Crb3影响小鼠睾丸间质细胞分泌睾酮的研究

发布时间：2018-04-22 05:38

  本文选题：Crb3 + 睾丸间质细胞　； 参考：《南昌大学》2014年硕士论文

【摘要】：目的：检测TM3(小鼠睾丸间质细胞)中是否有Crb3基因的表达，如有表达，则通过转染，使其Crb3表达上调，检测基因表达调控后睾酮分泌功能的变化。探索Crb3对睾丸间质细胞睾酮分泌功能影响的相关性。为临床中睾酮缺乏综合征的治疗提供向导。 方法： 1、通过培养传代细胞，从中提取RNA。将RNA反转录成C-DNA，利用普通PCR检测Crb3在睾丸间质细胞中的基因表达。 2、利用Western-Blot检测Crb3在睾丸间质细胞中蛋白翻译表达情况，证明Crb3在TM3中翻译蛋白。 3、转染细胞，使Crb3在TM3细胞中过表达，设置转染组、空转组、非转染组，利用荧光定量PCR检测Crb3基因转录表达改变。 4、同时以化学发光法检测转染组、空转组、非转染组培养基中睾酮的值。 5、统计学分析：利用spss19软件系统建立数据库和进行统计学分析，多样板均数间比较采用方差分析，P0.05认为差异用统计学意义。 结果： 1、睾丸间质细胞中Crb3的DNA的表达及其意义普通PCR结果显示：Crb3在睾丸间质细胞中呈现出低表达。 2、睾丸间质细胞中Crb3的蛋白质的翻译及其临床意义 Western-blot结果显示：Crb3在睾丸间质细胞中，Crb3蛋白质翻译亦呈现出低表达，，正好符合Crb3DNA的表达情况。 3、转染实验结果显示：Crb3过表达转染组与空转组及正常未转染组相比，转染组CT小于其他两组，可见DNA表达明显高于空转组与未转染组，且有统计学意义（P0.05） 4、转染实验结果显示：Crb3过表达转染组与空转组及正常未转染组相比，转染组培养基中睾酮含量低于空转组与未转染组，且有统计学意义（P0.05） 结论：Crb3蛋白基因在睾丸间质细胞中的过表达将导致睾丸间质细胞分泌睾酮功能下降。
[Abstract]:Aim: to detect the expression of Crb3 gene in mouse testicular stromal cells (TM3) and, if so, to up-regulate the expression of Crb3 by transfection, and to detect the changes of testosterone secretion after the regulation of gene expression. To explore the correlation between Crb3 and testosterone secretion in stromal cells of testis. To provide guidance for the treatment of testosterone deficiency syndrome. Methods: 1. RNA was extracted from the cultured cells. RNA was reverse transcribed into C-DNA, and the expression of Crb3 gene in Leydig cells of testis was detected by ordinary PCR. 2. Western-Blot was used to detect the protein translation and expression of Crb3 in Leydig cells of testis, which proved that Crb3 translated protein in TM3. (3) Crb3 was overexpressed in TM3 cells after transfection. The transcriptional changes of Crb3 gene were detected by fluorescence quantitative PCR in transfection group, empty group and non-transfection group. 4. At the same time, the testosterone in the culture medium of transfection group, empty group and non-transfection group was detected by chemiluminescence method. 5, statistical analysis: using spss19 software system to establish database and statistical analysis, the comparison of multi-sample mean using ANOVA (P0.05) that the difference is statistically significant. Results: 1. The expression of Crb3 DNA in Leydig cells of testis and its significance. The results of ordinary PCR showed that the expression of Crb3 was low in Leydig cells of testis. Translation of Crb3 protein in Leydig cells of Testis and its Clinical significance Western-blot results showed that the translation of Crb3 protein in Leydig cells of testis also showed a low expression, which coincided with the expression of Crb3DNA. 3. The results of transfection experiment showed that the DNA expression in the transfected group was lower than that in the other two groups, and the expression of DNA in the transfected group was significantly higher than that in the empty group and the untransfected group, and there was statistical significance (P0.05). 4. The results of transfection experiment showed that the testosterone content in the culture medium of the transfected group was lower than that of the empty group and the untransfected group, and had statistical significance (P0.05). Conclusion the overexpression of the 1: Crb3 protein gene in the Leydig cells of the testis may lead to the decrease of testosterone secretion in the stromal cells of the testis.
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R698

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