性控分离精子凋亡程度对牛体外受精的影响

doi:10.11843/j.issn.0366-6964.2014.06.006

畜牧兽医学报 ›› 2014, Vol. 45 ›› Issue (6): 893-900.doi: 10.11843/j.issn.0366-6964.2014.06.006

性控分离精子凋亡程度对牛体外受精的影响

郝海生¹，任晶晶¹，赵学明¹，王玲玲²，杜卫华¹，王长法²，朱化彬^1*

(1.中国农业科学院北京畜牧兽医研究所，北京 100193; 2.山东奥克斯生物技术有限公司，济南 250000)

收稿日期:2013-12-17 出版日期:2014-06-23 发布日期:2014-06-23
通讯作者: 朱化彬，博士，研究员，E-mail:zhuhuabin@iascaas.net.cn
作者简介:郝海生（1977-），男，北京市人，助理研究员，主要从事家畜胚胎工程研究，E-mail: haohaisheng@caas.cn
基金资助:
现代农业产业技术体系建设专项资金（CARS-37）;十二五科技支撑项目（2011BAD19B04-6）;引智项目（Y20110326005）

Effects of the Degree of Apoptosis in Sexed Sperms on Bovine Fertilization in vitro

HAO Hai-sheng¹，REN Jing-jing¹，ZHAO Xue-ming¹，WANG Ling-ling²，DU Wei-hua¹，WANG Chang-fa²，ZHU Hua-bin^1*

(1.Institute of Animal Science，Chinese Academy of Agricultural Sciences，Beijing 100193, China;2.Shandong Auox Biotechnology Company，Jinan 250000， China)

Received:2013-12-17 Online:2014-06-23 Published:2014-06-23

摘要/Abstract

摘要：

采用Annexin-V、JC-1和TUNEL对A、B、C、D 4头种公牛性控X冻精进行了凋亡分析，并对凋亡分析结果与性控精液体外受精卵裂率和囊胚率的关系进行了相关分析。结果表明：（1）公牛A的活精子（AN^-•PI^-）百分率（89.30%±3.04%）高于其他公牛（(74.39%±3.24%)~(81.17%±4.25%)），差异显著（P<0.05），而其DNA片段化精子百分率（12.24%±1.25%）低于其他公牛（(19.09%±1.27%)~(27.56%±3.17%)），差异显著（P<0.05）；（2）公牛A 的X冻精体外受精（IVF）卵裂率、囊胚率（(76.82%±3.00%)~(20.45%±1.44%)）均高于其他3头公牛（(22.00%±6.85%)~(31.88%±6.69%))，(3.33%±2.67%)~(4.00%±3.26%)），差异显著（P<0.05）；（3）活精子百分率与其IVF卵裂率和囊胚率呈正相关，而DNA片段化百分率与之呈负相关。综上表明：Annexin-V分析能够准确地预测牛X性控冻精的体外受精效率，这对于促进牛性控体外胚胎的生产效率有着重要意义。

关键词: 体外受精, 不同种公牛, Annexin-V, JC-1, TUNEL, 精子凋亡, 精液质量

Abstract:

The present study utilized three staining assays (Annexin-V，mitochondrial membrane potential (JC-1)，and TUNEL) for flow cytometric analysis of apoptosis in sex-sorted sperm from four different bulls (A，B，C and D).Correlations between sperm quality and IVF efficiency were then assessed to determine which assay provided the best prediction of IVF efficiency.The results showed that: (1) the percentage of viable sperm (AN^-•PI^-) in X-sorted semen from bull A (89.30%±3.04%) was significantly higher than that from other bulls ((74.39%±3.24%)-(81.17%±4.25%); P<0.05)，while the percentage of DNA fragmentation sperm from bull A (12.24%±1.25%) was significantly lower than that from other bulls ((19.09%±1.27%)-(27.56%±3.17%); P<0.05); (2) the percentage of cells reaching the cleavage and blastocyst stages after in vitro fertilization (IVF) in sex-sorted sperm from bull A（（76.82%±3.00%），（20.45%±1.44%）） were significantly greater than those from the other bulls ((22.00%±6.85%)-(31.88%±6.69%)，(3.33%±2.67%)-(4.00%±3.26%); P<0.05); (3) a significant positive correlation was observed between viable sperm and the percentage of cells at the cleavage or blastocyst stages (P<0.05) and a negative correlation was found between early apoptotic sperm and the percentage of cells at the cleavage or blastocyst stages (P<0.05).In conclusion，these results indicated that the Annexin-V assay was the most reliable technique for the prediction of the IVF success of sex-sorted bovine sperm.

Key words: in vitro fertilization, different bulls, Annexin-V, JC-1, TUNEL, sperm apoptosis, semen quality

中图分类号:

S823.3

郝海生，任晶晶，赵学明，王玲玲，杜卫华，王长法，朱化彬. 性控分离精子凋亡程度对牛体外受精的影响[J]. 畜牧兽医学报, 2014, 45(6): 893-900.

HAO Hai-sheng，REN Jing-jing，ZHAO Xue-ming，WANG Ling-ling，DU Wei-hua，WANG Chang-fa，ZHU Hua-bin. Effects of the Degree of Apoptosis in Sexed Sperms on Bovine Fertilization in vitro[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2014, 45(6): 893-900.

参考文献

［1］UNDERWOOD S L，BATHGATE R，PEREIRA D C，et al.Embryo production after in vitro fertilization with frozen-thawed，sex-sorted，re-frozen-thawed bull sperm ［J］.Theriogenology，2010，73(1): 97-102.
［2］XU J，GUO Z，SU L，et al.Developmental potential of vitrified holstein cattle embryos fertilized in vitro with sex-sorted sperm ［J］.J Dairy Sci，2006，89(7): 2510-2518.
［3］吴冬生，刘敬浩，孙伟，等.不同种公牛X/Y精子分离效果及对体外受精能力的影响［J］.中国奶牛，2009，4:11-16.
［4］ZHANG M，LU K H，SEIDEL G E.Development of bovine embryos after in vitro fertilization of oocytes with flow cytometrically sorted，stained and unsorted sperm from different bulls ［J］.Theriogenology，2003，60(9): 1657-1663.
［5］LIU D Y，BAKER H W.Defective sperm-zona pellucida interaction: a major cause of failure of fertilization in clinical in-vitro fertilization ［J］.Hum Reprod，2000,15(3): 702-708.
［6］GLANDER H J,SCHALLER J.Binding of annexin V to plasma membranes of human spermatozoa: a rapid assay for detection of membrane changes after cryostorage ［J］.Mol Hum Reprod，1999，5(2): 109-115.
［7］WENG S L,TAYLOR S L,MORSHEDI M，et al.Caspase activity and apoptotic markers in ejaculated human sperm ［J］.Mol Hum Reprod,2002，8(11): 984-991.
［8］ZHANG H B，LU S M,MA C Y,et al.Early apoptotic changes in human spermatozoa and their relationships with conventional semen parameters and sperm DNA fragmentation ［J］.Asian J Androl，2008,10(2): 227-235.
［9］ANZAR M,HE L,BUHR M M,et al.Sperm apoptosis in fresh and cryopreserved bull semen detected by flow cytometry and its relationship with fertility ［J］.Biol Reprod,2002，66(2): 354-360.
［10］MARCHETTI C,OBERT G,DEFFOSEZ A,et al.Study of mitochondrial membrane potential，reactive oxygen species,DNA fragmentation and cell viability by flow cytometry in human sperm ［J］.Hum Reprod，2002，17(5): 1257-1265.
［11］AGARWAL A，SAIC T M.Role of sperm chromatin abnormalities and DNA damage in male infertility ［J］.Hum Reprod Update，2003，9(4): 331-345.
［12］SAKKAS D，MARIETHOZ E，MANICARDI G，et al.Origin of DNA damage in ejaculated human spermatozoa ［J］.Rev Reprod，1999，4(1): 31-37.
［13］ERENPREISS J，SPANO M，ERENPREISA J，et al.Sperm chromatin structure and male fertility: biological and clinical aspects ［J］.Asian J Androl，2006，8(1):11-29.
［14］SAID T，AGARWAL A，GRUNEWALD S，et al.Selection of nonapoptotic spermatozoa as a new tool for enhancing assisted reproduction outcomes: an in vitro model ［J］.Biol Reprod，2006,74(3): 530-537.
［15］RICCI G，PERTICARARI S，FRAGONAS E，et al.Apoptosis in human sperm: its correlation with semen quality and the presence of leukocytes ［J］.Hum Reprod，2002,17(10):2665-2672.
［16］SAKKAS D，MOFFATT O,MANICARDI G C，et al.Nature of DNA damage in ejaculated human spermatozoa and the possible involvement of apoptosis ［J］.Biol Reprod，2002，66(4):1061-1067.
［17］梁小军,张秀陶,宁小波，等.奶牛性控冻精与普通冻精的活力对比试验［J］.四川畜牧兽医，2008，35(4): 29-30.
［18］PALMA G A，OLIVIER N S，NEUMULLER C，et al.Effects of sex-sorted spermatozoa on the efficiency of in vitro fertilization and ultrastructure of in vitro produced bovine blastocysts［J］.Anat Histol Embryol，2008，37(1): 67-73.
［19］BLONDIN P，BEAULIEU M，FOURNIER V，et al.Analysis of bovine sexed sperm for IVF from sorting to the embryo ［J］.Theriogenology，2009，71(1): 30-38.
［20］LU K H,SERDEL G E.Effects of heparin and sperm concentration on cleavage and blastocyst development rates of bovine oocytes inseminated with flow cytometrically-sorted sperm ［J］.Theriogenology，2004，62(5): 819-830.
［21］RODRIGUEZ I，ODY C，ARAKI K，et al.An early and massive wave of germinal cell apoptosis is required for the development of functional spermatogenesis［J］.EMBO J,1997,16(9):2262-2270.
［22］CHAVEIRO A，SANTOS P，DA SILVA F M.Assessment of sperm apoptosis in cryopreserved bull semen after swim-up treatment: a flow cytometric study［J］.Reprod Domest Anim，2007，42(1):17-21.

[1]	蒲思颖, 郑杰, 杨远潇, 王琴, 杨绕芬, 字向东. 牦牛新鲜囊胚与玻璃化冻融囊胚转录组的比较分析[J]. 畜牧兽医学报, 2018, 49(4): 709-717.
[2]	蒲思颖, 郑杰, 杨远潇, 王琴, 杨绕芬, 字向东. 牦牛新鲜囊胚与玻璃化冻融囊胚转录组的比较分析[J]. 畜牧兽医学报, 2018, 49(4): 709-717.
[3]	黄自强, 王玲玲, 庞云渭, 郝海生, 杜卫华, 赵学明, 刘岩, 朱化彬. 表没食子儿茶素没食子酸酯对牛冷冻精液品质及受精能力的影响[J]. 畜牧兽医学报, 2018, 49(10): 2154-2162.
[4]	黄自强, 王玲玲, 庞云渭, 郝海生, 杜卫华, 赵学明, 刘岩, 朱化彬. 表没食子儿茶素没食子酸酯对牛冷冻精液品质及受精能力的影响[J]. 畜牧兽医学报, 2018, 49(10): 2154-2162.
[5]	字向东, 刘霜, 夏威, 熊显荣, 黄林, 张正帆, 李志雄, 李键, 钟金城, 王利, 朱江江. 犏牛体外受精胚胎的发育转录组分析[J]. 畜牧兽医学报, 2018, 49(1): 92-101.
[6]	字向东, 刘霜, 夏威, 熊显荣, 黄林, 张正帆, 李志雄, 李键, 钟金城, 王利, 朱江江. 犏牛体外受精胚胎的发育转录组分析[J]. 畜牧兽医学报, 2018, 49(1): 92-101.
[7]	赵天，李谷月，潘阳阳，李月琴，陈平，温泽星，崔燕，余四九. Oct 4 在牦牛卵母细胞及早期胚胎发育过程中的表达[J]. 畜牧兽医学报, 2017, 48(3): 577-584.
[8]	赵天，李谷月，潘阳阳，李月琴，陈平，温泽星，崔燕，余四九. Oct 4 在牦牛卵母细胞及早期胚胎发育过程中的表达[J]. 畜牧兽医学报, 2017, 48(3): 577-584.
[9]	梁学超, 蒋明, 罗玉茹, 张为民, 卿素珠. 猪卵巢发育的组织学变化及卵泡闭锁规律研究[J]. 畜牧兽医学报, 2017, 48(10): 1863-1870.
[10]	郑杰, 蒲思颖, 杨远潇, 王琴, 杨绕芬, 字向东. 基于高通量测序的犏牛囊胚玻璃化冷冻损伤机制研究[J]. 畜牧兽医学报, 2017, 48(10): 1871-1881.
[11]	郑杰, 蒲思颖, 杨远潇, 王琴, 杨绕芬, 字向东. 基于高通量测序的犏牛囊胚玻璃化冷冻损伤机制研究[J]. 畜牧兽医学报, 2017, 48(10): 1871-1881.
[12]	梁学超, 蒋明, 罗玉茹, 张为民, 卿素珠. 猪卵巢发育的组织学变化及卵泡闭锁规律研究[J]. 畜牧兽医学报, 2017, 48(10): 1863-1870.
[13]	胡庭溪，刘霞，孙尉峻，郝海生，赵学明，王宗礼，杜卫华，朱化彬. 精子动力学分析与其体内、体外受精能力的相关性[J]. 畜牧兽医学报, 2016, 47(8): 1727-1732.
[14]	胡庭溪，刘霞，孙尉峻，郝海生，赵学明，王宗礼，杜卫华，朱化彬. 精子动力学分析与其体内、体外受精能力的相关性[J]. 畜牧兽医学报, 2016, 47(8): 1727-1732.
[15]	郝海生，任晶晶，赵学明，王玲玲，杜卫华，王长法，朱化彬. 性控分离精子凋亡程度对牛体外受精的影响[J]. 畜牧兽医学报, 2014, 45(6): 893-900.

性控分离精子凋亡程度对牛体外受精的影响

Effects of the Degree of Apoptosis in Sexed Sperms on Bovine Fertilization in vitro

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价