ABSTRACT
We report an electrochemical device for portable on-site detection of gaseous CH3I based on PVIm-F for the first time. The device achieves detection of gaseous CH3I with a significant selectivity and a low detection limit (0.474 ppb) in 20 min at 50 °C and 50% relative humidity, which is of great significance for achieving real-time on-site monitoring of radioactive hazardous environments.
ABSTRACT
The ability to effectively detect bacterial infection in human tissues is important for the timely treatment of the infection. However, traditional techniques fail to visualize bacterial species adhered to host cells in situ in a target-specific manner. Dihydropteroate synthase (DHPS) exclusively exists in bacterial species and metabolically converts p-aminobenzoic acid (PABA) to folic acid (FA). By targeting this bacterium-specific metabolism, we have developed a fluorescent imaging probe, PABA-DCM, based on the conjugation of PABA with a long-wavelength fluorophore, dicyanomethylene 4H-pyran (DCM). We confirmed that the probe can be used in the synthetic pathway of a broad spectrum of Gram-positive and negative bacteria, resulting in a significantly extended retention time in bacterial over mammalian cells. We validated that DHPS catalytically introduces a dihydropteridine group to the amino end of the PABA motif of PABA-DCM, and the resulting adduct leads to an increase in the FA levels of bacteria. We also constructed a hydrogel dressing containing PABA-DCM and graphene oxide (GO), termed PABA-DCM@GO, that achieves target-specific fluorescence visualization of bacterial infection on the wounded tissues of mice. Our research paves the way for the development of fluorescent imaging agents that target species-conserved metabolic pathways of microorganisms for the in situ monitoring of infections in human tissues.
Subject(s)
4-Aminobenzoic Acid , Bacterial Infections , 4-Aminobenzoic Acid/metabolism , Animals , Bacterial Infections/diagnostic imaging , Dihydropteroate Synthase/metabolism , Folic Acid/metabolism , Humans , Mammals/metabolism , MiceABSTRACT
In plants, NO has been proved the function of improving abiotic stress resistance. However, the role of NO in the lignin metabolism of okra under cold stress has not been clarified. Here, histochemical staining and lignin content analysis showed that cold stress promoted the lignin accumulation of cold stored okra pods, and NO inhibited the lignin accumulation and delayed lignification process. To better understand the roles of NO in okra cold stress resistance mechanism, the full-length transcriptome data of 'Hokkaido' was analyzed. The SNP-treated okra transcriptome and cPTIO-treated okra transcriptome were obtained. A total of 41957 unigenes were screened out from three groups at 10 d, among which, 33, 78 and 18 DEGs were found in ddH2O-treat, SNP-treat and cPTIO-treat group, respectively. Transcriptomic data suggested that the genes involved in lignin biosynthesis showed downregulation under SNP treatment. Transcriptomic data and enzyme activity showed that exogenous NO significantly promoted the biosynthesis of endogenous NO by enhancing NOS activity. Transcriptomic data and plant hormone data showed that NO played an important role in the process of inhibiting the ethylene and ABA synthesis mechanism of okra and thereby reducing the endogenous ethylene and ABA content under chilling stress. Relevant physiological data showed that NO helped to the protection of ROS scavenging system and removed the MDA and H2O2 induced by cold stress. These results provided a reference for studying the molecular mechanism of nitric oxide delaying the lignification of okra, and also provided a theoretical basis for postharvest storage of vegetables.
Subject(s)
Abelmoschus , Gene Expression Profiling , Hydrogen Peroxide , Lignin , Nitric Oxide , TranscriptomeABSTRACT
Oncolytic viruses (OV) have shown excellent safety and efficacy in preclinical and clinical studies. Influenza A virus (IAV) is considered a promising oncolytic virus. In this report, we generated a recombinant influenza virus expressing an immune checkpoint blockade agent targeting CTLA4. Using reverse genetics, a recombinant influenza virus, termed rFlu-CTLA4, encoding the heavy chain of a CTLA4 antibody on the PB1 segment and the light chain of the CTLA4 antibody on the PA segment was produced. RFlu-CTLA4 could replicate to high titers, and antibodies were produced in the allantoic fluid of infected eggs. Furthermore, the selective cytotoxicity of the virus was higher in various hepatocellular carcinoma cancer cell lines than in the normal cell line L02 in vitro, as indicated by MTS assays. More importantly, in a subcutaneous H22 mouse hepatocarcinoma model, intratumoral injections of rFlu-CTLA4 inhibited the growth of treated tumors and increased the overall survival of mice compared with injections of the PR8 virus. Taken together, these results warrant further exploration of this novel recombinant influenza virus for its potential use as a single or combination agent for cancer immunotherapy.
Subject(s)
CTLA-4 Antigen/immunology , Immunotherapy/methods , Influenza A virus/immunology , Neoplasms/therapy , Oncolytic Virotherapy/methods , Oncolytic Viruses/immunology , Animals , Hep G2 Cells , Humans , Mice , Mice, Inbred BALB CABSTRACT
Ischemic stroke is a life-threatening complication with a high rate of morbidity. Circulating fatty acid binding protein 4 (FABP4) has been reported to be associated with the outcome of acute ischemic stroke. The present study aimed to illustrate the function of FABP4 in ischemic stroke. PC12 cells exposed to oxygen glucose deprivation/reoxygenation (OGD/R) were used to mimic ischemia-reperfusion (I/R) injury in ischemic stroke. Cell viability was estimated using a Cell Counting Kit-8 assay. The expression of FABP4 in PC12 cells under OGD/R was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). PC12 cells were transfected with FABP4 small interfering RNA (siRNA), inflammatory cytokines and reactive oxygen species (ROS) were determined via RT-qPCR and ROS assay kit. Western blotting was performed to detect endoplasmic reticulum stress (ERS)-related proteins and peroxisome proliferator-activated receptor γ (PPARγ). Flow cytometry was used to evaluate the cell apoptotic rate. The expression of FABP4 increased gradually with the prolongation of reoxygenation within 8 h. FABP4-knockdown inhibited the transcription of inflammatory cytokines, the production of ROS and decreased cell apoptosis. Furthermore, decreased ERS-related proteins and increased PPARγ were estimated in PC12 cells transfected with FABP4 siRNA. PPARγ inhibitor GW9662 weakened the anti-apoptotic effect of FABP4-knockdown. Taken together, these results indicated that FABP4-knockdown suppressed cell apoptosis via relieving ERS; this effect was reversed by treatment of GW9662.
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OBJECTIVE: To explore potential mechanisms and effects of gallincin on a mouse model of colitis induced by dextran sulfate sodium (DSS). METHODS: Network pharmacology analysis was used to predict the molecular mechanism of action of gallincin for treatment of colitis. Gallincin was administered orally to mice with DSS-induced colitis. Expression of tumor necrosis factor α (TNF-α), D-lactate, and interleukin-1ß (IL-1ß) and myeloperoxidase activity were assessed with real-time quantitative PCR and an enzyme-linked immunoassay, respectively. Expression of occludin, zonula occludens 1 (ZO-1), and phosphorylated extracellular signal-regulated protein kinase1/2 (p-ERK1/2) was analyzed with immunohistochemical staining and/or western blot assays. RESULTS: Using a network pharmacology approach, 12 mapping targets between gallincin and colitis were obtained, including ERK/mitogen-activated protein kinase. Further investigations in an experimental colitis mouse model showed that gallincin significantly ameliorated experimental colitis, reduced D-lactate levels, and remarkably increased occludin and ZO-1 expression, possibly in part by decreasing IL-1ß, TNF-α, and p-ERK1/2 levels and inhibiting leukocyte penetration. CONCLUSIONS: Gallincin regulated colonic barrier function and reduced colitis-associated inflammation, suggesting it is a promising drug for the treatment of ulcerative colitis.
Subject(s)
Colitis, Ulcerative , Colitis , Animals , Colitis/chemically induced , Colitis/drug therapy , Colitis, Ulcerative/chemically induced , Colitis, Ulcerative/drug therapy , Dextran Sulfate/toxicity , Disease Models, Animal , Inflammation/drug therapy , Mice , Mice, Inbred C57BLABSTRACT
In plants, NAC (NAM, ATAF, CUC) is a class of transcription factors (TFs) involved in growth regulation and is associated with abiotic stress, morphogenesis, and metabolism. Celery (Apium graveolens L.) is an important leafy vegetable and its yield and quality are considerably influenced by environmental factors. Currently, the characterization of NAC genes in celery is still limited. In this study, a total of 111 putative NAC TFs were determined based on the celery transcriptome and genome database. They were divided into 18 subfamilies on the basis of their NAC domain. NAC TFs in celery account for a moderate number compared with other species, similar to that in carrot. Real-time quantitative PCR (RT-qPCR) showed that some AgNAC genes were differentially expressed under adverse conditions (heat, cold, drought, and salt). AgNAC63 (ortholog of ANAC072/RD26) was highly induced by heat, cold, and salt conditions. The expression levels of AgNAC47 in leaf blades were 105.25- and 123.14-fold those of petioles and roots, respectively. AgNAC63 and AgNAC47 showed significant tissue specificity, high expression in leaves, and varying degrees of response under the four treatments. This study provides a basis for the improved investigation of the structure and function of AgNAC TFs and celery stress resistance.
Subject(s)
Apium/genetics , Genes, Plant , Genome , Plant Proteins/analysis , Stress, Physiological/genetics , Transcription Factors/analysis , Arabidopsis/genetics , Droughts , Gene Expression Regulation, Plant , Organ Specificity/genetics , Phylogeny , Plant Leaves/genetics , Plant Proteins/genetics , Plant Roots/genetics , Temperature , Transcription Factors/genetics , TranscriptomeABSTRACT
BACKGROUND: The aim of this study was to determine the association between hypertriglyceridemia and hyperuricemia (HUA). METHODS: The study was conducted in 3884 subjects who had not received medication enrolled as a baseline. Each participant received at least three annual health check-ups between 2011 and 2017. The risk of hyperuricemia was assessed in four Quartiles (Q1 to Q4) according to TG levels using multivariate-adjusted logistic regression models. RESULTS: The total incidence rate of HUA was 62.3/1000 person-years. In the univariate analysis, the risk of hyperuricemia in people with hypertriglyceridemia was 2.353 times that of normal triglycerides, with a 95% confidence interval of (2.011, 2.754), and the risk of hyperuricemia in men was 1.86 times of female, and the 95% confidence interval is (1.634, 2.177). After adjusting the potential confounders, the relative risk RR of TG at Q2 Q3 Q4 was 1.445 (95%CI:1.114, 1.901), 2.075 (1.611, 2.674), 2.972 (2.322, 3.804). CONCLUSIONS: TG is an independent risk factor for hyperuricemia. As the level of TG increases, the risk of HUA increases.
Subject(s)
Hypertriglyceridemia/epidemiology , Hyperuricemia/epidemiology , Lipid Metabolism , Triglycerides/blood , Adult , Aged , Female , Humans , Hypertriglyceridemia/blood , Hypertriglyceridemia/pathology , Hyperuricemia/blood , Hyperuricemia/pathology , Male , Middle Aged , Risk Factors , Sex Characteristics , Urban Population , Uric Acid/bloodABSTRACT
Background Clinical evidence indicates that genetic variations may interfere with the mechanism of drug action. Recently, it has been reported that the single nucleotide polymorphisms (SNPs) of STAT4, PTPN2, PSORS1C1 and TRAF3IP2RA genes are associated with the clinical efficacy of tumor necrosis factor (TNF) inhibitors in the treatment of rheumatoid arthritis (RA) patients. Therefore, the detection of the SNPs linked with TNF inhibitor efficacy may provide an important basis for the treatment of RA. This study intended to establish molecular diagnostic methods for genotyping the linked SNPs based on high resolution melting (HRM) curve analysis. Methods The polymerase chain reaction-HRM (PCR-HRM) curve analysis detecting systems were established by designing the primers of the four SNPs, rs7574865G>T, rs7234029A>G, rs2233945C>A and rs33980500C>T, and the performance and clinical applicability of which were evaluated by using the Sanger sequencing method and genotyping test for 208 clinical samples. Results The self-developed molecular diagnostic methods of PCR-HRM were confirmed to be able to correctly genotype the four SNPs, the sensitivity and specificity of which were 100% in this study. The repeatability and reproducibility tests showed that there is little variable in intra-assay and inter-assay (the coefficient of variation ranged from 0.01% to 0.07%). The slight changes of DNA template and primer concentrations, PCR cycle number and reaction system volume had no significant effect on the genotyping performance of the method. The PCR-HRM assays were also applied to other PCR thermocyclers with HRM function and use different saturation fluorescent dyes. Conclusions The PCR-HRM genotyping method established in this study can be applied to the routine molecular diagnosis of rs7574865, rs7234029, rs2233945 and rs33980500.
Subject(s)
Genotyping Techniques , Molecular Diagnostic Techniques , Polymerase Chain Reaction , Polymorphism, Single Nucleotide , Tumor Necrosis Factor Inhibitors , Arthritis, Rheumatoid/drug therapy , Arthritis, Rheumatoid/genetics , Female , Genotype , Humans , Male , Middle Aged , Pathology, Molecular , Protein Tyrosine Phosphatase, Non-Receptor Type 2/genetics , Proteins/genetics , Reproducibility of Results , STAT4 Transcription Factor/genetics , Sensitivity and SpecificityABSTRACT
PURPOSE: Magnetic submicron particles (MSPs) are pivotal biomaterials for magnetic separations in bioanalyses, but their preparation remains a technical challenge. In this report, a facile one-step coating approach to MSPs suitable for magnetic separations was investigated. METHODS: Polyethylene glycol) (PEG) was derived into PEG-bis-(maleic monoester) and maleic monoester-PEG-succinic monoester as the monomers. Magnetofluids were prepared via chemical co-precipitation and dispersion with the monomers. MSPs were prepared via one-step coating of magnetofluids in a water-in-oil microemulsion system of aerosol-OT and heptane by radical co-polymerization of such monomers. RESULTS: The resulting MSPs contained abundant carboxyl groups, exhibited negligible nonspecific adsorption of common substances and excellent suspension stability, appeared as irregular particles by electronic microscopy, and had submicron sizes of broad distribution by laser scattering. Saturation magnetizations and average particle sizes were affected mainly by the quantities of monomers used for coating magnetofluids, and steric hindrance around carboxyl groups was alleviated by the use of longer monomers of one polymerizable bond for coating. After optimizations, MSPs bearing saturation magnetizations over 46 emu/g, average sizes of 0.32 µm, and titrated carboxyl groups of about 0.21 mmol/g were obtained. After the activation of carboxyl groups on MSPs into N-hydroxysuccinimide ester, biotin was immobilized on MSPs and the resulting biotin-functionalized MSPs isolated the conjugate of streptavidin and alkaline phosphatase at about 2.1 mg/g MSPs; streptavidin was immobilized at about 10 mg/g MSPs and retained 81% ± 18% (n = 5) of the specific activity of the free form. CONCLUSION: The facile approach effectively prepares MSPs for magnetic separations.
Subject(s)
Magnetite Nanoparticles/chemistry , Maleates/chemistry , Polyethylene Glycols/chemistry , Adsorption , Biotin/chemistry , Immobilized Proteins/chemistry , Immobilized Proteins/metabolism , Molecular Conformation , Particle Size , Polymerization , Protein Binding , Streptavidin/chemistry , Streptavidin/metabolismABSTRACT
Limonium sinense (Girard) Kuntze is a traditional Chinese folk medicine used for the treatment of fever, hemorrhage, hepatitis and other disorders. The study focused on the antitumor and immunomodulatory activities of L. sinense polysaccharides (LSP) which was obtained from the root of the plant. The antitumor effects of only LSP and LSP in combination with 5-fluorouracil (5-FU) were both evaluated with Heps-bearing tumor mice models. In addition, the macrophage phagocytosis assay, splenocyte proliferation and cytokines production tests were used to assess the immunomodulatory activities of LSP. The results revealed that the LSP (at the dose of 200 and 400 mg/kg) had an obvious inhibition on the growth of transplanted mouse tumor. It also exhibited a significant synergistic effect of antitumor activity when combined with 5-FU (p<0.05). Furthermore, the LSP (at the dose of 100 and 200 mg/kg) remarkably improved macrophage phagocytosis function in immune suppressed mice. In addition, LSP (at the dose of 50-200 µg/ml) showed significant synergistic effects on ConA-stimulated proliferation and IFN-γ and IL-2 production of splenocyte in vitro (p<0.05). These findings suggest that LSP had clear antitumor activity which might be related to its regulation of immune function in mice.
Subject(s)
Antineoplastic Agents/pharmacology , Immunologic Factors/pharmacology , Plant Roots/chemistry , Plumbaginaceae/chemistry , Polysaccharides/pharmacology , Animals , Antineoplastic Agents/isolation & purification , Cell Line, Tumor , Cell Proliferation/drug effects , Cytokines/biosynthesis , Drug Synergism , Fluorouracil/pharmacology , Humans , Immunologic Factors/isolation & purification , Macrophages/drug effects , Macrophages/immunology , Mice , Phagocytosis/drug effects , Polysaccharides/isolation & purification , Spleen/cytology , Xenograft Model Antitumor AssaysABSTRACT
Modification of accessible amino acid residues with poly(ethylene glycol) [PEG] is a widely used technique for formulating therapeutic proteins. In practice, site-specific PEGylation of all selected/engineered accessible nonessential reactive residues of therapeutic proteins with common activated PEG derivatives is a promising strategy to concomitantly improve pharmacokinetics, allow retention of activity, alleviate immunogenicity, and avoid modification isomers. Specifically, through molecular engineering of a therapeutic protein, accessible essential residues reactive to an activated PEG derivative are substituted with unreactive residues provided that protein activity is retained, and a limited number of accessible nonessential reactive residues with optimized distributions are selected/introduced. Subsequently, all accessible nonessential reactive residues are completely PEGylated with the activated PEG derivative in great excess. Branched PEG derivatives containing new PEG chains with negligible metabolic toxicity are more desirable for site-specific PEGylation. Accordingly, for the successful formulation of therapeutic proteins, optimization of the number and distributions of accessible nonessential reactive residues via molecular engineering can be integrated with the design of large-sized PEG derivatives to achieve site-specific PEGylation of all selected/engineered accessible reactive residues.
Subject(s)
Amino Acids/chemistry , Drug Carriers/chemistry , Polyethylene Glycols/chemistry , Protein Engineering/methods , Proteins , Binding Sites , Molecular Structure , Mutagenesis, Site-Directed , Proteins/administration & dosage , Proteins/chemistry , Proteins/genetics , Sulfhydryl Compounds/chemistryABSTRACT
At 0.12 mmol/L γ-glutamyl p-nitroaniline (GGPNA), an improved integrated method was developed for kinetic analysis of γ-glutamyltransferase (GGT) reaction process and the integration with the classical initial rate method to measure serum GGT. For the improved integrated method, an integrated rate equation, which used the predictor variable of reaction time and considered inhibitions by both GGPNA and products, was nonlinearly fit to GGT reaction processes. For the integration strategy, classical initial rates were estimated when GGPNA consumption percentages were below 50%; otherwise, maximal reaction rates of GGT were estimated by the improved integrated method and converted into initial rates according to the differential rate equation at 0.11 mmol/L GGPNA. The integration strategy was validated using optimized GGT kinetic parameters and 10-s intervals to record reaction curves within 8.0 min. By the integration strategy, there was a linear response from 0.9 to 32.0 U/L GGT, coefficients of variation were below 3.5% for GGT from 8.0 to 32.0 U/L (n=5), and GGT activities in clinical sera responded linearly to their classical initial rates at 2.00 mmol/L GGPNA with an expected slope. Therefore, the integration strategy was successful in measuring GGT at 0.12 mmol/L GGPNA.
Subject(s)
Aniline Compounds/chemistry , gamma-Glutamyltransferase/genetics , Animals , Biochemistry/methods , Kidney/enzymology , Kinetics , Models, Statistical , Swine , Time FactorsABSTRACT
OBJECTIVE: To investigate the clinical efficacy of cryotherapy ablation treatment for advanced hepatocellular carcinoma, and to analyse the predictive factors of cryotherapy ablation treatment. METHODS: 190 patients of hepatitis B-related advanced HCC from 2005 to 2008 in our hospital underwent curative cryoablation. We used clinical cohort method to analyze cryoablation group (147 cases) and control group (43 cases). The median OS (over survival time) and TTP (time to disease progression) were compared. We also evaluated the clinical significance of age, gender, location of portal vein tumor thrombus, HBeAg, tumor histological grade, Child-Pugh classification, end-stage liver disease (MELD) score, advanced liver cancer prediction system (ALCPS) score and the Eastern Cooperative Oncology Group performance status (ECOG PS) score for predicting the efficacy of cryoablation. Two Groups were compared with the x² test. Survival rates were estimated by the Kaplan-Meier method and compared by the log rank test. The Cox proportional hazards model was used to determine the independent factors on survival based on the variables selected in univariate analysis. RESULTS: Median survival time of cryoablation group and Control group were 7.5 (4.2 to 14.6) months and 3.2 (1.2 to 8.6) months, median TTP were 3.5 (2.5 to 4.5) months and 1.5 (1.0 to 3.5 months), the differences between were statistically significant (P < 0.05). Median OS and TTP of advanced HCC patients who had Well-differentiated tumor, Child-pugh A-class and low score of MELD score, ALCPS score; ECOG PS score were significantly longer than that of the poorly differentiated tumor, Child-pugh B-class and the high those scores (P < 0.05). ECOG PS (P less than 0.05, 95% CI 1.074 - 2.143) and ALCPS (P < 0.05, 95% CI 1.005-2.121) were independent predictors for OS of advanced HCC. CONCLUSIONS: Cryoablation treatment can prolong median OS and TTP of advanced HCC. ECOG PS and ALCPS are important predictors for survival time of advanced HCC.
Subject(s)
Azo Compounds/pharmacology , Carcinoma, Hepatocellular/metabolism , Gonanes/pharmacology , Liver Neoplasms/metabolism , Cell Line, Tumor/drug effects , Cell Line, Tumor/metabolism , Humans , ProteomicsABSTRACT
BACKGROUND: Knowledge about factors influencing the prognosis of resective epilepsy surgery can be used to identify which patients are most suitable for surgical treatment. The aim of this study was to identify preoperative prognostic factors associated with the chance of achieving long-term seizure freedom. METHODS: We retrospectively reviewed seizure outcomes and clinical, electroencephalography (EEG), magnetic resonance imaging (MRI), histopathology, and surgical variables from 99 epilepsy surgery patients with at least one year of postoperative follow-up. Seizure outcomes were categorized based on the modified classification by the International League Against Epilepsy. RESULTS: We found that the seizure-free rate was 27.9% after one year, and that it stabilized at about 20.0% between two and six years after surgery. Univariate analysis showed that medial temporal lobe epilepsy with hippocampal sclerosis, MRI with visible focal lesions concordant with EEG, and regional ictal EEG and electrocorticography patterns were associated with a favorable surgical outcome. On the other hand, seizure recurrence within six months, incomplete focus resection, and surgical complications were associated with a poor outcome. Multivariate analysis showed that medial temporal lobe epilepsy with hippocampal sclerosis and MRI with visible focal lesions were independent presurgical predictors of a favorable outcome (P < 0.01). Seizure recurrence within six months was the only significant independent predictor associated with a poor outcome (P < 0.01). CONCLUSION: Hippocampal sclerosis and abnormal MRI findings are strongly associated with a favorable surgical outcome, whereas seizure recurrence within six months is associated with a poor outcome.
Subject(s)
Epilepsy/surgery , Adolescent , Adult , Child , Female , Humans , Male , Middle Aged , Retrospective Studies , Treatment Outcome , Young AdultABSTRACT
Mechanisms underlying the mitochondrial protection of Limonium sinense extracts (LSE) was studied in lipopolysaccharide and D-galactosamine (LPS/D-GalN) intoxicated mice. It was found that increased activities of serum aspartate aminotransferase and alanine aminotransferase induced by LPS/D-GalN were significantly inhibited by pretreatment with LSE. The obvious disruption of membrane potential, intramitochondrial Ca (2+) overload and suppression in mitochondrial Ca (2+) -ATPase activity induced by LPS/D-GalN were significantly blocked by pretreatment with LSE. It was concluded that mechanisms underlying protection of LSE against liver mitochondria damage might be related to the preservation on mitochondrial Ca (2+) homeostasis through the preservation on mitochondrial Ca (2+) -ATPase activity.
ABSTRACT
The effects of industrial storage on the changes of the cell viability and the activities of intracellular alcohol dehydrogenase (ADH) and glucose-6-phosphate dehydrogenase (G6PDH) in brewer's yeast, and the corresponding capacity for the bioconversion of ethyl-3-oxobutanoate (EOB) to ethyl (S)-3-hydroxybutanoate ((S)-EHB), were investigated. The viability of fresh brewer's yeast cells stored in industrial circulating cooling water at 1-2 degrees C showed 4 and 15% drop after the storage of 7 and 15 days, respectively, after which cells died rapidly. The pretreatment of the stored brewer's yeast cells by washing and screening significantly enhanced cell viability during industrial storage. The intracellular levels of ADH and G6PDH after permeabilization of these stored cells with cetyltrimetylammonium bromide (CTAB) were much higher, which showed only slight decrease within 2 weeks during the industrial storage. When the stored cells after the permeabilization treatment was used as the biocatalyst at 90-120 g/L, EOB was converted almost completely into enantiopure (S)-EHB with an enantiomeric excess (ee) more than 99% and a yield of over 96%, by fed-batch bioconversion of 560 mM EOB within 6 h.