Development of Brillouin Spectroscopy for Mechanotransduction Research

用于力传导研究的布里渊光谱学的发展

• 批准号: BB/N021576/1
• 负责人: Che Connon
• 金额: $ 19.21万
• 依托单位: Newcastle University
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2016
• 资助国家: 英国
• 起止时间: 2016 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FN021576%2F1
• 关键词: Development; Brillouin; Spectroscopy; Mechanotransduction; Research; Development; Brillouin; Spectroscopy; Mechanotransduction; Research

Mechanobiology is a rapidly developing area of science with the potential to improve significantly our knowledge of how cells function at a tissue/organ level. A better understanding of how cells react to their local mechanical environment (extracellular matrix, fluid flow, etc.) will also lead to advanced biomedical applications such as improved biomaterials and cell therapy. For such new therapeutic strategies, it is necessary to provide cells with a confined environment (niche) that enhances and regulates their proliferation and differentiation. In this regard, biomaterial technology currently leads the way in trying to fulfil these requirements by artificially recreating native-like three-dimensional environments. However, understanding how, why, and in what environment these cells differentiate in a lineage-specific manner is essential for understanding regenerative biology and developing better tissue engineering and stem cell therapy approaches. We believe that this underlying biology (i.e., the cell's responses to local mechanical stimuli) has not yet been properly investigated, due to a lack of appropriate tools, and that this is likely to undermine current attempts to i) understand mechanobiology; and ii) recreate the stem cell niche using purposeful biomaterials. Therefore, we plan to understand the role substrate stiffness plays in maintaining/directing cell phenotype at an unprecedented level by developing a sophisticated microscope that can quantify mechanical properties of the cells and their immediate environment whilst simultaneously measuring the cells' response to this environment at a gene and protein level. Our novel approach uses only light, and thus requires no direct physical contact with the sample and can probe deep into tissue structures. Most importantly, it can be performed under physiological conditions on live cells, facilitating real-time measurements under dynamic conditions. Such experiments have not previously been possible.However, based on our recent success in building a confocal imaging system to measure tissue stiffness in 3D, we think this is now possible. Using the principle of Brillouin spectroscopy, the system detects light that has been scattered inelastically from acoustic phonons in the sample in a confocal optical arrangement to facilitate a non-contact, direct readout of the mechanical properties of tissues. We now wish to explore if this same confocal system can be used simultaneously to excite fluorescent molecules within the probed tissues. As such, our aims are to modify our existing machine so that confocal images of fluorescently-labelled proteins (from cells, tissues or tissue-engineered constructs) can be resolved and overlaid with simultaneous measurements of the mechanical properties of said cells/tissues. Ultimately, this will allow, for the first time and with unprecedented detail, the direct, real-time account of a cell's molecular response to differing local mechanical environments.

机械生物学是一个迅速发展的科学领域，具有显着提高我们对细胞在组织/器官水平的功能的潜力。更好地理解细胞对其局部机械环境的反应（细胞外基质，流体流量等）也将导致先进的生物医学应用，例如改善的生物材料和细胞疗法。对于这种新的治疗策略，有必要为细胞提供一个狭窄的环境（利基），以增强和调节其扩散和分化。在这方面，生物材料技术目前通过人为地重新创建类似土著的三维环境来实现这些要求的方式。但是，了解这些细胞以谱系特异性方式分化的环境如何，为什么以及在哪个环境中对于理解再生生物学以及开发更好的组织工程和干细胞治疗方法至关重要。我们认为，由于缺乏适当的工具，尚未对这种潜在的生物学（即细胞对局部机械刺激的反应）进行适当的研究，并且这可能会破坏当前尝试的尝试I）理解机械生物学； ii）使用有目的的生物材料重新创建干细胞生态位。因此，我们计划通过开发一个可以量化细胞的机械性能及其直接环境的复杂显微镜，同时测量细胞对基因和蛋白质水平上的环境的反应，同时测量细胞的响应，从而了解底物质量在保持前所未有的水平上保持/指导细胞表型的作用。我们的新方法仅使用光，因此不需要与样品直接接触，并且可以深入组织结构。最重要的是，它可以在生理细胞的生理条件下进行，从而促进动态条件下的实时测量。以前没有这样的实验。但是，基于我们最近在建立共聚焦成像系统来测量3D组织刚度的成功，我们认为这是可能的。使用布里鲁因光谱的原理，该系统检测到在共焦光学排列中从样品中的声音子中散布的光，以促进组织机械性能的非接触，直接读数。现在，我们希望探索是否可以同时使用相同的共聚焦系统来激发探测的组织中的荧光分子。因此，我们的目的是修改现有的机器，以便可以解决荧光标记蛋白（来自细胞，组织或组织工程构建体）的共聚焦图像，并通过同时测量所述细胞/组织的机械性能的同时测量。最终，这将首次允许，并具有前所未有的细节，即细胞对不同局部机械环境的分子响应的直接实时说明。