促黄体生成素和17β-雌二醇对牦牛输卵管糖蛋白表达的影响  

Effects of LH and E_2 on the Expression of Oviductin in Bos grunniens

作　　者：彭秀梅[1] 温泽星[1] 张译夫[1] 潘阳阳[1] 余四九[1] 

机构地区：[1]甘肃农业大学动物医学院/甘肃省牛羊胚胎工程技术研究中心,兰州730070

出　　处：《农业生物技术学报》2016年第10期1466-1473,共8页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(No.31272616和No.31472244)

摘　　要：为研究相关激素是否通过体细胞参与哺乳动物胚胎发育的调控,本研究探讨了促黄体生成素（luteinizing hormone,LH）和17β-雌二醇（17β-estradiol,E2）对牦牛（Bos grunniens）输卵管上皮细胞分泌输卵管糖蛋白（oviductin,Ovn）的影响。体外培养牦牛输卵管上皮细胞,分别添加0、10、25、50、100和200 ng/m L LH和E2,作用72 h后,运用q RT-PCR和免疫荧光技术,从基因和蛋白水平检测Ovn表达。获得如下结果：（1）LH和E2可以提高牦牛输卵管上皮细胞的Ovn m RNA相对表达量;当LH浓度为10 ng/m L时,Ovn m RNA的相对表达量最高,显著高于LH浓度为25、50、100和200 ng/m L组（P〈0.05）,对照组Ovn m RNA的表达量最低（P〈0.05）;当E2浓度为25 ng/m L时,输卵管上皮细胞的Ovn m RNA相对表达量最高,显著高于E2浓度为10、50、100和200 ng/m L组（P〈0.05）,对照组Ovn m RNA表达量最低（P〈0.05）。（2）Ovn主要分布于核膜,细胞质也有表达;LH和E2可以提高输卵管上皮细胞的蛋白相对表达量,当LH浓度为10ng/m L时,Ovn的蛋白表达量达到最高,LH浓度达到25 ng/m L时,Ovn的蛋白表达量降低;当E2浓度为10-25 ng/m L时,Ovn的蛋白表达量随着E2浓度的上升不断增加,在25 ng/m L的E2作用后,Ovn的蛋白表达量达到最高,E2浓度达到50 ng/m L时,Ovn的蛋白表达量降低。上述结果表明,LH和E2对牦牛输卵管上皮细胞分泌Ovn具有明显的促进作用,LH的最佳作用浓度为10 ng/m L,E2的最佳作用浓度为25ng/m L。本研究为揭示LH和E2对输卵管上皮细胞分泌Ovn的作用机制提供了基础资料,为进一步研究相关激素对牦牛胚胎发育的调控提供了一定理论依据。The aim of this study is to verify whether involved in mammalian embryonic development by Somatic cell. This paper discusses the effects of luteinizing hormone（LH） and 17β-estradiol（E2） on oviductin（Ovn） by oviduct epithelial cells. We culture yak（Bos grunniens） oviduct epithelial cells in vitro with different concentration LH and E2（10, 25, 50, 100 and 200 ng/m L）. The relative expression level of Ovn gene and protein in different groups were detected by q RT- PCR and immunofluorescence, and location of ovn protein was detected by immunofluorescence after 72 h. The result obtained in this study was as follows that：（1） LH improved the relative levels of Ovn m RNA; These improvements were highest in the 10 ng/m L LH group（P〈0.05）, followed by 25, 50, 100 and 200 ng/m L groups, the relative levels of Ovn m RNA were the lowest incontrol group（0 ng/m L LH）; E2 improved the relative levels of Ovn m RNA; These improvements were the highest in the 25 ng/m L E2group（P〈0.05）, followed by 50, 10, 100 and 200 ng/m L groups, the relative levels of Ovn m RNA were the lowest in control group（0 ng/m L E2）.（2） Detection by immunofluorescence showed that Ovn protein could be observed in yak cumulus cells cytoplasm as well as within nuclear membrane, and the expression of nuclear membrane was higher than the cytoplasm. LH and E2 could improve the relative expression levels of Ovn protein in yak oviduct epithelial cells; The improvements were the highest in the 10 ng/m L LH group（P〈0.05）. However, the Ovn protein expression was decreased when LH concentration over 25 ng /m L. The relative levels of Ovn protein were the lowest in control group（0 ng/m L LH）; The effective range of E2（10~25 ng/m L）, and the highest Ovn protein copy numbers at 25 ng/m L（P〈0.05）, and reduced protein expression during all others. However, the Ovn protein expression was decreased when E2 concentration over 50 ng/m L. The relative levels of Ovn protein were the lowest in control group�

关 键 词：促黄体生成素 17Β-雌二醇 输卵管糖蛋白 QRT-PCR 免疫荧光技术 

分 类 号：S857.21[农业科学—临床兽医学]