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Fig. 8. Inhibition of contractility during keratinocyte polarization perturbs the acquisition of maximum height at lateral domains. E-cadherin (red) and phalloidin (green) staining was performed after 1 hour of cell-cell contact formation in the absence (control) or presence of blebbistatin (blebbistatin). Confocal sections (0.3 µm) were collected at different levels through a keratinocyte monolayer and processed as follows. (A) Three dimensional reconstruction. After blebbistatin treatment, F-actin is diffusely localized and cadherin staining is found as a wavy line between neighbouring cells. Small ruffles/projections are seen at the cell surface and on top of junctions (arrows). (B) Confocal sections taken from basal to top of the monolayer at 0, 1.5, 3, 4.5 and 6 µm. Arrow points to small ruffles/projections sees at the top of junctions. Arrowhead shows cadherin staining in controls whereas no similar linear cadherin staining is observed in blebbistatin-treated cells. (C) Quantification of the maximum height of the lateral domains (basal to apical distance). Confocal sections showing linear cadherin staining were quantified and values expressed relative to control, untreated cells polarized for 1 hour (see Materials and Methods for details). Maximum height seen in lateral domain of control cells is 4.5 µm compared with 3 µm in blebbistatin-treated cells and 2 µm in cells maintained in low calcium (Braga et al., 1998). After blebbistatin treatment, lateral domain height is decreased by 35% (*P<0.001). Results represent the mean of seven different Z-series collected from two independent experiments.
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