甘加型藏羊发情周期血浆孕酮动态变化及HPO轴PGR的表达

Dynamic changes of plasma progesterone and expression of HPO axis PGR in Ganjia Tibetan sheep during estrus cycle

为研究甘加型藏羊(Ovis arise)发情周期血浆孕激素(P4)的分泌及孕激素受体(Progesterone receptor,PGR)在HPO轴中的表达,分析其在藏羊繁殖活动中的调控作用,本研究应用酶联免疫吸附试验、蛋白免疫印迹法、实时荧光定量和免疫组织化学染色法检测甘加型藏羊发情周期血浆中孕酮含量的动态变化、HPO组织中PGR mRNA及其蛋白的表达和组织分布。结果显示:1)甘加型藏羊发情周期血浆孕酮呈波动式和脉冲式分泌,在间情期达到最高峰值,发情期最低,二者差异显著(P<0.05)。2)PGR mRNA及其蛋白在HPO组织中均有表达;下丘脑中,PGR mRNA及其蛋白相对表达量在间情期最高,各时期之间差异显著(P<0.05);垂体中,PGR mRNA表达量在发情后期达到最高,显著高于发情前期和间情期(P<0.05),发情前期蛋白相对表达量高于其他时期;卵巢中,PGR mRNA在发情期最高,与间情期差异显著(P<0.05),蛋白表达水平在发情前期最高。3)PGR免疫阳性反应产物表达在下丘脑神经胶质细胞和大神经元胞体中,腺垂体嗜酸性细胞和嗜碱性细胞的胞浆中,卵巢生长卵泡的颗粒层细胞、卵泡膜细胞以及黄体细胞胞浆中。综上,甘加型藏羊发情周期血浆中孕酮含量的动态变化、HPO轴中PGR mRNA、蛋白的差异性表达表明孕酮通过与分布在HPO生殖轴上的PGR受体结合,调节甘加型藏羊周期性发情和生殖器官的功能活动。

To study the secretion of plasma progesterone(P4)and the expression of progesterone receptor(Progesterone receptor,PGR)in the HPO axis of Ganjia Tibetan sheep(Ovis arise)during the estrus cycle,and to analyze its regulatory effect on the reproductive activity of Tibetan sheep,in this study,enzyme-linked immunosorbent assay(ELISA),Western blotting,Real-time fluorescence quantification and immunohistochemistry staining were used to detect the dynamic changes of progesterone in plasma of Ganjia Tibetan sheep during estrus cycle and the expression and distribution of PGR mRNA and its protein in HPO tissues.The results showed that:1)The secretion of progesterone in the plasma of Ganjia Tibetan sheep showed fluctuating and pulsed secretion during the estrus cycle,with the highest value in diestrus and the lowest in estrus,with significant differences(P<0.05).2)PGR mRNA and its protein were expressed in HPO tissues.In hypothalamus,the relative expression level of PGR mRNA and its protein was the highest in diestrus,significant differences among different periods(P<0.05).In pituitary,PGR mRNA reached the highest in metestrus,which was significantly higher than that in proestrus and diestrus(P<0.05),and the relative expression level of proestrus protein was higher than that in other periods.Among the ovaries,PGR mRNA was the highest in estrus,which was significantly different from that in diestrus(P<0.05),and the protein expression level was the highest in proestrus.3)PGR immune reactive products were expressed in hypothalamic glial cells and large neuronal bodies,in the cytoplasm of eosinophils and basophils in adenohypophysis,and in the cytoplasm of granular layer cells,theca cells and luteal cells of growing follicles in ovaries.In summary,the dynamic changes of plasma progesterone content and the differential expression of PGR mRNA and protein in the HPO axis indicate that progesterone regulates the periodic estrus and reproductive organ function of Ganjia Tibetan sheep during the estrus cycle by binding to PGR receptors distributed on the reproductive axis of HPO.