Moor AN, Vachon DJ, Gould LJProteolytic activity in wound fluids and tissues derived from chronic venous leg ulcers. Wound Repair Regen 17:832-839

Department of Surgery, University of South Florida, Tampa, Florida, USA.
Wound Repair and Regeneration (Impact Factor: 2.75). 11/2009; 17(6):832-9. DOI: 10.1111/j.1524-475X.2009.00547.x
Source: PubMed


Venous leg ulcers affect approximately 1% of the general population and 3.6% of those over the age of 65. The goal of the research described herein is to shorten the time to healing by developing wound care alternatives that are based on a comprehensive understanding of the venous ulcer wound environment. The proteolytic and inflammatory components in wound fluids and tissue biopsy samples were characterized in subjects with documented long-standing venous ulcers that had showed resistance to standard therapy. All wounds showed polymicrobial colonization with greater than 10(6) CFU/g. Myeloperoxidase, a measure of leukocyte infiltration, was also markedly elevated in these wounds. Zymography revealed the presence of both pro-matrix metalloproteinase (MMP)-2 and pro-MMP-9 in wound fluids and to a lesser extent in tissue biopsies. Using an immunocapture activity assay we reveal a sevenfold excess of MMP-9 in wound fluid as compared to tissue, with 73% in the activated form. In contrast, MMP-8 total protein levels were nearly equal in wound fluids and biopsies. Fibronectin, a critical component of the extracellular matrix, was shown to be degraded in both wound fluids and biopsy samples. Finally, the potential of a novel wound dressing to neutralize several constituents of this hostile wound environment is shown.

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    • "The analysis of wound fluid (WF) provides a direct insight into the local extracellular microenvironment of a wound [2]. In the last years, several proteins, especially inflammatory mediators, growth factors and cytokines [3] [4], proteases [5] [6], and oxidative stress related proteins [7] [8], were identified in WF which can be used to characterize and classify the effect on different phases of the wound healing process and thereby can be used as predictive and diagnostic tools. Nevertheless, the underlying mechanisms in wound healing disturbance are not completely understood and so far none of the potential biomarkers for dysregulated wound healing was established obtained desorbates were highly concentrated, contained up to 100 micrograms of total protein, required no further purification, and were sufficient for a reproducible analysis of more than 500 proteins by in-gel separation and proteolytic digestion followed by liquid chromatography-tandem mass spectrometry (GeLC-MS/MS). "
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    ABSTRACT: Wound healing of soft tissue and bone defects is a complex process in which cellular differentiation and adaption are regulated by internal and external factors, among them are many different proteins. In contrast to insights into the significance of various single proteins based on model systems, the knowledge about the processes at the actual site of wound healing is still limited. This is caused by a general lack of methods that allow sampling of extracellular factors, metabolites, and proteins in situ. Sampling of wound fluids in combination with proteomics and metabolomics is one of the promising approaches to gain comprehensive and time resolved data on effector molecules. Here, we describe an approach to sample metabolites by microdialysis and to extract proteins simultaneously by adsorption. With this approach it is possible (i) to collect, enrich, and purify proteins for a comprehensive proteome analysis; (ii) to detect more than 600 proteins in different defects including more than 100 secreted proteins, of which many proteins have previously been demonstrated to have diagnostic or predictive power for the wound healing state; and (iii) to combine continuous sampling of cytokines and metabolites and discontinuous sampling of larger proteins to gain complementary information of the same defect.
    BioMed Research International 08/2014; 2014:934848. DOI:10.1155/2014/934848 · 1.58 Impact Factor
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    • "Moor et al.[33] reported that wound fluid myeloperoxidase levels were associated with nonhealing and infected wounds. Studies are also on to determine if wound analysis for volatile organic compounds (e.g., various esters, alcohols, and organic compounds) may help in prognostication.[34] "
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    ABSTRACT: Researchers have identified several of the cellular events associated with wound healing. Platelets, neutrophils, macrophages, and fibroblasts primarily contribute to the process. They release cytokines including interleukins (ILs) and TNF-α, and growth factors, of which platelet-derived growth factor (PDGF) is perhaps the most important. The cytokines and growth factors manipulate the inflammatory phase of healing. Cytokines are chemotactic for white cells and fibroblasts, while the growth factors initiate fibroblast and keratinocyte proliferation. Inflammation is followed by the proliferation of fibroblasts, which lay down the extracellular matrix. Simultaneously, various white cells and other connective tissue cells release both the matrix metalloproteinases (MMPs) and the tissue inhibitors of these metalloproteinases (TIMPs). MMPs remove damaged structural proteins such as collagen, while the fibroblasts lay down fresh extracellular matrix proteins. Fluid collected from acute, healing wounds contains growth factors, and stimulates fibroblast proliferation, but fluid collected from chronic, nonhealing wounds does not. Fibroblasts from chronic wounds do not respond to chronic wound fluid, probably because the fibroblasts of these wounds have lost the receptors that respond to cytokines and growth factors. Nonhealing wounds contain high levels of IL1, IL6, and MMPs, and an abnormally high MMP/TIMP ratio. Clinical examination of wounds inconsistently predicts which wounds will heal when procedures like secondary closure are planned. Surgeons therefore hope that these chemicals can be used as biomarkers of wounds which have impaired ability to heal. There is also evidence that the application of growth factors like PDGF will help the healing of chronic, nonhealing wounds.
    Indian Journal of Plastic Surgery 05/2012; 45(2):220-8. DOI:10.4103/0970-0358.101282
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    • "MMPs roles in pathology may be grouped into the following main types: (1) tissue destruction (e.g., cancer invasion and metastasis, arthritis, ulcers, periodontal diseases, brain degenerative diseases)(Gottschall and Deb, 1996; Yong et al., 2001; Malemud, 2006; Agrawal et al., 2008; Moor et al., 2009; Oyarzun et al., 2010; Gialeli et al., 2011; Hainard et al., 2011; Kessenbrock et al., 2011; Sexton et al., 2011; Skarmoutsou et al., 2011); (2) fibrosis (e.g., liver cirrhosis, fibrotic lung disease, otosclerosis, atherosclerosis, and multiple sclerosis)(Starckx et al., 2003; Fiotti et al., 2004; Chakrabarti and Patel, 2005; Roderfeld et al., 2007; Rybakowski, 2009; Lim et al., 2010; Ragino et al., 2010); (3) weakening of matrix (e.g., dilated cardiomyopathy, aortic aneurysm and varicose veins)(Papalambros et al., 2003; Reddy et al., 2004; Mannello and Raffetto, 2011). As for most biological processes, matrix degradation is a precise event, attributed to proteinases that are produced and released on demand from activated cells. "
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    ABSTRACT: Matrix metalloproteinases (MMPs) were originally identified as matrixin proteases that act in the extracellular matrix. Recent works have uncovered nontraditional roles for MMPs in the extracellular space as well as in the cytosol and nucleus. There is strong evidence that subspecialized and compartmentalized matrixins participate in many physiological and pathological cellular processes, in which they can act as both degradative and regulatory proteases. In this review, we discuss the transcriptional and translational control of matrixin expression, their regulation of intracellular sorting, and the structural basis of activation and inhibition. In particular, we highlight the emerging roles of various matrixin forms in diseases. The activity of matrix metalloproteinases is regulated at several levels, including enzyme activation, inhibition, complex formation and compartmentalization. Most MMPs are secreted and have their function in the extracellular environment. MMPs are also found inside cells, both in the nucleus, cytosol and organelles. The role of intracellular located MMPs is still poorly understood, although recent studies have unraveled some of their functions. The localization, activation and activity of MMPs are regulated by their interactions with other proteins, proteoglycan core proteins and / or their glycosaminoglycan chains, as well as other molecules. Complexes formed between MMPs and various molecules may also include interactions with noncatalytic sites. Such exosites are regions involved in substrate processing, localized outside the active site, and are potential binding sites of specific MMP inhibitors. Knowledge about regulation of MMP activity is essential for understanding various physiological processes and pathogenesis of diseases, as well as for the development of new MMP targeting drugs.
    Progress in Histochemistry and Cytochemistry 03/2012; 47(1):27-58. DOI:10.1016/j.proghi.2011.12.002 · 3.64 Impact Factor
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