RESUMEN
Germinal center (GC) B cells undergo proliferation at very high rates in a hypoxic microenvironment but the cellular processes driving this are incompletely understood. Here we show that the mitochondria of GC B cells are highly dynamic, with significantly upregulated transcription and translation rates associated with the activity of transcription factor A, mitochondrial (TFAM). TFAM, while also necessary for normal B cell development, is required for entry of activated GC precursor B cells into the germinal center reaction; deletion of Tfam significantly impairs GC formation, function and output. Loss of TFAM in B cells compromises the actin cytoskeleton and impairs cellular motility of GC B cells in response to chemokine signaling, leading to their spatial disorganization. We show that B cell lymphoma substantially increases mitochondrial translation and that deletion of Tfam in B cells is protective against the development of lymphoma in a c-Myc transgenic mouse model. Finally, we show that pharmacological inhibition of mitochondrial transcription and translation inhibits growth of GC-derived human lymphoma cells and induces similar defects in the actin cytoskeleton.
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Linfoma de Células B , Linfoma , Ratones , Humanos , Animales , Linfocitos B/patología , Centro Germinal/patología , Transcripción Genética , Linfoma de Células B/genética , Linfoma de Células B/patología , Ratones Transgénicos , Microambiente TumoralRESUMEN
NP105-113-B*07:02-specific CD8+ T cell responses are considered among the most dominant in SARS-CoV-2-infected individuals. We found strong association of this response with mild disease. Analysis of NP105-113-B*07:02-specific T cell clones and single-cell sequencing were performed concurrently, with functional avidity and antiviral efficacy assessed using an in vitro SARS-CoV-2 infection system, and were correlated with T cell receptor usage, transcriptome signature and disease severity (acute n = 77, convalescent n = 52). We demonstrated a beneficial association of NP105-113-B*07:02-specific T cells in COVID-19 disease progression, linked with expansion of T cell precursors, high functional avidity and antiviral effector function. Broad immune memory pools were narrowed postinfection but NP105-113-B*07:02-specific T cells were maintained 6 months after infection with preserved antiviral efficacy to the SARS-CoV-2 Victoria strain, as well as Alpha, Beta, Gamma and Delta variants. Our data show that NP105-113-B*07:02-specific T cell responses associate with mild disease and high antiviral efficacy, pointing to inclusion for future vaccine design.
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Antígeno HLA-B7/inmunología , Epítopos Inmunodominantes/inmunología , Proteínas de la Nucleocápside/inmunología , SARS-CoV-2/inmunología , Linfocitos T Citotóxicos/inmunología , Anciano , Secuencia de Aminoácidos , Anticuerpos Antivirales/inmunología , Afinidad de Anticuerpos/inmunología , COVID-19/inmunología , COVID-19/patología , Línea Celular Transformada , Femenino , Perfilación de la Expresión Génica , Humanos , Memoria Inmunológica/inmunología , Masculino , Persona de Mediana Edad , Receptores de Antígenos de Linfocitos T/inmunología , Índice de Severidad de la Enfermedad , Virus Vaccinia/genética , Virus Vaccinia/inmunología , Virus Vaccinia/metabolismoRESUMEN
Intestinal mesenchymal cells play essential roles in epithelial homeostasis, matrix remodeling, immunity, and inflammation. But the extent of heterogeneity within the colonic mesenchyme in these processes remains unknown. Using unbiased single-cell profiling of over 16,500 colonic mesenchymal cells, we reveal four subsets of fibroblasts expressing divergent transcriptional regulators and functional pathways, in addition to pericytes and myofibroblasts. We identified a niche population located in proximity to epithelial crypts expressing SOX6, F3 (CD142), and WNT genes essential for colonic epithelial stem cell function. In colitis, we observed dysregulation of this niche and emergence of an activated mesenchymal population. This subset expressed TNF superfamily member 14 (TNFSF14), fibroblastic reticular cell-associated genes, IL-33, and Lysyl oxidases. Further, it induced factors that impaired epithelial proliferation and maturation and contributed to oxidative stress and disease severity in vivo. Our work defines how the colonic mesenchyme remodels to fuel inflammation and barrier dysfunction in IBD.
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Enfermedades Inflamatorias del Intestino/fisiopatología , Mesodermo/fisiología , Animales , Proliferación Celular , Colitis/genética , Colitis/fisiopatología , Colon/fisiología , Células Epiteliales/metabolismo , Fibroblastos/fisiología , Heterogeneidad Genética , Homeostasis , Humanos , Inflamación , Mucosa Intestinal/inmunología , Mucosa Intestinal/fisiología , Intestinos/inmunología , Intestinos/fisiología , Células Madre Mesenquimatosas/fisiología , Mesodermo/metabolismo , Ratones , Ratones Endogámicos C57BL , Miofibroblastos , Pericitos , Células RAW 264.7 , Factores de Transcripción SOXD/fisiología , Análisis de la Célula Individual/métodos , Tromboplastina/fisiología , Miembro 14 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/genética , Vía de Señalización Wnt/fisiologíaRESUMEN
Resident memory B (BRM) cells develop and persist in the lungs of influenza-infected mice and humans; however, their contribution to recall responses has not been defined. Here, we used two-photon microscopy to visualize BRM cells within the lungs of influenza -virus immune and reinfected mice. Prior to re-exposure, BRM cells were sparsely scattered throughout the tissue, displaying limited motility. Within 24 h of rechallenge, these cells increased their migratory capacity, localized to infected sites, and subsequently differentiated into plasma cells. Alveolar macrophages mediated this process, in part by inducing expression of chemokines CXCL9 and CXCL10 from infiltrating inflammatory cells. This led to the recruitment of chemokine receptor CXCR3-expressing BRM cells to infected regions and increased local antibody concentrations. Our study uncovers spatiotemporal mechanisms that regulate lung BRM cell reactivation and demonstrates their capacity to rapidly deliver antibodies in a highly localized manner to sites of viral replication.
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Gripe Humana , Infecciones por Orthomyxoviridae , Orthomyxoviridae , Animales , Anticuerpos , Humanos , Memoria Inmunológica , Células B de Memoria , RatonesRESUMEN
The identification of lymphocyte subsets with non-overlapping effector functions has been pivotal to the development of targeted therapies in immune-mediated inflammatory diseases (IMIDs)1,2. However, it remains unclear whether fibroblast subclasses with non-overlapping functions also exist and are responsible for the wide variety of tissue-driven processes observed in IMIDs, such as inflammation and damage3-5. Here we identify and describe the biology of distinct subsets of fibroblasts responsible for mediating either inflammation or tissue damage in arthritis. We show that deletion of fibroblast activation protein-α (FAPα)+ fibroblasts suppressed both inflammation and bone erosions in mouse models of resolving and persistent arthritis. Single-cell transcriptional analysis identified two distinct fibroblast subsets within the FAPα+ population: FAPα+THY1+ immune effector fibroblasts located in the synovial sub-lining, and FAPα+THY1- destructive fibroblasts restricted to the synovial lining layer. When adoptively transferred into the joint, FAPα+THY1- fibroblasts selectively mediate bone and cartilage damage with little effect on inflammation, whereas transfer of FAPα+ THY1+ fibroblasts resulted in a more severe and persistent inflammatory arthritis, with minimal effect on bone and cartilage. Our findings describing anatomically discrete, functionally distinct fibroblast subsets with non-overlapping functions have important implications for cell-based therapies aimed at modulating inflammation and tissue damage.
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Artritis Reumatoide/patología , Fibroblastos/patología , Animales , Huesos/patología , Endopeptidasas , Femenino , Fibroblastos/clasificación , Fibroblastos/metabolismo , Gelatinasas/metabolismo , Humanos , Inflamación/patología , Articulaciones/patología , Masculino , Proteínas de la Membrana/metabolismo , Ratones , RNA-Seq , Serina Endopeptidasas/metabolismo , Análisis de la Célula Individual , Membrana Sinovial/patología , Antígenos Thy-1/metabolismoRESUMEN
Thymic epithelial cells (TEC) control the selection of a T cell repertoire reactive to pathogens but tolerant of self. This process is known to involve the promiscuous expression of virtually the entire protein-coding gene repertoire, but the extent to which TEC recapitulate peripheral isoforms, and the mechanisms by which they do so, remain largely unknown. We performed the first assembly-based transcriptomic census of transcript structures and splicing factor (SF) expression in mouse medullary TEC (mTEC) and 21 peripheral tissues. Mature mTEC expressed 60.1% of all protein-coding transcripts, more than was detected in any of the peripheral tissues. However, for genes with tissue-restricted expression, mTEC produced fewer isoforms than did the relevant peripheral tissues. Analysis of exon inclusion revealed an absence of brain-specific microexons in mTEC. We did not find unusual numbers of novel transcripts in TEC, and we show that Aire, the facilitator of promiscuous gene expression, promotes the generation of long "classical" transcripts (with 5' and 3' UTRs) but has only a limited impact on alternative splicing in mTEC. Comprehensive assessment of SF expression in mTEC identified a small set of nonpromiscuously expressed SF genes, among which we confirmed RBFOX to be present with AIRE in mTEC nuclei. Using a conditional loss-of-function approach, we show that Rbfox2 promotes mTEC development and regulates the alternative splicing of promiscuously expressed genes. These data indicate that TEC recommission a small number of peripheral SFs, including members of the RBFOX family, to generate a broad but selective representation of the peripheral splice isoform repertoire.
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Perfilación de la Expresión Génica , Empalme del ARN , Animales , Diferenciación Celular/genética , Células Epiteliales/metabolismo , Ratones , Ratones Endogámicos C57BL , Factores de Empalme de ARN/genética , Factores de Empalme de ARN/metabolismo , Timo/metabolismo , TranscriptomaRESUMEN
Parkinson's disease (PD) is the second most common neurodegenerative disorder and a central role for α-synuclein (αSyn; SNCA) in disease aetiology has been proposed based on genetics and neuropathology. To better understand the pathological mechanisms of αSyn, we generated induced pluripotent stem cells (iPSCs) from healthy individuals and PD patients carrying the A53T SNCA mutation or a triplication of the SNCA locus and differentiated them into dopaminergic neurons (DAns). iPSC-derived DAn from PD patients carrying either mutation showed increased intracellular αSyn accumulation, and DAns from patients carrying the SNCA triplication displayed oligomeric αSyn pathology and elevated αSyn extracellular release. Transcriptomic analysis of purified DAns revealed perturbations in expression of genes linked to mitochondrial function, consistent with observed reduction in mitochondrial respiration, impairment in mitochondrial membrane potential, aberrant mitochondrial morphology and decreased levels of phosphorylated DRP1Ser616. Parkinson's iPSC-derived DAns showed increased endoplasmic reticulum stress and impairments in cholesterol and lipid homeostasis. Together, these data show a correlation between αSyn cellular pathology and deficits in metabolic and cellular bioenergetics in the pathology of PD.
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Neuronas Dopaminérgicas/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Mitocondrias/metabolismo , Enfermedad de Parkinson/genética , alfa-Sinucleína/genética , Diferenciación Celular , Dinaminas/metabolismo , Estrés del Retículo Endoplásmico/genética , Metabolismo Energético/genética , Humanos , Metabolismo de los Lípidos/genética , Potencial de la Membrana Mitocondrial , Mitocondrias/ultraestructura , Mutación , Enfermedad de Parkinson/metabolismo , RNA-Seq , Sinucleinopatías/metabolismo , alfa-Sinucleína/metabolismoRESUMEN
RNA-seq is the standard method for profiling gene expression in many biological systems. Due to the wide dynamic range and complex nature of the transcriptome, RNA-seq provides an incomplete characterization, especially of lowly expressed genes and transcripts. Targeted RNA sequencing (RNA CaptureSeq) focuses sequencing on genes of interest, providing exquisite sensitivity for transcript detection and quantification. However, uses of CaptureSeq have focused on bulk samples and its performance on very small populations of cells is unknown. Here we show CaptureSeq greatly enhances transcriptomic profiling of target genes in ultra-low-input samples and provides equivalent performance to that on bulk samples. We validate the performance of CaptureSeq using multiple probe sets on samples of iPSC-derived cortical neurons. We demonstrate up to 275-fold enrichment for target genes, the detection of 10% additional genes and a greater than 5-fold increase in identified gene isoforms. Analysis of spike-in controls demonstrated CaptureSeq improved both detection sensitivity and expression quantification. Comparison to the CORTECON database of cerebral cortex development revealed CaptureSeq enhanced the identification of sample differentiation stage. CaptureSeq provides sensitive, reliable and quantitative expression measurements on hundreds-to-thousands of target genes from ultra-low-input samples and has the potential to greatly enhance transcriptomic profiling when samples are limiting.
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Perfilación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Análisis de Secuencia de ARN , Transcriptoma , Diferenciación Celular/genética , Biología Computacional/métodos , Perfilación de la Expresión Génica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Neuronas/citología , Neuronas/metabolismo , Análisis de Secuencia de ARN/métodos , Factores de Transcripción/metabolismoRESUMEN
Frozen shoulder is a spontaneously self-resolving chronic inflammatory fibrotic human disease, which distinguishes the condition from most fibrotic diseases that are progressive and irreversible. Using single-cell analysis, we identify pro-inflammatory MERTKlowCD48+ macrophages and MERTK + LYVE1 + MRC1+ macrophages enriched for negative regulators of inflammation which co-exist in frozen shoulder capsule tissues. Micro-cultures of patient-derived cells identify integrin-mediated cell-matrix interactions between MERTK+ macrophages and pro-resolving DKK3+ and POSTN+ fibroblasts, suggesting that matrix remodelling plays a role in frozen shoulder resolution. Cross-tissue analysis reveals a shared gene expression cassette between shoulder capsule MERTK+ macrophages and a respective population enriched in synovial tissues of rheumatoid arthritis patients in disease remission, supporting the concept that MERTK+ macrophages mediate resolution of inflammation and fibrosis. Single-cell transcriptomic profiling and spatial analysis of human foetal shoulder tissues identify MERTK + LYVE1 + MRC1+ macrophages and DKK3+ and POSTN+ fibroblast populations analogous to those in frozen shoulder, suggesting that the template to resolve fibrosis is established during shoulder development. Crosstalk between MerTK+ macrophages and pro-resolving DKK3+ and POSTN+ fibroblasts could facilitate resolution of frozen shoulder, providing a basis for potential therapeutic resolution of persistent fibrotic diseases.
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Bursitis , Humanos , Tirosina Quinasa c-Mer/metabolismo , Inflamación/metabolismo , Membrana Sinovial/metabolismo , FibrosisRESUMEN
BACKGROUND: The Nextera protocol, which utilises a transposome based approach to create libraries for Illumina sequencing, requires pure DNA template, an accurate assessment of input concentration and a column clean-up that limits its applicability for high-throughput sample preparation. We addressed the identified limitations to develop a robust workflow that supports both rapid and high-throughput projects also reducing reagent costs. RESULTS: We show that an initial bead-based normalisation step can remove the need for quantification and improves sample purity. A 75% cost reduction was achieved with a low-volume modified protocol which was tested over genomes with different GC content to demonstrate its robustness. Finally we developed a custom set of index tags and primers which increase the number of samples that can simultaneously be sequenced on a single lane of an Illumina instrument. CONCLUSIONS: We addressed the bottlenecks of Nextera library construction to produce a modified protocol which harnesses the full power of the Nextera kit and allows the reproducible construction of libraries on a high-throughput scale reducing the associated cost of the kit.
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Cartilla de ADN/aislamiento & purificación , Biblioteca de Genes , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Automatización de Laboratorios , Clostridioides difficile/genética , Cartilla de ADN/genética , Análisis de Secuencia de ADN/métodos , Flujo de TrabajoRESUMEN
IFNγ is an immune mediator with concomitant pro- and anti-tumor functions. Here, we provide evidence that IFNγ directly acts on intra-tumoral CD8 T cells to restrict anti-tumor responses. We report that expression of the IFNγ receptor ß chain (IFNγR2) in CD8 T cells negatively correlates with clinical responsiveness to checkpoint blockade in metastatic melanoma patients, suggesting that the loss of sensitivity to IFNγ contributes to successful antitumor immunity. Indeed, specific deletion of IFNγR in CD8 T cells promotes tumor control in a mouse model of melanoma. Chronic IFNγ inhibits the maintenance, clonal diversity and proliferation of stem-like T cells. This leads to decreased generation of T cells with intermediate expression of exhaustion markers, previously associated with beneficial anti-tumor responses. This study provides evidence of a negative feedback loop whereby IFNγ depletes stem-like T cells to restrict anti-tumor immunity. Targeting this pathway might represent an alternative strategy to enhance T cell-based therapies.
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Melanoma , Linfocitos T Citotóxicos , Ratones , Animales , Linfocitos T Citotóxicos/metabolismo , Linfocitos T CD8-positivos , Melanoma/terapia , Melanoma/tratamiento farmacológico , Células Clonales/metabolismoRESUMEN
The first immune-activating changes within joint resident cells that lead to pathogenic leukocyte recruitment during articular inflammation remain largely unknown. In this study, we employ state-of-the-art confocal microscopy and image analysis in a systemic, whole-organ, and quantitative way to present evidence that synovial inflammation begins with the activation of lining macrophages. We show that lining, but not sublining macrophages phagocytose immune complexes containing the model antigen. Using the antigen-induced arthritis (AIA) model, we demonstrate that on recognition of antigen-antibody complexes, lining macrophages undergo significant activation, which is dependent on interferon regulatory factor 5 (IRF5), and produce chemokines, most notably CXCL1. Consequently, at the onset of inflammation, neutrophils are preferentially recruited in the vicinity of antigen-laden macrophages in the synovial lining niche. As inflammation progresses, neutrophils disperse across the whole synovium and form swarms in synovial sublining during resolution. Our study alters the paradigm of lining macrophages as immunosuppressive cells to important instigators of synovial inflammation.
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Artritis , Humanos , Infiltración Neutrófila , Artritis/patología , Macrófagos , Membrana Sinovial/patología , Inflamación/patología , AntígenosRESUMEN
[This corrects the article DOI: 10.1038/s44161-023-00295-x.].
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The immune system is integral to cardiovascular health and disease. Targeting inflammation ameliorates adverse cardiovascular outcomes. Atherosclerosis, a major underlying cause of cardiovascular disease (CVD), is conceptualised as a lipid-driven inflammation where macrophages play a non-redundant role. However, evidence emerging so far from single cell atlases suggests a dichotomy between lipid associated and inflammatory macrophage states. Here, we present an inclusive reference atlas of human intraplaque immune cell communities. Combining scRNASeq of human surgical carotid endarterectomies in a discovery cohort with bulk RNASeq and immunohistochemistry in a validation cohort (the Carotid Plaque Imaging Project-CPIP), we reveal the existence of PLIN2hi/TREM1hi macrophages as a toll-like receptor-dependent inflammatory lipid-associated macrophage state linked to cerebrovascular events. Our study shifts the current paradigm of lipid-driven inflammation by providing biological evidence for a pathogenic macrophage transition to an inflammatory lipid-associated phenotype and for its targeting as a new treatment strategy for CVD.
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Salmonella enterica represent a major disease burden worldwide. S. enterica serovar Typhi (S. Typhi) is responsible for potentially life-threatening Typhoid fever affecting 10.9 million people annually. While non-typhoidal Salmonella (NTS) serovars usually trigger self-limiting diarrhoea, invasive NTS bacteraemia is a growing public health challenge. Dendritic cells (DCs) are key professional antigen presenting cells of the human immune system. The ability of pathogenic bacteria to subvert DC functions and prevent T cell recognition contributes to their survival and dissemination within the host. Here, we adapted dual RNA-sequencing to define how different Salmonella pathovariants remodel their gene expression in tandem with that of infected DCs. We find DCs harness iron handling pathways to defend against invading Salmonellas, which S. Typhi is able to circumvent by mounting a robust response to nitrosative stress. In parallel, we uncover the alternative strategies invasive NTS employ to impair DC functions.
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Reprogramación Celular/fisiología , Células Dendríticas/metabolismo , Salmonella enterica/clasificación , Células Dendríticas/inmunología , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , MutaciónRESUMEN
The immunological synapse is a molecular hub that facilitates the delivery of three activation signals, namely antigen, costimulation/corepression and cytokines, from antigen-presenting cells (APC) to T cells. T cells release a fourth class of signaling entities, trans-synaptic vesicles (tSV), to mediate bidirectional communication. Here we present bead-supported lipid bilayers (BSLB) as versatile synthetic APCs to capture, characterize and advance the understanding of tSV biogenesis. Specifically, the integration of juxtacrine signals, such as CD40 and antigen, results in the adaptive tailoring and release of tSV, which differ in size, yields and immune receptor cargo compared with steadily released extracellular vesicles (EVs). Focusing on CD40L+ tSV as model effectors, we show that PD-L1 trans-presentation together with TSG101, ADAM10 and CD81 are key in determining CD40L vesicular release. Lastly, we find greater RNA-binding protein and microRNA content in tSV compared with EVs, supporting the specialized role of tSV as intercellular messengers.
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Ligando de CD40 , Vesículas Extracelulares , Ligando de CD40/metabolismo , Vesículas Extracelulares/metabolismo , Sinapsis Inmunológicas , Vesículas Sinápticas , Linfocitos TRESUMEN
Current inflammatory bowel disease (IBD) therapies are ineffective in a high proportion of patients. Combining bulk and single-cell transcriptomics, quantitative histopathology and in situ localization across three cohorts of patients with IBD (total n = 376), we identify coexpressed gene modules within the heterogeneous tissular inflammatory response in IBD that map to distinct histopathological and cellular features (pathotypes). One of these pathotypes is defined by high neutrophil infiltration, activation of fibroblasts and vascular remodeling at sites of deep ulceration. Activated fibroblasts in the ulcer bed display neutrophil-chemoattractant properties that are IL-1R, but not TNF, dependent. Pathotype-associated neutrophil and fibroblast signatures are increased in nonresponders to several therapies across four independent cohorts (total n = 343). The identification of distinct, localized, tissular pathotypes will aid precision targeting of current therapeutics and provides a biological rationale for IL-1 signaling blockade in ulcerating disease.
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Enfermedades Inflamatorias del Intestino/patología , Interleucina-1/metabolismo , Infiltración Neutrófila/inmunología , Neutrófilos/inmunología , Células del Estroma/inmunología , Adulto , Anciano , Femenino , Fibroblastos/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/genética , Masculino , Persona de Mediana Edad , Receptores de Interleucina-1/metabolismo , Transducción de Señal/fisiología , Remodelación Vascular/fisiologíaRESUMEN
Mononuclear phagocytes (MNPs) are vital for maintaining intestinal homeostasis but, in response to acute microbial stimulation, can also trigger immunopathology, accelerating recruitment of Ly6Chi monocytes to the gut. The regulators that control monocyte tissue adaptation in the gut remain poorly understood. Interferon regulatory factor 5 (IRF5) is a transcription factor previously shown to play a key role in maintaining the inflammatory phenotype of macrophages. Here, we investigate the impact of IRF5 on the MNP system and physiology of the gut at homeostasis and during inflammation. We demonstrate that IRF5 deficiency has a limited impact on colon physiology at steady state but ameliorates immunopathology during Helicobacter hepaticus-induced colitis. Inhibition of IRF5 activity in MNPs phenocopies global IRF5 deficiency. Using a combination of bone marrow chimera and single-cell RNA-sequencing approaches, we examined the intrinsic role of IRF5 in controlling colonic MNP development. We demonstrate that IRF5 promotes differentiation of Ly6Chi monocytes into CD11c+ macrophages and controls the production of antimicrobial and inflammatory mediators by these cells. Thus, we identify IRF5 as a key transcriptional regulator of the colonic MNP system during intestinal inflammation.
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Antígenos CD11/inmunología , Inflamación/inmunología , Factores Reguladores del Interferón/inmunología , Macrófagos/inmunología , Monocitos/inmunología , Animales , Helicobacter hepaticus/inmunología , Inflamación/patología , Factores Reguladores del Interferón/deficiencia , Macrófagos/patología , Ratones , Ratones Noqueados , Monocitos/patología , FenotipoRESUMEN
Immune-regulatory mechanisms of drug-free remission in rheumatoid arthritis (RA) are unknown. We hypothesized that synovial tissue macrophages (STM), which persist in remission, contribute to joint homeostasis. We used single-cell transcriptomics to profile 32,000 STMs and identified phenotypic changes in patients with early/active RA, treatment-refractory/active RA and RA in sustained remission. Each clinical state was characterized by different frequencies of nine discrete phenotypic clusters within four distinct STM subpopulations with diverse homeostatic, regulatory and inflammatory functions. This cellular atlas, combined with deep-phenotypic, spatial and functional analyses of synovial biopsy fluorescent activated cell sorted STMs, revealed two STM subpopulations (MerTKposTREM2high and MerTKposLYVE1pos) with unique remission transcriptomic signatures enriched in negative regulators of inflammation. These STMs were potent producers of inflammation-resolving lipid mediators and induced the repair response of synovial fibroblasts in vitro. A low proportion of MerTKpos STMs in remission was associated with increased risk of disease flare after treatment cessation. Therapeutic modulation of MerTKpos STM subpopulations could therefore be a potential treatment strategy for RA.
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Artritis Reumatoide/metabolismo , Inflamación/metabolismo , Macrófagos/inmunología , Líquido Sinovial/metabolismo , Artritis Reumatoide/genética , Artritis Reumatoide/inmunología , Artritis Reumatoide/patología , Biopsia , Linaje de la Célula/genética , Humanos , Inflamación/genética , Inflamación/inmunología , Inflamación/patología , Articulaciones/inmunología , Articulaciones/metabolismo , Articulaciones/patología , Lectinas Tipo C/genética , Lectinas Tipo C/inmunología , Macrófagos/metabolismo , Receptor de Manosa , Lectinas de Unión a Manosa/genética , Lectinas de Unión a Manosa/inmunología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/inmunología , Receptores Inmunológicos/genética , Receptores Inmunológicos/inmunología , Líquido Sinovial/inmunología , Membrana SinovialRESUMEN
The capsular group B meningococcal (MenB) four component vaccine (4CMenB) has been licensed for the prevention of invasive disease caused by MenB. The vaccine causes fever in infants, particularly when given in combination (concomitant) with other routinely-administered vaccines (routine), such as the standard diphtheria, tetanus, pertussis (DTP)-containing vaccine. To assess the suitability of a mouse immunisation model to study this phenomenon, we monitored temperature in mice after a second dose of routine vaccines, with or without 4CMenB, and compared the results with those in humans. Using this mouse model, we explored the reactogenicity of 4CMenB components by measuring changes in temperature, cytokines, and gene expression induced by 4CMenB, one of its components, wild-type or attenuated endotoxin outer membrane vesicles (OMVs), or lipopolysaccharide (LPS). A significant rise (p < 0.01) in temperature was observed in mice immunised with 4CMenB, wild-type OMVs, and LPS. RNA-sequencing of mouse whole blood revealed a gene signature shared by the 4CMenB, OMV, and LPS groups consisting of bacterial pattern recognition receptors and neutrophil activation marker genes. Sequencing of neutrophils isolated after concomitant 4CMenB identified cells expressing the OMV-associated genes Plek and Lcp1. Immunisation with 4CMenB or OMVs led to increased IL-6 in serum and significant upregulation (p < 0.0001) of prostaglandin-synthesising enzymes on brain tissue. These data demonstrate the suitability of a mouse model for assessing vaccine reactogenicity and strongly indicate that the fever following vaccination with 4CMenB in human infants is induced by endotoxin contained in the OMV component of the vaccine.