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| GDF-9对牦牛COCs体外成熟培养及其相关基因mRNA表达的影响 |
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王玉恒1,2,3, 徐业芬1,2, 索朗斯珠1,2, 强巴央宗1, 郭敏4, 程玲华4, 牛家强1,2, 贡嘎1,2, 王刚1,2, 孔庆辉1,2
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| (1.西藏农牧学院 动物科学学院, 西藏 林芝 860000;2.西藏农牧学院 西藏高原动物疫病研究自治区高校重点实验室, 西藏 林芝 860000;3.西藏自治区兽医生物药品制造厂, 拉萨 850000;4.中国农业大学 动物科技学院, 北京 100193) |
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| 摘要: |
| 为探明生长分化因子-9（Growth differentiation factor 9，GDF-9）对牦牛卵丘-卵母细胞复合体（Cumulus-oocyte complexes，COCs）体外培养的卵母细胞成熟率及相关基因mRNA表达水平的影响：采取健康牦牛的卵巢，分离COCs进行体外成熟培养；在牦牛COCs成熟培养液中添加终浓度分别为0、200、400、600 ng/mL的GDF-9，统计卵母细胞成熟率；采用Real-time PCR方法检测GDF-9处理的COCs中Smad信号通路下游分子Smad 4以及卵丘扩展相关基因（HAS 2、PTX 3、PTGS 2）的mRNA表达水平。结果表明：添加0、200、400、600 ng/mL GDF-9的COCs成熟率差异不显著（P>0.05）。Smad 4和卵丘扩展相关基因（HAS 2、PTX 3、PTGS 2）的mRNA表达量随着GDF-9浓度升高呈上升趋势，其中600 ng/mL GDF-9处理组的Smad 4和PTX 3 mRNA表达量最高，与其他处理组差异显著（P<0.05）； 600 ng/mL GDF-9处理组的HAS 2、PTGS 2 mRNA表达量与400 ng/mL GDF-9处理组差异不显著（P>0.05），与其他实验组差异显著（P<0.05）。研究发现，在牦牛COCs体外培养过程中，添加不同浓度的GDF-9对牦牛卵母细胞成熟率的影响差异不显著；GDF-9显著提高其信号通路下游分子Smad 4基因以及卵丘细胞扩展相关基因（HAS 2、PTX 3、PTGS 2）的mRNA表达，说明600 ng/mL GDF-9对牦牛体外培养过程中卵丘扩展有作用，作用机制可能与激活Smad信号通路有关。 |
| 关键词: 牦牛 卵丘卵母细胞复合体 体外成熟 生长分化因子-9 |
| DOI：10.11841/j.issn.1007-4333.2018.11.09 |
| 投稿时间：2018-02-18 |
| 基金项目:西藏自治区科技厅厅校联合基金项目（XZ2017ZRG-25）；国家自然科学基金项目（31460604）；中西部高校综合实力提升计划（502000105）；国家肉牛牦牛产业体系项目（CARS-37） |
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| Effect of GDF-9 on in vitro maturation culture and the expression of related genes in yak COCs |
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WANG Yuheng1,2,3, XU Yefen1,2, Suolangsizhu1,2, Changbayangzong1, GUO Min4, CHENG Linghua4, NIU Jiaqiang1,2, GONG Ga1,2, WANG Gang1,2, KONG Qinhui1,2
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| (1.Department of Animal Science, Tibet Agricultural and Animal Husbandry College, Linzhi 860000, China;2.Provincial Key Laboratory of Tibet Plateau Animal Epidemic Disease Research, Tibet Agricultural and Animal Husbandry College, Linzhi 860000, China;3.Tibet Veterinary Biological Pharmaceutical Factory, Lhasa 850000, China;4.College of Animal Science and Technology, China Agricultural University, Beijing 100193, China) |
| Abstract: |
| The aim of this study was to investigate the effects of growth differentiation factor 9 (GDF-9)on culture and the related gene mRNA expression of yak cumulus-oocyte complexes (COCs) in vitro. The healthy yak ovaries were collected and COCs were isolated for maturation culture in vitro. GDF-9 with final concentrations of 0, 200, 400, and 600 ng/mL was separately added to the maturation medium for yak COCs culture in vitro, and the oocyte maturation rate was separately counted. The mRNA expression levels of Smad 4 and cumulus cell extension related genes (HAS 2, PTX 3, PTGS 2) in COCs were detected by Real-time PCR. The results showed that there was no significant difference in the maturation rates of COCs treated with 0, 200, 400 and 600 ng/mL GDF-9 (P>0.05). The mRNA expression of Smad 4 gene and extension related genes (HAS 2, PTX 3, PTGS 2) were increased with the increasing of GDF-9 concentration. The mRNA expression of Smad 4 and PTX 3 in group treated with 600 ng/mL GDF-9 was the highest, which was significantly different from other treatments (P<0.05). Compared with group treated with 400 ng/mL GDF-9, the mRNA expression of HAS 2 and PTGS 2 in 600 ng/mL GDF-9 group had no significant difference (P>0.05), which was significantly different from other experimental groups (P<0.05). It was found that the maturation rates of yak COCs treated with GDF-9 showed no significant differences, but GDF-9 significantly increased the expression of Smad 4which was the downstream molecule gene of GDF-9 in the cell signaling pathway. GDF-9 also increased the mRNA expression of cumulus cell extension genes (HAS 2, PTX 3, PTGS 2). These results suggested that GDF-9 might play an important role in the extension of cumulus oocytes during culturing yak COCs in vitro, and the mechanism might be related to the activation of Smad signaling pathway, but its mechanisms remained to be further studied. |
| Key words: yak cumulus-oocyte complexes in vitro maturation GDF-9 |
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