Figure 4.

tnfα+ macrophages respond to cardiac injury and LPS treatment affects Collagen I scarring and results in changes in macrophage phenotype. (A and B) Representative images and quantification of whole hearts from TgBAC(tnfα:GFP) fish at different time-points post-injury. The dashed line in A demarks the extent of the injured region. L-plastin, mpx, and mpeg1 data from Figure 2C and F (greys) are provided as a reference (B). (C) Representative image of a heart from a Tg(mpeg1:mCherry); TgBAC(tnfα:GFP) double transgenic fish at 7 dpi (higher magnification shown in F). The boxed region demarks the approximate area imaged in D–F and all subsequent ventricular apex images. (D–F) Representative images of the ventricular apex of unwounded Tg(mpeg1:mCherry); TgBAC(tnfα: GFP) fish (D) and the injury area at 3 (E) and 7 dpi (F). Very few tnfα+ mpeg1+ cells are observed in unwounded hearts (D). (D′–F′) For each image, single channel panels and the merge of the boxed region is provided. (G) Quantification of the percentage of mpeg1+ macrophages at the injury site expressing tnfα at 3, 7, and 14 dpi with and without LPS treatment. (H and I) Representative images of the injury site of Tg(mpeg1:mCherry); TgBAC(tnfα:GFP) fish (H) and following LPS treatment (I) at 14 dpi. (J) Quantification of the Collagen I fold change as compared to unwounded fish in control and LPS treated fish. (K–L) Representative images of sections through the injury site of control (K) and LPS treated fish at 14 dpi (L). Quantification in B and G; statistical analysis by Kruskal–Wallis/Dunn’s multiple comparisons tests. n numbers for mpx+, mpeg1+, and L-plastin+ are shown in Figure 2. Quantification in J; statistical analysis by Mann–Whitney tests at each time-point. un-w, unwounded. Scale bars: A, C, K, L = 250 μm; D–F, H, I = 50 μm; inset in A, D′–F′ = 20 μm.