Fig. 4: RPE polarization is associated with nuclear changes.

A Representative 3D reconstructions of iRPE cell lines expressing fluorescently-tagged FBL (nucleolus) and LMNB1 (nuclear envelope). Images from W1 constitute the baseline, while images at W4 show the endpoint for cells that were grown in the presence of PGE2 or HPI4 (magenta = phalloidin, cyan = Hoechst, yellow = GFP-tagged protein; scale bar = 5 μm). Note that the signal from phalloidin was stripped from the top of the cells in these images to better show the reconstruction of the GFP-tagged protein. B Panel of maximum intensity projected fluorescence images displaying wrinkling of the nuclear envelope occurring with polarization (scale bar = 10 μm). C Raincloud plots indicating the changes over time and between treatments of FBL for count per cell, X/Y distribution, and Z distribution (the points indicate the average values for each FOV; the diamonds represent the median, and the error bars indicate the 5th and 95th percentiles). Two-sided Welch’s t-test was performed to compare PGE2- and HPI4-treated cells at each timepoint. The solid lines indicate the generalized linear model maximum likelihood fit for the two treatments with week and the square of week as dependent variables. The tables at the bottom display the results of Tukey’s HSD pairwise comparison (N = 30; ***P < 0.005, **P < 0.01, *P < 0.05).