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1.
J Cancer Res Clin Oncol ; 145(12): 2969-2982, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31612319

RESUMO

PURPOSE: Non-canonical NFκB (NC-NFκB) pathway plays an influential role in metastasis, which promotes cancer proliferation and progression. The aim of the study was to examine the expression of NC-NFκB proteins and their correlation with clinicopathological factors associated with metastatic cases of uveal melanoma (UM) and with the patient outcome. METHOD: Expression of NC-NFκB proteins (p52, RelB, and co-expression of p52/RelB) was evaluated in 75 formalin-fixed cases of uveal melanoma by immunohistochemistry. Validation of nuclear immunoreactivity was done by western blotting. Transcriptional status of NC-NFκB genes was assessed in 60 fresh tumor tissues by quantitative real-time PCR. Co-immunoprecipitation was performed to determine the presence of native p52/RelB heterodimer in UM. Prognostic relevance was determined using Cox proportional hazard and Kaplan-Meier methods. RESULTS: Immunohistochemical expression of p52, RelB, and their co-expression was observed in 81%, 68.7%, 56.2% of metastatic cases, respectively, while their expression was seen only in 38%, 33% and 30% of non-metastatic cases. Loss of BAP-1 was correlated with expression of p52 and RelB proteins. Co-immunoprecipitation assay confirmed the putative interaction of p52 with RelB protein in metastatic cases of uveal melanoma. Co-expression of p52/RelB and expression of p52 protein was significantly correlated with decreased metastasis-free survival (MFS) (p = 0.004; p = 0.002) and overall survival (OS) (p = 0.004; p = 0.032), while the RelB expression only correlated with reduced MFS (p = 0.003). CONCLUSION: Our data showed that non-canonical NFκB proteins were significantly higher in metastatic cases and associated with poor outcome of the patients. Furthermore, the p52 protein could be used as a potential therapeutic biomarker for metastatic cases in uveal melanoma.


Assuntos
Regulação Neoplásica da Expressão Gênica/genética , Melanoma/genética , Subunidade p52 de NF-kappa B/genética , Metástase Neoplásica/genética , Fator de Transcrição RelB/genética , Neoplasias Uveais/genética , Adulto , Intervalo Livre de Doença , Feminino , Humanos , Imuno-Histoquímica/métodos , Masculino , Melanoma/patologia , Metástase Neoplásica/patologia , Prognóstico , Estudos Prospectivos , Transcrição Genética/genética , Neoplasias Uveais/patologia
2.
Am J Pathol ; 189(12): 2516-2530, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31539516

RESUMO

NF-κB signals through canonical transcription factor p65 (RelA)/p50 and noncanonical avian reticuloendotheliosis viral oncogene related B (RelB)/p52 pathways. The RelA/p50 is involved in basal and inflammatory lymphangiogenesis. However, the role of RelB/p52 in lymphatic vessel biology is unknown. Herein, we investigated changes in lymphatic vessels (LVs) in mice deficient in noncanonical NF-κB signaling and the function of RelB in lymphatic endothelial cells (LECs). LVs were examined in Relb-/-, p52-/-, or control mice, and the gene expression profiles in LECs with RelB knockdown. Relb-/-, but not p52-/-, mice exhibited multiple LV abnormalities. They include the following: i) increased capillary vessel diameter, ii) reduced smooth muscle cell (SMC) coverage of mature vessels, iii) leakage, and iv) loss of active and passive lymphatic flow. Relb-/- mature LVs had thinner vessel walls, more apoptotic LECs and SMCs, and fewer LEC junctions. RelB knockdown LECs had decreased growth, survival, and adhesion, and dysregulated signaling pathways involving these cellular events. These results suggest that Relb-/- mice have abnormal LVs, mainly in mature vessels with reduced SMC coverage, leakage, and loss of contractions. RelB knockdown in LECs leads to reduced growth, survival, and adhesion. RelB plays a vital role in LEC-mediated LV maturation and function.


Assuntos
Proliferação de Células , Células Endoteliais/patologia , Vasos Linfáticos/patologia , Fator de Transcrição RelB/fisiologia , Animais , Apoptose , Movimento Celular , Células Cultivadas , Células Endoteliais/metabolismo , Vasos Linfáticos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NF-kappa B , Transdução de Sinais
3.
Mol Immunol ; 114: 395-409, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31476634

RESUMO

Inflammation is a response to injury and infection. Although protective under physiological conditions, excessive and persistent inflammation is linked to numerous diseases. As the lungs are continuously exposed to the external environment, the respiratory system is particularly liable to damage from inflammation. RelB is a member of the non-canonical NF-κB pathway that may control lung inflammation caused by cigarette smoke (CS), a leading cause of morbidity and mortality worldwide. Our lab has previously shown that RelB protects against CS-induced inflammation in vitro, leading us to hypothesize that RelB would protect against acute CS-induced pulmonary inflammation in vivo. We exposed wild-type (Relb+/+) and RelB-deficient mice (Relb-/-) mice to room air or to CS and found that CS exposure caused a sustained decrease in pulmonary granulocytes in Relb-/- mice that was predominated by a decrease in neutrophils. Pulmonary inflammation caused by other irritants, including chlorine, ovalbumin (OVA; to mimic features of asthma) and lipopolysaccharide (LPS) was not controlled by RelB. Differential cytokine analysis suggests that alterations in chemotactic cytokines do not fully account for the CS-specific decrease in neutrophils in Relb-/- mice. Flow cytometric analysis of the bronchoalveolar lavage and bone marrow cells also reveal that it is unlikely that the sustained decrease is caused by excessive cell death or decreased hematopoiesis from the bone marrow. Overall, our results indicate that RelB regulates acute CS-induced pulmonary inflammation. Understanding how RelB regulates CS-induced inflammation may potentiate the discovery of new therapeutic strategies for many of the inflammatory diseases caused by CS.


Assuntos
Pulmão/imunologia , NF-kappa B/imunologia , Neutrófilos/imunologia , Pneumonia/imunologia , Fumaça/efeitos adversos , Tabaco/imunologia , Fator de Transcrição RelB/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Citocinas/imunologia , Modelos Animais de Doenças , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Doença Pulmonar Obstrutiva Crônica/imunologia , Transdução de Sinais/imunologia , Fumar/efeitos adversos , Fumar/imunologia , Tabaco/efeitos adversos
4.
J Neuroinflammation ; 16(1): 161, 2019 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-31362762

RESUMO

BACKGROUND: Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system (CNS). It is firmly established that overactivation of the p65 (RelA) nuclear factor kappa B (NF-κB) transcription factor upregulates expression of inflammatory mediators in both immune and non-immune resident CNS cells and promotes inflammation during MS. In contrast to p65, NF-κB family member RelB regulates immune cell development and can limit inflammation. Although RelB expression is induced during inflammation in the CNS, its role in MS remains unknown. METHODS: To examine the role of RelB in non-immune CNS cells, we generated mice with RelB specifically deleted in astrocytes (RelBΔAST), oligodendrocytes (RelBΔOLIGO), or neural progenitor-derived cells (RelBΔNP). We used experimental autoimmune encephalomyelitis (EAE), an accepted mouse model of MS, to assess the effect of RelB deletion on disease outcomes and performed analysis on the histological, cellular, and molecular level. RESULTS: Despite being a negative regulator of inflammation, conditional knockout of RelB in non-immune resident CNS cells surprisingly decreased the severity of EAE. This protective effect was recapitulated by conditional deletion of RelB in oligodendrocytes but not astrocytes. Deletion of RelB in oligodendrocytes reduced disease severity, promoted survival of mature oligodendrocytes, and correlated with increased activation of p65 NF-κB. CONCLUSIONS: These findings suggest that RelB fine tunes inflammation and cell death/survival during EAE. Importantly, our data points out the detrimental role RelB plays in controlling survival of mature oligodendrocytes, which could be explored as a viable option to treat MS in the future.


Assuntos
Encéfalo/metabolismo , Encefalomielite Autoimune Experimental/metabolismo , Oligodendroglia/metabolismo , Fator de Transcrição RelB/metabolismo , Animais , Astrócitos/metabolismo , Encéfalo/patologia , Encefalomielite Autoimune Experimental/patologia , Camundongos , NF-kappa B/metabolismo , Células-Tronco Neurais/metabolismo , Fator de Transcrição RelB/genética
5.
J Biol Chem ; 294(38): 14009-14019, 2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31362988

RESUMO

Lipid phosphate phosphatase 3 (LPP3), encoded by the PLPP3 gene, is an integral membrane enzyme that dephosphorylates phosphate esters of glycero- and sphingophospholipids. Cell surface LPP3 can terminate the signaling actions of bioactive lysophosphatidic acid (LPA) and sphingosine 1 phosphate, which likely explains its role in developmental angiogenesis, vascular injury responses, and cell migration. Heritable variants in the final intron PLPP3 associate with interindividual variability in coronary artery disease risk that may result from disruption of enhancer sequences that normally act in cis to increase expression of the gene. However, the mechanisms regulating PLPP3 expression are not well understood. We show that the human PLPP3 promoter contains three functional NF-κB response elements. All of these are required for maximal induction of PLPP3 promoter activity in reporter assays. The identified sequences recruit RelA and RelB components of the NF-κB transcription complex to chromatin, and these transcription factors bind to the identified target sequences in two different cell types. LPA promotes binding of Rel family transcription factors to the PLPP3 promoter and increases PLPP3 gene expression through mechanisms that are attenuated by an NF-κB inhibitor, LPA receptor antagonists, and inhibitors of phosphoinositide 3 kinase. These findings indicate that up-regulation of PLPP3 during inflammation and atherosclerosis results from canonical activation of the NF-κB signaling cascade to increase PLPP3 expression through nuclear import and binding of RelA and RelB transcription factors to the PLPP3 promoter and suggest a mechanism by which the LPP3 substrate, LPA, can regulate PLPP3 expression.


Assuntos
NF-kappa B/genética , NF-kappa B/metabolismo , Fosfatidato Fosfatase/biossíntese , Fosfatidato Fosfatase/genética , Regulação da Expressão Gênica , Humanos , Proteínas I-kappa B/metabolismo , Lisofosfolipídeos/metabolismo , Subunidade p50 de NF-kappa B/metabolismo , Fosfatidato Fosfatase/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Regiões Promotoras Genéticas , Transdução de Sinais , Esfingolipídeos/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Células THP-1 , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelB/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Genética
6.
Nat Commun ; 10(1): 3589, 2019 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399573

RESUMO

Overcoming cellular growth restriction, including the evasion of cellular senescence, is a hallmark of cancer. We report that PAK4 is overexpressed in all human breast cancer subtypes and associated with poor patient outcome. In mice, MMTV-PAK4 overexpression promotes spontaneous mammary cancer, while PAK4 gene depletion delays MMTV-PyMT driven tumors. Importantly, PAK4 prevents senescence-like growth arrest in breast cancer cells in vitro, in vivo and ex vivo, but is not needed in non-immortalized cells, while PAK4 overexpression in untransformed human mammary epithelial cells abrogates H-RAS-V12-induced senescence. Mechanistically, a PAK4 - RELB - C/EBPß axis controls the senescence-like growth arrest and a PAK4 phosphorylation residue (RELB-Ser151) is critical for RELB-DNA interaction, transcriptional activity and expression of the senescence regulator C/EBPß. These findings establish PAK4 as a promoter of breast cancer that can overcome oncogene-induced senescence and reveal a selective vulnerability of cancer to PAK4 inhibition.


Assuntos
Neoplasias da Mama/patologia , Fator de Transcrição RelB/metabolismo , Quinases Ativadas por p21/metabolismo , Animais , Mama/citologia , Mama/patologia , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Senescência Celular/genética , Células Epiteliais , Feminino , Técnicas de Silenciamento de Genes , Humanos , Glândulas Mamárias Animais/patologia , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Cultura Primária de Células , Prognóstico , Ensaios Antitumorais Modelo de Xenoenxerto , Quinases Ativadas por p21/genética
7.
Nutrients ; 11(7)2019 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-31248019

RESUMO

Intracellular Ca2+ homeostasis is commonly disrupted in acute pancreatitis. Sustained Ca2+ release from internal stores in pancreatic acinar cells (PACs), mediated by inositol triphosphate receptor (IP3R) and the ryanodine receptor (RyR), plays a key role in the initiation and propagation of acute pancreatitis. Pancreatitis induced by cerulein, an analogue of cholecystokinin, causes premature activation of digestive enzymes and enhanced accumulation of cytokines and Ca2+ in the pancreas and, as such, it is a good model of acute pancreatitis. High concentrations of the omega-3 fatty acid docosahexaenoic acid (DHA) inhibit inflammatory signaling pathways and cytokine expression in PACs treated with cerulein. In the present study, we determined the effect of DHA on key regulators of Ca2+ signaling in cerulein-treated pancreatic acinar AR42 J cells. The results of RNA-Sequencing (RNA-Seq) analysis showed that cerulein up-regulates the expression of IP3R1 and RyR2 genes, and that pretreatment with DHA blocks these effects. The results of real-time PCR confirmed that DHA inhibits cerulein-induced IP3R1 and RyR2 gene expression, and demonstrated that DHA pre-treatment decreases the expression of the Relb gene, which encodes a component of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) transcriptional activator complex, and the c-fos gene, which encodes a component of activator protein-1 (AP-1) transcriptional activator complex. Taken together, DHA inhibits mRNA expression of IP3R1, RyR2, Relb, and c-fos, which is related to Ca2+ network in cerulein-stimulated PACs.


Assuntos
Células Acinares/efeitos dos fármacos , Sinalização do Cálcio/efeitos dos fármacos , Ceruletídeo/toxicidade , Ácidos Docosa-Hexaenoicos/farmacologia , Perfilação da Expressão Gênica/métodos , Pâncreas Exócrino/efeitos dos fármacos , Pancreatite/tratamento farmacológico , Análise de Sequência de RNA , Células Acinares/metabolismo , Animais , Sinalização do Cálcio/genética , Linhagem Celular , Regulação da Expressão Gênica , Receptores de Inositol 1,4,5-Trifosfato/genética , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Pâncreas Exócrino/metabolismo , Pancreatite/induzido quimicamente , Pancreatite/genética , Pancreatite/metabolismo , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Fator de Transcrição RelB/genética , Fator de Transcrição RelB/metabolismo
8.
Int J Mol Sci ; 20(11)2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-31151139

RESUMO

Here, we investigate the role of RelB in the regulation of genes which were identified to be induced in an aryl hydrocarbon receptor (AhR)-dependent manner and critically involved in regulation of immune responses. We analyzed the expression of genes of the AhR gene battery, cytokines, and immune regulatory enzymes in bone marrow-derived macrophages (BMM) and thymus of B6 wildtype (wt) mice and RelB knockout (RelB-/-) mice after treatment with various AhR ligands. The 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced expression of indoleamine 2,3-dioxygenase 1 (IDO1) and IDO2 was significantly repressed in thymus of RelB-/- mice but not in BMM derived from RelB-/- mice. Interestingly, the induced and basal expression of the cytokines interleukin (IL)-17A, IL-22, and CCL20 required the functional expression of RelB. The RelB-dependent expression of CCL20 was induced by the AhR ligands TCDD and 6-formylindolo[3,2-b]carbazole (FICZ), whereas indole-3-carbinol (I3C) suppressed CCL20 in lipopolysaccharide (LPS)-activated wt BMM. The LPS-induced expression of IL-6 and IL-10 was enhanced by TCDD and FICZ, whereas I3C significantly suppressed these cytokines in BMM. The exposure to FICZ led to higher increases of IL-17A and IL-22 mRNA compared to the effect of TCDD or I3C in thymus of wt mice. On the other hand, TCDD was the strongest inducer of CYP1A1, AhR Repressor (AhRR), and IDO2. In summary, these findings provide evidence for the important role of RelB in the transcriptional regulation of cytokines and enzymes induced by AhR ligands.


Assuntos
Receptores de Hidrocarboneto Arílico/metabolismo , Fator de Transcrição RelB/metabolismo , Animais , Citocinas/genética , Citocinas/metabolismo , Expressão Gênica , Regulação da Expressão Gênica , Imunomodulação/genética , Ligantes , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Ligação Proteica , Receptores de Hidrocarboneto Arílico/genética , Timo/imunologia , Timo/metabolismo , Fator de Transcrição RelB/genética
9.
Immunobiology ; 224(5): 687-696, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31200979

RESUMO

Regulatory T cells (Tregs) maintain immune homeostasis and play an important role in tissue regeneration after injury. Mutations affecting development or homeostasis of Tregs lead to immune pathologies in humans and are often fatal in mouse models. Although the pathways required for Treg development are being increasingly characterized, factors crucial for Treg homeostasis are not completely understood. Previously we have found a role for alternative NF-κB pathway in restricting T cell activation and Th17 differentiation. Here, by using the mouse model of uncontrolled alternative NF-κB signaling we identify a crucial intrinsic role of RelB signaling in regulating homeostasis and competitive fitness of Tregs. The failure of p100-/- Tregs to maintain the population of effector Tregs and efficiently suppress immune reactions results in lethal multiorgan Th1-mediated inflammation in Rag1-/- recipients. This inflammation is combined with severe lymphopenia and could be rescued by adoptive transfer of wild type Tregs. Thus in addition to its role in Th17 differentiation, RelB acts as a potent inhibitor of Treg effector functions. Our results point to RelB as a potential therapeutic target for Treg manipulation.


Assuntos
Homeostase , NF-kappa B/metabolismo , Transdução de Sinais , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Animais , Autoimunidade , Biomarcadores , Citocinas/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Imuno-Histoquímica , Imunomodulação/genética , Imunofenotipagem , Ativação Linfocitária , Camundongos , Camundongos Knockout , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Fator de Transcrição RelB/metabolismo , Proteína p120 Ativadora de GTPase/genética , Proteína p120 Ativadora de GTPase/metabolismo
10.
J BUON ; 24(2): 720-728, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31128029

RESUMO

PURPOSE: The purpose of this study was to find out the activity and molecular mechanism of NF-kB subunits in cervical cancer which in turn were used as a molecular marker in diagnosing cancer progression. METHODS: Different cervical cancer biopsies were obtained from patients after obtaining proper written consent and approval, which were subjected to immunohistochemistry. Performed were Western blot analysis and electrophoretic mobility shift assays. RESULTS: Immunohistochemical analysis of low-grade, high-grade and squamous cell carcinoma (SCC) (stage IIIA, IIIB and IV) showed low to high nuclear expression of p52/RelB compared with p50/RelA, whereas in normal cells, c-Rel was expressed in the cytosol. p52/RelB expression was further validated by Western blot analysis. The binding ability of NF-κB to p52/RelB was increased during the progression of cervical cancer. All cervical carcinoma biopsies showed increased expression of p50/RelA and p52/RelB, but the p52/RelB NF-κB protein complex showed elevated nuclear expression and binding ability, indicating a pathway other than the classical pathway. The non-canonical NF-κB pathway also played an important role in cervical cancer progression by activating the p52/RelB NF-κB complex. CONCLUSIONS: This study provides a new approach for diagnosing, and establishing an appropriate treatment against cervical cancer progression.


Assuntos
Carcinoma de Células Escamosas/genética , NF-kappa B/genética , Fator de Transcrição RelA/genética , Fator de Transcrição RelB/genética , Neoplasias do Colo do Útero/genética , Transporte Ativo do Núcleo Celular , Adulto , Idoso , Biópsia , Carcinoma de Células Escamosas/patologia , Núcleo Celular/genética , Citoplasma/genética , Proteínas de Ligação a DNA/genética , Detecção Precoce de Câncer , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-rel/genética , Neoplasias do Colo do Útero/patologia
11.
Proc Natl Acad Sci U S A ; 116(21): 10592-10597, 2019 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-31048505

RESUMO

Signaling pathways often share molecular components, tying the activity of one pathway to the functioning of another. In the NFκB signaling system, distinct kinases mediate inflammatory and developmental signaling via RelA and RelB, respectively. Although the substrates of the developmental, so-called noncanonical, pathway are induced by inflammatory/canonical signaling, crosstalk is limited. Through dynamical systems modeling, we identified the underlying regulatory mechanism. We found that as the substrate of the noncanonical kinase NIK, the nfkb2 gene product p100, transitions from a monomer to a multimeric complex, it may compete with and inhibit p100 processing to the active p52. Although multimeric complexes of p100 (IκBδ) are known to inhibit preexisting RelA:p50 through sequestration, here we report that p100 complexes can inhibit the enzymatic formation of RelB:p52. We show that the dose-response systems properties of this complex substrate competition motif are poorly accounted for by standard Michaelis-Menten kinetics, but require more detailed mass action formulations. In sum, although tonic inflammatory signaling is required for adequate expression of the noncanonical pathway precursors, the substrate complex competition motif identified here can prevent amplification of the active RelB:p52 dimer in elevated inflammatory conditions to ensure reliable RelB-dependent developmental signaling independent of inflammatory context.


Assuntos
Modelos Químicos , Transdução de Sinais , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelB/metabolismo
12.
J Immunol Res ; 2019: 7026067, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30949517

RESUMO

Aim: The RelB gene plays an important role in guiding the progression of arthritis. We have previously demonstrated that the expression of the RelB gene is decreased significantly in bone marrow DCs of CD38-/- mice. In this study, we demonstrate that the cluster of the differentiation (CD38) gene could be a potentially therapeutic target for autoimmune arthritis. Method: Collagen-induced arthritis (CIA) models were generated with both the wild-type (WT) C57BL/6 and CD38-/- mice. The expression of the RelB gene and maturation of bone marrow-derived dendritic cells (DCs) from the WT and CD38-/- mice were detected. Antigen-specific T cell responses, joint damage, and expression of proinflammatory cytokines were assessed. The effects of the Nuclear Factor Kappa B (NF-κB) transcription factor and its mechanisms were characterized. Results: We demonstrated that in CD38-/- mice, the expression of the RelB gene and major histocompatibility complex II (MHC II) was decreased, accompanied with the inhibited T cell reaction in a mixed lymphocyte reaction (MLR) in bone marrow-derived DCs. Compared to the serious degeneration of the cartilage and the enlarged gap of the cavum articular in WT CIA mice, joint pathological changes of the CD38-/- CIA mice revealed marked attenuation, while the joint structures were well preserved. The preserved effects were observed by the inhibition of proinflammatory cytokines and promotion of anti-inflammatory cytokines. Furthermore, decreased phosphorylation of NF-κB was also observed in CD38-/- CIA mice. Conclusion: We demonstrate that CD38 could regulate CIA through NF-κB and this regulatory molecule could be a novel target for the treatment of autoimmune inflammatory joint disease.


Assuntos
ADP-Ribosil Ciclase 1/genética , Artrite Experimental/fisiopatologia , Glicoproteínas de Membrana/genética , NF-kappa B/metabolismo , Transdução de Sinais , ADP-Ribosil Ciclase 1/imunologia , Animais , Artrite Experimental/induzido quimicamente , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/imunologia , Diferenciação Celular , Colágeno , Citocinas/imunologia , Regulação para Baixo , Genes MHC da Classe II , Masculino , Glicoproteínas de Membrana/imunologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , NF-kappa B/genética , Fosforilação , Linfócitos T/imunologia , Fator de Transcrição RelB/genética , Fator de Transcrição RelB/imunologia
13.
J Cancer Res Clin Oncol ; 145(6): 1437-1448, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30941572

RESUMO

PURPOSE: Despite considerable evidence that supports the NF-kB role in the immune system and lymphomagenesis, it is unclear whether specific NF-kB dimers control a particular set of genes that account for their biological functions. Our previous work showed that Hodgkin Lymphoma (HL) is unique, among germinal center (GC)-derived lymphomas, with respect to its dependency on Rel-B to survive. In contrast, diffuse large B-Cell lymphoma (DLBCL) including both Activated B-Cell-Like and Germinal Center B-Cell-Like, requires cREL and Rel-A to survive and it is not affected by Rel-B depletion. These findings highlighted the activity of specific NF-kB subunits in different GC-derived lymphomas. METHODS: Sequenced chromatin immunoprecipitated DNA fragments (ChIP-Seq) analysis revealed an extensive NF-kB DNA-binding network in DLBCL and HL. The ChIP-Seq data was merged with microarray analysis following the Rel-A, Rel-B or cRel knockdown to determine effectively regulated genes. RESULTS: Downstream target analysis showed enrichment for cell cycle control, among other signatures. Rel-B and cRel controlled different genes within the same signature in HL and DLBCL, respectively. BCL2 was exclusively controlled by Rel-B in HL. Both mRNA and protein levels decreased following Rel-B depletion meanwhile there was no change upon cRel knock-down. BCL2 exogenous expression partially rescued the death induced by decreased Rel-B in HL cells. CONCLUSION: The Rel-B hierarchical network defined HL and the cRel hierarchical network characterized DLBCL. Each Rel member performs specific functions in distinct GC-derived lymphomas. This result should be considered for the development of targeted therapies that are aimed to selectively inhibit individual NF-kB dimers.


Assuntos
DNA de Neoplasias/metabolismo , Doença de Hodgkin/metabolismo , Linfoma Difuso de Grandes Células B/metabolismo , NF-kappa B/metabolismo , Apoptose/genética , Linfócitos B/metabolismo , Linfócitos B/patologia , Linhagem Celular Tumoral , DNA de Neoplasias/genética , Perfilação da Expressão Gênica , Células HEK293 , Doença de Hodgkin/genética , Humanos , Linfoma Difuso de Grandes Células B/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-rel/genética , Proteínas Proto-Oncogênicas c-rel/metabolismo , Transdução de Sinais , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelB/genética , Fator de Transcrição RelB/metabolismo , Transcrição Genética
14.
Glia ; 67(8): 1449-1461, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30957303

RESUMO

In response to brain injury or infections, astrocytes become reactive, undergo striking morphological and functional changes, and secrete and respond to a spectrum of inflammatory mediators. We asked whether reactive astrocytes also display adaptive responses during sterile IL-1ß-induced neuroinflammation, which may limit tissue injury associated with many disorders of the central nervous system. We found that astrocytes display days-to-weeks long specific tolerance of cytokine genes, which is coordinated by NF-κB family member, RelB. However, in contrast to innate immune cells, astrocytic tolerance does not involve epigenetic silencing of the cytokine genes. Establishment of tolerance depends on persistent higher levels of RelB in tolerant astrocytes and its phosphorylation on serine 472. Mechanistically, this phosphorylation prevents efficient removal of RelB from cytokine promoters by IκBα and helps to establish tolerance. Importantly, ablation of RelB from astrocytes in mice abolishes tolerance during experimental neuroinflammation in vivo.


Assuntos
Imunidade Adaptativa/fisiologia , Astrócitos/imunologia , Inflamação/metabolismo , Fator de Transcrição RelB/metabolismo , Animais , Encéfalo/imunologia , Citocinas/metabolismo , Epigênese Genética , Células HEK293 , Humanos , Tolerância Imunológica/fisiologia , Camundongos Transgênicos , Neuroimunomodulação , Fosforilação , Sirtuína 1/metabolismo , Fator de Transcrição RelB/genética
15.
Nature ; 568(7751): 249-253, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30894749

RESUMO

The non-canonical NF-κB signalling cascade is essential for lymphoid organogenesis, B cell maturation, osteoclast differentiation, and inflammation in mammals1,2; dysfunction of this system is associated with human diseases, including immunological disorders and cancer3-6. Although expression of NF-κB-inducing kinase (NIK, also known as MAP3K14) is the rate-limiting step in non-canonical NF-κB pathway activation2,7, the mechanisms by which transcriptional responses are regulated remain largely unknown. Here we show that the sine oculis homeobox (SIX) homologue family transcription factors SIX1 and SIX2 are integral components of the non-canonical NF-κB signalling cascade. The developmentally silenced SIX proteins are reactivated in differentiated macrophages by NIK-mediated suppression of the ubiquitin proteasome pathway. Consequently, SIX1 and SIX2 target a subset of inflammatory gene promoters and directly inhibit the trans-activation function of the transcription factors RELA and RELB in a negative feedback circuit. In support of a physiologically pivotal role for SIX proteins in host immunity, a human SIX1 transgene suppressed inflammation and promoted the recovery of mice from endotoxic shock. In addition, SIX1 and SIX2 protected RAS/P53-driven non-small-cell lung carcinomas from inflammatory cell death induced by SMAC-mimetic chemotherapeutic agents (small-molecule activators of the non-canonical NF-κB pathway). Our findings identify a NIK-SIX signalling axis that fine-tunes inflammatory gene expression programs under both physiological and pathological conditions.


Assuntos
Proteínas de Homeodomínio/metabolismo , Inflamação/metabolismo , NF-kappa B/deficiência , NF-kappa B/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Animais , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Feminino , Fibroblastos , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Proteínas de Homeodomínio/imunologia , Humanos , Inflamação/genética , Listeria monocytogenes/imunologia , Masculino , Camundongos , NF-kappa B/genética , Proteínas do Tecido Nervoso/imunologia , Regiões Promotoras Genéticas , Shigella flexneri/imunologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelB/metabolismo
16.
Front Immunol ; 10: 173, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30800127

RESUMO

The multistep differentiation process from hematopoietic stem cells through common myeloid progenitors into committed dendritic cell (DC) subsets remains to be fully addressed. These studies now show that Allograft Inflammatory Factor-1 (AIF1) is required for differentiation of classical DC type 1 (cDC1) subsets and monocyte-derived DC (Mo-DC). Phenotypic studies found that AIF1 expression increased in committed subsets differentiating from common myeloid progenitors (CMP). However, silencing AIF1 expression in hematopoietic stem progenitors restrained the capacity to differentiate into Mo-DC and cDC1 cell subsets under GM-CSF or Flt3-L stimuli conditions, respectively. This was further marked by restrained expression of IRF8, which is critical for development of Mo-DC and cDC1 subsets. As a result, absence of AIF1 restrained the cells at the Lin-CD117+FcγR-CD34+ CMP stage. Further biochemical studies revealed that abrogating AIF1 resulted in inhibition of the NFκB family member RelB expression and p38 MAPK phosphorylation during differentiation of Mo-DC. Lastly, protein binding studies identified that AIF1 interacts with protein kinase C (PKC) to influence downstream signaling pathways. Taken together, this is the first report showing a novel role of AIF1 as a calcium-responsive scaffold protein that supports IRF8 expression and interacts with PKC to drive NFκB-related RelB for successfully differentiating hematopoietic progenitor cells into cDC and Mo-DC subsets under Flt3-L and GM-CSF stimuli, respectively.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Diferenciação Celular/fisiologia , Células Dendríticas/citologia , Células-Tronco Hematopoéticas/fisiologia , Fatores Reguladores de Interferon/metabolismo , Proteínas dos Microfilamentos/metabolismo , Monócitos/citologia , Fator de Transcrição RelB/metabolismo , Animais , Células da Medula Óssea/citologia , Proteínas de Ligação ao Cálcio/genética , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Feminino , Técnicas de Silenciamento de Genes , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Hematopoese/efeitos dos fármacos , Masculino , Proteínas de Membrana/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas dos Microfilamentos/genética , Subunidade p50 de NF-kappa B/metabolismo , Proteína Quinase C/metabolismo , RNA Interferente Pequeno/genética , Transfecção , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
17.
PLoS One ; 14(2): e0211746, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30716099

RESUMO

Antiretroviral therapy (ART) suppresses HIV replication, but does not cure the infection because replication-competent virus persists within latently infected CD4+ T cells throughout years of therapy. These reservoirs contain integrated HIV-1 genomes and can resupply active virus. Thus, the development of strategies to eliminate the reservoir of latently infected cells is a research priority of global significance. In this study, we tested efficacy of a new inhibitor of apoptosis protein antagonist (IAPa) called Debio 1143 at reversing HIV latency and investigated its mechanisms of action. Debio 1143 activates HIV transcription via NF-kB signaling by degrading the ubiquitin ligase baculoviral IAP repeat-containing 2 (BIRC2), a repressor of the non-canonical NF-kB pathway. Debio 1143-induced BIRC2 degradation results in the accumulation of NF-κB-inducing kinase (NIK) and proteolytic cleavage of p100 into p52, leading to nuclear translocation of p52 and RELB. Debio 1143 greatly enhances the binding of RELB to the HIV-1 LTR. These data indicate that Debio 1143 activates the non-canonical NF-kB signaling pathway by promoting the binding of RELB:p52 complexes to the HIV-1 LTR, resulting in the activation of the LTR-dependent HIV-1 transcription. Importantly, Debio 1143 reverses viral latency in HIV-1 latent T cell lines. Using knockdown (siRNA BIRC2), knockout (CRIPSR NIK) and proteasome machinery neutralization (MG132) approaches, we found that Debio 1143-mediated HIV latency reversal is BIRC2 degradation- and NIK stabilization-dependent. Debio 1143 also reverses HIV-1 latency in resting CD4+ T cells derived from ART-treated patients or HIV-1-infected humanized mice under ART. Interestingly, daily oral administration of Debio 1143 in cancer patients at well-tolerated doses elicited BIRC2 target engagement in PBMCs and induced a moderate increase in cytokines and chemokines mechanistically related to NF-kB signaling. In conclusion, we provide strong evidences that the IAPa Debio 1143, by initially activating the non-canonical NF-kB signaling and subsequently reactivating HIV-1 transcription, represents a new attractive viral latency reversal agent (LRA).


Assuntos
Fármacos Anti-HIV/farmacologia , Azocinas/farmacologia , Compostos Benzidrílicos/farmacologia , Linfócitos T CD4-Positivos/metabolismo , HIV-1/fisiologia , Proteínas Inibidoras de Apoptose/antagonistas & inibidores , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Latência Viral/efeitos dos fármacos , Animais , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Feminino , Humanos , Proteínas Inibidoras de Apoptose/genética , Proteínas Inibidoras de Apoptose/metabolismo , Camundongos , Subunidade p52 de NF-kappa B/genética , Subunidade p52 de NF-kappa B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fator de Transcrição RelB/genética , Fator de Transcrição RelB/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
18.
Placenta ; 76: 40-50, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30642670

RESUMO

BACKGROUND: Chromatin alterations are important mediators of gene expression changes. We have recently shown that activated non-canonical NF-κB signaling (RelB/p52) recruits histone acetyltransferase CBP and deacetylase HDAC1 to selectively acetylate H3K9 (H3K9ac) to induce expression of corticotropin-releasing hormone (CRH) and prostaglandin-endoperoxide synthase-2 (PTGS2) in the human placenta. Both of these genes play a role in initiating parturition in human pregnancy. METHODS: We performed chromatin immunoprecipitation followed by gene sequencing (ChIP-seq) in primary term human cytotrophoblast (CTB) with use of antibodies to RelB, CBP, HDAC1 and H3K9ac. We further associated these chromatin alterations with gene expression changes from mid-trimester to term in CTB by RNA sequencing (RNA-seq). RESULTS: We detected a genome-wide differential gene enrichment between mid-trimester and term human placenta. Pathway analysis identified that cytokine-cytokine receptor interaction, NF-κB, and TNF are the leading pathways enriched in term placenta and associated with these chromatin alterations. DISCUSSIONS: Our analysis has provided the first-time characterization of the key players of human placental origin with molecular changes resulting from chromatin modifications, which could drive human labor.


Assuntos
Histona Desacetilase 1/metabolismo , Parto/metabolismo , Fator de Transcrição RelB/metabolismo , Trofoblastos/metabolismo , Sequenciamento de Cromatina por Imunoprecipitação , Feminino , Histona Acetiltransferases/metabolismo , Humanos , Gravidez , Análise de Sequência de RNA , Transdução de Sinais
19.
Mucosal Immunol ; 12(1): 277-289, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30327534

RESUMO

Cyclic dinucleotides (CDNs), including cyclic di-GMP (CDG), are promising vaccine adjuvants in preclinical/clinical trials. The in vivo mechanisms of CDNs are not clear. Here we investigated the roles of lung DC subsets in promoting CDG mucosal adjuvant responses in vivo. Using genetically modified mice and adoptive cell transfer, we identified lung conventional DC 2 (cDC2) as the central player in CDG mucosal responses. We further identified two functionally distinct lung cDC2 subpopulations: TNFR2+pRelB+ and TNFR2-pRelB- cDC2. The TNFR2+ cDC2 were mature and migratory upon intranasal CDG administration while the TNFR2- cDC2 were activated but not mature. Adoptive cell transfer showed that TNFR2- cDC2 mediate the antibody responses of CDG, while the TNFR2+ cDC2 generate Th1/17 responses. Mechanistically, immature TNFR2- cDC2 activate monocyte-derived DCs (moDCs), which do not take up intranasally administered CDG. moDCs promote CDG-induced generation of T follicular helper- and germinal center B cells in the lungs. Our data revealed a previously undescribed in vivo mode of DCs action, whereby an immature lung TNFR2- cDC2 subpopulation directs the non-migratory moDCs to generate CDG mucosal responses in the lung.


Assuntos
GMP Cíclico/análogos & derivados , Células Dendríticas/fisiologia , Pulmão/imunologia , Membrana Mucosa/imunologia , Receptores Tipo II do Fator de Necrose Tumoral/genética , Células Th1/imunologia , Células Th17/imunologia , Adjuvantes Imunológicos , Transferência Adotiva , Animais , Diferenciação Celular , Células Cultivadas , GMP Cíclico/genética , Citocinas/metabolismo , Ativação Linfocitária , Camundongos , Monócitos/fisiologia , Fator de Transcrição RelB/metabolismo
20.
Mol Carcinog ; 58(3): 411-425, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30488488

RESUMO

Head and neck squamous cell carcinomas (HNSCC) preferentially spread to regional cervical tissues and lymph nodes. Here, we hypothesized that lymphotoxin-ß (LTß), receptor LTßR, and NF-κB-inducing kinase (NIK), promote the aberrant activation of alternative NF-κB2/RELB pathway and genes, that enhance migration and invasion of HNSCC. Genomic and expression alterations of the alternative NF-kB pathway were examined in 279 HNSCC tumors from The Cancer Genome Atlas (TCGA) and a panel of HNSCC lines. LTßR is amplified or overexpressed in HNSCC of the larynx or oral cavity, while LTß, NIK, and RELB are overexpressed in cancers arising within lymphoid oropharyngeal and tonsillar sites. Similarly, subsets of HNSCC lines displayed overexpression of LTßR, NIK, and RELB proteins. Recombinant LTß, and siRNA depletion of endogenous LTßR and NIK, modulated expression of LTßR, NIK, and nuclear translocation of NF-κB2(p52)/RELB as well as functional NF-κB promoter reporter activity. Treatment with a NIK inhibitor (1,3[2H,4H]-Iso-Quinoline Dione) reduced the protein expression of NIK and NF-κB2(p52)/RELB, and blocked LTß induced nuclear translocation of RELB. NIK and RELB siRNA knockdown or NIK inhibitor slowed HNSCC migration or invation in vitro. LTß-induces expression of migration and metastasis related genes, including hepatocyte growth/scatter factor receptor MET. Knockdown of NIK or MET similarly inhibited the migration of HNSCC cell lines. This may help explain why HNSCC preferentially migrate to local lymph nodes, where LTß is expressed. Our findings show that LTß/LTßR promotes activation of the alternative NIK-NF-κB2/RELB pathway to enhance MET-mediated cell migration in HNSCC, which could be potential therapeutic targets in HNSCC.


Assuntos
Carcinoma de Células Escamosas/secundário , Neoplasias de Cabeça e Pescoço/patologia , Receptor beta de Linfotoxina/metabolismo , Linfotoxina-alfa/metabolismo , Linfotoxina-beta/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Fator de Transcrição RelB/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Receptor beta de Linfotoxina/genética , Linfotoxina-alfa/genética , Linfotoxina-beta/genética , Prognóstico , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais , Fator de Transcrição RelB/genética , Células Tumorais Cultivadas
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