Fathi Mohamed, Ashry Mohamed, Salama Ali, Badr Magdy R
Department of Theriogenology,Faculty of Veterinary Medicine,Cairo University,Giza,Egypt.
B275 Anthony Hall,department of Animal Science,Michigan State University,East Lansing,MI 48824,USA.
Zygote. 2017 Aug;25(4):529-536. doi: 10.1017/S0967199417000387. Epub 2017 Jul 11.
The objectives of the present studies were to investigate the developmental capacity of dromedary camel oocytes selected by brilliant cresyl blue (BCB) staining and to investigate the expression of select transcripts in germinal vesicle (GV) stage oocytes. These transcripts included BMP15 and GDF9 as important transcripts for folliculogenesis and oocyte development, Zar1 and Mater as maternal transcripts required for embryonic development, Cyclin B1 and CDK1 as cell cycle regulators and Oct4 and STAT3 as transcription factors. Dromedary camel oocytes were retrieved from ovaries collected at a local slaughterhouse. After exposure to BCB staining, cumulus-oocyte complexes (COCs) from BCB+, BCB- and control (selected based on morphological criteria) groups were subjected to in vitro maturation, in vitro fertilization and in vitro culture. For gene expression studies, after BCB staining cumulus cells were stripped off and the completely denuded GV stage oocytes were used for RT-PCR analysis of selected transcripts. BCB+ oocytes showed higher maturation, and fertilization rates compared with BCB- and control groups. Indices of early embryonic development, namely, cleavage at 48 hours post insemination (hpi), and development to morula at day 5 and day 7 blastocyst rates were also significantly higher in the BCB+ group. RT-PCR revealed a higher expression of BMP15, GDF9, Zar1, Mater, Cyclin B1, CDK1, OCT4 and STAT3 in good quality oocytes that stained positively for BCB (BCB+). Collectively, results provide novel information about the use of BCB screening for selecting good quality oocytes to improve in vitro embryo production in the dromedary camel.
本研究的目的是调查经灿烂甲酚蓝(BCB)染色筛选的单峰骆驼卵母细胞的发育能力,并研究生发泡(GV)期卵母细胞中特定转录本的表达。这些转录本包括作为卵泡发生和卵母细胞发育重要转录本的BMP15和GDF9、作为胚胎发育所需母体转录本的Zar1和Mater、作为细胞周期调节因子的细胞周期蛋白B1和细胞周期蛋白依赖性激酶1以及作为转录因子的Oct4和STAT3。单峰骆驼卵母细胞从当地屠宰场收集的卵巢中获取。经BCB染色后,将来自BCB+、BCB-和对照组(根据形态学标准选择)的卵丘-卵母细胞复合体(COC)进行体外成熟、体外受精和体外培养。对于基因表达研究,BCB染色后去除卵丘细胞,将完全裸化的GV期卵母细胞用于所选转录本的逆转录聚合酶链反应(RT-PCR)分析。与BCB-组和对照组相比,BCB+卵母细胞显示出更高的成熟率和受精率。BCB+组早期胚胎发育指标,即授精后48小时(hpi)的卵裂率以及第5天的桑椹胚发育率和第7天的囊胚率也显著更高。RT-PCR显示,在BCB染色呈阳性的优质卵母细胞(BCB+)中,BMP15、GDF9、Zar1、Mater、细胞周期蛋白B1、细胞周期蛋白依赖性激酶1、OCT4和STAT3的表达更高。总体而言,研究结果提供了关于使用BCB筛选来选择优质卵母细胞以提高单峰骆驼体外胚胎生产的新信息。