In vivo label free optical imaging of immune cells in human skin

人体皮肤免疫细胞体内无标记光学成像

• 批准号: 10664746
• 负责人: Mihaela Balu
• 金额: $ 17.27万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10664746
• 关键词: Address; Animal Experiments; Animal Model; Antibodies; Architecture; Area; Atopic Dermatitis; Autoimmune; Autoimmune Diseases; Autoimmune Process; Basal cell carcinoma; Binding Proteins; Biological Assay; Biological Markers; Blood Vessels; Cell Communication; Cells; Cellular Metabolic Process; Cellular Morphology; Chemicals; Clinical; Clinical Research; Collaborations; Cutaneous; Dermal; Detection; Devices; Disease; Disease remission; Early Diagnosis; Environment; Fluorescence; Germany; Goals; Heterogeneity; Histologic; Human; Image; Imaging Device; Immune; Immune response; Immune system; Immunologic Monitoring; Immunotherapy; Inflammatory; Investigation; Knowledge; Label; Lesion; Malignant Neoplasms; Measurement; Measures; Melanins; Metabolic; Methods; Microscopic; Modeling; Molecular; Morphology; Mus; NADH; Optics; Pathogenesis; Performance; Phase; Pigments; Play; Population; RNA Probes; Remission Induction Therapy; Reporter; Reporting; Resolution; Role; Scanning; Signal Transduction; Skin; Skin Cancer; Skin Carcinoma; Spatial Distribution; Specificity; Speed; Structure; Techniques; Technology; Three-Dimensional Image; Time; Tissues; Validation; Visualization; Work; cancer therapy; cell type; clinical imaging; combinatorial; density; design; detection sensitivity; fluorophore; human imaging; imaging approach; imaging platform; immune activation; immune imaging; immunoreaction; improved; in vivo; in vivo imaging; microscopic imaging; mouse model; multi-photon; multiphoton microscopy; optical imaging; response; skin disorder; spatiotemporal; success; tumor-immune system interactions

Summary. The study of mammalian immune cells and their interactions under native conditions is critical for understanding the mechanisms initiating many diseases, including cancer, and for improving their treatments. Intravital multiphoton microscopy (MPM), fluorescent reporter mouse models and in vivo cell and tissue labeling techniques have made possible the investigation of immune cells interactions at a cellular/subcellular level in their native environment. However, the significant differences in the skin and the immune system of mice and humans preclude studies of uniquely human immune responses. The goal of this proposal is to evaluate the ability of intravital MPM imaging to identify and distinguish immune cell populations in human skin, based on label-free molecular contrast. We will utilize a fast, large area multiphoton exoscope (FLAME), an imaging platform recently designed and developed by our group to have unique performance features optimized for clinical skin imaging. FLAME has the ability to generate in vivo 3D images of human skin over macroscopic areas (up to 8x8 mm2) with microscopic resolution (0.5-1µm) at fast acquisition rates (tens of seconds). FLAME features label-free specificity for melanin and NADH based on their spectral and rapid time-resolved endogenous fluorescence detection. Selective detection of melanin allows for imaging of pigment-rich cells such as melanophages with high specificity, while time-resolved NADH fluorescence detection reports on the protein binding activity of these molecules within the immune cells and on their metabolic heterogeneity. We have demonstrated the ability of FLAME to detect resident immune cells in normal human skin as well as an activation of a cellular immune response, based on endogenous molecular contrast. In this proposal, we will develop methods for the in vivo characterization of the spatial and metabolic heterogeneity of immune cells in human skin by using atopic dermatitis as an inflammatory skin condition model and Multi Omic Single-scan Assay with Integrated Combinatorial Analysis (MOSAICA) for validation. We will also evaluate the feasibility of FLAME to quantify changes in immune cell metabolism and spatiotemporal heterogeneity in response to immune activation. If successfully validated in a larger clinical study, this approach would be a critical first step in making it possible to characterize cellular-level immune responses in human skin at the bedside, with broad applications ranging from detecting early immune reactions to developing improved cancer treatments.

概括。 对哺乳动物免疫细胞及其在自然条件下相互作用的研究对于理解至关重要 引发包括癌症在内的许多疾病的机制，并改善其治疗方法。活体内 多光子显微镜 (MPM)、荧光报告小鼠模型以及体内细胞和组织标记 技术使得在细胞/亚细胞水平上研究免疫细胞相互作用成为可能 他们的原生环境。然而，小鼠和小鼠的皮肤和免疫系统存在显着差异。 人类排除了对人类独特免疫反应的研究。该提案的目标是评估 活体 MPM 成像能够识别和区分人体皮肤中的免疫细胞群，基于 无标记分子对比。我们将利用快速、大面积的多光子外窥镜 (FLAME)，一种成像技术 我们团队最近设计和开发的平台具有针对以下方面进行优化的独特性能特征 临床皮肤成像。 FLAME 能够在宏观区域生成人体皮肤的活体 3D 图像 （高达 8x8 mm2），在快速采集速率（数十秒）下具有显微分辨率 (0.5-1μm)。火焰特点 基于黑色素和 NADH 的光谱和快速时间分辨内源性，对黑色素和 NADH 具有无标记特异性 荧光检测。选择性检测黑色素可以对富含色素的细胞进行成像，例如 黑色素噬菌体具有高特异性，而时间分辨 NADH 荧光检测报告蛋白质 这些分子在免疫细胞内的结合活性及其代谢异质性。我们有 证明了 FLAME 能够检测正常人体皮肤中的常驻免疫细胞以及激活 基于内源性分子对比的细胞免疫反应。在本提案中，我们将开发 体内表征人体免疫细胞空间和代谢异质性的方法 通过使用特应性皮炎作为炎症性皮肤病模型和多组学单扫描测定 用于验证的综合组合分析 (MOSAICA)。我们还将评估 FLAME 的可行性 量化响应免疫激活的免疫细胞代谢和时空异质性的变化。 如果在更大规模的临床研究中成功得到验证，这种方法将是实现这一目标的关键的第一步 在床边表征人体皮肤细胞水平的免疫反应，具有广泛的应用范围 从检测早期免疫反应到开发改进的癌症治疗方法。