scholarly journals Secondary Rearrangements and Hypermutation Generate Sufficient B Cell Diversity to Mount Protective Antiviral Immunoglobulin Responses

1999 ◽  
Vol 189 (11) ◽  
pp. 1791-1798 ◽  
Author(s):  
Constantino López-Macías ◽  
Ulrich Kalinke ◽  
Marilia Cascalho ◽  
Matthias Wabl ◽  
Hans Hengartner ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Gene Replacement ◽  
Antibody Responses ◽  
Single Chain ◽  
Cell Repertoire ◽  
Region Gene ◽  
B Cell Repertoire ◽  
Single Chain Fv ◽  
Cell Diversity

Variable (V) region gene replacement was recently implicated in B cell repertoire diversification, but the contribution of this mechanism to antibody responses is still unknown. To investigate the role of V gene replacements in the generation of antigen-specific antibodies, we analyzed antiviral immunoglobulin responses of “quasimonoclonal” (QM) mice. The B cells of QM mice are genetically committed to exclusively express the anti-(4-hydroxy-3-nitrophenyl) acetyl specificity. However, ∼20% of the peripheral B cells of QM mice undergo secondary rearrangements and thereby potentially acquire new specificities. QM mice infected with vesicular stomatitis virus (VSV), lymphocytic choriomeningitis virus, or poliovirus mounted virus-specific neutralizing antibody responses. In general, kinetics of the antiviral immunoglobulin responses were delayed in QM mice; however, titers similar to control animals were eventually produced that were sufficient to protect against VSV-induced lethal disease. VSV neutralizing single-chain Fv fragments isolated from phage display libraries constructed from QM mice showed VH gene replacements and extensive hypermutation. Thus, our data demonstrate that secondary rearrangements and hypermutation can generate sufficient B cell diversity in QM mice to mount protective antiviral antibody responses, suggesting that these mechanisms might also contribute to the diversification of the B cell repertoire of normal mice.

scholarly journals Effects of IgM Allotype Suppression on Serum IgM Levels, B-1 and B-2 Cells, and Antibody Responses ir Allotype Heterozygous F1 Mice

1994 ◽  
Vol 4 (1) ◽  
pp. 27-41 ◽  
Author(s):  
Ann Marie Hamilton ◽  
John F. Kearney
Keyword(s):  
B Cells ◽  
B Cell ◽  
Antibody Responses ◽  
Treatment Results ◽  
Cell Repertoire ◽  
Antibody Treatment ◽  
B Cell Repertoire ◽  
Dependent Form

IgM allotype heterozygous F1 mice were independently suppressed for Igh6a or Igh6b to evaluate the contribution of B-1 and B-2 cells to natural serum IgM levels and Ab responses. B-2 B cells expressing IgM of the suppressed allotype were evident in the spleens of suppressed mice 4 to 6 weeks after cessation of the suppression regimen, whereas B-1 B cells of the suppressed allotype were undetectable for up to 9 months. Although serum IgM of the suppressed allotype was initially depleted in mice suppressed for either allotype, by 7 months of age, there were detectable levels of IgM of the suppressed allotype in the serum; however, the levels were significantly below that found in nonsuppressed mice. When mice were immunized with either the T-independent or T-dependent form of phosphorylcholine, those suppressed for either allotype, and consequently depleted of B-1 B cells of that allotype, did not respond with phosphorylcholine-specific IgM of the suppressed allotype. In contrast, when mice were immunized with α1-3 dextran, the Igh6a allotype-suppressed mice were able to produce dextran-specific IgM of that allotype. These results show that allotype-bearing B-1 cells of both allotypes can be effectively suppressed by this suppression protocol and this produces long-lasting effects on B-1 cell levels and serum IgM of the suppressed allotype. These observations reflect the derivation of the majority of B-1 cells from fetal-neonatal precursors, which cannot be replaced by newly emerging B-2 cells of adult origin. Their ablation by antibody treatment results in permanent alterations to the adult B-cell repertoire.

scholarly journals Role of variable region gene expression and environmental selection in determining the antiphosphorylcholine B cell repertoire.

1983 ◽  
Vol 158 (6) ◽  
pp. 1948-1961 ◽  
Author(s):  
N R Klinman ◽  
M R Stone
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Bone Marrow Cells ◽  
Marrow Cell ◽  
Variable Region ◽  
Precursor Cells ◽  
Cell Repertoire ◽  
Region Gene ◽  
B Cell Repertoire

To evaluate the role of environmental selective processes, as opposed to variable region gene expression, in the determination of B cell repertoire expression, we have assessed the phosphorylcholine (PC)-specific repertoire of precursor cells that remain in bone marrow cell populations after the removal of surface immunoglobulin (sIg)-bearing cells. Such cells are assumed to represent a stage in B cell maturation before the expression of sIg, and thus at a time when they have not as yet interfaced with environmental influences that operate through sIg receptors such as antigenic stimulation, tolerance, or antiidiotypic regulation. The repertoire as expressed in these cells, therefore, should reflect the readout of immunoglobulin variable region genes as they are expressed in progenitors to B cells. The results of these studies indicate that, as in mature primary B cell pools of BALB/c mice, the majority of PC-responsive sIg- bone marrow cells are of the T15 clonotype. Thus, environmental selective mechanisms would not appear to be required for the high frequency of B cells of the T15 idiotype in the primary B cell repertoire of BALB/c mice. Analysis of the sIg- bone marrow cells in (CBA/N X BALB/c)F1 male mice demonstrated that the deficit of PC-responsive mature B cells, which is a characteristic of this murine strain, must occur after receptor expression, since a normal frequency of PC-responsive and T15-expressing cells is present in their sIg- bone marrow population. Finally, these same mice were used to obtain bone marrow cell preparations from individual leg bones, so as to permit an analysis of the occurrence of T15+ and T15- clonotypes within individual bone marrow populations. The findings from these studies indicate that T15+ B cells occur as a high frequency event within bone marrow generative cell pools. Furthermore, bone marrow populations that are positive for PC-responsive precursor cells often display multiple copies of such precursor cells that are exclusively either T15+ or T15-. This finding indicates that clonal expansion of cells within the B cell lineage apparently occurs before immunoglobulin receptor acquisition.

scholarly journals Efficient Peripheral Clonal Elimination of B Lymphocytes in MRL/lpr Mice Bearing Autoantibody Transgenes

1998 ◽  
Vol 188 (5) ◽  
pp. 909-917 ◽  
Author(s):  
Jennifer A. Kench ◽  
David M. Russell ◽  
David Nemazee
Keyword(s):  
B Cells ◽  
B Cell ◽  
Genetic Background ◽  
Cell Tolerance ◽  
Cell Repertoire ◽  
B Cell Tolerance ◽  
B Cell Repertoire ◽  
Nuclear Antigens ◽  
Large Numbers ◽  
Liver And Kidney

Peripheral B cell tolerance was studied in mice of the autoimmune-prone, Fas-deficient MRL/ lpr.H-2d genetic background by introducing a transgene that directs expression of membrane-bound H-2Kb antigen to liver and kidney (MT-Kb) and a second transgene encoding antibody reactive with this antigen (3-83μδ, anti-Kk,b). Control immunoglobulin transgenic (Ig-Tg) MRL/lpr.H-2d mice lacking the Kb antigen had large numbers of splenic and lymph node B cells bearing the transgene-encoded specificity, whereas B cells of the double transgenic (Dbl-Tg) MRL/lpr.H-2d mice were deleted as efficiently as in Dbl-Tg mice of a nonautoimmune B10.D2 genetic background. In spite of the severely restricted peripheral B cell repertoire of the Ig-Tg MRL/lpr.H-2d mice, and notwithstanding deletion of the autospecific B cell population in the Dbl-Tg MRL/lpr.H-2d mice, both types of mice developed lymphoproliferation and exhibited elevated levels of IgG anti-chromatin autoantibodies. Interestingly, Dbl-Tg MRL/lpr.H-2d mice had a shorter lifespan than Ig-Tg MRL/lpr.H-2d mice, apparently as an indirect result of their relative B cell lymphopenia. These data suggest that in MRL/lpr mice peripheral B cell tolerance is not globally defective, but that certain B cells with receptors specific for nuclear antigens are regulated differently than are cells reactive to membrane autoantigens.

scholarly journals Idiotypic vaccination against human B-cell lymphoma. Rescue of variable region gene sequences from biopsy material for assembly as single-chain Fv personal vaccines

Blood ◽  
1994 ◽  
Vol 83 (11) ◽  
pp. 3279-3288 ◽  
Author(s):  
RE Hawkins ◽  
D Zhu ◽  
M Ovecka ◽  
G Winter ◽  
TJ Hamblin ◽  
...  
Keyword(s):  
B Cell ◽  
Tumor Antigens ◽  
Variable Region ◽  
B Cell Lymphoma ◽  
Repeated Sequences ◽  
Expression Vectors ◽  
Single Chain ◽  
Region Gene ◽  
Biopsy Material ◽  
Single Chain Fv

Abstract Idiotypic determinants on neoplastic B cells could provide tumor antigens for vaccination of patients with B-cell tumors. Because this approach requires an individual vaccine for each patient, simple methods for obtaining idiotypic antigen are desirable. Using polymerase chain reaction (PCR) with family-based V-gene and J-region primers, the variable region genes of heavy and light chains (VH and VL) of Ig have been obtained from biopsy material from 13 patients with B-cell tumors. In each case, analysis of random clones derived from the PCR product showed repeated, clonally-related sequences, whereas normal lymphoid tissue generated no repeated sequences. In 3/3 cases, the repeated sequences were found to be the same as those in a tumor-derived hybridoma. Mutational patterns in the V-genes differed among the tumors, with follicular lymphoma tending to be more highly mutated. The individual VH and VL sequences have been assembled with a flexible linker sequence to encode single-chain Fv (scFv). The scFv sequences can be cloned into bacterial expression vectors to produce protein, or into vectors suitable for direct vaccination using naked DNA. In a model system, expressed scFv protein retained all idiotypic determinants defined by a panel of five anti-idiotypic monoclonal antibodies (MoAbs). Similarly, expressed scFv proteins from two patients were shown to react with anti-idiotypic antibodies. This approach allows production of potential vaccines from surgical biopsies within 2 to 3 weeks.

scholarly journals High-frequency representation of a single VH gene in the expressed human B cell repertoire.

1993 ◽  
Vol 177 (2) ◽  
pp. 409-418 ◽  
Author(s):  
A K Stewart ◽  
C Huang ◽  
B D Stollar ◽  
R S Schwartz
Keyword(s):  
B Cells ◽  
B Cell ◽  
High Frequency ◽  
Gene Segment ◽  
Normal Subjects ◽  
Cell Repertoire ◽  
B Cell Repertoire ◽  
V Genes ◽  
Vh Gene ◽  
Vh Genes

Idiotype (Id) 16/6 marks a variable (V) region structure that occurs frequently in the human immunoglobulin repertoire. The basis of the Id has been traced to a germline heavy chain gene segment, VH18/2 (VH26). To pursue the molecular basis for the frequency of Id 16/6, we have analyzed polymerase chain reaction-generated C mu, C gamma, and VH3 family V gene libraries derived from the circulating and tonsillar B cells of four normal individuals and from the B cells of two patients with active systemic lupus erythematosus (SLE). The frequency of VH18/2 in these libraries was compared with three control VH genes, VH56P1, VH21/28, and VHA57. Plaque lifts from C mu and C gamma VH cDNA libraries were screened with gene-specific oligonucleotide probes. The frequency of VH18/2 ranged from 4 to 10% of JH+ plaques (two of five times that of control VH genes). In four VH3 family-specific libraries derived from rearranged DNA, VH18/2 represented 19-33% of VH3+ plaques. Hybridizing VH18/2 plaques were 98-100% homologous to the germline VH gene; mutations when present were often in framework 3. Extensive variation was seen in the complementarity determining region 3 sequences of these rearranged V genes. The high frequency of VH18/2 expression in the B cell repertoire was confirmed by sequencing randomly picked JH+ plaques. In two patients with active SLE the frequency of use of VH18/2 was not greater than that observed in normal subjects. These results show that VH18/2 is overrepresented in the B cell repertoire of normal subjects and suggest that the immune repertoire may be dominated by relatively few V genes.

scholarly journals Shared HIV envelope-specific B cell clonotypes induced by a pox-protein vaccine regimen

2021 ◽  
Author(s):  
Kristen W. Cohen ◽  
Lamar Ballweber-Fleming ◽  
Michael Duff ◽  
Rachael E. Whaley ◽  
Aaron Seese ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Neutralizing Antibodies ◽  
Rational Design ◽  
Critical Role ◽  
Memory B Cells ◽  
Protein Vaccine ◽  
Cell Repertoire ◽  
B Cell Repertoire ◽  
Specific Igg

An effective HIV-1 vaccine will likely induce potent, broad neutralizing antibodies. No candidate vaccines have elicited these responses presumably because they fail to activate human B cell precursors that can affinity mature to generate broad neutralizing antibodies. To identify the B cell clonotypes that are elicited, we conducted in-depth analyses of the envelope-specific B cell repertoire in recipients of ALVAC-HIV vector (vCP2438) and bivalent subtype C gp120 protein (HVTN100). We observed high frequencies of envelope-specific IgG+ memory B cells with restricted immunogenetic diversity, relative to non-vaccine induced memory B cells, with preferential expansions of distinct variable genes but limited accumulation of mutations. Many envelope-specific clonotypes were shared across vaccinees, but did not overlap with the envelope-negative memory repertoire, within and across subjects. Single-cell sequencing of envelope-specific IgG+ memory B cells often revealed VH1-2*02 and VK3-20 sequence co-expression and in one case, contained a 5 amino acid CDRL3, the canonical signature of VRC01-class antibodies, confirming that these B cells are extremely rare but detectable. Our study provides evidence that immunogens play a critical role in selecting and restricting the responding B cell repertoire and supports the rational design of HIV vaccines targeting specific B cell lineages for induction of broadly-reactive neutralizing antibodies.

scholarly journals Opposite Effects of Bruton Tyrosine Kinase (BTK) Inhibitor Therapy with Ibrutinib and FCR Chemo-Immunotherapy on the Normal B Lymphocyte Repertoire in Patients with Chronic Lymphocytic Leukemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4402-4402
Author(s):  
Simon Schliffke ◽  
Mariela Sivina ◽  
Ekaterina Kim ◽  
Benjamin Thiele ◽  
Nuray Akyüz ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Research Funding ◽  
Treatment Initiation ◽  
Lymphocytic Leukemia ◽  
Cell Repertoire ◽  
B Cell Repertoire ◽  
Btk Inhibitor ◽  
V Gene ◽  
Immunoglobulin Levels

Abstract Disease-inherent and treatment-related immune dysfunction remain leading causes for morbidity and mortality in patients with chronic lymphocytic leukemia (CLL). The advent of kinase inhibitors that target B cell receptor (BCR) signaling, which lack myelo- and T lymphocyte toxicity, raised hopes that these new agents may be less immunosuppressive and allow for better immune reconstitution when compared to chemo-immunotherapy (CIT). The effects of the BTK inhibitor ibrutinib or CIT with fludarabine, cyclophosphamide and rituximab (FCR) on the normal B cell repertoire have not been well characterized. Here, we used state-of-the-art immunosequencing technology to investigate how ibrutinib treatment affects the regeneration of non-malignant B-cells when compared to patients treated with FCR. Clinical data on infection rates and immunoglobulin levels was analyzed from 40 CLL patients treated with ibrutinib (median number of two pre-treatments) or frontline CIT with FCR at MD Anderson Cancer Center. In a representative subset of 20 patients, flow cytometry and next generation sequencing (NGS) of the immunoglobulin heavy chain (IGH) gene locus was used to monitor non-malignant B-cell immune reconstitution for 24 months after start of treatment with ibrutinib or FCR. Comparison of ibrutinib treatment with CIT revealed that immunoglobulin levels remained stable and relatively low in both cohorts, except for an increase in IgA during ibrutinib treatment, as previously reported. NGS results showed that ibrutinib treatment significantly decreased the non-malignant B-cells count after 24 months of treatment, while the counts were quantitatively stable in the FCR cohort. Next, we determined the dynamics of non-malignant B-cell immune repertoire composition over treatment. Based on the mutational status of the V gene, non-malignant B-cells were classified as IGH hypermutated (<98% identity to the corresponding germline V gene, corresponding to antigen-experienced B-cells) or IGH unmutated (≥98% identity to the corresponding germline V gene, corresponding to antigen-naïve B-cells). Before treatment initiation, the mean percentage of antigen-experienced B-cells did not significantly differ between the groups (ibrutinib 39%, FCR 48%). After 24 months, a significant decrease of antigen-experienced B-cells was observed in the FCR cohort, while the ratio of antigen-experienced and antigen-naïve B-cells remained unchanged in ibrutinib treated patients (ibrutinib 39%, FCR 22%, p=0.01). Analysis of the IGH clonotype repertoire using the Shannon-Wiener and the inverse Simpson diversity indices confirmed these results, showing that the non-malignant IGH repertoire was composed of balanced numbers of antigen-experienced and antigen-naïve medium sized clones before treatment initiation in both cohorts. In line with the IGH repertoire shift towards antigen-naïve B-cells in FCR treated patients, the medium-sized clones disappeared after treatment, with large numbers of small-sized unmutated clones dominating after 24 months (p<0.0001). In ibrutinib treated patients, the repertoire diversity remained stable throughout the course of treatment. Taken together, our data indicate that continuous treatment with ibrutinib preserves preexisting (partially antigen-experienced) B-cells but impairs de-novo generation of naive B-cells. In contrast, FCR leads to a deletion of memory B-cells but also a subsequent substantial renewal of the B-cell repertoire. Both patterns may differentially affect immune-competence towards infections. Disclosures Bokemeyer: Karyopharm: Research Funding. Jain:Pfizer: Consultancy, Honoraria, Research Funding; Incyte: Research Funding; Genentech: Research Funding; Abbvie: Research Funding; Pharmacyclics: Consultancy, Honoraria, Research Funding; Infinity: Research Funding; Novartis: Consultancy, Honoraria; Servier: Consultancy, Honoraria; Novimmune: Consultancy, Honoraria; ADC Therapeutics: Consultancy, Honoraria, Research Funding; BMS: Research Funding; Celgene: Research Funding; Seattle Genetics: Research Funding. Wierda:Gilead: Research Funding; Abbvie: Research Funding; Novartis: Research Funding; Acerta: Research Funding; Genentech: Research Funding. Burger:Pharmacyclics: Research Funding.

Bim establishes the B-cell repertoire from early to late in the immune response

2020 ◽  
Author(s):  
Akiko Sugimoto-Ishige ◽  
Michishige Harada ◽  
Miho Tanaka ◽  
Tommy Terooatea ◽  
Yu Adachi ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Plasma Cells ◽  
Affinity Maturation ◽  
Late Response ◽  
Cell Repertoire ◽  
High Affinity ◽  
B Cell Repertoire

Abstract In T cell-dependent antibody responses, some of the activated B cells differentiate along extrafollicular pathways into low-affinity memory and plasma cells, whereas others are involved in subsequent germinal center (GC) formation in follicular pathways, in which somatic hypermutation and affinity maturation occur. The present study demonstrated that Bim, a proapoptotic BH3-only member of the Bcl-2 family, contributes to the establishment of the B-cell repertoire from early to late stages of immune responses to T cell-dependent antigens. Extrafollicular plasma cells grew in the spleen during the early immune response, but their numbers rapidly declined with the appearance of GC-derived progeny in wild-type mice. By contrast, conditional Bim deficiency in B cells resulted in expansion of extrafollicular IgG1+ antibody-forming cells (AFCs) and this expansion was sustained during the late response, which hampered the formation of GC-derived high-affinity plasma cells in the spleen. Approximately 10% of AFCs in mutant mice contained mutated VH genes; thus, Bim deficiency appears not to impede the selection of high-affinity AFC precursor cells. These results suggest that Bim contributes to the replacement of low-affinity antibody by high-affinity antibody as the immune response progresses.

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