Figure 2. Collagen formation, vascularity, and monocyte/macrophage infiltration at the wound site.

Collagen formation, vascularity, and monocyte/macrophage infiltration were evaluated 14 days, 14 days, and 7 days, respectively, after wounds were treated with 6 mg/kg AMD3100 in 30 μL saline or saline alone. (A, B) Collagen formation was assessed in sections of wound tissue stained with Masson trichrome or picrosirius red; picosirius red stain accentuates the birefringence of collagen fibers when viewed under polarized light. The boxed regions in the Masson-trichrome–stained panels (left) are displayed with picrosirius red staining in the right panels. Bar=100 μm. (B) Fiber formation at the wound center was quantified as the percentage of the digitized image area that fluoresced red (mature fibers) or yellow-green (immature fibers). (C) Wound tissue sections were stained with FITC-conjugated BS1 lectin (green) to identify endothelial tissue and with alkaline-phosphatase αSMA (red) to identify smooth-muscle cells. αSMA-positive vessels are identified with arrows. Bar=100 μm. (D) Capillary density was quantified as the percentage of the image area that fluoresced positively for BS1 lectin, and (E) smooth muscle–containing vessels were quantified as the number of structures in the dermis that stained positively for both BS1 lectin and αSMA. (F) Sections of wound tissue were stained for expression of the monocyte/macrophage marker CD68 (red); nuclei were counterstained with DAPI (blue); bar=100 μm. The boxed regions were evaluated for quantification of (G) monocyte/macrophage infiltration (i.e., the number of CD68-positive cells).