Influence of Vitrification Device, Warming Protocol, and Subsequent <i>In Vitro</i> Culture on Structural Integrity of Testicular Fragments from Adult Domestic Cats

Macente, Beatrice Ingrid; Fonseca‐Alves, Carlos Eduardo; Magalhães, Georgia Modé; Tavares, Mariana Riboli; Mansano, Cleber Fernando Menegasso; Mouttham, Lara; Apparício, M.; Toniollo, Gilson Hélio; Comizzoli, Pierre

doi:10.1089/bio.2021.0092

Cited by 4 publications

(4 citation statements)

References 36 publications

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“…Although plastic straws showed the highest number of apoptotic cells than OTC and SSV, a recent study showed fewer caspase-positive testicular cells in adult cats when the Cryotop system was tested [12], like plastic in the lower conductivity material. Additionally, the direct and permanent contact of testicular tissue with the vitrification solution during the storage period can increase the toxic effects on cells because of the high concentration of the medium by cryoprotectant agents [40].…”

Section: Discussionmentioning

confidence: 96%

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Effect of Different Vitrification Techniques on Viability and Apoptotic Index of Domestic Cat Testicular Tissue Cells

Carvalho,

Soares,

Leão

et al. 2023

Animals

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Vitrification is essential for successful tissue cryopreservation and biobanking in wild cats. This study aimed to compare different methods of vitrification (Ovarian Tissue Cryosystem—OTC, Straws—STW, and Solid Surface vitrification—SSV) for testicular fragment vitrification in tom cats. Testicular fragments were recovered from five adult tom cats and subjected to equilibrium vitrification using different cryovials and methods under the same conditions of vitrification solutions and cryoprotectants. The efficiencies of the methods were evaluated using histological analysis of spermatogonia and Sertoli cell nuclei, seminiferous tubular basement membrane detachment, and the gonadal epithelium shrinkage score scale. Cell viability was assessed using Hoechst PI and Terminal deoxynucleotidyl transferase nick end labeling (TUNEL) assay. The results showed that OTC is an effective vitrification method for maintaining the distinction between spermatogonia and Sertoli cells. OTC was similar to the control for basal membrane detachment parameters (p = 0.05). Epithelial shrinkage was low in the SSV group, which showed the highest percentage of viable cells among the vitrified groups (p = 0.0023). The OTC and SSV vitrification methods were statistically similar in terms of the percentage of TUNEL-positive cells (p = 0.05). Therefore, OTC and SSV provide favorable conditions for maintaining viable cat testicular tissue cells after vitrification.

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Section: Discussionmentioning

confidence: 96%

“…In domestic cats, studies have compared vitrification and slow freezing [9], the use of cryoprotectants for prepubertal cats [10], and the effects of warming temperatures [1,11] on test tissue. Recently, some results have been described for adult cat testicular fragment vitrification protocols and conditions of warming [12].…”

Section: Introductionmentioning

confidence: 99%

Effect of Different Vitrification Techniques on Viability and Apoptotic Index of Domestic Cat Testicular Tissue Cells

Carvalho,

Soares,

Leão

et al. 2023

Animals

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“…As mentioned before, the higher dose of cryoprotectants during vitrification could be the reason, in our study, of the lower viability and DNA integrity of vitrified/warmed elongated spermatids/spermatozoa compared to those slow frozen/thawed. Other factors to consider are the cooling rate and the high temperature used for warming, since spermatozoa seem to be more sensitive to them than other germ cells ( 30 , 33 ). In fact, ejaculated spermatozoa from wild ungulates have shown a poor tolerance to the fast cooling rate of vitrification ( 30 , 31 ) and warming vitrified-needle samples at 50°C resulted in DNA damage in testicular spermatozoa from adult cats ( 33 ).…”

Section: Discussionmentioning

confidence: 99%

“…Other factors to consider are the cooling rate and the high temperature used for warming, since spermatozoa seem to be more sensitive to them than other germ cells ( 30 , 33 ). In fact, ejaculated spermatozoa from wild ungulates have shown a poor tolerance to the fast cooling rate of vitrification ( 30 , 31 ) and warming vitrified-needle samples at 50°C resulted in DNA damage in testicular spermatozoa from adult cats ( 33 ). Despite slow freezing provided better results for elongated spermatids/spermatozoa than vitrification, the viability of these testicular cells was remarkably lower than fresh ones.…”

Section: Discussionmentioning

confidence: 99%

DNA integrity and viability of testicular cells from diverse wild species after slow freezing or vitrification

Peris-Frau

Benito-Blanco²,

Martínez‐Nevado³

et al. 2023

Front. Vet. Sci.

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Introduction and objectiveCryopreservation of testicular tissues offers new possibilities to protect endangered species, genetically valuable individuals or even the fertility potential of prepubertal individuals who have died unexpectedly. However, the use of this technique still remains a challenge. In this study, slow freezing and vitrification of testicular tissue was investigated to find out which cryopreservation method could better preserve the viability and DNA integrity of testicular germ cells in diverse wild species.MethodsTestes were obtained post-mortem from 18 artiodactyls (wild boar, roe deer, dwarf goat, mhor gazelle, European mouflon, African forest buffalo, Malayan tapir, dorcas gazelle, Iberian ibex, gnu, red river hog), 5 primates (colobus monkey, capuchin monkey, mandrill), 8 carnivores (gray wolf, Persian leopard, binturong, European mink, American black bear, suricata), and 2 rodents (Patagonian mara). The testicles belonged to adult individuals and were cut into small pieces and cryopreserved by needle immersed vitrification or uncontrolled slow freezing using a passive cooling device. After warming or thawing, testicular tissues were enzymatically digested and two germ cell types were differentiated based on their morphology: rounded cells (spermatogonia, spermatocytes, and early spermatids) and elongated cells (elongated spermatids and spermatozoa). Cell viability was assessed by SYBR-14/propidium iodide while DNA fragmentation by TUNEL assay with fluorescence microscope.Results and discussionOur preliminary results revealed that our uncontrolled slow freezing method better preserved the viability and DNA integrity of elongated cells than vitrification. Such trend was observed in all species, being significant in artiodactyls, carnivores, and primates. Similarly, the viability and DNA integrity of rounded cells was also better maintained in primates by uncontrolled slow freezing, while in carnivores, vitrification by needle immersion showed better results in this type of cells. In artiodactyls and rodents both techniques preserved the viability of rounded cells in a similar manner, although the DNA integrity of these cells was greater after needle immersed vitrification in artiodactyls.ConclusionsIn conclusion, the effectiveness of each cryopreservation method is affected by the phylogenetic diversity between species and cell type.

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Definition of protocols for cryopreservation and three-dimensional in vitro culture of prepubertal goat testicular tissue after histomorphological, ultrastructural, and functional analysis

Gomes,

Ñaupas,

Palomino

et al. 2023

Theriogenology

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Influence of Vitrification Device, Warming Protocol, and Subsequent In Vitro Culture on Structural Integrity of Testicular Fragments from Adult Domestic Cats

Cited by 4 publications

References 36 publications

Effect of Different Vitrification Techniques on Viability and Apoptotic Index of Domestic Cat Testicular Tissue Cells

Effect of Different Vitrification Techniques on Viability and Apoptotic Index of Domestic Cat Testicular Tissue Cells

DNA integrity and viability of testicular cells from diverse wild species after slow freezing or vitrification

Definition of protocols for cryopreservation and three-dimensional in vitro culture of prepubertal goat testicular tissue after histomorphological, ultrastructural, and functional analysis

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