Wound debridement potential of glycosidases of the wound-healing maggot, Lucilia sericata.

The wound-healing maggot, Lucilia sericata Meigen (Diptera: Calliphoridae), degrades extracellular matrix components by releasing enzymes. The purpose of this study was to investigate the glycosylation profiles of wound slough/eschar from chronic venous leg ulcers and the complementary presence of glycosidase activities in first-instar excretions/secretions (ES1) and to define their specificities. The predominant carbohydrate moieties present in wound slough/eschar were determined by probing one-dimensional Western blots with conjugated lectins of known specificities. The presence of specific glycosidase activities in ES1 was determined using chromogenic and fluorogenic substrates. The removal of carbohydrate moieties from slough/eschar proteins by glycosidases in ES1 was determined by two-dimensional electrophoresis and Emerald 300 glycoprotein staining. α-D-glucosyl, α-D-mannosyl and N-acetylglucosamine residues were detected on slough/eschar-derived proteins. Furthermore, it was demonstrated that the treatment of slough/eschar with ES1 significantly reduced uptake of the carbohydrate-specific stain. Subsequently, α-D-glucosidase, α-D-mannosidase and N-acetylglucosaminidase activities were identified in ES1. Specific chromogenic and fluorogenic substrates and gel filtration chromatography showed that these activities result from distinct enzymes. These activities were mirrored in the removal of α-D-glucosyl, α-D-mannosyl and N-acetylglucosamine residues from proteins of slough/eschar from maggot-treated wounds. These data suggest that maggot glycosidases remove sugars from slough/eschar proteins. This may contribute to debridement, which is ultimately accomplished by a suite of biochemically distinct enzymes present in ES1.

Maggot chymotrypsin I from Lucilia sericata is resistant to endogenous wound protease inhibitors.

BACKGROUND: A chymotrypsin found in the secretions of Lucilia sericata and manufactured as a recombinant enzyme degrades chronic wound eschar ex vivo. OBJECTIVES: To characterize the inhibition profile of the L. sericata recombinant chymotrypsin I. METHODS: Activity of recombinant chymotrypsin I and its sensitivity to endogenous inhibitors were determined enzymatically using the fluorogenic substrate succinyl-alanyl-alanyl-prolyl-phenylalanyl-aminomethyl coumarin. RESULTS: We report the presence of high concentrations of two endogenous inhibitors, α1-antichymotrypsin and α1-antitrypsin, in wound eschar and a trace of a third, α2-macroglobulin, with the potential to inhibit this debridement process. However, the addition of a soluble and inhibitor-containing extract of chronic wound eschar to chymotrypsin I did not affect activity of the enzyme, neither did the addition of purified native α1-antichymotrypsin or α1-antitrypsin, although chymotrypsin I was inhibited by α2-macroglobulin. Conversely, the mammalian equivalent, α-chymotrypsin, was inhibited by the purified native α1-antichymotrypsin, α1-antitrypsin and α2-macroglobulin and by the soluble extract of wound eschar. CONCLUSIONS: The data suggest that the maggot-derived chymotrypsin I is biochemically distinct from human α-chymotrypsin and the lack of inhibition by wound eschar suggests a means by which chymotrypsin I activity survives within the wound to contribute towards debridement during maggot biotherapy.

Degradation of eschar from venous leg ulcers using a recombinant chymotrypsin from Lucilia sericata.

BACKGROUND: Larvae of the greenbottle Lucilia sericata are used to debride nonhealing wounds and stimulate the production of fresh granulation tissue. Previous publications have shown that secretions from L. sericata contain a number of proteolytic activities including a chymotrypsin that degrades a number of extracellular matrix components such as fibronectin, laminin and collagen. OBJECTIVES: To produce a recombinant L. sericata chymotrypsin (chymotrypsin I) and determine its effects on the degradation of patient wound eschar. METHODS: An active recombinant chymotrypsin I from L. sericata was cloned and expressed in Sf9 cells and its subsequent effects ex vivo on eschar from venous leg ulcers were determined by two-dimensional electrophoresis. RESULTS: The recombinant enzyme had the attributes of a chymotrypsin, possessing sequence homology with other chymotrypsins and demonstrating attributes of the native enzyme including cleavage of the chymotrypsin substrate succinyl-alanyl-alanyl-prolyl-phenylalanyl-7-amino-4-methyl coumarin, inhibition by phenylmethylsulphonyl fluoride and lack of inhibition by amidinophenylmethylsulphonyl fluoride. Importantly, the recombinant chymotrypsin cleaved the majority of proteins from slough/eschar from venous leg ulcers in a superior manner to chymotrypsins from human and bovine sources. CONCLUSIONS: The ex vivo degradation of eschar from venous leg ulcers indicates the potential value of recombinant chymotrypsin I as a novel, stand-alone debridement agent.

Updates from the British Association of Dermatologists 87 th annual meeting, 10-13 July 2007, Birmingham, U.K.

This is a synopsis of the main research and clinical findings presented at the British Association of Dermatologists meeting held during 10-13 July 2007 in Birmingham, U.K. The conference highlighted the recent biological, epidemiological and therapeutic advances that have been made recently in the field of dermatology. The authors focus on the more important advances or summaries of findings, but this is not meant as a substitute for reading the conference proceedings and related references quoted in this article.

Updates from the British Society for Investigative Dermatology Annual Meeting, 16-18 April 2007, Nottingham, U.K.

This is a synopsis of the main research findings presented at the British Society for Investigative Dermatology meeting held during 16-18 April 2007 in Nottingham, U.K. The conference highlighted the recent biological, epidemiological and therapeutic advances that have been made in the field of dermatology. The authors focus on the more important advances or summaries of findings, but this is not meant as a substitute for reading the conference proceedings and related references quoted in this article.

Enzymes, detergents and skin: facts and fantasies.

In their raw state, enzymes of bacterial/fungal origin cause allergic reactions in the lung. Proteolytic enzymes also cause irritation to skin, eyes and the respiratory tract. For 40 years, encapsulated enzymes have been used worldwide in detergent products, especially laundry formulations, and have increasing importance due to biodegradability and functionality at low temperatures, offering environmental benefits. Uniquely to the U.K., for years it has been suggested that the inclusion of enzymes in such products leads to adverse skin reactions, including erythema, pruritus and exacerbation of eczema. In this review, we look at the facts, asking whether there is evidence that the hazards identified for enzymes translate into any risk for consumer health. By considering the actual exposures in consumer use and exaggerated product usage, it is concluded that the irritating and allergenic hazards of enzyme raw materials do not translate into a risk of skin reactions, either irritant or allergic. Investigations of numerous individuals with skin complaints attributed to laundry products demonstrate convincingly that enzymes were not responsible. Indeed, enzyme-containing laundry products have an extensive history of safe use. Thus, the supposed adverse effects of enzymes on skin seem to be a consequence of a mythology. The important practical lesson is that when primary or secondary care practitioners are presented with a skin complaint, it should not be dismissed as a result of using an enzyme-containing laundry product as the diagnosis will certainly lie elsewhere. Education for healthcare professionals could usefully be enhanced to take this on board.

Updates from the British Association of Dermatologists 86th annual meeting, 4-7 July 2006, Manchester, U.K.

Here we provide a synopsis of the main clinical and research advances in clinical, epidemiological and biological dermatology that were presented at the meeting of the British Association of Dermatologists (BAD) held during 4-7 July 2006, in Manchester, U.K. Only the more important advances or summaries of findings are mentioned. The meeting was held at the Manchester International Conference Centre (Fig. 1). The annual dinner was held at Manchester Town Hall, in the Great Hall decorated with magnificent murals by Ford Madox Brown, with Dr Susan Burge as host.

Updates from the British Association of Dermatologists 85th annual meeting, 5-8 July 2005, Glasgow, U.K.

The conference highlighted the progress made in understanding recent biological, epidemiological and therapeutic advances in dermatology. Here we provide a synopsis of the main research and clinical findings presented at the meeting of the British Association of Dermatologists (BAD) held during 5-8 July 2005, in Glasgow, U.K., drawing attention to the most important advances and summaries. The BAD meeting was held at the Scottish Exhibition and Conference Centre, Glasgow (Fig. 1). The annual dinner was held in the wonderful setting of Stirling Castle, with Dr Robin Graham-Brown as host.

Intervention development in occupational research: an example from the printing industry.

BACKGROUND: Intervention development research is an essential prerequisite of any study that attempts to determine whether specific interventions work to prevent work related injury and illness. METHODS: Focus groups (n = 5) and direct observational studies (n = 21) of printers were used to elicit key issues that would aid the development of subsequent interventions. Transcripts from these were analysed by standard qualitative methods to identify common and related themes. RESULTS: The views of managers differed significantly from those of print workers in a number of areas, and working practices did not always follow policy. The majority of printers did not perceive dermatitis to be a major problem, although many complained of dry hands. Other key results included: the lack of skin care policy in most companies; poor understanding of the nature, causes, and treatment of dermatitis; low priority of dermatitis within health and safety concerns; little or no provision of occupational health services, particularly skin checks; variability in provision of and access to appropriate skin protection; and lack of accessible washing facilities. CONCLUSIONS: As a result it was decided to evaluate the implementation of four INTERVENTIONS: provision of (1) skin checks and treatment advice; (2) gloves of the correct type and size, and use of an after-work cream; (3) information on dermatitis within the printing industry; and (4) development of best practice skin care policy.

Updates from the British Association of Dermatologists 84th annual meeting, 6-9 July 2004, Belfast, U.K.

Herein is a synopsis of the main research and clinical findings presented at the British Association of Dermatologists meeting held during 6-9 July 2004, in Belfast, U.K. The conference highlighted the progress that has been made in understanding the increasing biological, epidemiological and therapeutic advances that have been made recently in the field of dermatology. The authors highlight the more important advances or summaries, but this is not meant as a substitute for reading the conference proceedings and related references quoted in this article.