Conjunctival Goblet Cell Mucin Secretion in Inflammation and Its Resolution

炎症中结膜杯状细胞粘蛋白的分泌及其解决

基本信息

批准号：
9920424
负责人：
Darlene A Dartt
金额：
$ 5.97万
依托单位：
SCHEPENS EYE RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-30 至 2019-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9920424
关键词：
1-Phosphatidylinositol 3-Kinase ADRBK1 gene Acute Adenoviruses Allergens Allergic Allergic Conjunctivitis American Anti-inflammatory Antihistamines Aspirin Cell Proliferation Cell secretion Cells Chinese Hamster Ovary Cell Chronic Clinical Clinical Trials Conjunctival Epithelium Cornea Cyclic AMP-Dependent Protein Kinases Disease Dominant-Negative Mutation Epithelial Eye GPR32 gene Generations Goals Goblet Cells Grant HRH2 gene Histamine Histamine H1 Receptors Histamine H2 Receptors Histamine Receptor Human Hypersensitivity Individual Infiltration Inflammation Inflammation Mediators Inflammatory Inflammatory Response Laboratories Leukotriene Receptor Leukotrienes Leukotrienes A Lipids Lung MAPK3 gene MUC5AC gene Mast Cell Stabilizer Measurement Measures Mediator of activation protein Molecular Mucins Mucous body substance Mus Nose Ovalbumin Pathway interactions Penetration Peptides Pharmacology Phospholipase C Phospholipase D Production Prostaglandin D2 Protein Isoforms Protein Kinase Protein Kinase C Proto-Oncogene Proteins c-akt Protocols documentation Pruritus Receptor Signaling Redness Regulation Research Resolution Role Series Signal Pathway Signal Transduction Skin Small Interfering RNA Source Stains Swelling Symptoms Work conjunctiva cytokine delta opioid receptor effective therapy eye dryness first responder improved in vivo inhibitor/antagonist lipid mediator mast cell mouse model novel ocular surface prevent receptor receptor function recruit reduce symptoms response symptomatic improvement

项目摘要

Project Summary Although 15-20% of Americans suffer from allergic conjunctivitis, current treatments are limited to anti- histamines and mast cell stabilizers that are often ineffective. The long-term goal of the proposal is to develop a new treatment for allergic conjunctivitis that could also be used for allergies of the lung, skin, and nose along with other inflammatory diseases of the ocular surface such as dry eye. The new treatment is an anti- inflammatory and pro-resolution lipid mediator, the D-series resolvins compared to E-series resolvins currently in a clinical trial. Conjunctival goblet cell oversecretion of mucous including the mucin MUC5AC is one of the symptoms of allergic conjunctivitis that include itching, conjunctival redness, tearing, and chemosis. D-, but not E-, series resolvins have two modes of action. They block histamine and leukotriene over stimulation of goblet cell secretion and themselves activate a small amount of secretion to protect the ocular surface while the over secretion is being reset. E-series do not themselves increase secretion. The current proposal will focus on determining how pro-resolution mediators act at a molecular level to counter-regulate proinflammatory mediator stimulation of goblet cell mucin secretion both in culture and in vivo in a mouse model of allergic conjunctivitis. Research will focus on the following aims: 1. Determine the cellular mechanisms the D- and E- series resolvins use to stimulate goblet cell mucin secretion and if goblet cells are the source of the resolvins and other lipid mediators; 2. Unravel the molecular mechanisms D- and E-series resolvins use to activate protein kinases to interact with histamine and leukotriene receptors to counter-regulate their action; and 3. Ascertain the efficacy of D- compared to E-series resolvins to terminate goblet cell secretion and improve symptoms in a murine model of allergic conjunctivitis. For aim 1 RvD1, aspirin triggered (AT)-RvD1, and RvE1 will be used to stimulate human conjunctival goblet cells in culture. The receptors and cellular signaling pathways activated by the resolvins including phospholipase C and D and PI-3K will be investigated using siRNA, pharmacological inhibitors, and adenovirus constructs. In addition the production of pro-inflammatory and pro-resolution mediators will be determined by lipidomic measurements on goblet cells in culture. For aim 2 CHO cells transfected with a pro-inflammatory histamine or leukotriene receptor and a pro-resolution receptor and cultured human goblet cells will be used. Protein kinases predicted to phosphorylate the pro- inflammatory mediator receptors will be studied to determine if RvD1, AT-RvD1, and RvE1 activate the protein kinases and counter-regulate the histamine or leukotriene receptor to block its activity and terminate mucin secretion. siRNA, pharmacological inhibitors, and adenovirus constructs will be used for this aim. For aim 3 a mouse model of severe allergic conjunctivitis will be used with ovalbumin as the allergen. Resolvins compared to anti-histamines will be used to determine if they decrease MUC5AC secretion and goblet cell proliferation, ameliorate the clinical symptoms of allergic conjunctivitis, and terminate infiltration of inflammatory cells.

项目摘要尽管有15-20％的美国人患有过敏性结膜炎，但目前的治疗仅限于抗通常无效的组胺和肥大细胞稳定剂。该提案的长期目标是发展一种针对过敏性结膜炎的新疗法，也可以用于沿肺，皮肤和鼻子的过敏与眼表面的其他炎症性疾病（如干眼）。新疗法是一种抗与当前的E系列Resolvins相比在临床试验中。连接的杯状细胞细胞对包括粘蛋白MUC5AC在内的粘液过度分泌是其中之一过敏性结膜炎的症状，包括瘙痒，结膜发红，撕裂和化学。 D-，但不是 E-，系列分辨率具有两种作用模式。它们阻止组胺和白三烯刺激杯状细胞分泌，自身激活了少量分泌以保护眼表时分泌正在重置。电子系列本身不会增加分泌物。当前的提议将重点放在确定促分辨率介体如何在分子水平上作用以对抗促炎性在培养物和体内刺激杯状细胞粘蛋白分泌的小鼠过敏模型结膜炎。研究将侧重于以下目的：1。确定D-和E-的细胞机制系列溶胶素用于刺激杯状细胞粘蛋白分泌，如果杯状细胞是溶质的来源和其他脂质介体； 2。阐明分子机制D-和电子系列溶质用于激活蛋白激酶与组胺和白三烯受体相互作用以对抗其作用；和3。与电子系列溶质终止杯状细胞分泌相比，确定D-的功效在过敏性结膜炎的鼠模型中的症状。对于AIM 1 RVD1，阿司匹林触发（AT）-RVD1和RVE1 将用于刺激培养物中的人类结膜杯细胞。受体和细胞信号传导将使用包括磷脂酶C和D和PI-3K在内的溶质激活的途径。 siRNA，药理抑制剂和腺病毒构建体。此外，促炎的产生促介质将由培养物中的杯状细胞上的脂质细胞测量结果确定。目的 2个CHO细胞用促炎性组胺或白三烯受体转染和促分辨率将使用受体和培养的人杯细胞。蛋白激酶预测磷酸化将研究炎症介质受体，以确定RVD1，AT-RVD1和RVE1是否激活蛋白质激酶并对组胺或白三烯受体进行反调节以阻断其活性并终止粘蛋白分泌。 siRNA，药理抑制剂和腺病毒构建体将用于此目的。对于目标3 a 严重的过敏性结膜炎的小鼠模型将与卵蛋白一起用作过敏原。比较分解将抗抗胺用于确定它们是否减少MUC5AC分泌和杯状细胞增殖，缓解过敏性结膜炎的临床症状，并终止炎症细胞的浸润。