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Biomedical subjects

B Fleischer

Publications and source records attributed to B Fleischer.

At least 217 records · Page 12Linked to original sources
PubMed1986

Function of the CD4 and CD8 molecules on human cytotoxic T lymphocytes: regulation of T cell triggering.

The function of the T cell differentiation antigens CD4 (Leu-3/T4) and CD8 (Leu-2/T8) on human cytotoxic T lymphocytes (CTL) is presently seen only in conjugate formation between CTL and target cell via class II or class I MHC antigens rather than in the later killing steps. In this study, human CD4+ and CD8+ CTL clones were used to investigate the effects of monoclonal antibodies against these differentiation antigens on nonspecific triggering of cytotoxicity. Cytotoxicity was induced either by antibodies against the CD3 (T3) antigen or by the lectins Con A and PHA. Anti-CD4 or anti-CD8 antibodies specifically inhibited all types of cytotoxicity of CD4+ or CD8+ CTL, respectively, regardless of the specificity of the CTL for class I or class II HLA antigens and regardless of whether target cells expressed class I or class II antigens. These results are incompatible with an exclusive role of the CD4 and CD8 molecules in MHC class recognition and are discussed with respect to a function as negative signal receptors for these molecules on CTL.

Antibodies, Monoclonal↗
PubMed1986

Triggering of the lethal hit in human cytotoxic T lymphocytes: a functional role for a 103-kDa T cell-specific activation antigen.

A monoclonal antibody (CB.1) is described that defines a new triggering signal for human cytotoxic T lymphocytes (CTL). The antibody precipitates a 103-kDa surface antigen from activated normal human T cells. The antigen is undetectable or present in only low amounts on resting T lymphocytes but its expression increases strongly after activation and proliferation on T4+ and T8+ T lymphocytes. Binding of antibody CB.1 to CTL results in triggering of the lethal hit. This induction of cytotoxicity is dependent on cross-linking of CTL and an Fc receptor-bearing target cell with CB.1 and requires Ca2+ like antigen-specific triggering. CB.1-induced triggering can be specifically inhibited by binding of antibodies to the T8 or T4 molecules on T8+ or T4+ CTL.

Antibodies, Monoclonal↗
PubMed1986

Lysis of bystander target cells after triggering of human cytotoxic T lymphocytes.

Human cloned cytotoxic T lymphocytes (CTL) specific for class I HLA antigens were used to investigate whether triggering of CTL leads to killing of innocent bystander target cells. After triggering of CTL by their specific target cells, lysis of bystander targets was detected in a 7-h cytotoxicity assay. Considerable differences were found in the susceptibility of various target cells to this type of lysis. Targets susceptible to this bystander lysis were also susceptible to lysis by CTL triggered by F(ab')2 fragments of an anti-T3 monoclonal antibody, whereas other targets were resistant to both types of cytotoxicity. Triggering of CTL by oxidized target cells or via a T3-independent activation pathway led to bystander lysis detectable already after 4 h. Bystander lysis was considerably enhanced under conditions that facilitated a non-specific cell contact between CTL and bystander target. We conclude that a function besides antigen recognition of the T cell receptor on CTL is to direct killing to the target cell. This directing, however, is incomplete and destruction of innocent bystanders can be detected under appropriate conditions.

Antibodies, Monoclonal↗
PubMed1986

Stimulus-dependent triggering or inhibition of cytotoxicity in human cytotoxic T lymphocytes by activators of protein kinase C.

The influence of activators of protein kinase C (PKC) on the delivery of the lethal hit by human cytotoxic T lymphocyte (CTL) clones was studied. In the absence of other signals, short-term incubation with two structurally unrelated activators of PKC, but not with a non-activating phorbolester, resulted in significant triggering of CTL, whereas overnight incubation with PKC activators led to reduction of cytotoxic activity. Furthermore, activation of PKC had an inhibitory effect on simultaneous triggering by anti-T3 monoclonal antibodies or by phytohaemagglutinin, but strongly enhanced the activating effect of anti-T11 antibodies. These results suggest that PKC is part of the cascade of signals transmitted within a CTL after triggering.

Antibodies, Monoclonal↗
PubMed1985

Detection of serum antibodies to Borna disease virus in patients with psychiatric disorders.

Borna disease virus causes a rare meningoencephalitis in horses and sheep and has been shown to produce behavioral effects in some species. The possibility that the Borna virus is associated with mental disorders in humans was evaluated by examining serum samples from 979 psychiatric patients and 200 normal volunteers for the presence of Borna virus-specific antibodies. Antibodies were detected by the indirect immunofluorescence focus assay. Antibodies to the virus were demonstrated in 16 of the patients but none of the normal volunteers. The patients with the positive serum samples were characterized by having histories of affective disorders, particularly of a cyclic nature. Further studies are needed to define the possible involvement of Borna virus in human psychiatric disturbances.

Adult↗
PubMed1985

Activation of human T lymphocytes. III. Triggering of bystander cytotoxicity in cytotoxic T cell clones by antibodies against the T3 antigen or by a calcium ionophore.

The role of T cell differentiation antigens in antigen-specific and nonspecific cytotoxicity by human cytotoxic T lymphocyte (CTL) clones was investigated. In contrast to other reports, several monoclonal antibodies (mAb) against the T3 antigen only marginally blocked antigen-specific cytotoxicity at high concentrations but induced cytotoxicity against third party cells at concentrations from 10 to 0.001 micrograms/ml. Susceptibility to anti-T3-induced lysis was variable but was found with all target cells. Incubation of CTL with anti-T3 mAb even led to self-destruction of the CTL. The effect was independent of the presence of Fc receptors on the target cell and could be obtained with F(ab')2 fragments of the antibody as well. Only activated but not resting T cells could be induced to lyse by anti-T3. Furthermore, this type of bystander killing of target cells could also be induced by the Ca2+ ionophore A23187. Antibodies against the T8 differentiation antigen inhibited antigen-specific, oxidation-induced and anti-T3-induced cytotoxicity by T8+ CTL clones, whereas triggering by the ionophore A23187 was not inhibited. These results show that undirected killing can be triggered in CTL by activating a transducing molecule directly without involving the antigen receptor. Since this triggering of the lethal hit can still be inhibited by mAb against the T8 molecule, the T8 molecule probably has a regulatory role in a late phase of CTL triggering.

Antibodies, Monoclonal↗
PubMed1985

Interferon-beta in patients with low-grade astrocytomas--a phase I study.

In 3 patients with low-grade astrocytomas clinical pharmacology of interferon-beta (10(7) U/mg protein) was investigated. Interferon-beta with escalating dosage (2.3, 6.9, 23, 69 X 10(6) U/patient) was given to each patient in 4 infusions at weekly time intervals. In these patients dose-dependent plasma-levels of interferon-beta of up to 5800 IU/ml were achieved. Plasma concentrations showed a biphasic decline (T1 1/2:0.095-0.49 hrs and T2 1/2: 5-14.5 hrs). Side effects were: mild fatigue, myalgia, tachycardia, hypertension, and fever; the latter was well controlled by pretreatment application of paracetamol. Hematological changes included lymphopenia (2-6 hrs after infusion) and granulocytosis (3-6 hrs after infusion). Natural Killer cell activity was also monitored: 6 hours after infusion a drop of activity - not clearly dose dependent - was observed to a minimum of 1% pretreatment activity; 24 hrs after infusion activity increased up to a maximum of 400%. In this phase I study high biological activity of interferon-beta could be detected in plasma of astrocytoma patients - clinical tolerance was good and only mild toxicity was observed.

Astrocytoma↗
PubMed1985

Homozygous typing cell-defined HLA-Dw specificities correlate better than serologically defined HLA-DR specificities with restriction elements for influenza virus-specific proliferative human T lymphocyte clones.

Human T lymphocyte clones with specific proliferative response to influenza A virus were derived by limiting dilution from peripheral blood lymphocytes (PBL) after in vitro stimulation with autologous irradiated, virus-infected PBL. Four OKT3+4+8- T lymphocyte clones (TLC) that showed HLA-restricted antigen-specific proliferative responses were used for a detailed analysis of the restriction elements for antigen presentation. None of the clones showed alloreactivity and all required the presence on the antigen-presenting cell of HLA class II antigens of one or other haplotype of the donor. Restriction elements for two clones were correlated with Dw1 rather than DR1, and for two others with Dw6 rather than DRw6. These latter clones showed differential recognition of HLA-Dw6 subtypes as defined tentatively by homozygous typing cells, without relationship to putative serological "splits" of DRw6. One of the Dw6-restricted clones was specific for a Dw6.1 (now Dw18) "subtype," confirmed by family segregation analysis, the other for a broad Dw6 (Dw18 and Dw19) specificity. Studies with a panel of monoclonal antibodies against monomorphic determinants of HLA class II antigens revealed heterogeneous patterns of blocking activity, distinguishing between clones of different restriction specificity. Inhibition patterns were partly as predictable from the known activity of the monoclonal antibody in alloantigeneic PLT systems. These results provide evidence that certain structures that function as restriction elements for antigen presentation also carry alloantigeneic determinants.

Antibodies, Monoclonal↗
PubMed1985

A human helper-dependent influenza virus-specific cytotoxic T lymphocyte clone restricted by HLA-DR2.

A human influenza A virus-specific cytotoxic T lymphocyte (CTL) clone is described that is restricted by a class II HLA-antigen. Of a large panel of B-lymphoblastoid cell lines (B-LCL) as target cells the clone lysed only infected cells sharing HLA-DR2 with the donor of the clone. In cold target inhibition studies only infected cells bearing HLA-DR2 could inhibit cytotoxicity. Infected PHA-blasts that express less DR-antigen than B-LCL cells were lysed to a much lesser extent though they could be infected equally well. Trypan blue staining and postlabelling of the target cells showed that the clone indeed killed the target cells. The clone did not proliferate in response to infected irradiated autologous LCL cells and had the OKT3+4+8-DR+ phenotype. The possible significance of class II restricted CTL is discussed.

Antibodies, Monoclonal↗
PubMed1985

Isolation and characterization of an enriched Golgi fraction from neurons of developing rat brains.

We report a method for the isolation of enriched fractions of intact Golgi apparatus from neurons of 10- to 12-day-old rat brains. Neurons were prepared according to a modified method of Farooq and Norton [J. Neurochem. 31, 887-894 (1978)]. Golgi-enriched fractions were obtained after centrifugation of postmitochondrial supernatants in a discontinuous sucrose gradient. Golgi fractions 1 and 2, recovered at the interfaces of 28-34% and 34-36% sucrose densities, respectively, were examined with morphometric and enzymatic methods. Morphometric analyses showed that 21-34% of fraction 1 and 11-29% of fraction 2 consisted of intact Golgi apparatus. Lysosomes, mitochondria, ribosomes, and rough endoplasmic reticulum contaminated fraction 1 (6-10%) and fraction 2 (14-26%). Golgi fraction 1 showed a 25- to 65-fold enrichment over neurons of UDP Gal:GlcNAc galactosyltransferase, CMP-sialic acid:lactosylceramide sialyltransferase, and PAPS:cerebroside sulfotransferase activities. Golgi fraction 2 showed a 8- to 23-fold enrichment over neurons of the activities of the above glycolipid- and glycoprotein-synthesizing enzymes. The activities of the possible marker enzymes rotenone-insensitive NADH-cytochrome c reductase, succinate-cytochrome c reductase, and arylsulfatase were low or minimally elevated in the Golgi fractions. A sevenfold enrichment of Na+, K+-ATPase activities was found in the Golgi fractions. This is consistent either with significant plasma membrane contamination or with the presence of this enzyme in the neuronal Golgi apparatus.

Animals↗
PubMed1985

Recognition of viral antigens by human influenza A virus-specific T lymphocyte clones.

The nature of the viral antigens recognized by influenza A virus-immune cytotoxic T lymphocytes (CTL) is still a matter of debate. We have used four human influenza A virus-specific T lymphocyte clones with antigen-specific cytotoxic and proliferative activity to investigate the requirements for recognition of viral antigens on infected cells. One clone recognized a cross-reactive determinant on the viral hemagglutinin, and two clones were specific for different epitopes on the viral nucleoprotein (NP). A fourth clone seemed to be specific for the viral M protein. Target cell recognition was abrogated by the addition, during infection, of the lysosomotropic drug chloroquine, known to inhibit antigen processing. Furthermore, target cells that had been pulsed with soluble purified NP were recognized and were lysed by the NP-specific clone. This reaction could also be abrogated by the addition of chloroquine during pulsing. These results were obtained irrespective of whether EBV-transformed B lymphoblastoid cells or Ia antigen-expressing T cell blasts were used as target cells. It is concluded that CTL can recognize internal viral proteins that are actively presented at the surface of the target cell. These data indicate that probably every viral protein can function as a target molecule for virus-immune CTL.

Antigen-Presenting Cells↗
PubMed1984

Subcellular distribution of bile acids, bile salts, and taurocholate binding sites in rat liver.

We have quantitated bile acids and their conjugates in rat liver using high-pressure liquid chromatography. Over 95% of the hepatic bile acid pool in rat liver homogenates is present as taurocholate and tauromuricholate. Although over 60% of the bile acid pool is recovered in the supernatant, evidence is presented suggesting that taurocholate redistributes among the subcellular fractions during their isolation. Taurocholate (TC) binding to purified subcellular fractions from rat liver was determined by using equilibrium dialysis in a TC concentration range from 0.1 to 100 microM. This is well below the critical micellar concentration of taurocholate (3 mM). All of the fractions investigated exhibited low-affinity binding with dissociation constants from 80 to 240 microM as did membrane lipid vesicles. Therefore, low-affinity binding appears referable to taurocholate nonspecifically partitioning into the lipid bilayer. High-affinity binding is present in plasma membranes, Golgi, and cell supernatant. The high-affinity binding sites in Golgi have a mean dissociation constant (A1) of 1.0 microM and bind 0.15 nmol of TC/mg of protein. Similarly, the high-affinity binding sites of plasma membrane have an A1 of 1.3 microM and bind 0.15 nmol of TC/mg of protein. For cell supernatant, the A1 was 4.8 microM, and 0.35 nmol of TC was bound per mg of protein. Mitochondria, smooth and rough microsomes, and Golgi liposomes showed no detectable amounts of high-affinity binding. These results are compatible with a role for the Golgi complex, cytoplasmic component(s), and plasma membranes in transhepatic bile acid transport.

Animals↗
PubMed1984

Two distinct mechanisms for taurocholate uptake in subcellular fractions from rat liver.

As part of the enterohepatic circulation, hepatocytes take up bile acids from the intestines via the hepatic portal blood using a sodium-dependent carrier mechanism and resecrete the bile acids into the bile. In order to assess whether intracellular organelles are involved in the transcellular secretion of bile acids, we measured directly the ability of purified subcellular fractions of rat liver to take up taurocholate using a Millipore filtration assay. Two distinct uptake mechanisms can be discerned, one localized in the plasma membranes and the other in the Golgi and smooth microsomal fractions. Plasma membranes prepared by the method of Fleischer and Kervina (Fleischer, S., and Kervina, M. (1974) Methods Enzymol. 31, 6) take up taurocholate in a saturable manner with an apparent Vmax of 2.4 nmol min-1 mg protein-1 and a Km of 190 microM at 37 degrees C. After preincubation of the membranes with K+ ions, a sodium gradient (100 mM outside) stimulates the uptake rate by 90% with the observed Km unchanged. The stimulation is inhibited by phalloidin but not by bromosulfophthalein. Bile canalicular plasma membranes made according to Kramer et al. (Kramer, W., Bickel, U., Buscher, H. P., Gerok, W., and Kurz, G. (1982) Eur. J. Biochem. 129, 13-24) do not take up taurocholate. The transport by Golgi vesicles and smooth microsomes differs from that in the plasma membrane fraction in that it is not stimulated by a sodium gradient, has a Vmax of 12 nmol min-1 mg protein-1 and a Km of 440 microM at 37 degrees C, and is inhibited by bromosulfophthalein but not by phalloidin. Taurocholate uptake into smooth microsomes is abolished by filipin, an antibiotic that complexes with cholesterol to disrupt the membrane. This suggests that taurocholate uptake occurs into a nonendoplasmic reticulum subfraction since endoplasmic reticulum membranes contain negligible amounts of cholesterol. Little uptake was observed using rough microsomes or mitochondria. A model of transhepatic transport compatible with our observations is that taurocholate uptake into the cytoplasm occurs via the plasma membranes on the sinusoidal side of the hepatocyte; taurocholate is then taken up into smooth vesicles and the Golgi complex and is secreted into the bile by exocytosis as the vesicles fuse with the canalicular plasma membranes.

Animals↗
PubMed1984

Activation of human T lymphocytes II. Involvement of the T3 antigen in polyclonal T cell activation by mitogenic lectins and oxidation.

The effect of monoclonal antibodies against the T cell surface antigens T3 and T4 on accessory cell-dependent mitogen-induced proliferation of human antigen-specific T4+ T lymphocyte clones and purified peripheral blood T cells was studied. To avoid the Fc receptor-dependent mitogenic effects of the OKT3 antibodies, monocytes were replaced by Epstein-Barr virus-transformed B lymphoblastoid cells as accessory cells. Monoclonal antibody OKT3 but not OKT4 inhibited the response of both T lymphocyte clones and purified T cells to mitogenic lectins and oxidation. The inhibition was not due to nonspecific effects of the monoclonal antibodies since it affected only the initial triggering but not the proliferation of activated T cells and it could be overcome by higher concentrations of lectin indicating a competition between OKT3 antibody and lectin. Furthermore, OKT3 antibody at the same concentration that was inhibitory in this system was mitogenic in the presence of Fc receptor-bearing monocytes. Surface modulation of T3 but not of T4 antigens led to unresponsiveness to a mitogen pulse given directly after modulation. These findings suggest that antigen-specific and mitogen-induced T cell triggering are due to interaction with the same receptors of the T lymphocyte.

Humans↗
PubMed1984

Ionic requirements for taurocholate transport in rat liver plasma membrane vesicles.

As part of the enterohepatic circulation, taurocholate is taken up by hepatocytes by a Na+-gradient-dependent, carrier-mediated process. The dependence of taurocholate uptake on the presence of a Na+ gradient, outside greater than inside, has been studied in isolated rat liver plasma membranes. The uptake is specific for sodium, and a cotransport stoichiometry of 2 Na+ per taurocholate taken up was found. The presence of K+ ions inside the vesicles was also found to be essential for maximum Na+-stimulated uptake of taurocholate, although a K+ gradient is not required. Mg2+ was almost as effective as K+ in this regard. The symport of Na+ and taurocholate during uptake was shown to be electrogenic, so that K+ may act as an exchange counterion preventing the accumulation of positive charge within the vesicles.

Animals↗
PubMed1984

Phenotypic markers and functional characteristics of T lymphocyte clones from cerebrospinal fluid in multiple sclerosis.

We have recently developed a technique for direct expansion of human T lymphocyte clones from cerebrospinal fluid (CSF) of patients with acute infections of the central nervous system (CNS). In the present study, T lymphocyte clones were established directly from the CSF of 4 patients with multiple sclerosis by limiting dilution in the presence of T cell growth factor and irradiated feeder cells. In 3 patients the CSF was obtained during an exacerbation of their disease. Cloning efficiencies ranged between 4 and 6%. About 40 clones per patient were available for surface marker analysis and functional studies. Typing of the clones for membrane antigens revealed the following results: 75-100% had the OKT4+8- and 0-25% the OKT4-8+ phenotype. Only one clone expressed both surface markers. When tested for PHA-dependent cytotoxicity, all OKT8+ clones and about 50% of the OKT4+ clones were found to express cytotoxic activity. Studies on the proliferative response showed that all OKT4+ and the majority of OKT8+ clones were capable of TCGF-independent, mitogen-induced proliferation. Screening of the clones for specific reactivity against a panel of antigens including measles virus, mumps virus, Epstein-Barr virus and myelin basic protein (MBP) did not reveal significant specific reactivity.

Adult↗
PubMed1984

Immunogenic properties of the small chain HA2 of the haemagglutinin of influenza viruses.

The small chain of influenza virus haemagglutinin, HA2 was isolated by a selective enzymic removal of HA1 or by preparative SDS-polyacrylamide gel electrophoresis. Anti-HA2 specific antisera and monoclonal antibodies were subtype-specific in immunodiffusion tests and radioimmunoassays. These antibodies did not inhibit haemagglutination or haemolysis, did not prevent virus release, did not neutralize infectivity, and HA2 did not induce a protective immunity. HA2-specific antigenic determinants could not be demonstrated on the surface of infected cells. Lymphocytes from pre-immunized mice could not be stimulated by HA2 to exert a cytotoxic effect.

Animals↗