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ETHNOPHARMACOLOGICAL RELEVANCE: The rhizome of Dryopteris crassirhizoma Nakai (Dryopteridaceae, RDC), a traditional East Asian herbal medicine, possesses a broad spectrum of medicinal properties, including anti-inflammatory, anticancer, antibacterial, and antiviral activities. AIM OF THE STUDY: This study investigates the 30% ethanolic extract of RDC's antiviral potential against human coronavirus OC43 (HCoV-OC43), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and its variants infections. MATERIALS AND METHODS: A 30% ethanolic extract of RDC or its components, filixic acid ABA (PubChem CID: 15081408) and dryocrassin ABBA (PubChem CID: 3082025) were treated with Human Coronavirus infection (HCoV-OC43, SARS-CoV-2 and its variants). The base peak chromatogram of RDC was evaluated using UPLC-Q/TOF Mass to identify the RDC, and the quantitative analysis of RDC compounds was performed using LC-MS/MS. A cytopathic effect (CPE) reduction assay, Western blot, immunofluorescence staining of viral protein expression, and qRT-PCR were performed to quantify the viral RNA copy numbers and determine the antiviral activity. The time-of-addition assay, the virus attachment, penetration, and virucidal assays, and SARS-CoV-2 Mpro and PLpro activity assay were used to elucidate the mode of action. RESULTS: RDC exhibited dose-dependent inhibition of HCoV-OC43-induced cytopathic effects, reducing viral RNA copy numbers and viral protein levels. Time-of-addition assays indicated that RDC targets the early stages of the HCoV-OC43 life cycle, inhibiting virion attachment and penetration with virucidal activity. Notably, filixic acid ABA and dryocrassin ABBA, constituents of RDC, reduced HCoV-OC43 viral RNA loads. Furthermore, RDC effectively blocked viral entry in pseudotyped lentivirus assays, involving spike proteins of SARS-CoV-2 Delta plus and South Africa variants, as well as control lentiviral particles expressing vesicular stomatitis virus glycoprotein G. Additionally, RDC demonstrated inhibition of SARS-CoV-2 infection and its variants by targeting viral proteases, namely main protease (Mpro) and papain-like protease (PLpro). CONCLUSIONS: These findings underscore RDC's multistage approach to targeting viral infections by impeding virus entry and inhibiting viral protease activity. Therefore, RDC holds promise as a potent, broad-spectrum anticoronaviral therapeutic agent.
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Antivirales , Dryopteris , Extractos Vegetales , Rizoma , SARS-CoV-2 , Internalización del Virus , Antivirales/farmacología , Antivirales/aislamiento & purificación , Internalización del Virus/efectos de los fármacos , Extractos Vegetales/farmacología , Dryopteris/química , Humanos , SARS-CoV-2/efectos de los fármacos , Coronavirus Humano OC43/efectos de los fármacos , Animales , Proteasas 3C de Coronavirus/antagonistas & inhibidores , Proteasas 3C de Coronavirus/metabolismo , Chlorocebus aethiops , Células VeroRESUMEN
Auraptene (7-geranyloxycoumarin) is the abundant prenyloxycoumarin found in the fruits of Citrus spp. Auraptene has a variety of pharmacological and therapeutic functions, such as anticancer, antioxidant, immunomodulatory, and anti-inflammation activities, with excellent safety profiles. In this study, we evaluated the anticoronaviral activity of auraptene in HCoV-OC43-infected human lung fibroblast MRC-5 cells. We found that auraptene effectively inhibited HCoV-OC43-induced cytopathic effects with 4.3 µM IC50 and 6.1 µM IC90, resulting in a selectivity index (CC50/IC50) of >3.5. Auraptene treatment also decreased viral RNA levels in HCoV-OC43-infected cells, as detected through quantitative real-time PCR, and decreased the expression level of spike proteins and nucleocapsid proteins in virus-infected cells, as detected through the Western blot analysis and immunofluorescence staining. Time-of-addition analysis showed auraptene's inhibitory effects at the post-entry stage of the virus life cycle; however, auraptene did not induce the antiviral interferon families, IFN-α1, IFN-ß1, and IFN-λ1. Additionally, auraptene-treated MRC-5 cells during HCoV-OC43 infection decreased the MMP-9 mRNA levels which are usually increased due to the infection, as auraptene is a previously reported MMP-9 inhibitor. Therefore, auraptene showed antiviral activity against HCoV-OC43 infection, and we suggest that auraptene has the potential to serve as a therapeutic agent against human coronavirus.
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Infecciones por Coronavirus , Coronavirus Humano OC43 , Humanos , Antivirales/farmacología , Antivirales/uso terapéutico , Coronavirus Humano OC43/genética , Metaloproteinasa 9 de la MatrizRESUMEN
A natural chalcone, cardamonin (2',4'-dihydroxy-6'-methoxychalcone; CDN) was isolated from the seeds of Alpinia katsumadai Hayata, which has been traditionally used to treat stomach aches. CDN has been reported to possess various pharmacological properties, including anticancer and anti-inflammatory effects. This study evaluated the antiviral activity of CDN against human coronavirus HCoV-OC43 and determined the mode of action in HCoV-OC43-infected human lung cell lines (MRC-5 and A549 cells). CDN significantly inhibited HCoV-OC43-induced cytopathic effects with an IC50 of 3.62 µM and a CC50 of >50 µM, resulting in a selectivity index of >13.81. CDN treatment reduced the level of viral RNA and the expression of spike and nucleocapsid proteins in HCoV-OC43-infected cells as determine through qRT-PCR and Western blot analysis. Additionally, the activation of p38 mitogen-activated protein kinase (MAPK) by anisomycin decreased viral protein expression, whereas an inhibitor of p38 MAPK signaling, SB202190, increased viral protein expression. CDN also amplified and extended the p38 MAPK signaling pathway in HCoV-OC43-infected cells. In conclusion, CDN inhibited HCoV-OC43 infection by activating the p38 MAPK signaling pathway and has potential as a therapeutic agent against human coronavirus.
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Chalconas , Infecciones por Coronavirus , Coronavirus Humano OC43 , Humanos , Coronavirus Humano OC43/genética , Chalconas/farmacología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Transducción de Señal , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Pulmón/metabolismo , Proteínas ViralesRESUMEN
The natural plant dietary polyphenols 1,2,3,4,6-O-Pentagalloylglucose (PGG) and proanthocyanidin (PAC) have potent antioxidant activity and a variety of pharmacological activities, including antiviral activity. In this study, we examined the inhibitory effect of PGG and PAC on SARS-CoV-2 virus infection, and elucidated its mode of action. PGG and PAC have dose-dependent inhibitory activity against SARS-CoV-2 infection in Vero cells. PGG has a lower IC50 (15.02 ± 0.75 µM) than PAC (25.90 ± 0.81 µM), suggesting that PGG has better inhibitory activity against SARS-CoV-2 than PAC. The PGG and PAC inhibit similar Mpro activities in a protease activity assay, with IC50 values of 25-26 µM. The effects of PGG and PAC on the activity of the other essential SARS-CoV-2 viral protein, RdRp, were analyzed using a cell-based activity assay system. The activity of RdRp is inhibited by PGG and PAC, and PGG has a lower IC50 (5.098 ± 1.089 µM) than PAC (21.022 ± 1.202 µM), which is consistent with their inhibitory capacity of SARS-CoV-2 infection. PGG and PAC also inhibit infection by SARS-CoV and MERS-CoV. These data indicate that PGG and PAC may be candidate broad-spectrum anticoronaviral therapeutic agents, simultaneously targeting the Mpro and RdRp proteins of SARS-CoV-2.
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BACKGROUND: Bavachin, a flavonoid compound isolated from the seeds and fruits of Psoralea corylifolia l. (family Fabaceae), is used as a traditional medicine in Asia. Indeed, it is reported to have various medicinal functions such as estrogenic and antiinflammatory activities among others. However, to date, the effects of bavachin on T cell activation have yet to be reported. PURPOSE AND STUDY DESIGN: We aimed to determine the effects of bavachin on the activation of a human T cell line in vitro and on antigen-specific immune responses in mice in vivo. METHODS: In a nuclear factor of activated T cells (NFAT) activity assay, the Jurkat T cell line expressing a luciferase reporter driven by an NFAT-response element was stimulated with antihuman CD3/CD28 antibody and bavachin. Furthermore, the level of cytokine production was measured in the Jurkat T cell line stimulated with phorbol 12-myristate 13-acetate/ionomycin and bavachin using an IL-2 ELISA and a cytometric bead array assay. For in vivo analyses, mice were subcutaneously immunized with an antigen (ovalbumin protein) and bavachin, and the immune responses of mice were analyzed by FACS analysis, a T cell proliferation assay, a cytokine ELISA, and an antiovalbumin-specific antibody ELISA. RESULTS: We found that bavachin activated NFAT-mediated transcription in the human T cell line in vitro. In mice, when bavachin was administered with the antigen, an increase in T cell responses and antibody production specific to the antigen was observed. CONCLUSION: Our results suggest that bavachin has immunoadjuvant and immunomodulation effects, which arise through activation of the NFAT signaling pathway.
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Adyuvantes Inmunológicos , Factores de Transcripción NFATC , Animales , Flavonoides , Humanos , Interleucina-2 , Células Jurkat , Ratones , Transducción de SeñalRESUMEN
The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), urgently needs effective prophylactic and therapeutic drugs. RNA-dependent RNA polymerase (RdRp), essential for replicating and transcribing a viral RNA genome, is highly conserved in coronaviruses; thus, it is a potential target for inhibiting coronavirus infection. In this study, we generated the cell-based SARS-CoV-2 RdRp activity assay system by modifying a previously reported cell-based MERS-CoV RdRp activity assay system to screen for SARS-CoV-2 RdRp inhibitors. The assay system consisted of an expression plasmid encoding SARS-CoV-2 RdRp and an RdRp activity reporter plasmid. RdRp activity in the cells could be conveniently detected by luminescence after transfection. We confirmed that SARS-CoV-2 RdRp replicated double-stranded RNA using immunofluorescence staining and the inhibition of RdRp activity by remdesivir and lycorine using this system. Moreover, the Z-factor of this system was calculated to be 0.798, suggesting the reproducibility and reliability of the high-throughput screening system. Finally, we screened nucleoside and nucleotide analogs and identified adefovir dipivoxil, emtricitabine, telbivudine, entecavir hydrate, moroxydine and rifampin as novel SARS-CoV-2 RdRp inhibitors and therapeutic candidates for COVID-19 This system provides an effective high-throughput screening system platform for developing potential prophylactic and therapeutic drugs for COVID-19 and emerging coronavirus infections.
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BACKGROUND: Highly effective novel treatments need to be developed to suppress emerging coronavirus (CoV) infections such as COVID-19. The RNA dependent RNA polymerase (RdRp) among the viral proteins is known as an effective antiviral target. Lycorine is a phenanthridine Amaryllidaceae alkaloid isolated from the bulbs of Lycoris radiata (L'Hér.) Herb. and has various pharmacological bioactivities including antiviral function. PURPOSE: We investigated the direct-inhibiting action of lycorine on CoV's RdRp, as potential treatment for emerging CoV infections. METHODS: We examined the inhibitory effect of lycorine on MERS-CoV, SARS-CoV, and SARS-CoV-2 infections, and then quantitatively measured the inhibitory effect of lycorine on MERS-CoV RdRp activity using a cell-based reporter assay. Finally, we performed the docking simulation with lycorine and SARS-CoV-2 RdRp. RESULTS: Lycorine efficiently inhibited these CoVs with IC50 values of 2.123 ± 0.053, 1.021 ± 0.025, and 0.878 ± 0.022 µM, respectively, comparable with anti-CoV effects of remdesivir. Lycorine directly inhibited MERS-CoV RdRp activity with an IC50 of 1.406 ± 0.260 µM, compared with remdesivir's IC50 value of 6.335 ± 0.731 µM. In addition, docking simulation showed that lycorine interacts with SARS-CoV-2 RdRp at the Asp623, Asn691, and Ser759 residues through hydrogen bonding, at which the binding affinities of lycorine (-6.2 kcal/mol) were higher than those of remdesivir (-4.7 kcal/mol). CONCLUSIONS: Lycorine is a potent non-nucleoside direct-acting antiviral against emerging coronavirus infections and acts by inhibiting viral RdRp activity; therefore, lycorine may be a candidate against the current COVID-19 pandemic.
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Alcaloides de Amaryllidaceae/farmacología , Antivirales/farmacología , Fenantridinas/farmacología , ARN Polimerasa Dependiente del ARN/antagonistas & inhibidores , SARS-CoV-2/efectos de los fármacos , Adenosina Monofosfato/análogos & derivados , Alanina/análogos & derivados , Animales , Chlorocebus aethiops , Enlace de Hidrógeno , Coronavirus del Síndrome Respiratorio de Oriente Medio/efectos de los fármacos , Simulación del Acoplamiento Molecular , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/efectos de los fármacos , Células Vero , Proteínas ViralesRESUMEN
Severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), and coronavirus disease 2019 (COVID-19) are emerging zoonotic diseases caused by coronavirus (CoV) infections. The viral RNA-dependent RNA polymerase (RdRp) has been suggested as a valuable target for antiviral therapeutics because the sequence homology of CoV RdRp is highly conserved. We established a cell-based reporter assay for MERS-CoV RdRp activity to test viral polymerase inhibitors. The cell-based reporter system was composed of the bicistronic reporter construct and the MERS-CoV nsp12 plasmid construct. Among the tested nine viral polymerase inhibitors, ribavirin, sofosbuvir, favipiravir, lamivudine, zidovudine, valacyclovir, vidarabine, dasabuvir, and remdesivir, only remdesivir exhibited a dose-dependent inhibition. Meanwhile, the Z-factor and Z'-factor of this assay for screening inhibitors of MERS-CoV RdRp activity were 0.778 and 0.782, respectively. Ribavirin and favipiravir did not inhibit the MERS-CoV RdRp activity, and non-nucleoside HCV RdRp inhibitor, dasabuvir, partially inhibited MERS-CoV RdRp activity. Taken together, the cell-based reporter assay for MERS-CoV RdRp activity confirmed remdesivir as a direct inhibitor of MERS-CoV RdRp in cells. A cell-based MERS-CoV RdRp activity reporter assay is reliable and accurate for screening MERS-CoV RdRp-specific inhibitors. It may provide a valuable platform for developing antiviral drugs for emerging CoV infections.
RESUMEN
Kurarinone is a prenylated flavonone isolated from the roots of Sophora flavescens. Among its known functions, kurarinone has both anti-apoptotic and anti-inflammatory properties. Coronaviruses (CoVs), including HCoV-OC43, SARS-CoV, MERS-CoV, and SARS-CoV-2, are the causative agents of respiratory virus infections that range in severity from the common cold to severe pneumonia. There are currently no effective treatments for coronavirus-associated diseases. In this report, we examined the anti-viral impact of kurarinone against infection with the human coronavirus, HCoV-OC43. We found that kurarinone inhibited HCoV-OC43 infection in human lung fibroblast MRC-5 cells in a dose-dependent manner with an IC50 of 3.458 ± 0.101 µM. Kurarinone inhibited the virus-induced cytopathic effect, as well as extracellular and intracellular viral RNA and viral protein expression. Time-of-addition experiments suggested that kurarinone acted at an early stage of virus infection. Finally, we found that HCoV-OC43 infection increased the autophagic flux in MRC-5 cells; kurarinone inhibited viral replication via its capacity to impair the virus-induced autophagic flux. As such, we suggest that kurarinone may be a useful therapeutic for the treatment of diseases associated with coronavirus infection.
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Stephaniatetrandra and other related species of Menispermaceae are the major sources of the bis-benzylisoquinoline alkaloids tetrandrine (TET), fangchinoline (FAN), and cepharanthine (CEP). Although the pharmacological properties of these compounds include anticancer and anti-inflammatory activities, the antiviral effects of these compounds against human coronavirus (HCoV) remain unclear. Hence, the aims of the current study were to assess the antiviral activities of TET, FAN, and CEP and to elucidate the underlying mechanisms in HCoV-OC43-infected MRC-5 human lung cells. These compounds significantly inhibited virus-induced cell death at the early stage of virus infection. TET, FAN, and CEP treatment dramatically suppressed the replication of HCoV-OC43 as well as inhibited viral S and N protein expression. The virus-induced host response was reduced by compound treatment as compared with the vehicle control. Taken together, these findings demonstrate that TET, FAN, and CEP are potential natural antiviral agents for the prevention and treatment of HCoV-OC43 infection.
Asunto(s)
Antivirales/farmacología , Bencilisoquinolinas/farmacología , Infecciones por Coronavirus/virología , Coronavirus Humano OC43/efectos de los fármacos , Coronavirus Humano OC43/fisiología , Extractos Vegetales/farmacología , Stephania tetrandra/química , Bencilisoquinolinas/química , Línea Celular , Infecciones por Coronavirus/genética , Infecciones por Coronavirus/metabolismo , Coronavirus Humano OC43/genética , Citocinas/genética , Citocinas/metabolismo , Humanos , Extractos Vegetales/químicaRESUMEN
3-Acyl-2-phenylamino-1,4-dihydroquinolin-4(1H)-one derivatives were synthesized and evaluated to show high anti-MERS-CoV inhibitory activities. Among them, 6,8-difluoro-3-isobutyryl-2-((2,3,4-trifluorophenyl)amino)quinolin-4(1H)-one (6u) exhibits high inhibitory effect (IC50â¯=â¯86â¯nM) and low toxicity (CC50â¯>â¯25⯵M). Moreover, it shows good metabolic stability, low hERG binding affinity, no cytotoxicity, and good in vivo PK properties.
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Antivirales/farmacología , Coronavirus del Síndrome Respiratorio de Oriente Medio/efectos de los fármacos , Quinolonas/farmacología , Animales , Antivirales/síntesis química , Antivirales/química , Células CHO , Supervivencia Celular/efectos de los fármacos , Chlorocebus aethiops , Cricetulus , Relación Dosis-Respuesta a Droga , Humanos , Ratones , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Células 3T3 NIH , Quinolonas/síntesis química , Quinolonas/química , Relación Estructura-Actividad , Células VeroRESUMEN
Study on the 2-phenylchroman-4-one derivatives and their anti-MERS-CoVactivities.
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SIRT2, a member of the class III histone deacetylase family, has been identified as a tumor suppressor, which is associated with various cellular processes including metabolism and proliferation. However, the effects of SIRT2 on cancer cell migration caused by cytoskeletal rearrangement remain uncertain. Here we show that SIRT2 inhibits cell motility by suppressing actin polymerization. SIRT2 regulates actin dynamics through HSP90 destabilization and subsequent repression of LIM kinase (LIMK) 1/cofilin pathway. SIRT2 directly interacts with HSP90 and regulates its acetylation and ubiquitination. In addition, the deacetylase activity of SIRT2 is required for the regulation of actin polymerization and the ubiquitin-mediated proteasomal degradation of HSP90 induced by SIRT2.
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Actinas/química , Proteínas HSP90 de Choque Térmico/química , Proteínas HSP90 de Choque Térmico/metabolismo , Neoplasias/metabolismo , Sirtuina 2/metabolismo , Acetilación , Factores Despolimerizantes de la Actina/metabolismo , Línea Celular Tumoral , Movimiento Celular , Células HCT116 , Células HeLa , Humanos , Quinasas Lim/metabolismo , Neoplasias/genética , Multimerización de Proteína , Proteolisis , Transducción de Señal , Sirtuina 2/genética , UbiquitinaciónRESUMEN
von Hippel-Lindau-binding protein 1 (VBP1) physically interacts with pVHL, an E3-ubiquitin ligase, which degrades HIF-1α in an oxygen-dependent manner. HIF-1 is a key regulator of adaptive responses to a lack of oxygen that controls glucose metabolism, angiogenesis, proliferation, invasion, and metastasis. However, the role of VBP1 in pVHL-mediated degradation of HIF-1α is not yet known. In this study, we show that VBP1 enhances the stability of pVHL and facilitates pVHL-mediated ubiquitination of HIF-1α. Furthermore, VBP1 suppresses HIF-1α-induced epithelial-mesenchymal transition in vitro and tumor metastasis in vivo. These findings suggest that VBP1 is a bona fide tumor suppressor protein associated with HIF-1α regulation.
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Proteínas Portadoras/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Animales , Proteínas Portadoras/genética , Línea Celular Tumoral , Proteínas del Citoesqueleto , Células HCT116 , Células HEK293 , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Ratones Endogámicos C57BL , Chaperonas Moleculares , Metástasis de la Neoplasia , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Neoplasias Experimentales/genética , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/patología , Unión Proteica , Interferencia de ARN , Ubiquitinación , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genéticaRESUMEN
The subcellular localization of Bax plays a crucial role during apoptosis. In response to apoptotic stimuli, Bax translocates from the cytoplasm to the mitochondria, where it promotes the release of cytochrome c to the cytoplasm. In cells infected with HSV-1, apoptosis is triggered or blocked by diverse mechanisms. In this study, we demonstrate how HSV-1 ICP27 induces apoptosis and modulates mitochondrial membrane potential in HEK 293T cells. We found that ICP27 interacts with 14-3-3θ which sequesters Bax to the cytoplasm. In addition, ICP27 promotes the translocation of Bax to the mitochondria by inhibiting the interaction between 14-3-3θ and Bax. Our findings may provide a novel apoptotic regulatory pathway induced by ICP27 during HSV-1 infection. [BMB Reports 2017; 50(5): 257-262].
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Proteínas 14-3-3/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Proteínas 14-3-3/genética , Apoptosis/genética , Apoptosis/fisiología , Proteínas Portadoras/metabolismo , Caspasas/metabolismo , Citocromos c/metabolismo , Citoplasma , Células HEK293 , Herpesvirus Humano 1 , Humanos , Mitocondrias , Sistemas de Translocación de Proteínas/metabolismo , Sistemas de Translocación de Proteínas/fisiología , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
Herpes simplex virus type 1 ICP27 is a multifunctional protein responsible for viral replication, late gene expression, and reactivation from latency. ICP27 interacts with various cellular proteins, including Daxx. However, the role of interaction between ICP27 and Daxx is largely unknown. Since Daxx is known to repress NF-κB activity, there is a possibility that ICP27 may influence the inhibitory effect of Daxx on NF-κB activity. In this study, we tested whether ICP27 affects the NF-κB activity through its interaction with Daxx. Interestingly, ICP27 enhanced the Daxx-mediated repression of NF-κB activity. In addition, we found that sumoylation of Daxx regulates its interaction with p65. ICP27 binds to Daxx, inhibits Daxx sumoylation, and enhances p65 deacetylation induced by Daxx. Consequently, ICP27 represses the NF-ï«B activity, by elevating the inhibitory effect of Daxx on NF-κB activity through desumoylation of Daxx. [BMB Reports 2017; 50(5): 275-280].
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Proteínas Inmediatas-Precoces/fisiología , Proteínas Nucleares/metabolismo , Proteínas Adaptadoras Transductoras de Señales/fisiología , Proteínas Co-Represoras , Expresión Génica , Regulación Viral de la Expresión Génica/genética , Células HEK293 , Herpesvirus Humano 1 , Humanos , Chaperonas Moleculares , FN-kappa B/metabolismo , FN-kappa B/fisiología , Proteínas Nucleares/fisiología , Transducción de Señal , Sumoilación/fisiología , Factor de Transcripción ReIA/metabolismo , Factores de Transcripción/metabolismo , Proteínas Virales/genética , Replicación Viral/efectos de los fármacosRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: A medicinal herb Tribulus terrestris Linn has been used to treat various diseases including hepatocellular carcinoma. The aim of the present study was to investigate the anticancer activity of Tribulus terrestris Linn (TT) in liver cancer cells. MATERIALS AND METHODS: The antitumor activity of aqueous TT extract was analyzed by testing the cytotoxicity and the effect on clonogenecity in HepG2 cells. Apoptosis and cell cycle arrest induced by TT were dissected by flow cytometry and its inhibitory effect on NF-κB activity was determined by analyzing the expression levels of NF-κB/IκB subunit proteins. The suppression of NF-κB-regulated gene expression by TT was assessed by RT-PCR. RESULTS: TT extract repressed clonogenecity and proliferation, induced apoptosis, and enhanced accumulation in the G0/G1 phase of liver cancer cells. It also turned out that TT extract inhibited NF-κB-dependent reporter gene expression and NF-κB subunit p50 expression, while it enhanced the cellular level of IκBα by inhibiting the phosphorylation and degradation of IκBα. In addition, IKK activity was inhibited in a dose-dependent manner. Furthermore, TT extract suppressed the transcription of genes associated with cell cycle regulation, anti-apoptosis, and invasion. CONCLUSION: These data showed that TT extract blocks proliferation and induces apoptosis in human liver cancer cells through the inhibition of NF-κB signaling. Aqueous TT extract can be used as an anticancer drug for hepatocellular carcinoma patients.
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Antineoplásicos Fitogénicos/uso terapéutico , Proliferación Celular/efectos de los fármacos , Hepatoblastoma/tratamiento farmacológico , Neoplasias Hepáticas/tratamiento farmacológico , FN-kappa B/metabolismo , Extractos Vegetales/uso terapéutico , Tribulus , Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo , Expresión Génica , Genes Reporteros , Células Hep G2 , Hepatoblastoma/genética , Hepatoblastoma/metabolismo , Humanos , Quinasa I-kappa B/antagonistas & inhibidores , Proteínas I-kappa B/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Inhibidor NF-kappaB alfa , Invasividad Neoplásica , Fosforilación , Fitoterapia , Extractos Vegetales/farmacología , Transducción de Señal/efectos de los fármacos , Ensayo de Tumor de Célula MadreRESUMEN
Silica nanorods were fabricated with single-walled carbon nanotubes (SWCNTs) via ultrasound. The diameter of the resulting SWCNT-silica particles ranged from 60 to 70 nm. The morphology of this composite material was investigated via scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The individual SWCNTs are uniformly coated with silica and formed a unique nanocomposite material. The important role of ultrasound and the mechanism of silica layer formation on SWCNTs were explained via the hydrolysis of the silica source and the adsorption of the siloxane groups on the SWCNT surfaces under ultrasound irradiation. The amino-functionalized silica nanorods were demonstrated as non-viral vectors for gene delivery.
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Técnicas de Transferencia de Gen , Nanocompuestos/química , Nanotubos de Carbono/química , Dióxido de Silicio/química , Supervivencia Celular/efectos de los fármacos , Células HeLa , Humanos , Microscopía Confocal , Microscopía Electrónica de Transmisión , Nanotecnología , Tamaño de la Partícula , Dióxido de Silicio/farmacología , Espectrometría por Rayos X , Espectrometría Raman , UltrasonidoRESUMEN
A new method is proposed for the fabrication of fluorescence-labeled and amine-modified silica nanoparticles for application as nonviral vectors in gene delivery. Highly monodisperse, stable fluorescent silica nanoparticles were prepared using 2,5-bis(5-tert-butyl-2-benzoxazolyl)thiophene and the water-in-oil microemulsion method. The green-fluorescent-protein gene can be easily combined onto the positively charged surfaces of nanoparticles to form a nanoparticle-DNA complex. The nanoparticle-DNA complex successfully passed through various barriers into the HeLa and HEK 293 K cells. The cytotoxicity of the PEI-coated and BBOT-encapsulated silica nanoparticles on both the HeLa and HEK 293T cell lines was found to be at an acceptable level for use as gene carriers when the particle concentration was below 125 microg/ml. The fluorescence intracellular images confirm the successful delivery of the nanoparticle-DNA complex and gene expression. The present work suggests the potential use of dye-incorporated silica nanoparticles in nonviral gene delivery.
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ADN/administración & dosificación , ADN/química , Colorantes Fluorescentes/química , Técnicas de Transferencia de Gen , Nanopartículas/química , Dióxido de Silicio/química , Proteínas Fluorescentes Verdes/química , Proteínas Fluorescentes Verdes/genética , Plásmidos/genéticaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: A medicinal herb Curcuma longa Linn has been used for treating various liver diseases caused by hepatitis B virus (HBV) in Asia. AIM OF THE STUDY: The study was performed in order to investigate the antiviral activity of Curcuma longa Linn against HBV replication in liver cells. MATERIALS AND METHODS: Aqueous extract of Curcuma longa Linn (CLL) was prepared and used to analyze its antiviral activity against HBV replication in HepG 2.2.15 cells, which contain HBV genomes. The inhibitory effect of CLL on HBV replication was examined by testing the levels of secreted HBV surface antigens (HBsAg), HBV DNAs, and HBV RNAs in HepG 2.2.15 cells using ELISA, Southern blot, and Northern blot analyses. Cytotoxic activities of CLL extract on various liver cells were analyzed by MTT assay. To dissect the inhibitory mechanism of CLL extract on HBV replication, the levels of p53 protein and p53 mRNAs were analyzed by Western blot and RT-PCR in HepG 2.2.15 cells. The repression of CLL extract on HBV transcription was analyzed by RT-PCR and CAT assay. RESULTS: CLL extract repressed the secretion of HBsAg from HepG 2.2.15 cells. CLL extract also suppressed the production of HBV particles and the level of intracellular HBV RNAs in HepG 2.2.15 cells, suggesting that CLL extract inhibits HBV replication. We found that the anti-HBV activity of CLL extract is mediated through enhancing the cellular accumulation of p53 protein by transactivating the transcription of p53 gene as well as increasing the stability of p53 protein. It turned out that CLL extract repressed the transcription of HBx gene by suppressing HBV enhancer I and X promoter through p53 protein. In addition, CLL extract did not have any cytotoxic effects on liver cells. CONCLUSION: These data showed that CLL extract represses HBV replication through enhancing the level of p53 protein. CLL extract can be used as a safe and specific drug for patients with liver diseases caused by HBV infection.