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.

总结。 研究哺乳动物免疫细胞及其在自然条件下的相互作用对于理解 启动包括癌症在内的许多疾病的机制，并改进其治疗方法。活体内 多光子显微镜(MPM)、荧光报告小鼠模型和体内细胞和组织标记 技术已经使在细胞/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，小鼠皮肤和免疫系统的显著差异 人类排除了对人类独特免疫反应的研究。这项提案的目标是评估 活体MPM成像识别和区分人类皮肤免疫细胞群的能力，基于 无标记分子对比度。我们将利用快速、大面积的多光子外窥镜(FLAME)，一种成像 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功能 临床皮肤成像。火焰能够在宏观区域生成人体皮肤的活体3D图像 (高达毫米)，显微分辨率(0.5-1微米)，快速采集速率(几十秒)。火焰特征 基于黑色素和NADH光谱和快速时间分辨内源性的无标记特异性 荧光检测。黑色素的选择性检测允许对富含色素的细胞进行成像，例如 高特异性的黑素噬菌体，而时间分辨的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的异质性。我们有 证明了火焰能够检测到正常人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。在这项提案中，我们将发展 人体免疫细胞空间和代谢异质性的体内表征方法 特应性皮炎作为炎症性皮肤状况模型的皮肤和多组单次扫描分析 用于验证的综合组合分析(Mosaica)。我们还将评估火焰发电的可行性 量化免疫细胞新陈代谢的变化和免疫激活的时空异质性。 如果在更大的临床研究中成功验证，这种方法将是使其成为可能的关键的第一步。 表征床边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。