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Biomedical subjects

J Banchereau

Publications and source records attributed to J Banchereau.

At least 289 records · Page 16Linked to original sources
PubMed1988

Identification of cell subpopulations by dual-color surface immunofluorescence using biotinylated and unlabeled monoclonal antibodies.

A new method of dual-color immunofluorescence is presented for analysis of surface antigen distribution among heterogeneous cell suspensions. It involves flow cytometric analysis of cells stained with a biotinylated first monoclonal antibody and/or with an unlabeled second monoclonal antibody. After addition of streptavidin-phycoerythrin and/or fluoresceinated goat antimouse immunoglobulin antibody, single-cell fluorescence intensities are measured and biparametric graphic representations are obtained, allowing one to determine the percentage of cells stained by each of the monoclonal antibodies or both. The validity of the method was assessed on human peripheral blood mononuclear cells by using three sets of two monoclonal antibodies: CD8 and CD5, CD3 and CD4, CD11 and HLA-DR. The results showed that dual staining did not induce significant quenching or competition between pairs of antibodies. The procedure is simple and sensitive. It requires only minute amounts of monoclonal antibodies. It is readily applicable to the screening of hybridoma supernatants and to the characterization of new antibodies to cell surface antigens with respect to well-defined markers.

Antibodies, Monoclonal↗
PubMed1988

Human recombinant interleukin 4 induces normal B cells to produce soluble CD23/IgE-binding factor analogous to that spontaneously released by lymphoblastoid B cell lines.

Surface-labeled Epstein-Barr virus (EBV)-transformed lymphoblastoid RPMI 8866 cells release in their supernatant a radiolabeled 25-kDa polypeptide which reacts with the Fc epsilon RL/CD23-specific monoclonal antibody (mAb) 25 and which binds to IgE but not IgG (IgE BF/sCD23). IgE BF/sCD23 had an isoelectric point of 4.5-5.0. The reactivity of mAb 25 with IgE BF/sCD23 allowed us to set up a radioimmunoassay for detection of IgE BF/sCD23 in cell culture supernatants. Supernatants from Fc epsilon RL/CD23+ cell lines were found to contain IgE BF/sCD23. Addition of human recombinant interleukin 4 (IL 4) to normal human B cells cultures induced the production of IgE BF/sCD23. Activation of B cells with anti-IgM antibody coupled to beads enhanced the IL 4-induced production of IgE BF/sCD23 when compared to nonactivated B cells. This correlates with the finding that anti-IgM antibody-activated B cells cultured with IL 4 express more Fc epsilon RL/CD23 than B cells cultured with IL 4 alone. The biochemical characteristics of radiolabeled IgE BF/sCD23 immunoprecipitated by mAb 25 from the supernatants of normal B cells cultured with IL 4 were identical to those of the IgE BF/sCD23 isolated from EBV-transformed cell line supernatants. Addition of interferon-gamma to B cells cultured with IL 4 strongly decreased the level of IgE BF/sCD23 in culture supernatants correlating with the observed decrease of Fc epsilon RL/CD23 on B cell surface. These data demonstrate that normal human B cells cultured in the presence of IL 4 produce an IgE-binding factor (sCD23) biochemically and antigenically equivalent to that spontaneously produced by EBV-transformed lymphoblastoid cell lines.

B-Lymphocytes↗
PubMed1988

Response of LFA-1-deficient B cells to interleukin 4 (BSF-1) and low molecular weight B cell growth factor (BCGFlow).

T cell-depleted B cells from a patient with LFA-1 deficiency were tested in costimulation assays for responsiveness to recombinant human IL4 (BSF-1) and purified low molecular weight B cell growth factor (BCGFlow). In both cases the response of LFA-1-deficient B cells was comparable with normal controls. Monoclonal antibodies to LFA-1 alpha (CD11a) and beta (CD18) chains were unable to mimic the action of IL4 on normal B cells in costimulation assays with anti-IgM, and did not inhibit normal B cell proliferation in response to IL4 and anti-IgM. Epstein-Barr virus-transformed lymphoblastoid B cell lines (LCL) from normal and LFA-1-deficient donors both responded in proliferation assays to BCGFlow but not IL4. Similarly, both normal and LFA-1-deficient LCL increased IgM secretion in response to BCDF, BCGFlow and, interestingly, IL4. The normal LCL also increased IgG secretion in response to these factors, but no IgG was detected in supernatants from the LFA-1-deficient LCL. These results show that LFA-1 expression is not essential for B cell responses to B cell growth and differentiation factors.

Antibodies, Monoclonal↗
PubMed1988

Induction of interleukin-4-dependent IgE synthesis and interleukin-5-dependent eosinophil differentiation by supernatants of a human helper T-cell clone.

The human alloreactive T-cell clone A1 produces interleukin (IL)-4, IL-5, and granulocyte macrophage-colony stimulating factor (GM-CSF), but not IL-2 or interferon-gamma (IFN-gamma), as assessed by probing for mRNA transcripts, immunoassays, or bioassays. Supernatants from clone A1 induced IgE synthesis by normal human peripheral blood mononuclear cells. IL-4 was shown to be necessary and sufficient for the induction of IgE by clone A1 supernatants. In contrast, IgE induction by clone A1 supernatants and recombinant (r) IL-4 was inhibited by IFN-gamma. This suggests that the absence of IFN-gamma from the IL-4-containing A1 supernatants was important for their IgE-inducing ability. Supernatants from clone A1 could also specifically induce the growth of cord blood cell progenitors and their differentiation into eosinophils but not into basophils. rIL-5, but not rIL-4 or rGM-CSF, also induced eosinophil maturation. These data suggest that IL-5 secreted by clone A1 was responsible for its ability to induce eosinophil differentiation. The implications of the concomitant production of IL-4 and IL-5 by a single T-cell clone are discussed.

Cell Differentiation↗
PubMed1988

IgE production by normal human lymphocytes is induced by interleukin 4 and suppressed by interferons gamma and alpha and prostaglandin E2.

The effect of human recombinant interleukin 4 (IL-4) on antibody production by normal peripheral blood mononuclear cells enriched for B cells was investigated. IL-4 preferentially induced IgE synthesis in vitro. In addition, a low induction of IgG production was observed, whereas IL-4 had no effect on IgA and IgM synthesis. The IL-4-induced IgE production by B cells required T cells and monocytes but was specifically inhibited by an anti-IL-4 antiserum indicating that, although IL-4 acts indirectly, it is responsible for the induction of IgE synthesis. IL-4-induced IgE production was blocked in a dose-dependent way by interferon gamma (IFN-gamma), interferon alpha (IFN-alpha), and prostaglandin E2. IFN-gamma also inhibited IL-4-induced IgG production. These inhibitory effects of IFN-gamma and IFN-alpha on IgE production cannot be attributed to toxic effects since IFN-alpha induced IgM production in the presence of IL-4, whereas IFN-gamma was ineffective in inhibiting IgG production induced by IL-2. IFN-gamma, IFN-alpha, and prostaglandin E2 also inhibited IL-4-induced expression of the low-affinity receptor for the Fc portion of IgE (CD23) on B cells, indicating that there is an association between CD23 expression and IL-4-induced IgE production. This theory was supported by the finding that IL-4-induced IgE production was inhibited by F(ab')2 fragments of an anti-CD23 monoclonal antibody.

Antibodies, Monoclonal↗
PubMed1988

The molecular cloning of interleukins 4, 5 and 6: multifunctional hemopoietic growth factors.

We summarize here our recent studies on the cloning and characterization of three lymphokines which are produced by activated T cells. Interleukins 4, 5 and 6 are involved in the regulation of B cell activation, proliferation and differentiation. IL-4 can activate resting B cells, while IL-5 stimulates the proliferation of activated B cells. Both of these factors also have a role in regulating the isotype of immunoglobulin produced by cultures of D cells. IL-6 appears to induce the differentiation of B cells to secrete high levels of immunoglobulin. In addition, each of these factors is involved in the regulation of other lineages of hemopoietic cells. Thus, T cells control multiple lineages of myeloid and lymphoid cells through the diverse actions of multiple lymphokines. IL-6 is exceptional because it is produced by a variety of cell types, and its action is not restricted to hemopoietic cells.

B-Lymphocytes↗
PubMed1987

High affinity binding of human interleukin 4 to cell lines.

Purified human recombinant interleukin 4 (IL-4) was radio iodinated to high specific radioactivity without loss of biological activity. 125I-IL-4 bound specifically to the Burkitt lymphoma Jijoye cells and other cell lines. Jijoye cells showed a high affinity for 125I-IL-4 (Kd approximately equal to 7 10(-11) M) and displayed 1200-1400 specific receptors per cell at 4 degrees C or 37 degrees C. The equilibrium dissociation constant (Kd) corresponds to the IL-4 concentration which induces 50% maximal expression of the low affinity IgE receptor (Fc epsilon RL/CD23) on Jijoye cells. At 4 degrees C the rate constant of association K1 is 1.7 x 10(6) M-1 s-1 and the rate contant of dissociation k -1 is 1.3 x 10(-4) s-1 (t 1/2 = 91 min.) No human recombinant lymphokines other than IL-4 were able to compete for the binding of 125I-IL-4 to its receptor.

Binding, Competitive↗
PubMed1987

Recombinant interleukin 4 promotes the growth of human T cells.

Recently, we reported the isolation of a cDNA clone that encodes a polypeptide which has B cell and T cell growth factor activities. The amino acid sequence of this polypeptide deduced from the nucleotide sequence of the cDNA clone showed significant homology with mouse B cell stimulating factor-1. Because of its multiple biologic activities, it was designated interleukin 4 (IL-4). Here we describe the effects of supernatants of Cos-7 mouse cells transfected with the IL-4 coding cDNA clone in a mammalian expression vector, on human thymocyte T cells and T cell clones. The T cell growth-promoting effect of IL-4 on preactivated T cells was not inhibited by monoclonal antibodies against IL-2 or the IL-2 receptor, indicating that the IL-4 activity is independent from IL-2 or the IL-2 receptor. IL-4 induces a low proliferative response in thymocytes and peripheral blood lymphocytes, but the response was considerably enhanced by preactivation of the thymocytes or peripheral blood T cells. Both T4+ and T8+ antigen-specific proliferative and cytotoxic T cell clones and T3 natural killer clones proliferated in response to IL-4. But one of six T4+ and one of four T8+ T cell clones were consistently found to be unresponsive. The proliferative responses to IL-4 were always lower than those obtained with IL-2. Most of the T cell clones generally became unresponsive to IL-4 10 days after stimulation, but still responded well to IL-2. These results indicate that the responsiveness to IL-4 is relatively short lasting and is regulated by activation signals. Interestingly, IL-4 acted in synergy with IL-2 in promoting the growth of T cell clones. Our results establish that IL-4 can act as a T cell growth factor independently of IL-2.

Animals↗
PubMed1987

Production and characterization of a monoclonal antibody specific for the human lymphocyte low affinity receptor for IgE: CD 23 is a low affinity receptor for IgE.

In the present study a gamma 1 kappa monoclonal antibody, Mab 25, specific for the receptor for the Fc fragment of IgE on lymphocytes (Fc epsilon RL) was established. This antibody was generated after fusion of spleen cells from mice immunized with the EBV-transformed lymphoblastoid cell line RPMI 8866, which is known to express Fc epsilon RL at high density. Mab 25 inhibits strongly the binding of IgE to RPMI 8866 cells and to other Fc epsilon RL-positive EBV-transformed lymphoblastoid cell lines. A 50% inhibition of IgE binding was observed at a Mab 25 concentration of 10 ng/ml. The binding of IgE was also inhibited by Fab fragments of Mab 25, suggesting that the inhibition is not simply due to steric hindrance or to an eventual binding through its Fc portion. Mab 25 only binds to cell lines expressing Fc epsilon RL. Mab 25 immunoprecipitated a single polypeptide with an apparent m.w. of 42 Kd, pI 4.9. The membrane molecule bound to and eluted from a Mab 25 immunoabsorbent had the same apparent m.w. and pI as the Fc epsilon RL purified from an IgE immunoabsorbent. Additionally, when RPMI 8866 cell lysates were cleared with Mab 25, no Fc epsilon RL could be bound to or eluted from an IgE immunoabsorbent. Mab 25 was found to weakly bind to a minor proportion of blood (1 to 4%), tonsil (2 to 9%) and spleen (4 to 5%) mononuclear cells with a low intensity. By double fluorescence analysis, most of the Fc epsilon RL-positive cells were found to be CD 20-positive B lymphocytes. The staining pattern of Mab 25 and the biochemical characteristics of the antigen detected by Mab 25 were comparable to those of the CD 23 Mab. The four CD 23 Mab MHM 6, PL 13, HD 50, and Tü 1 were found to inhibit the binding of IgE. PL 13 was found to totally inhibit the binding of Mab 25 to RPMI 8866 cells, whereas Tü 1 and MHM 6 only partially inhibited Mab 25 binding. HD 50 was unable to block the binding of Mab 25. The finding that different CD 23/Fc epsilon RL-specific monoclonal antibodies recognizing distinct epitopes have in common the capacity of inhibiting the binding of IgE suggests that upon binding they induce a conformational alteration of the Fc epsilon RL resulting in a loss of the IgE binding capacity. In conclusion, our data demonstrate that the CD 23 antigen is a low affinity receptor for IgE on lymphocytes.

Antibodies, Monoclonal↗
PubMed1987

The response of selected human B cell lines to B cell growth and differentiation factors.

Fifteen human B cell lines were tested for their ability to respond to B cell growth and differentiation factors present in phytohemagglutinin-conditioned medium. Five lines responded significantly: CESS showed an increase in IgG production only, HFB1 and BALM1 showed an increase in proliferation only and L4 and BALM4 showed an increase in both IgG production and proliferation. When four of the responding lines (CESS, HFB1, L4 and BALM4) were cultured with human recombinant-derived interleukin 1, interleukin 2, interleukin 4 or interferon-gamma no significant response was seen. CESS, L4 and BALM4 all increased IgG production in response to partially purified B cell growth factor (Cellular Products, Inc., Sera-Lab., Crawley Down, GB) and B cell differentiation factor-containing supernatant from the T24 bladder carcinoma cell line. HFB1, L4 and BALM4 all showed increased tritiated thymidine incorporation in response to purified B cell growth factor but not in response to B cell differentiation factor-containing supernatant. These lines may prove useful in the study of B cell growth and differentiation factors and their receptors.

B-Lymphocytes↗
PubMed1987

Supernatant from an activated human CD4+ T-cell clone modulates the proliferation and collagen synthesis of human dental pulp fibroblasts.

Several studies indicate a relationship between immunocompetent cells and fibroblast functions such as proliferation and collagen synthesis, which may be of importance in understanding the process of fibrosis. We demonstrate here the activity of supernatant from an activated human CD4+ proliferative T-cell clone (2F1) in stimulating the proliferation of human dental pulp fibroblasts, while inhibiting their soluble type I collagen secretion in either growing or confluent cultures. Taken together, our results indicate that T-cell derived lymphokines may be of importance in regulating normal dental pulp fibroblast functions.

Cell Division↗
PubMed1987

Epstein-Barr virus (EBV) induces expression of B-cell activation markers on in vitro infection of EBV-negative B-lymphoma cells.

A set of B-cell activation markers, including the EBV/C3d receptor [complement receptor type 2 (CR2) (CD21)], the 45-kDa lymphoblastoid cell-associated (Blast-2) antigen (CD23), and the B-cell restricted activation (Bac-1) antigen (which was recently identified as a potential B-cell growth factor receptor) can be turned on by infecting lymphoma cells that are genome negative for Epstein-Barr virus (EBV) with the B95-8 immortalizing strain of the virus. The nonimmortalizing EBV variant, strain P3HR-1, which possesses a deletion within the BamHI WYH region of the genome containing the coding sequence for the EBV-determined nuclear antigen 2, does not induce expression of these markers. Other lymphoblastoid cell-associated antigen markers can be activated by infection with either immortalizing or nonimmortalizing viruses. These results suggest that the immortalizing potential of EBV is correlated with its ability to induce expression of B-cell activation markers, which are suspected to play a major role in the physiological pathway leading to lymphoid cell proliferation. The viral genomic region deleted in the nonimmortalizing strain of EBV seems to be required for activation of some of these markers. Human lymphoma cell lines, such as those used in this study, can thus help identify the specific EBV genes involved in lymphoid B-cell proliferation and the mechanism of action of these genes.

Antigens, Neoplasm↗
PubMed1986

Human interferon-gamma acts as a B cell growth factor in the anti-IgM antibody co-stimulatory assay but has no direct B cell differentiation activity.

In this study it is illustrated that recombinant human interferon-gamma (IFN-gamma) acts as a B cell growth factor (BCGF) in the anti-IgM antibody co-stimulatory assay. A monoclonal antibody that specifically inhibits the biological activities of IFN-gamma blocks its BCGF activity supporting the specificity of the IFN-gamma effect. Various IFN-gamma obtained from different sources displayed the same BCGF activity. Nonactivated B lymphocytes do not proliferate in response to IFN-gamma. IFN-gamma acts directly on B cells because highly purified B cells obtained after standard purification procedures coupled to cell sorting could still proliferate in response to IFN-gamma. Blood B lymphocytes were found to be more sensitive to the BCGF activity of IFN-gamma than B cells obtained from spleens or tonsils. The IFN-gamma-induced proliferation of B cells was short lasting when compared with that of recombinant IL 2 or BCGF containing T cell clone supernatants. B cells preactivated with either Staphylococcus aureus strain Cowan I (SAC) or optimal concentrations of anti-IgM antibodies coupled to beads did not proliferate in response to IFN-gamma, whereas they proliferated in response to IL 2 or T cell clone supernatants. IFN-gamma did not stimulate nor inhibit the proliferative response of human B lymphocytes stimulated with optimal concentrations of anti-IgM antibodies or SAC. Additionally none of the different IFN-gamma tested had B cell differentiation factor activity in the standard SAC assay. These results indicate that IFN-gamma sensitizes B cells to suboptimal mitogenic concentrations of anti-IgM antibody.

Antibodies, Anti-Idiotypic↗
PubMed1986

A flow cytometric micromethod for the detection of Fc epsilon receptors and IgE binding factors using fluorescent microspheres.

Two assays based on the use of fluorescent microspheres have been developed in order to detect Fc epsilon receptors on human cells and human IgE binding factors. A direct assay using microspheres to which IgE was coupled permitted the detection of Fc epsilon receptors on RPMI 8866 cells. However the fluorescence intensity was relatively low and made it difficult to discriminate between positive and negative cells. To increase the sensitivity of the assay, an indirect 3-step test was developed, in which the cells were subsequently incubated with soluble IgE, a polyclonal or monoclonal anti-IgE antibody and fluorescent microspheres to which anti-mouse or anti-rabbit immunoglobulin was coupled. This indirect assay resulted in an increased fluorescence intensity. Optimal discrimination between positive and negative cells was obtained. This assay permitted the detection of human IgE binding factors produced by RPMI 8866 cells. The binding of IgE was not dependent on the origin of the latter. Among the different cell lines tested, the EBV positive lymphoblastoid cells were found to express Fc epsilon receptors and to release IgE binding factors in their supernatants.

Antibodies, Anti-Idiotypic↗
PubMed1986

Isolation and characterization of a human interleukin cDNA clone, homologous to mouse B-cell stimulatory factor 1, that expresses B-cell- and T-cell-stimulating activities.

A cDNA sequence coding for a human interleukin has been isolated from a concanavalin A-activated human T-cell cDNA library based on homology with a mouse interleukin cDNA that expresses B-cell stimulatory factor 1 (BSF-1) activity and T-cell- and mast-cell-stimulating activities. The human cDNA contains a single open reading frame encoding a protein of 153 amino acid residues including a putative signal peptide. Amino acid sequences of the mouse and human polypeptides, deduced from their cDNAs, share extensive homology with the exception of about 40 amino acid residues near the middle portion, which share little homology. Supernatant of COS-7 monkey cells transfected with the human cDNA clone stimulated proliferation of human helper T-cell clones and of anti-IgM-activated human B cells, two properties of mouse BSF-1 on mouse cells. These results indicate that this human cDNA clone encodes a protein structurally and functionally homologous to mouse BSF-1.

Amino Acid Sequence↗
PubMed1986

Cooperation between major histocompatibility complex mismatched mononuclear cells from a human chimera in the production of antigen-specific antibody.

Fetal liver and thymus transplantation can be successfully employed for the treatment of severe combined immunodeficiency disease. In virtually all cases, donor and recipient cells are HLA mismatched. In a patient suffering from a severe combined immunodeficiency disease, full immunological reconstitution was obtained after fetal liver and thymus transplantation. HLA typing revealed that the patient's T cells were of donor origin, while the B cells and monocytes were of host origin. Despite this complete HLA mismatch, the patient was found to mount a subnormal to normal antibody response in vivo. This finding is in contrast with the concept that antigen recognition by T cells is major histocompatibility complex (MHC) restricted. To define the mechanism responsible for this in vivo antibody response, antibody production by peripheral blood mononuclear cells from the patient was tested in vitro after in vivo booster. The in vitro anti-tetanus toxoid antibody production was similar to that of the control group. In addition, specific proliferative responses to tetanus toxoid were obtained. Immunoglobulin allotype determination showed that antibodies were synthetized by host B cells. The results of the present study indicate that transplanted T lymphocytes and recipient cells cooperate despite complete HLA mismatch.

Antibody Formation↗