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Biomedical subjects

G Kelsoe

Publications and source records attributed to G Kelsoe.

At least 19 recordsLinked to original sources
PubMed1992

In situ studies of the primary immune response to (4-hydroxy-3-nitrophenyl)acetyl. II. A common clonal origin for periarteriolar lymphoid sheath-associated foci and germinal centers.

In the genetically restricted response that follows immunization with (4-hydroxy-3-nitrophenyl)acetyl coupled to protein carriers, two distinct populations of B cells are observed in the spleens of C57BL/6 mice. By 48 h postimmunization, foci of antigen-binding B cells appear along the periphery of the periarteriolar lymphoid sheaths. These foci expand to contain large numbers of antibody-forming cells that neither bind the lectin, peanut agglutinin, nor mutate the rearranged immunoglobulin variable region loci. Germinal centers containing peanut agglutinin-positive B cells can be observed by 96-120 h after immunization. Although specific for the immunizing hapten, these B cells do not produce substantial amounts of antibody, but are the population that undergoes somatic hypermutation and affinity-driven selection. Both focus and germinal center populations are pauciclonal, founded, on average, by three or fewer B lymphocytes. Despite the highly specialized roles of the focus (early antibody production) and germinal center (higher affinity memory cells) B cell populations, analysis of VH to D to JH joins in neighboring foci and germinal centers demonstrate that these B cell populations have a common clonal origin.

Animals↗
PubMed1992

Aging and humoral immunity.

If human antibody responses undergo molecular shifts similar to those identified in mice, the appropriate immunization strategy for the elderly would be a passive administration of the protective antibody from young donors rather than an attempt to boost the individual's own response with a more potent vaccine, because the shifted immune system can no longer make the right kind of antibody.

Aging↗
PubMed1991

Intraclonal generation of antibody mutants in germinal centres.

The generation and selection of somatic antibody mutants are key elements of acquired immunity, essential for the affinity maturation of antibody responses dependent on T cells. The mutants are generated through a mechanism that introduces point mutations at high rate into rearranged variable (V) region genes in the course of cell proliferation. Their appearance coincides with the generation of germinal centres, which are characterized by oligoclonal B-cell proliferation and have been suggested to be the microenvironment in which antibody mutants are generated. We report here direct evidence for this hypothesis. Rearranged V-region genes were amplified from the genomic DNA of cells picked from individual germinal centres. The sequence analysis of these genes revealed that most represent cells of distinct B-cell clones which expanded locally, generating somatic antibody mutants at high rate. By contrast, antigen-induced proliferation of B cells at another site, periarteriolar lymphocyte sheath-associated foci, was not associated with somatic hypermutation.

Amino Acid Sequence↗
PubMed1991

In situ studies of the primary immune response to (4-hydroxy-3-nitrophenyl)acetyl. I. The architecture and dynamics of responding cell populations.

After primary immunization with an immunogenic conjugate of (4-hydroxy-3-nitrophenyl)acetyl, two anatomically and phenotypically distinct populations of antibody-forming cells arise in the spleen. As early as 2 d after immunization, foci of antigen-binding B cells are observed along the periphery of the periarteriolar lymphoid sheaths. These foci expand, occupying as much as 1% of the splenic volume by day 8 of the response. Later, foci grow smaller and are virtually absent from the spleen by day 14. A second responding population, germinal center B cells, appear on day 8-10 and persist at least until day 16 post-immunization. Individual foci and germinal centers represent discrete pauciclonal populations that apparently undergo somatic evolution in the course of the primary response. We suggest that foci may represent regions of predominantly interclonal competition for antigen among unmutated B cells, while germinal centers are sites of intraclonal clonal competition between mutated sister lymphocytes.

Animals↗
PubMed1991

Cloning of murine splenic T lymphocytes and natural killer (NK) cells on filter paper discs: detection of a novel NK/T phenotype.

Discrete colonies of splenocytes were grown on filter paper discs in the presence of concanavalin A and interleukin 2. Phenotypic analysis of the colonies indicated that the majority expressed the Thy-1.2 marker and 72% of these co-expressed the CD3 molecule. Of the colonies 20%-25% were NK 1.1+ and they developed regardless of the presence of Con A in the culture medium, a property of the NK lineage. In addition, Thy-1.2+ colonies developed when splenocytes from scid mice, which lack mature T and B cells, were grown both in the presence and absence of concanavalin A. These results demonstrate that colonies of murine splenic T lymphocytes and NK cells could be successfully grown on filter paper discs and phenotypically characterized. With this colonies technique, it was possible to identify a novel subset of NK 1.1+ colonies that co-expresses CD3 and shares growth properties with T cell colonies.

Animals↗
PubMed1991

Contribution of the VH11 gene family to mitogen-responsive B cell repertoire in C57BL/6 mice.

The contribution of VH11 gene family to the development of the primary B cell repertoire has been studied by analyzing 1.8 x 10(4) mitogen induced B lymphocyte colonies. The data demonstrate that VH11 family is predominantly expressed among neonatal splenic as well as adult peritoneal B cell colonies, both rich in Ly-1+ B cells. VH11 gene family expression among B splenocytes decreases during ontogeny and VH11 family pairs stochastically with different V kappa families among mitogen-activated neonatal B cell colonies, which are representative of an antigen unselected B cell repertoire. Thus, an increased VH11 expression among peritoneal and neonatal B cells points towards its biased expression among Ly-1+ B lymphocytes. The restricted V gene rearrangements and VH11-V kappa 9 pairing observed among anti-bromelain-treated mouse red blood cells autoantibodies are likely to be an outcome of both intrinsic gene recombination processes per se as well as selection by an autoantigen and/or local selective environmental factors.

Age Factors↗
PubMed1990

Antigen-binding repertoire and Ig H chain gene usage among B cell hybridomas from normal and autoimmune mice.

LPS-stimulated B cells were used to generate a panel of mAb that were a random sample of the preimmune repertoire of C57BL/6 and highly autoimmune, viable motheaten mice. These mAb were tested for reactivity to a number of "self" and foreign Ag. Binding that could be detected only at nM mAb concentrations or less was considered significant. We found that a surprisingly high number of the mAb bound one or more of the Ag tested, and many mAb bound more than a single Ag. Ag-induced mAb were likewise tested and found to have greatly reduced cross-reactivities. We found no significant differences, either in frequency of Ag binding or degree of cross-reactivity, between normal and autoimmune mice. Furthermore, the frequency with which a given Ag was bound by our panel of mAb was found to be proportional to the size of the Ag. The frequency with which individual VH gene families were expressed by our panel was consistent with a stochastic usage of VH genes in the preimmune repertoire. We interpret these data as showing that the preimmune repertoire is highly cross-reactive and that the activation of autoreactive clones in autoimmune animals is due to a defect in cellular regulation rather than a difference in repertoire.

Animals↗
PubMed1990

Stochastic pairing of heavy-chain and kappa light-chain variable gene families occurs in polyclonally activated B cells.

Frequencies of 25 immunoglobulin heavy-chain and kappa light-chain variable (VH + V kappa) gene-family pairings expressed in splenic B-cell populations were determined by hybridization of VH- and V kappa-family-specific DNA probes to mitogen-induced B-cell colonies from C57BL/6 mice or hybridomas derived from BALB/c and NZB mice. Both analyses support the conclusion that VH and V kappa gene families pair without bias; as would be expected for random association, the frequencies of specific VH + V kappa pairs may be estimated by the product of the independent VH and V kappa frequencies. Based upon the frequencies at which 9 VH and 12 V kappa gene families are expressed, we calculated the expected usage for approximately 100 VH + V kappa family pairings in neonatal and adult C57BL/6 mice. Variability in the expression of such VH + V kappa pairings is considerable; pairs representing greater than 10% to less than 0.01% of the splenic B-cell population occur. This variability is most pronounced in the neonate, where 6 VH + V kappa family pairs account for nearly 40% of all mitogen-reactive B cells. As the neonate matures, the distribution of frequencies for VH + V kappa pairings becomes more nearly uniform. This process may underlie the patterned acquisition of humoral immune responsiveness.

Animals↗
PubMed1989

Murine V kappa gene expression does not follow the VH paradigm.

V kappa gene family expression among LPS-reactive murine B lymphocytes, unlike that of VH gene families, is not proportional to genomic complexity, i.e., nonstoichiometric. Furthermore, no positional bias for the overexpression of J-proximal V kappa genes (V kappa 21) is observed among neonatal B lymphocytes. Yet, the V kappa 1 and V kappa 9 families located in the center of V kappa locus are preferentially used by neonatal B splenocytes. Thus, the mechanisms of V kappa gene rearrangement and expression appear to differ significantly from those controlling the VH locus.

Animals↗
PubMed1989

Mapping of antibody specificities to VH gene families.

VH gene segments represent the products of the repeated duplication and subsequent diversification of a primordial V gene element. It is widely assumed that natural selection, operating via pathogens, has played the dominant role in this process. Here, we screen some 3.7 x 10(4) C mu+ colonies of mitogen-activated B cells for the production of antibodies specific for phosphorylcholine or hen egg lysozyme and expression of the VH X-24, S107, Q52, or J558 gene families. These gene families were expressed at frequencies proportional to their genomic complexity among both unselected and antigen-specific C mu+ colonies. Thus, the capacity to encode equivalent antibody-combining sites is dispersed uniformly among VH families. This result suggests that individual VH genes have not evolved to address specific antigens.

Animals↗
PubMed1987

Genotypic analysis of B cell colonies by in situ hybridization. Stoichiometric expression of three VH families in adult C57BL/6 and BALB/c mice.

The filter paper disc method for cloning inducible lymphocytes was used to census the splenic B cell population of C57BL/6 and BALB/c mice for the expression of three VH gene-families, VH X-24, -Q52, and -J558. B cell colonies, arising from single founder lymphocytes, were identified by in situ hybridization with VH family- and C mu-specific cDNA probes. Some 6.7 X 10(4) C mu+ colonies were screened. Among C57BL/6- or BALB/c-derived colonies, approximately 3% were VH X-24+, approximately 19% were VH Q52+, and approximately 54% were VH J558+. These frequencies are consistent with a process of equiprobable expression for individual VH segments, and provide direct evidence that normal splenic B lymphocytes use a process of random genetic combinatorics to generate the antibody repertoire.

Animals↗
PubMed1986

Cloning of mitogen- and antigen-reactive B lymphocytes on filter paper discs. II. Paratope frequencies within the mitogen-selected repertoire.

Paratopic frequencies of C57BL/6 (Igh-Vb) and BALB/c (Igh-Va) mice were compared by determining the frequency of lipopolysaccharide-reactive, splenic B lymphocytes secreting antibody specific for (4-hydroxy-5-iodo-3-nitrophenyl) acetyl (NIP), trinitrophenyl (TNP), phosphorylcholine (PC), NIP/TNP, NIP/PC, and sheep erythrocytes. Despite the known genotypic and phenotypic differences between the two Igh-V loci, no significant differences in paratope frequencies were demonstrated. Similar determinations in C.B-20 mice, Ighb congenics of the BALB/c strain, and in C57BL/10 nude mice indicated that the mitogen-generated paratope frequencies directly reflected the capacity of immunoglobulin variable region elements rather than complex interactive or regulatory controls to generate diversity. We conclude that at least for the paratopic repertoire, the role of the somatic processes for the generation of antibody diversity exceeds the influence of germ-line differences between the Ighb and Igha haplotypes.

Analysis of Variance↗
PubMed1986

Regulation of the immune response. II. Concomitant idiotope-specific enhancement and suppression can result in a phenotypically normal response.

Idiotope-specific immunoenhancement or suppression was induced in C57BL/6 mice by the injection of physiological amounts (100 ng-10 micrograms) of monoclonal anti-idiotope antibody. As previously described, nanogram doses enhanced idiotope expression while a 10-micrograms dose of anti-idiotope antibody induced the activation of a population of Thy 1.2+, Lyt 1-, 2+ suppressors. Both positive and negative regulatory activities were confined to the non-mu, idiotope+ compartment of the plaque-forming cell response. Administration of intermediate doses of anti-idiotope antibody resulted in an immune state indistinguishable from that of naive mice. This apparently normal response was in fact the product of a simultaneous activation of balanced enhancing and suppressive activities. When treated with anti-Lyt 2 or Lyt 1 and complement, spleen cell populations taken from such phenotypically "naive" mice revealed latent idiotope-specific immunoenhancement or suppression, demonstrating the components of a functional regulatory equilibrium.

Animals↗
PubMed1985

Cloning of mitogen- and antigen-reactive B lymphocytes on filter paper discs. I. A description of the technique and of methods for the analysis of colonies.

A novel technique for establishing short term clones of antigen- or mitogen-activated splenic B lymphocytes is described. Spleen cells are plated onto the surface of filter paper discs and subsequently stimulated by antigen or mitogen in situ; activated B cells proliferate and differentiate into pure colonies of cells analogous to bacterial colonies growing on agar. These colonies of lymphocytes may be characterized in a series of replica hemolytic-plaque, autoradiographic, or immunoenzyme assays making possible a full characterization of the frequency of secreted idiotopes and paratopes and of the cells that produce them. Colony induction by either antigen or mitogen occurs under identical conditions, thus a rigorous comparison between the mitogen-selected and antigen-selected antibody repertoires may be made.

Animals↗