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 炎症性肠病（IBD），包括克罗恩病（CD）和溃疡性结肠炎，影响1/200 美国人抗肿瘤坏死因子（TNF）-α治疗仍然是患者治疗的基石 但是大约40%的患者将经历原发性或继发性应答丧失，并且“抗- 在目前的临床实践中，“经历过TNF的”患者不太可能对所有后续治疗有反应。 单细胞RNA测序揭示了与抗TNF无效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 我们对易患这种异常巨噬细胞的患者特异性因素的理解存在重大差距- 成纤维细胞活化，它如何与抗TNF无反应性联系起来，以及调节它以改变成纤维细胞的潜力。 病程和治疗反应。我们的工作集中在一种致病的 一种金属转运蛋白ZIP 8 A391 T的变异体，它使锰的体内平衡失调，与 复杂性（狭窄性和穿透性）克罗恩病。在小鼠模型（Zip 8）中研究ZIP 8 391-Thr 393 T-knock-in（KI）），我们已经显示出对结肠炎的易感性增加，但更有趣的是， 观察到纤维化增强。这些小鼠还表现出显著的IL-11诱导，IL-11是一种促炎性和促炎性因子。 纤维化细胞因子是炎性巨噬细胞和活化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-特别是抗TNF难治性疾病。因此我们 假设异常巨噬细胞-成纤维细胞信号传导研究是 Zip 8 393 T-KI小鼠。建立潜在的疾病机制很重要，因为（1）高达25% 在某些人群中，100%的克罗恩病患者携带ZIP 8 391-Thr;（2）ZIP 8 393 T-KI可作为治疗克罗恩病的有效药物。 作为一种新的翻译模型的机制研究，特别是有关抗TNF无反应。我们将 研究这一假说有两个目的：在目的1中，我们将确定Zip 8 393 T-KI是否干扰巨噬细胞先天性 在目的2中，我们将比较Zip 8 393 T-KI与WT Zip 8诱导的成纤维细胞活化 巨噬细胞长期的目标是确定ZIP 8 391-Thr基因型是否具有临床意义， 克罗恩病患者的预防和治疗以及ZIP 8 391-Thr相关病理的应用研究 作为阐明锰稳态在人类疾病中的作用的关键机会。此R 03应用程序基于 K 08在Zip 8 393 T-KI中建立了改变的Mn稳态和结肠炎纤维炎表型 小鼠这种具有强烈翻译相关性的应用将提供ZIP 8的关键机制见解 基因型对巨噬细胞-成纤维细胞信号传导的影响以及与Mn稳态的相互作用， 治疗靶点，包括STAT 3或IL-11抑制，以及作为R 01应用一部分的患者研究。