RESUMEN
PURPOSE: To define the clinical and histological characteristics of nephritis in patients with X-linked agammaglobulinemia (XLA) and their immunological profiles. METHODS: The clinical, immunological, and histological findings of nine patients with XLA and nephritis were retrospectively analyzed. RESULTS: Based on kidney histological findings, patients with XLA and nephritis could be divided into two groups, viz., chronic glomerulonephritis (CGN) and tubulointerstitial nephritis (TIN). The two groups showed different immunological profiles. Patients in the CGN group exhibited an atypical immunological profile of XLA, with pathogenic leaky B cells producing immunoglobulins that may play a role in forming immune complexes and causing immune-mediated glomerulonephritis. In contrast, patients in the TIN group exhibited a typical immunological profile of XLA, suggesting that antibody-independent/other BTK-dependent mechanisms, or immunoglobulin replacement therapy (IgRT)-related immune/nonimmune-mediated nephrotoxicity causes TIN. CONCLUSION: Nephritis occurring in patients with XLA could have links between their renal pathology and immunological status. Careful observation is recommended to detect kidney pathology in patients with XLA on IgRT.
Asunto(s)
Agammaglobulinemia , Enfermedades Genéticas Ligadas al Cromosoma X , Fenotipo , Humanos , Agammaglobulinemia/inmunología , Agammaglobulinemia/diagnóstico , Agammaglobulinemia/genética , Enfermedades Genéticas Ligadas al Cromosoma X/inmunología , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Enfermedades Genéticas Ligadas al Cromosoma X/diagnóstico , Enfermedades Genéticas Ligadas al Cromosoma X/complicaciones , Masculino , Adolescente , Niño , Adulto , Estudios Retrospectivos , Preescolar , Adulto Joven , Agammaglobulinemia Tirosina Quinasa/genética , Nefritis Intersticial/inmunología , Nefritis Intersticial/diagnóstico , Riñón/patología , Riñón/inmunología , Linfocitos B/inmunología , Femenino , Glomerulonefritis/inmunología , Glomerulonefritis/diagnóstico , Nefritis/inmunología , Nefritis/diagnóstico , Nefritis/etiologíaRESUMEN
T cell receptor beta chain (TCRß) diversity (Dß) gene segments are highly conserved across evolution, with trout Dß1 sequence identical to human and mouse Dß1. A key conserved feature is enrichment for glycine in all three Dß reading frames (RFs). Previously, we found that replacement of mouse Dß1 with a typical immunoglobulin DH sequence, which unlike Dß is enriched for tyrosine, leads to an increase in the use of tyrosine in TCRß complementarity determining region 3 (CDR-B3) after thymic selection, altering T cell numbers, CDR-B3 diversity, and T cell function. To test whether the incorporation of charged amino acids into the Dß sequence in place of glycine would also influence T cell biology, we targeted the TCRß locus with a novel glycine-deficient DßDKRQ allele that replaces Dß1 coding sequence with charged amino acids in all three reading frames. Developing T cells using DßDKRQ expressed TCR CDR-B3s depleted of tyrosine and glycine and enriched for germline-encoded lysine, arginine, and glutamine. Total thymocytes declined in number during the process of ß selection that occurs during the transition from the DN3bc to DN4 stage. Conventional thymocyte and T cell numbers remained reduced at all subsequent thymic stages and in the spleen. By contrast, regulatory T cell numbers were increased in Peyer's patches and the large intestine. In terms of functional consequences, T cell reactivity to an ovalbumin immunodominant epitope was reduced. These findings buttress the view that natural selection of Dß sequence is used to shape the pre-immune TCRß repertoire, affecting both conventional and regulatory T cell development and influencing epitope recognition.
Asunto(s)
Aminoácidos , Regiones Determinantes de Complementariedad , Ratones , Animales , Humanos , Regiones Determinantes de Complementariedad/genética , Aminoácidos/genética , Aminoácidos/metabolismo , Secuencia de Aminoácidos , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Epítopos Inmunodominantes , Células Germinativas/metabolismo , Tirosina/metabolismo , Glicina/metabolismoRESUMEN
Although at first glance the diversity of the immunoglobulin repertoire appears random, there are a number of mechanisms that act to constrain diversity. For example, key mechanisms controlling the diversity of the third complementarity determining region of the immunoglobulin heavy chain (CDR-H3) include natural selection of germline diversity (DH ) gene segment sequence and somatic selection upon passage through successive B-cell developmental checkpoints. To test the role of DH gene segment sequence, we generated a panel of mice limited to the use of a single germline or frameshifted DH gene segment. Specific individual amino acids within core DH gene segment sequence heavily influenced the absolute numbers of developing and mature B-cell subsets, antibody production, epitope recognition, protection against pathogen challenge, and susceptibility to the production of autoreactive antibodies. At the tip of the antigen-binding loop (PDB position 101) in CDR-H3, both natural (germline) and somatic selection favored tyrosine while disfavoring the presence of hydrophobic amino acids. Enrichment for arginine in CDR-H3 appeared to broaden recognition of epitopes of varying hydrophobicity, but led to diminished binding intensity and an increased likelihood of generating potentially pathogenic dsDNA-binding autoreactive antibodies. The phenotype of altering the sequence of the DH was recessive for T-independent antibody production, but dominant for T-cell-dependent responses. Our work suggests that the antibody repertoire is structured, with the sequence of individual DH selected by evolution to preferentially generate an apparently preferred category of antigen-binding sites. The result of this structured approach appears to be a repertoire that has been adapted, or optimized, to produce protective antibodies for a wide range of pathogen epitopes while reducing the likelihood of generating autoreactive specificities.
Asunto(s)
Diversidad de Anticuerpos/genética , Subgrupos de Linfocitos B/inmunología , Sitios de Unión de Anticuerpos/genética , Regiones Determinantes de Complementariedad/genética , Cadenas Pesadas de Inmunoglobulina/genética , Secuencia de Aminoácidos , Animales , Sitios de Unión de Anticuerpos/inmunología , Epítopos/inmunología , Humanos , Cadenas Pesadas de Inmunoglobulina/inmunología , Activación de Linfocitos/inmunología , Ratones , Ratones Transgénicos , Linfocitos T/inmunologíaRESUMEN
Immunoglobulin replacement therapy (IGRT) can protect against lung function decline in CVID. We tested whether increasing IgG dosage was beneficial in patients who exhibited a decline in forced expiratory flow at 25-75% (FEF25-75%) even though they were receiving IgG doses within the therapeutic range. Of 189 CVID patients seen over 12 years, 38 patients met inclusion criteria, were seen on ≥ 3 visits, and demonstrated a ≥ 10% decrease in FEF25-75% from visits 1 to 2. FEF25-75%, forced expiratory flow at 1 s (FEV1), and FEV1/FVC at visit 3 were compared among those with non-dose adjustment (non-DA) versus additional IgG dose adjustment (DA). Three FEF25-75% tiers were identified: top (> 80% predicted), middle (50-80%), and bottom (< 50%). DA and non-DA groups did not differ in clinical infections or bronchodilator use, although the non-DA group tended to use more antibiotics. In the top, normal tier, FEF25-75% increased in DA, but the change did not achieve statistical significance. In the middle moderate obstruction tier, visit 3 FEF25-75% increased among DA but not non-DA sets (11.8 ± 12.4%, p = 0.003 vs. 0.3 ± 9.9%, p = 0.94). Improvement in FEV1/FVC at visit 3 was also significant among DA vs. non-DA (7.2 ± 12.4%, p = 0.04 vs. - 0.2 ± 2.7%, p = 0.85). In the bottom, severe tier, FEF25-75% was unchanged in DA (- 0.5 ± 5.2%, p = 0.79), but increased in non-DA (5.1 ± 5.2%, p = 0.02). Among IGRT CVID patients with moderate but not severe obstruction as assessed by spirometry, increasing IgG dosage led to an increase in FEF25-75% and FEV1/FVC.
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Inmunodeficiencia Variable Común/tratamiento farmacológico , Inmunoglobulina G/uso terapéutico , Inmunoglobulinas Intravenosas/uso terapéutico , Pulmón/metabolismo , Pruebas de Función Respiratoria/métodos , Adulto , Profilaxis Antibiótica , Biomarcadores Farmacológicos , Cálculo de Dosificación de Drogas , Femenino , Humanos , Pulmón/patología , Masculino , Flujo Espiratorio Medio Máximo , Persona de Mediana Edad , Resultado del TratamientoAsunto(s)
Hipersensibilidad , Humanos , Hipersensibilidad/terapia , Alérgenos , Inmunoglobulina E , Inmunoterapia , Proteínas de PlantasRESUMEN
Phelan-McDermid syndrome (PMS, OMIM 606232) is a heterozygous contiguous gene microdeletion syndrome occurring at the distal region of chromosome 22q13. This deletion encompasses the SHANK3 gene at 22q13.33, which is thought to be the critical gene for the neurodevelopmental features seen in this syndrome. PMS is typically characterized by intellectual disability, autism spectrum disorder, absent to severely delayed speech, neonatal hypotonia, and dysmorphic features. Two patients presenting with classic clinical features of PMS have been reported to have interstitial microdeletions in the 22q13.2 region that map proximal to the SHANK3 gene (0.54 and 0.72 Mb, respectively). Here, we describe a 13-month-old girl with a de novo 1.16 Mb interstitial deletion in the 22q13.2 region who presented with global developmental delay, subtle dysmorphic features, and immunodeficiency. This deletion overlaps with the two previously published cases and five cases from the DECIPHER database. All eight patients share features common to patients with PMS including developmental delay and language delay, which suggests that this represents a previously unrecognized microdeletion syndrome in the 22q13.2 region. Our patient's deletion encompasses the TCF20 and TNFRSF13C genes, which are thought to play causative roles in the patient's neurodevelopmental and immunological features, respectively.
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Receptor del Factor Activador de Células B/genética , Trastornos de los Cromosomas/diagnóstico , Trastornos de los Cromosomas/genética , Fenotipo , Factores de Transcripción/genética , Alelos , Deleción Cromosómica , Cromosomas Humanos Par 22/genética , Hibridación Genómica Comparativa , Análisis Citogenético , Femenino , Estudios de Asociación Genética , Humanos , Lactante , MutaciónRESUMEN
PURPOSE: Gain-of-function (GOF) mutations in the signal transducer and activator of transcription 1 (STAT1) result in unbalanced STAT signaling and cause immune dysregulation and immunodeficiency. The latter is often characterized by the susceptibility to recurrent Candida infections, resulting in the clinical picture of chronic mucocutaneous candidiasis (CMC). This study aims to assess the frequency of GOF STAT1 mutations in a large international cohort of CMC patients. METHODS: STAT1 was sequenced in genomic DNA from 57 CMC patients and 35 healthy family members. The functional relevance of nine different STAT1 variants was shown by flow cytometric analysis of STAT1 phosphorylation in patients' peripheral blood cells (PBMC) after stimulation with interferon (IFN)-α, IFN-γ or interleukin-27 respectively. Extended clinical data sets were collected and summarized for 26 patients. RESULTS: Heterozygous mutations within STAT1 were identified in 35 of 57 CMC patients (61%). Out of 39 familial cases from 11 families, 26 patients (67%) from 9 families and out of 18 sporadic cases, 9 patients (50%) were shown to have heterozygous mutations within STAT1. Thirteen distinct STAT1 mutations are reported in this paper. Eight of these mutations are known to cause CMC (p.M202V, p.A267V, p.R274W, p.R274Q, p.T385M, p.K388E, p.N397D, and p.F404Y). However, five STAT1 variants (p.F172L, p.Y287D, p.P293S, p.T385K and p.S466R) have not been reported before in CMC patients. CONCLUSION: STAT1 mutations are frequently observed in patients suffering from CMC. Thus, sequence analysis of STAT1 in CMC patients is advised. Measurement of IFN- or IL-induced STAT1 phosphorylation in PBMC provides a fast and reliable diagnostic tool and should be carried out in addition to genetic testing.
Asunto(s)
Candidiasis Mucocutánea Crónica/diagnóstico , Síndromes de Inmunodeficiencia/diagnóstico , Leucocitos Mononucleares/inmunología , Mutación/genética , Factor de Transcripción STAT1/metabolismo , Adulto , Candidiasis Mucocutánea Crónica/genética , Células Cultivadas , Citocinas/metabolismo , Análisis Mutacional de ADN , Femenino , Humanos , Síndromes de Inmunodeficiencia/genética , Masculino , Linaje , Fenotipo , Estructura Terciaria de Proteína/genética , Factor de Transcripción STAT1/genéticaRESUMEN
Ag activation of the BCR may play a role in the pathogenesis of human follicular lymphoma (FL) and other B cell malignancies. However, the nature of the Ag(s) recognized by tumor BCRs has not been well studied. In this study, we used unbiased approaches to demonstrate that 42 (19.35%) of 217 tested FL Igs recognized vimentin as a shared autoantigen. The epitope was localized to the N-terminal region of vimentin for all vimentin-reactive tumor Igs. We confirmed specific binding to vimentin by using recombinant vimentin and by performing competitive inhibition studies. Furthermore, using indirect immunofluorescence staining, we showed that the vimentin-reactive tumor Igs colocalized with an anti-vimentin mAb in HEp-2 cells. The reactivity to N-terminal vimentin of IgG FL Igs was significantly higher than that of IgM FL Igs (30.4 versus 10%; p = 0.0022). However, vimentin-reactive FL Igs did not share CDR3 motifs and were not homologous. Vimentin was expressed in the T cell-rich regions of FL, suggesting that vimentin is available for binding with tumor BCRs within the tumor microenvironment. Vimentin was also frequently recognized by mantle cell lymphoma and multiple myeloma Igs. Our results demonstrate that vimentin is a shared autoantigen recognized by nonstereotyped FL BCRs and by the Igs of mantle cell lymphoma and multiple myeloma and suggest that vimentin may play a role in the pathogenesis of multiple B cell malignancies. These findings may lead to a better understanding of the biology and natural history of FL and other B cell malignancies.
Asunto(s)
Autoantígenos/inmunología , Linfoma de Células B/inmunología , Receptores de Antígenos de Linfocitos B/inmunología , Vimentina/inmunología , Secuencia de Aminoácidos , Anticuerpos Monoclonales/inmunología , Linfocitos B/inmunología , Linfocitos B/patología , Línea Celular Tumoral , Humanos , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Linfoma de Células B/patología , Linfoma Folicular/inmunología , Linfoma Folicular/patología , Datos de Secuencia Molecular , Mieloma Múltiple/inmunología , Mieloma Múltiple/patología , Linfocitos T/inmunología , Linfocitos T/patologíaAsunto(s)
Enfermedades del Sistema Inmune , Inmunoglobulina E , Alérgenos , Niño , Humanos , Inmunoglobulina G , LactanteRESUMEN
To test whether mechanisms controlling the range of diversity of the developing antibody repertoire in C57BL/6 mice (IgH(b)) operate similarly to those identified in BALB/c mice (IgH(a)), we compared the sequences of VH 7183-containing H-chain transcripts from sorted adult bone marrow C57BL/6 B-cell subsets with those previously obtained from BALB/c mice. Patterns of VDJ gene segment utilization and CDR-H3 amino acid composition, charge, and average length in C57BL/6 pro-B cells were similar, although not identical, to BALB/c pro-B cells. However, C57BL/6 mature, recirculating B cells failed to demonstrate the reduction in the use of VH81X and the narrowing in the range of variance of CDR-H3 hydrophobicity that characterizes B-cell maturation in BALB/c mice. To further test the ability of the C57BL/6 strain to discard B cells expressing highly charged CDR-H3s, we introduced a mutant IgH(a) DH allele that forces use of arginine, asparagine, and histidine. Unlike BALB/c mice, C57BL/6 mice congenic for the charged DH maintained normal numbers of mature, recirculating B cells that were enriched for charged CDR-H3s. Together these findings indicate that the mature C57BL/6 B-cell pool permits expression of immunoglobulins with antigen-binding sites that are typically discarded during late-stage bone marrow B-cell development in BALB/c mice.
Asunto(s)
Linfocitos B/inmunología , Linfocitos B/metabolismo , Células de la Médula Ósea/inmunología , Células de la Médula Ósea/metabolismo , Regiones Determinantes de Complementariedad/química , Cadenas Pesadas de Inmunoglobulina/química , Animales , Diversidad de Anticuerpos/genética , Diversidad de Anticuerpos/inmunología , Subgrupos de Linfocitos B/citología , Subgrupos de Linfocitos B/inmunología , Subgrupos de Linfocitos B/metabolismo , Linfocitos B/citología , Células de la Médula Ósea/citología , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Codón , Regiones Determinantes de Complementariedad/genética , Regiones Determinantes de Complementariedad/inmunología , Femenino , Reordenamiento Génico de Cadena Pesada de Linfocito B/inmunología , Interacciones Hidrofóbicas e Hidrofílicas , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Sistemas de LecturaRESUMEN
After birth, contact to environmental Ags induces the production of IgA, which represents a first line of defense for the neonate. We sought to characterize the maturation of the repertoire of IgA H chain transcripts in circulating blood B cells during human ontogeny. We found that IgA H chain transcripts were present in cord blood as early as 27 wk of gestation and that the restrictions of the primary Ab repertoire (IgM) persisted in the IgA repertoire. Thus, B cells harboring more "mature" V(H) regions were not preferred for class switch to IgA. Preterm and term neonates expressed a unique IgA repertoire, which was characterized by short CDR-H3 regions, preference of the J(H) proximal D(H)7-27 gene segment, and very few somatic mutations. During the first postnatal months, these restrictions were slowly released. Preterm birth did not measurably accelerate the maturation of the IgA repertoire. At a postconceptional age of 60 wk, somatic mutation frequency of IgA H chain transcripts reached 25% of the adult values but still showed little evidence of Ag-driven selection. These results indicate that similar to IgG, the IgA repertoire expands in a controlled manner after birth. Thus, the IgA repertoire of the newborn has distinctive characteristics that differ from the adult IgA repertoire. These observations might explain the lower affinity and specificity of neonatal IgA Abs, which could contribute to a higher susceptibility to infections and altered responses to vaccinations, but might also prevent the development of autoimmune and allergic diseases.
Asunto(s)
Antígenos/inmunología , Linfocitos B/inmunología , Regulación del Desarrollo de la Expresión Génica/inmunología , Inmunoglobulina A/genética , Cadenas Pesadas de Inmunoglobulina/genética , ARN Mensajero/inmunología , Adulto , Afinidad de Anticuerpos , Diversidad de Anticuerpos , Especificidad de Anticuerpos , Linfocitos B/metabolismo , Secuencia de Bases , Sangre Fetal , Edad Gestacional , Humanos , Inmunidad Innata , Cambio de Clase de Inmunoglobulina , Inmunoglobulina M/genética , Región Variable de Inmunoglobulina/genética , Lactante , Recién Nacido , Recien Nacido Prematuro , Datos de Secuencia Molecular , ARN Mensajero/biosíntesisRESUMEN
Antigen affinity is commonly viewed as the driving force behind the selection for dominant clonotypes that can occur during the T-cell-dependent processes of class switch recombination (CSR) and immune maturation. To test this view, we analyzed the variable gene repertoires of natural monoclonal antibodies to the hapten 2-phenyloxazolone (phOx) as well as those generated after phOx protein carrier-induced thymus-dependent or Ficoll-induced thymus-independent antigen stimulation. In contrast to expectations, the extent of IgM heterogeneity proved similar and many IgM from these three populations exhibited similar or even greater affinities than the classic Ox1 clonotype that dominates only after CSR among primary and memory IgG. The population of clones that were selected during CSR exhibited a reduced VH/VL repertoire that was enriched for variable domains with shorter and more uniform CDR-H3 lengths and almost completely stripped of variable domains encoded by the large VH1 family. Thus, contrary to the current paradigm, T-cell-dependent clonal selection during CSR appeared to select for VH family and CDR-H3 loop content even when the affinity provided by alternative clones exhibited similar to increased affinity for antigen.
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Anticuerpos Monoclonales de Origen Murino/inmunología , Cambio de Clase de Inmunoglobulina/inmunología , Receptores de Antígenos de Linfocitos B/inmunología , Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales de Origen Murino/genética , Regiones Determinantes de Complementariedad/genética , Regiones Determinantes de Complementariedad/inmunología , Femenino , Haptenos/inmunología , Haptenos/farmacología , Cambio de Clase de Inmunoglobulina/efectos de los fármacos , Cambio de Clase de Inmunoglobulina/genética , Inmunoglobulina G/genética , Inmunoglobulina G/inmunología , Inmunoglobulina M/genética , Inmunoglobulina M/inmunología , Memoria Inmunológica/efectos de los fármacos , Memoria Inmunológica/genética , Memoria Inmunológica/inmunología , Ratones , Ratones Endogámicos BALB C , Receptores de Orexina , Oxazoles/inmunología , Oxazoles/farmacología , Receptores de Antígenos de Linfocitos B/genética , Receptores de Neuropéptido/genética , Receptores de Neuropéptido/inmunologíaRESUMEN
Anti-polysaccharide Ab responses in mice are often oligoclonal, and the mechanisms involved in Ag-specific clone production and selection remain poorly understood. We evaluated the relative contribution of D(H) germline content versus N nucleotide addition in a classic oligoclonal, T-independent Ab response (α 1â3 dextran [DEX]) by challenging adult TdT-sufficient (TdT(+/+)) and TdT-deficient (TdT(-/-)) gene-targeted mice, limited to the use of a single D(H) gene segment (D-limited mice), with Enterobacter cloacae. D-limited mice achieved anti-DEX-specific levels of Abs that were broadly comparable to those of wild-type (WT) BALB/c mice. Sequence analysis of the third CDR of the H chain intervals obtained by PCR amplification of V(H) domain DNA from DEX-specific plasmablasts revealed the near universal presence of an aspartic acid residue (D99) at the V-D junction, irrespective of the composition of the D(H) locus. Although WT mice were able to use germline D(H) (DQ52, DSP, or DST) gene segment sequence, TdT activity, or both to produce D99, all three D-limited mouse strains relied exclusively on N addition. Additionally, in the absence of TdT, D-limited mice failed to produce a DEX response. Coupled with previous studies demonstrating a reduced response to DEX in TdT(-/-) mice with a WT D(H) locus, we concluded that in the case of the anti-DEX repertoire, which uses a short third CDR of the H chain, the anti-DEX response relies more intensely on sequences created by postnatal N nucleotide addition than on the germline sequence of the D(H).
Asunto(s)
Anticuerpos Antibacterianos/biosíntesis , Diversidad de Anticuerpos , Regiones Determinantes de Complementariedad/biosíntesis , Regiones Determinantes de Complementariedad/inmunología , Dextranos/inmunología , Cadenas Pesadas de Inmunoglobulina/biosíntesis , Polisacáridos Bacterianos/inmunología , Sustitución de Aminoácidos/genética , Sustitución de Aminoácidos/inmunología , Animales , Anticuerpos Antibacterianos/genética , Diversidad de Anticuerpos/genética , Secuencia de Bases , Regiones Determinantes de Complementariedad/genética , Dextranos/administración & dosificación , Dextranos/genética , Enterobacter cloacae/inmunología , Reordenamiento Génico de Linfocito B/inmunología , Cadenas Pesadas de Inmunoglobulina/genética , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Polisacáridos Bacterianos/administración & dosificación , Polisacáridos Bacterianos/genéticaRESUMEN
Developing T cells face a series of cell fate choices in the thymus and in the periphery. The role of the individual T cell receptor (TCR) in determining decisions of cell fate remains unresolved. The stochastic/selection model postulates that the initial fate of the cell is independent of TCR specificity, with survival dependent on additional TCR/coreceptor "rescue" signals. The "instructive" model holds that cell fate is initiated by the interaction of the TCR with a cognate peptide-MHC complex. T cells are then segregated on the basis of TCR specificity with the aid of critical coreceptors and signal modulators [Chan S, Correia-Neves M, Benoist C, Mathis (1998) Immunol Rev 165: 195-207]. The former would predict a random representation of individual TCR across divergent T cell lineages whereas the latter would predict minimal overlap between divergent T cell subsets. To address this issue, we have used high-throughput sequencing to evaluate the TCR distribution among key T cell developmental and effector subsets from a single donor. We found numerous examples of individual subsets sharing identical TCR sequence, supporting a model of a stochastic process of cell fate determination coupled with dynamic patterns of clonal expansion of T cells bearing the same TCR sequence among both CD4(+) and CD8+ populations.
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Receptores de Antígenos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/inmunología , Secuencia de Aminoácidos , Diferenciación Celular , Linaje de la Célula , Humanos , Receptores de Antígenos de Linfocitos T/química , Subgrupos de Linfocitos T/química , Subgrupos de Linfocitos T/citologíaRESUMEN
Intracellular trafficking of organelles often involves cytoskeletal track switching. Organelles such as melanosomes are transported by multiple motors including kinesin-2, dynein, and myosin-V, which drive switching between microtubules and actin filaments during dispersion and aggregation. Here, we used optical trapping to determine the unitary and ensemble forces of kinesin-2, and to reconstitute cargo switching at cytoskeletal intersections in a minimal system with kinesin-2 and myosin-V motors bound to beads. Single kinesin-2 motors exerted forces up to â¼5 pN, similar to kinesin-1. However, kinesin-2 motors were more likely to detach at submaximal forces, and the duration of force maintenance was short as compared to kinesin-1. In multimotor assays, force increased with kinesin-2 density but was not affected by the presence of myosin-V. In crossed filament assays, switching frequencies of motor-bound beads were dependent on the starting track. At equal average forces, beads tended to switch from microtubules onto overlying actin filaments consistent with the relatively faster detachment of kinesin-2 at near-maximal forces. Thus, in addition to relative force, switching probability at filament intersections is determined by the dynamics of motor-filament interaction, such as the quick detachment of kinesin-2 under load. This may enable fine-tuning of filament switching in the cell.
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Citoesqueleto de Actina/fisiología , Cinesinas/fisiología , Microtúbulos/fisiología , Proteínas de Xenopus/fisiología , Citoesqueleto de Actina/química , Animales , Cinesinas/química , Microscopía Fluorescente , Simulación de Dinámica Molecular , Miosina Tipo V/química , Miosina Tipo V/fisiología , Conformación Proteica , Conejos , Xenopus , Proteínas de Xenopus/químicaRESUMEN
Tyrosine and glycine constitute 40% of complementarity determining region 3 of the immunoglobulin heavy chain (CDR-H3), the center of the classic antigen-binding site. To assess the role of D(H) RF1-encoded tyrosine and glycine in regulating CDR-H3 content and potentially influencing B cell function, we created mice limited to a single D(H) encoding asparagine, histidine, and arginines in RF1. Tyrosine and glycine content in CDR-H3 was halved. Bone marrow and spleen mature B cell and peritoneal cavity B-1 cell numbers were also halved, whereas marginal zone B cell numbers increased. Serum immunoglobulin G subclass levels and antibody titers to T-dependent and T-independent antigens all declined. Thus, violation of the conserved preference for tyrosine and glycine in D(H) RF1 alters CDR-H3 content and impairs B cell development and antibody production.
Asunto(s)
Formación de Anticuerpos/inmunología , Linfocitos B/inmunología , Cadenas Pesadas de Inmunoglobulina/inmunología , Secuencia de Aminoácidos , Aminoácidos/inmunología , Animales , Cadenas Pesadas de Inmunoglobulina/genética , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia MolecularRESUMEN
To assess the extent and nature of somatic categorical selection of CDR-3 of the Ig H chain (CDR-H3) content in peritoneal cavity (PerC) B cells, we analyzed the composition of V(H)7183DJCµ transcripts derived from sorted PerC B-1a, B-1b, and B-2 cells. We divided these sequences into those that contained N nucleotides (N(+)) and those that did not (N(-)) and then compared them with sequences cloned from sorted IgM(+)IgD(+) B cells from neonatal liver and both wild-type and TdT-deficient adult bone marrow. We found that the PerC B-1a N(-) repertoire is enriched for the signatures of CDR-H3 sequences present in neonatal liver and shares many features with the B-1b N(-) repertoire, whereas the PerC B-1a N(+), B-1b N(+), and B-2 N(+) repertoires are enriched for adult bone marrow sequence signatures. However, we also found several sequence signatures that were not shared with other mature perinatal or adult B cell subsets but were either unique or variably shared between the two or even among all three of the PerC subsets that we examined. These signatures included more sequences lacking N nucleotides in the B-2 population and an increased use of D(H) reading frame 2, which created CDR-H3s of greater average hydrophobicity. These findings provide support for both ontogenetic origin and shared Ag receptor-influenced selection as the mechanisms that shape the unique composition of the B-1a, B-1b, and B-2 repertoires. The PerC may thus serve as a general reservoir for B cells with Ag binding specificities that are uncommon in other mature compartments.
Asunto(s)
Subgrupos de Linfocitos B/inmunología , Linfocitos B/inmunología , Regiones Determinantes de Complementariedad/inmunología , Cavidad Peritoneal/citología , Células Precursoras de Linfocitos B/inmunología , Animales , Diversidad de Anticuerpos , Subgrupos de Linfocitos B/química , Linfocitos B/química , Secuencia de Bases , Separación Celular , Regiones Determinantes de Complementariedad/química , Regiones Determinantes de Complementariedad/genética , Citometría de Flujo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Células Precursoras de Linfocitos B/química , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Compared with adult bone marrow (BM), the composition of the perinatal liver CDR-3 of the Ig H chain (CDR-H3) repertoire is marked by a paucity of N nucleotides and by enrichment for use of J(H) proximal DQ52 and D(H) proximal V(H) and J(H) gene segments. To test the extent to which these differences reflect limited perinatal TdT activity versus differences in the fetal/adult environment, we used the Hardy scheme to sort fractions B-F B lineage cells from TdT-deficient BALB/c adult BM. V(H)7183-containing VDJCµ transcripts from these cells were amplified, cloned, sequenced, and compared with transcripts from wild-type perinatal liver and adult BM. The pattern of V(H)DJ(H) usage in TdT-deficient BM largely matched that of TdT-sufficient adult cells. What minor differences were detected in the pro-B cell stage tended to diminish with B cell maturation, suggesting strong environmental or Ag-driven pressure to achieve a specific range of V(H)DJ(H) usage regardless of the extent of N nucleotide addition. However, although the patterns of V(H)DJ(H) usage in the TdT-deficient B lineage cells paralleled that of wild-type adult cells, the length distribution, global amino acid composition, and charge distribution of the CDR-H3 repertoire proved to be a close, although not exact, homologue of the CDR-H3 repertoire first expressed by late pre-B cells in the TdT-insufficient perinatal liver. Thus, although differing in V(H) content, TdT-deficient mice appear to represent a good, although not perfect, model for testing the role of perinatal CDR-H3 limitations on late B cell development and Ab responses.
Asunto(s)
Médula Ósea/inmunología , Regiones Determinantes de Complementariedad/genética , ADN Nucleotidilexotransferasa/deficiencia , Hígado/inmunología , Células Precursoras de Linfocitos B/inmunología , Secuencia de Aminoácidos , Animales , Linfocitos B/química , Linfocitos B/inmunología , Separación Celular , Regiones Determinantes de Complementariedad/química , Regiones Determinantes de Complementariedad/inmunología , Citometría de Flujo , Cadenas Pesadas de Inmunoglobulina/química , Cadenas Pesadas de Inmunoglobulina/inmunología , Hígado/citología , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Células Precursoras de Linfocitos B/química , Reacción en Cadena de la Polimerasa de Transcriptasa InversaAsunto(s)
Inmunodeficiencia Variable Común/genética , Inmunodeficiencia Variable Común/inmunología , Receptores KIR/genética , Receptores KIR/inmunología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Preescolar , Femenino , Genes MHC Clase I/inmunología , Genotipo , Humanos , Células Asesinas Naturales/inmunología , Masculino , Persona de Mediana Edad , Subgrupos de Linfocitos T/inmunología , Adulto JovenRESUMEN
The early B cell protein λ5 is an essential component of the surrogate light chain and the preB cell receptor (preBCR), which is critical for optimal B cell development. To investigate the effect of λ5 and/or B cells on bone acquisition over time, we developed a panel of JH -/- , λ5-/-, JH -/- λ5-/-, and wild-type (WT) BALB/c mice and then studied postnatal bone development and aging in these mice at one, six, twelve, and twenty-two months of age. The trabecular bone volume over total volume (BV/TV) in JH -/- mice was similar to WT mice at all ages. In contrast, at six months of age and thereafter, λ5-/- and JH -/- λ5-/- mice demonstrated a severe decrease in trabecular bone mass. Surprisingly, bone mass in six-month-old λ5-/- and JH -/- λ5-/- mice was similar to or even lower than in aged (twenty-two-months) WT mice, suggesting accelerated skeletal aging. The postnatal development and the acquisition of cortical bone mass in JH -/- λ5-/- mice were generally comparable to WT. However, JH -/- λ5-/- mice showed a significant decrease in cortical BV/TV at six- and twelve months of age. To examine the contribution of λ5 and B cells to postnatal bone synthesis, we separately transplanted whole bone marrow cells from JH -/- λ5-/- and WT mice into irradiated JH -/- λ5-/- and WT recipients. WT recipients of JH -/- λ5-/- marrow cells failed to show acquisition of trabecular bone mass, whereas transplanting WT marrow cells into JH -/- λ5-/- recipients led to the recovery of trabecular bone mass. Transfer of WT marrow cells into JH -/- λ5-/- mice promoted synthesis of new cortical and trabecular bone. Our findings indicate that λ5 plays a major role in preserving bone mass during postnatal development and skeletal aging which is distinct from its role in B cell development. The absence of both λ5 and B cells in JH -/- λ5-/- mice leads to delayed acquisition of cortical bone during postnatal development. Dissecting the mechanism(s) by which λ5 regulates bone homeostasis may provide new avenues for the treatment of age-related loss of bone mass and osteoporosis.