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L C Burkly

Publications and source records attributed to L C Burkly.

At least 55 records · Page 3Linked to original sources
PubMed1992

Regions of the CD4 molecule not involved in virus binding or syncytia formation are required for HIV-1 infection of lymphocytes.

Cell surface-expressed CD4 binds to the envelope glycoprotein of HIV-1 and mediates syncytia formation through interacting with membrane expressed HIV-1 gp120. Further possible roles of the CD4 molecule in the process of cell infection by HIV-1 remain poorly understood. In our study we describe two mAb that recognize the V3/V4 domain of the CD4 molecule. Although these mAb do not inhibit gp120-CD4 binding or HIV-1-induced syncytia formation, they inhibit HIV-1 infection of human PBL. These findings suggest that discrete, definable domains of the CD4 molecule may be involved in interactions after HIV-1 envelope binding that lead to virus entry into the cell.

Antibodies, Monoclonal↗
PubMed1992

A monoclonal antibody to CD4 domain 2 blocks soluble CD4-induced conformational changes in the envelope glycoproteins of human immunodeficiency virus type 1 (HIV-1) and HIV-1 infection of CD4+ cells.

The murine monoclonal antibody (MAb) 5A8, which is reactive with domain 2 of CD4, blocks human immunodeficiency virus type 1 (HIV-1) infection and syncytium formation of CD4+ cells (L. C. Burkly, D. Olson, R. Shapiro, G. Winkler, J. J. Rosa, D. W. Thomas, C. Williams, and P. Chisholm, J. Immunol., in press). Here we show that, in contrast to the CD4 domain 1 MAb 6H10, 5A8 and its Fab fragment do not block soluble CD4 (sCD4) binding to virions, whereas they do inhibit sCD4-induced exposure of cryptic epitopes on gp41 and dissociation of gp120 from virions. Two other MAbs, OKT4 and L120, which are reactive with domains 3 and 4 of CD4, have little or no effect on HIV-1 infection, syncytium formation, or sCD4-induced conformational changes in the envelope glycoproteins. The mechanisms of action of 5A8 and 6H10 can be further distinguished in syncytium inhibition assays: 6H10 blocks competitively, while 5A8 does not. We opine that 5A8 blocks HIV-1 infection and fusion by interfering with conformational changes in gp120/gp41 and/or CD4 that are necessary for virus-cell fusion.

Animals↗
PubMed1991

Signaling by vascular cell adhesion molecule-1 (VCAM-1) through VLA-4 promotes CD3-dependent T cell proliferation.

Vascular cell adhesion molecule, VCAM-1, is an adhesion molecule expressed on activated endothelium thought to play a role in leukocyte migration to sites of inflammation. VCAM-1 adheres to leukocytes through the VLA-4 integrin. Recombinant soluble VCAM-1 (rsVCAM) and anti-CD3 mAb OKT3 were utilized to address the role of the VCAM-1/VLA-4 pathway in antigen-dependent T cell activation. Monocyte-depleted T cells proliferated upon exposure to co-immobilized OKT3 and rsVCAM but to neither alone. In contrast, an anti-VLA-4 mAb HP1/2 failed to co-activate with OKT3, despite the fact that both rsVCAM and HP1/2 support T cell adhesion comparably. These data indicate that adhesive function is not sufficient for co-stimulatory activity. They also reveal that VCAM-1 may play a role in regulating T cell immune responses as well as migration in vivo.

Antigens, CD↗
PubMed1990

Tolerance in transgenic mice expressing major histocompatibility molecules extrathymically on pancreatic cells.

Transgenic mice with defined expression of major histocompatibility complex (MHC) proteins provide novel systems for understanding the fundamental question of T cell tolerance to nonlymphoid self components. The MHC class II I-E and I-A and class I H-2K molecules expressed specifically on pancreatic islet or acinar cells serve as model self antigens. In these systems, transgenic proteins are not detected in the thymus or other lymphoid tissues. Yet mice are tolerant to the pancreatic MHC products in vivo; this tolerance is not induced by clonal deletion. These studies have been aided by monoclonal antibodies specific for I-E-reactive T cells and indicate that clonal anergy may be an important mechanism of tolerance to peripheral proteins.

Animals↗
PubMed1989

T-cell tolerance by clonal anergy in transgenic mice with nonlymphoid expression of MHC class II I-E.

T-cell reactivity to the class II major histocompatibility complex I-E antigen is associated with T-cell antigen receptors containing the V beta gene segments V beta 17a and V beta 5. Mice expressing I-E with the normal tissue distribution (on B cells, macrophages, dendritic cells and thymic epithelium) induce tolerance to self I-E by clonal deletion in the thymus. By contrast, we find that transgenic INS-I-E mice that express I-E on pancreatic beta-cells, but not in the thymus or peripheral lymphoid organs, are tolerant to I-E but have not deleted V beta 5- and V beta 17a-bearing T cells. Moreover, whereas T-cell populations from nontransgenic mice proliferate in response to receptor crosslinking with V beta 5- and V beta 17a-specific antibodies, T cells from INS-I-E mice do not. Thus, our experiments provide direct evidence that T-cell tolerance by clonal paralysis does occur during normal T-cell development in vivo.

Animals↗
PubMed1989

Tolerance in transgenic mice expressing class II major histocompatibility complex on pancreatic acinar cells.

To study the nature of tolerance to antigens not expressed by cells of the lymphoid system, expression of class II MHC I-E was targeted to the acinar cells of the exocrine pancreas in transgenic mice (elastase [EL]-I-E). Despite the absence of detectable I-E in the thymus of EL-I-E transgenic mice, both thymocytes and peripheral T lymphocytes were tolerant to I-E, and the pancreas was free of autoimmune infiltrates. Nontolerant T cells adoptively transferred into irradiated or T-depleted transgenic mice rapidly destroy the I-E+ components of the pancreas; however, adoptive transfer of nontolerant T lymphocytes into nonirradiated transgenic mice do not. These results suggest that tolerance in transgenic mice is maintained by some peripheral tolerance mechanism. However, further studies indicate that tolerance in transgenic mice is not maintained by specific Ts cells. For example, cell mixing experiments both in vitro and in vivo fail to reveal dominant unresponsiveness. Furthermore, nontolerant T cells injected into otherwise unmanipulated EL-I-E mice can be primed in situ (by injections of I-E+ spleen cells) to destroy the I-E+ acinar cells.

Animals↗
PubMed1989

Selective expression of class II E alpha d gene in transgenic mice.

Class II genes of the MHC must be expressed by APC for activation of CD4+ T cells and efficient delivery of T cell help to B lymphocytes. Class II genes have restricted tissue expression and are under complex regulation. By using various deletion constructs of the class II E alpha d gene in transgenic mice we have mapped different 5' flanking regions which control E alpha d gene expression in distinct cell types. We demonstrate dissociate expression of E alpha d within the macrophage lineage as well as within the B cell lineage, and present evidence for a repressive element operative in B cells and macrophages. We describe the generation of novel transgenic lines with limited constitutive and inducible E alpha mRNA and I-E protein.

Animals↗
PubMed1989

NF-kappa B binds within a region required for B-cell-specific expression of major histocompatibility complex class II gene E alpha d.

A region upstream of the murine major histocompatibility complex gene, E alpha d, has been shown previously to be required for B-cell expression. Binding of the B-cell-specific factor, NF-kappa B, to a site within this region is indistinguishable from that observed with the kappa enhancer binding site. NF-kappa B may be responsible for E alpha d B-cell expression.

Animals↗
PubMed1989

Tolerance to class II MHC in transgenic mice.

Transgenic mice offer new possibilities in experimental techniques for understanding the familiar questions of T cell development and the selection of the T cell repertoire. As a powerful method to manipulate gene expression in the whole animal, precisely defined in vivo models can be developed. In our own studies, we have used transgenic mice with targeted expression of I-E and new monoclonal antibodies defining T cell receptors specific for class II I-E molecules. In the thymus, our results suggest that thymic epithelium has significant tolerance inducing capability, but the mechanism may be different from the clonal deletion induced by bone marrow derived cells. In the periphery, our results suggest that tolerance to tissue restricted antigens is not induced by clonal deletion. Instead, clonal paralysis may be an important mechanism for both inducing and maintaining peripheral tolerance.

Animals↗
PubMed1988

Diabetes and tolerance in transgenic mice expressing class II MHC molecules in pancreatic beta cells.

Insulin-dependent diabetes is caused by the loss of insulin-producing beta cells in pancreatic islets. It has been proposed that aberrant expression of Class II Major Histocompatibility Complex (MHC) molecules on beta cells stimulates an autoimmune attack against beta cell antigens. To test this hypothesis, we generated transgenic mice that express Class II MHC molecules (E alpha d/E beta b, or I-Eb) on beta cells. Diabetes was found in 100% of transgenic progeny from three expressing transgenic mouse lines, but without evidence for lymphocytic infiltrates. Furthermore, T lymphocytes appeared to be tolerant to the transgene I-Eb molecule, despite the absence of expression of I-Eb in the thymus or any other lymphoid tissue. The results suggest that novel expression of Class II MHC molecules on nonlymphoid cells is by itself insufficient to initiate autoimmune responses against tissue-specific antigens.

Animals↗
PubMed1987

Transgenic mice selectively lacking MHC class II (I-E) antigen expression on B cells: an in vivo approach to investigate Ia gene function.

The E alpha MHC class II gene with 1.4 kb of 5'-flanking and 0.5 kb of 3'-flanking sequences was introduced into (H-2b X s)F2 mice, which do not express their endogenous E alpha gene. The transgene was expressed in thymic tissue and in adherent spleen cells and was induced in peritoneal exudate cells by gamma-interferon. In contrast to the normal E alpha gene, there was no expression in B lymphocytes. Since transgenic animals made with constructs containing 3.2 kb and 2 kb of 5'-flanking sequences show normal expression pattern of the E alpha gene, it appears that deletion of 5'-flanking sequences between -1.4 kb and -2 kb inactivated or eliminated regulatory sequences required for expression of E alpha specifically in B cells. The presence of pBR327 DNA linked to the -1.4 kb E alpha transgene suppresses expression in peripheral adherent cells, yielding mice expressing E alpha only in the thymus. These mice appear to be tolerant to I-E, as measured in mixed leukocyte response experiments.

Animals↗
PubMed1986

Molecular biology of the H-2 histocompatibility complex.

The H-2 histocompatibility complex of the mouse is a multigene family, some members of which are essential for the immune response to foreign antigens. The structure and organization of these genes have been established by molecular cloning, and their regulation and function is being defined by expression of the cloned genes.

Animals↗
PubMed1985

T cell regulation of light chain expression: preferential enhancement of Ig kappa in a primary thymus-dependent response does not require affinity-based selection.

Previously we reported that the proportion of Ig kappa to Ig lambda anti-DNP antibodies produced by day 5 of a primary response to thymus-dependent antigens was fivefold greater than the Ig kappa:Ig lambda ratios found in responses to thymus-independent antigens. To determine whether an affinity-based process explained the differences in kappa/lambda ratios seen, we measured the affinity of Ig kappa and Ig lambda antibody by using the binding ratio radioimmunoassay method, which measures the affinity of antibodies of particular isotypes without the need for prior purification. By the addition of a mild reduction step, the affinities of IgG and IgM antibody could be simultaneously measured. The affinity of day 5 Ig kappa anti-DNP evoked by DNP-KLH is not significantly higher than that of Ig lambda anti-DNP evoked by DNP-Ficoll, nor is it significantly different from that of Ig lambda antibody produced. As an alternative to affinity-based selection, it is suggested that T cells can preferentially augment Ig kappa responses.

Animals↗
PubMed1985

T cell regulation of light chain expression: preferential enhancement of Ig kappa production by T cells in the response to DNP.

We have reported that the relative amounts of Ig kappa and Ig lambda anti-DNP antibodies produced depend on the molecular form in which the hapten was presented. Here we confirm this finding by demonstrating that 1) the effects of adjuvant are not sufficient to result in differential light chain production and 2) the presence of some DNP-containing epitopes unique to DNP-KLH are not sufficient to give rise to enhancement of Ig kappa production. We further demonstrate that preferential augmentation of Ig kappa in the response to DNP is a T cell-mediated effect. Although we cannot rule out the possibility that some Ig kappa antibodies specific for protein-conjugated DNP require T cells for their expression, one of several other possible mechanisms whereby T cells influence light chain production is considered likely.

Adjuvants, Immunologic↗
PubMed1982

Influence of the nude and X-linked immune deficiency genes on expression of kappa and lambda light chains.

The relative amounts of Ig kappa and Ig lambda1 anti-2,4-dinitrophenyl antibodies were measured at various times after immunizing mice with prototype thymus-dependent (TD), thymus-independent type 1 (TI-1) and thymus-independent type 2 (TI-2) antigens. Similar amounts of Ig lambda1 were produced after TD and TI-2 immunization and somewhat less was produced after a TI-1 stimulus. In contrast, Ig kappa levels were much greater after TD than after TI-1 or TI-2 antigen. The amount of light chain isotype produced appeared to depend on the molecular form in which the hapten was presented, although possible adjuvant effects were not ruled out. Levels of Ig kappa and Ig lambda present in nonimmune sera were measured in normal, xid and nude mice. The kappa/lambda ratio was higher in xid than in normal mice and the difference was demonstrated by F1 analysis to be due to an X-linked gene. Conversely, the kappa/lambda ratio was lower in nude than in normal mice. This was true for the CBA/Tufts (Ighj), CBA.Ighb and C57BL/10 strains. However, there were no detectable differences in the relative frequencies of surface Ig kappa- and Ig lambda-bearing B cells in adult CBA/Tufts, CBA/N and nude mice. Hence, serum ratios may reflect differences at the level of B cell triggering. Two possible explanations for these differences are discussed. Ig kappa and Ig lambda may be expressed on functionally distinct B cell subsets. (For instance Ig lambda-producing cells might be readily triggered by T1 antigens whereas Ig kappa-producing cells are more dependent on T cell signals. Such functional subsets could be determined by light chain expression). Alternatively, cells producing Ig kappa antibody are selected for because they have a higher affinity for antigen. If so, triggering of cells producing high affinity Ig kappa or their subsequent selection is T cell-dependent.

Animals↗