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Biomedical subjects

R A Seder

Publications and source records attributed to R A Seder.

At least 55 records · Page 3Linked to original sources
PubMed1994

Anti-IL-4 treatment of Schistosoma mansoni-infected mice inhibits development of T cells and non-B, non-T cells expressing Th2 cytokines while decreasing egg-induced hepatic fibrosis.

Increasing evidence suggests that schistosome egg granulomas are primarily Th2 cellular reactions. Mice infected with Schistosoma mansoni were treated with a neutralizing mAb against IL-4 to evaluate the role of this cytokine in the generation of parasite egg-induced cell-mediated responses and hepatic pathology. Animals treated with anti-IL-4 before egg deposition showed decreased IL-4, IL-5, and IL-10 production in response to in vitro antigenic stimulations and decreased IL-5 and IL-13 mRNA levels in the liver. As observed previously, non-B, non-T cells were a major source of IL-4 in infected mice treated with control mAb, and the diminished IL-4 response in anti-IL-4-treated animals was shown to be caused at least in part by a reduction in the number of these cells, as well as by decreased secretion of IL-4 per cell. In contrast, production of the Th1 cytokines IL-2 and IFN-gamma was elevated in anti-IL-4-treated infected mice in vitro, and the corresponding mRNAs in the liver were increased. Anti-IL-4 treatment did not consistently reduce the size of hepatic granulomas around S. mansoni eggs, but markedly inhibited granuloma formation in the lungs of the same animals after i.v. egg injection. Nevertheless, anti-IL-4-treated infected mice showed consistent and marked reductions in hepatic collagen deposition. These findings indicate that IL-4 plays a major role in the development of the Th2 response in S. mansoni-infected mice and contributes to the pathogenesis of hepatic fibrosis.

Animals↗
PubMed1994

CD28-mediated costimulation of interleukin 2 (IL-2) production plays a critical role in T cell priming for IL-4 and interferon gamma production.

Naive T cells require interleukin 4 (IL-4) to develop into IL-4-producing T cells and IL-4 blocks development of such cells into interferon gamma (IFN-gamma) producers. Prior studies in accessory cell-independent priming systems using antireceptor antibodies as agonists have demonstrated that IL-2 is also necessary for the development of IL-4-producing cells under these culture conditions. Here we have examined the role of IL-2 and the CD28 costimulation pathway in priming for IL-4 and IFN-gamma production using a more physiologic model. This involved antigen presentation by accessory cells to naive CD4+ T cells from transgenic mice whose cells express a T cell receptor (TCR) specific for a cytochrome c peptide in association with I-Ek. With splenic antigen-presenting cells (APCs), inhibition of CD28 costimulation by the fusion protein CTLA4-immunoglobulin (Ig) blocked effective priming. Similarly, transfected fibroblasts expressing both MHC class II and the CD28 ligand B7 could prime for IL-4 production and such priming also was blocked by CTLA4-Ig. However, APCs deficient in CD28 ligands also could prime TCR transgenic T cells to become IL-4 producers if an exogenous source of IL-2, as well as IL-4, was provided, and the inhibition of priming seen with splenic or transfected fibroblast APCs in the presence of CTLA4-Ig could be reversed by addition of IL-2. Likewise, priming for IFN-gamma production could be blocked by CTLA4-Ig and reversed by IL-2. Thus, we conclude that IL-2 plays a critical role in priming naive CD4+ T cells to become IL-4 or IFN-gamma producers. Engagement of the CD28 pathway, although normally important in such priming, is unnecessary in the presence of exogenous IL-2.

Animals↗
PubMed1994

Acquisition of lymphokine-producing phenotype by CD4+ T cells.

Naive CD4+ T cells when stimulated produce IL-2 as their major lymphokine. Upon priming, these cells develop into cells that produce either IFN gamma, TNF beta, and IL-2 or IL-4 and its congeners. The former cells are designated TH1-like, and the latter TH2-like. Here we review the regulation of the differentiation of naive CD4 cells into IFN gamma- or IL-4-producers. The dominant factors that determine such differentiation are lymphokines and other cytokines. IL-2 itself appears to be required for naive cells to develop into TH1- or TH2-like cells but is not deterministic of their differentiation fate. If IL-4 is also present during the priming period, the resultant CD4+ T cells produce IL-4 upon restimulation; the development of IFN gamma-producing cells is strikingly inhibited by IL-4. In the absence of IL-4, priming for IFN gamma-production occurs, but this is markedly enhanced by IL-12. The role of IFN gamma in enhancing priming for IFN gamma-production is not fully resolved. In some in vitro systems, it appears to act together with IL-12 to enhance such production. Anti-IFN gamma diminishes priming for IFN gamma production in vivo. Lymphokines also exert a "cross-regulatory" or inhibitory effect. As noted above, IL-4 strikingly diminishes priming for IFN gamma production, although this inhibitory effect is blunted in the presence of IL-12. IFN gamma similarly diminishes priming for IL-4 production; this effect is principally observed when low concentrations of IL-4 are used in the priming culture. Although other factors may play a role in the determination of lymphokine-producing phenotype, such as antigen dose, type of antigen-presenting cell, and expression of accessory molecules and hormones, these effects appear to be secondary to the dominant role of the lymphokines and cytokines.

Animals↗
PubMed1994

Effects of interleukin-3 with or without the c-kit ligand, stem cell factor, on the survival and cytoplasmic granule formation of mouse basophils and mast cells in vitro.

We assessed the ultrastructure and the cell-surface expression of receptors for immunoglobulin E (Fc epsilon R), and c-kit, the receptor for stem cell factor (SCF), in mouse basophils and mast cells present in short-term cultures of mouse bone marrow cells in interleukin-3 (IL-3) with or without SCF. Basophils did not develop increased numbers of cytoplasmic granules and underwent apoptosis in cultures containing IL-3 and SCF, whereas mast cells thrived and developed increased numbers of granules. Basophils were nearly all Fc epsilon R+ c-kit- when sorted after culture in IL-3 and SCF; most mast cells were Fc epsilon R+ c-kit+. However, a second population of Fc epsilon R+ c-kit- mast cells was present after culture in IL-3 and SCF. These c-kit receptor-negative mast cells were less mature than c-kit+ mast cells and contained significantly fewer cytoplasmic granules than the c-kit+ mast cells present in the same cultures (P < 0.001). Thus, mouse basophils express little or no c-kit receptor on their surface, nor can they survive for long periods in SCF-supplemented cultures. By contrast, mouse mast cells seem to express the Fc epsilon R early in their development, even before they express detectable c-kit receptors on their surface. IL-3 promotes cytoplasmic granule formation in immature mast cells, but even more granules are formed when c-kit receptor-positive immature mast cells are cultured in both SCF and IL-3.

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PubMed1993

Interleukin 12 acts directly on CD4+ T cells to enhance priming for interferon gamma production and diminishes interleukin 4 inhibition of such priming.

Naive CD4+ T cells produce interleukin 2 (IL-2) but little IL-4 or interferon gamma (IFN-gamma). In vitro, they develop into IL-4 or IFN-gamma producers depending on the conditions of the priming culture. Using T-cell receptor transgenic CD4+ T cells, the role of IL-12 and IL-4 in antigen-specific priming was examined. IL-12 substantially enhanced the ability of naive CD4+ T cells to develop into cells that produced IFN-gamma upon restimulation. However, it was not essential since anti-IL-12 antibodies failed to block the priming for IFN-gamma observed in the absence of exogenous IL-12. When both IL-12 and IL-4 were present in the priming culture, IL-12 did not inhibit priming for IL-4 production. In contrast, IL-4 diminished but did not abolish priming for IFN-gamma production. In an accessory cell-independent priming system, IL-12 strikingly augmented priming for IFN-gamma production, indicating that it acts directly on T cells. IFN-gamma itself did not enhance priming for IFN-gamma production in either accessory cell-dependent or independent systems. In an accessory cell-dependent system, the IL-12-mediated enhancement was not blocked by adding neutralizing anti-IFN-gamma monoclonal antibody. However, in an accessory cell-independent system, anti-IFN-gamma antibody did inhibit priming for IFN-gamma production leaving open a role for IFN-gamma in the priming process. These data indicate that IL-12 has a major effect on the inductive phase of T-cell priming by enhancing commitment to IFN-gamma production and thus can profoundly influence the state of immunity that develops.

Animals↗
PubMed1993

Interleukin 4 suppresses interleukin 2 and interferon gamma production by naive T cells stimulated by accessory cell-dependent receptor engagement.

Interleukin 2 (IL-2) and interferon gamma (IFN-gamma) production by CD4+ T cells and IFN-gamma production by CD8+ T cells from naive mice in response to soluble anti-CD3 and antigen-presenting cells (APCs) were strikingly inhibited by culture in the presence of IL-4. IL-4 decreased IL-2 and IFN-gamma mRNA levels after 15-24 hr but gave relatively little decrease in these mRNAs at 6-12 hr after stimulation with soluble anti-CD3. A 16-hr preculture of T cells with anti-CD3, APCs, and IL-4 was sufficient to inhibit subsequent production of IL-2 and IFN-gamma in response to restimulation in the absence of IL-4. Furthermore, IL-4 treatment of T cells purified 24 hr after stimulation inhibited their capacity to subsequently produce IL-2 in response to anti-CD3 and APCs, indicating that T cells were targets of IL-4-mediated inhibition. IL-4 blocked acute IL-2 production in response to a cytochrome c peptide of T cells derived from transgenic mice expressing T-cell receptors specific for cytochrome c but it did not block IL-2 production by such cells after they had been primed in vitro. Nor did IL-4 inhibit production of IFN-gamma by cloned T cells in response to antigen and APCs or production of IL-2 and IFN-gamma by naive T cells in response to phorbol ester and calcium ionophore. These results indicate that IL-4 strikingly inhibits IL-2 and IFN-gamma production by naive T cells in response to accessory cell-dependent, receptor-mediated stimulation (i.e., soluble anti-CD3 and APCs or antigen and APCs) but does not inhibit accessory cell-independent stimulation of naive T cells or accessory cell-dependent receptor-mediated stimulation of recently primed T cells or cloned T-cell lines.

Animals↗
PubMed1993

Fc epsilon receptor-positive cells are a major source of antigen-induced interleukin-4 in spleens of mice infected with Schistosoma mansoni.

When cultured in vitro with either mitogen or parasite antigens, spleen cells from mice infected with Schistosoma mansoni produce significantly higher levels of IL-4 than splenocytes from control animals. Previous studies suggested that this increase in IL-4 production occurs because of a selective expansion of T helper type 2 (Th2) cells in infected mice. However, these experiments employed unfractionated spleen populations rather than purified T lymphocytes. Here we demonstrate that T-depleted spleen cells from infected animals synthesize high levels of interleukin-4 (IL-4), but no IL-5 when stimulated with parasite antigen in vitro. Nevertheless, when purified by sorting, T cells and non-B, non-T (NBNT) populations produced similar amounts of IL-4 in response to parasite antigen. The IL-4 producing NBNT cells were found to belong to an Fc epsilon receptor (Fc epsilon R)-positive population which after sort purification produced high levels of IL-4 (between 1000 and 2000 U of per 5 x 10(3) cells). FACS analysis revealed that these Fc epsilon R+ cells make up 0.53% of splenic NBNT cells in control animals while in 8-9-week-infected animals they increase to 3.8% of that population. In contrast, in mice with 8-week unisexual worm infections these cells comprise only 1.71% of NBNT cells, indicating that eggs are a major stimulus of the response. The expansion of Fc epsilon R+ cells and their production of IL-4 could be an important factor regulating the selection and induction of different CD4+ subsets in schistosome-infected hosts.

Animals↗
PubMed1993

Ultrastructural characteristics of Fc epsilon R-positive basophils in the spleen and bone marrow of mice immunized with goat anti-mouse IgD antibody.

BACKGROUND: Previous work showed that injection of mice with goat anti-mouse IgD antibodies results in increased numbers of Fc epsilon R-positive, non-B, non-T cells in the spleen and Fc epsilon R-positive cells in the bone marrow, and that some of these cells had ultrastructural features of basophils. Fc epsilon R-positive, non-B, non-T cells express virtually all of the capacity of mouse splenic "non-B, non-T cells" to produce interleukin-4 in response to stimulation by cross-linking of Fc epsilon R or Fc gamma R, or by the calcium ionophore, ionomycin. EXPERIMENTAL DESIGN: The present study is a detailed ultrastructural analysis of Fc epsilon R-positive bone marrow cells or Fc epsilon R-positive splenic non-B, non-T cells sorted from mice injected with goat anti-mouse IgD antibody and of Fc epsilon R-positive bone marrow cells or spleen cells pooled from normal mice not injected with goat anti-IgD. RESULTS: Basophils represented the majority (90%) of the granulated cells present in the Fc epsilon R-positive splenic non-B, non-T cells or Fc epsilon R-positive bone marrow cells of goat anti-IgD-injected mice. In contrast, the cytoplasmic granule-containing Fc epsilon R-negative cells sorted from spleen or bone marrow of goat anti-IgD-injected animals contained predominantly a mixture of neutrophils, eosinophils, monocytes and their precursors. Both the Fc epsilon R-positive and -negative preparations contained rare (< 5%) cells with ultrastructural features of very immature mast cells. Basophils were also identified in Fc epsilon R-positive cells sorted from total bone marrow cells or spleen cells of normal mice not injected with goat anti-IgD. CONCLUSIONS: Taken together with data concerning the numbers of Fc epsilon R-positive, non-B, non-T cells in the spleen, and Fc epsilon R-positive B220-negative cells in the bone marrow, these ultrastructural findings indicate that injection of mice with goat anti-IgD results in increased numbers of basophils, particularly in the spleen, that exhibit an 8-fold increase in basophils as a result of injection of goat anti-IgD.

Animals↗
PubMed1992

The presence of interleukin 4 during in vitro priming determines the lymphokine-producing potential of CD4+ T cells from T cell receptor transgenic mice.

To study the factors that determine whether CD4+ T cells produce interleukin 4 (IL-4) or interferon gamma (IFN-gamma) upon stimulation we used a system allowing naive T cells to be primed in vitro by specific antigen. Dense CD4+ T cells were purified from mice that expressed transgenes encoding a T cell receptor specific for pigeon cytochrome C peptide 88-104 in association with I-Ek. These T cells produced very limited amounts of IL-4 and IFN-gamma upon immediate challenge with 88-104 and antigen-presenting cells (APC). However, after an initial "priming" culture in which they were incubated for 4 d in the presence of 88-104, APC, and 1,000 U/ml IL-4, the T cells acquired the capacity to produce substantial amounts of IL-4 upon rechallenge but made very little IFN-gamma. Cells primed in the absence of IL-4 produced IFN-gamma upon rechallenge but virtually no IL-4. The inhibitory effect of IL-4 on IFN-gamma production did not appear to be mediated by the induction of IL-10 production since IL-10 addition to initial cultures did not suppress priming for IFN-gamma production, nor did anti-IL-10 block the inhibitory effect of IL-4. IFN-gamma itself did not increase priming for IFN-gamma production, nor did anti-IFN-gamma reduce such priming. IFN-gamma did, however, diminish priming for IL-4 production when limiting amounts of IL-4 (100 U/ml) were used in the initial culture. The dominant effect of IL-4 in determining the lymphokine-producing phenotype of primed cells was observed with dendritic cells (DC), activated B cells, and I-Ek-transfected fibroblasts as APC. However, the different APC did vary in their potency, with DC being superior to activated B cells, which were superior to transfected fibroblasts.

Amino Acid Sequence↗
PubMed1992

CD8+ T cells can be primed in vitro to produce IL-4.

IL-4 production by T lymphocytes from naive mice in response to stimulation by plate-bound anti-CD3 is concentrated among CD4+ T cells. In vitro stimulation of lymph node T cells with anti-CD3 plus IL-2 and IL-4 strikingly increases the frequency of cells that produce IL-4 in response to subsequent stimulation with anti-CD3 plus IL-2. Separation of these primed cell populations into CD4+ and CD8+ T cell by cell sorting reveals that the frequency of IL-4-producing cells in both population is similar. Verification that CD8+ T cells produce IL-4 is provided by the capacity of anti-IL-4 mAb to inhibit the response of the indicator cell line to the growth factor produced by the primed cells and by detection of IL-4 by an IL-4-specific ELISA. The in vitro "priming" of CD8+ T cells to produce IL-4 is not dependent on the presence of CD4+ T cells because highly purified CD8+ T cells can be stimulated to develop into cells capable of producing IL-4 by culture with plate-bound anti-CD3 plus IL-2 and IL-4.

Animals↗
PubMed1992

Activation events during thymic selection.

During their differentiation in the mouse thymus, CD4+8- cells undergo several of the sequential changes observed upon normal activation of mature, peripheral CD4+ lymphocytes. Expression of CD69, an early activation marker, is first observed on a minority of cells at the T cell receptor (TCR)lo/med double-positive stage, is maximal (50-90%) on heat-stable antigen (HSA)hi TCRhi double-positive, HSAhi TCRmed CD4+8lo, and HSAhi TCRhi CD4+8- cells, and is downmodulated at the mature HSAlo CD4+8- stage. In contrast, CD44, a late activation marker, is selectively expressed at the HSAlo stage. The set of lymphokines that CD4+8- thymocytes can produce upon stimulation also characteristically expands from mainly interleukin 2 (IL-2) at the HSAhi stage, to IL-2 and very large amounts of IL-4, IL-5, IL-10, and interferon gamma (IFN-gamma) at the HSAlo stage. 1 in 30 HSAlo CD4+8- adult thymocytes secrete IL-4 upon stimulation through their TCR. This frequency is 25% of the frequency of IL-2 producers, about 100-fold above that of peripheral (mainly resting) CD4+ T cells. With time after their generation in organ culture, CD4+8- thymocytes lose their capacity to secrete IL-4, IL-5, and IFN-gamma, but not IL-2. Similarly, the frequency of IL-4, but not of IL-2, producers progressively decreases after emigration to the periphery as judged by direct comparison between thymic and splenic CD4+ cells in newborns, or by following the fate of intrathymically labeled CD4+8- cells in adults after their migration to the spleen. This sequence suggests that thymic selection results from an activation process rather than a simple rescue from death at the double-positive stage, and shows that the functional changes induced after intrathymic activation, although transient, are still evident after export to the periphery.

Animals↗
PubMed1991

Purified Fc epsilon R+ bone marrow and splenic non-B, non-T cells are highly enriched in the capacity to produce IL-4 in response to immobilized IgE, IgG2a, or ionomycin.

Non-B, non-T cells from spleen and bone marrow cells produce IL-4 in response to cross-linkage of high affinity receptors for Fc epsilon R or Fc gamma RII, and to treatment with calcium ionophores. Cells bearing high affinity Fc epsilon R constituted 1 to 2% of non-B, non-T cells of spleen and of total bone marrow cells from naive donors. In mice whose immune systems had been polyclonally activated by injection with anti-IgD antibodies or had been infected with Nippostrongylus brasiliensis larvae, the frequency of Fc epsilon R+ cells in splenic non-B, non-T cells was also 1 to 2% but in bone marrow from anti-IgD-injected mice donors the frequency was approximately 5%. Cell sorting experiments revealed that all of the capacity to produce IL-4 in response to immobilized IgE or IgG2a or to ionomycin was found in the Fc epsilon R+ fraction. Among the Fc epsilon R+ spleen cells from naive donors, the frequency of IL-4-producing cells was 1/20 to 1/40 whereas in mice that had been injected with anti-IgD or infected with N. brasiliensis, the frequency of IL-4 producing cells in the Fc epsilon R+ population was approximately 1/5.

Animals↗
PubMed1991

Mouse splenic and bone marrow cell populations that express high-affinity Fc epsilon receptors and produce interleukin 4 are highly enriched in basophils.

Splenic and bone marrow cells from normal mice, and from mice that have been polyclonally activated by injection of anti-IgD antibody, contain cells that produce interleukin 4 (IL-4) in response to crosslinkage of Fc epsilon receptors (Fc epsilon R) or Fc gamma R or to ionomycin. Isolated Fc epsilon R+ cells have recently been shown to contain all of the IL-4-producing capacity of the nonlymphoid compartment of spleen and bone marrow. Here, purified Fc epsilon R+ cells are shown to be enriched in cells that contain histamine and express alcian blue-positive cytoplasmic granules. By electron microscopy, the vast majority of cytoplasmic granule-containing cells are basophils; they constitute approximately 25% and approximately 50%, respectively, of Fc epsilon R+ spleen and bone marrow cells from anti-IgD-injected mice. The Fc epsilon R- populations contain cells that form colonies typical of mast cells. The Fc epsilon R+ populations also contain cells that, upon culture with IL-3, form colonies of alcian blue-positive cells, but (in contrast to colonies derived from Fc epsilon R- populations) the colonies are small, and all the cells die within 2-3 weeks. The Fc epsilon R+ cells synthesize histamine during a 60-hr culture with IL-3, while the Fc epsilon R- cells do not. These results indicate that IL-4-producing Fc epsilon R+ cells are highly enriched in basophils.

Animals↗
PubMed1991

Increased frequency of interleukin 4-producing T cells as a result of polyclonal priming. Use of a single-cell assay to detect interleukin 4-producing cells.

A limiting dilution assay capable of detecting interleukin 4 (IL4) production by a single cell has been developed. This assay is based on the stimulation of T cells, in the presence of IL2 (5 U/ml), with anti-CD3 antibody bound to the surface of Terasaki wells. Cells of the IL4-selective indicator line, CT.4S, are added 24-36 h later and IL4 production is determined based on their survival 24-48 h thereafter. A frequency of 0.98 was obtained for IL4 production by T cells of the D10.G4 cell line. T cells from naive donors capable of producing IL4 in response to anti-CD3 plus IL2 were quite rare, with a frequency in four experiments ranging between 0.0003 and 0.0018. Treatment of mice with polyclonal activators known to increase the IL 4-producing capacity of T cells when assayed in bulk culture caused striking increases in the frequency of IL4-producing cells. Similarly, culturing cells in vitro with anti-CD3, IL2 and IL4 for 5 days caused a marked increase in the frequency of cells capable of producing IL4, to 0.031.IL4 production by individual T cells is dependent upon IL2. Thus, in naive T cell populations, the frequency of IL4-producing cells in response to stimulation with anti-CD3 in the absence of IL2 was below the limit of detection. T cells from primed donors showed a striking inhibition in the frequency of IL4-producing cells in response to anti-CD3 when IL2 was not present. The availability of a simple assay to measure the frequency of cells capable of producing IL4 should have substantial utility in allowing the evaluation of conditions that regulate IL4 production in vivo and in vitro.

Animals↗
PubMed1991

Production of interleukin-4 and other cytokines following stimulation of mast cell lines and in vivo mast cells/basophils.

Interleukin-3 (IL-3)-dependent mast cell lines, upon stimulation by calcium ionophores or by Fc epsilon RI cross-linking, express mRNA for, and secrete, a distinct pattern of cytokines, similar to those secreted by cloned mouse T cells of the TH2 type. The mast-cell-derived cytokines include IL-3, IL-4, IL-5 and IL-6. Not only in vitro mast cell lines, but also in vivo derived peritoneal mast cells secrete cytokines. An in vivo derived cell, in mouse spleen and bone marrow, secretes IL-4 and other cytokines upon stimulation with calcium ionophores or by Fc epsilon RI cross-linking or Fc gamma RII cross-linking. The IL-4-producing cells are highly enriched in the Fc epsilon R+ subset of spleen and bone marrow cells. These Fc epsilon R+ cells produce large amounts of IL-4, and they have characteristics similar to those of immature mast cells and/or basophils. It is possible that cytokines produced by mast cells and/or basophils participate in allergic inflammatory diseases.

Animals↗