ABSTRACT
The emerging demand for device miniaturization and integration prompts the patterning technique of micronano-cross-scale structures as an urgent desire. Lithography, as a sufficient patterning technique, has been playing an important role in achieving functional micronanoscale structures for decades. As a promising alternative, we have proposed and demonstrated the maskless optical projection nanolithography (MLOP-NL) technique for efficient cross-scale patterning. A minimum feature size of 32 nm, which is λ/12 super resolution breaking the optical diffraction limit, has been achieved by a single exposure. Furthermore, multiscale two-dimensional micronano-hybrid structures with the size over hundreds of micrometers and the precision at tens of nanometers have been fabricated by simply controlling the exposure conditions. The proposed MLOP-NL technique provides a powerful tool for achieving cross-scale patterning with both large-scale and precise configuration with high efficiency, which can be potentially used in the fabrication of multiscale integrated microsystems.
Subject(s)
PrintingABSTRACT
OBJECTIVE: To investigate the incidence rate of infectious diseases during hospitalization in late preterm infants in Beijing, China, as well as the risk factors for infectious diseases and the effect of breastfeeding on the development of infectious diseases. METHODS: Related data were collected from the late preterm infants who were hospitalized in the neonatal wards of 25 hospitals in Beijing, China, from October 23, 2015 to October 30, 2017. According to the feeding pattern, they were divided into a breastfeeding group and a formula feeding group. The two groups were compared in terms of general status and incidence rate of infectious diseases. A multivariate logistic regression analysis was used to investigate the risk factors for infectious diseases. RESULTS: A total of 1 576 late preterm infants were enrolled, with 153 infants in the breastfeeding group and 1 423 in the formula feeding group. Of all infants, 484 (30.71%) experienced infectious diseases. The breastfeeding group had a significantly lower incidence rate of infectious diseases than the formula feeding group (22.88% vs 31.55%, P=0.033). The multivariate logistic regression analysis showed that breastfeeding was an independent protective factor against infectious diseases (OR=0.534, P=0.004), while male sex, premature rupture of membranes, gestational diabetes mellitus, and asphyxia were risk factors for infectious diseases (OR=1.328, 5.386, 1.535, and 2.353 respectively, P < 0.05). CONCLUSIONS: Breastfeeding can significantly reduce the incidence of infectious diseases and is a protective factor against infectious diseases in late preterm infants. Breastfeeding should therefore be actively promoted for late preterm infants during hospitalization.
Subject(s)
Breast Feeding , Communicable Diseases , Hospitalization , Infant, Premature , Beijing/epidemiology , China/epidemiology , Communicable Diseases/epidemiology , Female , Hospitals , Humans , Incidence , Infant , Infant, Newborn , Male , PregnancyABSTRACT
Cellulose can be converted to ethanol via the fermentation of glucose, which is considered as a promising green alternative for transportation fuels. The conversion of cellulose to glucose needs three enzymes, in which ß-glucosidase (BGL) plays an essential role. However, BGL is inhibited by its own product glucose, greatly limiting its applications in industry. We previously obtained a novel BGL named Bgl6 with a high glucose tolerance. Further engineering through random mutagenesis produced a triple mutant M3 with improved thermostability. This enzyme shows promising properties for wide applications but the structural basis of the unusual properties of Bgl6 is not clear. In this study, we determined the crystal structures of Bgl6 and variants at high resolution, which provide insights into its glucose-tolerant mechanism and thermostability. Particularly, Bgl6 forms an extra channel that could be used as a secondary binding site for glucose, which may contribute to glucose tolerance. Additionally, the triple mutations could strengthen the hydrophobic interactions within the enzyme and may be responsible for the enhanced thermostability exhibited by M3, which was further confirmed by dynamic light scattering data. Lastly, structural comparison to other orthologs allows us to formulate new strategies on how to improve the catalytic efficiency of Bgl6.
Subject(s)
Glucose/chemistry , beta-Glucosidase/chemistry , Bacterial Proteins , Binding Sites , Cellulose/metabolism , Cloning, Molecular , Escherichia coli/genetics , Molecular Structure , Mutation , Protein Engineering , Protein Stability , beta-Glucosidase/antagonists & inhibitors , beta-Glucosidase/geneticsABSTRACT
The endoglucanase gene endo753 from Aspergillus flavus NRRL3357 strains was cloned, and the recombinant Endo753 was displayed on the cell surface of Saccharomyces cerevisiae EBY100 strain by the C-terminal fusion using Aga2p protein as anchor attachment tag. The results of indirect immunofluorescence and Western blot confirmed the expression and localization of Endo753 on the yeast cell surface. The hydrolytic activity test of the whole-cell enzyme revealed that Endo753 immobilized on the yeast cell surface had high endoglucanase activity. The functional characterization of the whole-cell enzyme was investigated, and the whole-cell enzyme displayed the maximum activity at pH 8 and 50 °C. The enzyme was stable in a pH range of 7.0-10.0. Furthermore, the whole-cell enzyme displayed high thermostability below 50 °C and moderate stability between 50 and 70 °C. These properties make endo753 a good candidate in bioethanol production from lignocellulosic materials after displaying on the yeast cell surface.
Subject(s)
Aspergillus flavus/enzymology , Cell Membrane/enzymology , Cell Membrane/metabolism , Cellulase/metabolism , Recombinant Proteins/metabolism , Saccharomyces cerevisiae/enzymology , Amino Acid Sequence , Aspergillus flavus/genetics , Base Sequence , Cellulase/genetics , Cloning, Molecular , Enzyme Activation , Enzyme Assays , Enzyme Stability , Hydrogen-Ion Concentration , Recombinant Proteins/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Sequence Alignment , Sequence Analysis , TemperatureABSTRACT
OBJECTIVE: To investigate the early predictors of necrotizing pneumonia in children. METHODS: The clinical data of 43 children with necrotizing pneumonia and 83 children with lobar pneumonia were retrospectively analyzed. Sex, age, the number of days with fever, laboratory examination results, and bronchoscopic findings were compared between the two groups. The multiple logistic regression analysis was used to identify the early predictors of necrotizing pneumonia. RESULTS: The necrotizing pneumonia group had a higher percentage of girls than the lobar pneumonia group (P<0.05). Compared with the lobar pneumonia group, the necrotizing pneumonia group had a larger number of days with fever, a higher peripheral blood white blood cell count (WBC), a higher percentage of neutrophils (NE%), and higher serum levels of high-sensitivity C-reactive protein (hs-CRP), albumin (Alb), and lactate dehydrogenase (LDH) (P<0.05). The necrotizing pneumonia group also had higher percentages of children with a large amount of sputum bolt under a bronchoscope which needed to be removed with biopsy forceps and children with rice-water-like bronchoalveolar lavage fluid (P<0.05). The multiple logistic regression analysis showed that being a female, the presence of sputum bolt under a bronchoscope which needed to be removed with biopsy forceps, the number of days with fever, WBC, hs-CRP, and LDH were independent predictors of necrotizing pneumonia. The receiver operating characteristic curve analysis showed that the cut-off values of the latter 4 predictors were 18.5â d, 15.1×10(9)/L, 121.5 mg/L, and 353.5 U/L, respectively. CONCLUSIONS: Increased WBC (≥15.1×10(9)/L), increased hs-CRP (≥121.5â mg/L), increased serum LDH (≥353.5 U/L), and the presence of sputum bolt under a bronchoscope which needs to be removed with biopsy forceps and rice-water-like bronchoalveolar lavage fluid may be the early predictors of necrotizing pneumonia in children.
Subject(s)
Pneumonia/diagnosis , C-Reactive Protein/analysis , Child , Child, Preschool , Female , Humans , L-Lactate Dehydrogenase/blood , Leukocyte Count , Logistic Models , Male , Necrosis , Pneumonia/bloodABSTRACT
BACKGROUND: ß-Galactosidases can be used to produce low-lactose milk and dairy products for lactose intolerant people. Although commercial ß-galactosidases have outstanding lactose hydrolysis ability, their thermostability is low, and reaction products have strong inhibition to these enzymes. In addition, the ß-galactosidases possessing simultaneously high thermostability and tolerance of galactose and glucose are still seldom reported until now. Therefore, identification of novel ß-galactosidases with high thermostability and tolerance to reaction products from unculturable microorganisms accounting for over 99% of microorganisms in the environment via metagenomic strategy is still urgently in demand. RESULTS: In the present study, a novel ß-galactosidase (Gal308) consisting of 658 amino acids was identified from a metagenomic library from soil samples of Turpan Basin in China by functional screening. After being overexpressed in Escherichia coli and purified to homogeneity, the enzymatic properties of Gal308 with N-terminal fusion tag were investigated. The recombinant enzyme displayed a pH optimum of 6.8 and a temperature optimum of 78 °C, and was considerably stable in the temperature range of 40 °C - 70 °C with almost unchangeable activity after incubation for 60 min. Furthermore, Gal308 displayed a very high tolerance of galactose and glucose, with the highest inhibition constant K(i,gal) (238 mM) and K(i,glu) (1725 mM) among ß-galactosidases. In addition, Gal308 also exhibited high enzymatic activity for its synthetic substrate o-nitrophenyl-ß-D-galactopyranoside (ONPG, 185 U/mg) and natural substrate lactose (47.6 U/mg). CONCLUSION: This study will enrich the source of ß-galactosidases, and attract some attentions to ß-galactosidases from extreme habitats and metagenomic library. Furthermore, the recombinant Gal308 fused with 156 amino acids exhibits many novel properties including high activity and thermostability at high temperatures, the pH optimum of 6.8, high enzyme activity for lactose, as well as high tolerance of galactose and glucose. These properties make it a good candidate in the production of low-lactose milk and dairy products after further study.
Subject(s)
Enzyme Inhibitors/metabolism , Galactose/metabolism , Glucose/metabolism , Metagenomics/methods , Soil Microbiology , beta-Galactosidase/isolation & purification , beta-Galactosidase/metabolism , Amino Acid Sequence , China , Cloning, Molecular , Enzyme Stability , Escherichia coli/genetics , Gene Expression , Gene Library , Molecular Sequence Data , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Sequence Alignment , Sequence Analysis, DNA , Temperature , beta-Galactosidase/chemistry , beta-Galactosidase/geneticsABSTRACT
Bisphenol A (BPA) is an endocrine disrupting chemical and broadly used in plastics. The leakage of BPA in food and water cycles poses a significant risk to the environment and human health. Thus, monitoring the concentration of BPA to avoid its potential risk is highly important. In this work, a simple and efficient oxygen deficient molecularly imprinted TiO2 electrochemical sensor was proposed for the detection of BPA. The introduction both oxygen vacancies and molecular imprinting evidently enhanced the electrochemical oxidation signal of BPA. The sensor had a good linear response ranging from 0.01 µM to 20 µM with a limit of detection of 3.6 nM. Additionally, the sensor showed remarkable stability, reproducibility and interference resistant ability. It also exhibits excellent recovery during the detection of real water. These findings suggested that the sensor has the potential to be developed as a simple, efficient and low-cost monitoring system for the monitoring of BPA in water.
Subject(s)
Biosensing Techniques , Molecular Imprinting , Humans , Oxygen , Reproducibility of Results , Water , Electrochemical Techniques , Limit of DetectionABSTRACT
OBJECTIVE: This study assessed the necessity of surgical re-staging in women with borderline ovarian tumors (BOTs) and evaluated the impact of complete surgical staging, lymphadenectomy, and omentectomy on disease recurrence and survival. METHODS: We retrospectively reviewed the medical records of patients with BOTs. A total of 901 patients were eligible for inclusion in the study, and we evaluated some of the variables and clinical/surgical characteristics of the cases. The effects of the type of surgical procedure, surgical staging, and complete or incomplete staging on recurrence were calculated. The rates of disease-free survival, overall survival, and recurrence were compared according to complete surgical staging. A Cox regression analysis was performed to identify potential prognostic factors, and survival curves were constructed using the Kaplan-Meier method. RESULTS: The overall recurrence rate was 13.9%, and recurrence was comparable between the complete surgical staging group and the incomplete groups (P>0.05). The performance of complete surgical staging did not show an effect on long-term survival, and complete surgical staging, omentectomy, and lymphadenectomy had no effect on recurrence. In multivariate analyses, only radical surgery and adjuvant chemotherapy were risk factors for the recurrence of BOTs. Furthermore, we found that omentectomy led to a relatively low recurrence rate in patients with International Federation of Gynecology and Obstetrics (FIGO) stage > I (P=0.022). CONCLUSION: Our results suggest that complete surgical staging should be considered a standard treatment for patients with advanced stage BOTs but not for those at FIGO stage I. It might be safe to reduce the scope of surgical procedures in patients with early-stage BOTs. However, it is not necessary to perform re-staging operations for BOTs with a macroscopically normal extra-ovarian appearance.
Subject(s)
Ovarian Neoplasms , Pregnancy , Humans , Female , Ovarian Neoplasms/pathology , Neoplasm Staging , Retrospective Studies , Neoplasm Recurrence, Local/surgery , Neoplasm Recurrence, Local/pathology , Disease-Free SurvivalABSTRACT
OBJECTIVE: To evaluate the relationship between gastric cancer and its precancerous lesions and gastric xanthoma. METHODS: Medical records of 47 736 patients who underwent gastroscopy in our center from January 2020 to December 2021 were reviewed. Patients' age, sex, endoscopic and histopathological findings, and the presence, number and location of gastric xanthoma were recorded. To investigate the detection rate of gastric xanthoma at different stages of gastric lesions, the participants were further divided into the chronic gastritis group (n = 42 758), the precancerous lesion group (n = 3672), and the gastric cancer group (n = 1306), respectively. RESULTS: The overall detection rate of gastric xanthoma was 2.85%, and it was most commonly observed in the gastric antrum (52.50%). In addition, gastric xanthoma was more common in men and more likely to be single lesion. It was most detected in the precancerous lesion group (8.39%), followed by the gastric cancer group (5.44%), and least in the chronic gastritis group (2.29%). Multivariate analysis showed that gastric xanthoma was closely related to precancerous lesions (odds ratio [OR] 3.197, 95% confidence interval [CI] 2.791-3.662, P < 0.001) and gastric cancer (OR 1.794, 95% CI 1.394-2.309, P < 0.001). CONCLUSION: Gastric xanthoma is closely related to gastric precancerous lesions and gastric cancer.
Subject(s)
Gastritis, Atrophic , Helicobacter Infections , Helicobacter pylori , Precancerous Conditions , Stomach Neoplasms , Xanthomatosis , Male , Humans , Stomach Neoplasms/etiology , Stomach Neoplasms/pathology , Retrospective Studies , Gastric Mucosa/pathology , Gastritis, Atrophic/pathology , Precancerous Conditions/pathology , Xanthomatosis/complications , Xanthomatosis/pathology , MetaplasiaABSTRACT
The extensive application of plastic products leads to the increasingly significant harm of plastic wastes to the ecological environment, which is also a focus of global environmental issues. Due to the lack of a sound plastic waste management system, most plastic waste is still treated by the traditional mode or remains in the environment, with low recycling efficiency, and the plastic life cycle has not yet formed. Plastics in the environment will age and degrade under the actions of physical (wear, waves), chemical (ultraviolet radiation, hydrolysis), and biological (fungi, bacteria) factors for a long time and generate micro (nano) plastics. Due to their small particle size, large specific surface area, and charged characteristics, in addition to their own toxicity, they can also be used as carriers or covert carriers of pollutants (heavy metals, persistent organic pollutants, polycyclic aromatic hydrocarbons, bacteria, etc.) to migrate in the environment through runoff, sewage discharge, and hydrometeorology, causing ecological environmental pollution. MPs pollution has been listed as the second largest scientific problem in the field of environmental and ecological science by the United Nations Environment Programme. MPs are widely distributed, and there are different degrees of MPs pollution in the global water (freshwater, ocean), soil, and atmospheric environment. Traces of MPs have also been found in human placentas, human breastmilk, living lungs, and blood in recent years. Therefore, the formation mechanisms of MPs under the actions of physics, chemistry, and microorganisms, as well as their abundance levels and migration characteristics in water, soil, and atmosphere environment were comprehensively reviewed, with the hope of providing reference for monitoring the pollution levels of MPs in the environment, exploring their transport laws in the environment, proposing the management strategy of MPs pollution, and revealing the degradation mechanisms of MPs under different effects.
Subject(s)
Microplastics , Plastics , Humans , Female , Pregnancy , Ultraviolet Rays , Atmosphere , EnvironmentABSTRACT
To obtain better performing laccases for textile dyes decolorization, random mutagenesis of Lac591, a metagenome-derived alkaline laccase, was carried out. After three rounds of error-prone PCR and high-throughput screening by assaying enzymatic activity toward the phenolic substrate 2,6-dimethoxyphenol (2,6-DMP), a mutant (Lac3T93) with remarkably improved enzymatic activity was obtained. Sequence analysis revealed that four amino acid substitutions (N40S, V55A, F62L, and E316V) were accumulated in the Lac3T93. Compared to the wild-type enzyme, the specific activity of Lac3T93 toward 2,6-DMP was increased to 4.8-fold (61.22 U/mg), and its optimal temperature and pH were changed to 60°C and 8.0 from 55°C and 7.5 of the wild-type enzyme, respectively. Furthermore, the degradation ability of Lac3T93 for textile dyes was investigated, and the new variant represented improved decolorization percentage for four industrial dyes with complex phenyl structure (Basic Blue 3, Methylene Blue, Bromophenol Blue, and Crystal Violet) and higher decolorization efficiency for Indigo Carmine than that of the parent enzyme. Furthermore, the decolorization percentage of Lac3T93 for five dyes in the absence of hydroxybenzotrizole (HBT) is clearly higher than those of the wild-type enzyme with 1 mM HBT, and HBT can further improve its decolorization ability.
Subject(s)
Laccase/metabolism , Metagenome , Amino Acid Sequence , Base Sequence , Coloring Agents/chemistry , Coloring Agents/metabolism , Escherichia coli/genetics , High-Throughput Screening Assays , Laccase/genetics , Mutagenesis , Mutation , Polymerase Chain Reaction , Pyrogallol/analogs & derivatives , Pyrogallol/metabolism , Sequence Analysis, DNA , TextilesABSTRACT
Lac591, a gene encoding a novel multicopper oxidase with laccase activity, was identified through activity-based functional screening of a metagenomic library from mangrove soil. Sequence analysis revealed that lac591 encodes a protein of 500 amino acids with a predicted molecular mass of 57.4 kDa. Lac591 was overexpressed heterologously as soluble active enzyme in Escherichia coli and purified, giving rise to 380 mg of purified enzyme from 1 l induced culture, which is the highest expression report for bacterial laccase genes so far. Furthermore, the recombinant enzyme demonstrated activity toward classical laccase substrates syringaldazine (SGZ), guaiacol, and 2, 6-dimethoxyphenol (2, 6-DMP). The purified Lac591 exhibited maximal activity at 55 degrees C and pH 7.5 with guaiacol as substrate and was found to be stable in the pH range of 7.0-10.0. The substrate specificity on different substrates was studied with the purified enzyme, and the optimal substrates were in the order of 2, 6-DMP > catechol > alpha-naphthol > guaiacol > SGZ > 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid). The alkaline activity and highly soluble expression of Lac591 make it a good candidate of laccases in industrial applications for which classical laccases are unsuitable, such as biobleaching of paper pulp and dyestuffs processing.
Subject(s)
Bacterial Proteins/chemistry , Cloning, Molecular , Laccase/chemistry , Metagenome , Oxidoreductases/chemistry , Soil Microbiology , Amino Acid Sequence , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Enzyme Stability , Gene Expression , Kinetics , Laccase/genetics , Laccase/metabolism , Molecular Sequence Data , Oxidoreductases/genetics , Oxidoreductases/metabolism , Sequence Alignment , Solubility , Substrate SpecificityABSTRACT
A novel beta-galactosidase gene, zd410, was isolated by screening a soil metagenomic library. Sequence analysis revealed that zd410 encodes a protein of 672 amino acids with a predicted molecular weight of 78.6 kDa. The recombinant ZD410 was expressed and purified in Pichia pastoris, with a yield of ca. 300 mg from 1 L culture. The purified enzyme displayed optimal activity at 38 degrees C and pH 7.0. Given that the enzyme had 54% of the maximal activity at 20 degrees C and 11% of the maximal activity at close to 0 degrees C, ZD410 was regarded as a cold-adapted beta-galactosidase. ZD410 displays high enzymatic activity for its synthetic substrate-ONPG (o-nitrophenyl-beta-D-galactopyranoside, 243 U/mg) and its natural substrate-lactose (25.4 U/mg), while its activity was slightly stimulated by addition of Na(+), K(+), or Ca(2+) at low concentrations. ZD410 is a good candidate of beta-galactosidases for food industry after further study.
Subject(s)
Metagenome , Soil Microbiology , beta-Galactosidase/genetics , beta-Galactosidase/metabolism , Calcium/pharmacology , Cloning, Molecular , Cluster Analysis , Coenzymes/pharmacology , Enzyme Stability , Gene Expression , Hydrogen-Ion Concentration , Lactose/metabolism , Molecular Sequence Data , Molecular Weight , Nitrophenylgalactosides/metabolism , Phylogeny , Pichia/genetics , Potassium/pharmacology , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Sodium/pharmacology , Temperature , beta-Galactosidase/chemistry , beta-Galactosidase/isolation & purificationABSTRACT
BACKGROUND: Pyrethroids and pyrethrins are widely used insecticides. Extensive applications not only result in pest resistance to these insecticides, but also may lead to environmental issues and human exposure. Numerous studies have shown that very high exposure to pyrethroids might cause potential problems to man and aquatic organisms. Therefore, it is important to develop a rapid and efficient disposal process to eliminate or minimize contamination of surface water, groundwater and agricultural products by pyrethroid insecticides. Bioremediation is considered to be a reliable and cost-effective technique for pesticides abatement and a major factor determining the fate of pyrethroid pesticides in the environment, and suitable esterase is expected to be useful for potential application for detoxification of pyrethroid residues. Soil is a complex environment considered as one of the main reservoirs of microbial diversity on the planet. However, most of the microorganisms in nature are inaccessible as they are uncultivable in the laboratory. Metagenomic approaches provide a powerful tool for accessing novel valuable genetic resources (novel enzymes) and developing various biotechnological applications. RESULTS: The pyrethroid pesticides residues on foods and the environmental contamination are a public safety concern. Pretreatment with pyrethroid-hydrolyzing esterase has the potential to alleviate the conditions. To this end, a pyrethroid-hydrolyzing esterase gene was successfully cloned using metagenomic DNA combined with activity-based functional screening from soil, sequence analysis of the DNA responsible for the pye3 gene revealed an open reading frame of 819 bp encoding for a protein of 272 amino acid residues. Extensive multiple sequence alignments of the deduced amino acid of Pye3 with the most homologous carboxylesterases revealed moderate identity (45-49%). The recombinant Pye3 was heterologously expressed in E. coli BL21(DE3), purified and characterized. The molecular mass of the native enzyme was approximately 31 kDa as determined by gel filtration. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the deduced amino acid sequence of the Pye3 indicated molecular mass of 31 kDa and 31.5 kDa, respectively, suggesting that the Pye3 is a monomer. The purified Pye3 not only degraded all pyrethroid pesticides tested, but also hydrolyzed rho-nitrophenyl esters of medium-short chain fatty acids, indicating that the Pye3 is an esterase with broader specificity. The Km values for trans-Permethrin and cis-permethrin are 0.10 muM and 0.18 muM, respectively, and these catalytic properties were superior to carboxylesterases from resistant insects and mammals. The catalytic activity of the Pye3 was strongly inhibited by Hg2+, Ag+, rho-chloromercuribenzoate, whereas less pronounced effect was observed in the presence of divalent cations, the chelating agent EDTA and phenanthroline. CONCLUSION: A novel pyrethroid-hydrolyzing esterase gene was successfully cloned using metagenomic DNA combined with activity-based functional screening from soil, the broader substrate specificities and higher activity of the pyrethroid-hydrolyzing esterase (Pye3) make it an ideal candidate for in situ for detoxification of pyrethroids where they cause environmental contamination problems. Consequently, metagenomic DNA clone library offers possibilities to discover novel bio-molecules through the expression of genes from uncultivated bacteria.
ABSTRACT
Lignin polymers in bamboo (Phyllostachys pubescens) were decomposed into polyphenols at high temperatures and oxidized for the introduction of quinone groups from peroxidase extracted from bamboo shoots and catalysis of UV. According to the results of FT-IR spectra analysis, neutral proteases (NPs) can be immobilized on the oxidized lignin by covalent bonding formed by amine group and quinone group. The optimum condition for the immobilization of NPs on the bamboo bar was obtained at pH 7.0, 40 °C, and duration of 4 h; the amount of immobilized enzyme was up to 5 mg g-1 bamboo bar. The optimal pH for both free NP (FNP) and INP was approximately 7.0, and the maximum activity of INP was determined at 60 °C, whereas FNP presented maximum activity at 50 °C. The Km values of INP and FNP were determined as 0.773 and 0.843 mg ml-1, respectively; INP showed a lower Km value and Vmax, than FNP, which demonstrated that INP presented higher affinity to substrate. Compared to FNP, INP showed broader thermal and storage stability under the same trial condition. With respect to cost, INP presented considerable recycling efficiency for up to six consecutive cycles.
Subject(s)
Bambusa/chemistry , Enzymes, Immobilized/metabolism , Lignin/chemistry , Peptide Hydrolases/metabolism , Enzyme Stability , Hot Temperature , Hydrogen-Ion Concentration , Oxidation-Reduction , Protein Binding , Spectroscopy, Fourier Transform Infrared , Time Factors , Ultraviolet RaysABSTRACT
OBJECTIVES: Protein arginine methyltransferase 2 (PRMT2) protects against vascular injury-induced intimal hyperplasia; however, little is known about the role of PRMT2 in angiotensin II (Ang II)-induced VSMCs proliferation and inflammation. This research aims to determine whether PRMT2 inhibits Ang II-induced proliferation and inflammation of vascular smooth muscle cells (VSMCs). MATERIALS AND METHODS: PRMT2 overexpression was used to elucidate the role of PRMT2 in Ang II-induced VSMCs proliferation and inflammation. Western blotting and reverse transcriptional PCR were adopted to detect protein and mRNA expression severally. Cell viability was evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay and cell cycle distribution by flow cytometry. RESULTS: Ang II significantly reduced mRNA and protein levels of PRMT2 in VSMCs in time-dependent and dose-dependent manner. Results of PRMT2 overexpression indicated that PRMT2 inhibited proliferation of VSMCs stimulated with 100 nmol/L Ang II for 24 hours. Furthermore, overexpression of PRMT2 reduced Ang II-induced production of proinflammatory cytokines such as interleukin 6 (IL-6) and interleukin 1ß (IL-1ß) in VSMCs. CONCLUSIONS: These findings suggest that PRMT2 alleviates Ang II-induced VSMCs proliferation and inflammation, providing a new mechanism about how Ang II mediated VSMCs proliferation and inflammation.
Subject(s)
Cell Proliferation/physiology , Inflammation , Intracellular Signaling Peptides and Proteins/physiology , Muscle, Smooth, Vascular/physiology , Protein-Arginine N-Methyltransferases/physiology , Angiotensin II , Cells, Cultured , Humans , Myocytes, Smooth MuscleABSTRACT
In this work, a cultivation system with daily recycling of the post-harvest culture broth was set up and performed in order to reuse the water and nutrients in pretreated anaerobically digested swine manure, which was used as media to cultivate Chlorella vulgaris (UTEX 2714) at different recycling ratios. Results showed that the alga grew well in the system with an accumulative algal biomass and productivity of 1.68-3.47g/L and 234.1-532.2mg/L/d, respectively, at the end of the cultivation. Additionally, chemical compositions in this alga varied with the change of recycling ratios, and the highest productivities of carbohydrate, protein and lipids (76.4, 257.2 and 183.7mg/L/d, respectively) were obtained in the system with a recycling ratio of 1/4 or 1/6. Fatty acid profiles indicated that this alga could be used as a good-quality biodiesel feedstock with a biodiesel productivity of 9.65-40.1mg/L/d.
Subject(s)
Biofuels , Chlorella vulgaris , Manure , Recycling , Animals , Biomass , Chlorella , Lipids , SwineABSTRACT
Liquid swine manure was subjected to thermophilic anaerobic digestion, ammonia stripping and centrifugation in order to increase the available carbon sources and decrease the ammonia concentration and turbidity. Chlorella vulgaris (UTEX 2714) was grown on minimally diluted (2×, 3× and 4×) autoclaved and non-autoclaved pretreated anaerobic digestion swine manure (PADSM) in a batch-culture system for 7days. Results showed that C. vulgaris (UTEX 2714) grew best on 3× PADSM media, and effectively removed NH4+-N, TN, TP and COD by 98.5-99.8%, 49.2-55.4%, 20.0-29.7%, 31.2-34.0% and 99.8-99.9%, 67.4-70.8%, 49.3-54.4%, 73.6-78.7% in differently diluted autoclaved and non-autoclaved PADSM, respectively. Results of chemical compositions indicated that contents of pigment, carbohydrate, protein and lipid in C. vulgaris (UTEX 2714) changed with the culture conditions. Moreover, its fatty acid profiles suggested that this alga could be used as animal feed if cultivated in autoclaved PADSM or as good-quality biodiesel feedstock if cultivated in non-autoclaved PADSM.
Subject(s)
Biofuels , Chlorella vulgaris , Manure , Anaerobiosis , Animals , Biomass , SwineABSTRACT
OBJECTIVES: To evaluate the efficacy and safety of Bushen Huoxue prescription (BSHXP) for endometriosis. METHODS: A meta-analysis was performed, and studies were searched from the seven databases from the date of database establishment to April 30, 2017. Randomized controlled trials (RCTs) that explored the efficacy and safety of BSHXP for patients with endometriosis were included. Two assessors independently reviewed each trial. The Cochrane Risk of Bias assessment tool was used for quality assessment. RESULTS: In the 13 included studies, the total effectiveness rates of BSHXP were higher than those of Western medicine (RR, 1.55; 95% CI, 1.03-2.32; P = 0.04), but the dysmenorrhea alleviation rates of the two treatments did not significantly differ (RR, 1.28; 95% CI, 0.70-2.34; P = 0.42). The pregnancy rates of BSHXP were also higher than those of hormone therapy (RR, 1.99; 95% CI, 1.17-3.39; P = 0.01). However, whether BSHXP is more effective than Western medicine in diminishing endometriotic cyst remains unknown. CONCLUSIONS: Our study provides evidence that BSHXP is effective and safe for endometriosis, but this evidence is inconclusive because of the low methodological quality of the included RCTs. Our findings suggest that BSHXP is an alternative drug for endometriosis, but it should be further examined in future clinical research.
ABSTRACT
OBJECTIVE: To investigate whether the human endometrial carcinoma cells, RL95-2 infected with recombinant Ad-PTEN can steadily produce PTEN protein and enter apoptosis. METHODS: The recombinant adenovirus containing PTEN cDNA was constructed using the method of homologous recombination in bacteria. The viral titer was examined by plaque assay and the expression of PTEN protein was detected by western blot assay. The apoptosis of RL95-2 cells was evaluated as following: flipping of membrane phosphatidylserine (PS) and identification of activating caspase-3 positive cells was determined by flow cytometer (FCM), and furthermore genomic DNA fragmentation was detected by agarose electrophoresis. RESULTS: The recombinant adenovirus encoding PTEN cDNA was successfully constructed, and viral titers of Ad-PTEN were 5 x 10(9) pfu/ml. After infected by Ad-PTEN, the expression of PTEN protein was steady in human RL95-2 cells. After infected by Ad-PTEN for 24, 48, 72 and 96 h, the relative cell number of membrane PS flipping were (6.09 +/- 1.01)%, (9.98 +/- 2.17)%, (11.74 +/- 2.65)%, (27.69 +/- 8.67)%, which significantly increased than control group (P < 0.05), the relative cell number of activated caspase-3 positive were (2.6 +/- 0.5)%, (18.0 +/- 4.4)%, (21.8 +/- 5.1)%, (33.7 +/- 9.9)%, respectively, which significantly increased than control group (P < 0.05), and genomic DNA fragmentation was verified also. CONCLUSIONS: The recombinant Ad-PTEN vector is constructed successfully and the expression of specific PTEN is steady in RL95-2 cell line. The expression of PTEN induces RL95-2 cells to apoptosis. PTEN gene may be a novel therapeutic target in endometrial carcinoma.