RESUMEN
Human polyomaviruses cause a common childhood infection worldwide and typically elicit a neutralizing antibody and cellular immune response, while establishing a dormant infection in the kidney with minimal clinical manifestations. However, viral reactivation can cause severe pathology in immunocompromised individuals. We developed a high-throughput, functional antibody screen to examine the humoral response to BK polyomavirus. This approach enabled the isolation of antibodies from all peripheral B cell subsets and revealed the anti-BK virus antibody repertoire as clonally complex with respect to immunoglobulin sequences and isotypes (both IgM and IgG), including a high frequency of monoclonal antibodies that broadly neutralize BK virus subtypes and the related JC polyomavirus. Cryo-electron microscopy of a broadly neutralizing IgG single-chain variable fragment complexed with BK virus-like particles revealed the quaternary nature of a conserved viral epitope at the junction between capsid pentamers. These features unravel a potent modality for inhibiting polyomavirus infection in kidney transplant recipients and other immunocompromised patients.
Asunto(s)
Anticuerpos Neutralizantes/inmunología , Linfocitos B/inmunología , Virus BK/inmunología , Memoria Inmunológica/inmunología , Virus JC/inmunología , Infecciones por Polyomavirus/inmunología , Poliomavirus/inmunología , Anticuerpos Monoclonales/inmunología , Anticuerpos Antivirales/inmunología , Cápside/inmunología , Línea Celular , Epítopos/inmunología , Células HEK293 , Humanos , Inmunidad Celular/inmunología , Riñón/inmunologíaRESUMEN
Reactivation of BK polyomavirus (BKPyV) can cause significant kidney and bladder disease in immunocompromised patients. There are currently no effective, BKPyV-specific therapies. MAU868 is a novel, human immunoglobulin (Ig) G1 monoclonal antibody that binds the major capsid protein, VP1, of BKPyV with picomolar affinity, neutralizes infection by the 4 major BKPyV genotypes (EC50 ranging from 0.009-0.093 µg/mL; EC90 ranging from 0.102-4.160 µg/mL), and has comparable activity against variants with highly prevalent VP1 polymorphisms. No resistance-associated variants were identified in long-term selection studies, indicating a high in vitro barrier-to-resistance. The high-resolution crystal structure of MAU868 in complex with VP1 pentamer identified 3 key contact residues in VP1 (Y169, R170, and K172). A first-in-human study was conducted to assess the safety, tolerability, and pharmacokinetics of MAU868 following intravenous and subcutaneous administration to healthy adults in a randomized, placebo-controlled, double-blinded, single ascending dose design. MAU868 was safe and well-tolerated. All adverse events were grade 1 and resolved. The pharmacokinetics of MAU868 was typical of a human IgG, with dose-proportional systemic exposure and an elimination half-life ranging between 23 and 30 days. These results demonstrate the potential of MAU868 as a first-in-class therapeutic agent for the treatment or prevention of BKPyV disease.
RESUMEN
Microbial colonization of the gut induces the development of gut-associated lymphoid tissue (GALT). The molecular mechanisms that regulate GALT function and result in gut-commensal homeostasis are poorly defined. T follicular helper (Tfh) cells in Peyer's patches (PPs) promote high-affinity IgA responses. Here we found that the ATP-gated ionotropic P2X7 receptor controls Tfh cell numbers in PPs. Lack of P2X7 in Tfh cells enhanced germinal center reactions and high-affinity IgA secretion and binding to commensals. The ensuing depletion of mucosal bacteria resulted in reduced systemic translocation of microbial components, lowering B1 cell stimulation and serum IgM concentrations. Mice lacking P2X7 had increased susceptibility to polymicrobial sepsis, which was rescued by Tfh cell depletion or administration of purified IgM. Thus, regulation of Tfh cells by P2X7 activity is important for mucosal colonization, which in turn results in IgM serum concentrations necessary to protect the host from bacteremia.
Asunto(s)
Mucosa Intestinal/inmunología , Ganglios Linfáticos Agregados/inmunología , Receptores Purinérgicos P2X7/inmunología , Simbiosis/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Adenosina Trifosfato/metabolismo , Animales , Linfocitos B/inmunología , Bacteriemia/inmunología , Predisposición Genética a la Enfermedad , Centro Germinal/inmunología , Humanos , Inmunoglobulina A/inmunología , Inmunoglobulina M/sangre , Mucosa Intestinal/microbiología , Depleción Linfocítica , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microbiota/inmunología , Ganglios Linfáticos Agregados/citología , Receptores Purinérgicos P2X7/genética , Sepsis/inmunología , Sepsis/microbiologíaRESUMEN
Adenosine deaminase 2 deficiency (DADA2) is an autoinflammatory disease characterized by inflammatory vasculopathy, early strokes associated often with hypogammaglobulinemia. Pure red cell aplasia, thrombocytopenia, and neutropenia have been reported. The defect is due to biallelic loss of function of ADA2 gene, coding for a protein known to regulate the catabolism of extracellular adenosine. We therefore investigated immune phenotype and B- and T-cell responses in 14 DADA2 patients to address if ADA2 mutation affects B- and T-cell function. Here, we show a significant decrease in memory B cells, in particular class switch memory, and an expansion of CD21low B cells in DADA2 patients. In vitro stimulated B lymphocytes were able to secrete nonfunctional ADA2 protein, suggesting a cell intrinsic defect resulting in an impairment of B-cell proliferation and differentiation. Moreover, CD4+ and CD8+ T cells were diminished; however, the frequency of circulating T follicular helper cells was significantly increased but they had an impairment in IL-21 production possibly contributing to an impaired B cell help. Our findings suggest that ADA2 mutation could lead to a B-cell intrinsic defect but also to a defective Tfh cell function, which could contribute to the immunodeficient phenotype reported in DADA2 patients.
Asunto(s)
Adenosina Desaminasa/deficiencia , Agammaglobulinemia/inmunología , Linfocitos B/inmunología , Péptidos y Proteínas de Señalización Intercelular/deficiencia , Inmunodeficiencia Combinada Grave/inmunología , Células T Auxiliares Foliculares/inmunología , Adenosina Desaminasa/genética , Adenosina Desaminasa/inmunología , Adolescente , Adulto , Agammaglobulinemia/enzimología , Agammaglobulinemia/genética , Linfocitos B/enzimología , Linfocitos B/patología , Estudios de Casos y Controles , Diferenciación Celular , Proliferación Celular , Niño , Preescolar , Femenino , Humanos , Memoria Inmunológica , Inmunofenotipificación , Técnicas In Vitro , Lactante , Recién Nacido , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/inmunología , Interleucinas/biosíntesis , Activación de Linfocitos , Masculino , Mutación , Inmunodeficiencia Combinada Grave/enzimología , Inmunodeficiencia Combinada Grave/genética , Células T Auxiliares Foliculares/patologíaRESUMEN
[This corrects the article DOI: 10.1371/journal.ppat.1008477.].
RESUMEN
Post-transplant lymphoproliferative disorder (PTLD) is a potentially fatal complication after organ transplantation frequently associated with the Epstein-Barr virus (EBV). Immunosuppressive treatment is thought to allow the expansion of EBV-infected B cells, which often express all eight oncogenic EBV latent proteins. Here, we assessed whether HLA-A2 transgenic humanized NSG mice treated with the immunosuppressant FK506 could be used to model EBV-PTLD. We found that FK506 treatment of EBV-infected mice led to an elevated viral burden, more frequent tumor formation and diminished EBV-induced T cell responses, indicative of reduced EBV-specific immune control. EBV latency III and lymphoproliferation-associated cellular transcripts were up-regulated in B cells from immunosuppressed animals, akin to the viral and host gene expression pattern found in EBV-PTLD. Utilizing an unbiased gene expression profiling approach, we identified genes differentially expressed in B cells of EBV-infected animals with and without FK506 treatment. Upon investigating the most promising candidates, we validated sCD30 as a marker of uncontrolled EBV proliferation in both humanized mice and in pediatric patients with EBV-PTLD. High levels of sCD30 have been previously associated with EBV-PTLD in patients. As such, we believe that humanized mice can indeed model aspects of EBV-PTLD development and may prove useful for the safety assessment of immunomodulatory therapies.
Asunto(s)
Trastornos Linfoproliferativos/inmunología , Trastornos Linfoproliferativos/virología , Tacrolimus/farmacología , Animales , Linfocitos B/metabolismo , ADN Viral , Modelos Animales de Enfermedad , Infecciones por Virus de Epstein-Barr/virología , Femenino , Perfilación de la Expresión Génica/métodos , Antígeno HLA-A2 , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/metabolismo , Herpesvirus Humano 4/patogenicidad , Humanos , Huésped Inmunocomprometido , Inmunosupresores/farmacología , Masculino , Ratones , Ratones Endogámicos NOD , Ratones Transgénicos , Trasplante de Órganos/efectos adversos , Transcriptoma/genética , Carga ViralRESUMEN
RAS-associated autoimmune leukoproliferative disease (RALD) is a rare immune dysregulation syndrome caused by somatic gain-of-function mutations of either NRAS or KRAS gene in hematopoietic cells. We describe a 27-year-old patient presenting at 5 months of age with recurrent infections and generalized lymphadenopathy who developed a complex multi-organ autoimmune syndrome with hypogammaglobulinemia, partially controlled with oral steroids, hydroxichloroquine, mofetil mycophenolate and IVIG prophylaxis. Activation of type I interferon pathway was observed in peripheral blood. Since 18 years of age, the patient developed regenerative nodular hyperplasia of the liver evolving into hepatopulmonary syndrome. Whole exome sequencing analysis of the peripheral blood DNA showed the NRAS p.Gly13Asp mutation validated as somatic. Our report highlights the possibility of detecting somatic NRAS gene mutations in patients with inflammatory immune dysregulation and type I interferon activation.
Asunto(s)
Síndrome Linfoproliferativo Autoinmune/genética , Síndrome Linfoproliferativo Autoinmune/inmunología , GTP Fosfohidrolasas/genética , Interferón Tipo I/inmunología , Hepatopatías/genética , Proteínas de la Membrana/genética , Adulto , Síndrome Linfoproliferativo Autoinmune/complicaciones , Humanos , Hepatopatías/inmunología , MutaciónRESUMEN
In muscular dystrophies, muscle membrane fragility results in a tissue-specific increase of danger-associated molecular pattern molecules (DAMPs) and infiltration of inflammatory cells. The DAMP extracellular ATP (eATP) released by dying myofibers steadily activates muscle and immune purinergic receptors exerting dual negative effects: a direct damage linked to altered intracellular calcium homeostasis in muscle cells and an indirect toxicity through the triggering of the immune response and inhibition of regulatory T cells. Accordingly, pharmacologic and genetic inhibition of eATP signaling improves the phenotype in models of chronic inflammatory diseases. In α-sarcoglycanopathy, eATP effects may be further amplified because α-sarcoglycan extracellular domain binds eATP and displays an ecto-ATPase activity, thus controlling eATP concentration at the cell surface and attenuating the magnitude and/or the duration of eATP-induced signals. Herein, we show that in vivo blockade of the eATP/P2X purinergic pathway by a broad-spectrum P2X receptor-antagonist delayed the progression of the dystrophic phenotype in α-sarcoglycan-null mice. eATP blockade dampened the muscular inflammatory response and enhanced the recruitment of forkhead box protein P3-positive immunosuppressive regulatory CD4+ T cells. The improvement of the inflammatory features was associated with increased strength, reduced necrosis, and limited expression of profibrotic factors, suggesting that pharmacologic purinergic antagonism, altering the innate and adaptive immune component in muscle infiltrates, might provide a therapeutic approach to slow disease progression in α-sarcoglycanopathy.
Asunto(s)
Adenosina Trifosfato/inmunología , Distrofia Muscular Animal , Miofibrillas , Sarcoglicanos/deficiencia , Linfocitos T Reguladores , Adenosina Trifosfato/genética , Animales , Calcio/inmunología , Enfermedad Crónica , Inflamación/genética , Inflamación/inmunología , Inflamación/patología , Ratones , Ratones Noqueados , Distrofia Muscular Animal/genética , Distrofia Muscular Animal/inmunología , Distrofia Muscular Animal/patología , Miofibrillas/inmunología , Miofibrillas/patología , Receptores Purinérgicos P2X/genética , Receptores Purinérgicos P2X/inmunología , Sarcoglicanos/inmunología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/patologíaRESUMEN
Fas is a transmembrane receptor involved in the maintenance of tolerance and immune homeostasis. In murine models, it has been shown to be essential for deletion of autoreactive B cells in the germinal center. The role of Fas in human B-cell selection and in development of autoimmunity in patients carrying FAS mutations is unclear. We analyzed patients with either a somatic FAS mutation or a germline FAS mutation and somatic loss-of-heterozygosity, which allows comparing the fate of B cells with impaired vs normal Fas signaling within the same individual. Class-switched memory B cells showed: accumulation of FAS-mutated B cells; failure to enrich single V, D, J genes and single V-D, D-J gene combinations of the B-cell receptor variable region; increased frequency of variable regions with higher content of positively charged amino acids; and longer CDR3 and maintenance of polyreactive specificities. Importantly, Fas-deficient switched memory B cells showed increased rates of somatic hypermutation. Our data uncover a defect in B-cell selection in patients with FAS mutations, which has implications for the understanding of the pathogenesis of autoimmunity and lymphomagenesis of autoimmune lymphoproliferative syndrome.
Asunto(s)
Síndrome Linfoproliferativo Autoinmune/inmunología , Subgrupos de Linfocitos B/inmunología , Selección Clonal Mediada por Antígenos , Mutación , Receptor fas/fisiología , Apoptosis , Autoinmunidad , Línea Celular Transformada , Transformación Celular Neoplásica , Niño , Codón sin Sentido , Femenino , Mutación del Sistema de Lectura , Mutación de Línea Germinal , Heterocigoto , Humanos , Memoria Inmunológica , Pérdida de Heterocigocidad , Masculino , Análisis de Secuencia de ADN , Hipermutación Somática de Inmunoglobulina , Recombinación V(D)J , Receptor fas/deficiencia , Receptor fas/genéticaRESUMEN
Autosomal recessive osteopetrosis (ARO) is a severe bone disease characterized by increased bone density due to impairment in osteoclast resorptive function or differentiation. Hematopoietic stem cell transplantation is the only available treatment; however, this therapy is not effective in RANKL-dependent ARO, since in bone this gene is mainly expressed by cells of mesenchymal origin. Of note, whether lack of RANKL production might cause a defect also in the bone marrow (BM) stromal compartment, possibly contributing to the pathology, is unknown. To verify this possibility, we generated and characterized BM mesenchymal stromal cell (BM-MSC) lines from wild type and Rankl-/- mice, and found that Rankl-/- BM-MSCs displayed reduced clonogenicity and osteogenic capacity. The differentiation defect was significantly improved by lentiviral transduction of Rankl-/- BM-MSCs with a vector stably expressing human soluble RANKL (hsRANKL). Expression of Rankl receptor, Rank, on the cytoplasmic membrane of BM-MSCs pointed to the existence of an autocrine loop possibly activated by the secreted cytokine. Based on the close resemblance of RANKL-defective osteopetrosis in humans and mice, we expect that our results are also relevant for RANKL-dependent ARO patients. Data obtained in vitro after transduction with a lentiviral vector expressing hsRANKL would suggest that restoration of RANKL production might not only rescue the defective osteoclastogenesis of this ARO form, but also improve a less obvious defect in the osteoblast lineage, thus possibly achieving higher benefit for the patients, when the approach is translated to clinics. Stem Cells 2017;35:1365-1377.
Asunto(s)
Diferenciación Celular , Vectores Genéticos/metabolismo , Lentivirus/metabolismo , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Osteogénesis , Ligando RANK/deficiencia , Animales , Biomarcadores/metabolismo , Células Clonales , Inmunofenotipificación , Ratones Endogámicos C57BL , Ligando RANK/metabolismo , Transducción de Señal , Transducción GenéticaRESUMEN
The immune and the skeletal system are tightly interconnected, and B lymphocytes are uniquely endowed with osteo-interactive properties. In this context, receptor activator of NF-κB (RANK) ligand (RANKL) plays a pivotal role in lymphoid tissue formation and bone homeostasis. Although murine models lacking RANK or RANKL show defects in B cell number, the role of the RANKL-RANK axis on B physiology is still a matter of debate. In this study, we have characterized in detail B cell compartment in Rankl(-/-) mice, finding a relative expansion of marginal zone B cells, B1 cells, and plasma cells associated with increased Ig serum levels, spontaneous germinal center formation, and hyperresponse to CD40 triggering. Such abnormalities were associated with an increased frequency of regulatory B cells and augmented B cell-derived IL-10 production. Remarkably, in vivo IL-10-R blockade reduced T cell-triggered plasma cell differentiation and restrained the expansion of regulatory B cells. These data point to a novel role of the RANKL-RANK axis in the regulation of B cell homeostasis and highlight an unexpected link between IL-10 CD40 signaling and the RANKL pathway.
Asunto(s)
Linfocitos B/inmunología , Interleucina-10/inmunología , Ligando RANK/deficiencia , Ligando RANK/inmunología , Animales , Ratones , Ratones NoqueadosRESUMEN
The paracaspase MALT1 plays an important role in immune receptor-driven signaling pathways leading to NF-κB activation. MALT1 promotes signaling by acting as a scaffold, recruiting downstream signaling proteins, as well as by proteolytic cleavage of multiple substrates. However, the relative contributions of these two different activities to T and B cell function are not well understood. To investigate how MALT1 proteolytic activity contributes to overall immune cell regulation, we generated MALT1 protease-deficient mice (Malt1(PD/PD)) and compared their phenotype with that of MALT1 knockout animals (Malt1(-/-)). Malt1(PD/PD) mice displayed defects in multiple cell types including marginal zone B cells, B1 B cells, IL-10-producing B cells, regulatory T cells, and mature T and B cells. In general, immune defects were more pronounced in Malt1(-/-) animals. Both mouse lines showed abrogated B cell responses upon immunization with T-dependent and T-independent Ags. In vitro, inactivation of MALT1 protease activity caused reduced stimulation-induced T cell proliferation, impaired IL-2 and TNF-α production, as well as defective Th17 differentiation. Consequently, Malt1(PD/PD) mice were protected in a Th17-dependent experimental autoimmune encephalomyelitis model. Surprisingly, Malt1(PD/PD) animals developed a multiorgan inflammatory pathology, characterized by Th1 and Th2/0 responses and enhanced IgG1 and IgE levels, which was delayed by wild-type regulatory T cell reconstitution. We therefore propose that the pathology characterizing Malt1(PD/PD) animals arises from an immune imbalance featuring pathogenic Th1- and Th2/0-skewed effector responses and reduced immunosuppressive compartments. These data uncover a previously unappreciated key function of MALT1 protease activity in immune homeostasis and underline its relevance in human health and disease.
Asunto(s)
Linfocitos B Reguladores/inmunología , Caspasas/inmunología , Diferenciación Celular/inmunología , Proliferación Celular , Encefalomielitis Autoinmune Experimental/inmunología , Proteínas de Neoplasias/inmunología , Linfocitos T Reguladores/inmunología , Animales , Linfocitos B Reguladores/patología , Caspasas/genética , Diferenciación Celular/genética , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/patología , Humanos , Inmunoglobulina E/genética , Inmunoglobulina E/inmunología , Inmunoglobulina G/genética , Inmunoglobulina G/inmunología , Inflamación/genética , Inflamación/inmunología , Inflamación/patología , Interleucina-10/genética , Interleucina-10/inmunología , Interleucina-2/genética , Interleucina-2/inmunología , Ratones , Ratones Noqueados , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas , Proteínas de Neoplasias/genética , Linfocitos T Reguladores/patología , Células TH1/inmunología , Células TH1/patología , Células Th17/inmunología , Células Th17/patologíaRESUMEN
Infiltration of immune cells and chronic inflammation substantially affect skeletal and cardiac muscle degeneration in Duchenne muscular dystrophy. In the immune system, extracellular adenosine triphosphate (ATP) released by dying cells is sensed as a danger associated molecular pattern through P2 purinergic receptors. Specifically, the P2X7 subtype has a prominent role in regulating immune system physiology and contributes to inflammasome activation also in muscle cells. Here, we show that in vivo blockade of the extracellular ATP/P2X purinergic signaling pathway by periodate-oxidized ATP delayed the progression of the dystrophic phenotype and dampened the local inflammatory response in mdx mice, a spontaneous mouse model of dystrophin deficiency. Reduced infiltration of leukocytes and macrophages and decreased expression of IL-6 were revealed in the muscles of periodate-oxidized ATP-treated mdx mice. Concomitantly, an increase in Foxp3(+) immunosuppressive regulatory T cells was observed and correlated with enhanced myofiber regeneration. Moreover, we detected reduced concentrations of profibrotic cytokines, including transforming growth factor-ß and connective tissue growth factor, in muscles of periodate-oxidized ATP-treated mdx mice. The improvement of inflammatory features was associated with increased strength and reduced necrosis, thus suggesting that pharmacologic purinergic antagonism altering the adaptive immune component in the muscle infiltrates might represent a promising therapeutic approach in Duchenne muscular dystrophy.
Asunto(s)
Músculo Esquelético/inmunología , Distrofia Muscular de Duchenne/inmunología , Receptores Purinérgicos P2X/fisiología , Linfocitos T Reguladores/inmunología , Adenosina Trifosfato/análogos & derivados , Adenosina Trifosfato/inmunología , Adenosina Trifosfato/farmacología , Adenosina Trifosfato/uso terapéutico , Animales , Progresión de la Enfermedad , Evaluación Preclínica de Medicamentos/métodos , Masculino , Ratones Endogámicos C57BL , Ratones Endogámicos mdx , Músculo Esquelético/patología , Músculo Esquelético/fisiología , Distrofia Muscular de Duchenne/patología , Distrofia Muscular de Duchenne/prevención & control , Condicionamiento Físico Animal , Antagonistas del Receptor Purinérgico P2X/farmacología , Antagonistas del Receptor Purinérgico P2X/uso terapéutico , Receptores Purinérgicos P2X/metabolismo , Regeneración/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Linfocitos T Reguladores/efectos de los fármacosRESUMEN
BACKGROUND: Adenosine deaminase (ADA) deficiency causes severe cellular and humoral immune defects and dysregulation because of metabolic toxicity. Alterations in B-cell development and function have been poorly studied. Enzyme replacement therapy (ERT) and hematopoietic stem cell (HSC) gene therapy (GT) are therapeutic options for patients lacking a suitable bone marrow (BM) transplant donor. OBJECTIVE: We sought to study alterations in B-cell development in ADA-deficient patients and investigate the ability of ERT and HSC-GT to restore normal B-cell differentiation and function. METHODS: Flow cytometry was used to characterize B-cell development in BM and the periphery. The percentage of gene-corrected B cells was measured by using quantitative PCR. B cells were assessed for their capacity to proliferate and release IgM after stimulation. RESULTS: Despite the severe peripheral B-cell lymphopenia, patients with ADA-deficient severe combined immunodeficiency showed a partial block in central BM development. Treatment with ERT or HSC-GT reverted most BM alterations, but ERT led to immature B-cell expansion. In the periphery transitional B cells accumulated under ERT, and the defect in maturation persisted long-term. HSC-GT led to a progressive improvement in B-cell numbers and development, along with increased levels of gene correction. The strongest selective advantage for ADA-transduced cells occurred at the transition from immature to naive cells. B-cell proliferative responses and differentiation to immunoglobulin secreting IgM after B-cell receptor and Toll-like receptor triggering were severely impaired after ERT and improved significantly after HSC-GT. CONCLUSIONS: ADA-deficient patients show specific defects in B-cell development and functions that are differently corrected after ERT and HSC-GT.
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Adenosina Desaminasa/deficiencia , Linfocitos B/fisiología , Terapia de Reemplazo Enzimático , Terapia Genética , Trasplante de Células Madre Hematopoyéticas , Adenosina Desaminasa/genética , Adenosina Desaminasa/uso terapéutico , Adolescente , Factor Activador de Células B/fisiología , Linfocitos B/inmunología , Niño , Preescolar , Humanos , LactanteRESUMEN
Wiskott-Aldrich Syndrome protein (WASp) regulates the cytoskeleton in hematopoietic cells and mutations in its gene cause the Wiskott-Aldrich Syndrome (WAS), a primary immunodeficiency with microthrombocytopenia, eczema and a higher susceptibility to develop tumors. Autoimmune manifestations, frequently observed in WAS patients, are associated with an increased risk of mortality and still represent an unsolved aspect of the disease. B cells play a crucial role both in immune competence and self-tolerance and defects in their development and function result in immunodeficiency and/or autoimmunity. We performed a phenotypical and molecular analysis of central and peripheral B-cell compartments in WAS pediatric patients. We found a decreased proportion of immature B cells in the bone marrow correlating with an increased presence of transitional B cells in the periphery. These results could be explained by the defective migratory response of WAS B cells to SDF-1α, essential for the retention of immature B cells in the BM. In the periphery, we observed an unusual expansion of CD21(low) B-cell population and increased plasma BAFF levels that may contribute to the high susceptibility to develop autoimmune manifestations in WAS patients. WAS memory B cells were characterized by a reduced in vivo proliferation, decreased somatic hypermutation and preferential usage of IGHV4-34, an immunoglobulin gene commonly found in autoreactive B cells. In conclusion, our findings demonstrate that WASp-deficiency perturbs B-cell homeostasis thus adding a new layer of immune dysregulation concurring to the increased susceptibility to develop autoimmunity in WAS patients.
Asunto(s)
Autoinmunidad , Linfocitos B/inmunología , Susceptibilidad a Enfermedades/inmunología , Proteína del Síndrome de Wiskott-Aldrich/deficiencia , Síndrome de Wiskott-Aldrich/inmunología , Factor Activador de Células B/sangre , Factor Activador de Células B/genética , Factor Activador de Células B/inmunología , Linfocitos B/patología , Médula Ósea/inmunología , Médula Ósea/patología , Diferenciación Celular , Movimiento Celular , Quimiocina CXCL12/genética , Quimiocina CXCL12/inmunología , Expresión Génica , Homeostasis/inmunología , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/inmunología , Memoria Inmunológica , Receptores de Complemento 3d/genética , Receptores de Complemento 3d/inmunología , Síndrome de Wiskott-Aldrich/genética , Síndrome de Wiskott-Aldrich/patología , Proteína del Síndrome de Wiskott-Aldrich/genética , Proteína del Síndrome de Wiskott-Aldrich/inmunologíaRESUMEN
Omenn syndrome (OS) is an atypical primary immunodeficiency characterized by severe autoimmunity because of activated T cells infiltrating target organs. The impaired recombinase activity in OS severely affects expression of the pre-T-cell receptor complex in immature thymocytes, which is crucial for an efficient development of the thymic epithelial component. Anti-CD3ε monoclonal antibody (mAb) treatment in RAG2(-/-) mice was previously shown to mimic pre-TCR signaling promoting thymic expansion. Here we show the effect of anti-CD3ε mAb administration in the RAG2(R229Q) mouse model, which closely recapitulates human OS. These animals, in spite of the inability to induce the autoimmune regulator, displayed a significant amelioration in thymic epithelial compartment and an important reduction of peripheral T-cell activation and tissue infiltration. Furthermore, by injecting a high number of RAG2(R229Q) progenitors into RAG2(-/-) animals previously conditioned with anti-CD3ε mAb, we detected autoimmune regulator expression together with the absence of peripheral immunopathology. These observations indicate that improving epithelial thymic function might ameliorate the detrimental behavior of the cell-autonomous RAG defect. Our data provide important therapeutic proof of concept for future clinical applications of anti-CD3ε mAb treatment in severe combined immunodeficiency forms characterized by poor thymus function and autoimmunity.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Enfermedades Autoinmunes/prevención & control , Complejo CD3/inmunología , Inmunodeficiencia Combinada Grave/terapia , Timo/efectos de los fármacos , Animales , Animales Recién Nacidos , Autoinmunidad/efectos de los fármacos , Autoinmunidad/genética , Proteínas de Unión al ADN/genética , Modelos Animales de Enfermedad , Técnicas de Sustitución del Gen , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Tamaño de los Órganos/efectos de los fármacos , Inmunodeficiencia Combinada Grave/inmunología , Inmunodeficiencia Combinada Grave/patología , Timo/inmunología , Timo/patología , Timo/ultraestructuraRESUMEN
Adenosine acts as anti-inflammatory mediator on the immune system and has been described in regulatory T cell (Treg)-mediated suppression. In the absence of adenosine deaminase (ADA), adenosine and other purine metabolites accumulate, leading to severe immunodeficiency with recurrent infections (ADA-SCID). Particularly ADA-deficient patients with late-onset forms and after enzyme replacement therapy (PEG-ADA) are known to manifest immune dysregulation. Herein we provide evidence that alterations in the purine metabolism interfere with Treg function, thereby contributing to autoimmune manifestations in ADA deficiency. Tregs isolated from PEG-ADA-treated patients are reduced in number and show decreased suppressive activity, whereas they are corrected after gene therapy. Untreated murine ADA(-/-) Tregs show alterations in the plasma membrane CD39/CD73 ectonucleotidase machinery and limited suppressive activity via extracellular adenosine. PEG-ADA-treated mice developed multiple autoantibodies and hypothyroidism in contrast to mice treated with bone marrow transplantation or gene therapy. Tregs isolated from PEG-ADA-treated mice lacked suppressive activity, suggesting that this treatment interferes with Treg functionality. The alterations in the CD39/CD73 adenosinergic machinery and loss of function in ADA-deficient Tregs provide new insights into a predisposition to autoimmunity and the underlying mechanisms causing defective peripheral tolerance in ADA-SCID.
Asunto(s)
5'-Nucleotidasa/inmunología , Adenosina/inmunología , Agammaglobulinemia/inmunología , Antígenos CD/inmunología , Apirasa/inmunología , Inmunodeficiencia Combinada Grave/inmunología , Linfocitos T Reguladores/inmunología , 5'-Nucleotidasa/metabolismo , Adenosina/metabolismo , Adenosina Desaminasa/deficiencia , Adenosina Desaminasa/genética , Adenosina Desaminasa/inmunología , Adenosina Desaminasa/metabolismo , Adenosina Desaminasa/uso terapéutico , Adolescente , Adulto , Agammaglobulinemia/genética , Agammaglobulinemia/terapia , Animales , Antígenos CD/metabolismo , Apirasa/metabolismo , Autoanticuerpos/inmunología , Niño , Preescolar , Femenino , Factores de Transcripción Forkhead/inmunología , Factores de Transcripción Forkhead/metabolismo , Terapia Genética/métodos , Trasplante de Células Madre Hematopoyéticas/métodos , Humanos , Hipotiroidismo/enzimología , Hipotiroidismo/genética , Hipotiroidismo/inmunología , Inmunohistoquímica , Lactante , Masculino , Ratones , Ratones Noqueados , Polietilenglicoles/química , Inmunodeficiencia Combinada Grave/genética , Inmunodeficiencia Combinada Grave/terapia , Linfocitos T Reguladores/metabolismoRESUMEN
Dendritic cells (DC) are highly specialized antigen-presenting cells characterized by the ability to prime T-cell responses. Mesenchymal stem cells (MSC) are adult stromal progenitor cells displaying immunomodulatory activities including inhibition of DC maturation in vitro. However, the specific impact of MSC on DC functions, upon in vivo administration, has never been elucidated. Here we show that murine MSC impair Toll-like receptor-4 induced activation of DC resulting in the inhibition of cytokines secretion, down-regulation of molecules involved in the migration to the lymph nodes, antigen presentation to CD4(+) T cells, and cross-presentation to CD8(+) T cells. These effects are associated with the inhibition of phosphorylation of intracellular mitogen-activated protein kinases. Intravenous administration of MSC decreased the number of CCR7 and CD49dß1 expressing CFSE-labeled DC in the draining lymph nodes and hindered local antigen priming of DO11.10 ovalbumin-specific CD4(+) T cells. Upon labeling of DC with technetium-99m hexamethylpropylene amine oxime to follow their in vivo biodistribution, we demonstrated that intravenous injection of MSC blocks, almost instantaneously, the migration of subcutaneously administered ovalbumin-pulsed DC to the draining lymph nodes. These findings indicate that MSC significantly affect DC ability to prime T cells in vivo because of their inability to home to the draining lymph nodes and further confirm MSC potentiality as therapy for immune-mediated diseases.
Asunto(s)
Células Dendríticas/inmunología , Células Madre Mesenquimatosas/inmunología , Linfocitos T/inmunología , Animales , Presentación de Antígeno , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Diferenciación Celular/inmunología , Movimiento Celular/inmunología , Técnicas de Cocultivo , Citocinas/genética , Células Dendríticas/citología , Células Dendríticas/fisiología , Células Dendríticas/trasplante , Expresión Génica , Ganglios Linfáticos/citología , Ganglios Linfáticos/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Transducción de Señal/inmunología , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/inmunologíaRESUMEN
Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease characterized by recurring suppurating lesions of the intertriginous areas, resulting in a substantial impact on patients' QOL. HS pathogenesis remains poorly understood. An autoimmune component has been proposed, but disease-specific autoantibodies, autoantigens, or autoreactive T cells have yet to be described. In this study, we identify a high prevalence of IgM, IgG, and IgA antibodies directed against Nε-carboxyethyl lysine (CEL), a methylglyoxal-induced advanced glycation end-product, in the sera of patients with HS. Titers of anti-CEL IgG and IgA antibodies were highly elevated in HS compared with those in healthy controls and individuals with other inflammatory skin diseases. Strikingly, the majority of anti-CEL IgG was of the IgG2 subclass and correlated independently with both disease severity and duration. Both CEL and anti-CELâproducing plasmablasts could be isolated directly from HS skin lesions, further confirming the disease relevance of this autoimmune response. Our data point to an aberration of the methylglyoxal pathway in HS and support an autoimmune axis in the pathogenesis of this debilitating disease.
Asunto(s)
Hidradenitis Supurativa , Humanos , Autoanticuerpos , Lisina , Calidad de Vida , Piruvaldehído , Inmunoglobulina GRESUMEN
Regulated expression of positive and negative regulatory factors controls the extent and duration of T cell adaptive immune response preserving the organism's integrity. Calreticulin (CRT) is a major Ca2+ buffering chaperone in the lumen of the endoplasmic reticulum. Here we investigated the impact of CRT deficiency on T cell function in immunodeficient mice reconstituted with fetal liver crt-/- hemopoietic progenitors. These chimeric mice displayed severe immunopathological traits, which correlated with a lower threshold of T cell receptor (TCR) activation and exaggerated peripheral T cell response to antigen with enhanced secretion of inflammatory cytokines. In crt-/- T cells TCR stimulation induced pulsatile cytosolic elevations of Ca2+ concentration and protracted accumulation of nuclear factor of activated T cells in the nucleus as well as sustained activation of the mitogen-activated protein kinase pathways. These observations support the hypothesis that CRT-dependent shaping of Ca2+ signaling critically contributes to the modulation of the T cell adaptive immune response.