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1.
Int J Mol Sci ; 22(6)2021 Mar 17.
Artículo en Inglés | MEDLINE | ID: mdl-33802660

RESUMEN

Proximal tubular (PT) acidosis, which alkalinizes the urinary filtrate, together with Ca2+ supersaturation in PT can induce luminal calcium phosphate (CaP) crystal formation. While such CaP crystals are known to act as a nidus for CaP/calcium oxalate (CaOx) mixed stone formation, the regulation of PT luminal Ca2+ concentration ([Ca2+]) under elevated pH and/or high [Ca2+] conditions are unknown. Since we found that transient receptor potential canonical 3 (TRPC3) knockout (KO; -/-) mice could produce mild hypercalciuria with CaP urine crystals, we alkalinized the tubular pH in TRPC3-/- mice by oral acetazolamide (0.08%) to develop mixed urinary crystals akin to clinical signs of calcium nephrolithiasis (CaNL). Our ratiometric (λ340/380) intracellular [Ca2+] measurements reveal that such alkalization not only upsurges Ca2+ influx into PT cells, but the mode of Ca2+ entry switches from receptor-operated to store-operated pathway. Electrophysiological experiments show enhanced bicarbonate related current activity in treated PT cells which may determine the stone-forming phenotypes (CaP or CaP/CaOx). Moreover, such alkalization promotes reactive oxygen species generation, and upregulation of calcification, inflammation, fibrosis, and apoptosis in PT cells, which were exacerbated in absence of TRPC3. Altogether, the pH-induced alteration of the Ca2+ signaling signature in PT cells from TRPC3 ablated mice exacerbated the pathophysiology of mixed urinary stone formation, which may aid in uncovering the downstream mechanism of CaNL.


Asunto(s)
Acetazolamida/farmacología , Calcio/metabolismo , Túbulos Renales Proximales/metabolismo , Túbulos Renales Proximales/patología , Nefrolitiasis/metabolismo , Nefrolitiasis/patología , Animales , Transporte Biológico/efectos de los fármacos , Calcinosis/complicaciones , Estrés del Retículo Endoplásmico/efectos de los fármacos , Fibrosis , Concentración de Iones de Hidrógeno , Inflamación/patología , Túbulos Renales Proximales/efectos de los fármacos , Ratones , Nefrolitiasis/orina , Estrés Oxidativo/efectos de los fármacos , Canales Catiónicos TRPC/metabolismo , Regulación hacia Arriba/efectos de los fármacos
2.
Int J Mol Sci ; 22(6)2021 Mar 17.
Artículo en Inglés | MEDLINE | ID: mdl-33802742

RESUMEN

New anti-inflammatory treatments are needed for CF airway disease. Studies have implicated the endoplasmic reticulum stress transducer inositol requiring enzyme 1α (IRE1α) in CF airway inflammation. The activation of IRE1α promotes activation of its cytoplasmic kinase and RNase, resulting in mRNA splicing of X-box binding protein-1 (XBP-1s), a transcription factor required for cytokine production. We tested whether IRE1α kinase and RNase inhibition decreases cytokine production induced by the exposure of primary cultures of homozygous F508del CF human bronchial epithelia (HBE) to supernatant of mucopurulent material (SMM) from CF airways. We evaluated whether IRE1α expression is increased in freshly isolated and native CF HBE, and couples with increased XBP-1s levels. A FRET assay confirmed binding of the IRE1α kinase and RNase inhibitor, KIRA6, to the IRE1α kinase. F508del HBE cultures were exposed to SMM with or without KIRA6, and we evaluated the mRNA levels of XBP-1s, IL-6, and IL-8, and the secretion of IL-6 and IL-8. IRE1α mRNA levels were up-regulated in freshly isolated CF vs. normal HBE and coupled to increased XBP-1s mRNA levels. SMM increased XBP-1s, IL-6, and IL-8 mRNA levels and up-regulated IL-6 and IL-8 secretion, and KIRA6 blunted these responses in a dose-dependent manner. Moreover, a triple combination of CFTR modulators currently used in the clinic had no effect on SMM-increased XBP-1s levels coupled with increased cytokine production in presence or absence of KIRA6. These findings indicate that IRE1α mediates cytokine production in CF airways. Small molecule IRE1α kinase inhibitors that allosterically reduce RNase-dependent XBP-1s may represent a new therapeutic strategy for CF airway inflammation.


Asunto(s)
Fibrosis Quística/tratamiento farmacológico , Fibrosis Quística/patología , Endorribonucleasas/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/patología , Pulmón/patología , Terapia Molecular Dirigida , Proteínas Serina-Treonina Quinasas/metabolismo , Células Cultivadas , Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Citocinas/biosíntesis , Endorribonucleasas/genética , Epitelio/efectos de los fármacos , Epitelio/patología , Humanos , Imidazoles/química , Imidazoles/farmacología , Inflamación/genética , Modelos Biológicos , Naftalenos/química , Naftalenos/farmacología , Proteínas Serina-Treonina Quinasas/genética , Pirazinas/química , Pirazinas/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética , Proteína 1 de Unión a la X-Box/metabolismo
3.
Int J Nanomedicine ; 16: 2461-2475, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33814910

RESUMEN

Aim: To explore the effects of Radix Sophorae Flavescentis carbonisata-based carbon dots (RSFC-CDs) on an ethanol-induced acute gastric ulcer rat model. Methods: The structure, optical properties, functional groups and elemental composition of RSFC-CDs synthesized by one-step pyrolysis were characterized. The gastric protective effects of RSFC-CDs were evaluated and confirmed by applying a rat model of ethanol-induced acute gastric ulcers. The underlying mechanisms were investigated through the nuclear factor-kappa B (NF-κB) signalling pathway and oxidative stress. Results: RSFC-CDs with a diameter ranging from 2-3 nm mainly showed gastric protective effects by reducing the levels of NF-κB, tumour necrosis factor-α (TNF-α), interleukin (IL)-6, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione (GSH), malondialdehyde (MDA) and inducible nitric oxide synthase (iNOS) to inhibit ethanol-induced inflammation and oxidative stress. Conclusion: RSFC-CDs have anti-inflammatory and anti-oxidative effects, making them promising for application in ethanol-induced gastric injury.


Asunto(s)
Antiinflamatorios/uso terapéutico , Antioxidantes/uso terapéutico , Carbono/química , Medicamentos Herbarios Chinos/uso terapéutico , Sustancias Protectoras/uso terapéutico , Úlcera Gástrica/inducido químicamente , Úlcera Gástrica/tratamiento farmacológico , Enfermedad Aguda , Animales , Antiinflamatorios/farmacología , Antioxidantes/farmacología , Regulación hacia Abajo/efectos de los fármacos , Medicamentos Herbarios Chinos/farmacología , Etanol/efectos adversos , Interleucina-6/metabolismo , Masculino , FN-kappa B/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Estrés Oxidativo/efectos de los fármacos , Espectroscopía de Fotoelectrones , Sustancias Protectoras/farmacología , Ratas Sprague-Dawley , Úlcera Gástrica/patología , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia Arriba/efectos de los fármacos
4.
Nat Commun ; 12(1): 2136, 2021 04 09.
Artículo en Inglés | MEDLINE | ID: mdl-33837198

RESUMEN

Osteoclastic bone resorption and osteoblastic bone formation/replenishment are closely coupled in bone metabolism. Anabolic parathyroid hormone (PTH), which is commonly used for treating osteoporosis, shifts the balance from osteoclastic to osteoblastic, although it is unclear how these cells are coordinately regulated by PTH. Here, we identify a serine protease inhibitor, secretory leukocyte protease inhibitor (SLPI), as a critical mediator that is involved in the PTH-mediated shift to the osteoblastic phase. Slpi is highly upregulated in osteoblasts by PTH, while genetic ablation of Slpi severely impairs PTH-induced bone formation. Slpi induction in osteoblasts enhances its differentiation, and increases osteoblast-osteoclast contact, thereby suppressing osteoclastic function. Intravital bone imaging reveals that the PTH-mediated association between osteoblasts and osteoclasts is disrupted in the absence of SLPI. Collectively, these results demonstrate that SLPI regulates the communication between osteoblasts and osteoclasts to promote PTH-induced bone anabolism.


Asunto(s)
Resorción Ósea/tratamiento farmacológico , Osteogénesis/fisiología , Hormona Paratiroidea/administración & dosificación , Inhibidor Secretorio de Peptidasas Leucocitarias/metabolismo , Animales , Resorción Ósea/patología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Línea Celular , Modelos Animales de Enfermedad , Femenino , Fémur/citología , Fémur/diagnóstico por imagen , Fémur/efectos de los fármacos , Fémur/patología , Humanos , Masculino , Ratones , Ratones Noqueados , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Osteoclastos/efectos de los fármacos , Osteoclastos/metabolismo , Osteogénesis/efectos de los fármacos , Cultivo Primario de Células , RNA-Seq , Inhibidor Secretorio de Peptidasas Leucocitarias/genética , Regulación hacia Arriba/efectos de los fármacos , Microtomografía por Rayos X
5.
Nat Commun ; 12(1): 2163, 2021 04 12.
Artículo en Inglés | MEDLINE | ID: mdl-33846331

RESUMEN

γδ T cells are a distinct subgroup of T cells that bridge the innate and adaptive immune system and can attack cancer cells in an MHC-unrestricted manner. Trials of adoptive γδ T cell transfer in solid tumors have had limited success. Here, we show that DNA methyltransferase inhibitors (DNMTis) upregulate surface molecules on cancer cells related to γδ T cell activation using quantitative surface proteomics. DNMTi treatment of human lung cancer potentiates tumor lysis by ex vivo-expanded Vδ1-enriched γδ T cells. Mechanistically, DNMTi enhances immune synapse formation and mediates cytoskeletal reorganization via coordinated alterations of DNA methylation and chromatin accessibility. Genetic depletion of adhesion molecules or pharmacological inhibition of actin polymerization abolishes the potentiating effect of DNMTi. Clinically, the DNMTi-associated cytoskeleton signature stratifies lung cancer patients prognostically. These results support a combinatorial strategy of DNMTis and γδ T cell-based immunotherapy in lung cancer management.


Asunto(s)
Citoesqueleto/metabolismo , Citotoxicidad Inmunológica/genética , Epigénesis Genética , Sinapsis Inmunológicas/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Citoesqueleto de Actina/efectos de los fármacos , Citoesqueleto de Actina/metabolismo , Animales , Línea Celular Tumoral , Citoesqueleto/efectos de los fármacos , Citotoxicidad Inmunológica/efectos de los fármacos , ADN (Citosina-5-)-Metiltransferasas/antagonistas & inhibidores , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Decitabina/farmacología , Inhibidores Enzimáticos/farmacología , Epigénesis Genética/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Sinapsis Inmunológicas/efectos de los fármacos , Marcaje Isotópico , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/genética , Subgrupos Linfocitarios/efectos de los fármacos , Subgrupos Linfocitarios/metabolismo , Masculino , Ratones Endogámicos NOD , Fosfotirosina/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Análisis de Supervivencia , Proteína p53 Supresora de Tumor/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
6.
Nat Commun ; 12(1): 2183, 2021 04 12.
Artículo en Inglés | MEDLINE | ID: mdl-33846348

RESUMEN

Here we show that FTO as an N6-methyladenosine (m6A) RNA demethylase is degraded by selective autophagy, which is impaired by low-level arsenic exposure to promote tumorigenesis. We found that in arsenic-associated human skin lesions, FTO is upregulated, while m6A RNA methylation is downregulated. In keratinocytes, chronic relevant low-level arsenic exposure upregulated FTO, downregulated m6A RNA methylation, and induced malignant transformation and tumorigenesis. FTO deletion inhibited arsenic-induced tumorigenesis. Moreover, in mice, epidermis-specific FTO deletion prevented skin tumorigenesis induced by arsenic and UVB irradiation. Targeting FTO genetically or pharmacologically inhibits the tumorigenicity of arsenic-transformed tumor cells. We identified NEDD4L as the m6A-modified gene target of FTO. Finally, arsenic stabilizes FTO protein through inhibiting p62-mediated selective autophagy. FTO upregulation can in turn inhibit autophagy, leading to a positive feedback loop to maintain FTO accumulation. Our study reveals FTO-mediated dysregulation of mRNA m6A methylation as an epitranscriptomic mechanism to promote arsenic tumorigenicity.


Asunto(s)
Adenosina/análogos & derivados , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/metabolismo , Arsénico/toxicidad , Autofagia , Carcinogénesis/genética , Adenosina/metabolismo , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/genética , Animales , Autofagia/efectos de los fármacos , Autofagia/genética , Secuencia de Bases , Carcinogénesis/efectos de los fármacos , Transformación Celular Neoplásica/efectos de los fármacos , Transformación Celular Neoplásica/patología , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/genética , Epidermis/metabolismo , Ontología de Genes , Células HEK293 , Humanos , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Lisosomas/efectos de los fármacos , Lisosomas/metabolismo , Ratones , FN-kappa B/metabolismo , Ubiquitina-Proteína Ligasas Nedd4/metabolismo , Estabilidad Proteica/efectos de los fármacos , Estabilidad del ARN/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteína Sequestosoma-1/metabolismo , Transcriptoma/genética , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética , Vacuolas/efectos de los fármacos , Vacuolas/metabolismo , Vacuolas/ultraestructura
7.
Eur J Med Chem ; 216: 113250, 2021 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-33691258

RESUMEN

Inflammatory bowel disease (IBD) describes a set of disorders involving alterations to gastrointestinal physiology and mucosal immunity. Unravelling its complex pathophysiology is important since many IBD patients are refractory to or suffer adverse side effects from current treatments. Isothiocyanates (ITCs), such as 6-(methylsulfinyl)hexyl ITC (6-MITC) in Wasabia japonica, have potential anti-inflammatory activity. We aimed to elucidate the pathways through which 6-MITC alleviates inflammation by examining its role in the nuclear factor-kappa B (NF-κB) pathway through inhibition of glycogen synthase kinase 3 beta (GSK-3ß) using a chemically induced murine model of IBD, cell-based and in silico techniques. The effects of 6-MITC and two NF-κB inhibitors, sulfasalazine (SS), pyrrolidine dithiolcarbamate (PDTC) were investigated on a dextran sulfate sodium (DSS)-induced murine mouse model of acute and chronic colitis using macroscopic measurements and pro-inflammatory markers. The effect of 6-MITC on NF-κB induction was assessed using a murine macrophage cell line. Complexes of GSK-3ß-6-MITC and GSK-3ß-ATP were generated in silico to elucidate the mechanism of 6-MITC's direct inhibition of GSK-3ß. Changes in pro-inflammatory markers, inducible nitric oxide synthase (iNOS) (increased) and interleukin-6 (IL-6) (decreased) demonstrated that iNOS regulation occurred at the translational level. Intraperitoneal (ip) injection of 6-MITC to the colitis-induced mice ameliorated weight loss whereas oral administration had negligible effect. Fecal blood and colon weight/length ratio parameters improved on treatment with 6-MITC and the other NF-κB inhibitors. Levels of NF-κB decreased upon addition of 6-MITC in vitro while structural studies showed 6-MITC acts competitively to inhibit GSK-3ß at the ATP binding site. In this study we demonstrated that 6-MITC inhibits NF-κB signaling via GSK-3ß inhibition ameliorating fecal blood, colonic alterations and DSS-induced weight loss indirectly indicating reduced intestinal stress. Taken together these results suggest a role for 6-MITC in the treatment of IBD acting to alleviate inflammation through the GSK-3ß/NF-κB pathway. Furthermore, the GSK-3ß-6-MITC model can be utilized as a basis for development of novel therapeutics targeting GSK-3ß for use in other disorders including cancer.


Asunto(s)
Antiinflamatorios/química , Glucógeno Sintasa Quinasa 3 beta/antagonistas & inhibidores , Isotiocianatos/química , Wasabia/química , Animales , Antiinflamatorios/metabolismo , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Línea Celular , Sulfato de Dextran/toxicidad , Regulación hacia Abajo/efectos de los fármacos , Femenino , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/inducido químicamente , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/patología , Interleucina-6/metabolismo , Isotiocianatos/metabolismo , Isotiocianatos/farmacología , Isotiocianatos/uso terapéutico , Lipopolisacáridos/farmacología , Macrófagos/citología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Ratones Endogámicos BALB C , FN-kappa B/antagonistas & inhibidores , FN-kappa B/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Transducción de Señal/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Wasabia/metabolismo
8.
Chem Biol Interact ; 340: 109450, 2021 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-33775688

RESUMEN

The emergence of multidrug resistance (MDR) is among the crucial obstacles to breast cancer therapy success. The transcription factor nuclear factor (NF)-κB is correlated to the pathogenesis of breast cancer and resistance to therapy. NF-κB augments the expression of MDR1 gene, which encodes for the membrane transporter P-glycoprotein (P-gp) in cancer cells. Since NF-κB activity is considered to be relatively high in particular when it comes to breast cancer, in the present work, we proposed that the inhibition of NF-κB activity can augment and enhance the sensitivity of breast cancer cells to chemotherapy such as doxorubicin (DOX) by virtue of MDR modulation. Our results demonstrated that the DOX-resistant MCF-7 and MDA-MB-231 clones exhibit higher NF-κB (p65) activity, which is linked to the upregulated expression of ABCB1 and ABCC1 transporter proteins. Combined treatment with NF-kB inhibitors (pentoxifylline and bortezomib) sensitized the resistant breast cancer cells to DOX. Such synergy was compromised by forced overexpression of p65. The DOX/NF-κB inhibitor combinations hampered NF-κB (p65) activation and downregulated MDR efflux transporters' level. Breast cancer cell migration was sharply suppressed in cells co-treated with DOX/NF-κB inhibitors. The same treatments successfully enhanced DOX-mediated induction of apoptosis, which is reflected by the elevated ratio of annexin-V/PI positively stained cells, along with the activation of other apoptotic markers. In conclusion, the data generated from this study provide insights for future translational investigations introducing the use of the clinically approved NF-κB inhibitors as an adjuvant in the treatment protocols of resistant breast cancer to overcome the multidrug resistance and enhance the therapeutic outcomes.


Asunto(s)
Antineoplásicos/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/metabolismo , Resistencia a Múltiples Medicamentos/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , FN-kappa B/metabolismo , Transducción de Señal/efectos de los fármacos , Apoptosis/efectos de los fármacos , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Doxorrubicina/farmacología , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Células MCF-7 , Regulación hacia Arriba/efectos de los fármacos
9.
Cells ; 10(3)2021 02 27.
Artículo en Inglés | MEDLINE | ID: mdl-33673459

RESUMEN

The article describes the rationale for the administration of zinc-chelating agents in COVID-19 patients. In a previous work I have highlighted that the binding of the SARS-CoV spike proteins to the zinc-metalloprotease ACE2 has been shown to induce ACE2 shedding by activating the zinc-metalloprotease ADAM17, which ultimately leads to systemic upregulation of ACE2 activity. Moreover, based on experimental models, it was also shown the detrimental effect of the excessive systemic activity of ACE2 through its downstream pathways, which leads to "clinical" manifestations resembling COVID-19. In this regard, strong upregulation of circulating ACE2 activity was recently reported in COVID-19 patients, thus supporting the previous hypothesis that COVID-19 may derive from upregulation of ACE2 activity. Based on this, a reasonable hypothesis of using inhibitors that curb the upregulation of both ACE2 and ADAM17 zinc-metalloprotease activities and consequent positive feedback-loops (initially triggered by SARS-CoV-2 and subsequently sustained independently on viral trigger) is proposed as therapy for COVID-19. In particular, zinc-chelating agents such as citrate and ethylenediaminetetraacetic acid (EDTA) alone or in combination are expected to act in protecting from COVID-19 at different levels thanks to their both anticoagulant properties and inhibitory activity on zinc-metalloproteases. Several arguments are presented in support of this hypothesis and based on the current knowledge of both beneficial/harmful effects and cost/effectiveness, the use of chelating agents in the prevention and therapy of COVID-19 is proposed. In this regard, clinical trials (currently absent) employing citrate/EDTA in COVID-19 are urgently needed in order to shed more light on the efficacy of zinc chelators against SARS-CoV-2 infection in vivo.


Asunto(s)
/tratamiento farmacológico , Quelantes/farmacología , Ácido Cítrico/farmacología , Ácido Edético/farmacología , Sistema Renina-Angiotensina/efectos de los fármacos , Zinc/metabolismo , Proteína ADAM17/metabolismo , Anticoagulantes/farmacología , /terapia , Descubrimiento de Drogas , Humanos , Inmunización Pasiva/efectos adversos , Regulación hacia Arriba/efectos de los fármacos
10.
Chem Biol Interact ; 339: 109430, 2021 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-33676887

RESUMEN

Connexin-40 (Cx40) and Cx43 are the principal components of gap junctions. Dysregulation of connexin expression is clinically related to cardiac pathologies. 25-Hydroxy protopanaxadiol [25-OH-PPD, 20 (R)-dammarane-3ß, 12ß, 20, 25-tetrol], known as AD2, is a novel protopanaxadiol extracted from Panax ginseng that exhibits many pharmacological activities, but its effects on cardiac gap junctions are poorly understood. The aim of this study was to evaluate the effects of AD2 on angiotensin II (Ang II)-induced Cx40 and Cx43 dysregulation. In this study, isolated beating rat atria were perfused with Ang II (5 µM) for 1 h to induce Cx40 and Cx43 dysregulation. The effects of AD2 (1.6, 16, and 160 µg/100 g body weight) on Ang II-induced hemodynamics in rats were analyzed by biological recorder, and changes in proteins levels were analyzed by western blotting. The results showed that AD2 ameliorated Ang II-induced hyper hemodynamics and abnormal P-waves, and prevented fibrotic collagen deposition (3.77% ± 1.64%-26.31% ± 1.64% with Ang II, 5.76% ± 0.94% with AD2). Ang II upregulated expression of nuclear factor kappa B, activator protein 1, and transforming growth factor ß1, and downregulated of Cx40 and Cx43 expression, which were inhibited by AD2 concomitantly with increased of AMP-activated protein kinase (AMPK) expression via liver kinase B1 activation. The present findings suggest that AD2 inhibited Ang II-induced dysregulation of Cx40 and Cx43 via activation of AMPK signaling, thus highlighting the promise and utility of AD2 for treatment of connexin dysregulation-related heart disease.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Angiotensina II/farmacología , Conexina 43/metabolismo , Conexinas/metabolismo , Ginsenósidos/farmacología , Transducción de Señal/efectos de los fármacos , Animales , Regulación hacia Abajo/efectos de los fármacos , Fibrosis/metabolismo , Uniones Comunicantes/efectos de los fármacos , Uniones Comunicantes/metabolismo , Ginsenósidos/metabolismo , FN-kappa B/metabolismo , Ratas , Ratas Sprague-Dawley , Factor de Transcripción AP-1/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Regulación hacia Arriba/efectos de los fármacos
11.
Molecules ; 26(4)2021 Feb 19.
Artículo en Inglés | MEDLINE | ID: mdl-33669666

RESUMEN

Preliminary bioassay-guided fractionation was performed to identify cytotoxic compounds from Hechtia glomerata, a plant that is used in Mexican ethnomedicine. Organic and aqueous extracts were prepared from H. glomerata's leaves and evaluated against two cancer cell lines. The CHCl3/MeOH (1:1) active extract was fractionated, and the resulting fractions were assayed against prostate adenocarcinoma PC3 and breast adenocarcinoma MCF7 cell lines. Active fraction 4 was further analyzed by high-performance liquid chromatography-quadrupole time-of-flight-mass spectrometry analysis to identify its active constituents. Among the compounds that were responsible for the cytotoxic effects of this fraction were flavonoids, phenolic acids, and aromatic compounds, of which p-coumaric acid (p-CA) and its derivatives were abundant. To understand the mechanisms that underlie p-CA cytotoxicity, a microarray assay was performed on PC3 cells that were treated or not with this compound. The results showed that mitogen-activated protein kinases (MAPKs) that regulate many cancer-related pathways were targeted by p-CA, which could be related to the reported effects of reactive oxygen species (ROS). A molecular docking study of p-CA showed that this phenolic acid targeted these protein active sites (MAPK8 and Serine/Threonine protein kinase 3) at the same binding site as their inhibitors. Thus, we hypothesize that p-CA produces ROS, directly affects the MAPK signaling pathway, and consequently causes apoptosis, among other effects. Additionally, p-CA could be used as a platform for the design of new MAPK inhibitors and re-sensitizing agents for resistant cancers.


Asunto(s)
Bromeliaceae/química , Ácidos Cumáricos/farmacología , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Extractos Vegetales/química , Inhibidores de Proteínas Quinasas/farmacología , Bioensayo , Muerte Celular/efectos de los fármacos , Cromatografía Líquida de Alta Presión , Ácidos Cumáricos/química , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/genética , Humanos , Células MCF-7 , Proteínas Quinasas Activadas por Mitógenos/química , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Células PC-3 , Fenoles/farmacología , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética
12.
Molecules ; 26(4)2021 Feb 19.
Artículo en Inglés | MEDLINE | ID: mdl-33669807

RESUMEN

Dental papilla cells (DPCs), precursors of odontoblasts, are considered promising seed cells for tissue engineering. Emerging evidence suggests that melatonin promotes odontoblastic differentiation of DPCs and affects tooth development, although the precise mechanisms remain unknown. Retinoid acid receptor-related orphan receptor α (RORα) is a nuclear receptor for melatonin that plays a critical role in cell differentiation and embryonic development. This study aimed to explore the role of RORα in odontoblastic differentiation and determine whether melatonin exerts its pro-odontogenic effect via RORα. Herein, we observed that RORα was expressed in DPCs and was significantly increased during odontoblastic differentiation in vitro and in vivo. The overexpression of RORα upregulated the expression of odontogenic markers, alkaline phosphatase (ALP) activity and mineralized nodules formation (p < 0.05). In contrast, odontoblastic differentiation of DPCs was suppressed by RORα knockdown. Moreover, we found that melatonin elevated the expression of odontogenic markers, which was accompanied by the upregulation of RORα (p < 0.001). Utilising small interfering RNA, we further demonstrated that RORα inhibition attenuated melatonin-induced odontogenic gene expression, ALP activity and matrix mineralisation (p < 0.01). Collectively, these results provide the first evidence that RORα can promote odontoblastic differentiation of DPCs and mediate the pro-odontogenic effect of melatonin.


Asunto(s)
Diferenciación Celular , Papila Dental/citología , Melatonina/farmacología , Miembro 1 del Grupo F de la Subfamilia 1 de Receptores Nucleares/metabolismo , Odontoblastos/citología , Odontoblastos/metabolismo , Odontogénesis , Animales , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Odontoblastos/efectos de los fármacos , Odontogénesis/efectos de los fármacos , Ratas Sprague-Dawley , Regulación hacia Arriba/efectos de los fármacos
13.
Int J Mol Sci ; 22(4)2021 Feb 19.
Artículo en Inglés | MEDLINE | ID: mdl-33669634

RESUMEN

Little is known about the effects on hyaluronan (HA) metabolism of UVA radiation. This study demonstrates that the secretion of HA by human dermal fibroblasts (HDFs) is downregulated by UVA, accompanied by the down- and upregulation of mRNA and protein levels of the HA-synthesizing enzyme (HAS2) and the HA-degrading protein, HYaluronan Binding protein Involved in HA Depolymerization(HYBID), respectively. Signaling analysis revealed that the exposure distinctly elicits activation of the p38/MSK1/CREB/c-Fos/AP-1 axis, the JNK/c-Jun axis, and the p38/ATF-2 axis, but downregulates the phosphorylation of NF-kB and JAK/STAT3. A signal inhibition study demonstrated that the inhibition of p38 significantly abrogates the UVA-accentuated mRNA level of HYBID. Furthermore, the inhibition of STAT3 significantly downregulates the level of HAS2 mRNA in non-UVA exposed HDFs. Analysis using siRNAs demonstrated that transfection of ATF-2 siRNA but not c-Fos siRNA abrogates the increased protein level of HYBID in UVA-exposed HDFs. An inhibitor of protein tyrosine phosphatase but not of protein serine/threonine phosphatase restored the diminished phosphorylation level of STAT3 at Tyr 705, accompanied by a significant abolishing effect on the decreased mRNA expression level of HAS2. Silencing with a protein tyrosine phosphatase PTP-Meg2 siRNA revealed that it abrogates the decreased phosphorylation of STAT3 at Tyr 705 in UVA-exposed HDFs. These findings suggest that the UVA-induced decrease in HA secretion by HDFs is attributable to the down- and upregulation of HAS2 and HYBID expression, respectively, changes that are mainly ascribed to the inactivated signaling of the STAT3 axis due to the activated tyrosine protein phosphatase PTP-Meg2 and the activated signaling of the p38/ATF2 axis, respectively.


Asunto(s)
Regulación hacia Abajo/efectos de la radiación , Fibroblastos/efectos de la radiación , Hialuronano Sintasas/metabolismo , Ácido Hialurónico/metabolismo , Hialuronoglucosaminidasa/metabolismo , Transducción de Señal/efectos de la radiación , Rayos Ultravioleta , Regulación hacia Arriba/efectos de la radiación , Factor de Transcripción Activador 2/metabolismo , Muerte Celular/efectos de los fármacos , Muerte Celular/efectos de la radiación , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Dermis/citología , Regulación hacia Abajo/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Janus Quinasa 2/metabolismo , Masculino , Modelos Biológicos , Peso Molecular , Fosforilación/efectos de los fármacos , Fosforilación/efectos de la radiación , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Transcripción STAT3/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo
14.
Int J Mol Sci ; 22(3)2021 Jan 29.
Artículo en Inglés | MEDLINE | ID: mdl-33572938

RESUMEN

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is currently a global public health emergency. Periodontitis, the most prevalent disease that leads to tooth loss, is caused by infection by periodontopathic bacteria. Periodontitis is also a risk factor for pneumonia and the exacerbation of chronic obstructive pulmonary disease, presumably because of the aspiration of saliva contaminated with periodontopathic bacteria into the lower respiratory tract. Patients with these diseases have increased rates of COVID-19 aggravation and mortality. Because periodontopathic bacteria have been isolated from the bronchoalveolar lavage fluid of patients with COVID-19, periodontitis may be a risk factor for COVID-19 aggravation. However, the molecular links between periodontitis and COVID-19 have not been clarified. In this study, we found that the culture supernatant of the periodontopathic bacterium Fusobacterium nucleatum (CSF) upregulated the SARS-CoV-2 receptor angiotensin-converting enzyme 2 in A549 alveolar epithelial cells. In addition, CSF induced interleukin (IL)-6 and IL-8 production by both A549 and primary alveolar epithelial cells. CSF also strongly induced IL-6 and IL-8 expression by BEAS-2B bronchial epithelial cells and Detroit 562 pharyngeal epithelial cells. These results suggest that when patients with mild COVID-19 frequently aspirate periodontopathic bacteria, SARS-CoV-2 infection is promoted, and inflammation in the lower respiratory tract may become severe in the presence of viral pneumonia.


Asunto(s)
/metabolismo , Medios de Cultivo Condicionados/química , Citocinas/metabolismo , Fusobacterium nucleatum/metabolismo , /genética , /virología , Línea Celular , Medios de Cultivo Condicionados/farmacología , Células Epiteliales/citología , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Humanos , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Regulación hacia Arriba/efectos de los fármacos
15.
Arch Biochem Biophys ; 701: 108806, 2021 04 15.
Artículo en Inglés | MEDLINE | ID: mdl-33587903

RESUMEN

Clostridium perfringens (C. perfringens) is a globally recognized zoonotic pathogen. It has been reported that the beta2-toxin produced by C. perfringens can cause a variety of gastrointestinal diseases and even systemic inflammation. MicroRNA-124a (miR-124a) has been reported to play important roles in the host response to pathogenic infection. Although C. perfringens beta2-toxin induced injury in intestinal porcine epithelial (IPEC-J2) cells has been established, the underlying molecular mechanism is not completely unraveled. Here we show that a significant upregulation of ssc-miR-124a in IPEC-J2 cells after beta2-toxin stimulation was associated with the MiR-124A-1 and MiR-124A-2 gene promoter demethylation status. Importantly, overexpression of ssc-miR-124a significantly increased cell proliferation and decreased apoptosis and cytotoxicity in beta2-toxin treated IPEC-J2 cells. Transfection of IPEC-J2 cells with ssc-miR-124a mimic suppressed beta2-toxin induced inflammation. On the contrary, ssc-miR-124a inhibitor promoted aggravation of cell apoptosis and excessive damage. Furthermore, rho-associated coiled-coil-containing protein kinase 1 (ROCK1) was identified as the direct target gene of ssc-miR-124a in IPEC-J2 cells and its siRNA transfection reversed the promotion of apoptosis and aggravation of cellular damage induced by ssc-miR-124a inhibitor. Overall, we speculated that the miR-124A-1/2 gene was epigenetically regulated in IPEC-J2 cells after beta2-toxin treatment. Upregulation of ssc-miR-124a may restrain ROCK1, and attenuate apoptosis and inflammation induced by beta2-toxin that prevent IPEC-J2 cells from severe damages. We discover a new molecular mechanism by which IPEC-J2 cells counteract beta2-toxin-induced damage through the ssc-miR-124a/ROCK1 axis partially.


Asunto(s)
Apoptosis/efectos de los fármacos , Toxinas Bacterianas/toxicidad , Epigénesis Genética/efectos de los fármacos , Células Epiteliales/metabolismo , Mucosa Intestinal/metabolismo , MicroARNs/biosíntesis , Regulación hacia Arriba/efectos de los fármacos , Animales , Línea Celular , Clostridium perfringens , Células Epiteliales/patología , Inflamación/inducido químicamente , Inflamación/metabolismo , Inflamación/patología , Mucosa Intestinal/patología , MicroARNs/genética , Porcinos
16.
J Med Food ; 24(2): 188-196, 2021 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-33617363

RESUMEN

Isoamylamine (IA) is an aliphatic monoamine molecule present in cheese, eggs, and wine. It belongs to the family of polyamines and also can be synthesized endogenously. It has been known that regulation of polyamines in cells is related to cell cycle and tumor formation. Malignant melanoma is difficult to treat and easily resistant to chemotherapy/radiotherapy through autophagy. In this study, we aim to clarify whether IA has a growth control effect on melanoma tumor cells and the regulatory mechanism. We treated B16-F1 melanoma cells with IA at concentrations of 0, 200, 400, and 600 ppm for 24 h. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was checked for cell viability and results showed that IA has an inhibitory effect on B16-F1 melanoma cells. The signaling molecules, which included Raf/MEK/ERK, were activated, while MSK1 and protein kinase B (AKT) were suppressed. Autophagy was also confirmed to be induced by IA. The acridine orange stain-positive cells were increased and BECN-1/LC3 upregulated. The data also showed that the autophagy regulatory molecule, 5'-adenosine monophosphate-activated protein kinase (AMPK), was induced after IA treatment, so we used dorsomorphin to inhibit AMPK and found that it could suppress autophagy. In conclusion, IA has an effect of inducing autophagy in B16-F1 cells and it is regulated through AMPK.


Asunto(s)
Proteínas Quinasas Activadas por AMP , Aminas , Autofagia , Regulación hacia Arriba , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Aminas/farmacología , Animales , Antineoplásicos/farmacología , Autofagia/efectos de los fármacos , Línea Celular Tumoral , Ratones , Transducción de Señal/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos
17.
Chem Biol Interact ; 339: 109424, 2021 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-33617803

RESUMEN

OBJECTIVE: To reveal the effects and related mechanism of cisatracurium on colorectal cancer (CRC) development. METHODS: HCT116 and SW480 cells were treated with various concentrations of cisatracurium or transforming growth factor-ß (TGF-ß). Chemokine C-X-C-Motif Receptor 4 (CXCR4) was overexpressed and let-7a-5p was silenced in cells by transfection with pcDNA3.1-CXCR4 or let-7a-5p inhibitor. Cell Counting Kit-8 (CCK-8) assay measured cell viability, and transwell and wound healing assays evaluated cell invasion and migration, respectively. The expression levels of let-7a-5p and CXCR4 were measured using quantitative reverse transcription polymerase chain reaction (qRT-PCR). Western blotting was conducted to test the levels of CXCR4, TGF-ß/SMAD2/3 signalling and metastasis-related proteins. A tumour xenograft assay was performed to assess tumour growth. RESULTS: Cisatracurium treatment suppressed the viability and metastasis of HCT116 and SW480 cells in a concentration-dependent manner, whereas activating TGF-ß/SMAD2/3 signalling significantly reversed these effects. Cisatracurium treatment markedly reduced CXCR4 expression by inhibiting TGF-ß/SMAD2/3 signalling. Besides, let-7a-5p was identified as a target of CXCR4 and could be upregulated by cisatracurium. Both CXCR4 overexpression and let-7a-5p knockdown alleviated the biological roles of cisatracurium in CRC cells. Moreover, a tumour xenograft assay further confirmed that cisatracurium inhibited tumour growth and metastasis by increasing let-7a-5p expression. CONCLUSION: Cisatracurium suppressed the viability, metastasis and tumour growth of CRC by regulating the CXCR4/let-7a-5p axis via inhibiting TGF-ß/SMAD2/3 signalling. These findings provide a theoretical basis for the role of cisatracurium in the prognosis of CRC patients.


Asunto(s)
Atracurio/análogos & derivados , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , MicroARNs/genética , Receptores CXCR4/genética , Transducción de Señal/efectos de los fármacos , Animales , Atracurio/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Células HCT116 , Xenoinjertos/efectos de los fármacos , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Transducción de Señal/genética , Proteína Smad2/genética , Proteína smad3/genética , Factor de Crecimiento Transformador beta/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética , Ensayos Antitumor por Modelo de Xenoinjerto/métodos
18.
Nutrients ; 13(2)2021 Feb 03.
Artículo en Inglés | MEDLINE | ID: mdl-33546195

RESUMEN

Peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) is expressed in skeletal muscles and regulates systemic metabolism. Thus, nutraceuticals targeting skeletal muscle PGC-1α have attracted attention to modulate systemic metabolism. As auraptene contained in citrus fruits promotes lipid metabolism and improves mitochondrial respiration, it could increase mitochondrial function through PGC-1α. Therefore, we hypothesized that PGC-1α is activated by auraptene and investigated its effect using Citrus hassaku extract powder (CHEP) containing >80% of auraptene. C2C12 myotubes were incubated with vehicle or CHEP for 24 h; C57BL/6J mice were fed a control diet or a 0.25% (w/w) CHEP-containing diet for 5 weeks. PGC-1α protein level and mitochondrial content increased following CHEP treatment in cultured myotubes and skeletal muscles. In addition, the number of oxidative fibers increased in CHEP-fed mice. These findings suggest that CHEP-mediated PGC-1α upregulation induced mitochondrial biogenesis and fiber transformation to oxidative fibers. Furthermore, as CHEP increased the expression of the protein sirtuin 3 and of phosphorylated AMP-activated protein kinase (AMPK) and the transcriptional activity of PGC-1α, these molecules might be involved in CHEP-induced effects in skeletal muscles. Collectively, our findings indicate that CHEP mediates PGC-1α expression in skeletal muscles and may serve as a dietary supplement to prevent metabolic disorders.


Asunto(s)
Citrus/química , Mitocondrias Musculares/efectos de los fármacos , Fibras Musculares de Contracción Rápida/efectos de los fármacos , Músculo Esquelético/efectos de los fármacos , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/genética , Extractos Vegetales/farmacología , Animales , Línea Celular , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Mitocondrias Musculares/fisiología , Fibras Musculares de Contracción Rápida/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/ultraestructura , Mioblastos , Oxidación-Reducción , Polvos , Regulación hacia Arriba/efectos de los fármacos
20.
Leuk Res ; 101: 106511, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33517186

RESUMEN

Chronic myelomonocytic leukemia (CMML) is characterized by myelomonocytic bias and monocytic proliferation. Whether cell-intrinsic innate immune or inflammatory upregulation mediate disease pathogenesis and phenotype or whether the degree of aberrant monocytic differentiation influences outcomes remains unclear. We compared the transcriptomic features of bone marrow CD34+ cells from 19 patients with CMML and compared to healthy individuals. A total of 1495 genes had significantly differential expression in CMML (q<0.05, fold change>2), including 1271 genes that were significantly upregulated and 224 that were significantly downregulated in CMML. Top upregulated genes were associated with interferon (IFN) alpha and beta signaling, chemokine receptors, IFN gamma, G protein-coupled receptor ligand signaling, and genes involved in immunomodulatory interactions between lymphoid and non-lymphoid cells. Additionally, 6 gene sets were differentially upregulated and 139 were significantly downregulated in patients with myeloproliferative compared to myelodysplastic CMML. A total of 23 genes involved in regulation of monopoiesis were upregulated in CMML compared to healthy controls. We developed a prediction model using Cox regression including 3 of these genes, which differentiated patients into two prognostic subsets with distinct survival outcomes. This data warrants further evaluation of the roles and therapeutic potential of type I IFN signaling and monopoiesis in CMML.


Asunto(s)
Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Interferón Tipo I/administración & dosificación , Leucemia Mielomonocítica Crónica , Mielopoyesis/efectos de los fármacos , Proteínas de Neoplasias , Regulación hacia Arriba/efectos de los fármacos , Femenino , Humanos , Leucemia Mielomonocítica Crónica/tratamiento farmacológico , Leucemia Mielomonocítica Crónica/genética , Leucemia Mielomonocítica Crónica/metabolismo , Leucemia Mielomonocítica Crónica/patología , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/genética
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