Figure 4

Liposome encapsulation of PLP increased its inhibitory effect on neutrophil recruitment upon laser wounding. Tg(mpx:GFP) embryos (at 3 dpf) were subjected to the laser wounding procedure and injected with different doses of free or liposome-encapsulated PLP, and fluorescence microscopy images were taken at 4 hpw. (A) Schematic drawing of a zebrafish embryo at 3 dpf indicating the area in which the recruited neutrophils were counted (red box). (B) Representative image showing the accumulation of neutrophils (green) near the wound. Scale bar = 100 μm. (C–E) The number of neutrophils recruited to the wounded area at 4 hpw are shown after injection of different doses of free PLP (C), PLP encapsulated in AmbiMACs (20% DSPG) (D), and PLP encapsulated in PEGylated liposomes (E); H₂O and empty liposomes were injected as control. Statistical analysis was performed by one-way ANOVA with Bonferroni’s post hoc test. A significant inhibition of the neutrophil migration was observed when embryos had been injected with 5 or 25 pmol of free PLP, 0.2 or 1 pmol of PLP encapsulated in AmbiMACs (20% DSPG) and 1 pmol of PLP encapsulated in PEGylated liposomes. Each data point represents a single embryo, and the means ± SEM of data accumulated from three independent experiments are shown in red. Statistically significant differences between groups are indicated by: ns, non-significant; * p < 0.05; ** p < 0.01; *** p < 0.001.