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Minimal volume vitrification of epididymal spermatozoa results in successful in vitro fertilization and embryo development in mice

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Asian Journal of Andrology 2017年第1期19卷 107-112页

作者： fabrizzio horta Hamida Alzobi Sutthipat Jitanantawittaya Sally Catt Penny Chen Mulyoto Pangestu Peter Temple-Smith Education Program in Reproduction and Development Department Obstetrics and Gynaecology Monash University Clayton VlC 3168 Australia Department of Obstetrics and Gynaecology Unit of Reproductive Medicine Clinica Las Condes Lo Fentecilla 441 Las Condes Santiago Region Metropolitana 7550000 Chile.

This study compared three cryopreservation protocols on sperm functions, IVF outcomes, and embryo development. Epididymal spermatozoa cryopreserved using slow-cooling （18% w/v raffinose, RS-C） were compared with spermatozoa vitrified using 0.25 M sucrose （SV） or 18% w/v raffinose （RV）. The motility, vitality, and DNA damage （TUNEL assay） of fresh control （FC） spermatozoa were compared with post-thawed or warmed RS-C, RV, and SV samples. Mouse oocytes （n = 267） were randomly assigned into three groups for insemination： RV （n = 102）, RS-C （n = 86）, and FC （n = 79）. The number and the proportion of two-cell embryos and blastocysts from each treatment were assessed. Sperm motility （P 〈 0.01） and vitality （P 〈 0.05） were significantly reduced after vitrification compared with slow-cooled spermatozoa. However, DNA fragmentation was significantly reduced in spermatozoa vitrified using sucrose （15 - 1.8% [SV] vs 26 - 2.8% [RV] and 27 - 1.2% [RS-C]; P 〈 0.01）. Although the number of two-cell embryos produced by RS-C, RV, and FC spermatozoa was not significantly different, the number of blastocysts produced from two-cell embryos using RV spermatozoa was significantly higher than FC spermatozoa （P = 0.0053）. This simple, small volume vitrification protocol and standard insemination method allows successful embryo production from small numbers of epididymal spermatozoa and may be applied clinically to circumvent the need for ICSI, which has the disadvantage of bypassing sperm selection.

关键词： cryopreservation epididymal sperm in vitro fertilization/intracytoplasmic sperm injection reproductive techniques sperm DNA damage vitrification

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