Neutrophil recruitment and motility is hindered in the miR-722-overexpressing zebrafish line. (A) A schematic of the Tol2-lyzC:miR-722/Dendra2 plasmid, injected into wild-type AB zebrafish embryos to generate the transgenic line Tg(lyzC:miR-722/Dendra2)^pu6 (miR-722). Tg(lyzC:Dendra2)^pu7 was generated using the same configuration without the miR-722 insertion (vector). All experiments were performed with F2 larvae at 3 dpf. (B) Relative expression level of precursor and mature miR-722, miR-223 and let-7e (normalized to U6 expression) in vector and miR-722 lines determined by RT-qPCR. Data are means±s.d. (N=3 biological repeats with 10 larvae at each time point in each group). P values were calculated with unpaired Student's t-test. (C) Representative images and quantification of total neutrophils in vector and miR-722 lines. One representative result of three independent experiments is shown (n=20). Scale bar: 500 µm. (D) Representative images and quantification of neutrophil recruitment to localized ear infection in vector or miR-722 larvae at 1 hpi. The infected ear is indicated with the circle. One representative result of three independent experiments is shown (n=20). P value was calculated with unpaired Student's t-test. Scale bar: 100 µm. (E) Representative images and quantification of neutrophil recruitment to tail transection site in vector or miR-722 larvae at 1 h post-injury. Neutrophils in the boxed region were quantified. One representative result of three independent experiments is shown (n=20). P value was calculated with unpaired Student's t-test. Scale bar: 100 µm. (F) Tracks and quantification of neutrophil motility in vector or miR-722 larvae. Results were pooled from three independent larvae (n=60). P value was calculated with unpaired Student's t-test. Scale bar: 50 µm. (G-I) Quantification of total number of neutrophils (G), neutrophils recruited to the ear 1 h post P. aeruginosa infection (H) and to the wound 1 h post tail transection (I). One representative result of three independent experiments is shown (n=20). No statistical difference among the results from separate founders were observed with unpaired one-way ANOVA.

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.

Overexpression of rac2 rescues neutrophil recruitment in miR-722-expressing larvae. (A) Tg(lyzC:miR-722/Dendra2)^pu6 (miR-722) was crossed with Tg(mpx:mCherry-2A-Rac2) (Rac2) and the offspring were separated into four groups. For control, Tg(lyzC:miR-722/Dendra2)^pu6 was crossed with Tg(mpx:mCherry) (mCherry). All experiments were performed with F2 larvae at 3 dpf. (B) Tracks and quantification of neutrophil motility in indicated lines. Results were pooled from three independent larvae. P values were calculated with unpaired Student's t-test. Scale bar: 100 µm. (C,D) Quantification of neutrophil recruitment to tail wounding (C) or localized ear infection (D) in siblings separated into the four groups as depicted in A. One representative experiment of three independent biological repeats is shown (n=20 for each group). P values were calculated with unpaired two-way ANOVA.

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.

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.