RESUMEN
The force-balanced state of magnetically confined plasmas heated up to 100 million degrees Celsius must be sustained long enough to achieve a burning-plasma state, such as in the case of ITER, a fusion reactor that promises a net energy gain. This force balance between the Lorentz force and the pressure gradient force, known as a plasma equilibrium, can be theoretically portrayed together with Maxwell's equations as plasmas are collections of charged particles. Nevertheless, identifying the plasma equilibrium in real time is challenging owing to its free-boundary and ill-posed conditions, which conventionally involves iterative numerical approach with a certain degree of subjective human decisions such as including or excluding certain magnetic measurements to achieve numerical convergence on the solution as well as to avoid unphysical solutions. Here, we introduce GS-DeepNet, which learns plasma equilibria through solely unsupervised learning, without using traditional numerical algorithms. GS-DeepNet includes two neural networks and teaches itself. One neural network generates a possible candidate of an equilibrium following Maxwell's equations and is taught by the other network satisfying the force balance under the equilibrium. Measurements constrain both networks. Our GS-DeepNet achieves reliable equilibria with uncertainties in contrast with existing methods, leading to possible better control of fusion-grade plasmas.
RESUMEN
It is not completely clear whether "the lower, the better" cholesterol hypothesis for cardiovascular disease (CVD) and ischemic heart disease (IHD) can be applied to general populations with a low risk of heart disease mortality.We prospectively followed up 503,340 Koreans who participated in routine health checkups during 2002-2003 until 2013 via linkage to national mortality records.Nonlinear associations with total cholesterol (TC) were found: U-curves for overall CVD (I00-I99; nadir at 180-200âmg/dL) and a reverse-L-curve for IHD (I20-I25). Assuming a linear association in the lower range (<200âmg/dL), TC was inversely associated with CVD mortality (HR per 39âmg/dL [1âmmol/L] increaseâ=â0.90). In the upper range (200-349âmg/dL), TC was positively associated with CVD mortality, largely due to IHD (HRâ=â1.19), especially acute myocardial infarction (HRâ=â1.23). The associations were generally similar in men versus women and in middle-aged (40-64âyears) versus elderly (≥65âyears) adults.TC levels of 180-200âmg/dL were associated with the lowest CVD mortality. Below 200âmg/dL, TC had no graded positive associations with IHD mortality. It remains unclear whether the lowest cholesterol levels are associated with the least mortality from CVD and IHD in Korean adults with a low risk of heart disease.
Asunto(s)
Colesterol/sangre , Isquemia Miocárdica/mortalidad , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Isquemia Miocárdica/sangre , Estudios Prospectivos , República de Corea/epidemiologíaRESUMEN
BACKGROUND: The effect of drugs on ATP-binding cassette transporters, especially permeabilityglycoprotein (P-gp), is an important consideration during new anti-cancer drug development. OBJECTIVE: In this context, the effects of a newly synthesized artemisinin derivative, 10-(4-phenyl-1H-1,2,3- triazol)-artemisinin (5a), were evaluated on P-gp expression and function. METHODS: Reverse transcript polymerase chain reaction and immunoblotting techniques were used to determine the effect of 5a on P-gp expression in LS174T cells. In addition, the ability of 5a to work as either a substrate or an inhibitor of P-gp was investigated through different methods. RESULTS: The results revealed that 5a acts as a novel P-gp inhibitor that dually suppresses the overexpression and function of P-glycoprotein. Co-treatment of LS174T cell line, human colon adenocarcinoma cell line, with 5a and paclitaxel recovered the anticancer effect of paclitaxel by controlling the acquired drug resistance pathway. The overexpression of P-gp induced by rifampin and paclitaxel in a colorectal cell line was suppressed by 5a which could be a novel inhibitory substrate inhibiting the transport of paclitaxel by P-gp. CONCLUSION: The results revealed that 5a can be classified as a type B P-gp inhibitor (with both substrate and inhibitor activities) with an additional function of suppressing P-gp overexpression. The results might be clinically useful in the development of anticancer drugs against cancers with multidrug resistance.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Subfamilia B de Transportador de Casetes de Unión a ATP/antagonistas & inhibidores , Artemisininas/química , Artemisininas/farmacología , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Subfamilia B de Transportador de Casetes de Unión a ATP/metabolismo , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Línea Celular , Supervivencia Celular/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Estructura Molecular , Paclitaxel , ARN Mensajero/metabolismoRESUMEN
The meningococcus carriage rate is age-dependent, with a high prevalence in adolescents and young adults. This cross-sectional study aimed to estimate the oropharyngeal carriage rate of meningococcus among healthy Korean adolescents and its relationship with several population characteristics. The survey was conducted from April to May 2015 among 1,460 first-year high-school students in 9 high schools located in Gyeonggi province, Korea. Each student answered a short questionnaire assessing risk factors for carriage, and posterior pharyngeal wall swab samples were obtained. These samples were cultured on meningococcus-selective media, with colonies resembling meningococci identified using the Vitek® MS system (bioMérieux, Marcy l'Etoile, France). All isolates were characterized by molecular serogrouping and multilocus sequence typing (MLST). Meningococci were identified from 3.4% (49/1,460) swabs. Current smokers had significantly higher carriage rates than non-smokers (8.2% vs. 2.9%, P = 0.002), and boys had significantly higher carriage rates than girls (4.4% vs. 1.6%, P = 0.004). Serogroup B was the most common serogroup, followed by serogroup C, then 29E and Y. Twenty-seven different sequence types (STs) were identified; the most common were ST-3091, ST-11278, and ST-44. These belonged to clonal complexes (CCs) 269, 32, and 41/44, respectively, known as the hypervirulent clones. Evaluating meningococcal carriage is important to understand the epidemiology of meningococcal disease; however, little data exist in Korea. Similar to western countries, meningococcal serogroup B has emerged in Korea, and hypervirulent clones were identified. It is necessary to monitor the genetic and serologic characteristics of circulating meningococci and to assess the potential strain coverage of meningococcal vaccines.
Asunto(s)
Portador Sano/epidemiología , Infecciones Meningocócicas/epidemiología , Neisseria meningitidis/aislamiento & purificación , Adolescente , Portador Sano/diagnóstico , Portador Sano/microbiología , Estudios Transversales , Femenino , Humanos , Masculino , Infecciones Meningocócicas/diagnóstico , Infecciones Meningocócicas/microbiología , Tipificación de Secuencias Multilocus , Neisseria meningitidis/clasificación , Prevalencia , República de Corea/epidemiología , Serogrupo , Serotipificación , Factores Sexuales , Fumar/efectos adversos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Encuestas y CuestionariosRESUMEN
We synthesized a library of curcumin mimics with diverse alkylsulfonyl and substituted benzenesulfonyl modifications through a simple addition reaction of important intermediate, 1-(3-Amino-phenyl)-3-(4-hydroxy-3-methoxy-phenyl)-propenone (10), with various sulfonyl chloride reactants and then tested their vasodilatation effect on depolarization (50 mM K(+))- and endothelin-1 (ET-1)-induced basilar artery contraction. Generally, curcumin mimics with aromatic sulfonyl groups showed stronger vasodilation effect than alkyl sulfonylated curcumin mimics. Among the tested compounds, six curcumin mimics (11g, 11h, 11i, 11j, 11l, and 11s) in a depolarization-induced vasoconstriction and seven compounds (11g, 11h, 11i, 11j, 11l, 11p, and 11s) in an ET-1-induced vasoconstriction showed strong vasodilation effect. Based on their biological properties, synthetic curcumin mimics can act as dual antagonist scaffold of L-type Ca(2+) channel and endothelin A/B2 receptor in vascular smooth muscle cells. In particular, compounds 11g and 11s are promising novel drug candidates to treat hypertension related to the overexpression of L-type Ca(2+) channels and ET peptides/receptors-mediated cardiovascular diseases.
Asunto(s)
Bloqueadores de los Canales de Calcio/farmacología , Curcumina/farmacología , Antagonistas de los Receptores de la Endotelina A/farmacología , Antagonistas de los Receptores de la Endotelina B/farmacología , Animales , Bloqueadores de los Canales de Calcio/síntesis química , Bloqueadores de los Canales de Calcio/química , Canales de Calcio Tipo L/metabolismo , Curcumina/síntesis química , Curcumina/química , Relación Dosis-Respuesta a Droga , Antagonistas de los Receptores de la Endotelina A/síntesis química , Antagonistas de los Receptores de la Endotelina A/química , Antagonistas de los Receptores de la Endotelina B/síntesis química , Antagonistas de los Receptores de la Endotelina B/química , Masculino , Estructura Molecular , Conejos , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Relación Estructura-ActividadRESUMEN
The high frequency of intrinsic resistance to TNF-related apoptosisinducing ligand (TRAIL) in tumor cell lines has necessitated the development of strategies to sensitize tumors to TRAIL-induced apoptosis. We previously showed that elevated pressure applied as a mechanical stressor enhanced TRAIL-mediated apoptosis in human lung carcinoma cells in vitro and in vivo. This study focused on the effect of elevated pressure on the sensitization of TRAIL-resistant cells and the underlying mechanism. We observed elevated pressure-induced sensitization to TRAIL-mediated apoptosis in Hep3B cells, accompanied by the activation of several caspases and the mitochondrial signaling pathway. Interestingly, the enhanced apoptosis induced by elevated pressure was correlated with suppression of extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) phosphorylation and CREB without any change to other MAPKs. Phosphorylation of Bcl-2-associated death promoter (BAD) also decreased, leading to inhibition of the mitochondrial pathway. To confirm whether the activation of pERK1/2 plays a key role in the TRAIL-sensitizing effect of elevated pressure, Hep3B cells were pre-treated with the ERK1/2-specific inhibitor PD98059 instead of elevated pressure. Co-treatment with PD98059 and TRAIL augmented TRAIL-induced apoptosis and decreased BAD phosphorylation. The inhibition of ERK1/2 activation by elevated pressure and PD98059 also reduced BH3 interacting-domain death agonist (BID), thereby amplifying apoptotic stress at the mitochondrial level. Our results suggest that elevated pressure enhances TRAIL-induced apoptosis of Hep3B cells via specific suppression of ERK1/2 activation among MAPKs.
Asunto(s)
Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/patología , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Apoptosis/fisiología , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/metabolismo , Caspasas/metabolismo , Línea Celular Tumoral/efectos de los fármacos , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Resistencia a Antineoplásicos , Flavonoides/farmacología , Humanos , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Proteína Quinasa 1 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 3 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Presión , Inhibidores de Proteínas Quinasas/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/genética , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Proteína Letal Asociada a bcl/metabolismoRESUMEN
2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) can induce drug transporter genes such as the ATP-binding cassette G member 2 (ABCG2), which contributes to multidrug resistance. We investigated the effect of TCDD pretreatment on drug transporters induction from cancer cells of various origins. Cell viabilities after treatment of cisplatin were measured to evaluate acquiring cisplatin resistance by TCDD. Acquring cisplatin resistance was found only in cisplatin senstivie cancer cells including gastric SNU601, colon LS180, brain CRT-MG and lymphoma Jurkat cells which showed a significant increase in cell viability after combined treatment with TCDD and cisplatin. High increase of ABCG2 gene expression was found in SNU601 and LS180 cells with a mild increase in the expression of the ABCC3, ABCC5,and SLC29A2 genes in SNU601 cells, and of major vault protein (MVP) in LS180 cells. The AhR inhibitor kaempferol suppressed the upregulation of ABCG2 expression and reversed the TCDD-induced increase in cell viability in LS180 cells. However, in CRT-MG cells, other transporter genes including ABCC1, ABCC5, ABCA3, ABCA2, ABCB4, ABCG1, and SLC29A1 were up-regulated. These findings suggested the acquiring cisplatin resistance by TCDD associated with cancer cell-type-specific induction of drug transporters.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Cisplatino/farmacología , Proteínas de Neoplasias/metabolismo , Dibenzodioxinas Policloradas/farmacología , Regulación hacia Arriba/efectos de los fármacos , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/genética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Transportador Equilibrativo 2 de Nucleósido/genética , Transportador Equilibrativo 2 de Nucleósido/metabolismo , Humanos , Células Jurkat , Células K562 , Quempferoles/farmacología , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Proteínas de Neoplasias/genética , ARN Mensajero/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Partículas Ribonucleoproteicas en Bóveda/genética , Partículas Ribonucleoproteicas en Bóveda/metabolismoRESUMEN
In vivo and in vitro effects of TIS21 gene on the mature T cell activation and antitumor activities were explored by employing MO5 melanoma orthograft and splenocytes isolated from the TIS21-knockout (KO)(2) mice. Proliferation and survival of mature T cells were significantly increased in the KO than the wild type (WT3)e cells, indicating that TIS21 inhibits the rate of mature T cell proliferation and its survival. In MO5 melanoma orthograft model, the KO mice recruited much more CD8(+) T cells into the tumors at around day 14 after tumor cell injection along with reduced tumor volumes compared with the WT. The increased frequency of granzyme B+ CD8+ T cells in splenocytes of the KO mice compared with the WT may account for antitumor-immunity of TIS21 gene in the melanoma orthograft. In contrast, reduced frequencies of CD107a+ CD8+ T cells in the splenocytes of KO mice may affect the loss of CD8+ T cell infiltration in the orthograft at around day 19. These results indicate that TIS21 exhibits antiproliferative and proapoptotic effects in mature T cells, and differentially affects the frequencies of granzyme B+ CD8+ T-cells and CD107a+ CD8+ T-cells, thus transiently regulating in vivo anti-tumor immunity.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Proliferación Celular , Proteínas Inmediatas-Precoces/fisiología , Melanoma Experimental/inmunología , Receptores de Antígenos de Linfocitos T/metabolismo , Linfocitos T Citotóxicos/inmunología , Proteínas Supresoras de Tumor/fisiología , Animales , Apoptosis , Western Blotting , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Granzimas/genética , Granzimas/metabolismo , Técnicas para Inmunoenzimas , Activación de Linfocitos , Masculino , Melanoma Experimental/metabolismo , Melanoma Experimental/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Antígenos de Linfocitos T/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Bazo/citología , Bazo/inmunología , Bazo/patología , Linfocitos T Citotóxicos/metabolismo , Linfocitos T Citotóxicos/patología , Células Tumorales CultivadasRESUMEN
A newly designed curcumin mimic library (11a-11k) with 2-ethylamino groups in a chalcone structure and variously substituted triazole groups as side chains was synthesized using the Huisgen 1,3-cycloaddition reaction between various alkynes (a-k) and an intermediate (10), with CuSO4 and sodium ascorbate in a solution mixture of chloroform, ethanol, and water (5:3:1) at room temperature for 5h. In the lactate dehydrogenase (LDH) release assay involving co-treatment with tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and/or synthetic curcumin derivatives using TRAIL-resistant human CRT-MG astroglioma cells, the novel curcumin mimic library was found to effectively stimulate the cytotoxicity of TRAIL, causing mild cytotoxicity when administered alone. In particular, 11a and 11j are promising candidates for TRAIL-sensitizers with potential use in combination chemotherapy for brain tumors.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias Encefálicas/tratamiento farmacológico , Curcumina/química , Dietilaminas/química , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Triazoles/química , Protocolos de Quimioterapia Combinada Antineoplásica/síntesis química , Protocolos de Quimioterapia Combinada Antineoplásica/química , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Estructura Molecular , Relación Estructura-Actividad , Ligando Inductor de Apoptosis Relacionado con TNF/síntesis química , Ligando Inductor de Apoptosis Relacionado con TNF/químicaRESUMEN
A specific blocker of L-type Ca(2+) channels may be useful in decreasing arterial tone by reducing the open-state probability of L-type Ca(2+) channels. The aim of the present study was to evaluate the farnesylacetones, which are major active constituents of Sargassum siliquastrum, regarding their vasodilatation efficacies, selectivities toward L-type Ca(2+) channels, and in vivo antihypertensive activities. The application of 5E-(farnesylacetone 311) or 5Z-farnesylacetone (farnesylacetone 312) induced concentration-dependent vasodilatation effects on the basilar artery that was pre-contracted with depolarization and showed an ignorable potential role of endothelial-derived nitric oxide. We also tested farnesylacetone 311 or 312 to determine their pharmacological profiles for the blockade of native L-type Ca(2+) channels in basilar arterial smooth muscle cells (BASMCs) and ventricular myocytes (VMCs), cloned L- (α1C/ß2a/α2δ), N- (α1B/ß1b/α2δ), and T-type Ca(2+) channels (α1G, α1H, and α1I). Farnesylacetone 311 or 312 showed greater selectivity toward the L-type Ca(2+) channels among the tested voltage-gated Ca(2+) channels. The ranked order of the potency for farnesylacetone 311 was cloned α1CâL-type (BASMC)âL-type (VMCs)>α1B>α1H>α1I>α1G and that for farnesylacetone 312 was cloned α1CâL-type (BASMCs)âL-type (VMCs)>α1H>α1G>α1B>α1I. The oral administration of the farnesylacetone 311 (80mg/kg) conferred potent, long-lasting antihypertensive activity in spontaneous hypertensive rats, but it did not alter the heart rate.
Asunto(s)
Bloqueadores de los Canales de Calcio/farmacología , Canales de Calcio Tipo L/efectos de los fármacos , Sargassum/química , Terpenos/farmacología , Administración Oral , Animales , Antihipertensivos/química , Antihipertensivos/aislamiento & purificación , Antihipertensivos/farmacología , Arteria Basilar/efectos de los fármacos , Arteria Basilar/metabolismo , Tiempo de Circulación Sanguínea , Bloqueadores de los Canales de Calcio/química , Bloqueadores de los Canales de Calcio/aislamiento & purificación , Canales de Calcio Tipo L/metabolismo , Relación Dosis-Respuesta a Droga , Células HEK293 , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Hipertensión/tratamiento farmacológico , Masculino , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Óxido Nítrico/metabolismo , Conejos , Ratas , Ratas Endogámicas SHR , Ratas Sprague-Dawley , Terpenos/química , Terpenos/aislamiento & purificación , Vasodilatación/efectos de los fármacosRESUMEN
Protein arginine methyltransferases (PRMTs) generate asymmetric and symmetric dimethyl-arginines by catalyzing the transfer of methyl groups from S: -adenosyl-L-methionine to arginines in target proteins. Previously, we observed that the expression and activity of PRMTs were significantly down-regulated in replicatively senescent fibroblasts compared to young fibroblasts. In this study, we determined the level of three PRMT family members (PRMT1, PRMT4, and PRMT5) and the arginine methylation status in eight tissues from 6- and 24-month-old rats. We observed tissue-specific down-regulation of individual PRMT members in testis, thymus, kidney, lung, and heart from 24-month-old as compared to 6-month-old rats. Specifically, we observed reduced levels of PRMT1 in thymus and lung, reduced levels of PRMT4 in testis, thymus, and hearts, and reduced levels of PRMT5 in all five tissues. PRMT enzyme activity on histones generally correlated with PRMT expression. Furthermore, we observed a reduction in asymmetric and symmetric dimethylation on proteins in aged thymus and lung, and a reduction in symmetric dimethylation in aged testes relative to the testes harvested from young rats. These results suggest that individual PRMT proteins have tissue-specific functions and are regulated in a tissue-specific and age-dependent manner.
Asunto(s)
Envejecimiento/metabolismo , Proteína-Arginina N-Metiltransferasas/metabolismo , Animales , Western Blotting , Electroforesis en Gel de Poliacrilamida , Masculino , Ratas , Ratas Sprague-Dawley , Distribución TisularRESUMEN
Naegleria fowleri is a pathogenic free-living amoeba which causes primary amoebic meningoencephalitis in humans and experimental animals. To investigate the mechanisms of such inflammatory diseases, potential chemokine gene activation in human astroglial cells was investigated following treatment with N. fowleri lysates. We demonstrated that N. fowleri are potent inducers for the expression of interleukin-8 (IL-8) genes in human astroglial cells which was preceded by activation of extracellular signal-regulated kinase (ERK). In addition, N. fowleri lysates induces the DNA binding activity of activator protein-1 (AP-1), an important transcription factor for IL-8 induction. The specific mitogen-activated protein kinase kinase/ERK inhibitor, U0126, blocks N. fowleri-mediated AP-1 activation and subsequent IL-8 induction. N. fowleri-induced IL-8 expression requires activation of ERK in human astroglial cells. These findings indicate that treatment of N. fowleri on human astroglial cells leads to the activation of AP-1 and subsequent expression of IL-8 which are dependent on ERK activation. These results may help understand the N. fowleri-mediated upregulation of chemokine and cytokine expression in the astroglial cells.
Asunto(s)
Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Interleucina-8/metabolismo , Naegleria fowleri/química , Animales , Butadienos , Línea Celular , Ensayo de Inmunoadsorción Enzimática , Quinasas MAP Reguladas por Señal Extracelular/genética , Humanos , Interleucina-8/genética , Nitrilos , Factor de Transcripción AP-1/genética , Factor de Transcripción AP-1/metabolismoRESUMEN
Protein-arginine methylation is one of the modifications that yields mono and dimethyl (asymmetric or symmetric) arginine residues in proteins. Previously, we found that asymmetric arginine methylation is decreased proportionately with a decrease of cell proliferation potential of cells, and such arginine methylation is greatest in immortalized cells, followed by normal young cells, and lowest in replicatively senescent cells. Using an asymmetric dimethyl-arginine-specific antibody, we identified arginine-methylated proteins in these cell types by immunoprecipitation and 2-D immunoblotting followed by MS. As a result, arginine methylation of chaperone molecules and RNA-binding proteins was differentially regulated between immortalized or young cells and senescent cells. Immortalized cells had significantly higher levels of methyl-accepting proteins, such as cleavage stimulation factor 2 (CstF2) and heterogeneous nuclear ribonucleoprotein (hnRNP) R, than young cells. However, senescent cells contained hypomethylated CstF2, hnRNP K, and chaperone containing TCP1 subunit 7, as well as decreased hnRNP R level. Further, significant reduction of arginine modification in CstF2 and chaperone containing TCP1 subunit 7 was observed in prematurely senescent fibroblasts, induced by treatment with adenosine dialdehyde, a transmethylation inhibitor, or subcytotoxic concentration of H(2)O(2). These results suggest that asymmetric modification of RNA-binding proteins and molecular chaperones plays an essential role in maintaining cell proliferation capability.
Asunto(s)
Arginina/metabolismo , Senescencia Celular/fisiología , Chaperonas Moleculares/análisis , Proteína-Arginina N-Metiltransferasas/metabolismo , Proteínas de Unión al ARN/análisis , Adenosina/análogos & derivados , Adenosina/farmacología , Western Blotting , Línea Celular Transformada/metabolismo , Proliferación Celular , Chaperonina con TCP-1/metabolismo , Factor de Estimulación del Desdoblamiento , Humanos , Peróxido de Hidrógeno/farmacología , Inmunoprecipitación , Metilación/efectos de los fármacos , Chaperonas Moleculares/metabolismo , Proteínas de Unión al ARN/metabolismoRESUMEN
TNF-related apoptosis-inducing ligand (TRAIL, Apo2L) is a promising anticancer agent with high specificity for cancer cells. Many strategies have been proposed to enhance the sensitivity of cancer cells to TRAIL-mediated apoptosis, including the use of combination treatment with conventional cancer therapies. However, few reports have evaluated the effects of TRAIL in combination with mechanical stress, which can also cause apoptosis of cancer cells. In the present study, we describe a custom-designed culture system that delivers two atmospheres of elevated pressure (EP) by using compressed air, and which enhances the sensitivity of cancer cells to TRAIL-mediated apoptosis. The combination of TRAIL and EP significantly increased apoptosis of human H460 lung cancer cells more than hyperbaric normoxia or normobaric mild hyperoxia. EP-potentiating TRAIL-mediated apoptosis of H460 cells was accompanied by up-regulated death receptor 5 (DR5), activation of caspases, decreased mitochondrial membrane potential, and reactive oxygen species production. We also observed EP-induced sensitization of TRAIL-mediated apoptosis in other cancer cell types. In contrast, human normal cells showed no DNA damage or cell death when exposed to the combined treatment. In a chicken chorioallantoic membrane model, EP enhanced TRAIL-mediated apoptosis of tumors that developed from transplanted H460 cells. Collectively, EP enhanced TRAIL-induced apoptosis of human lung carcinoma cells in vitro and in vivo. These findings suggest that EP is a mechanical and physiological stimulus that might have utility as a sensitizing tool for cancer therapy.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Proteínas Recombinantes , Ligando Inductor de Apoptosis Relacionado con TNF , Animales , Antineoplásicos/uso terapéutico , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Caspasas/metabolismo , Línea Celular Tumoral , Embrión de Pollo , Terapia Combinada , Femenino , Feto , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Potencial de la Membrana Mitocondrial/fisiología , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/uso terapéutico , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Estrés Mecánico , Ligando Inductor de Apoptosis Relacionado con TNF/genética , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/uso terapéuticoRESUMEN
Intensive cancer therapy strategies have thus far focused on sensitizing cancer cells to anticancer drug-mediated apoptosis to overcome drug resistance, and this strategy has led to more effective cancer therapeutics. Cisplatin (cis-diamminedichloroplatinum(II), CDDP) is an effective anticancer drug used to treat many types of cancer, including non-small cell lung carcinoma (NSCLC), and can be used in combination with various chemicals to enhance cancer cell apoptosis. Here, we introduce the use of elevated pressure (EP) in combination with CDDP for cancer treatment and explore the effects of EP on CDDP-mediated apoptosis in NSCLC cells. Our findings demonstrate that preconditioning NSCLC cells with EP sensitizes cells for CDDP-induced apoptosis. Enhanced apoptosis was dependent on p53 and HO-1 expression, and was associated with increased DNA damage and down-regulation of genes involved in nucleotide excision repair. The transcriptional levels of transporter proteins indicated that the mechanism by which EP-induced CDDP sensitization was intracellular drug accumulation. The protein levels of some antioxidants, such as hemeoxygenase-1 (HO-1), glutathione (GSH) and glutathione peroxidase (Gpx), were decreased in A549 cells exposed to EP via the down-regulation of the transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf-2). Furthermore, normal human fibroblasts were resistant to EP treatment, with no elevated DNA damage or apoptosis. Collectively, these data show that administration of EP is a potential adjuvant tool for CDDP-based chemosensitivity of lung cancer cells that may reduce drug resistance.
Asunto(s)
Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/terapia , Cisplatino/uso terapéutico , Resistencia a Antineoplásicos , Neoplasias Pulmonares/terapia , Presión , Estrés Mecánico , Apoptosis , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Línea Celular Tumoral , Daño del ADN , Reparación del ADN/genética , Hemo-Oxigenasa 1/antagonistas & inhibidores , Hemo-Oxigenasa 1/genética , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Factor 2 Relacionado con NF-E2/antagonistas & inhibidoresRESUMEN
Matrix metalloproteinase-9 (MMP-9) is an important angiogenic and prognostic factor in malignant tumors. Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is known as the death ligand, which induces preferential apoptosis of transformed tumor cells. In this study, we investigated the biological functions of TRAIL, other than its role in induction of apoptosis. We demonstrated that TRAIL induces MMP-9 expression in human astrocytoma cells, which is preceded by activation of extracellular signal-regulated protein kinase (ERK). In addition, TRAIL induces the DNA-binding activity of NF-kappaB, an important transcription factor for MMP-9 induction. The specific MEK inhibitor, U0126, significantly blocks TRAIL-mediated NF-kappaB activation and subsequent MMP-9 induction. These findings indicate that TRAIL treatment in human astrocytoma cells leads to the activation of NF-kappaB and subsequent expression of MMP-9, which are dependent on ERK activation. Collectively, these results suggest that TRAIL has alternative biological functions in addition to its role in inducing apoptosis in human malignant astrocytoma cells.
Asunto(s)
Astrocitoma/enzimología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Metaloproteinasa 9 de la Matriz/biosíntesis , Ligando Inductor de Apoptosis Relacionado con TNF/fisiología , Butadienos/farmacología , Línea Celular Tumoral , Activación Enzimática , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Humanos , Metaloproteinasa 9 de la Matriz/genética , FN-kappa B/metabolismo , Nitrilos/farmacología , Inhibidores de Proteínas Quinasas , Ligando Inductor de Apoptosis Relacionado con TNF/farmacologíaRESUMEN
Inhalation of freshwater containing the free-living amoeba Naegleria fowleri leads to a potentially fatal infection known as primary amoebic meningoencephalitis (PAME). Amphotericin B is the only agent with clinical efficacy in the treatment of PAME in humans, however this drug is often associated with adverse effects on the kidney and other organs. In an attempt to select other useful therapeutic agents for treating PAME, the amoebicidal activities of antibacterial agents including clarithromycin, erythromycin, hygromycin B, neomycin, rokitamycin, roxithromycin and zeocin were examined. Results showed that the growth of amoeba was effectively inhibited by treatment with hygromycin B, rokitamycin and roxithromycin. Notably, when N. fowleri trophozoites were treated with rokitamycin, the minimal inhibitory concentration was 6.25 microg/mL on Day 2. In the treatment of experimental meningoencephalitis due to N. fowleri, survival rates of mice treated with roxithromycin and rokitamycin were 25% and 80%, respectively, over 1 month. The mean time to death for roxithromycin and rokitamycin treatment was 16.2 days and 16.8 days, respectively, compared with 11.2 days for control mice. Finally, rokitamycin showed both in vitro and in vivo therapeutic efficacy against N. fowleri and may be a candidate drug for the treatment of PAME.
Asunto(s)
Amebiasis/tratamiento farmacológico , Amebicidas/uso terapéutico , Infecciones Protozoarias del Sistema Nervioso Central/tratamiento farmacológico , Miocamicina/análogos & derivados , Naegleria fowleri , Amebiasis/microbiología , Amebicidas/farmacología , Animales , Antibacterianos/uso terapéutico , Nitrógeno de la Urea Sanguínea , Infecciones Protozoarias del Sistema Nervioso Central/microbiología , Femenino , Riñón/microbiología , Riñón/patología , L-Lactato Deshidrogenasa/metabolismo , Hígado/microbiología , Hígado/patología , Ratones , Ratones Endogámicos BALB C , Pruebas de Sensibilidad Microbiana , Miocamicina/farmacología , Miocamicina/uso terapéutico , Naegleria fowleri/efectos de los fármacos , Análisis de SupervivenciaRESUMEN
Naegleria fowleri is a ubiquitous, pathogenic free-living amoeba; it is the most virulent Naegleria species and causes primary amoebic meningoencephalitis (PAME) in laboratory animals and humans. Although amphotericin B is currently the only agent available for the treatment of PAME, it is a very toxic antibiotic and may cause many adverse effects on other organs. In order to find other potentially therapeutic agents for N. fowleri infection, the present study was undertaken to evaluate the in vitro and in vivo efficacies of miltefosine and chlorpromazine against pathogenic N. fowleri. The result showed that the growth of the amoeba was effectively inhibited by treatment with amphotericin B, miltefosine, and chlorpromazine. When N. fowleri trophozoites were treated with amphotericin B, miltefosine, and chlorpromazine, the MICs of the drug were 0.78, 25, and 12.5 microg/ml, respectively, on day 2. In experimental meningoencephalitis of mice that is caused by N. fowleri, the survival rates of mice treated with amphotericin B, miltefosine, and chlorpromazine were 40, 55, and 75%, respectively, during 1 month. The average mean time to death for the amphotericin B, miltefosine, and chlorpromazine treatments was 17.9 days. In this study, the effect of drugs was found to be optimal when 20 mg/kg was administered three times on days 3, 7, and 11. Finally, chlorpromazine had the best therapeutic activity against N. fowleri in vitro and in vivo. Therefore, it may be a more useful therapeutic agent for the treatment of PAME than amphotericin B.
Asunto(s)
Amebiasis/tratamiento farmacológico , Amebicidas/farmacología , Infecciones Protozoarias del Sistema Nervioso Central/tratamiento farmacológico , Clorpromazina/farmacología , Naegleria fowleri/efectos de los fármacos , Fosforilcolina/análogos & derivados , Amebiasis/parasitología , Amebiasis/patología , Anfotericina B/farmacología , Animales , Infecciones Protozoarias del Sistema Nervioso Central/parasitología , Infecciones Protozoarias del Sistema Nervioso Central/patología , Femenino , Técnicas In Vitro , Ratones , Ratones Endogámicos BALB C , Naegleria fowleri/patogenicidad , Pruebas de Sensibilidad Parasitaria , Fosforilcolina/farmacologíaRESUMEN
The central nervous system (CNS) is particularly vulnerable to reactive oxygen species (ROS), which have been implicated in the pathogenesis of various neurological disorders. The TNF superfamily of cytokines, especially tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), induces caspase-dependent cell death and is also implicated in various neurodegenerative diseases. In this study, we investigated the relationship between ROS and TRAIL-induced cell death. Exposure to hydrogen peroxide (H(2)O(2)) (100 microM) sensitized human astrocytic cells to TRAIL-induced cell death (up to 7-fold induction). To delineate the molecular mechanisms responsible for H(2)O(2)-induced sensitization, we examined expression of various genes (Caspase-8, Fas, FasL, DR4, DR5, DcR1, DcR2, TRAIL, TNFRp55) related to TRAIL-induced cell death. Treatment with H(2)O(2) significantly increased DR5 mRNA and protein expression in a time- and dose-dependent manner. H(2)O(2)-mediated cell death was blocked upon treatment with DR5:Fc protein, a TRAIL-specific antagonistic protein. These findings collectively suggest that oxidative stress sensitizes human astroglial cells to TRAIL-induced cell death through up-regulation of DR5 expression.
Asunto(s)
Apoptosis , Astrocitos/metabolismo , Estrés Oxidativo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Apoptosis/genética , Astrocitos/citología , Astrocitos/efectos de los fármacos , Células Cultivadas , Regulación de la Expresión Génica , Humanos , Peróxido de Hidrógeno/metabolismo , Peróxido de Hidrógeno/toxicidad , ARN Mensajero/metabolismo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/genética , Regulación hacia ArribaRESUMEN
A gene encoding a cytosolic heat shock protein 70 from pathogenic Naegleria fowleri (Nf-cHSP70) was identified. The Nf-cHSP70 was 2,062 bp in length with an open reading frame of 1,980 bp encoding 659 amino acid residues. The deduced amino acid sequence of the gene shared high sequence identities with HSP70s from other parasitic organisms and mammals. The characteristic domains, including N-terminal ATPase domain, calmodulin-binding domain, and EE(D)VD motif, found in HSP70s were also well conserved in this gene. The recombinant Nf-cHSP70 protein showed strong antigenicity against the sera from mice experimentally infected with N. fowleri. Immunofluorescence assay showed that Nf-cHSP70 localized in cytosol of the parasite. The results from semi-quantitative RT-PCR and Western blot analyses demonstrated the expression levels of gene transcripts, and its products were significantly increased at high temperature (42 degrees C). The definitive biological roles of Nf-cHSP70 are not clear, but it may protect the parasite under environmental changes especially high temperature.