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1.
Chem Biol Interact ; 320: 109029, 2020 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-32119866

RESUMEN

Geraniol (GOH), like other plant-derived natural bioactive compounds, has been found to possess antiproliferative properties that are essential to cope with malignant tumors. However, the mechanisms of molecular action of GOH are not fully elucidated. The aim of this study was to evaluate the effect of GOH on some oxidative parameters in human tumor cell lines (HepG2 and A549). Cytotoxicity evaluated in cell lines by the MTT assay, genotoxicity by the comet assay, and lipid peroxidation by the TBARS. The activities of antioxidant the enzymes, superoxide dismutase (SOD), catalase (CAT) and glutathione-S-transferase (GST), were also analyzed. Additionally, intracellular reactive oxygen species (ROS), nitric oxide, and lactate production were determined in HepG2 cells. Both tumor cell lines showed a clear concentration-dependent response to GOH in several of the parameters evaluated. Lipids turned out to be more sensitive than DNA to oxidative damage induced by GOH. TBARS levels increased with respect to control (p < 0.05) by 33% and 122% in HepG2 and A549 cells, respectively treated with 200 µM GOH. However, GOH caused a statistically significant decrease in SOD and CAT activities in HepG2 cells only. GST was not affected in any cell lines. GOH induced the production of ROS but not nitric oxide in HepG2, which shows that ROS were mainly responsible for oxidative damage. Lactate release increased statistically significantly compared to control (p < 0.001), by 41% and 86% at 200 and 800 µM GOH respectively, showing that this monoterpene also affected the glycolytic pathway in HepG2 cells. These results suggest that oxidative stress could mediate the anti-proliferative effects of GOH in HepG2 and A549 cells.


Asunto(s)
/farmacología , Proliferación Celular , Estrés Oxidativo/efectos de los fármacos , Células A549 , /química , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Hep G2 , Humanos , Estructura Molecular
2.
Nature ; 579(7798): 260-264, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32132711

RESUMEN

The production of pore-forming toxins that disrupt the plasma membrane of host cells is a common virulence strategy for bacterial pathogens such as methicillin-resistant Staphylococcus aureus (MRSA)1-3. It is unclear, however, whether host species possess innate immune mechanisms that can neutralize pore-forming toxins during infection. We previously showed that the autophagy protein ATG16L1 is necessary for protection against MRSA strains encoding α-toxin4-a pore-forming toxin that binds the metalloprotease ADAM10 on the surface of a broad range of target cells and tissues2,5,6. Autophagy typically involves the targeting of cytosolic material to the lysosome for degradation. Here we demonstrate that ATG16L1 and other ATG proteins mediate protection against α-toxin through the release of ADAM10 on exosomes-extracellular vesicles of endosomal origin. Bacterial DNA and CpG DNA induce the secretion of ADAM10-bearing exosomes from human cells as well as in mice. Transferred exosomes protect host cells in vitro by serving as scavengers that can bind multiple toxins, and improve the survival of mice infected with MRSA in vivo. These findings indicate that ATG proteins mediate a previously unknown form of defence in response to infection, facilitating the release of exosomes that serve as decoys for bacterially produced toxins.


Asunto(s)
Proteínas Relacionadas con la Autofagia/metabolismo , Toxinas Bacterianas/metabolismo , Exosomas/metabolismo , Células A549 , Proteína ADAM10/metabolismo , Animales , Toxinas Bacterianas/farmacología , Supervivencia Celular/efectos de los fármacos , ADN Bacteriano/farmacología , Exosomas/efectos de los fármacos , Exosomas/ultraestructura , Femenino , Células HEK293 , Humanos , Masculino , Staphylococcus aureus Resistente a Meticilina/patogenicidad , Staphylococcus aureus Resistente a Meticilina/fisiología , Ratones , Ratones Endogámicos C57BL , Infecciones Estafilocócicas/mortalidad
3.
Med Sci Monit ; 26: e918216, 2020 Mar 04.
Artículo en Inglés | MEDLINE | ID: mdl-32129321

RESUMEN

BACKGROUND Chemoresistance is a primary hindrance for current cancer treatments. The influence of abnormal mitochondria in chemotherapy resistance is not well known. To explore the correlation between mitochondria and acquired chemoresistance, this work studied alterations in mitochondrial dynamics, biogenesis, and functions for paclitaxel-resistant cancer cell line A549/Taxol and its parental line A549. MATERIAL AND METHODS Mitochondrial morphology was observed by transmission electron microscopy and confocal microscopy. We measured the mitochondrial mass and mitochondrial membrane potential using fluorescent dyes. The glucose metabolic profile and ATP (adenosine triphosphate) content were determined by bioluminescent cell assays. Seahorse bio-energy analyzer XF24 was used to detect the mitochondrial respiratory function. The expressions of mitochondrial dynamics and biogenesis related genes were quantified using real-time polymerase chain reaction. RESULTS We observed fusion morphology of the mitochondrial network in A549/Taxol cells, with upregulation of fusion genes (Mfn1 and Mfn2) and downregulation of fission gene Fis1. In A549/Taxol cells, mitochondrial mass showed a significant decrease, while the mitochondrial biogenesis pathway was strongly activated. Despite the decreased mitochondrial membrane potential, the capability for mitochondrial respiration was not impaired in A549/Taxol cells. CONCLUSIONS Our study revealed a series changes of mitochondrial characteristics in paclitaxel-resistant cells. Mfn1 and Mfn2 and PGC-1alpha increased, while Fis1 expression and mitochondrial oxidative phosphorylation decreased in A549/Taxol cell lines. These changes to mitochondrial fusion, fission, and biological function contributed to the occurrence of paclitaxel resistance in tumor cells which induced paclitaxel resistance.


Asunto(s)
Resistencia a Antineoplásicos , Dinámicas Mitocondriales , Biogénesis de Organelos , Paclitaxel/farmacología , Células A549 , GTP Fosfohidrolasas/metabolismo , Humanos , Potencial de la Membrana Mitocondrial , Microscopía Confocal , Microscopía Electrónica de Transmisión , Mitocondrias/ultraestructura , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Proteínas Mitocondriales/metabolismo , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/metabolismo
4.
Chem Biol Interact ; 319: 109024, 2020 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-32097614

RESUMEN

Silicosis is an occupational pulmonary fibrosis that is caused by inhalation of silica (SiO2), and there are no effective drugs to treat this disease. Tanshinone IIA (Tan IIA), a natural product, has been reported to possess antioxidant and anti-fibrotic properties in various diseases. The purpose of the current study was to examine Tan IIA's protective effects against silica-induced pulmonary fibrosis and to explore the underlying mechanisms. We found that in vivo treatment with Tan IIA significantly relieved silica-induced lung fibrosis in a silicosis rat model by histological and immunohistochemical analyses. Further, in vitro mechanistic investigations, mainly using western blot and immunofluorescence analyses, revealed that Tan IIA administration markedly inhibited the silica-induced epithelial-mesenchymal transition (EMT) and transforming growth factor-ß1 (TGF-ß1)/Smad signaling pathway and also reduced silica-induced oxidative stress and activated the nuclear factor erythroid 2-related factor-2 (Nrf2) signaling pathway in A549 and human bronchial epithelial (HBE) cells. Furthermore, through transfection with siRNA, we demonstrate that Nrf2 activation partially mediates the suppression effects of Tan IIA on EMT and TGF-ß1/Smad signaling pathway activation induced by silica exposure, thus mediating the anti-fibrotic effects of Tan IIA against silica-induced pulmonary fibrosis. In our study, Tan IIA has been identified as a possible anti-oxidative and anti-fibrotic drug for silicosis.


Asunto(s)
/farmacología , Transición Epitelial-Mesenquimal/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Fibrosis Pulmonar/tratamiento farmacológico , Transducción de Señal/efectos de los fármacos , Proteínas Smad/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Células A549 , Animales , Antioxidantes/metabolismo , Línea Celular , Línea Celular Tumoral , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Humanos , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Masculino , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/metabolismo , Ratas , Ratas Sprague-Dawley , Dióxido de Silicio/farmacología
5.
Life Sci ; 246: 117366, 2020 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-32001266

RESUMEN

BACKGROUND: Hydroxychloroquine exhibits synergistic anticancer properties as an adjuvant. However, the role and molecular mechanisms underlying of HCQ as monotherapy for lung adenocarcinoma (LUAD) have yet to be elucidated. METHODS: We assessed the antitumor effects of HCQ in LUAD cells through a series of in vitro and in vivo assays. GEO database and R packages were used to predict molecular mechanisms of HCQ in the treatment of lung adenocarcinoma, followed by verification of gene expression and subcellular localization via immunoblotting, immunofluorescent and immunohistochemistry assays. RESULTS: We showed the phenotypic effects that HCQ inhibited cell growth, induced apoptosis and cell cycle arrest at G1/S transition in A549 and PC-9 cells, which was associated with inhibition of CDK2, CDK4, CyclinD1 and CyclinE, but up-regulation of p21 and p27Kip1. Bioinformatic analysis predicted that 63 targets related to HCQ and LUAD were mainly enriched in JAK-STAT and FoxO pathways. Then, we observed that HCQ decreased the phosphorylation of STAT3, but increased the expression of FoxO3a and its accumulation in the nucleus. The specific STAT3 inhibitor cryptotanshinon augmented the HCQ-induced upregulation and nuclear translocation of FoxO3a. In addition, HCQ increased the expression of p27Kip1, which was impaired by FoxO3a blockade with siRNA. Finally, ablation of p27Kip1 expression abrogated the cytotoxicity of HCQ. More importantly, similar results were further confirmed in vivo. CONCLUSIONS: Taken together, this study suggests that STAT3/FoxO3a/p27Kip1 signaling pathway is involved in the anticancer effects of HCQ, and provides preliminary evidence for therapeutic prospects of HCQ alone in LUAD.


Asunto(s)
Adenocarcinoma/tratamiento farmacológico , Antineoplásicos/uso terapéutico , Proteína Forkhead Box O3/metabolismo , Hidroxicloroquina/uso terapéutico , Neoplasias Pulmonares/tratamiento farmacológico , Factor de Transcripción STAT3/metabolismo , Células A549 , Transporte Activo de Núcleo Celular/efectos de los fármacos , Adenocarcinoma/metabolismo , Animales , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Pulmonares/metabolismo , Ratones Desnudos , Trasplante de Neoplasias
6.
Life Sci ; 246: 117428, 2020 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-32057901

RESUMEN

PURPOSE: Arl4c is overexpressed in several cancer tissues and is involved in cancer development. Nevertheless, the exact mechanism that regulates Arl4c expression in lung cancer has not been fully elucidated. The aim of this study was to investigate the regulatory mechanism of Arl4c and to explore potential chemotherapeutic drugs targeting Arl4c. METHODS: Immunohistochemistry was used to examine Arl4c expression levels in human lung adenocarcinoma cancer specimens. Protein expression was detected by western blot. Overexpression of Arl4c-Flag protein was used to detect the ubiquitination of Arl4c. A short interfering RNA against Arl4c was used for gene silencing. RESULTS: Arl4c was overexpressed in lung cancer tissues, and knockdown of Arl4c expression by siRNA decreased lung cancer A549 and 95-D cell proliferation. In addition, Arl4c expression was downregulated via inhibition of the AKT pathway in A549 and 95-D cells, whereas exposure to benzo (a) pyrene (a carcinogen in smoke) increased Arl4c expression in 16HBE cells via AKT activation. Finally, we found that chemotherapy drug hydroxycamptothecin (HCPT) could decrease Arl4c expression levels by inhibiting the activation of the AKT pathway in A549 and 95-D cells. Moreover, accumulation of ubiquitinated Arl4c protein was increased by HCPT and LY294002 (an AKT inhibitor) treatment whereas the proteasome inhibitor MG-132 attenuated the inhibitory effect of HCPT and LY294002 on Arl4c expression. CONCLUSION: Here, we highlighted the AKT pathway as an important regulatory pathway for Arl4c expression in lung cancer cells and identified HCPT as a promising drug for lung adenocarcinoma treatment that functioned by targeting Arl4c expression.


Asunto(s)
Factores de Ribosilacion-ADP/metabolismo , Adenocarcinoma del Pulmón/metabolismo , Adenocarcinoma/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Células A549 , Western Blotting , Camptotecina/análogos & derivados , Camptotecina/farmacología , Línea Celular Tumoral , Cromonas/farmacología , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Leupeptinas/farmacología , Morfolinas/farmacología , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Fumar/efectos adversos
7.
Chem Commun (Camb) ; 56(18): 2695-2698, 2020 Mar 03.
Artículo en Inglés | MEDLINE | ID: mdl-32030397

RESUMEN

A BODIPY-based fluorescent sensor PS with an NO4S2 podand ligand was studied for the selective detection of Pt2+ over 21 cations as well as selected platinum drugs in aqueous medium. The platinum sensor PS shows 28-fold, 22-fold and 14-fold fluorescence turn-on enhancements to Pt2+, cisplatin and nedaplatin, and was thereby employed to detect platinum drugs in A-549 human lung cancer cells.


Asunto(s)
Compuestos de Boro/química , Cisplatino/análisis , Colorantes Fluorescentes/química , Neoplasias Pulmonares/diagnóstico por imagen , Platino (Metal)/análisis , Células A549 , Cisplatino/uso terapéutico , Humanos , Ligandos , Neoplasias Pulmonares/tratamiento farmacológico , Estructura Molecular , Imagen Óptica , Espectrometría de Fluorescencia
9.
J Enzyme Inhib Med Chem ; 35(1): 555-564, 2020 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31967481

RESUMEN

In this paper, a series of novel 3-methyl-quinazolinone derivatives was designed, synthesised and evaluated for antitumor activity in vitro on wild type epidermal growth factor receptor tyrosine kinase (EGFRwt-TK) and three human cancer cell lines including A549, PC-3, and SMMC-7721. The results displayed that some of the compounds had good activities, especially 2-{4-[(3-Fluoro-phenylimino)-methyl]-phenoxymethyl}-3-methyl-3H-quinazolin-4-one (5 g), 2-{4-[(3,4-Difluoro-phenylimino)-methyl]-phenoxymethyl}-3-methyl-3H-quinazolin-4-one (5k) and 2-{4-[(3,5-Difluoro-phenylimino)-methyl]-phenoxymethyl}-3-methyl-3H-quinazolin-4-one (5 l) showed high antitumor activities against three cancer cell lines. Moreover, compound 5k could induce late apoptosis of A549 cells at high concentrations and arrest cell cycle of A549 cells in the G2/M phase at tested concentrations. Also, compound 5k could inhibit the EGFRwt-TK with IC50 value of 10 nM. Molecular docking data indicates that the compound 5k may exert inhibitory activity by forming stable hydrogen bonds with the R817, T830 amino acid residues and cation-Π interaction with the K72 residue of EGFRwt-TK.


Asunto(s)
Antineoplásicos/farmacología , Diseño de Drogas , Inhibidores de Proteínas Quinasas/farmacología , Quinazolinonas/farmacología , Células A549 , Antineoplásicos/síntesis química , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Humanos , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Quinazolinonas/síntesis química , Quinazolinonas/química , Relación Estructura-Actividad
10.
Chem Biol Interact ; 317: 108962, 2020 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-31982400

RESUMEN

Quaternary ammonium compounds (e.g., benzalkonium chloride (BAC) and cetylpyridinium chloride (CPC)) constitute a group of cationic surfactants are widely used for personal hygiene and medical care despite the potential pulmonary toxicity. To examine whether BAC and CPC aerosols deposited in the alveolar region alter pulmonary function, we studied the effects on pulmonary surfactant using two-step in vitro models; cytotoxicity using A549 alveolar epithelial cell and changes in surface activity of the pulmonary surfactant monolayer using both Surfacten® and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Cell viability was decreased with BAC and CPC dose-dependently. A comparison of cytotoxicity among BAC homologues with different length of alkyl chain showed that C16-BAC, which has the longest alkyl chain, was more cytotoxic than C12- or C14-BAC. Caspase-3/7 activity and cleaved form of caspase-3 and PARP were increased in BAC- and CPC-exposed cells. The elevated caspase-3/7 activity and their cleaved active forms were abolished by caspase-3-inhibitor. Furthermore, we examined the features of the surface pressure/trough area (π-A) isotherm by the Langmuir-Wilhelmy method and atomic force microscopy (AFM) images of lipid monolayers on a subphase containing BAC, CPC, or pyridinium chloride (PC, as a control). The π-A isotherms showed that addition of BAC or CPC yielded dose-dependent increases in surface pressure without compression, indicating that BAC and CPC expand the isotherm to larger areas at lower pressure. The collapse pressure diminished with increasing concentration of CPC. Topographic images indicated that BAC and CPC resulted in smaller condensed lipid domains compared to the control. Conversely, PC without hydrocarbon tail group, showed no cytotoxicity and did not change the isotherms and AFM images. These results indicate that BAC and CPC cause cell death via caspase-3-dependent apoptotic pathway in A549 cells and alter the alveolar surfactant activity. These effects can be attributed to the long alkyl chain of BAC and CPC.


Asunto(s)
Apoptosis/efectos de los fármacos , Compuestos de Benzalconio/farmacología , Cetilpiridinio/farmacología , Células Epiteliales/efectos de los fármacos , Pulmón/citología , Mucosa Respiratoria/citología , Células A549 , Compuestos de Benzalconio/química , Supervivencia Celular/efectos de los fármacos , Cetilpiridinio/química , Humanos , Tensoactivos/metabolismo
11.
Anal Bioanal Chem ; 412(3): 601-609, 2020 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-31897558

RESUMEN

Numerous studies have shown that exosomes are closely related to the pathogenesis of various diseases, especially cancers. Therefore, a rapid and sensitive method for exosome detection will be of great importance for the diagnosis and prognosis of diseases. We report here a method for exosome detection based on the CD63 aptamer and clustered regular interspaced short palindromic repeats (CRISPR)/Cas12a system. This method consists mainly of exosomal membrane protein recognition based on the CD63 aptamer and signal amplification based on CRISPR/Cas12a. The CD63 aptamer, as an easily adaptable nucleic acid strand, is responsible for the conversion of the amounts of exosomes into nucleic acid detection, whereas CRISPR/Cas12a is responsible for highly specific nucleic acid signal amplification. The detection range of the method was determined as 3 × 103-6 × 107 particles per microliter. Additionally, we successfully applied this method to detect exosomes in clinical samples from both healthy individuals and patients with lung cancer, and the results were highly consistent with those obtained by nanoparticle tracking analysis. In general, this method provides a highly sensitive and specific method for the detection of exosomes and offers an avenue toward future exosome-based diagnosis of diseases.


Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Exosomas/química , Tetraspanina 30/análisis , Células A549 , Sistemas CRISPR-Cas , Exosomas/patología , Humanos , Neoplasias Pulmonares/patología , Técnicas de Amplificación de Ácido Nucleico/métodos
12.
Anticancer Res ; 40(1): 323-333, 2020 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-31892583

RESUMEN

BACKGROUND/AIM: Despite the Warburg effect, mitochondria play an essential role in the survival and maintenance of cancer cells. Thus, mitochondria have been considered a target for anticancer agents. Here, we identified a mitochondria-targeting anticancer agent from natural products. MATERIALS AND METHODS: Morphological and functional changes in mitochondria were determined by a fluorescence-based High Content Imaging System. Using human non-small cell lung cancer (NSCLC) cell lines (H1299, H226B, and A549), cell viability and colony formation assays, cell cycle analysis, and immunoblotting were performed to determine cytotoxic and proapoptotic effects of papuamine. RESULTS: Using a natural product chemical library, we identified papuamine as an active compound to inhibit viability and ATP production of NSCLC cells. Papuamine depleted intracellular ATP by causing mitochondrial dysfunction, as indicated by the loss of the mitochondrial membrane potential and increased mitochondrial superoxide generation. Papuamine significantly inhibited viability and colony formation of NSCLC cells by inducing apoptosis. CONCLUSION: Papuamine has a potential as a novel mitochondria-targeting anticancer agent.


Asunto(s)
Alcaloides/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Mitocondrias/patología , Células A549 , Adenosina Trifosfato/metabolismo , Adenilato Quinasa/metabolismo , Alcaloides/química , Alcaloides/farmacología , Carcinoma de Pulmón de Células no Pequeñas/patología , Supervivencia Celular/efectos de los fármacos , Humanos , Neoplasias Pulmonares/patología , Mitocondrias/efectos de los fármacos , Ensayo de Tumor de Célula Madre , Regulación hacia Arriba/efectos de los fármacos
13.
Phytochemistry ; 171: 112247, 2020 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-31927201

RESUMEN

Four previously undescribed acylated iridoid glucosides, linaburiosides A‒D, one undescribed iridoid, 7-deoxyiridolactonic acid, and one known acylated iridoid glucoside, iridolinarin C, were isolated from the aerial parts of a Mongolian traditional herbal medicine, Linaria buriatica. Linaburiosides A‒D had an acyl moiety corresponding to 7-deoxyiridolactonic acid. Detailed spectroscopic analyses of linaburiosides A‒D and 7-deoxyiridolactonic acid led to the assignment of their structures. The absolute configuration of 7-deoxyiridolactonic acid was elucidated by application of the PGME method; those of linaburiosides A‒D were assigned on the basis of chemical conversions, as well as application of the modified Mosher's method. The absolute configuration of iridolinarin C was also elucidated in this study. Anti-inflammatory and antiproliferative activities of isolated compounds and their derivatives were evaluated.


Asunto(s)
Antiinflamatorios/farmacología , Glucósidos/farmacología , Iridoides/farmacología , Linaria/química , Fitoquímicos/farmacología , Células A549 , Acilación , Antiinflamatorios/química , Antiinflamatorios/aislamiento & purificación , Glucósidos/química , Glucósidos/aislamiento & purificación , Humanos , Interleucina-1beta/antagonistas & inhibidores , Interleucina-1beta/biosíntesis , Iridoides/química , Iridoides/aislamiento & purificación , Lipopolisacáridos/antagonistas & inhibidores , Lipopolisacáridos/farmacología , Células MCF-7 , Microglía/efectos de los fármacos , Microglía/metabolismo , Conformación Molecular , Fitoquímicos/química , Fitoquímicos/aislamiento & purificación , Células Tumorales Cultivadas
14.
Chem Commun (Camb) ; 56(7): 1082-1084, 2020 Jan 23.
Artículo en Inglés | MEDLINE | ID: mdl-31894763

RESUMEN

G-quadruplexes (G4) are non-canonical nucleic acid structures with important implications in biology. Based on an α-helical fragment of the RHAU helicase that displays high specificity for parallel-stranded G-quadrplexes, herein we demonstrate its head-to-tail cyclization by a high-efficiency ligase. The cyclic peptide exhibits superior stability and binding affinity to a G-quadruplex, and can serve as an excellent investigational tool for chemical biology applications.


Asunto(s)
ARN Helicasas DEAD-box/metabolismo , ADN/metabolismo , G-Cuádruplex , Fragmentos de Péptidos/metabolismo , Péptidos Cíclicos/metabolismo , Células A549 , Ciclización , ARN Helicasas DEAD-box/química , ADN/genética , Humanos , Oldenlandia/enzimología , Fragmentos de Péptidos/química , Péptido Sintasas/química , Péptidos Cíclicos/síntesis química , Unión Proteica , Estabilidad Proteica
15.
Bratisl Lek Listy ; 121(1): 31-36, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-31950837

RESUMEN

OBJECTIVES: One of the best approaches for recognition of protein function is the induction of mutations for a gene knockout. In line with this strategy, gene editing tools allow researchers to induce these mutations. Lung cancer is one of the leading causes of death worldwide. ZEB1 and ZEB2 genes are the candidates for this disease. METHODS: The ZEB1 and ZEB2 knockout in the non-small cell lung cancer cell line (A549 cell) was investigated. Purification of recombination plasmids was performed from bacteria and then was transported to the A549 cell line. The deletion of ZEB1 and ZEB2 were examined by PCR. RESULTS: The results demonstrated the mutation and deletion in ZEB1 and ZEB2 genes. Based on the findings of this study, A549 cells were transfected with the vectors carrying the sgRNA/Cas9, simultaneously. The DNA fragment demonstrated the presence of indels in target sites as well as provided the potential of CRISPR/Cas9 system. CONCLUSION: CRISPR/Cas9 offers a great potential as an efficient technique for editing of ZEB1 and ZEB2 genes in A549 cell line (Tab. 1, Fig. 6, Ref. 44).


Asunto(s)
Sistemas CRISPR-Cas , Carcinoma de Pulmón de Células no Pequeñas , Edición Génica , Neoplasias Pulmonares , Caja Homeótica 2 de Unión a E-Box con Dedos de Zinc , Homeobox 1 de Unión a la E-Box con Dedos de Zinc , Células A549 , Carcinoma de Pulmón de Células no Pequeñas/genética , Edición Génica/métodos , Técnicas de Inactivación de Genes , Humanos , Neoplasias Pulmonares/genética , Caja Homeótica 2 de Unión a E-Box con Dedos de Zinc/genética , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/genética
16.
Cancer Sci ; 111(3): 951-961, 2020 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-31925985

RESUMEN

Lung adenocarcinoma is the most common histological type of lung cancer and is classified into adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA) and invasive adenocarcinoma (IA). Atypical adenomatous hyperplasia (AAH) lesions are possible precursors to adenocarcinoma. However, the mechanism underlying the stepwise continuum of lung adenocarcinoma is unclear. In this study, the involvement of ADP-ribosylation factor (ARF)-like (ARL) 4C (ARL4C), a member of the small GTP-binding protein family, in the progression of lung adenocarcinoma and the possibility of ARL4C as a molecular target for lung cancer therapy were explored. ARL4C was frequently expressed in AAH and ARL4C expression in immortalized human small airway epithelial cells promoted cell proliferation and suppressed cell death. In addition, ARL4C was expressed with increased frequency in AIS, MIA and IA in a stage-dependent manner, and the expression was correlated with histologic grade, fluorine-18 fluorodeoxyglucose uptake and poor prognosis. An anti-sense oligonucleotide (ASO) against ARL4C (ARL4C ASO-1316) inhibited RAS-related C3 botulinum toxin substrate activity and nuclear import of Yes-associated protein and transcriptional coactivator with PDZ-binding motif, and suppressed in vitro proliferation and migration of lung cancer cells with KRAS or epidermal growth factor receptor (EGFR) mutations. In addition, transbronchial administration of ARL4C ASO-1316 suppressed orthotopic tumor formation induced by these cancer cells. Thus, ARL4C is involved in the initiation of the premalignant stage and is associated with the stepwise continuum of lung adenocarcinoma. ARL4C ASO-1316 would be useful for lung adenocarcinoma patients expressing ARL4C regardless of the KRAS or EGFR mutation.


Asunto(s)
Factores de Ribosilacion-ADP/genética , Adenocarcinoma del Pulmón/genética , Neoplasias Pulmonares/genética , Células A549 , Adenocarcinoma/genética , Adenocarcinoma/patología , Adenocarcinoma del Pulmón/patología , Anciano , Animales , Movimiento Celular/genética , Proliferación Celular/genética , Progresión de la Enfermedad , Células Epiteliales/patología , Femenino , Humanos , Hiperplasia/genética , Hiperplasia/patología , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Lesiones Precancerosas/genética , Lesiones Precancerosas/patología , Activación Transcripcional/genética
17.
Nat Commun ; 11(1): 164, 2020 01 09.
Artículo en Inglés | MEDLINE | ID: mdl-31919360

RESUMEN

Host dependency factors that are required for influenza A virus infection may serve as therapeutic targets as the virus is less likely to bypass them under drug-mediated selection pressure. Previous attempts to identify host factors have produced largely divergent results, with few overlapping hits across different studies. Here, we perform a genome-wide CRISPR/Cas9 screen and devise a new approach, meta-analysis by information content (MAIC) to systematically combine our results with prior evidence for influenza host factors. MAIC out-performs other meta-analysis methods when using our CRISPR screen as validation data. We validate the host factors, WDR7, CCDC115 and TMEM199, demonstrating that these genes are essential for viral entry and regulation of V-type ATPase assembly. We also find that CMTR1, a human mRNA cap methyltransferase, is required for efficient viral cap snatching and regulation of a cell autonomous immune response, and provides synergistic protection with the influenza endonuclease inhibitor Xofluza.


Asunto(s)
Predisposición Genética a la Enfermedad/genética , Interacciones Huésped-Patógeno/genética , Virus de la Influenza A/patogenicidad , Gripe Humana/genética , Gripe Humana/patología , Células A549 , Proteínas Adaptadoras Transductoras de Señales/genética , Antivirales/farmacología , Sistemas CRISPR-Cas , Línea Celular , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genética , Estudio de Asociación del Genoma Completo , Humanos , Proteínas de la Membrana/genética , Metiltransferasas/metabolismo , Proteínas del Tejido Nervioso/genética , Oxazinas/farmacología , Piridinas/farmacología , Tiepinas/farmacología , Triazinas/farmacología , ATPasas de Translocación de Protón Vacuolares/metabolismo , Internalización del Virus
18.
Toxicol Lett ; 322: 20-31, 2020 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-31923465

RESUMEN

Particulate matter (PM) from combustion processes has been associated with oxidative stress to DNA, whereas effects related to telomere dysfunction are less investigated. We collected air-borne PM from a passenger cabin of a diesel-propelled train and at a training facility for smoke diving exercises. Effects on oxidative stress biomarkers, genotoxicity measured by the comet assay and telomere length in PM-exposed A549 cells were compared with the genotoxicity and telomere length in peripheral blood mononuclear cells (PBMCs) from human volunteers exposed to the same aerosol source. Although elevated levels of DNA strand breaks and oxidatively damaged DNA in terms of Fpg-sensitive sites were observed in PBMCs from exposed humans, the PM collected at same locations did not cause genotoxicity in the comet assay in A549 cells. Nevertheless, A549 cells displayed telomere length shortening after four weeks exposure to PM. This is in line with slightly shorter telomere length in PBMCs from exposed humans, although it was not statistically significant. In conclusion, the results indicate that genotoxic potency measured by the comet assay of PM in A549 cells may not predict genotoxicity in exposed humans, whereas telomere length measurements may be a novel indicator of genotoxic stress in cell cultures and humans.


Asunto(s)
Daño del ADN , Exposición por Inhalación/efectos adversos , Material Particulado/toxicidad , Humo/efectos adversos , Homeostasis del Telómero/efectos de los fármacos , Emisiones de Vehículos/toxicidad , Células A549 , Contaminantes Ocupacionales del Aire/toxicidad , Supervivencia Celular/efectos de los fármacos , Bomberos , Humanos , Exposición por Inhalación/análisis , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/patología , Exposición Profesional/efectos adversos , Exposición Profesional/análisis , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Tamaño de la Partícula , Homeostasis del Telómero/genética
19.
Gene ; 731: 144348, 2020 Mar 20.
Artículo en Inglés | MEDLINE | ID: mdl-31927006

RESUMEN

Mounting evidence demonstrates that N6-methyladenosine (m6A) play critical roles of m6A in the epigenetic regulation, especially for human cancer. The m6A modification is installed by methyltransferase and erased demethylases, leading to the significant modification for gene expression and cell fate. Here, we investigated the biological roles and mechanism of demethylase alkylation repair homolog protein 5 (ALKBH5) in the non-small cell lung cancer (NSCLC). Results revealed that ALKBH5 was ectopically up-regulated in the NSCLC tissue and cells, and closely correlated with the poor prognosis. Functionally, ALKBH5 promoted the proliferation and reduced apoptosis of NSCLC cells in vitro, and knockdown of ALKBH5 repressed the tumor growth in vivo. Mechanistically, RNA immunoprecipitation sequencing (RIP-Seq) revealed that ALKBH5 targeted the TIMP3. Moreover, ALKBH5 repressed TIMP3 mRNA stability and protein production. In conclusion, the present research confirmed the ALKBH5/TIMP3 pathway in the NSCLC oncogenesis progress, providing a novel insight for the epitranscriptome and potential therapeutic target for NSCLC.


Asunto(s)
Adenosina/análogos & derivados , Desmetilasa de ARN, Homólogo 5 de AlkB/metabolismo , Desmetilasa de ARN, Homólogo 5 de AlkB/fisiología , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/genética , Estabilidad del ARN/genética , Inhibidor Tisular de Metaloproteinasa-3/genética , Células A549 , Adenosina/metabolismo , Desmetilasa de ARN, Homólogo 5 de AlkB/genética , Animales , Carcinogénesis/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Progresión de la Enfermedad , Epigénesis Genética/fisiología , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , ARN Mensajero/metabolismo
20.
Chem Biodivers ; 17(1): e1900547, 2020 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-31916685

RESUMEN

Four previously unreported chromones, 5-hydroxy-2-(hydroxymethyl)-8-methoxy-4H-chromen-4-one (1), (5R,7S)-5,7-dihydroxy-2-propyl-5,6,7,8-tetrahydro-4H-chromen-4-one (2), (5R,7S)-5,7-dihydroxy-2-methyl-5,6,7,8-tetrahydro-4H-chromen-4-one (3), and (5R,7S)-5,7-dihydroxy-2-[(E)-prop-1-en-1-yl]-5,6,7,8-tetrahydro-4H-chromen-4-one (4), as well as one known analogue 5-hydroxy-2-methyl-4H-chromen-4-one (5) were isolated from the fermentation broth of the endophytic fungus Colletotrichum gloeosporioides derived from the mangrove Ceriops tagal. Their structures were elucidated based on extensive spectroscopic analyses. The absolute configurations of 2-4 were determined by comparison the experimental and calculated electronic circular dichroism (ECD) spectra. Compound 2 showed cytotoxic activity against A549 cell line with the IC50 value of 0.094 mm.


Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Cromonas/aislamiento & purificación , Cromonas/farmacología , Colletotrichum/química , Células A549 , Antineoplásicos/aislamiento & purificación , Proliferación Celular/efectos de los fármacos , Cromonas/química , Teoría Funcional de la Densidad , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Estructura Molecular , Relación Estructura-Actividad
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