Characterization of a dendritic cell specific receptor critical for SJS

对 SJS 至关重要的树突状细胞特异性受体的表征

• 批准号: 9982185
• 负责人: Xinzhong Dong
• 金额: $ 40.94万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-16 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9982185
• 关键词: Affect; Alleles; Allopurinol; Animal Model; Antigen Presentation; Antigen-Presenting Cells; Antigens; Binding; Biological Assay; Bulla; Calcium; Cell Death; Cell Surface Receptors; Cell physiology; Cell surface; Cells; Characteristics; Coculture Techniques; Complex; Cornea; Cytotoxic T-Lymphocytes; Data; Dendritic Cells; Development; Dose; Drug Exposure; Epidermis; Excision; Exposure to; Eye; Eyelid structure; Family; Flow Cytometry; G-Protein-Coupled Receptors; Genetic; Goals; Hand; Hemorrhage; Homologous Gene; Hospitalization; Human; Image; Immune; Immune response; Immune system; Immunohistochemistry; Inflammation Mediators; Inflammatory; Knock-out; Knockout Mice; Knowledge; Lead; Lesion; Life; Major Histocompatibility Complex; Mediating; Mediator of activation protein; Molecular; Molecular Target; Mucous Membrane; Mus; Mutation; Necrosis; Open Reading Frames; Oral; Oral Ingestion; Pathway interactions; Patients; Pharmaceutical Preparations; Pharmacotherapy; Phenotype; Play; Process; Reaction; Recovery; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Site; Spleen; Stevens-Johnson Syndrome; Symptoms; Syndrome; T-Cell Activation; T-Cell Proliferation; T-Lymphocyte; Tissues; adaptive immune response; adverse drug reaction; cellular targeting; conjunctiva; cytokine; cytotoxic; experimental study; foot; genetic approach; in vivo; keratinocyte; lamotrigine; lymph nodes; medication safety; member; mouse model; mutant; novel; novel therapeutics; prevent; receptor; receptor expression; response

Project Summary The goal of our research is to understand the cellular and molecular mechanisms behind the development of Stevens-Johnson syndrome. Adverse drug reactions (ADRs) are a serious drug safety concern that can be hard to predict and treat. Stevens-Johnson syndrome (SJS) is one of the most serious and life- threatening ADRs characterized by keratinocyte cell death and mucosal breakdown; patients often form lesions on their palms of hands and soles of their feet and blistering and necrosis of their eyelids, conjunctiva and cornea. The exact cellular and molecular mechanisms behind drugs triggering SJS is unknown. Using a Ca2+ imaging assay, we found certain members of the Mrgpr GPCR family are activated by multiple known SJS- causing drugs. To examine the immune effects of these SJS drugs, we developed a novel mouse treatment where daily oral ingestion of a SJS-causing drug resulted in WT mice forming mucosal secretion in their eyes and blister bleeding in their hindpaws, similar to symptoms seen in patients suffering from SJS. We generated a mouse line where the specific Mrgpr's open reading frame was deleted and replaced with GFP and this genetic removal prevented formation of the SJS-like phenotype. Strikingly, heterozygous Mrgpr +/GFP mice, mice retaining 1 functional copy of the certain Mrgpr allele, developed the SJS-like phenotype, implying an important role for this Mrgpr in drug-caused SJS. Further experiments revealed the Mrgpr was expressed in a subset of dendritic cells, cells known to play an important role in antigen-presentation and immune response initiation. In this proposal, we will use molecular, cellular, genetic approaches and a potential novel in vivo animal model to uncover the role of the Mrgpr-expressing dendritic cells in drug-activated SJS. Aim I will focus on establishing our novel drug-induced SJS mouse model. We will examine what immune changes occur as mice are dosed with specific SJS-causing drugs and then we will determine what role the specific Mrgprs play in these immune responses. In Aim II, we will examine the specific characteristics of Mrgpr-expressing dendritic cells. We will find what type of dendritic cell expresses the Mrgprs and the role they play in the immune system. We will then examine how the cells change upon exposure to SJS-causing drugs and how these changes cause a cytotoxic immune response and SJS phenotype. We will also confirm the pivotal role these Mrgpr-expressing cells play in the development of SJS by rescuing the phenotype in the Mrgpr KO mouse. In Aim III, we will continue examining the immune cascade triggered by the SJS drugs. Dendritic cells are known for their important role of presenting antigens to T cells, which activate and in turn cause an immune response. We will determine what T cells are being activated by the Mrgpr-expressing dendritic cells and what cellular effects and mediators are released by these activated T cells. We will establish the necessity of T cells in continuing the immune response that is initiated and maintained by activation of the Mrgpr-expressing dendritic cell. Data collected from this project will help reveal some of the cellular and molecular reasons behind certain drugs causing SJS; these new cellular and molecular targets could lead to new treatments and drug therapies for SJS and adverse drug reactions.

项目摘要 我们这项研究的目的是要了解这背后的细胞和分子机制 史蒂文斯-约翰逊综合征的发展。药品不良反应(ADRs)是一个严重的药品安全问题 这可能很难预测和治疗。史蒂文斯-约翰逊综合征(Stevens-Johnson综合征，SJS)是世界上最严重、最致命的疾病之一。 以角质形成细胞死亡和粘膜破坏为特征的具有威胁性的不良反应；患者经常形成病变 手掌和脚底上，眼皮、结膜和眼皮起水泡和坏死 眼角膜。药物引发SJS背后的确切细胞和分子机制尚不清楚。使用钙离子 成像分析，我们发现某些成员的mrgpr gpr家族被多个已知的SJS激活- 导致吸毒。为了检验这些SJS药物的免疫效果，我们开发了一种新的小鼠治疗方法 每天口服导致SJS的药物导致WT小鼠眼睛产生粘膜分泌物 和后爪起泡出血，类似于SJS患者的症状。我们生成了 一个小鼠品系，其中特定的Mr gpr的开放阅读框被删除，并被GFP和这个取代 基因去除阻止了SJS样表型的形成。引人注目的是，杂合的mrgpr+/gfp小鼠， 保留1个功能拷贝的特定mrgpr等位基因的小鼠，出现了SJS样表型，这意味着 该基因在药物引起的SJS中的重要作用。进一步的实验表明，mrgpr在 树突状细胞亚群，已知在抗原提呈和免疫反应中发挥重要作用的细胞 入会仪式。在这项提案中，我们将使用分子、细胞、遗传方法和一种潜在的体内新技术 揭示表达mrgpr的树突状细胞在药物激活的SJS中的作用的动物模型。目标我会集中精力 关于建立我们的新型药物诱导的SJS小鼠模型。我们将检查在以下情况下发生的免疫变化 给小鼠灌服特定的引起SJS的药物，然后我们将确定特定的MRGPR起什么作用 在这些免疫反应中。在AIM II中，我们将研究mrgpr表达的特殊特征。 树突状细胞。我们将发现哪些类型的树突状细胞表达MRGPR，以及它们在 免疫系统。然后我们将研究细胞在接触引起SJS的药物时如何变化，以及如何 这些变化导致细胞毒性免疫反应和SJS表型。我们还将确认关键作用 这些表达mrgpr的细胞通过挽救mrgpr KO的表型在SJS的发展中发挥作用 老鼠。在AIM III中，我们将继续研究SJS药物引发的免疫级联反应。树突状细胞 它们以向T细胞递送抗原的重要作用而闻名，T细胞激活并继而引起免疫 回应。我们将确定哪些T细胞被表达mrgpr的树突状细胞激活，以及什么 这些激活的T细胞释放细胞效应和介质。我们将确定T细胞的必要性 在继续通过激活mrgpr表达而启动和维持的免疫反应中 树突状细胞。从这个项目收集的数据将有助于揭示一些细胞和分子原因 在某些导致SJS的药物背后；这些新的细胞和分子靶点可能导致新的治疗方法和 SJS的药物治疗和药物不良反应。