Interference Traction Force Microscopy (iTFM) for Bioimaging of Cellular Forces

人体由超过10万亿个独立细胞组成，每个细胞在某种程度上都是一个独立的实体。这个极其复杂的系统的协调依赖于一个错综复杂的信号网络，其中的许多方面仍然知之甚少。虽然化学信号（如激素）参与这种具有挑战性的协调已被广泛接受，但现在人们对细胞如何在机械水平上相互作用以及与环境相互作用的兴趣迅速增加，例如通过推和拉或通过感知局部环境中的刚度或机械应力。这是因为有强有力的证据表明，机械信号传导（即机械转导）在许多重要过程中起着关键作用，从控制人类胚胎生长中的干细胞分化到影响重要疾病的进展。然而，虽然机械转导对改善人类健康显然既迷人又重要，但由于缺乏适当的方法来“成像”细胞或细胞成分施加的力，我们在细胞水平上研究力学的能力仍然有限。(已经开发了几种研究细胞力的显微镜方法，但它们的应用需要特殊的专业知识，而这些专业知识在生物学实验室中通常是不具备的，即使有这种专业知识，在细胞生物学中许多相关过程发生的较长时间内跟踪细胞的机械行为仍然是非常具有挑战性的。此外，现有的方法不够灵敏，无法研究较弱细胞的活性。在这个项目中，光被用作光学尺子，以准确而稳健地成像细胞施加的力模式。利用一种称为光学干涉的效应，可见光的短波长将使力成像的灵敏度比现有技术提高5到10倍。此外，光波将为新方法提供内部参考，这意味着它将能够在很长一段时间（长达几周）内跟踪力模式，而不需要许多现有方法目前需要的破坏性参考测量。在细胞生物学的许多领域，这项新技术可能被证明是非常有用的。由于这个项目的短期性质，它最初将应用于一个具体的例子：研究通常在肾脏中发现的细胞粘附力，这些细胞负责血液过滤，从而从体内清除废物。这些细胞处于恒定的机械压力下，它们的衰竭与肾功能衰竭有关。因此，研究相关力量有望提高我们对肾脏功能的理解，并可能在长期内找到早期发现和治疗肾衰竭的方法，肾衰竭通常与肥胖和糖尿病有关。

The human body consists of over 100,000 billion individual cells, with each cell being on some level an independent entity. Coordination of this hugely complex system relies on an intricate network of signalling, many aspects of which are still poorly understood. While the involvement of chemical signals (e.g. hormones) in this challenging coordination is widely accepted, there is now rapidly increasing interest in how cells interact with each other and with their environment on the mechanical level, for example by pushing and pulling or by sensing the stiffness or mechanical stress in their local environment. This is motivated by strong evidence that mechanical signalling (known as mechanotransduction) plays a critical role in a number of important processes, ranging from controlling how stem cells differentiate in the growing human embryo to affecting the progression of important diseases. However, while mechanotransduction is therefore obviously both fascinating and important for improving human health, our ability to study mechanics on a cellular level remains limited due to a lack of suitable methods to 'image' the forces that cells or components of cells apply. (Several microscopy methods to investigate cell force have been developed but their application requires special expertise that is often not available in a biology lab and even if this expertise is available it remains very challenging to follow the mechanical behaviour of cells over the extended time periods over which many relevant processes in cell biology occur. In addition, existing methods are not sensitive enough to study the activity of weaker cells.) In this project, light is used as an optical ruler to accurately and robustly image the force pattern applied by cells. Using an effect known as optical interference, the short wavelength of visible light will facilitate an increase in the sensitivity of force imaging by 5 to 10-fold over existing techniques. In addition, light-waves will provide an internal reference for the new method which means that it will be able to track force patterns over long periods of time (up to several weeks) without a need for disruptive reference measurements which many of the existing methods currently require. There are many areas of cell biology where the new technique may prove to be highly useful. Due to the short-term nature of this project, it will initially be applied to one specific example: a study of the forces involved in the adhesion of cells normally found in the kidney where they are responsible for blood filtration and thus for removal of waste products from the body. These cells are under constant mechanical pressure and their failure is associated with kidney malfunction. Studying the involved forces therefore promises to improve our functional understanding of the kidney and may in the long-run lead to methods for early detection and treatment of kidney failure, a condition often associated with obesity and diabetes.