Instrumenting blood platelets: nanosensors for cumulative shear and compression measurement

在这里，我们开发了新的原位和循环纳米级传感技术，到目前为止还不能直接 测量血小板在循环过程中所经历的累积流体结构力。高架剪力 对植入式心血管设备的无数不良事件负有责任，包括血栓形成和 在流血。我们建议利用利用DNA纳米技术制造的机械传感器来量化物理力 循环细胞在穿过高切变流动路径时所经历的扰动。通过创建和 这一新的检测工具系统的特征，这项工作有可能改善和提高安全性和 心血管治疗设备的疗效。此重新提交已根据有用的反馈进行了更新。 在纳米尺度上，推力、拉力和剪切力驱动着发育中的生化有益过程。 而重塑，也是疾病进展中的病理过程。在血小板中，剪切激活作为 在脑室辅助装置或狭窄的动脉粥样硬化中的经历会增加发生 血栓形成，但没有工具来测量给定循环将给予的剪切剂量，即强度x时间， 我们开发治疗干预措施的能力是有限的。纳米级的传感器模块将允许我们询问 不仅是血小板负荷如何影响血栓形成潜能，而且还可以预测植入性装置如何与 具有特定血管系统的患者将影响中风或其他血栓栓塞症的可能性，并最终通知 增强血液相容性的未来设计。在一个涉及生物工程师的高风险、高回报的合作中 和医生，我们建议应用DNA折纸方法来创造两种荧光和可调的 双稳态纳米传感器：一种对剪切载荷敏感(表面平行)，另一种对剪切载荷敏感 压缩载荷(表面垂直)。我们将装饰微米级的聚合物微珠以及 利用这些传感器，以及使用微流控平台，我们将调整这些机制 粒子传感器系统，包括它们的连接，以最大限度地提高对剪切和冲击的敏感度。 基于DNA的机械传感器的分子精度和可编程性可以实现高度并行 对几十飞牛顿的力具有可调灵敏度的测量。这一跨学科领域有 历史上是由蜂窝技术的精确应用和测量的发展推动的 和分子力；每一个新的工具都使大量的新的调查路线成为可能，通过这项拟议的工作，DNA- 基于纳米技术可以导致心血管机械生物学领域的另一场变革。 这一努力的成功将导致创造用于测量累积剪切和 循环细胞在高剪切微环境中移动时所经历的压缩载荷。 这种测量的洞察力将使改进的植入性设备、药理学 缓解剪切效应的试剂和新的研究思路，以提高我们对剪切和 血小板活化中的压缩负荷。

Here we develop novel in situ and circulating nanoscale sensing technologies heretofore unavailable to directly measure the cumulative fluid-structure forces experienced by blood platelets during circulation. Elevated shear is responsible for the myriad of adverse events in implantable cardiovascular devices including thrombosis and bleeding. We propose to utilize mechanosensors made using DNA nanotechnology to quantify physical force perturbations experienced by circulating cells as they traverse high shear flow paths. By creating and characterizing this new assay tools system, this work has potential to improve and enhance the safety and efficacy of cardiovascular therapeutic devices. This resubmission has been updated based on helpful feedback. At the nanoscale, pushing, pulling, and shearing forces drive biochemical beneficial processes in development and remodeling, but also pathological processes in disease progression. In blood platelets, shear activation as experienced in a ventricular assist device or in stenotic atherosclerotic artery leads to an increased likelihood of thrombosis, but without a tool to measure the shear dose, i.e. intensity x time, that a given circulation will impart, our capability to develop therapeutic interventions is limited. Nanoscale sensor modules would allow us to ask not only how platelet loads affects thrombotic potential, but also to predict how implantable devices in conjunction with specific vasculature will affect potential for stroke or other thromboembolic events, and ultimately inform future designs for enhanced hemocompatibility. In a high-risk, high-reward collaboration involving bioengineers and physicians, we propose to apply the DNA origami approach to create two varieties of fluorescent and tunable bistable nanosensors: one that is sensitive to shear loading (surface parallel) and another that is sensitive to compressive loading (surface perpendicular). We will decorate micron-scale polymer microbeads as well as human blood platelets with these sensors, and using microfluidic platforms, we will tune the mechanics of these sensor-on-particle systems, including their linkages, to maximize their sensitivity to both shear and impact. The molecular precision and programmability of DNA-based mechanosensors can enable highly parallel measurements with tunable sensitivity to forces as small at tens of femtonewtons. This interdisciplinary field has historically been driven by the development of technologies for precise application and measurement of cellular and molecular forces; each new tool has enabled vast new lines of inquiry, and with this proposed work, DNA- based nanotechnologies can lead to another transformation in the field of cardiovascular mechanobiology. Success in this endeavor will result in the creation of novel tools for measuring the cumulative shear and compressive loading that circulating cells experience as they move through high shear microenvironments. Insights from such measurements will enable the development of improved implantable devices, pharmacologic agents to mitigate shear effects and new lines of inquiry for improving our understanding of the role of shear and compressive loading in platelet activation.