The development of anisotropic behaviours of 3T3 fibroblasts on microgrooved patterns

3T3 fibroblasts cultured on microgrooved polydimethylsiloxane (PDMS) surfaces of two different widths (25 μm and 55 μm) were individually tracked using confocal microscopy with a novel live-cell staining technique over several hours without noticeable cytotoxic effects. By quantifying the cell morphology, orientation, and migration over time, we identified the timescale (about 2-4 h after seeding) over which cell behaviours transitioned from isotropy to anisotropy, where the preference is in the direction parallel to the pattern. The development of anisotropy occurred more rapidly and distinctly when a narrower ridge width was used, suggesting that it is the ridge width that imposed a physical barrier on the cells' morphology and motility. Furthermore, while we found a weak but statistically significant correlation between cell orientation and morphology on the single-cell level, there is a lack of correlation on the same level between cell orientation and migratory direction. This suggests that while morphology and migration are affected anisotropically by topographical patterns in a similar way, the underlying processes giving rise to the anisotropy is slightly different in the two cases.