树突状细胞亚群吞噬凋亡细胞并诱导免疫耐受的分子机制研究

In multicellular organisms, unnecessary or harmful cells are eliminated by apoptotic cell death. This elimination process is required to preserve tissue integrity and protect organisms from viral infection. After undergoing apoptosis, cell corpses are rapidly recognized and phagocytosed by phagocytic cells, such as macrophages and dendritic cells (DCs). Apoptotic cell clearance plays a critical role in the maintenance of self-tolerance, and that the impaired clearance of apoptotic cells is, at least, partly responsible for the etiology of autoimmune diseases...Intravenous injection of apoptotic cells could be dominantly engulfed by splenic CD8a + CD103 + DC located in the marginal zone of spleen. And the transient deletion of this DC subset caused the abberant immune tolerance induced by apoptotic cells clearance. However, the mechanism of the recognition of apoptotic cells by CD8a + CD103 + DC is still unclear. The previous research results showed that the phagocytic cells recognize apoptotic cells through their own phagocytic receptors, but CD8a + CD103 + DC subsets do not express the known phagocytic receptors: Tim4 and MFG-E8. Therefore, we speculate that the CD8a + CD103 + DC subsets express specific moleculars through which they can recognize apoptotic cells rapidly. ..To clarify the molecular mechanism of apoptotic cell clearance by DC subset, we plan to produce monoclonal antibodies against CD8a + CD103 + DC subsets, and establish hybridoma library. The hybridomas are used for phagocytosis assay to select the specific receptor involved in the phagocytosis of apoptotic cell. We also aim to understand the molecular mechanism of the self tolerance induction by apoptotic cell clearance. The expected results are helpful to understand the the etiology, pathology and treatment of the autoimmune diseases caused by the impairment of apoptotic cell clearance .

机体内的凋亡细胞如果不能及时被清除则会导致自身免疫性疾病。经静脉注射进入小鼠体内的凋亡细胞，会随着静脉血进入脾脏的边缘区，并被脾脏的CD8a+CD103+DC亚群特异性地吞噬。而且该类细胞亚群的暂时缺失能够破坏机体针对凋亡细胞的自身免疫耐受，但是其识别凋亡细胞并参与免疫耐受的机制仍不清楚。已有的研究结果证明吞噬细胞能够通过自身表达的Tim4或MFG-E8等吞噬受体识别凋亡细胞，但是CD8a+CD103+DC亚群却不表达这些已知受体。因此，我们推测CD8a+CD103+DC亚群通过其自身的特定分子识别凋亡细胞。本项目旨在利用DC亚群免疫动物的方法获得针对这一细胞表面分子的系列抗体，并利用抗体阻断细胞表面分子的原理，筛选能够调控DCs识别凋亡细胞的特异性受体，从而明确DC识别凋亡细胞并诱导免疫耐受的相关机制。本项目的研究成果可以为凋亡细胞的清除障碍导致自身免疫性疾病的病因，病理及治疗提供线索。

机体内每天都会产生大量的凋亡细胞，这些机体不需要的细胞会及时被机体的吞噬细胞清除掉。如果这些细胞不能被及时清除而堆积在体内，则会对周围组织器官造成伤害，并最终导致自身免疫性疾病。我们的前期结果证明脾脏的CD8a+CD103+ DC亚群能够特异性地吞噬经静脉注射进入小鼠体内的凋亡细胞。而且该类细胞亚群的暂时缺失能够破坏机体针对凋亡细胞的自身免疫耐受，但是其识别凋亡细胞并参与免疫耐受的机制仍不清楚。本研究项目主要致力于探讨脾脏的CD8a+CD103+ DC亚群吞噬凋亡细胞及其诱导免疫耐受的分子机制。.项目组围绕重点解决的科学问题设计实验方案和技术路线，取得了一系列研究成果。1. 确定了脾脏CD8a+CD103+ DC亚群吞噬凋亡细胞的特异性。通过脾脏DC吞噬异物实验验证了CD8a+CD103+ DC亚群具有吞噬经静脉注射的凋亡细胞的特异性，而且这种特性并不被其表面表达的已知分子所调控。2. 鉴定了脾脏DC亚群吞噬凋亡细胞的分子机制。吞噬实验证明了CD8a+CD103+ DC亚群吞噬凋亡细胞具有PS依赖性，并初步推测该亚群依靠特异性的分子识别凋亡细胞。3. 探讨了脾脏DCs诱导免疫耐受的新机制。项目组分析了机体清除凋亡细胞过程中趋化因子及其受体的变化规律，明确了CCL22/CCR4在CD8a+CD103+ DC亚群吞噬凋亡细胞并诱导免疫耐受过程中发挥关键调控作用。.DCs能够迅速摄取进入体内的外来抗原，通过发挥强大的抗原提呈作用，诱导T细胞的活化或耐受。然而到目前为止，有关DCs识别并吞噬凋亡细胞的研究报道相对较少。其原因之一在于没有一种具有强吞噬能力的DCs亚群，可以作为体内研究凋亡细胞调控过程的实验对象。该项目以脾脏的CD8a+CD103+ DC亚群为研究对象，取得了一系列研究成果。这些研究成果可以为凋亡细胞的清除障碍导致自身免疫性疾病的病因，病理及治疗提供线索。