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R R Hardy

Publications and source records attributed to R R Hardy.

116 records · Page 7Linked to original sources
PubMed1982

Surface phenotype of Peyer's patch germinal center cells: implications for the role of germinal centers in B cell differentiation.

The surface phenotype of Peyer's patch germinal center lymphoid cells in the mouse is described. It is confirmed that most germinal center lymphocytes bind high levels of peanut agglutination (PNA), a lectin with specificity for terminal galactosyl residues. It is shown that germinal center lymphocytes can be identified in cell suspensions as a discrete PNAhi population distinct from other B cells, plasma cells, and most T cells, which bind only low levels of PNA. Using fluorescence-labeled PNA as a marker in dual fluorescence studies, we found that the majority of Peyer's patch germinal center cells are B lymphocytes: PNAhi Peyer's patch cells express B220, the B lineage-specific form of the T200 family of molecules, as well as low levels of surface Ig. They do not express the T cell-lineage antigens Thy-1, Lyt-1, or Lyt-2 (only 1 to 3% positive). They bear lower levels of H2-K than PNAlo B cells, but two to three times the level of surface I-A-encoded determinants. A discrete but variable subpopulation of PNAhi Peyer's patch cells bear ThB in AKR/c mice, but BALB/c PNAhi lymphocytes are ThB-. About 10 to 30% bear surface IgM or IgG, but in contrast to essentially all PNAlo B lymphocytes in this site, they express no detectable surface IgD. The majority of Peyer's patch germinal center cells bear surface IgA, and this IgA is allelically excluded in F1 mice, indicating it is synthesized by the germinal center cells themselves. In fact, germinal centers contain most of the IgA-bearing cells in Peyer's patches (70 to 85%). These findings lend considerable support to the concept that germinal centers in Peyer's patches are the site of generation of precursors of the IgA-secreting plasma cells that characterize mucosal immune responses, and also suggest that germinal centers may play an important role in the process of heavy chain class switching.

Alleles↗
PubMed1978

Relation between structure and specificity of antibodies: nuclear magnetic resonance study of binding fluorine-19 labeled nitrophenyl haptens to myeloma immunoglobulins M315, M460, and X25.

The relation between structure and specificity of antibodies has been explored by 19F NMR studies of the binding of trifluoromethyl analogues of nitrophenyl haptens to the three mouse myeloma immunoglobulins M315, M460, and X25. We have used haptens with trifluoromethyl groups located at the ortho or para positions of the phenyl ring or attached to the side chain, two atoms removed from the ring (i.e.,-NHCH2CF3). The changes in chemical shift between hapten free in solution and bound to antibody are sensitive to microenvironment and range from 1.7-ppm downfield to 1-ppm upfield. The shifts of p-trifluoromethylnitrophenyl haptens bound to M315 and M460 are both large downfield shifts, which are likely caused by van der Waals interaction and ring-current effects, particularly from tyrosine-34(L); these haptens do not show similar shifts when bound to X25 which has a deletion of tyrosine34(L). Other differences in the binding of the aromatic rings of haptens by M315, M460, and X25 are observed and their origins considered. The importance of hydrogen bonding in the thermodynamic affinity of antibody for hapten has been estimated by comparisons of binding affinities for haptens with trifluoromethyl groups in place of nitro groups.

Amino Acid Sequence↗
PubMedDate not supplied

Demonstration of B-cell maturation in X-linked immunodeficient mice by simultaneous three-colour immunofluorescence.

CBA/N mice carrying the X-linked immune deficiency gene (xid) have fewer splenic B cells than normal CBA mice and are unresponsive to a certain class of antigens. Studies of B-cell surface-marker expression and immune responsiveness have led to the commonly accepted idea that the B cells in adult xid mice are immature and resemble the B cells of young (1-3 week old) normal mice. That is, like young animals, xid mice lack cells in the most numerous of three IgM/IgD B-cell subpopulations (designated I in Fig. 1a, b) present in adult spleen. We now report, however, that this picture is an oversimplification and that in fact the B cells in adult xid mice differ from those present in either adult or young normal mice. Using quantitative three-colour fluorescence-activated cell sorter (FACS) analyses, we have compared the correlated expression of IgM, IgD and a newly discovered B-lymphocyte antigen (BLA-1) on splenic B cells in normal and xid mice. We show here (1) that most B cells in adult xid mice (as in normals) are BLA-1- whereas all B cells in young animals are BLA-1+; (2) that the major difference in the IgM/IgD B-cell subpopulations found between xid and normal mice is limited to the BLA-1- cells; and (3) that xid mice have increased numbers of BLA-1+ population III B cells.

Animals↗
PubMedDate not supplied

Reconstitution of functional receptor for human interleukin-2 in mouse cells.

Interleukin-2 (IL-2) has a key role in the antigen-specific clonal growth of T lymphocytes, by virtue of its interaction with a specific cell-surface receptor (IL-2R). The growth signal seems to be delivered by IL-2 bound to the high-affinity, but not the low-affinity, receptor. Genes encoding IL-2 and its receptor (that is, Tac-antigen) have been cloned and analysed in detail. We have now achieved cell-type-specific reconstitution of the high-affinity human IL-2R by expressing the complementary DNA cloned from normal lymphocytes. A mouse T-lymphocytic line, EL-4, expressed human IL-2R with high (dissociation constant (Kd) = 160-220 pM) and low (Kd = 2.1-2.2 nM) affinity for recombinant human IL-2, while mouse L929 cells expressed only a single class of the IL-2R with lower affinity (Kd = 34.5 nM) for the ligand. We also show that the human IL-2R expressed in EL-4 cells responds to IL-2 and mediates reversed signal transduction: growth of the EL-4 cells harbouring the IL-2R is inhibited specifically by human recombinant IL-2. The approach described here may provide a general experimental framework for elucidating the molecular basis of signal transduction mediated by specific receptor-ligand interaction.

Animals↗
PubMedDate not supplied

Recombination between an expressed immunoglobulin heavy-chain gene and a germline variable gene segment in a Ly 1+ B-cell lymphoma.

The early stages of murine B-cell differentiation are characterized by a series of immunoglobulin gene rearrangements which are required for the assembly of heavy(H) and light(L)-chain variable regions from germline gene segments. Rearrangement at the heavy-chain locus is initiated first and consists of the joining of a diversity (DH) gene segment to a joining (JH) gene segment. This forms a DJH intermediate to which a variable (VH) gene segment is subsequently added. Light-chain gene rearrangement follows and consists of the joining of a VL gene segment to a JL gene segment: once a productive light-chain gene has been formed the cell initiates synthesis of surface immunoglobulin M (sIgM) receptors (reviewed in ref. 1). These receptors are clonally distributed and may undergo further diversification either by somatic mutation or possibly by continued recombinational events. Such recombinational events have been detected in the Ly 1+ B-cell lymphoma NFS-5, which has been shown to rearrange both lambda and H-chain genes subsequent to the formation of sIgM (mu kappa) molecules. Here we have analysed a rearrangement of the productive allele of NFS-5 and found that it is due to a novel recombination event between VH genes which results in the replacement of most or all of the coding sequence of the initial VHQ52 rearrangement by a germline VH7183 gene. Embedded in the VH coding sequence close to the site of the cross-over is the sequence 5' TACTGTG 3', which is identical to the signal heptamer found 5' of many DH gene segments. This embedded heptamer is conserved in over 70% of known VH genes. We suggest that this heptamer mediates VH gene replacement and may play an important part in the development of the antibody repertoire.

Animals↗