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Closed-system solid surface vitrification versus slow programmable freezing of mouse 2-cell embryos.

ABSTRACT: PURPOSE: To compare closed-system solid surface vitrification with slow freezing. METHODS: Mouse 2-cell embryos (n = 348) were divided into vitrification, slow freezing and non-frozen groups. For vitrification, embryos were exposed to 10% ethylene glycol (EG), 10% dimethylsulfoxide (DMSO) and 10% fetal bovine serum (FBS) in phosphate-buffered saline (PBS) for 10 min, then transferred into 17.5% EG, 17.5% DMSO, 0.25 M trehalose and 10% FBS in PBS. They were placed on hemi-straws and inserted into 0.5 ml straws inside a previously cooled aluminum cylinder. Slow freezing was done in straws by the conventional method. RESULTS: Vitrified embryos had significantly higher survival, further cleavage and blastocyst formation rates than those in the slow freezing group (p < 0.001) and were comparable to controls. Blastocysts in the vitrification and control groups had significantly more cells than those in the slow freezing group (p < 0.05). CONCLUSIONS: Closed-system vitrification was more effective than conventional slow freezing.

SUBMITTER: Vutyavanich T

PROVIDER: S-EPMC2719071 | biostudies-other | 2009 May

REPOSITORIES: biostudies-other

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Project description:Cryopreservation is known for its marked deleterious effects on embryonic health. Bovine compact morulae were vitrified or slow-frozen, and post-warm morulae were cultured to the expanded blastocyst stage. Blastocysts developed from vitrified and slow-frozen morulae were subjected to microarray analysis and compared with blastocysts developed from unfrozen control morulae for differential gene expression. Morula to blastocyst conversion rate was higher (P < 0.05) in control (72%) and vitrified (77%) than in slow-frozen (34%) morulae. Total 20 genes were upregulated and 44 genes were downregulated in blastocysts developed from vitrified morulae (fold change ? ± 2, P < 0.05) in comparison with blastocysts developed from control morulae. In blastocysts developed from slow-frozen morulae, 102 genes were upregulated and 63 genes were downregulated (fold change ? ± 1.5, P < 0.05). Blastocysts developed from vitrified morulae exhibited significant changes in gene expression mainly involving embryo implantation (PTGS2, CALB1), lipid peroxidation and reactive oxygen species generation (HSD3B1, AKR1B1, APOA1) and cell differentiation (KRT19, CLDN23). However, blastocysts developed from slow-frozen morulae showed changes in the expression of genes related to cell signaling (SPP1), cell structure and differentiation (DCLK2, JAM2 and VIM), and lipid metabolism (PLA2R1 and SMPD3). In silico comparison between blastocysts developed form vitrified and slow-frozen morulae revealed similar changes in gene expression as between blastocysts developed from vitrified and control morulae. In conclusion, blastocysts developed form vitrified morulae demonstrated better post-warming survival than blastocysts developed from slow-frozen morulae but their gene expression related to lipid metabolism, steroidogenesis, cell differentiation and placentation changed significantly (? 2 fold). Slow freezing method killed more morulae than vitrification but those which survived up to blastocyst stage did not express ? 2 fold change in their gene expression as compared with blastocysts from control morulae.

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Closed-system solid surface vitrification versus slow programmable freezing of mouse 2-cell embryos.

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