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1.
Stud Health Technol Inform ; 315: 577-578, 2024 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-39049330

RESUMO

Taiwan has a well-structured healthcare insurance system that offers accessible medical resources to the public through nominal health insurance fees. Consequently, individuals in need of care willingly pay nominal charges for medical services, including rehabilitation treatment. This study delves into the rehabilitation department of a medical center in southern Taiwan. Despite offering comprehensive traditional rehabilitation services covering neurological, musculoskeletal, pediatric, cardiopulmonary, communication, and swallow disorders, the demand for appointments significantly surpasses the number of available therapists. Therefore, this paper proposes an efficiently method to optimize patient-therapist appointment. With a Complex Conditional Logic that we have designed in this paper, we aim to simplify the scheduling processing for patient seeking appointment either online or via phone calls. More than 50,000 cases have been treated since the system's launch within a year, facilitates hospital resource allocation and enhancing patient medical experiences.


Assuntos
Agendamento de Consultas , Taiwan , Humanos , Eficiência Organizacional , Centros de Reabilitação
2.
Stud Health Technol Inform ; 315: 663-664, 2024 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-39049371

RESUMO

In the demanding realm of the Emergency Department, addressing every emergency incident efficiently becomes paramount, with time emerging as a critical factor. This paper is dedicated to investigating innovative approaches aimed at diminishing the preparation time required when a patient arrives at the hospital. Our methodology involves the integration of pre-hospital warning data, a valuable resource provided by the National Science and Technology Center for Disaster Reduction (NCDR), with an advanced flashing light alert system. This integration specifically targets a reduction in preparation time for major traumas, Cerebrovascular Accidents (CVAs), Out-Of-Hospital Cardiac Arrests (OHCAs), and Acute Myocardial Infarctions (AMIs) at the South Medical Center in Taiwan. By streamlining these critical processes, our research endeavors to enhance overall efficiency and responsiveness within emergency medical care, achieving an average preparation time of 6.61 minutes.


Assuntos
Serviços Médicos de Emergência , Taiwan , Humanos , Integração de Sistemas , Sistemas de Comunicação entre Serviços de Emergência
4.
Biomed Pharmacother ; 176: 116864, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38865847

RESUMO

BACKGROUND: DNA repair allows the survival of cancer cells. Therefore, the development of DNA repair inhibitors is a critical need for sensitizing cancers to chemoradiation. Sae2CtIP has specific functions in initiating DNA end resection, as well as coordinating cell cycle checkpoints, and it also greatly interacts with the DDR at different levels. RESULTS: In this study, we demonstrated that corylin, a potential sensitizer, causes deficiencies in DNA repair and DNA damage checkpoints in yeast cells. More specifically, corylin increases DNA damage sensitivity through the Sae2-dependent pathway and impairs the activation of Mec1-Ddc2, Rad53-p and γ-H2A. In breast cancer cells, corylin increases apoptosis and reduces proliferation following Dox treatment by inhibiting CtIP. Xenograft assays showed that treatment with corylin combined with Dox significantly reduced tumor growth in vivo. CONCLUSIONS: Our findings herein delineate the mechanisms of action of corylin in regulating DNA repair and indicate that corylin has potential long-term clinical utility as a DDR inhibitor.


Assuntos
Dano ao DNA , Reparo do DNA , Recombinação Homóloga , Humanos , Animais , Reparo do DNA/efeitos dos fármacos , Recombinação Homóloga/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Feminino , Camundongos Nus , Linhagem Celular Tumoral , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/efeitos dos fármacos , Doxorrubicina/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética
5.
Sci Rep ; 14(1): 12451, 2024 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-38816469

RESUMO

The FZP gene plays a critical role in the formation of lateral branches and spikelets in rice panicle architecture. This study investigates the qSBN7 allele, a hypomorphic variant of FZP, and its influence on panicle architectures in different genetic backgrounds. We evaluated two backcross inbred lines (BILs), BC5_TCS10sbn and BC3_TCS10sbn, each possessing the homozygous qSBN7 allele but demonstrating differing degrees of spikelet degeneration. Our analysis revealed that BC5_TCS10sbn had markedly low FZP expression, which corresponded with an increase in axillary branches and severe spikelet degeneration. Conversely, BC3_TCS10sbn exhibited significantly elevated FZP expression, leading to fewer secondary and tertiary branches, and consequently decreased spikelet degeneration. Compared to BC5_TCS10sbn, BC3_TCS10sbn carries three additional chromosomal substitution segments from its donor parent, IR65598-112-2. All three segments significantly enhance the expression of FZP and reduce the occurrence of tertiary branch and spikelet degeneration. These findings enhance our understanding of the mechanisms regulating FZP and aid rice breeding efforts.


Assuntos
Oryza , Oryza/genética , Oryza/crescimento & desenvolvimento , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Regulação da Expressão Gênica de Plantas , Alelos , Patrimônio Genético , Melhoramento Vegetal , Genes de Plantas , Genoma de Planta , Fenótipo
6.
Nat Commun ; 13(1): 1238, 2022 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-35264584

RESUMO

In the long history of traditional Chinese medicine, single herbs and complex formulas have been suggested to increase lifespan. However, the identification of single molecules responsible for lifespan extension has been challenging. Here, we collected a list of traditional Chinese medicines with potential longevity properties from pharmacopeias. By utilizing the mother enrichment program, we systematically screened these traditional Chinese medicines and identified a single herb, Psoralea corylifolia, that increases lifespan in Saccharomyces cerevisiae. Next, twenty-two pure compounds were isolated from Psoralea corylifolia. One of the compounds, corylin, was found to extend the replicative lifespan in yeast by targeting the Gtr1 protein. In human umbilical vein endothelial cells, RNA sequencing data showed that corylin ameliorates cellular senescence. We also examined an in vivo mammalian model, and found that corylin extends lifespan in mice fed a high-fat diet. Taken together, these findings suggest that corylin may promote longevity.


Assuntos
Células Endoteliais , Longevidade , Animais , Flavonoides/farmacologia , Mamíferos , Medicina Tradicional Chinesa , Camundongos
7.
Int J Mol Sci ; 22(18)2021 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-34575923

RESUMO

Molecules involved in DNA damage response (DDR) are often overexpressed in cancer cells, resulting in poor responses to chemotherapy and radiotherapy. Although treatment efficacy can be improved with the concomitant use of DNA repair inhibitors, the accompanying side effects can compromise the quality of life of patients. Therefore, in this study, we identified a natural compound that could inhibit DDR, using the single-strand annealing yeast-cell analysis system, and explored its mechanisms of action and potential as a chemotherapy adjuvant in hepatocellular carcinoma (HCC) cell lines using comet assay, flow cytometry, Western blotting, immunofluorescence staining, and functional analyses. We developed a mouse model to verify the in vitro findings. We found that hydroxygenkwanin (HGK) inhibited the expression of RAD51 and progression of homologous recombination, thereby suppressing the ability of the HCC cell lines to repair DNA damage and enhancing their sensitivity to doxorubicin. HGK inhibited the phosphorylation of DNA damage checkpoint proteins, leading to apoptosis in the HCC cell lines. In the mouse xenograft model, HGK enhanced the sensitivity of liver cancer cells to doxorubicin without any physiological toxicity. Thus, HGK can inhibit DDR in liver cancer cells and mouse models, making it suitable for use as a chemotherapy adjuvant.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Dano ao DNA/efeitos dos fármacos , Flavonoides/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Reparo do DNA/efeitos dos fármacos , Modelos Animais de Doenças , Sinergismo Farmacológico , Medicamentos de Ervas Chinesas , Regulação da Expressão Gênica , Recombinação Homóloga/efeitos dos fármacos , Humanos , Camundongos , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Leveduras/efeitos dos fármacos , Leveduras/genética , Leveduras/metabolismo
8.
Int J Mol Sci ; 22(9)2021 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-33922657

RESUMO

BRCA mutation, one of the most common types of mutations in breast and ovarian cancer, has been suggested to be synthetically lethal with depletion of RAD52. Pharmacologically inhibiting RAD52 specifically eradicates BRCA-deficient cancer cells. In this study, we demonstrated that curcumin, a plant polyphenol, sensitizes BRCA2-deficient cells to CPT-11 by impairing RAD52 recombinase in MCF7 cells. More specifically, in MCF7-siBRCA2 cells, curcumin reduced homologous recombination, resulting in tumor growth suppression. Furthermore, a BRCA2-deficient cell line, Capan1, became resistant to CPT-11 when BRCA2 was reintroduced. In vivo, xenograft model studies showed that curcumin combined with CPT-11 reduced the growth of BRCA2-knockout MCF7 tumors but not MCF7 tumors. In conclusion, our data indicate that curcumin, which has RAD52 inhibitor activity, is a promising candidate for sensitizing BRCA2-deficient cells to DNA damage-based cancer therapies.


Assuntos
Proteína BRCA2/deficiência , Neoplasias da Mama/tratamento farmacológico , Curcumina/farmacologia , Dano ao DNA , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Recombinação Homóloga , Proteína Rad52 de Recombinação e Reparo de DNA/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Apoptose , Proteína BRCA2/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células , Reparo do DNA , Feminino , Humanos , Irinotecano/farmacologia , Camundongos , Camundongos Nus , Mutação , Inibidores da Topoisomerase I/farmacologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Front Oncol ; 10: 1319, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32850418

RESUMO

Magnolia officinalis is widely used in Southeast Asian countries for the treatment of fever, headache, diarrhea, and stroke. Magnolol is a phenolic compound extracted from M. officinalis, with proven antibacterial, antioxidant, anti-inflammatory, and anticancer activities. In this study, we modified magnolol to synthesize a methoxylated derivative, 2-O-methylmagnolol (MM1), and investigated the use of MM1, and magnolol in the treatment of liver cancer. We found that both magnolol and MM1 exhibited inhibitory effects on the growth, migration, and invasion of hepatocellular carcinoma (HCC) cell lines and halted the cell cycle at the G1 phase. MM1 also demonstrated a substantially better tumor-suppressive effect than magnolol. Further analysis suggested that by inhibiting class I histone deacetylase expression in HCC cell lines, magnolol and MM1 induced p21 expression and p53 activation, thereby causing cell cycle arrest and inhibiting HCC cell growth, migration, and invasion. Subsequently, we verified the significant tumor-suppressive effects of magnolol and MM1 in an animal model. Collectively, these findings demonstrate the anti-HCC activities of magnolol and MM1 and their potential for clinical use.

10.
Artigo em Inglês | MEDLINE | ID: mdl-27420085

RESUMO

Since 1989, blue-collar foreign workers have been permitted to work in Taiwanese industries. Most blue-collar foreign workers apply for jobs in Taiwan through blue-collar foreign workers' agencies. Because blue-collar foreign workers are not familiar with the language and culture in Taiwan, in occupational accident education and hazard prevention, the agencies play an important role in the coordination and translation between employees and blue-collar foreign workers. The purpose of this study is to establish the agencies' role in the occupational accidents education and hazard prevention for blue-collar foreign workers in Taiwan. This study uses a qualitative method-grounded theory-to collect, code, and analyze the data in order to understand the agencies' role in occupational accident education and hazard prevention for blue-collar foreign workers in Taiwan. The results show that the duty of agencies in occupational accident education and hazard prevention includes selecting appropriate blue-collar foreign workers, communicating between employees and blue-collar foreign workers, collecting occupational safety and health information, assisting in the training of occupational safety and health, and helping blue-collar foreign workers adapt to their lives in Taiwan. Finally, this study suggests seven important points and discusses the implementation process necessary to improve governmental policies. The government and employees should pay attention to the education/training of occupational safety and health for blue-collar foreign workers to eliminate unsafe behavior in order to protect the lives of blue-collar foreign workers.


Assuntos
Acidentes de Trabalho/prevenção & controle , Emigrantes e Imigrantes , Indústrias , Ocupações , Adulto , Comunicação , Educação em Saúde , Humanos , Internacionalidade , Masculino , Pessoa de Meia-Idade , Saúde Ocupacional , Políticas , Taiwan/epidemiologia
11.
Arch Virol ; 161(1): 19-31, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26446888

RESUMO

Influenza A viruses (IAV) are widespread in birds and domestic poultry, occasionally causing severe epidemics in humans and posing health threats. Hence, the need to develop a strategy for prophylaxis or therapy, such as a broadly neutralizing antibody against IAV, is urgent. In this study, single-chain variable fragment (scFv) phage display technology was used to select scFv fragments recognizing influenza envelope proteins. The Tomlinson I and J scFv phage display libraries were screened against the recombinant HA2 protein (rHA2) for three rounds. Only the third-round elution sample of the Tomlinson J library showed high binding affinity to rHA2, from which three clones (3JA18, 3JA62, and 3JA78) were chosen for preparative-scale production as soluble antibody by E. coli. The clone 3JA18 was selected for further tests due to its broad affinity for influenza H1N1, H3N2 and H5N1. Simulations of the scFv 3JA18-HA trimer complex revealed that the complementarity-determining region of the variable heavy chain (VH-CDR2) bound the stem region of HA. Neutralization assays using a peptide derived from VH-CDR2 also supported the simulation model. Both the selected antibody and its derived peptide were shown to suppress infection with H5N1 and H1N1 viruses, but not H3N2 viruses. The results also suggested that the scFvs selected from rHA2 could have neutralizing activity by interfering with the function of the HA stem region during virus entry into target cells.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Fragmentos Fc das Imunoglobulinas/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A Subtipo H3N2/imunologia , Virus da Influenza A Subtipo H5N1/imunologia , Influenza Humana/virologia , Anticorpos de Cadeia Única/imunologia , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/imunologia , Especificidade de Anticorpos , Humanos , Fragmentos Fc das Imunoglobulinas/genética , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Virus da Influenza A Subtipo H5N1/genética , Influenza Humana/imunologia , Anticorpos de Cadeia Única/genética
12.
Int J Occup Saf Ergon ; 20(4): 537-49, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25513790

RESUMO

This study examines the roles and functions of businesses, labor-exporting countries' representative offices in Taiwan, religious organizations, and manpower agencies in promoting occupational safety and health (OSH). It also offers advice to Taiwanese authorities on making policies and improvements regarding the oversight mechanism mandated by the Labor Safety and Health Act, giving them an idea of what to focus on when enforcing control over blue-collar foreign workers' OSH conditions. This study also proposes that Taiwanese authorities may serve not only as an overseer/inspector of those hiring blue-collar foreign workers in Taiwan, but also expand their role to lay down policies regarding a variety of OSH teaching materials in the blue-collar foreign workers' native languages (spoken or written), the qualifications of translators in blue-collar foreign workers' OSH training programs, and regulations concerning the longer hours such training programs take.


Assuntos
Emigrantes e Imigrantes , Promoção da Saúde/organização & administração , Saúde Ocupacional , Gestão da Segurança/organização & administração , Acidentes de Trabalho/prevenção & controle , Conscientização , Humanos , Capacitação em Serviço/organização & administração , Relações Interinstitucionais , Gestão de Recursos Humanos , Taiwan
13.
J Phys Chem B ; 118(45): 12827-36, 2014 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-25333416

RESUMO

Entry of HIV-1 into the target cell is mediated by the envelope glycoprotein consisting of noncovalently associated surface subunit gp120 and transmembrane subunit gp41. To form a functional gp41 complex, the protein undergoes hairpin formation and self-assembly. The fusion event can be inhibited by gp41-derived peptides at nanomolar concentration and is highly dependent on the time of addition, implying a role of folding kinetics on the inhibitory action. Oligomerization of the gp41 ectodomain was demonstrated by light scattering measurements. Kinetic study by stopped-flow fluorescence and absorption measurements (i) revealed a multistate folding pathway and stable intermediates; (ii) showed a dissection of fast and slow components for early and late stages of folding, respectively, with 3 orders of magnitude difference in the time scale; (iii) showed the slow process was attributed to misfolding and unzipping of the hairpin; and (iv) showed retardation of the native hairpin formation is assumed to lead to coupling of the correctly registered hairpin and self-assembly. This coupling allows the deduction on the time scale of intrachain folding (0.1-1 s) for the protein. The folding reaction was illustrated by a free energy profile to explain the temporal dichotomy of fast and slow steps of folding as well as effective inhibition by gp41-derived peptide.


Assuntos
Proteína gp120 do Envelope de HIV/química , Proteína gp41 do Envelope de HIV/química , HIV-1/metabolismo , Substituição de Aminoácidos , Espectroscopia de Ressonância de Spin Eletrônica , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/metabolismo , Proteína gp41 do Envelope de HIV/genética , Proteína gp41 do Envelope de HIV/metabolismo , Cinética , Luz , Redobramento de Proteína , Estrutura Quaternária de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Rodaminas/química , Espalhamento de Radiação
14.
J Chromatogr A ; 1330: 61-8, 2014 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-24480734

RESUMO

Synthetic musk is widely used in various scented consumer products. However, the exposure via inhalation is often ignored due to pleasant smells. In addition, the information regarding the distribution of synthetic musk in air is limited. Hence, this research is aimed to develop a highly sensitive and widely applicable method for the determination of airborne synthetic musk. In this study, polyurethane foam (PUF) and filter were employed for active air sampling. Microwave assisted extraction (MAE) and nitrogen evaporator were performed for sample preparation. A gas chromatography coupled with triple quadrupole tandem mass spectrometer (GC/MS-MS) with specific multiple reaction monitoring (MRM) transition pairs was applied for sample analysis. Compared with using selected ion monitoring (SIM) mode traditionally, the sensitivities were improved in this study about an order at least. In terms of air concentration, as low as 0.48ngm(-3) can be determined when sampling at 3.5Lmin(-1) for 8h. The method established was further applied to the analysis of synthetic musk compounds in air samples collected in a cosmetics plant. The results showed that the airborne concentrations of gaseous polycyclic musk, gaseous nitro-musk, and particle-phase polycyclic musk were 6.4×10(2), 4.0×10(1) and 3.1×10(2)ngm(-3), respectively. Meanwhile, Cashmeran, Celstolide, Galaxolide, and Tonalide were found as the dominant musk compounds in the factory investigated.


Assuntos
Poluentes Ocupacionais do Ar/análise , Cosméticos/análise , Cromatografia Gasosa , Cromatografia Gasosa-Espectrometria de Massas/métodos , Gases , Poliuretanos , Sensibilidade e Especificidade , Espectrometria de Massas em Tandem/métodos
15.
Sci Total Environ ; 461-462: 317-22, 2013 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-23738985

RESUMO

Ten indoor swimming pools in Taipei, Taiwan were included in the study to assess the exposure of people to airborne trichloramine (NCl3) and also to discover the factors that might affect the associated concentrations. An active air sampling method was performed to determine the levels of NCl3, while questionnaires were administered to swimming pool workers, including lifeguards, swimming instructors, and management employees. The results show that the concentrations of trichloramine ranged from 0.017 to 0.15 mg m(-3), which were generally lower than what have been reported from other studies. Symptoms of sore throat and phlegm were more frequent among lifeguards and swimming instructors (exposure group) than management employees (reference group) (odds ratios were 11.28 and 4.22 for sore throat and phlegm, respectively). It seems that the current exposure limit for airborne NCl3, which was recommended by WHO, was not lower enough to protect the health of pool attendants. Regulated level of free available chlorine in Taipei (i.e., 0.3-0.7 ppm) is lower than what is required in other countries (e.g., 1-3 ppm in the UK). This might be the main reason why the concentrations of NCl3 reported elsewhere were higher than what were found in this research. Further international comparisons will help to elucidate if low free chlorine concentration should be adopted as an operating standard. For the indoor swimming pools in Taipei, the air quality is suggested to be improved, since even with the low concentrations of NCl3, higher respiratory ailments among pool workers were observed.


Assuntos
Poluição do Ar em Ambientes Fechados/análise , Cloretos/análise , Cloretos/toxicidade , Compostos de Nitrogênio/análise , Compostos de Nitrogênio/toxicidade , Exposição Ocupacional/análise , Doenças Respiratórias/induzido quimicamente , Piscinas , Poluição do Ar em Ambientes Fechados/efeitos adversos , Humanos , Exposição Ocupacional/efeitos adversos , Inquéritos e Questionários , Taiwan
16.
Anal Bioanal Chem ; 405(6): 1969-76, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23263519

RESUMO

This study describes an on-line stacking CE approach by sweeping with whole capillary sample filling for analyzing five anabolic androgenic steroids in urine samples. The five anabolic steroids for detection were androstenedione, testosterone, epitestosterone, boldenone, and clostebol. Anabolic androgenic steroids are abused in sport doping because they can promote muscle growth. Therefore, a sensitive detection method is imperatively required for monitoring the urine samples of athletes. In this research, an interesting and reliable stacking capillary electrophoresis method was established for analysis of anabolic steroids in urine. After liquid-liquid extraction by n-hexane, the supernatant was dried and reconstituted with 30 mM phosphate buffer (pH 5.00) and loaded into the capillary by hydrodynamic injection (10 psi, 99.9 s). The stacking and separation were simultaneously accomplished at -20 kV in phosphate buffer (30 mM, pH 5.0) containing 100 mM sodium dodecyl sulfate and 40 % methanol. During the method validation, calibration curves were linear (r≥0.990) over a range of 50-1,000 ng/mL for the five analytes. In the evaluation of precision and accuracy for this method, the absolute values of the RSD and the RE in the intra-day (n=3) and inter-day (n=5) analyses were all less than 6.6 %. The limit of detection for the five analytes was 30 ng/mL (S/N=5, sampling 99.9 s at 10 psi). Compared with simple MECK, this stacking method possessed a 108- to 175-fold increase in sensitivity. This simple and sensitive stacking method could be used as a powerful tool for monitoring the illegal use of doping.


Assuntos
Androstenodiona/urina , Eletroforese Capilar/métodos , Epitestosterona/urina , Detecção do Abuso de Substâncias/métodos , Testosterona/análogos & derivados , Testosterona/urina , Atletas , Calibragem , Dopagem Esportivo/prevenção & controle , Análise de Injeção de Fluxo , Hexanos/química , Humanos , Extração Líquido-Líquido , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
17.
J Struct Biol ; 175(3): 406-14, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21722736

RESUMO

The envelope glycoprotein gp41 of HIV-1 undergoes structural rearrangement to form a helix hairpin during the virus-mediated fusion. Previous studies to investigate the folding and stability of hairpin did not monitor the end-to-end distance of the molecule. To directly probe the distance change, rhodamine dye was conjugated to the gp41 recombinant near the N- and C-terminal regions to detect the UV absorption and fluorescence intensity changes induced by the chemical denaturant guanidinium chloride (GdmCl). Using the singly- and doubly-labeled constructs allowed us to distinguish between the hairpin formation and protein oligomerization. A biphasic transition of helical structure for the wild type protein was revealed by circular dichroism measurements while unfolding of the hairpin occurred at 6M GdmCl. The relevance of our study to the fusion inhibitor for HIV-1 was borne out by results on the mutants at the positions within the N-terminal heptad repeat (NHR) and the C-terminal heptad repeat (CHR) regions. A monophasic transition at lower denaturant concentration was detected for the NHR mutant supporting the concept of differential stability of NHR and CHR helical structure. The conclusion that the observed unstacking of doubly-labeled variant arises principally from the intra-molecular dimers was drawn from the unstacking of the protein labeled in the loop. Remarkably, it is deduced that the hairpin is more stable than the CHR helical structure. A model for denaturation of the helix hairpin bundle was proposed from these results. The biological implications of the findings and further applications of the distance-based approach were discussed.


Assuntos
Dicroísmo Circular/métodos , Proteína gp41 do Envelope de HIV/química , Proteína gp41 do Envelope de HIV/metabolismo , Modelos Moleculares , Simulação de Dinâmica Molecular , Multimerização Proteica , Estrutura Secundária de Proteína
18.
Anal Chim Acta ; 697(1-2): 75-82, 2011 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-21641421

RESUMO

This article reports the integration of the fiber optic-particle plasmon resonance (FO-PPR) biosensor with a microfluidic chip to reduce response time and improve detection limit. The microfluidic chip made of poly(methyl methacrylate) had a flow-channel of dimensions 4.0 cm × 900 µm × 900 µm. A partially unclad optical fiber with gold or silver nanoparticles on the core surface was placed within the flow-channel, where the volume of the flow space was about 14 µL. Results using sucrose solutions of various refractive indexes show that the refractive index resolution improves by 2.4-fold in the microfluidic system. The microfluidic chip is capable of delivering a precise amount of biological samples to the detection area without sample dilution. Several receptor/analyte pairs were chosen to examine the biosensing capability of the integrated platform: biotin/streptavidin, biotin/anti-biotin, DNP/anti-DNP, OVA/anti-OVA, and anti-MMP-3/MMP-3. Results show that the response time to achieve equilibrium can be shortened from several thousand seconds in a conventional liquid cell to several hundred seconds in a microfluidic flow-cell. In addition, the detection limit also improves by about one order of magnitude. Furthermore, the normalization by using the relative change of transmission response as the sensor output alleviate the demand on precise optical alignment, resulting in reasonably good chip-to-chip measurement reproducibility.


Assuntos
Técnicas Biossensoriais/instrumentação , Técnicas Analíticas Microfluídicas/instrumentação , Fibras Ópticas , Integração de Sistemas , Estudos de Viabilidade , Ouro/química , Humanos , Indicadores e Reagentes/química , Limite de Detecção , Metaloproteinase 3 da Matriz/análise , Nanopartículas Metálicas/química , Líquido Sinovial/enzimologia , Fatores de Tempo
19.
J Biotechnol ; 153(1-2): 48-55, 2011 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-21419812

RESUMO

We demonstrated a high level expression and purification of recombinant human immunodeficiency virus type 1 gp41 ectodomain (gp41e-FP) using glass bead approach with a final yield of 12±2mg/L bacterial culture. The proper folding of gp41e-FP encompassing the fusion peptide (FP) was ascertained by circular dichroism (CD) measurement and recognition by NC-1 antibody. The latter assay revealed stabilization of the gp41 coiled coil structure in the presence of liposome dispersion. The differential affinity of gp41e-FP and gp41e (devoid of FP) by NC-1 suggested an aggregated state for gp41e-FP and/or possible proximity of the fusion peptide domain to the coiled coil structure of gp41 ectodomain. Perfluorooctanoate (PFO)-PAGE electrophoresis experiment revealed the trimeric propensity of the recombinant gp41e-FP. In comparison to gp41e, the lipid mixing activity of gp41e-FP was two-fold higher suggesting a role of FP in promoting membrane fusion. The present approach to efficiently and quantitatively preparing the functional full-length recombinant gp41 ectodomain protein can be employed for structural and biomedical investigations and the extraction of other inclusion body-embedded recombinant proteins.


Assuntos
Biotecnologia/métodos , Escherichia coli/metabolismo , Proteína gp41 do Envelope de HIV/química , Proteína gp41 do Envelope de HIV/metabolismo , HIV-1/metabolismo , Peptídeos/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Humanos , Lipídeos de Membrana/metabolismo , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/isolamento & purificação , Ressonância de Plasmônio de Superfície
20.
Ann Occup Hyg ; 55(5): 519-25, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21430133

RESUMO

This study assessed the relationships between ethylbenzene exposure and levels of 8-hydroxydeoxyguanosine (8-OHdG) among spray painters. Sixty-four male workers employed at a large shipyard were recruited for this investigation. Fifteen spray painters exposed to paint, together with two non-exposed groups, namely 19 sandblasting workers and 30 office staffs were selected as the subjects. Personal exposure to xylene and ethylbenzene in air were collected using diffusive samplers. Urine samples of the spray painters were collected after a month-long holiday leave and during the pre- and post-workshifts. Urine samples of sandblasting workers and office staffs were gathered after their shift. Urinary mandelic acid and methyl hippuric acid were used as biological indices of dose of ethylbenzene and xylene, respectively. Urinary 8-OHdG was used as biomarker of oxidative DNA damage. The post-workshift concentration of urinary 8-OHdG for 10 spray painters (30.3 ± 9.28 µg g(-1) creatinine) significantly exceeded that of holiday leave (7.20 ± 1.08 µg g(-1) creatinine; P = 0.001). The post-workshift concentration of urinary 8-OHdG was higher among 15 spray painters (29.0 ± 6.52 µg g(-1) creatinine) than sandblasting workers (9.14 ± 2.05 µg g(-1) creatinine; P = 0.01) and office staffs (8.35 ± 0.84 µg g(-1) creatinine; P = 0.007). A stepwise regression model revealed an 8.11 µg g(-1) creatinine increase per 1 p.p.m. increase in ethylbenzene [95% confidence interval (CI) 4.13-12.1]. A stepwise regression model revealed an increase of 6.04 µg g(-1) creatinine (95% CI 2.23-9.84) per 1 p.p.m. in ethylbenzene after adjustment of age (95% CI 2.23-9.84). This pilot study suggests that occupational exposure to paint increases oxidative DNA injury. Moreover, urinary 8-OHdG levels displayed greater DNA damage in spray painters compared to other unexposed groups and their holiday leave samples. A significant correlation was found between urinary 8-OHdG and the exposure to ethylbenzene. The ethylbenzene exposure could not explain all urinary 8-OHdG measured. Other components of paint deserve further investigation.


Assuntos
Derivados de Benzeno/toxicidade , Dano ao DNA , Desoxiguanosina/análogos & derivados , Exposição Ocupacional/análise , Estresse Oxidativo , 8-Hidroxi-2'-Desoxiguanosina , Adulto , Poluentes Ocupacionais do Ar/análise , Derivados de Benzeno/análise , Biomarcadores/urina , Creatinina/urina , Desoxiguanosina/urina , Hipuratos/urina , Humanos , Masculino , Ácidos Mandélicos/urina , Pessoa de Meia-Idade , Projetos Piloto , Análise de Regressão , Xilenos/análise
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