Assessing how ocular surface nerves, immune cells, and epithelial cells communicate to encourage neuro-immune homeostasis

评估眼表神经、免疫细胞和上皮细胞如何沟通以促进神经免疫稳态

• 批准号: 10595234
• 负责人: Daniel H Kaplan
• 金额: $ 167.85万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-30 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10595234
• 关键词: ATAC-seq; Address; Affect; Afferent Neurons; Anatomy; Antibodies; Architecture; Axon; Back; Binding Proteins; Bioinformatics; Cells; Colon; Communication; Cornea; Corneal Diseases; Data; Data Set; Databases; Deposition; Dermatology; Designer Drugs; Development; Disease; Disease model; Dry Eye Syndromes; Environment; Epigenetic Process; Epithelial; Epithelial Cells; Esthesia; Eye; Flow Cytometry; Fostering; Future; Ganglia; Gastrointestinal tract structure; Gene Expression; Genetic; Goals; Graft Rejection; Grant; Hand; Health; Herpesvirus 1; Homeostasis; Human; Imaging Techniques; Immune; Immunohistochemistry; Immunology; Infiltration; Interdisciplinary Study; Intestines; Keratitis; Keratopathy; Knowledge; Label; Lead; Link; Location; Machine Learning; Mass Spectrum Analysis; Mediating; Membrane; Microbiology; Modeling; Molecular; Morphology; Mus; Nature; Nerve; Neuroglia; Neuroimmune; Neurology; Neurons; Neurotransmitters; Ophthalmology; Pain; Pathologic; Pathway interactions; Pattern; Play; Population; Process; Property; Proteins; Proteomics; Records; Regulation; Research; Role; Sensory; Shotguns; Skin; Stimulus; Structure; Structure of superior cervical ganglion; System; Technology; Tissues; Transgenic Mice; Vascular Endothelial Growth Factors; Viral; Wheat Germ Agglutinins; Work; afferent nerve; aqueous; base; cell type; chemokine; conjunctiva; cytokine; design; designer receptors exclusively activated by designer drugs; gain of function; immune checkpoint; in vivo; large datasets; member; nerve supply; neuroimmunology; neuron loss; neurotrophic factor; novel; ocular surface; prevent; programs; protein expression; proteomic signature; public database; receptor; recruit; relating to nervous system; response; single-cell RNA sequencing; small molecule; transcriptomics; web portal

PROJECT SUMMARY The nature and function of ocular surface nerves play a critical role in maintaining ocular surface health while preventing disease. Disruption of ocular surface sensory nerves can lead to blinding keratitis, graft rejection, dry eye disease, and ocular surface pain. The advent of “-omics” studies have provided a platform to understand how ocular surface nerves participate in the broader context of ocular health and disease. Here, we outline a proposal that will create large datasets detailing the nature of ocular surface innervation. We will take a systems-level approach towards analyzing our data using bioinformatics and machine learning platforms, so that we can gain a practical understanding of the ocular surface environment during health and disease. Specifically, this proposal will address how ocular surface nerves, epithelial cells, and immune cells interact. We have recruited a multidisciplinary research team that consists experts in ophthalmology, neuroimmunology, neurology, proteomics, systems immunology, and bioinformatics. With this team, we plan to comprehensively analyze all three levels of research outlined in the RFA, which includes anatomical and morphological characterization, defining cellular and molecular properties of neuronal and non-neuronal cell types, and assessing functional properties of neuronal and non-neuronal cells. Because innervation patterns and functionality changes with disease, we will use multiple ocular surface disease models (viral keratitis and aqueous dry eye disease) to better understand how neurons, epithelial cells, and immune cells affect neuronal functionality and subsequent disease. We will achieve our goal by pursuing four specific aims: 1) We will perform single-cell (sc)-omics on neuonrs and non-neuronal cells of the cornea and ocular surface innervating ganglia. These data will characterize neuronal identities and describe their functionality during disease. Additionally, we will be able to molecularly characterize immune and epithelial cells that influence the functionality of ocular surface afferents during disease. 2) We will use mass spectrometry to analyze the proteomic signatures of the cornea during health and disease. An unbiased shotgun approaches and a targeted approach focusing on neurotrophic factors will be used to identify proteins that influence ocular surface nerve functionality. 3) We will use our imaging techniques and histocytometry to comprehensively analyze the location and morphology of neuronal and non-neuronal cells within the cornea, so that we may be able to make conclusions about how nerves, epithelial, and immune cells interact in close proximity at the ocular surface. 4) We will use DREADD (designer receptor exclusively activated by designer drugs) technology to perturb neuronal sensitivity, so that we can assess how altering neuronal responsiveness to stimuli may affect the ocular surface homeostasis and progress of disease. A systems-level analysis of our observations using machine-learning and bioinformatics will lead to the prediction of novel interaction networks that can be interrogated in future studies. All data will be deposited into a web portal that we will develop.

项目概要 眼表神经的性质和功能对于维持眼表健康起着至关重要的作用，同时 预防疾病。眼表感觉神经的破坏可导致致盲性角膜炎、移植物排斥、 干眼症和眼表疼痛。 “组学”研究的出现提供了一个平台 了解眼表神经如何参与更广泛的眼部健康和疾病。这里， 我们概述了一项提案，该提案将创建详细说明眼表神经支配性质的大型数据集。我们将 采用生物信息学和机器学习的系统级方法来分析我们的数据 平台，使我们能够实际了解健康和生活期间的眼表环境 疾病。具体来说，该提案将解决眼表神经、上皮细胞和免疫细胞如何 相互影响。我们招募了一支由眼科专家组成的多学科研究团队， 神经免疫学、神经病学、蛋白质组学、系统免疫学和生物信息学。通过这个团队，我们计划 全面分析 RFA 中概述的所有三个层次的研究，其中包括解剖学和 形态学表征，定义神经元和非神经元细胞的细胞和分子特性 类型，并评估神经元和非神经元细胞的功能特性。因为神经支配模式 以及功能随疾病的变化，我们将使用多种眼表疾病模型（病毒性角膜炎和 水性干眼病），以更好地了解神经元、上皮细胞和免疫细胞如何影响神经元 功能和随后的疾病。我们将通过追求四个具体目标来实现我们的目标：1）我们将 对角膜和眼表神经支配的神经元和非神经元细胞进行单细胞 (sc) 组学 神经节。这些数据将表征神经元的特性并描述它们在疾病期间的功能。 此外，我们将能够对影响免疫和上皮细胞的分子特征进行描述。 疾病期间眼表传入的功能。 2) 我们将使用质谱分析 健康和疾病期间角膜的蛋白质组学特征。一支公正的霰弹枪接近了， 专注于神经营养因子的有针对性的方法将用于识别影响眼部的蛋白质 表面神经功能。 3）我们将利用我们的成像技术和组织细胞计数来全面 分析角膜内神经元和非神经元细胞的位置和形态，以便我们可以 能够得出关于神经、上皮细胞和免疫细胞如何在邻近区域相互作用的结论 眼表面。 4）我们将使用DREADD（专门由设计药物激活的设计受体）技术 扰乱神经元的敏感性，以便我们可以评估改变神经元对刺激的反应性可能会如何 影响眼表稳态和疾病进展。对我们观察结果的系统级分析 使用机器学习和生物信息学将导致新型交互网络的预测 在以后的学习中受到质疑。所有数据都将存入我们将开发的门户网站中。