RESUMEN
Inflammatory bowel disease is a chronic, relapsing condition with two subtypes, Crohn's disease (CD) and ulcerative colitis (UC). Genome-wide association studies (GWASs) in UC implicate a FCGR2A variant that alters the binding affinity of the antibody receptor it encodes, FcγRIIA, for immunoglobulin G (IgG). Here, we aimed to understand the mechanisms whereby changes in FcγRIIA affinity would affect inflammation in an IgA-dominated organ. We found a profound induction of anti-commensal IgG and a concomitant increase in activating FcγR signaling in the colonic mucosa of UC patients. Commensal-IgG immune complexes engaged gut-resident FcγR-expressing macrophages, inducing NLRP3- and reactive-oxygen-species-dependent production of interleukin-1ß (IL-1ß) and neutrophil-recruiting chemokines. These responses were modulated by the FCGR2A genotype. In vivo manipulation of macrophage FcγR signal strength in a mouse model of UC determined the magnitude of intestinal inflammation and IL-1ß-dependent type 17 immunity. The identification of an important contribution of IgG-FcγR-dependent inflammation to UC has therapeutic implications.
Asunto(s)
Anticuerpos Antibacterianos/inmunología , Colitis Ulcerosa/inmunología , Microbioma Gastrointestinal/inmunología , Inmunoglobulina G/inmunología , Interleucina-1beta/inmunología , Células Th17/inmunología , Animales , Colitis/inducido químicamente , Colitis/inmunología , Colitis/microbiología , Colitis/patología , Colitis Ulcerosa/microbiología , Colitis Ulcerosa/patología , Sulfato de Dextran/toxicidad , Regulación de la Expresión Génica , Genotipo , Humanos , Inflamación , Interleucina-8/biosíntesis , Interleucina-8/genética , Mucosa Intestinal/inmunología , Mucosa Intestinal/microbiología , Macrófagos/inmunología , Ratones , Fagocitos/inmunología , ARN Mensajero/biosíntesis , Especies Reactivas de Oxígeno , Receptores de IgG/biosíntesis , Receptores de IgG/genética , Receptores de IgG/inmunologíaRESUMEN
OBJECTIVE: Epigenetic mechanisms, including DNA methylation (DNAm), have been proposed to play a key role in Crohn's disease (CD) pathogenesis. However, the specific cell types and pathways affected as well as their potential impact on disease phenotype and outcome remain unknown. We set out to investigate the role of intestinal epithelial DNAm in CD pathogenesis. DESIGN: We generated 312 intestinal epithelial organoids (IEOs) from mucosal biopsies of 168 patients with CD (n=72), UC (n=23) and healthy controls (n=73). We performed genome-wide molecular profiling including DNAm, bulk as well as single-cell RNA sequencing. Organoids were subjected to gene editing and the functional consequences of DNAm changes evaluated using an organoid-lymphocyte coculture and a nucleotide-binding oligomerisation domain, leucine-rich repeat and CARD domain containing 5 (NLRC5) dextran sulphate sodium (DSS) colitis knock-out mouse model. RESULTS: We identified highly stable, CD-associated loss of DNAm at major histocompatibility complex (MHC) class 1 loci including NLRC5 and cognate gene upregulation. Single-cell RNA sequencing of primary mucosal tissue and IEOs confirmed the role of NLRC5 as transcriptional transactivator in the intestinal epithelium. Increased mucosal MHC-I and NLRC5 expression in adult and paediatric patients with CD was validated in additional cohorts and the functional role of MHC-I highlighted by demonstrating a relative protection from DSS-mediated mucosal inflammation in NLRC5-deficient mice. MHC-I DNAm in IEOs showed a significant correlation with CD disease phenotype and outcomes. Application of machine learning approaches enabled the development of a disease prognostic epigenetic molecular signature. CONCLUSIONS: Our study has identified epigenetically regulated intestinal epithelial MHC-I as a novel mechanism in CD pathogenesis.
Asunto(s)
Enfermedad de Crohn , Metilación de ADN , Epigénesis Genética , Mucosa Intestinal , Organoides , Humanos , Enfermedad de Crohn/genética , Enfermedad de Crohn/patología , Enfermedad de Crohn/metabolismo , Organoides/metabolismo , Organoides/patología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Ratones , Animales , Femenino , Masculino , Ratones Noqueados , Bancos de Muestras Biológicas , Adulto , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/metabolismo , Modelos Animales de Enfermedad , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismoRESUMEN
IgG antibodies cause inflammation and organ damage in autoimmune diseases such as systemic lupus erythematosus (SLE). We investigated the metabolic profile of macrophages isolated from inflamed tissues in immune complex (IC)-associated diseases, including SLE and rheumatoid arthritis, and following IgG Fcγ receptor cross-linking. We found that human and mouse macrophages undergo a switch to glycolysis in response to IgG IC stimulation, mirroring macrophage metabolic changes in inflamed tissue in vivo. This metabolic reprogramming was required to generate a number of proinflammatory mediators, including IL-1ß, and was dependent on mTOR and hypoxia-inducible factor (HIF)1α. Inhibition of glycolysis, or genetic depletion of HIF1α, attenuated IgG IC-induced activation of macrophages in vitro, including primary human kidney macrophages. In vivo, glycolysis inhibition led to a reduction in kidney macrophage IL-1ß and reduced neutrophil recruitment in a murine model of antibody-mediated nephritis. Together, our data reveal the molecular mechanisms underpinning FcγR-mediated metabolic reprogramming in macrophages and suggest a therapeutic strategy for autoantibody-induced inflammation, including lupus nephritis.
Asunto(s)
Reprogramación Celular/fisiología , Nefritis Lúpica/metabolismo , Animales , Células Cultivadas , Dinoprostona/genética , Dinoprostona/metabolismo , Metabolismo Energético , Regulación de la Expresión Génica , Glucólisis/fisiología , Humanos , Inmunoglobulina G/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Riñón/citología , Macrófagos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Especies Reactivas de Oxígeno , Receptores de IgG/genética , Receptores de IgG/metabolismoRESUMEN
The gastrointestinal tract is colonized by trillions of commensal microorganisms that collectively form the microbiome and make essential contributions to organism homeostasis. The intestinal immune system must tolerate these beneficial commensals, whilst preventing pathogenic organisms from systemic spread. Humoral immunity plays a key role in this process, with large quantities of immunoglobulin (Ig)A secreted into the lumen on a daily basis, regulating the microbiome and preventing bacteria from encroaching on the epithelium. However, there is an increasing appreciation of the role of IgG antibodies in intestinal immunity, including beneficial effects in neonatal immune development, pathogen and tumour resistance, but also of pathological effects in driving chronic inflammation in inflammatory bowel disease (IBD). These antibody isotypes differ in effector function, with IgG exhibiting more proinflammatory capabilities compared with IgA. Therefore, the process that leads to the generation of different antibody isotypes, class-switch recombination (CSR), requires careful regulation and is orchestrated by the immunological cues generated by the prevalent local challenge. In general, an initiating signal such as CD40 ligation on B cells leads to the induction of activation-induced cytidine deaminase (AID), but a second cytokine-mediated signal determines which Ig heavy chain is expressed. Whilst the cytokines driving intestinal IgA responses are well-studied, there is less clarity on how IgG responses are generated in the intestine, and how these cues might become dysfunctional in IBD. Here, we review the key mechanisms regulating class switching to IgA vs IgG in the intestine, processes that could be therapeutically manipulated in infection and IBD.
Asunto(s)
Inmunoglobulina A/inmunología , Cambio de Clase de Inmunoglobulina/inmunología , Inmunoglobulina G/inmunología , Intestinos/inmunología , Linfocitos B/inmunología , Células Cultivadas , Citidina Desaminasa/metabolismo , Citocinas/metabolismo , Microbioma Gastrointestinal/inmunología , Humanos , Inmunidad Humoral/inmunologíaRESUMEN
Transplantation is the optimal treatment for most patients with end-stage kidney disease but organ shortage is a major challenge. Normothermic machine perfusion (NMP) has been used to recondition marginal organs; however, mechanisms by which NMP might benefit organs are not well understood. Using pairs of human kidneys obtained from the same donor, we compared the effect of NMP with that of cold storage on the global kidney transcriptome. We found that cold storage led to a global reduction in gene expression, including inflammatory pathway genes and those required for energy generation processes, such as oxidative phosphorylation (OXPHOS). In contrast, during NMP, there was marked upregulation OXPHOS genes, but also of a number of immune and inflammatory pathway genes. Using biopsies from kidneys undergoing NMP that were subsequently transplanted, we found that higher inflammatory gene expression occurred in organs with prolonged delayed graft function (DGF). Therefore, we used a hemoadsorber (HA) to remove pro-inflammatory cytokines. This attenuated inflammatory gene expression increased OXPHOS pathway genes and had potentially clinically important effects in reducing the expression of a DGF-associated gene signature. Together, our data suggest that adsorption of pro-inflammatory mediators from the perfusate represents a potential intervention which may improve organ viability.
Asunto(s)
Funcionamiento Retardado del Injerto , Trasplante de Riñón , Citocinas/genética , Funcionamiento Retardado del Injerto/genética , Supervivencia de Injerto , Humanos , Riñón , Preservación de Órganos , Perfusión , Donantes de TejidosRESUMEN
Information about lipid-protein interactions for G protein-coupled receptors (GPCRs) is scarce. Here, we use electron spin resonance (ESR) and spin-labelled lipids to study lipid interactions with the rat neurotensin receptor 1 (NTS1). A fusion protein containing rat NTS1 fully able to bind its ligand neurotensin was reconstituted into phosphatidylcholine (PC) bilayers at specific lipid:protein molar ratios. The fraction of motionally restricted lipids in the range of 40:1 to 80:1 lipids per receptor suggested an oligomeric state of the protein, and the result was unaffected by increasing the hydrophobic thickness of the lipid bilayer from C-18 to C-20 or C-22 chain length PC membranes. Comparison of the ESR spectra of different spin-labelled lipids allowed direct measurement of lipid binding constants relative to PC (Kr), with spin-labelled phosphatidylethanolamine (PESL), phosphatidylserine (PSSL), stearic acid (SASL), and a spin labelled cholesterol analogue (CSL) Kr values of 1.05±0.05, 1.92±0.08, 5.20±0.51 and 0.91±0.19, respectively. The results contrast with those from rhodopsin, the only other GPCR studied this way, which has no selectivity for the lipids analysed here. Molecular dynamics simulations of NTS1 in bilayers are in agreement with the ESR data, and point to sites in the receptor where PS could interact with higher affinity. Lipid selectivity could be necessary for regulation of ligand binding, oligomerisation and/or G protein activation processes. Our results provide insight into the potential modulatory mechanisms that lipids can exert on GPCRs.
Asunto(s)
Lípidos/química , Receptores de Neurotensina/metabolismo , Animales , Espectroscopía de Resonancia por Spin del Electrón , Interacciones Hidrofóbicas e Hidrofílicas , Membrana Dobles de Lípidos , Simulación de Dinámica Molecular , Unión Proteica , Ratas , Receptores de Neurotensina/químicaRESUMEN
A role for vitamin D in immune modulation and in cancer has been suggested. In this work, we report that mice with increased availability of vitamin D display greater immune-dependent resistance to transplantable cancers and augmented responses to checkpoint blockade immunotherapies. Similarly, in humans, vitamin D-induced genes correlate with improved responses to immune checkpoint inhibitor treatment as well as with immunity to cancer and increased overall survival. In mice, resistance is attributable to the activity of vitamin D on intestinal epithelial cells, which alters microbiome composition in favor of Bacteroides fragilis, which positively regulates cancer immunity. Our findings indicate a previously unappreciated connection between vitamin D, microbial commensal communities, and immune responses to cancer. Collectively, they highlight vitamin D levels as a potential determinant of cancer immunity and immunotherapy success.
Asunto(s)
Bacteroides fragilis , Microbioma Gastrointestinal , Inhibidores de Puntos de Control Inmunológico , Neoplasias , Vitamina D , Animales , Femenino , Humanos , Masculino , Ratones , Bacteroides fragilis/metabolismo , Microbioma Gastrointestinal/efectos de los fármacos , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inhibidores de Puntos de Control Inmunológico/farmacología , Inmunoterapia , Mucosa Intestinal/inmunología , Mucosa Intestinal/microbiología , Mucosa Intestinal/metabolismo , Ratones Endogámicos C57BL , Neoplasias/inmunología , Neoplasias/microbiología , Neoplasias/terapia , Vitamina D/administración & dosificación , Vitamina D/metabolismo , Dieta , Línea Celular Tumoral , Calcifediol/administración & dosificación , Calcifediol/metabolismo , Proteína de Unión a Vitamina D/genética , Proteína de Unión a Vitamina D/metabolismoRESUMEN
Bladder infection affects a hundred million people annually, but our understanding of bladder immunity is incomplete. We found type 17 immune response genes among the most up-regulated networks in mouse bladder following uropathogenic Escherichia coli (UPEC) challenge. Intravital imaging revealed submucosal Rorc+ cells responsive to UPEC challenge, and we found increased Il17 and IL22 transcripts in wild-type and Rag2 -/- mice, implicating group 3 innate lymphoid cells (ILC3s) as a source of these cytokines. NCR-positive and negative ILC3 subsets were identified in murine and human bladders, with local proliferation increasing IL17-producing ILC3s post infection. ILC3s made a more limited contribution to bladder IL22, with prominent early induction of IL22 evident in Th17 cells. Single-cell RNA sequencing revealed bladder NCR-negative ILC3s as the source of IL17 and identified putative ILC3-myeloid cell interactions, including via lymphotoxin-ß-LTBR. Altogether, our data provide important insights into the orchestration and execution of type 17 immunity in bladder defense.
RESUMEN
B cells, which are critical for intestinal homeostasis, remain understudied in ulcerative colitis (UC). In this study, we recruited three cohorts of patients with UC (primary cohort, n = 145; validation cohort 1, n = 664; and validation cohort 2, n = 143) to comprehensively define the landscape of B cells during UC-associated intestinal inflammation. Using single-cell RNA sequencing, single-cell IgH gene sequencing and protein-level validation, we mapped the compositional, transcriptional and clonotypic landscape of mucosal and circulating B cells. We found major perturbations within the mucosal B cell compartment, including an expansion of naive B cells and IgG+ plasma cells with curtailed diversity and maturation. Furthermore, we isolated an auto-reactive plasma cell clone targeting integrin αvß6 from inflamed UC intestines. We also identified a subset of intestinal CXCL13-expressing TFH-like T peripheral helper cells that were associated with the pathogenic B cell response. Finally, across all three cohorts, we confirmed that changes in intestinal humoral immunity are reflected in circulation by the expansion of gut-homing plasmablasts that correlates with disease activity and predicts disease complications. Our data demonstrate a highly dysregulated B cell response in UC and highlight a potential role of B cells in disease pathogenesis.
Asunto(s)
Colitis Ulcerosa , Células Plasmáticas , Linfocitos B , Colitis Ulcerosa/genética , Humanos , Mucosa Intestinal/patología , Recuento de Linfocitos , Linfocitos T Colaboradores-InductoresRESUMEN
B cells are critical to immune homeostasis at mucosal surfaces including those of the gastrointestinal tract. B cell-related abnormalities, comprising of a lympho-plasmacytic infiltrate, as well as anti-microbial antibodies, are well reported in patients with inflammatory bowel disease (IBD). However, B cell-targeting is not part of the therapeutic armamentarium in IBD. Recently, driven by the identification of genetic associations between IgG Fc receptors and IBD susceptibility, there has been renewed interest in defining the immunobiology of B cells during mucosal inflammation. Functional studies have demonstrated mechanisms of IgG-mediated disease pathogenesis and deep mucosal immunophenotyping using single cell RNA sequencing has elaborated a significant remodelling of the B cell compartment in IBD. In light of these novel data, here we discuss potential strategies to target B cell immunity in IBD. Finally, we discuss potential risks and pitfalls of these approaches and emphasize on distinguishing between homeostatic and pathological B cell signatures, allowing for a data-based, prudent therapeutic approach.
Asunto(s)
Linfocitos B/inmunología , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/inmunología , Animales , HumanosRESUMEN
Immunoglobulins (Igs) form a cornerstone of mucosal immunity. In the gastrointestinal tract, secretory IgA and IgM bind to commensal microorganisms within the intestinal lumen to prevent them from breaching the intestinal epithelium - a process known as immune exclusion. In recent years, there has been renewed interest in the role of IgG in intestinal immunity, driven in part by a genetic association of an affinity-lowering variant of an IgG receptor, FcγRIIA, with protection from ulcerative colitis (UC), a subclass of inflammatory bowel disease (IBD). We recently demonstrated a role for IgG and Fcγ receptor signalling in driving pathogenic IL-1ß production by colonic mononuclear phagocytes and the subsequent induction of a local type 17 response in UC. Here, we discuss the potential relevance of our observations to the other major subclass of IBD - Crohn's disease (CD) - where the genetic association with FCGR variants is less robust and consider how this may impact therapeutic interventions in these disease subsets.
Asunto(s)
Colitis Ulcerosa/inmunología , Enfermedad de Crohn/inmunología , Inmunoglobulina G/inmunología , Mucosa Intestinal/inmunología , Especificidad de Anticuerpos , Colitis Ulcerosa/genética , Colitis Ulcerosa/microbiología , Enfermedad de Crohn/genética , Enfermedad de Crohn/microbiología , Microbioma Gastrointestinal , Glicosilación , Humanos , Inmunidad Mucosa , Inmunoglobulina G/metabolismo , Receptores de IgG/genética , Receptores de IgG/metabolismoRESUMEN
Macrophages play a central role in intestinal immunity, but inappropriate macrophage activation is associated with inflammatory bowel disease (IBD). Here, we identify granulocyte-macrophage colony stimulating factor (GM-CSF) as a critical regulator of intestinal macrophage activation in patients with IBD and mice with dextran sodium sulfate (DSS)-induced colitis. We find that GM-CSF drives the maturation and polarization of inflammatory intestinal macrophages, promoting anti-microbial functions while suppressing wound-healing transcriptional programs. Group 3 innate lymphoid cells (ILC3s) are a major source of GM-CSF in intestinal inflammation, with a strong positive correlation observed between ILC or CSF2 transcripts and M1 macrophage signatures in IBD mucosal biopsies. Furthermore, GM-CSF-dependent macrophage polarization results in a positive feedback loop that augmented ILC3 activation and type 17 immunity. Together, our data reveal an important role for GM-CSF-mediated ILC-macrophage crosstalk in calibrating intestinal macrophage phenotype to enhance anti-bacterial responses, while inhibiting pro-repair functions associated with fibrosis and stricturing, with important clinical implications.
Asunto(s)
Infecciones por Enterobacteriaceae/patología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Inflamación/patología , Intestinos/patología , Macrófagos/patología , Cicatrización de Heridas , Animales , Polaridad Celular , Citrobacter rodentium/fisiología , Colitis/complicaciones , Colitis/inmunología , Colitis/patología , Humanos , Inmunidad Innata , Linfocitos/inmunología , Activación de Macrófagos , Ratones Endogámicos C57BL , FenotipoRESUMEN
Fcγ receptors (FcγR) are cell surface glycoproteins that mediate cellular effector functions of immunoglobulin G (IgG) antibodies. Genetic variation in FcγR genes can influence susceptibility to a variety of antibody-mediated autoimmune and inflammatory disorders, including systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). More recently, however, genetic studies have implicated altered FcγR signaling in the pathogenesis of inflammatory bowel disease (IBD), a condition classically associated with dysregulated innate and T cell immunity. Specifically, a variant of the activating receptor, FcγRIIA, with low affinity for IgG, confers protection against the development of ulcerative colitis, a subset of IBD, leading to a re-evaluation of the role of IgG and FcγRs in gastrointestinal tract immunity, an organ system traditionally associated with IgA. In this review, we summarize our current understanding of IgG and FcγR function at this unique host-environment interface, from the pathogenesis of colitis and defense against enteropathogens, its contribution to maternal-fetal cross-talk and susceptibility to cancer. Finally, we discuss the therapeutic implications of this information, both in terms of how FcγR signaling pathways may be targeted for the treatment of IBD and how FcγR engagement may influence the efficacy of therapeutic monoclonal antibodies in IBD.
Asunto(s)
Susceptibilidad a Enfermedades , Inflamación/etiología , Inflamación/metabolismo , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Factores de Edad , Animales , Autoinmunidad , Biomarcadores , Manejo de la Enfermedad , Susceptibilidad a Enfermedades/inmunología , Tracto Gastrointestinal/inmunología , Tracto Gastrointestinal/metabolismo , Tracto Gastrointestinal/patología , Humanos , Inmunoglobulina G/inmunología , Inflamación/patología , Inflamación/terapia , Enfermedades Inflamatorias del Intestino/etiología , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Terapia Molecular Dirigida , Unión Proteica , Receptores de IgG/metabolismo , Transducción de SeñalRESUMEN
In the current era, one of the major factors limiting graft survival is chronic antibody-mediated rejection (ABMR), whilst patient survival is impacted by the effects of immunosuppression on susceptibility to infection, malignancy and atherosclerosis. IgG antibodies play a role in all of these processes, and many of their cellular effects are mediated by Fc gamma receptors (FcγRs). These surface receptors are expressed by most immune cells, including B cells, natural killer cells, dendritic cells and macrophages. Genetic variation in FCGR genes is likely to affect susceptibility to ABMR and to modulate the physiological functions of IgG. In this review, we discuss the potential role played by FcγRs in determining outcomes in solid organ transplantation, and how genetic polymorphisms in these receptors may contribute to variations in transplant outcome.