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Here, we report a bienzymatic cascade to produce ß-amino acids as an intermediate for the synthesis of the leading oral antidiabetic drug, sitagliptin. A whole-cell biotransformation using recombinant Escherichia coli coexpressing a esterase and transaminase were developed, wherein the desired expression level of each enzyme was achieved by promotor engineering. The small-scale reactions (30 ml) performed under optimized conditions at varying amounts of substrate (100-300 mM) resulted in excellent conversions of 82%-95% for the desired product. Finally, a kilogram-scale enzymatic reaction (250 mM substrate, 220 L) was carried out to produce ß-amino acid (229 mM). Sitagliptin phosphate was chemically synthesized from ß-amino acids with 82% yield and > 99% purity.
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Escherichia coli , Esterasas , Ingeniería Genética , Microorganismos Modificados Genéticamente , Regiones Promotoras Genéticas , Fosfato de Sitagliptina/metabolismo , Transaminasas , Escherichia coli/genética , Escherichia coli/metabolismo , Esterasas/genética , Esterasas/metabolismo , Microorganismos Modificados Genéticamente/genética , Microorganismos Modificados Genéticamente/metabolismo , Transaminasas/genética , Transaminasas/metabolismoRESUMEN
In this study, vertical nanowire arrays of MoO(3-x) grown on metallic substrates with diameters of ~90 nm show high-capacity retention of ~630 mAhg(-1) for up to 20 cycles at 50 mAg(-1) current density. Particularly, they exhibit a capacity retention of ~500 mAhg(-1) in the voltage window of 0.7-0.1 V, much higher than the theoretical capacity of graphite. In addition, 10 nm Si-coated MoO(3-x) nanowire arrays have shown a capacity retention of ~780 mAhg(-1), indicating that hybrid materials are the next generation materials for lithium ion batteries.
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STUDY DESIGN: Institutional review board-approved retrospective cohort study. OBJECTIVES: Failure to achieve alignment goals may result in accelerated adjacent segment degeneration and poorer outcomes. In "open" spine surgery, intraoperative tools can fine tune alignment; minimally invasive spine surgery techniques may not allow for this type of intraoperative adjustment. The aim of this study was to identify pre-operative radiographic parameters that accurately predict post-operative alignment after minimally invasive lumbar spine surgery. We hypothesized that pre-operative supine and extension sagittal alignment would predict post-operative standing alignment. METHODS: 50 consecutive patients underwent lateral or anterior lumbar interbody fusion with or without percutaneous posterior instrumentation by a single-surgeon. Sagittal alignment parameters were evaluated on pre-operative standing scoliosis radiographs, dynamic radiographs, supine CT scout, and 6-week post-operative standing radiographs. Demographic and perioperative data were analyzed. RESULTS: The mean age was 67.8 years. The mean BMI was 29.7. On average, 3 levels were instrumented (range, 2-6). Surgical time was 4.5 ± 1.8 hours. Following surgery, global lordosis increased from 44.7 ± 17° to 48.6 ± 16° (P = .001). However, there was no significant difference between the pre-operative supine (48.5 ± 15°), pre-operative extension (49.2 ± 18°), or 6-week post-operative standing radiographs (48.6 ± 16°). There were strong correlations between post-operative alignment and pre-operative supine (r = .825) and extension (r = .851) alignment. CONCLUSIONS: Our results suggest that pre-operative supine and extension radiographs could be a gold standard for minimally invasive lumbar spine surgery alignment correction as they predict post-operative alignment. The extension alignment was the strongest predictor of post-operative alignment.
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BACKGROUND AND AIM: Gastric cancers present with distinctive carcinogenesis pathways that vary with the mucin phenotypes. We attempted to elucidate the relations between the characteristics of the mucin phenotypes of gastric cancer and the tumor invasiveness. METHODS: Gastric adenocarcinomas that were resected surgically between August 2005 and April 2007 were included in the present study. The gastric cancers were subclassified into gastric and intestinal mucin phenotypes if more than 10% of cancer cells exhibited gastric (MUC5AC and/or MUC6) and intestinal (MUC2 or CD10) markers, respectively. RESULTS: The mucin phenotypes of 123 gastric cancers were gastric (n = 31), intestinal (n = 43), mixed (n = 28) and unclassified (n = 21). The mucin phenotype was related to histological type (P < 0.001), Lauren's classification (P = 0.001) and size (P = 0.014) of the gastric adenocarcinoma, but not to its location or to the presence of Helicobacter pylori infection. The unclassified mucin phenotype exhibited the largest number of lymph node metastases (P = 0.007), lymphatic invasions (P < 0.001) and neural invasions (P = 0.026), whereas the intestinal mucin phenotype exhibited the lowest invasiveness. CONCLUSION: The mucin phenotype reflects the biological behavior of gastric cancer, with the intestinal and unclassified mucin phenotypes exhibiting the lowest and highest invasiveness, respectively.
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Adenocarcinoma/química , Mucinas/análisis , Neoplasias Gástricas/química , Adenocarcinoma/genética , Adenocarcinoma/microbiología , Adenocarcinoma/patología , Adulto , Anciano , Diferenciación Celular , Femenino , Regulación Neoplásica de la Expresión Génica , Helicobacter pylori/aislamiento & purificación , Humanos , Inmunohistoquímica , Metástasis Linfática , Masculino , Inestabilidad de Microsatélites , Persona de Mediana Edad , Mucina 5AC/análisis , Mucina 2/análisis , Mucina 6/análisis , Invasividad Neoplásica , Neprilisina/análisis , Fenotipo , Neoplasias Gástricas/genética , Neoplasias Gástricas/microbiología , Neoplasias Gástricas/patologíaAsunto(s)
Biomarcadores/metabolismo , Hígado Graso/diagnóstico , Modelos Biológicos , Adulto , Factores de Edad , Alanina Transaminasa/metabolismo , Consumo de Bebidas Alcohólicas , Aspartato Aminotransferasas/metabolismo , Biomarcadores/sangre , Glucemia , Presión Sanguínea , Índice de Masa Corporal , Proteína C-Reactiva/metabolismo , Femenino , Hemoglobinas/análisis , Humanos , Lipoproteínas/sangre , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Curva ROC , República de Corea , Estudios Retrospectivos , Factores de Riesgo , Factores Sexuales , Fumar , Triglicéridos/sangre , gamma-Glutamiltransferasa/metabolismoRESUMEN
Depression has been strongly associated with poor HIV treatment adherence, but little research has explored how individuals manage to follow their regimens despite symptoms of major depression. Using a sample of antiretroviral therapy patients with clinically significant depressive symptoms (n = 84), we examined whether patients with optimal adherence differed from those with suboptimal adherence in terms of the types of depressive symptoms experienced and treatment self-efficacy. There were no significant differences between participants with regard to types of depressive symptoms. Findings indicate that patients with high treatment self-efficacy were more likely to report optimal levels of adherence than patients with low self-efficacy.
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Terapia Antirretroviral Altamente Activa/psicología , Depresión/psicología , Infecciones por VIH/psicología , Cumplimiento de la Medicación/psicología , Autoeficacia , Adulto , Femenino , Infecciones por VIH/tratamiento farmacológico , Humanos , Masculino , Persona de Mediana EdadRESUMEN
A panel of 17 tetraploid and 11 diploid potato genotypes was screened by comparative sequence analysis of polymerase chain reaction (PCR) products for single nucleotide polymorphisms (SNPs) and insertion-deletion polymorphisms (InDels), in regions of the potato genome where genes for qualitative and/or quantitative resistance to different pathogens have been localized. Most SNP and InDel markers were derived from bacterial artificial chromosome (BAC) insertions that contain sequences similar to the family of plant genes for pathogen resistance having nucleotide-binding-site and leucine-rich-repeat domains (NBS-LRR-type genes). Forty-four such NBS-LRR-type genes containing BAC-insertions were mapped to 14 loci, which tag most known resistance quantitative trait loci (QTL) in potato. Resistance QTL not linked to known resistance-gene-like (RGL) sequences were tagged with other markers. In total, 78 genomic DNA fragments with an overall length of 31 kb were comparatively sequenced in the panel of 28 genotypes. 1498 SNPs and 127 InDels were identified, which corresponded, on average, to one SNP every 21 base pairs and one InDel every 243 base pairs. The nucleotide diversity of the tetraploid genotypes (pi = 0.72 x 10(-3)) was lower when compared with diploid genotypes (pi = 2.31 x 10(-3)). RGL sequences showed higher nucleotide diversity when compared with other sequences, suggesting evolution by divergent selection. Information on sequences, sequence similarities, SNPs and InDels is provided in a database that can be queried via the Internet.
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PURPOSE: The Leber congenital amaurosis (LCA) protein AIPL1 is present only in the rod photoreceptors of the adult human retina and is excluded from the cone photoreceptors. LCA, however, is characterized by an absence of both rod and cone function at birth or shortly thereafter. Therefore, this study was conducted to determine whether AIPL1 is present in the rod and cone photoreceptors of the developing human retina. In addition, the expression of NUB1, a putative AIPL1-interacting partner, was examined. METHODS: A comprehensive spatiotemporal examination of AIPL1 distribution during development was performed by immunohistochemistry, using a previously characterized AIPL1 anti-serum. Immunofluorescence confocal microscopy was used to examine the coexpression of AIPL1 with the long/medium (L/M) and short (S) wavelength-sensitive cone photoreceptors in the developing human retina. The spatiotemporal distribution of NUB1 was also examined by immunohistochemistry, using a newly developed anti-serum to the C terminus of NUB1. RESULTS: AIPL1 protein was detected by 11.8 fetal weeks in the central fetal human retina. With continued development, AIPL1 expression spread gradually toward peripheral retina. AIPL1 was expressed in the L/M and S cone photoreceptors in addition to the rods of the developing human retina. NUB1 was localized in cell nuclei throughout the human fetal and adult eye at all time points. CONCLUSIONS: The pattern of AIPL1 expression closely follows the centroperipheral gradient in photoreceptor development. The data suggest that AIPL1 is essential for the normal development of both rod and cone photoreceptor cells and that mutations in the AIPL1 gene cause the death or dysfunction of photoreceptors early in development resulting in blindness or severely impaired vision at birth.
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Células Fotorreceptoras Retinianas Conos/embriología , Células Fotorreceptoras Retinianas Bastones/embriología , Proteínas Adaptadoras Transductoras de Señales , Anciano , Western Blotting , Proteínas Portadoras , Desarrollo Embrionario y Fetal , Proteínas del Ojo , Edad Gestacional , Humanos , Inmunohistoquímica , Microscopía Confocal , Microscopía Fluorescente , Factores de Transcripción/metabolismoRESUMEN
First discovered as a circulating secreted molecule expressed by activated T lymphocytes, attractin was examined as a potential marker of immune activity. The discovery that a transmembrane form not only controls neuropeptide regulation of hair pigmentation in animals but also affects basal metabolism led to proposals that attractin may also be an extracellular target amenable for the development of obesity-regulating drugs. Examination of several animal mutants used as models ofjuvenile-onset neurodegeneration revealed mutations at the attractin locus as the cause, and the reassessment of earlier attractin mutants demonstrated that neurodegeneration, alterations in pigmentation regulation, and basal metabolic rate were common to all the allelic variants. The presentation and severity of the symptoms differ depending upon the mutation, and some may be variably penetrant even within an allelic line. In this report, we review our rapidly altering perception of the functional activity of attractin with each further addition to its sphere of physiological involvement. We further reappraise our concepts of the subcellular location of attractin, leading to a new proposal that provides a unifying mechanism for attractin's pleiotropic activities. Progress in elucidating each new aspect of attractin function provides a case study in the evolution of possible therapeutic interventions as well as illuminating some of the pitfalls of studying molecular pathways isolated from the whole organism physiology.
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Dipeptidil Peptidasa 4/fisiología , Proteínas de la Membrana/fisiología , Obesidad , Terapias en Investigación , Animales , Humanos , Melanocitos/metabolismo , Mutación , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/metabolismo , Neuronas/metabolismo , Obesidad/etiología , Obesidad/metabolismo , Obesidad/prevención & control , Linfocitos T/metabolismoRESUMEN
Conjugated linoleic acid (CLA) inclusion complexes with alpha-cyclodextrin (alpha-CD), beta-cyclodextrin (beta-CD), and gamma-cyclodextrin (gamma-CD) (designated CLA/CDs inclusion complexes) were prepared to determine the mole ratio of CLA complexed with CDs and the oxidative stability of CLA in the CLA/CDs inclusion complexes. When measured by GC, (1)H NMR, and T(1) value analyses, 1 mole of CLA was complexed with 5 mol of alpha-CD, 4 mol of beta-CD, and 2 mol of gamma-CD. The oxidation of CLA induced at 35 degrees C for 80 h was completely prevented by the formation of CLA/CDs inclusion complexes.
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Ciclodextrinas/química , Ácido Linoleico/química , alfa-Ciclodextrinas , beta-Ciclodextrinas , gamma-Ciclodextrinas , Cromatografía de Gases , Estabilidad de Medicamentos , Espectroscopía de Resonancia Magnética , Oxidación-ReducciónRESUMEN
Mixtures of t,t conjugated linoleic acid methylester (t,t CLA-Me) isomers were prepared from synthetic CLA, consisting of 47.8% t10,c12 CLA; 45.5% c9,t11 CLA; 2.0% t,t CLA; and 4.7% others, by methylation with BF(3)/methanol (designated TT-TC/CT) in conjunction with purification at -68 degrees C for 24 h. The amount or composition of the TT-TC/CT was greatly affected by the concentration of BF(3) in methanol and the duration of methylation. The methylation of 50 mg of synthetic CLA for 30 min with 1 mL of 7.0% BF(3)/methanol produced a TT-TC/CT (21.54 mg) with the composition of 1.3% t12,t14; 5.9% t11,t13; 42.7% t10,t12; 44.0% t9,t11; 5.0% t8,t10; and 1.1% t 7,t9 CLA, whereas the methylation for 60 min with 14.0% BF(3)/methanol produced a TT-TC/CT (28.62 mg) with the composition of t,t CLA isomers different from that of TT-TC/CT by methylation for 30 min with 7.0% BF(3)/methanol. A large quantity of TT-TC/CT (14.15 g) with the composition similar to that of TT-TC/CT prepared from 50 mg of synthetic CLA was also prepared from 25 g of synthetic CLA. The purity of TT-TC/CT samples was greater than 98%. These results suggest that TT-TC/CT with a purity greater than 98% was easily prepared from synthetic CLA by BF(3)-catalyzed methylation, and the amount and composition of t,t CLA isomers of TT-TC/CT samples could be controlled by methylation conditions.
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Boratos/química , Ácidos Linoleicos/síntesis química , Metanol/química , Isomerismo , Ácidos Linoleicos/química , Metilación , Factores de TiempoRESUMEN
A newly fabricated plasma-polymerized poly(ethylene glycol) (PP-PEG) film shows extremely low toxicity, low fouling, good durability, and chemical similarity to typical PEG polymers, enabling live cell patterning as well as various bioapplications using bioincompatible materials. The PP-PEG film can be overlaid on any materials via the capacitively coupled plasma chemical vapor deposition (CCP-CVD) method using nontoxic PEG200 as a precursor. The biocompatibility of the PP-PEG-coated surface is confirmed by whole blood flow experiments where no thrombi and less serum protein adsorption are observed when compared with bare glass, polyethylene (PE), and polyethylene terephthalate (PET) surfaces. Furthermore, unlike bare PE films, less fibrosis and inflammation are observed when the PP-PEG-coated PE film is implanted into subcutaneous pockets of mice groin areas. The cell-repellent property of PP-PEG is also verified via patterning of mammalian cells, such as fibroblasts and hippocampal neurons. These results show that our PP-PEG film, generated by the CCP-CVD method, is a biocompatible material that can be considered for broad applications in biomedical and functional materials fields.
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Materiales Biocompatibles Revestidos/química , Polietilenglicoles/química , Ingeniería de Tejidos/instrumentación , Andamios del Tejido/química , Animales , Células Sanguíneas/citología , Humanos , Ensayo de Materiales , Ratones , Ratones Endogámicos C57BL , PolimerizacionRESUMEN
Here, we show that GaN nanowires (diameter <30 nm) can be used as strain relaxing substrates for the heteroepitaxial growth of stable In(x)Ga(1-x)N alloys of controlled composition and thickness. Thinner nanowires with their smaller interfacial area reduce the heteroepitaxial stress. Also, the limited adatom diffusion length scales on the thinner nanowires aid in reducing the kinetic segregation effects. In addition to being single crystal templates for heteroepitaxial growth, these thick single crystal overlayers on nanowire substrates can provide suitable architectures for photoelectrochemical applications. The stability and crystallinity of the In(x)Ga(1-x)N layers are preserved by the nanowires acting as compliant substrates. Photoelectrochemical water splitting requires In(x)Ga(1-x)N alloys with a 2.2-1.6 eV band gap (i.e. 0.45 < x < 0.65) and 150-200 nm film thickness for efficient light absorption and carrier generation. At such compositions, the In(x)Ga(1-x)N alloys are inherently unstable, the thickness-dependent stress builds up during the commonly employed heteroepitaxial growth methods, and adds to the instability causing phase segregation and property degradation. A dependence of the growth morphology on the GaN nanowire growth orientation was observed and a growth mechanism is presented for the observed orientation dependent growth on a-plane and c-plane GaN nanowires. Photoactivity of GaN and In(x)Ga(1-x)N films on GaN nanowires is also investigated which shows a distinct difference attributable to GaN and In(x)Ga(1-x)N, demonstrating the advantages of using nanowires as strain relaxing substrates.
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We investigate the efficacy of a multisectoral road safety campaign initiated at Luang Namtha Provincial Hospital (LNPH), North-Laos. Road safety days (RSD) with helmet promotion were organised prior to the Lao and International New Year 2007. Motorbike helmet protectiveness was demonstrated by dropping peeled versus unpeeled coconuts from 6-m high sticks simulating city speed-limit (40 km/h). The primary outcome was the number of road traffic injuries (RTI) needing admission (severe RTI) before and after the first RSD. Secondary outcomes were helmet usage, total RTI, hospital staff's behavioural changes and law enforcement. Neighbouring Bokeo Provincial Hospital (BPH) had no intervention. Severe RTI dropped by 34.7% within 12 months (from 449 to 293, p ≤ 0.0001, BPH +123% [from 88 to 196]). Total RTI increased by 10.8% (from 772 to 855, p = 0.0396, BPH +260.8% [from 186 to 671]). Police started the law enforcement right after the RSD. Helmet use increased from 11.2 to 42.5% (p < 0.0001). Coconut test was the main reason for safer behaviour among RSD participants seven months later (16/32). The intervention effectively prevented severe RTI. The speed-adapted coconut drop test was a simple, convincing and efficient educational tool, easily adaptable to other settings.
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Accidentes de Tránsito/prevención & control , Promoción de la Salud/métodos , Hospitales/estadística & datos numéricos , Seguridad/estadística & datos numéricos , Mercadeo Social , Accidentes de Tránsito/estadística & datos numéricos , Cocos , Dispositivos de Protección de la Cabeza/estadística & datos numéricos , Conductas Relacionadas con la Salud , Promoción de la Salud/estadística & datos numéricos , Humanos , Laos , Motocicletas/estadística & datos numéricos , Desarrollo de Programa , Asunción de Riesgos , Encuestas y Cuestionarios , Factores de Tiempo , Heridas y Lesiones/epidemiología , Heridas y Lesiones/prevención & controlRESUMEN
OBJECTIVES: To understand the relationship between the -202 A/C single-nucleotide polymorphism (SNP) of the insulin-like growth factor-binding protein 3 (IGFBP3) gene, IGFBP3 serum levels, and risk of prostate cancer (PCa) in a Korean population, as a potential reason for the lower incidence of PCa in the Korean population. METHODS: The IGFBP3 levels were measured and the -202 A/C SNP of the IGFBP3 gene was typed for 225 PCa cases and the same number of matched controls. Linear regression analysis and unconditional logistic regression analysis were used to test for the associations between the genotypes and the circulating IGFBP3 levels and the risk of PCa, respectively. To adjust for the potential bias introduced by the hospital cohort, the result of the genotyping was compared with that of 683 community-indwelling healthy men. RESULTS: Significantly lower plasma levels of IGFBP3 were noted in the PCa cases. Lower IGFBP3 plasma levels were associated with an increased number of C alleles (P<.001). Compared with the PCa cases, a lower frequency of the C allele was found in the hospital and community controls (P<.05). Compared with AA genotype, logistic regression analysis revealed an increased risk of PCa in subjects who were CC genotype (odds ratio: 2.39, 95% confidence interval: 1.05-5.48). Larger odds (odds ratio: 3.37, 95% confidence interval: 1.35-8.43) for PCa were associated with CC genotype when the analysis was confined to those who had high-risk PCa. CONCLUSIONS: The results supported the protective role of -202 A/C SNP of the IGFBP3 gene against PCa in Korean men.
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Predisposición Genética a la Enfermedad/epidemiología , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/sangre , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Polimorfismo Genético , Neoplasias de la Próstata/genética , Distribución por Edad , Anciano , Alelos , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , Estudios de Casos y Controles , Estudios de Cohortes , Humanos , Incidencia , Corea (Geográfico)/epidemiología , Masculino , Persona de Mediana Edad , Probabilidad , Neoplasias de la Próstata/sangre , Neoplasias de la Próstata/epidemiología , Neoplasias de la Próstata/patología , Valores de Referencia , Medición de RiesgoRESUMEN
Polyphosphazenes are a class of hybrid organic-inorganic macromolecules with high thermo-oxidative stability and good solubility in many solvents. Fluoroalkoxy phosphazene polymers also have high surface hydrophobicity. A method is described to tune this surface property while maintaining the advantageous bulk materials characteristics. The polyphosphazene single-substituent polymer, poly[bis(2,2,2-trifluoroethoxy)phosphazene], with flat film, fiber mat, or bead mat morphology was surface functionalized using an atmospheric plasma treatment with oxygen, nitrogen, methane, or tetrafluoromethane/hydrogen gases. Surface chemistry changes were detected by static water contact angle (WCA) measurements as well as X-ray photon spectroscopy (XPS). It was found that changes in the WCA of as much as 150 degrees occurred, accompanied by shifts in the ratio of elements on the polymer surface as detected by XPS. Overall this plasma technique provides a convenient method for the generation of specific surface characteristics while maintaining the hydrophobicity of the bulk material.
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BCR-ABL oncoprotein-expressing cells are associated with a relative increase of intracellular reactive oxygen species (ROS), which is thought to play a role in transformation. Elevated ROS levels in BCR-ABL-transformed cells were found to be blocked by the mitochondrial complex I inhibitor rotenone as well as the glucose transport inhibitor phloretin, suggesting that the source of increased ROS might be related to increased glucose metabolism. The glucose analog 2-deoxyglucose (2-DOG) reduced ROS to levels found in non-BCR-ABL-transformed cells and inhibited cell growth alone or in cooperation with imatinib mesylate (Gleevec). A mutant of BCR-ABL that is defective in transformation of myeloid cells, Tyr177Phe, was also found to be defective in raising intracellular ROS levels. Glucose metabolism in BCR-ABL-transformed cells is likely to be mediated by activation of the phosphatidylinositol-3'-kinase (PI3K) pathway, which is regulated through this site. Inhibition of PI3K or mTOR led to a significant decrease in ROS levels. Overall, our results suggest that elevated levels of ROS in BCR-ABL-transformed cells are secondary to a transformation-associated increase in glucose metabolism and an overactive mitochondrial electron transport chain and is specifically regulated by PI3K. Finally, these results hint at novel targets for drug development that may aid traditional therapy.
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Transformación Celular Neoplásica/metabolismo , Proteínas de Fusión bcr-abl/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Quinasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/fisiología , Regulación hacia Arriba , Benzamidas , Línea Celular , Línea Celular Tumoral , Desoxiglucosa/farmacología , Activación Enzimática , Humanos , Mesilato de Imatinib , Células Jurkat , Células K562 , Leucemia Mielógena Crónica BCR-ABL Positiva/enzimología , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Proteínas de Transporte de Monosacáridos/antagonistas & inhibidores , Proteínas de Transporte de Monosacáridos/metabolismo , Floretina/farmacología , Fosfatidilinositol 3-Quinasas/fisiología , Piperazinas/farmacología , Proteínas Quinasas/fisiología , Pirimidinas/farmacología , Serina-Treonina Quinasas TOR , Tirosina/genética , Tirosina/fisiologíaRESUMEN
Thermobifida fusca grows well on cellulose and xylan, and produces a number of cellulases and xylanases. The gene encoding a previously unstudied endoxylanase, xyl10B, was overexpressed in E. coli, and the protein was purified and characterized. Mature Xyl10B is a 43-kDa glycohydrolase with a short basic domain at the C-terminus. It has moderate thermostability, maintaining 50% of its activity after incubation for 16 h at 62 degrees C, and is most active between pH 5 and 8. Xyl10B is produced by growth of T. fusca on xylan or Solka Floc but not on pure cellulose. Mass spectroscopic analysis showed that Xyl10B produces xylobiose as the major product from birchwood and oat spelts xylan and that its hydrolysis products differ from those of T. fusca Xyl11A. Xyl10B hydrolyzes various p-nitrophenyl-sugars, including p-nitrophenyl alpha-D-arabinofuranoside, p-nitrophenyl-beta-D-xylobioside, p-nitrophenyl-beta-D-xyloside, and p-nitrophenyl-beta-D-cellobioside. Xyl11A has higher activity on xylan substrates, but Xyl10B produced more reducing sugars from corn fiber than did Xyl11A.