Meinig School of Biomedical Engineering, Weill Institute for Cell and Molecular Biology, Cornell University, USA.
Laboratoire MSSMat UMR CNRS 8579, CentraleSupelec, Université Paris-Saclay, France.
Lab Chip. 2019 Nov 7;19(21):3652-3663. doi: 10.1039/c9lc00444k. Epub 2019 Sep 27.
The mechanical properties of the cell nucleus are increasingly recognized as critical in many biological processes. The deformability of the nucleus determines the ability of immune and cancer cells to migrate through tissues and across endothelial cell layers, and changes to the mechanical properties of the nucleus can serve as novel biomarkers in processes such as cancer progression and stem cell differentiation. However, current techniques to measure the viscoelastic nuclear mechanical properties are often time consuming, limited to probing one cell at a time, or require expensive, highly specialized equipment. Furthermore, many current assays do not measure time-dependent properties, which are characteristic of viscoelastic materials. Here, we present an easy-to-use microfluidic device that applies the well-established approach of micropipette aspiration, adapted to measure many cells in parallel. The device design allows rapid loading and purging of cells for measurements, and minimizes clogging by large particles or clusters of cells. Combined with a semi-automated image analysis pipeline, the microfluidic device approach enables significantly increased experimental throughput. We validated the experimental platform by comparing computational models of the fluid mechanics in the device with experimental measurements of fluid flow. In addition, we conducted experiments on cells lacking the nuclear envelope protein lamin A/C and wild-type controls, which have well-characterized nuclear mechanical properties. Fitting time-dependent nuclear deformation data to power law and different viscoelastic models revealed that loss of lamin A/C significantly altered the elastic and viscous properties of the nucleus, resulting in substantially increased nuclear deformability. Lastly, to demonstrate the versatility of the devices, we characterized the viscoelastic nuclear mechanical properties in a variety of cell lines and experimental model systems, including human skin fibroblasts from an individual with a mutation in the lamin gene associated with dilated cardiomyopathy, healthy control fibroblasts, induced pluripotent stem cells (iPSCs), and human tumor cells. Taken together, these experiments demonstrate the ability of the microfluidic device and automated image analysis platform to provide robust, high throughput measurements of nuclear mechanical properties, including time-dependent elastic and viscous behavior, in a broad range of applications.
细胞核的力学特性在许多生物学过程中被越来越多地认为是至关重要的。细胞核的可变形性决定了免疫细胞和癌细胞穿过组织和内皮细胞层的迁移能力，而细胞核力学性质的变化可以作为癌症进展和干细胞分化等过程中的新型生物标志物。然而，目前测量核粘弹性力学特性的技术往往耗时较长，一次只能探测一个细胞，或者需要昂贵的、高度专业化的设备。此外，许多当前的测定方法没有测量具有粘弹性材料特征的时变特性。在这里，我们提出了一种易于使用的微流控装置，该装置应用了已建立的微管吸吮方法，适用于并行测量多个细胞。该装置设计允许快速加载和清洗细胞进行测量，并最大限度地减少大颗粒或细胞团堵塞。与半自动图像分析流水线相结合，微流控装置方法可显著提高实验通量。我们通过将设备中的流体力学计算模型与实验测量的流体流动进行比较，验证了实验平台的有效性。此外，我们还对缺乏核膜蛋白 lamin A/C 的细胞和野生型对照细胞进行了实验，这些细胞具有特征明确的核力学性质。对时变核变形数据进行幂律和不同粘弹性模型拟合表明， lamin A/C 的缺失显著改变了核的弹性和粘性性质，导致核的可变形性显著增加。最后，为了展示设备的多功能性，我们在各种细胞系和实验模型系统中对核的粘弹性力学性质进行了表征，包括来自与扩张型心肌病相关的 lamin 基因突变个体的人皮肤成纤维细胞、健康对照成纤维细胞、诱导多能干细胞 (iPSC) 和人肿瘤细胞。总之，这些实验证明了微流控装置和自动图像分析平台能够提供强大的、高通量的核力学性质测量，包括广泛应用中的时变弹性和粘性行为。
