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.

Apparent loss of 2H4+ T cells in peripheral blood lymphocytes of normal donors: a 2H4-specific artefact unique to T cells.

The antigenic cluster designated CD45R is recognized by a family of monoclonal antibodies. However, one of those most frequently used because of its commercial availability is 2H4. In a series of experiments we show that 2H4 immunofluorescence is almost completely lost from CD4+ or CD8+ T cells of human origin if they are fixed after staining with 2H4.

CD45 regulates signal transduction and lymphocyte activation by specific association with receptor molecules on T or B cells.

Evidence is presented that the leukocyte common antigen CD45 can regulate both signal transduction by lymphocyte receptor molecules and T- and B-cell proliferation in a manner dependent on specific interactions between these receptors on the cell surface. Formation of homoaggregates of CD3, CD2, or CD28 on the surface of T cells induced by crosslinking with monoclonal antibodies (mAbs) results in an increase in cytoplasmic free calcium concentration ([Ca2+]i). This increase in [Ca2+]i was abolished when these receptors were crosslinked to CD45 on the cell surface.

Immunobiologic differences between normal and leukemic human B-cell precursors.

The early stages of normal human B-cell differentiation were studied by flow cytometry and cell sorting based on expression of CD10 (CALLA) and CD19 antigens in fetal liver. Both CD10+ CD19+ and CD10+ CD19- precursor populations proliferated in vitro to form B-cell precursor colonies under stimulation from low molecular weight B-cell growth factor (L-BCGF) or recombinant interleukin 3 but did not respond to high molecular weight B-cell growth factor (H-BCGF). The colonies derived from the CD10+ CD19- fraction showed induction of CD19 expression in 10-50% of growing cells, suggesting that CD10 expression precedes CD19 expression in B-cell ontogeny.

Triggering of neoplastic B cells via surface IgM and the cell surface antigens CD20 and CDw40. Responses differ from normal blood B cells and are restricted to certain morphologic subsets.

By raising monoclonal antibodies (MAbs) against B cells, a number of cell surface molecules have recently been identified which after binding by their specific antibody can trigger B cells, either alone or in co-operation with antibodies to surface immunoglobulin (sIg). The anti-CD20 (Bp35) MAb IF5 can deliver a strong activation signal to resting normal B cells, and the anti-CDw40 (Bp50) MAb G28-5 can promote activated G1 B cells to enter S phase. These antibodies were tested for their functional effects in vitro on suspended cells from 17 follicle-center-cell (FCC) lymphomas, 5 cases of chronic lymphatic B-cell leukemia (B-CLL) and 8 cases of various histological types.

Role of CD2 cross-linking in cytoplasmic calcium responses and T cell activation.

The relationship between the increase of intracellular free Ca2+ concentration ([Ca2+]i) in resting T cells after stimulation with monoclonal antibody (mAb) to CD2 (E rosette receptor) and the subsequent proliferation response was investigated. Although the combination of 9.6 plus 9-1 mAb to CD2 was both mitogenic and induced an increase in [Ca2+]i, cross-linking of an individual CD2 mAb on the cell surface induced an increase in [Ca2+]i without directly stimulating T cell proliferation.

Effects of anti-gp120 monoclonal antibodies on CD4 receptor binding by the env protein of human immunodeficiency virus type 1.

Monoclonal antibodies (MAbs) to defined peptide epitopes on gp120 from human immunodeficiency virus type 1 were used to investigate the involvement of their epitopes in gp120 binding to the CD4 receptor. Recombinant vaccinia viruses were constructed that expressed either full-length gp120 (v-ED6), or a truncated gp120 lacking 44 amino acids at the carboxyl terminus (v-ED4). Binding of these glycoproteins to the CD4 receptor was detected directly with metabolically labeled gp120 or indirectly with the gp120 MAbs.

The Fourier transforms of the laser-induced absorption decay from glycogen phosphorylase and DOPA decarboxylase.

Fourier analysis of the laser-induced absorption decay curves of 3,4-dihydroxyphenylalanine (DOPA) decarboxylase and glycogen phosphorylase demonstrates a powerful technique in the analysis of complicated decay behavior. Phosphorylase which uses the pyridoxal 5'-phosphate cofactor in an unknown manner exhibits over weak absorption an intense decay while decarboxylase demonstrates only weak absorption. Fourier analysis of the decay curves clearly shows that phosphorylase has an intense absorption decay in the midst of three weaker ones and that decarboxylase only has three weak decays.

Fluctuations of CD4+ T-cell subsets in remitting-relapsing multiple sclerosis.

Patients with multiple sclerosis (MS) frequently have selective depletion of the CD45R+CD4+ T-cell subset during active phases of disease. To study the relationship between changes in this subset and the onset of objective clinical exacerbations of disease, a longitudinal study was undertaken. Two CD4+ T-cell subsets and two CD8+ T-cell subsets were monitored by two-color immunofluorescence using a fluorescence-activated cell sorter.

HIV-1-infected T cells show a selective signaling defect after perturbation of CD3/antigen receptor.

The binding of antigen or monoclonal antibody to the T cell receptor for antigen or the closely associated CD3 complex causes increases in the concentration of intracellular ionized calcium and subsequent cell proliferation. By measuring second messenger production in primary cultures of human immunodeficiency virus (HIV-1)--infected T cells stimulated with monoclonal antibodies specific for either CD3 or CD2, a specific impairment of membrane signaling was revealed. The HIV-1--infected T cells were unable to mobilize Ca2+ after stimulation with anti-CD3, whereas CD2-induced calcium mobilization remained intact.

Selective loss of CD4+ CD45R+ T cells in peripheral blood of multiple myeloma patients.

The phenotypic distribution of T-lymphocyte subsets in peripheral blood from multiple myeloma (MM) patients shows a reduced proportion of CD4+ cells and a normal proportion of CD8+ cells. The decrease in CD4+ cells could be due to a random process, with all types of CD4+ cells being equally affected, or it could reflect a nonrandom process with selected subsets preferentially reduced. In order to distinguish between these possibilities, double immunofluorescence analysis was performed on blood samples from patients with MM, patients with monoclonal gammopathy of unknown significance (MGUS), and age-matched normal donors, using monoclonal anti-CD4 or anti-CD8 paired with antibodies to the common leukocyte marker Lp220 (CD45R) or 4B4 (CDw29).

Antibody to a novel 95-kDa surface glycoprotein on human B cells induces calcium mobilization and B cell activation.

These findings characterize a 95-kDa glycoprotein on the surface of B lymphocytes recognized by the mAb G28-8. This protein (designated Bgp95), previously classified as a CD39 molecule, is unique based on functional, cell distribution, and immunochemical criteria. Biochemical analyses revealed that Bgp95 is a 95-kDa glycoprotein with N-linked carbohydrate and is reduced to about 70-kDa after treatment with endoglycopeptidase F.

Lysis of cells infected with HIV-1 by human lymphocytes targeted with monoclonal antibody heteroconjugates.

The present study was undertaken to determine whether human PBL can be specifically focused to lyse cells infected with HIV-1 by mAb heteroconjugates that can bridge target and effector cells. A mAb directed against the central portion of HIV-1 glycoprotein gp110 was chemically cross-linked to a mAb directed against the CD3/TCR complex or to a mAb directed against the CD16 Fc gamma-R expressed on large granular lymphocytes (LGL). HIV-1-infected cells, but not uninfected cells, were found to be lysed to a greater extent by PBL in the presence of the gp110 X CD3 or the gp110 X CD16 antibody heteroconjugate than in the presence of the single antibodies or a mixture of the mAb comprising the heteroconjugates.

Signal transduction through CD4 receptors: stimulatory vs. inhibitory activity is regulated by CD4 proximity to the CD3/T cell receptor.

The binding of antibody to the CD4 molecule inhibits mobilization of cytoplasmic free calcium ([Ca2+]i) in response to CD3 cross-linking on resting T cells. Similarly, when CD3 and CD4 are independently and simultaneously cross-linked, calcium mobilization is inhibited when compared to that induced by cross-linking CD3 alone. In contrast, when anti-CD4 and anti-CD3 are cross-linked together, calcium mobilization is substantially higher than from CD3 cross-linking alone.

Use of colony assays and anti-T cell immunotoxins to elucidate the immunobiologic features of leukemic progenitor cells in T-lineage acute lymphoblastic leukemia.

The specific binding of radioiodinated rIL-2 to fresh marrow blasts from T-lineage acute lymphoblastic leukemia (ALL) patients was initially investigated. The estimated number of radioiodinated rIL-2 molecules bound per blast ranged from undetectable to 1948. In colony assays, 72% of 32 cases analyzed showed a significant proliferative response to rIL-2, which depended on PHA-stimulated lymphocyte conditioned medium activation.

Differential regulatory signals delivered by antibody binding to the CD28 (Tp44) molecule during the activation of human T lymphocytes.

Molecule CD28 (Tp44) is expressed on the surface of majority of human T cells and has been implicated to play an active role in the regulation of T cell growth. The present study examines the effect of antibody binding to the CD28 molecule during T cell activation. Anti-CD28 but not isotype-matched anti-CD5 mAb consistently augmented anti-CD3-induced and IL-2-induced T cell proliferation and subsequent release of soluble CD25 molecule.

Resting B lymphocytes can be triggered directly through the CDw40 (Bp50) antigen. A comparison with IL-4-mediated signaling.

Resting tonsillar B lymphocytes were shown to enlarge and become more buoyant when exposed to either IL-4 or a mAb (G28-5) to the 50-kDa CDw40 Ag. A striking feature of activation through CDw40 was the promotion of strong homotypic adhesions which did not occur in populations cultured with IL-4. Whereas the CDw40 antibody down-regulated its target Ag, an increased expression of CDw40 accompanied IL-4 stimulation.

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.

Soluble CD23 is released by B lymphocytes cycling in response to interleukin 4 and anti-Bp50 (CDw40).

An enzyme-linked immunoassay was developed to quantitate the production of soluble CD23 from cycling B lymphocytes. This molecule has identity both with B cell-derived B cell growth factor and with an IgE-binding factor. B lymphocytes, which had been stimulated for 3 days with phorbol dibutyrate and calcium ionophore, washed and recultured, failed to produce detectable levels of CD23 over the following 3 days.

Loss of CD45R (Lp220) represents a post-thymic T cell differentiation event.

CD45R+ and CDw29+ CD4+ T cells are widely regarded as separate functionally defined T cell lineages. The work described here indicates that they represent maturation stages within the same differentiation pathway. Purified populations of CD4+ or CD8+ T cells, after stimulation with PHA, lose cell surface expression of CD45R (Lp220) and gain an increased surface density of CDw29 (4B4).

Molecular cloning of the human B cell CD20 receptor predicts a hydrophobic protein with multiple transmembrane domains.

CD20 is an antigen expressed on normal and malignant human B cells that is thought to function as a receptor during B cell activation. Here we report the isolation of a CD20-specific cDNA clone from a lambda gt11 library using a polyclonal antiserum raised against purified CD20 antigen. Additional cDNA clones were then isolated from a lambda gt10 library.

CDw40 and BLCa-specific monoclonal antibodies detect two distinct molecules which transmit progression signals to human B lymphocytes.

Two monoclonal antibodies (mAb), MA6 and G28-5, have the common property of detecting markers expressed on both B lymphocytes and carcinomas: BLCa (B lymphocyte carcinoma cross-reacting antigen) and CDw40 (Bp50). A comparison of the reactivity of these mAb revealed that MA6 and G28-5 detect distinct epitopes with different cell line and tissue distributions. L cell transfectants expressing CDw40 were not bound by MA6 anti-BLCa, but were bound by G28-5 anti-CDw40.

Antigen-independent regulation of cytoplasmic calcium in B cells with a 12-kDa B-cell growth factor and anti-CD19.

Increases in cytoplasmic free calcium ([Ca2+]i) can be induced in resting B cells either by a low molecular weight (12-kDa) B-cell growth factor (LMW-BCGF) or by crosslinking the B-cell antigen CD19 with monoclonal antibody (mAb). LMW-BCGF causes a slow [Ca2+]i increase in peripheral blood and tonsillar B cells but has no effect on [Ca2+]i in resting T cells. B-cell [Ca2+]i responses mediated by anti-surface immunoglobulin (sIg) or anti-CD19 are potentiated by LMW-BCGF, but anti-sIg and anti-CD19 do not show additive [Ca2+]i responses.

Reducing the risks of laryngoscopy in anaesthetised infants.

We have evaluated the use of oxygen insufflation during laryngoscopy with an Oxyscope laryngoscope blade compared to conventional laryngoscopy for maintenance of transcutaneous PO2 during intubation of anaesthetised, spontaneously breathing infants. Twenty healthy children aged between 1 and 24 months were anaesthetised with halothane in oxygen. Laryngoscopy and intubation were performed in a double-blind fashion using a Miller No.