Imaging immune cell type and behavior in the living retina using adaptive optics

使用自适应光学对活体视网膜中的免疫细胞类型和行为进行成像

• 批准号: 10521626
• 负责人: Jesse Barrett Schallek
• 金额: $ 59.2万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-30 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10521626
• 关键词: Acute; Affect; Age related macular degeneration; Antigens; Arteries; Autoimmune; Autoimmune Diseases; Behavior; Behavior monitoring; Binding Proteins; Biological Markers; Biopsy; Blindness; Blood Cell Count; Blood Cells; Blood Vessels; Blood capillaries; Blood flow; CD4 Positive T Lymphocytes; Caliber; Cell Communication; Cell physiology; Cells; Collection; Contrast Media; Diabetic Retinopathy; Disease model; Edema; Endotoxins; Evolution; Eye; Flow Cytometry; Fluorescence; Foundations; Geographic Distribution; Glaucoma; Histology; Hour; Human; Image; Immune; Immune Cell Suppression; Immune response; Immune system; Individual; Infiltration; Inflammation; Inflammatory; Inflammatory Response; Injections; Injury; Label; Laboratories; Lead; Leukocytes; Light; Lipopolysaccharides; Location; Measures; Mediating; Microglia; Microscopic; Microscopy; Modality; Modeling; Monitor; Mus; Natural History; Nature; Ophthalmoscopes; Ophthalmoscopy; Optical Coherence Tomography; Optics; Pathogenicity; Perfusion; Pharmacologic Substance; Phase; Play; Population; Proxy; Reporter; Reporting; Research; Resolution; Retina; Retinal Diseases; Role; Scanning; Speed; Swelling; T-Lymphocyte; Techniques; Thick; Thinness; Time; Tissues; Uveitis; Vascular System; Veins; Viral; Vision; Work; adaptive optics; arteriole; autoimmune uveitis; blood flow measurement; cell behavior; cell type; cellular imaging; cellular targeting; clinical application; clinical translation; contrast imaging; high resolution imaging; imaging approach; imaging modality; imaging platform; immune imaging; in vivo; in vivo imaging; innovation; insight; interstitial retinol-binding protein; intravitreal injection; microscopic imaging; millisecond; neutrophil; non-invasive imaging; novel; novel therapeutics; optical imaging; perfusion imaging; response; sight restoration; time use; transcriptome sequencing; treatment response; venule; visual tracking

Abstract: Immune cell response to ocular inflammation is involved in various prevalent retinal diseases that lead to blindness such as glaucoma, age related macular degeneration, uveitis and diabetic retinopathy. However, the microscopic and translucent nature of circulating and resident immune cells has made study of such cells challenging in the living eye. Further complicating matters, immune cells move at fast speeds (centimeters/second) when circulating in big vessels and glacial pace at micrometers/minute when in tissue. This project will combine a number of innovations that overcome many of these challenges to study single immune cells in the living eye in their response to inflammation. The use of adaptive optics scanning light ophthalmoscope (AOSLO) to image mouse retina will provide a noninvasive in vivo imaging method that can resolve single immune cells and track their behavior through the entire course of inflammation. With our AOSLO, we have recently demonstrated that phase contrast combined with both fast and time-lapse videography now enables us to visualize these microscopic immune cells using near infrared light alone and without fluorescence contrast. Therefore, this project will seek to study immune cell dynamics in two models of inflammation in the living mouse eye and track the cellular responses to initiation, escalation and resolution. And while the approach is non-invasive and does not require use of fluorescence, we will combine fluorescence confirmation of specific leukocyte populations that contribute to the inflammatory response. Our project tackles three synergistic aims to track the dynamic nature of inflammation using two established models: Model 1) An endotoxin induced uveitis model using lipopolysaccharide (LPS) injection in vitreous of the eye. This results in an acute, short-term form of inflammation. We will track the initial escalation of inflammation in the retina by monitoring behavior of tissue resident microglia and systemic early responder neutrophils. And Model 2) An autoimmune uveitis condition that is induced in a healthy host mouse by injecting a subpopulation of fluorescent CD4+ T cells that are reactive to interphotoreceptor retinoid binding protein from a different donor mouse. Here, we will characterize the behavior of these foreign reactive T cells in healthy retina, and their interaction with the host immune system. In both inflammatory models, we will characterize the initiation, escalation, infiltration and resolution of immune cells in the retina by longitudinally tracking their behavior from hours to days to months. Finally, in a third aim, we will also study the changes in blood flow in response to these inflammatory models. For both endotoxin mediated and autoimmune inflammatory models described above, we will monitor structural and functional changes in retinal vasculature from the largest arteries and veins to single file flow capillaries to understand the interplay between blood flow and immune cell arrival into the retina. Already, we have observed that the vascular response is not homogeneous, and arterioles, venules and capillaries respond in different ways to the inflammatory insult and likely mediate the location, arrival and type of immune cell passage. Together, the longitudinal monitoring of inflammation and blood flow will provide new insights to the retinal immune response using innovations in high-resolution ophthalmoscopy.

摘要： 对眼部炎症的免疫细胞应答涉及导致眼部炎症的各种流行性视网膜疾病。 失明如青光眼、年龄相关性黄斑变性、葡萄膜炎和糖尿病视网膜病变。但 循环和驻留免疫细胞的显微镜和半透明性质使得对这些细胞的研究具有挑战性， 活着的眼睛更复杂的是，免疫细胞在体内循环时以快速（厘米/秒）移动， 组织中的大血管和微米/分钟的缓慢速度。该项目将结合联合收割机的一些创新， 克服了许多这些挑战，以研究活眼中的单个免疫细胞对炎症的反应。的 利用自适应光学扫描光检眼镜（AOSLO）对小鼠视网膜进行成像将提供一种无创的活体 这种成像方法可以分辨单个免疫细胞，并在整个炎症过程中跟踪它们的行为。 最近，我们的AOSLO证明，相位对比结合快速和延时成像， 现在，摄像技术使我们能够单独使用近红外光来观察这些微观免疫细胞， 荧光对比度因此，本项目将寻求在两种炎症模型中研究免疫细胞动力学， 活的老鼠眼睛，并跟踪细胞反应的启动，升级和解决。虽然这种方法 非侵入性，不需要使用荧光，我们将结合联合收割机荧光确认特定的白细胞 导致炎症反应的人群。我们的项目处理三个协同目标，以跟踪动态 模型1）内毒素诱导的葡萄膜炎模型， 眼玻璃体内注射脂多糖（LPS）。这会导致急性、短期的炎症。我们将 通过监测组织驻留的小胶质细胞和系统的行为来跟踪视网膜中炎症的初始升级， 早期反应中性粒细胞和模型2）在健康宿主小鼠中通过以下诱导的自身免疫性葡萄膜炎病症： 注射荧光CD 4 + T细胞亚群，所述荧光CD 4 + T细胞对来自细胞的感光器间类视色素结合蛋白具有反应性 不同的供体小鼠在这里，我们将描述这些外源性反应性T细胞在健康视网膜中的行为，以及它们在视网膜中的表达。 与宿主免疫系统的相互作用。在这两种炎症模型中，我们将描述炎症的起始，升级， 通过纵向跟踪免疫细胞在视网膜中的行为，从数小时到数天， 个月最后，在第三个目标中，我们还将研究响应于这些炎症模型的血流变化。为 在上述内毒素介导的和自身免疫性炎症模型中，我们将监测结构和功能 视网膜血管系统的变化，从最大的动脉和静脉到单列流动毛细血管，以了解相互作用 血液流动和免疫细胞到达视网膜之间的联系。我们已经观察到，血管反应不是 小动脉、小静脉和毛细血管以不同的方式对炎性损伤作出反应， 介导免疫细胞通过的位置、到达和类型。总之，炎症的纵向监测和 血流将使用高分辨率检眼镜检查中的创新提供对视网膜免疫反应的新见解。