Slow freezing, but not vitrification supports complete spermatogenesis in cryopreserved, neonatal sheep testicular xenografts.
The ability to spur growth of early stage gametic cells recovered from neonates could lead to significant advances in rescuing the genomes of rare genotypes or endangered species that die unexpectedly. The purpose of this study was to determine, for the first time, the ability of two substantially different cryopreservation approaches, slow freezing versus vitrification, to preserve testicular tissue of the neonatal sheep and subsequently allow initiation of spermatogenesis post-xenografting. Testis tissue from four lambs (3-5 wk old) was processed and then untreated or subjected to slow freezing or vitrification. Tissue pieces (fresh, n = 214; slow freezing, then thawing, n = 196; vitrification, then warming, n = 139) were placed subcutaneously under the dorsal skin of SCID mice and then grafts recovered and evaluated 17 wk later. Grafts from fresh and slow frozen tissue contained the most advanced stages of spermatogenesis, including normal tubule architecture with elongating spermatids in ~1% (fresh) and ~10% (slow frozen) of tubules. Fewer than 2% of seminiferous tubules advanced to the primary spermatocyte stage in xenografts derived from vitrified tissue. Results demonstrate that slow freezing of neonatal lamb testes was far superior to vitrification in preserving cellular integrity and function after xenografting, including allowing ~10% of tubules to retain the capacity to resume spermatogenesis and yield mature spermatozoa. Although a first for any ruminant species, findings also illustrate the importance of preemptive studies that examine cryo-sensitivity of testicular tissue before attempting this type of male fertility preservation on a large scale.
Center for Species Survival, Smithsonian Conservation Biology Institute, National Zoological Park, Front Royal, Virginia, United States of America.
Budhan S
BS
Pukazhenthi
Center for Species Survival, Smithsonian Conservation Biology Institute, National Zoological Park, Front Royal, Virginia, United States of America; The Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca, New York, United States of America.
Jennifer
J
Nagashima
The Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca, New York, United States of America; Atkinson Center for a Sustainable Future, Cornell University, Ithaca, New York, United States of America.
Alexander J
AJ
Travis
Laboratory of Cellular Biology, Department of Morphology, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Guilherme M
GM
Costa
Department of Agriculture, Food and Resource Sciences School of Agriculture and Natural Sciences, University of Maryland-Eastern Shore, Princess Anne, Maryland, United States of America.
Enrique N
EN
Escobar
Laboratory of Cellular Biology, Department of Morphology, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Luiz R
LR
França
Center for Species Survival, Smithsonian Conservation Biology Institute, National Zoological Park, Front Royal, Virginia, United States of America.
David E
DE
Wildt
D000818:Animals; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D005615:Freezing; D006085:Graft Survival; D064593:Heterografts; D008297:Male; D016513:Mice, SCID; D009926:Organ Preservation / Q000379:methods / Q000662:veterinary*; D034561:Sheep, Domestic / Q000502:physiology*; D013091:Spermatogenesis; D013737:Testis / Q000502:physiology / Q000637:transplantation* / Q000648:ultrastructure; D014183:Transplantation, Heterologous; D058989:Vitrification