AFM / Confocal images and dataset

This dataset consist on AFM/confocal confinement at 3µm of HeLa Kyoto MYH9-eGFP / lifeact-mCherry.<br> Dishes with cells were mounted in a dish heater (JPK Instruments) and kept at 37 °C under an inverted light microscope (Axio Observer.Z1; Zeiss) equipped with a confocal microscope unit (LSM 700; Zeiss) and the atomic force microscope (AFM) head (CellHesion 200; JPK Instruments). Focused ion beam (FIB)-sculpted, flat silicon microcantilevers were processed and calibrated as described in (Cattin et al., 2015). The microcantilevers were fixed on a standard JPK glass block and mounted in the AFM head. The cantilever was lowered on the cell to a preset height with a constant speed of 0.5 μm·s-1, and the resulting varying force and cantilever height were recorded over time. At the same time, differential interference contrast (DIC) and fluorescence images at the confined cell's midplane were recorded every 5 seconds using a 63× water immersion objective. All microscopy equipment was placed, and experiments were carried out in a custom-made isolation box at 37 °C (The Cube; Life Imaging Services).<br> The following pharmacological inhibitors and chemical compounds were used:10 µM ROCK-mediated contractility inhibitor <b>Y-27632</b> (Y27) (ED Millipore),20 µM <b>AACOCF3</b> (AA) inhibiting the nuclear envelope stretch-sensitive enzyme cPLA2 (Tocris Bioscience),100 µM <b>2APB</b> blocking stretch-activated inositol triphosphate receptors (InsP3Rs) on the ER/nuclear membranes (Tocris Bioscience).

本数据集包含HeLa Kyoto MYH9-eGFP/lifeact-mCherry细胞在3μm约束条件下的原子力显微镜（AFM）与共聚焦成像相关实验数据。
将铺有细胞的培养皿安装至培养皿加热器（JPK Instruments）中，维持37℃恒温环境，并置于搭载共聚焦成像单元（LSM 700；蔡司（Zeiss））与原子力显微镜（AFM）探头（CellHesion 200；JPK Instruments）的倒置光学显微镜（Axio Observer.Z1；蔡司（Zeiss））系统中。聚焦离子束（Focused Ion Beam, FIB）刻蚀制备的扁平硅微悬臂梁，按照Cattin等（2015）的报道方法完成加工与校准。将微悬臂梁固定于标准JPK玻璃基座后，安装至AFM探头内。以0.5 μm·s⁻¹的恒定速率将悬臂梁降至细胞上方的预设高度，实时记录随时间变化的作用力与悬臂梁位移。与此同时，采用63×水浸物镜，每5秒采集一次受约束细胞中部平面的微分干涉差（DIC）与荧光图像。所有显微成像设备均安置于定制恒温隔离箱（The Cube；Life Imaging Services）内，实验全程维持37℃环境。
本实验使用如下药理学抑制剂与化学试剂：10 μM Rho相关卷曲螺旋蛋白激酶（ROCK）介导的收缩力抑制剂Y-27632（Y27，购自EMD Millipore）；20 μM AACOCF3（AA），可抑制核被膜牵张敏感性酶胞质磷脂酶A2（cytosolic phospholipase A2, cPLA2），购自Tocris Bioscience；100 μM 2APB，可阻断内质网（ER）与核膜上的牵张激活型肌醇三磷酸受体（InsP3Rs），购自Tocris Bioscience。