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

• 批准号: 9195092
• 负责人: Darlene A Dartt
• 金额: $ 52.82万
• 依托单位: SCHEPENS EYE RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2018-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9195092
• 关键词: 1-Phosphatidylinositol 3-Kinase; ADRBK1 gene; Acute; Adenoviruses; Adrenergic Receptor; Allergens; Allergic; Allergic Conjunctivitis; American; Anti-Inflammatory Agents; 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; 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; Recruitment Activity; Redness; Regulation; Research; Resolution; Role; STK6 gene; Series; Signal Pathway; Signal Transduction; Skin; Small Interfering RNA; Source; Staining method; Stains; Swelling; Symptoms; Work; conjunctiva; cytokine; effective therapy; emergency service responder; eye dryness; improved; in vivo; inhibitor/antagonist; lipid mediator; mast cell; mouse model; novel; ocular surface; prevent; public health relevance; receptor; receptor function; reduce symptoms; response; symptomatic improvement

DESCRIPTION (provided by applicant): Although 15-20% of Americans suffer from allergic conjunctivitis, current treatments are limited to anti-histamines and mast cell stabilizers that ar 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 over-secretion of mucous includin 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 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 infiltraton of inflammatory cells.

描述（由申请人提供）：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激素和肥大细胞稳定剂，而AR通常无效。该提案的长期目标是为过敏性结膜炎开发新的治疗方法，该治疗方法也可用于肺，皮肤和鼻子的过敏，以及其他眼部表面的炎症性疾病，例如干眼。与当前在临床试验中的电子系列溶质相比，新的治疗方法是一种抗炎和促进脂质介质的D系列溶质。粘液包括粘蛋白MUC5AC的结膜杯状细胞过度分泌是过敏性结膜炎的症状之一，其中包括瘙痒，结膜发红，撕裂和化学症。 D-但不是E-系列分辨率具有两种作用模式。它们阻止了杯状细胞分泌的组胺和白三烯过度刺激，并自身激活了少量分泌以保护眼表，而在重置过度分泌的同时。电子系列不会增加分泌物。当前的提案将重点侧重于确定促介质如何在分子水平上作用，以应对培养物和体内的润滑细胞粘蛋白分泌的促炎性介体刺激在过敏性结膜炎的小鼠模型中。研究将侧重于以下目的：1。确定D-和E系列Resolvins用来刺激杯状细胞粘蛋白分泌的细胞机制，以及如果杯状细胞是Resolvins和其他脂质介质的来源； 2。解开 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，以抵抗其作用。和3。与E系列分辨蛋白相比，确定D-的功效终止了杯状细胞分泌并改善了过敏性结膜炎模型中的症状。对于AIM 1 RVD1，阿司匹林触发（AT）-RVD1，RVE1将用于刺激培养物中的人类结膜杯细胞。通过siRNA，药理抑制剂和腺病毒构建体，将研究由磷脂酶C和D和PI-3K（包括磷脂酶C和D和PI-3K）激活的受体和细胞信号传导途径。另外，促炎和促促介质的产生将由培养物中的杯状细胞上的脂质组学测量结果确定。对于AIM 2，将使用促炎性组胺或白三烯受体转染的CHO细胞以及促促分辨率受体和培养的人杯细胞。将研究将研究促炎性介质受体的蛋白激酶，以确定RVD1，AT-RVD1和RVE1是否激活蛋白激酶，并对抗组胺或白细胞受体调节其活性并终止粘液蛋白分泌。 siRNA，药理抑制剂和腺病毒构建体将用于此目的。对于AIM 3，严重过敏性结膜炎的小鼠模型将与卵蛋白一起用作过敏原。与抗抗药酶相比，将分辨率用于确定它们是否减少MUC5AC分泌和杯状细胞增殖，改善过敏性结膜炎的临床症状，并终止炎性细胞的浸润。