- Title
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Rescue of hematopoietic stem/progenitor cells formation in plcg1 zebrafish mutant
- Authors
- Ferri-Lagneau, K.F., Haider, J., Sang, S., Leung, T.
- Source
- Full text @ Sci. Rep.
Ginger rescues the dorsal aorta (DA) and the formation of hematopoietic stem/progenitor cells (HSPCs) in plcg1−/−embryos. (A) Real-time imaging of the vasculature in tg(fli1:GFP) embryos at 30 hpf. Red rectangle shows the location of ISV. (B) In situ hybridization of the DA marker ephrin-B2a (efnb2a) at 1dpf (32 hpf). Black arrow indicates the artery, red arrow shows absence of artery in mutant fish. (C) In situ hybridization of the HSPC marker myb at 1 (32 hpf) vs 2dpf (54hpf). Black arrow points to myb expression in hemogenic endothelium (1 dpf) and CHT region (2 dpf), red arrow indicates absence of expression in mutant fish. (D) In situ hybridization of notch3 (normally expressed in the DA at 1 dpf (32 hpf)) and bmp7a (shown here for the first time to be expressed in the DA from 1(32 hpf) to 2 dpf (48 hpf)). These marker expressions are lost in plcg1−/− embryos and rescued following exposure to ginger. Red arrow points to DA. Scale bars = 250 μm. |
Signaling pathways involved in the rescue of arteriogenesis by ginger in plcg1−/− embryos. (A) Pharmacological inhibition of Bmp (DMH-1) and Notch (LY411575) signaling pathways affect the rescue of dorsal aorta, analyzed by efnb2a in situ at 1 dpf. (B) Representative images of different treatment. White arrow indicates the dorsal aorta, underneath is the vein indicated by red arrow. Scale bars = 250 μm. |
Signaling pathways involved in the rescue of definitive hematopoiesis by ginger in plcg1−/− embryos. (A) Pharmacological inhibition of Bmp (DMH-1) and Notch (LY411575) signaling pathways affect the rescued HSPCs, analyzed by myb in situ at 2 dpf. (B) Representative images of different treatment. Encircled area indicates mybexpression in the CHT region, whereas red arrows indicate the CHT region only. Scale bars = 250 μm. |
Ginger expands the Caudal Vein Plexus (CVP). (A) Ginger increases or rescues the CVP sinusoids in tg(fli1:GFP) WT siblings or plcg1−/− mutant embryos respectively (2 dpf). Yellow arrows show enlarged CVP with big sinusoid. White arrow indicates dorsal aorta and pink arrow cardinal vein. (B) Bmp is necessary for the effect of ginger on the CVP in tg(fli1:GFP) WT siblings and plcg1−/− mutant embryos (2 dpf). Vertical red bar represents the width of CVP. (C) Inhibition of Notch compromised the effect of ginger on the developing CVP at the sinusoidal level (1 dpf). Yellow arrows show enlarged CVP with big sinusoid. (D) runx1 and scl are ectopically expressed in the CVP/CHT area following ginger treatment. (E,F) Bmp7a/bmp2b expression (E) and notch3 expression (F), are highly promoted by ginger in the CVP/CHT of WT and plcg1−/−mutant embryos. Red arrow indicates high expression in CVP/CHT (E,F). Pictures are the representative images of ≥30 embryos per condition. Red box indicates the CHT region. Scale bars = 250 μm. |
Ginger increases the level of nitric oxide (NO) in non-transgenic WT and plcg1−/− mutant embryos. (A) Effect of ginger on NO staining at 1 dpf WT and plcg1−/− embryos. (B,C) Effect of NO donor (SNAP) and 2 Nos inhibitors (L-NAME, L-NMMA) on 2 dpf WT (B) and plcg1−/− embryos (C). Red arrows show ginger effect in CVP region. The effect of the NO donor SNAP is shown for comparison. Non-transgenic WT and mutant embryos are used to see the NO fluorescent dye. Pericardial edema, a typical phenotype occurred in plcg1−/− mutants at 2 dpf (C). Pictures are the representative images of ≥30 embryos per condition. Scale bars = 250 μm. |
Effects of nitric oxide on arterial and hematopoietic gene marker expression. (A) Vasculature visualized by tg(fli1:GFP)at 1 dpf and 2 dpf embryos. (B–D) The graphical results are shown in 1 dpf (B) and 2 dpf (C,D) embryos. NO synthase inhibitors L-NAME and L-NMMA, NO donor SNAP, and NO scavenger c-PTIO. White arrow indicates dorsal aorta and pink arrow indicates cardinal vein. Scale bars = 250 μm. |