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Biomedical subjects

D Redelman

Publications and source records attributed to D Redelman.

At least 37 records · Page 2Linked to original sources
PubMed1986

Characterization of Tritrichomonas foetus antigens, using bovine antiserum.

Tritrichomonas foetus antigens were identified, using the serum of an Angus heifer that had been repeatedly immunized with suspensions of 1 X 10(8) organisms in Freund's complete adjuvant. Antibody activity against T foetus was determined by dot-blot analysis, using horse-radish peroxidase-conjugated anti-bovine immunoglobulin to detect bound antibody. The antiserum contained antibodies against surface and flagellar components of live or fixed T foetus, as determined by use of immunofluorescence. The antiserum reacted with approximately 38 proteins in a pool of 55 to 60 components resolvable by polyacrylamide-gel electrophoresis of T foetus extracts.

Animals↗
PubMed1985

The role of xenoantigen in the human autologous mixed lymphocyte reaction: ability to respond in the absence of exposure to xenoantigen depends on the method of T-cell preparation.

Recent reports have suggested that the autologous mixed lymphocyte reaction (AMLR) is attributable to contaminating xenoantigen. T-cell populations separated from peripheral blood mononuclear cells (PBM) over Percoll fail to produce an AMLR in the absence of xenoantigen. However, T-cell populations that do not adhere to plastic and to nylon wool will in the absence of added xenoantigen produce an AMLR comparable to that obtained with E-rosetted populations. In the nonadherent cells separated sequentially by passage through nylon wool and Percoll, the presence of two lymphocyte subpopulations was demonstrated. One is xenoantigen dependent, the other xenoantigen independent in the AMLR. Further, the xenoantigen-independent population may provide help to the dependent population. In the previous reports with Percoll-separated cells that attributed the AMLR to contaminating xenoantigen, the xenoantigen-independent T-cell population was excluded from the responding fraction.

Animals↗
PubMed1984

The mechanism of cell-mediated cytotoxicity. IV. K-76 COONa, which inhibits the activity of Factor I and of C5, inhibits early events in cytotoxic T-lymphocyte-mediated cytolysis and in T-lymphocyte activation.

K-76 COONa is a derivative of a fungal product which blocks complement (C)-mediated lysis by combining with C5 and preventing its activation to C5b. K-76 COONa can also combine with Factor I and inhibit its ability to hydrolyze C3b to iC3b. The inclusion of K-76 COONa at concentrations similar to those which inhibit C lysis blocked both murine cytotoxic-T-lymphocyte (CTL)-mediated lysis (CML) and the lectin-stimulated proliferative response of murine and human T lymphocytes. A modified cation pulse procedure has been used to determine which phases of CML were most sensitive to the drug. K-76 COONa was inhibitory when it was added to CML prior to the early Mg+2-dependent binding phase, but was much less effective when it was added at any time after the formation of CTL-target conjugates. The principal effect of the drug on the proliferative response was also exerted during an early phase of the response. K-76 COONa did not appreciably decrease the production of T-cell growth factor (TCGF), but it did inhibit the induction of TCGF receptor expression by both functional criteria, i.e., induction of responsiveness to TCGF, and by morphological criteria, i.e., the expression of the Tac antigen. Later events, such as the TCGF-dependent proliferation of cycling T cells, were less sensitive to the drug. Evidence is discussed suggesting that molecules similar to Factor I and to C3 may be involved both in the early events of CML and of T-lymphocyte activation.

Cells, Cultured↗
PubMed1984

Deoxyadenosine modulates human suppressor T cell function and B cell differentiation stimulated by Staphylococcus aureus protein A.

Adenosine deaminase (ADA) deficiency and the resultant accumulation of deoxyadenosine (AdR) are associated with profound T cell dysfunction and variable B cell dysfunction in vivo. We examined the effects of AdR on the in vitro function of normal human peripheral blood B and T lymphocytes whose ADA activity was inhibited by 2'-deoxycoformycin. We found that OKT8+ T cell-mediated suppression of SPA-induced Ig production was markedly reduced by concentrations of AdR (3 to 10 microM) that did not affect helper T cell function. Because the lectin-induced proliferative responses of OKT8+ T cells and OKT8- T cells were equally susceptible to AdR, modulation of in vitro immune responses by low-dose AdR probably reflected different proliferative requirements for the expression of T cell helper or suppressor functions. Although low doses of AdR did not inhibit Ig production in SPA-stimulated cultures, we found that T cell-dependent, SPA-stimulated B cell proliferation was blocked by 3 to 10 microM AdR. Therefore, it appeared that B cell proliferation was not required for the induction of Ig synthesis in this system. Higher doses (30 to 100 microM) of AdR did block the induction of Ig synthesis, presumably by interfering with T-helper functions via a mechanism other than inhibition of proliferation and/or by inhibiting B cell differentiation events.

Adult↗
PubMed1984

The requirement for proteinase activity for human lymphocyte-mediated natural cytotoxicity (NK): evidence that the proteinase is serine dependent and has aromatic amino acid specificity of cleavage.

We used reagents specific for serine-dependent proteinases to verify that a proteinase of this class is necessary for natural cytotoxicity (NK). NK was inhibited by phenylmethylsulfonylfluoride (PMSF), by diisopropylfluorophosphate (DFP), and by the plasma antiproteinase alpha-1-antichymotrypsin (alpha-1-X), all of which are specific for serine-dependent proteinases. Substrate specificity was then determined on the basis of the specificity of the plasma and fungal anti-proteinases and synthetic alternate substrates that affected NK. alpha-1-X, which inhibits only serine proteinases with aromatic amino acid specificity, blocked NK. Chymostatin, but not other fungal inhibitors, also blocked NK activity. Furthermore, the only synthetic substrates that effectively reduced NK were those derived from aromatic amino acids. The ester derivatives of these substrates inhibited NK better than the amides. NK inhibition with these alternate substrates was also stereospecific, with the L forms twofold more active than the D forms. These reagents did not block initial lymphocyte-target cell binding. Therefore we propose that the "NK-proteinase" is involved in either the initiation of cytolysis, perhaps as part of stimulus and secretion of cytolytic molecules, or in the cascade of events that may lead to the formation of final lytic substance.

Amino Acids↗
PubMed1984

Deoxyadenosine (AdR) inhibition of newly activated lymphocytes: blockade at the G0-G1 interface.

The mechanism responsible for the lymphocytotoxicity associated with congenital adenosine deaminase (ADA) deficiency has been ascribed to an accumulation of dATP. Elevated levels of dATP can then lead to inhibition of DNA synthesis by inhibiting ribonucleotide reductase and causing a depletion of the other deoxynucleotide triphosphates (dNTP). This hypothesis was derived principally from studies with murine and human lymphoblastoid cell lines (LCL) and apparently confirmed in a limited number of investigations with lectin-stimulated lymphocytes. Our biochemical studies of lectin-stimulated mouse and human lymphocytes were not consistent with the dATP model and suggested that AdR exerted effects on lymphocyte activation that preceded the initiation of DNA synthesis. In the current studies, we focused on the effects of AdR on the early events in T lymphocyte activation, because we found they were the most sensitive to AdR toxicity. AdR blocked neither the production of T cell growth factor (TCGF) by lectin-stimulated lymphocytes nor the expression of TCGF receptors as detected by the anti-Tac monoclonal antibody that recognizes the human TCGF receptor. AdR did, however, block the early TCGF-dependent events leading to the entry into the cell cycle. By using the metachromatic fluorescence stain acridine orange, we found that AdR blocked the increased synthesis of RNA that characterizes the entry into the G1 phase of the cell cycle from the G0, resting state. Because these early effects were caused by the lowest doses of AdR, and because they preceded the synthesis of DNA by 15 to 20 hr, it suggested that these effects may be principally responsible for the in vivo toxicity associated with ADA deficiency. Furthermore, none of the other proposed biochemical mechanisms, e.g., inhibition of methylation, diminution of ATP levels, or incorporation of AdR into polyadenylated RNA, appeared adequate to explain AdR toxicity during T lymphocyte activation.

Animals↗
PubMed1984

Inhibition of human lymphocyte natural cytotoxicity and antibody-dependent cell-mediated cytotoxicity by K-76 COONa, a reagent that blocks complement activity.

K-76 COONa is a 440 m.w. fungal product that can inhibit complement activity of C5 and Factor I. K-76 COONa abrogated both human natural killer (NK) cell activity and antibody-dependent cell-mediated cytotoxicity (ADCC) (ID50 approximately 1.5 mM). To be effective, K-76 COONa had to be present during the assay, because pretreatment of lymphocytes with highly inhibitory concentrations of K-76 COONa did not inhibit cytolysis. The monocarboxylic K-76 derivative was more inhibitory to NK than the dicarboxylic derivative. This relative efficacy is similar to that observed for inhibition of complement lysis. K-76 COONa inhibited NK when added before NK conjugate formation, but had little effect when added after conjugate formation. The compound also inhibited the formation of conjugates by NK and K cells. Therefore, this reagent selectively affected events that occurred between the initial effector-target cell interactions and the formation of stable conjugates. It had little influence on the post-binding "lethal hit" stage of cytolysis. These data imply a) that if any molecules similar to C5 are activated during the "lethal hit" stage of cytolysis, then they are inaccessible to K-76 COONa , and b) that C3bi-like molecules may be involved in lymphocyte binding.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1983

The mechanism of cell-mediated cytotoxicity. III. Protease-specific inhibitors preferentially block later events in cytotoxic T lymphocyte-mediated lysis than do inhibitors of methylation or thiol-reactive agents.

Highly active mouse cytotoxic T lymphocytes (CTL) generated in secondary mixed-lymphocyte responses were used to examine the manner in which adenosine derivatives, thiol-specific reagents, or protease-specific probes affected CTL-mediated lysis (CML). The adenosine deaminase inhibitor deoxycoformycin (dCF) enhanced inhibition by adenosine (AR) or by deoxyadenosine (AdR), but not by 7-deazaadenosine (tubercidin). L-Homocysteinethiolactone (L-Hcy) acted synergistically with AR, but not with AdR or tubercidin, to block CML. Thus, AR derivatives may act both by affecting cellular methylation reactions, as demonstrated by the synergism between AR and L-Hcy, and by inhibiting other events required for CML. Conditions were then established to determine whether these reagents preferentially affected either the Ca2+-independent initial stage of cytolysis or the subsequent Ca2+-dependent events. Methylation inhibitors blocked lysis most effectively if added before effector-target binding. Similarly, the nonpenetrating thiol-specific reagent quaternary ammonium monobromobimane (qBBr) was more inhibitory when added prior to the Ca2+-dependent stage. Protease inhibitors such as alpha-1-antichymotrypsin and protease substrates such as acetyltyrosine ethyl ester (ATEE) or tyrosine ethyl ester (TEE) also inhibited CML. But, in contrast to qBBr or methylation inhibitors, neither TEE nor ATEE was more effective when added prior to the initial effector-target interaction. Furthermore, TEE did not appreciably affect CTL binding to target cells at concentrations that nearly abrogated CML. Thus, the implicated protease step is unique in that it does not appear to participate in recognition or binding.

Adenosine↗
PubMed1983

In vitro studies of the rabbit immune system. VIII. The production of rabbit T cell growth factor (TCGF) and its relationship to mouse and human TCGF.

Rabbit spleen cells (1 X 10(6)/ml) stimulated with Con A (2.5 micrograms/ml) begin producing T cell growth factor (TCGF) activity within 2-3 h at 37 degrees C and reach a plateau after 20-24 h. Lymphocytes from the mesenteric lymph nodes (MLN) also produce TCGF activity but require higher lectin and/or cell concentrations for optimal production. TCGF generation by MLN can be improved by including a small proportion of spleen cells (10-40%). Rabbit lymphocytes also produce TCGF in serum-free, protein-free medium, but the Con A concentration must be reduced to 0.5-1.25 micrograms/ml. Rabbit TCGF activity eluted from gel filtration columns at a volume equivalent to approximately 15,000 molecular weight, similar to human TCGF activity. However, when rabbit, mouse and human TCGFs were compared for their ability to support proliferation by activated cells from the 3 species, rabbit and mouse TCGFs appeared more similar. In all cases, the homologous combinations of cells and TCGF were most efficient. Human TCGF would support some proliferative activity in all 3 cell types but rabbit and mouse TCGFs failed to support the growth of human cells.

Animals↗
PubMed1983

In vitro studies of the rabbit immune system. IX. Extended culture of rabbit cytotoxic cells and the generation of TCGF-dependent T lymphocyte lines.

Rabbit T lymphocytes with cytotoxic activity for mouse target cells can be maintained in culture for several weeks by periodically restimulating with fresh mouse cells or by culturing with a source of TCGF. The former procedure greatly enriches the cultured cells for cytotoxic activity but has the disadvantage of introducing cellular debris. Cell growth and cytotoxic activity waned after 4-6 weeks using either procedure. However, T lymphocyte lines (TLL) could be isolated from limiting dilution cultures containing mitomycin C-treated mouse 'feeder' cells and a source of TCGF. These cells grew out with a frequency of one in 50-150 and approximately 15% of these growing cells had cytotoxic activity. Some cytotoxic cells lost their lytic activity after 4-5 months but others have persisted for over 7 months. Some TLL have now been growing for over 20 months.

Animals↗
PubMed1983

Studies of rheumatoid synovial fluid lymphocytes. II. A comparison of their behavior with blood mononuclear cells in the autologous mixed lymphocyte reaction and response to TCGF.

Synovial fluid lymphocytes (SFL) and peripheral blood lymphocytes (PBL) from patients with rheumatoid arthritis were compared for their response to lectin stimulation and for their behavior in the autologous mixed lymphocyte reaction (AMLR). The SFL proliferative response to phytohemagglutinin (PHA), as measured by tritiated thymidine incorporation at 72 hr, was lower than that of PBL (P less than 0.001). When T-cell growth factor (TCGF) was added to the medium, there was an increase in the SFL proliferative response to PHA (P less than 0.05). In contrast, TCGF did not alter significantly the PBL proliferative response to PHA. Mixing experiments were performed to determine whether the poor SFL proliferative response was due to passive absorption and removal of in situ-generated TCGF by "suppressor" cells. When cultured together, SFL did not suppress the PBL proliferative response to PHA, suggesting that decreased production of TCGF rather than competitive binding of TCGF results in the poor SFL proliferative response to lectin stimulation. In the AMLR, synovial fluid non-T cells were found to be more stimulatory to peripheral blood T cells than were peripheral blood non-T cells (P less than 0.001). In comparison to peripheral blood T cells, synovial fluid T cells were poor responders in the AMLR. Repetitive in vitro autologous stimulation of peripheral blood T cells resulted in proliferative responsiveness analogous to that of SFL, i.e., a relatively poor proliferative response in the AMLR and a poor response to PHA. The latter could be augmented by TCGF. The SFL requirement for exogenous TCGF is consistent with a state of immune activation. In vivo stimulation by non-T cells may play an important role in the immune activation which characterizes rheumatoid SFL.

Adult↗
PubMed1981

Inhibition of human natural cytotoxicity by macromolecular antiproteases.

Human natural cell-mediated cytotoxicity is inhibited by macromolecular protease inhibitors. Human plasma alpha 1 antiproteases are more effective than the plant antiproteases lima bean trypsin inhibitor and soybean trypsin inhibitor for reduction of cytotoxicity to the "slow" targets T24 human bladder carcinoma and NKI-1 melanoma. This inhibition of natural cytotoxicity is more readily demonstrable in serum-free medium containing crystalline bovine serum albumin than in medium containing fetal calf serum. Although electrophoretically homogeneous plasma alpha 1 antitrypsin inhibits natural cytotoxicity, partially purified alpha 1 antitrypsin preparations that contain several apha 1 proteins are more inhibitory at equivalent trypsin inhibitory capacities. Partially purified alpha 1 antichymotrypsin with no antitrypsin activity is an extremely potent inhibitor. Thus, it seems likely that several of the plasma antiproteases, including alpha 1 antitrypsin and alpha 1 antichymotrypsin, are capable of influencing natural cytotoxicity. These data indicate that serine-dependent proteases hae a critical role in triggering and/or effecting cell-mediated cytolysis. Furthermore, since alpha 1 antichymotrypsin and alpha 1 antitrypsin are acute phase proteins, the increase in plasma concentration or turnover rates of the proteins could influence natural killer cell activity in vivo.

Animals↗
PubMed1981

Active tumor cell resistance to human natural killer lymphocyte attack.

Inhibition of protein synthesis by cycloheximide, puromycin, or emetine increased tumor lysis mediated by human natural killer (NK) cells to "slow" but not "fast" tumor targets. Human T24 bladder carcinoma cells were used as slow targets which are killed after a approximately 3-hr lag period, and K562 cells derived from a patient with myelogenous leukemia were used as fast targets which are killed without a lag period. This enhancement of killing exceeded that which would have been expected simply from a reduction of tumor cell growth during the time of assay. Pretreatment of the NK effector cells and the tumor cells separately showed that the effect was on the tumor cells and not due to enhancement of NK cell activity. These observations imply that some tumor cells can actively resist NK attack. The discovery that some but not all human tumor cells actively resist cellular immune attack, perhaps by repair mechanisms dependent upon protein synthesis, provides a new model for evaluation of tumor cells in their resistance to host defenses.

Cell Line↗