BACKGROUND Acute liver failure(ALF)has a high mortality with widespread hepatocyte death involving ferroptosis and pyroptosis.The silent information regulator sirtuin 1(SIRT1)-mediated deacetylation affects multiple b...BACKGROUND Acute liver failure(ALF)has a high mortality with widespread hepatocyte death involving ferroptosis and pyroptosis.The silent information regulator sirtuin 1(SIRT1)-mediated deacetylation affects multiple biological processes,including cellular senescence,apoptosis,sugar and lipid metabolism,oxidative stress,and inflammation.AIM To investigate the association between ferroptosis and pyroptosis and the upstream regulatory mechanisms.METHODS This study included 30 patients with ALF and 30 healthy individuals who underwent serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)testing.C57BL/6 mice were also intraperitoneally pretreated with SIRT1,p53,or glutathione peroxidase 4(GPX4)inducers and inhibitors and injected with lipopolysaccharide(LPS)/D-galactosamine(D-GalN)to induce ALF.Gasdermin D(GSDMD)^(-/-)mice were used as an experimental group.Histological changes in liver tissue were monitored by hematoxylin and eosin staining.ALT,AST,glutathione,reactive oxygen species,and iron levels were measured using commercial kits.Ferroptosis-and pyroptosis-related protein and mRNA expression was detected by western blot and quantitative real-time polymerase chain reaction.SIRT1,p53,and GSDMD were assessed by immunofluorescence analysis.RESULTS Serum AST and ALT levels were elevated in patients with ALF.SIRT1,solute carrier family 7a member 11(SLC7A11),and GPX4 protein expression was decreased and acetylated p5,p53,GSDMD,and acyl-CoA synthetase long-chain family member 4(ACSL4)protein levels were elevated in human ALF liver tissue.In the p53 and ferroptosis inhibitor-treated and GSDMD^(-/-)groups,serum interleukin(IL)-1β,tumour necrosis factor alpha,IL-6,IL-2 and C-C motif ligand 2 levels were decreased and hepatic impairment was mitigated.In mice with GSDMD knockout,p53 was reduced,GPX4 was increased,and ferroptotic events(depletion of SLC7A11,elevation of ACSL4,and iron accumulation)were detected.In vitro,knockdown of p53 and overexpression of GPX4 reduced AST and ALT levels,the cytostatic rate,and GSDMD expression,restoring SLC7A11 depletion.Moreover,SIRT1 agonist and overexpression of SIRT1 alleviated acute liver injury and decreased iron deposition compared with results in the model group,accompanied by reduced p53,GSDMD,and ACSL4,and increased SLC7A11 and GPX4.Inactivation of SIRT1 exacerbated ferroptotic and pyroptotic cell death and aggravated liver injury in LPS/D-GalNinduced in vitro and in vivo models.CONCLUSION SIRT1 activation attenuates LPS/D-GalN-induced ferroptosis and pyroptosis by inhibiting the p53/GPX4/GSDMD signaling pathway in ALF.展开更多
Organoseleniums are a class of compounds attracting attention across the globe owing to their Glutathione peroxidase(GPx)mimicry,which confers on them a strong antioxidant activity.Diphenyl diselenide(DPDS)is an Organ...Organoseleniums are a class of compounds attracting attention across the globe owing to their Glutathione peroxidase(GPx)mimicry,which confers on them a strong antioxidant activity.Diphenyl diselenide(DPDS)is an Organoselenium whose GPx mimetic property has been suggested to rely on the oxidation of non-protein or protein thiols critical to the activities of some sulfhydryl enzymes.This study,therefore investigated the GPx mimic/antioxidant property of DPDS as well as the role of thiols of two key sulfhydryl enzymes,cerebral Na^(+)/K^(+)-ATPase(sodium pump)and hepatic delta-aminolevulinic acid dehydratase(δ-ALAD)in the GPx mimicry of DPDS.Albino Wistar rats were euthanized,and the liver and brain were removed and used to assay for the effect of DPDS on lipid peroxidation induced by two prooxidants[Fe2^(+)(10μM)and H2O2,(1 mM)]as well as the activities of the sulfhydryl enzymes.The results revealed that DPDS profoundly(P<0.05)counteracted Fe2^(+)and H2O2-induced lipid peroxidation in the rats’hepatic and cerebral tissues.Furthermore,the results of assay systems for lipid peroxidation and sodium pump revealed that DPDS inhibited Na^(+)/K^(+)-ATPase and lipid peroxidation in the brain tissue homogenates in the same reaction system.A similar result was obtained in the assay system for lipid peroxidation and hepaticδ-ALAD as DPDS simultaneously inhibited the enzyme’s activity and lipid peroxidation.This suggests that the GPx mimetic property of DPDS may be linked to the enzymes’loss of activity,which further validates the suggestions that the enzymes’inhibition,as well as the antioxidant action of DPDS,rely on the oxidation of critical thiols of the enzymes.However,the GPx mimicry of DPDS should be investigated in the presence of thiol-blocking or oxidizing agents in biological systems in order to further ascertain the role of protein thiols.展开更多
Glutathione peroxidase (GPx) is an antioxidant that plays an important role in the maintenance of male fertility. The aim of this study was to compare the profile of enzymatic activity of glutathione peroxidase in the...Glutathione peroxidase (GPx) is an antioxidant that plays an important role in the maintenance of male fertility. The aim of this study was to compare the profile of enzymatic activity of glutathione peroxidase in the seminal plasma of normozoosperm and those of pathological sperm. Thus, the activity of glutathione peroxidase was determined in the seminal plasma of 20 normozoosperms, 9 azoosperms and 31 oligoasthenoteratozoosperms. It was 37.58 ± 3.14 U/L in normozoosperms, 39.39 ± 2.27 U/L in oligoasthenoteratozoosperms, and 29.77 ± 2.62 U/L in azoosperms. The mean GPx enzyme activity of normozoosperms did not differ significantly from that of oligoasthenoteratozoosperms and azoosperms. In contrast, comparison of enzyme activity between abnormal sperms gave a significant difference. This study showed that glutathione peroxidase enzymatic activity is not related to sperm quality.展开更多
In order to enhance the glutathione peroxidase(GPX) catalytic activity of the selenium-containing single-chain variable fragments(Se-scFv), a novel human scFv was designed on the basis of the structure of human an...In order to enhance the glutathione peroxidase(GPX) catalytic activity of the selenium-containing single-chain variable fragments(Se-scFv), a novel human scFv was designed on the basis of the structure of human antibody and optimized via bioinformatics methods such as homologous sequence analysis, three-dimensional(3D) model building, binding-site analysis and docking. The DNA sequence of the new human scFv was synthesized and cloned into the expression vector pET22b(+), then the scFv protein was expressed in soluble form in Escherichia coli BL21(DE3) and purified by Ni2+-immobilized metal affinity chromatography(IMAC). The serine residue of scFv in the active site was converted into selenocysteine(Sec) with the chemical modification method, thus, the human Se-scFv with GPX activity was obtained. The GPX activity of the Se-scFv protein was characterized. Compared with other Se-scFv, the new human Se-scFv showed similar efficiency for catalyzing the reduction of hydrogen peroxide by glutathione. It exhibited pH and temperature dependent catalytic activity and a typical ping-pong kinetic mechanism.展开更多
A versatile peroxidase (VP-Peco60-7 ) was generated and purified from the liquid culture of Pleurotus eryngii. The purification procedure included ammonium sulfate precipitation, ion exchange chromatography, and gel c...A versatile peroxidase (VP-Peco60-7 ) was generated and purified from the liquid culture of Pleurotus eryngii. The purification procedure included ammonium sulfate precipitation, ion exchange chromatography, and gel chromatography. The molecular weight and isoelectric point (pI) of VP-Peco60-7 were determined to be approxi-mately 40 kDa and 4.1, respectively. By N-terminal sequence determination and peptide mapping analysis, VP-Peco60-7 was found to be similar to the versatile peroxidase isoenzyme VPL1, which was previously isolated from liquid cultures of the same species. However, the molecular weight and pI of VP-Peco60-7 were different from those of versatile peroxidases of liquid cultures, implying that the VP-Peco60-7 in this study is of a novel type. With 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) as a substrate, the maximal enzyme activity was obtained at 50 °C and pH 3.0. The catalysis of ABTS by VP-Peco60-7 was expressed by the Michaelis-Menten equa-tion. At 50 °C and pH 3.0, the maximum velocity (V max ) was 188.68 U·mg-1 and the michaelis constant (K m ) was 203.09 μmol·L-1 .展开更多
This study aims to analyze the clinical significance and mechanism of nuclear factor erythroid 2-related factor 2(NRF2)and glutathione peroxidase 4(GPX4)in primary hepatic carcinoma(PHC).Methods:The expression of NRF2...This study aims to analyze the clinical significance and mechanism of nuclear factor erythroid 2-related factor 2(NRF2)and glutathione peroxidase 4(GPX4)in primary hepatic carcinoma(PHC).Methods:The expression of NRF2 and GPX4 in peripheral blood of patients with PHC was determined to analyze the diagnostic value of the two combined for PHC.The prognostic significance of NRF2 and GPX4 was evaluated by 3-year followup.Human liver epithelial cells THLE-2 and human hepatocellular carcinoma cells HepG2 were purchased,and the expression of NRF2 and GPX4 in the cells was determined.NRF2 and GPX4 aberrant expression vectors were constructed and transfected into HepG2,and changes in cell proliferation and invasion capabilities were observed.Results:The expression of NRF2 and GPX4 in patients with PHC was higher than that in patients with LC or VH(p<0.05),and the two indicators combined was excellent in diagnosing PHC.Moreover,patients with high expression of NRF2 and GPX4 had a higher risk of death(p<0.05).In in vitro experiments,both NRF2 and GPX4 expression was elevated in HepG2(p<0.05).HepG2 activity was enhanced by increasing the expression of the two,vice versa(p<0.05).Conclusion:NRF2 and GPX4 combined is excellent in diagnosing PHC,and promotes the malignant development of PHC.展开更多
A third-generation horseradish peroxidase (HRP) biosensor has been developed by adsorbing HRP on multi-wall carbon nanotube (MWNTs) monolayer modified gold electrode surface. The assembly process was investigated by...A third-generation horseradish peroxidase (HRP) biosensor has been developed by adsorbing HRP on multi-wall carbon nanotube (MWNTs) monolayer modified gold electrode surface. The assembly process was investigated by electrochemical and spectroscopic techniques. Results showed that the immobilized HRP exhibited direct electrochemical behavior toward the reduction of H2O2. The resulting biosensor shows a fast amperometric response (<2 s) to H2O2. The linear response range was from 5.0×10-7~1.0×10-5 mol/L with a detection limit of 1.0×10-7mol/L. Moreover, the biosensor has a good reproducibility, and long-term stability.展开更多
A novel hydrogen peroxide biosensor has been fabricated based on covalently linked horseradish peroxidase (HRP) onto L- glutathione self-assembled monolayers (SAMs). The SAMs-based electrode was characterized by e...A novel hydrogen peroxide biosensor has been fabricated based on covalently linked horseradish peroxidase (HRP) onto L- glutathione self-assembled monolayers (SAMs). The SAMs-based electrode was characterized by electrochemical methods, and direct electrochemistry of HRP can be achieved with formal potential of-0.242 V (vs. saturated Ag/AgCl) in pH 7 phosphate buffer solution (PBS), the redox peak current is linear to scan rate and rate constant can be calculated to be 0.042 s^-1. The HRP-SAMs- based biosensors show its better electrocatalysis to hydrogen peroxide in the concentration range of 1 × 10^-6 mol/L to 1.2 × 10^-3 mol/L with a detection limit of 4 × 10^-7 mol/L. The apparent Michealis-Menten constant is 3.12 mmol/L. The biosensor can effectively eliminate the interferences of dopamine, ascorbic acid, uric acid, catechol and p-acetaminophen.展开更多
The bioavailability of selenium(Se)from ground beef has been previously found in this laboratory to be greater than that of selenite or selenate when fed to female Fischer 344 rats(B. Shi,J.E.Spallholz,J Am Coil Nutr,...The bioavailability of selenium(Se)from ground beef has been previously found in this laboratory to be greater than that of selenite or selenate when fed to female Fischer 344 rats(B. Shi,J.E.Spallholz,J Am Coil Nutr,13:95 ̄101,1994).In the present study we examined the bioavailability of Se from various commercial portions of beef,the liver,striploin,round, shoulder and brisket.All beef was cooked, freeze-dried,finely powdered and mixed with the other dietary ingredients.The experimental diets were fed to the weanling Fischer 344 rats which had been subjected to dietary depletion of Se for 6 weeks.The bioavailability of Se from the beef diets was compared with that of Se as se lenite or L-seienomethionine(SeMet)added to torula-yeast diets.Each experimental diet contained 0'10mg Se/kg.After 8 weeks of dietary Se repletion,relative activity of liver glutathione peroxidase (EC 1.11.1.9;GSHPx) from the different dietary groups colllpared with that of control animals(100%)was(%):selenite 91,SeMet 122 (P<0.05),liver 108, striploin 105,round 106, shoulder 106,brisket 103.Se recovery in liver was generally highest from SeMet>beef muscle=beef liver>selenite.Muscle tissue deposition of Se was highest from SeMet>beef muscle>selenite=beef liver.In addition, the feeal excretion of Se was lowest from the SeMet dietary group and highest from the selenite dietary group.The experimetal results suggest that all cuts of beef appear to be highly bioavailable sources of dietary Se when compared with selenite or L-SeMet.展开更多
Objective To oberve the change in blood glutathione peroxidase (GSH-Px) protein levels of residents in the low-selenium (Se) area by contrasting the blood GSH-Px protein level of the children in the Keshan disease are...Objective To oberve the change in blood glutathione peroxidase (GSH-Px) protein levels of residents in the low-selenium (Se) area by contrasting the blood GSH-Px protein level of the children in the Keshan disease area with those in the Kashin-Beck disease and non-endemic areas. Methods GSH-Px protein levels were measured by enzyme-linked immunosorbent assays (ELISA). The Se content and GSH-Px activity were assayed by the 2,3-diaminonaphthalene spectrofluorimetric method and glutathione reductase-coupled method respectively. Results ①The blood Se content and GSH-Px protein level of children in Keshan disease area (Moding) were significantly lower than those in Xi’an non-endemic area, however, there was no significant difference when compared with the low-Se non-endemic area; ②The blood Se content, GSH-Px activity and GSH-Px protein level of children in the Kashin-Beck disease area (Yulin) were significantly lower than those of children in two non-endemic areas and in the Keshan disease area; ③The blood Se content and GSH-Px activity were positively correlated to the GSH-Px protein level respectively. Conclusion These results indicate that the blood GSH-Px protein level is decreased in the low-Se residents. The Se status not only affects the GSH-Px activity but also regulate the GSH-Px protein level.展开更多
[ Objective ] To respectively compare esterase isozymes and peroxidase isozymes among black worm Tenebrio molitor Linnaeus ( Tenebno molitor L. ) at different developmental stages and thus provide a theoretical basi...[ Objective ] To respectively compare esterase isozymes and peroxidase isozymes among black worm Tenebrio molitor Linnaeus ( Tenebno molitor L. ) at different developmental stages and thus provide a theoretical basis for species identification and large-scale farming. [ Method] The esterase isozymes and peroxidase isozymes of black worm Tenebno molitor L. at different developmental stages were respectively analyzed by polyacrylamide gel electrophoresis. [ Result] Both esterase isozymes and peroxidase isozymes were different among black worm Tenebrio molitor L. at developmental stages, larvae in different instars or between the male adults and the female adults. [ Conclusion] The esterase isozymes, peroxidase isozymes and physiological characteristics of black worm Tenebrio molitor L. varies with developmental stages.展开更多
Based on the collected multiwavelength data, namely in the radio(NVSS, FIRST, RATAN-600), IR(WISE),optical(Pan-STARRS), UV(GALEX), and X-ray(ROSAT, Swift-XRT) ranges, we have performed a cluster analysis for the blaza...Based on the collected multiwavelength data, namely in the radio(NVSS, FIRST, RATAN-600), IR(WISE),optical(Pan-STARRS), UV(GALEX), and X-ray(ROSAT, Swift-XRT) ranges, we have performed a cluster analysis for the blazars of the Roma-BZCAT catalog. Using two machine learning methods, namely a combination of PCA with k-means clustering and Kohonen's self-organizing maps(SOMs), we have constructed an independent classification of the blazars(five classes) and compared the classes with the known Roma-BZCAT classification(FSRQs, BL Lacs, galaxy-dominated BL Lacs, and blazars of an uncertain type) as well as with the high synchrotron peaked(HSP) blazars from the 3HSP catalog and blazars from the TeVCat catalog. The obtained groups demonstrate concordance with the BL Lac/FSRQ classification along with a continuous character of the change in the properties. The group of HSP blazars stands out against the overall distribution. We examine the characteristics of the five groups and demonstrate distinctions in their spectral energy distribution shapes. The effectiveness of the clustering technique for objective analysis of multiparametric arrays of experimental data is demonstrated.展开更多
Methanolic extracts from the leaves of <em>Manihot esculenta </em>(Two cultivars) and <em>Manihot glaziovii</em>, consumed as traditional vegetables in DR. Congo was chemically characterized by...Methanolic extracts from the leaves of <em>Manihot esculenta </em>(Two cultivars) and <em>Manihot glaziovii</em>, consumed as traditional vegetables in DR. Congo was chemically characterized by Thin layer Chromatography and High Performance Liquid Chromatography. <em>In vitro</em> biochemical activities of extracts against Radical Oxidative Species (ROS) production were assessed in cellular models, on enzymes, Myeloperoxidase (MPO) and Horseradish Peroxidase (HRP) involved in inflammation. The microscopic analysis of the powder of leaves showed that each species displays specific and discriminating botanical microscopic features. Varieties of<em> M. esculenta</em> had a chemical fingerprint different from <em>M. glaziovii</em>. The majority of compounds were polyphenols, represented mainly by rutin, kaempferol-3-O-rutinoside, amentoflavone, phenolic acids such as gallic acid. All extracts exhibited high cellular antioxidant activity in the range of 0.1 to 10 μg<span style="white-space:nowrap;">·</span>mL<sup>-1</sup> using lucigenin with neutrophils, but a moderate cellular antioxidant activity ranging between 10 and 100 μg<span style="white-space:nowrap;">·</span>mL<sup>-1</sup> with DCFDA on HL60 monocytes. Extracts from <em>Manihot</em> leaves showed a pronounced inhibitory effect on the production of extracellular ROS, on HRP and myeloperoxidase activity. Cellular antioxidant activities, the inhibitory effect on HRP of extracts from <em>M. glaziovii</em>, <em>M. esculenta</em> cultivar <em>Mwambu </em>were significantly higher, but their inhibitory effect on the activity of MPO was lower than those of <em>M. esculenta</em> cultivar TEM 419. The biological activities of <em>Manihot esculenta</em> and <em>Manihot glaziovii </em>were well correlated to their phytochemicals that could justify their traditional use as vegetables, potential functional foods or nutraceutical resources and medicines.展开更多
基金Supported by National Natural Science Foundation of China,No.82060123Doctoral Start-up Fund of Affiliated Hospital of Guizhou Medical University,No.gysybsky-2021-28+1 种基金Fund Project of Guizhou Provincial Science and Technology Department,No.[2020]1Y299Guizhou Provincial Health Commission,No.gzwjk2019-1-082。
文摘BACKGROUND Acute liver failure(ALF)has a high mortality with widespread hepatocyte death involving ferroptosis and pyroptosis.The silent information regulator sirtuin 1(SIRT1)-mediated deacetylation affects multiple biological processes,including cellular senescence,apoptosis,sugar and lipid metabolism,oxidative stress,and inflammation.AIM To investigate the association between ferroptosis and pyroptosis and the upstream regulatory mechanisms.METHODS This study included 30 patients with ALF and 30 healthy individuals who underwent serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)testing.C57BL/6 mice were also intraperitoneally pretreated with SIRT1,p53,or glutathione peroxidase 4(GPX4)inducers and inhibitors and injected with lipopolysaccharide(LPS)/D-galactosamine(D-GalN)to induce ALF.Gasdermin D(GSDMD)^(-/-)mice were used as an experimental group.Histological changes in liver tissue were monitored by hematoxylin and eosin staining.ALT,AST,glutathione,reactive oxygen species,and iron levels were measured using commercial kits.Ferroptosis-and pyroptosis-related protein and mRNA expression was detected by western blot and quantitative real-time polymerase chain reaction.SIRT1,p53,and GSDMD were assessed by immunofluorescence analysis.RESULTS Serum AST and ALT levels were elevated in patients with ALF.SIRT1,solute carrier family 7a member 11(SLC7A11),and GPX4 protein expression was decreased and acetylated p5,p53,GSDMD,and acyl-CoA synthetase long-chain family member 4(ACSL4)protein levels were elevated in human ALF liver tissue.In the p53 and ferroptosis inhibitor-treated and GSDMD^(-/-)groups,serum interleukin(IL)-1β,tumour necrosis factor alpha,IL-6,IL-2 and C-C motif ligand 2 levels were decreased and hepatic impairment was mitigated.In mice with GSDMD knockout,p53 was reduced,GPX4 was increased,and ferroptotic events(depletion of SLC7A11,elevation of ACSL4,and iron accumulation)were detected.In vitro,knockdown of p53 and overexpression of GPX4 reduced AST and ALT levels,the cytostatic rate,and GSDMD expression,restoring SLC7A11 depletion.Moreover,SIRT1 agonist and overexpression of SIRT1 alleviated acute liver injury and decreased iron deposition compared with results in the model group,accompanied by reduced p53,GSDMD,and ACSL4,and increased SLC7A11 and GPX4.Inactivation of SIRT1 exacerbated ferroptotic and pyroptotic cell death and aggravated liver injury in LPS/D-GalNinduced in vitro and in vivo models.CONCLUSION SIRT1 activation attenuates LPS/D-GalN-induced ferroptosis and pyroptosis by inhibiting the p53/GPX4/GSDMD signaling pathway in ALF.
文摘Organoseleniums are a class of compounds attracting attention across the globe owing to their Glutathione peroxidase(GPx)mimicry,which confers on them a strong antioxidant activity.Diphenyl diselenide(DPDS)is an Organoselenium whose GPx mimetic property has been suggested to rely on the oxidation of non-protein or protein thiols critical to the activities of some sulfhydryl enzymes.This study,therefore investigated the GPx mimic/antioxidant property of DPDS as well as the role of thiols of two key sulfhydryl enzymes,cerebral Na^(+)/K^(+)-ATPase(sodium pump)and hepatic delta-aminolevulinic acid dehydratase(δ-ALAD)in the GPx mimicry of DPDS.Albino Wistar rats were euthanized,and the liver and brain were removed and used to assay for the effect of DPDS on lipid peroxidation induced by two prooxidants[Fe2^(+)(10μM)and H2O2,(1 mM)]as well as the activities of the sulfhydryl enzymes.The results revealed that DPDS profoundly(P<0.05)counteracted Fe2^(+)and H2O2-induced lipid peroxidation in the rats’hepatic and cerebral tissues.Furthermore,the results of assay systems for lipid peroxidation and sodium pump revealed that DPDS inhibited Na^(+)/K^(+)-ATPase and lipid peroxidation in the brain tissue homogenates in the same reaction system.A similar result was obtained in the assay system for lipid peroxidation and hepaticδ-ALAD as DPDS simultaneously inhibited the enzyme’s activity and lipid peroxidation.This suggests that the GPx mimetic property of DPDS may be linked to the enzymes’loss of activity,which further validates the suggestions that the enzymes’inhibition,as well as the antioxidant action of DPDS,rely on the oxidation of critical thiols of the enzymes.However,the GPx mimicry of DPDS should be investigated in the presence of thiol-blocking or oxidizing agents in biological systems in order to further ascertain the role of protein thiols.
文摘Glutathione peroxidase (GPx) is an antioxidant that plays an important role in the maintenance of male fertility. The aim of this study was to compare the profile of enzymatic activity of glutathione peroxidase in the seminal plasma of normozoosperm and those of pathological sperm. Thus, the activity of glutathione peroxidase was determined in the seminal plasma of 20 normozoosperms, 9 azoosperms and 31 oligoasthenoteratozoosperms. It was 37.58 ± 3.14 U/L in normozoosperms, 39.39 ± 2.27 U/L in oligoasthenoteratozoosperms, and 29.77 ± 2.62 U/L in azoosperms. The mean GPx enzyme activity of normozoosperms did not differ significantly from that of oligoasthenoteratozoosperms and azoosperms. In contrast, comparison of enzyme activity between abnormal sperms gave a significant difference. This study showed that glutathione peroxidase enzymatic activity is not related to sperm quality.
基金Supported by the National Natural Science Foundation of China(No.30970608)the Applicative Technological Project of Bureau of Science and Technology of Changchun City, China(No.2009045)+1 种基金the Development and Planning Major Program of Jilin Provincial Science and Technology Department, China(No.20100948)the Innovation Method Fund of China (No.2008IM040800)
文摘In order to enhance the glutathione peroxidase(GPX) catalytic activity of the selenium-containing single-chain variable fragments(Se-scFv), a novel human scFv was designed on the basis of the structure of human antibody and optimized via bioinformatics methods such as homologous sequence analysis, three-dimensional(3D) model building, binding-site analysis and docking. The DNA sequence of the new human scFv was synthesized and cloned into the expression vector pET22b(+), then the scFv protein was expressed in soluble form in Escherichia coli BL21(DE3) and purified by Ni2+-immobilized metal affinity chromatography(IMAC). The serine residue of scFv in the active site was converted into selenocysteine(Sec) with the chemical modification method, thus, the human Se-scFv with GPX activity was obtained. The GPX activity of the Se-scFv protein was characterized. Compared with other Se-scFv, the new human Se-scFv showed similar efficiency for catalyzing the reduction of hydrogen peroxide by glutathione. It exhibited pH and temperature dependent catalytic activity and a typical ping-pong kinetic mechanism.
基金Supported by the Special Funds for Major State Basic Research Program of China (2007CB707805) the Natural Science Foundation of Zhejiang Province (Y505334)
文摘A versatile peroxidase (VP-Peco60-7 ) was generated and purified from the liquid culture of Pleurotus eryngii. The purification procedure included ammonium sulfate precipitation, ion exchange chromatography, and gel chromatography. The molecular weight and isoelectric point (pI) of VP-Peco60-7 were determined to be approxi-mately 40 kDa and 4.1, respectively. By N-terminal sequence determination and peptide mapping analysis, VP-Peco60-7 was found to be similar to the versatile peroxidase isoenzyme VPL1, which was previously isolated from liquid cultures of the same species. However, the molecular weight and pI of VP-Peco60-7 were different from those of versatile peroxidases of liquid cultures, implying that the VP-Peco60-7 in this study is of a novel type. With 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) as a substrate, the maximal enzyme activity was obtained at 50 °C and pH 3.0. The catalysis of ABTS by VP-Peco60-7 was expressed by the Michaelis-Menten equa-tion. At 50 °C and pH 3.0, the maximum velocity (V max ) was 188.68 U·mg-1 and the michaelis constant (K m ) was 203.09 μmol·L-1 .
文摘This study aims to analyze the clinical significance and mechanism of nuclear factor erythroid 2-related factor 2(NRF2)and glutathione peroxidase 4(GPX4)in primary hepatic carcinoma(PHC).Methods:The expression of NRF2 and GPX4 in peripheral blood of patients with PHC was determined to analyze the diagnostic value of the two combined for PHC.The prognostic significance of NRF2 and GPX4 was evaluated by 3-year followup.Human liver epithelial cells THLE-2 and human hepatocellular carcinoma cells HepG2 were purchased,and the expression of NRF2 and GPX4 in the cells was determined.NRF2 and GPX4 aberrant expression vectors were constructed and transfected into HepG2,and changes in cell proliferation and invasion capabilities were observed.Results:The expression of NRF2 and GPX4 in patients with PHC was higher than that in patients with LC or VH(p<0.05),and the two indicators combined was excellent in diagnosing PHC.Moreover,patients with high expression of NRF2 and GPX4 had a higher risk of death(p<0.05).In in vitro experiments,both NRF2 and GPX4 expression was elevated in HepG2(p<0.05).HepG2 activity was enhanced by increasing the expression of the two,vice versa(p<0.05).Conclusion:NRF2 and GPX4 combined is excellent in diagnosing PHC,and promotes the malignant development of PHC.
文摘A third-generation horseradish peroxidase (HRP) biosensor has been developed by adsorbing HRP on multi-wall carbon nanotube (MWNTs) monolayer modified gold electrode surface. The assembly process was investigated by electrochemical and spectroscopic techniques. Results showed that the immobilized HRP exhibited direct electrochemical behavior toward the reduction of H2O2. The resulting biosensor shows a fast amperometric response (<2 s) to H2O2. The linear response range was from 5.0×10-7~1.0×10-5 mol/L with a detection limit of 1.0×10-7mol/L. Moreover, the biosensor has a good reproducibility, and long-term stability.
文摘A novel hydrogen peroxide biosensor has been fabricated based on covalently linked horseradish peroxidase (HRP) onto L- glutathione self-assembled monolayers (SAMs). The SAMs-based electrode was characterized by electrochemical methods, and direct electrochemistry of HRP can be achieved with formal potential of-0.242 V (vs. saturated Ag/AgCl) in pH 7 phosphate buffer solution (PBS), the redox peak current is linear to scan rate and rate constant can be calculated to be 0.042 s^-1. The HRP-SAMs- based biosensors show its better electrocatalysis to hydrogen peroxide in the concentration range of 1 × 10^-6 mol/L to 1.2 × 10^-3 mol/L with a detection limit of 4 × 10^-7 mol/L. The apparent Michealis-Menten constant is 3.12 mmol/L. The biosensor can effectively eliminate the interferences of dopamine, ascorbic acid, uric acid, catechol and p-acetaminophen.
文摘The bioavailability of selenium(Se)from ground beef has been previously found in this laboratory to be greater than that of selenite or selenate when fed to female Fischer 344 rats(B. Shi,J.E.Spallholz,J Am Coil Nutr,13:95 ̄101,1994).In the present study we examined the bioavailability of Se from various commercial portions of beef,the liver,striploin,round, shoulder and brisket.All beef was cooked, freeze-dried,finely powdered and mixed with the other dietary ingredients.The experimental diets were fed to the weanling Fischer 344 rats which had been subjected to dietary depletion of Se for 6 weeks.The bioavailability of Se from the beef diets was compared with that of Se as se lenite or L-seienomethionine(SeMet)added to torula-yeast diets.Each experimental diet contained 0'10mg Se/kg.After 8 weeks of dietary Se repletion,relative activity of liver glutathione peroxidase (EC 1.11.1.9;GSHPx) from the different dietary groups colllpared with that of control animals(100%)was(%):selenite 91,SeMet 122 (P<0.05),liver 108, striploin 105,round 106, shoulder 106,brisket 103.Se recovery in liver was generally highest from SeMet>beef muscle=beef liver>selenite.Muscle tissue deposition of Se was highest from SeMet>beef muscle>selenite=beef liver.In addition, the feeal excretion of Se was lowest from the SeMet dietary group and highest from the selenite dietary group.The experimetal results suggest that all cuts of beef appear to be highly bioavailable sources of dietary Se when compared with selenite or L-SeMet.
文摘Objective To oberve the change in blood glutathione peroxidase (GSH-Px) protein levels of residents in the low-selenium (Se) area by contrasting the blood GSH-Px protein level of the children in the Keshan disease area with those in the Kashin-Beck disease and non-endemic areas. Methods GSH-Px protein levels were measured by enzyme-linked immunosorbent assays (ELISA). The Se content and GSH-Px activity were assayed by the 2,3-diaminonaphthalene spectrofluorimetric method and glutathione reductase-coupled method respectively. Results ①The blood Se content and GSH-Px protein level of children in Keshan disease area (Moding) were significantly lower than those in Xi’an non-endemic area, however, there was no significant difference when compared with the low-Se non-endemic area; ②The blood Se content, GSH-Px activity and GSH-Px protein level of children in the Kashin-Beck disease area (Yulin) were significantly lower than those of children in two non-endemic areas and in the Keshan disease area; ③The blood Se content and GSH-Px activity were positively correlated to the GSH-Px protein level respectively. Conclusion These results indicate that the blood GSH-Px protein level is decreased in the low-Se residents. The Se status not only affects the GSH-Px activity but also regulate the GSH-Px protein level.
基金supported by Hebei Education Commission Project (2003308) Qinhuangdao Science and Technology Commission Project
文摘[ Objective ] To respectively compare esterase isozymes and peroxidase isozymes among black worm Tenebrio molitor Linnaeus ( Tenebno molitor L. ) at different developmental stages and thus provide a theoretical basis for species identification and large-scale farming. [ Method] The esterase isozymes and peroxidase isozymes of black worm Tenebno molitor L. at different developmental stages were respectively analyzed by polyacrylamide gel electrophoresis. [ Result] Both esterase isozymes and peroxidase isozymes were different among black worm Tenebrio molitor L. at developmental stages, larvae in different instars or between the male adults and the female adults. [ Conclusion] The esterase isozymes, peroxidase isozymes and physiological characteristics of black worm Tenebrio molitor L. varies with developmental stages.
文摘Based on the collected multiwavelength data, namely in the radio(NVSS, FIRST, RATAN-600), IR(WISE),optical(Pan-STARRS), UV(GALEX), and X-ray(ROSAT, Swift-XRT) ranges, we have performed a cluster analysis for the blazars of the Roma-BZCAT catalog. Using two machine learning methods, namely a combination of PCA with k-means clustering and Kohonen's self-organizing maps(SOMs), we have constructed an independent classification of the blazars(five classes) and compared the classes with the known Roma-BZCAT classification(FSRQs, BL Lacs, galaxy-dominated BL Lacs, and blazars of an uncertain type) as well as with the high synchrotron peaked(HSP) blazars from the 3HSP catalog and blazars from the TeVCat catalog. The obtained groups demonstrate concordance with the BL Lac/FSRQ classification along with a continuous character of the change in the properties. The group of HSP blazars stands out against the overall distribution. We examine the characteristics of the five groups and demonstrate distinctions in their spectral energy distribution shapes. The effectiveness of the clustering technique for objective analysis of multiparametric arrays of experimental data is demonstrated.
文摘Methanolic extracts from the leaves of <em>Manihot esculenta </em>(Two cultivars) and <em>Manihot glaziovii</em>, consumed as traditional vegetables in DR. Congo was chemically characterized by Thin layer Chromatography and High Performance Liquid Chromatography. <em>In vitro</em> biochemical activities of extracts against Radical Oxidative Species (ROS) production were assessed in cellular models, on enzymes, Myeloperoxidase (MPO) and Horseradish Peroxidase (HRP) involved in inflammation. The microscopic analysis of the powder of leaves showed that each species displays specific and discriminating botanical microscopic features. Varieties of<em> M. esculenta</em> had a chemical fingerprint different from <em>M. glaziovii</em>. The majority of compounds were polyphenols, represented mainly by rutin, kaempferol-3-O-rutinoside, amentoflavone, phenolic acids such as gallic acid. All extracts exhibited high cellular antioxidant activity in the range of 0.1 to 10 μg<span style="white-space:nowrap;">·</span>mL<sup>-1</sup> using lucigenin with neutrophils, but a moderate cellular antioxidant activity ranging between 10 and 100 μg<span style="white-space:nowrap;">·</span>mL<sup>-1</sup> with DCFDA on HL60 monocytes. Extracts from <em>Manihot</em> leaves showed a pronounced inhibitory effect on the production of extracellular ROS, on HRP and myeloperoxidase activity. Cellular antioxidant activities, the inhibitory effect on HRP of extracts from <em>M. glaziovii</em>, <em>M. esculenta</em> cultivar <em>Mwambu </em>were significantly higher, but their inhibitory effect on the activity of MPO was lower than those of <em>M. esculenta</em> cultivar TEM 419. The biological activities of <em>Manihot esculenta</em> and <em>Manihot glaziovii </em>were well correlated to their phytochemicals that could justify their traditional use as vegetables, potential functional foods or nutraceutical resources and medicines.