Successful cryopreservation of whole sheep ovary by using DMSO-free cryoprotectant

Abstract

Purpose The study aims to assess the protective effects of dimethyl sulfoxide (DMSO)-free solution based on trehalose on the cryopreservation of a whole sheep ovary and evaluate its use as an efficient cryoprotectant. Method Twenty-one ovaries collected from 6-to 8-month-old non-pregnant female sheep were randomly distributed into three groups, namely, a fresh group, a DMSO-free group, and a DMSO group. The morphology, cell apoptosis (by hematoxylin and eosin (HE) staining and terminal deoxynucleotidyltransferasemediated dUTP nick-end labeling (TUNEL) assay), and mRNA transcript of Bcl-2-associated X protein (BAX) and cold inducible RNA-binding protein (CIRP) (by real-time PCR) of the thawed sheep ovaries and fresh controls were tested to establish a criterion for appraising the results of the cryopreservation.Results (i) The histological assessment indicated that the structure of the DMSO-free ovaries remained largely intact and comparable to those of the fresh control groups; whereas, significant damage was observed in the ovaries of the DMSO group (P<0.05). (ii) The TUNEL assay and mRNA transcript of the BAX assessment showed that the apoptosis parameter in the fresh group was the lowest among all the groups (P<0.05), and the parameter in the DMSO-free group was significantly lower than that in the DMSO group (P<0.05). (iii) The level of the CIRP transcripts increased the most in the DMSO-free group followed by the DMSO group and the fresh control group (P<0.05). Conclusions These results indicate that a DMSO-free cryoprotectant solution, especially a trehalose cryoprotectant, is an efficient cryoprotectant and has a beneficial effect on the cryopreservation of whole sheep ovaries.Capsule These results indicate that a DMSO-free and trehalose-containing cryoprotectant solution has a beneficial effect on the cryopreservation of intact sheep ovaries.Tianqi Du and Lan Chao contributed equally to this work.

Cold-inducible proteins CIRP and RBM3, a unique couple with activities far beyond the cold

“…CIRP is also associated with the enhancement of developmental competence of vitrified-warmed oocytes [ 66 ]. In addition, high CIRP expression level has been found in cryopreservation of reproductive organs, such as calf testis and sheep ovary [ 154 , 155 ].…”

Complete protection against cryodamage of cryopreserved whole bovine and human ovaries using DMSO as a cryoprotectant

“…In addition, the majority of children that were born after autotransplantation of ovarian cortical tissue were derived from tissue that was cryopreserved using DMSO as a cryoprotectant. Should, however, major issues concerning the use DMSO arise in the future, the use of less or even non-toxic alternatives such as trehalose should be further investigated [ 47 ].…”

Umbilical cord tissue cryopreservation: a short review

“…Moreover, it is impossible to remove it by washing, even with the use of a specialized system for washing cell transplants. The reasonable alternative is a medium supplemented with a cocktail of non-penetrating cryoprotectants (e.g., glucose, sucrose, galactose, or trehalose) and intracellular cryoprotectants (e.g., ethylene glycol, propylene glycol (1,2-propanediol), glycerol, formamide, methanol, and butanediol) [ 39 – 41 ]. According to pilot studies, this cocktail provides better tissue preservation than DMSO [ 41 , 42 ].…”