RESUMEN
OBJECTIVE: Cisplatin is a widely used anticancer drug that provokes various side effects. Nephrotoxicity is one of the well-known major side effects in the chemotherapeutic use of cisplatin. Reactive oxygen species (ROS) and p53 play important roles in cisplatin-induced nephrotoxicity. AMP-activated protein kinase (AMPK) is known to be sensitively activated by ROS and can directly activate p53. The present study investigated the role of AMPK on cisplatin-induced apoptosis in rat renal epithelial NRK-52E cells. MATERIALS AND METHODS: NRK-52E cells were treated with cisplatin in the absence or presence of specific ROS scavenger and AMPK inhibitor for indicated times under the serum-free condition. The expression and phosphorylation levels of proteins were evaluated by Western blot and densitometry analysis. RESULTS: Cisplatin induced apoptotic cell death through ROS-mediated p53 activation, which is associated with AMPK activation. AMPK inhibitor suppressed cisplatin-induced p53 activation, as well as AMPK activation. Interestingly, ROS scavenger also diminished cisplatin-induced p53 activation and AMPK activation. Furthermore, cisplatin induced phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), which attenuated p53 activation, but did not affect the expression levels of total p53, cleaved caspase-3 and PARP. Meanwhile, inhibition of AMPK induced premature phosphorylation of eIF2α in cisplatin-treated cells. CONCLUSIONS: Taken together, these suggest that AMPK may be required for activation of p53 by oxidative stress in cisplatin-induced nephrotoxicity. Moreover, eIF2α phosphorylation may interrupt the AMPK-activated p53 in NRK-52E cells exposed to cisplatin, but does not critically affect cisplatin-induced nephrotoxicity because AMPK activation can be disrupted eIF2α phosphorylation.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Antineoplásicos/toxicidad , Cisplatino/toxicidad , Especies Reactivas de Oxígeno/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Apoptosis/efectos de los fármacos , Línea Celular , Factor 2 Eucariótico de Iniciación/metabolismo , Enfermedades Renales/inducido químicamente , Enfermedades Renales/metabolismo , Estrés Oxidativo/efectos de los fármacos , RatasRESUMEN
Cigarette smoking is a major risk factor for pulmonary diseases, including chronic obstructive pulmonary disease (COPD) and cancer. Cigarette smoke is reported to contain over 4,000 chemical compounds. Therefore, it needs to study the effects of cigarette smoke extract (CSE) administration on intracellular calcium concentration. In this study, we investigated how CSE influences intracellular calcium concentration in human lung adenocarcinoma A549 cells. The CSE concentrations used (0.4, 2, 3%) did not influence cell viability. However, at these CSE concentrations, calcium influx transient receptor potential vanilloid 4 (TRPV4) and transient receptor potential vanilloid 6 (TRPV6) proteins significantly increased, whereas calcium efflux sodium-calcium exchanger (NCX1) and plasma membrane Ca2+ ATPase (PMCA1) proteins significantly decreased from those of the control cells. The 3% CSE treatment produced an intracellular calcium concentration higher than that of the control treatment through methods of co-transfection of pGP-CMV-GCaMP6f/CMV-R-GECO1.2 and Rhod-4 Assay. CSE induced concentration-dependent increments in hypoxia-inducible factor (HIF)-1α and HIF-2α protein levels. Moreover, phosphorylation of ERK and Akt was induced by CSE treatment. Also, mitochondrial marker B-cell lymphoma 2 (Bcl-2) protein level decreased and Bcl-2-associated X (Bax) protein level increased following CSE treatment. Also, endoplasmic reticulum (ER) stress markers BiP, CHOP, p-SAPK, and p-eIF2α levels were increased by CSE treatment. These results suggest that CSE may increase the concentration of intracellular calcium, thus increasing mitochondrial and ER stress.
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Calcio/metabolismo , Nicotiana/efectos adversos , Humo/efectos adversos , Células A549 , Estrés del Retículo Endoplásmico/efectos de los fármacos , Humanos , Intercambiador de Sodio-Calcio/fisiologíaRESUMEN
Dexamethasone inhibits mucin secretion considering the primary option for treating acute asthma exacerbation. However, the mechanism underlying dexamethasone-induced decreased in mucosecretion is unclear. Recent studies have reported that dexamethasone exerts an inhibitory effect on mucosecretion in the lung by modulating the expression of calcium processing genes. However, the expression of the calcium processing genes in the trachea is not examined yet. Thus, the present study is the first to report the localization of calcium processing proteins such as transient receptor potential vanilloid-4 (Trpv4), transient receptor potential vanilloid-6 (Trpv6), calbindin-D9k (CaBP-9k) and plasma membrane Ca2+-ATPase 1 (Pmca1) in the mouse trachea and their glucocorticoid-induced response. In this study, mice were subcutaneously injected with dexamethasone for 5 days, and their tracheal samples were collected by dividing the trachea into the cervical, and thoracic sections based on its anatomical structure. The localization of TRPV4, TRPV6, CaBP-9k, and PMCA1 proteins was detected in the tracheal epithelium, submucosal glands, cartilages and muscles. Dexamethasone treatment downregulated the mRNA expression of the four calcium processing genes and mucin producing genes. The dexamethasone-induced decrease in the secretion of mucosubstances in the trachea was determined by performing Alcian blue-periodic acid-Schiff staining. Thus, the findings of the present study suggest that glucocorticoids simultaneously can regulate the expression of calcium processing genes and tracheal mucosecretion.
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Dexametasona/farmacología , Glucocorticoides/farmacología , Mucosa Respiratoria/efectos de los fármacos , Animales , Canales de Calcio/genética , Canales de Calcio/metabolismo , Femenino , Masculino , Ratones Endogámicos C57BL , ATPasas Transportadoras de Calcio de la Membrana Plasmática/genética , ATPasas Transportadoras de Calcio de la Membrana Plasmática/metabolismo , Mucosa Respiratoria/metabolismo , Proteína G de Unión al Calcio S100/genética , Proteína G de Unión al Calcio S100/metabolismo , Canales Catiónicos TRPV/genética , Canales Catiónicos TRPV/metabolismo , Tráquea/efectos de los fármacos , Tráquea/metabolismoRESUMEN
OBJECTIVE: Nephrotoxicity is one of the major side effects that limit the use of cisplatin in cancer therapy. Cisplatin-induced apoptosis in renal cells is associated with reactive oxygen species (ROS)-mediated p53 activation. Licorice (Glycyrrhiza uralensis Fischer) is one of the most widely used medicinal herbs in Korea, China and Japan. The aim of the study was to evaluate the protective effects of licorice extract (LE) and its active compound glycyrrhizic acid (GA) against cisplatin-induced nephrotoxicity in human renal proximal tubular epithelial (HK-2) cells. MATERIALS AND METHODS: HK-2 cells were pretreated with LE or GA for 1 h and then treated with 40 µM of cisplatin for indicated times under the serum-free condition. Cell viability was evaluated by MTT assay. Apoptosis was evaluated by flow cytometric analysis and caspase-3 activity. The intracellular ROS levels were determined by DCFH-DA assay. The expression and phosphorylation levels of protein were evaluated by Western blot and densitometry analysis. RESULTS: When treating HK-2 cells with LE or GA, both of them alleviated cisplatin-induced cytotoxicity and apoptosis. LE and GA inhibited caspase-3 activity and polymerase (PARP) cleavage in cisplatin-treated cells. LE and GA also inhibited p53 expression and its phosphorylation as well as ROS production in cells exposed to cisplatin. Meanwhile, LE and GA enhanced cisplatin-induced p21 expression, which then led to S-phase arrest in cell cycle and limited cell growth. Presumably, increased p21 expression may contribute to cellular prevention from cisplatin-induced apoptosis, because p21 is the key molecule to cytoprotection during cisplatin-induced nephrotoxicity. CONCLUSIONS: These results suggest that LE and GA ameliorate cisplatin-induced apoptosis through reduction of ROS-mediating p53 activation and promotion of p21 expression in HK-2 cells.
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Cisplatino/efectos adversos , Células Epiteliales/efectos de los fármacos , Ácido Glicirrínico/farmacología , Extractos Vegetales/farmacología , Especies Reactivas de Oxígeno/metabolismo , Antineoplásicos/efectos adversos , Apoptosis/efectos de los fármacos , Línea Celular , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Células Epiteliales/metabolismo , Glycyrrhiza/química , Humanos , Túbulos Renales Proximales/citología , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
BACKGROUND/OBJECTIVES: Activation of Notch signaling pathologically enhances lipogenesis and gluconeogenesis in the liver causing non-alcoholic fatty liver disease (NAFLD) and diabetes. Delta-like 1 homolog (DLK1), an imprinted gene that can modulate adipogenesis and muscle development in mice, was found as an inhibitory regulator of Notch signaling. Therefore, we investigated the metabolic effect of exogenous DLK1 in vitro and in vivo. SUBJECTS/METHODS: A soluble DLK1 peptide was generated with fusion between a human Fc fragment and extracellular domain of DLK1. Male db/db mice were randomly assigned to two groups: vehicle treated and DLK1-treated group (25 mg kg(-1), intraperitoneal injection, twice a week for 4 weeks). Primary mice hepatocytes and HepG2 cells were used for in vitro experiments. RESULTS: After 4 weeks of DLK1 administration, hepatic triglyceride content and lipid droplets in liver tissues, as well as serum levels of liver enzymes, were markedly decreased in db/db mice. DLK1 treatment induced phosphorylation of AMPK and ACC and suppressed nuclear expression of SREBP-1c in the mouse liver or hepatocytes, indicating regulation of fatty acid oxidation and synthesis pathways. Furthermore, DLK1-treated mice showed significantly lower levels of fasting and random glucose, with improved glucose and insulin tolerance compared with the vehicle-treated group. Macrophage infiltration and proinflammatory cytokine levels in the epididymal fat were decreased in DLK1-treated db/db mice. Moreover, DLK1 suppressed glucose production from hepatocytes, which was blocked after co-administration of an AMPK inhibitor, compound C. DLK1-treated hepatocytes and mouse liver tissues showed lower PEPCK and G6Pase expression. DLK1 triggered AKT phosphorylation followed by cytosolic translocation of FOXO1 from the nucleus in hepatocytes. CONCLUSIONS: The present study demonstrated that exogenous administration of DLK1 reduced hepatic steatosis and hyperglycemia via AMPK activation in the liver. This result suggests that DLK1 may be a novel therapeutic approach for treating NAFLD and diabetes.
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Péptidos y Proteínas de Señalización Intercelular/farmacología , Hígado/patología , Enfermedad del Hígado Graso no Alcohólico/patología , Receptores Notch/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Proteínas Quinasas Activadas por AMP , Animales , Proteínas de Unión al Calcio , Modelos Animales de Enfermedad , Gluconeogénesis , Inyecciones Intraperitoneales , Metabolismo de los Lípidos , Ratones , Ratones Endogámicos C57BL , Receptores Notch/metabolismoRESUMEN
We investigated the microscopic mechanism underlying the double-state lasing behavior (simultaneous lasing at the ground state [GS] and excited state [ES]) in InAs/GaAs quantum dot (QD) laser diodes. The ES and GS lasing processes that contributed to double-state lasing were examined experimentally and theoretically. Experiments were conducted in which spontaneous emission from a window of a QD laser diode was examined under lasing conditions, and numerical simulations were performed using a coupled rate equation model of the QD microstates. The findings showed that, when carrier relaxation from the ES to the GS was sufficiently slow, double-state lasing occurred. Additionally, ES lasing was found to arise not from the QD group undergoing GS lasing, but rather from another QD group in which the states were lower in energy and outside of the homogeneous bandwidth.
RESUMEN
OBJECTIVE: Indoxyl sulfate (IS) has been reported to induce endoplasmic reticulum (ER) stress in tubular cells and to inhibit the cell proliferation via ER stress and ERK/IL-6/p21 pathways. This study has investigated the effect of apigenin on IS-induced ER stress in immortalized human renal proximal tubular HK-2 cells. MATERIALS AND METHODS: Human Kidney 2 (HK-2) cells were treated with IS (5 mM) in the absence or presence of apigenin (10 µM) or salubrinal (20 µM) for indicated times under the serum-free condition. Cell viability was evaluated by MTT assay. The levels of protein expression and phosphorylation were evaluated by Western blot analysis. RESULTS: In HK-2 cells, apigenin completely inhibited IS-induced ER stress, as indicated by decreased expression of CHOP, ATF4 and GRP78, although the phosphorylated level of eIF2α did not decrease. IS-induced expression levels of IL-6 and p21 proteins were also inhibited by apigenin, with no significant changes in ERK activation. The suppression of cell proliferation by IS was abolished by salubrinal, an ER stress inhibitor, but not by apigenin. Apigenin inhibited the phosphorylation of Akt and GSK-3ß in IS-treated HK-2 cells. The phosphorylation of GSK-3ß, which was inhibited by apigenin, resulted in hypo-phosphorylation of retinoblastoma (Rb) protein, which was associated with the decrease in cyclin D1 expression. CONCLUSIONS: These results suggest that apigenin may inhibit IS-induced ER stress and expression of IL-6 and p21 proteins in HK-2 cells. It is most likely that apigenin, together with its inhibitory effect on ER stress, may also suppress the cell growth by inducing the loss of Rb phosphorylation, which was associated with the decrease in cyclin D1 expression by GSK-3ß activation through the inhibition of PI3K/Akt pathway.
Asunto(s)
Apigenina/farmacología , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/biosíntesis , Estrés del Retículo Endoplásmico/fisiología , Indicán/toxicidad , Interleucina-6/biosíntesis , Túbulos Renales Proximales/metabolismo , Factor de Transcripción CHOP/biosíntesis , Ciclo Celular/efectos de los fármacos , Ciclo Celular/fisiología , Línea Celular Transformada , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/antagonistas & inhibidores , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Humanos , Indicán/antagonistas & inhibidores , Interleucina-6/antagonistas & inhibidores , Túbulos Renales Proximales/efectos de los fármacos , Factor de Transcripción CHOP/antagonistas & inhibidoresRESUMEN
UNLABELLED: This study investigated the effect of pesticide chlorpyrifos (CP) on a freshwater cyanobacterium Chroococcus turgidus NTMS12. The changes in chlorophyll-a, proline, superoxide dismutase (SOD), catalase (CAT) activities and fatty acid composition of the test organism were analysed. Organism was grown at 6, 9 and 12 mg l(-1) of CP, and based on the chlorophyll-a content, 6 mg l(-1) of CP was found to be the tolerable concentration. Hence, 6 mg l(-1) of CP was taken to evaluate the concentration of proline and activities of SOD and CAT at 48-h exposure. The changes in the fatty acid profile were analysed after 7 days of exposure. Upon pesticide exposure, increased concentration of proline and activities of SOD and CAT were found. Significant changes in fatty acid profile have also been observed. However, polyunsaturated fatty acid content was decreased in treated cultures when compared with the untreated control. Changes in biochemical activities indicate that cyanobacteria C. turgidus NTMS12 undergo adaptive changes against CP-induced oxidative stress. SIGNIFICANCE AND IMPACT OF THE STUDY: : Chlorpyrifos induces oxidative stress in Chroococcus turgidus NTMS12. A strong inference was made on increased activity of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT) and proline content and lowering the level of unsaturated fatty acids under the pesticide-exposed condition. These significant changes are the defence mechanisms against the oxidative stress. Thus, this organism holds great promise in resisting toxic pesticide.
Asunto(s)
Antioxidantes/metabolismo , Cloropirifos/toxicidad , Cianobacterias/efectos de los fármacos , Cianobacterias/metabolismo , Ácidos Grasos/metabolismo , Insecticidas/toxicidad , Catalasa/metabolismo , Clorofila/metabolismo , Clorofila A , Cianobacterias/enzimología , Agua Dulce/microbiología , Estrés Oxidativo , Prolina/metabolismo , Superóxido Dismutasa/metabolismoRESUMEN
BACKGROUND AND OBJECTIVES: Nephrotoxicity is one of the main side effects of the anticancer drug cisplatin, and one of its main therapeutic limitations. It has been suggested that p53 activation plays important roles in renal cell injury by cisplatin. However, the mechanism of p53 activation by cisplatin is unclear. This study examined whether reactive oxygen species (ROS) production by cisplatin would be linked to p53 activation in rat mesangial cells. MATERIALS AND METHODS: Renal cells were incubated with cisplatin in the absence or presence of pifithrin-a (PFT), N-acetyl-cysteine (NAC), or dimethylthiourea (DMT). Cell viability was evaluated by 3-(4,5-dimethyl-2-thiazol yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis was evaluated by caspase-3 activity and cleavage of poly (ADP-ribose) polymerase (PARP). The relative levels of ROS and p53 phosphorylation were determined by fluorometric assay and Western blot analysis, respectively. RESULTS: Cisplatin induced apoptotic cell death via caspase-3 activation and PARP cleavage, and also increased p53 activation and ROS production. The p53 inhibitor PFT inhibited cisplatin-induced apoptosis. NAC and DMT, two antioxidants, also inhibited cisplatin-induced apoptosis. Interestingly, NAC and DMT reduced ROS production and suppressed p53 activation in renal cells exposed to cisplatin. CONCLUSIONS: Our results suggest that the ability of cisplatin to induce apoptosis of rat mesangial cells requires ROS-dependent p53 activation, thus, supporting the potential therapeutic role of antioxidants in preventing the cisplatin nephrotoxicity.
Asunto(s)
Antineoplásicos/toxicidad , Apoptosis/efectos de los fármacos , Cisplatino/toxicidad , Células Mesangiales/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Antioxidantes/farmacología , Caspasa 3/metabolismo , Línea Celular , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Mesangiales/metabolismo , Células Mesangiales/patología , Fosforilación , Poli(ADP-Ribosa) Polimerasas/metabolismo , Ratas , Transducción de Señal/efectos de los fármacos , Factores de TiempoRESUMEN
In supraoptic nucleus (SON) magnocellular neurosecretory cells (MNCs), γ-GABA, via activation of GABAA receptors (GABAA Rs), mediates persistent tonic inhibitory currents (Itonic ), as well as conventional inhibitory postsynaptic currents (IPSCs, Iphasic ). In the present study, we examined the functional significance of Itonic in SON MNCs challenged by 24-h water deprivation (24WD). Although the main characteristics of spontaneous IPSCs were similar in 24WD compared to euhydrated (EU) rats, Itonic , measured by bicuculline (BIC)-induced Iholding shifts, was significantly smaller in 24WD compared to EU rats (P < 0.05). Propofol and diazepam prolonged IPSC decay time to a similar extent in both groups but induced less Itonic in 24WD compared to EU rats, suggesting a selective decrease in GABAA receptors mediating Itonic over Iphasic in 24WD rats. THIP (4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol), a preferential δ subunit agonist, and L-655,708, a GABAA receptor α5 subunit selective imidazobenzodiazepine, caused a significantly smaller inward and outward shift in Iholding , respectively, in 24WD compared to EU rats (P < 0.05 in both cases), suggesting an overall decrease in the α5 subunit-containing GABAA Rs and the δ subunit-containing receptors mediating Itonic in 24WD animals. Consistent with a decrease in 24WD Itonic , bath application of GABA induced significantly less inhibition of the neuronal firing activity in 24WD compared to EU SON MNCs (P < 0.05). Taken together, the results of the present study indicate a selective decrease in GABAA Rs functions mediating Itonic as opposed to those mediating Iphasic in SON MNCs, demonstrating the functional significance of Itonic with respect to increasing neuronal excitability and hormone secretion in 24WD rats.
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Potenciales Postsinápticos Inhibidores/fisiología , Células Neuroendocrinas/fisiología , Receptores de GABA-A/fisiología , Núcleo Supraóptico/citología , Núcleo Supraóptico/fisiología , Privación de Agua/fisiología , Animales , Bicuculina/farmacología , Diazepam/farmacología , Agonistas del GABA/farmacología , Antagonistas de Receptores de GABA-A/farmacología , Hipnóticos y Sedantes/farmacología , Imidazoles/farmacología , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Isoxazoles/farmacología , Masculino , Células Neuroendocrinas/efectos de los fármacos , Propofol/farmacología , Subunidades de Proteína/efectos de los fármacos , Subunidades de Proteína/fisiología , Ratas , Núcleo Supraóptico/efectos de los fármacos , Ácido gamma-Aminobutírico/farmacologíaRESUMEN
BACKGROUND: Nitric oxide (NO) is known to act cytostatically on several tumor cell when functioning as an effector molecule of activated macrophages, but the differential effects of NO on immortalized and malignant oral keratinocytes have not been examined. METHODS: We investigated the influence of NO on the proliferation, cell cycle, apoptosis, and differentiation of immortalized human oral keratinocytes (IHOK) and primary oral cancer cells (HN4) using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, sulforhodamine B (SRB) assay, flow cytometry, nuclear DNA staining, and Western blotting. RESULTS: The MTT and SRB assays indicated inhibited growth of IHOK and HN4 cells that were treated with sodium nitroprusside (SNP) at concentrations higher than 1 mM but not at lower SNP concentrations. The higher concentrations of SNP up-regulated the apoptosis-related protein expression, which is consistent with the analyses of sub-G(1) phase arrest, annexin V-FITC (fluorescein isothiocynate) staining, nuclear staining, and DNA fragmentation. On the other hand, the lower concentrations of SNP enhanced the expression of keratinocyte differentiation markers in IHOK and HN4 cells. CONCLUSIONS: These data suggest that high concentrations of NO can inhibit the growth of IHOK and HN4 cells through the induction of apoptosis, while low concentrations of NO can induce cytodifferentiation. The dual effects of NO, namely, the induction of apoptosis or cytodifferentiation, have important implications for the possible anti-oral cancer treatment.
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Apoptosis/efectos de los fármacos , Carcinoma de Células Escamosas/metabolismo , Queratinocitos/efectos de los fármacos , Neoplasias de la Boca/metabolismo , Óxido Nítrico/farmacología , Western Blotting , Proteínas de Ciclo Celular/metabolismo , Diferenciación Celular/efectos de los fármacos , Línea Celular Transformada , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Citocromos c/metabolismo , Humanos , Queratinocitos/citología , Mucosa Bucal/citología , Óxido Nítrico/fisiologíaRESUMEN
OBJECTIVE: We evaluated the potential of 15 herbal medicines (HMs), commonly used in Korea, to inhibit the catalytic activities of several cytochrome P450 (CYP) isoforms and microsomal NADPH-CYP reductase. METHODS: The abilities of 1-1000 microg/mL of freeze-dried aqueous extracts of 15 HMs to inhibit phenacetin O-deethylation (CYP1A2), tolbutamide 4-methylhydroxylation (CYP2C9), S-mephenytoin 4'-hydroxylation (CYP2C19), dextromethorphan O-demethylation (CYP2D6), chlorzoxazone 6-hydroxylation (CYP2E1), midazolam 1-hydroxylation (CYP3A4) and NADPH-CYP reductase were tested using human liver microsomes. RESULTS: The HMs Epimedii herba, Glycyrrhizae radix and Leonuri herba inhibited one or more of the CYP isoforms or NADPH-CYP reductase. Of the three HMs, Epimedii herba extracts were the most potent inhibitors of several CYP isoforms (IC(50) 67.5 microg/mL for CYP2C19, 104.8 microg/mL for CYP2E1, 110.9 microg/mL for CYP2C9, 121.9 microg/mL for CYP3A4, 157.8 microg/mL for CYP2D6 and 168.7 microg/mL for CYP1A2) and NADPH-CYP reductase (IC(50) 185.9 microg/mL ). CONCLUSION: These results suggest that some of the HMs used in Korea have the potential to inhibit CYP isoforms in vitro. Although the plasma concentrations of the active constituents of the HMs were not determined, some herbs could cause clinically significant interactions because the usual doses of those individual herbs are several grams of freeze-dried extracts. Controlled trials to test the significance of these results are necessary.
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Inhibidores Enzimáticos del Citocromo P-450 , Microsomas Hepáticos/efectos de los fármacos , NADPH-Ferrihemoproteína Reductasa/antagonistas & inhibidores , Plantas Medicinales/química , Basidiomycota/química , Sistema Enzimático del Citocromo P-450/metabolismo , Interacciones Farmacológicas , Medicamentos Herbarios Chinos/farmacología , Humanos , Corea (Geográfico) , Magnoliopsida/química , Microsomas Hepáticos/enzimología , NADPH-Ferrihemoproteína Reductasa/metabolismo , Extractos Vegetales/farmacología , Quercetina/farmacologíaAsunto(s)
Iduronidasa/genética , Mucopolisacaridosis I/genética , Adolescente , Adulto , Alelos , Niño , Preescolar , Análisis Mutacional de ADN , Humanos , Iduronidasa/deficiencia , Corea (Geográfico) , Mutación , LinajeRESUMEN
The sorption kinetics of the divalent metals Zn, Co, Ni, and Cd to hematite were studied in single sorbate systems with high sorbate/sorbent ratios (from 1.67 to 3.33mol sorbate/mol sorption sites) in 10mM Na-piperazine N,N'-bis 2-ethane sulfonic acid (Na-PIPES) solution at pH 6.8. The experimental data showed a rapid initial sorption (half-time about 1min) followed by slower sorption that continued for 1-5 days. The sequence of fast to slow sorption kinetics was modeled by slow inner-sphere (IS) complexation in equilibrium with outer-sphere (OS) complexes. Although the OS reaction was fast and considered to be in equilibrium, the extent of OS complexation changed over time due to increased surface potential from the IS complexes. For example, the model showed that the dimensionless OS complexation function, K(os), decreased from 0.014 initially to 0.0016 at steady state due to sorption of 4x10(-5)M Zn(II) to 2gL(-1) hematite. Sorption rate constants, k(ads), for the various divalent metals ranged from 6.1 to 82.5M(-1)s(-1). Desorption rate constants, k(des), ranged from 5.2x10(-7) to 6.7x10(-5)s(-1). This study suggests that the conversion from OS to IS complex was the rate-determining step for the sorption of divalent metals on crystalline adsorbents.
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Compuestos Férricos/química , Metales Pesados/química , Purificación del Agua/métodos , Adsorción , Cationes Bivalentes , Cloro/química , Concentración de Iones de Hidrógeno , Cinética , Modelos Químicos , Sulfitos/química , Ácidos Sulfónicos/química , Contaminantes del Agua/aislamiento & purificaciónRESUMEN
Fasciola hepatica, a parasitic flatworm belonging to the Class Trematoda, is one of the first metazoan groups to possess a centralized nervous system. However, the electrophysiological properties of neurones in F. hepatica are largely unknown. In the present study, we acutely isolated viable neurones from F. hepatica and characterized their electrophysiological properties. A hyperpolarization-activated cation current was recorded in the cells using the whole-cell patch-clamp. The current was found to be activated slowly at membrane potentials negative to 0 mV and did not display any time-dependent inactivation. This current was reduced by 1 mM Gd3+ to the level of the leak current, while 3 mM of Cs+ had no effect. However, the current was inhibited by extracellular acidosis in the pH range 7.0-7.8, and the membrane potentials of these cells were depolarized by extracellular alkalosis in the pH range of 5.8 to 8.2. Gd3+ (1 mM), which inhibited the pH-sensitive hyperpolarization-activated cation current, also hyperpolarized the cells. In summary, we isolated single neurones from F. hepatica, and these were found to express a pH-sensitive hyperpolarization-activated cation current. This current may participate in the membrane depolarization of F. hepatica neurones during alkaline challenge.
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Fasciola hepatica/citología , Gadolinio/farmacología , Activación del Canal Iónico/efectos de los fármacos , Canales Iónicos/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Animales , Cationes/metabolismo , Cesio/farmacología , Canales Catiónicos Regulados por Nucleótidos Cíclicos , Conductividad Eléctrica , Concentración de Iones de Hidrógeno , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización , Potenciales de la Membrana/efectos de los fármacos , Técnicas de Placa-Clamp , Canales de Potasio , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The rac1 GTPase promotes oxidative stress through reactive oxygen species (ROS) production, whereas the DNA repair enzyme and transcriptional regulator redox factor-1 (ref-1) protects against cell death due to oxidative stimuli. However, the function of ref-1 in regulating intracellular oxidative stress, particularly that induced by rac1, has not been defined. We examined the role of ref-1 in vascular endothelial cell oxidative stress and apoptosis. Ref-1 was expressed in both the cytoplasm and nuclei of resting endothelial cells. Cytoplasmic ref-1 translocated to the nucleus with the oxidative trigger hypoxia/reoxygenation (H/R). Forced cytoplasmic overexpression of ref-1 suppressed H/R-induced oxidative stress (H(2)O(2) production), NF-kappaB activation, and apoptosis, and also mitigated rac1-regulated H(2)O(2) production and NF-kappaB transcriptional activity. We conclude that inhibition of oxidative stress is another mechanism by which ref-1 protects against apoptosis, and that this is achieved through modulation of cytoplasmic rac1-regulated ROS generation. This suggests a novel extra-nuclear function of ref-1.
Asunto(s)
Apoptosis , Liasas de Carbono-Oxígeno/metabolismo , ADN-(Sitio Apurínico o Apirimidínico) Liasa , Estrés Oxidativo , Adenoviridae , Liasas de Carbono-Oxígeno/biosíntesis , Liasas de Carbono-Oxígeno/genética , Hipoxia de la Célula , Línea Celular , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Endotelio Vascular/citología , Expresión Génica , Vectores Genéticos , Humanos , FN-kappa B/metabolismo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
We cloned and sequenced a gene, kpcA (Kex2p-like proprotein convertase A), from a genomic library of Aspergillus nidulans. The kpcA gene encodes an 820-residue protein, named KpcA, which contains a putative subtilisin-like catalytic domain (residues 136-466) homologous to that of the subtilisin serine protease family. KpcA shows 56, 73, and 47% amino acid identities with Saccharomyces cerevisiae Kex2p, Aspergillus niger KexB, and mouse furin within the subtilisin-like catalytic domain, respectively. The sequences around the proposed active site Asp, His, and Ser residues of KpcA are similar to those of other Kex2p family members. The KpcA mRNA transcript with an expected size of approximately 2.8 kb was detected in A. nidulans. The substrate specificity of KpcA, expressed in CHO cells, is similar to that of A. niger KexB and yeast Kex2p. We conclude that KpcA is a resident Kex2p-like proprotein that processes endoprotease in A. nidulans.
Asunto(s)
Aspergillus nidulans/enzimología , Endopeptidasas/genética , Proteínas Fúngicas/genética , Proproteína Convertasas , Proteínas de Saccharomyces cerevisiae , Secuencia de Aminoácidos , Animales , Aspergillus nidulans/fisiología , Secuencia de Bases , Clonación Molecular , Endopeptidasas/química , Endopeptidasas/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Humanos , Datos de Secuencia Molecular , Estructura Terciaria de Proteína , Alineación de Secuencia , Especificidad por Sustrato , Subtilisinas/química , Subtilisinas/genética , Subtilisinas/metabolismoRESUMEN
To investigate whether alpha1-adrenoceptors are involved in pain behaviors in streptozotocin (STZ)-induced diabetic rats, we measured the effects of phenylephrine or prazosin on allodynia in the diabetic rats. Phenylephrine aggravated allodynia, while prazosin alleviated allodynia in the diabetic rats. We also measured alpha1-adrenoceptors gene expression or density of [3H]-prazosin binding sites in the dorsal root ganglia (DRG) and spinal cord in painful diabetic rats. Alpha1-adrenoceptors mRNA and density of [3H]prazosin binding sites were increased in the DRG of the diabetic rats, however there were no significant differences in alpha1-adrenoceptors expression in the spinal cord between the control and diabetic rats. These results suggest increased alpha1-adrenoceptors in the DRG may play a role in the pathogenesis of painful diabetic neuropathy.
Asunto(s)
Neuropatías Diabéticas/fisiopatología , Dolor/fisiopatología , Receptores Adrenérgicos alfa 1/metabolismo , Antagonistas Adrenérgicos alfa/metabolismo , Antagonistas Adrenérgicos alfa/farmacología , Animales , Diabetes Mellitus Experimental/fisiopatología , Ganglios Espinales/química , Ganglios Espinales/fisiología , Expresión Génica/fisiología , Masculino , Umbral del Dolor/efectos de los fármacos , Prazosina/metabolismo , Prazosina/farmacología , ARN Mensajero/análisis , Ensayo de Unión Radioligante , Ratas , Ratas Sprague-Dawley , Receptores Adrenérgicos alfa 1/análisis , Receptores Adrenérgicos alfa 1/genética , Médula Espinal/química , Médula Espinal/fisiología , TritioRESUMEN
The change of blood pressure and heart rate after intravenous injection of Korea red ginseng (KRG) were studied in the conscious normotensive and one-kidney, one-clip Goldblatt hypertensive (1K, 1C-GBH) rats. Crude saponin (CS) of KRG (50, 100 mg/kg i.v.) induced a hypotensive effect and bradycardia in a dose-dependent manner in the anesthetized rats. On the other hand, CS of KRG (100 mg/kg) induced a hypotensive effect and reflex tachycardia in the conscious rats. Saponin-free fraction (SFF) of KRG did not affect them in the anesthetized normotensive rats (P>.05). The maximal hypotensive effect by CS of KRG in the conscious 1K, 1C-GBH hypertensive rats and L-nitroarginine methyl ester (L-NAME, 40 mg/kg)-treated conscious hypertensive rats was not different from that of conscious normotensive rats (Delta 31.6+/-6.3, Delta 27.5+/-5.8 vs. Delta 26.7+/-4.3 mmHg, P>.05). However, pretreatment of L-NAME significantly inhibited the reflex tachycardia by CS of KRG (70.8+/-7.0 vs. 30.6+/-15.0 bpm, P<.05). Hemolysate-sensitive nitric oxide (NO) current by the CS of KRG was greater than that of the SFF of KRG (651.9+/-128.2 pA for CS and 164.9+/-92.5 pA for SFF, P<.001). These findings suggest that KRG has a hypotensive effect and its effect may be due to saponin fraction of KRG in the conscious rats. The releasing effect of NO of KRG, like NO donor, may be partly contributed to the hypotensive effect of KRG.
Asunto(s)
Presión Sanguínea/efectos de los fármacos , Hipertensión/tratamiento farmacológico , Panax/química , Fitoterapia/métodos , Animales , Presión Sanguínea/fisiología , Estado de Conciencia/fisiología , Relación Dosis-Respuesta a Droga , Frecuencia Cardíaca/efectos de los fármacos , Frecuencia Cardíaca/fisiología , Masculino , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , Ratas , Ratas Sprague-DawleyRESUMEN
The effect of high blood flow on the expression of endothelial nitric oxide synthase has been investigated in the femoral arteriovenous shunt (AVS) rats created by inserting U-shaped polyurethane tubes in the left femoral arteries and veins. Three days after inserting the femoral AVS, the mean aortic blood flow rate in the abdominal aorta of the AVS rats was about 2.0 times higher than that in the control rats (110.0 +/- 8.4 ml/min vs 52.7 +/- 2.7 ml/min, p < 0.001). The competitive reverse transcriptase-polymerase chain reaction (RT-PCR) data revealed that the mRNA expression level of the endothelial constitutive nitric oxide synthase (ecNOS) was increased in the aortas of the femoral AVS rats compared to that in the control rats. Western blot analysis using a monoclonal antibody against ecNOS revealed that the ecNOS protein levels were markedly increased in the aortas of femoral AVS rats, but ecNOS protein levels in aortas without endothelium were not significantly increased. Inducible nitric oxide synthase (iNOS) protein was not expressed in the aortic tissues with and without endothelium in the control rats. This iNOS expression was not increased by the high blood flow in the femoral AVS rats. These findings suggest that high blood flow could up-regulate the expression levels of ecNOS mRNA and proteins in femoral arteriovenous shunt rats.