Urrego R, Herrera-Puerta E, Chavarria N A, Camargo O, Wrenzycki C, Rodriguez-Osorio N
Grupo CENTAURO, Universidad de Antioquia, Medellín, Colombia; Grupo INCA-CES, Facultad de Medicina Veterinaria y Zootecnia, Universidad CES, Medellín, Colombia.
Grupo INCA-CES, Facultad de Medicina Veterinaria y Zootecnia, Universidad CES, Medellín, Colombia; Grupo Biología CES-EIA, Universidad CES, Medellín, Colombia.
Theriogenology. 2015 Apr 15;83(7):1179-87. doi: 10.1016/j.theriogenology.2014.12.024. Epub 2014 Dec 30.
The ability of bovine embryos to develop to the blastocyst stage and to implant and generate healthy offspring depends greatly on the competence of the oocyte. Oocyte competence is attributed to its close communication with the follicular environment and to its capacity to synthesize and store substantial amounts of messenger RNA. Higher developmental competence of bovine oocytes has been associated with both the expression of a cohort of developmental genes and the concentration of sex steroids in the follicular fluid. The aim of this study was to identify differences in the expression of FST in cumulus cells and OCT-4 and MATER in oocytes and the influence of the follicular progesterone and follicular estrogen concentration on the competence of bovine oocytes retrieved 30 minutes or 4 hours after slaughter. Cumulus-oocyte complexes (COCs) were left in postmortem ovaries for 30 minutes (group I) or 4 hours (group II) at 30 °C. Aspirated oocytes were then subjected to IVM, IVF, and IVC or were evaluated for MATER and OCT-4 messenger RNA abundance by quantitative real-time polymerase chain reaction. Total RNA was isolated from pools of 100 oocytes for each experimental replicate. Progesterone and estradiol concentration in follicular fluid was evaluated by immunoassay using an IMMULITE 2000 analyzer. Three repeats of in vitro embryo production were performed with a total of 455 (group I) and 470 (group II) COCs. There were no significant differences between the cleavage rates (72 hours postinsemination [hpi]) between both groups (63.5% vs. 69.1%). However, blastocyst (168 hpi) and hatching (216 hpi) rates were higher (P < 0.05) in group II compared with those of group I (21.3% vs. 30.7% and 27.6% vs. 51.5%, respectively). Group II oocytes exhibited the highest MATER and OCT-4 abundance (P < 0.05). Follicular estradiol concentration was not different between both the groups, whereas the progesterone concentration was lower (P ≤ 0.05) in group II follicles. These results indicate that retrieving COCs 4 hours after slaughter could increase bovine in vitro developmental competence, which is linked to higher levels of oocyte MATER and OCT-4 transcripts and lower follicular progesterone concentration. Moreover, the results of the present study contribute to the identification of factors involved in the developmental competence of immature oocytes.
牛胚胎发育至囊胚阶段、着床并产生健康后代的能力在很大程度上取决于卵母细胞的质量。卵母细胞的质量归因于其与卵泡环境的密切交流以及合成和储存大量信使核糖核酸的能力。牛卵母细胞较高的发育能力与一组发育基因的表达以及卵泡液中性类固醇的浓度有关。本研究的目的是确定卵丘细胞中卵泡抑素(FST)的表达以及卵母细胞中八聚体结合转录因子4(OCT-4)和母源表达的胚胎停滞相关蛋白(MATER)的表达差异,以及卵泡孕酮和卵泡雌激素浓度对屠宰后30分钟或4小时采集的牛卵母细胞质量的影响。卵丘-卵母细胞复合体(COCs)在30℃下于死后卵巢中放置30分钟(I组)或4小时(II组)。然后将吸出的卵母细胞进行体外成熟(IVM)、体外受精(IVF)和体外培养(IVC),或通过定量实时聚合酶链反应评估MATER和OCT-4信使核糖核酸丰度。每次实验重复从100个卵母细胞的集合中分离总核糖核酸。使用IMMULITE 2000分析仪通过免疫测定法评估卵泡液中的孕酮和雌二醇浓度。进行了三次体外胚胎生产重复,I组共有455个COCs,II组共有470个COCs。两组之间的卵裂率(授精后72小时[hpi])无显著差异(63.5%对69.1%)。然而,与I组相比,II组的囊胚率(168 hpi)和孵化率(216 hpi)更高(P<0.05)(分别为21.3%对30.7%和27.6%对51.5%)。II组卵母细胞表现出最高的MATER和OCT-4丰度(P<0.05)。两组之间的卵泡雌二醇浓度无差异,而II组卵泡中的孕酮浓度较低(P≤0.05)。这些结果表明,屠宰后4小时采集COCs可提高牛的体外发育能力,这与卵母细胞中较高水平的MATER和OCT-4转录本以及较低的卵泡孕酮浓度有关。此外,本研究结果有助于确定与未成熟卵母细胞发育能力相关的因素。