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N Sharon

Publications and source records attributed to N Sharon.

At least 163 records · Page 9Linked to original sources

New clevable photoreactive heterobifunctional cross-linking reagents for studying membrane organization.

The synthesis is described of four new cleavable, photosensitive, heterobifunctional cross-linking reagents for use in examining membrane organization: 4'-azidoazobenzene-4oxysuccinimide ester (1), N-[4-(p-azidophenylazo)benzoyl]-3-aminopropyl-N'-oxysuccinimide ester (2), N-[4-(p-azidophenylazo)benzoyl]-6-aminohexyl-N'-oxysuccinimide ester (3), and N-[4-(p-azidophenylazo)-benzoyl]-11-aminoundecyl-N'-oxysuccinimide ester (4). Two photoaffinity-directed cross-linking agents were prepared by attaching reagents 1 and 2 via their activated ester groups to soybean agglutinin (subunit Mr 30 000). Irradiation of the lectin derivatives resulted in a decrease in their absorption spectra at 360 nm due to photolysis of the bound reagents. Cross-linking of soybean agglutinin subunits following irradiation of the soybean agglutinin derivative to which reagent 2 had been coupled was observed by the appearance of new Coomassie blue staining material (60 000, 90 000, and 120 000 daltons) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. On polyacrylamide gel electrophoresis in the absence of sodium dodecyl sulfate, soybean agglutinin oligomers were observed. Cleavage of the cross-linked soybean agglutinin with 0.1 M sodium dithionite for 25 min at room temperature resulted in the disappearance of the high molecular weight bands and an increase in the amount of uncross-linked material. The use of the photoaffinity-directed agents probing membrane organization is discussed.

Azides↗

Carbohydrates.

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Animals↗

Separation of antibody helper and antibody suppressor human T cells by using soybean agglutinin.

Soybean agglutinin (SBA) binds specifically to mouse B cells and has been used in the past to separate mouse B and T spleen cells by differential agglutination of the B cells. In the present study it was found that a major T-cell subpopulation of human peripheral blood mononuclear cells is agglutinated by SBA along with the B cells and monocytes. Tests of such cell surface markers as Fc receptors for IgG and IgM, as well as functional assays of antibody production by B cells, revealed that the SBA-agglutinated cell fraction contains the antibody helper T cells whereas the unagglutinated fraction is enriched with antibody suppressor T cells. Similar observations were made in tests of the proliferative response to mumps antigen. A recently prepared monoclonal antibody, anti-Leu 2a, which recognized the same thymus-dependent antigen previously defined by a heterologous anti-human T cell serum (alpha TH2), was found to define by indirect immunofluorescence a subpopulation of SBA- cells of intermediate staining intensity which was not detectable in the SBA+ population.

Antibody Formation↗

Alteration of lymphocyte surface properties by insertion of foreign functional components of plasma membrane.

We report the successful coreconstitution of solubilized lymphocyte plasma membranes and Sendai virus envelopes into membrane vesicles that possess the ability to fuse efficiently with mouse splenocytes of thymocytes to give fully viable cells with modified surface properties. Integration of donor membrane components into the acceptor cells was demonstrated by chemical, morphological, and immunological methods. Over 40% of the label from vesicles coreconstituted with radiolabeled (with 125I or 3H) lymphocyte membranes was incorporated into the fused cells. Insertion of biotin-labeled membrane components into the membranes of the acceptor cells was shown by electron microscopy with ferritin-conjugated avidin. Transfer of receptors to soybean agglutinin was also demonstrated. When vesicles from thymocyte membranes were fused with B splenocytes, over 50% of the cells were lysed with anti-theta antiserum and complement. Moreover, lymphocytes into which allogeneic membranes were incorporated triggered the autologous cells in the mixed lymphocyte reaction, indicating the functional transfer of major histocompatibility complex antigens. These findings strongly suggest that the transferred membranes were functionally incorporated into the acceptor cell membranes. The technique described opens new ways for elucidation of the role of the lymphocyte membrane in the immune response as well as for the understanding of the structure-function relationship of membrane components in other cells.

Animals↗

Expression of peanut agglutinin receptors on virus-induced preleukemic cells in mice.

Preleukemic bone marrow and spleen cells of irradiated C57BL/6 mice that were inoculated with the radiation-induced leukemia virus variant D-RadLV differ from autonomous end-stage leukemia cells in the expression of the receptor for peanut agglutinin. As a result, the preleukemic cells are agglutinated by peanut agglutinin and the end-stage cells are not. This observation provides further evidence that preleukemic cells possess surface markers similar to those of the prothymocyte. In vitro and in vivo thymocyte-virus interactions reveal that cells susceptible to D-RadLV transformation are present mainly among the peanut agglutinin-receptor-bearing thymocyte population which has previously been demonstrated to be immunologically immature.

Animals↗

Reactivity of human lymphoid and lymphoblastoid cells with peanut agglutinin: detection of a blood cell subset which lacks detectable membrane HLA.

The number of cells reacting with fluorescein-labelled peanut agglutinin varies widely among human lymphoid organs. By separation techniques, reactive cells from tonsils were characterized as E-negative, non-immunoglobulin-bearing lymphocytes. In peripheral blood, the lectin reacted with a small percentage of lymphoid and monocytoid cells which lacked HLA membrane determinants but expressed the beta 2-microglobulin antigens. The latter phenotype was also found on human lymphoblastoid cell lines. Lectin binding thus provides an effective means of characterizing these cell subsets.

Antigen-Antibody Reactions↗

Mannose-binding activity of Escherichia coli: a determinant of attachment and ingestion of the bacteria by macrophages.

Recently, it was suggested that a mannose-specific lectin on the bacterial cell surface is responsible for the recognition by phagocytic cells of certain nonopsonized Escherichia coli strains. In this study we assessed the interaction of two strains of E. coli at different phases of growth with a monolayer of mouse peritoneal macrophages and developed a direct method with [(14)C]mannan to quantitate the bacterial mannose-binding activity. Normal-sized bacteria were obtained from logarithmic and stationary phases of growth. Nonseptated filamentous cells were formed by growing the organisms in the presence of cephalexin or at a restrictive temperature. Attachment to macrophages of all bacterial forms was inhibited by methyl alpha-d-mannoside and mannan but not by other sugars tested. The attachment of stationary phase and filamentous bacteria to macrophages, as well as their mannose-binding activity, was similar, whereas in the exponential-phase bacteria they were markedly reduced. The results show a linear relation between the two parameters (R = 0.98, P < 0.001). The internalization of the filamentous cells attached to macrophages during 45 min of incubation was much less efficient (20%) compared to that of exponential-phase, stationary-phase, or antibody-coated filamentous bacteria (90%). The results indicate that the mannose-binding activity of E. coli determines the recognition of the organisms by phagocytes. They further suggest that administration of beta-lactam antibiotics may impair elimination of certain pathogenic bacteria by inducing the formation of filaments which are inefficiently internalized by the host's phagocytic cells.

Animals↗

Chemiluminescence and immune cell activation: general features of the thymocyte chemiluminescent responses to plant lectins.

Rat thymocytes respond to exposure to plant lectins by a burst of oxidant generation as detected by chemiluminescence (CL) (CL) in presence of luminol. All lectins tested were capable of evoking CL as long as sufficiently high concentration was used. There was no correlation between the capacity of a given lectin to evoke CL on one hand and its ability to cause mitogenic transformation or thymocyte agglutination on the other. During the first few minutes following exposure to Con A, cells in the thymocyte pool become committed to CL but once commitment is induced, removal of cell bound ligand by addition of excess methyl-alpha-D-mannoside does not significantly alter the response. Succinylated Con A is as effective as native Con A at lower concentrations but is much more effective at higher concentrations in evoking CL. Thymocyte CL can occur in absence of extracellular calcium but this calcium-independent CL is reduced by the addition of magnesium. In presence of calcium, CL response is greatly augmented and the calcium dependent response is not inhibited in presence of magnesium. The response is relatively radiosensitive. Thymocyte CL response is inhibitable by catalase and scavengers of oxidative radicals but is relatively resistant to the effects of superoxide dismutase, requiring high concentrations of significant effect.

Animals↗

Thyroid carcinoma. Immunology, irradiation, and lymphocytic infiltration.

Patients undergoing thyroidectomies at Evanston (III) Hospital, during a six-month period had immunological studies performed preoperatively. No differential could be found between those with carcinoma or benign pathologic findings. T- and B-cell distribution and lymphocytic response to mitogens varied widely. Quantitative immunoglobulins showed slightly increased levels of IgG in patients with carcinoma and thyroiditis in comparison with those patients with adenomas. Antithyroglobulin antibodies were negative in all patients. Pathology slides from 107 patients with thyroid carcinoma between 1972 and 1978 at Evanston Hospital were reviewed for the presence of thyroiditis, either focal or diffuse. It was found that 50% of all carcinomas had either diffuse or focal thyroiditis. Diffuse thyroiditis was more common in patients with no history of irradiation and papillary carcinoma, and in younger age groups.

Adenoma↗

Characterization of reactive and suppressive cells in the mouse embryonic liver by peanut agglutinin (PNA).

Embryonic liver cells suppressed the MLC response of adult mouse spleen cells and reactivity to mitogens Con A, PHA and LPS. Suppression was exerted by cells agglutinated by PNA (PNA+ cells). Cells reacting to LPS and to DxS were found in the nonagglutinated (PNA-) cell fraction. The PNA+ fraction did not react to DxS nor did it reduce the response of sdult spleen cells to this mitogen.

Animals↗

Interaction of peanut agglutinin with normal human lymphocytes and with leukemic cells.

The interaction of peanut agglutinin (PNA) with human thymocytes, peripheral blood lymphocytes, and peripheral blood cells of various types of leukemia was investigated by using fluorescein isothiocyanate-conjugated PNA. The majority of human thymocytes (60-80%) bind the lectin. The major subpopulation of thymocytes that is PNA-positive was separated from the PNA-negative cells by differential agglutination with the lectin. The two thymocyte subpopulations were tested in the mixed lymphocyte reaction and with the phytohemagglutinin of Phaseolus vulgaris. The poor response of the PNA-positive thymocytes to these stimuli indicates that these thymocytes are functionally immature. The fluorescein isothiocyanate-PNA-binding test with peripheral blood lymphocytes of leukemic patients revealed that in most acute leukemias the PNA receptor is exposed on the blastic cells, whereas in most cases of chronic leukemia the peripheral blood lymphocytes are PNA-negative. The validity of PNA as a marker of immature blood cells and its potential clinical application are discussed.

Arachis↗

Prevention of colonization of the urinary tract of mice with Escherichia coli by blocking of bacterial adherence with methyl alpha-D-mannopyranoside.

Methyl alpha-D-mannopyranoside (alpha MM), a competitor inhibitor of the binding of mannose by Escherichia coli, was tested for its ability to prevent infection of the urinary tract of mice with infective strains of the organisms. Injection of the bacteria in the presence of the drug resulted in a considerable reduction in the number of bacteriuric mice. In this system alpha MM was inactive against Proteus mirabilis in accordance with its inability to inhibit the adherence of this organism to epithelial cells in vitro, and methyl alpha-D-glucopyranoside proved inactive against both E. coli and P. mirabilis.

Adhesiveness↗