Illuminating IL-31-producing cells in allergic skin disease

照亮过敏性皮肤病中产生 IL-31 的细胞

基本信息

批准号：
10729476
负责人：
Marlys S Fassett
金额：
$ 16.15万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-15 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10729476
关键词：
Address Allergic Antibodies Atopic Dermatitis Bioinformatics Biological Products Blood CD3 Antigens CD4 Positive T Lymphocytes CD8B1 gene Cells Chronic Clinical Trials Cutaneous Cytokine Receptors Data Data Analyses Data Set Dermatitis Detection Disease Dropout Elements Enterobacteria phage P1 Cre recombinase Epithelium Family Fellowship Flow Cytometry Funding Funding Mechanisms Future Gene Expression Gene Targeting Genetic Recombination Genetic Transcription Grant Hematopoietic House Dust Mite Allergens Human IL7R gene Immunology In Vitro Individual Inflammation Inflammatory Interleukin Gene Interleukin-6 Interleukins Knock-in Knowledge Laboratories LoxP-flanked allele Lung Lymphoid Lymphoid Cell Maps Mediating Molecular Mouse Strains Mus Myelogenous Myeloid Cells National Institute of Arthritis, and Musculoskeletal, and Skin Diseases Pathology Pathway interactions Patients Phenotype Population Prurigo Pruritus Red Cross Reporter Reporting Research Research Project Grants Resolution Skin Sorting Source T-Lymphocyte T-Lymphocyte Subsets Technology Therapeutic Tissues Transgenes Transgenic Animals Transgenic Mice Transgenic Organisms Visualization cell type chronic inflammatory skin cytokine draining lymph node experimental study gene discovery in vivo inducible Cre knockout gene lymph nodes novel programs receptor single-cell RNA sequencing skin disorder therapeutic target tool transcription factor

Address Allergic Antibodies Atopic Dermatitis Bioinformatics Biological Products Blood CD3 Antigens CD4 Positive T Lymphocytes CD8B1 gene Cells Chronic Clinical Trials Cutaneous Cytokine Receptors Data Data Analyses Data Set Dermatitis Detection Disease Dropout Elements Enterobacteria phage P1 Cre recombinase Epithelium Family Fellowship Flow Cytometry Funding Funding Mechanisms Future Gene Expression Gene Targeting Genetic Recombination Genetic Transcription Grant Hematopoietic House Dust Mite Allergens Human IL7R gene Immunology In Vitro Individual Inflammation Inflammatory Interleukin Gene Interleukin-6 Interleukins Knock-in Knowledge Laboratories LoxP-flanked allele Lung Lymphoid Lymphoid Cell Maps Mediating Molecular Mouse Strains Mus Myelogenous Myeloid Cells National Institute of Arthritis, and Musculoskeletal, and Skin Diseases Pathology Pathway interactions Patients Phenotype Population Prurigo Pruritus Red Cross Reporter Reporting Research Research Project Grants Resolution Skin Sorting Source T-Lymphocyte T-Lymphocyte Subsets Technology Therapeutic Tissues Transgenes Transgenic Animals Transgenic Mice Transgenic Organisms Visualization cell type chronic inflammatory skin cytokine draining lymph node experimental study gene discovery in vivo inducible Cre knockout gene lymph nodes novel programs receptor single-cell RNA sequencing skin disorder therapeutic target tool transcription factor

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项目摘要

PROJECT SUMMARY/ABSTRACT Biologics targeting the receptor for cytokine interleukin-31 (IL-31) show promising results in advanced clinical trials for diverse inflammatory skin diseases. My new laboratory's skin immunology research program is focused on characterization of pathways that connect IL-31 producing cells and IL-31 responsive cells in chronic tissue inflammation. In this project, we will use the R03 funding mechanism and tools we developed during my NIAMS K08 fellowship-funded research to pursue fundamental open questions about the cellular identity of IL-31-producing skin cell populations. Experiments outlined here will illuminate specific IL-31 source populations and the cutaneous cellular and molecular networks they modulate. The first objective of this proposal is to perform unbiased high-resolution phenotypic analysis of all cutaneous IL-31-producing cells in vivo. To do so, we will take advantage of a novel IL31 reporter transgenic mouse strain we developed by a knock-in/knock-out gene targeting strategy. The IL31 reporter transgene enables visualization of individual live IL-31-producing cells. However, since cells that express IL-31 are sparse in the absence of tissue inflammation, IL31 reporter transgenic animals will be challenged topically with the well-characterized allergen house dust mite to provoke skin inflammation in our experiments. Readouts will include high-parameter flow cytometry plus scRNA-sequencing. CD4 T cells are expected to express the reporter transgene, but additional cells of both hematopoietic and non-hematopoietic lineages have also been proposed as IL-31 sources. If present in skin, IL31 reporter expression by these cells and other previously- undetected IL-31 sources such as innate lymphoid cells will be captured by our unbiased mapping efforts. Our second objective is to resolve downstream effects of IL-31-producing cells on other skin cell populations. To do so, we will take advantage of Cre recombinase embedded in the IL31 reporter transgene to selectively alter cells that express IL31. Specifically, mice double transgenic for the IL31 reporter and the Cre- inducible 'fate-mapper' transgene Ai14 will transgenically flag all cells that ever expressed IL31. Mice double transgenic for the IL31 reporter and Cre-inducible 'deleter' transgene Rosa26-DTA will cell-autonomously delete all cells that ever expressed IL31. By intersecting scRNA-sequencing datasets resolved from 'IL31 fate- mapper' and 'IL31 deleter' mouse skin, we will be able to distinguish IL31-expressing cells from additional dropout populations that represent direct or indirect responders. Planned bioinformatic analyses of datasets from responder cell populations can identify gene expression programs imposed by IL31-expressing cells on responder cell networks in vivo. When successful, experiments outlined in this proposal will generate significant new data for my new laboratory and identify angles for more selective therapeutic targeting of IL-31-mediated pathways in the future.

项目摘要/摘要靶向细胞因子白细胞介素-31（IL-31）受体的生物制剂显示出令人鼓舞的结果多种炎症性皮肤疾病的临床试验。我新实验室的皮肤免疫学研究计划是专注于连接IL-31产生细胞和IL-31响应细胞的途径的表征慢性组织炎症。在这个项目中，我们将使用我们开发的R03资金机制和工具在我的NIAMS K08奖学金资助的研究中，提出有关细胞的基本开放问题产生IL-31的皮肤细胞群体的身份。这里概述的实验将阐明特定的IL-31源它们调节的皮肤细胞和分子网络的种群和分子网络。该提案的第一个目的是对所有人进行公正的高分辨率表型分析皮肤IL-31在体内产生的细胞。为此，我们将利用一种新颖的IL31报告基因转基因我们通过敲入/敲除基因靶向策略开发的小鼠菌株。 IL31报告基因转基因使单个活产生的细胞可视化。但是，由于表达IL-31的细胞稀疏在没有组织炎症的情况下，IL31报告基因转基因动物将在局部挑战在我们的实验中，特征良好的过敏原房屋粉尘螨会引起皮肤炎症。读数会包括高参数流式细胞仪加上SCRNA测序。 CD4 T细胞有望表达记者转基因，但造血和非造血谱系的其他细胞也已经提议为IL-31来源。如果存在于皮肤中，则这些细胞和其他先前的IL31报告基因表达未发现的IL-31来源（例如先天淋巴样细胞）将被我们无偏图的绘制工作捕获。我们的第二个目标是解决IL-31产生细胞对其他皮肤细胞的下游影响人群。为此，我们将利用IL31报告基因中嵌入的CRE重组酶有选择地改变表达IL31的细胞。具体而言，小鼠的IL31记者双重转基因和Cre- 诱导的“命运 - 贴”转基因AI14将通过转基因标记所有表达IL31的细胞。小鼠双重 IL31记者的转基因和CRE诱导的“ deleter”转基因Rosa26-DTA将以细胞自主为单位删除所有表达IL31的细胞。通过与从'il31命运解决的scrna-seter-revering数据集 - mapper'和'il31 deleter'小鼠皮肤，我们将能够区分表达IL31的细胞与其他代表直接或间接响应者的辍学群体。计划的数据集的生物信息学分析来自响应者细胞群体可以识别由表达IL31的细胞施加的基因表达程序体内响应者细胞网络。成功后，该提案中概述的实验将为我的新数据产生重要的新数据实验室并确定将来IL-31介导的途径更具选择性的治疗靶向的角度。