An ultra-sensitive micro sensor for biophysical studies of single cells cultured in 3D extracellular matrix

用于 3D 细胞外基质中培养的单细胞生物物理研究的超灵敏微传感器

• 批准号: 1934991
• 负责人: Taher Saif
• 金额: $ 40.5万
• 依托单位: University of Illinois at Urbana-Champaign
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2024-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1934991&HistoricalAwards=false
• 关键词: ultra; sensitive; micro; sensor; biophysical

Considerable experimental evidence acquired during the last three decades has now established that living cells are contractile. They generate force on 3D tissues or 2D substrates. Forces and matrix stiffness have a profound influence on a wide range of cell behavior such as growth, motility, and differentiation. More recently, it is appreciated that cells not only respond to these cues from their 3D matrix, but also remodel the matrix and hence influence the subsequent cues. Cancer cells exploit such reciprocity within the tumor to support their metastatic journey. The precise role of mechanics in determining this reciprocity remains elusive. For example, how cells transduce mechanical cues into biochemical processes and vice versa. Which processes are involved in remodeling the matrix? Probing these fundamental questions need experimental platforms that allow to evaluate cell-forces in a 3D matrix and quantify the key parameters that determine the cell-matrix dynamics. Methods to measure single cell forces on 2D have been advanced over the last three decades. Currently, there is no method available to measure cell forces in 3D. Centennial High School (Champaign) drama teacher will assist in developing a drama with high school students representing cancer development where characters will represent cancer, fibroblast and immune cells. The scenes will mimic various stages of cancer. Research will be integrated with education through involvement of undergraduate students from minorities in research, and web dissemination. The PI has a long record of outreach activities, including multiple videos series development for students and public. The goal of this project is to close this gap. It will design, develop and test an ultra-sensitive (~10nN/um) micro force sensor that allows to quantify single cell force, dynamics, and extra-cellular matrix remodeling in 3D matrix. The sensor will be applied to quantify force generation and matrix remodeling by fibroblasts, and to explore force-dependent cross talk between cancer and stromal cells leading to metastatic progression. The sensor consists of two grids, one connected to a soft spring and the other to a rigid support. A droplet of liquid cell-matrix (e.g., collagen) mixture is dispensed on the grids. It fills the grids due to capillarity and forms a free-standing tissue bridging between them. The tissue contains one or a discrete number of cells depending on the questions to be addressed. The cell(s) forms adhesion with the matrix and generates force, which is sensed by the spring. The extra cellular matrix stiffness can also be obtained by the sensor by stretching the matrix. The sensor will be used to study single fibroblasts in 3D collagen for their force, dynamics and matrix remodeling. Finally, the interactions between cancer cells and cancer associated fibroblasts in 3D matrix and the role of forces on the interaction will be explored. The outreach activities will include developing a drama with high school students representing cancer development where characters will represent cancer, fibroblast and immune cells. The scenes will mimic various stages of cancer.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

在过去的三十年中获得的大量实验证据现在已经确定活细胞是可收缩的。它们在3D组织或2D基质上产生力。力和基质刚度对广泛的细胞行为如生长、运动和分化具有深远的影响。最近，人们认识到，细胞不仅对来自其3D矩阵的这些线索做出反应，而且还重塑矩阵，从而影响后续线索。癌细胞利用肿瘤内的这种相互作用来支持它们的转移过程。力学在确定这种相互作用中的确切作用仍然是难以捉摸的。例如，细胞如何将机械信号转化为生化过程，反之亦然。哪些过程参与重塑基质？探索这些基本问题需要实验平台，允许在3D矩阵中评估细胞力，并量化决定细胞-基质动力学的关键参数。在过去的三十年中，测量二维单细胞力的方法已经取得了进展。目前，还没有方法可用于测量3D中的细胞力。百年高中（尚潘）戏剧老师将协助开发一个戏剧与高中学生代表癌症的发展，其中字符将代表癌症，成纤维细胞和免疫细胞。这些场景将模拟癌症的各个阶段。将通过少数民族本科生参与研究和网络传播，将研究与教育结合起来。PI有很长的推广活动记录，包括为学生和公众制作多个视频系列。 该项目的目标是缩小这一差距。它将设计，开发和测试一种超灵敏（~ 10 nN/um）的微力传感器，可以量化3D矩阵中的单细胞力，动力学和细胞外基质重塑。该传感器将被应用于量化成纤维细胞的力生成和基质重塑，并探索导致转移进展的癌症和基质细胞之间的力依赖性串扰。传感器由两个网格组成，一个连接到软弹簧，另一个连接到刚性支撑。将液体细胞基质的液滴（例如，胶原蛋白）混合物分配在网格上。由于毛细作用，它填充网格并在它们之间形成独立的组织桥接。根据要解决的问题，组织包含一个或离散数量的细胞。细胞与基质形成粘附并产生力，该力由弹簧感测。细胞外基质刚度也可以通过拉伸基质由传感器获得。该传感器将用于研究3D胶原中的单个成纤维细胞的力、动力学和基质重塑。最后，将探讨3D矩阵中癌细胞和癌相关成纤维细胞之间的相互作用以及力对相互作用的作用。宣传活动将包括与代表癌症发展的高中生一起制作一部戏剧，其中人物将代表癌症、成纤维细胞和免疫细胞。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Dose-independent threshold illumination for non-invasive time-lapse fluorescence imaging of live cells

用于活细胞非侵入性延时荧光成像的剂量无关阈值照明

• DOI: 10.1016/j.eml.2021.101249
• 发表时间: 2021
• 期刊: Extreme Mechanics Letters
• 影响因子: 4.7
• 作者: Emon, M.A. Bashar;Knoll, Samantha;Doha, Umnia;Ladehoff, Lauren;Lalonde, Luke;Baietto, Danielle;Sivaguru, Mayandi;Bhargava, Rohit;Saif, M. Taher
• 通讯作者: Saif, M. Taher