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1.
Exp Dermatol ; 31(6): 962-969, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35297512

RESUMO

Interleukin (IL)-12 and IL-23 are pro-inflammatory cytokines produced by dendritic cells (DCs) and associated with Psoriasis (Pso) and Psoriatic Arthritis (PsA) pathogenesis. Tofacitinib, a Janus kinase inhibitor, effectively suppresses inflammatory cascades downstream the IL-12/IL-23 axis in Pso and PsA patients. Here, we investigated whether Tofacitinib directly regulates IL-12/IL-23 production in DCs, and how this regulation reflects responses to Tofacitinib in Pso patients. We treated monocyte-derived dendritic cells and myeloid dendritic cells with Tofacitinib and stimulated cells with either lipopolysaccharide (LPS) or a combination of LPS and IFN-γ. We assessed gene expression by qPCR, obtained skin microarray and blood Olink data and clinical parameters of Pso patients treated with Tofacitinib from public data sets. Our results indicate that in DCs co-stimulated with LPS and IFN-γ, but not with LPS alone, Tofacitinib leads to the decreased expression of IL-23/IL-12 shared subunit IL12B (p40). In Tofacitinib-treated Pso patients, IL-12 expression and psoriasis area and severity index (PASI) are significantly reduced in patients with higher IFN-γ at baseline. These findings demonstrate for the first time that Tofacitinib suppresses IL-23/IL-12 shared subunit IL12B in DCs upon active IFN-γ signaling, and that Pso patients with higher IFN-γ baseline levels display improved clinical response after Tofacitinib treatment.


Assuntos
Interferon gama , Subunidade p40 da Interleucina-12 , Inibidores de Janus Quinases , Piperidinas , Psoríase , Pirimidinas , Pele , Artrite Psoriásica/tratamento farmacológico , Células Dendríticas/imunologia , Humanos , Interferon gama/metabolismo , Subunidade p40 da Interleucina-12/antagonistas & inibidores , Subunidade p40 da Interleucina-12/sangue , Subunidade p40 da Interleucina-12/metabolismo , Inibidores de Janus Quinases/farmacologia , Inibidores de Janus Quinases/uso terapêutico , Lipopolissacarídeos/imunologia , Piperidinas/farmacologia , Piperidinas/uso terapêutico , Psoríase/tratamento farmacológico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Pele/efeitos dos fármacos , Pele/imunologia
2.
J Autoimmun ; 111: 102444, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32284212

RESUMO

OBJECTIVE: To analyze how monocyte and macrophage exposure to CXCL4 induces inflammatory and fibrotic processes observed in Systemic sclerosis (SSc) patients. METHODS: In six independent experiments, monocytes of healthy controls (HC) and SSc patients were stimulated with CXCL4, TLR-ligands, IFNɑ or TGFß and the secretion of cytokines in the supernatant was assessed by multiplex immunoassays. PDGF-BB production by monocyte-derived macrophages was quantified using immunoassays. The number of monocytes and PDGF-BB in circulation was quantified in HC and SSc patients with the Sysmex XT-1800i haematology counter and immunoassays. Intracellular PDGF-BB was quantified in monocytes by Western blot. PDGF-receptor inhibition was achieved using siRNA-mediated knockdown or treatment with Crenolanib. The production of inflammatory mediators and extracellular matrix (ECM) components by dermal fibroblasts was analyzed by qPCR, ELISA and ECM deposition assays. RESULTS: SSc and HC monocytes released PDGF-BB upon stimulation with CXCL4. Conversely, TLR ligands, IFNɑ or TGFß did not induce PDGF-bb release. PDGF-BB plasma levels were significantly (P = 0.009) higher in diffuse SSc patients (n = 19), compared with HC (n = 21). In healthy dermal fibroblasts, PDGF-BB enhanced TNFɑ-induced expression of inflammatory cytokines and increased ECM production. Comparable results were observed in fibroblasts cultured in supernatant taken from macrophages stimulated with CXCL4. This effect was almost completely abrogated by inhibition of the PDGF-receptor using Crenolanib. CONCLUSION: Our findings demonstrate that CXCL4 can drive fibroblast activation indirectly via PDGF-BB production by myeloid cells. Hence, targeting PDGF-BB or CXCL4-induced PDGF-BB release could be clinically beneficial for patients with SSc.


Assuntos
Becaplermina/metabolismo , Fibroblastos/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Fator Plaquetário 4/metabolismo , Escleroderma Sistêmico/imunologia , Adulto , Idoso , Benzimidazóis/farmacologia , Células Cultivadas , Citocinas/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Piperidinas/farmacologia , Receptores do Fator de Crescimento Derivado de Plaquetas/antagonistas & inibidores
3.
Ann Rheum Dis ; 78(4): 529-538, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30793699

RESUMO

BACKGROUND AND OBJECTIVE: Systemic sclerosis (SSc) is a severe autoimmune disease, in which the pathogenesis is dependent on both genetic and epigenetic factors. Altered gene expression in SSc monocytes, particularly of interferon (IFN)-responsive genes, suggests their involvement in SSc development. We investigated the correlation between epigenetic histone marks and gene expression in SSc monocytes. METHODS: Chromatin immunoprecipitation followed by sequencing (ChIPseq) for histone marks H3K4me3 and H3K27ac was performed on monocytes of nine healthy controls and 14 patients with SSc. RNA sequencing was performed in parallel to identify aberrantly expressed genes and their correlation with the levels of H3K4me3 and H3K27ac located nearby their transcription start sites. ChIP-qPCR assays were used to verify the role of bromodomain proteins, H3K27ac and STATs on IFN-responsive gene expression. RESULTS: 1046 and 534 genomic loci showed aberrant H3K4me3 and H3K27ac marks, respectively, in SSc monocytes. The expression of 381 genes was directly and significantly proportional to the levels of such chromatin marks present near their transcription start site. Genes correlated to altered histone marks were enriched for immune, IFN and antiviral pathways and presented with recurrent binding sites for IRF and STAT transcription factors at their promoters. IFNα induced the binding of STAT1 and STAT2 at the promoter of two of these genes, while blocking acetylation readers using the bromodomain BET family inhibitor JQ1 suppressed their expression. CONCLUSION: SSc monocytes have altered chromatin marks correlating with their IFN signature. Enzymes modulating these reversible marks may provide interesting therapeutic targets to restore monocyte homeostasis to treat or even prevent SSc.


Assuntos
Epigênese Genética , Código das Histonas/genética , Monócitos/imunologia , Escleroderma Sistêmico/genética , Adulto , Idoso , Azepinas/farmacologia , Estudos de Casos e Controles , Montagem e Desmontagem da Cromatina/genética , Montagem e Desmontagem da Cromatina/imunologia , Feminino , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Histonas/genética , Humanos , Interferon-alfa/imunologia , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular/métodos , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/metabolismo , Escleroderma Sistêmico/imunologia , Triazóis/farmacologia
4.
J Autoimmun ; 89: 162-170, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29371048

RESUMO

OBJECTIVE: MicroRNAs (miRNAs) are regulatory molecules, which have been addressed as potential biomarkers and therapeutic targets in rheumatic diseases. Here, we investigated the miRNA signature in the serum of systemic sclerosis (SSc) patients and we further assessed their expression in early stages of the disease. METHODS: The levels of 758 miRNAs were evaluated in the serum of 26 SSc patients as compared to 9 healthy controls by using an Openarray platform. Three miRNAs were examined in an additional cohort of 107 SSc patients and 24 healthy donors by single qPCR. MiR-483-5p expression was further analysed in the serum of patients with localized scleroderma (LoS) (n = 22), systemic lupus erythematosus (SLE) (n = 33) and primary Sjögren's syndrome (pSS) (n = 23). The function of miR-483-5p was examined by transfecting miR-483-5p into primary human dermal fibroblasts and pulmonary endothelial cells. RESULTS: 30 miRNAs were significantly increased in patients with SSc. Of these, miR-483-5p showed reproducibly higher levels in an independent SSc cohort and was also elevated in patients with preclinical-SSc symptoms (early SSc). Notably, miR-483-5p was not differentially expressed in patients with SLE or pSS, whereas it was up-regulated in LoS, indicating that this miRNA could be involved in the development of skin fibrosis. Consistently, miR-483-5p overexpression in fibroblasts and endothelial cells modulated the expression of fibrosis-related genes. CONCLUSIONS: Our findings showed that miR-483-5p is up-regulated in the serum of SSc patients, from the early stages of the disease onwards, and indicated its potential function as a fine regulator of fibrosis in SSc.


Assuntos
Células Endoteliais/fisiologia , Fibroblastos/fisiologia , MicroRNAs/genética , Escleroderma Sistêmico/genética , Pele/patologia , Adulto , Idoso , Estudos de Coortes , Feminino , Fibrose , Testes Genéticos , Humanos , Masculino , Pessoa de Meia-Idade , Regulação para Cima
5.
Ann Rheum Dis ; 76(1): 277-285, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27457515

RESUMO

OBJECTIVES: Non-selective histone deacetylase (HDAC) inhibitors (HDACi) have demonstrated anti-inflammatory properties in both in vitro and in vivo models of rheumatoid arthritis (RA). Here, we investigated the potential contribution of specific class I and class IIb HDACs to inflammatory gene expression in RA fibroblast-like synoviocytes (FLS). METHODS: RA FLS were incubated with pan-HDACi (ITF2357, givinostat) or selective HDAC1/2i, HDAC3/6i, HDAC6i and HDAC8i. Alternatively, FLS were transfected with HDAC3, HDAC6 or interferon (IFN)-α/ß receptor alpha chain (IFNAR1) siRNA. mRNA expression of interleukin (IL)-1ß-inducible genes was measured by quantitative PCR (qPCR) array and signalling pathway activation by immunoblotting and DNA-binding assays. RESULTS: HDAC3/6i, but not HDAC1/2i and HDAC8i, significantly suppressed the majority of IL-1ß-inducible genes targeted by pan-HDACi in RA FLS. Silencing of HDAC3 expression reproduced the effects of HDAC3/6i on gene regulation, contrary to HDAC6-specific inhibition and HDAC6 silencing. Screening of the candidate signal transducers and activators of transcription (STAT)1 transcription factor revealed that HDAC3/6i abrogated STAT1 Tyr701 phosphorylation and DNA binding, but did not affect STAT1 acetylation. HDAC3 activity was required for type I IFN production and subsequent STAT1 activation in FLS. Suppression of type I IFN release by HDAC3/6i resulted in reduced expression of a subset of IFN-dependent genes, including the chemokines CXCL9 and CXCL11. CONCLUSIONS: Inhibition of HDAC3 in RA FLS largely recapitulates the effects of pan-HDACi in suppressing inflammatory gene expression, including type I IFN production in RA FLS. Our results identify HDAC3 as a potential therapeutic target in the treatment of RA and type I IFN-driven autoimmune diseases.


Assuntos
Artrite Reumatoide/metabolismo , Fibroblastos/metabolismo , Histona Desacetilases/fisiologia , Mediadores da Inflamação/metabolismo , Sinoviócitos/metabolismo , Acetilação , Adulto , Idoso , Artrite Reumatoide/genética , Artrite Reumatoide/imunologia , Células Cultivadas , Regulação para Baixo/fisiologia , Feminino , Regulação da Expressão Gênica/imunologia , Regulação da Expressão Gênica/fisiologia , Histona Desacetilases/genética , Humanos , Interferon beta/biossíntese , Interleucina-1beta/biossíntese , Interleucina-1beta/genética , Masculino , Pessoa de Meia-Idade , Fosforilação , Fator de Transcrição STAT1/metabolismo , Membrana Sinovial/metabolismo , Sinoviócitos/imunologia
6.
Ann Rheum Dis ; 75(2): 430-8, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25452308

RESUMO

OBJECTIVES: Epigenetic modifications play an important role in the regulation of gene transcription and cellular function. Here, we examined if pro-inflammatory factors present in the inflamed joint of patients with rheumatoid arthritis (RA) could regulate histone deacetylase (HDAC) expression and function in fibroblast-like synoviocytes (FLS). METHODS: Protein acetylation in synovial tissue was assessed by immunohistochemistry. The mRNA levels of HDAC family members and inflammatory mediators in the synovial tissue and the changes in HDAC expression in RA FLS were measured by quantitative (q) PCR. FLS were either transfected with HDAC5 siRNA or transduced with adenoviral vector encoding wild-type HDAC5 and the effects of HDAC5 manipulation were examined by qPCR arrays, ELISA and ELISA-based assays. RESULTS: Synovial class I HDAC expression was associated with local expression of tumour necrosis factor (TNF) and matrix metalloproteinase-1, while class IIa HDAC5 expression was inversely associated with parameters of disease activity (erythrocyte sedimentation rate, C-reactive protein, Disease Activity Score in 28 Joints). Interleukin (IL)-1ß or TNF stimulation selectively suppressed HDAC5 expression in RA FLS, which was sufficient and required for optimal IFNB, CXCL9, CXCL10 and CXCL11 induction by IL-1ß, associated with increased nuclear accumulation of the transcription factor, interferon regulatory factor 1(IRF1). CONCLUSIONS: Inflammatory cytokines suppress RA FLS HDAC5 expression, promoting nuclear localisation of IRF1 and transcription of a subset of type I interferon response genes. Our results identify HDAC5 as a novel inflammatory mediator in RA, and suggest that strategies rescuing HDAC5 expression in vivo, or the development of HDAC inhibitors not affecting HDAC5 activity, may have therapeutic applications in RA treatment.


Assuntos
Artrite Reumatoide/metabolismo , Citocinas/genética , Fibroblastos/metabolismo , Histona Desacetilases/metabolismo , Membrana Sinovial/citologia , Adulto , Idoso , Artrite Reumatoide/genética , Sedimentação Sanguínea , Proteína C-Reativa/análise , Epigênese Genética , Feminino , Humanos , Fator Regulador 1 de Interferon/genética , Interleucina-1beta/metabolismo , Masculino , Metaloproteinase 1 da Matriz/metabolismo , Pessoa de Meia-Idade , RNA Mensageiro/metabolismo , Índice de Gravidade de Doença , Transcrição Gênica , Fator de Necrose Tumoral alfa/metabolismo
7.
Ann Rheum Dis ; 74(9): 1763-71, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24812285

RESUMO

BACKGROUND: Forkhead box O (FoxO) transcription factors integrate environmental signals to modulate cell proliferation and survival, and alterations in FoxO function have been reported in rheumatoid arthritis (RA). OBJECTIVES: To examine the relationship between inflammation and FoxO expression in RA, and to analyse the mechanisms and biological consequences of FoxO regulation in RA fibroblast-like synoviocytes (FLS). METHODS: RNA was isolated from RA patient and healthy donor (HD) peripheral blood and RA synovial tissue. Expression of FoxO1, FoxO3a and FoxO4 was measured by quantitative PCR. FoxO1 DNA binding, expression and mRNA stability in RA FLS were measured by ELISA-based assays, immunoblotting and quantitative PCR. FLS were transduced with adenovirus encoding constitutively active FoxO1 (FoxO1ADA) or transfected with small interfering RNA targeting FoxO1 to examine the effects on cell viability and gene expression. RESULTS: FoxO1 mRNA levels were reduced in RA patient peripheral blood compared with HD blood, and RA synovial tissue FoxO1 expression correlated negatively with disease activity. RA FLS stimulation with interleukin 1ß or tumour necrosis factor caused rapid downregulation of FoxO1. This effect was independent of protein kinase B (PKB), but dependent on c-Jun N-terminal kinase (JNK)-mediated acceleration of FoxO1 mRNA degradation. FoxO1ADA overexpression in RA FLS induced apoptosis associated with altered expression of genes regulating cell cycle and survival, including BIM, p27(Kip1) and Bcl-XL. CONCLUSIONS: Our findings identify JNK-dependent modulation of mRNA stability as an important PKB-independent mechanism underlying FoxO1 regulation by cytokines, and suggest that reduced FoxO1 expression is required to promote FLS survival in RA.


Assuntos
Artrite Reumatoide/genética , Fibroblastos/metabolismo , Fatores de Transcrição Forkhead/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , RNA Mensageiro/metabolismo , Adulto , Idoso , Artrite Reumatoide/metabolismo , Proteínas de Ciclo Celular , Sobrevivência Celular , Regulação para Baixo/efeitos dos fármacos , Feminino , Proteína Forkhead Box O1 , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead/efeitos dos fármacos , Humanos , Interleucina-1beta/farmacologia , Interleucina-6/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-akt/metabolismo , Membrana Sinovial/citologia , Membrana Sinovial/metabolismo , Fatores de Transcrição/genética , Fator de Necrose Tumoral alfa/farmacologia
8.
J Invest Dermatol ; 142(2): 402-413, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34333017

RESUMO

Dermal fibroblasts are strategically positioned underneath the basal epidermis layer to support keratinocyte proliferation and extracellular matrix production. In inflammatory conditions, these fibroblasts produce cytokines and chemokines that promote the chemoattraction of immune cells into the dermis and the hyperplasia of the epidermis, two characteristic hallmarks of psoriasis. However, how dermal fibroblasts specifically contribute to psoriasis development remains largely uncharacterized. In this study, we investigated through which cytokines and signaling pathways dermal fibroblasts contribute to the inflammatory features of psoriatic skin. We show that dermal fibroblasts from lesional psoriatic skin are important producers of inflammatory mediators, including IL-6, CXCL8, and CXCL2. This increased cytokine production was found to be regulated by ZFP36 family members ZFP36, ZFP36L1, and ZFP36L2, RNA-binding proteins with mRNA-degrading properties. In addition, the expression of ZFP36 family proteins was found to be reduced in chronic inflammatory conditions that mimic psoriatic lesional skin. Collectively, these results indicate that dermal fibroblasts are important producers of cytokines in psoriatic skin and that reduced expression of ZFP36 members in psoriasis dermal fibroblasts contributes to their inflammatory phenotype.


Assuntos
Fator 1 de Resposta a Butirato/metabolismo , Fibroblastos/metabolismo , Psoríase/imunologia , Fatores de Transcrição/metabolismo , Tristetraprolina/metabolismo , Biópsia , Fator 1 de Resposta a Butirato/genética , Estudos de Casos e Controles , Epiderme/imunologia , Epiderme/metabolismo , Epiderme/patologia , Técnicas de Silenciamento de Genes , Voluntários Saudáveis , Humanos , Mediadores da Inflamação/metabolismo , Queratinócitos/imunologia , Queratinócitos/metabolismo , Psoríase/patologia , Fatores de Transcrição/genética , Tristetraprolina/genética
9.
J Clin Med ; 10(3)2021 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-33573268

RESUMO

Compelling evidence shows the involvement of plasmacytoid dendritic cells (pDCs) in systemic sclerosis (SSc) pathogenesis. This study investigated whether microRNAs (miRNAs) are involved in the dysregulation of pDCs in SSc patients already at early stages. RNA from circulating pDCs was isolated from two independent cohorts of SSc patients with different disease phenotypes, and individuals with Raynaud's phenomenon, for microRNA profiling and RNA-sequencing analysis. Proteomic analysis was exploited to identify novel direct miRNA targets at the protein level. Twelve and fifteen miRNAs were differentially expressed in at least one group of patients compared to healthy controls in discovery cohort I and II, respectively. Of note, miR-126 and miR-139-5p were upregulated in both preclinical and definite SSc patients and correlated with the expression of type I interferon (IFN)-responsive genes. Toll-like receptor 9 (TLR9) stimulation of healthy pDCs upregulated the expression of both miRNAs, similarly to what was observed in patients. The proteomic analysis identified USP24 as a novel target of miR-139-5p. The expression level of USP24 was inversely correlated with miR-139-5p expression in SSc patients and induced by TLR9 stimulation in healthy pDCs. These findings demonstrated that the miRNA profile is altered in pDCs of SSc patients already at early stages of the disease and indicate their potential contribution to pDC activation observed in patients.

10.
Front Immunol ; 11: 2149, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33042127

RESUMO

Fibrosis is a condition shared by numerous inflammatory diseases. Our incomplete understanding of the molecular mechanisms underlying fibrosis has severely hampered effective drug development. CXCL4 is associated with the onset and extent of fibrosis development in multiple inflammatory and fibrotic diseases. Here, we used monocyte-derived cells as a model system to study the effects of CXCL4 exposure on dendritic cell development by integrating 65 longitudinal and paired whole genome transcriptional and methylation profiles. Using data-driven gene regulatory network analyses, we demonstrate that CXCL4 dramatically alters the trajectory of monocyte differentiation, inducing a novel pro-inflammatory and pro-fibrotic phenotype mediated via key transcriptional regulators including CIITA. Importantly, these pro-inflammatory cells directly trigger a fibrotic cascade by producing extracellular matrix molecules and inducing myofibroblast differentiation. Inhibition of CIITA mimicked CXCL4 in inducing a pro-inflammatory and pro-fibrotic phenotype, validating the relevance of the gene regulatory network. Our study unveils that CXCL4 acts as a key secreted factor driving innate immune training and forming the long-sought link between inflammation and fibrosis.


Assuntos
Células Dendríticas/citologia , Fibrose/imunologia , Redes Reguladoras de Genes , Inflamação/imunologia , Fator Plaquetário 4/fisiologia , Transcriptoma , Células Cultivadas , Técnicas de Reprogramação Celular , Metilação de DNA , Árvores de Decisões , Decitabina/farmacologia , Fibroblastos , Fibrose/genética , Humanos , Inflamação/genética , Monócitos/citologia , Análise de Escalonamento Multidimensional , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/fisiologia , Poli I-C/farmacologia , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , RNA-Seq , Transativadores/antagonistas & inibidores , Transativadores/fisiologia
11.
Mol Immunol ; 114: 524-534, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31518856

RESUMO

The chemokine CXCL4 has been implicated in several immune diseases. Exposure of monocyte-derived dendritic cells (moDCs) to CXCL4 potentiates the production of inflammatory cytokines in the presence of TLR3 or TLR7/8 agonists. Here we investigated the transcriptional and post-transcriptional events underlying the augmented inflammatory responses in CXCL4-moDCs. Our results indicate that CXCL4-moDCs display an increased expression and secretion of IL-12, IL-23, IL-6 and TNF upon TLR3 activation. Analysis of the cytokine transcripts for the presence of AU-rich elements (ARE), motifs necessary for ARE-mediated mRNA decay, revealed that all these cytokine transcripts are, at least in silico, possibly regulated at the level of mRNA stability. In vitro assays confirmed that mRNA stability of IL6 and TNF, but not IL12B and IL23A, is increased in CXCL4-moDCs. We next screened the expression of ARE-binding proteins (ARE-BPs) and found that TLR stimulation of CXCL4-moDCs induced tristetraprolin (TTP or ZFP36). Increased TTP mRNA expression was found to be a consequence of TTP phospho-mediated inactivation, which over time causes the protein to degrade its own mRNA. Concomitantly with TTP inactivation, we observed increased MAPK p38 signalling, upstream of TTP, in stimulated CXCL4-moDCs. P38 inhibition restored TTP activation and subsequently reduced the production of inflammatory cytokines. Finally, TTP knockdown in moDCs resulted in an increased production of IL6 and TNF after TLR stimulation. Overall, our study shows that the pro-inflammatory phenotype of CXCL4-moDCs relies in part on enhanced cytokine mRNA stability dictated by TTP inactivation.


Assuntos
Citocinas/metabolismo , Células Dendríticas/metabolismo , Monócitos/metabolismo , Fator Plaquetário 4/metabolismo , RNA Mensageiro/metabolismo , Humanos , Interleucina-12/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Estabilidade de RNA/fisiologia , Transdução de Sinais/fisiologia , Tristetraprolina/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
12.
Nat Rev Rheumatol ; 14(11): 657-673, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30305700

RESUMO

Systemic sclerosis (SSc) is a severe autoimmune disease that is characterized by vascular abnormalities, immunological alterations and fibrosis of the skin and internal organs. The results of genetic studies in patients with SSc have revealed statistically significant genetic associations with disease manifestations and progression. Nevertheless, genetic susceptibility to SSc is moderate, and the functional consequences of genetic associations remain only partially characterized. A current hypothesis is that, in genetically susceptible individuals, epigenetic modifications constitute the driving force for disease initiation. As epigenetic alterations can occur years before fibrosis appears, these changes could represent a potential link between inflammation and tissue fibrosis. Epigenetics is a fast-growing discipline, and a considerable number of important epigenetic studies in SSc have been published in the past few years that span histone post-translational modifications, DNA methylation, microRNAs and long non-coding RNAs. This Review describes the latest insights into genetic and epigenetic contributions to the pathogenesis of SSc and aims to provide an improved understanding of the molecular pathways that link inflammation and fibrosis. This knowledge will be of paramount importance for the development of medicines that are effective in treating or even reversing tissue fibrosis.


Assuntos
Epigênese Genética , Predisposição Genética para Doença/genética , Escleroderma Sistêmico/genética , Metilação de DNA , Progressão da Doença , Estudos de Associação Genética , Humanos , Processamento de Proteína Pós-Traducional , RNA não Traduzido/genética
13.
Arthritis Res Ther ; 20(1): 148, 2018 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-30029685

RESUMO

BACKGROUND: Histone deacetylase inhibitors (HDACi) suppress cytokine production in immune and stromal cells of patients with rheumatoid arthritis (RA). Here, we investigated the effects of the HDACi givinostat (ITF2357) on the transcriptional and post-transcriptional regulation of inflammatory markers in RA fibroblast-like synoviocytes (FLS). METHODS: The effects of ITF2357 on the expression and messenger RNA (mRNA) stability of IL-1ß-inducible genes in FLS were analyzed using array-based qPCR and Luminex. The expression of primary and mature cytokine transcripts, the mRNA levels of tristetraprolin (TTP, or ZFP36) and other AU-rich element binding proteins (ARE-BP) and the cytokine profile of fibroblasts derived from ZFP36+/+ and ZFP36-/- mice was measured by qPCR. ARE-BP silencing was performed by small interfering RNA (siRNA)-mediated knockdown, and TTP post-translational modifications were analyzed by immunoblotting. RESULTS: ITF2357 reduced the expression of 85% of the analyzed IL-1ß-inducible transcripts, including cytokines (IL6, IL8), chemokines (CXCL2, CXCL5, CXCL6, CXCL10), matrix-degrading enzymes (MMP1, ADAMTS1) and other inflammatory mediators. Analyses of mRNA stability demonstrated that ITF2357 accelerates IL6, IL8, PTGS2 and CXCL2 mRNA degradation, a phenomenon associated with the enhanced transcription of TTP, but not other ARE-BP, and the altered post-translational status of TTP protein. TTP knockdown potentiated cytokine production in RA FLS and murine fibroblasts, which in the latter case was insensitive to inhibition by ITF2357 treatment. CONCLUSIONS: Our study identifies that regulation of cytokine mRNA stability is a predominant mechanism underlying ITF2357 anti-inflammatory properties, occurring via regulation of TTP. These results highlight the therapeutic potential of ITF2357 in the treatment of RA.


Assuntos
Artrite Reumatoide/metabolismo , Citocinas/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Sinoviócitos/efeitos dos fármacos , Animais , Artrite Reumatoide/imunologia , Células Cultivadas , Citocinas/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Camundongos Knockout , Estabilidade de RNA/efeitos dos fármacos , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Sinoviócitos/metabolismo , Tristetraprolina/biossíntese
14.
Epigenomics ; 9(4): 447-461, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28102705

RESUMO

Growing evidence supports the idea that aberrancies in epigenetic processes contribute to the onset and progression of human immune-mediated inflammatory diseases, such as rheumatoid arthritis (RA). Epigenetic regulators of histone tail modifications play a role in chromatin accessibility and transcriptional responses to inflammatory stimuli. Among these, histone deacetylases (HDACs) regulate the acetylation status of histones and nonhistone proteins, essential for immune responses. Broad-spectrum HDAC inhibitors are well-known anti-inflammatory agents and reduce disease severity in animal models of arthritis; however, selective HDAC inhibitors remain poorly studied. In this review, we describe emerging findings regarding the aberrant acetyl code in RA and other rheumatic disorders which may help identify not only novel diagnostic and prognostic clinical biomarkers for RA, but also new targets for epigenetic pharmacological applications.


Assuntos
Artrite Reumatoide/genética , Epigênese Genética , Histonas/metabolismo , Doenças Reumáticas/genética , Acetilação/efeitos dos fármacos , Animais , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Epigênese Genética/efeitos dos fármacos , Código das Histonas , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Histona Desacetilases/metabolismo , Humanos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Doenças Reumáticas/tratamento farmacológico , Doenças Reumáticas/metabolismo
15.
Arthritis Res Ther ; 19(1): 207, 2017 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-28923079

RESUMO

BACKGROUND: Endoplasmic reticulum (ER) stress has proinflammatory properties, and transgenic animal studies of rheumatoid arthritis (RA) indicate its relevance in the process of joint destruction. Because currently available studies are focused primarily on myeloid cells, we assessed how ER stress might affect the inflammatory responses of stromal cells in RA. METHODS: ER stress was induced in RA fibroblast-like synoviocytes (FLS), dermal fibroblasts, and macrophages with thapsigargin or tunicamycin alone or in combination with Toll-like receptor (TLR) ligands, and gene expression and messenger RNA (mRNA) stability was measured by quantitative polymerase chain reaction. Cellular viability was measured using cell death enzyme-linked immunosorbent assays and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, and signaling pathway activation was analyzed by immunoblotting. RESULTS: No cytotoxicity was observed in FLS exposed to thapsigargin, despite significant induction of ER stress markers. Screening of 84 proinflammatory genes revealed minor changes in their expression (fold change 90th percentile range 2.8-8.3) by thapsigargin alone, but the vast majority were hyperinduced during combined stimulation with thapsigargin and TLR ligands (35% greater than fivefold vs lipopolysaccharide alone). The synergistic response could not be explained by quantitative effects on nuclear factor-κB and mitogen-activated protein kinase pathways alone, but it was dependent on increased mRNA stability. mRNA stabilization was similarly enhanced by ER stress in dermal fibroblasts but not in macrophages, correlating with minimal cooperative effects on gene induction in macrophages. CONCLUSIONS: RA FLS are resistant to apoptosis induced by ER stress, but ER stress potentiates their activation by multiple TLR ligands. Interfering with downstream signaling pathway components of ER stress may be of therapeutic potential in the treatment of RA.


Assuntos
Artrite Reumatoide/metabolismo , Estresse do Retículo Endoplasmático/fisiologia , Sinoviócitos/metabolismo , Receptores Toll-Like/metabolismo , Artrite Reumatoide/patologia , Células Cultivadas , Humanos , Sinoviócitos/patologia
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