Elucidating mechanisms of fibrosis associated with the Crohns disease-associated pathogenic variant in the metal transporter ZIP8

阐明与克罗恩病相关的金属转运蛋白 ZIP8 致病性变异相关的纤维化机制

• 批准号: 10571146
• 负责人: Joanna Miller Peloquin Melia
• 金额: $ 12.28万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10571146
• 关键词: Affect; American; Anti-Tumor Necrosis Factor Therapy; Biological Assay; Bone Marrow; CRISPR/Cas technology; Cells; Clinical; Colitis; Collagen; Colon; Crohn' s disease; Data; Disease; Exhibits; Family; Fibroblasts; Fibrosis; Future; Genetic; Genotype; Goals; Homeostasis; Human; Human Genome; Hydroxyproline; Immune; Impairment; Inflammatory; Inflammatory Bowel Diseases; Innate Immune Response; Interleukin-11; Interleukin-6; Intestinal Fibrosis; Knock-in; Knock-in Mouse; Link; Macrophage; Macrophage Activation; Manganese; Measures; Mediating; Metals; Methods; Micronutrients; Modeling; Mucous Membrane; Mus; Operative Surgical Procedures; Orthologous Gene; PTPRC gene; Pathogenesis; Pathogenicity; Pathology; Patients; Penetration; Phenotype; Population; Positioning Attribute; Predisposing Factor; Predisposition; Prevention; Process; Production; Recovery; Refractory; Refractory Disease; Regulation; Reporting; Risk; Role; STAT3 gene; Signal Pathway; Signal Transduction; Sodium Dextran Sulfate; Stimulator of Interferon Genes; Stromal Cells; TNF gene; Tissues; Ulcerative Colitis; Variant; Western Blotting; Work; Zinc; chemokine; clinical practice; cytokine; dextran sulfate sodium induced colitis; differential expression; experience; human data; human disease; insight; member; mouse model; novel; overexpression; pre-clinical; profibrotic cytokine; programs; response; risk variant; single-cell RNA sequencing; therapeutic target; transcriptome sequencing; translational model; treatment response

PROJECT ABSTRACT Inflammatory bowel diseases (IBD), including Crohn’s disease (CD) and ulcerative colitis, affect 1 in 200 Americans. Anti-tumor necrosis factor (TNF)-alpha therapies remain the cornerstone of therapy for patients with IBD, but approximately 40% of patients will experience primary or secondary loss of response, and “anti- TNF-experienced” patients are less likely to respond to all subsequent therapies in current clinical practice. Single-cell RNA sequencing has uncovered key cellular signatures associated with anti-TNF refractoriness that strongly implicate signaling between inflammatory macrophages and activated fibroblasts. There remains a major gap in our understanding of the patient-specific factors that predispose to this aberrant macrophage- fibroblast activation, how it links to anti-TNF unresponsiveness, and the potential to modulate it to change to disease course and treatment response. Our work has focused on the functional implications of a pathogenic variant in a metal transporter, ZIP8 A391T, that dysregulates manganese homeostasis and is associated with complicated (stricturing and penetrating) Crohn’s disease. In studying ZIP8 391-Thr in a mouse model (Zip8 393T-knock-in (KI)), we have shown increased susceptibility to colitis, but even more interestingly, we have observed enhanced fibrosis. These mice also exhibit marked induction of Il-11, a pro-inflammatory and pro- fibrotic cytokine that is a hallmark of aberrant signaling between inflammatory macrophages and activated fibroblasts in patients with Crohn’s disease – particularly anti-TNF refractory disease. We therefore hypothesize that study of aberrant macrophage-fibroblast signaling is a key feature of disease pathogenesis in the Zip8 393T-KI mice. Establishing the underlying disease mechanisms are important because (1) up to 25% of patients with Crohn’s disease carry ZIP8 391-Thr in some populations and (2) the Zip8 393T-KI could serve as a novel translational model for mechanistic studies, particularly related to anti-TNF non-response. We will study this hypothesis in two aims: In Aim 1, we will determine if Zip8 393T-KI perturbs macrophage innate immune responses; in Aim 2, we will compare fibroblast activation induced by Zip8 393T-KI vs. WT Zip8 macrophages. The long-term goal is to establish if ZIP8 391-Thr genotype has clinical implications for the prevention of and treatment of patients with Crohn’s disease and use study of ZIP8 391-Thr-related pathology as key opportunity to elucidate the role of Mn homeostasis in human disease. This R03 application builds from a K08 that established the altered Mn homeostasis and fibroinflammatory phenotype in colitis in Zip8 393T-KI mice. This application with strong translational relevance will provide critical mechanistic insight of the ZIP8 genotypic effect on macrophage-fibroblast signaling and the interaction with Mn homeostasis to prioritize therapeutic targets, including STAT3 or IL-11 inhibition, and patient studies as part of an R01 application.

项目摘要 炎症性肠道疾病（IBD），包括克罗恩病（CD）和溃疡性结肠炎，有200份 美国人。抗肿瘤坏死因子（TNF） - α疗法仍然是患者治疗的基石 与IBD有关 经验丰富的”患者对当前临床实践中随后的所有疗法的反应较小。 单细胞RNA测序具有与抗TNF耐火性相关的关键细胞特征 强烈暗示炎症巨噬细胞和活化的成纤维细胞之间的信号传导。仍然有一个 我们对这种异常巨噬细胞的患者特异性因素的理解的主要差距 成纤维细胞激活，如何链接到抗TNF无反应性，并将其调节以更改为 疾病课程和治疗反应。我们的工作集中于致病性的功能含义 金属转运蛋白的变体ZIP8 A391T，使锰稳态失调，与 克罗恩病复杂（狭窄和穿透性）。在研究小鼠模型中zip8 391-thR时（zip8 393t-knock-in（Ki）），我们显示出对结肠炎的敏感性增加，但更有趣的是，我们有 观察到增强的纤维化。这些小鼠还表现出明显的IL-11诱导，这是一种促炎和促进性的 纤维化细胞因子是炎症巨噬细胞之间异常信号传导的标志 克罗恩病患者的成纤维细胞，尤其是抗TNF难治性疾病。因此，我们 假设对异常巨噬细胞 - 纤维细胞信号传导的研究是疾病发病机理的关键特征 ZIP8 393T-KI小鼠。建立潜在的疾病机制很重要，因为（1）高达25％ 在克罗恩病的患者中，有些人群携带Zip8 391-thR，（2）Zip8 393T-KI可以使用 作为机械研究的新型翻译模型，尤其是与抗TNF无反应有关的模型。我们将 在两个目的中研究了这一假设：在AIM 1中，我们将确定Zip8 393t-Ki是否存在巨噬细胞先天 免疫反应；在AIM 2中，我们将比较Zip8 393T-KI与WT ZIP8诱导的成纤维细胞激活 巨噬细胞。长期目标是确定ZIP8 391-THR基因型是否对 预防和治疗克罗恩病患者并使用Zip8 391-thR相关病理研究 作为阐明MN稳态在人类疾病中的作用的关键机会。此R03应用程序由 在Zip8 393T-KI中建立了MN稳态变化和肌炎表型的K08 老鼠。具有强大转化相关性的应用程序将提供Zip8的关键机理见解 基因型对巨噬细胞纤维细胞信号传导以及与MN稳态的相互作用以优先级 作为R01应用的一部分，包括STAT3或IL-11抑制以及患者研究在内的治疗靶标。