AIM:To assess expression of matrix metalloproteinases 2(MMP2)and MMP9 in gastric cancer,superficial gastritis and normal mucosa,and to measure metalloproteinase activity.METHODS:MMP2 and MMP9 mRNA expression was deter...AIM:To assess expression of matrix metalloproteinases 2(MMP2)and MMP9 in gastric cancer,superficial gastritis and normal mucosa,and to measure metalloproteinase activity.METHODS:MMP2 and MMP9 mRNA expression was determined by quantitative real-time polymerase chain reaction.Normalization was carried out using three different factors.Proteins were analyzed by quantitative gelatin zymography(qGZ).RESULTS:18S ribosomal RNA(18SRNA)was very highly expressed,while hypoxanthine ribosyltransferase-1(HPRT-1)was moderately expressed.MMP2 was highly expressed,while MMP9 was not detected or lowly expressed in normal tissues,moderately or highly expressed in gastritis and highly expressed in cancer.Relative expression of 18SRNA and HPRT-1 showed no significant differences.Significant differences in MMP2 and MMP9 were found between cancer and normal tissue,but not between gastritis and normal tissue.Absolute quantification of MMP9 echoed this pattern,but differential expression of MMP2 proved conflictive.Analysis by qGZ indicated significant differences between cancer and normal tissue in MMP-2,total MMP-9,250 and 110 kDa bands.CONCLUSION:MMP9 expression is enhanced in gastric cancer compared to normal mucosa;interpretation of differential expression of MMP2 is difficult to establish.展开更多
Objective: To explore whether a DNA immunization approach targeting the major haemorrhage molecule of a prothrombin activator-like metalloproteinase from Echis ocellatus(E. ocellatus) venom could be conceived to inspi...Objective: To explore whether a DNA immunization approach targeting the major haemorrhage molecule of a prothrombin activator-like metalloproteinase from Echis ocellatus(E. ocellatus) venom could be conceived to inspire antibodies with more prominent specificity and equal adequacy to current conventional antivenoms systems.Methods: The isolated DNA Eo MP-6 was used as the template for PCR amplification using the Eo DC-2-specific forward and reverse primers. A PCR product of approximately700 bp was obtained and cloned into p Sec Tag-B expression vector where anti-Eo DC-2antibodies were generated and analysed for their efficacy to neutralise local haemorrhage in vitro and in vivo.Results: Our results suggest that the generated anti-Eo DC-2 showed a remarkable efficacy by(a) interfering with the interaction of the recombinant disintegrin "Eo DC-2" isolated from the E. ocellatus as well as other viper species to the a2b1-integrins on platelets;(b) complete inhibition of the catalytic site of the metalloproteinase molecules in vitro using an adaptation antibody zymography assay. Furthermore, it has a polyspecific potential and constitutively expressed significant inhibition by cross-reaction and neutralised venom-induced local haemorrhage exerted by different viper species in vivo. The potential characteristic of Eo DC-2 against one part(the non-catalytic domain) as opposed to the whole molecule to neutralise its haemorrhagic activity is of crucial importance as it represents a novel approach with greater immunological specificity and fewer hazards, if any, than conventional systems of antivenom production, by exposure large animals that usually being used for the current antivenom production to a less injurious than expression of the whole molecule containing the catalytic metalloprotease domain. Hence, we report for the first time that our preliminary results hold a promising future for antivenom development.Conclusions: Antibodies generated against the E. ocellatus venom prothrombin activatorlike metalloprotease and disintegrin-cysteine-rich domains modulated and inhibited the catalytic activity both in vitro and in vivo of venom metalloproteinase disintegrin cysteine rich molecules. Thus, generating of venom specific-toxin antibodies by DNA immunization offer a more rational treatment of snake envenoming than conventional antivenom.展开更多
The activity of matrix metalloproteinases (MMPs) in the uterine flushing and endometrial tissue of normal adult women wearing FCu IUD (fixed Cu IUD) or FICu IUD (indomethacin releasing FCu IUD) was observed by using z...The activity of matrix metalloproteinases (MMPs) in the uterine flushing and endometrial tissue of normal adult women wearing FCu IUD (fixed Cu IUD) or FICu IUD (indomethacin releasing FCu IUD) was observed by using zymography on SDS PAGE containing gelatin. The results showed that the activity and kinds of MMPs in FCu IUD group were increased significantly as compared with themselves before being inserted FCu IUD. However, compared with the FCu IUD group, the activity of some kinds of MMPs in the FICu IUD group was decreased significantly. These data suggest that IUD can enhance the activity of MMPs in human endometrium, intermediated by prostaglandins, and MMPs may have relation to IUD induced menorrhagia and indomethacin reduces IUD induced menorrhagia by partly inhibiting MMPs synthesis.展开更多
Matrix metalloproteinases (MMPs) are matrix-degrading enzymes that are over-expressed in joints of rheumatoid arthritis (RA) patients. However, the contribution of specific MMPs for the development of arthritic joints...Matrix metalloproteinases (MMPs) are matrix-degrading enzymes that are over-expressed in joints of rheumatoid arthritis (RA) patients. However, the contribution of specific MMPs for the development of arthritic joints is unknown. This study is aimed at studying the role of matrix metalloproteinase-9 (MMP-9) in mice, using the K/BxN serum-transfer model of RA. Arthritis was induced in Balb/c mice by injecting K/BxN serum. Development of arthritis was followed in these mice by measuring ankle thickness and clinical index score. MMP-9 expression in the joints of mice killed at various time points during the disease progression was determined by gelatin zymography using ankle lysates. We found that MMP-9 expression increased with the severity of arthritis. Importantly MMP-9 deficient mice injected with K/BxN serum showed a milder form of arthritis in comparison to the control C57BL/6 mice injected with K/BxN serum. We therefore conclude that MMP-9 promotes arthritis in mice.展开更多
To investigate whether the calcium channel blocker amlodipine could inhibit macrophage matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) expression and secretion. Methods Peritoneal macrophages...To investigate whether the calcium channel blocker amlodipine could inhibit macrophage matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) expression and secretion. Methods Peritoneal macrophages were isolated from BALB/C mice and incubated with low (5μg/L), middle (15μg/L) and high (305μg/L) concentrations of amlodipine, or in the medium alone (controls) for 24 hours, and the expression and secretion of MMP-2 and MM-9 of the cells were analyzed by RT-PCR and gelatin zymography. Results Compared with controls, amlodipine at low concentration had no significant effects on the expression and secretion of either MMP-2 and MMP-9 (P＞0.05);at middle concentrationit it could inhibited MMP-2 and MMP-9 expressions completely and significantly reduced the secretion of MMP-9 (P＜0.05); but it had no effect on the secretion of MMP-2. At high concentration it also inhibited MMP-2 and MMP-9 expression completely. Conclusion Amlodipine at 15 ìg/L inhibited the expression of MMP-2 and MMP-9 and reduced the secretion of MMP-9, suggesting that amlodipine may stabilize atherosclerotic plaque.展开更多
Microplastics provide a new ecological niche for microorganisms,and the accumulation levels of microplastics(MPs)in terrestrial ecosystems are higher than those in marine ecosystems.Here,we applied the zymography to i...Microplastics provide a new ecological niche for microorganisms,and the accumulation levels of microplastics(MPs)in terrestrial ecosystems are higher than those in marine ecosystems.Here,we applied the zymography to investigate how MPs–polyethylene[PE],and polyvinyl chloride[PVC])at two levels(0.01%and 1%soil weight)impacted the spatial distribution of soil hydrolases,nutrient availability,and rice growth in paddy soil.MPs increased the above-ground biomass by 13.0%–15.5%and decreased the below-ground biomass by 8.0%–15.1%.Addition of 0.01%and 1%MPs reduced soil NH4+content by 18.3%–63.2%and 52.2%–80.2%,respectively.The average activities of N-and P-hydrolases increased by 0.8%–4.8%and 1.9%–6.3%with addition of MPs,respectively.The nutrient uptake by rice plants and the enzyme activities in hotspots increased with MP content in soil.The accumulation of MPs in paddy soil could provide an ecological niche that facilitates microbial survival,alters the spatial distribution of soil hydrolases,and decreases nutrient availability.展开更多
基金Supported by The National Council on Science and Technology (CONACYT:85675 and 79628)Institute of Public Health(POA: 2008-2010)Research Office of Veracruzana University and Public Education Secretariat(SEP-PROMEP-UV:PTC-319)
文摘AIM:To assess expression of matrix metalloproteinases 2(MMP2)and MMP9 in gastric cancer,superficial gastritis and normal mucosa,and to measure metalloproteinase activity.METHODS:MMP2 and MMP9 mRNA expression was determined by quantitative real-time polymerase chain reaction.Normalization was carried out using three different factors.Proteins were analyzed by quantitative gelatin zymography(qGZ).RESULTS:18S ribosomal RNA(18SRNA)was very highly expressed,while hypoxanthine ribosyltransferase-1(HPRT-1)was moderately expressed.MMP2 was highly expressed,while MMP9 was not detected or lowly expressed in normal tissues,moderately or highly expressed in gastritis and highly expressed in cancer.Relative expression of 18SRNA and HPRT-1 showed no significant differences.Significant differences in MMP2 and MMP9 were found between cancer and normal tissue,but not between gastritis and normal tissue.Absolute quantification of MMP9 echoed this pattern,but differential expression of MMP2 proved conflictive.Analysis by qGZ indicated significant differences between cancer and normal tissue in MMP-2,total MMP-9,250 and 110 kDa bands.CONCLUSION:MMP9 expression is enhanced in gastric cancer compared to normal mucosa;interpretation of differential expression of MMP2 is difficult to establish.
基金Supported by the Wellcome Trust,UK(RAH,Grant No.061325)the University of Science and Technology,Yementhe Gunter Trust,UK
文摘Objective: To explore whether a DNA immunization approach targeting the major haemorrhage molecule of a prothrombin activator-like metalloproteinase from Echis ocellatus(E. ocellatus) venom could be conceived to inspire antibodies with more prominent specificity and equal adequacy to current conventional antivenoms systems.Methods: The isolated DNA Eo MP-6 was used as the template for PCR amplification using the Eo DC-2-specific forward and reverse primers. A PCR product of approximately700 bp was obtained and cloned into p Sec Tag-B expression vector where anti-Eo DC-2antibodies were generated and analysed for their efficacy to neutralise local haemorrhage in vitro and in vivo.Results: Our results suggest that the generated anti-Eo DC-2 showed a remarkable efficacy by(a) interfering with the interaction of the recombinant disintegrin "Eo DC-2" isolated from the E. ocellatus as well as other viper species to the a2b1-integrins on platelets;(b) complete inhibition of the catalytic site of the metalloproteinase molecules in vitro using an adaptation antibody zymography assay. Furthermore, it has a polyspecific potential and constitutively expressed significant inhibition by cross-reaction and neutralised venom-induced local haemorrhage exerted by different viper species in vivo. The potential characteristic of Eo DC-2 against one part(the non-catalytic domain) as opposed to the whole molecule to neutralise its haemorrhagic activity is of crucial importance as it represents a novel approach with greater immunological specificity and fewer hazards, if any, than conventional systems of antivenom production, by exposure large animals that usually being used for the current antivenom production to a less injurious than expression of the whole molecule containing the catalytic metalloprotease domain. Hence, we report for the first time that our preliminary results hold a promising future for antivenom development.Conclusions: Antibodies generated against the E. ocellatus venom prothrombin activatorlike metalloprotease and disintegrin-cysteine-rich domains modulated and inhibited the catalytic activity both in vitro and in vivo of venom metalloproteinase disintegrin cysteine rich molecules. Thus, generating of venom specific-toxin antibodies by DNA immunization offer a more rational treatment of snake envenoming than conventional antivenom.
基金This project was supported by a grantfrom Hubei Scienceand Technology Comm ittee (No.96 1P190 1)
文摘The activity of matrix metalloproteinases (MMPs) in the uterine flushing and endometrial tissue of normal adult women wearing FCu IUD (fixed Cu IUD) or FICu IUD (indomethacin releasing FCu IUD) was observed by using zymography on SDS PAGE containing gelatin. The results showed that the activity and kinds of MMPs in FCu IUD group were increased significantly as compared with themselves before being inserted FCu IUD. However, compared with the FCu IUD group, the activity of some kinds of MMPs in the FICu IUD group was decreased significantly. These data suggest that IUD can enhance the activity of MMPs in human endometrium, intermediated by prostaglandins, and MMPs may have relation to IUD induced menorrhagia and indomethacin reduces IUD induced menorrhagia by partly inhibiting MMPs synthesis.
文摘Matrix metalloproteinases (MMPs) are matrix-degrading enzymes that are over-expressed in joints of rheumatoid arthritis (RA) patients. However, the contribution of specific MMPs for the development of arthritic joints is unknown. This study is aimed at studying the role of matrix metalloproteinase-9 (MMP-9) in mice, using the K/BxN serum-transfer model of RA. Arthritis was induced in Balb/c mice by injecting K/BxN serum. Development of arthritis was followed in these mice by measuring ankle thickness and clinical index score. MMP-9 expression in the joints of mice killed at various time points during the disease progression was determined by gelatin zymography using ankle lysates. We found that MMP-9 expression increased with the severity of arthritis. Importantly MMP-9 deficient mice injected with K/BxN serum showed a milder form of arthritis in comparison to the control C57BL/6 mice injected with K/BxN serum. We therefore conclude that MMP-9 promotes arthritis in mice.
文摘To investigate whether the calcium channel blocker amlodipine could inhibit macrophage matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) expression and secretion. Methods Peritoneal macrophages were isolated from BALB/C mice and incubated with low (5μg/L), middle (15μg/L) and high (305μg/L) concentrations of amlodipine, or in the medium alone (controls) for 24 hours, and the expression and secretion of MMP-2 and MM-9 of the cells were analyzed by RT-PCR and gelatin zymography. Results Compared with controls, amlodipine at low concentration had no significant effects on the expression and secretion of either MMP-2 and MMP-9 (P＞0.05);at middle concentrationit it could inhibited MMP-2 and MMP-9 expressions completely and significantly reduced the secretion of MMP-9 (P＜0.05); but it had no effect on the secretion of MMP-2. At high concentration it also inhibited MMP-2 and MMP-9 expression completely. Conclusion Amlodipine at 15 ìg/L inhibited the expression of MMP-2 and MMP-9 and reduced the secretion of MMP-9, suggesting that amlodipine may stabilize atherosclerotic plaque.
基金We are grateful for financial support from the National Key Research and Development Plan(2019YFC1805100)the National Natural Science Foundation of China(42107341,21767012)+2 种基金the UK Natural Environment Research Council and the Global Challenges Research Fund(NE/V005871/1)sponsorship by K.C.Wong Magna Fund in Ningbo Universitythe Science and Technology Plan of Ganzhou City(GSKF201850).
文摘Microplastics provide a new ecological niche for microorganisms,and the accumulation levels of microplastics(MPs)in terrestrial ecosystems are higher than those in marine ecosystems.Here,we applied the zymography to investigate how MPs–polyethylene[PE],and polyvinyl chloride[PVC])at two levels(0.01%and 1%soil weight)impacted the spatial distribution of soil hydrolases,nutrient availability,and rice growth in paddy soil.MPs increased the above-ground biomass by 13.0%–15.5%and decreased the below-ground biomass by 8.0%–15.1%.Addition of 0.01%and 1%MPs reduced soil NH4+content by 18.3%–63.2%and 52.2%–80.2%,respectively.The average activities of N-and P-hydrolases increased by 0.8%–4.8%and 1.9%–6.3%with addition of MPs,respectively.The nutrient uptake by rice plants and the enzyme activities in hotspots increased with MP content in soil.The accumulation of MPs in paddy soil could provide an ecological niche that facilitates microbial survival,alters the spatial distribution of soil hydrolases,and decreases nutrient availability.
