Analysis of the acute and chronic wound environments: the role of proteases and their inhibitors

Abstract

To assess the differences in proteolytic activity of acute and chronic wound environments, wound fluids were collected from acute surgical wounds (22 samples) and chronic wounds (25 samples) of various etiologies, including mixed vessel disease ulcers, decubiti and diabetic foot ulcers. Matrix metalloproteinase (MMP) activity measured using the Azocoll assay was significantly elevated by 30 fold in chronic wounds (median 22.8 microg MMP Eq/ml) compared to acute wounds (median 0.76 microg MMP Eq/ml) (p < 0.001). The addition of the matrix metalloproteinase inhibitor Illomostat decreased the matrix metalloproteinase activity by approximately 90% in all samples, confirming that the majority of the activity measured was due to matrix metalloproteinases. Gelatin zymograms indicated predominantly elevated matrix metalloproteinase-9 with smaller elevations of matrix metalloproteinase-2. In addition tissue inhibitor of metalloproteinase-1 levels were analyzed in a small subset of acute and chronic wounds. When tissue inhibitor of metalloproteinase-1 levels were compared to protease levels there was an inverse correlation (p = 0.02, r = - 0.78). In vitro degradation of epidermal growth factor was measured by addition of 125I labelled epidermal growth factor to acute and chronic wound fluid samples. There was significantly higher degradation of epidermal growth factor in chronic wound fluid samples (mean 28.1%) compared to acute samples (mean 0.6%). This also correlated to the epidermal growth factor activity of these wound fluid samples (p < 0. 001, r = 0.64). Additionally, the levels of proteases were assayed in wound fluid collected from 15 venous leg ulcers during a nonhealing and healing phase using a unique model of chronic wound healing in humans. Patients with nonhealing venous leg ulcers were admitted to the hospital for bed rest and wound fluid samples were collected on admission (nonhealing phase) and after 2 weeks (healing phase) when the ulcers had begun to heal as evidenced by a reduction in size (median 12%). These data showed that the elevated levels of matrix metalloproteinase activity decreased significantly as healing occurs in chronic leg ulcers (p < 0.01). This parallels the processes observed in normally healing acute wounds. This data also supports the case for the addition of protease inhibitors in chronic wounds in conjunction with any treatments using growth factors.

Nano-formulated curcumin accelerates acute wound healing through Dkk-1-mediated fibroblast mobilization and MCP-1-mediated anti-inflammation

“…Studies have also shown that the prolonged exposure of wound tissues to pro-inflammatory cytokines, for example, interleukin and tumor necrosis factor-α, may act to stimulate the production of MMPs, while inhibiting the synthesis of tissue inhibitors of metalloproteinases. 31 Furthermore, MCP-1 has recently been shown to up-regulate MMP-1 and MMP-2 mRNA expression in human skin fibroblasts in which interleukin-1α has been found to take part as a mediator through an autocrine loop. 32 As a matter of fact, curcumin itself possesses distinct chemical properties that allow it to use the metal atoms of metalloproteins as a means to interact with these proteins and become a natural MMP inhibitor.…”

Fibroblasts from chronic wounds show altered TGF‐β‐signaling and decreased TGF‐β Type II Receptor expression

“…It has been speculated that topical application of these agents, especially in the harsh microenvironment of a chronic wound, is fraught with problems. There are questions about the appropriate topical dose and the delivery vehicle, penetration of the polypeptide into the ulcer bed and its breakdown by tissue metalloproteinases (Robson et al, 1998;Trengove et al, 1999;Lobmann et al, 2002). Also, single growth factors may not be enough (Robson et al, 2000;Smith et al, 2000a,b).…”

“…In the case of venous ulcers, a number of hypotheses have been proposed. Some of these hypotheses have emphasized failure of the fibrinolytic system and the formation of pericapillary fibrin cuffs (Browse and Burnand, 1982;Van de Scheur and Falanga, 1997), while others have addressed the possibility of trapping of growth factors (Falanga and Eaglstein, 1993) and breakdown of extracellular matrix and cytokines by excessive amounts of tissue metalloproteinases (Trengove et al, 1999;Lobmann et al, 2002). More recently, however, a novel line of investigation has been centered on the hypothesis that chronic wounds fail to heal because their resident cells are senescent and/or have become unresponsive to the action of growth factors.…”

Adeno-associated viral vector-mediated human vascular endothelial growth factor gene transfer stimulates angiogenesis and wound healing in the genetically diabetic mouse

“…This would suggest that exogenous presentation can lead to clearance, sequestration [41] or destruction [42] of the factor.…”