A dual-modality quantitative phase and polarized light microscope to assess cell motility and extracellular matrix remodeling during invasion

双模态定量相和偏光显微镜评估侵袭过程中的细胞运动和细胞外基质重塑

• 批准号: 9924599
• 负责人: Christopher B Raub
• 金额: $ 7.95万
• 依托单位: CATHOLIC UNIVERSITY OF AMERICA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-01 至 2022-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9924599
• 关键词: 3-Dimensional; Actins; Biological Assay; Biopsy; Birefringence; Breast Cancer cell line; Caliber; Calibration; Cancer Biology; Cancer Model; Cancer cell line; Cells; Cessation of life; Characteristics; Clinical; Collagen; Complex; Confocal Microscopy; Development; Disease; Dose-Limiting; Effectiveness; Environment; Extracellular Matrix; F-Actin; Fibrosis; Future; Goals; Health; Holography; Hour; Hydrogels; Image; Immunofluorescence Immunologic; In Vitro; Interstitial Collagenase; Invaded; Knowledge; Label; Laser Scanning Microscopy; Length; Light Microscope; Link; MDA MB 231; Malignant Neoplasms; Maps; Measurement; Microscope; Microscopy; Microspheres; Modality; Modeling; Neoplasm Metastasis; Neural Tube Development; Optics; Pathologic Processes; Phase; Phenotype; Phototoxicity; Polarization Microscopy; Polystyrenes; Process; Proteins; Publishing; Research; Resources; Sampling; Signal Transduction; Slide; Solid; Specimen; Stains; Stress; System; Testing; Time; Tissues; Toxic effect; Wound models; base; cancer cell; cancer therapy; cell motility; cellular imaging; digital; drug candidate; flexibility; gastrulation; imaging biomarker; imaging modality; in vitro Model; indexing; inhibitor/antagonist; innovation; insight; instrument; microscopic imaging; non-invasive imaging; novel; polarized light; prognostic value; reconstruction; scale up; success; targeted treatment; therapeutic candidate; three-dimensional modeling; tumor microenvironment

7. Project Summary/Abstract Cancer metastases are responsible for most deaths from the disease. However, most current cancer therapies are anti-proliferative, rather than anti-metastatic. Challenges to the clinical realization of anti-metastatic therapies include dose-limiting toxicity to non-cancer cells, effective targeting, effective timing of administration given that metastasis is an early cancer process, and effectiveness in the face of adaptive invasion strategies by cancer cells. An ideal platform to test anti-metastatic therapeutic candidates would mimic the tumor microenvironment, and accurately assess cancer cell motile phenotypes as well as pro-invasion microstructural signatures in the extracellular matrix. This proposal aims to 1) develop a dual-modality quantitative phase and polarized light microscopy system capable of 2) evaluating the effects of inhibitors of matrix invasion and microenvironmental factors on the spread of cancer cells in a tissue-like in vitro environment. Quantitative optical indices from the proposed system accurately assess cell phenotype and microstructural signatures of invasion, by evaluating cell phase and matrix birefringence signals. These imaging modalities also deliver low optical power to the sample, allowing for long-term, serial microscopy without phototoxic effects on cancer cell movement. Finally, an innovative but simple culture set-up creates collagen networks with alignment and pre-stress similar to the tumor microenvironment. Steps to achieve these aims include addition of polarizing optics to an existing digital holographic microscope, signal calibration, channel co-registration, and initial time-lapse imaging of an in vitro 3D model of cancer invasion. Optical indices of invasion will be evaluated in a scaled-up study. After installing a second camera and full polarization state generator and analyzer on the existing digital holographic microscope, phase and birefringence signals will be evaluated and co-registered using a polystyrene microsphere standard (n=1.59) set in solid mounting media (n=1.52). Phase and polarized light parameters will be calibrated by computing phase maps of standard beads of fixed diameter, and optical retardance of a zero-order waveplate. The invasion of the breast cancer cell line MDA-MB-231 will be evaluated from serial time-lapse imaging over 24 hours, in the presence and absence of 30 nM chondramide, an actin-stabilizing anti-metastatic therapeutic candidate. The effects of pre-stress and extracellular matrix alignment will also be evaluated. The health-relatedness of this proposal lies in development of a quantitative phase and polarized light microscope that computes parameter maps for invading cancer cells and their microenvironmental surroundings. The proposed microscope reduces phototoxicity and provides quantitative metrics for accurate assessment of the mechanisms and aggressiveness of cancer cell invasion, thus enabling testing of anti- metastatic drug candidates.

7。项目摘要/摘要 癌症转移造成大多数疾病死亡。但是，大多数目前的癌症疗法 是抗增殖的，而不是抗转移性的。抗复位的临床实现挑战 疗法包括对非癌细胞的剂量限制毒性，有效的靶向，有效的给药时间 鉴于转移是一个早期的癌症过程，并且面对适应性入侵策略的有效性 由癌细胞。测试抗转移性治疗候选者的理想平台将模仿肿瘤 微环境，并准确评估癌细胞运动表型以及促侵袭微结构 细胞外基质中的特征。 该建议的目的是1）开发双模式定量阶段和极化光显微镜系统 2）评估基质侵袭和微环境因素对抑制剂对 癌细胞在组织样的体外环境中的传播。来自提议的定量光学索引 系统通过评估细胞相准确地评估侵袭的细胞表型和微结构特征 和矩阵双向信号。这些成像方式还为样品提供低光功率， 允许长期的连续显微镜对癌细胞运动没有光毒性影响。最后，一个 创新但简单的文化设置创建了胶原蛋白网络，并具有一致性和预感类似 肿瘤微环境。实现这些目标的步骤包括将两极化光学添加到现有数字 全息显微镜，信号校准，通道共注册和体外初始延时成像 3D癌症入侵模型。入侵的光学指标将在一项扩展研究中评估。 在现有数字上安装了第二个摄像头和完整的极化状态生成器和分析仪之后 全息显微镜，相位和双发性信号将通过A进行评估和注册 聚苯乙烯微球标准（n = 1.59）设置在固体安装介质中（n = 1.52）。相和偏振光 参数将通过计算固定直径和光学标准珠的相位图来校准 零级波板的延迟。乳腺癌细胞系MDA-MB-231的入侵将是 在存在和不存在30 nm软骨胺的情况下，通过在24小时内的连续延时成像进行评估， 肌动蛋白稳定的抗转移治疗候选者。预压力和细胞外基质的影响 还将评估对齐方式。 该提案的健康相关性在于定量阶段和两极分化的光 计算入侵癌细胞及其微环境的参数图的显微镜 环境。所提出的显微镜降低了光毒性，并提供了定量指标以准确 评估癌细胞侵袭的机制和攻击性，从而实现了抗 转移性药物候选人。