cathepsin-g and Wounds-and-Injuries

Topics: Bandages; Cathepsin G; Cathepsins; Chronic Disease; Fibrinolysin; Humans; Hydrogen-Ion Concentration; Matrix Metalloproteinase 2; Ointments; Pancreatic Elastase; Peptide Hydrolases; Serine Endopeptidases; Skin Care; Wound Healing; Wounds and Injuries

Human neutrophil elastase (HNE) and cathepsin G (CatG) are involved in the pathogenesis of a number of inflammatory disorders. These serine proteinases are released by neutrophils and monocytes in case of infection. Wound infection is a severe complication regarding wound healing causing diagnostic and therapeutic problems. In this study we have shown the potential of HNE and CatG to be used as markers for early detection of infection. Significant differences in HNE and CatG levels in infected and non-infected wound fluids were observed. Peptide substrates for these two enzymes were successfully immobilised on different surfaces, including collagen, modified collagen, polyamide polyesters and silica gel. HNE and CatG activities were monitored directly in wound fluid via hydrolysis of the chromogenic substrates. Infected wound fluids led to significant higher substrate hydrolysis compared with non-infected ones. These different approaches could be used for the development of devices which are able to detect elevated enzyme activities before manifestation of infection directly on bandages. This would allow a timely intervention by medical doctors thus preventing severe infections.

Topics: Bandages; Biomarkers; Cathepsin G; Chromogenic Compounds; Exudates and Transudates; Humans; Leg Ulcer; Leukocyte Elastase; Pressure Ulcer; Surgical Wound Infection; Wound Healing; Wound Infection; Wounds and Injuries

High elastase and cathepsin G activities have been observed in chronic wounds to inhibit healing through degradation of growth factors, cytokines, and extracellular matrix proteins. Oleic acid is a non-toxic elastase inhibitor. Cotton wound dressing material was characterized as a transfer carrier for affinity uptake of oleic acid by albumin under conditions mimicking chronic wounds. The mechanism of oleic acid uptake from cotton and binding by albumin was examined with both intact dressings and cotton fiber-designed chromatography. Raman spectra of the albumin-oleic acid complexes under liquid equilibrium conditions revealed fully saturated albumin-oleic acid complexes with a 1:1 weight ratio of albumin:oleic acid. Liquid-solid equilibrium conditions revealed oleic acid transfer from cotton to albumin at 27 mole equivalents of oleic acid per mole albumin. Comparing oleic acid formulated wound dressings for dose dependent ability to lower elastase activity, we found cotton gauze>hydrogel>hydrocolloid. In contrast, the cationic serine protease cathepsin G was inhibited by oleic acid within a narrow range of oleic acid-cotton formulations. 2% albumin was sufficient to transfer quantities of oleic acid necessary to achieve a significant elastase-lowering effect. Oleic acid bound to cotton wound dressings may have promise in the selective lowering of cationic serine protease activity useful in topical application for chronic inflammatory pathogenesis.

Topics: Bandages; Bandages, Hydrocolloid; Cathepsin G; Cathepsins; Chemistry, Pharmaceutical; Chromatography, High Pressure Liquid; Chronic Disease; Cotton Fiber; Dose-Response Relationship, Drug; Drug Compounding; Humans; Leukocyte Elastase; Occlusive Dressings; Oleic Acid; Protein Binding; Serine Endopeptidases; Serine Proteinase Inhibitors; Serum Albumin, Bovine; Solubility; Spectrum Analysis, Raman; Wound Healing; Wounds and Injuries

The host defense roles of neutrophil elastase in a porcine skin wound chamber model were explored. Analysis of wound fluid by acid-urea polyacrylamide gel electrophoresis, Western blot, and bacterial overlay confirmed that the neutrophil-derived protegrins constituted the major stable antimicrobial polypeptide in the wound fluid. The application to the wound of 0.10 and 0.25 mM N-methoxysuccinyl-alanine-alanine-proline-valine (AAPV) chloromethyl ketone, a specific neutrophil elastase inhibitor (NEI), blocked the proteolytic activation of protegrins and diminished the associated antimicrobial activity as detected by radial diffusion assay against Staphylococcus epidermidis, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and Candida albicans or by bacterial gel overlay against S epidermidis and E coli. The application of the related cathepsin G inhibitor (CGI), benzyloxycarbonyl-glycine-leucine-phenylalanine (ZGLF) chloromethyl ketone, had no effect. In wound chambers that received 10(6) colony-forming unit (CFU)/mL of S epidermidis, the presence of NEI significantly decreased the 24-hour clearance of bacteria from the wound compared to wounds treated with CGI or solvent only. Neither inhibitor, at 0.10 or 0.25 mM concentration, affected leukocyte accumulation or degranulation in the wound chambers. The in vitro microbicidal decrement due to NEI was restored by an amount of the specific protegrin (PG-1), which was equivalent to the measured difference of protegrin between control and inhibited chambers. Administration of 1 microg/mL exogenous PG-1 4 hours after chamber preparation was sufficient to normalize in vivo antimicrobial activity. Although pharmacologic NEIs are promising candidates as anti-inflammatory drugs, they may impair host defense in part by inhibiting the activation of cathelicidins by neutrophil elastase.

Topics: Animals; Anti-Bacterial Agents; Antimicrobial Cationic Peptides; Bacteria; Body Fluids; Cathelicidins; Cathepsin G; Cathepsins; Dose-Response Relationship, Drug; Leukocyte Elastase; Microbial Sensitivity Tests; Models, Animal; Neutrophil Infiltration; Proteins; Serine Endopeptidases; Serine Proteinase Inhibitors; Serpins; Superoxides; Swine; Wounds and Injuries