RESUMEN
Twelve compounds were isolated from Liquidambaris Resina by silica gel column chromatography and thin layer chromatography. Their structures were identified on the basis of spectral data, electron capture detector data, and physicochemical properties as(2'R, 3'R)-2',3'-dihydroxy-hydrocinnamyl-(E)-cinnamate(1),(E)-cinnamyl-(E)-cinnamate(2), cinnamic acid(3), 28-norlup-20(29)-en-3-one-17ß-hydroperoxide(4), erythrodiol(5), 13ß,28-epoxy-30-hydroxyolean-1-en-3-one(6),(3ß)-olean-12-ene-3,23-diol(7), 2α,3α-dihydroxy-olean-12-en-28-oic acid(8), 28-hydroxyolean-12-en-3-one(9), 3-epi-oleanolic acid(10), 3-oxo-oleanolic acid(11), and hederagenin(12). Compound 1 was a new cinnamic acid ester derivative and compounds 2-4,6-8, and 12 were isolated from Liquidambaris Resina for the first time. Compounds 4, 5, 10, and 12 exerted inhibitory effects on the proliferation of human umbilical vein endothelial cells(HUVEC) with the IC_(50) values of(17.43±2.17),(35.32±0.61),(27.50±0.80), and(46.30±0.30) µmol·L~(-1), respectively.
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Ácido Oleanólico , Triterpenos , Humanos , Células Endoteliales , Ésteres , Cinamatos , Triterpenos/química , Estructura MolecularRESUMEN
A novel mutation, HBB: c.393T>G on the HBB gene, was detected in two hypochromic microcytic anemia patients from Yulin, in the Guangxi Province of the People's Republic of China (PRC), by next-generation sequencing (NGS). It is a nonsense mutation causing a stop codon at amino acid 131 in exon 3 of the HBB gene. It was found in a heterozygous state in two patients who both presented severe anemia during pregnancy and moderate anemia before pregnancy; Hb A2 levels were slightly increased (more than 4.0%) in both patients. It was also detected in the father of one of the patients. This mutation was pathogenic, and caused the dominant thalassemia-like phenotypes in the two patients.
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Globinas beta , Talasemia beta , Anemia Hipocrómica , China , Codón sin Sentido , Femenino , Humanos , Masculino , Globinas beta/genética , Talasemia beta/genéticaRESUMEN
Combinatorial approach of adsorbent resin HP20 addition and metabolic profiling analysis were carried out to enhance ascomycin production. Under the optimal condition of 5 % m/v HP20 added at 24 h, ascomycin production was increased to 380 from 300 mg/L. To further rationally guide the improvement of ascomycin production, metabolic profiling analysis was employed to investigate the intracellular metabolite changes of Streptomyces hygroscopicus var. ascomyceticus FS35 in response to HP20 addition. A correlation between the metabolic profiles and ascomycin accumulation was revealed by partial least-squares to latent structures discriminant analysis, and 11 key metabolites that most contributed to metabolism differences and ascomycin biosynthesis were identified. Based on the analysis of metabolite changes together with their pathways, the potential key factors associated with ascomycin overproduction were determined. Finally, rationally designed fermentation strategies based on HP20 addition were performed as follows: 2 % v/v n-hexadecane was added at 24 h; 1.0 g/L valine was supplemented at 48 h; 1.0 g/L lysine was added at 72 h. The ascomycin production was ultimately improved to 460 mg/L, a 53.3 % enhancement compared with that obtained in initial condition. These results demonstrated that the combination of HP20 addition and metabolic profiling analysis could be successfully applied to the rational guidance of production improvement of ascomycin, as well as other clinically important compounds.
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Antibacterianos/biosíntesis , Metaboloma , Resinas Sintéticas/química , Streptomyces/metabolismo , Tacrolimus/análogos & derivados , Antibacterianos/química , Medios de Cultivo/química , Medios de Cultivo/metabolismo , Fermentación , Tacrolimus/química , Tacrolimus/metabolismoRESUMEN
INTRODUCTION: The human epidermal growth factor receptor 2 (HER2) receptor tyrosine kinase (RTK) oncogene is an attractive therapeutic target for the treatment of HER2-addicted tumors. Although lapatinib, an FDA-approved small-molecule HER2 and epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), represents a significant therapeutic advancement in the treatment of HER2+ breast cancers, responses to lapatinib have not been durable. Consequently, elucidation of mechanisms of acquired therapeutic resistance to HER-directed therapies is of critical importance. METHODS: Using a functional protein-pathway activation mapping strategy, along with targeted genomic knockdowns applied to a series of isogenic-matched pairs of lapatinib-sensitive and resistant cell lines, we now report an unexpected mechanism of acquired resistance to lapatinib and similar TKIs. RESULTS: The signaling analysis revealed that whereas HER2 was appropriately inhibited in lapatinib-resistant cells, EGFR tyrosine phosphorylation was incompletely inhibited. Using a targeted molecular knockdown approach to interrogate the causal molecular underpinnings of EGFR-persistent activation, we found that lapatinib-resistant cells were no longer oncogene addicted to HER2-HER3-PI3K signaling, as seen in the parental lapatinib-sensitive cell lines, but instead were dependent on a heregulin (HRG)-driven HER3-EGFR-PI3K-PDK1 signaling axis. Two FDA-approved EGFR TKIs could not overcome HRG-HER3-mediated activation of EGFR, or reverse lapatinib resistance. The ability to overcome EGFR-mediated acquired therapeutic resistance to lapatinib was demonstrated through molecular knockdown of EGFR and treatment with the irreversible pan-HER TKI neratinib, which blocked HRG-dependent phosphorylation of HER3 and EGFR, resulting in apoptosis of resistant cells. In addition, whereas HRG reversed lapatinib-mediated antitumor effects in parental HER2+ breast cancer cells, neratinib was comparatively resistant to the effects of HRG in parental cells. Finally, we showed that HRG expression is an independent negative predictor of clinical outcome in HER2+ breast cancers, providing potential clinical relevance to our findings. CONCLUSIONS: Molecular analysis of acquired therapeutic resistance to lapatinib identified a new resistance mechanism based on incomplete and "leaky" inhibition of EGFR by lapatinib. The selective pressure applied by incomplete inhibition of the EGFR drug target resulted in selection of ligand-driven feedback that sustained EGFR activation in the face of constant exposure to the drug. Inadequate target inhibition driven by a ligand-mediated autocrine feedback loop may represent a broader mechanism of therapeutic resistance to HER TKIs and suggests adopting a different strategy for selecting more effective TKIs to advance into the clinic.
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Comunicación Autocrina , Neoplasias de la Mama/metabolismo , Receptores ErbB/metabolismo , Neurregulina-1/metabolismo , Receptor ErbB-2/metabolismo , Receptor ErbB-3/metabolismo , Transducción de Señal , Neoplasias de la Mama/genética , Neoplasias de la Mama/mortalidad , Proteína Tirosina Quinasa CSK , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Resistencia a Antineoplásicos , Receptores ErbB/genética , Femenino , Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Lapatinib , Neurregulina-1/genética , Fosfatidilinositol 3-Quinasas , Fosfoproteínas Fosfatasas/metabolismo , Fosforilación , Pronóstico , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora , Quinazolinas/farmacología , Receptor ErbB-3/genética , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/metabolismo , Familia-src Quinasas/antagonistas & inhibidoresRESUMEN
Rapamycin is a clinically important macrocyclic polyketide with immunosuppressive activity produced by Streptomyces hygroscopicus. To rationally guide the improvement of rapamycin production, comparative metabolic profiling analysis was performed in this work to investigate the intracellular metabolic changes in S. hygroscopicus U1-6E7 fermentation in medium M1 and derived medium M2. A correlation between the metabolic profiles and rapamycin accumulation was revealed by partial least-squares to latent structures analysis, and 16 key metabolites that most contributed to the metabolism differences and rapamycin production were identified. Most of these metabolites were involved in tricarboxylic acid cycle, fatty acids, and shikimic acid and amino acids metabolism. Based on the analysis of key metabolites changes in the above pathways, corresponding exogenous addition strategies were proposed as follows: 1.0 g/L methyl oleate was added at 0 h; 1.0 g/L lysine was added at 12 h; 0.5 g/L shikimic acid was added at 24 h; 0.5 g/L sodium succinate, 0.1 g/L phenylalanine, 0.1 g/L tryptophan, and 0.1 g/L tyrosine were added at 36 h, successively, and a redesigned fermentation medium (M3) was obtained finally on the basis of M2. The production of rapamycin in M3 was increased by 56.6 % compared with it in M2, reaching 307 mg/L at the end of fermentation (120 h). These results demonstrated that metabolic profiling analysis was a successful method applied in the rational guidance of the production improvement of rapamycin, as well as other industrially or clinically important compounds.
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Inmunosupresores/metabolismo , Metaboloma , Sirolimus/metabolismo , Streptomyces/metabolismo , Biotecnología/métodos , Medios de Cultivo/química , Tecnología Farmacéutica/métodosRESUMEN
Long noncoding RNAs (lncRNAs) have been associated with a variety of biological activities, including immune responses. However, the function of lncRNAs in antiviral innate immune responses are not fully understood. Here, we identified a novel lncRNA, termed dual function regulating influenza virus (DFRV), elevating in a dose- and time-dependent manner during influenza A virus (IAV) infection, which was dependent on the NFκB signaling pathway. Meanwhile, DFRV was spliced into two transcripts post IAV infection, in which DFRV long suppress the viral replication while DFRV short plays the opposite role. Moreover, DFRV regulates IL-1ß and TNF-α via activating several pro-inflammatory signaling cascades, including NFκB, STAT3, PI3K, AKT, ERK1/2 and p38. Besides, DFRV short can inhibit DFRV long expression in a dose-dependent manner. Collectively, our studies reveal that DFRV may act as a potential dual-regulator to preserve innate immune homeostasis in IAV infection.
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In this work a chip-based nano HPLC coupled MS (HPLC-chip/MS) method with a simple sample preparation procedure was developed for the flavonoid profiling of soybean. The analytical properties of the method including the linearity (R(2) , 0.992-0.995), reproducibility (RSD, 1.50-7.66%), intraday precision (RSD, 1.41-5.14%) and interday precision (RSD, 2.76-16.90%) were satisfactory. Compared with the conventional HPLC/MS method, a fast extraction and analysis procedure was applied and more flavonoids were detected in a single run. Additionally, 13 flavonoids in soybean seed were identified for the first time. The method was then applied to the profiling of six varieties of soybean sowed at the same place. A clear discrimination was observed among different cultivars, three isoflavones, accounting for nearly 80% of total flavonoid contents, were found increased in the spring soybeans compared with the summer cultivars.
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Cromatografía Líquida de Alta Presión/métodos , Flavonoides/análisis , Glycine max/química , Procedimientos Analíticos en Microchip/métodos , Flavonoides/aislamiento & purificación , Análisis de los Mínimos Cuadrados , Modelos Lineales , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Glycine max/clasificación , Espectrometría de Masas en Tándem/métodosRESUMEN
Spinal muscular atrophy (SMA) is one of the most common and severe genetic diseases. SMA carrier screening is an effective way to identify couples at risk of having affected children. Next-generation sequencing (NGS)-based expanded carrier screening could detect SMN1 gene copy number without extra experiment and with high cost performance. However, its performance has not been fully evaluated. Here we conducted a systematic comparative study to evaluate the performance of three common methods. 478 samples were analyzed with multiplex ligation probe amplification (MLPA), real-time quantitative polymerase chain reaction (qPCR) and NGS, simultaneously. Taking MLPA-based results as the reference, for 0 copy, 1 copy and ≥ 2 copy SMN1 analysis with NGS, the sensitivity, specificity and precision were all 100%. Using qPCR method, the sensitivity was 100%, 97.52% and 94.30%, respectively; 98.63%, 95.48% and 100% for specificity; and 72.72%, 88.72% and 100% for precision. NGS repeatability was higher than that of qPCR. Moreover, among three methods, NGS had the lowest retest rate. Thus, NGS is a relatively more reliable method for SMN1 gene copy number detection. In expanded carrier screening, compared with the combination of multiple methods, NGS method could reduce the test cost and simplify the screening process.
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Exones , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Atrofia Muscular Espinal/genética , Eliminación de Secuencia , Proteína 1 para la Supervivencia de la Neurona Motora/genética , Dosificación de Gen , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteína 2 para la Supervivencia de la Neurona Motora/genéticaRESUMEN
The aim of this study was to use a two steps strategy metabolomics to screen/identify and validate novel metabolic biomarker(s) for epithelial ovarian cancer (EOC). In the screening step, serum samples from 27 healthy women, 28 benign ovarian tumors, and 29 EOCs were analyzed by using LC-MS based nontargeted metabolomics. The three groups were separated with OSC filtered PLS-DA model, and six metabolites (27-nor-5ß-cholestane-3,7,12,24,25 pentol glucuronide (CPG), phenylalanine, glycocholic acid, propionylcarnitine, Phe-Phe and Lyso PC (18:2)) were considered as potential biomarker candidates. In the validation step, the six metabolites were analyzed in targeted metabolomics by LC-selective ion monitoring mass spectrometry in another 685 serum samples with various clinical backgrounds. As a result, CPG was evaluated to be a potential biomarker and its content was elevated in EOC tissues compared with benign ovarian tumor tissues (p = 0.0005). Besides, CPG levels were found to be up-regulated in early stage EOC and in the three types of EOC histological types. Other variables such as nonovarian diseases, medicine consumption, gynecological inflammations, and menopausal state did not interfere in using CPG as diagnosis marker. CPG was found to be complementary to CA125. Our findings suggest that CPG can be considered a statistical relevant biomarker of EOC, ready for early stage detection.
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Biomarcadores de Tumor/sangre , Colestanoles/sangre , Regulación Neoplásica de la Expresión Génica/fisiología , Glucurónidos/sangre , Metabolómica/métodos , Antígeno Ca-125/metabolismo , Carcinoma Epitelial de Ovario , Cromatografía Liquida , Femenino , Humanos , Espectrometría de Masas , Proteínas de la Membrana/metabolismo , Neoplasias Glandulares y Epiteliales/sangre , Neoplasias Glandulares y Epiteliales/diagnóstico , Neoplasias Ováricas/sangre , Neoplasias Ováricas/diagnóstico , Curva ROCRESUMEN
BACKGROUND: Noninvasive prenatal testing (NIPT) of recessive monogenic diseases depends heavily on knowing the correct parental haplotypes. However, the currently used family-based haplotyping method requires pedigrees, and molecular haplotyping is highly challenging due to its high cost, long turnaround time, and complexity. Here, we proposed a new two-step approach, population-based haplotyping-NIPT (PBH-NIPT), using α-thalassemia and ß-thalassemia as prototypes. METHODS: First, we deduced parental haplotypes with Beagle 4.0 with training on a large retrospective carrier screening dataset (4356 thalassemia carrier screening-positive cases). Second, we inferred fetal haplotypes using a parental haplotype-assisted hidden Markov model (HMM) and the Viterbi algorithm. RESULTS: With this approach, we enrolled 59 couples at risk of having a fetus with thalassemia and successfully inferred 94.1% (111/118) of fetal alleles. We confirmed these alleles by invasive prenatal diagnosis, with 99.1% (110/111) accuracy (95% CI, 95.1-100%). CONCLUSIONS: These results demonstrate that PBH-NIPT is a sensitive, fast, and inexpensive strategy for NIPT of thalassemia.
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Haplotipos/genética , Pruebas Prenatales no Invasivas , Padres , Talasemia alfa/genética , Talasemia beta/genética , Genética de Población , Humanos , Tamaño de la MuestraRESUMEN
In this study, we performed a spinal muscular atrophy carrier screening investigation with NGS-based method. First, the validation for NGS-based method was implemented in 2255 samples using real-time PCR. The concordance between the NGS-based method and real-time PCR for the detection of SMA carrier and patient were up to 100%. Then, we applied this NGS-based method in 10,585 self-reported normal couples (34 Chinese ethnic groups from 5 provinces in South China) for SMA carrier screening. The overall carrier frequency was 1 in 73.8 (1.4%). It varied substantially between ethnic groups, highest in Dai ethnicity (4.3%), and no significant difference was found between five provinces. One couple was detected as carriers with an elevated risk of having an SMA affected baby. The distribution of SMN1:SMN2 genotype was also revealed in this study. Among the individuals with normal phenotype, the exon 7 copy-number ratio of SMN1 to SMN2 proved the gene conversion between them. With NGS-based method, we investigated SMA carrier status in Chinese population for the first time, and our results demonstrated that it is a promising alternative for SMA carrier screening and could provide data support and reference for future clinical application.
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Etnicidad/genética , Frecuencia de los Genes , Tamización de Portadores Genéticos/estadística & datos numéricos , Atrofia Muscular Espinal/genética , China , Femenino , Conversión Génica , Secuenciación de Nucleótidos de Alto Rendimiento/estadística & datos numéricos , Humanos , Masculino , Atrofia Muscular Espinal/etnología , Análisis de Secuencia de ADN/estadística & datos numéricos , Proteína 1 para la Supervivencia de la Neurona Motora/genética , Proteína 2 para la Supervivencia de la Neurona Motora/genéticaRESUMEN
Defects in tight junction barrier have been considered as an important etiologic factor of Crohn's disease. n-3 polyunsaturated fatty acids (PUFAs) exert beneficial effects on inflammatory bowel disorders. However, the mechanisms remain unknown. We found that docosahexaenoic acid (DHA, 22:6 n-3) and eicosapentaenoic acid (EPA, 20:5 n-3) changed lipid environment in membrane microdomains of tight junction in vitro. n-3 PUFAs treatment effectively prevented the redistribution of occludin and ZO-1 and distortion of TJ morphology, reduced transepithelial electrical resistance induced by IFN-gamma and TNF-alpha. We also observed dramatic reorganization of TJ proteins in epithelial lateral membrane following treatment with these cytokines. Our findings for first time indicate that n-3 PUFAs play an important role in proinflammatory cytokines-induced permeability defects and epithelial barrier dysfunction by modifying lipid environment in membrane microdomains of tight junction.
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Permeabilidad de la Membrana Celular , Citocinas/farmacología , Epitelio/fisiología , Ácidos Grasos Insaturados/fisiología , Microdominios de Membrana/química , Uniones Estrechas/química , Humanos , Inflamación , Interferón gamma/farmacología , Proteínas de la Membrana/metabolismo , Ocludina , Fosfoproteínas/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Proteína de la Zonula Occludens-1RESUMEN
Expanded carrier screening (ECS) has been demonstrated to increase the detection rate of carriers compared with traditional tests. The aim of this study was to assess the potential value of ECS for clinical application in Southern China, a region with high prevalence of thalassemia and with diverse ethnic groups, and to provide a reference for future implementations in areas with similar population characteristics. A total of 10,476 prenatal/preconception couples from 34 self-reported ethnic groups were simultaneously tested and analyzed anonymously for 11 Mendelian disorders using targeted next-generation sequencing. Overall, 27.49% of individuals without self-reported family history of disorders were found to be carriers of at least 1 of the 11 conditions, and the carrier frequency varied greatly between ethnic groups, ranging from 4.15% to 81.35%. Furthermore, 255 couples (2.43%) were identified as carrier couples at an elevated risk having an affected baby, sixty-five of which would not have been identified through the existing screening strategy, which only detects thalassemia. The modeled risk of fetuses being affected by any of the selected disorders was 531 per 100,000 (95% CI, 497-567 per 100,000). Our data demonstrate the feasibility of ECS, and provide evidence that ECS is a promising alternative to traditional one-condition screening strategies. The lessons learned from this experience should be applicable for other countries or regions with diverse ethnic groups.
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Etnicidad/genética , Tamización de Portadores Genéticos/métodos , Adulto , Femenino , Frecuencia de los Genes , Genes Recesivos , Tamización de Portadores Genéticos/normas , Humanos , Masculino , Proyectos Piloto , Tamaño de la MuestraRESUMEN
Tight junctions of epithelial cells determine epithelial membrane integrity and play an important role in selective paracellular permeability to ions and macromolecules. In this work, we investigated the effect of one of n-3 series polyunsaturated fatty acids, eicosapentaenoic acid (EPA) on the phospholipid composition of membrane microdomains of tight junctions. After treated by EPA, membrane microdomains of tight junctions were isolated by discontinuous sucrose density gradient ultracentrifugation, and raft phospholipids were extracted. The PE, PI, PS, PC and SM were separated and determined by high-performance liquid chromatography/quadrupole-linear ion trap mass spectrometry (HPLC Qtrap-MS), and were further identified by HPLC-MS/MS. It was found that EPA altered the fatty acyl substitution of phospholipids that constituted membrane microdomains of tight junctions by enriching the unsaturated fatty acyl chains of the phospholipids. It provides a new visual angle to explaining the intracellular mechanism involved in n-3 polyunsaturated fatty acids (PUFAs) modulation of intestinal tight junction barrier.
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Ácido Eicosapentaenoico/farmacología , Células Epiteliales/metabolismo , Microdominios de Membrana , Fosfolípidos/análisis , Uniones Estrechas/química , Línea Celular , Cromatografía Líquida de Alta Presión , Células Epiteliales/fisiología , Humanos , Microdominios de Membrana/química , Microdominios de Membrana/efectos de los fármacos , Microdominios de Membrana/fisiología , Fosfolípidos/química , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
In breast cancer, overexpression of ErbB2 or aberrant regulation of survivin, a member of the inhibitor of apoptosis family, is associated with resistance to chemo/hormone therapy and predicts for a poor clinical outcome. A functional link between the two predictive factors has not been previously shown. Here, using genetic and pharmacologic approaches to block ErbB2 signaling, we show that ErbB2 regulates survivin protein expression in ErbB2-overexpressing breast cancer cells. Selective knockdown of ErbB2 using small interfering RNA markedly reduced survivin protein, resulting in apoptosis of ErbB2-overexpressing breast cancer cell lines such as BT474. Alternatively, inhibition of ErbB2 signaling using lapatinib (GW572016), a reversible small-molecule inhibitor of ErbB1/ErbB2 tyrosine kinases, at pharmacologically relevant concentrations, leads to marked inhibition of survivin protein with subsequent apoptosis. The effect of lapatinib on survivin seems to be predominantly posttranslational, mediated by ubiquitin-proteosome degradation as lactacystin, a proteosome inhibitor, reverses these effects. Furthermore, lapatinib down-regulated the expression of His-tagged survivin, which was under the transcriptional control of a heterologous promoter, providing additional evidence supporting a posttranslational mechanism of regulation. In contrast, trastuzumab and gefitinib failed to down-regulate survivin in ErbB2-overexpressing breast cancer cells. Importantly, the clinical relevance of these findings was illustrated in patients with ErbB2-overexpressing breast cancer whose clinical response to lapatinib was associated with marked inhibition of survivin in their tumors. These findings shed new light on the mechanism by which ErbB2 overexpression protects against apoptotic stimuli in breast cancer and identifies therapeutic interventions to improve clinical outcomes in these aggressive tumors.
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Neoplasias de la Mama/metabolismo , Proteínas Asociadas a Microtúbulos/biosíntesis , Proteínas de Neoplasias/biosíntesis , Receptor ErbB-2/fisiología , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Neoplasias de la Mama/genética , Línea Celular Tumoral , Regulación hacia Abajo , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Humanos , Proteínas Inhibidoras de la Apoptosis , Lapatinib , Proteínas Asociadas a Microtúbulos/antagonistas & inhibidores , Proteínas Asociadas a Microtúbulos/genética , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Quinazolinas/farmacología , Interferencia de ARN , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/biosíntesis , Receptor ErbB-2/genética , Receptor ErbB-3/fisiología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , SurvivinRESUMEN
High-performance electromagnetic interference (EMI)-shielding materials featuring lightweight, flexibility, excellent conductivity, and shielding properties, as well as superior mechanical robustness, are highly required, yet their development still remains a daunting challenge. Here, a flexible and exceptional EMI-shielding polydimethylsilane/reduced graphene oxide/single-wall carbon nanotube (PDMS/rGO/SWCNT) nanocomposite was developed by a facile backfilling approach utilizing a preformed rGO/SWCNT aerogel as the three-dimensional (3D) conducting and reinforcement skeleton. Pristine SWCNTs acting as secondary conductive fillers showed intriguing advantages, whose intrinsically high conductivity could be well preserved in the composites because of no surface acidification treatment. The robust and interconnected 3D network can not only serve as fast channels for electron transport but also effectively transfer external load. Accordingly, a prominent electrical conductivity of 1.2 S cm-1 and an outstanding EMI-shielding effectiveness of around 31 dB over the X-band frequency range were achieved for the resultant composite with an ultralow loading of 0.28 wt %, which is among the best results for currently reported conductive polymer nanocomposites. Moreover, the composite displayed excellent mechanical properties and bending stability; for example, a 233% increment in the compression strength was obtained compared with that of neat PDMS. These observations indicate the unrivalled effectiveness of 3D rGO/SWCNT aerogel as a reinforcement to endow the polymer composites with outstanding conductive and mechanical properties toward high-performance EMI-shielding application.
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Endothelial nitric oxide synthase (eNOS) plays a crucial role in the regulation of a variety of cardiovascular functions. Many studies have shown that dietary n-3 polyunsaturated fatty acids (PUFAs) have beneficial effects on coronary atherosclerosis. However, the mechanisms of n-3 PUFAs regulation in eNOS activation remain unknown. In the present study we investigated the effects of eicosapentaenoic acid (EPA, 20:5 n-3) on subcellular distribution of eNOS and lipid composition of caveolae. We demonstrated for the first time that EPA treatment profoundly altered lipid composition and fatty acyl substitutions of phospholipids in caveolae. We found that caveolin-1 was solely located in caveolae fractions in control cells, and EPA treatment displaced caveolin-1 from caveolae. eNOS was detected in the caveolin-enriched fractions and noncaveolae fractions in control cells. EPA treatment induced the translocation of eNOS from caveolae fractions to soluble fractions. P-eNOS was also distributed in both fractions. After EPA treatment, the level of p-eNOS in each fraction was increased but the distribution of which was unaffected. Moreover, the results of immunofluorescence confirmed that EPA could redistribute caveolin-1 and eNOS in plasma membrane. eNOS activity in HUVEC cells was increased after EPA treatment, which was in a dose dependent manner. And incubation with 50 microM EPA had the maximum effect on eNOS activity. Our results suggested that eNOS translocation was paralleled by a stimulated capacity for NO production in the cells. We found that total Akt and p-Akt were primarily presented in heavy membranes in control cells, and the relative level of p-Akt increased but the distribution did not change after EPA treatment. The distribution of CaM was slightly changed after EPA treatment. Our results indicated that n-3 PUFAs profoundly altered caveolae microenvironment, thereby modifying location and function of proteins in caveolae. EPA-induced alterations of lipid and proteins in caveolae may be an important mechanism in the pathophysiologic process of atherosclerosis.
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Caveolas/metabolismo , Ácido Eicosapentaenoico/metabolismo , Ácidos Grasos/metabolismo , Óxido Nítrico Sintasa de Tipo III/metabolismo , Western Blotting , Caveolas/química , Técnica del Anticuerpo Fluorescente , Humanos , Espectrometría de Masas , Transporte de ProteínasRESUMEN
Plasma phospholipids metabolic profile of chronic glomerulonephritis was investigated using high performance liquid chromatography/mass spectrometry (LC/MS) and principal component analysis. The plasma samples of 18 patients with chronic glomerulonephritis, 17 patients with chronic renal failure (CRF) without renal replacement therapy and 18 healthy persons were collected and analyzed. It was found that combination of the LC/MS technique with PCA can be successfully applied to phospholipid profile analysis. The results showed that primary chronic glomerulonephritis and CRF had phospholipids metabolic abnormality. Nineteen phospholipid species were identified as possible biomarkers in plasma samples of chronic glomerulonephritis and chronic renal failure. Moreover, the identification of the molecular structure of the potential phospholipid markers was obtained by ESI-MS/MS experiment. It suggests that phospholipids can be used as potential biomarkers on the progress of primary chronic glomerulonephritis.
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Biomarcadores/sangre , Cromatografía Líquida de Alta Presión/métodos , Glomerulonefritis/metabolismo , Espectrometría de Masas/métodos , Fosfolípidos/metabolismo , Adulto , Enfermedad Crónica , Femenino , Glomerulonefritis/sangre , Humanos , Fallo Renal Crónico/sangre , Masculino , Persona de Mediana Edad , Fosfolípidos/sangreRESUMEN
Hemoglobinopathies are among the most common autosomal-recessive disorders worldwide. A comprehensive next-generation sequencing (NGS) test would greatly facilitate screening and diagnosis of these disorders. An NGS panel targeting the coding regions of hemoglobin genes and four modifier genes was designed. We validated the assay by using 2522 subjects affected with hemoglobinopathies and applied it to carrier testing in a cohort of 10,111 couples who were also screened through traditional methods. In the clinical genotyping analysis of 1182 ß-thalassemia subjects, we identified a group of additional variants that can be used for accurate diagnosis. In the molecular screening analysis of the 10,111 couples, we detected 4180 individuals in total who carried 4840 mutant alleles, and identified 186 couples at risk of having affected offspring. 12.1% of the pathogenic or likely pathogenic variants identified by our NGS assay, which were undetectable by traditional methods. Compared with the traditional methods, our assay identified an additional at-risk 35 couples. We describe a comprehensive NGS-based test that offers advantages over the traditional screening/molecular testing methods. To our knowledge, this is among the first large-scale population study to systematically evaluate the application of an NGS technique in carrier screening and molecular diagnosis of hemoglobinopathies.
Asunto(s)
Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Pruebas Genéticas , Técnicas de Genotipaje , Hemoglobinopatías/diagnóstico , Hemoglobinopatías/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Adolescente , Adulto , Estudios de Casos y Controles , China/epidemiología , Índices de Eritrocitos , Estudios de Asociación Genética/métodos , Tamización de Portadores Genéticos , Pruebas Genéticas/métodos , Variación Genética , Genotipo , Geografía Médica , Hemoglobinopatías/epidemiología , Hemoglobinas Anormales/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Persona de Mediana Edad , Fenotipo , Polimorfismo de Nucleótido Simple , Vigilancia de la Población , Prevalencia , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Índice de Severidad de la Enfermedad , Adulto JovenRESUMEN
To rationally guide the improvement of isobutanol production, metabolic network and metabolic profiling analysis were performed to provide global and profound insights into cell metabolism of isobutanol-producing Bacillus subtilis. The metabolic flux distribution of strains with different isobutanol production capacity (BSUL03, BSUL04 and BSUL05) drops a hint of the importance of NADPH on isobutanol biosynthesis. Therefore, the redox pathways were redesigned in this study. To increase NADPH concentration, glucose-6-phosphate isomerase was inactivated (BSUL06) and glucose-6-phosphate dehydrogenase was overexpressed (BSUL07) successively. As expected, NADPH pool size in BSUL07 was 4.4-fold higher than that in parental strain BSUL05. However, cell growth, isobutanol yield and production were decreased by 46%, 22%, and 80%, respectively. Metabolic profiling analysis suggested that the severely imbalanced redox status might be the primary reason. To solve this problem, gene udhA of Escherichia coli encoding transhydrogenase was further overexpressed (BSUL08), which not only well balanced the cellular ratio of NAD(P)H/NAD(P)+, but also increased NADH and ATP concentration. In addition, a straightforward engineering approach for improving NADPH concentrations was employed in BSUL05 by overexpressing exogenous gene pntAB and obtained BSUL09. The performance for isobutanol production by BSUL09 was poorer than BSUL08 but better than other engineered strains. Furthermore, in fed-batch fermentation the isobutanol production and yield of BSUL08 increased by 11% and 19%, up to the value of 6.12 g/L and 0.37 C-mol isobutanol/C-mol glucose (63% of the theoretical value), respectively, compared with parental strain BSUL05. These results demonstrated that model-driven complemented with metabolic profiling analysis could serve as a useful approach in the strain improvement for higher bio-productivity in further application.