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Biomedical subjects

M Teodorescu

Publications and source records attributed to M Teodorescu.

At least 37 records · Page 2Linked to original sources
PubMed1985

Immunobiological function of normal rabbit synovial cells.

The ability of enzyme-dissociated primary cultures of synovial cells (Sy) to present antigen was investigated. Adult rabbits were immunized in foot pads with bovine serum albumin (BSA) in complete Freund's adjuvant (CFA) or with CFA alone. Four to six weeks later draining popliteal lymph node cells (LNC) and synovial cells were obtained. Synovial cells were cultured overnight with or without antigen. About 20% of these synovial cells had Fc receptors and 15% C3 receptors. As a positive control splenic adherent cells (SAC) were similarly treated. Next day, autologous lymph node cells were added to the extensively washed and irradiated synovial cells or splenic adherent cells. Lymphocyte proliferation was measured by [3H]thymidine uptake. Synovial cells as well as splenic adherent cells induced mixed lymphocyte reaction (MLR) and autologous mixed lymphocyte reaction (AMLR) and effectively presented antigen for specific immune response to the priming antigen. Thus, the synovium contains macrophage-like cells that can effectively interact with lymphocytes and participate in the immune phenomena in the joints of patients with rheumatoid arthritis.

Animals↗
PubMed1985

The use of bacteria as probes for lectins in preparations of solubilized human tonsil cell membranes.

In earlier work we have shown that some bacteria bind naturally to lymphocyte subpopulations and that this binding may be due to lectin-carbohydrate interactions. Here we determined the possibility of using bacteria to probe for these lectins in solubilized tonsil cell membrane preparations. Since lectins are capable of agglutination, we determined the ability of human tonsil cell membrane extract (TCME) to agglutinate bacteria. We used Escherichia coli strain YS57 which does not bind to human lymphocytes and a mutant strain derived from it, E. coli UI 2023, which binds to about 50 percent of human lymphocytes. The UI 2023 was agglutinated while the YS57 was not; this agglutination was not due to antibodies or DNA. When E. coli UI 2023 was treated with periodate, it lost its ability to be agglutinated. The agglutination of E. coli UI 2023 was not blocked by any of the monosaccharides and disaccharides used but was blocked by the E. coli LPS, more specifically, by its carbohydrate moiety. Also, the E. coli UI 2023 absorbed the agglutinating factor while its parental strain, YS57, did not. Sodium dodecylsulfate-polyacrylamide gel electrophoresis of TCME after absorption with bacteria showed that a band around 67kD was absent in the TCME absorbed by E. coli prevented the absorption by E. coli UI 2023 whereas Na2IO4-treated LPS did not. In addition, tonsil cell membrane was radioiodinated before obtaining the TCME; sodium dodecylsulfate-polyacrylamide gel electrophoresis of the radioiodinated TCME recovered after elution from E. coli UI 2023, but not from E. coli YS57, showed again a band around 67 kD.(ABSTRACT TRUNCATED AT 250 WORDS)

Agglutination↗
PubMed1985

Allogeneic lymphocyte stimulation in rabbits: induction of a low MW inhibitor for trypsin and for a concurrently induced alpha-macroglobulin-proteinase complex.

We have shown previously that the i.v. inoculation of allogeneic lymph node cells in rabbits induces the appearance in the serum of an alpha M-serine proteinase complex which behaves in an Ig-turnover assay as any polyclonal B-cell activator (PBA), and that this PBA activity is due to the enzyme. Here, we show that the allogeneic stimulation also induces the appearance in the low molecular weight fraction of the serum (1000-110,000 MW) of an inhibitor which blocks the PBA activity of the complex without affecting the PBA activity of LPS or dextran sulphate. The inhibitor blocked the ability of the enzyme associated with alpha M to degrade Chromozym TRY, a low MW trypsin substrate. The inhibitor also blocked the enzymatic activity of trypsin for large as well as for low MW substrates. Thus, allogeneic stimulation in vivo results in the production, not only of an alpha M-proteinase complex, but also of an inhibitor for this proteinase as well as for trypsin. The appearance of the inhibitor, along with the alpha M-serine proteinase complex as a result of allogeneic stimulation in rabbits, is of interest since a similar alpha M-serine proteinase complex and inhibitor may appear in the serum of patients with rheumatoid arthritis.

Animals↗
PubMed1985

Specific cytotoxicity of human lymphocyte subpopulations defined by bacterial adherence.

It has been shown that lymphocytes can be subdivided into subpopulations based on their binding of bacteria. Monolayers of immobilized and fixed bacteria were used here to separate T cells into BA-T1T2, adherent to Escherichia coli-2 (EC-2+) and BA-T3T4, non-adherent to this strain of bacteria (Ec-2-) (our denomination). The cells were activated in mixed lymphocyte cultures (MLC) and tested for cytotoxic activity. The BA-T1T2 cells developed the same cytotoxic activity as the sham-separated T cells whereas BA-T3T4 cells did not become cytotoxic. When the T cells were separated into BA-T2 cells, adherent to Bacillus globigii (Bg+), and BA-T1T3T4, non-adherent, (Bg- cells became cytotoxic. Since BA-T1 cells, which represent 10-20% of T cells, are common to the two populations they appear to contain all T cells needed to develop the specific cytotoxicity for allogeneic cells. When the cells were first activated in MLC for 6 days and then separated by adherence to E. coli-2 or B. globigii, all cytotoxic cells were in the non-adherent fraction. We concluded that the subpopulation of T cells which are Ec-2+Bg- (less than 20%) contain all the cells required for the development of cytoxic cell function and that after activation they become Ec-2-Bg-.

Bacillus↗
PubMed1985

Class II histocompatibility antigen-mediated immunologic function of normal articular chondrocytes.

Using monoclonal antibodies, we examined the display of rabbit Ia by articular chondrocytes. We found that 29 to 46% of chondrocytes displayed Ia antigen compared with 46 to 60% of spleen cells. Ia antigen expression was not likely to be the result of enzyme treatment. To investigate antigen presenting activity of enzyme dissociated normal articular chondrocytes, adult rabbits were immunized in the front foot pads with ovalbumin (OVA) in complete Freund's adjuvant. Four to six weeks later, draining popliteal lymph node cells (LNC) were obtained. Articular chondrocytes were obtained by overnight collagenase, DNase, and hyaluronidase digestion of cartilage from both ends of femurs and proximal end of tibias. Antigen-presenting cells from spleen were used as positive controls. LNC and nylon wool-purified T cells were cultured with OVA pulsed and mitomycin C-treated chondrocytes or spleen cells, and lymphocyte proliferation was measured by 3H-TdR uptake. Both chondrocytes and spleen cells showed antigen presenting activity, and stimulation of lymphocyte proliferation was inhibited by murine monoclonal anti-rabbit Ia antibody (2C4), whereas control plasmacytoma cell supernatants had no effect. When T cells were purified first by Sephadex G-10 and later by nylon wool columns, these cells were dependent on antigen-presenting cells for immunogen (OVA)-induced lymphocyte proliferation. Again, chondrocytes under these strict experimental conditions presented antigen to T cells. Chondrocytes also stimulated autologous and allogeneic normal lymphocytes. Thus, normal chondrocytes have Ia antigens on their surface and can function as antigen-presenting cells. These results are significant for the understanding of local cellular interaction in the pathogenesis of rheumatoid arthritis.

Animals↗
PubMed1985

The role of intercellular contacts in the activation of B lymphocytes by anti-immunoglobulin antibodies.

It has been proposed that anti-Ig antibody activates B cells in a way analogous to the antigens, i.e., it delivers its signal by cross-linking and clustering the Ig receptors on the surface of one cell. However, the cross-linking of different B cells which may deliver to each other a signal has not been considered. Thus, we examined the effect of preventing cell contacts on the response of rabbit B cells to anti-Ig allotype antibody by using a solid agarose medium. First, we examined the 14C-uridine incorporation in liquid medium and in liquid or solid agarose in cultures stimulated with anti-Ig antibody (Ab). The response was high in liquid agarose or liquid medium but was absent when the agarose-containing medium was solidified at the start of the cultures. If the agarose was solidified 6 hr after the start, a good response was obtained. Moreover, if the cells were sedimented at the start before solidifying the agarose-containing medium, a good response was also obtained. Similar results were obtained when T cells were activated by Con A. To examine whether B cells require contacts with other B cells or with non-B cells, we examined their response to anti-Ig Ab in the absence of macrophages or T cells, and found that purified B cells from lymphoid organs or from thoracic duct responded well in the absence of T cells and/or macrophages. The response was also absent when the cells were cultured in agar at a "local" concentration close to 10(8) cell/ml. Also, concentrated supernatants of anti-Ig-stimulated cells did not increase the response to anti-Ig in solid agarose. These two last observations suggest that the lack of response in solid agarose is not due to a lack of diffusible factors or to a lack of feeder cells. Therefore, because the only difference between the cultures that responded to anti-Ig or Con A and those that did not was the distance between the cells, we concluded that the contact between B cells or between T cells is essential to their activation by their respective mitogens. We speculate that the anti-Ig Ab or the antigen cross-links B cells, which then provide each other with the activating signal or with one of the activating signals.

Animals↗
PubMed1984

Enhancement of human MLR by very low concentrations of lipopolysaccharide and blocking of this enhancement by polymyxin B.

Addition of 50 micrograms LPS/ml to as little as 5 pg of LPS/ml to the allogeneic human mixed leukocyte reaction (MLR) enhanced [3H]thymidine incorporation 2-4-fold. The same concentrations of LPS had no effect on thymidine incorporation in cultures without the allogeneic cells. The Limulus amebocyte lysate (LAL) test we used was sensitive only to 15 pg LPS/ml indicating that the MLR is more sensitive to LPS than the LAL test. A commercially available RPMI 1640 culture medium used by many investigators was found to contain concentrations of LPS that would significantly enhance the MLR. Polymyxin B blocked the MLR-enhancing effect of LPS while having no inhibitory or stimulating effect itself. We concluded that low concentrations of LPS are helpful in revealing MLR incompatibilities by enhancing the lymphocyte responses and that the addition of polymyxin B is essential in the study of factors affecting the MLR by eliminating any LPS contribution.

Cells, Cultured↗
PubMed1984

Degradation of a chromogenic substrate by alpha 2-macroglobulin from plasma of patients with rheumatoid arthritis.

We have shown previously that serum from patients with rheumatoid arthritis (RA) contains a polyclonal B cell activator that is associated with alpha 2-macroglobulin (alpha 2M). Some biologic effects of this activator appear to be due to a trypsin-like protease attached to alpha 2M. Therefore, in the present study, we used an anti-alpha 2M antibody solid-phase assay, with Chromozym-Try as a substrate, to determine the level of alpha 2M-protease complexes in plasma alpha 2M. We found higher levels of these complexes in RA patients than in 2 control groups. Since alpha 2M-protease complexes have been shown to induce RA-like inflammation in experimental animals and to be produced by lymphoid cells, we speculate that they may be involved in the pathogenesis of RA. However, the role of the other cells or enzyme systems in the formation of these complexes has not yet been ruled out. Results of these investigations could lead to another link between activation of the immune system and joint inflammation.

Adult↗
PubMed1984

Binding of bacteria in lymphocyte subpopulations: role of lectin-carbohydrate interactions.

Bacteria have been found to bind to lymphocyte subpopulations in a highly reproducible manner. Some of these bacteria such as B. melitensis and a strain of E. coli binds to mammalian B. cells. The binding of B. melitensis and other bacteria is due, at least in part, to lectins on lymphocytes interacting with the carbohydrates on the LPS or LTA of the bacteria. These receptors for bacteria give some indications regarding the functional potential of the cells, suggesting the possibility that the receptors identified by bacteria are used in cellular interactions with normal or malignant cells.

Animals↗
PubMed1984

Detection of alpha 2-macroglobulin-associated proteases in the plasma of patients with rheumatoid arthritis.

In previous work a polyclonal B cell activator has been detected in the serum of patients with rheumatoid arthritis (RA). This activator is associated with alpha 2-macroglobulin (alpha 2M) and its activity is blocked by low-Mr trypsin inhibitors, which suggests that it may be a protease-alpha 2M complex. Here we determined the possibility of developing a routine clinical chemistry test for detection of this complex in patients' blood. We measured with chromogenic substrates the total proteolytic activity of citrated plasma and of the alpha 2M immunoabsorbed from plasma. Low-Mr substrates containing Arg were degraded much better by plasma from RA patients than by plasma from patients with other arthritides. Low-Mr substrates containing Leu, Lys, or Gly or the large-Mr substrate Azocoll were not degraded by RA patients' plasma. alpha 2M from RA patients' plasma attached to a solid-phase immunoabsorbent degraded an Arg-containing tripeptide much better than did the alpha 2M from normal donors, from patients with systemic lupus erythematosus, or from those with joint inflammation of other, "non-autoimmune" origin. Although the enzyme associated with alpha 2M in the plasma from RA patients appeared to be similar to trypsin, the differences in optimal pH, cation concentration, degradation of Lys-containing substrates, and biological activity suggest otherwise. We speculate that the alpha 2M-protease complexes are generated in the immune system and contribute to the inflammatory and autoimmune phenomena in RA.

Arthritis, Rheumatoid↗
PubMed1983

The mechanism of natural binding of bacteria to human lymphocyte subpopulations.

Some bacteria bind naturally to human lymphocyte subpopulations. Here we investigated the nature of the interacting factors" on bacterial cells and on lymphocytes. First, surface Ig was not involved in the binding of B. melitensis to B cells, since capping of surface Ig did not affect the binding of this bacteria. We put forward the hypothesis that the binding is due to an interaction between a protein (probably a lectin) on the lymphocyte surface and a carbohydrate on the bacteria. The following observations were consistent with this hypothesis: (1) the binding of B. melitensis to B cells was prevented by alpha-methyl-D-mannoside (alpha-MM) but not by other sugars; (2) the binding of B. melitensis to B cells was prevented by pretreatment of the peripheral blood lymphocytes (PBL) with 257 mM of alpha-MM but pretreatment of the bacteria had no effect; (2) an Escherichia coli mutant (strain 2023) which binds to B cells and part of the T cells was also agglutinated by Concanavalin A (Con A) but the nonbinding parental strain was not; the binding of this mutant to B cells was also inhibited by 257 mM of alpha-MM; (4) bacteria that bind to human lymphocytes were agglutinated at high titers by various plant lectins while those that do not bind were not agglutinated or were agglutinated only at low titers; (5) bacteria that bind to B cells as well as those that bind to B and T cells all were agglutinated by Con A, Lens culinaris agglutinin and Pisum sativum agglutinin, whose carbohydrate specificities were alpha-D-mannosyl- and alpha-D-glucosyl-residues; (6) the "receptors" on lymphocytes but not those on bacteria were sensitive to pronase digestion; and (7) bacteria still bound after being heated at 121 degrees C or being fixed with formaldehyde. Thus, we suggest that one of the mechanisms by which bacteria bind to lymphocytes may be by the interaction between a lectin on the lymphocyte and a carbohydrate on the bacteria.

Agglutination↗
PubMed1983

The use of mutants of Escherichia coli for the identification of human lymphocyte subpopulations in blood smears.

In previous work we have shown that some bacteria can bind to human lymphocytes and can be used to identify lymphocyte subpopulations in conventionally stained blood smears. These bacteria are of different species or genera, which makes it difficult to study the binding mechanism. Also, the main marker for B cells, Brucella melitensis, is of very small size and highly pathogenic. Here we show that B cells as well as some of the T cell subpopulations can be identified by different mutants obtained from a strain of an Escherichia coli. Two procedures were used to generate mutants. First, E. coli-YS57 (pro-his-trp-) was mutagenized with N-methyl-N'-nitro-N-nitrosoguanidine and the binding to mouse spleen cells was used as a selective pressure. Second, phage-resistant mutants of E. coli-YS57 were obtained and tested for the ability to bind to lymphocytes. Out of 10 strains selected by the former procedure, 5 bound to a significant number of human lymphocytes. All four phage-resistant mutants bound to human lymphocytes. Out of the total of nine mutants that bound to lymphocytes, six bound consistently, i.e., they bound to similar percentages of peripheral blood lymphocytes from different normal donors. One phage-resistant mutant, E. coli USC-106, bound only to B cells. The subpopulations of lymphocytes identified by the mutants were essentially the same as those identified by different species or genera of bacteria. We concluded that E. coli mutants can be obtained that identify human lymphocyte subpopulations and that one of these mutants recognizes B cells; these mutants may be used to study the nature of the receptors for bacteria on lymphocytes, which appear to have a lectin-like nature.

Animals↗
PubMed1982

The effect of protease inhibitors on the polyclonal B cell activator from the serum of patients with rheumatoid arthritis.

It has been reported that polyclonal B cell stimulation results in formation of autoantibodies and immune complexes. We have previously reported that a polyclonal B cell activator (PBA) associated with alpha 2-macroglobulin (alpha 2M) is present in the serum of patients with rheumatoid arthritis and related diseases. Here we studied the possibility that patient alpha 2M (Pt-alpha 2M) carries a trypsin-like protease responsible for the PBA activity. This activity was determined by the Ig-turnover assay developed in our laboratories. The small molecular weight protease inhibitors, aprotinin (Trasylol, Bayer) and phenylmethylsulfonylfluoride (PMSF), and the large molecular weight soybean trypsin inhibitor (SBTI) were used. These inhibitors did not affect the PBA activity of dextran sulfate of LPS. However, as expected, trypsin had a PBA-activity which was blocked by all of the above mentioned inhibitors. A trypsin-normal alpha 2M complex (Tr-N alpha 2M) and PBA activity which was inhibited by PMSF or aprotinin but not by SBTI. The PBA associated with Pt-alpha 2M was also inhibited by PMSF or aprotinin but not by SBTI. Moreover, the Tr-N alpha 2M complex and the Pt-alpha 2M, but not that from normal donors, had esterase activity for p-toluenesulfonyl-L-argininemethyl ester. These data suggest a similarity between the Pt-alpha 2M and Tr-N alpha 2M complex. Thus, we concluded that the esterolytic activity is sufficient for PBA activity, that Pt-alpha 2M has esterolytic activity and that this PBA activity can be blocked by small molecular weight protease inhibitors.

Animals↗
PubMed1981

Enumeration of lymphocytes and their subpopulations identified by bacterial adherence in blood smears of patients with breast tumors.

Peripheral blood lymphocyte counts were determined in 57 breast clinic patients. These patients were grouped into four clinical/pathological groups: 18 had benign breast disease, 13 had a history of breast cancer but were free of disease at the time of the study, 7 had a history of breast cancer and were free of disease at the time of the study but were on adjuvant chemotherapy, and 19 had active metastatic breast cancer. Two parameters were investigated in a double blind study: 1) the absolute lymphocyte counts and 2) the percentages of lymphocytes that bound different bacteria as markers of lymphocyte subpopulations in conventionally stained blood smears. A significant reduction in mean lymphocyte counts was demonstrated in patients with advanced disease. A significant increase in these counts was found in patients who were free of disease following surgical treatment alone. The T1T2 cells (our denomination), the subpopulation of cells responsible for specific killing in vitro and for suppression of natural cytotoxic cells, was significantly reduced in patients with advanced disease under treatment. The T/B cell ratio in all groups was the same and within normal range. These observations suggest that the observed decrease in the total lymphocytes is due not only to an overall decrease in all classes of lymphocytes but especially in those responsible for specific cell-mediated reactions. They also show that bacteria can be used as reliable reagents for the identification of lymphocyte subpopulations in blood smears.

Adult↗