In vitro cell on cell dry eye model incorporating normal stress control

We aim to improve the mechanical understanding of dry eye disease (DED). The data are laser scanning confocal images (raw data) of stratified layers of corneal cells (cornea) in contact with stratified layers of conjunctival cells (eyelid) in situ. The nuclei have been stained with Hoechst (blue), the plasma membrane with CellMask Deep Red(red), and the green dots are a fluorescent coating on the cell culture surfaces. This geometry is an in-vitro dry eye model. The long-term goal of this project is to incorporate different normal stresses into this dry eye model. In the project's first step (this dataset), we compare the response to uniaxial stresses of healthy and dry eye diseased cell layers. We expect marked differences in normal stress response between healthy and dry eyes. The change in distances between the cell layers (represented by a fluorescent coating on the cell culture dish) is interpreted as the strain acting on the cell layer. Compression and tensile moduli of healthy and dry eyes can hence be extracted from these images by elaborating XZ-views. The intensity profiles of the green fluorescence channel (fluorescent coating on the cell culture surface) are fitted with Gaussians. The peak-to-peak distance corresponds to the separation distance between the cell layers. We assess the collective behavior of an entire cell layer in-situ. Methodologically it is different from single-cell techniques (like AFM). We observe compression moduli on the order of 5 kPa. We find a non-significant change in the compression modulus of the dry eye cell layers. However, there is embrittlement in dry eye cell layers upon decompression. Healthy cell layers with intact mucins show strong ductility and viscoelasticity, and their original states are not recovered upon decompression.本研究旨在加深对干眼症（dry eye disease, DED）的力学认知。本数据集包含共聚焦激光扫描显微原始图像，拍摄对象为原位状态下，与复层结膜细胞层（眼睑）接触的复层角膜细胞层（角膜）。其中，细胞核经Hoechst染色后呈蓝色，细胞膜经CellMask Deep Red染色后呈红色，图像中的绿色亮点为细胞培养表面的荧光涂层。该体外培养体系即为干眼症体外模型。本项目的长期目标是在该干眼症模型中引入不同的法向应力。本项目的第一步（即本数据集对应的研究内容）中，我们对比了健康组与干眼症组细胞层对单轴应力的响应，并预期两组在法向应力响应方面存在显著差异。细胞层之间的间距变化（以细胞培养皿上的荧光涂层为标识）被视为作用于细胞层的应变。因此，通过重构XZ切面视图，可从这些图像中提取健康组与干眼症组细胞层的压缩模量与拉伸模量。绿色荧光通道（对应细胞培养表面的荧光涂层）的强度分布可通过高斯函数进行拟合，其峰间距即为细胞层之间的分隔距离。本研究可评估原位状态下整个细胞层的集体行为，从方法学角度而言，该技术与单细胞技术（如原子力显微镜，Atomic Force Microscopy，缩写AFM）存在差异。我们测得的压缩模量约为5千帕（kPa），且发现干眼症组细胞层的压缩模量无显著变化。但在卸压过程中，干眼症组细胞层出现了脆化现象；而带有完整黏蛋白的健康细胞层则表现出优异的延展性与黏弹性，且在卸压后无法恢复至初始状态。