Fig 4

Fig 4

(A) Snapshots from time-lapse imaging of pdgfrb^NTR-mCherry; col1a2:GFP embryos from 54 to 73 hpf. Newly differentiated pericytes were retrospectively traced to identify their cell of origin. The ISV is outlined by dotted lines at the first time point. One perivascular fibroblast (green, arrows) traced can be seen gradually upregulating pdgfrb^NTR-mCherry expression and extending pericyte-like cellular processes (notched arrowheads). The time stamps are indicated in the hh:mm format. Schematic drawings of the merged images at each time point are shown at the bottom. The corresponding time-lapse movie is shown in S2 Video. n = 7 embryos. (B) Schematic showing experimental timeline of perivascular fibroblast ablation. col1a2^NTR-mCherry; pdgfrb:GFP embryos were incubated in either water or metronidazole (MTZ) from 25 to 54 hpf following which embryos were washed at 54 hpf and imaged at 4 dpf to visualize pericytes. (C) Representative images of water (left) or MTZ (right) treated col1a2^NTR-mCherry; pdgfrb:GFP embryos at 4 dpf. Compared to water-treated control embryos, MTZ-treated embryos showed complete ablation of mCherry⁺ cells with only residual mCherry⁺ debris (notched arrowheads). Fewer pdgfrb:GFP^high pericytes (arrowheads) can be seen in MTZ-treated embryos compared to water-treated controls. (D) Quantification of pericyte number after perivascular fibroblast ablation along aISVs and vISVs. Pericytes were identified based on high level expression of the pdgfrb:GFP reporter as shown in (C) and S3A and S3C Fig. Arterial versus venous ISVs were determined based on direction of the blood flow in each vessel. From each embryo, 8–10 ISVs in the mid-trunk region were imaged and scored. The average number of pericytes on aISVs, vISVs, or all ISVs were plotted in the graph with each data point representing one individual embryo. Pericytes were lost along both aISVs and vISVs after MTZ treatment. n = 10 (water) and 13 (MTZ) embryos. Data are plotted as mean ± SEM. Statistics: Mann-Whitney U test. Asterisk representation: p-value < 0.0001 (****). Scale bars: (A) 10 μm; (C) 50 μm.