Uncover the new subsets of epidermal Langerhans cells

发现表皮朗格汉斯细胞的新亚群

• 批准号: 9912418
• 负责人: Indra Adrianto
• 金额: $ 37.63万
• 依托单位: HENRY FORD HEALTH SYSTEM
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-17 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9912418
• 关键词: Adult; Aging; Antigen-Presenting Cells; Autoimmune Diseases; Autoimmune Process; Bioinformatics; Biological Markers; Biology; Birth; Bone Marrow; C-Type Lectins; Cell Differentiation process; Cell Maturation; Cell model; Cells; Cellular Assay; Cellular biology; Chromatin; Data; Dendritic Cells; Development; Disease model; Embryo; Embryonic Development; Epidermis; Fetal Liver; Goals; Growth; Heterogeneity; Human; Hypersensitivity; Immune; Immune Tolerance; Immunity; Immunologist; In Vitro; Individual; Infection; Inflammation; Inflammatory; Intervention; Knockout Mice; Langerhans cell; Lead; Leishmaniasis; Maps; Mediating; Minor; Modeling; Molecular Profiling; Mus; Mutation; Mycoses; Names; Outcome; Peripheral; Phase; Phenotype; Play; Population; Procedures; Regulatory Element; Regulatory T-Lymphocyte; Reporter; Research; Research Personnel; Resolution; Role; Skin; Skin Cancer; Sorting - Cell Movement; T-Lymphocyte; Testing; Time; Transposase; Variant; Vitiligo; Work; Yolk Sac; activating transcription factor 3; adaptive immunity; base; epigenomics; experience; genetic signature; immune function; immunoregulation; in vivo; insight; langerin; macrophage; melanoma; monocyte; mouse model; new technology; progenitor; single cell analysis; single-cell RNA sequencing; skin disorder; transcriptome; transcriptome sequencing; transcriptomics

PROJECT SUMMARY Langerhans cells (LCs) are skin-resident dendritic cells (DCs) expressing the C-type lectin Langerin (CD207) that mediate both adaptive immunity and immune tolerance in skin and are involved in various types of skin diseases. Adult LCs are originated from embryonic yolk-sac-derived macrophages and fetal liver monocytes in the steady state. Interestingly, LCs could also be derived from the bone marrow or peripheral monocytes and repopulate the skin under inflammatory conditions. However, due to the lack of molecular profiles at individual LC level, a significant gap remains in our understanding on how a single CD207+ epidermal LC population can induce both immunity and tolerance. Fortunately, new technologies such as the single-cell RNA-sequencing (scRNA-seq) can evaluate cell-to-cell transcriptomic variation, while the single-cell assay for transposase- accessible chromatin using sequencing (scATAC-seq) can assess the epigenomic heterogeneity at single-cell resolution in an unbiased manner. Recently, we identified two major LC subsets in mice, ATF3+Bal2a1b- (mLC1) and ATF3-Bal2a1b+(mLC2) subsets, and three major LC subsets in human including ATF3+ (hLC1) subset using scRNA-seq. We also found in ATF3 knockout mice that lack of ATF3 enhances LC maturation and promotes LCs-induced Th1 and Th17 cell differentiation suggesting immune suppressive function induced by ATF3+LC1. Hence, these preliminary data support our hypothesis that LCs are heterogeneous consisting of distinct subsets with different immune functions. Our objective is to use single-cell analysis platforms plus the LC fate-mapping and mutation mouse models to further validate this. We will pursue two Specific Aims in the R61 phase: Aim 1) Characterize the gene signatures and regulatory elements of mLC1 and mLC2 by profiling LCs during embryonic, young, and aging development at steady-state and repopulated LCs at inflamed state using scRNA-seq and scATAC-seq; Aim 2) Generate ATF3negEGFP reporter mice to fate-map ATF3+LC1 embryonic development and the dynamic change of ATF3+LC1 and ATF3-LC2 subset at steady state during adult and aging development and at inflammatory state and functionally characterize LC subsets in vitro by sorting ATF3EGFP+ LC1 and ATF3- LC2 cells and rederiving ATF3.loxp mice, which will be crossed with hLangerin-Cre mice to generate LC- specific/time-induced ATF3KO for in vivo functional study. In the R33 phase, we will pursue the following Specific Aim: Aim 3) Functionally characterize ATF3+LC1 subset in vivo using LC-specific ATF3 deletion hLCcre.ATF3KO mice to evaluate the potential immune regulation function of ATF3+LC1 subset in the different disease models, including autoimmune vitiligo, melanoma, and fungi infection models. Our work will uncover the mystery of LC subsets with their specific functions, which will provide new insights into the biology of LCs and lead to the development of LC-based intervention strategies for skin diseases.

项目摘要 朗格汉斯细胞（LC）是皮肤驻留的树突状细胞（DC），表达C型凝集素朗格林（CD 207） 其介导皮肤中的适应性免疫和免疫耐受，并涉及各种类型的皮肤 疾病成人LC来源于胚胎卵黄囊来源的巨噬细胞和胎肝单核细胞， 稳定状态有趣的是，LC也可以来源于骨髓或外周单核细胞， 在炎症条件下重新填充皮肤。然而，由于缺乏个体的分子特征， LC水平，在我们理解单个CD 207+表皮LC群体如何 诱导免疫和耐受。幸运的是，新技术，如单细胞RNA测序， （scRNA-seq）可以评估细胞间转录组变异，而转座酶-RNA的单细胞测定法可以评估细胞间转录组变异。 使用测序的可接近染色质（scATAC-seq）可以评估单细胞中的表观基因组异质性， 以公正的方式解决问题。最近，我们在小鼠中鉴定了两种主要的LC亚群，ATF 3 + Bal 2a 1b-（mLC 1）。 和ATF 3-Bal 2a 1b+（mLC 2）亚群，以及包括ATF 3+（hLC 1）亚群在内的人中的三种主要LC亚群， scRNA-seq.我们还发现，在ATF 3基因敲除小鼠中，缺乏ATF 3可增强LC成熟， LC诱导的Th 1和Th 17细胞分化表明ATF 3 + LC 1诱导的免疫抑制功能。 因此，这些初步数据支持我们的假设，即LC是由不同的子集组成的异质性 不同的免疫功能。我们的目标是使用单细胞分析平台加上LC命运图 和突变小鼠模型来进一步验证这一点。我们将在R61阶段实现两个具体目标： 通过分析胚胎期间的LC来表征mLC 1和mLC 2的基因特征和调控元件， 在稳态下的年轻和老化发育和在发炎状态下的再增殖LC， scATAC-seq;目的2）产生ATF 3 + EGFP报告小鼠以对ATF 3 + LC 1胚胎发育进行命运图谱绘制， ATF 3 + LC 1和ATF 3-LC 2亚群在成年和衰老过程中稳态动态变化 并通过分选ATF 3 EGFP + LC 1和ATF 3-EGFP + LC 1在体外功能上表征LC亚群。 LC 2细胞和再衍生的ATF3.loxp小鼠，其将与hLangerin-Cre小鼠杂交以产生LC-1细胞。 特异性/时间诱导的ATF 3 KO用于体内功能研究。在R33阶段，我们将追求以下具体目标： 目的：目的3）使用LC特异性ATF 3缺失hLCcre.ATF3KO在体内功能表征ATF 3 + LC 1亚群 评价ATF 3 + LC 1亚群在不同疾病模型中的潜在免疫调节功能， 包括自身免疫性白癜风、黑素瘤和真菌感染模型。我们的工作将揭开LC的神秘面纱 亚群及其特定功能，这将为LC的生物学提供新的见解，并导致 开发基于LC的皮肤病干预策略。