FSH对支持细胞增殖相关基因表达的影响

发布时间：2018-05-24 08:32

本文选题：支持细胞 + FSH　；参考：《东北农业大学》2015年硕士论文

【摘要】：睾丸支持细胞位于曲精细管中,是精子发生的微环境的主要构成。许多激素和影响因子都参与了睾丸支持细胞增殖调控,其中FSH是调节睾丸支持细胞增殖进程的重要激素之一。因此,研究FSH对犊牛睾丸支持细胞增殖的调节机制不仅能为人工调控支持细胞增殖奠定一定基础,而且对提高雄性动物的生殖能力也具有十分重要的意义。本实验以新生犊牛睾丸为实验材料,利用胰酶、胶原酶两步消化法对支持细胞进行分离,差速贴壁法纯化支持细胞,进行孚尔根染色法、HE染色法以及油红O染色法鉴定支持细胞;以培养的犊牛支持细胞为实验材料,用不同浓度的FSH(0、0.01、0.02、0.04、0.08 IU/ml)对支持细胞进行处理,利用CCK8检测五组浓度6个时间点支持细胞增殖情况并用RealtimePCR检测一定浓度FSH处理后CDC25等基因表达情况。两步酶法可以对犊牛睾丸支持细胞进行分离纯化,经孚尔根染色等方法鉴定纯度大于95%;体外培养条件下的支持细胞能够传至第20代,并且随着传代次数的增加,支持细胞的增殖速度越来越慢。FSH可以显著提高睾丸支持细胞体外增殖的效率,0.04 IU/ml的FSH能显著地促进支持细胞体外增殖。FSH显著促进WNT/β-catenin信号通路相关基因表达量的提高,可以通过该信号通路促进支持细胞的增殖。FSH对CDC25A基因表达影响不显著,在一定浓度范围内,FSH显著促进CDC25B的表达。CDC25B可能参与促进支持细胞增殖作用,其可能决定支持细胞的增殖速度。在一定浓度范围内,FSH显著促进P53的表达量,P53可能在FSH刺激性下对支持增殖起到控制作用;FSH显著抑制FAS和CDC25C的表达量,同时显著提高FASL的表达量。
[Abstract]:Testicular Sertoli cells, located in the tubules, are the main components of the microenvironment of spermatogenesis. Many hormones and influencing factors are involved in the regulation of testicular Sertoli cell proliferation, and FSH is one of the important hormones to regulate the proliferation of testicular Sertoli cells. Therefore, the study of the regulatory mechanism of FSH on the proliferation of testicular Sertoli cells in calves can not only lay a foundation for artificial regulation of Sertoli cell proliferation, but also play an important role in improving the reproductive ability of male animals. Sertoli cells were isolated from newborn calf testis by trypsin and collagenase digestion, and Sertoli cells were purified by differential adhesion method. Sertoli cells were identified by HE staining and oil red O staining, and Sertoli cells were treated with different concentrations of FSHO 0. 01 and 0. 02O 0. 04U / ml, respectively, and cultured calf Sertoli cells were used as experimental materials. CCK8 was used to detect the proliferation of Sertoli cells at 6 time points in five groups and RealtimePCR was used to detect the expression of CDC25 and other genes after treatment with certain concentration of FSH. Two-step enzymatic method was used to isolate and purify testicular Sertoli cells from calves. The purity of Sertoli cells in vitro was higher than 95. The Sertoli cells cultured in vitro could be transmitted to the 20th passage, and with the increase of passage times, the purity of Sertoli cells was higher than 95%. The proliferation of Sertoli cells was slower and slower. FSH could significantly improve the proliferation efficiency of testicular Sertoli cells in vitro. FSH of 0.04 IU/ml could significantly promote the proliferation of Sertoli cells in vitro. FSH significantly promoted the expression of genes related to WNT/ 尾 -catenin signaling pathway. FSH can promote the proliferation of Sertoli cells through this signaling pathway. FSH has no significant effect on the expression of CDC25A gene. In a certain concentration range, FSH can significantly promote the expression of CDC25B. CDC25B may play a role in promoting the proliferation of Sertoli cells. It may determine the rate of proliferation of Sertoli cells. In a certain concentration range, FSH significantly promoted the expression of p53. The expression of p53 may play a controlling role in supporting proliferation under the stimulation of FSH. FSH significantly inhibited the expression of FAS and CDC25C, and significantly increased the expression of FASL.
【学位授予单位】：东北农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S814

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