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Pazhakh et al., 2017 - A GCSFR/CSF3R zebrafish mutant models the persistent basal neutrophil deficiency of severe congenital neutropenia. Scientific Reports   7:44455 Full text @ Sci. Rep.

Fig. 2

Neutrophil deficiency in csf3r null F2 embryos.

(ad) Fluorescence micrographs of 3 dpf Tg(mpx:EGFP) embryos showing that several representative csf3r mutant null allelotypes (bd) have substantially reduced numbers of fluorescent neutrophils compared to WT (a). Panel b is a representative homozygous csf3r3/3 null embryo, and panels c-d show representative compound heterozygous null embryos of the two allelotypes shown. (e) Quantification over 2–5 days post-fertilisation (dpf) of tail region neutrophil numbers in embryos of control (WT) matings and incrosses of 3 different pairs of csf3r null parents of the csf3r allelotypes shown. Data are mean ± SD. Within-day comparisons across genotypes are analysed by unpaired two-tailed t-tests, pooling data from the mutant compound heterozygous allelotypes shown, which are not significantly different to each other. These parental allelotype combinations were randomly selected, being those pairs that laid of those that were set up. They are not intended to be comprehensive, but they do demonstrate a consistent non-complementing neutrophil-depletion phenotype encompassing five embryonic allelotypes (1/2, 1/3, 2/2, 2/3 and 3/3). (f) Embryos carrying single csf3r null alleles have an intermediate neutrophil deficiency. A mix of heterozygous csf3rwt/(1or2) embryos in a 1:1 ratio were generated by outcrossing a parent of genotype csf3r1/2 to WT. Their neutrophil numbers at 3 dpf are compared with the non-contemporaneous csf3rWT/WT and pooled csf3r−/− mutant 3 dpf groups of panel (e). The mutant data were pooled as none of the mutant allelotype groups is significantly different to any other. p-values are from one-way ANOVA with Tukey’s multiple comparisons test. p < 0.0001. (g) csf3r null embryos have equivalent survival to WT embryos up to 5 days post-fertilisation (dpf). Kaplan-Meier plots compared by Wilcoxon rank sum test.

Fig. 3

Normal macrophage numbers in csf3r null F2 embryos.

(a) Photomicrographs of representative 3 dpf WT and csf3r null embryos stained by whole mount in situ hybridisation (WISH) for the macrophage-specific marker csf1ra/cfms. (b) Quantification of torso-located macrophage numbers in csf1ra/cfms WISH embryos in shows no difference between genotypes (the torso being the region distal to the dotted red line in (a), selected for scoring for its lower density of macrophages and lower incidence of overlapping cells).

Fig. 4

Neutrophil deficiency in csf3r null adults.

(a,b) Fluorescent photomicrographs of 5 month old adult Tg(mpx:EGFP) zebrafish tail fins showing reduced numbers of EGFP-positive neutrophils in csf3r null (b) vs WT (a) animals. (c) Quantification of tail fin neutrophil density. n = 4–5 animals/genotype as shown.

EXPRESSION / LABELING:
Gene:
Fish:
Knockdown Reagent:
Anatomical Term:
Stage: Adult
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage: Adult

Fig. 5 ZFIN is incorporating published figure images and captions as part of an ongoing project. Figures from some publications have not yet been curated, or are not available for display because of copyright restrictions.

Acknowledgments:
ZFIN wishes to thank the journal Scientific Reports for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Sci. Rep.