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1.
Environ Sci Technol ; 53(16): 9636-9645, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31347357

RESUMO

California methane (CH4) emissions are quantified for three years from two tower networks and one aircraft campaign. We used backward trajectory simulations and a mesoscale Bayesian inverse model, initialized by three inventories, to achieve the emission quantification. Results show total statewide CH4 emissions of 2.05 ± 0.26 (at 95% confidence) Tg/yr, which is 1.14 to 1.47 times greater than the anthropogenic emission estimates by California Air Resource Board (CARB). Some of differences could be biogenic emissions, superemitter point sources, and other episodic emissions which may not be completely included in the CARB inventory. San Joaquin Valley (SJV) has the largest CH4 emissions (0.94 ± 0.18 Tg/yr), followed by the South Coast Air Basin, the Sacramento Valley, and the San Francisco Bay Area at 0.39 ± 0.18, 0.21 ± 0.04, and 0.16 ± 0.05 Tg/yr, respectively. The dairy and oil/gas production sources in the SJV contribute 0.44 ± 0.36 and 0.22 ± 0.23 Tg CH4/yr, respectively. This study has important policy implications for regulatory programs, as it provides a thorough multiyear evaluation of the emissions inventory using independent atmospheric measurements and investigates the utility of a complementary multiplatform approach in understanding the spatial and temporal patterns of CH4 emissions in the state and identifies opportunities for the expansion and applications of the monitoring network.


Assuntos
Poluentes Atmosféricos , Metano , Aeronaves , Teorema de Bayes , California , São Francisco
2.
J Ginseng Res ; 42(2): 165-174, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29719463

RESUMO

Background: Extended endoplasmic reticulum (ER) stress may initiate apoptotic pathways in cancer cells, and ER stress has been reported to possibly increase tumor death in cancer therapy. We previously reported that caspase-8 played an important role in compound K-induced apoptosis via activation of caspase-3 directly or indirectly through Bid cleavage, cytochrome c release, and caspase-9 activation in HL-60 human leukemia cells. The mechanisms leading to apoptosis in A549 and SK-MES-1 human lung cancer cells and the role of ER stress have not yet been understood. Methods: The apoptotic effects of compound K were analyzed using flow cytometry, and the changes in protein levels were determined using Western blot analysis. The intracellular calcium levels were monitored by staining with Fura-2/AM and Fluo-3/AM. Results: Compound K-induced ER stress was confirmed through increased phosphorylation of eIF2α and protein levels of GRP78/BiP, XBP-1S, and IRE1α in human lung cancer cells. Moreover, compound-K led to the accumulation of intracellular calcium and an increase in m-calpain activities that were both significantly inhibited by pretreatment either with BAPTA-AM (an intracellular Ca2+ chelator) or dantrolene (an RyR channel antagonist). These results were correlated with the outcome that compound K induced ER stress-related apoptosis through caspase-12, as z-ATAD-fmk (a specific inhibitor of caspase-12) partially ameliorated this effect. Interestingly, 4-PBA (ER stress inhibitor) dramatically improved the compound K-induced apoptosis. Conclusion: Cell survival and intracellular Ca2+ homeostasis during ER stress in human lung cancer cells are important factors in the induction of the compound K-induced apoptotic pathway.

3.
Lab Anim Res ; 34(4): 166-175, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30671102

RESUMO

Recombination activating gene-2 (RAG-2) plays a crucial role in the development of lymphocytes by mediating recombination of T cell receptors and immunoglobulins, and loss of RAG-2 causes severe combined immunodeficiency (SCID) in humans. RAG-2 knockout mice created using homologous recombination in ES cells have served as a valuable immunodeficient platform, but concerns have persisted on the specificity of RAG-2-related phenotypes in these animals due to the limitations associated with the genome engineering method used. To precisely investigate the function of RAG-2, we recently established a new RAG-2 knockout FVB mouse line (RAG-2 -/-) manifesting lymphopenia by employing a CRISPR/Cas9 system at Center for Mouse Models of Human Disease. In this study, we further characterized their phenotypes focusing on histopathological analysis of lymphoid organs. RAG-2 -/- mice showed no abnormality in development compared to their WT littermates for 26 weeks. At necropsy, gross examination revealed significantly smaller spleens and thymuses in RAG-2 -/- mice, while histopathological investigation revealed hypoplastic white pulps with intact red pulps in the spleen, severe atrophy of the thymic cortex and disappearance of follicles in lymph nodes. However, no perceivable change was observed in the bone marrow. Moreover, our analyses showed a specific reduction of lymphocytes with a complete loss of mature T cells and B cells in the lymphoid organs, while natural killer cells and splenic megakaryocytes were increased in RAG-2 -/- mice. These findings indicate that our RAG-2 -/- mice show systemic lymphopenia with the relevant histopathological changes in the lymphoid organs, suggesting them as an improved Rag-2-related immunodeficient model.

4.
Lab Anim Res ; 34(4): 279-287, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30671116

RESUMO

Placenta specific 8 (PLAC8, also known as ONZIN) is a multi-functional protein that is highly expressed in the intestine, lung, spleen, and innate immune cells, and is involved in various diseases, including cancers, obesity, and innate immune deficiency. Here, we generated a Plac8 knockout mouse using the CRISPR/Cas9 system. The Cas9 mRNA and two single guide RNAs targeting a region near the translation start codon at Plac8 exon 2 were microinjected into mouse zygotes. This successfully eliminated the conventional translation start site, as confirmed by Sanger sequencing and PCR genotyping analysis. Unlike the previous Plac8 deficient models displaying increased adipose tissue and body weights, our male Plac8 knockout mice showed rather lower body weight than sex-matched littermate controls, though the only difference between these two mouse models is genetic context. Differently from the previously constructed embryonic stem cell-derived Plac8 knockout mouse that contains a neomycin resistance cassette, this knockout mouse model is free from a negative selection marker or other external insertions, which will be useful in future studies aimed at elucidating the multi-functional and physiological roles of PLAC8 in various diseases, without interference from exogenous foreign DNA.

5.
Lab Anim Res ; 34(4): 302-310, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30671119

RESUMO

CD47 (integrin-associated protein), a multi-spanning transmembrane protein expressed in all cells including red blood cells (RBCs) and leukocytes, interacts with signal regulatory protein α (SIRPα) on macrophages and thereby inhibits phagocytosis of RBCs. Recently, we generated a novel C57BL/6J CD47 knockout (CD47 -/- hereafter) mouse line by employing a CRISPR/Cas9 system at Center for Mouse Models of Human Disease, and here report their hematological phenotypes. On monitoring their birth and development, CD47 -/- mice were born viable with a natural male-to-female sex ratio and normally developed from birth through puberty to adulthood without noticeable changes in growth, food/water intake compared to their age and sex-matched wild-type littermates up to 26 weeks. Hematological analysis revealed a mild but significant reduction of RBC counts and hemoglobin in 16 week-old male CD47 -/- mice which were aggravated at the age of 26 weeks with increased reticulocyte counts and mean corpuscular volume (MCV), suggesting hemolytic anemia. Interestingly, anemia in female CD47 -/- mice became evident at 26 weeks, but splenomegaly was identified in both genders of CD47 -/- mice from the age of 16 weeks, consistent with development of hemolytic anemia. Additionally, helper and cytotoxic T cell populations were considerably reduced in the spleen, but not in thymus, of CD47 -/- mice, suggesting a crucial role of CD47 in proliferation of T cells. Collectively, these findings indicate that our CD47 -/- mice have progressive hemolytic anemia and splenic depletion of mature T cell populations and therefore may be useful as an in vivo model to study the function of CD47.

6.
Clin Ther ; 39(1): 138-149, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27989618

RESUMO

PURPOSE: Pelubiprofen is a novel nonsteroidal anti-inflammatory, analgesic, and antipyretic drug with at least similar efficacy and better tolerability compared with other nonsteroidal anti-inflammatory, analgesic, and antipyretic drugs such as naproxen and aceclofenac. Eperisone hydrochloride is a centrally acting muscle relaxant that performs by blocking calcium channels. The combined use of pelubiprofen and eperisone hydrochloride is increasingly anticipated to promote the clinical effectiveness of pelubiprofen in relieving musculoskeletal symptoms of osteoarthritis, rheumatoid arthritis, and low back pain. No published data are yet available, however, regarding the pharmacokinetic interactions between these 2 drugs when administered concurrently. The objective of this study was to evaluate any pharmacokinetic interactions between pelubiprofen and eperisone hydrochloride in healthy Korean male volunteers. METHODS: This was a randomized, open-label, crossover study. Each participant was randomly assigned to 1 of 6 treatment sequences and orally received either 45-mg sustained-release pelubiprofen, 75-mg sustained-release eperisone hydrochloride, or both as a single dose in each treatment period, with a 7-day washout period between each treatment. Serial blood samples were collected over 24 hours after dosing, and plasma concentrations of each drug and the major active metabolite of pelubiprofen (trans-alcohol pelubiprofen) were determined by using a validated HPLC-MS/MS system. Pharmacokinetic analyses were conducted by using noncompartmental methods. FINDINGS: A total of 24 men (mean ± standard deviation of: age, 29 ± 4 years; weight, 72.5 ± 7.8 kg; body mass index, 23.4 ± 1.9 kg/m2) were enrolled, and 23 participants completed the study. For pelubiprofen, the geometric mean ratios (90% CIs) of Cmax and AUC0-∞ were 1.02 (0.87-1.19) and 0.97 (0.88-1.07), respectively. For the major active metabolite of pelubiprofen (trans-alcohol pelubiprofen), the geometric mean ratios (90% CIs) of Cmax and AUC0-∞ were 1.05 (0.98-1.13) and 1.04 (1.01-1.07). For eperisone, the geometric mean ratios (90% CIs) of Cmax and AUC0-∞ were 0.87 (0.67-1.15) and 1.05 (0.85-1.30). None of the study participants experienced serious adverse events during the study. IMPLICATIONS: No clinically significant changes were noted in the pharmacokinetic interactions of pelubiprofen, the major active metabolite of pelubiprofen (trans-alcohol pelubiprofen), and eperisone hydrochloride between monotherapy and combination therapy with 45-mg sustained-release pelubiprofen and 75-mg sustained-release eperisone hydrochloride.


Assuntos
Anti-Inflamatórios não Esteroides/administração & dosagem , Bloqueadores dos Canais de Cálcio/administração & dosagem , Fenilpropionatos/administração & dosagem , Propiofenonas/administração & dosagem , Administração Oral , Adulto , Área Sob a Curva , Bloqueadores dos Canais de Cálcio/farmacocinética , Estudos Cross-Over , Humanos , Masculino , Fenilpropionatos/farmacocinética , Propiofenonas/farmacocinética , Espectrometria de Massas em Tandem/métodos , Adulto Jovem
7.
Acta Pharm ; 67(4): 479-494, 2017 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-29337669

RESUMO

In this study, we investigated the gastroprotective effect of an isopropanol extract from the aerial parts of Artemisia princeps (IPAP) and developed a gastroretentive floating tablet of IPAP (IPAP-FR) for maximized local gastroprotective effects. Pre-treatment with IPAP ameliorated the gastric mucosal hemorrhagic lesions in ethanol/HCl- or indomethacin- treated rats. IPAP decreased mucosal hemorrhage of gastric ulcers induced by ethanol or indomethacin plus pyloric ligation in rats. The optimized floating tablet, IPAP-FR, floated on medium surface with more sustained eupatilin release compared to the non-floating control tablet. X-ray photographs in beagle dogs showed that IPAPFR was retained for > 2 h in the stomach. In the ethanol-induced gastric ulcer rat model, the gastric hemorrhagic lesion was improved more substantially with IPAP-FR compared to the non-floating control tablet. Based on these data, our data suggest that IPAP-FR has an improved therapeutic potential for the treatment of gastric ulcer.


Assuntos
Artemisia/química , Mucosa Gástrica/efeitos dos fármacos , Extratos Vegetais/farmacologia , 2-Propanol , Animais , Antiulcerosos/farmacologia , Cães , Etanol/efeitos adversos , Flavonoides/farmacologia , Indometacina/efeitos adversos , Ligadura/efeitos adversos , Masculino , Úlcera Péptica Hemorrágica/induzido quimicamente , Úlcera Péptica Hemorrágica/etiologia , Úlcera Péptica Hemorrágica/prevenção & controle , Extratos Vegetais/administração & dosagem , Ratos , Ratos Sprague-Dawley , Úlcera Gástrica/induzido quimicamente , Úlcera Gástrica/complicações , Úlcera Gástrica/prevenção & controle , Comprimidos
8.
Artigo em Inglês | MEDLINE | ID: mdl-30984251

RESUMO

We report continuous surface observations of carbon dioxide (CO2) and methane (CH4) from the Los Angeles (LA) Megacity Carbon Project during 2015. We devised a calibration strategy, methods for selection of background air masses, calculation of urban enhancements, and a detailed algorithm for estimating uncertainties in urban-scale CO2 and CH4 measurements. These methods are essential for understanding carbon fluxes from the LA megacity and other complex urban environments globally. We estimate background mole fractions entering LA using observations from four "extra-urban" sites including two "marine" sites located south of LA in La Jolla (LJO) and offshore on San Clemente Island (SCI), one "continental" site located in Victorville (VIC), in the high desert northeast of LA, and one "continental/mid-troposphere" site located on Mount Wilson (MWO) in the San Gabriel Mountains. We find that a local marine background can be established to within ~1 ppm CO2 and ~10 ppb CH4 using these local measurement sites. Overall, atmospheric carbon dioxide and methane levels are highly variable across Los Angeles. "Urban" and "suburban" sites show moderate to large CO2 and CH4 enhancements relative to a marine background estimate. The USC (University of Southern California) site near downtown LA exhibits median hourly enhancements of ~20 ppm CO2 and ~150 ppb CH4 during 2015 as well as ~15 ppm CO2 and ~80 ppb CH4 during mid-afternoon hours (12:00-16:00 LT, local time), which is the typical period of focus for flux inversions. The estimated measurement uncertainty is typically better than 0.1 ppm CO2 and 1 ppb CH4 based on the repeated standard gas measurements from the LA sites during the last 2 years, similar to Andrews et al. (2014). The largest component of the measurement uncertainty is due to the single-point calibration method; however, the uncertainty in the background mole fraction is much larger than the measurement uncertainty. The background uncertainty for the marine background estimate is ~10 and ~15 % of the median mid-afternoon enhancement near downtown LA for CO2 and CH4, respectively. Overall, analytical and background uncertainties are small relative to the local CO2 and CH4 enhancements; however, our results suggest that reducing the uncertainty to less than 5 % of the median mid-afternoon enhancement will require detailed assessment of the impact of meteorology on background conditions.

9.
Oncol Rep ; 36(6): 3577-3587, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27748905

RESUMO

Previously, we reported that (E)-8-acetoxy-2-[2-(3,4-diacetoxyphenyl)ethenyl]-quinazoline (8-ADEQ), a synthetic analogue of resveratrol had anti-inflammatory and G2/M cell cycle arrest activities, but the underlying molecular mechanism of cytotoxic effects of this compound was not determined. In this study, 8-ADEQ displayed potent cytotoxicity and triggered apoptosis in HL-60 cells as evidenced by DNA fragmentation, DNA ladder formation, and the externalization of Annexin V-targeted phosphatidylserine residues in HL-60 cells. In addition, 8-ADEQ triggered activation of caspases-8, -9, -6 and -3 and cleavage of their substrates such as poly(ADP-ribose) polymerase (PARP). Moreover, 8-ADEQ induced loss of mitochondrial membrane potential (MMP) and release of cytochrome c to the cytosol. Caspase-3 inhibitor (z-DEVD-fmk), caspase-8 inhibitor (z-IETD-fmk), caspase-9 inhibitor (z-LEHD), and broad caspase inhibitor (z-VAD­fmk) significantly suppressed the 8-ADEQ-induced DNA fragmentation. Interestingly, pretreatment with z-IETD-fmk, a caspase-8 inhibitor, completely abolished 8-ADEQ-induced caspase-3 and -9 activation, and subsequent DNA fragmentation. 8-ADEQ also increased the expression of Fas, Fas-associated death domain (FADD) and FasL, and formation of death-inducing signaling complex (DISC). Further analysis revealed that 8-ADEQ-induced apoptosis was mediated by upregulation of reactive oxidative species (ROS) generation. Taken together, our data indicated that 8-ADEQ-stimulated apoptosis in HL-60 leukemia cells is due to a Fas-mediated caspase-8-dependent pathway via ROS generation, but also, to a lesser extent cytochrome c release and caspase-9 activation.


Assuntos
Apoptose/efeitos dos fármacos , Quinazolinas/farmacologia , Estilbenos/farmacologia , Clorometilcetonas de Aminoácidos , Caspase 3/metabolismo , Caspase 8/metabolismo , Caspase 9/metabolismo , Inibidores de Caspase/farmacologia , Citocromos c/metabolismo , Proteína Ligante Fas , Células HL-60 , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Oligopeptídeos , Poli(ADP-Ribose) Polimerases/metabolismo , Resveratrol
10.
Artigo em Inglês | MEDLINE | ID: mdl-25618252

RESUMO

A suitable liquid chromatography tandem mass spectrometry (LC-MS/MS) method is required to determine pelubiprofen and its active metabolite, trans-alcohol (M-D), in human plasma for pharmacokinetic studies of pelubiprofen preparations. After one-step liquid-liquid extraction (LLE) using methyl tert-butyl ether (MTBE), pelubiprofen, M-D, and tolbutamide (the internal standard, IS) were eluted from a Capcellpak C18 ACR column using a gradient mobile phase consisting of 0.1% formic acid in water and acetonitrile at a flow rate 0.35mL/min. The achieved lower limits of quantitation (LLOQ) of pelubiprofen and M-D were both 15ng/mL (S/N>10) and the standard calibration curves for pelubiprofen and M-D were linear (correlation coefficients >0.99) over the studied concentration range (15-2000ng/mL). Intra- and inter-day precisions were within 7.62% for all analytes and the deviation of assay accuracies was within ±13.23%. The developed method was successfully applied to a pharmacokinetic study of pelubiprofen in healthy Korean male volunteers.


Assuntos
Álcoois/sangue , Álcoois/farmacocinética , Cromatografia Líquida/métodos , Fenilpropionatos/sangue , Fenilpropionatos/farmacocinética , Espectrometria de Massas em Tandem/métodos , Adulto , Álcoois/química , Humanos , Limite de Detecção , Masculino , Fenilpropionatos/química , Padrões de Referência , Reprodutibilidade dos Testes , Tolbutamida/química
11.
Environ Sci Technol ; 48(1): 491-8, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24298975

RESUMO

The sources of halogenated compounds in East Asia associated with stratospheric ozone depletion and climate change are relatively poorly understood. High-precision in situ measurements of 18 halogenated compounds and carbonyl sulfide (COS) made at Gosan, Jeju Island, Korea, from November 2007 to December 2011 were analyzed by a positive matrix factorization (PMF). Seven major industrial sources were identified from the enhanced concentrations of halogenated compounds observed at Gosan and corresponding concentration-based source contributions were also suggested: primary aluminum production explaining 37% of total concentration enhancements, solvent usage of which source apportionment is 25%, fugitive emissions from HCFC/HFC production with 11%, refrigerant replacements (9%), semiconductor/electronics industry (9%), foam blowing agents (6%), and fumigation (3%). Statistical trajectory analysis was applied to specify the potential emission regions for seven sources using back trajectories. Primary aluminum production, solvent usage and fugitive emission sources were mainly contributed by China. Semiconductor/electronics sources were dominantly located in Korea. Refrigerant replacement, fumigation and foam blowing agent sources were spread throughout East Asian countries. The specified potential source regions are consistent with country-based consumptions and emission patterns, verifying the PMF analysis results. The industry-based emission sources of halogenated compounds identified in this study help improve our understanding of the East Asian countries' industrial contributions to halogenated compound emissions.


Assuntos
Poluentes Atmosféricos/análise , Mudança Climática , Monitoramento Ambiental/métodos , Hidrocarbonetos Halogenados/análise , Indústrias , Modelos Teóricos , Extremo Oriente , Análise dos Mínimos Quadrados , Estações do Ano
12.
Proc Natl Acad Sci U S A ; 110(6): 2029-34, 2013 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-23341630

RESUMO

Nitrogen trifluoride (NF(3)) has potential to make a growing contribution to the Earth's radiative budget; however, our understanding of its atmospheric burden and emission rates has been limited. Based on a revision of our previous calibration and using an expanded set of atmospheric measurements together with an atmospheric model and inverse method, we estimate that the global emissions of NF(3) in 2011 were 1.18 ± 0.21 Gg⋅y(-1), or ∼20 Tg CO(2)-eq⋅y(-1) (carbon dioxide equivalent emissions based on a 100-y global warming potential of 16,600 for NF(3)). The 2011 global mean tropospheric dry air mole fraction was 0.86 ± 0.04 parts per trillion, resulting from an average emissions growth rate of 0.09 Gg⋅y(-2) over the prior decade. In terms of CO(2) equivalents, current NF(3) emissions represent between 17% and 36% of the emissions of other long-lived fluorinated compounds from electronics manufacture. We also estimate that the emissions benefit of using NF(3) over hexafluoroethane (C(2)F(6)) in electronics manufacture is significant-emissions of between 53 and 220 Tg CO(2)-eq⋅y(-1) were avoided during 2011. Despite these savings, total NF(3) emissions, currently ∼10% of production, are still significantly larger than expected assuming global implementation of ideal industrial practices. As such, there is a continuing need for improvements in NF(3) emissions reduction strategies to keep pace with its increasing use and to slow its rising contribution to anthropogenic climate forcing.

13.
Environ Sci Technol ; 45(13): 5668-75, 2011 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-21649439

RESUMO

High-frequency in situ measurements at Gosan (Jeju Island, Korea) during November 2007 to December 2008 have been combined with interspecies correlation analysis to estimate national emissions of halogenated compounds (HCs) in East Asia, including the chlorofluorocarbons (CFCs), halons, hydrochlorofluorocarbons (HCFCs), hydrofluorocarbons (HFCs), perfluorocarbons (PFCs), sulfur hexafluoride (SF(6)), and other chlorinated and brominated compounds. Our results suggest that overall China is the dominant emitter of HCs in East Asia, however significant emissions are also found in South Korea, Japan and Taiwan for HFC-134a, HFC-143a, C(2)F(6), SF(6), CH(3)CCl(3), and HFC-365mfc. The combined emissions of CFCs, halon-1211, HCFCs, HFCs, PFCs, and SF(6) from all four countries in 2008 are 25.3, 1.6, 135, 42.6, 3.6, and 2.0 kt/a, respectively. They account for approximately 15%, 26%, 29%, 16%, 32%, and 26.5% of global emissions, respectively. Our results show signs that Japan has successfully phased out CFCs and HCFCs in compliance with the Montreal Protocol (MP), Korea has started transitioning from HCFCs to HFCs, while China still significantly consumes HCFCs. Taiwan, while not directly regulated under the MP, is shown to have adapted the use of HFCs. Combined analysis of emission rates and the interspecies correlation matrix presented in this study proves to be a powerful tool for monitoring and diagnosing changes in consumption of HCs in East Asia.


Assuntos
Poluentes Atmosféricos/análise , Monitoramento Ambiental/estatística & dados numéricos , Hidrocarbonetos Halogenados/análise , Cromatografia Gasosa , Monitoramento Ambiental/métodos , Extremo Oriente , Geografia
14.
Drug Chem Toxicol ; 29(3): 303-12, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16777708

RESUMO

Some fluoroquinolones have been reported to induce QT interval prolongation associated with the onset of torsades de pointes (TdP), resulting in a life-threatening ventricular arrhythmia. We investigated the cardiac electrophysiological effects of two new fluoroquinolones, gemifloxacin and balofloxacin, by using conventional microelectrode recording techniques in isolated rabbit Purkinje fiber and whole-cell patch-clamp techniques in human ether-á-go-go related gene (hERG)-transient transfected CHO cells. Gemifloxacin had no significant effects on the resting membrane potential, total amplitude, action potential, and Vmax of phase 0 depolarization at concentrations up to 30 microM, but gemifloxacin at 100 microM significantly decreased total amplitude (p < 0.01). These values of gemifloxacin (30 and 100 microM) were approximately 25- and 83-fold more than the free plasma concentration of 1.2 microM in a single therapeutic injection in humans. For I(hERG), the IC(50) value was about 300 microM. Balofloxacin had also no significant effects on the resting membrane potential, total amplitude, action potential duration, and Vmax of phase 0 depolarization at concentrations up to 30 microM, but balofloxacin at 100 microM significantly (p < 0.01) prolonged action potentials at both 50% repolarization (APD(50)) and 90% repolarization (APD(90)). These values of balofloxacin (30 and 100 microM) were approximately 6.8- and 23-fold more than the free plasma concentration of 4.4 microM in a single therapeutic injection in humans. For I(hERG), the IC(50) value was 214 +/- 14 microM. Therefore, our data suggested that in the electrophysiological aspect, gemifloxacin and balofloxacin may have no torsadogenic potenties up to 30 microM.


Assuntos
Antibacterianos/toxicidade , Fluoroquinolonas/toxicidade , Naftiridinas/toxicidade , Potenciais de Ação/efeitos dos fármacos , Animais , Antibacterianos/sangue , Antibacterianos/farmacocinética , Disponibilidade Biológica , Canal de Potássio ERG1 , Eletrofisiologia , Canais de Potássio Éter-A-Go-Go/efeitos dos fármacos , Feminino , Fluoroquinolonas/sangue , Fluoroquinolonas/farmacocinética , Gemifloxacina , Coração/efeitos dos fármacos , Técnicas In Vitro , Naftiridinas/sangue , Naftiridinas/farmacocinética , Bloqueadores dos Canais de Potássio , Ramos Subendocárdicos/efeitos dos fármacos , Coelhos
15.
Toxicology ; 216(2-3): 140-6, 2005 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-16182432

RESUMO

UNLABELLED: The protective effect of an antioxidant, Vitamin E (dl-alpha-tocopherol, 100 mg/kg/day, 8 days p.o. in vivo and 10 and 50 microM in vitro) was tested against PCB-induced neurotoxicity. IN VIVO STUDIES: Microdialysis was used to investigate changes in the striatal extracellular dopamine level and in p-nNOS expression in PCB-treated (Aroclor 1254, 10 microg/ml, 2 microl/min, 5 h; 6 microg was infused by microdialysis probe) rats. IN VITRO STUDIES: Cell viability and levels of p-nNOS expression were observed in PCB-treated (Aroclor 1254, 5 microg/ml) immortalized dopaminergic cell line (CATH.a cells). RESULTS: Treatment with PCB: (1) decreased the extracellular dopamine level in rat striatum, (2) increased p-nNOS expression both in rat striatal tissue and in CATH.a cells, (3) reduced the cell viability of, and (4) increased LDH release by CATH.a cells. However, Vitamin E showed a protective effect against PCB-induced toxicity and downregulation of the extracellular dopamine level. These results indicate that Vitamin E may have neuroprotective effects by inhibiting PCB-induced nNOS phosphorylation.


Assuntos
Dopamina/metabolismo , Disruptores Endócrinos , Síndromes Neurotóxicas/prevenção & controle , Óxido Nítrico Sintase Tipo I/metabolismo , Bifenilos Policlorados/toxicidade , Vitamina E/farmacologia , Administração Oral , Animais , Técnicas de Cultura de Células , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Immunoblotting , Masculino , Microdiálise , Síndromes Neurotóxicas/etiologia , Síndromes Neurotóxicas/metabolismo , Óxido Nítrico Sintase Tipo I/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Bifenilos Policlorados/antagonistas & inibidores , Ratos , Ratos Sprague-Dawley , Fatores de Tempo , Regulação para Cima/efeitos dos fármacos , Vitamina E/administração & dosagem , Vitamina E/uso terapêutico
16.
Toxicology ; 200(2-3): 93-101, 2004 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-15212806

RESUMO

The relationship between depleting effects of polychlorinated biphenyls (PCBs) on the intracellular calcium store and PCBs-induced cell death in dopaminergic cells has not been fully evaluated. Here, we evaluated the effects of inhibitors of the release of ER-stored calcium on the cytotoxicities induced by 10 microg/ml of Aroclor 1254 (A1254; polychlorinated biphenyl mixture) in a catecholaminergic cell-line, CATH.a cells. Exposure to A1254 produced an elevation in free calcium ([Ca2+]i) in the presence or absence of extracellular calcium and decreased in cell viability. From our results, we deduced that the A1254-induced elevation of [Ca2+]i resulted from the depletion of ER-stored calcium. The [Ca2+)]i elevation was dramatically inhibited by an inositol 1,4,5-triphosphate receptor (IP3R) antagonist, and slightly inhibited by a ryanodine receptor (RyR) blocker. IP3R blockers conferred significant protection against A1254-induced cell death, as did RyR blockers, but calcium chelators or NMDA blockers did not. However, none of these reagents inhibited the depletion of intracellular dopamine by A1254 indicating that the mechanism of PCB-induced dopamine depletion may be independent of calcium alterations. Taken together, these data suggest that agents inhibiting the receptor-mediated depletion of stored calcium can prevent the A1254-induced cell death, but not modulate the A1254-induced intracellular dopamine depletion in CATH.a cells.


Assuntos
Cálcio/metabolismo , Catecolaminas/metabolismo , /antagonistas & inibidores , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio , Morte Celular/efeitos dos fármacos , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Maleato de Dizocilpina/farmacologia , Dopamina/metabolismo , Dopamina/fisiologia , Retículo Endoplasmático/metabolismo , Inibidores Enzimáticos/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Humanos , Receptores de Inositol 1,4,5-Trifosfato , L-Lactato Desidrogenase/metabolismo , Compostos Macrocíclicos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Oxazóis/farmacologia , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Canal de Liberação de Cálcio do Receptor de Rianodina/efeitos dos fármacos , Tapsigargina/farmacologia
17.
Bioconjug Chem ; 15(2): 326-32, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15025528

RESUMO

Receptor-associated protein (RAP) is a ligand for all members of low-density lipoprotein (LDL) receptor families. RAP is internalized into cells via receptor-mediated endocytic trafficking, making it an attractive mechanism for efficient gene delivery. In this study, we have developed a gene delivery system using RAP as a targeting ligand. A RAP cDNA lacking a C-terminal heparin-binding domain was amplified by polymerase chain reaction (PCR) from a human liver cDNA library and was reamplified by using a primer containing a cysteine codon at its carboxyl end to facilitate its conjugation to polylysine (polyK). RAP was purified using a bacterial expression system and coupled to poly-D-lysine (PDL) or poly-L-lysine (PLL) of average MW 50 kDa via the heterobifunctional cross-linker SPDP. Using fluorescence-labeled RAP ligand, cellular uptake of the transfection complexes into HepG2 cells was shown to be highly efficient and more specific to PDL-conjugated RAP compared with PLL-conjugated one. Plasmid DNA containing a luciferase reporter gene was condensed with either RAP-PDL or RAP-PLL. In vitro transfection into HepG2 cells with RAP-PDL conjugate resulted in significantly higher luciferase expression levels in comparison to either nonconjugated PDL, or RAP-PLL, or LipofecAMINE/DNA complexes in the presence of 10% fetal bovine serum. Luciferase expression was inhibited by the addition of excess RAP. Treatment of the cells with Lovastatin, which inhibits HMG-Co reductase and increases expression of LDL receptor, stimulates luciferase expression, suggesting that the gene delivery is specifically mediated by LDL receptor. Thus, RAP-PDL conjugates have the potential to be used as a new nonviral gene delivery vector.


Assuntos
Carcinoma Hepatocelular/genética , Técnicas de Transferência de Genes , Proteína Associada a Proteínas Relacionadas a Receptor de LDL/genética , Polilisina/genética , Ligação Competitiva , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Humanos , Proteína Associada a Proteínas Relacionadas a Receptor de LDL/metabolismo , Lovastatina/farmacologia , Polilisina/metabolismo , Transfecção/métodos
18.
Mol Cells ; 13(2): 221-7, 2002 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-12018843

RESUMO

Methamphetamine (METH) causes neurotoxic damages to the dopaminergic system in mammals, but whether it exerts toxicity to dopamine cells in culture has not been fully explored. In order to develop an in vitro model of METH-induced dopamine neurotoxicity toward more systemical examination of the mechanism, we investigated METH toxicity in a clonal dopamine producing cell line (CATH.a). We show in the present study that METH produces a time- and dose-dependent increase in cell death via a process similar to apoptosis. The METH toxicity seems to be produced by oxidative stress, as it was attenuated by the antioxidant glutathione, and to involve dopamine because dopamine release and synthesis inhibitors attenuated the toxicity. This catecholaminergic cell line derived from the central nervous system may become a useful in vitro model to elucidate the mechanism underlying the METH-induced dopaminergic neuronal damage.


Assuntos
Apoptose/efeitos dos fármacos , Sistema Nervoso Central/fisiologia , Dopamina/metabolismo , Metanfetamina/farmacologia , Tetra-Hidroisoquinolinas , Animais , Linhagem Celular , Sistema Nervoso Central/citologia , Cicloeximida/farmacologia , Dactinomicina/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/metabolismo , Glutationa/metabolismo , Humanos , Isoquinolinas/farmacologia , Metanfetamina/toxicidade , Estresse Oxidativo , Inibidores da Síntese de Proteínas/farmacologia , Fatores de Tempo
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