Stimulatory effect of ROCK inhibitor on revivability of in vitro-produced bovine blastocysts after vitrification
نویسندگان
چکیده
Inhibition of Rho-associated coiled-coil kinase (ROCK) activity has been reported to promote recovery and growth of frozen-thawed human embryonic stem cells. Since in vitro-produced and/or manipulated bovine embryos are sensitive to cryopreservation, the present study was conducted to investigate whether the presence of ROCK inhibitor (Y-27632) in post-thaw culture medium improve the revivability of bovine blastocysts produced in vitro and vitrified by minimum volume cooling procedure. Expanding or fully-expanded blastocysts were harvested 7 days after in vitro fertilization, and vitrified in the presence of 15% ethylene glycol, 15% DMSO and 0.5 M sucrose using Cryotop as cryodevice. When the post-warm blastocysts were cultured in mSOF medium supplemented with 10 μM Y-27632, the survival rate (94.9 ± 2.4%), assessed by re-expansion of blastocoel at 24 h of the culture, was higher than that cultured without Y-27632 (78.0 ± 6.0%; P < 0.05). Conversely, hatching rate and mean total cell number of surviving embryos at 48 h of the culture were comparable regardless of the Y-27632 supplementation (62.8 ± 11.1 vs 59.6 ± 9.4%, and 135.2 ± 13.1 vs 146.7 ± 13.3, respectively). In non-vitrified in vitro-produced blastocysts, hatching rate on Day-9 was improved by the presence of Y-27632 (91.7 ± 3.8 vs 54.7 ± 8.9%; P < 0.05), but the increase in mean total cell number of blastocysts was not significant (230.0 ± 23.0 vs 191.2 ± 22.2; P = 0.23). In an additional experiment, the Y-27632 was supplemented to culture medium on either Day-0, -2 or -4 until Day-8, resulting in no increase in the blastocyst yield (7.5 ± 2.1 to 36.2 ± 3.2% vs 28.6 ± 6.9%). In conclusion, the supplementation of ROCK inhibitor to post-thaw culture medium improved the revivability of in vitro-produced bovine blastocysts after vitrification and warming.
منابع مشابه
Effect of Culture System on Developmental Competence, Cryosurviva and DNA-Fragmentation of In Vitro Bovine Blastocysts
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