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T Owens

Publications and source records attributed to T Owens.

At least 73 records · Page 4Linked to original sources
PubMed1991

Requirement for noncognate interaction with T cells for the activation of B cell immunoglobulin secretion by IL-2.

The mechanism whereby noncognate contact with activated IL-2-producing Type 1 helper T cells (TH1) induces B cell activation was examined. Small resting B cells from C57B1/6 mice were cultured, in the absence of any ligand for surface Ig, with irradiated cells of the hapten-specific, CBA-derived, F23.1+ TH1 clone E9.D4 in F23.1 (anti-T cell receptor V-beta 8)-coated microwells. This induced polyclonal B cell activation to enter cell cycle (thymidine incorporation) at 2 days and to secrete immunoglobulin at 5 days. An anti-IL-2 mAb (S4B6) inhibited antibody production completely. Anti-IL-2 did not inhibit either LPS-induced B cell responses, or T cell activation (measured as IL-3 secretion). Anti-IL-2 receptor (anti-Tac) mAbs also inhibited T-dependent B cell responses, without affecting LPS responses. An anti-IFN-gamma mAb partially inhibited Ig secretion, without affecting entry into cycle. LPS responses or T cell activation. Other antibodies (anti-IL-3, IL-4, IL-5, Thy-1.2, CD5) were not inhibitory. After 2 days of culture with F23.1-activated T cells, B cells appeared to have become responsive to IL-2, in that they could be driven to immunoglobulin production by the addition of IL-2. Flow cytometry showed no expression by these B cells of 55-kDa (Tac) IL-2 receptors. Also, rigorous removal of T cells from 2-day cocultures prevented the response to IL-2, and readdition of T cells restored it. Because the reconstituted responses were inhibited both by anti-IL-2 and by anti-Tac, IL-2 must have acted indirectly, via the T cells that were present in these cultures. Continued contact with T cells was therefore necessary for the progression of B cells to antibody secretion.

Animals↗
PubMed1991

A role for adhesion molecules in contact-dependent T help for B cells.

The role of cell contact in T-dependent B cell activation was examined. Small resting B cells from C57BL/6 mice were cultured with CBA-derived, non-alloreactive cloned T helper cells in anti-T cell receptor V beta 8-coated microwells. This induced polyclonal B cell activation to enter cell cycle (as measured by thymidine incorporation at 2 days) and to secrete immunoglobulin (as measured by an enzyme-linked immunoassay detecting high-rate Ig secretion at 5 days). The inclusion of monoclonal antibodies against LFA-1. ICAM-1 and CD4 in these cultures strongly inhibited antibody responses, although proliferative responses were only inhibited to about 50%. Inhibitory monoclonal antibodies did not significantly affect lipopolysaccharide-induced responses. T cell activation to interleukin (IL) 3 secretion, nor did they inhibit the formation of multicellular clusters containing T and B cells. There was no correlation between the level of expression of adhesion molecules by T cells and their ability to induce B cell responses. Anti-LFA-1 abrogated T-dependent responses to IL2 which were inducible after 2 days in culture, but did not inhibit the induction of this IL2 responsiveness. These results suggest that continued cell contact involving adhesion/accessory molecules induces B cells to proliferate and to respond to T cell lymphokines. A signaling role for cell interaction molecules on B cells is proposed, similar to the role of these and analogous molecules on T cells.

Animals↗
PubMed1989

Detection of colony-stimulating factor messenger RNA in single T cells by in situ hybridization.

In situ hybridization has been used to study the accumulation of colony-stimulating factor (CSF) mRNA in single cells of a T lymphocyte clone (E9.D4) following antibody-mediated (F23.1) activation via the Ti-T3 complex in filler-independent bulk cultures. The specificity of hybridization for cellular RNA was demonstrated by pretreating the cells with the Ca2+-dependent enzyme micrococcal nuclease by using a novel protocol developed for use with riboprobes. Maximal levels of granulocyte-macrophage (GM) and multipotential-CSF (interleukin 3) mRNA were detected after 8-10 h, with GM-CSF mRNA being detected earlier and at a lower concentration of stimulus. The rise in intracellular mRNA was accompanied by an increase in the corresponding CSF bioactivity in the supernatant. In situ hybridization was of comparable sensitivity to Northern blot analysis and revealed significant heterogeneity in the accumulation of CSF mRNA within individual cells of the clone following stimulation with F23.1. This could account for the corresponding heterogeneity in CSF production by single cells. Under optimal conditions at least 25% of cells contained both transcripts. The method has been used to examine CSF production by normal spleen cells and should be useful in the further analysis of lymphokine gene regulation in single T cells.

Animals↗
PubMed1989

The cell biology of T-dependent B cell activation.

The requirement that CD4+ helper T cells recognize antigen in association with class II Major Histocompatibility Complex (MHC) encoded molecules constrains T cells to activation through intercellular interaction. The cell biology of the interactions between CD4+ T cells and antigen-presenting cells includes multipoint intermolecular interactions that probably involve aggregation of both polymorphic and monomorphic T cell surface molecules. Such aggregations have been shown in vitro to markedly enhance and, in some cases, induce T cell activation. The production of T-derived lymphokines that have been implicated in B cell activation is dependent on the T cell receptor for antigen and its associated CD3 signalling complex. T-dependent help for B cell activation is therefore similarly MHC-restricted and involves T-B intercellular interaction. Recent reports that describe antigen-independent B cell activation through coculture with T cells activated by anti-T-cell receptor or anti-CD3 antibodies suggest that cellular interaction with T cells, independent of antigen presentation or lymphokine secretion, induces or triggers B cells to become responsive to T-derived lymphokines, and that this may be an integral component of the physiological, antigen- and MHC-restricted T-dependent B cell activation that leads to antibody production.

Animals↗
PubMed1988

The role of CD4 in antigen-independent activation of isolated single T lymphocytes.

The membrane molecule CD4 (L3T4) is thought to facilitate activation of Class II H-2-restricted T cells by binding to Ia determinants on antigen-presenting cells. Recent reports suggest that CD4 can also contribute to antigen-independent activation by anti-T cell receptor (TCR) antibodies. An assay which measures the secretion of two lymphokines, granulocyte-macrophage colony-stimulating factor and interleukin 3 (IL-3), by single T cells activated with an anti-TCR antibody, F23.1, was used to analyze the effects of anti-CD4 antibodies on antigen-independent T cell activation. Single cells of a CD4+F23.1+ clone were micromanipulated into wells to which F23.1 had been immobilized, and their lymphokine secretion was measured 24 hr later. The frequency of lymphokine-secreting cells was consistently reduced up to 10-fold in the presence of soluble anti-CD4 antibody (GK1.5) but only up to 2.5-fold by an antibody to the cell adhesion molecule, LFA-1. In both bulk and single-cell cultures, responses to suboptimal concentrations of F23.1 were more susceptible to inhibition by GK1.5 than responses to optimal F23.1. The failure of GK1.5 to inhibit IL-2-stimulated lymphokine synthesis in bulk cultures suggested that CD4 ligation did not deliver a negative signal to the clone. By contrast, when either anti-CD4 or anti-LFA-1 was immobilized on the same surface as F23.1, the frequency of lymphokine-secreting cells could be increased up to 10-fold. It is concluded that anti-CD4 antibodies can act directly on the responding T cell to affect TCR-dependent activation, in the absence of interaction with antigen-presenting cells or any other cell type.

Antibodies, Monoclonal↗
PubMed1988

T cell antigen receptor expression by subsets of Ly-2-L3T4- (CD8-CD4-) thymocytes.

The V beta 8-specific mAb F23.1 and KJ16 were used as fluorescent stains to test for TCR expression on the surface of subpopulations of early, CD4-CD8- (L3T4-Ly-2-) thymocytes from adult CBA mice. A surprisingly high proportion (27%) of Ly-2-L3T4- thymocytes were strongly F23.1 and KJ16 positive. No positive cells were detected among Ly-2-L3T4- thymocytes from V beta 8-negative SJL mice. In contrast to the adult thymus, Ly-2-L3T4- cells from embryonic CBA thymus lacked F23.1-positive cells. Subsets of adult CBA Ly-2-L3T4- thymocytes were separated to determine which expressed V beta 8. The major subset, Ly-1 low B2A2-M1/69+Thy-1+Pgp-1-, representing a phenotype similar to embryonic Ly-2-L3T4- thymocytes and the phenotype commonly isolated from adult thymocytes as Ly-1 "dull," lacked cells strongly positive for F23.1. In contrast, a series of subsets of adult CBA Ly-2-L3T4- thymocytes which were B2A2-M1/69- and Pgp-1+ all included strongly F23.1-positive cells. A minor subset, negative for most markers except Pgp-1 and presumed on the basis of this phenotype and some reconstitution studies to include the earliest intrathymic precursors, contained 28% F23.1-positive cells. However, no F.23.1-positive cells were detected in equivalent "prethymic" populations from bone marrow or from athymic mouse spleen. The subsets of Ly-2-L3T4- thymocytes which were Ly-1 high, B2A2-M1/69-, and Pgp-1+ all contained about 70% F23.1-positive cells, indicating a V beta 8 usage much higher than the mature T cell average. These results indicate that a series of distinct developmental events have occurred within these CD4-CD8- thymocytes previously considered as a single group of early precursor cells, and that some aspects of repertoire selection may be occurring amongst thymocytes which lack CD4 or CD8.

Aging↗
PubMed1988

Production of two hemopoietic growth factors is differentially regulated in single T lymphocytes activated with an anti-T cell receptor antibody.

A method has been developed to measure the production by single activated T lymphocytes of two hemopoietic growth factors, granulocyte-macrophage CSF (GM-CSF) and multipotential CSF (multi-CSF or IL-3). When individual cells of the L3T4 (CD4)+ F23.1+ T cell clone E9.D4 were transferred by micromanipulation into wells coated with the monoclonal anti-T cell receptor antibody F23.1, up to 90% of cells produced CSF as detected by CSF-dependent hemopoietic cell lines. Production occurred in the absence of proliferation and did not require the addition of accessory cells or IL-2. Both the frequency of CSF-producing cells and the average production per positive cell depended on the density of the immobilized stimulating ligand, indicating that the response of each cell is not an all-or-none phenomenon but varies with the strength of stimulation. Individual cells of the clone varied over a 100-fold range in their total CSF titer with a mean value of about 0.05 U/cell. They also varied in their relative production of GM-CSF and multi-CSF. Thus, low producing cells secreted only GM-CSF whereas high producing cells also secreted multi-CSF. The failure of low producing cells to secrete multi-CSF was not genetically based since such cells could give rise to progeny that synthesized multi-CSF. These results suggest that the synthesis of these two lymphokines can be differentially regulated at the level of the single cell.

Antibodies, Monoclonal↗
PubMed1988

A noncognate interaction with anti-receptor antibody-activated helper T cells induces small resting murine B cells to proliferate and to secrete antibody.

Culture of small resting allogeneic B cells (of an irrelevant haplotype) with two clones of T helper (Th) cells that were activated by the F23.1 anti-T cell receptor antibody led to the activation of B cells to proliferate and to secrete antibody. Th cell supernatants by themselves had no effect on resting B cells (even in the presence of intact F23.1 antibody), but could induce antibody secretion by anti-Ig-preactivated B cells. Both F23.1+ clones (E9.D4 and 4.35F2) and one F23.1- clone (D2.2) could synergize with supernatants from activated E9.D4 T cells to induce B cell activation. F(ab')2 fragments of F23.1 induced E9.D4 to activate B cells as efficiently as intact F23.1 and B cell populations that had been incubated with F23.1 were not activated when cultured with E9.D4, although T cells recognized cell-presented F23.1 and were weakly activated. Reduction of the density of F23.1 adsorbed to plastic resulted in weak T cell activation, and these T cells did not induce B cell responses. Haptenated B cell populations, although recognized by E9.D4, were not activated. Separation of T and B cells by a 0.4-micron membrane prevented T-dependent B cell activation, although Th cell-derived B cell-activating lymphokines would be assayed across these membranes. These results suggest a polyclonal noncognate B cell activation that depends on physical contact between B cells and activated T cells. The requirement for a cognate interaction of Th with B cells for the production and delivery of B help can therefore be overcome by activating Th cells with high densities of T cell receptor ligands.

Animals↗
PubMed1988

A significant proportion of normal resting B cells are induced to secrete immunoglobulin through contact with anti-receptor antibody-activated helper T cells in clonal cultures.

This report describes single-cell techniques to address the nature of a cellular interaction in which activated T lymphocytes stimulate small resting B cells to develop into antibody-forming cell clones in the absence of any surface immunoglobulin ligand or an antigen bridge. The cloned T helper cell line E9.D4 was stimulated with the anti-V beta 8 antibody F23.1 bound to the plastic of Terasaki 10-ul culture wells. When an excess of T helper lymphocytes was used (1,000 X-irradiated or 600 unirradiated, stimulated E9.D4 cells), 10-25% of B cells responded by antibody formation as judged by an enzyme-linked immunosorbent assay performed after 5 days of culture. When one of a very small number of B cells were present, the rate-limiting step to antibody-forming cell formation was the number of T cells present. Far fewer T cells sufficed for stimulation when culture trays were tilted to force T and B cells into proximity at the sulcus formed at the bottom edge of the culture wells. When T cell numbers were limiting, unirradiated T cells out-performed irradiated T cells. Some cell clones held for 7 days switched to IgG antibody production. E9.D4 supernatants were virtually ineffective in causing B cell stimulation, even when 3T3 filler cells were added to support cultures. The results suggest that cell contact, and perhaps conjugate formation, with a strongly activated T cell can cause changes in the adjacent resting B cells akin to those of Ig receptor cross-linking, following which a lymphokine flux (even one not involving IL 4 and 5) promotes antibody-forming cell development.

Animals↗
PubMed1987

Participation of L3T4 in T cell activation in the absence of class II major histocompatibility complex antigens. Inhibition by anti-L3T4 antibodies is a function both of epitope density and mode of presentation of anti-receptor antibody.

The recognition of many class II major histocompatibility complex (MHC)-associated antigens by T cells requires the participation of the L3T4 molecule. It has been proposed that this molecule acts to stabilize low affinity binding to antigen in association with MHC and thereby increases the avidity of T cell/antigen interactions. By using antibodies against the T cell antigen receptor (TCR) to activate T cells, thereby circumventing the requirement for antigen presenting cells and MHC-associated antigen, we have been able to study the function of L3T4 in the absence of class II MHC. We have used two monoclonal antibodies, KJ16-133.18 and F23.1, that recognize a determinant encoded by the T cell receptor V beta 8 gene family. These antibodies were used to select two clones of T cells with surface phenotype Thy-1.2+, L3T4+, Lyt-2-, KJ16-133.18+, F23.1+, IA-, IE-. One of these clones (E9.D4) was hapten-specific (anti-ABA + Iak), the other (4.35F2) was alloreactive (anti-Iak). Activation of these clones by antigen, concanavalin A (Con A) or by the F23.1 antibody was studied by assaying the production of interleukin 3 (IL 3). Both soluble and solid phase-coupled F23.1 induced T cell activation in the complete absence of class II MHC, immobilized antibody (either Sepharose-coupled or plastic-adsorbed) being more effective. The induction of IL 3 production by suboptimal doses of either Con A or plastic-adsorbed F23.1 was inhibited by the anti-L3T4 antibody GK1.5, as was the response to F23.1 coupled to Sepharose-4B beads. However, the responses to optimal or superoptimal doses of these stimuli were not inhibited. In contrast, weak responses to non-TCR cross-linking stimuli such as phorbol myristate acetate (PMA) or low concentrations of soluble F23.1 were not inhibited by GK1.5 (the latter response was usually slightly enhanced). These results show that anti-L3T4 antibodies are not inherently inhibitory, but require both ligation and cross-linking of the TCR for their effect. We propose a model whereby L3T4 interacts with the TCR during T cell activation. Anti-L3T4 antibodies sterically hinder the formation of TCR complexes and so prevent activation. However, by increasing the epitope density of the activating ligand, the avidity of the T cell/ligand interaction can be increased sufficiently to prevent this disruption.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Interaction in vivo between hapten-specific suppressor T cells and an in vitro cultured helper T cell line.

The interaction in vivo between hapten-specific suppressor T cells (Ts) and a hapten-specific T helper (Th) cell line was examined. Antigen-specific Ts were induced in CBA mice by i.v. priming with 3 X 10(7) syngeneic spleen cells (SC) that were chemically coupled with the hapten azobenzenearsonate (ABA). Transfer of splenic T cells from these mice by i.v. injection suppressed the induction in syngeneic assay hosts of ABA-reactive helper and cytotoxic T cell (Tc) responses. Although the Th responses and their suppression were ABA specific, in that they were not induced or activated by trinitrophenyl (TNP)-coupled SC, both Tc responses and their suppression were occasionally nonspecific. Induction of Th was assayed by measuring the release from primed lymph node cells of IL 2 and IL 3 in response to haptenated SC in vitro. Both cytotoxic and Th responses could be made dependent on the provision of exogenous Th by reducing the antigen dose. This stratagem allowed the assay in vivo of a long-term cultured ABA-specific Th cell line (E9). Injection of 10(5) E9 cells/mouse (with antigen, in the rear footpad) helped the induction of both Tc and Th in response to a reduced dose of antigen. These responses, which were dependent on the E9 cell line, were also suppressed by i.v. transferred Ts. When normal doses of antigen were used, the injection of 10(5) E9 Th overcame suppression. These results show that Ts act by inhibiting the activation of Th, thereby suppressing Th-dependent responses generally. The fact that the E9 Th cell line could be suppressed also shows that long-term culture of T cells does not affect their capacity to be regulated in vivo.

Animals↗
PubMed1987

Coaggregation of the T-cell receptor with CD4 and other T-cell surface molecules enhances T-cell activation.

The CD4 molecule, expressed by T cells restricted by class II major histocompatibility complex (MHC) molecules, is believed to play a role in T-cell activation. We have previously suggested that CD4 interacts with the T-cell receptor for antigen (TCR) and with class II MHC and that this dual interaction stabilizes the bond between the TCR and antigen in association with MHC. To investigate the contribution of CD4-TCR interaction, we have used the murine monoclonal anti-TCR V beta 8 antibody F23.1 to activate cloned T cells. Weak activation by soluble biotinylated F23.1 was markedly enhanced by crosslinking with either avidin or with anti-immunoglobulin (anti-Ig). The monoclonal anti-L3T4 antibody GK1.5, which normally inhibits the activation induced by F23.1, did not inhibit when GK1.5 and F23.1 were coaggregated on T cells by anti-Ig, and in many experiments activation was enhanced. Coaggregation of anti-Thy-1.2, anti-H-2Kk, or anti-LFA-1 with F23.1 also enhanced T-cell activation, although, unlike GK1.5, these antibodies in soluble form had no effect on the response to F23.1. These results are consistent with a model for T-cell activation that proposes a primary interaction between L3T4 and the TCR to stabilize TCR complexes and so to enhance T-cell activation. A related but less specific accessory role for other T-cell surface molecules is also suggested. We propose that the cellular interaction that leads to physiological T-cell activation not only achieves TCR ligation but also promotes through their ligation or redistribution the interaction of other T-cell surface molecules, all of which contribute to the overall strength of the activation signal.

Animals↗
PubMed1985

Suppression induction in vivo by a T helper clone?

We have previously described a helper T cell clone which augments in vivo cytotoxic T cell responses when injected at 10(4) cells per mouse, but not at 10(5) per mouse (Crispe, I. N. et al., Immunology 1984. 52:55). To test whether this dose-response relationship was due to the induction of suppression, naive syngeneic mice were injected with 10(5) cloned T helper cells, and their spleen cells were subsequently assayed for suppressive activity in adoptive transfer experiments. Lymphocytes from such mice indeed suppressed an antigen-specific cytotoxic response, but only in the presence of the same T helper cell clone freshly added at the time of adoptive transfer. On this basis we argue that the distinction between T helper cell activity and T suppressor-inducer activity corresponds to differences in cell numbers, rather than to two separate cell lineages.

Animals↗
PubMed1985

Suppressor T cells, distinct from "veto cells," are induced by alloantigen priming and mediate transferable suppression of cytotoxic T lymphocyte responses in vivo.

Primary and secondary cytotoxic T lymphocyte responses to minor alloantigens can be suppressed by priming host mice with a high dose (10(8) cells) of alloantigenic donor spleen cells (SC). Such suppression is antigen specific and transferable into secondary hosts with T cells. One interpretation of this is that antigen-specific host suppressor T cells (Ts) are activated. Alternatively, donor Lyt-2+ T cells, introduced in the priming inoculum, may inactivate host CTL precursors (CTLp) that recognize the priming (donor) alloantigens. Donor cells that act in this way are termed veto T cells. The experiments described here exclude veto T cell participation in transferable alloantigen-specific suppression, and demonstrate the operation of an alloantigen-specific host-derived T suppressor (Ts) cell. The origin of the Ts has been studied directly by using Thy-1-disparate BALB/c mice. The cell responsible for the transfer of suppression of a secondary CTL response to B10 minors was of the host Thy-1 allotype, and so originated in the host spleen and was not introduced in the priming inoculum. Secondly, antigen-specific Ts generated in CBA female mice against B10 minors could act on CTL responses to an unequivocally non-cross-reactive-third party antigen (H-Y), provided the two antigens were expressed on the same cell membrane. Such third-party suppression is incompatible with the operation of veto T cells. Depletion of Thy-1.2+ or Lyt-2+ cells from the suppression-inducing donor SC inoculum did not abrogate suppression induction in BALB/c mice; instead, suppression was enhanced. The demonstration of veto cell activity in similarly primed mice by other groups of investigators indicates that both types of suppression may operate. However, our results show that only antigen-specific Ts can mediate the transferable suppression of CTL responses to alloantigens.

Animals↗
PubMed1984

Helper T cell lines to major and to minor histocompatibility antigens are equally effective in the regulation of B cell responses in vivo.

Long-term lines of helper T (Th) cells, reactive to minor histocompatibility (minor-H) antigens, were grown by antigen restimulation in the absence of exogenous interleukin-2. These lines were antigen specific and H-2b restricted. When introduced in vivo by adoptive transfer, these Th cells helped syngeneic B cells in an antibody response to other alloantigens. Linked recognition was required for effective help to occur, this suggests B cell presentation of antigen to Th cells in vivo. Parallel titration experiments performed with long-term cultured Th lines to MHC and to minor-H antigens showed that, on a per cell basis, they are equivalent in their ability to help in vivo B cell responses. This shows that any inability to produce antisera to minor-H antigens is not due to a Th or APC defect, but results from either a B cell defect or from suppression.

Animals↗
PubMed1984

Derivation from an alloreactive T-cell line of a clone which cross-reacts with a self H2-E-restricted minor alloantigen.

An alloreactive T-helper-cell line [(A.TH X Balb/c) anti-A.TL] was shown to recognize both H2-Ek and H2-Ed. Both proliferation and polyclonal B-cell activation (protein A plaques) were used in the analyses of specificity. On cloning, the H2-Ek/Ed cross-reaction was shown by one clonotype. This cross-reaction is interpreted in light of the P. Matzinger and M.J. Bevan model (Cell. Immunol. 29, 1, 1977) for antigen recognition by T cells. The antigen recognized was a cell surface non-major histocompatibility (MHC) (minor) alloantigen, shared by the DBA and B10 backgrounds, whose recognition was H2-Ed restricted. This clone is therefore both alloreactive and self-MHC (Ed) restricted minor reactive. Other clones from the cell line were either allospecific or autoreactive. It was shown by antibody blocking that the H2-E alpha (Ia7) determinants involved in the cross-reactive and allospecific recognition were not the same. By antibody blocking it was also shown that the antigenic determinant recognized by the autoreactive clone included H2-Ed. The alloreactive cell line therefore contained three H2-E-reactive clonotypes: allo-Ek specific, H2-Ek and DBA/B10 minor plus H2-Ed cross-reactive, and Ed-associated autoreactive.

Animals↗