RESUMEN
As emerging pollutants in the environment, nanoplastics (NPs) can cross biological barriers and be enriched in organisms, posing a greatest threat to the health of livestock and humans. However, the size-dependent toxic effects of NPs in higher mammals remain largely unknown. To determine the size-dependent potential toxicities of NPs, we exposed mouse (AML-12) and human (L02) liver cell lines in vitro, and 6-week-old C57BL/6 mice (well-known preclinical model) in vivo to five different sizes of polystyrene NPs (PS-NPs) (20, 50, 100, 200 and 500 nm). We found that ultra-small NPs (20 nm) induced the highest cytotoxicity in mouse and human liver cell lines, causing oxidative stress and mitochondrial membrane potential loss on AML-12 cells. Unexpectedly in vivo, after long-term oral exposure to PS-NPs (75 mg/kg), medium NPs (200 nm) and large NPs (500 nm) induced significant hepatotoxicity, evidenced by increased oxidative stress, liver dysfunction, and lipid metabolism disorders. Most importantly, medium or large NPs generated local immunotoxic effects via recruiting and activating more numbers of neutrophils and monocytes in the liver or intestine, which potentially resulted in increased proinflammatory cytokine secretion and the tissue damage. The discrepancy in in vitro-in vivo toxic results might be attributed to the different properties of biodistribution and tissue accumulation of different sized NPs in vivo. Our study provides new insights regarding the hepatotoxicity and immunotoxicity of NPs on human and livestock health, warranting us to take immense measures to prevent these NPs-associated health damage.
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Antineoplásicos , Enfermedad Hepática Inducida por Sustancias y Drogas , Leucemia Mieloide Aguda , Nanopartículas , Contaminantes Químicos del Agua , Humanos , Animales , Ratones , Ratones Endogámicos C57BL , Microplásticos/toxicidad , Poliestirenos/toxicidad , Distribución Tisular , Ganado , MamíferosRESUMEN
BACKGROUND AND AIMS: We aimed to investigate the relationship between the cumulative exposure to different lipid parameters and carotid intima-media thickness(CIMT), which is considered a marker for the early stage of atherosclerosis. This is due to the shift in research focus from assessing individual lipoproteins to considering their cumulative exposure. METHODS AND RESULTS: The study included 2,348 participants who had their lipid parameters measured biennially since 2006.To calculate the cumulative lipid burden, the weighted sum of the difference between the measured value and the cutoff value of each parameter, including total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDLC), high-density lipoprotein cholesterol (HDLC), and non-HDLC, was added. Carotid ultrasound was used to detect CIMT. The association between cumulative lipid burden and CIMT was evaluated using linear and logistic analyses. TC and LDLC burden were significantly associated with thickening CIMT (p<0.05). A 2.65-fold, 1.67-fold increased risk of abnormal CIMT was documented in the highest quartile of these two lipid burdens. Notably, a dose-dependent relationship was observed in the overall population when taking non-HDLC burden as a continuous variable (fully-adjusted ß coefficient=0.0013, 95%CI 0.0004-0.0022). CONCLUSIONS: Out of the five lipid parameters, TC and LDLC burden showed independent associations with abnormal CIMT. it is crucial to attain optimal lipid levels for the prevention and treatment of subclinical atherosclerosis.
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Enfermedades Asintomáticas , Biomarcadores , Enfermedades de las Arterias Carótidas , Grosor Intima-Media Carotídeo , Lípidos , Valor Predictivo de las Pruebas , Humanos , Masculino , Femenino , Persona de Mediana Edad , Enfermedades de las Arterias Carótidas/diagnóstico por imagen , Enfermedades de las Arterias Carótidas/sangre , Enfermedades de las Arterias Carótidas/epidemiología , Biomarcadores/sangre , Factores de Riesgo , Lípidos/sangre , Adulto , Medición de Riesgo , Dislipidemias/sangre , Dislipidemias/epidemiología , Dislipidemias/diagnóstico , Triglicéridos/sangre , Anciano , LDL-Colesterol/sangre , HDL-Colesterol/sangre , Factores de TiempoRESUMEN
Mitigation of biofouling and the host's foreign body response (FBR) is a critical challenge with biomedical implants. The surface coating with various anti-fouling materials provides a solution to overcome it, but limited options in clinic and their potential immunogenicity drive the development of more alternative coating materials. Herein, inspired by liquid-liquid phase separation of intrinsically disordered proteins (IDPs) to form separated condensates in physiological conditions, we develop a new type of low-fouling biomaterial based on flexible IDP of FUS protein containing rich hydrophilic residues. A chemical structure-defined FUS IDP sequence tagged with a tetra-cysteine motif (IDPFUS) was engineered and applied for covalent immobilization on various surfaces to form a uniform layer of protein tangles, which boosted strong hydration on surfaces, as revealed by molecular dynamics simulation. The IDPFUS-coated surfaces displayed excellent performance in resisting adsorption of various proteins and adhesion of different cells, platelets, and bacteria. Moreover, the IDPFUS-coated implants largely mitigated the host's FBR compared with bare implants and particularly outperformed PEG-coated implants in reducing collagen encapsulation. Thus, this novel low-fouling and anti-FBR strategy provides a potential surface coating material for biomedical implants, which will also shed light on exploring similar applications of other IDP proteins.
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Incrustaciones Biológicas , Cuerpos Extraños , Proteínas Intrínsecamente Desordenadas , Humanos , Incrustaciones Biológicas/prevención & control , Interacciones Hidrofóbicas e Hidrofílicas , Propiedades de SuperficieRESUMEN
Candida glabrata is currently the first or second most commonly encountered non-albicans Candida species worldwide. The potential severity of Candida resistance mandates the discovery of novel antifungal agents, including those that can be used in combination therapies. In this study, we evaluated the in vitro interactions of pyrogallol (PG) and azole drugs against 22 clinical C. glabrata isolates. The potential mechanism underlying the synergism between PG and fluconazole (FLC) was investigated by the rhodamine 6G efflux method and quantitative reverse transcription (qRT)-PCR analysis. In susceptibility tests, PG showed strong synergism with FLC, itraconazole (ITC), and voriconazole (VRC), with fractional inhibitory concentration index values of 0.18 to 0.375 for PG+FLC, 0.250 to 0.750 for PG+ITC, and 0.141 to 0.750 for PG+VRC. Cells grown in the presence of PG+FLC exhibited reduced rhodamine 6G extrusion and significantly downregulated expression of the efflux-related genes CgCDR1, CgCDR2, and CgPDR1 compared with cells grown in the presence of PG or FLC alone. PG did not potentiate FLC when tested against a ΔCgpdr1 strain. Restoration of a functional CgPDR1 allele also restored the synergism. These results indicate that PG is an antifungal agent that synergistically potentiates the activity of azoles. Furthermore, PG appears to exert its effects by inhibiting efflux pumps and downregulating CgCDR1, CgCDR2, and CgPDR1, with CgPDR1 probably playing a crucial role in this process.
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Candida glabrata , Fluconazol , Antifúngicos/farmacología , Candida glabrata/genética , Farmacorresistencia Fúngica/genética , Fluconazol/farmacología , Pruebas de Sensibilidad Microbiana , Pirogalol/farmacologíaRESUMEN
Reasonable design of non-noble metal catalysts with hollow open structure for hydrodeoxygenation (HDO) of lignin derivatives to value-added chemicals is of great significance but challenging. Herein, a novel MOF-derived multilayer hollow sphere coated nickeltungsten bimetallic catalyst (Ni2-WOx@CN-700) was fabricated via by confined pyrolysis strategy using bimetallic MOFs as a self-sacrificial template, which exhibits robust activity for the typical model HDO of vanillin to 2-methoxy-4-methylphenol (Yield of 100 % at 140 °C for no less than 10 cycles). The characterizations revealed that WOx facilitated the dispersion of Ni nanoparticles and adjusted the acidic capacity of the catalyst through the formed Ni-WOx heterojunction. Density functional theory (DFT) calculations confirms that WOx species enhanced the electron-rich nature of the active sites, while the adsorption energies of H2 and vanillin on Ni-WOx decreased from -0.572 eV and - 0.622 eV on Ni to -3.969 eV and - 4.922 eV, respectively. These results further indicated that the high activity of Ni2-WOx@CN-700 was attributed to the Ni-WOx heterojunction. Based on the characterizations and the thermodynamic calculations, the reaction mechanism was proposed. In addition, the catalyst shows good substrate universality, which enables its good commercial application prospect.
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Benzaldehídos , Níquel , Catálisis , Níquel/química , Benzaldehídos/química , Tungsteno/química , Lignina/química , Termodinámica , Estructuras Metalorgánicas/química , Adsorción , Teoría Funcional de la DensidadRESUMEN
Achieving reliable detection of trace levels of NO2 gas is essential for environmental monitoring and protection of human health protection. Herein, a thin-film gas sensor based on branched WO3/W18O49 heterostructures was fabricated. The optimized WO3/W18O49 sensor exhibited outstanding NO2 sensing properties with an ultrahigh response value (1038) and low detection limit (10 ppb) at 50 °C. Such excellent sensing performance could be ascribed to the synergistic effect of accelerated charge transfer and increased active sites, which is confirmed by electrochemical impedance spectroscopy and temperature-programmed desorption characterization. The sensor exhibited an excellent detection ability to NO2 under different air quality conditions. This work provides an effective strategy for constructing WO3/W18O49 heterostructures for developing NO2 gas sensors with an excellent sensing performance.
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Espectroscopía Dieléctrica , Dióxido de Nitrógeno , Humanos , Dominio Catalítico , Monitoreo del AmbienteRESUMEN
An aptamer targeting gliotoxin (GTX) was optimized to increase the binding affinity by approximately 20 times and achieve higher structural stability and targeting specificity. Molecular dynamics simulations were used to explore the molecular mechanism and key action sites underlying the recognition of GTX by the optimized aptamer. Subsequently, the optimized aptamer was split into two fragments and a convenient and rapid one-pot assay for GTX detection was successfully established using a target-driven split aptamer recognition and assembly strategy. The method exhibited a good linear range of 0.128 nM to 2 µM, a low detection limit of 0.07 nM, and excellent selectivity for GTX. Furthermore, the method had good accuracy and stability in real sample analysis. Therefore, the developed one-pot method provides a reliable, convenient, and cost-effective approach for the widespread application of GTX detection.
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Aptámeros de Nucleótidos , Gliotoxina , Aptámeros de Nucleótidos/química , Gliotoxina/química , Gliotoxina/análisis , Límite de Detección , Contaminación de Alimentos/análisis , Técnicas Biosensibles/métodos , Simulación de Dinámica Molecular , AnimalesRESUMEN
Therapeutic cancer vaccines fail to produce satisfactory outcomes against solid tumors since vaccine-induced anti-tumor immunity is significantly hampered by immunosuppression. Generating an in situ cancer vaccine targeting immunological cold tumor microenvironment (TME) appears attractive. Here, a type of free-field based whole-body ultrasound (US)-driven nanovaccines are constructed, named G5-CHC-R, by conjugating the sonosensitizer, Chenghai chlorin (CHC) and the immunomodulator, resiquimod (R848) on top of a super small-sized dendrimeric nanoscaffold. Once entering tumors, R848 can be cleaved from a hypoxia-sensitive linker, thus modifying the TME via converting macrophage phenotypes. The animals bearing orthotopic pancreatic cancer with intestinal metastasis and breast cancer with lung metastasis are treated with G5-CHC-R under a free-field based whole-body US system. Benefit from the deep penetration capacity and highly spatiotemporal selectiveness, G5-CHC-R triggered by US represented a superior alternative for noninvasive irradiation of deep-seated tumors and magnification of local immune responses via driving mass release of tumor antigens and "cold-warm-hot" three-state transformation of TME. In addition to irradiating primary tumors, a robust adaptive anti-tumor immunity is potentiated, leading to successful induction of systemic tumor suppression. The sono-nanovaccines with good biocompatibility posed wide applicability to a broad spectrum of tumors, revealing immeasurable potential for translational research in oncology.
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Vacunas contra el Cáncer , Neoplasias , Animales , Nanovacunas , Ultrasonografía , Inmunidad Adaptativa , Adyuvantes Inmunológicos , Neoplasias/diagnóstico por imagen , Neoplasias/terapiaRESUMEN
Reasonable construction of bi-function catalysts with well dispersed hydrogenation active sites and acidic sites are crucial for the hydrodeoxygenation (HDO) of biomass-derived compounds but still a huge challenge. Herein, a 3D Mo functionalized Ni-based bimetallic embedded catalyst with fine metal nanoparticles size (<6 nm) was prepared for the first time using dendritic mesoporous silica as a sacrificial template by one-pot hydrothermal synthesis and adopted in the HDO process of vanillin (VAN) upgrade to 2-methoxy-4-methylphenol (MMP). The characterization results illustrated that Mo species regulated the acidity of the catalyst and promoted the formation of Ni-Mo alloy sites. Density functional theory (DFT) calculations further unveiled that Ni-Mo alloy sites promoted the activation and dissociation of CO bond in VAN, enhanced the ability of protonation hydrogenolysis. Benefitting from the synergistic effect of the highly uniformly dispersed hydrogenation metal sites and acidic sites, nearly 100% yield of MMP could obtained over the designed catalyst under mild conditions (130 °C, 1.5 MPa H2, 3 h, 10 wt% catalyst dosage). Additionally, the NiMo0.1@MSN catalyst displayed robust activity for no less than 8 recycles and excellent universality for the HDO of a variety of lignin derivatives and biomass platform molecules, which provide a feasible strategy for the construction of 3D confined catalysts for the high-efficiency HDO of biomass derivatives.
RESUMEN
Candida albicans biofilms are resistant to several clinical antifungal agents. Thus, it is necessary to develop new antibiofilm intervention measures. Pseudolaric acid A (PAA), a diterpenoid mainly derived from the pine bark of Pseudolarix kaempferi, has been reported to have an inhibitory effect on C. albicans. The primary aim of the current study was to investigate the antibiofilm effect of PAA when combined with fluconazole (FLC) and explore the underlying mechanisms. Biofilm activity was assessed by tetrazolium {XTT [2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide salt]} reduction assays. PAA (4 µg/mL) combined with FLC (0.5 µg/mL) significantly inhibited early, developmental, and mature biofilm formation compared with the effect of PAA or FLC alone (P < 0.05). Furthermore, PAA (4 µg/mL) combined with FLC (0.5 µg/mL) produced a 56% reduction in C. albicans biofilm adhesion. The combination of PAA (4 µg/mL) and FLC (0.5 µg/mL) also performed well in inhibiting yeast-to-hypha transition. Transcriptome analysis using RNA sequencing and quantitative reverse transcription PCR indicated that the PAA-FLC combination treatment produced a strong synergistic inhibitory effect on the expression of genes involved in adhesion (ALS1, ALS4, and ALS2) and yeast-to-hypha transition (ECE1, PRA1, and TEC1). Notably, PAA, rather than FLC, may have a primary role in suppressing the expression of ALS1. In conclusion, these findings demonstrate, for the first time, that the combination of PAA and FLC has an improved antibiofilm effect against the formation of C. albicans biofilms by inhibiting adhesion and yeast-to-hypha transition; this may provide a novel therapeutic strategy for treating C. albicans biofilm-associated infection. IMPORTANCE Biofilms are the primary cause of antibiotic-resistant candida infections associated with medical implants and devices worldwide. Treating biofilm-associated infections is a challenge for clinicians because these infections are intractable and persistent. Candida albicans readily forms extensive biofilms on the surface of medical implants and mucosa. In this study, we demonstrated, for the first time, an inhibitory effect of pseudolaric acid A alone and in combination with fluconazole on C. albicans biofilms. Moreover, pseudolaric acid A in combination with fluconazole exerted an antibiofilm effect through multiple pathways, including inhibition of yeast-to-hypha transition and adhesion. This research not only provides new insights into the synergistic mechanisms of antifungal drug combinations but also brings new possibilities for addressing C. albicans drug resistance.
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Candida albicans , Fluconazol , Antifúngicos/farmacología , Biopelículas , Diterpenos , Sinergismo Farmacológico , Fluconazol/farmacología , Hifa/genética , Pruebas de Sensibilidad MicrobianaRESUMEN
African American (AA) families have the highest risk of prostate cancer. However, the genetic factors contributing to prostate cancer susceptibility in AA families remain poorly understood. We performed whole-exome sequencing of one affected and one unaffected brother in an AA family with hereditary prostate cancer. The novel non-synonymous variants discovered only in the affected individuals were further analyzed in all affected and unaffected men in 20 AA-PC families. Here, we report one rare recurrent ADPRHL1 germline mutation (c.A233T; p.D78V) in four of the 20 families affected by prostate cancer. The mutation co-segregates with prostate cancer in two families and presents in two affected men in the other two families, but was absent in 170 unrelated healthy AA men. Functional characterization of the mutation in benign prostate cells showed aberrant promotion of cell proliferation, whereas expression of the wild-type ADPRHL1 in prostate cancer cells suppressed cell proliferation and oncogenesis. Mechanistically, the ADPRHL1 mutant activates PARP1, leading to an increased H2O2 or cisplatin-induced DNA damage response for prostate cancer cell survival. Indeed, the PARP1 inhibitor, olaparib, suppresses prostate cancer cell survival induced by mutant ADPRHL1. Given that the expression levels of ADPRHL1 are significantly high in normal prostate tissues and reduce stepwise as Gleason scores increase in tumors, our findings provide genetic, biochemical, and clinicopathological evidence that ADPRHL1 is a tumor suppressor in prostate tissue. A loss of function mutation in ADPRHL1 induces prostate tumorigenesis and confers prostate cancer susceptibility in high-risk AA families. IMPLICATIONS: This study highlights a potential strategy for ADPRHL1 mutation detection in prostate cancer-risk assessment and a potential therapeutic application for individuals with prostate cancer in AA families.
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Mutación de Línea Germinal , Neoplasias de la Próstata , Humanos , Masculino , Negro o Afroamericano/genética , Peróxido de Hidrógeno , Clasificación del Tumor , Poli(ADP-Ribosa) Polimerasa-1/genética , Neoplasias de la Próstata/patologíaRESUMEN
Prostate cancer (PCa) is the second most common cancer in men. Androgen receptor (AR) signaling pathway plays a crucial role in prostate development and homeostasis. Dysregulation of this pathway activates AR leading to PCa pathogenesis and progression. AR binds testosterone and other male hormones, which then undergoes post-translational modification for AR nuclear translocation and transcriptional activation. AR activation by post-translational modification is thus imperative for PCa cell growth and survival. Identification and understanding of the pathological and mechanistic roles of AR modifications may increase our understanding of AR activation in PCa and provide new therapeutic options. Recently, AR acetylation has been described as an important step for AR activation. Upregulation of several acetyltransferases has been reported to be associated with PCa progression. Herein, we provide a general understanding of AR acetylation, with a special emphasis on ARD1, and potential therapies that may be exploited against the ARD1-AR axis for PCa treatment.
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Acetiltransferasa A N-Terminal/metabolismo , Acetiltransferasa E N-Terminal/metabolismo , Neoplasias de la Próstata/metabolismo , Acetilación , Animales , Humanos , Masculino , Acetiltransferasa A N-Terminal/genética , Acetiltransferasa E N-Terminal/genética , Procesamiento Proteico-Postraduccional , Receptores Androgénicos/metabolismoRESUMEN
Androgen receptor splice variants (AR-V) are implicated in resistance of prostate cancer to androgen-directed therapies. When expressed alone in cells, some AR-Vs (e.g., AR-V7) localize primarily to the nucleus, whereas others (e.g., AR-V1, AR-V4, and AR-V6) localize mainly to the cytoplasm. Significantly, the latter are often coexpressed with the nucleus-predominant AR-Vs and the full-length AR (AR-FL). An important question to be addressed is whether the cytoplasmic-localized AR-Vs play a role in castration-resistant prostate cancer (CRPC) through interaction with the nucleus-predominant AR-Vs and AR-FL. Here, it is demonstrated that AR-V1, -V4, and -V6 can dimerize with both AR-V7 and AR-FL. Consequently, AR-V7 and androgen-bound AR-FL induced nuclear localization of AR-V1, -V4, and -V6, and these variants, in turn, mitigated the ability of the antiandrogen enzalutamide to inhibit androgen-induced AR-FL nuclear localization. Interestingly, the impact of nuclear localization of AR-V4 and -V6 on AR transactivation differs from that of AR-V1. Nuclear localization leads to an increased ability of AR-V4 and -V6 to transactivate both canonical AR targets and AR-V-specific targets and to confer castration-resistant cell growth. However, although AR-V1, which lacks inherent transcriptional activity, appears to activate AR-FL in an androgen-independent manner, it significantly antagonizes AR-V7 transactivation. Together, these data demonstrate that the complex interactions among different AR-Vs and AR-FL play a significant role in castration-resistant disease. IMPLICATIONS: This study suggests important consequences for clinical castration resistance due to simultaneous expression of AR-FL and AR-Vs in patient tumors and suggests that dissecting these interactions should help develop effective strategies to disrupt AR-V signaling. Mol Cancer Res; 15(1); 59-68. ©2016 AACR.
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Empalme Alternativo/genética , Núcleo Celular/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/genética , Receptores Androgénicos/genética , Andrógenos/metabolismo , Línea Celular Tumoral , Células HEK293 , Humanos , Masculino , Modelos Biológicos , Neoplasias de la Próstata Resistentes a la Castración/patología , Multimerización de Proteína , Transporte de Proteínas , Receptores Androgénicos/metabolismo , Activación Transcripcional/genéticaRESUMEN
Prostate cancer is an androgen receptor (AR)-driven disease and post-translational modification of AR is critical for AR activation. We previously reported that Arrest-defective protein 1 (ARD1) is an oncoprotein in prostate cancer. It acetylates and activates AR to promote prostate tumorigenesis. However, the ARD1-targeted residue within AR and the mechanisms of the acetylation event in prostate tumorigenesis remained unknown. In this study, we show that ARD1 acetylates AR at lysine 618 (K618) in vitro and in vivo. An AR construct with the charged lysine substitution by arginine (AR-618R) reduces RNA Pol II binding, AR transcriptional activity, prostate cancer cell growth, and xenograft tumor formation due to attenuation of AR nuclear translocation, whereas, construct mimicking neutral polar substitution acetylation at K618 by glutamine (AR-618Q) enhanced these effects beyond that of the wild-type AR. Mechanistically, ARD1 forms a ternary complex with AR and HSP90 in vitro and in vivo. Expression of ARD1 increases levels of AR acetylation and AR-HSP90 dissociation in a dose dependent manner. Moreover, the AR acetylation defective K618R mutant is unable to dissociate from HSP90 while the HSP90-dissociated AR is acetylated following ligand exposure. This work identifies a new mechanism for ligand-induced AR-HSP90 dissociation and AR activation. Targeting ARD1-mediated AR acetylation may be a potent intervention for AR-dependent prostate cancer therapy.
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Proteínas HSP90 de Choque Térmico/química , Acetiltransferasa A N-Terminal/fisiología , Acetiltransferasa E N-Terminal/fisiología , Neoplasias de la Próstata/etiología , Procesamiento Proteico-Postraduccional , Receptores Androgénicos/metabolismo , Acetilación , Animales , Línea Celular Tumoral , Núcleo Celular/metabolismo , Humanos , Masculino , Ratones , Neoplasias de la Próstata/metabolismo , Receptores Androgénicos/químicaRESUMEN
Although men with testosterone deficiency are at increased risk for type 2 diabetes (T2D), previous studies have ignored the role of testosterone and the androgen receptor (AR) in pancreatic ß cells. We show that male mice lacking AR in ß cells (ßARKO) exhibit decreased glucose-stimulated insulin secretion (GSIS), leading to glucose intolerance. The AR agonist dihydrotestosterone (DHT) enhances GSIS in cultured male islets, an effect that is abolished in ßARKO(-/y) islets and human islets treated with an AR antagonist. In ß cells, DHT-activated AR is predominantly extranuclear and enhances GSIS by increasing islet cAMP and activating the protein kinase A. In mouse and human islets, the insulinotropic effect of DHT depends on activation of the glucagon-like peptide-1 (GLP-1) receptor, and accordingly, DHT amplifies the incretin effect of GLP-1. This study identifies AR as a novel receptor that enhances ß cell function, a finding with implications for the prevention of T2D in aging men.
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Núcleo Celular/metabolismo , Glucosa/farmacología , Insulina/metabolismo , Receptores Androgénicos/metabolismo , Animales , Línea Celular Tumoral , Núcleo Celular/efectos de los fármacos , AMP Cíclico/metabolismo , Intolerancia a la Glucosa/patología , Humanos , Células Secretoras de Insulina/metabolismo , Masculino , Ratones Noqueados , Modelos Biológicos , Receptores Androgénicos/deficiencia , Transducción de Señal/efectos de los fármacos , Testosterona/farmacologíaRESUMEN
Vascular endothelial growth factor (VEGF) plays an important role in both physiological and pathological angiogenesis. VEGF receptor1 (VEGFR1) acts as a decoy VEGF receptor that enables the regulation of VEGF on the vascular endothelium. In the present study, the recombinant human VEGFR1D13/Fc (rhVEGFR1), which contains key domains for VEGF binding, was cloned and expressed in Chinese hamster ovary (CHO) cells. The rhVEGFR1 protein was purified using proteinA affinity chromatography. The molecular weight of rhVEGFR1 was found to be ~162 and 81 kD in nonreducing and reducing SDSPAGE, respectively. The majority of the final protein products were in the dimeric conformation. Western blot analysis revealed that rhVEGFR1 was only capable of binding to the full glycan form of rhVEGF165 and rhVEGF121. The dissociation constant for the binding of rhVEGFR1 to VEGF165, detected using Biacore, was 285 pM. In addition, rhVEGFR1 inhibited the proliferation and migration of human microvascular endothelial cells. In vivo experiments also demonstrated that rhVEGFR1 inhibited chicken chorioallantoic membrane neovascularization and angiogenesis in nude mice. In conclusion, an antiangiogenic recombinant soluble VEGFR was expressed (up to 5 mg/l) in CHO cells and was shown to be capable of inhibiting neovascularization in vivo and in vitro.
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Inhibidores de la Angiogénesis/farmacología , Neovascularización Fisiológica/efectos de los fármacos , Proteínas Recombinantes de Fusión , Receptor 1 de Factores de Crecimiento Endotelial Vascular/farmacología , Animales , Células CHO , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Clonación Molecular , Cricetulus , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Humanos , Ratones , Ratones Desnudos , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genéticaRESUMEN
Docetaxel-based chemotherapy is established as a first-line treatment and standard of care for patients with metastatic castration-resistant prostate cancer. However, half of the patients do not respond to treatment and those do respond eventually become refractory. A better understanding of the resistance mechanisms to taxane chemotherapy is both urgent and clinical significant, as taxanes (docetaxel and cabazitaxel) are being used in various clinical settings. Sustained signaling through the androgen receptor (AR) has been established as a hallmark of CRPC. Recently, splicing variants of AR (AR-Vs) that lack the ligand-binding domain (LBD) have been identified. These variants are constitutively active and drive prostate cancer growth in a castration-resistant manner. In taxane-resistant cell lines, we found the expression of a major variant, AR-V7, was upregulated. Furthermore, ectopic expression of two clinically relevant AR-Vs (AR-V7 and ARV567es), but not the full-length AR (AR-FL), reduced the sensitivities to taxanes in LNCaP cells. Treatment with taxanes inhibited the transcriptional activity of AR-FL, but not those of AR-Vs. This could be explained, at least in part, due to the inability of taxanes to block the nuclear translocation of AR-Vs. Through a series of deletion constructs, the microtubule-binding activity was mapped to the LBD of AR. Finally, taxane-induced cytoplasm sequestration of AR-FL was alleviated when AR-Vs were present. These findings provide evidence that constitutively active AR-Vs maintain the AR signaling axis by evading the inhibitory effects of microtubule-targeting agents, suggesting that these AR-Vs play a role in resistance to taxane chemotherapy.
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Empalme Alternativo/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Microtúbulos/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Receptores Androgénicos/metabolismo , Transducción de Señal , Transporte Activo de Núcleo Celular , Animales , Células COS , Línea Celular Tumoral , Chlorocebus aethiops , Citoplasma/metabolismo , Docetaxel , Recuperación de Fluorescencia tras Fotoblanqueo , Eliminación de Gen , Humanos , Ligandos , Masculino , Estructura Terciaria de Proteína , Receptores Androgénicos/genética , Taxoides/química , Transcripción Genética , Regulación hacia ArribaRESUMEN
Upregulation of constitutively-active androgen receptor splice variants (AR-Vs) has been implicated in AR-driven tumor progression in castration-resistant prostate cancer. To date, functional studies of AR-Vs have been focused mainly on their ability to regulate gene expression independent of the full-length AR (AR-FL). Here, we showed that AR-V7 and ARv567es, two major AR-Vs, both facilitated AR-FL nuclear localization in the absence of androgen and mitigated the ability of the antiandrogen enzalutamide to inhibit AR-FL nuclear trafficking. AR-V bound to the promoter of its specific target without AR-FL, but co-occupied the promoter of canonical AR target with AR-FL in a mutually-dependent manner. AR-V expression attenuated both androgen and enzalutamide modulation of AR-FL activity/cell growth, and mitigated the in vivo antitumor efficacy of enzalutamide. Furthermore, ARv567es levels were upregulated in xenograft tumors that had acquired enzalutamide resistance. Collectively, this study highlights a dual function of AR-Vs in mediating castration resistance. In addition to trans-activating target genes independent of AR-FL, AR-Vs can serve as a "rheostat" to control the degree of response of AR-FL to androgen-directed therapy via activating AR-FL in an androgen-independent manner. The findings shed new insights into the mechanisms of AR-V-mediated castration resistance and have significant therapeutic implications.
Asunto(s)
Andrógenos/farmacología , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Neoplasias Hormono-Dependientes/genética , Neoplasias de la Próstata Resistentes a la Castración/genética , Empalme del ARN/genética , Receptores Androgénicos/genética , Animales , Apoptosis/efectos de los fármacos , Benzamidas , Núcleo Celular/metabolismo , Proliferación Celular/efectos de los fármacos , Inmunoprecipitación de Cromatina , Humanos , Masculino , Ratones , Ratones Desnudos , Neoplasias Hormono-Dependientes/tratamiento farmacológico , Neoplasias Hormono-Dependientes/patología , Nitrilos , Feniltiohidantoína/análogos & derivados , Feniltiohidantoína/farmacología , Regiones Promotoras Genéticas/genética , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/patología , Isoformas de Proteínas , Transporte de Proteínas/efectos de los fármacos , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Fracciones Subcelulares , Activación Transcripcional , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Significant advances in our understanding of continued androgen receptor (AR) signaling in castration-resistant prostate cancer have led to the development and FDA approval of two next-generation androgen-directed therapies, abiraterone and enzalutamide. These new therapies heralded a new era of prostate cancer therapy. However, disease progression during androgen-directed therapies remains the most critical challenge in the clinical management of prostate cancer. Accumulating evidence points to an important contribution of constitutively-active AR splice variants to AR-driven tumor progression during androgen-directed therapies. In this review, we will focus on the structure, activity, detection, clinical relevance, and mechanisms of production of AR splice variants.