Quantitative observations on cytoskeleton changes of osteocytes at different cell parts using digital holographic microscopy.

Cytoskeletons such as F-actin have different distributions in different cell parts and they are the cause of different degrees of cell collapse when the F-actin is disrupted. It is challenging to use conventional methods such as fluorescence microscopy and atomic force microscopy to conduct real-time and three-dimensional observations on the dynamic processes at different cell parts due to the slow measuring speed and the need for live-cell staining. In this study, the morphological variations of different bone cell parts caused by F-actin disruption are dynamically measured by using digital holographic microscopy (DHM). We separately analyze local parameters (cell height and cell width) and global parameters (cell projected area and cell volume) of cells to address variations of specific cell areas and quantify the changing process of the whole cell. We found significant differences in temporal variations of both local and global cell parameters between the cell body and cell process, which is consistent with the qualitative observation by fluorescence staining. Our study not only validates the unique ability of DHM to simultaneously investigate the dynamic process at different cell parts, but also provides sufficient experimental bases for exploring the mechanism for F-actin disruption.