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1.
JCI Insight ; 8(24)2023 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-37917179

RESUMO

Monocyte-derived macrophages, the major source of pathogenic macrophages in COVID-19, are oppositely instructed by macrophage CSF (M-CSF) or granulocyte macrophage CSF (GM-CSF), which promote the generation of antiinflammatory/immunosuppressive MAFB+ (M-MØ) or proinflammatory macrophages (GM-MØ), respectively. The transcriptional profile of prevailing macrophage subsets in severe COVID-19 led us to hypothesize that MAFB shapes the transcriptome of pulmonary macrophages driving severe COVID-19 pathogenesis. We have now assessed the role of MAFB in the response of monocyte-derived macrophages to SARS-CoV-2 through genetic and pharmacological approaches, and we demonstrate that MAFB regulated the expression of the genes that define pulmonary pathogenic macrophages in severe COVID-19. Indeed, SARS-CoV-2 potentiated the expression of MAFB and MAFB-regulated genes in M-MØ and GM-MØ, where MAFB upregulated the expression of profibrotic and neutrophil-attracting factors. Thus, MAFB determines the transcriptome and functions of the monocyte-derived macrophage subsets that underlie pulmonary pathogenesis in severe COVID-19 and controls the expression of potentially useful biomarkers for COVID-19 severity.


Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/metabolismo , COVID-19/metabolismo , Macrófagos/metabolismo , Macrófagos Alveolares/metabolismo , Biomarcadores/metabolismo , Fator de Transcrição MafB/genética , Fator de Transcrição MafB/metabolismo
2.
J Nonverbal Behav ; 47(2): 117-210, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37162792

RESUMO

Behavioural coding is time-intensive and laborious. Thin slice sampling provides an alternative approach, aiming to alleviate the coding burden. However, little is understood about whether different behaviours coded over thin slices are comparable to those same behaviours over entire interactions. To provide quantitative evidence for the value of thin slice sampling for a variety of behaviours. We used data from three populations of parent-infant interactions: mother-infant dyads from the Grown in Wales (GiW) cohort (n = 31), mother-infant dyads from the Avon Longitudinal Study of Parents and Children (ALSPAC) cohort (n = 14), and father-infant dyads from the ALSPAC cohort (n = 11). Mean infant ages were 13.8, 6.8, and 7.1 months, respectively. Interactions were coded using a comprehensive coding scheme comprised of 11-14 behavioural groups, with each group comprised of 3-13 mutually exclusive behaviours. We calculated frequencies of verbal and non-verbal behaviours, transition matrices (probability of transitioning between behaviours, e.g., from looking at the infant to looking at a distraction) and stationary distributions (long-term proportion of time spent within behavioural states) for 15 thin slices of full, 5-min interactions. Measures drawn from the full sessions were compared to those from 1-, 2-, 3- and 4-min slices. We identified many instances where thin slice sampling (i.e., < 5 min) was an appropriate coding method, although we observed significant variation across different behaviours. We thereby used this information to provide detailed guidance to researchers regarding how long to code for each behaviour depending on their objectives.

3.
Cell Mol Life Sci ; 80(4): 96, 2023 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-36930354

RESUMO

Monocyte-derived macrophages contribute to pathogenesis in inflammatory diseases and their effector functions greatly depend on the prevailing extracellular milieu. Whereas M-CSF primes macrophages for acquisition of an anti-inflammatory profile, GM-CSF drives the generation of T cell-stimulatory and pro-inflammatory macrophages. Liver X Receptors (LXRα and LXRß) are nuclear receptors that control cholesterol metabolism and regulate differentiation of tissue-resident macrophages. Macrophages from rheumatoid arthritis and other inflammatory pathologies exhibit an enriched LXR pathway, and recent reports have shown that LXR activation raises pro-inflammatory effects and impairs the acquisition of the anti-Inflammatory profile of M-CSF-dependent monocyte-derived macrophages (M-MØ). We now report that LXR inhibition prompts the acquisition of an anti-inflammatory gene and functional profile of macrophages generated within a pathological environment (synovial fluid from Rheumatoid Arthritis patients) as well as during the GM-CSF-dependent differentiation of human monocyte-derived macrophages (GM-MØ). Mechanistically, inhibition of LXR results in macrophages with higher expression of the v-Maf Avian Musculoaponeurotic Fibrosarcoma Oncogene Homolog B (MAFB) transcription factor, which governs the macrophage anti-inflammatory profile, as well as over-expression of MAFB-regulated genes. Indeed, gene silencing experiments on human macrophages evidenced that MAFB is required for the LXR inhibitor to enhance the anti-inflammatory nature of human macrophages. As a whole, our results demonstrate that LXR inhibition prompts the acquisition of an anti-inflammatory transcriptional and functional profile of human macrophages in a MAFB-dependent manner, and propose the use of LXR antagonists as potential therapeutic alternatives in macrophage re-programming strategies during inflammatory responses.


Assuntos
Artrite Reumatoide , Fator Estimulador de Colônias de Granulócitos e Macrófagos , Humanos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Fator Estimulador de Colônias de Macrófagos/genética , Regulação para Cima , Macrófagos/metabolismo , Artrite Reumatoide/patologia , Anti-Inflamatórios/metabolismo , Receptores X do Fígado/genética , Receptores X do Fígado/metabolismo , Fator de Transcrição MafB/genética , Fator de Transcrição MafB/metabolismo
4.
Immunobiology ; 228(2): 152334, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36641984

RESUMO

Helicobacter pylori is a gram-negative bacterium that is present in over half of the world's population. The colonization of the stomach́s gastric mucosa by H. pylori is related to the onset of chronic gastritis, peptic ulcer, and cancer. The estimated deaths from gastric cancer caused by this bacterial infection are in the 15,000-150,000 range. Current treatment for controlling the colonization of H. pylori includes the administration of two to four antibiotics and a gastric ATPase proton pump inhibitor. Nevertheless, the bacterium has shown increased resistance to antibiotics. Despite an extensive list of attempts to develop a vaccine, no approved vaccine against H. pylori is available. Recombinant viruses are a novel alternative for the control of primary pathogenic agents. In this work, we employed a baculovirus that carries a Thp1 transgene coding for nine H. pylori epitopes, some from the literature, and others were selected in silico from the sequence of H. pylori proteins (carbonic anhydrase, urease B subunit, gamma-glutamyl transpeptidase, Lpp20, Cag7, and CagL). We verified the expression of this hybrid multiepitopic protein in HeLa cells. Mice were inoculated with the recombinant baculovirus Bac-Thp1 using various administration routes: intranasal, intragastric, intramuscular, and a combination of intranasal and intragastric. We identified a strong adjuvant-independent IgG-antibody response in the serum of recombinant baculovirus-Thp1 inoculated mice, which was specific for a strain of H. pylori isolated from a human patient. The bacterium-specific IgG-antibodies were present in sera 125 days after the first vaccine administration. Also, H. pylori-specific IgA-antibodies were found in feces at 82 days after the first inoculation. A baculovirus-based vaccine for H. pylori is promising for controlling this pathogen in humans.


Assuntos
Infecções por Helicobacter , Helicobacter pylori , Humanos , Animais , Camundongos , Baculoviridae , Células HeLa , Vacinas Bacterianas , Imunoglobulina G , Anticorpos Antibacterianos
5.
J Clin Immunol ; 42(5): 1093-1105, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35486340

RESUMO

Common variable immunodeficiency disorders (CVID), the most common primary immune deficiency, includes heterogeneous syndromes characterized by hypogammaglobulinemia and impaired antibody responses. CVID patients frequently suffer from recurrent infections and inflammatory conditions. Currently, immunoglobulin replacement therapy (IgRT) is the first-line treatment to prevent infections and aminorate immune alterations in CVID patients. Intravenous Immunoglobulin (IVIg), a preparation of highly purified poly-specific IgG, is used for treatment of immunodeficiencies as well as for autoimmune and inflammatory disorders, as IVIg exerts immunoregulatory and anti-inflammatory actions on innate and adaptive immune cells. To determine the mechanism of action of IVIg in CVID in vivo, we determined the effect of IVIg infusion on the transcriptome of peripheral blood mononuclear cells from CVID patients, and found that peripheral blood monocytes are primary targets of IVIg in vivo, and that IVIg triggers the acquisition of an anti-inflammatory gene profile in human monocytes. Moreover, IVIg altered the relative proportions of peripheral blood monocyte subsets and enhanced the proportion of CD14+ cells with a transcriptional, phenotypic, and functional profile that resembles that of monocytic myeloid-derived suppressor cells (MDSC). Therefore, our results indicate that CD14 + MDSC-like cells might contribute to the immunoregulatory effects of IVIg in CVID and other inflammatory disorders.


Assuntos
Imunodeficiência de Variável Comum , Células Supressoras Mieloides , Imunodeficiência de Variável Comum/tratamento farmacológico , Humanos , Imunoglobulinas Intravenosas , Leucócitos Mononucleares , Monócitos
6.
Front Immunol ; 13: 835478, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35280993

RESUMO

Liver X Receptors (LXR) control cholesterol metabolism and exert anti-inflammatory actions but their contribution to human macrophage polarization remains unclear. The LXR pathway is enriched in pro-inflammatory macrophages from rheumatoid arthritis as well as in tumors-associated macrophages from human tumors. We now report that LXR activation inhibits the anti-inflammatory gene and functional profile of M-CSF-dependent human macrophages, and prompts the acquisition of a pro-inflammatory gene signature, with both effects being blocked by an LXR inverse agonist. Mechanistically, the LXR-stimulated macrophage polarization shift correlates with diminished expression of MAFB and MAF, which govern the macrophage anti-inflammatory profile, and with enhanced release of activin A. Indeed, LXR activation impaired macrophage polarization in response to tumor-derived ascitic fluids, as well as the expression of MAF- and MAFB-dependent genes. Our results demonstrate that LXR activation limits the anti-inflammatory human macrophage polarization and prompts the acquisition of an inflammatory transcriptional and functional profile.


Assuntos
Fator Estimulador de Colônias de Macrófagos , Macrófagos , Anti-Inflamatórios/metabolismo , Anti-Inflamatórios/farmacologia , Células Cultivadas , Humanos , Receptores X do Fígado/genética , Receptores X do Fígado/metabolismo , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos/metabolismo
7.
J Innate Immun ; 14(3): 243-256, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34670213

RESUMO

During inflammatory responses, monocytes are recruited into inflamed tissues, where they become monocyte-derived macrophages and acquire pro-inflammatory and tissue-damaging effects in response to the surrounding environment. In fact, monocyte-derived macrophage subsets are major pathogenic cells in inflammatory pathologies. Strikingly, the transcriptome of pathogenic monocyte-derived macrophage subsets resembles the gene profile of macrophage colony-stimulating factor (M-CSF)-primed monocyte-derived human macrophages (M-MØ). As M-MØ display a characteristic cytokine profile after activation (IL10high TNFlow IL23low IL6low), we sought to determine the transcriptional signature of M-MØ upon exposure to pathogenic stimuli. Activation of M-MØ led to the acquisition of a distinctive transcriptional profile characterized by the induction of a group of genes (Gene set 1) highly expressed by pathogenic monocyte-derived macrophages in COVID-19 and whose presence in tumor-associated macrophages (TAM) correlates with the expression of macrophage-specific markers (CD163, SPI1) and IL10. Indeed, Gene set 1 expression was primarily dependent on ERK/p38 and STAT3 activation, and transcriptional analysis and neutralization experiments revealed that IL-10 is not only required for the expression of a subset of genes within Gene set 1 but also significantly contributes to the idiosyncratic gene signature of activated M-MØ. Our results indicate that activation of M-CSF-dependent monocyte-derived macrophages induces a distinctive gene expression profile, which is partially dependent on IL-10, and identifies a gene set potentially helpful for macrophage-centered therapeutic strategies.


Assuntos
COVID-19 , Fator Estimulador de Colônias de Macrófagos , Diferenciação Celular , Células Cultivadas , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Interleucina-10/genética , Interleucina-10/metabolismo , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos/metabolismo , Monócitos/metabolismo
8.
Front Immunol ; 11: 603507, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33312178

RESUMO

Defective IFN production and exacerbated inflammatory and pro-fibrotic responses are hallmarks of SARS-CoV-2 infection in severe COVID-19. Based on these hallmarks, and considering the pivotal role of macrophages in COVID-19 pathogenesis, we hypothesize that the transcription factors MAFB and MAF critically contribute to COVID-19 progression by shaping the response of macrophages to SARS-CoV-2. Our proposal stems from the recent identification of pathogenic lung macrophage subsets in severe COVID-19, and takes into consideration the previously reported ability of MAFB to dampen IFN type I production, as well as the critical role of MAFB and MAF in the acquisition and maintenance of the transcriptional signature of M-CSF-conditioned human macrophages. Solid evidences are presented that link overexpression of MAFB and silencing of MAF expression with clinical and biological features of severe COVID-19. As a whole, we propose that a high MAFB/MAF expression ratio in lung macrophages could serve as an accurate diagnostic tool for COVID-19 progression. Indeed, reversing the macrophage MAFB/MAF expression ratio might impair the exacerbated inflammatory and profibrotic responses, and restore the defective IFN type I production, thus becoming a potential strategy to limit severity of COVID-19.


Assuntos
COVID-19/imunologia , Macrófagos/imunologia , Fatores de Transcrição Maf/imunologia , Fator de Transcrição MafB/imunologia , SARS-CoV-2/imunologia , COVID-19/genética , COVID-19/virologia , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Macrófagos/metabolismo , Fatores de Transcrição Maf/genética , Fatores de Transcrição Maf/metabolismo , Fator de Transcrição MafB/genética , Fator de Transcrição MafB/metabolismo , SARS-CoV-2/fisiologia , Índice de Gravidade de Doença
9.
J Immunol ; 204(10): 2808-2817, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32253244

RESUMO

Macrophages can either promote or resolve inflammatory responses, and their polarization state is modulated by peripheral serotonin (5-hydroxytryptamine [5-HT]). In fact, pro- and anti-inflammatory macrophages differ in the expression of serotonin receptors, with 5-HT2B and 5-HT7 expression restricted to M-CSF-primed monocyte-derived macrophages (M-MØ). 5-HT7 drives the acquisition of profibrotic and anti-inflammatory functions in M-MØ, whereas 5-HT2B prevents the degeneration of spinal cord mononuclear phagocytes and modulates motility of murine microglial processes. Because 5-HT2B mediates clinically relevant 5-HT-related pathologies (valvular heart disease, pulmonary arterial hypertension) and is an off target of anesthetics, antiparkinsonian drugs, and selective serotonin reuptake inhibitors, we sought to determine the transcriptional consequences of 5-HT2B engagement in human macrophages, for which 5-HT2B signaling remains unknown. Assessment of the effects of specific agonists and antagonist revealed that 5-HT2B engagement modifies the cytokine and gene signature of anti-inflammatory M-MØ, upregulates the expression of aryl hydrocarbon receptor (AhR) target genes, and stimulates the transcriptional activation of AhR. Moreover, we found that 5-HT dose dependently upregulates the expression of AhR target genes in M-MØ and that the 5-HT-mediated activation of AhR is 5-HT2B dependent because it is abrogated by the 5-HT2B-specific antagonist SB204741. Altogether, our results demonstrate the existence of a functional 5-HT/5-HT2B/AhR axis in human macrophages and indicate that 5-HT potentiates the activity of a transcription factor (AhR) that regulates immune responses and the biological responses to xenobiotics.


Assuntos
Macrófagos/fisiologia , Microglia/fisiologia , Receptor 5-HT2B de Serotonina/metabolismo , Receptores de Hidrocarboneto Arílico/metabolismo , Serotonina/metabolismo , Diferenciação Celular , Células Cultivadas , Humanos , Indóis/farmacologia , Fagocitose , RNA Interferente Pequeno/genética , Receptores de Hidrocarboneto Arílico/genética , Receptores de Serotonina/metabolismo , Agonistas do Receptor 5-HT2 de Serotonina/farmacologia , Transdução de Sinais , Tiofenos/farmacologia , Ativação Transcricional , Transcriptoma
10.
Biomedicines ; 8(3)2020 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-32143497

RESUMO

In hydroxychloroquine (HCQ) retinopathy, early detection of asymptomatic retinal changes and the interruption of the drug are essential to prevent permanent vision loss. Our purpose was to investigate the roles of ganglion cell layer (GCL) and outer nuclear layer (ONL) thicknesses measured by optical coherence tomography (OCT) in the early diagnosis of retinopathy. One hundred and fourteen eyes of 76 individuals with HCQ treatment were enrolled in the study (42 eyes with impaired visual field (VF) and 72 eyes with nondamaged VF). We found that ONL was significantly decreased in the HCQ retinopathy group compared with the control group in the nasal macula (p = 0.032) as well as in four sectors (p < 0.044), whereas no significant differences were found comparing GCL in both groups. If VF were altered superiorly or temporarily, ONL was significantly thinned inferiorly (p = 0.029) and nasally (p = 0.008), respectively. Duration of HCQ treatment was significantly related with ONL in seven sectors of ONL (p < 0.047). We suggest that ONL measured with OCT might be used to assess early HCQ retinal toxicity.

11.
PLoS One ; 14(9): e0222347, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31509597

RESUMO

PURPOSE: To evaluate the accuracy of the measurement of the ganglion cell layer (GCL) of the posterior pole analysis (PPA) software of the Spectralis spectral-domain (SD) optical coherence tomography (OCT) device (Heidelberg Engineering, Inc., Heidelberg, Germany), the asymmetry of paired GCL sectors, the total retinal thickness asymmetry (RTA), and the peripapillary retinal nerve fiber layer (pRNFL) test to discriminate between healthy, early and advanced glaucoma eyes. METHODS: Three hundred eighteen eyes of 161 individuals with reliable visual fields (VF) were enrolled in this study. All participants were examined using the standard posterior pole and the pRNFL protocols of the Spectralis OCT device. VF impairment was graded in hemifields, and the GCL sectors were correlated with this damage. Thicknesses of each GCL, the GCL map deviation asymmetry and the pRNFL were compared between control and glaucomatous eyes. The area under the receiver operating characteristic curve (AUC) of these analyses was assessed. RESULTS: Fourteen of the 16 sectors of the GCL and pRNFL were significantly thinner in eyes with glaucoma than in control eyes (p<0.006). Similarly, the GCL map deviation showed a significant difference between these eyes and both the control eyes as well as the eyes with early glaucoma (p = 0.001 and p = 0.039, respectively). The highest values of AUC to diagnose both early and advanced glaucoma corresponded to the average pRNFL analysis and the GCL map deviation (AUC>0.823, p<0.040 and AUC>0.708, p<0.188, respectively). CONCLUSIONS: Although 16 central sectors of the GCL observed with PPA showed good correlation with VF damage, the pRNFL and the GCL map deviation were more effective for discrimination of glaucomatous damage.


Assuntos
Glaucoma/diagnóstico por imagem , Células Ganglionares da Retina/fisiologia , Tomografia de Coerência Óptica/métodos , Campos Visuais/fisiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Alemanha , Humanos , Pressão Intraocular , Macula Lutea/fisiologia , Masculino , Pessoa de Meia-Idade , Fibras Nervosas/fisiologia , Disco Óptico/fisiologia , Estudos Prospectivos , Curva ROC , Retina/fisiologia , Testes de Campo Visual/métodos
12.
Eye (Lond) ; 33(11): 1741-1747, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31164729

RESUMO

OBJECTIVES: We investigated the effects of topical phenylephrine 2.5% instillation on choroidal thickness (CT), peripapillary choroidal thickness (pCT) and retinal nerve fibre layer (RNFL). METHODS: Healthy control patients underwent enhanced depth imaging (EDI) by spectral-domain optical coherence tomography (OCT) before and 30 min after phenylephrine instillation, using eye-tracking and follow-up software. Changes in 14 different locations of CT, 2 locations of pCT and RNFL were assessed. RESULTS: The study included 119 eyes of 62 patients (19 males and 43 females), with a mean age of 59.8 ± 15.3 years (range: 26-88 years). Within 30 min after instillation, the mean subfoveal CT both in vertical and horizontal scan were significantly thinned (p = 0.005 and p = 0.018, respectively). In total, 1500, 1000 and 500 µm temporal CT measurements showed also a significant thinning (p = 0.021, p = 0.037 and p = 0.020, respectively), as well as 500 µm both superior (p = 0.045) and inferior (p = 0.009). 1500, 1000 and 500 µm nasal CT, and 1500 and 1000 µm CT superior and inferior measurements showed no significant thinning after phenylephrine instillation. pCT was significantly thinned after phenylephrine in both superior (p = 0.016) and inferior (p = 0.050) measurements. RNFL analysis did not significantly change after phenylephrine instillation (p = 0.209). CONCLUSIONS: A significant thinning of CT and pCT occurred following phenylephrine instillation. Future studies analysing CT and pCT should detail if this mydriatic agent was used or not.


Assuntos
Corioide/anatomia & histologia , Midriáticos/administração & dosagem , Fenilefrina/administração & dosagem , Administração Oftálmica , Adulto , Idoso , Idoso de 80 Anos ou mais , Corioide/diagnóstico por imagem , Feminino , Voluntários Saudáveis , Humanos , Macula Lutea/anatomia & histologia , Macula Lutea/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Fibras Nervosas , Soluções Oftálmicas , Disco Óptico/anatomia & histologia , Disco Óptico/diagnóstico por imagem , Pupila/efeitos dos fármacos , Células Ganglionares da Retina/citologia , Tomografia de Coerência Óptica
14.
J Immunol ; 201(1): 41-52, 2018 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-29743313

RESUMO

IVIg is an approved therapy for immunodeficiency and for several autoimmune and inflammatory diseases. However, the molecular basis for the IVIg anti-inflammatory activity remains to be fully explained and cannot be extrapolated from studies on animal models of disease. We now report that IVIg impairs the generation of human monocyte-derived anti-inflammatory macrophages by inducing JNK activation and activin A production and limits proinflammatory macrophage differentiation by inhibiting GM-CSF-driven STAT5 activation. In vivo, IVIg provokes a rapid increase in peripheral blood activin A, CCL2, and IL-6 levels, an effect that can be recapitulated in vitro on human monocytes. On differentiating monocytes, IVIg promotes the acquisition of altered transcriptional and cytokine profiles, reduces TLR expression and signaling, and upregulates negative regulators of TLR-initiated intracellular signaling. In line with these effects, in vivo IVIg infusion induces a state tolerant toward subsequent stimuli that results in reduced inflammatory cytokine production after LPS challenge in human peripheral blood and significant protection from LPS-induced death in mice. Therefore, IVIg conditions human macrophages toward the acquisition of a state of cross-tolerance against inflammatory stimuli, an effect that correlates with the net anti-inflammatory action of IVIg in vivo.


Assuntos
Anti-Inflamatórios/imunologia , Tolerância Imunológica/imunologia , Imunoglobulinas Intravenosas/imunologia , Imunoglobulinas Intravenosas/farmacologia , Macrófagos/imunologia , Fator de Transcrição STAT5/metabolismo , Ativinas/sangue , Animais , Células Cultivadas , Quimiocina CCL2/sangue , Ativação Enzimática , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Inflamação/imunologia , Interleucina-6/sangue , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Lipopolissacarídeos/imunologia , Camundongos , Monócitos/citologia , Monócitos/imunologia
15.
Front Immunol ; 9: 31, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29434585

RESUMO

GM-CSF promotes the functional maturation of lung alveolar macrophages (A-MØ), whose differentiation is dependent on the peroxisome proliferator-activated receptor gamma (PPARγ) transcription factor. In fact, blockade of GM-CSF-initiated signaling or deletion of the PPARγ-encoding gene PPARG leads to functionally defective A-MØ and the onset of pulmonary alveolar proteinosis. In vitro, macrophages generated in the presence of GM-CSF display potent proinflammatory, immunogenic and tumor growth-limiting activities. Since GM-CSF upregulates PPARγ expression, we hypothesized that PPARγ might contribute to the gene signature and functional profile of human GM-CSF-conditioned macrophages. To verify this hypothesis, PPARγ expression and activity was assessed in human monocyte-derived macrophages generated in the presence of GM-CSF [proinflammatory GM-CSF-conditioned human monocyte-derived macrophages (GM-MØ)] or M-CSF (anti-inflammatory M-MØ), as well as in ex vivo isolated human A-MØ. GM-MØ showed higher PPARγ expression than M-MØ, and the expression of PPARγ in GM-MØ was found to largely depend on activin A. Ligand-induced activation of PPARγ also resulted in distinct transcriptional and functional outcomes in GM-MØ and M-MØ. Moreover, and in the absence of exogenous activating ligands, PPARγ knockdown significantly altered the GM-MØ transcriptome, causing a global upregulation of proinflammatory genes and significantly modulating the expression of genes involved in cell proliferation and migration. Similar effects were observed in ex vivo isolated human A-MØ, where PPARγ silencing led to enhanced expression of genes coding for growth factors and chemokines and downregulation of cell surface pathogen receptors. Therefore, PPARγ shapes the transcriptome of GM-CSF-dependent human macrophages (in vitro derived GM-MØ and ex vivo isolated A-MØ) in the absence of exogenous activating ligands, and its expression is primarily regulated by activin A. These results suggest that activin A, through enhancement of PPARγ expression, help macrophages to switch from a proinflammatory to an anti-inflammatory polarization state, thus contributing to limit tissue damage and restore homeostasis.


Assuntos
Ativinas/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Inflamação/imunologia , Macrófagos Alveolares/citologia , Macrófagos Alveolares/imunologia , PPAR gama/metabolismo , Animais , Diferenciação Celular/imunologia , Linhagem Celular , Meios de Cultivo Condicionados/farmacologia , Regulação da Expressão Gênica , Células HEK293 , Humanos , Inflamação/patologia , Macrófagos Alveolares/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , PPAR gama/genética , Interferência de RNA , RNA Interferente Pequeno/genética
16.
Ann Hepatol ; 17(6): 1026-1034, 2018 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-30600292

RESUMO

INTRODUCTION AND AIM: Hepatic encephalopathy (HE), caused by hyperammonemia resulting from liver disease, is a spectrum of neuropsychiatric and motor disorders that can lead to death. Existing therapies are deficient and alternative treatments are needed. We have shown that gene therapy with a baculovirus vector containing the glutamine synthetase (Bac-GS) gene is efficient for reducing ammonia levels in an acute hyperammonemia rat model. However, the most common condition resulting from liver disease is chronic hyperammonemia. In this work, Bac-GS was evaluated in bile-duct ligated rats, a chronic liver disease model with hyperammonemia and some characteristics of Type C HE. MATERIAL AND METHODS: Bac-GS was tested for mediating GS overexpression in HeLa cells and H9C2 myotubes. For determining the utility of Bac-GS for the reduction of ammonia levels in a chronic hyperammonemia animal model, four groups of rats were treated: control, sham, ligated with Bac-GS and ligated with Bac-GFP. Baculoviruses were injected i.m. 18 days post-surgery. Blood was drawn 2, 3 and 4 weeks post-surgery and plasma ammonia concentrations were quantified. RESULTS: In protein lysates of cells and myotubes transduced with Bac-GS, a 44 kDa band corresponding to GS was detected. Significant results were obtained in the hyperammonemic bile-duct ligated rat model, as plasma ammonia was reduced to normal levels 3 days after treatment with Bac-GS. Furthermore, a transitory effect of Bac-GS was observed. CONCLUSION: Our results show that gene therapy by delivering GS is a promising alternative for treatment of hyperammonemia in acute-on-chronic liver failure patients with HE.


Assuntos
Baculoviridae/genética , Terapia Genética/métodos , Encefalopatia Hepática/etiologia , Encefalopatia Hepática/terapia , Hiperamonemia/complicações , Análise de Variância , Animais , Western Blotting , Células Cultivadas , Doença Crônica , Modelos Animais de Doenças , Vetores Genéticos , Glutamato-Amônia Ligase/administração & dosagem , Células HeLa/citologia , Células HeLa/patologia , Encefalopatia Hepática/patologia , Humanos , Hiperamonemia/fisiopatologia , Distribuição Aleatória , Ratos , Fatores de Risco , Sensibilidade e Especificidade
17.
Sci Rep ; 7(1): 14761, 2017 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-29116249

RESUMO

Peripheral serotonin (5-hydroxytryptamine, 5-HT) regulates cell growth and differentiation in numerous cell types through engagement of seven types of cell surface receptors (HTR1-7). Deregulated 5-HT/HTR levels contribute to pathology in chronic inflammatory diseases, with macrophages being relevant targets for the physio-pathological effects of 5-HT. In fact, 5-HT skews human macrophage polarization through engagement of 5-HT2BR and 5-HT7R receptors. We now report that 5-HT primes macrophages for reduced pro-inflammatory cytokine production and IFN type I-mediated signaling, and promotes an anti-inflammatory and pro-fibrotic gene signature in human macrophages. The acquisition of the 5-HT-dependent gene profile primarily depends on the 5-HT7R receptor and 5-HT7R-initiated PKA-dependent signaling. In line with the transcriptional results, 5-HT upregulates TGFß1 production by human macrophages in an HTR7- and PKA-dependent manner, whereas the absence of Htr7 in vivo results in diminished macrophage infiltration and collagen deposition in a mouse model of skin fibrosis. Our results indicate that the anti-inflammatory and pro-fibrotic activity of 5-HT is primarily mediated through the 5-HT7R-PKA axis, and that 5-HT7R contributes to pathology in fibrotic diseases.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Citocinas/genética , Fibrose/genética , Perfilação da Expressão Gênica , Mediadores da Inflamação/metabolismo , Receptores de Serotonina/metabolismo , Serotonina/fisiologia , Transdução de Sinais , Dermatopatias/genética , Animais , Células Cultivadas , Humanos , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Receptores de Serotonina/genética
18.
J Immunol ; 199(11): 3858-3869, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-29061766

RESUMO

Obesity is associated with low-grade inflammation and elevated levels of circulating saturated fatty acids, which trigger inflammatory responses by engaging pattern recognition receptors in macrophages. Because tissue homeostasis is maintained through an adequate balance of pro- and anti-inflammatory macrophages, we assessed the transcriptional and functional profile of M-CSF-dependent monocyte-derived human macrophages exposed to concentrations of saturated fatty acids found in obese individuals. We report that palmitate (C16:0, 200 µM) significantly modulates the macrophage gene signature, lowers the expression of transcription factors that positively regulate IL-10 expression (MAFB, AhR), and promotes a proinflammatory state whose acquisition requires JNK activation. Unlike LPS, palmitate exposure does not activate STAT1, and its transcriptional effects can be distinguished from those triggered by LPS, as both agents oppositely regulate the expression of CCL19 and TRIB3 Besides, palmitate conditions macrophages for exacerbated proinflammatory responses (lower IL-10 and CCL2, higher TNF-α, IL-6, and IL-1ß) toward pathogenic stimuli, a process also mediated by JNK activation. All of these effects of palmitate are fatty acid specific because oleate (C18:1, 200 µM) does not modify the macrophage transcriptional and functional profiles. Therefore, pathologic palmitate concentrations promote the acquisition of a specific polarization state in human macrophages and condition macrophages for enhanced responses toward inflammatory stimuli, with both effects being dependent on JNK activation. Our results provide further insight into the macrophage contribution to obesity-associated inflammation.


Assuntos
Inflamação/imunologia , Macrófagos/imunologia , Obesidade/imunologia , Palmitatos/imunologia , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular , Células Cultivadas , Quimiocina CCL19/genética , Quimiocina CCL19/metabolismo , Citocinas/metabolismo , Regulação da Expressão Gênica , Humanos , Mediadores da Inflamação/metabolismo , MAP Quinase Quinase 4/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Ativação Transcricional , Transcriptoma
19.
J Immunol ; 198(5): 2070-2081, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-28093525

RESUMO

Macrophage phenotypic and functional heterogeneity derives from tissue-specific transcriptional signatures shaped by the local microenvironment. Most studies addressing the molecular basis for macrophage heterogeneity have focused on murine cells, whereas the factors controlling the functional specialization of human macrophages are less known. M-CSF drives the generation of human monocyte-derived macrophages with a potent anti-inflammatory activity upon stimulation. We now report that knockdown of MAFB impairs the acquisition of the anti-inflammatory profile of human macrophages, identify the MAFB-dependent gene signature in human macrophages and illustrate the coexpression of MAFB and MAFB-target genes in CD163+ tissue-resident and tumor-associated macrophages. The contribution of MAFB to the homeostatic/anti-inflammatory macrophage profile is further supported by the skewed polarization of monocyte-derived macrophages from multicentric carpotarsal osteolysis (Online Mendelian Inheritance in Man #166300), a pathology caused by mutations in the MAFB gene. Our results demonstrate that MAFB critically determines the acquisition of the anti-inflammatory transcriptional and functional profiles of human macrophages.


Assuntos
Diferenciação Celular , Síndrome de Hajdu-Cheney/imunologia , Macrófagos/fisiologia , Fator de Transcrição MafB/metabolismo , Monócitos/fisiologia , Animais , Anti-Inflamatórios , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Diferenciação Celular/genética , Células Cultivadas , Microambiente Celular , Citocinas/metabolismo , Técnicas de Silenciamento de Genes , Ontologia Genética , Síndrome de Hajdu-Cheney/genética , Homeostase , Humanos , Fator Estimulador de Colônias de Macrófagos/metabolismo , Fator de Transcrição MafB/genética , Camundongos , Mutação/genética , Receptores de Superfície Celular/metabolismo , Células Th2/imunologia , Transcriptoma
20.
J Immunol ; 196(3): 1327-37, 2016 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-26729812

RESUMO

Human CD14(++)CD16(-) and CD14(+/lo)CD16(+) monocyte subsets comprise 85 and 15% of blood monocytes, respectively, and are thought to represent distinct stages in the monocyte differentiation pathway. However, the differentiation fates of both monocyte subsets along the macrophage (Mϕ) lineage have not yet been elucidated. We have now evaluated the potential of CD14(++) CD16(-) and CD16(+) monocytes to differentiate and to be primed toward pro- or anti-inflammatory Mϕs upon culture with GM-CSF or M-CSF, respectively (subsequently referred to as GM14, M14, GM16, or M16). Whereas GM16 and GM14 were phenotypic and functionally analogous, M16 displayed a more proinflammatory profile than did M14. Transcriptomic analyses evidenced that genes associated with M-CSF-driven Mϕ differentiation (including FOLR2, IL10, IGF1, and SERPINB2) are underrepresented in M16 with respect to M14. The preferential proinflammatory skewing of M16 relative to M14 was found to be mediated by the secretion of activin A and the low levels of IL-10 produced by M16. In fact, activin A receptor blockade during the M-CSF-driven differentiation of CD16(+) monocytes, or addition of IL-10-containing M14-conditioned medium, significantly enhanced their expression of anti-inflammatory-associated molecules while impairing their acquisition of proinflammatory-related markers. Thus, we propose that M-CSF drives CD14(++)CD16- monocyte differentiation into bona fide anti-inflammatory Mϕs in a self-autonomous manner, whereas M-CSF-treated CD16(+) monocytes generate Mϕs with a skewed proinflammatory profile by virtue of their high activin A expression unless additional anti-inflammatory stimuli such as IL-10 are provided.


Assuntos
Ativinas/biossíntese , Diferenciação Celular/imunologia , Interleucina-10/biossíntese , Macrófagos/citologia , Monócitos/imunologia , Ativinas/imunologia , Western Blotting , Separação Celular , Citometria de Fluxo , Imunofluorescência , Humanos , Inflamação/imunologia , Interleucina-10/imunologia , Macrófagos/imunologia , Monócitos/citologia , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase em Tempo Real , Receptores de IgG/imunologia
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