[Diagnostic value of cerebrospinal fluid oligoclonal immunoglobulins in multiple sclerosis and inflammatory disease of the nervous system].
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Biomedical subjects
Publications and source records attributed to H Ovadia.
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Myelin basic protein (MBP) of guinea pig origin was incorporated into magnetically responsive albumin microspheres. Protein-protein bonding and stabilization of the GPMBP microspheres by heating at 120 degrees C did not adversely influence their capacity to bind anti-MBP antibodies or demonstrably alter the encephalitogenic activity of the incorporated GPMBP. The magnetic properties of the particles and the fact that immunodeterminants of some of the incorporated MBP fortuitously were distributed on the exterior surfaces of the microspheres allowed a number of experiments to be carried out in Lewis rats for the first time: (a) selective capture and deletion of that particular subpopulation of lymphoid cells responsible for transfer of experimental allergic encephalomyelitis (EAE) represented within the lymph node cells (LNC) of donor animals sensitized to neutral antigen, (b) enhancement of in vivo uptake of MBP by macrophages (M phi s) contained in oil-induced peritoneal cell exudates and exposed briefly to MBP microspheres, and (c) preparation of cell suspensions specifically enriched with respect to MBP-containing M phi s.
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Daily administration of indomethacin (5 mg/kg) to Lewis rats from day 7 through day 21 following sensitization to guinea-pig spinal cord neural antigen plus Freund's complete adjuvant (FCA) resulted in an aggravation of clinical signs of EAE accompanied by a higher mortality. In contrast, lymph node cells (LNC) from sensitized and drug-treated donor rats showed decreased EAE transfer activity. The in vitro Con A response of LNC from sensitized, indomethacin-treated rats was comparable to those of normal LNC or LNC from sensitized animals only. The spleen cell response of sensitized rats was significantly lower than the response seen in normal animals, but was returned to normal levels in sensitized, indomethacin-treated animals. We believe these observations reflect an indomethacin-mediated diminished accumulation of EAE effector cells within peripheral lymph node tissues together with abrogation of splenic suppressor cell activity by indomethacin-mediated inhibition of prostaglandin (PG) synthesis in Lewis rats sensitized and treated with this drug under the conditions defined.
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Protein A, a protein derived from Staphylococcus aureus, was incorporated into the matrix of magnetic albumin microspheres. Because staphylococcal protein A binds most subclasses of immunoglobulin G through their Fc portions, immunoglobulin may be rapidly bound to microspheres without chemical coupling agents or a diamino-heptane spacer group. Microspheres so prepared bind specifically to a given cell type when incubated in vitro with a heterogeneous cell population. The use of these microspheres as a drug carrier capable of cellular specificity as well as their ability to isolate homogeneous cell populations rapidly is discussed.
A method to rapidly perform immunofluorescence or light microscopic staining on formalin-fixed paraffin sections has been devised utilizing magnetic albumin microspheres containing Staphylococcal protein A. Because the protein A constituent of the microspheres has the property of binding the Fc portion of immunoglobulin G (IgG) class antibodies, the microspheres can be used to rapidly bind antigen-antibody complexes by the Fc portion of the antibody. Deparaffinized sections were stained with fluorescein isothiocyanate-conjugated antibody (IgG fractions) by standard techniques, after which the protein A microspheres were layered over the sections. Distinct fluorescence of sections was noted with the addition of the microspheres, whereas only autofluorescence was present with direct staining alone. The microspheres were also visualized by light microscopy by a subsequent Prussian blue reaction, staining the Fe3O4 within the microsphere matrix. This method represents a more rapid method for identifying antigens in tissues embedded in paraffin than has previously been reported.
Normal immunosuppressive protein, prepared from human plasma by DEAE-cellulose chromatography, inhibits DNA synthesis in human cell lines of lymphocytes of both T and B origin. It also inhibits [3H]thymidine incorporation in mouse cell lines. Normal immunosuppressive protein was able to inhibit the proliferation of these cells, although they were already transformed and had a high rate of DNA synthesis. On the other hand, it does not inhibit myeloid cells or fibroblasts.
An improved method for the preparation and purification of normal immunosuppressive protein (NIP) is described. The purified material has a molecular weight between 10,000 and 25,000. Its biological and serological activity is approximately 10--20 times higher than that of the crude fraction. An antibody to normal immunosuppressive protein prepared in rabbits made the quantitative estimation of NIP by a haemaggluination inhibition test possible. Similarly, a very sensitive assay for the quantitative determination of NIP by its inhibitory effect on the proliferation of EL-4 tumor cells is also described. Eluates prepared from polyacrylamide gels were active in inhibiting EL-4 tumor cell proliferation and neutralized the anti-NIP activity in the haemagglutination inhibition test.
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The biologic activity of normal immunosuppressive protein (NIP) isolated from human plasma was studied. NIP was found to inhibit the proliferation of both T and B lympohcytes in vitro. It suppressed the DNA synthesis of normal mouse lymphocytes responding to the mitogens phytohemagglutinin and lipopolysaccharide, as well as the [3H]Thymidine and [3H]leucine uptake by T and B lymphoid cell lines of human and murine origin. The lymphoid specificity of NIP was demonstrated by showing that DNA and protein synthesis of normal and transformed fibroblasts and other nonlympohid cell lines was not affected by NIP treatment. Furthermore, by using lymphoid cell lines we were able to show that 1) NIP inhibits the process of ongoing DNA and protein synthesis; 2) the duration of the cells' exposure to NIP is crucial for obtaining optimal effect; and 3) the inhibitory effect of NIP is totally reversible.
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Normal immunosuppressive protein isolated from human plasma was found to inhibit the generation of primary cytotoxic effector lymphocytes against allogeneic tumour cells in vitro. Total inhibition was observed when NIP was present during the early stages of the sensitization process. In contrast, the generation of secondary cytotoxic lymphocytes in vitro was only slightly inhibited even though large amounts of NIP were used. The inhibition of target cell lysis by sensitized lymphocytes required long preincubation of a relatively small number of effector cells with large amounts of NIP and was most significant when tested at low effector: target cell ratios. Under the same conditions NIP showed no inhibitory effect on the cytotoxic activity of immune macrophages. The present in vitro experiments suggest that NIP exerts its effect through inhibition of DNA synthesis and cellular proliferation and to a limited extent only, by inducing specific suppressor cells.
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The effect of restraint stress on the distribution of lymphocyte subsets were studied in young BALB/c male mice. Loss of whole body weight, a reduction in the weights of spleen and lymph nodes, and higher levels of serum corticosterone were evident after a single continuous restraint period of 16-18 h. Tissue sections of spleens from restrained animals revealed erythrocyte depletion in the contracted red pulp. Furthermore, there was a significantly higher proportion of CD4+, but not of CD8+, lymphocytes in the spleen. The proportion of the CD4+ subset was markedly diminished in peripheral blood, whereas no changes were detected in lymph nodes. Restraint resulted in enhanced allogeneic mixed lymphocyte reactivity and in altered expression of some CD4+, but not CD8+, splenocyte adhesion molecules (CD44, LFA-1 and VLA-4). Removal of circulating corticosteroids by surgical adrenalectomy abolished the restraint-induced changes in lymphocyte adhesion molecule expression. The findings suggest that the observed differences in lymphocytes subset distribution of lymphoid organs may be due to changes in the pattern of adhesion molecule expression.
In the present study we have shown for the first time that lymphocytes from the spleen, thymus and mesenteric lymph nodes can specifically bind and retain 3H-dexamethasone in the nuclear fraction. The use of the specific corticosteroid receptor antagonists, RU28362 and RU38486, for the displacement of labeled dexamethasone revealed that binding was mainly of type-II glucocorticoids receptors. In lymphocytes obtained from mesenteric lymph nodes, binding was significantly higher than in cells obtained from spleen and thymus. Long-term adrenalectomy (up to 30 days), which markedly elevated serum ACTH, caused a time-dependent increase in binding in all lymphoid cells. No increase in binding was found in lymphocytes obtained from rats hypophysectomized for 15 days, in which neither serum ACTH nor corticosterone could be measured. These results suggest that the cellular nuclear binding in lymphocytes may be regulated by a pituitary factor.