骨桥蛋白在长白公猪生殖细胞中的表达及其与繁殖性能的相关性

doi:10.16431/j.cnki.1671-7236.2020.08.022

中国畜牧兽医 ›› 2020, Vol. 47 ›› Issue (8): 2536-2543.doi: 10.16431/j.cnki.1671-7236.2020.08.022

骨桥蛋白在长白公猪生殖细胞中的表达及其与繁殖性能的相关性

陈云¹, 王凯¹, 周乐乐¹, 黄建豪¹, 叶超², 卫恒习¹, 李莉¹, 张守全¹

1. 华南农业大学动物科学学院, 国家生猪种业工程技术研究中心, 广东省农业动物基因组学与分子育种重点实验室, 广州 510642;
2. 广东温氏食品集团股份有限公司, 新兴 527439

收稿日期:2020-02-29 出版日期:2020-08-20 发布日期:2020-08-15
通讯作者: 张守全 E-mail:sqzhang@scau.edu.cn
作者简介:陈云(1986-),女,山东成武人,博士生,研究方向:动物遗传育种与繁殖,E-mail:luxixiworld@163.com
基金资助:
国家重点研究计划"经产母猪定时输精及批次化繁殖技术研发与示范"（2017YFD0501902）；2019年华南农业大学博士生创新人才（国内培养）培植项目（CX2019N005）

Expression of OPN in Germ Cells of Landrace Boar and Its Correlation with Reproductive Performance

CHEN Yun¹, WANG Kai¹, ZHOU Lele¹, HUANG Jianhao¹, YE Chao², WEI Hengxi¹, LI Li¹, ZHANG Shouquan¹

1. National Engineering Research Center for Breeding Swine Industry, Guangdong Provincial Key Laboratory of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agricultural University, Guangzhou 510642, China;
2. Guangdong Wens Foodstuff Group Co., Ltd., Xinxing 527439, China

Received:2020-02-29 Online:2020-08-20 Published:2020-08-15

摘要/Abstract

摘要： 本试验旨在研究骨桥蛋白（osteopontin，OPN）在长白公猪生殖细胞中的表达和定位，并探究其与公猪繁殖性能的相关性。试验采集了长白公猪精液和不同阶段（3日龄、3月龄、6月龄和12月龄）的睾丸组织，通过蛋白印迹的方法检测OPN蛋白在精液和不同月龄睾丸中的表达，通过免疫组化的方法对OPN蛋白在公猪睾丸细胞中进行定位；同时，根据配种胎次≥ 20胎，3次配种公猪为同一头的标准，筛选并采集17头长白种公猪精液，统计相对应的1 388头母猪的生产成绩，计算得到公猪繁殖性能指标（包括窝产总仔数、窝产活仔数、分娩率和繁殖力）。低温离心精液分离得到精子和精浆，丙酮法提取精浆蛋白，Lysis buffer方法提取精子蛋白，最后运用BCA和ELISA的方法检测精子和精浆中OPN蛋白的含量，分析OPN蛋白与公猪繁殖性能的相关性。蛋白印迹结果显示，OPN在精子、精浆和各月龄阶段的长白公猪睾丸中均以两种形式表达（67.4和33.7 ku），且67.4 ku的形式在3月龄公猪睾丸中表达量最高；免疫组化的结果显示，OPN在长白公猪的初级精母细胞、次级精母细胞和精子细胞中表达，在精母细胞、支持细胞和间质细胞中无表达；BCA和ELISA结果显示，精子中的OPN蛋白含量是精浆中的7倍（P<0.05），精液中的OPN蛋白与公猪窝产活仔数显著正相关（P<0.05）。本试验结果表明，OPN在各阶段的长白公猪睾丸中都有表达，且在精子和精浆中也有表达，这可能与公猪的繁殖性能有关，从而为后期OPN蛋白在公猪受精力的研究奠定基础。

关键词: 长白公猪; 睾丸; 繁殖性能; 骨桥蛋白

Abstract: The aim of this study was to investigate the expression and localization of osteopontin (OPN) in germ cells of Landrace boars,and their correlations with boar reproductive performance.The semen and testis in different age stages (3-day-old,3-month-old,6-month-old,and 12-month-old) from Landrace boars were collected,followed with the expression levels of OPN protein in semen and testis,and its location in boar testis cells detected by Western blotting and immunohistochemistry(IHC),respectively.At the same time,according to the standard of breeding parity ≥ 20 births and three times of breeding semen from the same boar,17 Landrace boars were selected along with their semen collected and evaluated.Further,productive performance data of the 1 388 boar-paired sows were calculated for evaluating the performance of those 17 breeding boars by total number born per litter (TNB),number born alive per litter (NBA),farrowing rate (FR) and fecundity.To evaluated the semen of those 17 breeding boars,sperm and seminal plasma were separated by low-temperature centrifugation of semen.Seminal plasma protein was extracted with acetone method,and sperm protein was extracted with Lysis buffer method.BCA (Bicinchoninic acid) and ELISA were used to detect OPN contention in sperm and seminal plasma,for subsequent correlation analysis between OPN protein and boar reproductive performance by SPSS software.Consequently,Western blotting results showed that two forms of OPN protein(67.4 and 33.7 ku) could be observed in sperm,seminal plasma and testis of Landrace boars at all age stages,and 67.4 ku form was highest expressed in testis of 3-month-old boars;IHC results showed that OPN were expressed in primary spermatocytes,secondary spermatocytes and sperm cells,but not in spermatogonia,sertoli cells and interstitial cells;BCA and ELISA results showed that the concentration of OPN protein in sperm was 7 times higher than that in seminal plasma (P<0.05).Additionally,correlation analysis between OPN protein and boar reproductive performance indicated the OPN protein in semen was significantly positively correlated with NBA (P<0.05).Collectively,our study suggested that OPN expresses in testis (at all age stages),sperm and seminal plasma of Landrace boars,and it might concern the reproductive performance of boars,thereby laying a foundation for the mechanism study of OPN protein in boar fertility.

Key words: Landrace boar; testis; reproductive performance; OPN

中图分类号:

S828.2

陈云, 王凯, 周乐乐, 黄建豪, 叶超, 卫恒习, 李莉, 张守全. 骨桥蛋白在长白公猪生殖细胞中的表达及其与繁殖性能的相关性[J]. 中国畜牧兽医, 2020, 47(8): 2536-2543.

CHEN Yun, WANG Kai, ZHOU Lele, HUANG Jianhao, YE Chao, WEI Hengxi, LI Li, ZHANG Shouquan. Expression of OPN in Germ Cells of Landrace Boar and Its Correlation with Reproductive Performance[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(8): 2536-2543.

参考文献

[1] 欧云文,阎传忠,张杰,等.非洲猪瘟病毒的分子病原学及致病机理研究进展[J].中国畜牧兽医,2017,44(7):2139-2146. OU Y W,YAN C Z,ZHANG J,et al.Research progress on molecular etiology and pathogenesis for African swine fever virus[J].China Animal Husbandry & Veterinary Medicine,2017,44(7):2139-2146.(in Chinese)
[2] ROCA J,BROEKHUIJSE M L,PARRILLA I,et al.Boar differences in artificial insemination outcomes:Can they be minimized?[J].Reproduction in Domestic Animals,2015,50 Suppl 2:48-55.
[3] BEZERRA M,SILVA M B,LOBO C H,et al.Gene and protein expression in the reproductive tract of Brazilian Somalis rams[J].Reproduction in Domestic Animals,2019,54(7):939-948.
[4] WANG X B,QI Q R,WU K L,et al.Role of osteopontin in decidualization and pregnancy success[J].Reproduction,2018,155(5):423-432.
[5] LV Y,JIN Y,ZHOU Y,et al.Deep sequencing of transcriptome profiling of GSTM2 knock-down in swine testis cells[J].Scientific Reports,2016,6:38254.
[6] ASLANKOC R,OZMEN O.The effects of high-fructose corn syrup consumption on testis physiopathology-the ameliorative role of melatonin[J].Andrologia,2019,51(8):e13327.
[7] KING A H,JIANG Z,GIBSON J P,et al.Mapping quantitative trait loci affecting female reproductive traits on porcine chromosome 8[J].Biology of Reproduction,2003,68(6):2172-2179.
[8] BUSKE B,STERNSTEIN I,BROCKMANN G.QTL and candidate genes for fecundity in sows[J].Animal Reproduction Science,2006,95(3-4):167-183.
[9] 孟庆利,刘铁铮.猪骨调素基因多态性对产仔性状的影响[J].家畜生态学报,2005,3:13-16. MENG Q L,LIU T Z.The effect of swine osteopontin gene polymorphism on litter size[J].Ecology of Domestic Animal,2005,3:13-16.(in Chinese)
[10] 刘乙,马力鹏,李涛,等.ESR、OPN和RBP4基因多态性对大白猪繁殖性状的影响[J].中国畜牧兽医,2017,44(1):123-130. LIU Y,MA L P,LI T,et al.Influence of ESR,OPN and RBP4 genes polymorphisms on reproductive traits of Yorkshire sows[J].China Animal Husbandry & Veterinary Medicine,2017,44(1):123-130.(in Chinese)
[11] NOVAK S,RUIZ-SANCHEZ A,DIXON W T,et al.Seminal plasma proteins as potential markers of relative fertility in boars[J].Journal of Andrology,2010,31(2):188-200.
[12] 张守全,陈云,卫恒习,等.一种重组慢病毒载体、重组慢病毒及其应用:201811476996.8[P].2019-03-08. ZHANG S Q,CHEN Y,WEI H X,et al.Recombinant lentivirus vector,recombinant lentivirus and application thereof:201811476996.8[P].2019-03-08.(in Chinese)
[13] 张守全,陈云,王凯,等.猪源OPN单克隆抗体、其杂交瘤细胞株及应用:201911323477.2[P].2020-05-19. ZHANG S Q,CHEN Y,WANG K,et al.Pig-derived OPN monoclonal antibody,its hybridoma cell line and its application:201911323477.2[P].2020-05-19.(in Chinese)
[14] HERNANDEZ S C,HOGG C O,BILLON Y,et al.Secreted phosphoprotein 1 expression in endometrium and placental tissues of hyperprolific large white and meishan gilts[J].Biology of Reproduction,2013,88(5):120.
[15] ZHANG G M,LAN S,JIA R X,et al.Age-associated and tissue-specific expression of osteopontin in male Hu sheep reproductive tract[J].Tissue Cell,2016,48(5):496-502.
[16] KIM S,SHIN T.Immunohistochemical study of osteopontin in boar testis[J].Journal of Veterinary Science,2007,8(2):107-110.
[17] STAUB C,JOHNSON L.Review:Spermatogenesis in the bull[J].Animal,2018,12(s1):s27-s35.
[18] CANNARELLA R,CONDORELLI R A,MONGIOI L M,et al.Molecular biology of spermatogenesis:Novel targets of apparently idiopathic male infertility[J].International Journal of Molecular Sciences,2020,21(5):1728.
[19] POTTER S J,DEFALCO T.Role of the testis interstitial compartment in spermatogonial stem cell function[J].Reproduction,2017,153(4):R151-R162.
[20] LIU Q,XIE Q Z,ZHOU Y,et al.Osteopontin is expressed in the oviduct and promotes fertilization and preimplantation embryo development of mouse[J].Zygote,2015,23(4):622-630.
[21] LIN C,THOLEN E,JENNEN D,et al.Evidence for effects of testis and epididymis expressed genes on sperm quality and boar fertility traits[J].Reproduction in Domestic Animals,2006,41(6):538-543.
[22] BAKER M A,NIXON B,NAUMOVSKI N,et al.Proteomic insights into the maturation and capacitation of mammalian spermatozoa[J].Systems Biology in Reproductive Medicine,2012,58(4):211-217.
[23] BROMFIELD J J.Seminal fluid and reproduction:Much more than previously thought[J].Journal of Assisted Reproduction and Genetics,2014,31(6):627-636.
[24] DRUART X,DE GRAAF S.Seminal plasma proteomes and sperm fertility[J].Animal Reproduction Science,2018,194:33-40.
[25] MARCELLO M R,DRUZHININA M,SINGSON A.Caenorhabditis elegans sperm membrane protein interactome[J].Biology of Reproduction,2018,98(6):776-783.
[26] GUIMARAES D B,BARROS T B,VAN TILBURG M F,et al.Sperm membrane proteins associated with the boar semen cryopreservation[J].Animal Reproduction Science,2017,183:27-38.
[27] PARRILLA I,PEREZ-PATINO C,LI J,et al.Boar semen proteomics and sperm preservation[J].Theriogenology,2019,137:23-29.

骨桥蛋白在长白公猪生殖细胞中的表达及其与繁殖性能的相关性

Expression of OPN in Germ Cells of Landrace Boar and Its Correlation with Reproductive Performance

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