Assessing the long-term consequences of inflammation on epidermal stem cell homeostasis and function

评估炎症对表皮干细胞稳态和功能的长期影响

• 批准号: 9191760
• 负责人: Samantha B. Larsen
• 金额: $ 4.36万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9191760
• 关键词: ATAC-seq; Acute; Address; Adult; Affect; Age; Aging; Atopic Dermatitis; Autoimmunity; Bioinformatics; C57BL/6 Mouse; Cell Death; Cells; Chromatin; Chronic; Cutaneous; Data; Development; Disease; Embryo; Enhancers; Epidermis; Epigenetic Process; Epithelial; Epithelium; Etiology; Exhibits; Gene Expression; Genes; Histones; Homeostasis; Imiquimod; Immune; Immune response; Inflammation; Inflammatory; Inflammatory Response; Injection of therapeutic agent; Injury; Intervention; Keratin; Knock-out; Label; Learning; Life; Malignant Neoplasms; Maps; Mediating; Memory; Migration Assay; Modeling; Molecular; Monitor; Mus; Organ; Organism; Pathology; Pathway interactions; Physiological; Play; Population; Process; Promoter Regions; Psoriasis; Reporter; Resolution; Role; Secondary to; Signal Transduction; Skin; Skin Tissue; Staging; Stem cells; Stress; Subfamily lentivirinae; Surface; System; T-Lymphocyte; Tamoxifen; Technology; Testing; Thick; Time; Tissues; Toxic Environmental Substances; Transgenes; Wound Healing; activating transcription factor; assault; base; cell type; chronic wound; differential expression; experience; fitness; improved; in utero; in vivo; insight; interest; mouse model; novel; pathogen; progenitor; promoter; response; self-renewal; small hairpin RNA; stem cell population; transcription factor; transcriptome; transcriptome sequencing; vector; wound

Project Summary The skin, the largest and most external organ in the body, is routinely subject to inflammatory insults. As such the skin, specifically the epidermis, is tasked with maintaining a physical barrier in the face of pathogenic and toxic insults. Importantly, cutaneous epithelia are replenished by a small pool of epidermal stem cells (EpdSCs) capable of self-renewing and differentiating into the various lineages of the tissue in healthy and disease states. Yet, to date, the lasting impact of inflammation on these precious tissue stem cells has not been examined. My preliminary data indicate that EpdSCs can sense and respond to inflammatory signals, and sustain epithelial hyperproliferation. Intriguingly, I find that inflammation-experienced epidermis is faster at full thickness wound closure than age-matched control skin indicating that inflammation causes long-term changes to tissue fitness and function. Based on these early findings, I hypothesize that EpdSCs are likely altered as a result of inflammation. Using a well-defined murine model of T cell mediated skin inflammation, I aim to systematically dissect specifically how EpdSCs are altered by inflammation and elucidate the consequences of such alterations on tissue fitness. To this end, I will 1) characterize the long-lasting transcriptional and epigenetic changes within EpdSCs and changes to the skin milieu, post-inflammation, 2) assess differences in tissue fitness and function within the post-inflamed epidermis, and lastly, 3) test the functional significance of identified genes and pathways for their contribution to the sustained differences within the inflammation-experienced epidermis. I propose using cell type/stage-specific CreER+; Rosa26YFPfl/fl mice to lineage trace and FACS purify the progenitor and differentiating populations of the epidermis as well as inflammation-experienced epidermal cells. To dissect intrinsic changes within the EpdSCs post-inflammation, I will perform ATAC-Seq in conjunction with transcriptome analysis (RNA-seq). Genes of interest will be those with sustained differential expression after inflammation and which are candidates for direct targets of the transcription factors activated in EpdSCs after inflammation. To determine their functional significance, I will knock them down in vivo using shRNA and an established in utero lentiviral injection system and test their consequences to the rapid immune response. Because EpdSCs are long-lived cells, they represent ideal targets for efficacious long-term interventions. Additionally, because these adult epidermal stem cells are central to the development of cancers, chronic wounds and epidermal aging, as well as disorders associated with accumulating inflammatory stresses, understanding how an epidermal stem cell is impacted by inflammation will provide novel mechanistic insights into the etiology and pathology of these epithelial diseases.

项目摘要 皮肤是人体最大和最外部的器官，经常受到炎症性损伤。作为 这样的皮肤，特别是表皮，的任务是在面对病原体时保持物理屏障。 有毒的侮辱。重要的是，皮肤上皮细胞由一小群表皮干细胞补充 EPDSC能够自我更新并分化成健康和健康人的各种组织谱系， 疾病状态。然而，到目前为止，炎症对这些珍贵的组织干细胞的持久影响还没有 被检查过了我的初步数据表明，EPDSC可以感知并响应炎症信号， 维持上皮细胞过度增殖有趣的是，我发现经历炎症的表皮在 与年龄匹配的对照皮肤相比，全层伤口闭合表明炎症导致长期 组织适应性和功能的变化。基于这些早期发现，我假设EPDSC可能是 由于炎症而改变。使用明确的T细胞介导的皮肤炎症小鼠模型， 旨在系统地剖析EPDSC如何被炎症改变，并阐明EPDSC的作用机制。 这种改变对组织适应性的影响。为此，我将1）描述持久的 EpdSC内的转录和表观遗传变化以及炎症后皮肤环境的变化，2） 评估发炎后表皮内组织适应性和功能的差异，最后，3）测试 鉴定的基因和途径的功能意义，因为它们对内部持续差异的贡献， 炎症表皮。我建议使用细胞类型/阶段特异性CreER+; Rosa 26 YFPfl/fl小鼠 为了谱系追踪和FACS纯化表皮的祖细胞和分化群体， 炎症经历的表皮细胞。为了剖析炎症后EpdSCs的内在变化，我 将进行ATAC-Seq结合转录组分析（RNA-seq）。感兴趣的基因将是那些 在炎症后具有持续的差异表达，并且是免疫调节剂的直接靶点的候选者。 炎症后EpdSC中激活的转录因子。为了确定它们的功能意义，我将 使用shRNA和建立的子宫内慢病毒注射系统在体内敲低它们，并测试它们的 快速免疫反应的后果。由于EpdSC是长寿细胞，它们代表了理想的 长期有效干预的目标。此外，由于这些成体表皮干细胞是 在癌症、慢性伤口和表皮老化以及相关疾病的发展中起着重要作用 随着炎症压力的积累，了解表皮干细胞如何受到 炎症将提供新的机制的见解，这些上皮细胞的病因和病理学， 疾病