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1.
Toxicol Lett ; 318: 1-11, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31618665

RESUMO

Triptolide (TP), a principal bioactive component extracted from traditional Chinese medicine Tripterygium wilfordii Hook. F. (TWHF), has attracted wide attention of its therapeutic effects on inflammation and autoimmune diseases. However, the therapeutic application of TP is hindered by severe cardiomyocyte toxicity and narrow therapeutic window. We previously identified that the p53 was an indispensable contributor in TP-induced myocardial injury. p53 has an inhibitory effect on IKKß-NF-κB pathway that regulates glucose transporters (GLUT) expression. Based on these evidences, we speculate that p53 mediates TP-disturbed glucose uptake by blocking IKKß-NF-κB signaling. This study focused on the effect of TP on cardiac glucose uptake and the role of p53 in glucose metabolism in cardiomyocytes, and p53 -/- mice. TP treatment depressed glucose consumption and ATP production resulting in myocardial damage. Incubation with ATP (5 mM) remarkably decreased the cellular damage. Immunoblotting and immunofluorescence identified that TP suppressed glucose uptake by restricting IKKß-NF-κB signaling activation, GLUT1 and GLUT4 expression. p53 inhibition alleviated the cell damage and the compromise of glucose uptake. Mechanistically, p53 antagonist PFTα abolished TP-induced the inhibition of IKKß, IκBα phosphorylation, p65 nuclear translocation, and GLUT1, GLUT4 expression. Consistently, in acute heart injury models, p53 deficiency upregulated IKKß-NF-κB activation and GLUT1, GLUT4 protein levels which was also indicated as amelioration of heart histological injury after 1.2 mg kg-1 TP administration. The present findings indicate that TP-induced p53 overactivation suppresses glucose uptake by inhibiting IKKß-NF-κB pathway and downregulating NF-κB-dependent GLUT1 and GLUT4 expression.


Assuntos
Diterpenos/toxicidade , Glucose/metabolismo , Cardiopatias/induzido quimicamente , Quinase I-kappa B/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , NF-kappa B/metabolismo , Fenantrenos/toxicidade , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/efeitos dos fármacos , Cardiotoxicidade , Linhagem Celular , Metabolismo Energético/efeitos dos fármacos , Compostos de Epóxi/toxicidade , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Transportador de Glucose Tipo 4/genética , Transportador de Glucose Tipo 4/metabolismo , Cardiopatias/genética , Cardiopatias/metabolismo , Cardiopatias/patologia , Camundongos Knockout , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genética
2.
Mol Cells ; 42(10): 702-710, 2019 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-31656063

RESUMO

Neuroinflammation is an important contributor to the pathogenesis of neurodegenerative disorders including Parkinson's disease (PD). We previously reported that our novel synthetic compound KMS99220 has a good pharmacokinetic profile, enters the brain, exerts neuroprotective effect, and inhibits NFκB activation. To further assess the utility of KMS99220 as a potential therapeutic agent for PD, we tested whether KMS99220 exerts an anti-inflammatory effect in vivo and examined the molecular mechanism mediating this phenomenon. In 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mice, oral administration of KMS99220 attenuated microglial activation and decreased the levels of inducible nitric oxide synthase and interleukin 1 beta (IL-1b) in the nigrostriatal system. In lipopolysaccharide (LPS)-challenged BV-2 microglial cells, KMS99220 suppressed the production and expression of IL-1b. In the activated microglia, KMS99220 reduced the phosphorylation of IκB kinase, c-Jun N-terminal kinase, and p38 MAP kinase; this effect was mediated by heme oxygenase-1 (HO-1), as both gene silencing and pharmacological inhibition of HO-1 abolished the effect of KMS99220. KMS99220 induced nuclear translocation of the transcription factor Nrf2 and expression of the Nrf2 target genes including HO-1. Together with our earlier findings, our current results show that KMS99220 may be a potential therapeutic agent for neuroinflammation-related neurodegenerative diseases such as PD.


Assuntos
Anti-Inflamatórios/farmacologia , Chalconas/farmacologia , Heme Oxigenase-1/metabolismo , Quinase I-kappa B/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Morfolinas/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/metabolismo , Modelos Biológicos , Transdução de Sinais/efeitos dos fármacos
3.
Nat Commun ; 10(1): 3974, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31481662

RESUMO

Tumor-associated macrophages (TAMs) usually express an M2 phenotype, which enables them to perform immunosuppressive and tumor-promoting functions. Reprogramming these TAMs toward an M1 phenotype could thwart their pro-cancer activities and unleash anti-tumor immunity, but efforts to accomplish this are nonspecific and elicit systemic inflammation. Here we describe a targeted nanocarrier that can deliver in vitro-transcribed mRNA encoding M1-polarizing transcription factors to reprogram TAMs without causing systemic toxicity. We demonstrate in models of ovarian cancer, melanoma, and glioblastoma that infusions of nanoparticles formulated with mRNAs encoding interferon regulatory factor 5 in combination with its activating kinase IKKß reverse the immunosuppressive, tumor-supporting state of TAMs and reprogram them to a phenotype that induces anti-tumor immunity and promotes tumor regression. We further establish that these nanoreagents are safe for repeated dosing. Implemented in the clinic, this immunotherapy could enable physicians to obviate suppressive tumors while avoiding systemic treatments that disrupt immune homeostasis.


Assuntos
Macrófagos/imunologia , Nanopartículas , Neoplasias/imunologia , RNA Mensageiro/administração & dosagem , Animais , Linhagem Celular Tumoral , Reprogramação Celular , Feminino , Glioblastoma/imunologia , Glioblastoma/patologia , Glioblastoma/terapia , Humanos , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Imunossupressão , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Ativação de Macrófagos , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Camundongos , Metástase Neoplásica , Neoplasias/genética , Neoplasias/patologia , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/terapia , RNA Mensageiro/genética , Linfócitos T/imunologia , Fatores de Transcrição/genética , Transfecção
4.
Artif Cells Nanomed Biotechnol ; 47(1): 3391-3398, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31394949

RESUMO

Nimbolide, a triterpenoid isolated from flower of neem tree possess various therapeutic properties. The objective of the study was to assess the anti-arthritic activity of nimbolide in arthritis induced rats. Nimbolide (20 mg/kg per day) was given orally to arthritic rats induced with Complete Freund's Adjuvant and changes in paw volume, body weight, organ indices (thymus and spleen), arthritic score, biochemical parameters and proinflammatory cytokines levels were determined. Histopathological analysis was also performed. Western blot analysis was also performed. Rats treated with nimbolide displayed marked reduction in arthritic score, organ indices, volume of paw, edema formation, along with substantial enhancement in body weight. Histopathological findings showed significant reduction in destruction of joints and inflammation following nimbolide treatment. The protective action of arthritic rats treated with nimbolide was also substantiated by molecular and biochemical studies. The results of the study show that nimbolide treatment has markedly enhanced health and reduced inflammation via lessening the proinflammatory cytokines expression in arthritic rats. Hence, nimbolide may be used as a potent therapeutic drug in treating rheumatoid arthritis.


Assuntos
Anti-Inflamatórios/farmacologia , Artrite Experimental/tratamento farmacológico , Adjuvante de Freund/efeitos adversos , Limoninas/farmacologia , Animais , Anti-Inflamatórios/uso terapêutico , Antioxidantes/metabolismo , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Peso Corporal/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Quinase I-kappa B/metabolismo , Articulações/efeitos dos fármacos , Articulações/metabolismo , Articulações/patologia , Limoninas/uso terapêutico , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Ratos
5.
Chem Biol Interact ; 311: 108798, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31433962

RESUMO

Natural products are a valuable source of anticancer agents, with many naturally derived compounds currently used in clinical and preclinical treatments. This study aims to investigate the antiproliferative activity and potential mechanism of action of the xanthoquinodin JBIR-99, isolated from fungi Parengyodontium album MEXU 30,054 and identified by single-crystal X-ray crystallography. Cytotoxicity of xanthoquinodin was evaluated in a panel of human cancer cells lines and CCD-112-CoN normal colon cells, using the sulforhodamine B assay. PC-3 prostate cancer cells were used in biochemical assays including cell cycle, mitochondrial transmembrane potential (MTP), reactive oxygen species (ROS) and caspase activity. Expression levels of apoptosis-pathway-related proteins were analyzed by Western blot. The in vivo toxicity of xanthoquinodin was determined using a zebrafish model. Xanthoquinodin showed cytotoxicity in all cancer cell lines but demonstrated relative selective potency against PC-3 cells with an IC50 1.7 µM. In CCD-112-CoN cells, xanthoquinodin was non-cytotoxic at 100 µM. In PC-3 cells, the compound induced loss of MTP, production of ROS, and cell cycle arrest in S phase. The expression and activity of caspase-3 was increased, which correlates with the upregulation of Cyt c, Bax, nuclear factor kappa-B (NF-κB) (p65) and IKKß, and downregulation of poly ADP ribose polymerase (PARP-1) and Bcl-2. Lastly, xanthoquinodin did not cause any visible developmental toxicity in zebrafish at 50 µM. These results demonstrate xanthoquinodin induces apoptosis in PC-3 prostate cancer cells by activation of both intrinsic and extrinsic apoptotic pathways. In addition, the non-toxic effect in vivo indicates that xanthoquinodin could be a useful lead in the development of a novel, anti-cancer agent that is selective for prostate cancer.


Assuntos
Apoptose/efeitos dos fármacos , Ascomicetos/química , Cromonas/farmacologia , Ascomicetos/metabolismo , Linhagem Celular Tumoral , Cromonas/química , Cristalografia por Raios X , Citocromos c/metabolismo , Humanos , Quinase I-kappa B/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Conformação Molecular , Poli(ADP-Ribose) Polimerase-1/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Repressoras/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
6.
Int J Mol Sci ; 20(17)2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31466215

RESUMO

Food peptides contain a very wide range of diversified structures, which explains their diverse range of functional activities. Proatherogenic endothelium is related to vasoconstriction, inflammation, and oxidative stress. In this line, four synthetic bioactive peptides from dry-cured pork ham, previously identified according to their Angiotensin I Converting Enzyme (ACE) inhibitory capacity and high bioavailability, were tested. Among them, KPVAAP displayed an estimated IC50 of 59.22 µM for human ACE inhibition, and docking simulations demonstrated the consistency of the noncompetitive binding with the protein. The addition of synthetic peptides to human endothelial cells significantly prevents the expression of genes related to endothelial dysfunction and inflammation (eNOS, ICAM-1, VCAM-1, IL-6) and lowers NF-κB activation (all p < 0.05). In silico dockings showed that the four bioactive peptides interact with the regulatory subunit NEMO of the NF-κB transcription factor at the same site as other characterized inhibitors (CC2-LZ region). This is the first study linking experimental and computational approaches that shows NF-κB to be the target of biopeptides of food origin. These multifunctional peptides from dry-cured pork ham make them good candidates for further research into their therapeutic or preventive use to attenuate the inflammatory atherosclerotic process.


Assuntos
Anti-Inflamatórios/farmacologia , Células Endoteliais/efeitos dos fármacos , Quinase I-kappa B/metabolismo , Proteínas de Carne/química , Simulação de Acoplamento Molecular , Oligopeptídeos/farmacologia , Anti-Inflamatórios/química , Sítios de Ligação , Linhagem Celular , Células Endoteliais/metabolismo , Endotélio Vascular/citologia , Endotélio Vascular/efeitos dos fármacos , Humanos , Quinase I-kappa B/química , Oligopeptídeos/química , Ligação Proteica
7.
Molecules ; 24(17)2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31466313

RESUMO

Fangchinoline (FCN) derived from Stephaniae tetrandrine S. Moore can be employed to treat fever, inflammation, rheumatism arthralgia, edema, dysuria, athlete's foot, and swollen wet sores. FCN can exhibit a plethora of anti-neoplastic effects although its precise mode of action still remains to be deciphered. Nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) can closely regulate carcinogenesis and thus we analyzed the possible action of FCN may have on these two signaling cascades in tumor cells. The effect of FCN on NF-κB and AP-1 signaling cascades and its downstream functions was deciphered using diverse assays in both human chronic myeloid leukemia (KBM5) and multiple myeloma (U266). FCN attenuated growth of both leukemic and multiple myeloma cells and repressed NF-κB, and AP-1 activation through diverse mechanisms, including attenuation of phosphorylation of IκB kinase (IKK) and p65. Furthermore, FCN could also cause significant enhancement in TNFα-driven apoptosis as studied by various molecular techniques. Thus, FCN may exhibit potent anti-neoplastic effects by affecting diverse oncogenic pathways and may be employed as pro-apoptotic agent against various malignancies.


Assuntos
Benzilisoquinolinas/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Mieloma Múltiplo/metabolismo , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Quinase I-kappa B/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Mieloma Múltiplo/tratamento farmacológico , NF-kappa B/metabolismo , Fosforilação/efeitos dos fármacos , Fator de Transcrição AP-1/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
8.
Transplant Proc ; 51(6): 2141-2145, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31307771

RESUMO

Indoleamine 2,3-dioxygenase (IDO) is an important enzyme in immune response regulation; cells that express IDO can suppress T-cell responses. Endothelial cells (ECs) play an important role in graft rejection. Therefore, we investigated the role of IDO expression by vascular ECs in immunoregulation. We demonstrated that interferon gamma can upregulate IDO expression in cultured ECs. Moreover, IDO induction by interferon gamma required IKKα activation, a part of the noncanonical NF-κB signaling pathway. In addition, IκB kinase α (IKKα) silencing resulted in significantly reduced IPO expression, demonstrating an essential role of NF-κB signaling pathway in IDO activation in vitro. These results may have an implication for regulating the immune response to alloantigens. The results obtained are important not only in understanding the role of ECs in the regulation of the transplantation immune response, but also in determining a potential therapeutic target for inhibiting allograft rejection.


Assuntos
Células Endoteliais/metabolismo , Rejeição de Enxerto/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Interferon gama/metabolismo , NF-kappa B/metabolismo , Animais , Regulação Enzimológica da Expressão Gênica , Rejeição de Enxerto/imunologia , Humanos , Quinase I-kappa B/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Linfócitos T/imunologia
9.
Mol Cell ; 75(4): 669-682.e5, 2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31302002

RESUMO

Phosphorylated IKKα(p45) is a nuclear active form of the IKKα kinase that is induced by the MAP kinases BRAF and TAK1 and promotes tumor growth independent of canonical NF-κB signaling. Insights into the sources of IKKα(p45) activation and its downstream substrates in the nucleus remain to be defined. Here, we discover that IKKα(p45) is rapidly activated by DNA damage independent of ATM-ATR, but dependent on BRAF-TAK1-p38-MAPK, and is required for robust ATM activation and efficient DNA repair. Abolishing BRAF or IKKα activity attenuates ATM, Chk1, MDC1, Kap1, and 53BP1 phosphorylation, compromises 53BP1 and RIF1 co-recruitment to sites of DNA lesions, and inhibits 53BP1-dependent fusion of dysfunctional telomeres. Furthermore, IKKα or BRAF inhibition synergistically enhances the therapeutic potential of 5-FU and irinotecan to eradicate chemotherapy-resistant metastatic human tumors in vivo. Our results implicate BRAF and IKKα kinases in the DDR and reveal a combination strategy for cancer treatment.


Assuntos
Dano ao DNA , Resistencia a Medicamentos Antineoplásicos , Fluoruracila/farmacologia , Quinase I-kappa B/metabolismo , Irinotecano/farmacologia , Sistema de Sinalização das MAP Quinases , Proteínas de Neoplasias , Neoplasias , Animais , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Células HCT116 , Humanos , Quinase I-kappa B/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Células MCF-7 , Camundongos , Camundongos Nus , Metástase Neoplásica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Telômero/genética , Telômero/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Mol Med Rep ; 20(2): 1418-1428, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31173238

RESUMO

An increasing body of evidence has demonstrated that microRNA (miR) deregulation serves pivotal roles in tumor progression and metastasis. However, the function of miR­379 in lung cancer remains understudied, particularly in non­small cell lung cancer (NSCLC). Bioinformatics and luciferase reporter analyses confirmed that conserved helix­loop­helix ubiquitous kinase (CHUK) is a target of miR­379, which may directly bind to the 3'­untranslated region of CHUK and significantly downregulate its expression in NSCLC cells. Transwell assays were used to evaluate the role of miR­379 in cell migration and invasion, and western blotting was used to address the association between miR­379 and epithelial­mesenchymal markers, including E­cadherin, cytokeratin and Vimentin. In the present study, miR­379 expression in NSCLC tissues and cell lines was downregulated, which may be associated with the poor survival of patients with NSCLC. miR­379 may act as a tumor suppressor in NSCLC, potentially by suppressing cell growth and proliferation, delaying G1­S transition, enhancing cell apoptosis and suppressing NSCLC cell migration and invasion. Furthermore, it was also observed that CHUK may function as an oncogene, and downregulation of CHUK induced by miR­379 may partially rescue the malignant characteristics of tumors, indicating that miR­379 may be suppressed in tumorigenesis. The overexpression of miR­379 may prevent the growth of NSCLC tumors via CHUK suppression and the downstream nuclear factor­κB pathway. The results of the present study demonstrated that miR­379 may act as a tumor suppressor, and may constitute a potential biomarker and a promising therapeutic agent for the treatment for NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Quinase I-kappa B/genética , Neoplasias Pulmonares/genética , MicroRNAs/genética , NF-kappa B/genética , Antígenos CD/genética , Antígenos CD/metabolismo , Sequência de Bases , Caderinas/genética , Caderinas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Humanos , Quinase I-kappa B/metabolismo , Queratinas/genética , Queratinas/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , MicroRNAs/agonistas , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Oligorribonucleotídeos/genética , Oligorribonucleotídeos/metabolismo , Transdução de Sinais , Técnicas de Cultura de Tecidos , Vimentina/genética , Vimentina/metabolismo
11.
Dev Cell ; 50(2): 229-246.e7, 2019 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-31204173

RESUMO

Serum starvation stimulates cilia growth in cultured cells, yet serum factors associated with ciliogenesis are unknown. Previously, we showed that starvation induces rapid Rab11-dependent vesicular trafficking of Rabin8, a Rab8 guanine-nucleotide exchange factor (GEF), to the mother centriole, leading to Rab8 activation and cilium growth. Here, we demonstrate that through the LPA receptor 1 (LPAR1), serum lysophosphatidic acid (LPA) inhibits Rab11a-Rabin8 interaction and ciliogenesis. LPA/LPAR1 regulates ciliogenesis initiation via downstream PI3K/Akt activation, independent of effects on cell cycle. Akt stabilizes Rab11a binding to its effector, WDR44, and a WDR44-pAkt-phosphomimetic mutant blocks ciliogenesis. WDR44 depletion promotes Rabin8 preciliary trafficking and ciliogenesis-initiating events at the mother centriole. Our work suggests disruption of Akt signaling causes a switch from Rab11-WDR44 to the ciliogenic Rab11-FIP3-Rabin8 complex. Finally, we demonstrate that Akt regulates downstream ciliogenesis processes associated with Rab8-dependent cilia growth. Together, this study uncovers a mechanism whereby serum mitogen signaling regulates Rabin8 preciliary trafficking and ciliogenesis initiation.


Assuntos
Cílios/fisiologia , Regulação da Expressão Gênica , Quinase I-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Animais , Humanos , Quinase I-kappa B/genética , Fosfatidilinositol 3-Quinases/genética , Transporte Proteico , Proteínas Proto-Oncogênicas c-akt/genética , Peixe-Zebra , Proteínas rab de Ligação ao GTP/genética
12.
Int Immunopharmacol ; 73: 193-200, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31103874

RESUMO

OBJECTIVE: This study was aimed to explore effect of baicalin on IKK/IKB/NF-kB signaling pathway and apoptosis-related proteins in rats with ulcerative colitis (UC). METHODS: Histopathological observation and scores of colon tissue were performed in the UC rat model. IKK/IKB/NF-kB signaling pathway and apoptosis-related proteins were measured by Western blotting. RESULTS: Baicalin significantly increased the activity of SOD, CAT and GSH-Px in colon tissue of rats with UC, but significantly decreased the content of MDA, IL-1ß, MPO, PEG2 and TNF-α in colon tissue of rats with UC. In the molecular mechanism, baicalin significantly decreased the expression of cleaved-caspase3, cleaved-caspase9, Bcl-2/Bax, cyt-c, NF-kB p-65, p-IKKß/IKKß and p-IKBα/IKBα. Baicalin could significantly inhibit p-IKBα/IKBα content change, but had no significant effect on p-IKKß/IKKß. CONCLUSION: Baicalin may have a regulating effect on IKK/IKB/NF-kB signaling pathway and apoptosis-related proteins in UC rats.


Assuntos
Colite Ulcerativa/metabolismo , Flavonoides/farmacologia , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Colite Ulcerativa/patologia , Colo/efeitos dos fármacos , Colo/metabolismo , Colo/patologia , Quinase I-kappa B/metabolismo , Interleucina-1beta/metabolismo , Masculino , Camundongos , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Peroxidase/metabolismo , Células RAW 264.7 , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo
13.
Neuroscience ; 409: 43-57, 2019 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-31047976

RESUMO

Inflammation aggravates the lethal consequences of intracerebral hemorrhage. Recently, many studies have found that nuclear factor-κB (NF-κB) is a crucial transcription factor that initiates inflammation in the perihematomal region of ICH. NF-κB essential modulator (NEMO)-binding domain (NBD) peptide, a cell-permeable peptide spanning the NBD of IKKα or IKKß, functions as a highly specific inhibitor of NF-κB. This peptide can negatively regulate the NF-κB pathway. The present study aimed to explore the effects and underlying pathomechanisms of NBD peptides after ICH. Striatum infusion of whole blood or saline was performed on C57BL/6 mice (n = 198). Experimental animals were administered NBD or control (mutated) peptides 2 h before or after ICH by intracerebroventricular injection (icv.). NBD peptides significantly inhibited edema formation, ameliorated the neurological deficits, markedly reduced IκBα and p65 phosphorylation, blocked nuclear translocation of p65, and upregulated IκBα expression by NF-κB after ICH induction. Using an in vitro hemin toxicity model, we investigated the effects of NBD peptides on microglial inflammation. We found that NBD peptides suppressed microglia inflammation and lowered the expression of TNF-α and IL-1ß in both in vivo and in vitro experiments. Further experiments were performed in mice and cultured microglia, which treated with NBD peptides in the presence of p65 siRNA confirmed that the specificity of NBD peptides inhibit ICH-induced NF-κB activation. This study demonstrated that NBD peptides exert a neuroprotective role after ICH and might be a potential candidate for a novel therapeutic strategy for ICH.


Assuntos
Edema Encefálico/tratamento farmacológico , Hemorragia Cerebral/tratamento farmacológico , Fármacos Neuroprotetores/uso terapêutico , Peptídeos/uso terapêutico , Regulação para Cima/efeitos dos fármacos , Animais , Edema Encefálico/metabolismo , Hemorragia Cerebral/metabolismo , Quinase I-kappa B/metabolismo , Interleucina-1beta/metabolismo , Camundongos , Microglia/efeitos dos fármacos , Microglia/metabolismo , NF-kappa B/metabolismo , Fármacos Neuroprotetores/farmacologia , Peptídeos/farmacologia , Fosforilação/efeitos dos fármacos , RNA Interferente Pequeno , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição RelA/metabolismo , Resultado do Tratamento , Fator de Necrose Tumoral alfa/metabolismo
14.
Mol Cell Biol ; 39(16)2019 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-31138664

RESUMO

Increased ribosomal biogenesis occurs during tissue hypertrophy, but whether ribosomal biogenesis is impaired during atrophy is not known. We show that hyperammonemia, which occurs in diverse chronic disorders, impairs protein synthesis as a result of decreased ribosomal content and translational capacity. Transcriptome analyses, real-time PCR, and immunoblotting showed consistent reductions in the expression of the large and small ribosomal protein subunits (RPL and RPS, respectively) in hyperammonemic murine skeletal myotubes, HEK cells, and skeletal muscle from hyperammonemic rats and human cirrhotics. Decreased ribosomal content was accompanied by decreased expression of cMYC, a positive regulator of ribosomal biogenesis, as well as reduced expression and activity of ß-catenin, a transcriptional activator of cMYC. However, unlike the canonical regulation of ß-catenin via glycogen synthase kinase 3ß (GSK3ß)-dependent degradation, GSK3ß expression and phosphorylation were unaltered during hyperammonemia, and depletion of GSK3ß did not prevent ammonia-induced degradation of ß-catenin. Overexpression of GSK3ß-resistant variants, genetic depletion of IκB kinase ß (IKKß) (activated during hyperammonemia), protein interactions, and in vitro kinase assays showed that IKKß phosphorylated ß-catenin directly. Overexpressing ß-catenin restored hyperammonemia-induced perturbations in signaling responses that regulate ribosomal biogenesis. Our data show that decreased protein synthesis during hyperammonemia is mediated via a novel GSK3ß-independent, IKKß-dependent impairment of the ß-catenin-cMYC axis.


Assuntos
Hiperamonemia/metabolismo , Subunidades Ribossômicas Menores/genética , Subunidades Ribossômicas Menores/metabolismo , beta Catenina/química , beta Catenina/genética , Animais , Linhagem Celular , Modelos Animais de Doenças , Fibrose , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Células HEK293 , Humanos , Hiperamonemia/genética , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Camundongos , Proteólise , Proteômica , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Ratos , Análise de Sequência de RNA , Transdução de Sinais
15.
Cardiovasc Pathol ; 41: 11-17, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31004933

RESUMO

Atherosclerosis is a chronic inflammatory disease with lipid accumulation. Apolipoprotein C3 (APOC3), which is an important regulator of human lipid metabolism, is associated with multiple vascular mechanisms in atherosclerosis and proinflammatory responses. We have previously reported that the expression of inflammatory cytokine TNF-α is elevated in human endothelial cells (HUVECs) after APOC3 treatment. This study investigates the APOC3 signaling pathway involved in TNF-α-mediated expression of JAM-1 in HUVECs. Cultured HUVECs were exposed to APOC3 (50 µg/ml) for 16 h. Mechanistic studies were carried out by silencing TNF-α gene with lentiviral TNF-α-shRNA. Our study was based on the eight signaling pathway inhibitors to block the effect of APOC3 in HUVECs. The expression of JAM-1 was determined by qRT-PCR, Western blotting, and flow cytometry. IKK2 degradation and NF-κB p65 phosphorylation were determined by Western blotting. Our results showed that APOC3 significantly promoted the TNF-α-induced expression of JAM-1 in HUVECs. Inhibiting APOC3 reversed the TNF-α-induced overexpression of JAM-1. Moreover, APOC3 induced the expression of NF-κB p65 and degraded IκB. In conclusion, APOC3 promoted the expression of JAM-1 via the NF-κB, IKK2, and PI3K signaling pathway.


Assuntos
Apolipoproteína C-III/farmacologia , Moléculas de Adesão Celular/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Quinase I-kappa B/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Receptores de Superfície Celular/metabolismo , Fator de Transcrição RelA/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Moléculas de Adesão Celular/genética , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/enzimologia , Humanos , Fosforilação , Proteólise , Interferência de RNA , Receptores de Superfície Celular/genética , Transdução de Sinais/efeitos dos fármacos , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/enzimologia , Fator de Necrose Tumoral alfa/genética
16.
Nat Commun ; 10(1): 1729, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30988283

RESUMO

RIPK1 regulates cell death and inflammation through kinase-dependent and -independent mechanisms. As a scaffold, RIPK1 inhibits caspase-8-dependent apoptosis and RIPK3/MLKL-dependent necroptosis. As a kinase, RIPK1 paradoxically induces these cell death modalities. The molecular switch between RIPK1 pro-survival and pro-death functions remains poorly understood. We identify phosphorylation of RIPK1 on Ser25 by IKKs as a key mechanism directly inhibiting RIPK1 kinase activity and preventing TNF-mediated RIPK1-dependent cell death. Mimicking Ser25 phosphorylation (S > D mutation) protects cells and mice from the cytotoxic effect of TNF in conditions of IKK inhibition. In line with their roles in IKK activation, TNF-induced Ser25 phosphorylation of RIPK1 is defective in TAK1- or SHARPIN-deficient cells and restoring phosphorylation protects these cells from TNF-induced death. Importantly, mimicking Ser25 phosphorylation compromises the in vivo cell death-dependent immune control of Yersinia infection, a physiological model of TAK1/IKK inhibition, and rescues the cell death-induced multi-organ inflammatory phenotype of the SHARPIN-deficient mice.


Assuntos
Apoptose , Modelos Imunológicos , Proteína Serina-Treonina Quinases de Interação com Receptores/fisiologia , Animais , Caspase 8/genética , Caspase 8/metabolismo , Caspase 8/fisiologia , Linhagem Celular , Quinase I-kappa B/metabolismo , Quinase I-kappa B/fisiologia , Imunidade/fisiologia , Camundongos , Fosforilação , Proteína Serina-Treonina Quinases de Interação com Receptores/química , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Serina/química , Serina/metabolismo , Yersinia , Yersiniose/imunologia
17.
Nat Commun ; 10(1): 1801, 2019 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-30996248

RESUMO

Macrophage-orchestrated, low-grade chronic inflammation plays a pivotal role in obesity and atherogenesis. However, the underlying regulatory mechanisms remain incompletely understood. Here, we identify major vault protein (MVP), the main component of unique cellular ribonucleoprotein particles, as a suppressor for NF-κB signaling in macrophages. Both global and myeloid-specific MVP gene knockout aggravates high-fat diet induced obesity, insulin resistance, hepatic steatosis and atherosclerosis in mice. The exacerbated metabolic disorders caused by MVP deficiency are accompanied with increased macrophage infiltration and heightened inflammatory responses in the microenvironments. In vitro studies reveal that MVP interacts with TRAF6 preventing its recruitment to IRAK1 and subsequent oligomerization and ubiquitination. Overexpression of MVP and its α-helical domain inhibits the activity of TRAF6 and suppresses macrophage inflammation. Our results demonstrate that macrophage MVP constitutes a key constraint of NF-κB signaling thereby suppressing metabolic diseases.


Assuntos
Aterosclerose/imunologia , Fígado Gorduroso/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Obesidade/imunologia , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Tecido Adiposo/patologia , Animais , Aterosclerose/etiologia , Aterosclerose/metabolismo , Biópsia , Células da Medula Óssea , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Fígado Gorduroso/etiologia , Fígado Gorduroso/metabolismo , Feminino , Técnicas de Inativação de Genes , Humanos , Quinase I-kappa B/metabolismo , Inflamação/etiologia , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , NF-kappa B/metabolismo , Obesidade/etiologia , Obesidade/metabolismo , Obesidade/patologia , Cultura Primária de Células , Transdução de Sinais/imunologia , Fator 6 Associado a Receptor de TNF/metabolismo , Ubiquitinação , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética , Partículas de Ribonucleoproteínas em Forma de Abóbada/imunologia
18.
Eur J Pharmacol ; 855: 10-19, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31028739

RESUMO

The emergence of resistance to EGF receptor (EGFR) inhibitor therapy is a significant challenge for patients with non-small cell lung cancer (NSCLC). During the past few years, a correlation between EGFR TKIs resistance and dysregulation of IKKß/NF-κB signaling has been increasingly suggested. However, few studies have focused on the effects of combining IKK/NF-κB and EGFR inhibitors to overcome EGFR TKIs resistance. In this study, we discovered that Schizandrin A (Sch A), a lignin compound isolated from Schisandra chinesnesis, could synergize with the EGFR receptor inhibitor Gefitinib to inhibit cell growth, induce cell cycle arrest and apoptosis of HCC827/GR cells. Sch A effectively suppressed the phosphorylation of IKKß and IκBα, as well as the nuclear translocation of NF-κB p65, and showed high and selective affinity for IKKß in surface plasmon resonance (SPR) experiments, indicating that Sch A was a selective IKKß inhibitor. Molecular modeling between IKKß and Sch A suggested that Sch A formed key hydrophobic interactions with IKKß, which may contribute to its potent IKKß inhibitory effect. These findings suggest a novel approach to improve poor clinical outcomes in EGFR TKIs therapy, by combining it with Sch A.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Ciclo-Octanos/farmacologia , Gefitinibe/farmacologia , Quinase I-kappa B/metabolismo , Lignanas/farmacologia , Neoplasias Pulmonares/patologia , NF-kappa B/metabolismo , Compostos Policíclicos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sinergismo Farmacológico , Humanos , Quinase I-kappa B/química , Simulação de Dinâmica Molecular , Conformação Proteica
19.
Can J Cardiol ; 35(4): 490-500, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30935640

RESUMO

BACKGROUND: The IκB kinase (IKK) complex has been found to have critical functions in cancer and the immune system. In particular, IKKα, which is a member of the IKK complex, has been shown to influence the inflammatory response and malignant diseases. However, the role of IKKα in macrophages after myocardial infarction (MI) remains largely unknown. METHODS: Sham or MI operations were performed on macrophage-specific IKKɑ knockout (mIKKɑ-/-) mice and IKKɑflox/flox littermates. We ligated the left anterior descending coronary artery of the MI group and observed the results at 3, 7, and 30 days after MI. RESULTS: We discovered more severe cardiac dysfunction with reduced angiogenesis, fibrosis, and collagen deposition in mIKKɑ-/- than in IKKɑflox/flox. In addition, we also observed that macrophages in mIKKɑ-/- were easier to polarize to the M1 phenotype and expressed more proinflammatory factors than IKKɑflox/flox. Mechanistically, IKKα deficiency in macrophages inhibited the alternative nuclear factor-κB/RelB pathway and enhanced the MEK1/2/ERK1/2 pathway. CONCLUSIONS: Overall, our data identified IKKɑ in the heart as a novel mediator that protected the heart from a severe inflammatory response and attenuated ventricular remodelling after MI by negatively regulating macrophage polarization to the M1 phenotype. Therefore, IKKα may serve as a potential therapeutic target for treatment after MI.


Assuntos
Quinase I-kappa B/metabolismo , Macrófagos/metabolismo , Infarto do Miocárdio/fisiopatologia , Remodelação Ventricular/fisiologia , Animais , Colágeno/metabolismo , Fibrose , Quinase I-kappa B/genética , MAP Quinase Quinase 1/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Macrófagos/imunologia , Camundongos Knockout , Miocárdio/metabolismo , Miocárdio/patologia , Neovascularização Patológica/metabolismo
20.
Eur J Pharmacol ; 854: 179-186, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-30940449

RESUMO

Doxorubicin is a chemotherapeutic agent commonly used to treat breast cancer. However, breast cancer often develops drug resistance, leading to disease recurrence and poor prognosis. Delineating the mechanisms underlying drug resistance is imperative for overcoming the challenge of treating doxorubicin-resistant breast cancer. In this study, by identifying the possible role of Sentrin/SUMO-specific proteases (SENPs) in doxorubicin resistance, we show here that among the 6 members of SENPs, only SENP2 is downregulated in doxorubicin-resistant MCF-7 (MCF-7/adr) and MDA-MB-231 (dr) breast cancer cells, as compared with sensitive counterparts. In addition, functionally, SENP2 overexpression resensitizes resistant breast cancer cells to doxorubicin treatment, and its knockdown confers doxorubicin resistance in sensitive ones. Moreover, NF-κB pathway is activated in MCF-7/adr cells, however, treatment with Bay 11-7085, one specific inhibitor of this pathway, reverses resistance to doxorubicin, suggesting that NF-κB pathway activation contributes to doxorubicin resistance in MCF-7/adr cells. We further show that SENP2 overexpression enhances NEMO deSUMOylation and suppresses NF-κB activation particularly in MCF-7/adr cells. Furthermore, SENP2 overexpression-induced sensitivity of MCF-7/adr cells to doxorubicin is drastically abrogated when treated with NF-κB pathway activator, thus establishing a causal link between SENP2-suppressed NF-κB pathway and enhanced doxorubicin sensitivity in breast cancer cells. Overall, this study reveals a novel function of SENP2 in counteracting doxorubicin resistance in breast cancer, and highlights the critical role of NF-κB suppression in mediating this effect.


Assuntos
Neoplasias da Mama/patologia , Cisteína Endopeptidases/metabolismo , Doxorrubicina/farmacologia , NF-kappa B/metabolismo , Cisteína Endopeptidases/genética , Regulação para Baixo/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Humanos , Quinase I-kappa B/metabolismo , Células MCF-7 , Transdução de Sinais/efeitos dos fármacos , Sumoilação/efeitos dos fármacos
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