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The development of CRISPR-based barcoding methods creates an exciting opportunity to understand cellular phylogenies. We present a compact, tunable, high-capacity Cas12a barcoding system called dual acting inverted site array (DAISY). We combined high-throughput screening and machine learning to predict and optimize the 60-bp DAISY barcode sequences. After optimization, top-performing barcodes had â¼10-fold increased capacity relative to the best random-screened designs and performed reliably across diverse cell types. DAISY barcode arrays generated â¼12 bits of entropy and â¼66,000 unique barcodes. Thus, DAISY barcodes-at a fraction of the size of Cas9 barcodes-achieved high-capacity barcoding. We coupled DAISY barcoding with single-cell RNA-seq to recover lineages and gene expression profiles from â¼47,000 human melanoma cells. A single DAISY barcode recovered up to â¼700 lineages from one parental cell. This analysis revealed heritable single-cell gene expression and potential epigenetic modulation of memory gene transcription. Overall, Cas12a DAISY barcoding is an efficient tool for investigating cell-state dynamics.
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Sistemas CRISPR-Cas , Código de Barras del ADN Taxonómico , Linaje de la Célula/genética , Código de Barras del ADN Taxonómico/métodos , Humanos , Aprendizaje Automático , FilogeniaRESUMEN
Gastric cancer (GC) is one of the most common malignant tumors in the world, and current treatments are still based on surgery and drug therapy. However, due to the complexity of immunosuppression and drug resistance, the treatment of gastric cancer still faces great challenges. Chemokine receptor 2 (CXCR2) is one of the most common therapeutic targets in targeted therapy. As a G protein-coupled receptor, CXCR2 and its ligands play important roles in tumorigenesis and progression. The abnormal expression of these genes in cancer plays a decisive role in the recruitment and activation of white blood cells, angiogenesis, and cancer cell proliferation, and CXCR2 is involved in various stages of tumor development. Therefore, interfering with the interaction between CXCR2 and its ligands is considered a possible target for the treatment of various tumors, including gastric cancer.
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Receptores de Interleucina-8B , Neoplasias Gástricas , Humanos , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/tratamiento farmacológico , Receptores de Interleucina-8B/metabolismo , Animales , Terapia Molecular Dirigida/métodos , Transducción de Señal , Proliferación CelularRESUMEN
BACKGROUND: Myeloid differentiation factor-88 (MyD88) is a crucial adapter protein that coordinates the innate immune response and establishes an adaptive immune response. The interaction of the Toll/Interleukin-1 receptor (IL-1R) superfamily with MyD88 triggers the activation of various signalling pathways such as nuclear factor-κB (NF-κB) and activator protein-1 (AP-1), promoting the production of a variety of immune and inflammatory mediators and potentially driving the development of a variety of diseases. OBJECTIVE: This article will explore the therapeutic potential and mechanism of the MyD88-specific inhibitor ST2825 and describe its use in the treatment of several diseases. We envision future research and clinical applications of ST2825 to provide new ideas for the development of anti-inflammatory drugs and disease-specific drugs to open new horizons for the prevention and treatment of related inflammatory diseases. MATERIALS AND METHODS: This review analysed relevant literature in PubMed and other databases. All relevant studies on MyD88 inhibitors and ST2825 that were published in the last 20 years were used as screening criteria. These studies looked at the development and improvement of MyD88 inhibitors and ST2825. RESULTS: Recent evidence using the small-molecule inhibitor of ST2825 has suggested that blocking MyD88 activity can be used to treat diseases such as neuroinflammation, inflammatory diseases such as acute liver/kidney injury, or autoimmune diseases such as systemic lupus erythematosus and can affect transplantation immunity. In addition, ST2825 has potential therapeutic value in B-cell lymphoma with the MyD88 L265P mutation. CONCLUSION: Targeting MyD88 is a novel therapeutic strategy, and scientific research is presently focused on the development of MyD88 inhibitors. The peptidomimetic compound ST2825 is a widely studied small-molecule inhibitor of MyD88. Thus, ST2825 may be a potential therapeutic small-molecule agent for modulating host immune regulation in inflammatory diseases and inflammatory therapy.
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Factor 88 de Diferenciación Mieloide , FN-kappa B , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , FN-kappa B/metabolismo , Transducción de Señal , Antiinflamatorios/uso terapéutico , Antiinflamatorios/farmacologíaRESUMEN
Several existing technologies enable short genomic alterations including generating indels and short nucleotide variants, however, engineering more significant genomic changes is more challenging due to reduced efficiency and precision. Here, we developed RecT Editor via Designer-Cas9-Initiated Targeting (REDIT), which leverages phage single-stranded DNA-annealing proteins (SSAP) RecT for mammalian genome engineering. Relative to Cas9-mediated homology-directed repair (HDR), REDIT yielded up to a 5-fold increase of efficiency to insert kilobase-scale exogenous sequences at defined genomic regions. We validated our REDIT approach using different formats and lengths of knock-in templates. We further demonstrated that REDIT tools using Cas9 nickase have efficient gene-editing activities and reduced off-target errors, measured using a combination of targeted sequencing, genome-wide indel, and insertion mapping assays. Our experiments inhibiting repair enzyme activities suggested that REDIT has the potential to overcome limitations of endogenous DNA repair steps. Finally, our REDIT method is applicable across cell types including human stem cells, and is generalizable to different Cas9 enzymes.
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Proteína 9 Asociada a CRISPR , Proteínas de Unión al ADN , Proteínas de Escherichia coli , Edición Génica/métodos , Línea Celular , Genoma , Humanos , Reparación del ADN por Recombinación , Células Madre/metabolismoRESUMEN
Nitrogen-doped carbon dots (NCDs) have been constructed in which coal washing wastewater is used as carbon precursor, tryptophan is added for nitrogen doping and surface functional together with polyethylene glycol. The nitrogen doping and surface functional with electron rich groups resulted in excellent fluorescent properties regarding stability, reversibility, printability with high quantum yield which not only enable the NCDs as fluorescent ink for advanced message encryption, but also realize specific on-off-on fluorescent sensing of Hg2+ and GSH as solution, hydrogel and filter paper sensors. The NCDs had a linear range of 0.01-100 µM and a detection limit of 6.27 nM (RSD 0.33%) for Hg2+ and the NCDs@Hg2+ had a linear range of 0.01-60 µM and a detection limit of 3.53 nM (RSD 1.53%) for GSH in sensing studies with aqueous solutions. In addition, with the low cytotoxicity and good biocompatibility NCDs have been successfully used for imaging Hg2+ and GSH in living MG-63 cells. The presented NCDs recycle waste coal washing water into worthwhile material which can be implemented as promising anti-counterfeiting and message encryption candidates as well as effective Hg2+ and GSH sensing, tracking and removing tools in complicated environmental and biological systems.
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Mercurio , Puntos Cuánticos , Carbono , Colorantes Fluorescentes , Glutatión , Mercurio/análisis , NitrógenoRESUMEN
Epstein Barr-virus (EBV) is related to several cancers. Long non-coding RNAs (lncRNAs) act by regulating target genes and are involved in tumourigenesis. However, the role of lncRNAs in EBV-associated cancers is rarely reported. Understanding the role and mechanism of lncRNAs in EBV-associated cancers may contribute to diagnosis, prognosis and clinical therapy in the future. EBV encodes not only miRNAs, but also BART lncRNAs during latency and the BHLF1 lncRNA during both the latent and lytic phases. These lncRNAs can be targeted regulate inflammation, invasion, and migration and thus tumourigenesis. The products of EBV also directly and indirectly regulate host lncRNAs, including LINC00312, NORAD CYTOR, SHNG8, SHNG5, MINCR, lncRNA-BC200, LINC00672, MALATI1, LINC00982, LINC02067, IGFBP7-AS1, LOC100505716, LOC100128494, NAG7 and RP4-794H19.1, to facilitate tumourigenesis using different mechanisms. Additionally, lncRNAs have been previously validated to interact with microRNAs (miRNAs), and lncRNAs and miRNAs mutually suppress each other. The EBV-miR-BART6-3p/LOC553103/STMN1 axis inhibits EBV-associated tumour cell proliferation. Additionally, H. pylori-EBV co-infection promotes inflammatory lesions and results in EMT. HPV-EBV co-infection inhibits the transition from latency to lytic replication. KSHV-EBV co-infection aggravates tumourigenesis in huNSG mice. COVID-19-EBV co-infection may activate the immune system to destroy a tumour, although this situation is rare and the mechanism requires further confirmation. Hopefully, this information will shed some light on tumour therapy strategies tumourigenesis. Additionally, this strategy benefits for infected patients by preventing latency to lytic replication. Understanding the role and expression of lnRNAs in these two phases of EBV is critical to control the transition from latency to the lytic replication phase. This review presents differential expressed lncRNAs in EBV-associated cancers and provides resources to aid in developing superior strategies for clinical therapy.
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Epstein-Barr virus (EBV) is closely associated with multiple human cancers. EBV-associated cancers are mainly lymphomas derived from B cells and T cells (Hodgkin lymphoma, Burkitt lymphoma, NK/T-cell lymphoma, and posttransplant lymphoproliferative disorder (PTLD)) and carcinomas derived from epithelial cells (nasopharyngeal carcinoma and gastric carcinoma). EBV can induce oncogenesis in its host cell by activating various signaling pathways, such as nuclear factor-κB (NF-κB), phosphoinositide-3-kinase/protein kinase B (PI3K/AKT), Janus kinase/signal transducer and transcription activator (JAK/STAT), mitogen-activated protein kinase (MAPK), transforming growth factor-ß (TGF-ß), and Wnt/ß-catenin, which are regulated by EBV-encoded proteins and noncoding RNA. In this review, we focus on the oncogenic roles of EBV that are mediated through the aforementioned signaling pathways.
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Anxiety disorders are the most prevalent psychiatric disorders, but their pathogenic mechanism remains poorly understood. Here, we report that transmembrane protein 74 (TMEM74), which contains two putative transmembrane domains and exhibits high levels of mRNA in the brain, is closely associated with the pathogenesis of anxiety disorders. TMEM74 was decreased in the serum of patients with anxiety and the basolateral amygdaloid nucleus (BLA) in chronic stress mice. Furthermore, genetic deletion of Tmem74 or selective knockdown of Tmem74 in BLA pyramidal neurons resulted in anxiety-like behaviors in mice. Whole-cell recordings in BLA pyramidal neurons revealed lower hyperpolarization-activated cation current (Ih) and greater input resistance and excitability in Tmem74-/- neurons than in wild-type neurons. Accordingly, surface expression of hyperpolarization-activated cyclic nucleotide-gated 1 (HCN1) channels was also lower in the BLA of Tmem74-/- mice. The Ih current blocker ZD7288 mimicked these effects in BLA pyramidal neurons in wild-type mice but not in Tmem74-/- mice. Consistent with the improvement in anxiety-like behaviors, Tmem74 overexpression restored HCN1 channel trafficking and pyramidal neuron excitability in the BLA of Tmem74-/- and chronic stress mice. Mechanistically, we demonstrate that interactions between Tmem74 and HCN1 are physiologically relevant and that transmembrane domain 1 (TM1) is essential for the cellular membrane localization of Tmem74 to enhance Ih. Together, our findings suggest that Tmem74 coupling with HCN1 acts as a critical component in the pathophysiology of anxiety and is a potential target for new treatments of anxiety disorders.
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Ansiedad/metabolismo , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/metabolismo , Proteínas de la Membrana/metabolismo , Animales , Ansiedad/genética , Trastornos de Ansiedad/genética , Trastornos de Ansiedad/metabolismo , Complejo Nuclear Basolateral/metabolismo , Encéfalo/metabolismo , Canales Catiónicos Regulados por Nucleótidos Cíclicos , Hipocampo/metabolismo , Humanos , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/genética , Potenciales de la Membrana/fisiología , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/metabolismo , Técnicas de Placa-Clamp , Canales de Potasio/genética , Transporte de Proteínas , Células Piramidales/metabolismoRESUMEN
Grb2-associated-binding protein 1 (Gab1) is a docking/scaffolding molecule known to play an important role in cell growth and survival. Here, we report that Gab1 is decreased in cholinergic neurons in Alzheimer's disease (AD) patients and in a mouse model of AD. In mice, selective ablation of Gab1 in cholinergic neurons in the medial septum impaired learning and memory and hippocampal long-term potentiation. Gab1 ablation also inhibited SK channels, leading to an increase in firing in septal cholinergic neurons. Gab1 overexpression, on the other hand, improved cognitive function and restored hippocampal CaMKII autorphosphorylation in AD mice. These results suggest that Gab1 plays an important role in the pathophysiology of AD and may represent a novel therapeutic target for diseases involving cholinergic dysfunction.
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Enfermedad de Alzheimer/patología , Corteza Cerebral/patología , Neuronas Colinérgicas/fisiología , Cognición/fisiología , Regulación de la Expresión Génica/genética , Fosfoproteínas/metabolismo , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/genética , Proteínas Adaptadoras Transductoras de Señales , Anciano de 80 o más Años , Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/diagnóstico por imagen , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animales , Encéfalo/citología , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/genética , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Colina O-Acetiltransferasa/genética , Colina O-Acetiltransferasa/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Ratones Transgénicos , Persona de Mediana Edad , Mutación/genética , Fosfoproteínas/genética , Presenilina-1/genética , Presenilina-1/metabolismoRESUMEN
Severe hypoglycemia has a detrimental impact on the cerebrovasculature, but the molecular events that lead to the disruption of the integrity of the tight junctions remain unclear. Here, we report that the microvessel integrity was dramatically compromised (59.41% of wild-type mice) in TP53-induced glycolysis and apoptosis regulator (TIGAR) transgenic mice stressed by hypoglycemia. Melatonin, a potent antioxidant, protects against hypoglycemic stress-induced brain endothelial tight junction injury in the dosage of 400 nmol/L in vitro. FRET (fluorescence resonance energy transfer) imaging data of endothelial cells stressed by low glucose revealed that TIGAR couples with calmodulin to promote TIGAR tyrosine nitration. A tyrosine 92 mutation interferes with the TIGAR-dependent NADPH generation (55.60% decreased) and abolishes its protective effect on tight junctions in human brain microvascular endothelial cells. We further demonstrate that the low-glucose-induced disruption of occludin and Caludin5 as well as activation of autophagy was abrogated by melatonin-mediated blockade of nitrosative stress in vitro. Collectively, we provide information on the detailed molecular mechanisms for the protective actions of melatonin on brain endothelial tight junctions and suggest that this indole has translational potential for severe hypoglycemia-induced neurovascular damage.
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Antioxidantes/farmacología , Células Endoteliales/efectos de los fármacos , Melatonina/farmacología , Proteínas/metabolismo , Uniones Estrechas/efectos de los fármacos , Animales , Proteínas Reguladoras de la Apoptosis , Barrera Hematoencefálica/efectos de los fármacos , Barrera Hematoencefálica/metabolismo , Línea Celular , Humanos , Hipoglucemia/complicaciones , Péptidos y Proteínas de Señalización Intracelular/efectos de los fármacos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Monoéster Fosfórico Hidrolasas , Proteínas/efectos de los fármacos , Uniones Estrechas/metabolismoRESUMEN
Epstein-Barr virus (EBV) is a human oncogenic herpesvirus associated with lymphoma and nasopharyngeal carcinoma. Because the susceptible hosts of EB virus are limited to human and cotton-top tamarins (Saguinus oedipus), there have been no appropriate animal models until the lymphoma model induced by EBV in human peripheral blood lymphocyte (hu-PBL)/SCID chimeric mice was reported. However, it is still controversial whether the EBV-associated lymphoma induced in hu-PBL/SCID mice is a monoclonal tumor. In this study, we transplanted normal human peripheral blood lymphocytes (hu-PBL) from six donors infected with EBV into SCID mice to construct hu-PBL/SCID chimeric mice. The induced tumors were found in the mediastinum or abdominal cavity of SCID mice. Microscopic observation exhibited tumor cells that were large and had a plasmablastic, centroblastic or immunoblastic-like appearance. Immunophenotyping assays showed the induced tumors were LCA-positive, CD20/CD79a-positive (markers of B cells), and CD3/CD45RO-negative (markers of T cells). A human-specific Alu sequence could be amplified by Alu-PCR. This confirmed that induced tumors were B-cell lymphomas originating from the transplanted human lymphocytes rather than mouse cells. EBER in situ hybridization detected positive signals in the nuclei of the tumor cells. Expression of EBV-encoded LMP1, EBNA-1, and EBNA-2 in the tumors was significantly positive. PCR-based capillary electrophoresis analysis of IgH gene rearrangement revealed a monoclonal peak and single amplification product in all six cases of induced tumors. This indicated that EBV can induce monoclonal proliferation of human B lymphocytes and promotes the development of lymphoma. J. Med. Virol. 88:1804-1813, 2016. © 2016 Wiley Periodicals, Inc.
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Infecciones por Virus de Epstein-Barr/complicaciones , Reordenamiento Génico , Cadenas Pesadas de Inmunoglobulina/genética , Inmunofenotipificación , Linfoma de Células B/virología , Elementos Alu , Animales , Modelos Animales de Enfermedad , Antígenos Nucleares del Virus de Epstein-Barr/genética , Herpesvirus Humano 4 , Humanos , Hibridación in Situ , Transfusión de Linfocitos , Ratones SCID , Proteínas de la Matriz Viral/genética , Proteínas Virales/genéticaRESUMEN
BACKGROUND: Chemoresistance is a major obstacle in successfully treating cancers, and the mechanisms responsible for drug resistance are still far from understood. Carbonic anhydrase 9 (CA9) has been shown to be upregulated in the drug-resistant tongue cancer cell line Tca8113/PYM and to be associated with drug resistance. However, the mechanisms regulating CA9 expression and its role in drug resistance remain unclear. METHODS: Bioinformatic and experimental analysis involving ChIP and luciferase reporter assays were used to validate Zinc finger E-box-binding homeobox 1 (ZEB1) as a transcriptional regulator of CA9. Gene expression and protein levels were evaluated by quantitative RT-PCR and western blotting, respectively. Sensitivity to chemotherapy was examined using the MTS assay and Hoechst staining and analysis caspase-3 activity to evaluate changes in apoptosis. Intracellular pH (pHi) was measured using fluorescent pH-indicator BCECF-AM. Protein expression in patient tissue samples was examined by immunohistochemistry and survival of tongue cancer patients from which these samples were derived was also analyzed. RESULTS: ZEB1 bound to the promoter of CA9 to positively regulate CA9 expression in tongue cancer cells. Knockdown of CA9 using short interfering RNA (siRNA) abolished the chemoresistance resulting from ZEB1 overexpression in Tca8113 and SCC-25 cells, and CA9 overexpression attenuated chemosensitivity induced by ZEB1 knockdown in Tca8113/PYM cells. CA9 knockdown also prevented maintenance of pHi mediated by overexpression of ZEB1 in Tca8113 and SCC-25 cells following chemotherapy, associated with increased apoptosis and caspase-3 activation. Conversely, ectopic expression of CA9 suppressed decrease in pHi mediated by ZEB1 knockdown in Tca8113/PYM cells following chemotherapy, accompanied by decreased apoptosis and caspase-3 activation. Importantly, a positive correlation was observed between ZEB1 and CA9 protein expression in tongue cancer tissues, and expression of these proteins associated with a poor prognosis for patients. CONCLUSION: Our finding that tumor cells regulate pHi in response to chemotherapy provides new insights into mechanisms of drug resistance during cancer treatment. Identification of the ZEB1-CA9 signaling axis as a biomarker of poor prognosis in tongue cancer will be valuable in future development of therapeutic strategies aimed at improving treatment efficacy, especially in terms of drug resistance associated with this disease.
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Antígenos de Neoplasias/genética , Anhidrasas Carbónicas/genética , Resistencia a Antineoplásicos/genética , Proteínas de Homeodominio/genética , Neoplasias de la Lengua/genética , Factores de Transcripción/genética , Transcripción Genética/genética , Apoptosis/genética , Anhidrasa Carbónica IX , Caspasa 3/genética , Línea Celular , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/genética , Células HEK293 , Humanos , Concentración de Iones de Hidrógeno , ARN Interferente Pequeño/genética , Homeobox 1 de Unión a la E-Box con Dedos de ZincRESUMEN
Cisplatin (DDP) is the first-line chemotherapy drug widely used for the treatment of lung cancer patients, whereas the majority of cancer patients will eventually show resistance to DDP. The mechanisms responsible for DDP resistance are not fully understood. Tongue cancer resistance-associated protein 1 (TCRP1) gene was recently cloned and reported to specially mediate DDP resistance in human oral squamous cell carcinoma (OSCC) cells. However, the mechanisms of TCRP1-mediated DDP resistance are far from clear, and whether TCRP1 participates in DDP resistance in lung cancer cells remains unknown. Here, we show that TCRP1 contributes to DDP resistance in lung cancer cells. Knockdown of TCRP1 sensitizes the cells to DDP and increases the DDP-induced DNA damage. We have identified that Pol ß is associated with DDP resistance, and Pol ß knockdown delays the repair of DDP-induced DNA damage in A549/DDP cells. We find TCRP1 interacts with Pol ß in lung cancer cells. Moreover, TCRP1 knockdown decreases the level of Pol ß and increases the level of its ubiquitination. These results suggest that TCRP1 contributes to DDP resistance through the prevention of Pol ß degradation in lung cancer cells. These findings provide new insights into chemoresistance and may contribute to prevention and reversal of DDP resistance in treatment of lung cancer in the future.
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Cisplatino/farmacología , ADN Polimerasa beta/metabolismo , Resistencia a Antineoplásicos , Proteínas/metabolismo , Antineoplásicos/farmacología , Western Blotting , Línea Celular Tumoral , Daño del ADN , ADN Polimerasa beta/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Microscopía Fluorescente , Unión Proteica , Proteínas/genética , Proteolisis , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , UbiquitinaciónRESUMEN
A sampling campaign was conducted monthly to investigate the occurrence of N-nitrosamines at a conventional water treatment plant in one city in North China. The yield of N-nitrosamines in the treated water indicated precursors changed greatly after the source water switching. Average concentrations of N-nitrosodimethylamine (NDMA), N-nitrosomorpholine (NMOR), and N-nitrosopyrrolidine (NPYR) in the finished water were 6.9, 3.3, and 3.1ng/L, respectively, from June to October when the Luan River water was used as source water, while those of NDMA, N-nitrosomethylethylamine (NMEA), and NPYR in the finished water were 10.1, 4.9, and 4.7ng/L, respectively, from November to next April when the Yellow River was used. NDMA concentration in the finished water was frequently over the 10ng/L, i.e., the notification level of California, USA, which indicated a considerable threat to public health. Weak correlations were observed between N-nitrosamine yield and typical water quality parameters except for the dissolved organic nitrogen.
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Contaminantes Químicos del Agua/análisis , Purificación del Agua , China , Dimetilnitrosamina/análogos & derivados , Dimetilnitrosamina/análisis , Nitrosaminas/análisisRESUMEN
Cisplatin is one of the major chemotherapeutic agents used against different human cancers. A better understanding of the downstream cellular targets of cisplatin will provide information on its mechanism of action. FOXO3a is a member of the FOXO transcription factor family, which modulates the expression of genes involved in cell cycle arrest, apoptosis, and other cellular processes. In this study, we have investigated the effects of cisplatin in a panel of lung cancer cell lines. The results showed that cisplatin inhibited the proliferation of these lung cancer cell lines by inhibiting the PI3K/AKT pathway, with evidence of decreasing phosphorylation of PI3K and AKT under cisplatin treatment, and constitutively activating AKT1 could reduce cisplatin-induced cell apoptosis. More importantly, cisplatin significantly inhibited FOXO3a phosphorylation (at Thr32, AKT phosphorylation site) and induced FOXO3a nuclear accumulation, which in turn increased the expression of FOXO3a-dependent apoptotic protein Bim. Knockdown of FOXO3a expression using small interfering RNA attenuated cisplatin-induced apoptosis. Furthermore, activation of FOXO3a induced cell apoptosis irrespective of p53 status, whereas p53 may act as FOXO3a downstream molecules involved in cisplatin-induced cell apoptosis. Together, our findings suggested that FOXO3a is a relevant mediator of the cytotoxic effects of cisplatin in lung cancer cells.
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Antineoplásicos/farmacología , Cisplatino/farmacología , Factores de Transcripción Forkhead/metabolismo , Neoplasias Pulmonares/patología , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/metabolismo , Proteína 11 Similar a Bcl2 , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Proteína Forkhead Box O3 , Factores de Transcripción Forkhead/genética , Humanos , Proteínas de la Membrana/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Transporte de Proteínas , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Treonina/metabolismo , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
This paper proposes an introspective deep metric learning (IDML) framework for uncertainty-aware comparisons of images. Conventional deep metric learning methods focus on learning a discriminative embedding to describe the semantic features of images, which ignore the existence of uncertainty in each image resulting from noise or semantic ambiguity. Training without awareness of these uncertainties causes the model to overfit the annotated labels during training and produce overconfident judgments during inference. Motivated by this, we argue that a good similarity model should consider the semantic discrepancies with awareness of the uncertainty to better deal with ambiguous images for more robust training. To achieve this, we propose to represent an image using not only a semantic embedding but also an accompanying uncertainty embedding, which describes the semantic characteristics and ambiguity of an image, respectively. We further propose an introspective similarity metric to make similarity judgments between images considering both their semantic differences and ambiguities. The gradient analysis of the proposed metric shows that it enables the model to learn at an adaptive and slower pace to deal with the uncertainty during training. Our framework attains state-of-the-art performance on the widely used CUB-200-2011, Cars196, and Stanford Online Products datasets for image retrieval. We further evaluate our framework for image classification on the ImageNet-1 K, CIFAR-10, and CIFAR-100 datasets, which shows that equipping existing data mixing methods with the proposed introspective metric consistently achieves better results (e.g., +0.44% for CutMix on ImageNet-1 K).
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The deficiency in available targeted agents and frequency of chemoresistance are primary challenges in clinical management of triple-negative breast cancer (TNBC). The aberrant expression of USP21 and JAK2 represents a characterized mechanism of TNBC progression and resistance to paclitaxel (PTX). Despite its clear that high expression of USP21-mediated de-ubiquitination leads to increased levels of JAK2 protein, we lack regulator molecules to dissect the mechanisms that the interaction between USP21 and JAK2 contributes to the phenotype and resistance of TNBC. Here, we report a USP21/JAK2/STAT3 axis-targeting regulator 13c featuring a N-anthraniloyl tryptamine scaffold that showed excellent anti-TNBC potency and promising safety profile. Importantly, the therapeutic potential of using 13c in combination with PTX in PTX-resistant TNBC was demonstrated. This study showcases N-anthraniloyl tryptamine derivatives as a novel anti-TNBC chemotype with a pharmacological mode of action targeting the USP21/JAK2/STAT3 axis and provides a potential therapeutic target for the treatment of TNBC.
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Antineoplásicos , Janus Quinasa 2 , Factor de Transcripción STAT3 , Neoplasias de la Mama Triple Negativas , Ubiquitina Tiolesterasa , Humanos , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/patología , Neoplasias de la Mama Triple Negativas/metabolismo , Factor de Transcripción STAT3/metabolismo , Factor de Transcripción STAT3/antagonistas & inhibidores , Janus Quinasa 2/metabolismo , Janus Quinasa 2/antagonistas & inhibidores , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/síntesis química , Ubiquitina Tiolesterasa/antagonistas & inhibidores , Ubiquitina Tiolesterasa/metabolismo , Relación Estructura-Actividad , Proliferación Celular/efectos de los fármacos , Animales , Descubrimiento de Drogas , Estructura Molecular , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Bibliotecas de Moléculas Pequeñas/síntesis química , Femenino , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Línea Celular Tumoral , Ratones , Paclitaxel/farmacología , Paclitaxel/químicaRESUMEN
The colonization of microbes in the gut is key to establishing a healthy host-microbiome symbiosis for newborns. We longitudinally profiled the gut microbiome in a model consisting of 36 neonatal oxen from birth up to 2 months postpartum and carried out microbial transplantation to reshape their gut microbiome. Genomic reconstruction of deeply sequenced fecal samples resulted in a total of 3931 metagenomic-assembled genomes from 472 representative species, of which 184 were identified as new species when compared with existing databases of oxen. Single nucleotide level metagenomic profiling shows a rapid influx of microbes after birth, followed by dynamic shifts during the first few weeks of life. Microbial transplantation was found to reshape the genetic makeup of 33 metagenomic-assembled genomes (FDR < 0.05), mainly from Prevotella and Bacteroides species. We further linked over 20 million microbial single nucleotide variations to 736 plasma metabolites, which enabled us to characterize 24 study-wide significant associations (P < 4.4 × 10-9) that identify the potential microbial genetic regulation of host immune and neuro-related metabolites, including glutathione and L-dopa. Our integration analyses further revealed that microbial genetic variations may influence the health status and growth performance by modulating metabolites via structural regulation of their encoded proteins. For instance, we found that the albumin levels and total antioxidant capacity were correlated with L-dopa, which was determined by single nucleotide variations via structural regulations of metabolic enzymes. The current results indicate that temporal colonization and transplantation-driven strain replacement are crucial for newborn gut development, offering insights for enhancing newborn health and growth.
Asunto(s)
Microbioma Gastrointestinal , Microbiota , Recién Nacido , Humanos , Femenino , Microbioma Gastrointestinal/fisiología , Nucleótidos , Levodopa , Heces , Metagenómica/métodosRESUMEN
Genistein (GEN) is a molecule of great interest as a potent chemopreventive agent against atherosclerosis and cancer. However, the bioavailability of GEN is very low in vivo. Our previous study showed that a GEN derivative, 7-difluoromethyl-5,4'-dimethoxygenistein (dFMGEN) has a better bioavailability than GEN in vivo. In this study, we further evaluated the efficacy of dFMGEN as a candidate for cancer therapy. We demonstrated that dFMGEN treatment decreased the viability of A549 cells in a concentration- and time-dependent manner and induced cell-cycle arrest at the G(1) phase. G(1) phase arrest was correlated with a significant reduction of Cdk4 and cyclin D1 protein level. Further studies showed that cyclin-dependent kinase (Cdk)4 and cyclin D1 protein-level decrease was caused by Cdk inhibitors p15, p21, and p27 level increase, and decreased protein level directly suppressed Rb protein phosphorylation and E2F-1 expression, then cell-cycle progression was arrested. Finally, we also found that dFMGEN has a dosage effect in suppressing tumor growth in vivo, and that dFMGEN was well tolerated by animals. In summary, our results suggest that dFMGEN has therapeutic potential for the treatment of human lung cancer.