Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA.
Rev Sci Instrum. 2021 Nov 1;92(11):114101. doi: 10.1063/5.0056366.
The mechanotransduction pathways that mediate cellular responses to contact forces are better understood than those that mediate response to distance forces, especially the force of gravity. Removing or reducing gravity for significant periods of time involves either sending samples to space, inducing diamagnetic levitation with high magnetic fields, or continually reorienting samples for a period, all in a manner that supports cell culturing. Undesired secondary effects due to high magnetic fields or shear forces associated with fluid flow while reorienting must be considered in the design of ground-based devices. We have developed a lab-friendly and compact random positioning machine (RPM) that fits in a standard tissue culture incubator. Using a two-axis gimbal, it continually reorients samples in a manner that produces an equal likelihood that all possible orientations are visited. We contribute a new control algorithm by which the distribution of probabilities over all possible orientations is completely uniform. Rather than randomly varying gimbal axis speed and/or direction as in previous algorithms (which produces non-uniform probability distributions of orientation), we use inverse kinematics to follow a trajectory with a probability distribution of orientations that is uniform by construction. Over a time period of 6 h of operation using our RPM, the average gravity is within 0.001 23% of the gravity of Earth. Shear forces are minimized by limiting the angular speed of both gimbal motors to under 42 °/s. We demonstrate the utility of our RPM by investigating the effects of simulated microgravity on adherent human osteoblasts immediately after retrieving samples from our RPM. Cytoskeletal disruption and cell shape changes were observed relative to samples cultured in a 1 g environment. We also found that subjecting human osteoblasts in suspension to simulated microgravity resulted in less filamentous actin and lower cell stiffness.
介导细胞对接触力反应的机械转导途径比介导对距离力（尤其是重力）反应的途径理解得更好。长时间去除或减少重力，要么将样品送到太空，要么用强磁场产生抗磁性悬浮，要么在一段时间内不断重新定向样品，所有这些都要支持细胞培养。在设计基于地面的设备时，必须考虑由于高磁场或与重新定向相关的剪切力而产生的不良次要效应。我们开发了一种便于实验室使用且紧凑的随机定位机（RPM），可放置在标准细胞培养孵育箱中。使用双轴万向节，它以一种使所有可能的方向都有相同机会被访问的方式不断重新定向样品。我们贡献了一种新的控制算法，通过该算法，所有可能方向的概率分布完全均匀。与以前的算法（其中产生方向的非均匀概率分布，该算法随机改变万向节轴的速度和/或方向）不同，我们使用运动学逆推来遵循具有均匀方向概率分布的轨迹，该轨迹的概率分布是构造性均匀的。使用我们的 RPM 运行 6 小时的过程中，平均重力在地球重力的 0.00123%以内。通过将两个万向节电机的角速度限制在 42°/s 以下，最大限度地减少了剪切力。通过从我们的 RPM 中取回样品后立即研究模拟微重力对贴壁人成骨细胞的影响，证明了我们的 RPM 的实用性。与在 1 g 环境中培养的样品相比，观察到细胞骨架的破坏和细胞形状的变化。我们还发现，将悬浮培养的人成骨细胞置于模拟微重力下会导致丝状肌动蛋白减少和细胞刚性降低。
