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F Bettens

Publications and source records attributed to F Bettens.

At least 37 records · Page 2Linked to original sources
PubMed1992

Use of antibody/peptide constructs of direct antigenic peptides to T cells: evidence for T cell processing and presentation.

Human T cells can express MHC-class II products and were shown to be potential antigen-presenting cells. However, they are unable to capture the antigen and only antigens, which bind to T cell membranes such as the gp120 glycoprotein of HIV, are internalized, processed, and presented by T cells. To better understand the role of T cells as antigen-presenting cells, we established a method which overcomes the lack of antigen capture by T cells. Antigen (tetanus toxoid, TT) or an antigenic peptide of TT (residue 830-843, P2) was coupled to antibodies directed to T cell surface molecules such as CD2, CD4, CD8. Antibody/TT and antibody/P2 constructs stimulated P2-specific T cell clones in the absence of accessory cells, if the antibody recognized a T cell surface structure. Compared to the peptide alone, a 100-500 times lower molar concentration of the antibody/peptide construct was required to achieve a similar proliferative response. T cell stimulation via the constructs involved intracellular processing, as nonspecific, glutaraldehyde fixed T cell lines pulsed with the constructs could present the peptide and processing inhibitors like Leupeptin or Chloroquine inhibited the development of a proliferative response to the constructs. Our data underline the ability of T cells to function as antigen-processing and -presenting cells and show that antibody/antigen or antibody/peptide constructs are able to direct a certain antigen or peptide to a T cell. Antibody/peptide constructs may be interesting tools to better understand antigen processing and to study the consequences of antigen presentation by different cells.

Antigen-Antibody Complex↗
PubMed1991

Selective stimulation of CD4+ versus CD8+ T-cell subsets in symptomatic and asymptomatic HIV-1-infected individuals.

To analyze the proliferative capacity of CD4+ or CD8+ T-cell subsets of individuals infected with human immunodeficiency virus type 1 (HIV-1) and to optimize the in vitro conditions for virus replication, CD4+ or CD8+ cells of HIV-1-infected patients were selectively activated inside the whole peripheral blood mononuclear cell (PMNC) population by dual antibody stimulation. To do so PMNC of HIV-1-infected individuals were stimulated with the per se nonmitogenic anti-CD3 antibody fragment BMA030 F(ab)2 crosslinked through goat antimouse antibodies with an anti-CD4 or an anti-CD8 antibody, which lead to selective proliferation of either the CD4+ or the CD8+ T-cell subset. In the presence of monocyte supernatant and recombinant interleukin-2 (rIL2) CD4+ cells of HIV-1 patients responded normally upon such stimulation as their proliferation correlated (r = 0.9) to the percentage CD4+ cells present in the PMNC population. Selective stimulation and proliferation of CD8+ cells could, however, only partially be elicited by dual antibody stimulation, even in the presence of rIL-2 and monocyte supernatant. Their proliferative response did not correspond (r = 0.1) to the percentage CD8+ cells present in the PMNC culture. A positive correlation (r = 0.7) was detected only between percentage CD8+ HLA-DR- cells and proliferation. This confirmed previous studies showing that the defective in vitro proliferative response of peripheral blood lymphocytes of HIV-infected individuals to mitogens, which is usually interpreted being due to a CD4 cell defect, is actually due to a failure of CD8+DR+ cells to proliferate.(ABSTRACT TRUNCATED AT 250 WORDS)

Antilymphocyte Serum↗
PubMed1991

Incorporation of biotinylated nucleotides for the quantification of PCR-amplified HIV-1 DNA by chemiluminescence.

Chemiluminescent detection of polymerase chain reaction (PCR-)amplified DNA was used as a quantitative method for detecting HIV-1. For this purpose, biotinylated dUMP was directly incorporated into the amplified DNA during the PCR reaction. Biotinylation was visualized in an enzymatic reaction using avidine-conjugated alkaline phosphatase and its chemiluminescent 1,2-dioxetane substrate AMPPD, which decomposes upon dephosphorylation and emits light. Light emission was either detected with X-ray films or quantified with a single-photon counting camera connected to a computer imaging system. The specificity of the method was shown by hybridization with a biotinylated or radiolabelled HIV-1-specific oligonucleotide probe. Besides being quantitative, this method represented a non-hazardous, rapid and sensitive technique for the detection of HIV-1 DNA.

Biotin↗
PubMed1990

Dual antibody stimulation: role of monocyte products and activation requirements of T cell subsets.

Using the model of dual antibody stimulation (cross-linking anti-CD3 BMA030-F(ab')2 antibodies with subset specific anti-T cell antibodies, i.e., anti-CD4 or anti-CD8 antibodies) we investigated the role of monocyte products in eliciting IL2 production and asked whether T cell subsets differ in their activation requirements. We found that activation mechanisms of dual antibody stimulation were equal for CD4 and CD8 as well as CD4 CDw29 and CD4 CD45R T cell subsets: dual antibody stimulation was necessary to induce responsiveness to soluble monocyte products which trigger IL2 synthesis. IL2 production induced by soluble monocyte products was measured in CD4 and CD8 T cells as specific IL2 mRNA expression and with a biological assay. Combined actions of IL1 beta and TNF alpha were identified as active monocyte products as (a) treatment of LPS-stimulated monocyte supernatants with anti-IL1 beta and anti-TNF alpha antibodies abrogated the proliferation inducing effect of monocyte supernatants and (b) addition of rIL1 beta and/or rTNF alpha enhanced proliferation of T cells stimulated by dual antibody cross-linking. If both recombinant monokines were added simultaneously a potentiated proliferative effect was observed. Whereas rIL1 beta plus/or rTNF alpha sustained proliferation, only rIL1 beta but not rTNF alpha induced IL2 synthesis in T cell subsets stimulated by dual antibody cross-linking.

Antigens, CD↗
PubMed1990

Dichotomous effect of monocyte Fc receptor interaction on anti-CD3-induced immunoglobulin synthesis.

Monoclonal antibodies against the TCR/CD3 complex are capable of activating T cells which in turn may induce immunoglobulin synthesis in B cells under appropriate conditions. Here we present evidence that distinct immune responses, induced by four commonly used TCR/CD3 mAb (Leu4, OKT3, BMA030, BMA031) were related to the mAb interaction with monocyte Fc receptors for IgG. Depending on their isotype and on the technique by which they were crosslinked, TCR/CD3 mAb induced variable IgM and IgG synthesis in PBMC: If the mAb were crosslinked by monocyte IgG-Fc receptors they induced a high Ig production, while crosslinking the same mAb by plastic-bound goat anti-mouse antibodies (panning) failed to do so. Nevertheless, both crosslinking techniques triggered a strong proliferation and IL-2, IL-4, and IFN gamma lymphokine gene expression. The lack of Ig production under panning conditions was due to an additional IgG-Fc receptor interaction with monocytes: (a) If namely mAb F(ab')2 fragments, or mAb isotypes unable to bind to monocyte Fc receptors (IgG2b, IgG1 in nonresponders) were crosslinked by panning, both a good proliferation as well as Ig production ensued; (b) if TCR/CD3 mAb isotypes which could additionally bind to monocyte Fc receptor (IgG2a) were crosslinked, no Ig production occurred; (c) if mAb F(ab')2 fragments were crosslinked with a second anti-T cell antibody of IgG2a isotype, which could bind to monocyte Fc receptors, Ig synthesis was reduced. Interestingly enough, this diminishing effect, due to monocyte Fc receptor interaction, was only observed if CD4-positive cells were proliferating, but not if CD8-positive cells were activated.

Adult↗
PubMed1989

Lymphokine gene expression related to CD4 T cell subset (CD45R/CDw29) phenotype conversion.

In this study, we investigated whether the phenotype-related differentiation of human 2H4+ (CD45R) naive cells to 2H4- (CDw29) memory CD4 cells corresponded to modulation of interleukin (IL) 2, IL 4, interferon (IFN)-gamma and granulocyte-monocyte colony-stimulating factor (GM-CSF) gene expression, a phenomenon which might correlate to the distinct functional activities of naive cells or memory cells. To mimic in vitro CD4 T cell subset differentiation, freshly isolated 2H4+ and 2H4- CD4 cells were stimulated with the anti-CD3 antibody (Leu-4), expanded in IL 2-containing medium and restimulated with Leu-4 after 7 and 13 days. Absence of monocyte-T cell interaction was compensated by adding monocyte supernatant to the culture medium and by cross-linking the anti-CD3 antibodies with goat anti-mouse antibody coated on culture dishes. It has been previously shown that in vitro stimulated 2H4+ cells acquire CDw29 surface antigens. Measurement of lymphokine gene expression by dot-blot hybridization revealed that although stimulated 2H4+ cells proliferated less than stimulated 2H4- cells, and expressed less actin mRNA, they expressed more IL 2 but less IL 4 and GM-CSF than 2H4- cells. No significant difference was observed between the two subsets for the expression of IFN-gamma. If subsets were restimulated with Leu-4 antibodies, expression of IL 2 was decreased and expression of IL 4 was increased in both subsets; however, the differences among the subsets persisted. They were even more enhanced for IL 2 but less pronounced for GM-CSF. Thus, in spite of phenotype conversion, CD4 T cell subsets maintained a distinct capacity to express IL 2 and IL 4 genes.

Antigens, Differentiation↗
PubMed1987

Activation of T cells by cross-linking an anti-CD3 antibody with a second anti-T cell antibody: mechanism and subset-specific activation.

Binding of anti-CD3 antibody BMA030-F(ab')2 to T cells is not able to induce T cell proliferation even after additional cross-linking by plastic-bound goat anti-mouse Ig antibodies (panning) and addition of either interleukin (IL) 1 or IL2. In search for agents able to complement the signals provided by BMA030-F(ab')2, several antibodies directed against CD2, CD4, CD5, CD6 and CD8 were used. Neither of these caused T cell proliferation either by themselves or when simply added together with BMA030-F(ab')2. However, if BMA030-F(ab')2 and any other of these anti-CD2, CD4, CD5, CD6 or CD8 antibodies were cross-linked, then proliferation of T cells occurred. The mitogenic effect of cross-linking BMA030-F(ab')2 and a second anti-T cell antibody is dependent on the presence of monocytes or exogenously added IL2. The enhancing effect of monocytes could not be replaced by addition of IL1, suggesting that other soluble factors or monocyte contact might be involved in induction of activation signals. The mechanism leading to this mitogenic effect is dependent on combining cross-linking of BMA030 with the second anti-T cell antibody, whereby the second antibody appears to act as an "anchor" for the CD3 antibody, controlling and/or changing the signal transduction via the CD3 structure. This "anchor" hypothesis could be substantiated by the following observations: the best stimulatory signals were obtained at a BMA030/anti-T cell antibody ratio of 1:1; the effect is independent of the antibody isotype and the epitope recognized by the second antibody; the binding of BMA030 and separate cross-linking of the anti-T cell antibody was not sufficient to trigger T cell mitogenesis; immobilization by direct binding of the CD3 antibody alone or of the CD3 and second membrane structure/antibody complex separately was not sufficient to stimulate T cell proliferation. A further interesting aspect of this finding is the fact that a selective T cell stimulation is possible, since cross-linking BMA030-F(ab')2 plus anti-CD4 antibodies induced a selective stimulation of T4 cells, while cross-linking BMA030-F(ab')2 plus anti-CD8 activated solely T8 cells. It is thus possible to selectively activate T cell subsets in vitro.

Antibodies↗
PubMed1987

T cell activation by cross-linking anti-CD3 antibodies with second anti-T cell antibodies: dual antibody cross-linking mimics physical monocyte interaction.

The anti-CD3 antibody BMA030 (IgG2a isotype) induces T cell activation and proliferation if an interaction with monocytes is provided. In contrast to other anti-CD3 antibodies, it is unable to induce interleukin (IL)2 responsiveness through cross-linking by plastic-bound goat anti mouse Ig antibodies (panning). Cross-linking BMA030 with a second anti-T cell antibody is, however, able to induce IL 2 responsiveness in monocyte-depleted T cell cultures. In this report we show that a large number of different antibodies are suitable for this dual antibody stimulation, and that the extent of proliferation corresponds to the percentage of T cells expressing the respective T cell antigen. Proliferation induced by low concentrations (0.1-1 ng/ml) of other anti-CD3 antibodies requires also cross-linking with second anti-T cell antibodies. The proliferative response of monocyte-depleted T cells to two cross-linked anti-T cell antibodies plus added IL 2 is of the same magnitude as the one induced by anti-CD3 antibodies plus monocytes. On the other hand, if monocytes are present, soluble anti-CD2, -CD4, -CD8, -LFA-1 antibodies (IgG1 or F(ab')2 fragments) can inhibit OKT3 or BMA030-induced T cell activation. Anti-CD6 antibodies do not interfere with this monocyte-dependent T cell stimulation. We conclude that dual antibody stimulation mimics the physical contact of T cells with monocyte membranes, where the T cell receptor CD3 complex is cross-linked with neighboring structures (mainly so-called adhesion molecules) through the interaction with respective counter-structures on monocyte membranes. Dual antibody cross-linking bypasses this interaction and can be used to stimulate IL 2 responsiveness of antibody-defined T cells.

Adult↗
PubMed1987

Differences of T-cell activation by the anti-CD3 antibodies Leu4 and BMA030.

Two anti-CD3 antibodies and their Fab/F(ab')2 fragments were compared with regard to their requirement for secondary signals and generations of intracellular messengers. The anti-CD3 antibody BMA030 was found to require monocyte contact to elicit T-cell mitogenesis. Cross-linking by plastic-bound goat anti-mouse antibodies (panning) failed to activate T cells, even in the presence of recombinant IL-1 or IL-2. In contrast, crosslinking of the anti-CD3 antibody Leu4 or Leu4 fragments was mitogenic in monocyte-free cultures. Measurements of intracellular Ca2+ ([Ca2+]i) and generation of inositol phosphates revealed that binding (+/- panning) of BMA030, Leu4, and their F(ab')2 fragments generated similar amounts of intracellular messengers and thus failed to explain the different responsiveness to passive crosslinking. Since the generation of these messengers was not necessarily followed by proliferation but was always observed when mitogenesis occurred, we conclude that the elevation of [Ca2+]i and the production of inositol phosphates are required but not sufficient to trigger mitogenesis.

Adult↗
PubMed1986

1,25-Dihydroxycholecalciferol-induced differentiation of myelomonocytic leukemic cells unresponsive to colony stimulating factors and phorbol esters.

The murine myelomonocytic leukemia cell line WEHI-3B D+, which differentiates in response to granulocyte colony stimulating factor (G-CSF), can also be induced to differentiate into monocyte-macrophages by phorbol myristate acetate (PMA) treatment, whereas the WEHI-3B D- subline, which is unresponsive to G-CSF and PMA, can be induced to differentiate to granulocytes as well as monocytes by 1,25-dihydroxycholecalciferol [1,25-(OH)2 D3], the biologically active metabolite of vitamin D3. A newly developed variant of the WEHI-3B D+ line, named WEHI-3B D+ G, which was responsive to G-CSF but not to PMA, was also differentiated to granulocytes by 1,25-(OH)2 D3. Although vitamin D3 has been reported to induce macrophage differentiation in responsive tumor cells, this is the first demonstration that 1,25-(OH)2 D3 can induce granulocyte differentiation. In both differentiation pathways, cessation of cellular proliferation accompanies changes in morphologic and cytochemical properties of the cells. This suggests that leukemic cell lines unresponsive to differentiation agents acting at the cell surface retain their ability to differentiate in response to agents that do not act via the plasma membrane such as 1,25-(OH)2 D3, which has cytosolic/nuclear receptors. Vitamin D3 could act through different cellular pathways inducing differentiation or by bypassing only the first step of a common differentiation cascade used by agents with cell surface receptors such as CSF. These results suggest that low doses of 1,25-(OH)2 D3 may be useful in combination with hemopoietic growth factors (CSFs) as therapeutic agent to induce leukemic cell differentiation in vivo.

Animals↗
PubMed1985

Effect of cyclosporin A on the early activation of human T helper lymphocytes: inhibition of RNA-synthesis and modification of the expression of activation antigens.

The ability of cyclosporin A (CS-A) to inhibit induced lymphocyte activation and to modify expression of membrane receptors was assessed on human T helper cells. Flow cytometric cell cycle analyses of acridine orange-stained cells showed that CS-A (0.5 micrograms/ml) inhibits the G0-G1 activation process of a substantial proportion of PHA- and Con A-stimulated lymphocytes. The expression of Tac, OKT9 and 4F2 antigens (previously shown to be expressed or increased on activated cells) was investigated by immunofluorescence. Fewer cells expressed the Tac and OKT9 antigens after activation in presence of CS-A, but the percentage of 4F2-positive cells remained unchanged. Analyses of receptor densities measured by fluorescence intensity revealed for all three investigated antigens a decreased receptor density on positive cells in presence of CS-A. Thus, CS-A not only inhibited cell activation (G0-G1 transition) and the expression of Tac, OKT9 and 4F2 antigens, but it also diminished the number of Tac, OKT9 and 4F2 antigens per cell. Assessing specifically the activation of OKT4 (helper) and OKT8 (cytotoxic) cells after 24 h, either by double-fluorescence or by cell fractionation with anti-OKT4 or anti-OKT8 antibodies plus complement, showed that preferentially OKT4 cell activation as well as expression of Tac and OKT9 antigens on those cells was inhibited in the presence of CS-A.

Antibodies, Monoclonal↗
PubMed1985

Effects of poly(I,C)-LC on growth and differentiation of normal and malignant myelopoietic progenitor cells.

Polyriboinosinic-polycytidylic acid with poly-L-lysine stabilized with carboxymethylcellulose [poly(I,C)-LC] augmented, in a dose- and time-dependent manner, secretion of colony-stimulating factor (CSF) by peritoneal macrophages (M phi) and bone marrow cells (BMC). Optimal effects were found after 2 days of in vitro culture of the cells with 50 micrograms/ml of poly(I,C)-LC or 14 h to 3 days after a single intraperitoneal injection of 1-2 mg/kg of poly(I,C)-LC into normal mice. The increase in CSF secretion by M phi and BMC was paralleled in vivo by an increase in serum CSF levels, followed by a rise in committed granulocyte and M phi progenitor cells (GM-CFU-C), nucleated BMC, and blood leukocytes of myelomonocytic origin. Poly(I,C)-LC at doses greater than 4 mg/kg, however, were strongly myelosuppressive. In vitro treatment of undifferentiated myelomonocytic leukemia cells from the WEHI-3B cell line with 10-1,000 micrograms/ml of poly(I,C)-LC resulted in a significant increase in CSF secretion by the leukemic cells and a concomitant inhibition of their proliferation. Incubation of cells from the WEHI-3B D+ subline, which differentiate in response to GM-CSF or G-CSF, with 50-100 micrograms/ml poly(I,C)-LC in agar cultures induced in approximately 45% of the leukemic colonies a differentiation into granulocytes and/or M phi. Poly(I,C)-LC, however, had no effect on differentiation of cells from the CSF unresponsive WEHI-3B D- subline. The CSF-inducing biological response modifier poly(I,C)-LC thus has the potential to stimulate growth and differentiation of normal, as well as differentiation of malignant myelopoietic progenitor cells.

Animals↗
PubMed1984

Interleukin 2-independent stimulation of rabbit chondrocyte collagenase and prostaglandin E2 production by an interleukin 1-like factor.

In the present study, interleukin 1 (IL 1)-containing media from different sources, namely a murine macrophage cell line (P388D1), rabbit peritoneal macrophages, and human peripheral blood mononuclear cells, were compared for their effect on thymocyte proliferation and on collagenase and PGE2 secretion by chondrocytes. A high correlation was found between the enhancement of thymocyte proliferation and the induction of collagenase and PGE2 secretion by chondrocytes. Furthermore, a highly purified IL 1-like factor, namely mononuclear cell factor (MCF) was also active on chondrocytes. The addition of highly purified IL 2 to rabbit chondrocytes had no effect on collagenase and PGE2 secretion induced by IL 1-containing media. Our findings suggest that the factor which induced collagenase and PGE2 secretion by rabbit chondrocytes was an IL 1-like factor. Thus, collagenase secretion by chondrocytes may be used as an IL 2-insensitive assay for the detection of IL 1-like factors.

Animals↗
PubMed1984

Lymphokine regulation of human lymphocyte proliferation: formation of resting G0 cells by removal of interleukin 2 in cultures of proliferating T lymphocytes.

The question of whether lymphocytes which have once been activated and have completed one or several cell cycle(s) can return to the G0 phase and stay ready for a new activation (G0-G1 transition), rather than simply die, was investigated. To do so interleukin 2 (IL-2) was removed from cultures of continuously proliferating human T lymphocytes and the formation of resting (G0) cells was measured. Kinetic analyses in freshly prepared peripheral blood lymphocytes (PBL) revealed that the onset of detectable RNA synthesis and the appearance of structures binding the anti-Tac antibody occurred simultaneously. This allowed the expansion of the definition of G0 T lymphocytes as cells having a low RNA (and DNA) content, and no Tac antigen. When cultured human T cells proliferating continuously by means of IL-2 were characterized in terms of their distribution in the cell cycle, 7 days after the initial PHA stimulation, it could be demonstrated that very few cells were in the G0 phase, supporting the concept of direct S/G2/M-G1 transition. However, when IL-2 was removed from the cultures, the [3H]thymidine incorporation per 10(4) cells and correspondingly the number of cells in the S/G2/M and G1 phases were reduced drastically and during the following 72-hr period, the number of G0 cells increased markedly. Restimulation of such in vitro formed G0 cells, under conditions permitting observation of their shift from the G0 to G0 phase, demonstrated that most cells could respond normally. Based on these observations, it was concluded that IL-2 not only ensures T-lymphocyte survival and proliferation, but IL-2 starvation induces many continuously proliferating T lymphocytes to stop cycling and to return to the G0 phase of the cell cycle where they remain functional.

Antibodies, Monoclonal↗