CAREER:The role of nuclear biomechanics during cell migration in 3-D environments

派：拉默丁。JanProposal ID：1254846智能优点：虽然细胞在二维(2D)底物上的迁移已经研究了几十年，但细胞在三维环境中移动时面临的物理挑战现在才刚刚开始出现。这一提议的中心假设是，在三维环境中，如细胞外基质或在细胞内或外渗过程中，大而硬的细胞核的变形是通过紧密收缩进行迁移的限速步骤。该项目将研究细胞核的变形能力在多大程度上控制细胞通过狭窄的运输，通过监测细胞通过限定狭窄的微流控通道迁移，同时可视化亚细胞变形和运输效率。此外，当细胞通过狭窄的狭窄时，将使用飞秒激光脉冲消融细胞骨架细丝，以确定细胞核是被细胞骨架拉动还是推动，以克服其在挤压通过小于细胞核大小的狭窄时的阻力。作为对这些实验的补充，由改良的核膜蛋白设计的分子传感器将通过荧光强度或荧光共振能量转移(FRET)效率的变化来报告核膜上的张力和局部压缩，并将应用于3D迁移过程中的细胞骨架力量化，并可视化它们在核外围的分布。广泛的影响：研究活动将与一个全面的教育和推广计划相辅相成，该计划建立在拟议研究的简化模型的基础上，向初中生介绍生物医学工程，目标是增加女学生对科学、技术、工程和数学(STEM)学科的参与。木薯珍珠是广受欢迎的泡泡茶的关键成分，它将被用作安全、容易获得和廉价的生物细胞替代品，而简单的“微流体”设备将取代技术要求苛刻的微流体设计，以探测细胞生物力学。作为康奈尔大学现有的和新的外展项目的一部分，康奈尔大学将向服务不足的初中和高中的学生介绍这些“木薯微流体”实验和相应的讲座，这些项目针对高中女生和代表不足的少数族裔，为她们提供生物医学工程方面的实践经验，以增强信心，满足榜样，并克服性别刻板印象。另一个教育组成部分是创建一个在线门户网站，用于自我指导学习，以应对向具有不同学术背景的学生教授跨学科生物医学工程课程的挑战。

PI: Lammerdng. JanProposal ID: 1254846Intellectual Merits: While cell migration on 2-dimensional (2-D) substrates has been studied for many decades, the physical challenges cells face when moving in 3-D environments are only beginning to emerge now. The central hypothesis of this proposal is that deformation of the large and stiff nucleus presents a rate-limiting step during migration through tight constriction encountered in 3-D environments such as extracellular matrixes or during intra- or extravasation. This project will address to what extent the deformability of the nucleus governs the transit of cells through narrow constrictions by monitoring cells as they migrate through microfluidic channels with defined constrictions while visualizing subcellular deformations and transit efficiency. In addition, ablation of cytoskeletal filaments with femtosecond laser pulses as cells pass through narrow constrictions will be used to determine whether the nucleus is being pulled or pushed by the cytoskeleton to overcome its resistance when squeezing through constrictions smaller than the size of the nucleus. Complementing these experiments, molecular sensors engineered from modified nuclear envelope proteins that report tensile forces and localized compression at the nuclear envelope by changes in fluorescence intensity or fluorescence resonance energy transfer (FRET) efficiency will be applied to cells during 3-D migration to quantify the cytoskeletal forces acting on the nucleus during 3-D migration and visualize their distribution along the nuclear periphery.Broader Impact:The research activities will be complemented with a comprehensive education and outreach program that builds on simplified models of the proposed research to introduce middle and high school students to biomedical engineering, with the goal of increasing participation of female students in science, technology, engineering, and math (STEM) subjects. Tapioca pearls, the key ingredient of the popular bubble tea, will be used as safe, easily accessible, and inexpensive stand-in for biological cells, while simple 'millifluidic' devices replace technically demanding microfluidics designs to probe cellular biomechanics. These 'tapioca millifluidics' experiments, along with accompanying lectures, will be introduced to students from underserved middle and high schools as part of existing and novel outreach programs at Cornell University that target high school girls and underrepresented minorities to provide them with hands-on experiences in biomedical engineering to boost confidence, meet role models, and overcome gender stereotypes. An additional education component is the creation of an online portal for self-directed learning to address the challenge of teaching interdisciplinary biomedical engineering courses to students with diverse academic backgrounds.