Figure 2.
C15 Inhibits Early Platelet and Neutrophil Inflammatory Events after Cutaneous Wounding
A single 580 ± 39 μm2 focal injury was induced on the surface of a dorsal skin flap with a modified electrocautery device. C15 (100 pg/wound) or vehicle (saline) was administered intradermally immediately after wounding. For sham experiments, mice were prepared for intravital microscopy and imaged identically to injured animals, but no injury was induced.
(A) Schematic diagram of the skin preparation with burn injury for multichannel fluorescence spinning-disk confocal microscopy.
(B) Schematic showing the wound as viewed with a 4× objective; necrotic cells stained with propidium iodide are shown in red.
(C) Low-power (4×) views of necrotic cells (propidium iodide, red) and neutrophil recruitment (blue) to vehicle- or C15-treated wounds 2 hr after injury. The scale bar indicates 150 μm. Dotted lines indicate the wound margin, 150 μm and 500 μm from the wound edge.
(D) Quantification of neutrophil recruitment to wounds treated with vehicle or C15.
(E) High-power (20x) views of neutrophil (Ly6G, blue) and platelet (CD49b, red) behavior within dermal postcapillary venules 2 hr after wounding. The scale bar indicates 30 μm. The arrow in “Sham” indicates a rolling neutrophil. The large arrow in “Burn + vehicle” indicates a neutrophil without interacting platelets. The small arrow indicates neutrophils with interacting platelets.
(F–H) Quantification of neutrophil rolling and adhesion (F), platelet rolling and adhesion (G), and platelet-neutrophil interactions (H) after wounding.
Data are expressed as means ± SEM; four to eight vessels were visualized per time point in each mouse, and there were four to eight animals per treatment group. Vehicle (n = 4), burn (n = 8), C15 (n = 7). ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001 relative to vehicle-treated mice. See also Movies S1, S2, and S3. The following abbreviation is used: FOV, field of view.