182 MATURATION OF BOVINE CUMULUS-OOCYTE COMPLEXES WITH FOLLICLE FLUID VARYING IN ESTRADIOL CONTENT AFFECTS CUMULUS CELL EXPANSION WITHOUT AFFECTING SUBSEQUENT EMBRYO DEVELOPMENT IN VITRO
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The objective of this work was to determine how characteristics of bovine follicle fluid (FF; especially oestradiol content) affect cumulus cell expansion and oocyte competence. In the first study, FF was collected from abattoir-derived ovaries and pooled separately for large follicles (10mm) and small follicles (3mm). A portion of the FF from each category was charcoal stripped. These 4 types of FF were then used as the primary ingredient (75% vol/vol) in oocyte maturation media. A separate control group lacking FF but containing BSA was included to monitor potential impacts of protein on outcomes (control; without FF). Some of the cumulus-oocyte complexes (COC; n=250) were matured in individual drops for analysis of cumulus expansion (photographed and measured at 0 and 21h of maturation). Other COC (n=770) were matured in groups of 12 to 25 in the previously described media, and then subjected to IVF procedures. Cleavage rates were recorded on Day 3, and blastocyst rates were recorded on Day 8 post-fertilization. Cumulus cell expansion was greatest when COC were matured in medium containing FF from large follicles, wherein it even exceeded the controls (P<0.02). Maturation in FF from small follicles resulted in cumulus expansion that was intermediate between large and control. Maturation in charcoal-stripped FF severely restricted cumulus cell expansion (P<0.05) compared with those matured in untreated FF. Despite the observed improvement in cumulus cell expansion, COC that had been matured in media containing FF were less likely to cleave (P<0.05) and also less likely to develop to the blastocyst stage (P<0.01) than those matured in control medium. Cleavage and blastocyst rates did not differ among any of the maturation media containing FF. In the second study, oestrous cycles of 9 crossbred cows were synchronized and FF samples were collected 36 to 42h after prostaglandin F2 injection. Samples from individual cows were categorized as having high oestradiol (>800,000pgmL1; H) or low oestradiol concentrations (<800,000pgmL1; L). The FF was retained for use in in vitro experiments, where it was added to maturation media (20% vol/vol). Cumulus-oocyte complexes (n=1,775) were randomly distributed into treatments across 12in vitro maturation/fertilization replicates (H and L, balanced within replicate; 4 replicates/cow). Each replicate included the following 3 control groups: maturation medium containing BSA without FF, maturation medium without BSA with abattoir-derived FF, and maturation medium without BSA and without FF. The COC were matured in their assigned medium for 21h, and then all COC were subjected to IVF procedures. Cleavage rates were recorded on Day 3, and blastocyst rates were recorded on Day 7 and 8 post-fertilization. Oestradiol content of the FF (H v. L) did not affect oocyte cleavage nor blastocyst rates on Day 7 or 8. The results of these studies indicate that although FF improves cumulus cell expansion during maturation in vitro, it does not result in higher rates of cleavage or blastocyst development regardless of oestradiol content.
