A novel CD4+ T cell subset associated with allergy protection

项目摘要 与过敏相关的疾病，如哮喘，是一个日益严重的健康问题，目前还没有治愈的解决方案。 对常见过敏原应答的CD4+辅助性T细胞（TH）在驱动气道炎症中发挥重要作用 哮喘病为了更好地了解过敏和哮喘患者T细胞亚群的多样性，我们分析了单细胞 来自哮喘患者和非哮喘患者的约50，000个屋尘螨（HDM）过敏原反应性TH细胞的转录组， 有无HDM过敏。根据我们的分析，除了典型的细胞群如TH2、TH17和TH1， 我们鉴定了一种新的过敏原反应性TH细胞亚群，其特征在于IFN应答基因标记， 我们称之为THIFNR细胞（Seumois等，Science Immunology，2020）。THIFNR细胞的比例为 与过敏性患者相比，非过敏性个体中显著增加，表明过敏原特异性 甚至在非过敏性个体中宿主特异性反应。此外，过敏原反应性的唯一存在 TH2细胞在过敏性患者中的作用表明THIFNR细胞在非过敏性患者中的保护作用（抗TH2应答）。 暴露于过敏原的过敏患者。这种潜在的保护作用在我们的体外研究中得到了加强 显示由THIFNR细胞产生的TNF相关凋亡诱导配体（TRAIL）直接抑制T细胞增殖， 由TCR接合触发的激活。在后续的研究中，我们在病毒反应性TH中发现了THIFNR细胞， 细胞直接针对流感或SARS-CoV2，表明这些细胞在免疫反应中的作用更广泛。还有， 我们在大量健康个体中发现THIFNR细胞作为稳定的TH亚群。因为最近 虽然THIFNR细胞的发现，但对其起源、分化、表型和功能知之甚少。我们 假设这些THIFNR HDM反应性T细胞可以通过TRAIL参与抑制 过敏和哮喘中的TH2炎症。 在目标1中，我们将定义THIFNR细胞的功能特性。我们将进行共培养实验， THIFNR细胞以TRAIL依赖性方式抑制TH2细胞增殖和功能的能力。在aim中 2、确定体外诱导THIFNR细胞分化的最佳条件，并比较THIFNR细胞在体外诱导分化的效果。 功能特性与体内产生的功能特性相同。最后，在目标3中，我们将确定HDM反应性THIFNR 细胞在体内表现出持久性和可塑性。我们将进行HDM反应性的单细胞TCR测序， 从HDM患者纵向采集的血样中分离的THIFNR细胞和其他记忆性CD4 + T细胞亚群 过敏和非过敏受试者。通过比较纵向样本中的TCR库，我们将评估 THIFNR细胞在体内的持久性。总的来说，R21计划中的功能研究将提供重要的见解 转化为这种新的CD4 + T细胞亚群;并形成未来R01应用的基础，以进行转化研究 发现诊所。

PROJECT SUMMARY Disease associated with allergies such as asthma are a rising health problem with no current curative solutions. CD4+ helper T cells (TH) that respond to common allergens play an important role in driving airway inflammation in asthma. To better understand the diversity of T cell subsets in allergy and asthma, we analyzed the single-cell transcriptome of ~50,000 house dust mite (HDM) allergen-reactive TH cells from asthmatics and non-asthmatics, with and without HDM allergy. From our analysis, besides canonical clusters of cells such as TH2, TH17, and TH1, we identified a novel subset of allergen-reactive TH cells characterized by an IFN responsive gene signature that we called THIFNR cells (Seumois et al. Science Immunology, 2020). Proportions of THIFNR cells were significantly increased in nonallergic individuals compared to allergic patients, suggesting an allergen-specific host specific response even in non-allergic individuals. Moreover, the exclusive presence of the allergen-reactive TH2 cells in the allergic patients suggests a protective role (anti-TH2 response) of the THIFNR cells in the non- allergic patients with exposure to allergen. This potential protective role was reinforced by our in vitro studies showing that TNF-related apoptosis-inducing ligand (TRAIL) produced by THIFNR cells directly inhibits T cell activation triggered by TCR engagement. In follow-up studies, we found THIFNR cells among viral-reactive TH cells directed towards Flu or SARS-CoV2, suggesting a broader role of those cells in immune responses. Also, we found THIFNR cells as a stable TH subset in a large cohort of healthy individuals. Because of the recent discovery of THIFNR cells, very little is known about their origins, differentiation, phenotype, and function. We hypothesize that these THIFNR HDM-reactive T cells could play a role through TRAIL engagement in dampening TH2 inflammation in allergy and asthma. In Aim 1, we will define the functional properties of THIFNR cells. We will perform co-culture experiments to test the ability of THIFNR cells to inhibit proliferation and function of TH2 cells in a TRAIL-dependent manner. In Aim 2 we will determine optimal conditions to induce differentiation of THIFNR cells in vitro and compare their functional properties with those generated in vivo. Finally, in Aim 3, we will determine if HDM-reactive THIFNR cells show persistence and plasticity in vivo. We will perform single-cell TCR sequencing of HDM-reactive THIFNR cells and other memory CD4+ T cell subsets isolated from longitudinally collected blood samples of HDM allergic and non-allergic subjects. By comparing TCR repertoire in longitudinal samples, we will assess persistence of THIFNR cells in vivo. Overall, functional studies in this R21 program will provide important insights into this novel CD4+ T cell subset; and form the basis of a future R01 application to conduct studies translating findings to the clinic.