Leydig细胞中Toll样受体（TLR）介导的睾丸天然免疫反应

发布时间：2018-03-06 00:35

本文选题：Leydig细胞　切入点：TLR受体　出处：《北京协和医学院》2011年博士论文　论文类型：学位论文

【摘要】：背景与目的：睾丸是免疫豁免组织,然而睾丸仍能有效的清除感染,说明睾丸自身具有抵抗病原体的机制。本论文的主要目的是研究Leydig细胞在睾丸中的天然免疫功能。睾丸巨噬细胞表现出较低的免疫活性,其机理尚不清楚,我们研究了Leydig细胞对巨噬细胞免疫反应的调节。材料与方法：利用C57/BL6小鼠为模型分离纯化小鼠Leydig细胞。利用定量实时RT-PCR (Q-PCR)分析基因mRNA的水平。使用Western blotting和免疫组化染色研究蛋白的表达定位。ELISA检测细胞因子产生和睾酮合成。利用基因敲除小鼠模型研究TAM受体在TLR介导天然免疫反应中的作用。结果：小鼠Leydig细胞中表达较高水平的TLR3和TLR4,以及较低水平的TLR2、TLR7、TLR9和TLR12,但不表达其它TLR。Leydig细胞中的TLR3和TLR4可被其受体激活,会引起核因子κB (NF-kB)和干扰素调节因子3(IRF3)的活化,进而诱导该细胞分泌炎症因子(IL-1β,IL-6和TNF-α)口I型干扰素(IFN-α及IFN-β),对入侵病原体产生免疫防御作用。为防止睾丸组织持续产生慢性炎症反应,Leydig细胞中TLR3和TLR4受体的激活必须受到严格的调控。研究发现Leydig细胞中同时表达Axl和Mer受体,它们以协同的方式抑制TLR3和TLR4信号传导,调节Leydig细胞的天然免疫反应。敲除Axl及Mer双基因敲除(Axl-/-Mer-/-)小鼠的Leydig细胞中TLR3和TLR4激活水平显著升高。Gas6是TAM受体的特异性配体,可以显著抑制Leydig细胞中TLR-介导的NF-κB和IRF-3的激活及炎症因子的表达,而这种抑制效应在Axl-/-Mer-/- Leydig细胞中则不存在。这些结果说明Axl和Mer受体酪氨酸激酶在Leydig细胞中以协同的方式负调控TLR信号。另外我们发现TLR3与TLR4的活化可以抑制Leydig细胞合成睾酮的能力。实验表明Leydig细胞可以抑制巨噬细胞的免疫反应能力,可能对维持睾丸免疫平衡和免疫豁免状态具有重要作用。结论：Leydig细胞可以通过TLR3和TLR4的激活启动睾丸天然免疫反应；TAM受体可以负调控Leydig细胞中TLR信号传导；Leydig细胞可以抑制巨噬细胞的免疫反应。这些结果表明Leydig细胞对睾丸抵抗入侵病原体,维持睾丸免疫平衡及免疫豁免状态可能发挥重要作用。
[Abstract]:Background & objective: the testis are immune free tissue, but the testis can still effectively clear the infection. The main purpose of this paper is to study the innate immune function of Leydig cells in testis. We studied the regulation of macrophage immune response by Leydig cells. Materials and methods: mouse Leydig cells were isolated and purified by C57 / BL6 mouse model. Quantitative real-time RT-PCR Q-PCR was used to analyze the level of gene mRNA. Western blotting and immunohistochemical staining were used to study the expression localization of protein. Elisa was used to detect cytokine production. The role of TAM receptor in the innate immune response mediated by TLR was studied by gene knockout mouse model. Results: higher levels of TLR3 and TLR4, and lower levels of TLR2TLR7, TLR9 and TLR12 were expressed in mouse Leydig cells, but no expression of TLR3 and TLR4 in other TLR.Leydig cells could be activated by their receptors, which resulted in activation of nuclear factor- 魏 B (NF-kB) and interferon regulatory factor 3IRF3 (IFR3). Furthermore, the cells were induced to secrete inflammatory cytokines such as IL-1 尾, IL-6 and TNF- 伪, and IFN- 伪 and IFN- 尾 were induced to secrete IFN- 伪 and IFN- 尾. In order to prevent the sustained chronic inflammatory response of testicular tissues and the activation of TLR3 and TLR4 receptors in Leydig cells, IFN- 伪 and IFN- 尾 were induced to induce the secretion of IFN- 伪 and IFN- 尾. It was found that both Axl and Mer receptors were expressed in Leydig cells. They synergistically inhibited TLR3 and TLR4 signal transduction and regulated the innate immune response of Leydig cells. TLR3 and TLR4 activation levels in Leydig cells of Axl and Mer knockout mice were significantly increased. Gas6 was a specific ligand of TAM receptor. TLR- mediated activation of NF- 魏 B and IRF-3 and the expression of inflammatory factors in Leydig cells were significantly inhibited. These results suggest that Axl and Mer receptor tyrosine kinase negatively regulate TLR signal in Leydig cells in a synergistic manner. In addition, we found that the activation of TLR3 and TLR4 can inhibit Leydig fine. The ability of cells to synthesize testosterone. Experiments have shown that Leydig cells inhibit the immune response of macrophages. It may play an important role in maintaining testicular immune balance and immune immunity. Conclusion TLR3 and TLR4 can activate testicular innate immunoreactive tam receptor, which can negatively regulate the TLR signal transduction in Leydig cells and inhibit the immune response of macrophages. These results suggest that Leydig cells can inhibit the immune response of macrophages. The testicles resist invading pathogens, Maintaining testicular immune balance and immune immunity may play an important role.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R392.1

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