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1.
Chem Biol Interact ; 332: 109304, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33164868

RESUMO

Cisplatin-based chemotherapy is a common first-line regimen for treating non-small cell lung cancer (NSCLC). However, drug resistance is still a major problem. The purposes of this study were to evaluate whether sclareol can reverse cisplatin resistance and to investigate its possible mechanisms. A549 cells, the human NSCLC cells with inherent cisplatin resistance, were used to investigate synergistic effect of sclareol with cisplatin in cell proliferation and migration as well as its regulatory mechanisms in expression of excision repair cross-complementation group 1 (ERCC1), a cisplatin resistance-associated molecule. Nude mice bearing subcutaneous A549 tumors were applied to investigate synergistic activity of sclareol in anti-tumor. As comparing to the cisplatin alone group, the treatment of cisplatin combined with sclareol significantly suppressed survival rate and cell migration of A549 cells. Besides, sclareol also exhibited suppression in ERCC1 expression by inhibiting AKT-GSK3ß-AP1/Snail and JNK-AP1 pathways. Furthermore, the experimental data from in vivo study also demonstrated that the combination group of cisplatin and sclareol showed the greatest anti-tumor activity, whose effect could be partially attributed to sclareol-mediated decrease in intratumoral level of ERCC1 protein. Accordingly, sclareol has potential as an adjuvant for the treatment in NSCLC patients with cisplatin resistance.


Assuntos
Adenocarcinoma de Pulmão/patologia , Cisplatino/farmacologia , Diterpenos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteína de Replicação C/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Células A549 , Animais , Movimento Celular/efeitos dos fármacos , Diterpenos/química , Sinergismo Farmacológico , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Camundongos , Modelos Biológicos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Fator de Transcrição AP-1/metabolismo
2.
Nat Commun ; 11(1): 5079, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033234

RESUMO

Tumor heterogeneity and lack of knowledge about resistant cell states remain a barrier to targeted cancer therapies. Basal cell carcinomas (BCCs) depend on Hedgehog (Hh)/Gli signaling, but can develop mechanisms of Smoothened (SMO) inhibitor resistance. We previously identified a nuclear myocardin-related transcription factor (nMRTF) resistance pathway that amplifies noncanonical Gli1 activity, but characteristics and drivers of the nMRTF cell state remain unknown. Here, we use single cell RNA-sequencing of patient tumors to identify three prognostic surface markers (LYPD3, TACSTD2, and LY6D) which correlate with nMRTF and resistance to SMO inhibitors. The nMRTF cell state resembles transit-amplifying cells of the hair follicle matrix, with AP-1 and TGFß cooperativity driving nMRTF activation. JNK/AP-1 signaling commissions chromatin accessibility and Smad3 DNA binding leading to a transcriptional program of RhoGEFs that facilitate nMRTF activity. Importantly, small molecule AP-1 inhibitors selectively target LYPD3+/TACSTD2+/LY6D+ nMRTF human BCCs ex vivo, opening an avenue for improving combinatorial therapies.


Assuntos
Carcinoma Basocelular/metabolismo , Proteínas Hedgehog/metabolismo , Transdução de Sinais , Neoplasias Cutâneas/metabolismo , Fator de Transcrição AP-1/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Cromatina/metabolismo , DNA de Neoplasias/metabolismo , Resistencia a Medicamentos Antineoplásicos , Matriz Extracelular/metabolismo , Ontologia Genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Folículo Piloso/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Células NIH 3T3 , Proteínas de Neoplasias/metabolismo , Ligação Proteica , Proteína Smad3/metabolismo , Transativadores/metabolismo , Regulação para Cima
3.
Medicine (Baltimore) ; 99(38): e22241, 2020 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-32957369

RESUMO

BACKGROUND: Quercetin, a major flavonol, wildly exists in plantage, which has been reported to have an anti-apoptosis and anti-inflammation effects on vascular endothelial cells, but its underlying molecular mechanisms remain unclear. OBJECTIVE: The aim of this study was to investigate the mechanisms of how quercetin inhibits tumor necrosis factor alpha (TNF-α) induced human umbilical vein endothelial cells (HUVECs) apoptosis and inflammation. METHODS AND RESULTS: HUVECs were preconditioned with quercetin for 18 hours, and subsequently treated with TNF-α for 6 hours to induce apoptosis. The expression of intercellular cell adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), E-selectin, ß-actin mRNA was then detected by RT-PCR. Flow cytometry was used to estimate the apoptosis rates, and the expression of activator protein 1 (AP-1) and nuclear factor kappa B (NF-κB) was measured by Western blot. TNF-α induced elevated apoptosis rates and upregulation of VCAM-1, ICAM-1, and E-selectin were meaningfully reduced in HUVECs by pretreatment with quercetin. In addition, quercetin also inhibited the activation of AP-1and NF-κB. CONCLUSION: Results indicate that quercetin could suppress TNF-α induced apoptosis and inflammation by blocking NF-κB and AP-1 signaling pathway in HUVECs, which might be one of the underlying mechanisms in treatment of coronary heart disease.


Assuntos
Anti-Inflamatórios/farmacologia , Apoptose/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/citologia , Inflamação/prevenção & controle , NF-kappa B/metabolismo , Quercetina/farmacologia , Fator de Transcrição AP-1/metabolismo , Regulação para Baixo , Selectina E/metabolismo , Humanos , Inflamação/metabolismo , Inflamação/patologia , Molécula 1 de Adesão Intercelular/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/farmacologia , Molécula 1 de Adesão de Célula Vascular/metabolismo
4.
Science ; 369(6508)2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32883834

RESUMO

Vertebrates vary in their ability to regenerate, and the genetic mechanisms underlying such disparity remain elusive. Comparative epigenomic profiling and single-cell sequencing of two related teleost fish uncovered species-specific and evolutionarily conserved genomic responses to regeneration. The conserved response revealed several regeneration-responsive enhancers (RREs), including an element upstream to inhibin beta A (inhba), a known effector of vertebrate regeneration. This element activated expression in regenerating transgenic fish, and its genomic deletion perturbed caudal fin regeneration and abrogated cardiac regeneration altogether. The enhancer is present in mammals, shares functionally essential activator protein 1 (AP-1)-binding motifs, and responds to injury, but it cannot rescue regeneration in fish. This work suggests that changes in AP-1-enriched RREs are likely a crucial source of loss of regenerative capacities in vertebrates.


Assuntos
Elementos Facilitadores Genéticos/fisiologia , Evolução Molecular , Peixes Listrados/genética , Peixes Listrados/fisiologia , Regeneração/genética , Motivos de Aminoácidos , Animais , Epigênese Genética , Perfilação da Expressão Gênica , Histonas/metabolismo , Subunidades beta de Inibinas/genética , RNA-Seq , Análise de Célula Única , Fator de Transcrição AP-1/química , Fator de Transcrição AP-1/metabolismo , Ativação Transcricional , Peixe-Zebra/genética , Peixe-Zebra/fisiologia
5.
Ceska Slov Farm ; 69(3): 130-136, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32972156

RESUMO

Geranylated flavanone diplacone is a flavanone iso- lated from Paulownia tomentosa (Thunb.) Steud. (Paulowniaceae) with anti-inflammatory and antioxidant properties, nevertheless showing high lipophilicity and low solubility in water. Diplacone was therefore used as a model molecule for incorporation into glucan particles (GPs). GPs are prepared by intensive washing of yeast (Saccharomyces cerevisiae) leading to hollow shells consisting of β-(13)/β-(16) glucan mainly. The aim of this study was to compare anti-inflammatory potential of GPs-diplacone composites with the compound itself, GPs themselves and the physical mixture of GPs and diplacone. The cell line THP1-XBlueTM-MD2-CD14 derived from human leukemic monocytes was stimulated with lipopolysaccharide (LPS) from Escherichia coli to trigger inflammatory reaction. The composites of GPs with diplacone significantly decreased the activity of pro-inflammatory transcription factors nuclear factor κB (NF-κB) and activator protein 1 (AP-1).


Assuntos
Anti-Inflamatórios/farmacologia , Flavanonas/farmacologia , Glucanos/química , Saccharomyces cerevisiae/química , Humanos , NF-kappa B/metabolismo , Células THP-1 , Fator de Transcrição AP-1/metabolismo
6.
Res Vet Sci ; 132: 462-465, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32799169

RESUMO

Porcine reproductive and respiratory syndrome virus (PRRSV) is a causative pathogen of PRRS that has generated a serious adverse impact on current global pork production. PRRSV primarily infects pig alveolar macrophages, but poor induction of innate immunity after infection often leads to more severe complications. Defining the functional role of each PRRSV non structural protein (NSP) within host cells might be helpful in understanding how PRRSV induces poor innate immunity in host cells. NSP1 of PRRSV exhibits papain like cysteine protease activity and may therefore modulate host cell signaling by degrading a target protein in host cells during infection. In this study, we demonstrated that NSP1 of PRRSV-2 indirectly blocked extracellular signal-regulated kinase (ERK) signaling in polyriboinosinic polyribocytidylic acid (Poly I:C) stimulated pig macrophages (3D4/31 cells). ERK which belongs to the mitogen-activated protein kinase family mediates many biological processes including inflammatory responses during viral infection. The blocking of ERK signaling by NSP1 of PRRSV-2 further leads to the transcriptional inhibition of inflammatory enhancers, cellular communication network factor 1 and 2 (ccn1 and ccn2) through down-regulation of v-fos FBJ murine osteosarcoma viral oncogene homolog (fos) and fosb, the component of activating protein-1 (AP-1) which is an ERK downstream transcription factor. Therefore, NSP1 of PRRSV-2 inhibited the mRNA transcription of ccn1 and ccn2 by blocking the ERK-AP-1 axis in Poly I:C stimulated pig macrophages. These results provide additional evidence supporting that NSP1 has anti-inflammatory function during PRRSV-2 infection.


Assuntos
Fator de Crescimento do Tecido Conjuntivo/metabolismo , Proteína Rica em Cisteína 61/metabolismo , Vírus da Síndrome Respiratória e Reprodutiva Suína , Fator de Transcrição AP-1/metabolismo , Proteínas não Estruturais Virais/metabolismo , Animais , Linhagem Celular , Fator de Crescimento do Tecido Conjuntivo/genética , Proteína Rica em Cisteína 61/genética , Regulação para Baixo , Imunidade Inata , Macrófagos , Macrófagos Alveolares , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais , Suínos , Fator de Transcrição AP-1/genética , Proteínas não Estruturais Virais/genética
7.
Proc Natl Acad Sci U S A ; 117(34): 20860-20867, 2020 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-32788362

RESUMO

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that latently infects hematopoietic cells and has the ability to reactivate when triggered by immunological stress. This reactivation causes significant morbidity and mortality in immune-deficient patients, who are unable to control viral dissemination. While a competent immune system helps prevent clinically detectable viremia, a portrait of the factors that induce reactivation following the proper cues remains incomplete. Our understanding of the complex molecular mechanisms underlying latency and reactivation continues to evolve. We previously showed the HCMV-encoded G protein-coupled receptor US28 is expressed during latency and facilitates latent infection by attenuating the activator protein-1 (AP-1) transcription factor subunit, c-fos, expression and activity. We now show AP-1 is a critical component for HCMV reactivation. Pharmacological inhibition of c-fos significantly attenuates viral reactivation. In agreement, infection with a virus in which we disrupted the proximal AP-1 binding site in the major immediate early (MIE) enhancer results in inefficient reactivation compared to WT. Concomitantly, AP-1 recruitment to the MIE enhancer is significantly decreased following reactivation of the mutant virus. Furthermore, AP-1 is critical for derepression of MIE-driven transcripts and downstream early and late genes, while immediate early genes from other loci remain unaffected. Our data also reveal MIE transcripts driven from the MIE promoter, the distal promoter, and the internal promoter, iP2, are dependent upon AP-1 recruitment, while iP1-driven transcripts are AP-1-independent. Collectively, our data demonstrate AP-1 binding to and activation of the MIE enhancer is a key molecular process controlling reactivation from latency.


Assuntos
Citomegalovirus/genética , Fator de Transcrição AP-1/metabolismo , Ativação Viral/genética , Citomegalovirus/metabolismo , Citomegalovirus/fisiologia , Infecções por Citomegalovirus/virologia , Genes Precoces/genética , Humanos , Proteínas Imediatamente Precoces/genética , Regiões Promotoras Genéticas/genética , Transdução de Sinais/genética , Fator de Transcrição AP-1/genética , Ativação Transcricional/genética , Latência Viral/genética
8.
Science ; 369(6505): 842-846, 2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32792398

RESUMO

How is neuropathic pain regulated in peripheral sensory neurons? Importins are key regulators of nucleocytoplasmic transport. In this study, we found that importin α3 (also known as karyopherin subunit alpha 4) can control pain responsiveness in peripheral sensory neurons in mice. Importin α3 knockout or sensory neuron-specific knockdown in mice reduced responsiveness to diverse noxious stimuli and increased tolerance to neuropathic pain. Importin α3-bound c-Fos and importin α3-deficient neurons were impaired in c-Fos nuclear import. Knockdown or dominant-negative inhibition of c-Fos or c-Jun in sensory neurons reduced neuropathic pain. In silico screens identified drugs that mimic importin α3 deficiency. These drugs attenuated neuropathic pain and reduced c-Fos nuclear localization. Thus, perturbing c-Fos nuclear import by importin α3 in peripheral neurons can promote analgesia.


Assuntos
Dor Crônica/fisiopatologia , Neuralgia/fisiopatologia , Células Receptoras Sensoriais/fisiologia , alfa Carioferinas/fisiologia , Transporte Ativo do Núcleo Celular , Animais , Benzofenonas/farmacologia , Dor Crônica/genética , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Isoxazóis/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Neuralgia/genética , Proteínas Proto-Oncogênicas c-fos/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-fos/metabolismo , Fator de Transcrição AP-1/metabolismo , alfa Carioferinas/genética
9.
Nature ; 584(7820): 268-273, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32728211

RESUMO

The ability of the skin to grow in response to stretching has been exploited in reconstructive surgery1. Although the response of epidermal cells to stretching has been studied in vitro2,3, it remains unclear how mechanical forces affect their behaviour in vivo. Here we develop a mouse model in which the consequences of stretching on skin epidermis can be studied at single-cell resolution. Using a multidisciplinary approach that combines clonal analysis with quantitative modelling and single-cell RNA sequencing, we show that stretching induces skin expansion by creating a transient bias in the renewal activity of epidermal stem cells, while a second subpopulation of basal progenitors remains committed to differentiation. Transcriptional and chromatin profiling identifies how cell states and gene-regulatory networks are modulated by stretching. Using pharmacological inhibitors and mouse mutants, we define the step-by-step mechanisms that control stretch-mediated tissue expansion at single-cell resolution in vivo.


Assuntos
Mecanotransdução Celular/fisiologia , Análise de Célula Única , Pele/citologia , Pele/crescimento & desenvolvimento , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Junções Aderentes/metabolismo , Animais , Sequência de Bases , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/efeitos dos fármacos , Autorrenovação Celular/efeitos dos fármacos , Cromatina/efeitos dos fármacos , Cromatina/genética , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Células Clonais/citologia , Células Clonais/efeitos dos fármacos , Células Clonais/metabolismo , Modelos Animais de Doenças , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Redes Reguladoras de Genes/efeitos dos fármacos , Hidrogéis/administração & dosagem , Hidrogéis/farmacologia , Mecanotransdução Celular/efeitos dos fármacos , Mecanotransdução Celular/genética , Camundongos , Camundongos Transgênicos , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Mutação , RNA Mensageiro/genética , RNA-Seq , Pele/efeitos dos fármacos , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Transativadores/antagonistas & inibidores , Transativadores/metabolismo , Fator de Transcrição AP-1/metabolismo , Transcrição Genética/efeitos dos fármacos
10.
Toxicol Appl Pharmacol ; 402: 115115, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32634518

RESUMO

Physalin A (PA), a withanolide, was isolated from Physalis angulata L. In this study, it is shown that PA can inhibit the production of inflammatory cytokines such as PGE2, NO, IL-1ß, IL-6, and TNF-α in LPS-induced RAW 264.7 cells. Furthermore, the results indicated that PA suppressed the IκB/NF-κB and JNK/ AP-1 inflammatory signaling pathways and inhibited the levels of pro-inflammatory factors iNOS and COX-2 in LPS-stimulated RAW 264.7 cells. In the carrageenan-induced mouse hind paw edema study, PA was shown to inhibit the production of inflammatory mediators such as NO, MDA, and TNF-α production. Conversely, the antioxidant factor levels of SOD, CAT, and GPx were all increased by the treated PA. According to the data, we are suggesting that the anti-inflammatory effects of PA may be through the suppressions of the JNK/AP-1 and IκB/NF-κB signaling pathways and up-regulation of the anti-oxidative activity.


Assuntos
Inflamação/tratamento farmacológico , Inflamação/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fator de Transcrição AP-1/metabolismo , Vitanolídeos/farmacologia , Animais , Antioxidantes/química , Antioxidantes/farmacologia , Carragenina/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas I-kappa B/genética , Proteínas I-kappa B/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Masculino , Malondialdeído , Camundongos , Camundongos Endogâmicos ICR , Estrutura Molecular , NF-kappa B/genética , NF-kappa B/metabolismo , Physalis/química , Células RAW 264.7 , Distribuição Aleatória , Fator de Transcrição AP-1/genética , Vitanolídeos/química
11.
Environ Toxicol ; 35(12): 1352-1363, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32677756

RESUMO

Although pipoxolan (PIPO) is a smooth muscle relaxant, its anti-inflammatory capability has not been studied. Therefore, we investigated the anti-inflammatory molecular mechanisms of PIPO in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. In this study, we used the MTT assay to evaluate the cytotoxicity, applied the enzyme-linked immunosorbent assay to determine the inflammatory cytokines, and performed Western blotting to assess protein expression. The results showed that PIPO significantly inhibited cytokine production, including nitric oxide, prostaglandin E2 , tumor necrosis factor-α, and interleukin-6. PIPO also suppressed the pro-inflammatory mediator expression with inducible nitric oxide synthase and cyclooxygenase-2. Moreover, PIPO prohibited the multiple inflammatory transcription factor pathways, including inhibitor kappa B/nuclear factor of the κ light chain enhancer of B cells (NF-κB), mitogen-activated protein kinase/activator protein-1 (AP-1), Janus kinase/signal transducer and activator of transcription (STAT), and toll-like receptor 4 (TLR4)/serine/threonine kinase (AKT). Besides, PIPO effectively activated the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 antioxidative pathway. Collectively, PIPO may attenuate the inflammatory effects via influencing the LPS/TLR4 receptor binding; suppress the expression of anti-inflammatory transcription factors NF-κB, AP-1, and STAT; and activating the antioxidative transcription factor Nrf2 in LPS-stimulated mouse RAW 264.7 cells.


Assuntos
Anti-Inflamatórios/farmacologia , Dioxolanos/farmacologia , Macrófagos/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Fatores de Transcrição STAT/metabolismo , Fator de Transcrição AP-1/metabolismo , Animais , Antioxidantes/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Óxido Nítrico Sintase Tipo II/metabolismo , Células RAW 264.7
12.
J Environ Pathol Toxicol Oncol ; 39(1): 13-21, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32479009

RESUMO

Breast cancer is a widespread disease that affects women globally. Diagnostic processes and remedial approaches to breast carcinogenesis have improved in recent decades, but continuous survival of patients with breast carcinogenesis is still lacking due to increased cell proliferation. The aim of the present work is to explore the anticell proliferative effects of nobiletin (NOB) against 7,12-dimethylbenz[a]anthracene (DMBA)-treated mammary tumorigenesis in rats. We stimulate mammary carcinogenesis using oral gavage of DMBA (25 mg/kg body weight) mixed with olive oil (1 mL). This results in reduced body weight; increased liver marker enzymes such as alkaline phosphatase, acid phosphatase, aspartate aminotransferase, and alanine aminotransferase; and cell proliferative markers such as c-Jun, proliferating cell nuclear antigen, c-Fos, cyclin D1, and activating protein-1 (AP-1) in the DMBA-treated cancer-bearing animals. NOB administration improved body weight, significantly reduced hepatic marker enzymes, and altered histopathological changes. Furthermore, NOB efficiently reduced tumor cell proliferation markers in DMBA-induced mammary carcinogenesis. Overall, these results suggest that NOB has an anticell proliferative effect on DMBA-induced mammary cancer via modulation of the AP-1 signaling pathway.


Assuntos
9,10-Dimetil-1,2-benzantraceno/toxicidade , Antioxidantes/farmacologia , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Flavonas/farmacologia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Animais , Carcinógenos/toxicidade , Transformação Celular Neoplásica/efeitos dos fármacos , Feminino , Ratos , Ratos Sprague-Dawley , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismo
13.
Nat Cell Biol ; 22(7): 842-855, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32514071

RESUMO

Senescent cells affect many physiological and pathophysiological processes. While select genetic and epigenetic elements for senescence induction have been identified, the dynamics, epigenetic mechanisms and regulatory networks defining senescence competence, induction and maintenance remain poorly understood, precluding the deliberate therapeutic targeting of senescence for health benefits. Here, we examined the possibility that the epigenetic state of enhancers determines senescent cell fate. We explored this by generating time-resolved transcriptomes and epigenome profiles during oncogenic RAS-induced senescence and validating central findings in different cell biology and disease models of senescence. Through integrative analysis and functional validation, we reveal links between enhancer chromatin, transcription factor recruitment and senescence competence. We demonstrate that activator protein 1 (AP-1) 'pioneers' the senescence enhancer landscape and defines the organizational principles of the transcription factor network that drives the transcriptional programme of senescent cells. Together, our findings enabled us to manipulate the senescence phenotype with potential therapeutic implications.


Assuntos
Senescência Celular , Cromatina/metabolismo , Epigênese Genética , Fibroblastos/citologia , Regulação da Expressão Gênica , Histonas/metabolismo , Fator de Transcrição AP-1/metabolismo , Transcriptoma , Animais , Cromatina/genética , Feminino , Fibroblastos/metabolismo , Histonas/genética , Humanos , Camundongos Endogâmicos C57BL , Fator de Transcrição AP-1/genética
14.
Biochim Biophys Acta Mol Cell Res ; 1867(10): 118766, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32504661

RESUMO

Amlexanox, an anti-inflammatory agent, is widely used for treating aphthous ulcers. Recently, amlexanox has received considerable attention because of its efficacy in mitigating metabolic inflammation via directly suppressing IKKε/TBK1. However, because the knockdown of IKKε/TBK1 has no anti-inflammatory effect on lipopolysaccharide (LPS)-primed RAW264.7 cells, the mechanism of amlexanox against classical inflammation is independent of IKKε/TBK1. In this study, we aim to examine the effects of amlexanox on LPS-treated macrophages and in a mouse model of endotoxemia. We found that amlexanox significantly inhibited the production of pro-inflammatory mediators, both in vitro and in vivo, while increased interleukin-10 level in LPS-activated macrophages. Mechanistically, amlexanox down-regulated nuclear factor κB and extracellular signal-regulated kinase/activator protein-1 signaling by elevating intracellular 3',5'-cyclic adenosine monophosphate (cAMP) level and subsequently activating protein kinase A. Molecular docking along with fluorescence polarization and enzyme inhibition assays revealed that amlexanox bound directly to phosphodiesterase (PDE) 4B to inhibit its activity. The anti-inflammatory effects of amlexanox could be abolished by the application of cAMP antagonist or PDE4B siRNA. In addition to PDE4B, the activities of PDE1C, 3A, and 3B were directly inhibited by amlexanox. Our results provide mechanistic insight into the clinical utility of amlexanox for the treatment of inflammatory disorders and might contribute to extending the clinical indications of amlexanox.


Assuntos
Aminopiridinas/farmacologia , Anti-Inflamatórios/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Aminopiridinas/química , Animais , Anti-Inflamatórios/química , AMP Cíclico/metabolismo , Inflamação/patologia , Interleucina-10/metabolismo , Lipopolissacarídeos , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos ICR , Milrinona/farmacologia , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Células RAW 264.7 , Transdução de Sinais/efeitos dos fármacos , Espectrometria de Fluorescência , Fator de Transcrição AP-1/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
15.
Anticancer Res ; 40(6): 3255-3264, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32487620

RESUMO

BACKGROUND/AIM: Rta, a transactivator of Epstein-Barr virus, is associated with progression of nasopharyngel carcinoma (NPC); however, its mechanism of contribution to the pathogenesis of NPC remains unclear. Interleukin-6 (IL-6), a tumor promoter, is detected in NPC. This in vitro study examined whether and how Rta promotes NPC progression by up-regulating IL-6. MATERIALS AND METHODS: Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR), quantitative real-time PCR, ELISA, immunoblotting assays, reporter gene assays, and transwell migration assays were performed. RESULTS: In NPC cells, Rta up-regulated IL-6 expression at the mRNA and protein levels, and the Rta's C-terminus was essential for promoter activation and expression of IL-6. The induction of IL-6 by Rta also required activation of extracellular signal-regulated kinase 1/2 and activator protein-1. Furthermore, IL-6 secreted from Rta-expressing NPC cells promoted migration of Rta-negative NPC cells by activating IL-6 receptor/Janus kinase/signal transducer and activator of transcription 3 pathway. CONCLUSION: Rta contributes to progression of NPC cells through induction of IL-6 in vitro.


Assuntos
Proteínas Imediatamente Precoces/metabolismo , Interleucina-6/metabolismo , Janus Quinases/metabolismo , Neoplasias/metabolismo , Receptores de Interleucina-6/metabolismo , Fator de Transcrição STAT3/metabolismo , Transativadores/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Neoplasias da Mama/virologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Humanos , Proteínas Imediatamente Precoces/genética , Interleucina-6/biossíntese , Interleucina-6/genética , Sistema de Sinalização das MAP Quinases , Células MCF-7 , Carcinoma Nasofaríngeo/genética , Carcinoma Nasofaríngeo/metabolismo , Carcinoma Nasofaríngeo/patologia , Carcinoma Nasofaríngeo/virologia , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patologia , Neoplasias Nasofaríngeas/virologia , Neoplasias/genética , Neoplasias/patologia , Neoplasias/virologia , Transdução de Sinais , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Neoplasias Gástricas/virologia , Transativadores/genética , Fator de Transcrição AP-1/metabolismo , Transfecção , Regulação para Cima
16.
Am J Hum Genet ; 106(6): 748-763, 2020 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-32442411

RESUMO

The identification of causal variants and mechanisms underlying complex disease traits in humans is important for the progress of human disease genetics; this requires finding strategies to detect functional regulatory variants in disease-relevant cell types. To achieve this, we collected genetic and transcriptomic data from the aortic endothelial cells of up to 157 donors and four epigenomic phenotypes in up to 44 human donors representing individuals of both sexes and three major ancestries. We found thousands of expression quantitative trait loci (eQTLs) at all ranges of effect sizes not detected by the Gene-Tissue Expression Project (GTEx) in human tissues, showing that novel biological relationships unique to endothelial cells (ECs) are enriched in this dataset. Epigenetic profiling enabled discovery of over 3,000 regulatory elements whose activity is modulated by genetic variants that most frequently mutated ETS, AP-1, and NF-kB binding motifs, implicating these motifs as governors of EC regulation. Using CRISPR interference (CRISPRi), allele-specific reporter assays, and chromatin conformation capture, we validated candidate enhancer variants located up to 750 kb from their target genes, VEGFC, FGD6, and KIF26B. Regulatory SNPs identified were enriched in coronary artery disease (CAD) loci, and this result has specific implications for PECAM-1, FES, and AXL. We also found significant roles for EC regulatory variants in modifying the traits pulse pressure, blood protein levels, and monocyte count. Lastly, we present two unlinked SNPs in the promoter of MFAP2 that exhibit pleiotropic effects on human disease traits. Together, this supports the possibility that genetic predisposition for complex disease is manifested through the endothelium.


Assuntos
Doença/genética , Células Endoteliais/metabolismo , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica/genética , Variação Genética/genética , Alelos , Epigênese Genética/genética , Feminino , Humanos , Cinesina/genética , Masculino , Mutação , NF-kappa B/metabolismo , Polimorfismo de Nucleotídeo Único/genética , Proteína Proto-Oncogênica c-ets-1/metabolismo , Locos de Características Quantitativas/genética , Fator de Transcrição AP-1/metabolismo , Regulador Transcricional ERG/metabolismo , Fator C de Crescimento do Endotélio Vascular/genética
17.
Vascul Pharmacol ; 131: 106690, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32407896

RESUMO

Cutaneous cold-induced vasoconstriction is a normal physiological reaction mediated by alpha 2C-adrenergic receptors (α2C-ARs) expressed in vascular smooth muscle cells (VSMCs). When this reaction is exaggerated, Raynaud's phenomenon (RP) ensues. RP is more prevalent in females compared to age-matched men. We previously established that 17-ß estradiol (estrogen) upregulates α2C-ARs in human VSMCs via a cAMP/Epac/Rap pathway. We also showed that cAMP acts through JNK to increase α2C-AR expression. However, whether estrogen employs JNK to regulate α2C-AR is not investigated. Knowing that the α2C-AR promoter harbors an activator protein-1 (AP-1) binding site that can be potentially activated by JNK, we hypothesized that estrogen regulates α2C-AR expression through an Epac/JNK/AP-1 pathway. Our results show that estrogen (10-10 M) activated JNK in human VSMCs extracted from cutaneous arterioles. Pretreatment with ESI09 (10 µM; an Epac inhibitor), abolished estrogen-induced JNK activation. In addition, pre-treatment with SP600125 (3 µM; a JNK specific inhibitor) abolished estrogen-induced expression of α2C-AR. Importantly, estrogen-induced activation of α2C-AR promoter was attenuated with SP600125. Moreover, transient transfection of VSMCs with an Epac dominant negative mutant (Epac-DN) abolished estrogen-induced activation of α2C-AR promoter. However, co-transfection of constitutively active JNK mutant overrode the inhibitory effect of Epac-DN on α2C-AR promoter. Moreover, estrogen caused a concentration-dependent increase in the activity of AP-1-driven reporter construct. Mutation of AP-1 site in the α2C-AR promoter abolished its activation by estrogen. This in vitro estrogen-increased α2C-AR expression was mirrored by an increase in the ex vivo functional responsiveness of arterioles. Indeed, estrogen potentiated α2C-AR-mediated cold-induced vasoconstriction, which was abolished by SP600125. Collectively, these results indicate that estrogen upregulates α2C-AR expression via an EPAC-mediated JNK/AP-1- dependent mechanism. These results provide an insight into the mechanism by which exaggerated cold-induced vasoconstriction occurs in estrogen-replete females and identify Epac and JNK as potential targets for the treatment of RP.


Assuntos
Temperatura Baixa , AMP Cíclico/metabolismo , Estradiol/farmacologia , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Receptores Adrenérgicos alfa 2/metabolismo , Cauda/irrigação sanguínea , Fator de Transcrição AP-1/metabolismo , Vasoconstrição/efeitos dos fármacos , Animais , Arteríolas/efeitos dos fármacos , Arteríolas/enzimologia , Células Cultivadas , Fatores de Troca do Nucleotídeo Guanina/genética , Humanos , Masculino , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/enzimologia , Miócitos de Músculo Liso/enzimologia , Doença de Raynaud/tratamento farmacológico , Doença de Raynaud/enzimologia , Doença de Raynaud/fisiopatologia , Receptores Adrenérgicos alfa 2/genética , Transdução de Sinais , Fator de Transcrição AP-1/genética , Regulação para Cima
18.
Nat Cell Biol ; 22(6): 701-715, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32424275

RESUMO

Acquired therapy resistance is a major problem for anticancer treatment, yet the underlying molecular mechanisms remain unclear. Using an established breast cancer cellular model, we show that endocrine resistance is associated with enhanced phenotypic plasticity, indicated by a general downregulation of luminal/epithelial differentiation markers and upregulation of basal/mesenchymal invasive markers. Consistently, similar gene expression changes are found in clinical breast tumours and patient-derived xenograft samples that are resistant to endocrine therapies. Mechanistically, the differential interactions between oestrogen receptor α and other oncogenic transcription factors, exemplified by GATA3 and AP1, drive global enhancer gain/loss reprogramming, profoundly altering breast cancer transcriptional programs. Our functional studies in multiple culture and xenograft models reveal a coordinated role of GATA3 and AP1 in re-organizing enhancer landscapes and regulating cancer phenotypes. Collectively, our study suggests that differential high-order assemblies of transcription factors on enhancers trigger genome-wide enhancer reprogramming, resulting in transcriptional transitions that promote tumour phenotypic plasticity and therapy resistance.


Assuntos
Adaptação Fisiológica , Neoplasias da Mama/tratamento farmacológico , Reprogramação Celular , Resistencia a Medicamentos Antineoplásicos , Fator de Transcrição GATA3/metabolismo , Regulação Neoplásica da Expressão Gênica , Fator de Transcrição AP-1/metabolismo , Animais , Antineoplásicos Hormonais/farmacologia , Apoptose , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Fator de Transcrição GATA3/genética , Humanos , Camundongos , Camundongos Nus , Tamoxifeno/farmacologia , Fator de Transcrição AP-1/genética , Ativação Transcricional , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
19.
J Biol Chem ; 295(21): 7274-7288, 2020 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-32284328

RESUMO

Hyaluronan (HA) is an extracellular matrix glycosaminoglycan that critically supports the physicochemical and mechanical properties of the skin. Here, we demonstrate that mycosporine-like amino acids (MAAs), which typically function as UV-absorbing compounds, can stimulate HA secretion from normal human fibroblasts. MAA-stimulated HA secretion was associated with significantly increased and decreased levels of mRNAs encoding HA synthase 2 (HAS2) and the HA-binding protein involved in HA depolymerization (designated HYBID), respectively. Using immunoblotting, we found that MAAs at 10 and at 25 µg/ml stimulate the phosphorylation of the mitogen-activated protein kinase (MAPK) p38, extracellular signal-regulated kinase (ERK)/c-Jun, and mitogen- and stress-activated protein kinase 1 (MSK1) (at Thr-581, Ser-360, and Ser-376, respectively) and activation of cAMP-responsive element-binding protein (CREB) and activating transcription factor 2 (ATF2), but not phosphorylation of JUN N-terminal kinase (JNK) or NF-κB (at Ser-276 or Ser-536, respectively), and increased c-Fos protein levels. Moreover, a p38-specific inhibitor, but not inhibitors of MAPK/ERK kinase (MEK), JNK, or NF-κB, significantly abrogated the increased expression of HAS2 mRNA, accompanied by significantly decreased MAA-stimulated HA secretion. These results suggested that the p38-MSK1-CREB-c-Fos-transcription factor AP-1 (AP-1) or the p38-ATF2 signaling cascade is responsible for the MAA-induced stimulation of HAS2 gene expression. Of note, siRNA-mediated ATF2 silencing failed to abrogate MAA-stimulated HAS2 expression, and c-Fos silencing abolished the increased expression of HAS2 mRNA. Our findings suggest that MAAs stimulate HA secretion by up-regulating HAS2 mRNA levels through activation of an intracellular signaling cascade consisting of p38, MSK1, CREB, c-Fos, and AP-1.


Assuntos
Aminoácidos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Hialuronan Sintases/biossíntese , Ácido Hialurônico/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Células Cultivadas , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Humanos , Proteínas Proto-Oncogênicas c-fos/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Fator de Transcrição AP-1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
20.
J Biol Chem ; 295(21): 7431-7441, 2020 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-32312752

RESUMO

Lytic replication of Epstein-Barr virus (EBV) is not only essential for its cell-to-cell spread and host-to-host transmission, but it also contributes to EBV-induced oncogenesis. Thus, blocking EBV lytic replication could be a strategy for managing EBV-associated diseases. Previously, we identified a series of natural lignans isolated from the roots of Saururus chinensis (Asian lizard's tail) that efficiently block EBV lytic replication and virion production with low cytotoxicity. In this study, we attempted to elucidate the molecular mechanism by which these lignans inhibit EBV lytic replication. We found that a representative compound, CSC27 (manassantin B), inhibits EBV lytic replication by suppressing the expression of EBV immediate-early gene BZLF1 via disruption of AP-1 signal transduction. Further analysis revealed that manassantin B specifically blocks the mammalian target of rapamycin complex 2 (mTORC2)-mediated phosphorylation of AKT Ser/Thr protein kinase at Ser-473 and protein kinase Cα (PKCα) at Ser-657. Using phosphoinositide 3-kinase-AKT-specific inhibitors for kinase mapping and shRNA-mediated gene silencing, we validated that manassantin B abrogates EBV lytic replication by inhibiting mTORC2 activity and thereby blocking the mTORC2-PKC/AKT-signaling pathway. These results suggest that mTORC2 may have utility as an antiviral drug target against EBV infections and also reveal that manassantin B has potential therapeutic value for managing cancers that depend on mTORC2 signaling for survival.


Assuntos
Furanos/farmacologia , Herpesvirus Humano 4/fisiologia , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/antagonistas & inibidores , Ativação Viral/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Proteína Quinase C/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Transativadores/metabolismo , Fator de Transcrição AP-1/metabolismo
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