Fig. 7

Supplement of metabolites that are able to produce acetyl-CoA significantly recovered the neutrophil numbers and migration toward wound in tdh−/− embryos. (A, B) Whole-mount in situ hybridization (WISH) analysis of lyz mRNA in CHT of WT and tdh−/− embryos treated with different metabolites at 3 dpf. A close-up view of boxed regions is shown in the lower panel. (C) The quantification analysis of the lyz+ cell numbers in CHT in (A). Note that only supplementation with Pyr largely recovered the number of neutrophils. (D) The quantification analysis of the lyz+ cell numbers in CHT in (B). Note that only supplementation with nutrients that are able to produce energy largely recovered the number of neutrophils. (E) Brightfield and fluorescent microscopy images of the wound site of 3 dpf WT//Tg (lyz:EGFP) and tdh−/−//Tg (lyz:EGFP) transgenic zebrafish embryos treated with different metabolites at 6 h post transection. (F, G) Quantification analysis of numbers of EGFP+ cell (F) and percentages of EGFP+ cells in wound site compared to the whole CHT in the WT//Tg (lyz:EGFP) and tdh−/−//Tg (lyz:EGFP) transgenic zebrafish embryos (G) (n = 5). Note that, supplementation with Pyr alone or with Glu efficiently rescued the migration defect of neutrophil. Glu, glucose; Gly, glycine; Ile, isoleucine; Leu, leucine; Met, methionine; Pyr, pyruvate; Ser, serine; Thr, threonine. Embryos were replenished with indicated metabolites at 24 h post fertilization (hpf) and culture media were refreshed every 24 h. Scale bar = 500 μm. Data are presented as the mean ± SD; two-tailed t-test; ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05.