Fig. 5

ncam1a-E4 directs injury-induced epicardial gene expression. (A) Larval expression of ncam1a-E4:EGFP lines. Scale bar: 500 µm. (B) Whole-mount image of a 6 dpf heart with tcf21:H2A-mCherry (red) labeling the epicardial cells. No EGFP expression was observed. V, ventricle. Scale bar: 20 µm. (C) Whole-mount images (maximum projections) of the ventricular surface showing expression of a ncam1a-E4:EGFP line in uninjured (Ctrl) and 3 dpa samples. Anti-EGFP antibody staining was used to detect the EGFP expression. Scale bar: 100 μm. (D) Top: the ncam1a-E4E4:EGFP reporter construct. Bottom: larval expression of ncam1a-E4E4:EGFP reporter line. Scale bar: 500 µm. (E) Optical section image of a 6 dpf heart (whole-mounted) showing EGFP expression in the muscle. tcf21:H2A-mCherry (red) labels the epicardial cells. V, ventricle. Scale bar: 20 µm. (F) Whole-mount images (maximum projections) of the ventricular surface showing expressions of ncam1a-E4E4:EGFP line 1 (green) in uninjured (Ctrl) and 3, 7 and 30 dpa samples. tcf21:H2A-mCherry (magenta) labels the epicardial cells. White dashed lines indicate the injury sites. The outlined regions are enlarged in G. Scale bar: 100 µm. (G) Single optical section of confocal images from the regions outlined in F. Single-channel images are shown in grayscale. Arrows indicate representative GFP+mCherry+ cells. Scale bar: 100 µm. (H) Section images demonstrating expression of ncam1a-E4E4:EGFP (green) in tcf21:H2A-mCherry+ (magenta) cells. Magnified views of the outlined regions are shown on the right (distal and wound regions). White dashed lines indicate the injury sites. Arrows indicate representative double-positive cells. Scale bar: 100 µm. (I,J) HCR staining of ncam1a (magenta dots) on ventricular sections (ctrl, 3 dpa) showing ncam1a-E4E4:EGFP expression (green) in ncam1a+ cells both in the wound (I) and distal (J) regions. The outlined regions are enlarged on the right. Arrows indicated EGFP+ncam1a+ cells. Scale bars: 50 µm.