摘要
Objective To investigate the expression regulation of thyrotrophin-releasing hormone (TRH) and TRH receptor (TRH-R), and their role in the development of rat testis.Methods Oligonucleotide primers were designed from the sequences of rat hypothalamus prepro TRH (ppTRH) and pituitary TRH-R cDNA for reverse transcription polymerase chain reaction (RT-PCR). Specific fragments of ppTRH and TRH-R cDNA were cloned and sequenced. Expression plasmids containing ppTRH and TRH-R genes were then constructed, and expression was found in E.coli DH5-α. ppTRH and TRH-R mRNA in the testis was quantitated in RNA samples prepared from rats at different developmental stages by real time quantitative RT-PCR.Results The quantitative analyses demonstrated that no ppTRH and TRH mRNA could be detected at the earliest stage (day 8). ppTRH and TRH mRNA signals were detected on day 15 and increased progressively on days 20, 35, 60 and 90. Conclusion Our results suggest that rat testis could specifically express TRH and TRH-R, and the transcriptions of ppTRH and TRH-R genes in the rat testis were development-dependent. The acquirement of expressed products for ppTRH and TRH-R can be used for further research on the physiological significance of TRH and TRH-R expression in rat testis.
目的 研究促甲状腺激素释放激素受体 (TRH R)在大鼠睾丸组织中的表达规律和在生殖发育调节中的作用。方法 依据大鼠垂体中的TRH RcDNA设计引物 ,采用RT PCR法从大鼠睾丸组织中获得了TRH R的cDNA克隆 ,测序表明其核苷酸序列与大鼠垂体中的TRH RcDNA序列完全一致。在大肠杆菌中表达了可溶性的ppTRH和TRH R的融合蛋白 ,并利用实时动态定量RT PCR法观察了TRH R在不同发育阶段大鼠睾丸中的表达变化。结果 在睾丸间质细胞发育的初期阶段 (第 8天 )没有TRH R的表达 ,但从第 15天起能观察到TRH R的表达 ,并且表达量在 2 0天、35天、6 0天、90天逐渐增加。结论 大鼠睾丸组织间质细胞能特异性表达TRH R ,并且表达量与发育过程相关。
