BioMediTech, Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön Katu 34, 33520, Tampere, Finland; INL - International Iberian Nanotechnology Laboratory, Ultrafast Bio- and Nanophotonics Group, Av. Mestre José Veiga s/n, 4715-330, Braga, Portugal.
BioMediTech, Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön Katu 34, 33520, Tampere, Finland.
J Mech Behav Biomed Mater. 2023 Oct;146:106069. doi: 10.1016/j.jmbbm.2023.106069. Epub 2023 Aug 9.
Cellular physiology has been mainly studied by using two-dimensional cell culture substrates which lack in vivo-mimicking extracellular environment and interactions. Thus, there is a growing need for more complex model systems in life sciences. Micro-engineered scaffolds have been proven to be a promising tool in understanding the role of physical cues in the co-regulation of cellular functions. These tools allow, for example, probing cell morphology and migration in response to changes in chemo-physical properties of their microenvironment. In order to understand how microtopographical features, what cells encounter in vivo, affect cytoskeletal organization and nuclear mechanics, we used direct laser writing via two-photon polymerization (TPP) to fabricate substrates which contain different surface microtopographies. By combining with advanced high-resolution spectral imaging, we describe how the constructed grid and vertical line microtopographies influence cellular alignment, nuclear morphology and mechanics. Specifically, we found that growing cells on grids larger than 10 × 20 μm and on vertical lines increased 3D actin cytoskeleton orientation along the walls of microtopographies and abolished basal actin stress fibers. In concert, the nuclei of these cells were also more aligned, elongated, deformed and less flattened, indicating changes in nuclear force transduction. Importantly, by using fluorescence lifetime imaging microscopy for measuring Förster resonance energy transfer for a genetically encoded nesprin-2 molecular tension sensor, we show that growing cells on these microtopographic substrates induce lower mechanical tension at the nuclear envelope. To conclude, here used substrate microtopographies modulated the cellular mechanics, and affected actin organization and nuclear force transduction.
细胞生理学主要通过使用二维细胞培养底物来研究，这些底物缺乏体内模拟的细胞外环境和相互作用。因此，生命科学领域越来越需要更复杂的模型系统。微工程支架已被证明是理解物理线索在细胞功能的共同调节中的作用的一种有前途的工具。这些工具例如允许探测细胞形态和迁移，以响应其微环境的化学物理性质的变化。为了了解体内细胞遇到的微形貌特征如何影响细胞骨架组织和核力学，我们使用双光子聚合（TPP）的直接激光写入来制造包含不同表面微形貌的底物。通过与先进的高分辨率光谱成像相结合，我们描述了构建的网格和垂直线微形貌如何影响细胞排列、核形态和力学。具体来说，我们发现，在大于 10×20 μm 的网格和垂直线上生长的细胞增加了 3D 肌动蛋白细胞骨架沿着微形貌壁的取向，并消除了基底肌动蛋白应力纤维。协调一致的是，这些细胞的细胞核也更加对齐、拉长、变形和更少扁平，表明核力转导发生变化。重要的是，通过使用荧光寿命成像显微镜测量遗传编码的 nesprin-2 分子张力传感器的Förster 共振能量转移，我们表明，在这些微形貌底物上生长的细胞在核膜处诱导的机械张力较低。总之，这里使用的底物微形貌调节了细胞力学，并影响了肌动蛋白组织和核力转导。
