High Hydrostatic Pressure (HHP) pre-treatment has been reported to increase cryosurvival of mouse and bovine oocytes and embryos. This treatment induces a sublethal stress that stimulates the expression of specific proteins which improves stress tolerance, thereby cross‐protecting the cells from subsequent stress conditions like cryopreservation. No information are available about HHP pre-treatment effect on ovarian tissue cryopreservation. Aim of the present experiments was to evaluate the impact of HHP treatment prior to freezing of ovarian fragment. Sheep ovaries were collected at the local slaughterhouse. Cortex was carefully dissected to obtain small pieces (5×5×2 mm), and exposed to 0; 200 and 400 bar for 30 min, 1 h or 2 h. Treatments were carried out at room temperature and 37°C in a HHP Machine 1400 (CryoInnovation). Ovarian fragments were then equilibrated at 38,5° C for 1 hour in 199H medium + 10%FCS. They were separated in two groups; one was frozen by slow freezing with a Planer The Kryo 560M, while the other was stored at -80°C for gene expression analysis. Ovarian cortex fragments not exposed to HHP were stored at -80°C as control. To evaluate HHP stress related effects, Hsp70, NFAT5, TauT and P53 gene expression levels were quantified by semi-quantitative PCR. The relative abundance of mRNA for the target genes was evaluated by ANOVA (P<0.05). Morphology was assessed by ematoxilin/eosin staining using a NIKON EclipseE600. No changes in transcript levels were observed for the genes considered, both before and after cryopreservation following all the experimental treatments. Microscopic evaluation also demonstrated no obvious variations of ovarian tissue morphology in the different experimental groups. The results obtained indicate that HHP at the conditions tested is unlikely to increase stress response molecule expression nor to induce morphological changes in cryopreserved ovarian tissues. Supported by AIRC IG 10376 and by Carraresi Foundation

Effect of High Hydrostatic Pressure on ovarian tissue fragments cryopreservation / S. Maffei, G. Pennarossa, T. Brevini, F. Gandolfi. ((Intervento presentato al 4. convegno Maternal Interactions with Gametes and Embryos : Gemini General Conference : COST Action FA0702 tenutosi a Gijon nel 2011.

Effect of High Hydrostatic Pressure on ovarian tissue fragments cryopreservation

S. Maffei
Primo
;
G. Pennarossa
Secondo
;
T. Brevini;F. Gandolfi
Ultimo
2011

Abstract

High Hydrostatic Pressure (HHP) pre-treatment has been reported to increase cryosurvival of mouse and bovine oocytes and embryos. This treatment induces a sublethal stress that stimulates the expression of specific proteins which improves stress tolerance, thereby cross‐protecting the cells from subsequent stress conditions like cryopreservation. No information are available about HHP pre-treatment effect on ovarian tissue cryopreservation. Aim of the present experiments was to evaluate the impact of HHP treatment prior to freezing of ovarian fragment. Sheep ovaries were collected at the local slaughterhouse. Cortex was carefully dissected to obtain small pieces (5×5×2 mm), and exposed to 0; 200 and 400 bar for 30 min, 1 h or 2 h. Treatments were carried out at room temperature and 37°C in a HHP Machine 1400 (CryoInnovation). Ovarian fragments were then equilibrated at 38,5° C for 1 hour in 199H medium + 10%FCS. They were separated in two groups; one was frozen by slow freezing with a Planer The Kryo 560M, while the other was stored at -80°C for gene expression analysis. Ovarian cortex fragments not exposed to HHP were stored at -80°C as control. To evaluate HHP stress related effects, Hsp70, NFAT5, TauT and P53 gene expression levels were quantified by semi-quantitative PCR. The relative abundance of mRNA for the target genes was evaluated by ANOVA (P<0.05). Morphology was assessed by ematoxilin/eosin staining using a NIKON EclipseE600. No changes in transcript levels were observed for the genes considered, both before and after cryopreservation following all the experimental treatments. Microscopic evaluation also demonstrated no obvious variations of ovarian tissue morphology in the different experimental groups. The results obtained indicate that HHP at the conditions tested is unlikely to increase stress response molecule expression nor to induce morphological changes in cryopreserved ovarian tissues. Supported by AIRC IG 10376 and by Carraresi Foundation
2011
Settore VET/01 - Anatomia degli Animali Domestici
Servicio Regional de Investigación y Desarrollo Agroalimentario
SERIDA
Effect of High Hydrostatic Pressure on ovarian tissue fragments cryopreservation / S. Maffei, G. Pennarossa, T. Brevini, F. Gandolfi. ((Intervento presentato al 4. convegno Maternal Interactions with Gametes and Embryos : Gemini General Conference : COST Action FA0702 tenutosi a Gijon nel 2011.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/563252
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