Fig. 4

Olaparib and IR can have modulating effects on the tumor vascular network. All cell lines were fluorescently labeled with CM-DiI (in red) and injected in the perivitelline space (PVS) of 2dpf Tg(fli1:eGFP)zebrafish embryos. At 24 hpi, zebrafish xenografts were screened and randomly distributed into the different experimental conditions: control, olaparib, IR, IR + olaparib. Xenografts were treated for 4 consecutive days, fixed at 5dpi and imaged by confocal microscopy for analysis of vessel density (A–T). The quantification of vessel density is represented by percentage of tumor area occupied by vessels (in green) (U). To analyze vessel functionality, TNBC cell line Hs578T was fluorescently labeled with CellTracker™ Deep Red Dye (in red-false color) and injected in the PVS of 2dpf Tg(gata1:RFP;fli1:eGFP). Hs578T xenografts were screened and randomly distributed in the different experimental conditions: control, olaparib, IR and IR + olaparib. Representative images of 5dpi Hs578T xenografts (V,V’). The presence of erythrocytes (in red) inside the tumor-associated vessels (in green) was scored as: absence of erythrocytes = no perfusion (W,W’) or presence of erythrocytes = with perfusion (X,X’) and quantified (Z). All images are anterior to the left, posterior to right, dorsal up, and ventral down (as depicted in the scheme on top left). The dashed lines delineate the tumor area. White arrowheads indicate erythrocytes inside the vasculature. Scale bar: 50 µm. Results are from three independent experiments and expressed as mean ± SEM, each dot represents one xenograft. The total number of xenografts analyzed is indicated in the images. Statistical results: ns > 0.05, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001.