Th Cell Clones Research Articles

Differential Requirement for Humoral Factors for IL-2R Expression of Murine T Cell Subsets, Th1, Th2, and CD8Th Clones

Three types of alloreactive T helper (Th) cell clones, Th1, Th2, and CD8Th, were examined for the requirement of costimulatory factors for IL-2 receptor (IL-2R) expression and proliferation. When stimulated with allogeneic spleen adherent cells, these Th cell clones all expressed IL-2R strongly, whereas they expressed the receptor only weakly when subjected to allogeneic B cell stimulation. The addition of partially purified IL-2RIF to Th1, Th2, or CD8Th cell clones stimulated with B cell antigen-presenting cells increased the IL-2R expression of Th1 and CD8Th cell clones but not of the Th2 cell clone. Conversely, IL-1 up-regulated IL-2R expression of Th2 cell clones stimulated with B cell APC but not of Th1 and CD8Th cell clones. Neither IL-2RIF nor IL-1 induced IL-2R expression of unstimulated Th cell clones. IL-2R expressed in the presence of these costimulators contributed to IL-2-dependent proliferation of these cells. Thus, the expression of IL-2R was indicated to be regulated differently in three types of Th cell subsets through different cytokines.

Gangliosides: differentiation markers for murine T helper lymphocyte subpopulations TH1 and TH2.

On the basis of the pattern of lymphokines they secrete, murine T helper clones can be divided into two subsets, TH1 and TH2. This concept of two different T helper effector cells helps to explain the diversity of immune reactions occurring in different parts of the body. The in vivo localization of T helper subtypes is of great interest, but up to now no biochemical or surface markers were available to distinguish between them. We analyzed the glycolipids from altogether 12 murine TH1 and TH2 cell lines or clones. A comparison of the gangliosides by thin-layer chromatography showed differences between the TH1 and TH2 cells. Binding studies with specific antibodies to asialo backbone structures after degradation by neuraminidases showed that the main gangliosides from these lymphocytes shared a common GgOse4 backbone and thus differed only in their degree or position of sialylation. Two disialogangliosides appeared to be characteristic. They were isolated from the D10.G4.1 TH2 cell clone and identified by fast atom bombardment mass spectrometry as IVNeuAc,IINeuAc-GgOse4Cer (GD1a) and IVNeuAc,IIINeuAc-GgOse4Cer (GD1 alpha), respectively. GD1a was characteristically only detected in TH2 cells, whereas GD1 alpha was preferably, but not exclusively, expressed by TH1 lymphocytes. Although GD1a was also found in the lung, heart, kidney, and spleen, its expression within the murine immune cells under investigation was unique to TH2 lymphocytes. Scarcely any GD1a was found in thymocytes, B cells, or CD8 positive (cytolytic) T cells, but significant expression was seen in CD4 positive (helper) T cells which include the TH2 subpopulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of transforming growth factor (TGF)-beta 1 on IgA isotype expression. TGF-beta 1 induces a small increase in sIgA+ B cells regardless of the method of B cell activation.

Transforming growth factor-beta (TGF-beta) has been reported to play an important role in IgA isotype expression when B cells are stimulated with LPS. The goal of the present study was to determine whether TGF-beta has similar effects on IgA isotype expression under more physiologic conditions utilizing a variety of B cell activation systems. As previously reported, in the LPS system TGF-beta caused a small, but significant, absolute increase in surface IgA (sIgA) expression and a very definite increase in IgA secretion; these effects were enhanced by IL-2 and IL-5. In the case of B cell stimulation with another B cell stimulant, the thymus-independent type II mitogen, anti-IgD-dextran, TGF-beta led to a similar small increase in sIgA expression, but caused suppression of IgA secretion. Using the Th2 cell clones CDC35 and D10 to stimulate resting B cells in a cognate and non-cognate T cell-dependent fashion, respectively, TGF-beta again increased sIgA expression to a similar small extent. TGF-beta at low doses (0.1 ng/ml) did not increase IgA secretion significantly and, at higher doses (1.0 ng/ml) caused significant suppression of IgA secretion. Addition of various cytokines (IL-2, -4, -5, D10sup) other than TGF-beta to stimulated B cells did not increase sIgA expression, but did give rise to increased amounts of IgA secretion, especially when activated D10 T cells were used as the B cell stimulant. Finally, the addition of an antibody against TGF-beta to cultures containing TGF-beta on day 2 led to partial or complete reversal of the inhibitory effects of TGF-beta on IgA secretion. In conclusion, TGF-beta causes a consistent, but small increase in sIgA+ B cells, in cultures of B cells stimulated by a variety of T cell-dependent or independent stimuli. In contrast, TGF-beta either promotes or inhibits B cell survival and IgA secretion, depending on the method of B cell activation. These results are most consistent with the view that TGF-beta provides only a partial or incomplete IgA switch signal but that additional factors are involved in IgA isotype switching and differentiation.

Isolation and characterization of an IL-1-dependent IL-4-producing bovine CD4 + T cell clone

An Ag-specific interleukin 1 (IL-1)-dependent bovine CD4 + Th cell clone, termed 300B1, was isolated and found to resemble the previously described IL-1-dependent murine CD4 + Th2 cell clone, D10.G4.1. Both the 300B1 and the D10.G4. 1 T cell clones proliferated to bovine (Bo) IL-1β human (Hu) IL-1α and IL-1β, and murine IL-1α when cells were costimulated with concanavalin A (Con A). Proliferation of the 300B1 clone, when costimulated with Con A, appeared to be IL-1 -specific in that proliferation could not be promoted by BoIL-2, HuIL-3, HuIL-4, HuIL-5, or HuIL-6. The 300B1 clone produced interferon-γ (IFN-γ), but not IL-2 following stimulation with either Con A, Con A plus phorbol 12-myristate 13-acetate or Ag plus antigen-presenting cells. Upon stimulation with Con A, the 300B1 clone expressed IL-4 mRNA and produced an automne growth factor (AGF) that could be inhibited by anti-HuIL-4 but not by anti-HuIL-2 Ab. The clonal derivation of the 300B1 clone was confirmed by isolating five 300B1 subclones, all of which produced IFN-γ and an AGF but not IL-2. Collectively, these results suggest the IL-1-dependent bovine 300B1 Th cell clone produces IL-4, but not IL-2, as an AGF. Furthermore, the bovine Th cell clone appeared to share many characteristics of previously described murine Th2 cell clones except that the bovine clone produced IFN-γ.

Junctional region sequences of T-cell receptor beta-chain genes expressed by pathogenic anti-DNA autoantibody-inducing helper T cells from lupus mice: possible selection by cationic autoantigens.

We rescued from the spleens of 10 (SWR x NZB)F1 (SNF1) mice with lupus nephritis the T cells that were activated in vivo and cloned 268 T-cell lines and hybridomas. Only 12% of these T-cell clones had the functional ability to preferentially augment the production of pathogenic anti-DNA autoantibodies. Among these, 16 helper T-cell (Th-cell) clones that were mostly CD4+ and had the strongest autoantibody-inducing ability were analyzed for T-cell receptor (TCR) beta-chain gene usage. Seven of the 16 Th-cell clones expressed beta-chain variable region (V beta) V beta 8 (8.2 or 8.3) genes and three expressed V beta 4, whereas two clones each used a V beta 1 or V beta 2 or V beta 14 gene, suggesting some restriction in TCR gene usage. Although heterogeneous, the V-D-J junctional region sequences of TCR beta-chain genes used by these Th-cell clones invariably encoded one or more negatively charged residues (aspartic or glutamic acid) that had been generated in most cases by unspecified nucleotide (N) additions. Representative pathogenic autoantibody-inducing Th-cell clones could rapidly induce the development of lupus nephritis when injected into young prenephritic SNF1 mice. The pathogenic autoantibody-inducing Th cells expressing the anionic residues in their TCR beta-chain junctions (complementarity-determining region CDR3) were probably selected by some cationic autoantigenic peptide presented by the anti-DNA B cells they preferentially helped. These results offer a clue regarding the nature of the primary autoantigen that may drive the pathogenic autoimmune response in lupus.

Membranes from both Th1 and Th2 T cell clones stimulate B cell proliferation and prepare B cells for lymphokine-induced differentiation to secrete Ig.

Plasma membranes from the mitogen-activated mouse Th2 cell clone D10.G4.1 have recently been shown to provide the cell contact-dependent signals necessary for the induction of small B cell proliferation. Together with the Th2-derived lymphokines IL-4 and IL-5, these membranes stimulate production of Ig isotypes identical to those produced when B cells were stimulated by intact Th2 cells. In contrast, Th1 clones are poor inducers of Ig production in vitro. This could be solely due to differences in the lymphokines released by Th1 and Th2 cells or to differences in the cell-cell contact signals delivered by activated Th1 and Th2 cells. We report that membranes from three different activated Th1 clones induced strong Ag-independent proliferation of small dense B cells. The level of B cell proliferation was enhanced approximately fourfold by the addition of lymphokine-containing supernatant from Con A-activated Th2 cells and was unaffected by any of the lymphokine-containing supernatants from Con A-activated Th1 clones. As with D10.G4.1 membranes, Th1 membranes alone induced B cell proliferation but not secretion of Ig. However, addition of supernatant from Con A-activated D10.G41 cells, but not any supernatants from Con A-activated Th1 cells, induced Ig secretion of all isotypes. These effects were shown to not simply result from increased B cell numbers after stimulation with Th2 lymphokines. Thus, Th1 cell clones seem to poorly induce antibody responses entirely because of their lymphokine repertoire and not because of differences or deficiencies in the ability of these cells to deliver cell contact-dependent signals to B cells.

T-helper cell and associated antibody response to synthetic peptides of the E glycoprotein of Murray Valley encephalitis virus

A battery of 16 synthetic peptides, selected primarily by computer analysis for predicted B- and T-cell epitopes, was prepared from the deduced amino acid sequence of the envelope (E) glycoprotein of Murray Valley encephalitis (MVE) virus. We examined all of the peptides for T-helper (Th)-cell recognition and antibody induction in three strains of mice: C57BL/6, BALB/c, and C3H. Lymphoproliferative and interleukin-2 assays were performed on splenic T cells from mice inoculated with peptides in Freund's incomplete adjuvant or with MVE virus. Several peptides found to contain predicted T-cell epitopes elicited a Th-cell response in at least one strain of mice, usually with a concomitant antibody response. Peptides 145 (amino acids 145 to 169) and 17 (amino acids 356 to 376) were strongly recognized by T cells from all three inbred strains of mice. Peptide 06 (amino acids 230 to 251) primed C57BL/6 mice for Th- and B-cell reactivity with native MVE virus, and T cells from virus-immune mice were stimulated by this peptide. Peptide 06 was recognized by several Th-cell clones prepared from mice immunized with MVE, West Nile, or Kunjin virus. These results indicate that it may be feasible to design synthetic flavivirus peptides that define T-cell epitopes capable of generating a helper cell response for B-cell epitopes involved in protective immunity.

IL-10 acts on the antigen-presenting cell to inhibit cytokine production by Th1 cells.

Murine IL-10 (cytokine synthesis inhibitory factor) inhibits cytokine production by Th1 cell clones when they are activated under conditions requiring the presence of APC. By preincubating APC with IL-10, we demonstrate that IL-10 acts principally on APC to inhibit IFN-gamma production by Th1 clones. Moreover, IL-10 is not active when Th1 cells are stimulated with glutaraldehyde-fixed APC, which also indicates that its action involves regulation of APC function. Furthermore, IL-10 inhibits cytokine synthesis by Th1 cells stimulated with the super-antigen Staphylococcus enterotoxin B, which does not appear to require processing. Flow microfluorimetry purified splenic or peritoneal B cells and macrophages, and B cell and macrophage cell lines can present Ag to Th1 clones. However, IL-10 acts only on sorted macrophages and the macrophage cell line to suppress IFN-gamma production by Th1 clones. IL-10 does not show this effect when B cells are used as APC. In contrast, IL-10 does not impair the ability of APC to stimulate cytokine production by Th2 cells. IL-10 does not decrease IFN-gamma-induced I-Ad levels on a macrophage cell line. Inasmuch as IL-10 also inhibits IL-2-induced IFN-gamma production by Th1 cells in an Ag-free system requiring only the presence of accessory cells, these data suggest that IL-10 may inhibit macrophage accessory cell function which is independent of TCR-class II MHC interactions.

A role for guanine-nucleotide-binding proteins in mediating T-cell-receptor coupling to inositol phospholipid hydrolysis in a murine T-helper (type II) lymphocyte clone.

Perturbation of the T-cell receptor (TCR) complex is followed by the rapid hydrolysis of inositol phospholipids (InsPL) by phospholipase C (PLC), producing diacylglycerol and inositol phosphates, which act as second messengers in signal transduction. The mechanism coupling the TCR to InsPL hydrolysis is not clearly defined, and no information is available on this mechanism in the CD4+ helper subset of T-lymphocytes (Th). We have tested the hypothesis that guanine-nucleotide-binding proteins (G-proteins) may couple the TCR to PLC in a murine Th type II (Th2) cell clone. Cell permeabilization with streptolysin O (SLO) or tetanolysin (TL) was used to allow membrane-impermeable nucleotides access to intracellular sites of action. Exposure of permeabilized Th2 cells to guanosine 5'-[gamma-thio]triphosphate (GTP gamma S), a non-hydrolysable GTP analogue, resulted in a 2.1-2.5-fold increase in inositol phosphate generation. Similarly, perturbation of the TCR with the monoclonal antibody 145.2C11 (directed against the epsilon-chain of the CD3 component of the TCR) resulted in a 3.1-4.2-fold increase in InsPL hydrolysis by permeabilized cells. Both lysins were similarly effective in allowing GTP gamma S induction of InsPL hydrolysis, but TL-permeabilized cells responded better to TCR perturbation than SLO-treated cells. A role for G-proteins in TCR coupling to PLC was further supported by the inhibition of TCR-induced InsPL hydrolysis by guanosine 5'-[beta-thio]diphosphate (GDP beta S), a guanine nucleotide analogue that inhibits G-protein function. ATP was required for TCR-mediated InsPL hydrolysis, and potentiated GTP gamma S-induced hydrolysis. Other nucleotides (i.e. CTP, GDP, GTP, ITP) did not affect the response. These data indicate that G-proteins may contribute to the regulation of PLC activation in Th2 cells, coupling it to the TCR.

Differential expression of the alternatively spliced exons of murine CD45 in Th1 and Th2 cell clones.

Antigen-specific murine CD4+ T cell clones can be divided into functionally distinct subsets known as Th1 and Th2. To date these cells have been indistinguishable by surface phenotype. This report identifies two anti-CD45R monoclonal antibodies (14.8 and C363.16A) that bind preferentially to Th2 cells. Further analysis of the CD45-specific mRNA in Th1 and Th2 cells shows clear differences between these two cell types. Th1 cell clones express mRNA for the two smallest forms of CD45 containing none or only one of the alternatively splices exons. In contrast, Th2 cell clones express predominantly the high molecular weight isoforms of CD45 containing two or three of the alternatively spliced exons.

Immunization of susceptible hosts with a soluble antigen fraction from Leishmania major leads to aggravation of murine leishmaniasis mediated by CD4+ T cells

This study was performed in order to define Leishmania major antigens that function as disease-modulating immunogens in susceptible BALB/c mice. A soluble leishmanial antigen preparation (S-SLA) derived from highly infective stationary-phase L. major parasites was fractionated by preparative gel electrophoresis. In vitro, the low molecular mass fraction (less than 31 kDa) of S-SLA fraction D (FR D) was found to be a potent stimulator of L. major-specific Th1 and Th2 helper cell clones. In vivo, immunization with FR D induced a Th2-biased immune response in BALB/c mice as determined by the numbers of splenic CD4+ cells secreting interleukin 4 and interferon-gamma according to limiting dilution analyses. In addition, FR D caused significant disease exacerbation in parasite-infected susceptible mice as assessed by the local lesion development and the numbers of parasites in lymph nodes and spleen. This effect was observed after local subcutaneous application of FR D as well as after systemic immunization (intrasplenic or intraperitoneal). Transfer experiments revealed, that the disease-aggravating effect of FR D was mediated by CD4+ T cells. From these results it is concluded that leishmanial protein preparations exist that not only fail to induce protective anitparasitic immunity, but can mediate disease exacerbation, independently of the primary application site of the immunogen. The existence of such structures may serve the parasite as a means to evade the host's immune attack and may also have implications for the development of vaccines.

Induction of macrophage membrane interleukin 1 expression by T-cell dependent and T-cell independent pathways is inhibited by cyclosporin A

Here we report that cyclosporin A (CsA) inhibits the induction of membrane interleukin 1 (mIL-1) expression on murine peritoneal macrophages. The inhibition of mIL-1 expression was noted in response to both autoreactive T-cell lines specific for class I or class II MHC determinants as well as bacterial endotoxin. The macrophages were the direct target of this inhibition as shown by pretreating T cells and macrophages separately with CsA. The effective suppression by CsA of endotoxin-induced mIL-1 expression was dependent not only on the concentration of endotoxin employed, but also on the relative time of addition of CsA and endotoxin. Furthermore, CsA pretreatment of macrophages abrogated their ability to stimulate synthesis of IL-4 by a Th2 cell clone. These data suggest that inhibition of induction of accessory molecules such as mIL-1 may be a mechanism by which CsA abrogates the capacity of macrophages to present antigen.

Participation of L3T4 in the avidity, activation and interactions of TH-cell clones

Participation of L3T4 in the avidity, activation and interactions of TH-cell clones

Molecular localization and polymorphism of HLA class II restriction determinants defined by Mycobacterium leprae-reactive helper T cell clones from leprosy patients.

MHC class II molecules carry the restriction determinants (RDs) for antigen presentation to antigen-specific Th lymphocytes. This restriction of T cell activation endows those molecules with a key role in the induction and regulation of antigen-specific immune responses. Moreover, class II molecules are the products of class II immune response (Ir) genes. The polymorphism of these Ir genes leads to genetically controlled differences in immuneresponsiveness between different individuals. An important human example is leprosy, in which HLA class II-linked Ir genes determine the immune response against Mycobacterium leprae, the causative organism of the disease. Since the immune response against M. leprae is entirely dependent on Th cells, the HLA class II-linked Ir gene products may well regulate the immune response by controlling the presentation of M. leprae antigens to Th cells. We therefore have investigated the HLA class II RD repertoire of M. leprae-reactive Th cell clones (TLC) by means of extensive panel and inhibition studies with fully class II-typed allogeneic APCs and well-defined HLA class II-specific mAbs. The TLC studied (n, 36) proliferated specifically towards M. leprae, produced IFN-gamma upon activation, and had the CD3+CD4+CD8- phenotype. The results show in the first place that the majority of the RDs for M. leprae reside on DR and not on DP or DQ molecules. This indicates a major role for DR molecules in the immune response to M. leprae and suggests that these molecules are the main products of M. leprae-specific Ir genes. Furthermore, since the expression of DR molecules is much stronger than that of DP and DQ molecules, these findings suggest that the localization of RDs for M. leprae on class II molecules correlates with the quantitative expression of these molecules. The observation that the RDs on DR molecules coded by a DR4 haplotype were situated only on those DR molecules that are known to be highest in expression can be explained in the same way. Second, four distinct RDs related with but not identical to the Dw13 allodeterminant were carried by the DR+DRw53- (alpha beta 1) molecules of a DR4Dw13 haplotype. Since the known amino acid residue differences between the allelic DR4 related Dw beta 1 chains cannot explain the observed RD-polymorphism, this observation suggests that multiple distinct RDs unique for the DR4Dw13 haplotype are expressed by these molecules. Only 2 of 36 TLC were not restricted by DR.(ABSTRACT TRUNCATED AT 400 WORDS)

T Helper Cell Control of B Cell Development and Isotype Expression

The supernatant of the T helper cell clone 52.3 (52.3-SN) was shown to induce polyclonal activation of resting B cells. 52.3-SN acts on most small B cells and through the allogeneic barrier. This supernatant induces cell size increase, RNA and DNA synthesis, and appearance of interleukin-2 and transferrin receptor. These results are interpreted as indicating the existence of a B Cell Activating Factor (BCAF) acting on resting B cells in an MHC-unrestricted way.TH cells can be obtained in an intermediate state of activation where they secrete lymphokines leading to B cell proliferation and not the biological activities leading to plasmocyte development. TH cell clones can induce sIgG− and sIgA− unprimed B cells to switch and express all classes and subclasses of immunoglobulin. The bulk of the response consists of IgM. Among the non IgM isotypes, IgG1 and IgA predominate. The supernatants prepared from TH cells reproduce these effects.

Influence of helper T cells on the expression of a murine intrastrain crossreactive idiotype.

The requirement for idiotype-specific helper T (Th) cells in the generation of a major intrastrain crossreactive idiotype was investigated. This idiotype, designated CRIA, is associated with a large proportion of anti-p-azobenzenearsonate (anti-Ar) antibodies in A/J mice. Secondary in vitro responses were studied. Using carrier-primed heterogeneous Th-cell populations, it was found that CRIA expression is determined by the mouse strain that provides the responding B cells and is independent of the strain of the Th cells functioning in vitro. Thus, A/J or A.BY (Ighe) B-plus-accessory-cell populations, primed in vivo to keyhole limpet hemocyanin-Ar (KLH-Ar), generated CRIA-dominant responses in vitro in the presence of KLH-Ar regardless of whether the KLH-primed Th cells were derived from CRIA+ strains (A/J or A.BY, Ighe) or CRIA- strains (B10.A or C57BL/10, Ighb). Further, when major histocompatibility complex-restricted, KLH-specific Th-cell clones were used, the CRIA dominance of the Ar-specific responses was again determined by the strain providing B plus accessory cells. Similar levels of expression of CRIA in Ar-specific antibodies were generated in the presence of heterogeneous or cloned Th cells. The results suggest that there is no absolute requirement for idiotype-specific Th cells in generating an Ar-specific secondary antibody response in vitro.

T cell regulation of B cell activation: MHC-restricted T augmenting cells enhance the B cell responses mediated by MHC-restricted cloned T helper cells.

The present studies demonstrated that unprimed populations of Lyt-1+2- T cells are able to augment the responses generated by optimal numbers of antigen-specific and MHC-restricted cloned TH cells. The TA cells function early in the course of B cell responses. Responses mediated by (A X B)F1 (B + accessory) cells and parentA restricted cloned TH cells are augmented by TA cells from (A X B)F1 leads to parentA radiation bone marrow chimeras, but not by TA cells from (A X B)F1 leads to parentB chimeras. Thus, TA cell activation and function are MHC-restricted, but this restriction is not related to recognition of genotypically expressed B cell and/or accessory cell MHC products alone. Rather, TA cell function is intimately related to the MHC-restricted interaction between TH cells and responding (B + accessory) cells. Specifically, it was shown that TH cell interaction with B cells via an MHC-restricted and carrier-hapten-linked pathway is required for the function of TA cells expressing the same MHC restriction. Moreover, neither MHC homology nor TH cell recognition of MHC determinants on TA cells is required for TA cell function, and the activity of TA cells is not demonstrably influenced by non-MHC genes. It was further demonstrated that the function of cloned TH cells in B cell activation consists of at least two distinct components, one is radiosensitive and the other is radioresistant. TA cells are able either to replace the radiosensitive function of cloned TH cells directly or to bypass this requirement through an alternative pathway.

The IgA response: inductive aspects, regulatory cells, and effector functions.

In this review, we have emphasized our current studies on the inductive aspects of the IgA immune response and homeostatic mechanisms involved in the induction of oral tolerance. By use of unique inbred mouse strains in restricted microbial environments, we have provided evidence for a central role of LPS in systemic unresponsiveness to orally encountered antigens. We have continued studies on characterization of GALT lymphoreticular cell types, including accessory cells, regulatory T-cells, and precursor IgA B-cells. We have placed recent emphasis on characterization of antigen-specific Th-cell clones derived from murine PP, which preferentially support IgA isotype responses. Relevant areas for continued research have been emphasized in this review.