Comprehensive Quantitative Profiling of Cellular Alterations Caused by Injury

损伤引起的细胞改变的全面定量分析

• 批准号: 10392403
• 负责人: PRAMOD K DASH
• 金额: $ 63.73万
• 依托单位: UNIVERSITY OF HOUSTON
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10392403
• 关键词: Adoption; Anatomy; Animals; Antibodies; Apoptotic; Astrocytes; Atlases; Biological Assay; Biological Models; Biology; Blood Vessels; Brain; Brain Concussion; Brain Diseases; Brain Injuries; Brain Pathology; Brain region; Cells; Characteristics; Clinical Trials; Communities; Complex; Computer software; Computers; Cortical Cell Layer; Data; Dementia; Development; Disease; Distant; Drug Combinations; Drug Side Effects; Drug usage; Endothelial Cells; Engineering; Failure; Goals; Image; Image Analysis; Individual; Inflammation; Injury; Intelligence; Internet; Investigation; Laboratories; Lighting; Measurement; Methods; Microglia; Modification; Molecular; Morphology; Neurons; Oligodendroglia; Outcome; Pathologic; Pathology; Pharmacotherapy; Photobleaching; Process; Proliferating; Proteins; Protocols documentation; Rattus; Reproducibility; Research; Research Personnel; Research Project Grants; Resolution; Rest; Slice; Software Engineering; Speed; Stains; System; Testing; Therapeutic Intervention; Tissues; Treatment Protocols; Validation; Vascular Endothelial Cell; Visual; Work; base; brain cell; cell type; cost; data acquisition; data mining; design; digital; effective therapy; effectiveness evaluation; excitatory neuron; flexibility; fluorescence microscope; fluorophore; functional status; high resolution imaging; improved outcome; inhibitory neuron; mild traumatic brain injury; next generation; novel drug combination; novel therapeutics; open source; regenerative; side effect; software systems; stem cells; tool; treatment effect; whole slide imaging

ABSTRACT Currently, cellular alterations associated with pathological conditions are studied using low complexity immunohistochemical (IHC) assays, typically utilizing 2-5 antibodies, that only reveal a tiny subset of the alterations that are occurring, lack comprehensive cellular context, and do not provide quantitative readouts of cellular changes throughout the tissue. For example, a injury or disease can initiate a complex web of pathological alterations across cell types, and at multiple scales ranging from individual cells to multi-cellular units and the layered brain cytoarchitecture. However, technological limitations are hindering a more comprehensive global understanding of these pathological changes. This lack of understanding is hampering our ability to intelligently design effective treatment regimens, and may have contributed to the failures of clinical trials that targeted a single cell type or specific protein. To bridge this gap in our understanding, we propose to develop a Comprehensive Brain Cellular Alteration Profiling Toolkit (CBAT), a carefully validated and broadly applicable image analysis toolkit with unprecedented potential to accelerate investigation & development of next-generation treatments for brain diseases. CBAT, in association with a flexible and modular protocol for highly multiplexed IHC, will enable simultaneous profiling of all major brain cell types and their functional/pathological status (e.g., resting, reactive, apoptotic) across whole brain sections. It will provide quantitative readouts of cellular alterations at multiple scales ranging from individual cells of all types to multi- cellular units (e.g. niches), brain cell layers, and brain regions. Comprehensive cellular profiling and measurements generated using CBAT will enable a deeper understanding of pathological cellular changes that will enable accelerated design, testing, and optimization of therapeutic interventions. Further, it will reduce overall experimental costs by replacing a large number of less-informative assays with a single comprehensive assay. In the longer term, it will enhance our ability to conduct the systems-level investigations that will be required for fully understanding, and successfully treating, multiple brain pathologies. To achieve these goals, we propose the following aims: Aim 1: Develop and validate a flexible, scalable, extensible, and reproducible method for comprehensive whole slide imaging of all the major brain cell types in stereotactically aligned rat whole brain sections; Aim 2: Develop and validate a turnkey software system profiling cell identify and status at multiple scales ranging from individual cells to multi-cellular units, brain cell layers, and brain anatomic regions; and Aim 3: Test the utility of the CBAT system to comprehensively profile concussion biology, and assess the effectiveness of a drug combination to reduce newly identified pathologies. After its development and validation, CBAT will be disseminated to the research community at no cost for use in their specific research projects.

抽象的 目前，与病理状况相关的细胞改变是使用低复杂度来研究的 免疫组织化学 (IHC) 检测通常使用 2-5 种抗体，仅揭示一小部分抗体 正在发生的改变，缺乏全面的细胞背景，并且不提供定量读数 整个组织的细胞变化。例如，受伤或疾病可能会引发一个复杂的网络 跨细胞类型以及从单个细胞到多细胞的多个尺度的病理改变 单位和分层脑细胞结构。然而，技术限制阻碍了更多 对这些病理变化的全面的全球了解。这种缺乏理解正在阻碍 我们有能力明智地设计有效的治疗方案，并且可能导致了失败 针对单一细胞类型或特定蛋白质的临床试验。为了弥合我们理解上的差距，我们 建议开发一个综合脑细胞改变分析工具包（CBAT），一个仔细的 经过验证且广泛适用的图像分析工具包，具有加速调查速度的前所未有的潜力 和开发下一代脑疾病治疗方法。 CBAT 与灵活且 用于高度多重 IHC 的模块化协议将能够同时分析所有主要脑细胞类型和 他们整个大脑切片的功能/病理状态（例如，休息、反应、凋亡）。它将提供 多种尺度的细胞变化的定量读数，范围从所有类型的单个细胞到多细胞 细胞单位（例如壁龛）、脑细胞层和大脑区域。全面的细胞分析和 使用 CBAT 生成的测量结果将有助于更深入地了解病理性细胞变化 将能够加速治疗干预措施的设计、测试和优化。此外，还将减少 通过用单一的综合分析代替大量信息较少的分析来降低总体实验成本 化验。从长远来看，它将增强我们进行系统级调查的能力 充分理解并成功治疗多种脑部疾病所必需的。为了实现这些目标， 我们提出以下目标： 目标 1：开发和验证灵活的、可扩展的、可扩展的和可重复的 立体定向排列大鼠所有主要脑细胞类型的全面全玻片成像方法 全脑切片；目标 2：开发并验证交钥匙软件系统分析单元识别和状态 从单个细胞到多细胞单位、脑​​细胞层和大脑解剖学的多个尺度 地区；目标 3：测试 CBAT 系统全面分析脑震荡生物学的实用性，以及 评估药物组合减少新发现的病理的有效性。其发展后 和验证后，CBAT 将免费传播给研究界，用于其特定领域 研究项目。