LCN-2基因在雄性小鼠睾丸中的表达与调节

发布时间：2018-03-17 08:36

本文选题：LCN-2　切入点：睾丸　出处：《华南农业大学》2016年硕士论文　论文类型：学位论文

【摘要】：LCN-2(lipocalin-2)名为脂质运载蛋白2,又名中性粒细胞白明胶酶相关脂质运载蛋白,属于转运蛋白中亲脂性的分泌型糖蛋白,参与动物机体多种生物功能。本研究主要采用原位杂交、免疫印迹、实时荧光定量PCR等方法,并利用发育性表达、隐睾、白消安处理等模型,分别检测LCN-2基因和蛋白在CD-1小鼠睾丸中的表达与分布情况,探讨LCN-2基因与雄性生殖的关系以及调节机制,为进一步的研究LCN-2基因在小鼠生殖发育过程中的功能提供理论依据。发育性表达模型中发现LCN-2基因在小鼠1日龄、10日龄和20日龄中未见表达,30日龄、40日龄、2月龄、4月龄、8月龄、12月龄中有表达,原位杂交结果显示在4月龄时表达量最高。隐睾模型中睾丸体积变小,HE染色曲细精管形态结构损伤,生精上皮和曲细精管出现不同程度的退化,曲细精管中只有少量或基本没有精原细胞,m RNA表达量明显上升,其中大部分在睾丸间质细胞中表达(P=0.0378)。WB结果显示蛋白水平也是明显增加(P=0.0005)。白消安模型中睾丸体积明显变小,曲细精管结构严重破坏,只剩下一层很薄的生精上皮细胞,管腔中几乎没有精子细胞。原位杂交结果显示,m RNA表达量明显上调(P=0.0232)。WB结果如同隐睾模型(P=0.0004)。实时荧光定量PCR结果显示隐睾模型和白消安模型中实验组相比于对照组变化显著,这也与原位杂交的结果相吻合。综上所述,LCN-2基因在小鼠睾丸中有表达,且在小鼠不同生长发育阶段的睾丸中表达量有所不同,在隐睾模型和白消安模型中LCN-2的表达量明显上调,说明无精子模型能够诱导LCN-2的表达,LCN-2基因可能参与精子的发生及成熟,睾丸中生殖细胞的分化和凋亡,有望给雄性不育症、睾丸炎等生殖障碍病提供一个新的研究方向,为治疗雄性不育症患者提供理论依据。
[Abstract]:LCN-2lipocalin-2 (LCN-2lipocalin-2), also known as neutrophil gelatinase-associated lipid transport protein (LCN-2lipocalin-2), belongs to the lipocalin-2 transport protein, which is a kind of lipocalin-2 protein, and is involved in many kinds of biological functions. The expression and distribution of LCN-2 gene and protein in the testis of CD-1 mice were detected by Western blot and real-time fluorescence quantitative PCR. To explore the relationship between LCN-2 gene and male reproduction and its regulatory mechanism. In order to further study the function of LCN-2 gene in mouse reproductive development, it was found in the developmental expression model that LCN-2 gene was not expressed at the age of 30 days or 40 days old or 2 months old in 10 days old and 20 days old mice. It was expressed in the age of 4 months and 8 months and 12 months of age. The results of in situ hybridization showed that the highest expression level was observed at 4 months old. In the model of cryptorchidism, the testis became smaller and the seminiferous tubules were damaged by HE staining, and the seminiferous epithelium and seminiferous tubules degenerated to different degrees. The expression of m RNA in seminiferous tubule was significantly increased in only a few or little or no spermatogonial cells, most of which were expressed in Leydig cells of testis. The results showed that the protein level was also significantly increased by 0.0005. The testicular volume in Baoxuan model became smaller obviously. The structure of the seminiferous tubules was severely damaged, leaving only a very thin layer of spermatogenic epithelial cells. There were almost no sperm cells in the lumen. The results of in situ hybridization showed that the expression of RNA was significantly up-regulated as that of the cryptorchidism model. The results of real-time fluorescence quantitative PCR showed that the experimental group was significantly different from the control group in the cryptorchidism model and the Baixiaan model. These results were consistent with the results of in situ hybridization. In conclusion, the expression of LCN-2 gene was found in mouse testis, and the expression of LCN-2 gene was different in mouse testis at different stages of growth and development, and the expression of LCN-2 was up-regulated in cryptorchidism model and Baoxuan model. These results suggest that the azoospermia model can induce the expression of LCN-2 and LCN-2 gene in spermatogenesis and maturation. The differentiation and apoptosis of germ cells in testis may provide a new research direction for male infertility, orchitis and other reproductive disorders. To provide theoretical basis for the treatment of male infertility.
【学位授予单位】：华南农业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S814.1

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