Fig. 2

Endocardial proliferation and endothelial addition accompany OFT endocardial growth. (A-D) Lateral slices through the OFT at 51 hpf (A-C) show incorporation of EdU (red) into the Tg(fli1a:negfp)-labeled (green) endocardium between 36 and 51 hpf, with the myocardium marked by immunofluorescence for Alcama (gray). Quantification (D) indicates an OFT endocardial proliferation index (PI) of 25.8±1.2% during this period of OFT expansion (n=6; N=1). (E-H) Lateral views of three-dimensional reconstructions at 51 hpf (E-G) show incorporation of BrdU (red) into the Tg(fli1a:negfp)-labeled (green) endocardium between 36 and 51 hpf. Quantification (H) indicates an OFT endocardial PI of 28.9±4.2% (n=14; N=2). Data are mean+s.e.m. (I,J) Cartoons illustrate the schemes for photoconversion experiments designed to assess the contribution of external sources (I) or endothelial cells from the AA1 (J) to the OFT endocardium. V, ventricle. (K-P) Three-dimensional reconstructions depict expression of Tg(kdrl:dendra), which drives the green-to-red photoconvertible protein Dendra in all endothelial cells. Photoconversion of the OFT endocardium (K-M) at 36 hpf was followed by detection of green cells, which had not been labeled by photoconversion, in the OFT endocardium at 51 hpf (N-P), indicating the contribution of cells from external sources to the OFT endocardium during this time period (n=3; N=2). Arrowheads indicate examples of cells that were not labeled by photoconversion (green only) and arrows indicate examples of cells that were labeled by photoconversion (red and green). (Q-V) Photoconversion of a portion of AA1 endothelium (red) at 36 hpf (Q-S) was followed by detection of labeled cells in the OFT at 51 hpf (T-V) (n=5; N=2). Scale bars: 5 μm.