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.
Steps to reproduce
One has to elaborate on the XZ-views (side views). We used ImageJ macros. 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 between the cell layers, translating into the strain values by reference to the initial separation distance.