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W Knapp

Publications and source records attributed to W Knapp.

At least 109 records · Page 6Linked to original sources
PubMed1992

Phenotype of human T cells expressing CD31, a molecule of the immunoglobulin supergene family.

The CD31 molecule is a leucocyte-surface glycoprotein of 130 kDa with homology to the immunoglobulin gene superfamily. In this study we report on the expression of CD31 on human T cells and demonstrate that it subdivides peripheral blood T lymphocytes into two novel subsets. CD31 is expressed by 38% of CD3+ lymphocytes. About 85% of CD31+ T cells display the CD45RA phenotype, 35% the CD45RO phenotype, 24% the CD4 phenotype and 72% the CD8 phenotype. There is also a correlation between CD31 expression and CD45RA expression in cord blood T cells; 89% of CD3+ cord blood cells express CD31, and most of them have the CD45RA phenotype. A discrepancy was found with thymocytes, which are positive for CD31 but negative for CD45RA. Stimulation of human T cells leads to down-regulation of CD45RA, while CD31 continues to be expressed. In functional studies, CD31 antibody binding to T lymphocytes does not lead to mobilization of intracellular calcium, proliferation or modulation of T-cell proliferation.

Antibodies, Monoclonal↗
PubMed1991

Synthesis and functional characterization of a recombinant monoclonal antibody directed against the alpha-chain of the human interleukin-2 receptor.

We have determined the sequence of the light and heavy chains of mAb 3G-10 (IgG1), a monoclonal antibody competing with interleukin 2 (IL2) for binding to the human IL2 receptor Tac protein. The antibody-encoding genes were chimerized by introducing splice donor and part of the intron sequences into the cDNA and subsequently linking it to the constant parts of the human IgG1 gene. The chimeric mAb was produced in mouse myeloma cells and purified. Murine and chimeric mAbs showed similar properties with respect to inhibition of T-cell proliferation. In contrast to its murine counterpart, the chimeric mAb exhibited Ab-dependent cellular cytotoxicity and, when combined with an Ab recognizing a different epitope on the IL2 receptor Tac protein, was able to activate human complement. The chimerized mAb might therefore have improved therapeutic efficacy.

Amino Acid Sequence↗
PubMed1991

GPI-anchored cell-surface molecules complexed to protein tyrosine kinases.

Binding of ligand or antibody to certain cell-surface proteins that are anchored to the membrane by glycophosphatidylinositol (GPI) can cause activation of leukocytes. However, it is not known how these molecules, which lack intracellular domains, can transduce signals. The GPI-linked human molecules CD59, CD55, CD48, CD24, and CD14 as well as the mouse molecules Thy-1 and Ly-6 were found to associate with protein tyrosine kinases, key regulators of cell activation and signal transduction. A protein tyrosine kinase associated with the GPI-linked proteins CD59, CD55, and CD48 in human T cells, and with Thy-1 in mouse T cells was identified as p56lck, a protein tyrosine kinase related to Src. This interaction of GPI-linked molecules with protein tyrosine kinases suggests a potential mechanism of signal transduction in cells.

Animals↗
PubMed1991

Analysis of function-associated receptor molecules on peripheral blood and synovial fluid granulocytes from patients with rheumatoid and reactive arthritis.

In this study we report the expression pattern of 13 different function-associated surface molecules on synovial fluid and peripheral blood granulocytes from rheumatoid and reactive arthritis patients. We found increased expression of the complement receptors 1 (CD35) and 3 (CD11b) and of the activation-associated antigens CD67, CD24, and M5 on synovial fluid granulocytes from rheumatoid and/or reactive arthritis patients compared to autologous peripheral blood granulocytes. In addition, synovial fluid granulocytes expressed IgG Fc receptor 1 (CD64) and complement receptor 4 (CD11c), neither of which can be found on peripheral blood granulocytes. Peripheral blood granulocytes from rheumatoid and reactive arthritis patients expressed higher levels of leucocyte function-associated antigen 1 (CD11a) and of the membrane proteins CD31, CD24, M5, and M6 compared to peripheral blood granulocytes from healthy controls and patients with degenerative joint disease. No significant differences in the expression of any of the molecules studied could be observed between cells from rheumatoid and cells from reactive arthritis patients, suggesting a similar activation process for granulocytes in these two diseases.

Antigens, CD↗
PubMed1991

Ratio of complement receptor over Fc-receptor III expression: a sensitive parameter to monitor granulocyte-macrophage colony-stimulating factor effects on neutrophils.

In vitro activation of human granulocytes leads to altered expression of distinct surface antigens. Compared with the changes observed with classic activating reagents such as the phorbol ester PMA similar, but less pronounced alterations of surface antigen expression were observed upon granulocyte activation with human recombinant granulocyte-macrophage colony-stimulating factor (hrGM-CSF). In particular, stimulation with hrGM-CSF is followed by an enhanced expression of the complement receptors CD35 (CR1) and CD11b (CR3) while the low affinity Fc-gamma receptor CD16 (FcRIII) is downregulated. In order to investigate whether there are similar effects under in vivo conditions, we studied the granulocytes from patients undergoing rhGM-CSF therapy before, during, and after treatment. We found a marked increase in CD35 (CR1) and CD11b (CR3) expression and a substantial decrease or even loss of CD16 (FcRIII) on these granulocytes. These changes correlated well with our in vitro data and occurred extremely rapidly after therapy onset. Furthermore, therapy monitoring using ratios calculated by the mean fluorescence channel numbers of CR and FcRIII stainings may combine the advantage of high sensitivity with high reproducibility as a result of the contrasting change in CR and FcRIII expression during granulocyte activation. Being nonparametric values, such ratios are not influenced by individual flow cytometry standardization. Taken together, these activation-associated changes of surface receptor expression and especially of CR over FcRIII ratios are useful parameters for monitoring the in vivo effects of rhGM-CSF.

Antigens, CD↗
PubMed1991

In vitro and in vivo activated T cells display increased sensitivity to PGE2.

In this study we report that in vitro activation of T cells increased the cyclic AMP response to subsequent prostaglandin E2 (PGE2) stimulation severalfold per cell. This sensitization of T cells to PGE2-induced cyclic AMP generation was observed when the T cells had been stimulated in vitro for 5 days with either the CD3 monoclonal antibody OKT3, phytohemagglutinin, or the combination phytohemagglutinin plus the phorbol ester PMA. Enhanced cyclic AMP generation following mitogenic activation was seen in response to both PGE2 and forskolin, direct activator of the adenylate cyclase, indicating that the amount of adenylate cyclase had increased during the in vitro activation course. In order to investigate whether various T cell subsets in general and in vivo activated T cells in particular would differ in their susceptibility to PGE2, we isolated CD4+, CD8+, CD4-CD8-, CD4+CD45RO+ ("memory"), and CD4+CD45RA+ ("virgin") T cells and studied PGE2-mediated inhibition of CD3-induced proliferation, as well as cyclic AMP generation in response to PGE2, respectively. We found that CD8+ T cells are more susceptible to PGE2 inhibition and produce more cyclic AMP than CD4+ T cells. Double-negative T cells (enriched for gamma delta T cell receptor positive cells) were found to be sensitive to PGE2 as well. Within the CD4+ T cell population, CD45RO+ ("memory") T cells were significantly more sensitive to PGE2-mediated suppression than CD45RA+ ("virgin") T cells. CD45RO+ cells required a 10-fold lower dose of PGE2 for half-maximum suppression of proliferation. However, no difference in cyclic AMP production could be demonstrated between these two subsets. We propose that substantial heterogeneity exists among peripheral blood T lymphocyte subsets regarding their sensitivity to the immunosuppressive action of PGE2 and that the sensitivity of individual cells changes in the course of an immune response.

Antibodies, Monoclonal↗
PubMed1991

Monoclonal antibodies VIB-E3, IB5 and HB9 to the leucocyte/epithelial antigen CD24 resemble BA-1 in recognizing sialic acid-dependent epitope(s). Evidence that VIB-E3 recognizes NeuAc alpha 2-6GalNAc and NeuAc alpha 2-6Gal sequences.

The specificities of seven monoclonal antibodies to the human B cell differentiation marker CD24 have been investigated with respect to sialic acid containing carbohydrates. These are antibodies HB8, HB9, VIB-C5, VIB-E3, AL1a, LC66 and IB5, which are known to bind to polydisperse sialoglycoprotein(s) on Nalm-6 B lymphoblastoid cells. Three of the antibodies, HB9, VIB-E3 and IB5, have been found to resemble the first described antibody in this series, BA-1, in binding also to bovine submaxillary mucin. As with BA-1, the binding of the antibodies is abolished or reduced markedly after desialylation of the epithelial glycoprotein, and the binding to neuraminidase-treated Nalm-6 cells is also reduced. There is evidence for the involvement of of non-O-acetylated sialic acid in the determinants recognized by these antibodies, since there is a substantially enhanced binding following mild-alkali treatment of the epithelial mucin which removes O-acetyl groups. One of the antibodies, VIB-E3, is deduced to recognize the oligosaccharide sequences NeuAc alpha 2-6GalNAc and NeuAc alpha 2-6Gal as part of larger antigenic structures. This conclusion has been reached from the results of inhibition-of-binding experiments using a series of structurally defined sialo-oligosaccharides and direct binding experiments using oligosaccharides chemically linked to lipid (neoglycolipids).

Animals↗
PubMed1991

[Autologous bone marrow transplantation in children with acute leukemia or non-Hodgkin's lymphoma].

More than 95% of children with acute lymphoblastic leukemia (ALL) achieve complete remission with conventional chemotherapy and 60 to 70% are cured. In contrast, only 70 to 80% of children with acute myeloid leukemia (AML) achieve remission and 30 to 40% are cured. In acute leukemia, ALL and AML, high initial white blood cell counts, poor initial response to therapy, certain structural chromosomal abnormalities, and diagnosis under 1 year of age indicate an increased risk of relapse. Prognosis is poor in children, who relapse during treatment. If no bone marrow donor is available, high dose chemo(radio)therapy and autologous stem cell rescue can be applied in these children to consolidate second remission. Furthermore, autologous bone marrow transplantation may be used to consolidate first remission of individual children, who suffer from acute leukemia with high risk criteria. 19 children with a median age of 8 years (0.5 to 19 years) underwent high dose chemo(radio)therapy and autologous bone marrow and/or blood derived stem cell rescue. 11 suffered from ALL, 4 had AML, and 4 suffered from Non Hodgkin's lymphoma (NHL). 7/13 children (54%), who were in first or second remission after early relapse, are alive in continued remission for median 24 months (9 to 44) after autografting. Children in more advanced stage of disease had no benefit from high dose therapy and autologous stem cell rescue.

Adolescent↗
PubMed1991

[Phenotypical and functional characterization of leukocytes--the CD-system].

In several workshops held over the past few years 89 different cluster molecules have been identified on the surface of human leukocytes by the use of monoclonal antibodies. In this review we describe the principal of the CD clustering system and present a short survey of its application in clinic and basic research along with a complete listing of the CD molecules.

Antibodies, Monoclonal↗
PubMed1990

A single laser flow cytometry method to evaluate the binding of three antibodies.

Here we describe a flow cytometry method which permits the collection of data regarding three staining parameters of a single cell from a suitable combination of dual parameter stainings using PE- and FITC-labelled mAbs and a single argon laser. The technique neither requires a third dye with non-overlapping emission spectra nor a second light source. In order to test the accuracy of this method, we compared data calculated using this triple parameter analysis with data obtained by double staining a presorted population homogeneously positive for one parameter. In experiments using either resting or in vitro activated T cells, the percentages obtained with both methods were identical (P greater than 0.05). Using this method we tested which T cell sub-subpopulation is responsible for the weak CD25 (IL-2R alpha) expression constantly associated with freshly isolated human T cells and concluded that it is predominantly expressed on the CD4+ CD45RO+ (CD4 positive memory) T cell subpopulation.

Adult↗
PubMed1990

Molecular characterization and functional analysis of the leukocyte surface protein CD31.

The CD31 Ag is a surface glycoprotein of 130 kDa with a broad cellular distribution. We show that among peripheral human blood cells, it is expressed on monocytes, granulocytes, platelets, and a subpopulation of lymphocytes. Activation of granulocytes leads to down-regulation of CD31 molecule expression. Sequence analysis and quantitative measurements of the relatedness of the CD31 molecule to other known proteins demonstrate that it consists of six Ig constant domains and that each domain bears substantial similarity to Fc gamma R domains. We find, however, that the CD31 molecule does not bind Ig Fc domains. On human monocytes we demonstrate that CD31 mAb recognizing certain epitopes of the CD31 molecule induce the generation of reactive oxygen metabolites. No such effect was seen with human granulocytes. By using two CD31 mAb, termed 1B5 and 7E4, we analyzed the requirements for activation of the monocyte respiratory burst via CD31 Ag in more detail. We show that signal transduction occurs via formation of a CD31 Ag-mAb-Fc gamma R complex involving either Fc gamma RI (CD64) or Fc gamma RII (CDw32) molecules.

Amino Acid Sequence↗
PubMed1990

Activated human T lymphocytes express MHC class I heavy chains not associated with beta 2-microglobulin.

We present here the molecular characterization of a new activation-induced surface structure on human T lymphocytes, termed LA45, with high homology (93% at protein level) to MHC class I molecules. Antigen modulation and sequential immunoprecipitation experiments revealed that LA45 and HLA class I proteins do not crossreact with the corresponding antibodies. Furthermore, LA45 is not associated with beta 2-m. On the other hand, we could show that the separation of HLA-A,B,C and beta 2m molecules, induced by SDS-denaturation, leads to a conformational change in the heavy chain in such a way that it becomes reactive with LA45. The 90/45 kD LA45 proteins thus appear to be non-beta 2m-associated MHC class I alpha chains that are selectively expressed by activated but not by resting human T lymphocytes.

Amino Acid Sequence↗
PubMed1990

Human blood basophils synthesize interleukin-2 binding sites.

Recent data suggest that basophils express receptors for a variety of lymphokines. In this study we present the biochemical characterization of the interleukin-2 (IL-2) receptor on the basophil surface membrane. Highly enriched populations (purity: 92% to 99%) of blood basophils were obtained from chronic granulocytic leukemia (CGL) patients (n = 3) by negative selection using monoclonal antibodies (MoAbs) and complement. CGL basophils were found to bind CD25 MoAbs (n = 4) directed against different epitopes of the 55- to 60-Kd subunit of the IL-2 receptor (= Tac peptide). Immunoprecipitation experiments with lysates of purified CGL basophils and CD25 MoAbs showed a protein with a molecular weight of 60 Kd, equivalent to the Tac peptide on human T blasts. Quantitative binding studies and Scatchard plot analysis using radiolabeled recombinant human (rh) IL-2 indicated the presence of 12,000 +/- 4,700 low affinity IL-2 binding sites (kd = 66 nmol/L) per purified CGL basophil. Northern blot analysis with enriched CGL basophils showed two messenger RNA bands of 3.5 and 1.5 kilobases hybridizing to radiolabeled Tac cDNA. Immunoprecipitation of the Tac peptide from enriched basophils metabolically labeled with 35S-methionine showed active synthesis of the IL-2 receptor. Our results show that human blood basophils synthesize and express receptors for IL-2.

Antibodies, Monoclonal↗
PubMed1990

Signal transduction in lymphocytic and myeloid cells via CD24, a new member of phosphoinositol-anchored membrane molecules.

The CD24 Ag is present on human B cells from the earliest stages of B lineage development and is lost on terminal B cell maturation to plasma cells. This Ag is also expressed on mature forms of granulocytes. We studied the effects of CD24 mAb on the function of both lymphocytic and myeloid cell types. We found a clear-cut increase in free cytoplasmic calcium when tonsil B cells or mononuclear cells from B cell chronic lymphatic leukemia patients were preincubated with CD24 mAb and further stimulated with goat F(ab')2 anti-mouse Ig antibodies. The same experimental setting with granulocytes instead of B lymphocytes led to the triggering of hydrogen peroxide production in these cells. CD24 Ag is expressed at higher levels on activated granulocytes and is anchored to the plasma membrane by a phosphoinositol linkage. In conclusion we provide evidence that the CD24 Ag are functional molecules in cells of different lineage, playing a signal transducing role in cell function.

Antibodies, Monoclonal↗
PubMed1990

CD27 expression by a distinct subpopulation of human B lymphocytes.

CD27 is present on the surface of a major subset of peripheral blood T lymphocytes. In this report we show that CD27 is also expressed on a subpopulation of the normal human B cell lineage which is absent from cord blood but present in tonsils and in the peripheral blood of adult individuals. CD27+ B lymphocytes are characterized by the following criteria: (a) in terms of physical properties, the CD27+ B cells form a population with an increased cell size combined with a decreased cell density; (b) the CD27 expression of tonsillar B lymphocytes is postively correlated with mIgA but negatively correlated with membrane IgM/membrane IgD positivity; (c) CD27 on B cells can be induced selectively by the combination of Staphylococcus aureus plus interleukin 2, but not by either treatment alone, and (d) CD27+ B lymphocytes express high levels of the adhesion structures LFA-1 (CD11a), ICAM-1 (CD54), LFA-3 (CD58) and of the lymphocyte homing receptor CD44. These latter findings suggest that CD27+ B cells are predispose to form cell-cell interactions. Accordingly, within 3 h of cell culture CD27+, but not CD27-, B lymphocytes were found to form LFA-1-mediated homotypic B cell clusters.

Antibodies, Monoclonal↗
PubMed1990

M5, a phosphoinositol-linked human myelomonocytic activation-associated antigen.

When investigating the previously described monoclonal antibody (MoAb) VIM-5, raised against THP1 cells and binding to human monocytes and granulocytes, we found that the antigen detected by this antibody, designated M5, becomes very strongly expressed on monocytes after overnight culture with phorbol myristate acetate (PMA) or lipopolysaccharide (LPS) but not with recombinant human interferon-gamma (rhIFN-gamma). Granulocytes stimulated with formyl-methionyl-leucyl-phenylalanine (FMLP) become negative for binding VIM-5. Immature granulocytes from bone marrow do not express M5, thus its expression on granulocytes is differentiation linked. The antigen bound by VIM-5 is sensitive to hydrolysis by phosphoinositol-specific phospholipase C (PI-PLC). The immunoprecipitated M5 antigen on monocytes is a broad band, with a peak of 50 kD (unreduced) and two bands of 53 kD and 44 kD (reduced). We have therefore detected an antigen that is upregulated on stimulated monocytes but, conversely, down-regulated on FMLP-stimulated granulocytes.

Antigens, Differentiation, Myelomonocytic↗
PubMed1990

The stimulation of T cells with anti-CD2 monoclonal antibodies facilitates the induction of polyclonal B-cell responses but does not enhance the activation of antigen-specific B cells.

In this report we compare the effect of stimulation of peripheral mononuclear cells (PBMC) by using two monoclonal antibodies (MoAb) directed against the CD2 receptor on T cells or by using autologous erythrocytes (E) which express on their surface lymphocyte function-associated antigen 3 (LFA3), a natural ligand for CD2. The addition of autologous erythrocytes to pokeweed mitogen (PWM)-stimulated PBMC results in the enhancement of polyclonal immunoglobulin synthesis and of antigen-specific B-cell responses. Because B cells lack the CD2 molecule, it can be concluded that their enhanced activity is a consequence of the delivery of activating signals by activated T lymphocytes. When PBMC cultures were stimulated with a pair of anti-CD2 MoAb (Leu5b and VIT13) we were able to induce polyclonal immunoglobulin synthesis, particularly IgM, in cultures supplemented with interleukin 2(IL-2). Specific responses to tetanus toxoid (TT) and keyhole limpet haemocyanin (KLH) were also enhanced by the addition of autologous E to PWM-stimulated PBMC. Significant anti-TT responses were observed in cultures stimulated with E + TT + IL-2. In contrast, stimulation of PBMC with VIT13 + Leu5b + IL-2 + antigen was not effective in inducing anti-TT antibody and only weakly effective in inducing anti-KLH antibodies. Replacing Leu5b by anti-CD3 had no effect on the induction of specific antibody responses; in contrast, replacement of Leu5b by E enhanced anti-TT antibody production while the effect on polyclonal production of IgM was minimal. Therefore, it appears that the signal delivered by the association of CD2 with LFA3 is a better potentiating signal for specific B-cell responses than the signal delivered by pairs of MoAb to different epitopes of CD2 or to CD2 and CD3 epitopes.

Antibodies, Monoclonal↗