摘要
目的应用实时反转录（Real-time）PCR和ELISA法检测不同青春期发育阶段Sprague-Dawley（sD）雌性大鼠下丘脑KisslmRNA、雌激素受体d（ERa）mRNA的表达水平以及血清黄体生成素（LH）、雌二醇（E2）水平，探讨Kissl基因和ERa基因在大鼠性发育过程中的作用。方法实验动物选用正常3日龄雌性SD大鼠35只，于出生22d断奶后每日观察阴道开口情况，分别于出生15d（幼年期组，n=19）和出生35d（青春期组，n=16）处死动物，分离下丘脑组织，心脏采血法提取血清，标本均置-80℃冰箱保存备测；提取下丘脑标本的RNA，反转录后所得eDNA进行Real—timePCR检测，计算Kissl基因和ERa基因的mRNA相对表达水平。ELISA方法检测其血清标本LH、E。水平。结果1．大体观察：观察到青春期组大鼠阴道开口的时间为（32．1±1．0）d，幼年期组在处死前一直未观察到阴道开口。2．Real-timePCR结果显示，青春期组大鼠的Kissl基因（5．394-2．52）和ERa基因（1．57±1．87）的表达，均显著高于幼年期组Kissl基因（1．06±1．09）和ERa基因（0．59±0．68），差异均有统计学意义（P均〈0．001）。3．ELISA结果显示，青春期组大鼠血清LH[（11．61±0．95）IU／L]和E2[（167．53±31．09）ng／L]水平，均显著高于幼年期组LH[（5．46±1．89）IU／L]和E2[（58．59±29．96）ng／L]，差异均有统计学意义（P均〈0．001）。结论Kissl基因和ERa基因参与雌性sD大鼠性发育的启动过程。
Objective To investigate the function of Kissl gene and estrogen receptor α gene （ERa gene） in puberty of rats, by detecting the expressions of Kissl mRNA and ERa mRNA in the hypothalamus and the serum luteini- zing hormone（LH） and estradiol（E2） level of female Sprague-Dawley（SD） rats at various stage of development with Real-time PCR and enzyme-linked immunosorbent assay （ELISA）. Methods Thirty-five female SD rats of 3 days were weaned on postnatal（PND）22 and then the vaginal opening condition was observed daily. The rats were sacrificed at PND 15（juvenile group,n = 19） and PND 35 （pubertal group,n = 16）. The hypothalamus were segregated and the serum were extracted from heart blood. All of the samples were stored at - 80 ℃ prepared. Then the mRNA were extrac- ted from the hypothalamus and the cDNA obtained by reverse transcription were tested with real-time PCR. The relative mRNA expression level of Kiss1 gene and ERa gene were calculated. Results 1. Entire level ： it was found that the pu- bertal group vaginal opening time was （32.1 ± 1.0） days, while the juvenile group was not found with vaginal opening until sacrificed. 2. Real-time PCR ： the expressions of Kiss1 and ERa gene were significantly increased in pubertal group （ Kissl gene ：5.39 ± 2.52, ERα gene ： 1.57 ± 1.87 ） compared with juvenile group （ Kissl gene ： 1.06 ±1.09, ERa gene ： 0.59 ± 0.68 ）, and the differences were statistically significant （ all P 〈 0.001 ）. 3. ELISA： the serum LH and E2 in pu- bertal group[ LH（ 11.61 ±0.95） IU/L,E2 （ 167.53 ±31.09） ng/L] were significantly higher than LH[ （5.46 ± 1.89） IU/L] and E2 [ （58.59 ± 29.96） ng/L 1 in juvenile group, and the differences were statistically significant （ all P 〈 0. 001 ）. Conclusion Kissl gene and ERa gene are involved in the start of the sexual development of female SD rat.
出处
《中华实用儿科临床杂志》
CAS
CSCD
北大核心
2013年第8期589-591,共3页
Chinese Journal of Applied Clinical Pediatrics
基金
江苏省自然科学基金（BK2009448）
