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The cytosolic DNA sensor cGMP-AMP synthase (cGAS) synthesizes the noncanonical cyclic dinucleotide 2'3'-cGAMP to activate the adaptor protein stimulator of IFN genes (STING), thus awakening host immunity in response to DNA pathogen infection. However, dengue virus (DENV), an RNA virus without a DNA stage in its life cycle, also manipulates cGAS-STING-mediated innate immunity by proteolytic degradation of STING. Here, we found that the sensitivity of STING to DENV protease varied with different human STING haplotypes. Exogenous DNA further enhanced DENV protease's ability to interact and cleave protease-sensitive STING. DNA-enhanced STING cleavage was reduced in cGAS-knockdown cells and triggered by the cGAS product 2'3'-cGAMP. The source of DNA may not be endogenous mitochondrial DNA but rather exogenous reactivated viral DNA. Cells producing 2'3'-cGAMP by overexpressing cGAS or with DNA virus reactivation enhanced STING cleavage in neighboring cells harboring DENV protease. DENV infection reduced host innate immunity in cells with the protease-sensitive STING haplotype, whose homozygote genotype frequency was found significantly reduced in Taiwanese people with dengue fever. Therefore, the human STING genetic background and DNA pathogen coinfection may be the missing links contributing to DENV pathogenesis.
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Dengue/enzimología , Endopeptidasas/metabolismo , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Nucleótidos Cíclicos/metabolismo , Células A549 , ADN Viral/genética , Dengue/inmunología , Endopeptidasas/genética , Haplotipos , Humanos , Evasión Inmune , Inmunidad Innata , Nucleótidos Cíclicos/genéticaRESUMEN
The SARS-CoV-2 pseudovirus is a commonly used strategy that mimics certain biological functions of the authentic virus by relying on biological legitimacy at the molecular level. Despite the fact that spike (S), envelope (E), and membrane (M) proteins together wrap up the SARS-CoV-2 virion, most of the reported pseudotype viruses consist of only the S protein. Here, we report that the presence of E and M increased the virion infectivity by promoting the S protein priming. The S, E, and M (SEM)-coated pseudovirion is spherical, containing crown-like spikes on the surface. Both S and SEM pseudoviruses packaged the same amounts of viral RNA, but the SEM virus bound more efficiently to cells stably expressing the viral receptor human angiotensin-converting enzyme II (hACE2) and became more infectious. Using this SEM pseudovirus, we examined the infectivity and antigenic properties of the natural SARS-CoV-2 variants. We showed that some variants have higher infectivity than the original virus and that some render the neutralizing plasma with lower potency. These studies thus revealed possible mechanisms of the dissemination advantage of these variants. Hence, the SEM pseudovirion provides a useful tool to evaluate the viral infectivity and capability of convalescent sera in neutralizing specific SARS-CoV-2 S dominant variants.
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Anticuerpos Antivirales/metabolismo , COVID-19/inmunología , Proteínas de la Envoltura de Coronavirus/metabolismo , SARS-CoV-2/patogenicidad , Proteínas de la Matriz Viral/metabolismo , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , Anticuerpos Antivirales/inmunología , COVID-19/sangre , COVID-19/virología , Línea Celular , Proteínas de la Envoltura de Coronavirus/genética , Proteínas de la Envoltura de Coronavirus/inmunología , Proteínas de la Envoltura de Coronavirus/ultraestructura , Cricetinae , Humanos , Microscopía Electrónica de Transmisión , Mutación , Pruebas de Neutralización , SARS-CoV-2/genética , SARS-CoV-2/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/metabolismo , Proteínas de la Matriz Viral/genética , Proteínas de la Matriz Viral/inmunología , Proteínas de la Matriz Viral/ultraestructura , Virión/genética , Virión/inmunología , Virión/metabolismo , Virión/ultraestructuraRESUMEN
OBJECTIVES: To analyze and present the demography, clinical behavior, especially the risk factors of tumor hemorrhage and management of sporadic angiomyolipoma (SAML), tuberous sclerosis complex associated angiomyolipoma (TSCAML) and epithelioid angiomyolipoma (EAML) in our institution. METHODS: A retrospective study of 587 patients who were diagnosed with renal angiomyolipoma in our institution between January 2000 and May 2015 was done. The AMLs were diagnosed by ultrasonography, CT, or MRI. EAML was confirmed by histopathology. Medical records and follow-up results were analyzed using the SPSS version 22 software. RESULTS: Out of 587 cases of renal AMLs, 87.4% were SAMLs, 8.7% were TSCAMLs and 3.9% were EAMLs. Most of the AML patients were asymptomatic. The most common presenting symptoms included flank pain and abdominal pain. The median tumor size of SAML, TSCAML, EAML were 4.7, 2.7, 10.5 cm respectively. Approximately half of SAMLs were conservatively treated, almost all TSCAMLs were treated conservatively, while all EAMLs were surgically treated. The median tumor size of hemorrhagic SAML cases was 8 cm versus non-hemorrhagic cases of 4.1 cm. The optimal cut-off point on the ROC curve for predicting SAML tumor hemorrhage was 7.35 cm. CONCLUSION: A larger tumor size, younger patient's age and higher BMI value correlated with a higher risk of tumor hemorrhage. For tumor sizes less than 7.35 cm, we recommend active surveillance or TAE for hemorrhage prevention. We also suggest that surgical management should be considered for patients with tumors larger than 7.35 cm, symptomatic and progressive AML, or suspicious EAML.
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Angiomiolipoma/clasificación , Angiomiolipoma/terapia , Neoplasias Renales/clasificación , Neoplasias Renales/terapia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Angiomiolipoma/patología , Niño , Preescolar , Diagnóstico Diferencial , Embolización Terapéutica , Femenino , Hemorragia/epidemiología , Hemorragia/terapia , Humanos , Lactante , Riñón/diagnóstico por imagen , Riñón/patología , Neoplasias Renales/patología , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Curva ROC , Estudios Retrospectivos , Taiwán , Tomografía Computarizada por Rayos X , Esclerosis Tuberosa/complicaciones , Ultrasonografía , Adulto JovenRESUMEN
Infection with Japanese encephalitis virus (JEV) can induce the expression of pro-inflammatory cytokines and cause acute encephalitis in humans. ß-oxidation breaks down fatty acids for ATP production in mitochondria, and impaired ß-oxidation can induce pro-inflammatory cytokine expression. To address the role of fatty-acid ß-oxidation in JEV infection, we measured the oxygen consumption rate of mock- and JEV-infected cells cultured with or without long chain fatty acid (LCFA) palmitate. Cells with JEV infection showed impaired LCFA ß-oxidation and increased interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α) expression. JEV nonstructural protein 5 (NS5) interacted with hydroxyacyl-CoA dehydrogenase α and ß subunits, two components of the mitochondrial trifunctional protein (MTP) involved in LCFA ß-oxidation, and NS5 proteins were detected in mitochondria and co-localized with MTP. LCFA ß-oxidation was impaired and higher cytokines were induced in cells overexpressing NS5 protein as compared with control cells. Deletion and mutation studies showed that the N-terminus of NS5 was involved in the MTP association, and a single point mutation of NS5 residue 19 from methionine to alanine (NS5-M19A) reduced its binding ability with MTP. The recombinant JEV with NS5-M19A mutation (JEV-NS5-M19A) was less able to block LCFA ß-oxidation and induced lower levels of IL-6 and TNF-α than wild-type JEV. Moreover, mice challenged with JEV-NS5-M19A showed less neurovirulence and neuroinvasiveness. We identified a novel function of JEV NS5 in viral pathogenesis by impairing LCFA ß-oxidation and inducing cytokine expression by association with MTP.
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Virus de la Encefalitis Japonesa (Especie)/metabolismo , Encefalitis Japonesa/metabolismo , Ácidos Grasos/metabolismo , Subunidad alfa de la Proteína Trifuncional Mitocondrial/metabolismo , Subunidad beta de la Proteína Trifuncional Mitocondrial/metabolismo , Proteínas no Estructurales Virales/metabolismo , Animales , Virus de la Encefalitis Japonesa (Especie)/genética , Encefalitis Japonesa/genética , Ácidos Grasos/genética , Células HEK293 , Humanos , Ratones , Subunidad alfa de la Proteína Trifuncional Mitocondrial/genética , Subunidad beta de la Proteína Trifuncional Mitocondrial/genética , Oxidación-Reducción , Mutación Puntual , Proteínas no Estructurales Virales/genéticaRESUMEN
BACKGROUND: Arsenic is a widely distributed metalloid compound that has biphasic effects on cultured cells. In large doses, arsenic can be toxic enough to trigger cell death. In smaller amounts, non-toxic doses may promote cell proliferation and induces carcinogenesis. Aberration of chromosome is frequently detected in epithelial cells and lymphocytes of individuals from arsenic contaminated areas. Overexpression of Aurora-A, a mitotic kinase, results in chromosomal instability and cell transformation. We have reported that low concentration (â¦1 µM) of arsenic treatment increases Aurora-A expression in immortalized bladder urothelial E7 cells. However, how arsenic induces carcinogenesis through Aurora-A activation remaining unclear. METHODS: Bromodeoxyuridine (BrdU) staining, MTT assay, and flow cytometry assay were conducted to determine cell proliferation. Messenger RNA and protein expression levels of Aurora-A were detected by reverse transcriptional-PCR and Western blotting, respectively. Centrosome of cells was observed by immunofluorescent staining. The transcription factor of Aurora-A was investigated by promoter activity, chromosome immunoprecipitation (ChIP), and small interfering RNA (shRNA) assays. Mouse model was utilized to confirm the relationship between arsenic and Aurora-A. RESULTS: We reveal that low dosage of arsenic treatment increased cell proliferation is associated with accumulated cell population at S phase. We also detected increased Aurora-A expression at mRNA and protein levels in immortalized bladder urothelial E7 cells exposed to low doses of arsenic. Arsenic-treated cells displayed increased multiple centrosome which is resulted from overexpressed Aurora-A. Furthermore, the transcription factor, E2F1, is responsible for Aurora-A overexpression after arsenic treatment. We further disclosed that Aurora-A expression and cell proliferation were increased in bladder and uterus tissues of the BALB/c mice after long-term arsenic (1 mg/L) exposure for 2 months. CONCLUSION: We reveal that low dose of arsenic induced cell proliferation is through Aurora-A overexpression, which is transcriptionally regulated by E2F1 both in vitro and in vivo. Our findings disclose a new possibility that arsenic at low concentration activates Aurora-A to induce carcinogenesis.
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Arsénico/toxicidad , Aurora Quinasa A/biosíntesis , Carcinoma de Células Transicionales/enzimología , Factor de Transcripción E2F1/metabolismo , Neoplasias de la Vejiga Urinaria/enzimología , Animales , Western Blotting , Carcinogénesis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Inmunoprecipitación de Cromatina , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Ratones , Ratones Endogámicos BALB C , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Neurocognitive (NC) complications continue to afflict a substantial proportion of HIV-infected people taking effective antiretroviral therapy (ART). One contributing mechanism for this is antiretroviral neurotoxicity. Efavirenz (EFV) is associated with short-term central nervous system (CNS) toxicity, but less is known about its long-term effects. Our objective was to compare NC functioning with long-term use of EFV to that of a comparator, lopinavir-ritonavir (LPV/r), in a cohort of well-characterized adults. Four hundred forty-five patients were selected from the CNS HIV Antiretroviral Therapy Effects Research (CHARTER) cohort based on their use of either EFV (n = 272, mean duration 17.9 months) or LPV/r (n = 173, mean duration 16.4 months) and the lack of severe NC comorbidities. All patients had undergone standardized comprehensive NC testing. Univariable and multivariable analyses to predict NC outcomes were performed. Compared with LPV/r users, EFV users were more likely to be taking their first ART regimen (p < 0.001), were less likely to have AIDS (p < 0.001) or hepatitis C virus (HCV) coinfection (p < 0.05), had higher CD4+ T cell nadirs (p < 0.001), had lower peak (p < 0.001) and current (p < 0.001) plasma HIV RNA levels, and were less likely to have detectable HIV RNA in cerebrospinal fluid (CSF) (p < 0.001). Overall, EFV users had worse speed of information processing (p = 0.04), verbal fluency (p = 0.03), and working memory (p = 0.03). An interaction with HCV serostatus was present: Overall among HCV seronegatives (n = 329), EFV users performed poorly, whereas among HCV seropositives (n = 116), LPV/r users had overall worse performance. In the subgroup with undetectable plasma HIV RNA (n = 269), EFV users had worse speed of information processing (p = 0.02) and executive functioning (p = 0.03). Substantial differences exist between EFV and LPV/r users in this observational cohort, possibly because of channeling by clinicians who may have prescribed LPV/r to more severely ill patients or as second-line therapy. Despite these differences, EFV users had worse functioning in several cognitive abilities. A potentially important interaction was identified that could indicate that the NC consequences of specific antiretroviral drugs may differ based on HCV coinfection. The complexity of these data is substantial, and findings would best be confirmed in a randomized clinical trial.
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Benzoxazinas/efectos adversos , Disfunción Cognitiva/fisiopatología , Función Ejecutiva/efectos de los fármacos , Infecciones por VIH/fisiopatología , Hepatitis C/fisiopatología , Memoria/efectos de los fármacos , Aprendizaje Verbal/efectos de los fármacos , Adulto , Alquinos , Fármacos Anti-VIH/uso terapéutico , Benzoxazinas/administración & dosificación , Recuento de Linfocito CD4 , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/virología , Disfunción Cognitiva/complicaciones , Disfunción Cognitiva/tratamiento farmacológico , Disfunción Cognitiva/inmunología , Coinfección , Ciclopropanos , Quimioterapia Combinada , Femenino , Infecciones por VIH/complicaciones , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/inmunología , VIH-1/fisiología , Hepacivirus/fisiología , Hepatitis C/complicaciones , Hepatitis C/tratamiento farmacológico , Hepatitis C/inmunología , Humanos , Lopinavir/uso terapéutico , Masculino , Persona de Mediana Edad , Pruebas Neuropsicológicas , Ritonavir/uso terapéuticoRESUMEN
Human MCP-1-induced protein 1 (MCPIP1, also known as ZC3H12A and Regnase-1) plays important roles in negatively regulating the cellular inflammatory response. Recently, we found that as an RNase, MCPIP1 has broad-spectrum antiviral effects by targeting viral RNA. In this study, we demonstrated that MCPIP1 expression was induced by hepatitis C virus (HCV) infection in Huh7.5 hepatoma cells. MCPIP1 expression was higher in liver tissue from patients with chronic HCV infection compared with those without chronic HCV infection. Knockdown of MCPIP1 increased HCV replication and HCV-mediated expression of proinflammatory cytokines, such as TNF-α, IL-6, and MCP-1. However, overexpression of MCPIP1 significantly inhibited HCV replication and HCV-mediated expression of proinflammatory cytokines. Various mutants of functional domains of MCPIP1 showed disruption of the RNA binding and oligomerization abilities, as well as RNase activity, but not deubiquitinase activity, which impaired the inhibitory activity against HCV replication. On immunocytochemistry, MCPIP1 colocalized with HCV RNA. Use of a replication-defective HCV John Cunningham 1/AAG mutant and in vitro RNA cleavage assay demonstrated that MCPIP1 could directly degrade HCV RNA. MCPIP1 may suppress HCV replication and HCV-mediated proinflammatory responses with infection, which might contribute to the regulation of host defense against the infection and virus-induced inflammation.
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Hepacivirus/fisiología , Hepatitis C Crónica/inmunología , Factores de Transcripción/fisiología , Replicación Viral , Línea Celular Tumoral , Quimiocina CCL2/biosíntesis , Células HEK293 , Hepacivirus/genética , Hepatitis C Crónica/patología , Hepatitis C Crónica/virología , Humanos , Interleucina-6/biosíntesis , Hígado/inmunología , Hígado/patología , Hígado/virología , Mutación , Estructura Terciaria de Proteína , Interferencia de ARN , ARN Interferente Pequeño , ARN Viral/metabolismo , Proteínas de Unión al ARN/genética , Ribonucleasas , Factores de Transcripción/genética , Factor de Necrosis Tumoral alfa/biosíntesis , Proteasas Ubiquitina-EspecíficasRESUMEN
Delivering synthetic protein-coding RNA bypassing the DNA stage for ectopic protein functioning is a novel therapeutic strategy. Joining the linear RNA head-to-tail covalently could be a state-of-the-art strategy for functioning longer. Here we enroll a cis-acting ligase ribozyme (RzL) to generate circular RNA (circRNA) in vitro for ectopic protein expression. The RNA circularization is confirmed by masking the 5' phosphate group, resisting exonuclease RNase R digestion, failing for further tailing, and sequencing the RT-PCR products of the joined region. Interestingly, one internal ribosome entry site (IRES) renders circRNA translation competent, but two IRES in cis, not trans, hamper the translation. The circRNA with highly potent in translation is conferred for antiviral functioning. Accompanying specific guided RNA, a circRNA expressing ribonuclease Cas13 shows excellent potential against the corresponding RNA virus, further extending circRNA functioning in its growing list of applications.
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ARN Catalítico , ARN Circular , ARN Circular/metabolismo , ARN Circular/genética , ARN Catalítico/metabolismo , ARN Catalítico/genética , Humanos , Sitios Internos de Entrada al Ribosoma , Biosíntesis de Proteínas , ARN/metabolismo , ARN/genética , Células HEK293 , ExorribonucleasasRESUMEN
For advanced hepatocellular carcinoma (HCC), the best second-line treatment after first-line treatment with sorafenib is unclear. This study aimed to compared the efficacy of second-line regorafenib (a tyrosine kinase inhibitor) and immune checkpoint inhibitors (ICIs) in patients with advanced HCC after sorafenib therapy. This retrospective study included 89 patients with HCC treated with sorafenib, and then regorafenib (n = 58) or an ICI (n = 31). Treatment response, overall survival (OS) and progression-free survival (PFS) of the 2 groups were compared, and factors associated with post-treatment mortality or disease progression were evaluated. During follow-up period, compared to regorafenib, treatment with an ICI results in a slight increase in a 20% decrease of AFP (35.7% vs. 31.8%), complete response rate (6.5% vs. 0%), objective response rate (16.1% vs. 6.9%), median overall survival (13.3 vs. 5 months), and median PFS (3.0 vs. 2.6 months). Combined locoregional treatment (LRT) (hazard ratio [HR] = 0.40, 95% confidence interval [CI]: 0.15-0.99) during second-line treatment was associated with a decreased risk of post-treatment mortality. After propensity scoring matching, combined LRT during second-line treatment had longer post-treatment OS than patients without combined LRT. A 20% decrease of AFP (HR = 0.54, 95% CI: 0.31-0.94) was associated with a decreased risk of post-treatment disease progression. In conclusions, second-line treatment with regorafenib or ICI prolongs OS in patients with advanced HCC treated with sorafenib. Combined LRT during second-line treatment is associated with decreased post-treatment mortality. A 20% decrease of AFP level may be predictive of a lower rate of disease progression.
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We present a centrifugal microfluidic cartridge for the eight-fold parallel generation of monodisperse water-in-oil droplets using standard laboratory equipment. The key element is interfacing centrifugal microfluidics with its design based on polar coordinates to the linear structures of standard high-throughput laboratory automation. Centrifugal step emulsification is used to simultaneously generate droplets from eight samples directly into standard 200 µl PCR 8-tube strips. To ensure minimal manual liquid handling, the design of the inlets allows the user to load the samples and the oil via a standard multichannel pipette. Simulation-based design of the cartridge ensures that the performance is consistent in each droplet generation unit despite the varying radial positions that originate from the interface to the linear oriented PCR 8-tube strip and from the integration of linear oriented inlet holes for the multichannel pipettes. Within 10 minutes, sample volumes of 50 µl per droplet generation unit are emulsified at a fixed rotation speed of 960 rpm into 1.47 × 105 monodisperse droplets with a mean diameter of 86 µm. The overall coefficient of variation (CV) of the droplet diameter was below 4%. Feasibility is demonstrated by an exemplary digital droplet polymerase chain reaction (ddPCR) assay which showed high linearity (R2 ≥ 0.999) across all of the eight tubes of the strip.
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Microfluídica , Técnicas de Amplificación de Ácido Nucleico , Reacción en Cadena de la Polimerasa , Emulsiones/química , AguaRESUMEN
A rapid, precise, and viability-retaining method for cytoplasmic molecule delivery is highly desired for cell engineering. Routine methods suffer from low throughput, lack of selectivity, requirement of helper compounds, predominant endosomal delivery, and/or are restricted to specific molecule classes. Photonic cell manipulation bears the potential to overcome these drawbacks. Here we investigated mammalian cell manipulation by single sub-nanosecond laser pulses. Axial beam waist positioning close to a cell monolayer induced culture vessel damage and zones of cell ablation. Cells at margins of ablation zones exhibited uptake of membrane-impermeant fluorophores and GFP expression plasmids. Increasing Rayleigh-length and beam waist diameter reduced the sensitivity to axial defocusing and resulted in robust molecule transfer. Serial application of single pulses focused over a moving cell monolayer yielded quantitative molecule transfer to cells at rates up to 40%. Our results could be basic to spatially and temporally controlled single laser pulse-mediated marker-free high throughput cell manipulation.
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Rayos Láser , Luz , Animales , Colorantes Fluorescentes , Endosomas , Fotones , MamíferosRESUMEN
Glioblastoma (GBM) is the most common aggressive malignant brain cancer and is chemo- and radioresistant, with poor therapeutic outcomes. The "double-edged sword" of virus-induced cell death could be a potential solution if the oncolytic virus specifically kills cancer cells but spares normal ones. Zika virus (ZIKV) has been defined as a prospective oncolytic virus by selectively targeting GBM cells, but unclear understanding of how ZIKV kills GBM and the consequences hinders its application. Here, we found that the cellular gasdermin D (GSDMD) is required for the efficient death of a human GBM cell line caused by ZIKV infection. The ZIKV protease specifically cleaves human GSDMD to activate caspase-independent pyroptosis, harming both viral protease-harboring and naive neighboring cells. Analyzing human GSDMD variants showed that most people were susceptible to ZIKV-induced cytotoxicity, except for those with variants that resisted ZIKV cleavage or were defective in oligomerizing the N terminus GSDMD cleavage product. Consistently, ZIKV-induced secretion of the pro-inflammatory cytokine interleukin-1ß and cytolytic activity were both stopped by a small-molecule inhibitor targeting GSDMD oligomerization. Thus, potential ZIKV oncolytic therapy for GBM would depend on the patient's GSDMD genetic background and could be abolished by GSDMD inhibitors if required.
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We demonstrate detection and quantification of bacterial load with a novel microfluidic one-pot wash-free fluorescence in situ hybridization (FISH) assay in droplets. The method offers minimal manual workload by only requiring mixing of the sample with reagents and loading it into a microfluidic cartridge. By centrifugal microfluidic step emulsification, our method partitioned the sample into 210 pL (73 µm in diameter) droplets for bacterial encapsulation followed by in situ permeabilization, hybridization, and signal detection. Employing locked nucleic acid (LNA)/DNA molecular beacons (LNA/DNA MBs) and NaCl-urea based hybridization buffer, the assay was characterized with Escherichia coli, Klebsiella pneumonia, and Proteus mirabilis. The assay performed with single-cell sensitivity, a 4-log dynamic range from a lower limit of quantification (LLOQ) at ~3 × 103 bacteria/mL to an upper limit of quantification (ULOQ) at ~3 × 107 bacteria/mL, anda linearity R2 = 0.976. The total time-to-results for detection and quantification was around 1.5 hours.
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ADN , Microfluídica , Escherichia coli/genética , Hibridación Fluorescente in Situ/métodos , Microfluídica/métodos , OligonucleótidosRESUMEN
Developing efficient bifunctional electrocatalysts in neutral media to avoid the deterioration of electrodes or catalysts under harsh environments has become the ultimate goal in electrochemical water splitting. This work demonstrates the fabrication of an on-chip bifunctional two-dimensional (2D) monolayer (ML) WSe2/graphene heterojunction microreactor for efficient overall water splitting in a neutral medium (pH = 7). Through the synergistic atomic growth of the metallic Cr dopant and graphene stitching contact on the 2D ML WSe2, the bifunctional WSe2/graphene heterojunction microreactor consisting of a full-cell configuration demonstrates excellent performance for overall water splitting in a neutral medium. Atomic doping of metallic Cr atoms onto the 2D ML WSe2 effectively facilitates the charge transfer at the solid-liquid interface. In addition, the direct growth of the self-stitching graphene contact with the 2D WSe2 catalyst largely reduces the contact resistance of the microreactor and further improves the overall water splitting efficiency. A significant reduction of the overpotential of nearly 1000 mV at 10 mA cm-2 at the Cr-doped WSe2/graphene heterojunction microreactor compared to the ML pristine WSe2 counterpart is achieved. The bifunctional WSe2/graphene self-stitching heterojunction microreactor is an ideal platform to investigate the fundamental mechanism of emerging bifunctional 2D catalysts for overall water splitting in a neutral medium.
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Drug substances are at risk of contamination with N-nitrosamines (NAs), well-known carcinogenic agents, during synthesis processes and/or long-term storage. Therefore, in this study, we developed an efficient data-based screening approach to systemically assess marketed products and investigated its scalability for benefiting both regulatory agencies and pharmaceutical industries. A substructure-based screening method employing DataWarrior, an open-source software, was established to evaluate the risks of NA impurities in drug substances. Eight NA substructures containing susceptible amino sources for N-nitrosation have been identified as screening targets: dimethylamine (DMA), diethylamine, isopropylethylamine, diisopropylamine, N-methyl-2-pyrrolidone, dibutylamine, methylphenylamine, and tetrazoles. Our method detected 192 drug substances with a theoretical possibility of NA impurity, 141 of which had not been reported previously. In addition, the DMA moiety was significantly dominant among the eight NA substructures. The results were validated using data from the literature, and a high detection sensitivity of 0.944 was demonstrated. Furthermore, our approach has the advantage of scalability, owing to which 31 additional drugs with suspected NA-contaminated substructures were identified using the substructures of 1-methyl-4-piperazine in rifampin and 1-cyclopentyl-4-piperazine in rifapentine. In conclusion, the reported substructure-based approach provides an effective and scalable method for the screening and investigation of NA impurities in various pharmaceuticals and might be used as an ancillary technique in the field of pharmaceutical quality control for risk assessments of potential NA impurities.
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Contaminación de Medicamentos , Nitrosaminas , Piperazinas , Control de Calidad , Medición de RiesgoRESUMEN
UDP glucuronosyltransferases (UGTs) of the gastrointestinal tract play a crucial role in protection against the toxic effects of xenobiotics in the environment. UGTs such as UGT1A8 and UGT1A10 are predominantly expressed in gastrointestinal tissues. In this study, we examined the phase II metabolism of raloxifene in differentiated Caco-2 monolayers by inducing UGT1A8 and UGT1A10 expression in these cells. The present study evaluated the following four flavonoids of Scutellaria baicalensis as model herbal compounds: scutellarein, salvigenin, baicalein, and wogonin. All test compounds, endpoint substrates, and their metabolites were quantified using liquid chromatography and high-resolution mass spectrometry. The transepithelial electrical resistance values for the individual compounds were comparable regardless of whether they were measured individually. Salvigenin significantly inhibited UGT1A8 and UGT1A10 activities in a concentration-dependent manner. All individual compounds except scutellarein inhibited UGT1A8 and UGT1A10 activity at a concentration of 100 µM. In addition, all individual flavonoids at 100 µM, except wogonin, significantly increased the amount of raloxifene in the basolateral chambers. The positive control, canagliflozin, significantly inhibited both UGT1A8 and UGT1A10 activities. These findings suggest that the Caco-2 assay can be utilized for identifying UGT1A8 and UGT1A10 inhibitors and indicate the potential of salvigenin for enhancing the pharmacological effects of UGT substrate drugs.
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Flavonoides/farmacología , Glucuronosiltransferasa/antagonistas & inhibidores , Interacciones de Hierba-Droga , Clorhidrato de Raloxifeno/farmacología , Scutellaria baicalensis , Moduladores Selectivos de los Receptores de Estrógeno/farmacología , Células CACO-2 , Glucurónidos/metabolismo , Glucuronosiltransferasa/metabolismo , Humanos , Intestinos/enzimologíaRESUMEN
AIMS AND BACKGROUND: Thyroid cancer is the most common endocrine neoplasm worldwide. Although differentiated thyroid cancers are associated with a favorable survival, the prognosis worsens dramatically for patients with distant metastasis. Metastases from follicular thyroid carcinoma (FTC) occur earlier and are more aggressive than those from papillary thyroid carcinoma. For FTC that is resistant to radioactive iodine, new treatments are urgently needed. Human alpha-fetoprotein (HAFP) is a tumor-associated fetal protein that has been demonstrated to regulate tumorigenesis. Growth inhibitory peptide (GIP), a synthetic 34-mer peptide isolated from the third domain of HAFP, has been shown to have antitumor growth ability in various human cancers. However, the effects of GIP in FTC have not yet been studied. The aim of this study was to investigate the antitumor ability of GIP in FTC. METHODS AND STUDY DESIGN: Using both PBS and GIP control peptide as a negative control, the antiproliferative activity of GIP in the WRO human FTC cell line was determined using a tetrazolium-based colorimetric assay. In addition, cell migration and invasion assays were used to measure tumor metastasis inhibition effects in vitro. RESULTS: GIP did not inhibit WRO cell proliferation in a time- or dose-dependent manner. However, in WRO cells treated with GIP for 4 days, migration was significantly inhibited at concentrations of 50 and 100 microM (33.3% and 19.5%, respectively; both P <0.05). Cell invasion was also significantly inhibited at 50 and 100 microM (67.1% and 39.0%, respectively; both P<0.05). CONCLUSIONS: Although GIP failed to suppress FTC cell growth, it effectively interrupted both FTC cell migration and invasion abilities in vitro. Further validation in an animal model and elucidation of the underlying mechanisms will be required. GIP may potentially serve as an anti-FTC metastasis agent aiding current chemotherapy regimens.
Asunto(s)
Adenocarcinoma Folicular/tratamiento farmacológico , Adenocarcinoma Folicular/patología , Antineoplásicos/farmacología , Proteínas de la Membrana/farmacología , Neoplasias de la Tiroides/tratamiento farmacológico , Neoplasias de la Tiroides/patología , Movimiento Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Humanos , Invasividad Neoplásica , Células Tumorales CultivadasRESUMEN
The alarming dynamics of antibiotic-resistant infections calls for the development of rapid and point-of-care (POC) antibiotic susceptibility testing (AST) methods. Here, we demonstrated the first completely stand-alone microfluidic system that allowed the execution of digital enumeration of bacteria and digital antibiograms without any specialized microfluidic instrumentation. A four-chamber gravity-driven step emulsification device generated â¼2000 monodisperse 2 nanoliter droplets with a coefficient of variation of 8.9% of volumes for 95% of droplets within less than 10 minutes. The manual workload required for droplet generation was limited to the sample preparation, the deposition into the sample inlet of the chip and subsequent orientation of the chip vertically without an additional pumping system. The use of shallow chambers imposing a 2D droplet arrangement provided superior stability of the droplets against coalescence and minimized the leakage of the reporter viability dye between adjacent droplets during long-term culture. By using resazurin as an indicator of the growth of bacteria, we were also able to reduce the assay time to â¼5 hours compared to 20 hours using the standard culture-based test.
Asunto(s)
Antibacterianos/farmacología , Enterococcus faecalis/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Gravitación , Dispositivos Laboratorio en un Chip , Staphylococcus aureus/efectos de los fármacos , Antibacterianos/química , Emulsiones/química , Imagen Óptica/instrumentación , Tamaño de la PartículaRESUMEN
Intensive energy demand urges state-of-the-art rechargeable batteries. Rechargeable aluminum-ion batteries (AIBs) are promising candidates with suitable cathode materials. Owing to high abundance of carbon, hydrogen, and oxygen and rich chemistry of organics (structural diversity and flexibility), small organic molecules are good choices as the electrode materials for AIB. Herein, a series of small-molecule quinone derivatives (SMQD) as cathode materials for AIB were investigated. Nonetheless, dissolution of small organic molecules into liquid electrolytes remains a fundamental challenge. To nullify the dissolution problem effectively, 1,4-benzoquinone was integrated with four bulky phthalimide groups to form 2,3,5,6-tetraphthalimido-1,4-benzoquinone (TPB) as the cathode materials and assembled to be the AI/TPB cell. As a result, the Al/TPB cell delivered capacity as high as 175 mA h/g over 250 cycles in the urea electrolyte system. Theoretical studies have also been carried out to reveal and understand the storage mechanism of the TPB electrode.
RESUMEN
Recently, aluminum ion batteries (AIBs) have attracted great attention across the globe by virtue of their massive gravimetric and volumetric capacities in addition to their high abundance. Though carbon derivatives are excellent cathodes for AIBs, there is much room for further development. In this study, flexuous graphite (FG) was synthesized by a simple thermal shock treatment, and for the first time, an Al/FG battery was applied as a cathode for AIBs to reveal the real-time intercalation of AlCl4- into FG with high flexibility by using in-situ scanning electron microscope (SEM) measurements exclusively. Similarly, in-situ X-ray diffraction (XRD) and in-situ Raman techniques have been used to understand the anomalous electrochemical behavior of FG. It was found that FG adopts a unique integrated intercalation-adsorption mechanism where it follows an intercalation mechanism potential above 1.5 V and an adsorption mechanism potential below 1.5 V. This unique integrated intercalation-adsorption mechanism allows FG to exhibit superior properties, like high capacity (≥140 mAh/g), remarkable long-term stability (over 8000 cycles), excellent rate retention (93 mAh/g at 7.5 A/g), and extremely rapid charging and slow discharging.