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1.
J Dermatol Sci ; 56(3): 168-80, 2009 Dec.
Article in English | MEDLINE | ID: mdl-19854029

ABSTRACT

BACKGROUND: Scar formation following skin trauma can have devastating consequences causing physiological and psychosocial concerns. Currently, there are no accepted predictable treatments to prevent scarring which emphasizes a need for a better understanding of the wound healing and scar formation process. OBJECTIVES: Previously it was shown that healing of small experimental wounds in the oral mucosa of red Duroc pigs results in significantly reduced scar formation as compared with equivalent full-thickness skin wounds. In the present study, scar formation was assessed in 17 times larger experimental wounds in both oral mucosa and skin of the red Duroc pigs. METHODS: Equivalent experimental wounds were created in the oral mucosa and dorsal skin of red Duroc pigs, and scar formation, localization and abundance of key wound healing cells, transforming growth factor-beta (TGF-beta) and phosphorylated Smad3 (pSmad3) were assessed. RESULTS: Oral mucosal wounds displayed significantly less clinical and histological scar formation than did the corresponding skin wounds. The number of macrophages, mast cells, TGF-beta and pSmad3 immunopositive cells was significantly reduced in the oral mucosal wounds as compared with skin wounds during the maturation stage of the healing process. Although the number of myofibroblasts was significantly elevated, the oral mucosal wounds showed significantly less contraction than did the skin wounds over time. CONCLUSIONS: Earlier resolution of the inflammatory reaction and reduced wound contraction may promote scarless oral mucosal wound healing. In addition, scar formation likely depends not only on the number of myofibroblasts but also on the extracellular environment which regulates their function.


Subject(s)
Cicatrix/physiopathology , Mouth Mucosa/physiopathology , Skin/physiopathology , Wound Healing , Animals , Blood Vessels/physiology , Cicatrix/pathology , Disease Models, Animal , Female , Fibroblasts/metabolism , Inflammation/pathology , Inflammation/physiopathology , Macrophages/metabolism , Mouth Mucosa/pathology , Skin/pathology , Smad3 Protein/metabolism , Swine , Transforming Growth Factor beta/metabolism
2.
Wound Repair Regen ; 17(5): 717-29, 2009.
Article in English | MEDLINE | ID: mdl-19769724

ABSTRACT

Scar formation is a common, unwanted result of wound healing in skin, but the mechanisms that regulate it are still largely unknown. Interestingly, wound healing in the oral mucosa proceeds faster than in skin and clinical observations have suggested that mucosal wounds rarely scar. To test this concept, we created identical experimental wounds in the oral mucosa and skin in red Duroc pigs and compared wound healing and scar development over time. We also compared the pig oral mucosal wound healing to similar experimental wounds created in human subjects. The findings showed significantly reduced scar formation at both clinical and histological level in the pig oral mucosa as compared with skin 49 days after wounding. Additionally, the skin scars contained a significantly increased number of type I procollagen immunopositive cells and an increased fibronectin content, while the oral mucosal wounds demonstrated a prolonged accumulation of tenascin-C. Furthermore, the pig oral mucosal wounds showed similar molecular composition and clinical and histological scar scores to human oral mucosal wounds. Thus, the reduced scar formation in the pig oral mucosa provides a model to study the biological processes that regulate scarless wound healing to find novel approaches to prevent scar formation in skin.


Subject(s)
Cicatrix/physiopathology , Disease Models, Animal , Mouth Mucosa/physiopathology , Regeneration/physiology , Skin/physiopathology , Swine , Wound Healing/physiology , Adult , Aged , Animals , Female , Humans , Male , Middle Aged , Mouth Mucosa/injuries , Young Adult
3.
J Immunol ; 179(8): 5137-45, 2007 Oct 15.
Article in English | MEDLINE | ID: mdl-17911599

ABSTRACT

A successful Th cell response to bacterial infections is induced by mature MHC class II molecules presenting specific Ag peptides on the surface of macrophages. In recent studies, we demonstrated that infection with the conventional vaccine Mycobacterium bovis bacillus Calmette-Guérin (BCG) specifically blocks the surface export of mature class II molecules in human macrophages by a mechanism dependent on inhibition of cathepsin S (Cat S) expression. The present study examined class II expression in macrophages infected with a rBCG strain engineered to express and secrete biologically active human Cat S (rBCG-hcs). Cat S activity was completely restored in cells ingesting rBCG-hcs, which secreted substantial levels of Cat S intracellularly. Thus, infection with rBCG-hcs, but not parental BCG, restored surface expression of mature MHC class II molecules in response to IFN-gamma, presumably as result of MHC class II invariant chain degradation dependent on active Cat S secreted by the bacterium. These events correlated with increased class II-directed presentation of mycobacterial Ag85B to a specific CD4(+) T cell hybridoma by rBCG-hcs-infected macrophages. Consistent with these findings, rBCG-hcs was found to accelerate the fusion of its phagosome with lysosomes, a process that optimizes Ag processing in infected macrophages. These data demonstrated that intracellular restoration of Cat S activity improves the capacity of BCG-infected macrophages to stimulate CD4(+) Th cells. Given that Th cells play a major role in protection against tuberculosis, rBCG-hcs would be a valuable tuberculosis vaccine candidate.


Subject(s)
Antigen Presentation/immunology , Cathepsins/metabolism , Gene Expression Regulation/immunology , HLA-D Antigens/biosynthesis , Macrophages/immunology , Macrophages/metabolism , Mycobacterium bovis/immunology , Mycobacterium bovis/metabolism , Acyltransferases/immunology , Acyltransferases/metabolism , Antigen Presentation/genetics , Antigens, Bacterial/immunology , Antigens, Bacterial/metabolism , Bacterial Proteins/immunology , Bacterial Proteins/metabolism , Cathepsins/genetics , Cell Line, Tumor , Genetic Engineering , HLA-D Antigens/genetics , HLA-D Antigens/metabolism , Humans , Intracellular Fluid/immunology , Intracellular Fluid/metabolism , Macrophages/microbiology , Mycobacterium bovis/genetics , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , T-Lymphocytes, Helper-Inducer/immunology , T-Lymphocytes, Helper-Inducer/metabolism , T-Lymphocytes, Helper-Inducer/microbiology , Tuberculosis Vaccines/administration & dosage , Tuberculosis Vaccines/genetics , Tuberculosis Vaccines/immunology
4.
Biosens Bioelectron ; 20(12): 2581-93, 2005 Jun 15.
Article in English | MEDLINE | ID: mdl-15854827

ABSTRACT

Sodium cyanide has been used in the Philippines to collect tropical marine fish for aquarium and food trades since the early 1960s. Cyanide fishing is a fast method to stun and collect fish. This practice is damaging the coral reefs irreversibly. In most countries cyanide fishing is illegal, but most of the exporting and importing countries do not have test and certificate systems. Many analytical methods are available for the detection of cyanide in environmental and biological samples. However, most of the techniques are time consuming, and some lack specificity or sensitivity. Besides, an ultra sensitive cyanide detection method is needed due to the rapid detoxification mechanisms in fish. The aim of this review is to give an overview of cyanide fishing problem in the south-east Asia and current strategies to combat this destructive practice, summarise some of the methods for cyanide detection in biological samples and their disadvantages. A novel approach to detect cyanide in marine fish tissues is briefly discussed.


Subject(s)
Biosensing Techniques/methods , Chemistry Techniques, Analytical/methods , Cyanides/analysis , Cyanides/pharmacokinetics , Fishes/metabolism , Water Pollutants, Chemical/analysis , Water Pollutants, Chemical/pharmacokinetics , Agriculture/methods , Animals , Anthozoa/metabolism , Biosensing Techniques/instrumentation , Chemistry Techniques, Analytical/instrumentation , Environmental Monitoring/instrumentation , Environmental Monitoring/methods , Food Contamination/analysis
5.
Appl Microbiol Biotechnol ; 67(5): 631-6, 2005 Jun.
Article in English | MEDLINE | ID: mdl-15630582

ABSTRACT

A partially purified preparation of cyanide hydrolase (cyanidase) from a bacterium, Klebsiella sp., was applied as a biocatalyst in a biosensor system for low-level cyanide detection. In the biosensor system cyanide hydrolase converts cyanide into formate and ammonia. The formate produced in the cyanide degradation was detected with a formate biosensor, in which formate dehydrogenase (FDH; E.C. 1.2.1.2) was co-immobilized with salicylate hydroxylase (SHL; E.C. 1.14.13.1) on a Clark electrode. The principle of the formate sensor is that FDH converts formate into carbon dioxide using beta-nicotinamide adenine dinucleotide hydrate (NAD(+)). The corresponding NADH produced is then oxidized to NAD(+) by SHL using salicylate and oxygen. The oxygen consumption is monitored with the Clark electrode. The optimum buffer pH and temperature for the enzymatic hydrolysis of potassium cyanide were studied. The preliminary experiments including the pretreatment of cyanide with cyanide hydrolase and then detection by the formate sensor gave a detection limit at 7.3 micromol l(-1) cyanide. The linear range of the calibration curve was between 30 micromol l(-1) and 300 micromol l(-1) cyanide.


Subject(s)
Biosensing Techniques/methods , Cyanides/analysis , Hydrolases/metabolism , Klebsiella/enzymology , Ammonia/metabolism , Carbon Dioxide/metabolism , Formate Dehydrogenases/metabolism , Formates/metabolism , Hydrogen-Ion Concentration , Mixed Function Oxygenases/metabolism , NAD/metabolism , Oxygen/metabolism , Oxygen Consumption , Salicylic Acid/metabolism , Sensitivity and Specificity , Temperature
6.
Biosens Bioelectron ; 19(12): 1745-52, 2004 Jul 15.
Article in English | MEDLINE | ID: mdl-15142609

ABSTRACT

A novel trienzyme sensor for the amperometric determination of lactate was constructed by immobilizing salicylate hydroxylase (SHL, E.C. 1.14.13.1), l-lactate dehydrogenase (LDH, E.C. 1.1.1.27), and pyruvate oxidase (PyOD, E.C. 1.2.3.3) on a Clark-type oxygen electrode. The enzymes were entrapped by a poly(carbamoyl) sulfonate (PCS) hydrogel on a Teflon membrane. LDH catalyzes the specific dehydrogenation of lactate consuming NAD(+). SHL catalyzes the irreversible decarboxylation and the hydroxylation of salicylate in the presence of oxygen and NADH produced by LDH. PyOD decarboxylates pyruvate using oxygen and phosphate. SHL and PyOD force the equilibrium of dehydrogenation of lactate by LDH to the product side by consuming NADH and pyruvate, respectively. Dissolved oxygen acts as an essential material for both PyOD and SHL during their respective enzymatic reactions. Therefore, an amplified signal, caused by the consumptions of dissolved oxygen by the two enzymes, was observed in the measurement of lactate. Regeneration of cofactor was found in the trienzyme system. A Teflon membrane was used to fabricate the sensor in order to avoid interferences. The sensor has a fast response (2s) and short recovery times (2 min). The total test time for a measurement by using this lactate sensor (4 min) was faster than using a commercial lactate testing kit (up to 10 min). The sensor has a linear range between 10 and 400 microM lactate, with a detection limit of 4.3 microM. A good agreement (R2 = 0.9984) with a commercial lactate testing kit was obtained in beverage sample measurements.


Subject(s)
Biosensing Techniques/instrumentation , Carbamates/chemistry , Electrochemistry/instrumentation , Lactic Acid/analysis , Lactic Acid/chemistry , Oxidoreductases/chemistry , Polyesters/chemistry , Biosensing Techniques/methods , Electrochemistry/methods , Enzymes, Immobilized/chemistry , Equipment Failure Analysis , Hydrogels/chemistry , Hydrogen-Ion Concentration , Multienzyme Complexes/chemistry , Reproducibility of Results , Sensitivity and Specificity
7.
Biosens Bioelectron ; 18(9): 1095-100, 2003 Aug 15.
Article in English | MEDLINE | ID: mdl-12788551

ABSTRACT

A biosensor for detection of formate at submicromolar concentrations has been developed by co-immobilizing formate dehydrogenase (FDH, E.C. 1.2.1.2), salicylate hydroxylase (SHL, E.C. 1.14.13.1) and NAD(+) linked to polyethylene glycol (PEG-NAD(+)) in a poly(vinyl alcohol) (PVA) matrix in front of a Clark-electrode. The principle of the bi-enzyme scheme is as follows: formate dehydrogenase converts formate into carbon dioxide using PEG-NAD(+). Corresponding PEG-NADH produced is then oxidized to PEG-NAD(+) by salicylate hydroxylase using sodium salicylate and oxygen. The oxygen consumption is monitored with the Clark-electrode. The advantages of this biosensor approach are the effective re-oxidation of PEG-NADH, and the entrapment of PEG-NAD(+) resulting in avoiding the addition of expensive cofactor to the working medium for each measurement. This bi-enzyme sensor has achieved a linear range of 1-300 microM and a detection limit of 1.98 x 10(-7) M for formate (S/N=3), with the response time of 4 min. The working stability is limited to 7 days due to the inactivation of the enzymes. Only sodium salicylate was needed in milli-molar amounts.


Subject(s)
Biosensing Techniques/instrumentation , Electrochemistry/instrumentation , Formate Dehydrogenases/chemistry , Formates/analysis , Microchemistry/instrumentation , Mixed Function Oxygenases/chemistry , NAD/chemistry , Biosensing Techniques/methods , Coenzymes/chemistry , Electrochemistry/methods , Enzyme Reactivators/chemistry , Enzyme Stability , Enzymes, Immobilized/chemistry , Equipment Design , Equipment Failure Analysis , Equipment Reuse , Feasibility Studies , Formates/chemistry , Microchemistry/methods , Multienzyme Complexes/chemistry , Reproducibility of Results , Sensitivity and Specificity
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