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Biomedical subjects

O Stutman

Publications and source records attributed to O Stutman.

At least 55 records · Page 3Linked to original sources
PubMed1983

Characterization of IL-2-dependent cytotoxic T-cell clones. III. Inhibition of killing activity by monosaccharides.

The effects of monosaccharides on the cytotoxic activity of cytotoxic T lymphocytes (CTL) and three cloned long-term cytotoxic T-lymphocyte lines (CTLL) are compared. Uncultured CTL and clones CTLL-A2 and CTLL-A11 were derived from the peritoneal cavity of C57BL/6 mice immunized against the H-2Dd determinants on the BALB/c sarcoma Meth A. Clone CTLL-R5 was derived from spleen of (BALB/c X C57BL)F1 mice immunized against a unique determinant on the BALB/c radiation-induced leukemia RL male 1. The cell-surface phenotype of the clones is Lyt-1+,2+,3+. Cytotoxic activity of CTLL-A2 and CTLL-R5 as determined by a 4-hr 51Cr-release assay was inhibited over 50% by 1 mM 2-deoxy-D-glucose. CTLL-A11 and the uncultured cytotoxic T cells were more resistant to inhibition by 2DG (40% at 20 mM). Surprisingly, it was found that the addition of D-mannose, D-galactose, D-glucose, L-fucose, alpha-methyl-D-mannose, and N-acetyl-D-glucosamine also inhibited, in a dose-related manner, the cytotoxicity of CTLL-A2 and CTLL-A11. CTLL-R5 showed a more restricted inhibition pattern: only D-mannose and D-galactose were inhibitory. The mechanism of inhibition remains to be clarified.

Animals↗
PubMed1983

Ia dependent lymphokine production in the syngeneic mixed lymphocyte response.

We have shown that Interleukin 2 (IL-2) is produced in the murine syngeneic mixed lymphocyte response (SMLR) by T cells from both young thymus and adult spleen responder populations. We extend these findings in this report by showing that: 1) IL-2 production is the function of an Lyt 1+, Lyt-2-, Qa-2,3+, Qa-5- and Ia- T cell; 2) Similar to IL-2 elicited following alloantigen or mitogen stimulation, it peaks at day 2-3 of culture in both thymic and splenic SMLR; 3) The presence of the SMLR/IL-2-producing cell in thymus is a transient event, disappearing by 4 weeks of age; 4) The production of cytotoxic T lymphocyte (CTL) helper factor activity in primary SMLR is under the same Ia control as is IL-2 production and 5) That significant levels of interferon (IFN) are produced in the splenic SMLR. These findings support our contention that the SMLR represents an in vitro model of an in vivo mechanism for the production of IL-2, and that such a mechanism may play a role in T cell differentiation.

Age Factors↗
PubMed1983

Characterization of T lymphocyte subsets with monoclonal antibodies: discovery of a distinct marker, Ly-m22, of T suppressor cells.

The study of cell surface antigens has accelerated in the few years since the advent of hybridoma technology to the point where many dozens of such markers have now been described. The functional heterogeneity of post-thymic T cells in the immune response, however, still exhibit complexities beyond the resolving power of our current repertoire of marker antigens. In this study, we investigated the surface phenotype of three types of effector T cells: helper cells, nonspecific suppressor cells, and cytotoxic T cells, using four recently discovered alloantigen systems of T cells: Ly-m10, Ly-m18, Ly-m19, and Ly-m22. SRBC-primed spleen cells were used as a source of specific helper T cells, and they were tested by their ability to promote antibody synthesis by B lymphocytes. Concanavalin A-activated suppressor cells were assayed by their ability to inhibit that response. Cytotoxic T cells were activated by alloantigen. We found Ly-m10 to be expressed on all three cell types tested, whereas Ly-m18 and Ly-m19 were absent from all three. Ly-m22, whose controlling locus is closely linked to the Mls region, had a unique distribution, being present solely on suppressor cells. Thus Ly-m22 emerged as a new marker that distinguishes nonspecific T suppressor cells (Ly-m22+) from T helper as well as cytotoxic T cells (Ly-m22-). Ly-m22 is the only antigen besides I-J so far known to be restricted to suppressor cells.

Animals↗
PubMed1983

Limiting dilution analysis of T helper cell heterogeneity: a single class of T cell makes both IL 2 and IL 3.

We have previously described a limiting dilution assay (LDA) for estimating the frequency, in the spleens of unprimed mice, of alloantigen-responsive cells that, together with their immediate clonal progeny, can produce interleukin 2 (IL 2) in short-term culture. In this paper, we provide further evidence that the limiting cell in these cultures is, in fact, the immediate precursor of the IL 2-producing cell rather than merely a participant in a multicellular cascade that ultimately leads to production of this lymphokine. We also demonstrate the usefulness of the LDA method for estimating the frequency of the limiting cell for production of IL 3. By analyzing the supernatants of these short-term microcultures for both IL 2 and IL 3, we show that essentially all wells that produce IL 3 also produce IL 2. Furthermore, the amount of IL 2 produced in any individual well correlates strongly with the IL 3 production in the same well. Our data suggest that both these lymphokines are produced by a single class of T cell, and further that the regulatory events that control IL 2 generation in allostimulated helper cells act to control IL 3 levels in parallel.

Animals↗
PubMed1983

Multiple lymphokine production by a phorbol ester-stimulated mouse thymoma: relationship to cell cycle events.

Interleukin 2 (IL-2) production was studied in a subclone of the murine thymoma EL 4. Phenotypic characterization revealed the EL 4-17-2 line to be Thy-1.2+, Lyt-1.2+, and Lyt-2.2-. Costimulation with 500 ng 12-O-tetradecanoylphorbol 13-acetate (TPA)/ml and 5 micrograms concanavalin A (Con A)/ml induced optimal levels of IL-2. Three related phorbol esters stimulated comparable levels of IL-2 when used in conjunction with Con A. Kinetic experiments indicated that IL-2 first became detectable at 2 hours in TPA-treated cultures, whereas in cultures stimulated with Con A alone IL-2 production was not evident until 8 hours. Flow cytometry indicated that TPA and its related phorbol esters cause a perturbation in the cycling of the cell which may be related to increased IL-2 production. Under the conditions examined, no interferon-gamma (IFN-gamma) was detectable. Conversely, both granulocyte-macrophage colony-stimulating factor (CSF-GM) and interleukin-3 (IL-3) were found under conditions that led to stimulation of IL-2 synthesis. CSF-GM was produced in cultures treated singly with 500 ng TPA/ml or with Con A. IL-3 production was similar to IL-2 production, because optimal levels were found in cultures after combined treatment with phorbol ester and mitogen.

Animals↗
PubMed1982

Suppressor T cells activated in a primary in vitro response to non-major histocompatibility alloantigens.

Normal mouse spleen cells are not capable of mounting a primary cytotoxic T lymphocyte (Tc) response to non-H-2 alloantigens in vitro, although a good secondary H-2-restricted response is observable after in vivo immunization of the responder animals. Suppressor cells are generated in such a primary responses provided a Mls incompatibility exists between the responder and stimulator. These suppressors are not antigen specific, are Thy-1+, Lyt-1+, 2-, I-J-, and are highly radiosensitive. The suppressor cell precursors in normal spleen express the same phenotype. These suppressor cells are probably implicated in the lack of a primary Tc response in a primary mixed lymphocyte reaction across non-H-2 incompatibilities that include an Mls difference.

Animals↗
PubMed1982

Natural cytotoxic cells against solid tumors in mice. IV. Natural cytotoxic (NC) cells are not activated natural killer (NK) cells.

Natural cell-mediated cytotoxicity (NCMC), measured against a variety of tumors, is mediated by at least two sub-populations of effector cells: natural cytotoxic (NC) and natural killer (NK). The studies described in this report show that target lysis by NC cells requires a prolonged (18- to 24 h) assay period, whereas NC-susceptible targets can be lysed in short-term (4h) [3H]-proline assays using allo-sensitized CTL or mitogen-activated cytotoxic populations. NC cells are not "activated" during the long-term assay as indicated by: (1) demonstrating that lysis of NC-susceptible targets in long-term (20 h) 51Cr assays is still the function of a Qa-5- effector cell (NC) and (2) the fact that preincubation of the NC effector cell with susceptible targets for 18 h did not result in the activation of an NK-like population (kinetics of target lysis were comparable to those noted with fresh NC cell preparations). We show that NC cell activity is preserved in both the beige mutant and the PL/J mouse strains, both of which exhibit low NK cell activity, even in long-term assays. These combined studies support the view that NC and NK cell activities are the function of distinct cell types and not the property of a single cell class under different states of activation.

Animals↗
PubMed1982

Primary in vitro cytotoxic T cell response to non-major histocompatibility complex alloantigens in normal mice.

We have shown for the first time that it is possible to consistently generate a primary in vitro cytotoxic T cell (Tc) response to non-major histocompatibility complex alloantigens using responder cells from a normal mouse strain. This was achieved by carrying out, in the generating phase, a limiting dilution procedure in which it appears that suppressor cells that inhibit Tc activation or expansion are too dilute to manifest their effect. Moreover, the response was observed in mouse serum-(MS) as well as fetal calf serum- (FCS) supplemented media, an important finding in the light of the anomalous nonspecific effects induced by FCS. The cytotoxic response produced in MS-supplemented media was shown to be highly specific in both the generating and effector phases, whereas the responses in FCS had a strong nonspecific component.

Animals↗
PubMed1982

Limiting dilution analysis of IL-2 production: studies of age, genotype, and regulatory interactions.

We have previously described a limiting dilution method for estimating both the frequency of mouse T helper cells that can respond to alloantigens by producing Interleukin-2 (IL-2) and the amount of IL-2 produced by each responding cell (together with its progeny). At these very low cell concentrations, the amount of IL-2 produced per cell is remarkably higher (30-300 X) than the amounts generated under conventional, high density, culture conditions. We show here that cells cultured at intermediate densities "regulate" the amounts of IL-2 produced, so that the IL-2 concentration becomes independent of helper cell dose. The much higher level of IL-2 produced at very low concentrations therefore probably represents the loss, by dilution, of a regulatory cell that is less frequent than the IL-2 producing cells themselves. We show also how limiting dilution methods can be applied to KLH-responsive IL-2 secreting cells, and to questions about the effects of genotype and age on IL-2 production (Key words: Interleukin-2, helper T cells, aging, suppressor cells).

Age Factors↗
PubMed1982

Ia-dependent interleukin 2 production in syngeneic cellular interactions.

In the murine syngeneic mixed lymphocyte response (SMLR), the co-culture of nylon wool-nonadherent spleen cells or young (4 to 6-day-old) thymocytes with syngeneic nylon wool-adherent adult spleen cells, results in proliferation and Interleukin 2 (IL 2) production. In both the thymus and spleen SMLR, proliferation and IL 2 production are the function of Lyt-1+2-3- (Lyt-1) responder cells and require the expression of IA (IB) and/or IE cell surface antigens on the stimulator cell. IL 2 production in both spleen and thymus SMLR is independent of heterologous serum factors (i.e., fetal calf serum), which, however, influences the magnitude of the in vitro proliferation. The IL 2-producing cell in spleen is capable of self-renewal in the absence of the thymus; it is detectable 6 to 7 mo after adult thymectomy. These results suggest the in vitro SMLR may measure a cellular interaction between the Lyt-1 subset of T cells and self Ia antigens on macrophage-like cells that produces IL 2 in an antigen-independent fashion. This interaction may have physiologic significance in the expression and regulation of T cell function.

Animals↗
PubMed1982

Enumeration of IL 2-secreting helper T cells by limiting dilution analysis, and demonstration of unexpectedly high levels of IL 2 production per responding cell.

We describe here a method for estimating the frequency of alloantigen-responsive precursors of interleukin 2-(IL 2) secreting mouse T helper cells in spleen cell populations, and for measuring the amount of IL 2 generated by the progeny of each responsive cell over a 5-day culture period. Depending on the particular stimulus, about 1 T cell in every 30 to 300 can generate detectable levels of IL 2. Mls-locus incompatibilities activate a higher fraction of cells than do differences at the H-2 complex. The responder cell is sensitive to anti-Thy-1.2 antibody and complement, and is highly enriched in Lyt-2- populations as compared to positively selected Lyt-2+ cells. The dilution curve obtained is consistent with the idea that IL 2 production depends on a single class of T cell. The amount of IL 2 produced in these limiting dilution cultures is unexpectedly high; about 30-fold greater than the levels predicted by extrapolation from conventional mixed lymphocyte cultures. These results suggest that conventional cultures may be very responsive to regulatory events that do not occur at very low responder cell concentrations. Thus, limiting dilution analyses will provide insights into the effects of drugs, age, subset manipulation, etc., on the IL 2-producing cell itself, information that cannot be provided by conventional methods alone.

Animals↗