Department of Animal Sciences, University of Florida, Gainesville, FL 32611, USA.
J Anim Sci. 2011 May;89(5):1338-46. doi: 10.2527/jas.2010-3559.
In the present study, we performed quantitative reverse-transcription PCR (qPCR) to examine changes in gene expression of prolactin receptor (long form: l-PRLR; short form: s-PRLR) and 20α-hydroxysteroid dehydrogenase (20α-HSD; EC 1.1.1.149) in the bovine corpus luteum (CL) throughout the estrous cycle and pregnancy. Western blotting was used to determine protein abundance. Bovine CL were collected and luteal stages (n = 6/stage) were classified by macroscopic observation as early (d 1 to 4 after ovulation), mid (d 5 to 10), late (d 11 to 17), and regressing (d 18 to 20). A CL of pregnancy (n = 6) was determined by the presence of conceptus (d 28 to term). The mRNA for both forms of PRLR were expressed at all the luteal stages. Expression of s-PRLR and l-PRLR mRNA was less (P < 0.01) during early and regressing luteal stages compared with mid and late stages. Expression of s-PRLR mRNA in CL of pregnancy was greater (P < 0.01) than early, mid, and regressing CL and did not differ from late luteal stage expression. A greater (P < 0.01) expression of l-PRLR mRNA was observed in pregnant vs. early and regressing CL. In addition, qPCR showed the presence of 20α-HSD mRNA during all luteal stages of the estrous cycle, with the greatest (P < 0.01) expression observed in the regressing luteal stage. Western blotting revealed protein abundance of both PRLR isoforms during all luteal stages and pregnancy, with a predominance of the s-PRLR protein. Densitometry analysis indicated that protein abundances of s-PRLR were greater (P < 0.05) than l-PRLR during early, mid, and late luteal stages and did not differ during the regressing luteal stage. Protein abundances of 20α-HSD were least (P < 0.05) during the early luteal stage. In conclusion, results of the current study suggest a possible involvement of PRLR, especially s-PRLR, in the regulation of progesterone secretion and metabolism during the bovine estrous cycle and pregnancy.
在本研究中,我们通过定量逆转录 PCR (qPCR) 检测了牛黄体(CL)中催乳素受体(长形式:l-PRLR;短形式:s-PRLR)和 20α-羟类固醇脱氢酶(20α-HSD;EC 1.1.1.149)基因表达在发情周期和妊娠过程中的变化。Western blot 用于确定蛋白质丰度。采集牛 CL,根据宏观观察将黄体阶段(n=6/阶段)分为早期(排卵后第 1 至 4 天)、中期(第 5 至 10 天)、晚期(第 11 至 17 天)和退化期(第 18 至 20 天)。妊娠 CL(n=6)通过存在胚胎(第 28 至足月)确定。两种形式的 PRLR mRNA 均在所有黄体阶段表达。与中期和晚期相比,早期和退化期黄体 s-PRLR 和 l-PRLR mRNA 的表达较少(P<0.01)。妊娠 CL 中 s-PRLR mRNA 的表达(P<0.01)高于早期、中期和退化期 CL,与晚期黄体阶段的表达没有差异。与早期和退化期 CL 相比,妊娠 CL 中 l-PRLR mRNA 的表达更高(P<0.01)。此外,qPCR 显示在发情周期的所有黄体阶段均存在 20α-HSD mRNA,退化期黄体阶段的表达最高(P<0.01)。Western blot 显示两种 PRLR 同工型在所有黄体阶段和妊娠期间的蛋白丰度,s-PRLR 蛋白占优势。密度计分析表明,早期、中期和晚期黄体阶段 s-PRLR 的蛋白丰度(P<0.05)大于 l-PRLR,退化期黄体阶段则无差异。20α-HSD 的蛋白丰度在早期黄体阶段最低(P<0.05)。总之,本研究结果表明,PRLR 特别是 s-PRLR,可能参与了牛发情周期和妊娠期间孕激素分泌和代谢的调节。