RESUMEN

Methicillin-resistant Staphylococcus aureus (MRSA) is an important human pathogen that is cross-resistant to most ß-lactam antibiotics. We investigated whether oxacillin, which is a ß-lactam antibiotic, alone or in combination with punicalagin can affect the penicillin binding protein 2a (PBP2a)-mediated resistance of MRSA. Susceptibility testing of punicalagin with oxacillin was performed using the microdilution and checkerboard assay and the growth curve assay. Binding affinity of punicalagin for cell wall peptidoglycan (PGN) was confirmed by an increased concentration of PGN in bacterial cultures containing punicalagin. The level of PBP2a was analyzed by western blotting. Punicalagin exhibited antimicrobial activity in the viability assay and increased the susceptibility of MRSA to oxacillin. PGN interfered with the antimicrobial activity of punicalagin and prevented the synergistic activity of punicalagin and oxacillin. Increasing the concentration of punicalagin and maintaining a constant concentration of oxacillin resulted in synergistic suppression of the expression of the mec operon (mecA, mecI, and mecR1). The production of PBP2a was suppressed by the addition of punicalagin to oxacillin. Our findings demonstrate that punicalagin potentiates the effect of oxacillin on MRSA by reducing the transcription of mecA (a gene marker for methicillin resistance), which resulted in a reduced level of PBP2a.

Asunto(s)

Antibacterianos/farmacología , Proteínas Bacterianas/genética , Taninos Hidrolizables/farmacología , Resistencia a la Meticilina/efectos de los fármacos , Resistencia a la Meticilina/genética , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Oxacilina/farmacología , Proteínas de Unión a las Penicilinas/genética , Pared Celular/metabolismo , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Expresión Génica/efectos de los fármacos , Taninos Hidrolizables/metabolismo , Staphylococcus aureus Resistente a Meticilina/citología , Staphylococcus aureus Resistente a Meticilina/genética , Staphylococcus aureus Resistente a Meticilina/ultraestructura , Pruebas de Sensibilidad Microbiana/métodos , Peptidoglicano/metabolismo , Transcripción Genética/efectos de los fármacos

RESUMEN

The objective of the present study was to investigate the antibacterial activity of a single constituent, ursolic acid 3-O-α-L-arabinopyranoside (URS), isolated from the leaves of Acanthopanax henryi (Oliv.) Harms, alone and in combination with oxacillin (OXA) against methicillin-resistant Staphylococcus aureus (MRSA). A broth microdilution assay was used to determine the minimal inhibitory concentration (MIC). The synergistic effects of URS and OXA were determined using a checkerboard dilution test and time-kill curve assay. The mechanism of action of URS against MRSA was analyzed using a viability assay in the presence of a detergent and an ATPase inhibitor. Morphological changes in the URS-treated MRSA strains were evaluated via transmission electron microscopy (TEM). In addition, the producing penicillin-binding protein 2a (PBP2a) protein level was analyzed using western blotting. The MIC value of URS against MRSA was found to be 6.25 µg/ml and there was a partial synergistic effect between OXA and URS. The time-kill growth curves were suppressed by OXA combined with URS at a sub-inhibitory level. Compared to the optical density at 600 nm (OD600) value of URS alone (0.09 µg/ml), the OD600 values of the suspension in the presence of 0.09 µg/ml URS and 0.00001% Triton X-100 or 250 µg/ml N,N'-dicyclohexylcarbodiimide reduced by 56.6 and 85.9%, respectively. The TEM images of MRSA indicated damage to the cell wall, broken cell membranes and cell lysis following treatment with URS and OXA. Finally, an inhibitory effect on the expression of PBP2a protein was observed when cells were treated with URS and OXA compared with untreated controls. The present study suggested that URS was significantly active against MRSA infections and revealed the potential of URS as an effective natural antibiotic.

Asunto(s)

Antibacterianos/farmacología , Pared Celular/efectos de los fármacos , Glicósidos/farmacología , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Oxacilina/farmacología , Triterpenos/farmacología , Antibacterianos/aislamiento & purificación , Pared Celular/metabolismo , Pared Celular/ultraestructura , Combinación de Medicamentos , Sinergismo Farmacológico , Eleutherococcus/química , Glicósidos/aislamiento & purificación , Staphylococcus aureus Resistente a Meticilina/crecimiento & desarrollo , Staphylococcus aureus Resistente a Meticilina/metabolismo , Staphylococcus aureus Resistente a Meticilina/ultraestructura , Pruebas de Sensibilidad Microbiana , Microscopía Electrónica de Transmisión , Extractos Vegetales/química , Hojas de la Planta/química , Triterpenos/aislamiento & purificación , Ácido Ursólico

RESUMEN

Staphylococcus aureus produces a number of virulence factors. The major virulence factors exhibited by S aureus include various antigens, enzymes, cytotoxins and exotoxins (e.g. hemolysins, enterotoxins and toxic shock syndrome toxin). In this report, we show the influence of punicalagin on the secretion of exoprotein from S aureus by western blotting, tumor necrosis factor (TNF) release assay and quantitative RT-PCR. When added to S aureus cultures at an OD600 of 0.9, graded subinhibitory concentrations of punicalagin reduced the production of α-toxin, SEA and SEB in methicillin-resistant Staphylococcus aureus in a dose-dependent manner. Consistently, punicalagin reduced TNF-inducing activity by S aureus culture supernatants. Here, the transcriptional level of agr (accessory gene regulator) in S aureus was inhibited by punicalagin, suggesting that the reduced transcription may affect the secretion of exotoxins. These findings suggest that the expression of α-toxin and enterotoxins in S aureus is sensitive to the action of punicalagin, which may be an advantageous candidate in the treatment of toxigenic staphylococcal disease.

Asunto(s)

Antibacterianos/farmacología , Toxinas Bacterianas/biosíntesis , Enterotoxinas/biosíntesis , Proteínas Hemolisinas/biosíntesis , Taninos Hidrolizables/farmacología , Staphylococcus aureus Resistente a Meticilina/metabolismo , Staphylococcus aureus Resistente a Meticilina/patogenicidad , Animales , Proteínas Bacterianas/genética , Línea Celular , Enterotoxinas/metabolismo , Proteínas Hemolisinas/metabolismo , Ratones , Células RAW 264.7 , Transactivadores/genética , Transcripción Genética/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo , Factores de Virulencia/biosíntesis

RESUMEN

Salidroside [2-(4-hydroxyphenyl)ethyl ß-D-gluco-pyranoside (SAS)] has been identified as the most potent ingredient of the plant Rhodiola rosea L. Previous studies have demonstrated that it possesses a number of pharmacological properties, including anti-aging, anti-fatigue, antioxidant, anticancer and anti-inflammatory properties. In this study, to ascertain the molecular mechanisms responsible for the anti-inflammatory activity of SAS, we used phorbol-12-myristate-13-acetate (PMA) plus A23187 to induce inflammation in human mast cell line-1 (HMC-1). The HMC-1 cells were treated with SAS prior to being stimulated with PMA plus A23187. Pro-inflammatory cytokine production was measured by enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). Western blot analysis was used to examine the activation of mitogen-activated protein kinases (MAPKs) and nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB). SAS inhibited the mRNA expression and production of interleukin (IL)-6, IL-8 and tumor necrosis factor (TNF). In cells stimulated with PMA plus A23187, SAS suppressed the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 and c-jun N-terminal kinase 1/2 (JNK1/2), but not that of p38 MAPK. SAS suppressed the expression of NF-κB in the nucleus. On the whole, our results suggest that SAS exerts an anti-inflammatory effect by inhibiting the production of pro-inflammatory cytokines through the blocking of the NF-κB and MAPK signaling pathways.

Asunto(s)

Antiinflamatorios/farmacología , Calcimicina/farmacología , Glucósidos/farmacología , Fenoles/farmacología , Ésteres del Forbol/farmacología , Línea Celular Tumoral , Citocinas/genética , Citocinas/metabolismo , Regulación de la Expresión Génica , Humanos , Inflamación/genética , Inflamación/metabolismo , Mediadores de Inflamación/metabolismo , Proteínas Quinasas Activadas por Mitógenos , FN-kappa B/metabolismo , Transducción de Señal/efectos de los fármacos

RESUMEN

Oxyresveratrol (ORV) is a naturally occurring compound found in mulberries that exhibits a wide spectrum of biological activities. However, the underlying mechanism of the action of ORV against the methicillin-resistant S. aureus (MRSA) pathogen has not yet been reported. MRSA is multidrug-resistant, causing skin and other types of infections. The aim of the present study was to examine the antimicrobial activity of ORV and the underlying mechanism of its action on MRSA. The antibacterial activity of ORV was evaluated using a minimum inhibitory concentration (MIC) assay, and the mechanism of its antibacterial action on S. aureus was investigated using a combination of ORV with detergent, ATPase inhibitors and peptidoglycan (PGN). In addition, the survival characteristics and changes in MRSA morphology were monitored using transmission electron microscopy (TEM). The MIC value of ORV against all S. aureus strains was found to be 125 µg/ml. The optical density at 600 nm of each suspension treated using a combination of ORV with Triton X-100, N,N'-dicyclohexylcarbodiimide or sodium azide was reduced by 68.9-89.8% compared with the value upon treatment with ORV alone. In the ORV and PGN combination assay, direct binding of ORV with PGN from S. aureus was evident. Furthermore, TEM examination of MRSA treated with ORV showed alterations in septa formation. In conclusion, these results showed that ORV has a strong antibacterial effect against S. aureus, mainly by increasing membrane permeability and inhibiting ATPase when combined with other drugs.

RESUMEN

The present study aimed to investigate the unknown mechanisms underlying the anti­inflammatory activity of Ciwujianoside C3 (CJS C3), extracted from the leaves of Acanthopanax henryi Harms, on lipopolysaccharide (LPS)­stimulated RAW 264.7 cells. Cells were treated with CJS C3 for 1 h prior to the addition of 200 ng/ml LPS. Cell viability was measured using the MTS assay. Nitric oxide levels were determined by Griess assay. Proinflammatory cytokine production was measured by enzyme­linked immunosorbent assay. The expression levels of cyclooxygenase (COX)­2, inducible nitric oxide synthase (iNOS), and mitogen­activated protein kinases (MAPKs) were investigated by western blotting, reverse transcription (RT)­polymerase chain reaction (PCR) and RT­quantitative PCR. Nuclear factor (NF)­κB/p65 localization, and interaction of the TLR4 receptor with LPS was examined by immunofluorescence assay. The results indicated that CJS C3 exhibited no cytotoxicity at the measured concentrations. Treatment with CJS C3 inhibited NO production, proinflammatory cytokine levels, including interleukin (IL)­6, tumor necrosis factor (TNF)­α, and prostaglandin E2 (PGE2), and protein and mRNA expression levels of iNOS and COX­2. Furthermore, CJS C3 suppressed phosphorylation of extracellular signal­regulated kinases and c­jun N­terminal kinases. It was also able to suppress activation of NF­κB via inhibition of the TLR4 signaling pathway. These results suggested that CJS C3 exerts inhibitory effects on LPS­induced PGE2, NO, IL­6 and TNF­α production. In addition, iNOS and COX­2 expression was decreased in murine macrophages. These inhibitory effects may be achieved via suppression of MAPKs and NF­κB phosphorylation following inhibition of the TLR4 signaling pathway.

Asunto(s)

Antiinflamatorios/farmacología , Lipopolisacáridos/inmunología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Saponinas/farmacología , Animales , Ciclooxigenasa 2/inmunología , Dinoprostona/inmunología , Eleutherococcus/química , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Interleucina-6/inmunología , Ratones , FN-kappa B/inmunología , Óxido Nítrico/inmunología , Óxido Nítrico Sintasa de Tipo II/inmunología , Células RAW 264.7 , Saponinas/química , Receptor Toll-Like 4/inmunología , Factor de Necrosis Tumoral alfa/inmunología

RESUMEN

Methicillin-resistant Staphylococcus aureus (MRSA) infection has become a serious clinical problem worldwide, and alternative natural or combination drug therapies are required for its treatment. The aim of the present study was to examined the antimicrobial activity of luteolin (LUT) against MRSA. Luteolin is a polyphenolic flavonoid compound with a wide spectrum of biological activities. The antimicrobial activities of LUT and the antibiotics ampicillin (AM), oxacillin (OX) and gentamicin (GT), used alone or in combination, were evaluated against five clinical MRSA isolates and two reference strains using a minimum inhibitory concentration (MIC) assay, MTT colorimetric assay, checkerboard dilution test and time-kill assay. The MIC of LUT against all strains was found to be 62.5 µg/ml. The combinations of LUT and antibiotics exhibited a synergistic effect against MRSA in the majority of cases, as determined by the checkerboard method. Time-kill curves revealed that a combination of LUT with AM, OX or GT significantly reduced bacterial counts, which dropped below the lowest detectable limit after 24 h. These results indicate that LUT potentiates the effects of ß-lactam and aminoglycoside antibiotics against MRSA.

RESUMEN

OBJECTIVE: To investigate the gene related to ß-lactam resistance and to confirm the mechanism about a synergy effect between CPZ and ß-lactam antibiotics. METHODS: To measure antibacterial activity, we performed a minimum inhibitory concentration (MIC) and synergy test. Transmission electron microscopy (TEM) was used in morphological analysis. To analyze gene expression, we conducted reverse transcriptase polymerase chain reaction (PCR). RESULTS: We confirmed a synergy effect between CPZ and ß-lactam antibiotics. Furthermore, we observed that CPZ affect the cell envelope of MRSA by using TEM. At the gene level, CPZ reduced the expression of resistance genes. CONCLUSIONS: Through this result, we hypothesize that a decrease of resistance factor expressions was caused by CPZ because it disrupts the activity of a sensor protein located in the cell membrane.

RESUMEN

OBJECTIVE: To investigate the mechanism of antibacterial activity of luteolin (LUT) against methicillin-resistant Staphylococcus aureus (MRSA). METHODS: The mechanism of anti-MRSA activity of LUT was analyzed by the viability assay in membrane permeabilizing agent, ATPase inhibitors, and peptidoglycan (PGN) derived from Staphylococcus aureus (S. aureus). Also, transmission electron microscopy was used to monitor survival characteristics and changes in S. aureus morphology. RESULTS: Compared to the LUT alone, the optical density of suspensions treated with the combination of 125 µg/mL Tris and 250 µg/mL N,N'-dicyclohexylcarbodiimide were reduced to 60% and 46% of the control, respectively. PGN (15.6 µg/mL) gradually impeded the activity of LUT, and PGN (62.5 µg/mL) completely blocked the activity of LUT on S. aureus. CONCLUSIONS: Increased susceptibility to LUT with the Tris-dicyclohexylcarbodiimide combinations is evident in all tested MRSA isolates. The results indicate LUT synergy in increasing cytoplasmic membrane permeability and inhibiting ATPase. S. aureus PGN directly blocks the antibacterial activity of LUT, suggesting the direct binding of LUT with PGN. These findings may be validated for the development of antibacterial agent for low MRSA resistance.

RESUMEN

OBJECTIVE: To investigate the antibacterial activity of SHH extracted with either water or ethanol against methicillin-resistant Staphylococcus aureus (MRSA) and combinatory antimicrobial effect with ciprofloxacin (CIP) by time kill assay and checkerboard dilution test. METHODS: The antibacterial activity determined by broth dilution method indicated that the antibacterial activity of Sami-Hyanglyun-Hwan (SHH) water extract (SHHW) and SHH ethanol extract (SHHE) ranged from 250 to 2000 µg/mL and 125 to 1000 µg/mL against MRSA, respectively. RESULTS: In the checkerboard method, the combinations of SHHE with CIP had a partial synergistic or synergistic effect against MRSA. The time-kill curves showed that a combined SHHE and CIP treatment reduced the bacterial counts dramatically after 24 h. CONCLUSIONS: The present study demonstrates the therapeutic ability of SHHE against MRSA infections.

RESUMEN

Morin, a plant-derived flavonol, is known to be an effective inhibitor of Gram-positive bacteria. In this study, we explored the combined effect of morin with ß-lactam antibiotics against methicillin-resistant Staphylococcus aureus (MRSA), a multidrug-resistant pathogen. The anti-MRSA activity of morin was investigated by the broth microdilution method, checkerboard dilution test, and time-kill curve assay. The expression of the resistant protein, penicillin-binding protein (PBP2a) encoded by mecA, was analyzed by the Western blotting method in the presence of morin and oxacillin. An increased susceptibility of MRSA toward oxacillin was observed in the presence of morin. The protein level of PBP2a was reduced when MRSA (ATCC 33591) was treated with the combination of morin and oxacillin, indicating that the combination of morin and oxacillin potentiates the killing effect against MRSA. The present study indicates that the killing effect by the combinative treatment of morin and ß-lactam antibiotic is dependent on the PBP2a-mediated resistance mechanism.

Asunto(s)

Antibacterianos/farmacología , Proteínas Bacterianas/antagonistas & inhibidores , Farmacorresistencia Bacteriana Múltiple/efectos de los fármacos , Flavonoides/farmacología , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Proteínas de Unión a las Penicilinas/antagonistas & inhibidores , beta-Lactamas/agonistas , Ampicilina/agonistas , Ampicilina/farmacología , Antibacterianos/química , Proteínas Bacterianas/metabolismo , Membrana Celular/efectos de los fármacos , Membrana Celular/ultraestructura , Pared Celular/efectos de los fármacos , Pared Celular/ultraestructura , Recuento de Colonia Microbiana , Citoplasma/efectos de los fármacos , Citoplasma/ultraestructura , Sinergismo Farmacológico , Humanos , Staphylococcus aureus Resistente a Meticilina/crecimiento & desarrollo , Staphylococcus aureus Resistente a Meticilina/aislamiento & purificación , Staphylococcus aureus Resistente a Meticilina/metabolismo , Pruebas de Sensibilidad Microbiana , Viabilidad Microbiana/efectos de los fármacos , Microscopía Electrónica de Transmisión , Oxacilina/agonistas , Oxacilina/farmacología , Proteínas de Unión a las Penicilinas/metabolismo , República de Corea , Infecciones Estafilocócicas/microbiología , beta-Lactamas/farmacología

RESUMEN

With the industrialization of society, the increase in the prevalence of obesity and metabolic disorders has become an important health concern in a number of countries. Quercetin (3,30,40,5,7-pentahydroxyflavone) is well known as a bioactive flavonoid in a variety of biological resources. The aim of the present study was to explore the machanisms responsible for the anti-adipogenic activity of quercetin and its effects on the lipolysis in OP9 mouse stromal cells which rapidly differentiate into adipocytes. The differentiation of OP9 cells into adipocytes was evaluated by the measurement of lipid accumulation by Oil Red O (ORO) staining; lipid accumulation was significantly impaired by treatment with quercetin. Reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis were used to measure the expression levels of CCAAT/enhancer binding protein α (C/EBPα), proliferator-activated receptor γ (PPARγ), sterol regulatory element-binding protein-1 (SREBP-1) and fatty acid synthase (FAS). The mRNA expression levels of lipases, such as adipose triglyceride lipase (ATGL), hormone sensitive lipase (HSL) and lipoprotein lipase (LPL) were also measured by RT-PCR. Quercetin significantly decreased the expression of transcription factors, including C/EBPα, PPARγ and SREBP-1c both at the protein and mRNA level. The results from the present study demonstrate that quercetin prevents adipogenesis by upregulating ATGL and HSL expression and downregulating FAS, LPL and adipocyte fatty acid-binding protein (aP2) expression, as well as the expression of transcription factors. Our data suggest that quercetin has therapeutic potential by regulating the expression of transcriptional factors and enzymes associated with adipogenesis.

Asunto(s)

Adipogénesis/efectos de los fármacos , Lipasa/metabolismo , Quercetina/farmacología , Factores de Transcripción/metabolismo , Animales , Línea Celular , Lipasa/genética , Ratones , Factores de Transcripción/genética

RESUMEN

Human skin is the first line of defense for the protection of the internal organs of the body from different stimuli. Ultraviolet B (UVB), one of the harmful radiations for skin, is widely known to induce abnormally increased cytokine release from keratinocytes leading to inflammatory skin disorders. IL-6 and IL-8 induce an acute-phase response and stimulate leukocyte infiltration in the skin. Previous studies have shown that chronic exposure to UVB radiation increases cyclooxygenase-2 (COX­2) expression through various cell signaling pathways, resulting in skin cancer. Recent studies have shown that the activation of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38 MAPK is strongly correlated with acute inflammation and development of skin cancer caused by an increased expression of COX-2. Ixerisoside A (IXA) is an active constituent of Ixeris dentata of the Compositae (Asteraceae) family. The effect of IXA on skin inflammation has yet to be elucidated. To determine the anti-inflammatory effects of IXA, we examined its effect on UVB-induced pro-inflammatory cytokine production in human keratinocytes (HaCaT cells) by observing these cells in the presence or absence of IXA. In this study, pro-inflammatory cytokine production was determined by enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (rt-pcr), and western blot analysis to evaluate the activation of mitogen-activated protein kinases (MAPKs). IXA inhibited UVB-induced production of the pro-inflammatory cytokines IL-6 and IL-8 in a dose-dependent manner. Moreover, IXA inhibited the expression of COX-2, ERK, JNK, and p38 MAPKs, indicating that the secretion of the pro-inflammatory cytokines IL-6 and IL-8, and COX-2 expression was inhibited by blocking MAPK phosphorylation. These results indicated that IXA potentially protects against UVB-induced skin inflammation.

Asunto(s)

Citocinas/biosíntesis , Mediadores de Inflamación/metabolismo , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Lactonas/farmacología , Sustancias Protectoras/farmacología , Sesquiterpenos/farmacología , Línea Celular , Supervivencia Celular/efectos de los fármacos , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Citocinas/genética , Activación Enzimática/efectos de los fármacos , Expresión Génica , Humanos , Queratinocitos/efectos de la radiación , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Extractos Vegetales/farmacología , Rayos Ultravioleta

RESUMEN

Methicillin-resistant Staphylococcus aureus (MRSA) infection is a serious clinical problem worldwide. The aim of the present study was to examine the antimicrobial activity of oxyresveratrol (ORV) against MRSA. The antimicrobial activity of ORV was evaluated against three strains of MRSA and one methicillin-susceptible S. aureus (MSSA) strain using a minimal inhibitory concentration (MIC) assay, MTT colorimetric assay, checkerboard dilution test and time-kill assay. The MIC of ORV for all strains was moderate at 125 µg/ml. Of note, the antimicrobial activity and fractional inhibitory concentration index values of ORV were markedly increased in the presence of a non-growth inhibitory dose of certain antibiotics. Time-kill curves revealed that a combination of ORV with ciprofloxacin or with gentamicin reduced bacterial counts to below the lowest detectable limit after 24 h. These effective combinations may be used as potential antimicrobial regimens for use in the management of MRSA.

Asunto(s)

Sinergismo Farmacológico , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Extractos Vegetales/administración & dosificación , Infecciones Estafilocócicas/tratamiento farmacológico , Estilbenos/administración & dosificación , Antibióticos Antituberculosos/administración & dosificación , Ciprofloxacina/administración & dosificación , Gentamicinas/administración & dosificación , Humanos , Staphylococcus aureus Resistente a Meticilina/patogenicidad , Infecciones Estafilocócicas/microbiología , Infecciones Estafilocócicas/patología

RESUMEN

Non-alcoholic fatty liver disease (NAFLD) is characterized by the hepatic manifestation of metabolic syndrome and is the leading cause of chronic liver disease. Steatohepatitis plays a critical role in the process resulting in liver fibrosis and cirrhosis. Puerarin is a herbal product widely used in Asia, and is believed to have therapeutic benefits for alleviating the symptoms of steatohepatitis. The present study was designed to investigate the effects and mechanisms of action of puerarin in reducing lipid accumulation in oleic acid (OA)-treated HepG2 cells. Hepatocytes were treated with OA with or without puerarin to observe lipid accumulation by Oil Red O staining. We also examined hepatic lipid contents (e.g., triacylglycerol and cholesterol) following treatment with puerarin. Western blot analysis and reverse transcription-polymerase chain reaction (RT-PCR) were used to measure sterol regulatory element binding protein (SREBP)-1, fatty acid synthase (FAS), peroxisome proliferator-activated receptor α (PPARα) and adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) protein and mRNA expression, respectively. Our results revealed that puerarin suppressed OA-induced lipid accumulation, and reduced the triacylglycerol and cholesterol levels. Furthermore, puerarin decreased the expression levels of lipogenic enzymes, such as FAS and SREBPs, and increased the expression levels of PPARα, which are critical regulators of hepatic lipid metabolism through the AMPK signaling pathway. These results indicate that puerarin has the same ability to activate AMPK, and reduce SREBP-1 and FAS expression, thus inhibiting hepatic lipogenesis and increasing hepatic antioxidant activity. We found that puerarin exerted a regulatory effect on lipid accumulation by decreasing lipogenesis in hepatocytes. Therefore, puerarin extract may have therapeutic benefits in the treatment of fatty liver and lipid-related metabolic disorders.

Asunto(s)

Proteínas Quinasas Activadas por AMP/metabolismo , Hígado Graso/metabolismo , Hígado Graso/patología , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Isoflavonas/farmacología , PPAR alfa/metabolismo , Transducción de Señal/efectos de los fármacos , Hígado Graso/tratamiento farmacológico , Hígado Graso/genética , Regulación de la Expresión Génica/efectos de los fármacos , Células Hep G2 , Humanos , Isoflavonas/administración & dosificación , Isoflavonas/química , Metabolismo de los Lípidos/efectos de los fármacos , Ácido Oléico/farmacología , PPAR alfa/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo

RESUMEN

Curcumin, a natural polyphenolic flavonoid extracted from the rhizome of Curcuma longa L., was shown to possess superior potency to resensitize methicillin-resistant Staphylococcus aureus (MRSA) to antibiotics. Previous studies have shown the synergistic activity of curcumin with ß-lactam and quinolone antibiotics. Further, to understand the anti-MRSA mechanism of curcumin, we investigated the potentiated effect of curcumin by its interaction in diverse conditions. The mechanism of anti-MRSA action of curcumin was analyzed by the viability assay in the presence of detergents, ATPase inhibitors and peptidoglycan (PGN) from S. aureus, and the PBP2a protein level was analyzed by western blotting. The morphological changes in the curcumin-treated MRSA strains were investigated by transmission electron microscopy (TEM). We analyzed increased susceptibility to MRSA isolates in the presence of curcumin. The optical densities at 600 nm (OD600) of the suspensions treated with the combinations of curcumin with triton X-100 and Tris were reduced to 63% and 59%, respectively, compared to curcumin without treatment. N,N'-dicyclohexylcarbodiimide (DCCD) and sodium azide (NaN3) were reduced to 94% and 55%, respectively. When peptidoglycan (PGN) from S. aureus was combined with curcumin, PGN (0-125 µg/mL) gradually blocked the antibacterial activity of curcumin (125 µg/mL); however, at a concentration of 125 µg/mL PGN, it did not completely block curcumin. Curcumin has a significant effect on the protein level of PBP2a. The TEM images of MRSA showed damage of the cell wall, disruption of the cytoplasmic contents, broken cell membrane and cell lysis after the treatment of curcumin. These data indicate a remarkable antibacterial effect of curcumin, with membrane permeability enhancers and ATPase inhibitors, and curcumin did not directly bind to PGN on the cell wall. Further, the antimicrobial action of curcumin involved in the PBP2a-mediated resistance mechanism was investigated.

Asunto(s)

Antiinflamatorios no Esteroideos/farmacología , Curcumina/farmacología , Resistencia a la Meticilina/efectos de los fármacos , Staphylococcus aureus Resistente a Meticilina/metabolismo , Viabilidad Microbiana/efectos de los fármacos

RESUMEN

Tectorigenin (TTR) is an O-methylated isoflavone derived from the rhizome of Belamacanda chinensis (L.) DC. It is known to perform a wide spectrum of biological activities such as antioxidant, anti-inflammatory, anti-tumor. The aim of this study is to examine the mechanism of antibacterial activity of TTR against methicillin-resistant Staphylococcus aureus (MRSA). The anti-MRSA activity of TTR was analyzed in combination assays with detergent, ATPase inhibitors, and peptidoglycan (PGN) derived from S. aureus. Transmission electron microscopy (TEM) was used to monitor survival characteristics and changes in S. aureus morphology. The MIC values of TTR against all the tested strains were 125 µ g/mL. The OD(600) of each suspension treated with a combination of Triton X-100, DCCD, and NaN3 with TTR (1/10 × MIC) had been reduced from 68% to 80%, compared to the TTR alone. At a concentration of 125 µ g/mL, PGN blocked antibacterial activity of TTR. This study indicates that anti-MRSA action of TTR is closely related to cytoplasmic membrane permeability and ABC transporter, and PGN at 125 µ g/mL directly bind to and inhibit TTR at 62.5 µ g/mL. These results can be important indication in study on antimicrobial activity mechanism against multidrug resistant strains.

RESUMEN

Galla rhois is a commonly used traditional medicine for the treatment of pathogenic bacteria in Korea as well as in other parts of Asia. Methyl gallate (MG), a major component of Galla Rhois, exhibits strong antibacterial activity, but its mechanism of action against Salmonella spp. is unclear. In the present study, we investigated the antibacterial actions of MG against Salmonella. The antibacterial activity determined by broth dilution method indicated that the antibacterial activity of MG against Salmonella strains ranged from 3.9 to 125 µg/ml. In vitro bacterial viability test indicated that MG significantly decreased the viability of Salmonella over 40% when combined with ATPase inhibitors. The time-kill curves showed that a combined MG and ATPase inhibitors (DCCD and NaN3) treatment reduced the bacterial counts dramatically after 24 h. Oral administration of MG showed a strong anti-bacterial activity against WS-5 infected BALB/c mice. In contrast to the untreated Salmonella infected control animals, MG treated groups showed no clinical symptoms of the disease, such as lethargy and liver damage. It was observed that MG treatment significantly increased the survival of animals from Salmonella infection, while in untreated groups all animal succumbed to disease by the sixth day post infection. Thus, the present study demonstrates the therapeutic ability of MG against Salmonella infections.

Asunto(s)

Antibacterianos/uso terapéutico , Ácido Gálico/análogos & derivados , Extractos Vegetales/uso terapéutico , Salmonelosis Animal/tratamiento farmacológico , Salmonella/efectos de los fármacos , Administración Oral , Anacardiaceae/química , Animales , Antibacterianos/administración & dosificación , Ácido Gálico/administración & dosificación , Ácido Gálico/química , Ácido Gálico/uso terapéutico , Masculino , Ratones , Ratones Endogámicos BALB C , Extractos Vegetales/administración & dosificación , Extractos Vegetales/química

RESUMEN

Sophoraflavanone B (SPF-B), a prenylated flavonoid, can be isolated from the roots of Desmodium caudatum. The aim of this study was to determine the mechanism of SPF-B's antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA). MRSA is a multidrug-resistant pathogen and the main cause of hospital- and community-acquired infections. The minimum inhibitory concentration (MIC) of SPF-B was assessed using the broth microdilution method. The mechanism of action of SPF-B on S. aureus was analyzed in combination assays incorporating detergents, ATPase inhibitors, and peptidoglycan (PGN) derived from S. aureus. Furthermore, morphological changes in the SPF-B-treated MRSA strains were investigated using transmission electron microscopy. The MIC of SPF-B for MRSA was in the range of 15.6-31.25 µg/mL. The mechanism of action of SPF-B on MRSA was investigated using combination assays with detergent and ATPase inhibitors. The optical density at 600 nm of MRSA suspensions treated with a combination of detergent and SPF-B reduced the MRSA by 63%-73%. In the SPF-B and PGN combination assay, direct binding of SPF-B with PGN from S. aureus was evident. These data may be validated for the development of new antibacterial drugs for low MRSA resistance.

Asunto(s)

Antiinfecciosos/farmacología , Flavanonas/farmacología , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Peptidoglicano/metabolismo , Antiinfecciosos/química , Antiinfecciosos/metabolismo , Pared Celular/efectos de los fármacos , Pared Celular/metabolismo , Pared Celular/ultraestructura , Detergentes/farmacología , Diciclohexilcarbodiimida/farmacología , Flavanonas/química , Flavanonas/metabolismo , Staphylococcus aureus Resistente a Meticilina/metabolismo , Staphylococcus aureus Resistente a Meticilina/ultraestructura , Pruebas de Sensibilidad Microbiana , Microscopía Electrónica de Transmisión

RESUMEN

Carvone is one of the naturally occurring monoterpenes, the largest class of secondary metabolites in plants, and exists in two enantiomers, R-carvone (R-car) and S-car. The objective of this study was to investigate the antimicrobial activity of R-car and S-car with gentamicin (GET) against methicillin-resistant Staphylococcus aureus (MRSA). MRSA is a major human pathogen that causes serious problems, including hospital-acquired pneumonia, abscesses and surgical wound infections. Nosocomial MRSA infections often exhibit multidrug resistance. In the present study, antimicrobial susceptibility testing was performed with R-car, S-car and GET using the broth microdilution method. Minimal inhibitory concentration values for R- and S-car against six different strains of S. aureus ranged between 500 and 1,000 µg/ml. Anti-MRSA activity was evaluated using the checkerboard and time-kill assays to investigate the potential synergistic effects of different combinations of the carvone enantiomers and GET. R-car plus S-car, R-car plus GET and S-car plus GET exhibited significant synergistic activity against MRSA. These findings suggest that the single-agent anti-MRSA activities of R-car, S-car and GET are effectively increased through combination therapy. This study showed that carvone may be a potential adjuvant antimicrobial agent.