Cryopreservation of caprine ovarian tissue using glycerol and ethylene glycol

Abstract

Cryopreservation of ovarian tissue may be a potential alternative for the conservation of genetically superior animals, including high milk-and meat-producing goat breeds. However, until now, no information was available concerning the cryopreservation of preantral follicles (PF) enclosed in caprine ovarian tissue. The objective of the present study was to evaluate the structural and ultrastructural characteristics of caprine PF after exposure to and cryopreservation of ovarian tissue in 1.5 and 3 M glycerol (GLY) and ethylene glycol (EG). At the slaughterhouse, each ovarian pair from five adult mixed breed goats was divided into nine fragments and randomly distributed into treatment groups. One fragment was immediately fixed for histological examination and ultrastructural analysis, after slaughter (control). Four of the ovarian fragments were equilibrated at 20 °C for 20 min in 1.8 ml of MEM containing 1.5 or 3 M GLY or EG for a toxicity test and the final four fragments were slowly frozen using these cryoprotectants at the concentrations above. After toxicity testing and freezing/thawing, the ovarian fragments were fixed for histological examination. Histological analysis showed that after toxicity testing and cryopreservation of the ovarian tissue in GLY or EG at both concentrations, the percentage of normal PF was significantly lower than controls. Ultrastructural analysis of PF frozen in 1.5 and 3 M GLY, as well as 3 M EG demonstrated that these follicles remained morphologically normal. In conclusion, we demonstrated cryopreservation of caprine PF in ovarian tissue.

Ultrastructural features of agouti (Dasyprocta aguti) preantral follicles cryopreserved using dimethyl sulfoxide, ethylene glycol and propanediol

“…In follicles included in ovarian tissue that were cryopreserved in the presence of EG, greater damage was observed in this study, manifested as reduced electron density and fewer organelles in the ooplasm. These findings differed from those in goats [2] and cattle [31], in which the cryoprotectant agent was highly efficient in maintaining follicle ultrastructure.…”

“…Similar changes were reported following exposure of ovarian tissue of sheep [19] and goats [2,3] to 1.5 M concentrations of the same cryoprotectants.…”

“…animals, including sheep [1], goats [2,3], primates [4], rodents [5], and humans [6]. Satisfactory results have been reported, especially when a 1.5 M concentration was used in slow-freezing protocols.…”

Preservation of bovine preantral follicle viability and ultra-structure after cooling and freezing of ovarian tissue

“…Most of the preservation protocols for ovarian tissue have been performed in presence of permeable cryoprotective substances such as glycerol (GLY), ethylene glycol (EG), propanediol (PROH) and dimethyl sulfoxide (DMSO), in concentrations ranging from 1.5 to 3.0 M. Besides their cryoprotective effect, these substances can also be toxic to cells, mainly during exposure period (Rodrigues et al, 2004a, Rodrigues et al, 2004band Santos et al, 2007b. Therefore, attention must be given to ovary transport before cryopreservation procedures.…”

Efecto de Tres Crioprotectores en la Criopreservación de Espermatozoides Epididimarios de Alpaca

“…Dentro de los crioprotectores, el glicerol (GL) ha sido extensamente usado en la criopreservación de varios tipos de células, incluyendo el esperma mamífero (Rasul et al, 2007). Por otro lado, el Dimetil Sulfóxido (DMSO) es un agente crioprotector prometedor para las células, pues su bajo peso molecular (78.13 g/mol) le permite atravesar la membrana celular de manera rápida (Ávila-Portillo et al, 2006); en tanto que el etilenglicol (EG) es un crioprotector que ha sido utilizado universalmente en la congelación de tejido ovárico y de embriones de muchas especies (Rodrigues et al, 2004), y con el semen de toro ejerce un menor efecto inhibitorio en la motilidad que el GL o el DMSO (Guthrie et al, 2002).…”