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R Burger

Publications and source records attributed to R Burger.

At least 145 records · Page 8Linked to original sources
PubMed1986

Quantitative analysis of the cellular constituents of the cerebrospinal fluid in chronic relapsing experimental allergic encephalomyelitis.

Cerebrospinal fluid (CSF) was taken from guinea pigs in various stages of chronic relapsing experimental allergic encephalomyelitis (CR-EAE). The leucocytes in CSF samples were counted and subjected to immunocytochemical analysis using monoclonal antibodies selectively recognising guinea pig T cells, macrophages or Ia antigens. The results showed that total leucocyte numbers and the proportion of macrophages in CSF were elevated in the acute phase of CR-EAE but samples of CSF from animals in early relapse did not show a significant elevation in leucocyte count or macrophage content. In addition the level of T cell activation was higher in CSF than in peripheral blood during disease and was highest in the acute and immediately post-acute phases of the CR-EAE.

Animals↗
PubMed1986

Expression of Ia-antigens on guinea pig Kupffer cells. Studies with monoclonal antibodies.

The expression of the Ia-antigen on guinea pig Kupffer cells was studied employing two monoclonal antibodies against two different determinants of the Ia-molecule. The study was performed in situ on liver sections and on isolated highly purified Kupffer cells kept in culture up to 6 days. The influence of guinea pig hepatocyte culture supernatant and of supernatants of phytohemoagglutinin (PHA)-stimulated human peripheral blood lymphocytes (PBL) on the Ia expression was measured. Immunofluorescence staining of cryostat sections revealed that the monoclonal antibodies used are able to detect Ia-antigens on liver macrophages in situ. The in vitro studies strongly suggest that all Kupffer cells are able to express Ia-antigens. The expression of Ia-antigens on Kupffer cells decreased during the culture time and was no longer detectable on day 5. At this time, incubation of the cell cultures with PBL conditioned medium induced the reexpression of Ia-antigens. Supernatants from guinea-pig hepatocyte cultures did not have any negative influence on the expression of the Ia-antigen on Kupffer cells in vitro. These data support the hypothesis that liver macrophages have the potential to behave immunologically as accessory cells and might contribute to local immune responses in acute as well as chronic states of liver diseases.

Animals↗
PubMed1986

Characterization of a T-lymphocyte membrane protein involved in T-cell function: its contribution to T-cell recognition or cellular interaction.

A monoclonal antibody (Ab188) specific for guinea-pig T lymphocytes recognizes a membrane heterodimer protein (alpha-chain, MW 43,000; beta-chain, 39,000) and inhibits efficiently the antigen-, mitogen- or alloantigen-induced T-cell proliferation. The role of this protein in T-cell activation was analysed in more detail with emphasis on the recognition or activation event of T cells. Quin 2, an intracellularly trapped calcium indicator, was used to measure calcium influx into T cells. The addition of the mitogen concanavalin A to T cells loaded previously with quin 2 induced an increase in fluorescence, revealing an increase in intracellular free calcium. This calcium increase is considered as one of the primary events in the initiation of T-cell activation and was blocked by Ab188. In contrast, other T-cell specific antibodies that react to a comparable extent with T cells had no effect on calcium increase. This indicates that Ab188 is directed to a protein involved in a very early step of T-cell activation. Alloreactive T-cell lines were established from a secondary mixed leucocyte culture by soft agar cloning. Single T-cell colonies were picked and were propagated by repeated restimulation with allogeneic macrophages differing in MHC-class II antigens. In a chromium release assay, the cytotoxic activity of several strain 13 T-cell lines directed against strain 2 Ia antigens was inhibited by Ab188 to about 50%. Similarly, Ab188 inhibited the cytotoxic activity of a MHC-class I-restricted TNP-specific T-cell line of strain 2 guinea-pigs to about 50%. In contrast, lectin-mediated cytotoxicity of the T-cell lines against murine P815 mastocytoma target cells remained unaffected. These results indicate that Ab188 interferes with the function of a protein contributing to the recognition event in the process of T-cell activation.

Animals↗
PubMed1985

Monoclonal antibodies against complement 3 neoantigens for detection of immune complexes and complement activation. Relationship between immune complex levels, state of C3, and numbers of receptors for C3b.

C3-bearing immune complexes and C3 activation products were detected by using two monoclonal antibodies, one specific for a neoantigenic determinant on C3c and the other for C3d. To quantitate immune complexes, the anti-C3c or anti-C3d antibodies were fixed to microtiter plates and reacted with test plasma. The binding of C3-bearing immune complexes in this plasma was then measured with radioisotope- or enzyme-labeled anti-human IgG. To test for C3 breakdown products, solid-phase monoclonal antibody to the C3d neoantigen was reacted with EDTA-plasma samples, and fixed iC3b or C3d was measured with a polyclonal anti-C3 antibody. Patients with autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis, and Sjogren's syndrome, and paracoccidioidomycosis were found to contain immune complexes bearing C3b/iC3b or C3d. In most conditions, there were more C3d-containing immune complexes than C3b/iC3b. Although CR1 (C3b receptors) rapidly converted immune complex-bound iC3b to C3dg/C3d and lupus patients had reduced CR1, no correlation between the state of C3 on circulating immune complexes or levels of immune complexes and CR1 numbers was seen. However, levels of C3-fixing ICs correlated with levels of C3 activation products. This assay system with monoclonal antibodies to neoantigens expressed on activated, but not native, C3 provides sensitive and specific means for detecting and classifying C3-fixing immune complexes and for assessing C3 activation.

Antibodies, Monoclonal↗
PubMed1985

Expression of polypeptide segments of the human complement component C3 in E. coli: genetic and immunological characterization of cDNA clones specific for the alpha-chain of C3.

The third component of complement C3 and its fragments have a central role in a variety of host defense mechanisms. The identification of functionally relevant C3 domains is important because of the marked functional versatility of the C3 molecule. Several human C3 cDNA clones from a human liver cDNA library were isolated and characterized. A bacterial expression vector system was used to express cDNA clones that were identified by an immunological screening procedure. The C3 cDNA clones produced in E. coli the hybrid proteins consisting of cro-beta-galactosidase and polypeptide segments of human C3, as revealed by Western blotting with antisera to human C3. The C3 moiety of the hybrid proteins had a m.w. of up to 46.000. Polyclonal antibodies against the C3 segments expressed by one of the C3 cDNA clones (ReC3-1) have been raised in mice and rabbit, and in addition, a monoclonal antibody was produced. The antisera and the monoclonal antibody reacted in Western blotting analysis selectively with the alpha-chain, but not the beta-chain of human C3. Restriction mapping of the different cDNA clones was performed, and revealed that the different clones were partially overlapping. The ReC3-1 cDNA clone included a 0.7 kb noncoding region at the 3' terminal end of the C3 cDNA. One of the restriction sites (Hind III) identified in the ReC3-1 cDNA clone was not present in the recently published sequence of human C3 cDNA. This difference in nucleotide sequence provides direct evidence for C3 polymorphism at the DNA level. The combination of immunologic procedures with recombinant DNA methodology should facilitate additional analysis of the structure-function relationship of the C3 molecule.

Animals↗
PubMed1985

Cell surface antigens on the guinea-pig macrophage: identification by monoclonal antibodies and association with the activation state.

Ten monoclonal antibodies (moAb) directed against cell surface antigens of guinea-pig monocytes and macrophages (mph) were produced and characterized. The corresponding antigens are not present on granulocytes, T lymphocytes, an Ia-positive B-cell line or other haematopoietic cells. In binding or cytotoxicity assays, the moAb demonstrated characteristics patterns of reactivity, with mph being in different stages of differentiation or activation. Three moAb (342, 322, 249) recognized 'lineage antigens' (i.e. antigens continuously expressed during maturation of monocytes to mph and after stimulation or activation of the cells). MoAb 342 possibly defines a major cell surface determinant, being present on 90% of mph. The antigens detected by moAb 305, 320, 321 and 344 characterize mature mph. They were not expressed on monocytes, but were expressed on the majority of resident, elicited or activated peritoneal mph. MoAb 253, 310 or 257 defined discrete subpopulations of elicited and--with the exception of moAb 257--activated mph. The corresponding antigens were not present on monocytes or resident mph, but appeared on the cell surface during in vivo or in vitro stimulation of the cells. There was no indication of a contribution of the moAb-defined antigens to the presentation of antigen, mitogen or alloantigen by the mph to T cells. The functional significance of the antigens thus remains to be elucidated. Our studies indicate that cells committed to the monocyte/mph lineage share a family of differentiation antigens, distinguishing them from other cell lines. The moAb provide useful tools for further investigation of the activation of mph and allow the rapid detection of mph in different tissues.

Animals↗
PubMed1984

[Dyspnea].

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Adult↗
PubMed1984

Kaposi's sarcoma and malignant lymphoma in AIDS.

A 48-year-old homosexual with contacts in different countries, including Haiti, presented with multiple pigmented or bluish nodules on both lower legs and upper arms. He had a history of secondary syphilis, hepatitis B and herpes zoster ophthalmicus. Biopsies of the skin tumors revealed a typical Kaposi's sarcoma of low grade malignancy. The endothelial origin of the tumor was indicated by the presence of specific endothelial organelles (Weibel-Palade bodies) in the cytoplasma of the tumor cells. Erythrocyte phagocytosis was found in tumor cells within and without the vascular channels. Laboratory tests were compatible with the clinical diagnosis of an acquired immune deficiency syndrome (AIDS) with a helper: suppressor T-lymphocyte ratio of 0.28 and a cutaneous anergy. In the course of the illness tumors of the stomach and duodenum were detected. Histology showed a malignant non-Hodgkin lymphoma of high grade malignancy. Within weeks the patient died in a cachectic state. Autopsy revealed a Kaposi's sarcoma of the skin with metastases in the stomach and a wide-spread malignant lymphoma in the gastrointestinal tract, in several visceral organs and in many lymph nodes.

Acquired Immunodeficiency Syndrome↗
PubMed1984

Non-activated guinea-pig T cells and thymocytes express Ia antigens: FACS analysis with alloantibodies and monoclonal antibodies.

Conventional alloantisera and monoclonal antibodies to guinea-pig Ia antigens were used for analysis of Ia expression by guinea-pig T cells and thymocytes. Indirect immunofluorescent staining was performed with alloantisera or with ascitic fluid as a source of monoclonal antibody followed by flow microfluorometry analysis on the fluorescence activated cell sorter. About 80% of normal, non-activated peritoneal exudate T cells, lymph node T cells and thymocytes expressed Ia antigens. These data are therefore in contrast to studies with human or murine T cells where Ia antigens were shown to be expressed predominantly on activated but not on non-activated T cells. All the reactivity of the anti-Ia alloantisera for strain 2 T cells could be removed by absorption with an Ia-bearing B cell leukaemia, EN-L2C, but not by its Ia-negative variant, BZ-L2C. Thus, the Ia determinants identified on T and B cells are probably identical. One monoclonal antibody, 25E3, which had previously been shown by serologic analysis to react exclusively with an alloantigenic determinant of strain 2 Ia antigens displayed an unusual pattern of reactivity in that it clearly stained strain 13 thymocytes, but not mature strain 13 T or B lymphocytes. The significance of this possible expression of inappropriate Ia determinants by thymocytes remains unclear. This phenomenon might be associated with differentiation processes in the thymus.

Animals↗
PubMed1984

Expression of Ia antigens on macrophages is reduced after stimulation with homologous C3b.

The complement cleavage product C3b was examined for its ability to influence the expression of Ia antigens on macrophages. Monoclonal anti-Ia antibodies were applied for quantitation of Ia antigens by using FACS analysis. Albumin-elicited guinea pig peritoneal macrophages were cultured for 48 hr in the presence of C3b. C3b at final concentrations varying from 10 to 80 micrograms/ml reduced the proportion of Ia-positive macrophages from 87 to 25%. The C3b-stimulated loss of Ia antigens from the macrophage surface was mediated by metabolites of the cyclooxygenase pathway as shown by the inhibitory effect of indomethacin. Exposure of macrophages to C3b for 5 min was sufficient to induce Ia reduction, seen after subsequent 48-hr incubation. C3b generated in vivo in the course of inflammatory reactions might therefore influence the interaction of immunocompetent cells by modulating macrophage Ia expression.

Animals↗
PubMed1984

Activation of the first component of complement, C1, by a monoclonal antibody recognizing the C chain of C1q.

The effect of a purified monoclonal anti-C1q anti-body (Ab 242 G3) on the function of C1q, a subcomponent of the first component of complement C1, was studied. No inhibition of purified activated C1 was observed, whereas binding of the Ab to fluid phase C1q, to C1q bound to immune complexes (EAC1q), or to serum C1 in fluid phase resulted in a dose-dependent inhibition of the hemolytic activity of C1. In contrast, when the effect of the Ab on serum C1 bound to immune complexes (EAC1) was measured, no inhibition, but a dose-dependent enhancement, of the hemolytic activity was obtained. The dose-response curve of the Ab-treated cell-bound serum C1 was indistinguishable from that of activated C1. Isolated Fab fragments of this Ab did not cause an increase in C1 activity. After separation of the A, B, and C chains of C1q by SDS-PAGE, Ab 242 G3 reacted in immunoblotting selectively with the C chain. These data indicate that cross-linking of C1q via the C chain of C1q might lead to an internal activation of C1.

Animals↗
PubMed1984

Quantitative determination of complement components produced by purified hepatocytes.

In this report we describe, on a quantitative basis, the secretion of complement components by hepatocytes. Primary cultures were established after isolation of the cells from guinea-pig liver and the synthesis of C3, C5, C4 and C2 was measured. The cells were isolated by collagenase perfusion of the liver followed by differential centrifugation. The contamination of the hepatocyte suspension with non-parenchymal cells was less than 1%. At 24 h after plating the cells the kinetics of complement production were measured. C3 and C5 content in the culture medium harvested at different time intervals was determined by a sensitive ELISA. Secretion of C2 and C4 was measured haemolytically using C2 or C4 deficient guinea-pig serum. Under the conditions used hepatocytes secreted C3 at a rate of about 100 ng/10(6) cells/h with a plateau of secretion after 24 h of culture corresponding to about 350,000 molecules/cell/h. C5 secretion was detectable after 3-6 h of culture. The C5 secretion rate was about 15 ng/10(6) cells/24 h. The functional activity of C4 and C2 in the supernatants amounted to about 80 SFU/cell/h if the culture medium was changed every 3 h but dropped significantly if the medium was changed every 12 h. The decrease of the haemolytic activity became stronger if the medium was changed every 24 h. Cycloheximide reversibly inhibited the complement production. Our results show that guinea-pig hepatocytes synthesize considerably more C3 and C5 compared to peritoneal macrophages supporting the hypothesis that hepatocytes provide the major source of plasma complement.

Animals↗
PubMed1983

Use of monoclonal and polyclonal antibodies as structural and topographical probes for hepatic epoxide hydrolase.

Monoclonal antibodies have been prepared against rat liver epoxide hydrolase (EH), some of which gave precipitation lines on immunodiffusion against pure EH suggesting the presence of repetitive structural domains on the enzyme. Using ELISA, with polyclonal antibodies to rat and rabbit liver EH, reactivity and therefore structural similarities between EH of all species tested, including human, were observed. This was in contrast to immunodiffusion results demonstrating the limitations of the latter technique. Using monoclonal antibodies in ELISA, greatest structural similarity was between rat, mouse, and Syrian hamster EH and relatively little between rat and human. Two of the antibodies reacted with nearly all species tested and may be directed towards critical sites on the enzyme. This and most of the EH molecule would appear to be localised on the cytoplasmic surface of the endoplasmic reticulum.

Animals↗
PubMed1983

Identification and characterization of gp TFA-1, a guinea pig T cell surface antigen associated with T cell function.

Monoclonal antibodies (Ab) were produced that specifically recognized guinea pig T cells. FACS analysis revealed that Ab 188 bound to the majority of peripheral T lymphocytes of strain 2 and strain 13 guinea pigs and to a minor population of thymocytes. It failed to react with the Ia-bearing guinea pig B cell leukemia line EN-L2C, with macrophages, bone marrow cells, erythrocytes, or thrombocytes. Treatment of T cells with Ab 188 and complement prevented T cell activation. Culturing primed T cells with antigen- or mitogen-pulsed syngeneic or with allogeneic macrophages in the continuous presence of Ab 188 produced a marked, dose-dependent inhibition of T cell proliferation. The antigen defined by Ab 188 was therefore designated guinea pig T lymphocyte function-associated antigen-1, gp TFA-1. The magnitude of inhibition by Ab 188 varied between 65 and 85% whereas three other antibodies to guinea pig T cells had no inhibitory effect on T cell proliferation. Time course experiments revealed that gp TFA-1 is critically involved in an early phase of T cell activation. Maximal inhibition was achieved only if the antibody was present from the beginning of the cell culture; the addition of antibody after 24 hr of culture no longer had an inhibitory effect. Ab 188 did not induce T cell mitogenesis. Two-dimensional analysis (one-dimensional, IEF; two-dimensional, SDS-PAGE) of immunoprecipitates obtained from NP40 lysates of [35S]methionine-labeled T cell blasts indicated that a molecule was specifically precipitated that consisted of two noncovalently associated polypeptide chains with apparent m.w. of 43,000 and 38,000. Both subunits displayed extensive charge heterogeneity focusing at an average isoelectric point of 5.0 and 6.5, respectively. The gp TFA-1 molecule exhibits striking similarities in its functional and structural properties to recently described clonotypically expressed T cell glycoproteins, which were shown to be involved in antigen recognition by T cells in the murine and human systems.

Animals↗