RESUMEN
BACKGROUND: Giant cell arteritis (GCA) is an immune-mediated large-vessels vasculitis with complex etiology. Although the pathogenic mechanisms remain poorly understood, a central role for CD4+ T cells has been demonstrated. In this context, understanding the transcriptome dysregulation in GCA CD4+ T cells will yield new insights into its pathogenesis. METHODS: Transcriptome analysis was conducted on CD4+ T cells from 70 patients with GCA with different disease activity and treatment status (active patients before treatment and patients in remission with and without glucocorticoid treatment), and 28 healthy controls. The study also evaluated potential impacts of DNA methylation on gene expression alterations and assessed cross-talk with CD14+ monocytes. RESULTS: This study has uncovered a substantial number of genes and pathways potentially contributing to the pathogenicity of CD4+ T cells in GCA. Specifically, CD4+ T cells from GCA patients with active disease exhibited altered expression levels of genes involved in multiple immune-related processes, including various interleukins (IL) signaling pathways. Notably, IL-2, a decisive interleukin for regulatory T cells homeostasis, was among the most significant. Additionally, impaired apoptotic pathways appear crucial in GCA development. Our findings also suggest that histone-related epigenetic pathways may be implicated in promoting an inflammatory phenotype in GCA active patients. Finally, our study observed altered signaling communication, such as the Jagged-Notch signaling, between CD4+ T cells and monocytes that could have pathogenic relevance in GCA. CONCLUSIONS: Our study suggests the participation of novel cytokines and pathways and the occurrence of a disruption of monocyte-T cell crosstalk driving GCA pathogenesis.
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Linfocitos T CD4-Positivos , Perfilación de la Expresión Génica , Arteritis de Células Gigantes , Monocitos , Transducción de Señal , Transcriptoma , Humanos , Arteritis de Células Gigantes/inmunología , Arteritis de Células Gigantes/genética , Monocitos/inmunología , Monocitos/metabolismo , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Femenino , Masculino , Anciano , Metilación de ADN , Persona de Mediana Edad , Anciano de 80 o más Años , Epigénesis Genética , Comunicación Celular/inmunología , Regulación de la Expresión GénicaRESUMEN
Multiple myeloma is a malignancy characterized by the accumulation of malignant plasma cells in bone marrow and the production of monoclonal immunoglobulin. A hallmark of cancer is the evasion of immune surveillance. Histone deacetylase inhibitors have been shown to promote the expression of silenced molecules and hold potential to increase the anti-MM efficacy of immunotherapy. The aim of the present work was to assess the potential effect of tinostamustine (EDO-S101), a first-in-class alkylating deacetylase inhibitor, in combination with daratumumab, an anti-CD38 monoclonal antibody (mAb), through different preclinical studies. Tinostamustine increases CD38 expression in myeloma cell lines, an effect that occurs in parallel with an increment in CD38 histone H3 acetylation levels. Also, the expression of MICA and MICB, ligands for the NK cell activating receptor NKG2D, augments after tinostamustine treatment in myeloma cell lines and primary myeloma cells. Pretreatment of myeloma cell lines with tinostamustine increased the sensitivity of these cells to daratumumab through its different cytotoxic mechanisms, and the combination of these two drugs showed a higher anti-myeloma effect than individual treatments in ex vivo cultures of myeloma patients' samples. In vivo data confirmed that tinostamustine pretreatment followed by daratumumab administration significantly delayed tumor growth and improved the survival of mice compared to individual treatments. In summary, our results suggest that tinostamustine could be a potential candidate to improve the efficacy of anti-CD38 mAbs.
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ADP-Ribosil Ciclasa 1 , Anticuerpos Monoclonales , Mieloma Múltiple , Subfamilia K de Receptores Similares a Lectina de Células NK , Animales , Humanos , Ratones , ADP-Ribosil Ciclasa 1/efectos de los fármacos , ADP-Ribosil Ciclasa 1/metabolismo , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Línea Celular Tumoral , Sinergismo Farmacológico , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase I/genética , Inhibidores de Histona Desacetilasas/farmacología , Inhibidores de Histona Desacetilasas/uso terapéutico , Glicoproteínas de Membrana/efectos de los fármacos , Glicoproteínas de Membrana/metabolismo , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/metabolismo , Mieloma Múltiple/patología , Subfamilia K de Receptores Similares a Lectina de Células NK/efectos de los fármacos , Subfamilia K de Receptores Similares a Lectina de Células NK/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Bencimidazoles/farmacología , Bencimidazoles/uso terapéuticoRESUMEN
In COVID-19, hyperinflammatory and dysregulated immune responses contribute to severity. Patients with pre-existing autoimmune conditions can therefore be at increased risk of severe COVID-19 and/or associated sequelae, yet SARS-CoV-2 infection in this group has been little studied. Here, we performed single-cell analysis of peripheral blood mononuclear cells from patients with three major autoimmune diseases (rheumatoid arthritis, psoriasis, or multiple sclerosis) during SARS-CoV-2 infection. We observed compositional differences between the autoimmune disease groups coupled with altered patterns of gene expression, transcription factor activity, and cell-cell communication that substantially shape the immune response under SARS-CoV-2 infection. While enrichment of HLA-DRlow CD14+ monocytes was observed in all three autoimmune disease groups, type-I interferon signaling as well as inflammatory T cell and monocyte responses varied widely between the three groups of patients. Our results reveal disturbed immune responses to SARS-CoV-2 in patients with pre-existing autoimmunity, highlighting important considerations for disease treatment and follow-up.
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Enfermedades Autoinmunes , COVID-19 , Humanos , SARS-CoV-2 , Leucocitos Mononucleares , Multiómica , Autoinmunidad , Análisis de la Célula IndividualRESUMEN
BACKGROUND: DNA methylation profiling of circulating cell-free DNA (cfDNA) has rapidly become a promising strategy for biomarker identification and development. The cell-type-specific nature of DNA methylation patterns and the direct relationship between cfDNA and apoptosis can potentially be used non-invasively to predict local alterations. In addition, direct detection of altered DNA methylation patterns performs well as a biomarker. In a previous study, we demonstrated marked DNA methylation alterations in brain tissue from patients with mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS). RESULTS: We performed DNA methylation profiling in cfDNA isolated from the serum of MTLE patients and healthy controls using BeadChip arrays followed by systematic bioinformatic analysis including deconvolution analysis and integration with DNase accessibility data sets. Differential cfDNA methylation analysis showed an overrepresentation of gene ontology terms and transcription factors related to central nervous system function and regulation. Deconvolution analysis of the DNA methylation data sets ruled out the possibility that the observed differences were due to changes in the proportional contribution of cortical neurons in cfDNA. Moreover, we found no overrepresentation of neuron- or glia-specific patterns in the described cfDNA methylation patterns. However, the MTLE-HS cfDNA methylation patterns featured a significant overrepresentation of the epileptic DNA methylation alterations previously observed in the hippocampus. CONCLUSIONS: Our results support the use of cfDNA methylation profiling as a rational approach to seeking non-invasive and reproducible epilepsy biomarkers.
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Ácidos Nucleicos Libres de Células , Epilepsia del Lóbulo Temporal , Humanos , Metilación de ADN , Hipocampo , Epilepsia del Lóbulo Temporal/genética , Encéfalo , Ácidos Nucleicos Libres de Células/genéticaRESUMEN
BACKGROUND: COVID-19 manifests with a wide spectrum of clinical phenotypes, ranging from asymptomatic and mild to severe and critical. Severe and critical COVID-19 patients are characterized by marked changes in the myeloid compartment, especially monocytes. However, little is known about the epigenetic alterations that occur in these cells during hyperinflammatory responses in severe COVID-19 patients. METHODS: In this study, we obtained the DNA methylome and transcriptome of peripheral blood monocytes from severe COVID-19 patients. DNA samples extracted from CD14 + CD15- monocytes of 48 severe COVID-19 patients and 11 healthy controls were hybridized on MethylationEPIC BeadChip arrays. In parallel, single-cell transcriptomics of 10 severe COVID-19 patients were generated. CellPhoneDB was used to infer changes in the crosstalk between monocytes and other immune cell types. RESULTS: We observed DNA methylation changes in CpG sites associated with interferon-related genes and genes associated with antigen presentation, concordant with gene expression changes. These changes significantly overlapped with those occurring in bacterial sepsis, although specific DNA methylation alterations in genes specific to viral infection were also identified. We also found these alterations to comprise some of the DNA methylation changes occurring during myeloid differentiation and under the influence of inflammatory cytokines. A progression of DNA methylation alterations in relation to the Sequential Organ Failure Assessment (SOFA) score was found to be related to interferon-related genes and T-helper 1 cell cytokine production. CellPhoneDB analysis of the single-cell transcriptomes of other immune cell types suggested the existence of altered crosstalk between monocytes and other cell types like NK cells and regulatory T cells. CONCLUSION: Our findings show the occurrence of an epigenetic and transcriptional reprogramming of peripheral blood monocytes, which could be associated with the release of aberrant immature monocytes, increased systemic levels of pro-inflammatory cytokines, and changes in immune cell crosstalk in these patients.
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COVID-19 , Monocitos , Humanos , Transcriptoma , Citocinas , COVID-19/genética , Interferones , Antivirales , Epigénesis GenéticaRESUMEN
Dendritic cells (DCs), the most potent antigen-presenting cells, are necessary for effective activation of naïve T cells. DCs' immunological properties are modulated in response to various stimuli. Active DNA demethylation is crucial for DC differentiation and function. Vitamin C, a known cofactor of ten-eleven translocation (TET) enzymes, drives active demethylation. Vitamin C has recently emerged as a promising adjuvant for several types of cancer; however, its effects on human immune cells are poorly understood. In this study, we investigate the epigenomic and transcriptomic reprogramming orchestrated by vitamin C in monocyte-derived DC differentiation and maturation. Vitamin C triggers extensive demethylation at NF-κB/p65 binding sites, together with concordant upregulation of antigen-presentation and immune response-related genes during DC maturation. p65 interacts with TET2 and mediates the aforementioned vitamin C-mediated changes, as demonstrated by pharmacological inhibition. Moreover, vitamin C increases TNFß production in DCs through NF-κB, in concordance with the upregulation of its coding gene and the demethylation of adjacent CpGs. Finally, vitamin C enhances DC's ability to stimulate the proliferation of autologous antigen-specific T cells. We propose that vitamin C could potentially improve monocyte-derived DC-based cell therapies.
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Ácido Ascórbico , Células Dendríticas , Epigénesis Genética , FN-kappa B , Humanos , Ácido Ascórbico/farmacología , Diferenciación Celular/genética , FN-kappa B/metabolismo , Linfocitos T/metabolismo , Reprogramación CelularRESUMEN
OBJECTIVES: Giant cell arteritis (GCA) is a complex systemic vasculitis mediated by the interplay between both genetic and epigenetic factors. Monocytes are crucial players of the inflammation occurring in GCA. Therefore, characterisation of the monocyte methylome and transcriptome in GCA would be helpful to better understand disease pathogenesis. METHODS: We performed an integrated epigenome-and transcriptome-wide association study in CD14+ monocytes from 82 patients with GCA, cross-sectionally classified into three different clinical statuses (active, in remission with or without glucocorticoid (GC) treatment), and 31 healthy controls. RESULTS: We identified a global methylation and gene expression dysregulation in GCA monocytes. Specifically, monocytes from active patients showed a more proinflammatory phenotype compared with healthy controls and patients in remission. In addition to inflammatory pathways known to be involved in active GCA, such as response to IL-6 and IL-1, we identified response to IL-11 as a new pathway potentially implicated in GCA. Furthermore, monocytes from patients in remission with treatment showed downregulation of genes involved in inflammatory processes as well as overexpression of GC receptor-target genes. Finally, we identified changes in DNA methylation correlating with alterations in expression levels of genes with a potential role in GCA pathogenesis, such as ITGA7 and CD63, as well as genes mediating the molecular response to GC, including FKBP5, ETS2, ZBTB16 and ADAMTS2. CONCLUSION: Our results revealed profound alterations in the methylation and transcriptomic profiles of monocytes from GCA patients, uncovering novel genes and pathways involved in GCA pathogenesis and in the molecular response to GC treatment.
RESUMEN
Common variable immunodeficiency (CVID), the most prevalent symptomatic primary immunodeficiency, displays impaired terminal B-cell differentiation and defective antibody responses. Incomplete genetic penetrance and ample phenotypic expressivity in CVID suggest the participation of additional pathogenic mechanisms. Monozygotic (MZ) twins discordant for CVID are uniquely valuable for studying the contribution of epigenetics to the disease. Here, we generate a single-cell epigenomics and transcriptomics census of naïve-to-memory B cell differentiation in a CVID-discordant MZ twin pair. Our analysis identifies DNA methylation, chromatin accessibility and transcriptional defects in memory B-cells mirroring defective cell-cell communication upon activation. These findings are validated in a cohort of CVID patients and healthy donors. Our findings provide a comprehensive multi-omics map of alterations in naïve-to-memory B-cell transition in CVID and indicate links between the epigenome and immune cell cross-talk. Our resource, publicly available at the Human Cell Atlas, gives insight into future diagnosis and treatments of CVID patients.
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Inmunodeficiencia Variable Común , Linfocitos B , Inmunodeficiencia Variable Común/diagnóstico , Inmunodeficiencia Variable Común/genética , Epigénesis Genética , Epigenómica , Centro Germinal , HumanosRESUMEN
Identifying predictive biomarkers at early stages of inflammatory arthritis is crucial for starting appropriate therapies to avoid poor outcomes. Monocytes (MOs) and macrophages, largely associated with arthritis, are contributors and sensors of inflammation through epigenetic modifications. In this study, we investigated associations between clinical features and DNA methylation in blood and synovial fluid (SF) MOs in a prospective cohort of patients with early inflammatory arthritis. DNA methylation profiles of undifferentiated arthritis (UA) blood MOs exhibited marked alterations in comparison with those from healthy donors. We identified additional differences both in blood and SF MOs after comparing patients with UA grouped by their future outcomes, i.e., good versus poor. Patient profiles in subsequent visits revealed a reversion toward a healthy level in both groups, those requiring disease-modifying antirheumatic drugs and those who remitted spontaneously. Changes in disease activity between visits also affected DNA methylation, which was partially concomitant in the SF of UA and in blood MOs of patients with rheumatoid arthritis. Epigenetic similarities between arthritis types allow a common prediction of disease activity. Our results constitute a resource of DNA methylation-based biomarkers of poor prognosis, disease activity, and treatment efficacy for the personalized clinical management of early inflammatory arthritis.
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Artritis Reumatoide , Epigenoma , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/genética , Biomarcadores , Humanos , Monocitos , Pronóstico , Estudios Prospectivos , Membrana SinovialRESUMEN
The active form of vitamin D, 1,25-dihydroxyvitamin D3, induces a stable tolerogenic phenotype in dendritic cells (DCs). This process involves the vitamin D receptor (VDR), which translocates to the nucleus, binds its cognate genomic sites, and promotes epigenetic and transcriptional remodeling. In this study, we report the occurrence of vitamin D-specific DNA demethylation and transcriptional activation at VDR binding sites associated with the acquisition of tolerogenesis in vitro. Differentiation to tolerogenic DCs associates with activation of the IL-6-JAK-STAT3 pathway. We show that JAK2-mediated STAT3 phosphorylation is specific to vitamin D stimulation. VDR and the phosphorylated form of STAT3 interact with each other to form a complex with methylcytosine dioxygenase TET2. Most importantly, pharmacological inhibition of JAK2 reverts vitamin D-induced tolerogenic properties of DCs. This interplay among VDR, STAT3, and TET2 opens up possibilities for modulating DC immunogenic properties in clinics.
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Proteínas de Unión al ADN/inmunología , Células Dendríticas/inmunología , Dioxigenasas/inmunología , Tolerancia Inmunológica/inmunología , Receptores de Calcitriol/inmunología , Factor de Transcripción STAT3/inmunología , Células Cultivadas , Proteínas de Unión al ADN/metabolismo , Células Dendríticas/metabolismo , Dioxigenasas/metabolismo , Humanos , Receptores de Calcitriol/metabolismo , Factor de Transcripción STAT3/metabolismoRESUMEN
Mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS) is the most common focal epilepsy in adults. It is characterized by alarming rates of pharmacoresistance. Epileptogenesis is associated with the occurrence of epigenetic alterations, and the few epigenetic studies carried out in MTLE-HS have mainly focused on the hippocampus. In this study, we obtained the DNA methylation profiles from both the hippocampus and anterior temporal neocortex of MTLE-HS patients subjected to resective epilepsy surgery and autopsied non-epileptic controls. We assessed the progressive nature of DNA methylation changes in relation to epilepsy duration. We identified significantly altered hippocampal DNA methylation patterns encompassing multiple pathways known to be involved in epileptogenesis. DNA methylation changes were even more striking in the neocortex, wherein pathogenic pathways and genes were common to both tissues. Most importantly, DNA methylation changes at many genomic sites varied significantly with epilepsy duration. Such progressive changes were associated with inflammation-related genes in the hippocampus. Our results suggest that the neocortex, relatively spared of extensive histopathological damage, may also be involved in epilepsy development. These results also open the possibility that the observed neocortical impairment could represent a preliminary stage of epileptogenesis before the establishment of chronic lesions or a consequence of prolonged seizure exposure. Our two-tissue multi-level characterization of the MTLE-HS DNA methylome suggests the occurrence of a self-propagating inflammatory wave of epigenetic dysregulation.
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Epilepsia del Lóbulo Temporal , Epilepsia , Adulto , Metilación de ADN/genética , Epilepsia del Lóbulo Temporal/genética , Hipocampo/patología , Humanos , Esclerosis/complicaciones , Esclerosis/patologíaRESUMEN
Glucocorticoids (GCs) exert potent anti-inflammatory effects in immune cells through the glucocorticoid receptor (GR). Dendritic cells (DCs), central actors for coordinating immune responses, acquire tolerogenic properties in response to GCs. Tolerogenic DCs (tolDCs) have emerged as a potential treatment for various inflammatory diseases. To date, the underlying cell type-specific regulatory mechanisms orchestrating GC-mediated acquisition of immunosuppressive properties remain poorly understood. In this study, we investigated the transcriptomic and epigenomic remodeling associated with differentiation to DCs in the presence of GCs. Our analysis demonstrates a major role of MAFB in this process, in synergy with GR. GR and MAFB both interact with methylcytosine dioxygenase TET2 and bind to genomic loci that undergo specific demethylation in tolDCs. We also show that the role of MAFB is more extensive, binding to thousands of genomic loci in tolDCs. Finally, MAFB knockdown erases the tolerogenic properties of tolDCs and reverts the specific DNA demethylation and gene upregulation. The preeminent role of MAFB is also demonstrated in vivo for myeloid cells from synovium in rheumatoid arthritis following GC treatment. Our results imply that, once directly activated by GR, MAFB plays a critical role in orchestrating the epigenomic and transcriptomic remodeling that define the tolerogenic phenotype.
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Células Dendríticas/inmunología , Epigénesis Genética , Tolerancia Inmunológica , Factor de Transcripción MafB/metabolismo , Receptores de Glucocorticoides/metabolismo , Adulto , Células Cultivadas , Metilación de ADN , Proteínas de Unión al ADN/metabolismo , Dioxigenasas/metabolismo , Femenino , Humanos , Factor de Transcripción MafB/genética , Masculino , Persona de Mediana EdadRESUMEN
Microbial challenges, such as widespread bacterial infection in sepsis, induce endotoxin tolerance, a state of hyporesponsiveness to subsequent infections. The participation of DNA methylation in this process is poorly known. In this study, we perform integrated analysis of DNA methylation and transcriptional changes following in vitro exposure to gram-negative bacterial lipopolysaccharide, together with analysis of ex vivo monocytes from septic patients. We identify TET2-mediated demethylation and transcriptional activation of inflammation-related genes that is specific to toll-like receptor stimulation. Changes also involve phosphorylation of STAT1, STAT3 and STAT5, elements of the JAK2 pathway. JAK2 pathway inhibition impairs the activation of tolerized genes on the first encounter with lipopolysaccharide. We then confirm the implication of the JAK2-STAT pathway in the aberrant DNA methylome of patients with sepsis caused by gram-negative bacteria. Finally, JAK2 inhibition in monocytes partially recapitulates the expression changes produced in the immunosuppressive cellular state acquired by monocytes from gram-negative sepsis, as described by single cell-RNA-sequencing. Our study evidences both the crucial role the JAK2-STAT pathway in epigenetic regulation and initial response of the tolerized genes to gram-negative bacterial endotoxins and provides a pharmacological target to prevent exacerbated responses.
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Tolerancia a Endotoxinas/genética , Bacterias Gramnegativas/inmunología , Infecciones por Bacterias Gramnegativas/genética , Infecciones por Bacterias Gramnegativas/inmunología , Monocitos/inmunología , Monocitos/microbiología , Sepsis/genética , Sepsis/inmunología , Estudios de Casos y Controles , Metilación de ADN/genética , Metilación de ADN/inmunología , Tolerancia a Endotoxinas/efectos de los fármacos , Tolerancia a Endotoxinas/inmunología , Endotoxinas/toxicidad , Epigénesis Genética , Femenino , Infecciones por Bacterias Gramnegativas/microbiología , Humanos , Técnicas In Vitro , Janus Quinasa 2/antagonistas & inhibidores , Janus Quinasa 2/genética , Janus Quinasa 2/inmunología , Lipopolisacáridos/toxicidad , Masculino , Monocitos/efectos de los fármacos , Factores de Transcripción STAT/genética , Factores de Transcripción STAT/inmunología , Sepsis/microbiología , Transducción de Señal/genética , Transducción de Señal/inmunología , Receptor Toll-Like 2/inmunología , Receptor Toll-Like 4/inmunologíaRESUMEN
Activation-induced deaminase (AID) initiates antibody diversification in germinal center B cells by deaminating cytosines, leading to somatic hypermutation and class-switch recombination. Loss-of-function mutations in AID lead to hyper-IgM syndrome type 2 (HIGM2), a rare human primary antibody deficiency. AID-mediated deamination has been proposed as leading to active demethylation of 5-methycytosines in the DNA, although evidence both supports and casts doubt on such a role. In this study, using whole-genome bisulfite sequencing of HIGM2 B cells, we investigated direct AID involvement in active DNA demethylation. HIGM2 naïve and memory B cells both display widespread DNA methylation alterations, of which â¼25% are attributable to active DNA demethylation. For genes that undergo active demethylation that is impaired in HIGM2 individuals, our analysis indicates that AID is not directly involved. We demonstrate that the widespread alterations in the DNA methylation and expression profiles of HIGM2 naïve B cells result from premature overstimulation of the B-cell receptor prior to the germinal center reaction. Our data support a role for AID in B cell central tolerance in preventing the expansion of autoreactive cell clones, affecting the correct establishment of DNA methylation patterns.
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Linfocitos B/inmunología , Citidina Desaminasa/fisiología , Metilación de ADN , Síndrome de Inmunodeficiencia con Hiper-IgM/genética , Síndrome de Inmunodeficiencia con Hiper-IgM/inmunología , Autoinmunidad , Linfocitos B/metabolismo , Citidina Desaminasa/deficiencia , Citidina Desaminasa/genética , Centro Germinal/inmunología , Humanos , Síndrome de Inmunodeficiencia con Hiper-IgM/metabolismo , Tolerancia Inmunológica , Memoria Inmunológica , Receptores de Antígenos de Linfocitos B/genética , Transcriptoma , Secuenciación Completa del GenomaRESUMEN
Multiple myeloma (MM) progression and myeloma-associated bone disease (MBD) are highly dependent on bone marrow mesenchymal stromal cells (MSCs). MM-MSCs exhibit abnormal transcriptomes, suggesting the involvement of epigenetic mechanisms governing their tumor-promoting functions and prolonged osteoblast suppression. Here, we identify widespread DNA methylation alterations of bone marrow-isolated MSCs from distinct MM stages, particularly in Homeobox genes involved in osteogenic differentiation that associate with their aberrant expression. Moreover, these DNA methylation changes are recapitulated in vitro by exposing MSCs from healthy individuals to MM cells. Pharmacological targeting of DNMTs and G9a with dual inhibitor CM-272 reverts the expression of hypermethylated osteogenic regulators and promotes osteoblast differentiation of myeloma MSCs. Most importantly, CM-272 treatment prevents tumor-associated bone loss and reduces tumor burden in a murine myeloma model. Our results demonstrate that epigenetic aberrancies mediate the impairment of bone formation in MM, and its targeting by CM-272 is able to reverse MBD.
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Antineoplásicos/farmacología , Enfermedades Óseas/tratamiento farmacológico , Metilación de ADN/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Células Madre Mesenquimatosas/efectos de los fármacos , Mieloma Múltiple/tratamiento farmacológico , Adulto , Anciano , Anciano de 80 o más Años , Animales , Antineoplásicos/uso terapéutico , Enfermedades Óseas/diagnóstico , Enfermedades Óseas/genética , Enfermedades Óseas/patología , Médula Ósea/patología , ADN (Citosina-5-)-Metiltransferasas/antagonistas & inhibidores , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Inhibidores Enzimáticos/uso terapéutico , Epigénesis Genética/efectos de los fármacos , Femenino , Fémur/diagnóstico por imagen , Fémur/patología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Antígenos de Histocompatibilidad/metabolismo , N-Metiltransferasa de Histona-Lisina/antagonistas & inhibidores , N-Metiltransferasa de Histona-Lisina/metabolismo , Humanos , Masculino , Células Madre Mesenquimatosas/patología , Ratones , Persona de Mediana Edad , Mieloma Múltiple/complicaciones , Mieloma Múltiple/genética , Mieloma Múltiple/patología , Osteogénesis/efectos de los fármacos , Osteogénesis/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Systemic sclerosis (SSc) is a genetically complex autoimmune disease mediated by the interplay between genetic and epigenetic factors in a multitude of immune cells, with CD4+ T lymphocytes as one of the principle drivers of pathogenesis. METHODS: DNA samples exacted from CD4+ T cells of 48 SSc patients and 16 healthy controls were hybridized on MethylationEPIC BeadChip array. In parallel, gene expression was interrogated by hybridizing total RNA on Clariom™ S array. Downstream bioinformatics analyses were performed to identify correlating differentially methylated CpG positions (DMPs) and differentially expressed genes (DEGs), which were then confirmed utilizing previously published promoter capture Hi-C (PCHi-C) data. RESULTS: We identified 9112 and 3929 DMPs and DEGs, respectively. These DMPs and DEGs are enriched in functional categories related to inflammation and T cell biology. Furthermore, correlation analysis identified 17,500 possible DMP-DEG interaction pairs within a window of 5 Mb, and utilizing PCHi-C data, we observed that 212 CD4+ T cell-specific pairs of DMP-DEG also formed part of three-dimensional promoter-enhancer networks, potentially involving CTCF. Finally, combining PCHi-C data with SSc GWAS data, we identified four important SSc-associated susceptibility loci, TNIP1 (rs3792783), GSDMB (rs9303277), IL12RB1 (rs2305743), and CSK (rs1378942), that could potentially interact with DMP-DEG pairs cg17239269-ANXA6, cg19458020-CCR7, cg10808810-JUND, and cg11062629-ULK3, respectively. CONCLUSION: Our study unveils a potential link between genetic, epigenetic, and transcriptional deregulation in CD4+ T cells of SSc patients, providing a novel integrated view of molecular components driving SSc pathogenesis.
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Linfocitos T CD4-Positivos/metabolismo , Epigenoma , Sitios Genéticos , Predisposición Genética a la Enfermedad , Esclerodermia Sistémica/etiología , Transcriptoma , Linfocitos T CD4-Positivos/inmunología , Biología Computacional/métodos , Metilación de ADN , Epigénesis Genética , Epigenómica/métodos , Perfilación de la Expresión Génica/métodos , Estudios de Asociación Genética , Humanos , FN-kappa B/metabolismo , Esclerodermia Sistémica/metabolismo , Transducción de SeñalRESUMEN
Sirtuins 1 and 2 (SIRT1/2) are two NAD-dependent deacetylases with major roles in inflammation. In addition to deacetylating histones and other proteins, SIRT1/2-mediated regulation is coupled with other epigenetic enzymes. Here, we investigate the links between SIRT1/2 activity and DNA methylation in macrophage differentiation due to their relevance in myeloid cells. SIRT1/2 display drastic upregulation during macrophage differentiation and their inhibition impacts the expression of many inflammation-related genes. In this context, SIRT1/2 inhibition abrogates DNA methylation gains, but does not affect demethylation. Inhibition of hypermethylation occurs at many inflammatory loci, which results in more drastic upregulation of their expression upon macrophage polarization following bacterial lipopolysaccharide (LPS) challenge. SIRT1/2-mediated gains of methylation concur with decreases in activating histone marks, and their inhibition revert these histone marks to resemble an open chromatin. Remarkably, specific inhibition of DNA methyltransferases is sufficient to upregulate inflammatory genes that are maintained in a silent state by SIRT1/2. Both SIRT1 and SIRT2 directly interact with DNMT3B, and their binding to proinflammatory genes is lost upon exposure to LPS or through pharmacological inhibition of their activity. In all, we describe a novel role for SIRT1/2 to restrict premature activation of proinflammatory genes.
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Metilación de ADN/genética , Inflamación/genética , Sirtuina 1/genética , Sirtuina 2/genética , Acetilación , Diferenciación Celular/genética , Cromatina/genética , Regulación de la Expresión Génica/genética , Histonas/genética , Humanos , Inflamación/inducido químicamente , Inflamación/patología , Lipopolisacáridos/toxicidad , Macrófagos/metabolismo , Regiones Promotoras Genéticas , Activación Transcripcional/genéticaRESUMEN
BACKGROUND: Sepsis, a life-threatening organ dysfunction caused by a dysregulated systemic immune response to infection, associates with reduced responsiveness to subsequent infections. How such tolerance is acquired is not well understood but is known to involve epigenetic and transcriptional dysregulation. METHODS: Bead arrays were used to compare global DNA methylation changes in patients with sepsis, non-infectious systemic inflammatory response syndrome, and healthy controls. Bioinformatic analyses were performed to dissect functional reprogramming and signaling pathways related to the acquisition of these specific DNA methylation alterations. Finally, in vitro experiments using human monocytes were performed to test the induction of similar DNA methylation reprogramming. RESULTS: Here, we focused on DNA methylation changes associated with sepsis, given their potential role in stabilizing altered phenotypes. Tolerized monocytes from patients with sepsis display changes in their DNA methylomes with respect to those from healthy controls, affecting critical monocyte-related genes. DNA methylation profiles correlate with IL-10 and IL-6 levels, significantly increased in monocytes in sepsis, as well as with the Sequential Organ Failure Assessment score; the observed changes associate with TFs and pathways downstream to toll-like receptors and inflammatory cytokines. In fact, in vitro stimulation of toll-like receptors in monocytes results in similar gains and losses of methylation together with the acquisition of tolerance. CONCLUSION: We have identified a DNA methylation signature associated with sepsis that is downstream to the response of monocytes to inflammatory signals associated with the acquisition of a tolerized phenotype and organic dysfunction.
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Citocinas/genética , Metilación de ADN , ADN/análisis , Mediadores de Inflamación/metabolismo , Inflamación/complicaciones , Monocitos/patología , Insuficiencia Multiorgánica/complicaciones , Sepsis/diagnóstico , Anciano , Estudios de Casos y Controles , ADN/genética , Femenino , Humanos , Inflamación/genética , Masculino , Persona de Mediana Edad , Monocitos/inmunología , Monocitos/metabolismo , Insuficiencia Multiorgánica/genética , Fenotipo , Sepsis/etiología , Sepsis/metabolismo , Transducción de SeñalRESUMEN
OBJECTIVE: Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease that mainly targets joints. Monocytes and macrophages are critical in RA pathogenesis and contribute to inflammatory lesions. These extremely plastic cells respond to extracellular signals which cause epigenomic changes that define their pathogenic phenotype. Here, we interrogated how DNA methylation alterations in RA monocytes are determined by extracellular signals. METHODS: High-throughput DNA methylation analyses of patients with RA and controls and in vitro cytokine stimulation were used to investigate the underlying mechanisms behind DNA methylation alterations in RA as well as their relationship with clinical parameters, including RA disease activity. RESULTS: The DNA methylomes of peripheral blood monocytes displayed significant changes and increased variability in patients with RA with respect to healthy controls. Changes in the monocyte methylome correlate with DAS28, in which high-activity patients are divergent from healthy controls in contrast to remission patients whose methylome is virtually identical to healthy controls. Indeed, the notion of a changing monocyte methylome is supported after comparing the profiles of same individuals at different stages of activity. We show how these changes are mediated by an increase in disease activity-associated cytokines, such as tumour necrosis factor alpha and interferons, as they recapitulate the DNA methylation changes observed in patients in vitro. CONCLUSION: We demonstrate a direct link between RA disease activity and the monocyte methylome through the action of inflammation-associated cytokines. Finally, we have obtained a DNA methylation-based mathematical formula that predicts inflammation-mediated disease activity for RA and other chronic immune-mediated inflammatory diseases.
Asunto(s)
Artritis Reumatoide/sangre , Artritis Reumatoide/genética , Citocinas/sangre , Epigenoma/inmunología , Mediadores de Inflamación/sangre , Biomarcadores/sangre , Metilación de ADN/inmunología , Humanos , Leucocitos Mononucleares/inmunología , Macrófagos/inmunología , Factor de Necrosis Tumoral alfa/sangreRESUMEN
Activation-induced cytidine deaminase (AID) triggers antibody diversification in B cells by catalysing deamination and subsequently mutating immunoglobulin (Ig) genes. Association of AID with RNA Pol II and occurrence of epigenetic changes during Ig gene diversification suggest participation of AID in epigenetic regulation. AID is mutated in hyper-IgM type 2 (HIGM2) syndrome. Here, we investigated the potential role of AID in the acquisition of epigenetic changes. We discovered that AID binding to the IgH locus promotes an increase in H4K20me3. In 293F cells, we demonstrate interaction between co-transfected AID and the three SUV4-20 histone H4K20 methyltransferases, and that SUV4-20H1.2, bound to the IgH switch (S) mu site, is replaced by SUV4-20H2 upon AID binding. Analysis of HIGM2 mutants shows that the AID truncated form W68X is impaired to interact with SUV4-20H1.2 and SUV4-20H2 and is unable to bind and target H4K20me3 to the Smu site. We finally show in mouse primary B cells undergoing class-switch recombination (CSR) that AID deficiency associates with decreased H4K20me3 levels at the Smu site. Our results provide a novel link between SUV4-20 enzymes and CSR and offer a new aspect of the interplay between AID and histone modifications in setting the epigenetic status of CSR sites.