of deadly diseases. In teleost fishes, spermatogonial stem cells have been effectively used to produce progeny of difficult-to-breed fish species and/or commercially valuable species through the surrogacy technique. The present study is the first report of an innovative application of stem cell therapy in teleostean fish species for revitalising the reproductive competence of senescent individuals. Senescent male goldfish, Carassius auratus aged approximately 10 years were procured from an ornamental fish-breeding farm and were reared locally for an additional 2 years. The senescence of the fish was evaluated and confirmed using histological analysis, gonadal index assessment, and germ-cell specific vasa gene expression. Analyses revealed the absence of spermatogonial cells and other germ cells in the testes of the senescent fish (n = 5). Spermatogonial cells from a prepubertal C. auratus male donor were isolated using discontinuous percoll gradients, labelled with the fluorescent dye PKH-26, and transplanted into the gonads of senescent C. auratus males through the urogenital papilla. Six months after the therapy, spermatozoa from males were collected through applying gentle manual pressure on the abdomen and were observed under the microscope. All the senescent therapy-treated C. auratus males produced spermatozoa from the transplanted cells; this was confirmed by retention of PKH-26 in the spermatozoa and diagnostic SSR locus. The senescent males were crossed with gravid C. auratus females through artificial insemination and natural spawning, and viable progeny was produced. These observations suggest that the reproductive competence of senescent individuals of commercially valuable and/or endangered fish species can be revitalised and extended through spermatogonia stem cell therapy to produce functional gametes.