Résumé
Tg(mpeg1:gal4/UAS:Kaede) larvae with mosaic expression of Kaede protein in macrophages were amputated at 3 dpf and imaged using high resolution Spinning Disk microscopy between 8 to 13 h,every 2.5 min (C) Quantification of perimeter (upper graph) and circularity value (lower graph) for individual macrophages present at the wound, during a time lapse sequence from 8 hpA to 13 hpA every 2.5 min. For quantification, we used automated image analysis, the image segmentation was carried out from 8 to 13 hpA. At first, the images were filtered by using space-time filtering that keeps the temporal coherence of moving macrophages (Sarti et al., 1999). Then, the filtered images were segmented by a combination of the local Otsu method (Otsu, 1979; Saddami et al., 2019) and (Park et al., unpublished) and the subjective surface segmentation (SUBSURF) method (Sarti et al., 2000) where the binarized images from the local Otsu method were considered as an initial condition of the SUBSURF equation (Park et al., 2023). Trajectories of macrophages in segmented images were reconstructed based on automatic cell tracking (Park et al., 2023), and shape descriptors (perimeter and circularity) were measured. Refernces: Sipka T, Park SA, Ozbilgic R, Balas L, Durand T, Mikula K, Lutfalla G, Nguyen-Chi M. (2022). Macrophages undergo a behavioural switch during wound healing in zebrafish. Free Radical Biology and Medicine 192:200–212. doi:10.1016/j.freeradbiomed.2022.09.021 and Park SA, Sipka T, Kriva Z, Lutfalla G, Nguyen-Chi M, Mikula K. (2023). Segmentation-based tracking of macrophages in 2D+time microscopy movies inside a living animal. Computers in Biology and Medicine 153, 106499. doi: 10.1016/j.compbiomed.2022.106499