The number of neutrophils recruited to the wounded area in the tlr2 and myd88 mutants and wild type sibling controls larvae. (A) Experimental scheme. Tlr2 and myd88 homozygous mutants and sibling control larvae were wounded at 3 dpf. Their tails were wounded to the tip of the notochord. The red dashed line shows the site of wounding. Recruited neutrophils at the wound were imaged at 1, 2, 4, and 6 hpw by using CLSM. For recruited cell counting analysis, cells localized within an area of 200 μm from the wounding edge toward the body trunk were counted as recruited cells. The red dashed box shows the area where neutrophils were counted as recruited neutrophils. (B,D) Representative images of 3 days dpf larvae at 1, 2, 4, and 6 h post-wounding (hpw). Scale bar: 50 μm. (C) Quantification of recruited neutrophil numbers to the wounded area at 1, 2, 4, and 6 hpw in 3 dpf tlr2+/+ and tlr2– /– larvae. Each point represents a different larva. Sample size (n): 45, 46, 82, 72, 74, 68, 50, 50. (E) Quantification of recruited neutrophil numbers to the wounded area at 1, 2, 4, and 6 hpw in 3 dpf myd88+/+ and myd88– /– larvae. Each point represents a different larva. Sample size (n): 29, 28, 37, 38, 45, 39, 51, 45. In all cases, statistical analyses were done from three independent experiments. An unpaired, two-tailed t-test was used to assess significance (ns, no significant difference, *P < 0.05, **P < 0.01, ***P < 0.001) and data are shown as mean ± SD.
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