Anchoring a cytoactive factor in a wound bed promotes healing

Abstract

Wound healing is a complex process that requires the intervention of cytoactive factors. The one-time application of soluble factors to a wound bed does not maintain a steady, sufficient concentration. Here we investigated the benefits of anchoring a factor in a wound bed via a tether to endogenous collagen. We used a collagen-mimetic peptide (CMP) as a pylon. The CMP binds to damaged but not intact collagen and thus localizes a pendant cytoactive factor in the regions of a wound bed that require intervention. As a model factor, we chose substance P, a peptide of the tachykinin family that promotes wound healing. Using splinted wounds in db/db mice, we found that the one-time application of a CMP-substance P conjugate enhances wound healing compared to unconjugated substance P and other controls. Specifically, all 16 wounds treated with the conjugate closed more thoroughly and, did so with extensive re-epithelialization and mitigated inflammatory activity. These data validate a simple and general strategy for re-engineering wound beds by the integration of beneficial cytoactive factors. Copyright © 2014 John Wiley & Sons, Ltd.

Keywords: Mus musculus; Substance P; collagen; extracellular matrix; peptide; synthetic biology.

Figure 1

Conceptual representation of a collagen mimetic peptide conjugate (CMP–X) annealed to a damaged collagen triple helix in a wound bed. “X” is a cytoactive factor that becomes anchored in the wound bed.

Figure 2

Photographs depicting the effect of SubP-immobilization on splinted mouse wounds. Representative images are from Day 0 immediately post-treatment, or Day 16 after removal of the splints but before euthanasia. Wounds (8-mm o.d.) were treated with (A) saline, (B) PEG (5% w/v) in saline, (C) SubP (25 nmol) in 5% w/v PEG/saline, (D) CMP (25 nmol) in 5% w/v PEG/saline, or (E) CMP–SubP (25 nmol) in 5% w/v PEG/saline.

Figure 3

Effect of SubP-immobilization on wound histology. Representative histological images are from Day 16 after treatment as described in Figure 2. Wounds were stained with hematoxylin + eosin and viewed under bright-field light (top or left panels; 2× objective or 4× objective for CMP–SubP), and stained with picrosirius red and viewed under polarized light (bottom or right panels; 2× objective or 4× objective for CMP–SubP). Wound edges are indicated by “*” symbols. Scale bars: 500 μm.

Figure 4

Effect of SubP-immobilization on wound healing. Data are from Day 16 after treatment as described in Figure 2. (A) Mean size of wounds. (B) Re-epithelialization of wounds. (C) Collagen deposition as a percentage of the total area of the wound. (D) Inflammation in wounds on scale of 0–4. Data are the mean ± SE from histopathological analyses of 80 wounds (= 5 regimens × 16 wounds/regimen); asterisks (*) indicate significant differences (p < 0.05).