亮甲酚蓝染色对牛卵母细胞体外成熟和凋亡的影响

›› 2014, Vol. 41 ›› Issue (11): 171-174.

亮甲酚蓝染色对牛卵母细胞体外成熟和凋亡的影响

乔利敏¹, 乔富强², 姚华³, 雷莉辉¹, 王金秋¹, 关伟军⁴, 马月辉⁴

1. 北京农业职业学院畜牧兽医系, 北京 102442;
2. 北京农学院科技产业集团, 北京 102206;
3. 北京农学院动物科学技术学院, 北京 102206;
4. 中国农业科学院北京畜牧兽医研究所, 北京 100193

收稿日期:2014-03-24 出版日期:2014-11-20 发布日期:2014-12-06
通讯作者: 关伟军, 马月辉 E-mail:wjguan86@126.com;Yuehui.ma@263.net
作者简介:乔利敏(1972-),女, 山西人,硕士,讲师,研究方向:胚胎工程与动物繁殖。
基金资助:
北京农业职业学院博士基金项目:牛瘤胃补液器的设计制作与推广(XY-BS-12-04);北京农业职业学院技术研发与示范推广基金项目:奶牛细管冻精解冻后的保存与应用研究(XY-YF-14-20)。

Effect of BCB Staining on the IVM Apoptosis of Cow Oocytes

QIAO Li-min¹, QIAO Fu-qiang², YAO Hua³, LEI Li-hui¹, WANG Jin-qiu¹, GUAN Wei-jun⁴, MA Yue-hui⁴

1. Beijing Vocational College of Agriculture, Beijing 102442, China;
2. Group of Science Technology and Industry, Beijing University of Agriculture, Beijing 102206, China;
3. Institute of Animal Science and Veterinary Medicine, Beijing University of Agriculture, Beijing 102206, China;
4. Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China

Received:2014-03-24 Online:2014-11-20 Published:2014-12-06

摘要/Abstract

摘要： 为了探讨亮甲酚蓝(brilliant cresyl blue,BCB)对牛未成熟卵母细胞染色选择后对其体外成熟(IVM)和凋亡的影响,本研究在牛卵母细胞成熟培养前用26 μmol/L BCB染色90 min作为处理组,以未染色卵母细胞作为对照组;将处理组依照胞质的颜色分为蓝色组(BCB⁺)和无色组(BCB^-),成熟培养后检测卵子的发育能力。结果表明:①经BCB染色卵母细胞在成熟后,BCB⁺组卵母细胞成熟率(80.92%)较对照组(60.00%)和BCB^-组(51.61%)差异显著(P<0.05);②BCB⁺组的凋亡率(6.50%)较对照组(28.21%)、BCB^-组(39.06%)差异显著(P <0.05),随着卵母细胞发育潜能的升高其凋亡率逐渐降低。由此可见,以亮甲酚蓝染色为基础的牛卵丘-卵母细胞复合体的区分可以用来有效的选择更具发育活力的牛卵母细胞。

关键词: 牛; 卵母细胞; 亮甲酚蓝染色; 体外成熟

Abstract: The aim of this investigation was to study the effect of brilliant cresyl blue (BCB) staining on IVM apoptosis in cow, COCs without BCB staining was considered as control group, and COCs stained with BCB in 26 μmol/L for 90 min as treatment group before mature culture. Treated oocytes were divided into BCB^-(colourless cytoplasm) and BCB⁺(coloured cytoplasm). After IVM, to assess the developmental potential of the oocytes. The results showed that, the BCB⁺ oocytes had a significantly higher proportion of mature rate (80.92%) than control group (60.00%) and BCB^-oocytes (51.61%)(P<0.05);The BCB⁺ oocytes yielded a significantly lower apoptosis rate (6.50%) than that in control (28.21%) and BCB^- oocytes (39.06%)(P<0.05). These results suggested that the staining of cow cumulus-oocyte complexes with BCB before in vitro maturation could be used to select developmentally competent oocytes.

Key words: cow; oocytes; brilliant cresyl blue staining; IVM

中图分类号:

Q813.1

乔利敏, 乔富强, 姚华, 雷莉辉, 王金秋, 关伟军, 马月辉. 亮甲酚蓝染色对牛卵母细胞体外成熟和凋亡的影响[J]. , 2014, 41(11): 171-174.

QIAO Li-min, QIAO Fu-qiang, YAO Hua, LEI Li-hui, WANG Jin-qiu, GUAN Wei-jun, MA Yue-hui. Effect of BCB Staining on the IVM Apoptosis of Cow Oocytes[J]. , 2014, 41(11): 171-174.

参考文献

1 乔利敏. 亮甲酚蓝(BCB)染色法预测牛卵母细胞未来发育能力的研究[D].兰州:甘肃农业大学,2009.
2 乔利敏,李向臣,乔富强,等. 利用亮甲酚蓝染色法预测牛卵母细胞发育潜能的研究[J]. 畜牧兽医学报,2011,42(9):1249～1255.
3 Alm H, Torner H, Lohrke B, et al. Bovine blastocyst development rate in vitro is influenced by selection of oocytes by brillant cresyl blue staining before IVM as indicator for glucose-6-phosphate dehydrogenase activity[J]. Theriogenology, 2005, 63(8):2194～2205.
4 Coucouvanis E C, Sherwood S W, Carswell-Crumpton C, et al. Evidence that the mechanism of prenatal germ cell death in the mouse is apoptosis[J]. Experimental Cell Research, 1993, 209:238～247.
5 Gloria I P, Perez I, Xiao J T, et al. Fragmentation and death (a.k.a.apoptosis) of ovulated oocytes[J].Mol Hum Reprod,1999,5(5):414～420.
6 Gordon I. Recovering the bovine oocyte[A]. In Laboratory Production of Cattle Embryos,2nd Edition, Biotechnology in Agriculture No.27 CAB International[C].Cambridge University Press:Cambridge,2003.
7 Long C R, Dobrinsky J R, Garrett W M, et al. Dual labeling of the cytoskeleton and DNA strand breaks in porcine embryos produced in vivo and in vitro[J]. Mol Reprod,1998,51(1):59～65.
8 Roca J,Martinez E,Vazquez J M,et al. Selection of immature pig oocytes for homologous in vitro penetration assays with the brilliant cresyl blue test[J].Reprod Fertil Dev, 1998, 10:479～485.
9 Rodrigiguez-Gonzalez E,Lopez-Bejar M,Velilla E,et al.Selection of prepubertal goat oocytes using the brilliant cresyl blue test[J].Theriogenology,2002,57:1397~1409.
10 Urdaneta A,Jimenez A R,Izquierdo D,et al. Effect of the addition of glutathione and glucose to the culture medium on embryo development of IVM-IVF prepubertal goat oocytes[J].Zygote, 2003,11:131～138.
11 Urdaneta A, Jimenez Macedo A R, Izquierdo D, et al. Supplementation with cysteamine during maturation and embryoculture on embryo development of prepubertal goat oocytes selected by the brilliant cresyl blue test[J]. Zygote, 2003,11(4):347～354.
12 Wang C, Youle R J. The role of mitochondria in apoptosis[J]. Annual Review of Genetics,2009,43: 95～118.
13 Wassarman M. The mammalian ovum.The physiology of reproduction (2nd)[M].New York,USA:Raven Press,1988.
14 Yang J, Liu X, Bhalla K, et al. Prevention of apoptosis by Bcl-2: Release of cytochrome C from mitoehondria blocked[J]. Science, 1997, 275(5303):1129～1132.

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