Presence of sucrose in the vitrification solution and exposure for longer periods of time improve post‐warming follicle integrity in cat ovarian tissues

Abstract

ContentsOvarian tissue cryopreservation followed by tissue culture is a promising approach to preserving the fertility of biomedical models and endangered species. The objective of this study was to investigate the impact of exposure time to vitrification solution and presence of sucrose using different exposure temperatures and base media on intra-ovarian follicle integrity. Peripubertal ovarian cortical pieces were obtained by isolating the cortex and dissecting it into 1 × 1 × 0.2 mm 3 pieces. The cortical pieces were then exposed to equilibration solution and then vitrification solutions (VS) in one of the conditions mentioned above, plunged directly into liquid nitrogen and stored for ≥24 hr in liquid nitrogen. After thawing, the cortical pieces were cultured in vitro for 0, 1 or 7 days to determine the follicle integrity (through histological assessment) and the ability of the tissue to recover from cryoinjury. Fresh controls maintained a constant level of normal morphology (>60% of the total follicles) throughout the culture period. Cortical pieces exposed to VS with sucrose for 10 min had the highest percentage of normal follicles (approximately 20% after 7 days of culture) throughout the culture period. Other conditions using different base medium, lower exposure temperatures or different thawing methods did not improve the follicle integrity. This protocol provides a solid foundation on which to optimize ovarian tissue cryopreservation in the domestic cat and to investigate the molecular effects of vitrification.

Influence of different warming temperatures on the vitrified testicular fragments from pre‐pubertal cat

“…A rapid warming process can increase tissue survival and reanimation (Lima et al., 2018; Seki et al., 2014) causing a positive impact in structural and functional properties due to the short exposure and high temperature during warming (Mouttham and Comizzoli, 2017; Lima et al., 2018). Avoiding formation of ice crystals in the post‐cryopreservation warming process is very important to preserve the biological integrity and quality of tissues.…”

Zinc Chloride Supplemented during Ovarian Tissue Vitrification Improves In Vitro Follicle Development and Fertilization in Pigs

“…There is a growing interest and need for cryopreservation techniques to be improved so that ovarian tissue and oocytes can be preserved in biomedical models and endangered species (Mouttham and Comizzoli 2016). Cryopreservation in the form of vitrification is a desirable type of preservation compared to other techniques, due to the ability of cryopreservation to preserve large numbers of oocytes at once.…”

Initial response of ovarian tissue transcriptome to vitrification or microwave-assisted dehydration in the domestic cat model

“…Prepubertal ovarian tissues have been used due to their high reserves of primordial and primary follicles and homogeneity compared to adult counterparts [ 19 ]. Our vitrification protocol has been optimized so there is no significant differences in percentage of morphologically normal follicles between vitrified/warmed and fresh cortical tissue, and only mild decrease in follicular viability and RNA synthesis [ 8 , 20 ]. Our laboratory also reported initial steps in the development of microwave-assisted dehydration technique for cat germinal vesicle [ 21 , 22 ], sperm cells [ 23 ], and most recently ovarian tissue [ 18 ], relying on intracellular delivery of trehalose as protection from dehydration/rehydration damage.…”