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Biomedical subjects

J Ledbetter

Publications and source records attributed to J Ledbetter.

16 recordsLinked to original sources
PubMed1995

Studies on the interdependence of gp39 and B7 expression and function during antigen-specific immune responses.

Interactions between T and B cells are dynamic and regulated by interacting receptor: co-receptors. Interactions between CD40 and its ligand, gp39, and the CD28/CTLA-4 and B7 family members play a decisive role in regulating the progression of cognate interactions. The interdependence of gp39-CD40 and CD28/CTLA-B7 expression and function was studied in vitro during an antigen-induced immune response using T cells from mice expressing a transgenic T cell receptor (TCR). gp39 was induced on pigeon cytochrome c (PCC)-transgenic T cells in the presence of antigen and antigen-presenting cells. The antigen-induced expression of gp39 on transgenic T cells was inhibited by antibodies to class II major histocompatibility complex, CD4 and LFA-1, but not by CTLA-4 Ig, anti-B7-1 or anti-B7-2. These data established that the antigen-induced expression of gp39 was not dependent on co-stimulation via CD28/CTLA-4. The addition of PCC also resulted in the modest expression of B7-1 and a more robust expression of B7-2 on the cognate B cells. The addition of anti-gp39 blocked the up-regulated expression of B7-1 and partially blocked the up-regulated expression of B7-2. The addition of anti-gp39 and anti-interleukin-4 inhibited antigen-induced expression of B7-2 on B cells to near background levels. Studies on the up-regulation of B7-1 and B7-2 on resting B cells showed that soluble gp39 up-regulated B7-1 and B7-2 expression on B cells. In addition, interleukin-4 and interferon-gamma up-regulated B7-2 expression on B cells. Taken together, these data demonstrate that the antigen-induced expression of gp39 is dependent on TCR-derived signals, yet independent of CD28/CTLA-4 co-stimulatory signals. Cognate interactions also resulted in the modest enhancement of B7-1 expression and a more profound expression of B7-2 which were completely or partially dependent on gp39-CD40 interactions.

Abatacept↗
PubMed1994

Antibody to the ligand of CD40, gp39, blocks the occurrence of the acute and chronic forms of graft-vs-host disease.

Chronic and acute graft-versus-host disease (cGVHD and aGVHD) result from donor cells responding to host disparate MHC alleles. In cGVHD (H-2d-->H-2bd), heightened polyclonal immunoglobulin production is due to the interaction of donor allospecific helper T cells (Th) and the host B cells. In vivo administration of antibody to the ligand for CD40, gp39, blocked cGVHD-induced serum anti-DNA autoantibodies, IgE production, spontaneous immunoglobulin production in vitro, and associated splenomegaly. Antibody production remained inhibited for extended periods of time after termination of anti-gp39 administration. Antiallogeneic CTL responses induced in a GVHD were also prevented by the in vivo administration of anti-gp39 as was the associated splenomegaly. These data suggest that CD40-gp39 interactions are critical in GVHD and that CD40-gp39 may be a valuable ligand-receptor pair for targeting immunotherapeutic agents to control GVHD.

Acute Disease↗
PubMed1992

Signal transduction in Sjögren's syndrome T cells. Abnormalities associated with a newly described human A-type retrovirus.

OBJECTIVE: To study the effects of a novel A-type retrovirus, detected in cocultures of lip biopsy specimens from Sjögren's syndrome (SS) patients and a human T cell line, on the infected T cells. METHODS: Interleukin-2 (IL-2) and IL-6 secretion were measured by bioassay and enzyme-linked immunosorbent assay, respectively, in the infected and noninfected cell lines. Surface antigen expression was determined by flow cytometry, using monoclonal antibodies. Protein kinase C (PKC) activity was measured using an enzyme assay kit, and calcium mobilization was assessed with a fluorescent probe. RESULTS: Infected cells expressed less CD4 and IL-6 receptor, but more HLA-DR, compared with noninfected cells. Infected cells also produced less IL-2 and displayed reduced PKC activation and calcium mobilization. A similar defect in calcium mobilization was detected in T cells from SS patients. CONCLUSION: These data suggest a possible involvement of the newly described retrovirus in T cell abnormalities.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1992

Functional roles of the V3 hypervariable region of HIV-1 gp160 in the processing of gp160 and in the formation of syncytia in CD4+ cells.

To study the roles of the V3 hypervariable region (amino acid residues 301-336) of the HIV-1 envelope glycoprotein (gp160) during infection, we constructed recombinant vaccinia viruses that expressed either wild-type gp160 (v-env10) or mutant gp160 in which the V3 region was deleted (v-dl29.1 and v-dl29.2). In v-dl29.1 the V3 loop, formed by disulfide bonding between cysteine residues 301 and 336, was deleted from cys301 to cys336 (inclusive) and replaced by one serine residue. In v-dl29.2 the V3 loop was deleted from arg303 to ala334 and replaced by three residues: gly-ala-gly. Cells infected with all three recombinant vaccinia viruses expressed gp160 on the cell surface, but v-dl29.1-derived gp160 was not cleaved into gp120 and gp41 and did not bind the CD4 glycoprotein. In contrast, gp160 produced by recombinant v-dl29.2 was cleaved normally, and the mutant gp120 produced was secreted and retained binding activity to CD4+ cells. However, both mutants failed to induce syncytia in HeLa CD4+ cells. Thus a disulfide loop at the V3 portion of gp160 is required for cleavage into gp120 and gp41, presumably because the loop is required for proper tertiary structure. The sequence within the V3 loop, however, is not required for cleavage and secretion of gp160, or for binding to CD4+, but this region is essential for gp120-mediated syncytia formation.

CD4 Antigens↗
PubMed1991

Effects of mutations in glycosylation sites and disulphide bonds on processing, CD4-binding and fusion activity of human immunodeficiency virus envelope glycoproteins.

Site-directed mutagenesis was used to study the biological significance of a disulphide bridge and two N-linked oligosaccharides in the CD4-binding region of the envelope glycoproteins of human immunodeficiency virus type 1. Mutagenesis was performed in a phage M13 system at sites corresponding to the cysteine residue (amino acid 402) and the asparagine residues (390 and 447) of the env gene. The mutated env gene was inserted into a recombinant vaccinia virus under the control of the vaccinia virus 7.5K promoter and the expression of mutated env proteins was analysed by SDS-PAGE, a conventional indirect immunofluorescence assay and by a fluorescence-activated cell sorter. Cysteine 402 was found to be essential for the specific cleavage of gp160 into gp120 and gp41, and for intracellular transport of the protein to the cell surface. CD4-binding and syncytium formation assays demonstrated that the disulphide bridge of cysteine 402 stabilized a conformation essential for receptor binding as well as syncytium formation by CD4+ cells. No altered biological activity compared to that of the wild-type proteins could be detected for the mutant proteins lacking the N-glycosylation sites. These data show that the two conserved glycans attached to asparagine residues 390 and 447 do not play any active role in the formation of the disulphide bridge involving cysteine 402 or in the maintenance of an active conformation of the protein, despite their location within the functionally important CD4-binding region.

Amino Acid Sequence↗
PubMed1989

Phosphatidyl inositol hydrolysis after CD3 binding in human peripheral blood T cells: inhibition by prostaglandin E2.

Hydrolysis of phosphatidyl inositol-4,5-bisphosphate, leading to generation of inositol phosphates (IPs), occurs after crosslinking CD3 antigens on the surface of murine and human T-cell lines and clones, and is thought to represent a basic mechanism of signal transduction after antigen receptor binding. Previous investigators have had difficulty demonstrating this phenomenon using human peripheral blood T-cells. In this paper we demonstrate significant IP generation after anti-CD3 stimulation of human peripheral blood lymphocytes and T-cells. The amount of IP generation is less and considerably more variable than what is obtained using a T-cell line. Also, exposure to ammonium chloride in the E-rosetting procedure totally inhibits IP generation, perhaps explaining previous unsuccessful attempts. Finally, prostaglandin E2 and other agents that raise cyclic AMP inhibit IP generation by anti-CD3 antibodies in human T-cells and a T-cell line.

Adult↗
PubMed1988

Comparison of circle absorber and Jackson-Rees systems for paediatric anaesthesia.

To evaluate effects of anaesthesia delivery systems on respiratory work in infants, we measured end-tidal PCO2 (PETCO2), transcutaneous PCO2 (PTcCO2) respiratory rate and arterial blood gases in infants (ages 0.2 to 23.8 months) anaesthetised with either a Jackson-Rees (n = 11) or paediatric circle absorber system (n = 15). Two hours after induction, with assisted ventilation there was no difference in PETCO2, PTcCO2, respiratory rate, pH or PaCO2 in infants anaesthetised with either system. In the laboratory, we measured inspiratory and expiratory resistances to breathing through each system at various tidal volumes and total gas flows into the circuit. These values were compared to resistances created by breathing through an appropriately sized endotracheal tube. Although there was lower resistance to ventilation through Jackson-Rees system, both systems had lower resistances for almost all flows tested than breathing through an endotracheal tube alone (p less than 0.05). The paediatric circle absorber system is an efficient apparatus for anaesthetising spontaneously breathing infants.

Anesthesia, Closed-Circuit↗
PubMed1988

Stimulation of B-chronic lymphocytic leukemia populations by recombinant interleukin-4 and other defined growth-promoting agents.

Monoclonal populations from 10 cases of phenotypically well-characterized B-chronic lymphocytic leukemia (B-CLL) and from a single case of hairy cell leukemia were assessed for their ability to respond by mitogenic stimulation to a number of agents described as growth-promoting for normal B cells. These included the recombinant factors interleukin-1 (IL1), IL2, IL4, IL5, and gamma-interferon, partially purified B cell growth factor (BCGF), B cell stimulatory factor 2 (BSF2), and a CDw40 antibody to the Bp50 antigen. With only few exceptions, no factor or combination of factors stimulated B-CLL populations directly to DNA synthesis. By marked contrast, the hairy cells were responsive to IL4, BCGF, and the CDw40 antibody. B-CLL cells could become responsive with the inclusion of the phorbol ester TPA (12-O-tetradecanoylphorbol-13-acetate) as co-stimulant such that half of the populations were now activated by IL4, particularly when BCGF was also present. Populations refractory to IL4 were, nonetheless, still responsive to BCGF. In only three cases was a significant effect seen with IL2. gamma-interferon could be either inhibitory or stimulatory and, in a few cases, modulated specifically the effects of IL4. In contrast to normal B cell activations, neither the CDw40 antibody nor a calcium ionophore synergized with TPA for stimulating the majority of B-CLL populations. BSF2 was stimulatory in the two cases examined while both IL1 and IL5 were ineffective where studied. No simple correlation was observed between the patterns of responsiveness and the expression of a panel of CD markers assayed on cells both freshly isolated and after TPA stimulation. The data demonstrate a functional heterogeneity not disclosed by simple phenotypic analysis and also indicate the range of activities which can impinge on the growth regulation of monoclonal B cell populations.

Antibodies, Monoclonal↗
PubMed1978

Identification of an FMR cell surface antigen associated with murine leukemia virus-infected cells.

FMR antigens are found on the surface of cells infected with Friend, Moloney, and Rauscher murine leukemia viruses (MuLV). These antigens are serologically distinct from the G cell surface antigens that are found on cells infected with endogenous MuLV (AKR and Gross virus). Cell surface antigens of both virus groups are immunogenic in mice, and immunization with appropriate virus-infected cells leads to the production of cytotoxic antisera. The cytotoxic activity of FMR antisera can be absorbed by disrupted preparations of Rauscher MuLV, but not by AKR MuLV. FMR antisera precipitate the viral envelope proteins gp70, pl5(E), and p12(E) from detergent-disrupted preparations of [3H]leucine-labeled MuLV. The reaction of these antisera with p15(E) and p12(E) proteins is directed against group-specific antigens and can be absorbed with AKR MuLV; in contrast, the reaction of these antisera with gp70 is directed against type-specific antigens and is absorbed only by viruses of the FMR group. In immune precipitation assays with detergent-disrupted 125I surface-labeled cells, FMR antisera react only with type-specific antigens of the viral envelpe protein. On the basis of these findings we conclude that the FMR cell surface antigen is a determinant on the MuLV env gene product.

Animals↗
PubMed1977

Viral proteins expressed on the surface of murine leukemia cells.

Leukemic cells of AKR mice contain as constituents of their membranes the murine leukemia virus envelope protein gp70 and the precursor polyprotein of the viral internal (core) structural proteins. Both gp70 and the core polyprotein are represented on the cell surface as glycoproteins, as evidenced by incorporation of [3H]glucosamine into their structure and the binding of these proteins to lectins. The glycosylated core polyprotein exists in at least two serologically distinguishable forms: the 95,000-dalton polyprotein reacts with antisera prepared against the viral proteins p30, p12, and p10, whereas the 85,000-dalton polyprotein reacts with antisera prepared against the viral proteins p30 and p12, but not p10. Additional heterogeneity in these cell surface polyproteins has been observed wtih leukemias induced by exogenous leukemia viruses. Spontaneous leukemia cells of AKR mice invariably express gp70 and the core polyprotein on their cell surface; normal thymocytes of young AKR mice express gp70, but not the core polyprotein on their surface.

AKR murine leukemia virus↗
PubMed1977

Identification of the Gross cell surface antigen associated with murine leukemia virus-infected cells.

The Gross cell surface antigen (GCSA) is produced by cells that are either exogenously infected with murine leukemia virus (MuLV) or are expressing endogenous MuLV genomes. In immune precipitation assays, GCSA was resolved into two serologically distinct 85,000- and 95,000-dalton viral proteins. These antigenic components are glycosylated forms of the polyprotein precursors of the MuLV internal structural proteins.

AKR murine leukemia virus↗