Fig. 4.

flt1 deletion limits myofibroblast differentiation. (A) Immunostaining for EGFP (green), Aldh1a2 (magenta) and α-SMA (myofibroblasts, red) with DAPI (blue) counterstaining on representative sections of cryoinjured egr3>EGFP ventricles at 7 dpci. White and orange arrowheads indicate the EGFP⁺Aldh1a2⁺ EdCs and α-SMA⁺ cells, respectively. (B) Proportion of EGFP⁺ EdCs within the wound of cryoinjured egr3>EGFP ventricles (n=3) at 7 dpci (see Table S4 for raw data). (C) Immunostaining for GFP (yellow), Aldh1a2 (cyan) and Vim (fibroblasts/myofibroblasts, magenta) with DAPI (blue) counterstaining on representative sections of cryoinjured egr3>EGFP ventricles at 7 dpci. Arrowheads indicate the EGFP⁺Aldha1a⁺Vim⁺ EdCs. (D-D″,F,F′) Immunostaining for MHC (magenta) and α-SMA (white) on sections of cryoinjured ventricles from Ctrl [n=5 for egr3 OE, n=6 for Tg(hsp70l:Cre);egr3^flox/flox, D], egr3 OE (n=5, D′) and Tg(hsp70l:Cre);egr3^flox/flox (n=5, D″) zebrafish, and from flt1^+/+ (n=5, F) and flt1^−/− (n=5, F′) zebrafish at 7 dpci. (E,G) Quantification of α-SMA⁺ cell number within the wound of the indicated genotypes at 7 dpci. (H,H′) Immunostaining for Aldh1a2 (magenta) and α-SMA (yellow) with DAPI (cyan) counterstaining on sections of cryoinjured flt1^+/+ (n=4, H) and flt1^−/− (n=4, H′) ventricles at 7 dpci. Arrowheads indicate endocardial-associated α-SMA⁺ cells. (I) Quantification of endocardial-associated α-SMA⁺ cell number within the wound on ventricular sections of the indicated genotypes at 7 dpci. White and orange dotted lines indicate the injured area. Areas outlined are shown at higher magnification on the right in A,C,H and H′. Scale bars: 100 µm. Data are mean±s.e.m. (two-tailed, unpaired Student's t-test with P values shown in the graphs).