Resumption of donor-origin spermatogenesis in senescent goldfish Carassius auratus (Linnaeus, 1758) following spermatogonial cell therapy

Abstract

Stem cell research has come into 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 surrogate gametes and progeny through germ cell transplantation technique. The present study is the first report of an innovative application of stem cell therapy in fish species for revitalising the reproductive competence of senescent individuals. Senescent male goldfish, Carassius auratus, approximately 10 years of age were procured from a fish-breeding farm and were reared locally in the lab for an additional two years. The senescence of the individuals was then evaluated and confirmed using histological analysis, gonadal index assessment, and germ-cell specific vasa gene expression. Analyses revealed absence of spermatogonial cells and other germ cells in the testes of the senescent fish (n = 5). Spermatogonial cells from sexually immature 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 urogenital papilla. Six months after the transplant, spermatozoa were collected through applying gentle manual pressure on the abdomen and were observed under a microscope. All C. auratus males with the transplant had produced spermatozoa from the transplanted cells. This was confirmed by the retention of PKH-26 in the spermatozoa and diagnostic SSR locus. Gravid C. auratus females were artificially inseminated with the spermatozoa of those senescent males and natural spawning was allowed. As a result viable progeny were produced. These observation suggests that the reproductive competence of senescent male fishes can be revitalised through spermatogonial stem cell therapy to produce functional gametes.