Septins in intestinal fibrosis

肠道纤维化中的脓毒症

• 批准号: 10656661
• 负责人: Andrei Ivanovich Ivanov
• 金额: $ 63.01万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10656661
• 关键词: Acceleration; Actomyosin; Affect; Animal Model; Anti-Inflammatory Agents; Atlases; Attenuated; Automobile Driving; Binding; Biology; Cell Differentiation process; Cell physiology; Cells; Collagen; Complex; Complication; Crohn' s disease; Cytoskeletal Proteins; Cytoskeleton; Data; Deposition; Development; Disease; Disease Progression; Effector Cell; Endocytosis; Excision; Exposure to; Extracellular Matrix; Extracellular Matrix Proteins; Fibroblasts; Fibronectins; Fibrosis; Filament; Flagellin; Fostering; GTP-Binding Proteins; Gene Expression; Generations; Genetic; Genetic Induction; Goals; Growth Factor; Health Benefit; Human; Immune; In Vitro; Incidence; Inflammation; Intestinal Fibrosis; Intestinal Obstruction; Intestines; Investigation; Link; LoxP-flanked allele; Maps; Mediating; Membrane; Microfilaments; Molecular; Motor; Mus; Myofibroblast; Myosin ATPase; Operative Surgical Procedures; Organ; Organelles; Patients; Phenotype; Polymers; Prevention; Production; Property; Protein Isoforms; Proteins; Receptor Signaling; Regulation; Resolution; Role; Signal Transduction; Small Interfering RNA; Structure; Testing; Thick; Tissue Sample; Tissues; Transforming Growth Factors; Translations; United States National Institutes of Health; Up-Regulation; Vesicle; antifibrotic treatment; cell motility; fibrogenesis; gain of function; genetic approach; gut inflammation; improved; in vivo; loss of function; migration; mouse model; novel; novel strategies; novel therapeutic intervention; novel therapeutics; overexpression; pharmacologic; posttranscriptional; prevent; scaffold; self assembly; single-cell RNA sequencing; therapeutically effective; trafficking; translatome

ABSTRACT Over their disease course more than half of Crohn’s disease (CD) patients develop fibrosis-induced intestinal obstruction and ultimately require surgery. No specific anti-fibrotic therapies are available. Despite advances of anti-inflammatory therapies the incidence of strictures remains high, suggesting that inflammation- independent mechanisms are crucial in the progression of the disease. The main effector cell mediating fibrosis is the myofibroblast that is activated by multiple pro-fibrotic growth factors, such as transforming growth factor (TGF)-1. Such activation results in accelerated secretion of extracellular matrix (ECM) and remodeling of the actomyosin cytoskeleton. Septins are understudied cytoskeletal proteins that regulate secretory and actomyosin- dependent cellular functions. No data on the roles and regulation of the septin cytoskeleton in intestinal fibrosis exists. Our preliminary data shows a high gene expression of septins 2, 6, 7, 8, 9, 10, 11 in human intestinal tissues with septin 7 as the most predominant isoform, which is upregulated in CD. Pharmacologic or genetic disruption of the septin cytoskeleton inhibited TGF-β1-dependent increase in ECM production (Collagen I & fibronectin) and migration in immortalized and primary human myo/fibroblasts. Preliminary evidence suggests this is post-transcriptionally regulated. Septin modulation improved experimental murine fibrosis. We hence propose to investigate the hypothesis that remodeling of the septin cytoskeleton is a driver of intestinal fibrosis and targeting the septin cytoskeleton is a novel approach to therapy of fibrostenosing Crohn’s disease. This hypothesis will be tested by three specific aims: AIM1. Characterization of alterations in septin expression and distribution in tissue samples of IBD patients. This includes development of a high-resolution map of septin expression profiles in human intestinal tissues and primary human intestinal myofibroblasts, including generation of the first full thickness single cell RNA sequencing gut atlas for stricturing CD and controls. AIM2. Investigation of the roles and mechanisms of septin dependent regulation of pro-fibrotic myofibroblast activation. We will test if septin disruption or overexpression modulates TGF-β1-signaling, intracellular vesicular trafficking or the translatome and post-transcriptionally regulated networks using a loss-of-function and gain-of- function approach. AIM3. Functional exploration if targeting septins ameliorates intestinal fibrosis in vivo. We will modulate septins in vivo using a pharmacologic and genetic approach and induce experimental fibrosis in two different animal models. We will temporally control the deletion of the central septin 7 prior to (prevention) and after induction (reversal) of experimental intestinal fibrosis specifically in Col I positive cells. If successful, this proposal will challenge the paradigm of purely immune-driven ECM deposition driving stricture formation and provide proof-of-concept for a novel mechanism to prevent or treat stricture associated intestinal obstruction in CD patients.

抽象的 在他们的疾病病程中，克罗恩氏病（CD）患者的一半以上是纤维化引起的肠道 阻塞并最终需要手术。没有特定的抗纤维化疗法可用。尽管进步 抗炎疗法的狭窄事件仍然很高，表明注射 - 独立机制对于疾病的发展至关重要。主要效应细胞介导纤维化 是由多种促纤维化生长因子激活的肌纤维细胞，例如转化生长因子 （TGF）-1。这种激活导致细胞外基质（ECM）的分泌加速和重塑 肌动蛋白细胞骨架。 septins被理解为调节秘密和肌动蛋白的细胞骨架蛋白 依赖性细胞功能。没有关于肠道septin细胞骨架在肠道中的作用和调节的数据 存在纤维化。我们的初步数据显示了人类中的Septins 2、6、6、6、7、8、9、10、11的高基因表达 具有SEPTIN 7的肠道组织是最主要的同工型，在CD中进行了更新。药理学或 Septin细胞骨架的遗传破坏抑制了ECM产生的TGF-β1依赖性增加（胶原蛋白 I＆fibronectin）和永生和原发性肌细胞中的迁移。初步证据表明 这是转录后的调节。 SEPTIN调节改善了实验性鼠纤维化。因此，我们 提议研究septin细胞骨架重塑的假设是肠道的驱动力 纤维化和靶向Septin细胞骨架是一种新型的治疗纤维蛋白的方法 疾病。该假设将通过三个特定目的进行检验：AIM1。 Septin的改变的表征 IBD患者组织样品的表达和分布。这包括开发高分辨率 人类肠道组织和原代人肠肌纤维细胞中的Septin表达谱图， 包括生成第一个完整厚度单细胞RNA测序肠道图集以进行狭窄的CD和对照。 AIM2。研究促纤维化成肌纤维细胞的SEPTIN依赖调节的作用和机制 激活。我们将测试SEPTIN的破坏或过表达是否调节TGF-β1信号，细胞内水泡 使用功能丧失和获取的贩运或转录后的翻译和转录后的网络 功能方法。 AIM3。功能探索如果靶向septins会在体内改善肠纤维化。我们将 使用药理学和遗传方法调节septins在体内，并诱导两个实验性纤维化 不同的动物模型。我们将在预防之前暂时控制中央9月7日的删除和 在Col I阳性细胞中专门的实验性肠纤维化诱导后（逆转）。如果成功，这 提案将挑战纯粹由免疫驱动的ECM存款驾驶狭窄的范式 形成并提供概念概念，以防止或治疗相关的狭窄机制 CD患者的肠反应。