RESUMEN
Memory B cells (MBCs) can persist for a lifetime, but the mechanisms that allow their long-term survival remain poorly understood. Here, we isolated and analyzed human splenic smallpox/vaccinia protein B5-specific MBCs in individuals who were vaccinated more than 40 years ago. Only a handful of clones persisted over such an extended period, and they displayed limited intra-clonal diversity with signs of extensive affinity-based selection. These long-lived MBCs appeared enriched in a CD21hiCD20hi IgG+ splenic B cell subset displaying a marginal-zone-like NOTCH/MYC-driven signature, but they did not harbor a unique longevity-associated transcriptional or metabolic profile. Finally, the telomeres of B5-specific, long-lived MBCs were longer than those in patient-paired naive B cells in all the samples analyzed. Overall, these results imply that separate mechanisms such as early telomere elongation, affinity selection during the contraction phase, and access to a specific niche contribute to ensuring the functional longevity of MBCs.
Asunto(s)
Memoria Inmunológica , Células B de Memoria , Linfocitos B/metabolismo , Centro Germinal , Humanos , Inmunoglobulina G/metabolismoRESUMEN
In the version of this article initially published, the name of one author was incorrect (James P. Santo). The correct name is James P. Di Santo. The error has been corrected in the HTML and PDF versions of the article.
RESUMEN
The quantification and characterization of circulating immune cells provide key indicators of human health and disease. To identify the relative effects of environmental and genetic factors on variation in the parameters of innate and adaptive immune cells in homeostatic conditions, we combined standardized flow cytometry of blood leukocytes and genome-wide DNA genotyping of 1,000 healthy, unrelated people of Western European ancestry. We found that smoking, together with age, sex and latent infection with cytomegalovirus, were the main non-genetic factors that affected variation in parameters of human immune cells. Genome-wide association studies of 166 immunophenotypes identified 15 loci that showed enrichment for disease-associated variants. Finally, we demonstrated that the parameters of innate cells were more strongly controlled by genetic variation than were those of adaptive cells, which were driven by mainly environmental exposure. Our data establish a resource that will generate new hypotheses in immunology and highlight the role of innate immunity in susceptibility to common autoimmune diseases.
Asunto(s)
Variación Genética/inmunología , Inmunidad Innata/genética , Inmunidad Adaptativa/genética , Adulto , Anciano , Femenino , Estudio de Asociación del Genoma Completo , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
dsRNA-dependent protein kinase R (PKR) is a key factor of innate immunity. It is involved in translation inhibition, apoptosis, and enhancement of the proinflammatory and IFN responses. However, how these antiviral functions are conserved during evolution remains largely unknown. Overexpression and knockout studies in a Chinook salmon (Oncorhynchus tshawytscha) cell line were conducted to assess the role of salmonid PKR in the antiviral response. Three distinct mRNA isoforms from a unique pkr gene, named pkr-fl (full length), pkr-ml (medium length) and pkr-sl (short length), were cloned and a pkr-/- clonal fish cell line was developed using CRISPR/Cas9 genome editing. PKR-FL includes an N-terminal dsRNA-binding domain and a C-terminal kinase domain, whereas PKR-ML and PKR-SL display a truncated or absent kinase domain, respectively. PKR-FL is induced during IFNA2 stimulation but not during viral hemorrhagic septicemia virus (VHSV) infection. Overexpression experiments showed that only PKR-FL possesses antiviral functions, including activation of apoptosis and inhibition of de novo protein synthesis. Knockout experiments confirmed that PKR is involved in apoptosis activation during the late stage of VHSV infection. Endogenous PKR also plays a critical role in translation inhibition upon poly(I:C) transfection after IFNA2 treatment. It is, however, not involved in translational arrest during VHSV infection. Extra- and intracellular titrations showed that endogenous PKR does not directly inhibit viral replication but apparently favors virion release into the supernatant, likely by triggering late apoptosis. Altogether, our data confirm that salmonid PKR has conserved molecular functions that VHSV appears to bypass with subversion strategies.
Asunto(s)
Apoptosis , Biosíntesis de Proteínas , Salmón , eIF-2 Quinasa , Animales , Apoptosis/inmunología , eIF-2 Quinasa/metabolismo , eIF-2 Quinasa/genética , Salmón/inmunología , Línea Celular , Biosíntesis de Proteínas/inmunología , Inmunidad Innata , Novirhabdovirus/fisiología , Novirhabdovirus/inmunología , Proteínas de Peces/genética , Proteínas de Peces/inmunología , Proteínas de Peces/metabolismo , ARN Bicatenario/inmunología , Enfermedades de los Peces/inmunología , Sistemas CRISPR-CasRESUMEN
BACKGROUND: Viperin, also known as radical S-adenosyl-methionine domain containing protein 2 (RSAD2), is an interferon-inducible protein that is involved in the innate immune response against a wide array of viruses. In mammals, Viperin exerts its antiviral function through enzymatic conversion of cytidine triphosphate (CTP) into its antiviral analog ddhCTP as well as through interactions with host proteins involved in innate immune signaling and in metabolic pathways exploited by viruses during their life cycle. However, how Viperin modulates the antiviral response in fish remains largely unknown. RESULTS: For this purpose, we developed a fathead minnow (Pimephales promelas) clonal cell line in which the unique viperin gene has been knocked out by CRISPR/Cas9 genome-editing. In order to decipher the contribution of fish Viperin to the antiviral response and its regulatory role beyond the scope of the innate immune response, we performed a comparative RNA-seq analysis of viperin-/- and wildtype cell lines upon stimulation with recombinant fathead minnow type I interferon. CONCLUSIONS: Our results revealed that Viperin does not exert positive feedback on the canonical type I IFN but acts as a negative regulator of the inflammatory response by downregulating specific pro-inflammatory genes and upregulating repressors of the NF-κB pathway. It also appeared to play a role in regulating metabolic processes, including one carbon metabolism, bone formation, extracellular matrix organization and cell adhesion.
Asunto(s)
Cyprinidae , Inflamación , Animales , Cyprinidae/metabolismo , Cyprinidae/genética , Inflamación/metabolismo , Inflamación/genética , Inmunidad Innata , Proteínas de Peces/genética , Proteínas de Peces/metabolismo , Línea Celular , Sistemas CRISPR-Cas , Interferón Tipo I/metabolismo , Edición Génica , Regulación de la Expresión GénicaRESUMEN
Flow cytometry is the method of choice for immunophenotyping in the context of clinical, translational, and systems immunology studies. Among the latter, the Milieu Intérieur (MI) project aims at defining the boundaries of a healthy immune response to identify determinants of immune response variation. MI used immunophenotyping of a 1000 healthy donor cohort by flow cytometry as a principal outcome for immune variance at steady state. New generation spectral cytometers now enable high-dimensional immune cell characterization from small sample volumes. Therefore, for the MI 10-year follow up study, we have developed two high-dimensional spectral flow cytometry panels for deep characterization of innate and adaptive whole blood immune cells (35 and 34 fluorescent markers, respectively). We have standardized the protocol for sample handling, staining, acquisition, and data analysis. This approach enables the reproducible quantification of over 182 immune cell phenotypes at a single site. We have applied the protocol to discern minor differences between healthy and patient samples and validated its value for application in immunomonitoring studies. Our protocol is currently used for characterization of the impact of age and environmental factors on peripheral blood immune phenotypes of >400 donors from the initial MI cohort.
Asunto(s)
Estudios de Seguimiento , Humanos , Inmunofenotipificación , Fenotipo , Citometría de Flujo/métodosRESUMEN
Upon infection, B lymphocytes develop clonal responses. In teleost fish, which lack lymph nodes, the kinetics and location of B cell responses remain poorly characterized. Fish pronephros is the site of B cell differentiation and the main niche for persistence of plasma cells. In this study, we undertook the analysis of the rainbow trout IgHµ repertoire in this critical tissue for humoral adaptive immunity after primary immunization and boost with a rhabdovirus, the viral hemorrhagic septicemia virus (VHSV). We used a barcoded 5' RACE-cDNA sequencing approach to characterize modifications of the IgHµ repertoire, including VH usage in expressed V(D)J rearrangements, clonal diversity, and clonotype sharing between individual fish and treatments. In the pronephros, our approach quantified the clonotype frequency across the whole IgH repertoire (i.e., with all VH), measuring the frequency of Ag-responding clonotypes. Viral infection led to extensive modifications of the pronephros B cell repertoire, implicating several VH subgroups after primary infection. In contrast, only modest changes in repertoire persisted 5 mo later, including VHSV-specific public expansions. The IgM public response implicating IgHV1-18 and JH5, previously described in spleen, was confirmed in pronephros in all infected fish, strongly correlated to the response. However, the distribution of top clonotypes showed that pronephros and spleen B cells constitute distinct compartments with different IgH repertoires. Unexpectedly, after boost, the frequency of anti-VHSV clonotypes decreased both in pronephros and spleen, raising questions about B cell circulation. A better monitoring of B cell response kinetics in lymphoid tissues will be an essential step to understand B memory and plasmocyte formation mechanisms in fish.
Asunto(s)
Enfermedades de los Peces , Septicemia Hemorrágica Viral , Novirhabdovirus , Oncorhynchus mykiss , Pronefro , Virosis , Animales , Septicemia Hemorrágica Viral/prevención & control , Oncorhynchus mykiss/genética , BazoRESUMEN
The repertoire of Abs is generated by genomic rearrangements during B cell differentiation. Although V(D)J rearrangements lead to repertoires mostly different between individuals, recent studies have shown that they contain a substantial fraction of overrepresented and shared "public" clones. We previously reported a strong public IgHµ clonotypic response against the rhabdovirus viral hemorrhagic septicemia virus in a teleost fish. In this study, we identified an IgL chain associated with this public response that allowed us to characterize its functionality. We show that this public Ab response has a potent neutralizing capacity that is typically associated with host protection during rhabdovirus infections. We also demonstrate that the public response is not restricted to a particular trout isogenic line but expressed in multiple genetic backgrounds and may be used as a marker of successful vaccination. Our work reveals that public B cell responses producing generic Abs constitute a mechanism of protection against infection conserved across vertebrates.
Asunto(s)
Formación de Anticuerpos/inmunología , Peces/inmunología , Mamíferos/inmunología , Animales , Linfocitos B/inmunología , Células Clonales/inmunología , Rhabdoviridae/inmunología , Infecciones por Rhabdoviridae/inmunología , Recombinación V(D)J/inmunología , Vacunación/métodosRESUMEN
HFE, an MHC class Ib molecule that controls iron metabolism, can be directly targeted by cytotoxic TCR αß T lymphocytes. Transgenic DBA/2 mice expressing, in a Rag 2 KO context, an αß TCR that directly recognizes mouse HFE (mHFE) were created to further explore the interface of HFE with the immune system. TCR-transgenic mHfe WT mice deleted mHFE-reactive T cells in the thymus, but a fraction of reprogrammed cells were able to escape deletion. In contrast, TCR-transgenic mice deprived of mHFE molecules (mHfe KO mice) or expressing a C282âY mutated mHFE molecule - the most frequent mutation associated with human hereditary hemochromatosis - positively selected mHFE-reactive CD8(+) T lymphocytes and were not tolerant toward mHFE. By engrafting these mice with DBA/2 WT (mHFE(+)) skin, it was established, as suspected on the basis of similar engraftments performed on DBA/2 mHfe KO mice, that mHFE behaves as an autonomous skin-associated histocompatibility antigen, even for mHFE-C282âY mutated mice. By contrast, infusion of DBA/2 mHFE(+) mice with naïve mHFE-reactive transgenic CD8(+) T lymphocytes did not induce GVHD. Thus, tolerance toward HFE in mHfe WT mice can be acquired at either thymic or peripheral levels but is disrupted in mice reproducing human familial hemochromatosis.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Enfermedades Genéticas Congénitas/inmunología , Hemocromatosis/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Tolerancia Inmunológica , Proteínas de la Membrana/inmunología , Sustitución de Aminoácidos , Animales , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Linfocitos T CD8-positivos/trasplante , Modelos Animales de Enfermedad , Enfermedades Genéticas Congénitas/genética , Enfermedades Genéticas Congénitas/metabolismo , Enfermedades Genéticas Congénitas/patología , Enfermedad Injerto contra Huésped/genética , Enfermedad Injerto contra Huésped/inmunología , Enfermedad Injerto contra Huésped/metabolismo , Enfermedad Injerto contra Huésped/patología , Hemocromatosis/genética , Hemocromatosis/metabolismo , Hemocromatosis/patología , Proteína de la Hemocromatosis , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos DBA , Ratones Noqueados , Mutación Missense , Piel/inmunología , Piel/metabolismo , Piel/patología , Timo/inmunología , Timo/metabolismo , Timo/patología , Trasplante HomólogoRESUMEN
Humoral memory is maintained by two types of persistent cells, memory B cells and plasma cells, which have different phenotypes and functions. Long-lived plasma cells can survive for a lifespan within a complex niche in the bone marrow and provide continuous protective serum antibody levels. Memory B cells reside in secondary lymphoid organs, where they can be rapidly mobilized upon a new antigenic encounter. Surface IgG has long been taken as a surrogate marker for memory in the mouse. Recently, however, we have brought evidence for a long-lived IgM memory B cell population in the mouse, while we have also argued that, in humans, these same cells are not classical memory B cells but marginal zone (MZ) B cells which, as opposed to their mouse MZ counterpart, recirculate and carry a mutated B cell receptor. In this review, we will discuss these apparently paradoxical results.
Asunto(s)
Linfocitos B/inmunología , Inmunoglobulina M/química , Memoria Inmunológica , Animales , Linfocitos B/fisiología , Humanos , Ratones , Receptores de Antígenos de Linfocitos B/genética , Especificidad de la EspecieRESUMEN
Growth factor independence-1B (Gfi-1B) is a transcriptional repressor essential for erythropoiesis and megakaryopoiesis. Targeted gene disruption of GFI1B in mice leads to embryonic lethality resulting from failure to produce definitive erythrocytes, hindering the study of Gfi-1B function in adult hematopoiesis. We here show that, in humans, Gfi-1B controls the development of erythrocytes and megakaryocytes by regulating the proliferation and differentiation of bipotent erythro-megakaryocytic progenitors. We further identify in this cell population the type III transforming growth factor-beta receptor gene, TGFBR3, as a direct target of Gfi-1B. Knockdown of Gfi-1B results in altered transforming growth factor-beta (TGF-beta) signaling as shown by the increase in Smad2 phosphorylation and its inability to associate to the transcription intermediary factor 1-gamma (TIF1-gamma). Because the Smad2/TIF1-gamma complex is known to specifically regulate erythroid differentiation, we propose that, by repressing TGF-beta type III receptor (TbetaRIotaII) expression, Gfi-1B favors the Smad2/TIF1-gamma interaction downstream of TGF-beta signaling, allowing immature progenitors to differentiate toward the erythroid lineage.
Asunto(s)
Diferenciación Celular/fisiología , Eritrocitos/metabolismo , Megacariocitos/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Transducción de Señal/fisiología , Factor de Crecimiento Transformador beta/metabolismo , Adulto , Proteínas Reguladoras de la Apoptosis/metabolismo , Proliferación Celular , Eritrocitos/citología , Eritropoyesis/fisiología , Regulación de la Expresión Génica/fisiología , Humanos , Células K562 , Células Progenitoras de Megacariocitos y Eritrocitos/citología , Células Progenitoras de Megacariocitos y Eritrocitos/metabolismo , Megacariocitos/citología , Complejos Multiproteicos/metabolismo , Proteínas Nucleares/metabolismo , Fosforilación/fisiología , Proteoglicanos/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Proteína Smad2/metabolismo , Trombopoyesis/fisiologíaRESUMEN
Stunting and environmental enteric dysfunction (EED) may be responsible for altered gut and systemic immune responses. However, their impact on circulating immune cell populations remains poorly characterized during early life. A detailed flow cytometry analysis of major systemic immune cell populations in 53 stunted and 52 non-stunted (2 to 5 years old) children living in Antananarivo (Madagascar) was performed. Compared to age-matched non-stunted controls, stunted children aged 2-3 years old had a significantly lower relative proportion of classical monocytes. No significant associations were found between stunting and the percentages of effector T helper cell populations (Th1, Th2, Th17, Th1Th17, and cTfh). However, we found that HLA-DR expression (MFI) on all memory CD4+ or CD8+ T cell subsets was significantly lower in stunted children compared to non-stunted controls. Interestingly, in stunted children compared to the same age-matched non-stunted controls, we observed statistically significant age-specific differences in regulatory T cells (Treg) subsets. Indeed, in 2- to 3-year-old stunted children, a significantly higher percentage of memory Treg, whilst a significantly lower percentage of naive Treg, was found. Our results revealed that both innate and adaptive systemic cell percentages, as well as activation status, were impacted in an age-related manner during stunting. Our study provides valuable insights into the understanding of systemic immune system changes in stunted children.
Asunto(s)
Monocitos , Linfocitos T Reguladores , Niño , Preescolar , Trastornos del Crecimiento , Humanos , Subgrupos de Linfocitos T , Células Th17RESUMEN
Allergic asthma is characterized by elevated levels of IgE antibodies, type 2 cytokines such as interleukin-4 (IL-4) and IL-13, airway hyperresponsiveness (AHR), mucus hypersecretion and eosinophilia. Approved therapeutic monoclonal antibodies targeting IgE or IL-4/IL-13 reduce asthma symptoms but require costly lifelong administrations. Here, we develop conjugate vaccines against mouse IL-4 and IL-13, and demonstrate their prophylactic and therapeutic efficacy in reducing IgE levels, AHR, eosinophilia and mucus production in mouse models of asthma analyzed up to 15 weeks after initial vaccination. More importantly, we also test similar vaccines specific for human IL-4/IL-13 in mice expressing human IL-4/IL-13 and the related receptor, IL-4Rα, to find efficient neutralization of both cytokines and reduced IgE levels for at least 11 weeks post-vaccination. Our results imply that dual IL-4/IL-13 vaccination may represent a cost-effective, long-term therapeutic strategy for the treatment of allergic asthma as demonstrated in mouse models, although additional studies are warranted to assess its safety and feasibility.
Asunto(s)
Asma/terapia , Interleucina-13/antagonistas & inhibidores , Interleucina-4/antagonistas & inhibidores , Vacunación/métodos , Animales , Asma/inmunología , Proteínas Bacterianas/administración & dosificación , Proteínas Bacterianas/inmunología , Enfermedad Crónica/terapia , Modelos Animales de Enfermedad , Femenino , Humanos , Inyecciones Intramusculares , Interleucina-13/genética , Interleucina-13/inmunología , Interleucina-4/genética , Interleucina-4/inmunología , Ratones , Ratones Transgénicos , Vacunas Conjugadas/administración & dosificación , Vacunas Conjugadas/inmunologíaRESUMEN
The immunoscope methodology has proven useful to analyze T-cell repertoires in mice and humans. We adapted it to the analysis of VH chains of human peripheral B cells by setting up a quantification of various VH and JH segments and the profiling of IgM-, IgG-, IgA- and IgE-expressing B cells. We then tested the hypothesis that the human B-cell and T-cell repertoires have a similar diversity of VH and V-beta rearrangements. We studied in more detail the VH5 family because it is not abundantly used, which facilitated the analysis. The data showed that the number of distinct VH5 rearrangements in all samples studied is close to the number of cells in the sample. This contrasts with T cells in which we previously showed that distinct V-beta rearrangements amount to a few percent of the number of T cells because each V-beta chain is on the average paired with approximately 25 alpha chains. Thus, in the VH5 family, the light chains add little quantitative diversity to that produced by the heavy chain alone. Whether this feature can be generalized to other VH chains is discussed.
Asunto(s)
Linfocitos B/metabolismo , Reordenamiento Génico de Cadena Pesada de Linfocito B , Cadenas Pesadas de Inmunoglobulina/metabolismo , Inmunoglobulina M/metabolismo , Región Variable de Inmunoglobulina/metabolismo , Adulto , Femenino , Citometría de Flujo , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Isotipos de Inmunoglobulinas/metabolismo , Región Variable de Inmunoglobulina/genética , Masculino , Persona de Mediana Edad , Reacción en Cadena de la PolimerasaRESUMEN
During somatic hypermutation (SHM) of immunoglobulin genes, uracils introduced by activation-induced cytidine deaminase are processed by uracil-DNA glycosylase (UNG) and mismatch repair (MMR) pathways to generate mutations at G-C and A-T base pairs, respectively. Paradoxically, the MMR-nicking complex Pms2/Mlh1 is apparently dispensable for A-T mutagenesis. Thus, how detection of U:G mismatches is translated into the single-strand nick required for error-prone synthesis is an open question. One model proposed that UNG could cooperate with MMR by excising a second uracil in the vicinity of the U:G mismatch, but it failed to explain the low impact of UNG inactivation on A-T mutagenesis. In this study, we show that uracils generated in the G1 phase in B cells can generate equal proportions of A-T and G-C mutations, which suggests that UNG and MMR can operate within the same time frame during SHM. Furthermore, we show that Ung-/-Pms2-/- mice display a 50% reduction in mutations at A-T base pairs and that most remaining mutations at A-T bases depend on two additional uracil glycosylases, thymine-DNA glycosylase and SMUG1. These results demonstrate that Pms2/Mlh1 and multiple uracil glycosylases act jointly, each one with a distinct strand bias, to enlarge the immunoglobulin gene mutation spectrum from G-C to A-T bases.
Asunto(s)
Emparejamiento Base , Reparación de la Incompatibilidad de ADN , Genes de Inmunoglobulinas , Endonucleasa PMS2 de Reparación del Emparejamiento Incorrecto/fisiología , Mutación , Uracil-ADN Glicosidasa/fisiología , Animales , Endodesoxirribonucleasas/fisiología , Fase G1 , Ratones , Ratones Endogámicos C57BLRESUMEN
We investigated the ability of monoclonal B cells to restore primary and secondary T-cell dependent antibody responses in adoptive immune-deficient hosts. Priming induced B cell activation and expansion, AID expression, antibody production and the generation of IgM+IgG- and IgM-IgG+ antigen-experienced B-cell subsets that persisted in the lymphopenic environment by cell division. Upon secondary transfer and recall the IgM-IgG+ cells responded by the production of antigen-specific IgG while the IgM+ memory cells secreted mainly IgM and little IgG, but generated new B cells expressing germinal center markers. The recall responses were more efficient if the antigenic boost was delayed suggesting that a period of adaptation is necessary before the transferred cells are able to respond. Overall these findings indicate that reconstitution of a functional and complete memory pool requires transfer of all different antigen-experienced B cell subsets. We also found that the size of the memory B cell pool did not rely on the number of the responding naïve B cells, suggesting autonomous homeostatic controls for naïve and memory B cells. By reconstituting a stable memory B cell pool in immune-deficient hosts using a monoclonal high-affinity B cell population we demonstrate the potential value of B cell adoptive immunotherapy.
Asunto(s)
Subgrupos de Linfocitos B/inmunología , Centro Germinal/inmunología , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Memoria Inmunológica , Linfocitos T/inmunología , Animales , Antígenos de Diferenciación/genética , Antígenos de Diferenciación/inmunología , Inmunoglobulina G/genética , Inmunoglobulina M/genética , Ratones , Ratones NoqueadosRESUMEN
Primary immune thrombocytopenia (ITP) is a disorder caused by autoantibody-mediated platelet destruction and decreased platelet production. Rituximab, a B cell-depleting agent, has become the first-line treatment for ITP; however, patients with refractory disease usually require splenectomy. We identified antibody-secreting cells as the major splenic B cell population that is resistant to rituximab. The phenotype, antibody specificity, and gene expression profile of these cells were characterized and compared to those of antibody-secreting cells from untreated ITP spleens and from healthy tissues. Antiplatelet-specific plasma cells (PC) were detected in the spleens of patients with ITP up to 6 months after rituximab treatment, and the PC population displayed a long-lived program similar to the one of bone marrow PC, thus explaining for most of these patients the absence of response to rituximab and the response to splenectomy. When analyzed by multiplex PCR at the single-cell level, normal splenic PC showed a markedly different gene expression profile, with an intermediate signature, including genes characteristic of both long-lived PC and proliferating plasmablasts. Surprisingly, long-lived PC were not detected in untreated ITP spleens. These results suggest that the milieu generated by B cell depletion promotes the differentiation and settlement of long-lived PC in the spleen.
Asunto(s)
Anticuerpos Monoclonales de Origen Murino/administración & dosificación , Factores Inmunológicos/administración & dosificación , Depleción Linfocítica , Células Plasmáticas/metabolismo , Púrpura Trombocitopénica Idiopática/metabolismo , Púrpura Trombocitopénica Idiopática/terapia , Bazo/metabolismo , Adulto , Anciano , Autoanticuerpos/sangre , Proliferación Celular , Femenino , Humanos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa Multiplex , Células Plasmáticas/patología , Púrpura Trombocitopénica Idiopática/patología , Rituximab , Bazo/patología , Bazo/cirugía , Esplenectomía , Factores de Tiempo , TranscriptomaRESUMEN
In most cells, the NF-kappaB transcription factor is sequestered in the cytoplasm by interaction with inhibitory proteins, the IkappaBs. Here, we show that combined IkappaBalpha/IkappaBepsilon deficiency in mice leads to neonatal death, elevated kappaB binding activity, overexpression of NF-kappaB target genes, and disruption of lymphocyte production. In IkappaBalpha/IkappaBepsilon-deficient fetuses, B220+IgM+ B cells and single-positive T cells die by apoptosis. In adults, IkappaBalpha-/-IkappaBepsilon-/- reconstituted chimeras exhibit a nearly complete absence of T and B cells that is not rescued by cotransfer with wild-type bone marrow. These findings demonstrate that IkappaBs tightly control NF-kappaB activity in vivo and that increased NF-kappaB activity intrinsically impairs lymphocyte survival. Because reduction or rise of NF-kappaB activity leads to similar dysfunction, they also reveal that only a narrow window of NF-kappaB activity is tolerated by lymphocytes.