FIGURE 3.

Expression of Cx43 significantly reduces total cell number and cell proliferation. A, 6-day growth curves revealed that Cx43-GFP-expressing melanomas displayed significantly reduced total cell number as early as day 4 (n = 8; *, p < 0.05), which was further reduced on days 5 and 6 (n = 8; ***, p < 0.001). Cx26-GFP-expressing melanomas displayed a slight reduction in total cell number only on day 6 (n = 8, *, p < 0.05). B, proliferation at day 4 was assessed using an EdU assay, which labels nuclei passing through the S phase of the cell cycle, where the proportion of EdU-labeled nuclei (green) to total nuclei labeled with Hoescht (blue) was determined. Cells were counterstained with an anti-GFP antibody (red) because the normal fluorescence of GFP-tagged Cx43 was quenched by the EdU signal. C, expression of Cx43 significantly reduced the percentage of EdU-positive cells by ∼25% in comparison to the controls (n = 40; **, p < 0.01). Bar, 20 μm.