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Fig. 1 Circadian disorder increases the recruitment of neutrophils to injured tissue (A) Zebrafish embryos were raised in LD, DD, and LL conditions for 4 consecutive days and then tested in LD and DD conditions at 5 dpf. (B and C) The rhythmic expression of clock1a, bmal1b, cry1ba, and per1b was significantly disrupted in larvae raised under LL and DD conditions. Relative expression levels of these genes were normalized to the control gene β-actin. The rhythmic mRNA expression of clock genes was analyzed by the JTK-CYCLE method. Two-way ANOVA was used to analyze the variation between different groups. Each experiment included three samples, and each sample contained 50 larvae. ZT, zeitgeber time; ZT0, light on; ZT14, light off. (D) The movements of zebrafish larvae were constantly monitored by behavioral instruments. (E and F) The locomotor behavior of larvae raised in LL and DD conditions showed irregular rhythms compared with the control group (n = 16, each group). (G) The tail fin of zebrafish was damaged by a surgical blade. (H and I) Neutrophil migration to the injured site at 3 hpw significantly increased in larvae raised in LL and DD conditions at ZT1 (n = 20, each group). The data were analyzed with one-way ANOVA. White boxes represent statistical areas. Scale bar: 10 μm (J and K) There was no significant difference in the total number of fluorescently labeled neutrophil in larvae raised in LD, LL, and DD conditions (n = 20, each group). Scale bar: 50 μm. The data were analyzed with one-way ANOVA. All experiments were repeated three times, and the results show the data from an independent experiment. Bar graphs represent the mean ± standard error of the mean (SEM) (∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001).