趋化因子CCL2调控辅助性T细胞亚群分化极性对RSV感染所致严重气道病变的影响及相关机制研究

Respiratory syncytial virus (RSV) is the leading cause of respiratory viral infection in infancy with the clinical illnesses varied from common cold-like symptoms to severe bronchiolitis and pneumonitis. The severity of pulmonary inflammmation is related to the extense of mononuclear cells infiltration and expression patterns of Th related cytokines. A Th1 to Th2 bias was believed to play a major role in severe RSV bronchiolitis. Recently the over-expression of IL17 was revealed related to severe RSV disease whereas regulatory T cells (iTregs) was proved to be important in controlling the disease, thus iTregs and Th17 cells are added as new players in controlling the severity of RSV bronchiolitis. We have established mice RSV infection models with mild or servere bronchiolitis. An overpression of chemokine CCL2 was detected in the bronchoalveloar lavage (BAL) from mice with severe bronchiolitis. CCL2 is a weill studied chemokine with its chemoattract bioactivity of recruiting monocytes into the site of infection. Interestingly, CCL2 might also directly affects T cells differentiation with the mechanisms kept unrevealed. We made the hypothesis that the CCL2 work as a co-stimulatory factor for na?ve T cell activation and/or contribute to the composition of microenvironments which is critial for both Th cell differentiation and funtion. In this study, we will 1.check the direct effects of CCL2 to Th0 cells in a in vitro differentiation culture model; 2. dynamicly check the patterns of cytokines and chemokines expression in lung as well as in BAL during the course of disease; 3. check the composition of Th subpopulations and profiles of their chemokine receptor expression both in lung and in draining lumph tissues at the early and late stages of the disease; 4. use ex-vivo stimulation conditions, check the differentiation of Th0 cells in lung and track the migration of Th cells; 5. use CFSE staining in vivo cell tracking technique, study the funtional switch of Th cells from draining lymph tissues to the lung. Due to the plasticity of Th cell subpopulation, we might be able to adjust the Th responses from Th2/Th17 to Th1/iTregs bias. Our results will help for the understanding of RSV pathogenesis and provide potential targets for anti-virus therapy.

呼吸道合胞病毒(RSV)感染可引起婴幼儿严重毛细支气管炎和肺炎，其病理特征为细支气管周围单核和淋巴细胞浸润以及辅助性T细胞（Th）相关细胞因子分泌谱改变。我们发现趋化因子CCL2异常高表达与RSV感染引起气道严重病变相关，但机制不明。我们推测CCL2异常高表达除了通过趋化作用，亦可直接刺激或通过调节细胞因子内环境影响Th亚群分化及功能转化，使Th亚群从Th1、iTregs样效应转变为Th2、Th17样效应，并抑制iTregs功能，导致RSV严重感染。本项目将以CCL2高表达RSV感染小鼠为模型，深入研究CCL2对免疫细胞的趋化作用、对肺组织局部细胞因子微环境及Th分化与功能转化影响，确定CCL2效应关键靶细胞和效应分子。通过本项目研究，不仅有助于了解RSV致病机理，也将为抗病毒治疗提供潜在的靶点。具有较高创新性和重要临床意义。

RSV感染在医学上的重要性仍未消弱，目前针对RSV病毒结构、复制过程、致病过程等研究均已经较为透彻，但是目前尚无有效药物、疫苗问世。RSV与细胞、宿主之间的复杂调控作用仍然是解决这一问题的关键。本研究证实了CCL2与RSV病毒复制相关，可能在VED中起了抑制病毒复制作用，但是与免疫应答Th1/Th2失衡无关；CCL2可与细胞内关键磷酸酶Shp-2发生协同作用，抑制RSV病毒复制。本研究首次阐明在上皮细胞中，Shp-2对IFN-α产生无影响，但对IFN-α效应起正向调控作用；Shp-2可促进α-干扰素诱导的Jak/Stat1信号通路从而抑制RSV的复制；在单核-巨噬细胞中发现Shp-2直接抑制RSV的复制；而RSV感染髓样细胞系Shp-2敲除小鼠，其RSV在小鼠体内复制没有影响。此外，本项目发现一种磷酸激酶Riok3在RSV/细胞/宿主相互作用中可能起到重要调控作用。本研究对CCL2，Shp-2，Riok3在不同来源细胞内功能的阐明将有助于加深对RSV病毒本身、病毒与宿主相关性，尤其是干扰素抗病毒效应的了解。在本项目研究研究结果基础上，利用这几年快速发展的RNA SEQ以及机器学习技术，我们的研究将为疫苗研究及药物靶点研究提供更充分的依据。