Epithelial cell complement production in the pathogenesis of chronic rhinosinusitis

慢性鼻窦炎发病机制中上皮细胞补体的产生

基本信息

批准号：
9886648
负责人：
Carl Atkinson
金额：
$ 42.6万
依托单位：
MEDICAL UNIVERSITY OF SOUTH CAROLINA
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-03 至 2020-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9886648
关键词：
Allergic Anatomy Carrier Proteins Cell Culture System Cell Line Cell physiology Cells Chronic Chronic Obstructive Airway Disease Clinical Research Clinical Trials Complement Complement 3a Complement Activation Complement Inactivators Development Disease Epithelial Epithelial Cells Epithelium Functional disorder Generations Goals Heart failure Human Immune Immune System Diseases In Vitro Infiltration Inflammation Inflammatory Intercellular Fluid Irritants Laboratories Location Lymph Mediating Modeling Mucositis Mucous Membrane Mus Nasal Polyps Nature Nose Outcome Study Pathogenesis Patients Play Production Proteins Quality of life Recording of previous events Regulation Reporter Reporting Role Serum Severity of illness Shapes Signal Transduction Sinus Sinusitis Source Stimulus Structure of respiratory epithelium Systemic Therapy T-Lymphocyte Tamoxifen Testing Therapeutic Therapeutic Agents Therapy Clinical Trials Tissues Tomatoes Topical application Translations adaptive immune response airway epithelium autocrine cell type chronic rhinosinusitis clinically relevant complement C3 precursor complement pathway complement system conditional knockout design early phase clinical trial extracellular innovation insight knockout animal mouse Cre recombinase mouse model novel novel strategies novel therapeutics pre-clinical receptor success tool tumor-immune system interactions

项目摘要

Sinonasal epithelial cells (SNEC) line the sinus cavity and play important roles in orchestrating innate and adaptive immune responses. We, and others, have demonstrated that SNEC from patients with chronic rhinosinusitis (CRSwNP), are endogenously pro-inflammatory, hyper-responsive to environmental stimuli, and promote immune cell infiltration and activation. Recently, multiple components of the complement (Cp) system have been shown to be activated and up regulated in the mucosa of CRSwNP patients, including C3, though how they contribute to CRS-related inflammation is unknown and thus the focus of these proposed studies. Traditionally, the observed effect of Cp factors and receptors on various cell types was thought to be mediated solely by Cp activation fragments generated in the serum, the lymph, or interstitial fluids. Yet, recent paradigm shifting studies have shown that Cp generated by the cell itself can function in an autocrine, and unexpectedly, an intracellular fashion, and that intracellular signaling is essential for regulating the cells functions. We have shown that SNEC cells are the primary sinonasal cavity producers of C3, the central protein in the Cp pathway, and that SNECs from CRSwNP have dysregulated intracellular stores of C3/C3a that can be rapidly mobilized upon stimulation with exogenous irritates. In these proposed studies we will test the hypothesis that dysregulated intracellular Cp signaling in SNEC plays a role in the epithelial cell dysfunction seen in CRSwNP. We will test our hypothesis by executing two inter-related but independent specific aims. In Aim 1 we will determine the role of local and intracellular Cp production in the regulation of human sinonasal epithelial cell inflammation. Sub-Aim 1.1 we will to determine the role of intracellular versus extracellular C3 signaling on SNEC functions. Sub-Aim 1.2 will test the hypothesis that inhibition of Cp-mediated signaling will reduce the pro-inflammatory and hyper-responsive nature of CRSwNP SNEC such that they will function similarly to control subject-derived SNEC. In Aim 2, determine the role local C3 production in the regulation of inflammation and disease severity in a murine model of atopic CRS. Sub-Aim 2.1 will determine the role of locally produced C3 in the sinonasal cavity using our C3-TdT tomato reporter mouse (C3-TdT), and by using a murine model of allergic fungal CRS (Af-CRS) test the key cell types that generate Cp at baseline and under disease conditions. To mimic the human Cp inhibitory therapies utilized in Aim 1, systemic and local C3 will be inhibited with Crry-Ig, and intracellular C3 will be inhibited through the use of intracellular delivery of Crry-Ig with a novel cell penetrating carrier protein (Feldan Shuttle). Using these novel approaches, we will dissect the impact of epithelial and systemic Cp activity on disease development (sub-Aim 2.2), and the therapeutic potential of inhibiting C3 to reverse established disease (sub-Aim 2.3) using the Af-CRS model. Together these studies will provide mechanistic insights into how elevated epithelial intracellular and locally produced Cp can shape the local immune microenvironment.

Sinonasal上皮细胞（SNEC）在鼻窦腔中排成一列，并在策划先天性方面发挥重要作用和自适应免疫反应。我们和其他人已经证明了来自慢性患者的SNEC 鼻孔炎（CRSWNP）具有内源性促炎，对环境刺激的反应性，并且促进免疫细胞浸润和激活。最近，补体系统的多个组件（CP）系统已显示在包括C3在内的CRSWNP患者的粘膜中被激活并受到调节它们如何促进与CRS相关的炎症是未知的，因此是这些提出的研究的重点。传统上，观察到的CP因子和受体对各种细胞类型的影响是介导的仅通过在血清，淋巴或间质液中产生的CP激活片段。然而，最近的范式转变研究表明，细胞本身生成的CP可以在自分泌中起作用，并且出乎意料地在细胞内的方式，而细胞内信号传导对于调节细胞功能至关重要。我们有表明SNEC细胞是C3的主要鼻腔腔腔生产剂，C3途径中的中心蛋白， CRSWNP的SNEC的C3/C3A细胞内存储失调，可以快速动员外源性刺激后，会发出刺激。在这些拟议的研究中，我们将检验以下假设 SNEC中的细胞内CP信号失调在CRSWNP中看到的上皮细胞功能障碍中起作用。我们将通过执行两个相互关联但独立的特定目的来检验我们的假设。在目标1中，我们将确定局部和细胞内CP产生在人鼻纳纳斯上皮细胞调节中的作用炎。 Sub-aim 1.1我们将确定细胞内和细胞外C3信号在 SNEC功能。 Sub-aim 1.2将检验以下假设：抑制CP介导的信号传导将减少 CRSWNP SNEC的促炎和超响应性质，使它们的功能与对照相似主题衍生的SNEC。在AIM 2中，确定局部C3产生在调节炎症和疾病的严重程度在特应crs的鼠模型中。 Sub-aim 2.1将确定本地生产的C3在使用我们的C3-TDT番茄记者小鼠（C3-TDT）和使用过敏性的鼠模型真菌CRS（AF-CRS）测试在基线和疾病条件下生成CP的关键细胞类型。模仿 AIM 1中使用的人类CP抑制疗法，全身和局部C3将被crry-ig抑制，并且细胞内C3将通过使用新颖的细胞穿透的细胞内递送来抑制细胞内C3 载体蛋白（Feldan Shuttle）。使用这些新颖的方法，我们将剖析上皮和全身性CP对疾病发育的活性（Sub-AIM 2.2），以及抑制C3的治疗潜力使用AF-CRS模型反向建立的疾病（Sub-Aim 2.3）。这些研究将共同提供关于上皮细胞内和局部生产的CP如何塑造局部的机械洞察力免疫微环境。