Toll-like受体在小鼠Sertoli细胞中的表达及功能

发布时间：2018-02-13 06:17

本文关键词： Toll-like受体 Sertoli细胞吞噬 Tyro3家族受体阴道闭锁　出处：《中国协和医科大学》2008年博士论文　论文类型：学位论文

【摘要】： 一、Toll-like受体在小鼠Sertoli细胞中的表达及功能 Toll-like受体(Toll like receptors,TLRs)是近来发现的一种模式识别受体,识别病原微生物进化中的保守分子。在人中已发现10个成员(TLR1-10),小鼠中发现了12个成员(TLR1-9,11-13)。TLRs在介导天然免疫和获得性免疫中发挥着重要的作用。本文旨在研究TLRs在小鼠Sertoli细胞的表达及其功能。应用RT-PCR,流式细胞仪和Western blots的方法检测基因表达。免疫荧光染色用于检测NF-κB的活化。油红O染色检测脂滴的形成用来评价Sertoli细胞的吞噬功能。结果显示,TLR2-TLR5在Sertoli细胞中高表达,TLR6,TLR7和TLR13表达相对较低,而TLR1,TLR8,TLR9,TLR11和TLR12在Sertoli细胞中不表达。我们选择表达水平较高的TLR2-5进行深入研究。结果表明Sertoli细胞中的TLR2-5均可被其配体酵母多糖,poly(I:C),脂多糖和鞭毛蛋白激活,进而活化NF-κB,诱导Sertoli细胞分泌主要炎症因子(IL-1α,IL-6,IFNα及IFNβ)。TLR2-5被激活后均可诱导IL-1α和IL-6的表达,而TLR3-4的激活还可诱导IFNα及IFNβ的分泌。另外,TLR3具有调节Sertoli细胞吞噬能力的作用,并且发现TLR介导的Sertoli细胞吞噬作用与清道夫受体的表达上调有关。上述结果表明TLR2-5可能在保护生精上皮抵抗外来病原体和自身抗原,维持睾丸的免疫获免特征中发挥着重要的作用。二、Tyro3家族受体敲除小鼠呈现高比率的末端阴道闭锁阴道闭锁是女性生殖系统的常见畸形,导致阴道发育缺陷的特异分子机制仍不清楚。我们报道缺失Tyro3家族受体的雌性小鼠表现高比率的末端阴道闭锁。Tyro3受体酪氨酸激酶家族包括三个成员:Tyro3、Axl与Mer。在分析三基因敲除小鼠(Tyro3~(-/-)Axl~(-/-)Mer~(-/-),TAM)表型的过程中,发现约16%的雌性小鼠表现为末端阴道闭锁。进而对双基因敲除及单基因敲除小鼠进行分析,发现Axl~(-/-) Mer~(-/-)(AM)与Tyro3~(-/-) Mer~(-/-)(TM)小鼠的阴道闭锁率分别为4.0%和3.7%,Mer~(-/-)(M)雌性小鼠的发病率为2.5%。其它基因型小鼠Axl~(-/-) Tyro3~(-/-)(AT)、Axl~(-/-)(A)、Tyro3~(-/-)(T)及野生型小鼠从未见此变化。提示Tyro3亚家族成员在调节阴道发育上相互协调,Mer起着关键作用,Axl和Tyro3起辅助Mer的作用。Tyro3家族受体敲除小鼠可做为研究阴道闭锁发生机理的有用模型。
[Abstract]:Expression and function of Toll-like receptor in murine Sertoli cells. Toll-like receptor Toll like receptor TLRs is a recently discovered pattern recognition receptor. Ten members of TLR1-10 have been found in humans and 12 members of TLR1-9 have been found in mice. TLRs play an important role in mediating innate and acquired immunity. The purpose of this paper is to study TLRs in small mice. Expression and function of mouse Sertoli cells. Detection of gene expression by RT-PCR, flow cytometry and Western blots. Immunofluorescence staining was used to detect the activation of NF- 魏 B. Oil red O staining was used to detect lipid droplet formation to evaluate the phagocytosis of Sertoli cells. The results showed that the expression of TLR2-TLR5 was relatively low in Sertoli cells. However, TLR1, TLR9, TLR11 and TLR12 were not expressed in Sertoli cells. We selected TLR2-5 with high expression level for further study. The results showed that the TLR2-5 in Sertoli cells could be activated by its ligand yeast polysaccharide, polylipopolysaccharide, lipopolysaccharide and flagellin. Furthermore, the activation of NF- 魏 B could induce the expression of IL-1 伪 and IL-6 in Sertoli cells after the activation of IL-1 伪, IL-6, IFN- 伪 and IFN 尾. TLR2-5, and the activation of TLR3-4 could also induce the secretion of IFN 伪 and IFN 尾. In addition, TLR3 could regulate the phagocytosis of Sertoli cells, and TLR3 could regulate the phagocytosis of Sertoli cells. It was also found that the phagocytosis of Sertoli cells mediated by TLR was related to the up-regulation of scavenger receptor expression. These results suggest that TLR2-5 may protect spermatogenic epithelium from foreign pathogens and autoantigens. It plays an important role in maintaining immune immunity characteristics of testis. The tyro3 receptor knockout mice present a high rate of terminal vaginal atresia. Vaginal atresia is a common malformation of the female reproductive system. The specific molecular mechanisms leading to vaginal dysplasia are still unclear. We report a high rate of terminal vaginal atresia. Tyro3 receptor tyrosine kinase family consists of three members: Tyro3 axl and Mer. in this study, we report the presence of a high rate of terminal vaginal atresia in female mice without Tyro3 family receptors. In the process of removing the phenotype of the Tyro3- /---tam in the tri-gene knockout mice, It was found that about 16% of the female mice showed terminal vaginal atresia, and then the double gene knockout and single gene knockout mice were analyzed. It was found that the vaginal atresia rates were 4.0% and 3.7% in the Axltr / R / M and Tyro3 / R / M mice, respectively.) the incidence of vaginal atresia in the female mice was 2.55.The other genotypic mice, Axl / R / R) Tyro-R / R / R) and wild-type mice, have never seen this change. It is suggested that the Tyro3 subfamily members are regulating the vagina development phase. Mer plays a key role in assisting Mer by axl and Tyro3. Tyro3 receptor knockout mice can be used as a useful model to study the mechanism of vaginal atresia.
【学位授予单位】：中国协和医科大学
【学位级别】：博士
【学位授予年份】：2008
【分类号】：R392

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