Sorting and characterization of mechanically heterogeneous cell populations based on cellular contractility

基于细胞收缩性的机械异质细胞群的分类和表征

• 批准号: 10728070
• 负责人: Jian Zhang
• 金额: $ 22.03万
• 依托单位: UNIVERSITY OF ARKANSAS AT FAYETTEVILLE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10728070
• 关键词: Actomyosin; Adopted; Aging; Attention; Biological; Biological Models; Breast Cancer Cell; Cardiovascular Diseases; Cell Line; Cell Separation; Cells; Coupled; Cultured Cells; Defect; Development; Disease; Disease Outcome; Disease Progression; Engineering; Environment; Exhibits; Fibrosis; Fluorescence-Activated Cell Sorting; Foundations; Future; Genetic Heterogeneity; Geometry; Goals; Heterogeneity; In Vitro; Individual; Invaded; Knowledge; Label; Malignant Neoplasms; Mechanics; Memory; Mesenchymal Stem Cells; Metabolism; Metastatic breast cancer; Methods; Microscopy; Modeling; Molecular; Mus; Neoplasm Metastasis; Organ; Outcome; Output; Pathway interactions; Pattern; Phenotype; Play; Population; Postdoctoral Fellow; Primary Neoplasm; Process; Proliferating; Proteins; Reporting; Research; Role; Signal Transduction; Sorting; Source; Subgroup; Surface; System; Testing; Time; Tissues; Traction; Traction Force Microscopy; Work; cell motility; design; experimental study; extracellular; high throughput screening; human disease; in vivo; innovation; mechanical properties; mechanotransduction; migration; mouse model; novel; photoactivation; population based; screening; spatiotemporal; stem cell population; targeted treatment; therapeutic target; therapy resistant; tool; tumor; tumor progression

Project Summary Cellular contractility plays a critical role in both development and disease. Recent evidence suggests that even within a cell population derived from the same source, vast heterogeneity exists in terms of cellular contractility. Dysregulation of spatiotemporally organized cellular contractility often results in developmental defects. In invasive diseases like cancer which are often highly contractile, the existence of a weakly contractile subpopulation is receiving increasing attention. Adherent cells are known for their ability to sense and dynamically adapt to their local microenvironment. Hence, the heterogeneity in mechanical phenotype may be a result of genetic heterogeneity or cellular plasticity and mechanical adaptation. Mechanomedicines or mechano- based therapies that target specific physical cellular and tissue interactions, including abnormal cellular contractility, in diseases like cancer, fibrosis, and cardiovascular disease, as well as aging, are emerging and hold great potential. While mechanical heterogeneity and plasticity are known to contribute to resistance to therapies that target a specific molecular pathway, it is not clear whether a change in mechanical phenotype predicts disease outcome or if mechanical adaption happens as a result of disease progression. This project proposes to phenotypically sort adherent cells into subpopulations with distinct contractile phenotypes and use these sorted subpopulations to test the hypothesis that the initial contractile phenotype and heterogeneity determine the disease outcome against the alternative hypothesis that mechanical adaptation to the local microenvironment and phenotypical switching contribute to disease progression regardless of the initial mechanical heterogeneity. Cancer metastasis will be used as the main biological model for hypothesis testing. In Aim 1, the engineering approach for cell sorting based on cellular contractility will be optimized. Fluorescence- activated cell sorting (FACS) will be coupled with an engineered high throughput cell contractility screening platform, automated microscopy, and photoactivation and fluorescent labeling of cells for cell separation. In Aim 2, the sorted contractile subpopulations will be used to test the main biological hypothesis in vitro and in vivo against the alternative hypothesis. Engineered systems mimicking the environmental conditions in cancer progression will be designed to characterize the migration, proliferation, survival, and metabolism of the subpopulations, as well as their mechanical adaptation. The metastatic potential of these subpopulations and their mechanical adaptations at various stages along the metastatic cascade will be examined in a mouse tumor model. The innovative aspect of this proposal is the concept to sort by cellular contractility with the goal of uncovering the role of initial mechanical phenotype in the progression of diseases like cancer and in development. This project will use the novel engineered cell sorting approach to dissect the respective roles of mechanical heterogeneity and adaptability in disease progression, thus laying the foundation for future work to identify the key molecular pathways to precisely target for the development of mechanomedicine.

项目概要 细胞收缩力在发育和疾病中都起着至关重要的作用。最近的证据表明，即使 在同一来源的细胞群中，细胞收缩性存在巨大的异质性。 时空组织的细胞收缩力失调通常会导致发育缺陷。在 侵袭性疾病，如癌症，通常具有高度收缩性，存在弱收缩性 亚人群越来越受到关注。贴壁细胞以其感知和感知的能力而闻名。 动态适应当地的微环境。因此，机械表型的异质性可能是 遗传异质性或细胞可塑性和机械适应的结果。机械医学或机械医学 基于特定物理细胞和组织相互作用的疗法，包括异常细胞 收缩性在癌症、纤维化、心血管疾病以及衰老等疾病中的作用正在不断出现， 拥有巨大的潜力。虽然已知机械异质性和塑性有助于抵抗 针对特定分子途径的疗法，尚不清楚机械表型是否发生变化 预测疾病结果或机械适应是否因疾病进展而发生。这个项目 提出将贴壁细胞按表型分类为具有不同收缩表型的亚群并使用 这些排序的亚群来检验初始收缩表型和异质性的假设 根据机械适应局部的替代假设确定疾病结果 无论最初的情况如何，微环境和表型转换都会导致疾病进展 机械异质性。癌症转移将作为假设检验的主要生物学模型。 在目标 1 中，将优化基于细胞收缩性的细胞分选工程方法。荧光- 活化细胞分选（FACS）将与工程化高通量细胞收缩性筛选相结合 平台、自动显微镜、以及用于细胞分离的细胞光活化和荧光标记。瞄准 2、分选的收缩亚群将用于体外和体内检验主要生物学假设 反对备择假设。模仿癌症环境条件的工程系统 进展将被设计为表征迁移、增殖、存活和代谢 亚种群及其机械适应。这些亚群的转移潜力和 将在小鼠肿瘤中检查它们在转移级联的各个阶段的机械适应 模型。该提案的创新之处在于通过细胞收缩性进行排序的概念，其目标是 揭示初始机械表型在癌症等疾病进展和发育中的作用。 该项目将使用新颖的工程细胞分选方法来剖析机械细胞的各自作用 疾病进展的异质性和适应性，从而为未来确定疾病进展的工作奠定基础 精确定位机械医学发展的关键分子途径。