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Fig. 2
Endocardium-derived cells contribute to transient fibrosis during zebrafish heart regeneration. (A) Analysis of the transcriptome of kdrl:mCherry+ cells FACS-sorted from the ventricular apex of hearts with no injury or at 7 dpi was performed. Volcano plot. Light blue, false discovery rate (FDR) >0.05, abs[log fold change (LFC)] <1; orange, FDR >0.05, abs(LFC) >1; green, FDR <0.05, abs(LFC) >1. (B) Ingenuity pathway analysis including the number of differentially expressed genes. (C–F) Immunofluorescence with anti-GFP and anti-mCherry of a heart section from an adult fli1a:GFP;col1a2:mCherry-NTR zebrafish. Yellow arrowheads mark double-positive cells at the injury area (IA; asterisk). Green arrowheads mark fli1a:GFP+ cells negative for mCherry. (G and H) FACS-sorted cells from fli1a:GFP;col1a2:mCherry hearts without injury or at 7 dpi. Shown are two representative examples from a total of four hearts per condition analyzed. (I) The percentage of fli1a:GFP+ cells that express more than 100 a.u. of mCherry, with the threshold corresponding to the maximum value of mCherry detected in endocardial cells from uninjured hearts. (J) The percentage of col1a2:mCherry+;fli1a:GFP+ cells according to the thresholds in H. Graphs show individual measurements from hearts as well as mean ± SD values. *P = 0.0363 by two-tailed t test. (K) Experimental scheme for visualizing collagen-producing endocardial cells. (L–N) Immunostaining of a heart section close to the IA. M is a zoomed-in view of L. (N and O) Zoomed-in views of M. mCherry marks fli1a-derived cells expressing col1a2, myosin heavy chain (MHC) marks the myocardium, and nuclei are DAPI-counterstained. [Scale bars, 25 µm (D, M, and N) and 100 µm (C and L).]