Chilled chicken is inevitably contaminated by microorganisms during slaughtering and processing,resulting in spoilage.Cutting parts of chilled chicken,especially wings,feet,and other skin-on products,are abundant in c...Chilled chicken is inevitably contaminated by microorganisms during slaughtering and processing,resulting in spoilage.Cutting parts of chilled chicken,especially wings,feet,and other skin-on products,are abundant in collagen,which may be the primary target for degradation by spoilage microorganisms.In this work,a total of 17 isolates of spoilage bacteria that could secrete both collagenase and lipase were determined by raw-chicken juice agar(RJA)method,and the results showed that 7 strains of Serratia,Aeromonas,and Pseudomonas could significantly decompose the collagen ingredients.The gelatin zymography showed that Serratia liquefaciens(F5)and(G7)had apparent degradation bands around 50 kDa,and Aeromonas veronii(G8)and Aeromonas salmonicida(H8)had a band around.65 and 95 kDa,respectively.The lipase and collagenase activities were detected isolate-by-isolate,with F5 showing the highest collagenase activity.For spoilage ability on meat in situ,F5 performed strongest in spoilage ability,indicated by the total viable counts,total volatile basic nitrogen content,sensory scores,lipase,and collagenase activity.This study provides a theoretical basis for spoilage heterogeneity of strains with high-producing collagenase in meat.展开更多
BACKGROUND Enzymatic fasciotomy with collagenase clostridium histolyticum(CCH)has revolutionized the treatment for Dupuytren’s contracture(DC).Despite its benefits,the long-term outcomes remain unclear.This study pre...BACKGROUND Enzymatic fasciotomy with collagenase clostridium histolyticum(CCH)has revolutionized the treatment for Dupuytren’s contracture(DC).Despite its benefits,the long-term outcomes remain unclear.This study presented a comprehensive 10-year follow-up assessment of the enduring effects of CCH on patients with DC.AIM To compare the short-term(12 wk)and long-term(10 years)outcomes on CCH treatment in patients with DC.METHODS A cohort of 45 patients was treated with CCH at the metacarpophalangeal(MCP)joint and the proximal interphalangeal(PIP)joint and underwent systematic reevaluation.The study adhered to multicenter trial protocols,and assessments were conducted at 12 wk,7 years,and 10 years post-surgery.RESULTS Thirty-seven patients completed the 10-year follow-up.At 10 years,patients treated at the PIP joint exhibited a 100%recurrence.However,patients treated at the MCP joint only showed a 50%recurrence.Patient satisfaction varied,with a lower satisfaction reported in PIP joint cases.Recurrence exceeding 20 degrees on the total passive extension deficit was observed,indicating a challenge for sustained efficacy.Significant differences were noted between outcomes at the 7-year and 10-year intervals.CONCLUSION CCH demonstrated sustained efficacy when applied to the MCP joint.However,caution is warranted for CCH treatment at the PIP joint due to a high level of recurrence and low patient satisfaction.Re-intervention is needed within a decade of treatment.展开更多
Background:Scar contractions caused by trauma or burns can cause secondary physical dysfunction and disfigurement.Many minimally invasive methods for scar contraction have shown limited applicability and efficacy.This...Background:Scar contractions caused by trauma or burns can cause secondary physical dysfunction and disfigurement.Many minimally invasive methods for scar contraction have shown limited applicability and efficacy.This study investigated the feasibility and efficacy of intralesional collagenase injections for scar contraction treatment.Methods:Patients with contracted scars who had limited joint movement and physical disfiguration for>1 year were enrolled in this single-blind,randomized clinical trial from July 2017 to February 2018 at Shanghai Ninth People’s Hospital.Collagenase was injected into the firm-contracted scar(15 U/cm^(2))three times at 4-week intervals in the multiple treatment group and once in the single treatment group,and a placebo injection was performed in the control group.Scar length and skin texture were documented at the 4-and 12-week follow-ups.The safety of the collagenase treatment was also evaluated.Results:The contracted scar was significantly elongated after both single and multiple collagenase treatments.The results showed that,compared to a one-time treatment,repeated injections were more effective at 12 weeks,with an average improvement of 26.83(15.79%).At 12 weeks,78.9% of the patients in the multiple group and 52.9%in the single group achieved significant improvement at 12 weeks.No severe adverse events were observed.Conclusion:Intralesional collagenase injection showed promising results in improving scar contraction and provides an alternative treatment for patients.展开更多
AIM： Type IV collagenase including MMP-2 and -9 plays an important role in cancer cell invasion and metastasis and is an attractive target for rnAb-directed therapy. The irnrnunoreactivity of rnAb 3G11, a rnAb direct...AIM： Type IV collagenase including MMP-2 and -9 plays an important role in cancer cell invasion and metastasis and is an attractive target for rnAb-directed therapy. The irnrnunoreactivity of rnAb 3G11, a rnAb directed against type Ⅳ collagenase in human colorectal carcinomas, was studied by irnrnuno-histochernical （IHC） staining, rnAb 3G11 was conjugated to an antiturnor antibiotic lidarnycin （LDM）. The antiturnor activity of 3G11-LDM conjugate against colon carcinoma was investigated in mice. METHODS： ELISA, gelatin zyrnography, and Western blot assay were used for the biological characterization of rnAb 3G11. The irnrnunoreactivity of rnAb 3G11 with human colorectal carcinomas was detected by IHC staining. The cytotoxicity of LDM and 3G11-LDM conjugate to human colon carcinoma HT-29 cells was examined by clonogenic assay and MTT assay. The therapeutic effect of conjugate 3G11-LDM was evaluated with colon carcinoma 26 in mice. RESULTS： As shown in ELISA, mAb 3Gll reacted specifically with type IV collagenase, while 3G11-LDM conjugate also recognized specifically its respective antigen. In IHC assay, mAb 3G11 showed positive irnrnunoreactivity in most cases of colorectal carcinoma, and negative irnrnunoreactivity in the adjacent non-malignant tissues. By gelatin zyrnography, the inhibition effect of rnAb 3G11 on the secretion activity of type IV collagenase was proved. In terms of IC50 values in MTT assay, the cytotoxicity of LDM to human colon carcinoma HT-29 cells was 10 000-fold more potent than that of rnitornycin C （MMC） and adriarnycin （ADM）. 3G11- LDM conjugate also displayed extremely potent cytotoxicity to human colon carcinoma HT-29 cells with an IC50 value of 5.6× 10^-19 mol/L. 3G11-LDM conjugate at the doses of 0.05 and 0.1 mg/kg inhibited the growth of colon carcinoma 26 in mice by 70.3 and 81.2%, respectively. CONCLUSION： mAb 3G11 is immunoreactive with human colorectal carcinoma and its conjugate with LDM is highly effective against colon carcinoma in mice.展开更多
Background: An ideal aneurysm model of cerebral aneurysm is of great importance for studying the pathogenesis of the lesion and testing new techniques for diagnosis and treatment. Several models have been created in r...Background: An ideal aneurysm model of cerebral aneurysm is of great importance for studying the pathogenesis of the lesion and testing new techniques for diagnosis and treatment. Several models have been created in rabbits and are now widely used in experimental studies; however, every model has certain intrinsic limitations. Here we report the development of a novel saccular aneurysm model in rabbits using an arterial pouch that is subject to in vitro pre-digestion with combined elastase and collagenase. Methods: A segment of right common carotid artery (CCA) was dissected out and treated with elastase (60 U/ml, 20 min) followed by type I collagenase (1 mg/ml, 15 min) in vitro. The graft was anastomosed to an arterial arch built with the left CCA and the remaining right CCA, while the other end of the graft was ligated. The dimension and tissue structure of the pouch were analysed immediately, 2 or 8 weeks after operation. Findings: Ten terminal aneurysms were produced. The gross mor-phology of the aneurysm resembles the human cerebral terminal aneurysms. We have observed the following pathological changes: (1) growth of the aneurysm (mean diameter increased from (2.0±0.1) to (3.2±0.3) mm at 2 weeks, P【0.001, n=7~10); (2) thinning of the aneurysmal wall (the mean wall thickness decreased to 44% at 2 weeks), which was accompanied by significant losses of elastic fibres, collagen and the cellular component; and (3) spontaneous rupture (3 out of 9, one aneurysm ruptured 24 h after operation with the other two at 2 and 4 weeks respectively). Conclusion: This rabbit arterial pouch model mimics human cerebral aneurysms in relation to morphology and histology. In particular, this model exhibited an increased tendency of spontaneous rupture.展开更多
The deleterious effect of vasculature damage on the outcome of spinal cord injury has long been recognized, and numerous clinical studies have shown that the presence of hemorrhage into the spinal cord is directly ass...The deleterious effect of vasculature damage on the outcome of spinal cord injury has long been recognized, and numerous clinical studies have shown that the presence of hemorrhage into the spinal cord is directly associated with a poorer neurological outcome. Vascular damage leads to de- creased blood flow to the cord and the release of potentially toxic blood-borne components. Here we consider the mechanisms that may be contributing to hemorrhage-induced damage and discuss the utility of a new model of spinal cord hemorrhage, which was urgently required as most of our current understanding has been extrapolated from intracerebral hemorrhage studies.展开更多
Collagenase was isolated from fish waste (a mixture of haddock, herring, ground fish and flounder) using a Tris-buffer system. The proteins in the crude extract were precipitated using ammonium sulfate (40% - 80%) and...Collagenase was isolated from fish waste (a mixture of haddock, herring, ground fish and flounder) using a Tris-buffer system. The proteins in the crude extract were precipitated using ammonium sulfate (40% - 80%) and purified with gel-filtration chromatography using Sephadex G-100. The results showed that the collagenase enzyme was produced as a latent enzyme and was activated with bovine trypsin and potassium thiocyanate (KSCN). The enzyme activity was affected by pH and temperature. Optimal enzyme activities were found at 35?C and a pH of 7.5 when insoluble collagene type I was used as substrate and the liberated amino acids were measured in relation to L-leucine in the presence of ninhydrin. The enzyme activity was completely inhibited by the action of ethylenediaminetetraacetic acid (EDTA) suggesting that the collagenase enzyme isolated from the fish waste is a metalloproteinase enzyme requiring metal ions for enzyme activity. Dialysis against KSCN decreased the enzyme total activity and increased its specific activity. Sodium dodecyl sulphate polyacryla-mide gel electrophoresis (SDS-PAGE) indicated that the purified procollagenase enzyme have only one band at molecular weight of 50 kilodaltons (kDa). When the enzyme was cleaved with trypsin, it was found to consist of two subunits: a large unit with a molecular weight of 50 kDa and a small unit with a molecular weight of 10 kDa.展开更多
The aim of this study was to determine the role of neutrophil collagenase in the pathogenesis of acute lung injury induced by endotoxin. 28 Sprague-Dawley were randomized into control group and LPS-enduced groups. Sam...The aim of this study was to determine the role of neutrophil collagenase in the pathogenesis of acute lung injury induced by endotoxin. 28 Sprague-Dawley were randomized into control group and LPS-enduced groups. Samples of left lung were obtained in 2 h (group L 1), 6 h (group L 2), 12 h (group L 3) after intravenous LPS. Immunohistochemsitry was employed for detection of expression of neutrophil collagenase. Pathological scores, lung wet/dry weight ratio and the number of neutrophils were measured. The results showed that the concentration of neutrophil collagenase in LPS-enduced groups (group L 1, L 2, L 3) were significantly higher than that of control group (P<0.01). Pathological scores, lung wet/dry weight ratio and the number of neutrophils in LPS-enduced groups (group L 1, L 2, L 3) were also significantly higher than that of control group (P<0.01). Moreover, among group L 1, L 2 and L 3, there were significant correlations in concentration of neutrophil collagenase and pathological scores, lung wet/dry weight ratio, the number of neutrophils (P<0.05). The present study showed that neutrophil collagenase play an important role in the pathogenesis and progress of endotoxic acute lung injury.展开更多
Clostridium histolyticum is used for production of several proteolytic enzymes such as elastase, neutral proteases, clostripain and in particular collagenase. Besides industrial applications, collagenase has been indi...Clostridium histolyticum is used for production of several proteolytic enzymes such as elastase, neutral proteases, clostripain and in particular collagenase. Besides industrial applications, collagenase has been indispensable for medical purposes including isolation of pancreatic islets for diabetes treatment. The aim of this study was to optimize the method for production and partial purification of a new collagenase blend and to test its suitability for successful pancreatic islets isolation in a rat model. Bacterial strain of C. histolyticum was sequenced for presence of the collagenase genes. Different fermentation conditions were tested and the process of collagenase extraction was modified and optimized. Samples of collagenases were taken for western blot detection, activity assessment, and ability for dissociation of pancreatic tissue. Findings indicate that concentrated trypton growth medium with pepton was the most suitable for Clostridium growth and collagenase production. Whole genome sequencing revealed two genes for collagenase and also gene for clostripain. Western blot specific detection helped to select useful production modifications. Following these modifications was also improved the yield, morphology and in vitro function of intact pancreatic islets which were finally comparable or better than those achieved using standard blends of collagenase. The results support the use of the new collagenase blend for islet isolation giving thus the opportunity to choose an alternative product. Our next steps would lead to further enzyme purification through scaling up of the production method for a wider use.展开更多
The epidermal collagenase activity and its induction by 20-hydroxyecdysone in Uca pugilator were investigated. Zymographic electrophoresis showed four bands of collagenase activity, 16, 19, 22 and 29 kDa in molecular ...The epidermal collagenase activity and its induction by 20-hydroxyecdysone in Uca pugilator were investigated. Zymographic electrophoresis showed four bands of collagenase activity, 16, 19, 22 and 29 kDa in molecular weight, with the former two accounting for 60% and 36% , respectively, of the total collagenase activity. The collagenase activity varies during the molting cycle. Among the molt stages tested, Premolt Stage Do exhibited the highest epidermal collagenase activity for both the 16 and 19 kDa isoenzymes and, as the molt stage proceeded, the enzymatic activity of these two isoenzymes decreased, with the lowest activity for both found in Premolt Stage D3 -4 - Injection of 20-hydroxyecdysone significantly induced the activity of the 16 kDa eollagenase in the epidermis of Uca pugilator, suggesting that the activity of this isoenzyme is under molting hormone control. Although 20-hydroxyeedysone injection did not result in a statistically significant increase in the activity of the 19 kDa isoenzyme, a tendency of the induction was nonetheless demonstrated. This is the first report on epidermal collagenase activity and its induction by the molting hormone in a crustacean .展开更多
Non-invasive therapeutic methods have recently been used in medical sciences. Enzymes have shown high activity at very low concentrations in laboratories and pharmaceutical,enabling them to play crucial roles in diffe...Non-invasive therapeutic methods have recently been used in medical sciences. Enzymes have shown high activity at very low concentrations in laboratories and pharmaceutical,enabling them to play crucial roles in different biological phenomena related to living organism, especially human medicine. Recently, using the therapeutic methods based on non-invasive approaches has been emphasized in medical society. Researchers have focused on producing medicines and tools reducing invasive procedures in medical.Collagenases are proteins which catalyze chemical processes and break the peptide bonds in collagen. Collagen may be generated more than the required amount or produced in unsuitable sites or may not degrade after a certain time. In such cases, using an injectable collagenase or its ointment can be helpful in collagen degradation. In both in vitro and in vivo tests, it has been revealed that collagenases have several therapeutic properties in wound healing, burns, nipple pain and some diseases including intervertebral disc herniation, keloid, cellulite, lipoma among others. This review describes the therapeutic application of collagenase in medical sciences and the process for its production using novel methods, paving the way for more effective and safe applications of collagenases.展开更多
A selective, sensitive, and convenient assay for human collagenase is required because of its implication in diseases such as rheumatoid arthritis, osteoarthritis, and tumors. Here, a novel assay for human collagenase...A selective, sensitive, and convenient assay for human collagenase is required because of its implication in diseases such as rheumatoid arthritis, osteoarthritis, and tumors. Here, a novel assay for human collagenase activity is described in which enzymatic degradation of native collagen or acetyl peptide is determined by using a fluorogenic reaction for oligopeptides. The oligopeptides are quantified spectrofluorometrically with either 3,4-dihydroxyphenylacetic acid or 1,2-dihydroxybenzen reaction in the presence of sodium periodate and sodium borate (pH 7 - 8). These reactions can selectively convert N-terminal Gly-containing oligopeptides and N-terminal Ile-containing oligopeptides to fluorescence (FL) compounds, respectively, but not proteins, acetyl peptides or amino acids. Under optimized conditions using 1.65 μM collagen IV or 1.5 mM Ac-GPQGI- AGQ as substrates, this assay exhibits a proportional relationship between FL intensities and human collagenase-3 (MMP-13) concentrations. It can assay endogenous collagenase activities in several biological samples, such as cultured human cells and cheek tissue.展开更多
AIM:To review the relevant literature in an effort to examine the body of evidence available to date.METHODS:Ovid MEDLINE search database was queried using MeS H terms"penile induration","peyronie’s di...AIM:To review the relevant literature in an effort to examine the body of evidence available to date.METHODS:Ovid MEDLINE search database was queried using MeS H terms"penile induration","peyronie’s disease","Collagenases"and"Collagenase"using various permutations.No temporal parameters were employed.RESULTS:In all,5 relevant clinical trials were isolated from 34 results.These trials were analyzed using the Oxford Centre for Evidence-Based Medicine criteria.They were further examined based on study design and methods;the primary and secondary outcomes were reviewed for treatment efficacy and collagenase-related side effects.CONCLUSION:Intralesional collagenase appears to be safe and effective in the non-surgical treatment of Peyronie’s disease.However,the data remains limited and further inquiries into the safety of collagenase,treatment standardization and standardized outcomesreporting remain necessary.Furthermore,studies comparing intralesional collagenase to alternative medical and surgical therapy will be important in guiding the future treatment decision process.展开更多
Hide and skin are complex tissue where the most abundant component is collagen.Matrix metalloproteinases and bacterial collagenases are two kinds of collagenases that can cleave the triple-helical domain of native fib...Hide and skin are complex tissue where the most abundant component is collagen.Matrix metalloproteinases and bacterial collagenases are two kinds of collagenases that can cleave the triple-helical domain of native fibrillar collagens.In this paper,the family members and domain composition of matrix metalloproteinases and bacterial collagenases are summarized.The catalytic mechanism of collagen hydrolysis by collagenases is described,and the methods adopted to date for investigating and regulating collagenases and their inhibitors are reviewed.Furthermore,the applications of collagenases and their inhibitors in biomedicine,food processing and the enzymatic unhairing process in the leather-making industry are presented.展开更多
Type IV collagenase plays an important role in tumor invasion and metastasis through cleaving type IV collagen in the basement membrane and extracellular matrix. In this study a molecule-downsized immunoconjugate (Fab...Type IV collagenase plays an important role in tumor invasion and metastasis through cleaving type IV collagen in the basement membrane and extracellular matrix. In this study a molecule-downsized immunoconjugate (Fab′-LDM) was constructed by linking lidamycin (LDM), a highly potent antitumor antibiotic, to the Fab′ fragment of a monoclonal antibody directed against type IV collagenase and its antitumor effect was investigated. As assayed in 10% SDS-PAGE gel, the molecular weight of Fab′-LDM conjugate was 65 kD with a 1:1 molecular ratio of Fab′ and LDM. The Fab′-LDM conjugate maintained most part of the immunoreactivity of Fab′ fragment to both type IV collagense and mouse hepatoma 22 cells by ELISA. By MTT assay, Fab′-LDM conjugate showed more potent cytotoxicity to hepatoma 22 cells than that of LDM. Administered intravenously, Fab′-LDM conjugate proved to be more effective against the growth of subcutaneously transplanted hepatoma 22 in mice than free LDM in two experiment settings. In Experiment I, the drugs were given intravenously on day 1 and day 8. Fab′-LDM at the doses of 0.025 mg/kg, 0.05 mg/kg and 0.1 mg/kg inhibited tumor growth by 76.7%, 93.3% and 94.8%, while free LDM at 0.05 mg/kg inhibited tumor growth by 76.1%, respectively. In experiment II, the drugs were given intravenously on day 4 and day 11, Fab′-LDM at the doses of 0.025 mg/kg and 0.05 mg/kg inhibited tumor growth by 74.2%, 80.9%, while free LDM at 0.05 mg/kg inhibited tumor growth by 60.5%, respectively. In terms of survival time, Fab′-LDM was more effective than free LDM. The results suggest that the molecule-downsized immunoconjugate directed against type IV collagenase is of high efficacy in experimental cancer therapy.展开更多
The over\|expression of collagenase IV in tumor tissues was found to be closely related to tumor metastasis. Collagenase IV has been therefore considered as one of the novel indicative molecules for tumor diagnosis an...The over\|expression of collagenase IV in tumor tissues was found to be closely related to tumor metastasis. Collagenase IV has been therefore considered as one of the novel indicative molecules for tumor diagnosis and treatment. Based on phage display antibody library technique, a single\|chain Fv specific for collagenase IV was successfully cloned. This antibody, referred to as hCo4, was mainly composed of variable regions from heavy and light chains, with its molecular weight of 27 ku. The engineered antibody bound to collagenase IV specifically. The affinity of hCo4 was found to be the same as that of a single\|chain antibody constructed from a monoclonal antibody to collagenase IV. Since hCo4 is the smallest among all the antibodies specific for collagenase IV and it is of human origin, it has a potential to be applied for tumor immunotherapy and for the study of the relationship between collagenase IV and tumor metastasis.展开更多
It was documented that type IV collagenase with two subtypes of 72 ku/MMP-2 and 92 ku/MMP-9 plays an important role in tumor invasion and metastasis. The endoplasmic reticulum (ER)- retained, single chain Fv antibody ...It was documented that type IV collagenase with two subtypes of 72 ku/MMP-2 and 92 ku/MMP-9 plays an important role in tumor invasion and metastasis. The endoplasmic reticulum (ER)- retained, single chain Fv antibody fragment (scFv) was used to inhibit the function of type IV collagenase. For expression in mammalian cells, the assembled scFv M97 gene with ER retention signal encoding 6 additional amino acids (SEKDEL) was reamplified by PCR. The amplified fragments were cloned into the pcDNA3.1 vector. The resulting plasmid was sequenced and then introduced into PG cells, a highly metastatic human lung cancer cell line, by lipofectAMINE method. The result of intrabody gene therapy showed that type IV collegenase expression was down regulated significantly as measured by ELISA. The biological behavior of PG cell, such as the ability of in vitro invasion through Matrigel, colony formation on soft agar, was also inhibited by scFv M97 transfection. Animal experiments in a xenograft model of human lung cancer showed that scFv M97 transfection significantly prolonged the survival time of nude mice. The results indicate that intracellular antibody technology represents a novel and efficient way to abrogate selectively the activity of type IV collagenase.展开更多
AIM:Overexpression of mucosal metalloproteinases(MMP) has been demonstrated recently in inflammatory bowel disease.Their activity can be counterbalanced by the tissue inhibitor of metalloproteinases(TIMP).The aim of t...AIM:Overexpression of mucosal metalloproteinases(MMP) has been demonstrated recently in inflammatory bowel disease.Their activity can be counterbalanced by the tissue inhibitor of metalloproteinases(TIMP).The aim of this study was to evaluate the effect of ulcerative colitis(UC)on MMP- 1 and TIMP-1 plasma concentrations,as two possible biomarkers of the disease activity. METHODS:MMP-1 and TIMP-1 plasma concentrations were measured with an enzyme immunoassay in 16 patients with endoscopically confirmed active UC. RESULTS:Plasma concentrations of both MMP-1(13.7±0.2 ng/ml)and TIMP-1(799±140 ng/ml)were significantly elevated in UC patients in comparison to healthy controls (11.9±0.9 ng/ml and 220±7 ng/ml respectively).There was no correlation between TIMP-1 and MMP-1 concentrations (r=0.02).TIMP-1 levels revealed significant positive correlations with scored endoscopic degree of mucosal injury, disease activity index and clinical activity index values as well as C-reactive protein concentration.There was no correlation between MMP-1 and laboratory,clinical or endoscopic indices of the disease activity.CONCLUSION: These results confirm the role of both MMP- 1 and TIMP-1 in the pathogenesis of ulcerative colitis. However only TIMP-1 can be useful as a biomarker of the disease activity, demonstrating association with clinical and endoscopic pictures.展开更多
基金financed by grants from the Natural Science Foundation of Jiangsu Province in China (BK20221515)the National Natural Science Foundation of China (32172266)。
文摘Chilled chicken is inevitably contaminated by microorganisms during slaughtering and processing,resulting in spoilage.Cutting parts of chilled chicken,especially wings,feet,and other skin-on products,are abundant in collagen,which may be the primary target for degradation by spoilage microorganisms.In this work,a total of 17 isolates of spoilage bacteria that could secrete both collagenase and lipase were determined by raw-chicken juice agar(RJA)method,and the results showed that 7 strains of Serratia,Aeromonas,and Pseudomonas could significantly decompose the collagen ingredients.The gelatin zymography showed that Serratia liquefaciens(F5)and(G7)had apparent degradation bands around 50 kDa,and Aeromonas veronii(G8)and Aeromonas salmonicida(H8)had a band around.65 and 95 kDa,respectively.The lipase and collagenase activities were detected isolate-by-isolate,with F5 showing the highest collagenase activity.For spoilage ability on meat in situ,F5 performed strongest in spoilage ability,indicated by the total viable counts,total volatile basic nitrogen content,sensory scores,lipase,and collagenase activity.This study provides a theoretical basis for spoilage heterogeneity of strains with high-producing collagenase in meat.
文摘BACKGROUND Enzymatic fasciotomy with collagenase clostridium histolyticum(CCH)has revolutionized the treatment for Dupuytren’s contracture(DC).Despite its benefits,the long-term outcomes remain unclear.This study presented a comprehensive 10-year follow-up assessment of the enduring effects of CCH on patients with DC.AIM To compare the short-term(12 wk)and long-term(10 years)outcomes on CCH treatment in patients with DC.METHODS A cohort of 45 patients was treated with CCH at the metacarpophalangeal(MCP)joint and the proximal interphalangeal(PIP)joint and underwent systematic reevaluation.The study adhered to multicenter trial protocols,and assessments were conducted at 12 wk,7 years,and 10 years post-surgery.RESULTS Thirty-seven patients completed the 10-year follow-up.At 10 years,patients treated at the PIP joint exhibited a 100%recurrence.However,patients treated at the MCP joint only showed a 50%recurrence.Patient satisfaction varied,with a lower satisfaction reported in PIP joint cases.Recurrence exceeding 20 degrees on the total passive extension deficit was observed,indicating a challenge for sustained efficacy.Significant differences were noted between outcomes at the 7-year and 10-year intervals.CONCLUSION CCH demonstrated sustained efficacy when applied to the MCP joint.However,caution is warranted for CCH treatment at the PIP joint due to a high level of recurrence and low patient satisfaction.Re-intervention is needed within a decade of treatment.
基金supported by the National Natural Science Foundation of China(grant nos.81501678,81971848,and 82272287)Clinical Research Plan of Shanghai Hospital Development Center(grant nos.SHDC2020CR1019B and SHDC2020CR4029)+1 种基金Shanghai Municipal Key Clinical Specialty(grant no.shslczdzk00901)Innovative Research Team of High-Level Local University in Shanghai(grant no.SSMUZDCX20180700).
文摘Background:Scar contractions caused by trauma or burns can cause secondary physical dysfunction and disfigurement.Many minimally invasive methods for scar contraction have shown limited applicability and efficacy.This study investigated the feasibility and efficacy of intralesional collagenase injections for scar contraction treatment.Methods:Patients with contracted scars who had limited joint movement and physical disfiguration for>1 year were enrolled in this single-blind,randomized clinical trial from July 2017 to February 2018 at Shanghai Ninth People’s Hospital.Collagenase was injected into the firm-contracted scar(15 U/cm^(2))three times at 4-week intervals in the multiple treatment group and once in the single treatment group,and a placebo injection was performed in the control group.Scar length and skin texture were documented at the 4-and 12-week follow-ups.The safety of the collagenase treatment was also evaluated.Results:The contracted scar was significantly elongated after both single and multiple collagenase treatments.The results showed that,compared to a one-time treatment,repeated injections were more effective at 12 weeks,with an average improvement of 26.83(15.79%).At 12 weeks,78.9% of the patients in the multiple group and 52.9%in the single group achieved significant improvement at 12 weeks.No severe adverse events were observed.Conclusion:Intralesional collagenase injection showed promising results in improving scar contraction and provides an alternative treatment for patients.
基金Supported by the National High Technology Research and Development Program of China, 863 Program, No. 2002AA2Z346D
文摘AIM： Type IV collagenase including MMP-2 and -9 plays an important role in cancer cell invasion and metastasis and is an attractive target for rnAb-directed therapy. The irnrnunoreactivity of rnAb 3G11, a rnAb directed against type Ⅳ collagenase in human colorectal carcinomas, was studied by irnrnuno-histochernical （IHC） staining, rnAb 3G11 was conjugated to an antiturnor antibiotic lidarnycin （LDM）. The antiturnor activity of 3G11-LDM conjugate against colon carcinoma was investigated in mice. METHODS： ELISA, gelatin zyrnography, and Western blot assay were used for the biological characterization of rnAb 3G11. The irnrnunoreactivity of rnAb 3G11 with human colorectal carcinomas was detected by IHC staining. The cytotoxicity of LDM and 3G11-LDM conjugate to human colon carcinoma HT-29 cells was examined by clonogenic assay and MTT assay. The therapeutic effect of conjugate 3G11-LDM was evaluated with colon carcinoma 26 in mice. RESULTS： As shown in ELISA, mAb 3Gll reacted specifically with type IV collagenase, while 3G11-LDM conjugate also recognized specifically its respective antigen. In IHC assay, mAb 3G11 showed positive irnrnunoreactivity in most cases of colorectal carcinoma, and negative irnrnunoreactivity in the adjacent non-malignant tissues. By gelatin zyrnography, the inhibition effect of rnAb 3G11 on the secretion activity of type IV collagenase was proved. In terms of IC50 values in MTT assay, the cytotoxicity of LDM to human colon carcinoma HT-29 cells was 10 000-fold more potent than that of rnitornycin C （MMC） and adriarnycin （ADM）. 3G11- LDM conjugate also displayed extremely potent cytotoxicity to human colon carcinoma HT-29 cells with an IC50 value of 5.6× 10^-19 mol/L. 3G11-LDM conjugate at the doses of 0.05 and 0.1 mg/kg inhibited the growth of colon carcinoma 26 in mice by 70.3 and 81.2%, respectively. CONCLUSION： mAb 3G11 is immunoreactive with human colorectal carcinoma and its conjugate with LDM is highly effective against colon carcinoma in mice.
基金the National Natural Science Foundation of China (No. 30772234)the Natural Science Foundation of Beijing(Nos. 7022008 and 7072016)the Outstanding Talent Foundation of the Organization Department of Municipal Party Committee of Beijing (No. 2006D0300400072), China
文摘Background: An ideal aneurysm model of cerebral aneurysm is of great importance for studying the pathogenesis of the lesion and testing new techniques for diagnosis and treatment. Several models have been created in rabbits and are now widely used in experimental studies; however, every model has certain intrinsic limitations. Here we report the development of a novel saccular aneurysm model in rabbits using an arterial pouch that is subject to in vitro pre-digestion with combined elastase and collagenase. Methods: A segment of right common carotid artery (CCA) was dissected out and treated with elastase (60 U/ml, 20 min) followed by type I collagenase (1 mg/ml, 15 min) in vitro. The graft was anastomosed to an arterial arch built with the left CCA and the remaining right CCA, while the other end of the graft was ligated. The dimension and tissue structure of the pouch were analysed immediately, 2 or 8 weeks after operation. Findings: Ten terminal aneurysms were produced. The gross mor-phology of the aneurysm resembles the human cerebral terminal aneurysms. We have observed the following pathological changes: (1) growth of the aneurysm (mean diameter increased from (2.0±0.1) to (3.2±0.3) mm at 2 weeks, P【0.001, n=7~10); (2) thinning of the aneurysmal wall (the mean wall thickness decreased to 44% at 2 weeks), which was accompanied by significant losses of elastic fibres, collagen and the cellular component; and (3) spontaneous rupture (3 out of 9, one aneurysm ruptured 24 h after operation with the other two at 2 and 4 weeks respectively). Conclusion: This rabbit arterial pouch model mimics human cerebral aneurysms in relation to morphology and histology. In particular, this model exhibited an increased tendency of spontaneous rupture.
基金supported by a grant from Medical Research Council,No.MRCG0300456
文摘The deleterious effect of vasculature damage on the outcome of spinal cord injury has long been recognized, and numerous clinical studies have shown that the presence of hemorrhage into the spinal cord is directly associated with a poorer neurological outcome. Vascular damage leads to de- creased blood flow to the cord and the release of potentially toxic blood-borne components. Here we consider the mechanisms that may be contributing to hemorrhage-induced damage and discuss the utility of a new model of spinal cord hemorrhage, which was urgently required as most of our current understanding has been extrapolated from intracerebral hemorrhage studies.
文摘Collagenase was isolated from fish waste (a mixture of haddock, herring, ground fish and flounder) using a Tris-buffer system. The proteins in the crude extract were precipitated using ammonium sulfate (40% - 80%) and purified with gel-filtration chromatography using Sephadex G-100. The results showed that the collagenase enzyme was produced as a latent enzyme and was activated with bovine trypsin and potassium thiocyanate (KSCN). The enzyme activity was affected by pH and temperature. Optimal enzyme activities were found at 35?C and a pH of 7.5 when insoluble collagene type I was used as substrate and the liberated amino acids were measured in relation to L-leucine in the presence of ninhydrin. The enzyme activity was completely inhibited by the action of ethylenediaminetetraacetic acid (EDTA) suggesting that the collagenase enzyme isolated from the fish waste is a metalloproteinase enzyme requiring metal ions for enzyme activity. Dialysis against KSCN decreased the enzyme total activity and increased its specific activity. Sodium dodecyl sulphate polyacryla-mide gel electrophoresis (SDS-PAGE) indicated that the purified procollagenase enzyme have only one band at molecular weight of 50 kilodaltons (kDa). When the enzyme was cleaved with trypsin, it was found to consist of two subunits: a large unit with a molecular weight of 50 kDa and a small unit with a molecular weight of 10 kDa.
文摘The aim of this study was to determine the role of neutrophil collagenase in the pathogenesis of acute lung injury induced by endotoxin. 28 Sprague-Dawley were randomized into control group and LPS-enduced groups. Samples of left lung were obtained in 2 h (group L 1), 6 h (group L 2), 12 h (group L 3) after intravenous LPS. Immunohistochemsitry was employed for detection of expression of neutrophil collagenase. Pathological scores, lung wet/dry weight ratio and the number of neutrophils were measured. The results showed that the concentration of neutrophil collagenase in LPS-enduced groups (group L 1, L 2, L 3) were significantly higher than that of control group (P<0.01). Pathological scores, lung wet/dry weight ratio and the number of neutrophils in LPS-enduced groups (group L 1, L 2, L 3) were also significantly higher than that of control group (P<0.01). Moreover, among group L 1, L 2 and L 3, there were significant correlations in concentration of neutrophil collagenase and pathological scores, lung wet/dry weight ratio, the number of neutrophils (P<0.05). The present study showed that neutrophil collagenase play an important role in the pathogenesis and progress of endotoxic acute lung injury.
基金supported by MH CZ-DRO(“Institute for Clinical and Experimental Medicine—IKEM,IN 00023001”)by grant no.TA04010038 from Technology Agency of the Czech Republicby grant no.FV20139 from Ministry of Industry and Trade.
文摘Clostridium histolyticum is used for production of several proteolytic enzymes such as elastase, neutral proteases, clostripain and in particular collagenase. Besides industrial applications, collagenase has been indispensable for medical purposes including isolation of pancreatic islets for diabetes treatment. The aim of this study was to optimize the method for production and partial purification of a new collagenase blend and to test its suitability for successful pancreatic islets isolation in a rat model. Bacterial strain of C. histolyticum was sequenced for presence of the collagenase genes. Different fermentation conditions were tested and the process of collagenase extraction was modified and optimized. Samples of collagenases were taken for western blot detection, activity assessment, and ability for dissociation of pancreatic tissue. Findings indicate that concentrated trypton growth medium with pepton was the most suitable for Clostridium growth and collagenase production. Whole genome sequencing revealed two genes for collagenase and also gene for clostripain. Western blot specific detection helped to select useful production modifications. Following these modifications was also improved the yield, morphology and in vitro function of intact pancreatic islets which were finally comparable or better than those achieved using standard blends of collagenase. The results support the use of the new collagenase blend for islet isolation giving thus the opportunity to choose an alternative product. Our next steps would lead to further enzyme purification through scaling up of the production method for a wider use.
文摘The epidermal collagenase activity and its induction by 20-hydroxyecdysone in Uca pugilator were investigated. Zymographic electrophoresis showed four bands of collagenase activity, 16, 19, 22 and 29 kDa in molecular weight, with the former two accounting for 60% and 36% , respectively, of the total collagenase activity. The collagenase activity varies during the molting cycle. Among the molt stages tested, Premolt Stage Do exhibited the highest epidermal collagenase activity for both the 16 and 19 kDa isoenzymes and, as the molt stage proceeded, the enzymatic activity of these two isoenzymes decreased, with the lowest activity for both found in Premolt Stage D3 -4 - Injection of 20-hydroxyecdysone significantly induced the activity of the 16 kDa eollagenase in the epidermis of Uca pugilator, suggesting that the activity of this isoenzyme is under molting hormone control. Although 20-hydroxyeedysone injection did not result in a statistically significant increase in the activity of the 19 kDa isoenzyme, a tendency of the induction was nonetheless demonstrated. This is the first report on epidermal collagenase activity and its induction by the molting hormone in a crustacean .
基金Supported by Pasteur Institute of Iran and Iran National Science Foundation(Grant No.93011174)
文摘Non-invasive therapeutic methods have recently been used in medical sciences. Enzymes have shown high activity at very low concentrations in laboratories and pharmaceutical,enabling them to play crucial roles in different biological phenomena related to living organism, especially human medicine. Recently, using the therapeutic methods based on non-invasive approaches has been emphasized in medical society. Researchers have focused on producing medicines and tools reducing invasive procedures in medical.Collagenases are proteins which catalyze chemical processes and break the peptide bonds in collagen. Collagen may be generated more than the required amount or produced in unsuitable sites or may not degrade after a certain time. In such cases, using an injectable collagenase or its ointment can be helpful in collagen degradation. In both in vitro and in vivo tests, it has been revealed that collagenases have several therapeutic properties in wound healing, burns, nipple pain and some diseases including intervertebral disc herniation, keloid, cellulite, lipoma among others. This review describes the therapeutic application of collagenase in medical sciences and the process for its production using novel methods, paving the way for more effective and safe applications of collagenases.
文摘A selective, sensitive, and convenient assay for human collagenase is required because of its implication in diseases such as rheumatoid arthritis, osteoarthritis, and tumors. Here, a novel assay for human collagenase activity is described in which enzymatic degradation of native collagen or acetyl peptide is determined by using a fluorogenic reaction for oligopeptides. The oligopeptides are quantified spectrofluorometrically with either 3,4-dihydroxyphenylacetic acid or 1,2-dihydroxybenzen reaction in the presence of sodium periodate and sodium borate (pH 7 - 8). These reactions can selectively convert N-terminal Gly-containing oligopeptides and N-terminal Ile-containing oligopeptides to fluorescence (FL) compounds, respectively, but not proteins, acetyl peptides or amino acids. Under optimized conditions using 1.65 μM collagen IV or 1.5 mM Ac-GPQGI- AGQ as substrates, this assay exhibits a proportional relationship between FL intensities and human collagenase-3 (MMP-13) concentrations. It can assay endogenous collagenase activities in several biological samples, such as cultured human cells and cheek tissue.
文摘AIM:To review the relevant literature in an effort to examine the body of evidence available to date.METHODS:Ovid MEDLINE search database was queried using MeS H terms"penile induration","peyronie’s disease","Collagenases"and"Collagenase"using various permutations.No temporal parameters were employed.RESULTS:In all,5 relevant clinical trials were isolated from 34 results.These trials were analyzed using the Oxford Centre for Evidence-Based Medicine criteria.They were further examined based on study design and methods;the primary and secondary outcomes were reviewed for treatment efficacy and collagenase-related side effects.CONCLUSION:Intralesional collagenase appears to be safe and effective in the non-surgical treatment of Peyronie’s disease.However,the data remains limited and further inquiries into the safety of collagenase,treatment standardization and standardized outcomesreporting remain necessary.Furthermore,studies comparing intralesional collagenase to alternative medical and surgical therapy will be important in guiding the future treatment decision process.
基金support of this study was from the National Natural Science Foundation of China(No.22178231).
文摘Hide and skin are complex tissue where the most abundant component is collagen.Matrix metalloproteinases and bacterial collagenases are two kinds of collagenases that can cleave the triple-helical domain of native fibrillar collagens.In this paper,the family members and domain composition of matrix metalloproteinases and bacterial collagenases are summarized.The catalytic mechanism of collagen hydrolysis by collagenases is described,and the methods adopted to date for investigating and regulating collagenases and their inhibitors are reviewed.Furthermore,the applications of collagenases and their inhibitors in biomedicine,food processing and the enzymatic unhairing process in the leather-making industry are presented.
文摘Type IV collagenase plays an important role in tumor invasion and metastasis through cleaving type IV collagen in the basement membrane and extracellular matrix. In this study a molecule-downsized immunoconjugate (Fab′-LDM) was constructed by linking lidamycin (LDM), a highly potent antitumor antibiotic, to the Fab′ fragment of a monoclonal antibody directed against type IV collagenase and its antitumor effect was investigated. As assayed in 10% SDS-PAGE gel, the molecular weight of Fab′-LDM conjugate was 65 kD with a 1:1 molecular ratio of Fab′ and LDM. The Fab′-LDM conjugate maintained most part of the immunoreactivity of Fab′ fragment to both type IV collagense and mouse hepatoma 22 cells by ELISA. By MTT assay, Fab′-LDM conjugate showed more potent cytotoxicity to hepatoma 22 cells than that of LDM. Administered intravenously, Fab′-LDM conjugate proved to be more effective against the growth of subcutaneously transplanted hepatoma 22 in mice than free LDM in two experiment settings. In Experiment I, the drugs were given intravenously on day 1 and day 8. Fab′-LDM at the doses of 0.025 mg/kg, 0.05 mg/kg and 0.1 mg/kg inhibited tumor growth by 76.7%, 93.3% and 94.8%, while free LDM at 0.05 mg/kg inhibited tumor growth by 76.1%, respectively. In experiment II, the drugs were given intravenously on day 4 and day 11, Fab′-LDM at the doses of 0.025 mg/kg and 0.05 mg/kg inhibited tumor growth by 74.2%, 80.9%, while free LDM at 0.05 mg/kg inhibited tumor growth by 60.5%, respectively. In terms of survival time, Fab′-LDM was more effective than free LDM. The results suggest that the molecule-downsized immunoconjugate directed against type IV collagenase is of high efficacy in experimental cancer therapy.
文摘The over\|expression of collagenase IV in tumor tissues was found to be closely related to tumor metastasis. Collagenase IV has been therefore considered as one of the novel indicative molecules for tumor diagnosis and treatment. Based on phage display antibody library technique, a single\|chain Fv specific for collagenase IV was successfully cloned. This antibody, referred to as hCo4, was mainly composed of variable regions from heavy and light chains, with its molecular weight of 27 ku. The engineered antibody bound to collagenase IV specifically. The affinity of hCo4 was found to be the same as that of a single\|chain antibody constructed from a monoclonal antibody to collagenase IV. Since hCo4 is the smallest among all the antibodies specific for collagenase IV and it is of human origin, it has a potential to be applied for tumor immunotherapy and for the study of the relationship between collagenase IV and tumor metastasis.
文摘It was documented that type IV collagenase with two subtypes of 72 ku/MMP-2 and 92 ku/MMP-9 plays an important role in tumor invasion and metastasis. The endoplasmic reticulum (ER)- retained, single chain Fv antibody fragment (scFv) was used to inhibit the function of type IV collagenase. For expression in mammalian cells, the assembled scFv M97 gene with ER retention signal encoding 6 additional amino acids (SEKDEL) was reamplified by PCR. The amplified fragments were cloned into the pcDNA3.1 vector. The resulting plasmid was sequenced and then introduced into PG cells, a highly metastatic human lung cancer cell line, by lipofectAMINE method. The result of intrabody gene therapy showed that type IV collegenase expression was down regulated significantly as measured by ELISA. The biological behavior of PG cell, such as the ability of in vitro invasion through Matrigel, colony formation on soft agar, was also inhibited by scFv M97 transfection. Animal experiments in a xenograft model of human lung cancer showed that scFv M97 transfection significantly prolonged the survival time of nude mice. The results indicate that intracellular antibody technology represents a novel and efficient way to abrogate selectively the activity of type IV collagenase.
文摘AIM:Overexpression of mucosal metalloproteinases(MMP) has been demonstrated recently in inflammatory bowel disease.Their activity can be counterbalanced by the tissue inhibitor of metalloproteinases(TIMP).The aim of this study was to evaluate the effect of ulcerative colitis(UC)on MMP- 1 and TIMP-1 plasma concentrations,as two possible biomarkers of the disease activity. METHODS:MMP-1 and TIMP-1 plasma concentrations were measured with an enzyme immunoassay in 16 patients with endoscopically confirmed active UC. RESULTS:Plasma concentrations of both MMP-1(13.7±0.2 ng/ml)and TIMP-1(799±140 ng/ml)were significantly elevated in UC patients in comparison to healthy controls (11.9±0.9 ng/ml and 220±7 ng/ml respectively).There was no correlation between TIMP-1 and MMP-1 concentrations (r=0.02).TIMP-1 levels revealed significant positive correlations with scored endoscopic degree of mucosal injury, disease activity index and clinical activity index values as well as C-reactive protein concentration.There was no correlation between MMP-1 and laboratory,clinical or endoscopic indices of the disease activity.CONCLUSION: These results confirm the role of both MMP- 1 and TIMP-1 in the pathogenesis of ulcerative colitis. However only TIMP-1 can be useful as a biomarker of the disease activity, demonstrating association with clinical and endoscopic pictures.
