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Slow freezing, but not vitrification supports complete spermatogenesis in cryopreserved, neonatal sheep testicular xenografts.

Pukazhenthi BS, Nagashima J, Travis AJ, Costa GM, Escobar EN, França LR, Wildt DE - PLoS ONE (2015)

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Center for Species Survival, Smithsonian Conservation Biology Institute, National Zoological Park, Front Royal, Virginia, United States of America.

ABSTRACT
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.

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Xenograft volume density traits for pieces of lamb testis transplanted fresh (control) versus after slow freezing and thawing versus vitrification and thawing.For each assessed metric, data for the slow freezing group were no different (P>0.05) from the control. Within a trait, lines above the error bar denote significance.
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pone.0123957.g003: Xenograft volume density traits for pieces of lamb testis transplanted fresh (control) versus after slow freezing and thawing versus vitrification and thawing.For each assessed metric, data for the slow freezing group were no different (P>0.05) from the control. Within a trait, lines above the error bar denote significance.

Mentions: Histological assessments of tissue grafts at 17 wk revealed more area comprised of tubular compartment, tubular lumen, seminiferous epithelium and tunica propria in fresh and slow frozen samples compared to vitrified counterparts (Fig 3A–3C). By contrast, intertubular compartment values were higher in vitrified compared to fresh (P<0.01) or slow frozen tissue (P<0.001; Fig 3D). The proportion of tissue volume comprised of Leydig cells did not differ among the three treatments (Fig 3E). Fresh and slow frozen tissue xenografts also expressed similar percentages of seminiferous cords (Fig 4A) and seminiferous tubules (Fig 4B). Xenografts derived from vitrified tissue retained mostly unchanged seminiferous cords with few seminiferous tubules.


Slow freezing, but not vitrification supports complete spermatogenesis in cryopreserved, neonatal sheep testicular xenografts.

Pukazhenthi BS, Nagashima J, Travis AJ, Costa GM, Escobar EN, França LR, Wildt DE - PLoS ONE (2015)

Xenograft volume density traits for pieces of lamb testis transplanted fresh (control) versus after slow freezing and thawing versus vitrification and thawing.For each assessed metric, data for the slow freezing group were no different (P>0.05) from the control. Within a trait, lines above the error bar denote significance.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4414479&req=5

pone.0123957.g003: Xenograft volume density traits for pieces of lamb testis transplanted fresh (control) versus after slow freezing and thawing versus vitrification and thawing.For each assessed metric, data for the slow freezing group were no different (P>0.05) from the control. Within a trait, lines above the error bar denote significance.
Mentions: Histological assessments of tissue grafts at 17 wk revealed more area comprised of tubular compartment, tubular lumen, seminiferous epithelium and tunica propria in fresh and slow frozen samples compared to vitrified counterparts (Fig 3A–3C). By contrast, intertubular compartment values were higher in vitrified compared to fresh (P<0.01) or slow frozen tissue (P<0.001; Fig 3D). The proportion of tissue volume comprised of Leydig cells did not differ among the three treatments (Fig 3E). Fresh and slow frozen tissue xenografts also expressed similar percentages of seminiferous cords (Fig 4A) and seminiferous tubules (Fig 4B). Xenografts derived from vitrified tissue retained mostly unchanged seminiferous cords with few seminiferous tubules.

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Center for Species Survival, Smithsonian Conservation Biology Institute, National Zoological Park, Front Royal, Virginia, United States of America.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus