Dual genetic regulation by H-2 and non-H-2-linked genes of the magnitude and isotypic pattern of the dinitrophenyl antibody response to DNP-GAT conjugate.
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Biomedical subjects
Publications and source records attributed to J Theze.
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In vitro cooperative responses between hapten-primed anti-Thy-1.2 plus C-treated spleen cells and carrier-primed T cells have different isotypic patterns depending on the source of the T helper cells. T helper cells from primed lymph node induce IgG1, IgG2a, and IgG2b PFC responses, whereas T helper cells from primed spleen induce only an IgG1 type of response. The addition of activated spleen cells to the lymph node cells suppresses their ability to generate IgG2a and IgG2b PFC responses. The suppressor cells involved have been characterized. Functionally, they appear as nonantigen specific and isotype specific, because they never reduce the IgG1 response. They are Lyt-2.2 positive, Lyt-1 negative, and radiosensitive. Their relative resistance to anti-Thy-1.2 plus C treatment indicates that they express low amounts of this antigen or that they are heterogeneous concerning the expression of Thy-1.2.
We obtained T cell clones specific for poly (Glu60, Ala30, Tyr10) (GAT) from GAT-primed lymph node cells of BALB/c mice. The clones consisted of helper cells that were carrier-specific and H-2-restricted and required a hapten-carrier bridge to activate B cells. They may be considered TH1 cells. They could all induce antibody responses of various isotypes in adoptive transfer in vitro. Isotypic patterns of the responses obtained with the clones were identical to those obtained with polyclonal T cells and were not dependent on the number of T helper cells added to the culture. Therefore, our observations demonstrate that classic TH1 cells do not regulate isotype expression on secondary responses. The expression of idiotype-like determinants on the GAT-specific clones was assayed. A syngeneic anti-idiotypic serum (B658), prepared in BALB/c mice against BALB/c anti-GAT antibodies previously characterized, was shown to block specifically the helper activity of GAT-specific T cell lines from BALB/c mice. Under the same experimental conditions the activity of none of the BALB/c clones was affected by serum B658.
NH2-terminal sections of H and L chains isolated from five monoclonal anti-GAT antibodies derived from BALB/c mice have been sequenced upon to residue 43. Four among these five antibodies, sharing similar public idiotypic determinants, possess extremely conserved sequences, both for the H, which is apparented to the VH II type, and the L chains, which belong to the V kappa I subgroup. VH sequences are identical up to residue 43 and contain the common sequences (residues 1 to 32) defined for the H chains derived from the DBA/2 IgM anti-GAT monoclonal antibodies. Light chains are also remarkably conserved, a rather unusual situation for kappa-chains. The fifth antibody that expresses only part of the public idiotypic determinants contains very distinctive H and L chains. Its heavy chains are close to the VH I subgroup, whereas its kappa-chains permit definition of a new V kappa subgroup. The repertoire appears to be highly conserved between BALB/c and DBA/2 mice, and does not seem larger in IgG than in IgM antibodies. This latter observation does not speak in favor of a switch-linked amplification of diversity.
Specific anti-dinitrophenyl (DNP) response to DNP-conjugated L-glutamine60-L-alanine30-L-tyrosine10 (DNP-GAT) was obtained in GAT-responder mice by using synthetic N-acetylmuramyl-L-alanyl-D-isoglutamine (MDP) as adjuvant. Significant levels of anti-DNP antibodies were observed during a secondary response to DNP-GAT, when both antigen and MDP were used for priming. In this system, MDP was able to prime the carrier-specific T cells but not the hapten specific B cells. The study of the isotypic pattern of the anti-DNP response shows that MDP stimulates only the appearance of specific anti-DNP IgG1 plaque-forming cells. Anti-DNP plaque-forming cells were stimulated in animals primed with DNP-GAT in Freund's complete adjuvant or in Maalox-pertussis and used as control IgG1, IgG2a, and IgG2b.
The synthetic terpolymer of L-glutamic acid60-L-alanine30-L-tyrosine10 (GAT) induced anti-GAT antibodies in D1.LP mice. Some of these antibodies shared common idiotypic specificities. The interaction between idiotypic antibody and anti-idiotypic antiserum was specifically inhibited by GAT and the closely related polymer of L-glutamic acid and L-tyrosine (GT) but not by the copolymer of L-glutamic acid and L-alanine (GA). The binding of labeled GAT by the anti-GAT antibodies was also specifically inhibited by the anti-idiotypic antiserum. These results indicated a close association of idiotypic determinants with the antibody-combining sites. Furthermore, such active site-related common idiotypic specificities were found in individual sera of both responder mice (D1.LP) and nonresponder mice (DBA/1) immunized with GAT and GAT-MBSA, respectively, indicating that both responder and nonresponder mice carry B cells with common clonotypes.
The activity of suppressor T cells has been demonstrated in almost every phase of the immune response. These regulatory cells modulate both humoral and cell-mediated immunity utilizing antigen-specific and nonspecific mechanisms. For comparative purposes two murine models are described, the nonspecific suppressor T cell stimulated by the mitogen concanavalin A and the antigen-specific suppressor T cell stimulated by injection of the synthetic terpolymer acid 60-L-alanine30-L-tyrosine10 (GAT) in nonresponder mice. These two T cells are similar to expression of Ly alloantigens, ability to inhibit antibody responses, and the mediation of suppression, at least in part, by soluble products. However, differences in radio-resistance and antigenic specificity of the suppressor T cells, as well as differences in molecular characteristics of the soluble factors and their targets suggest that these T cells regulate the immune response by different mechanisms. The relationship of these two suppressor T cells to other nonspecific and antigen-specific suppressor T cells is discussed.
The induction of new suppressor T cells (Ts2) by suppressive extracts (TsF) from L-glutamic acid50L-tyrosine50 (GT) nonresponder mice was examined. Incubation of normal spleen cells with allogeneic GT-TsF for 2 days in vitro led to the generation of Ts2 cells able to suppress subsequent responses to the immunogen GT-methylated bovine serum albumin (GT-MBSA) in vivo. This induction occurred efficiently when TsF donor and target cells differed at all of H-2, including the I-J subregion. B10.BR (H-2k) GT-TsF, adsorbed on, then acid eluted from GT-Sepharose and anti-I-Jk [B10.A (3R) anti-B10.A (5R)]-Sepharose in a sequential fashion could induce BALB/c (H-2d) spleen cells to become Ts2 only if nanogram quantities of GT were added to the purified GT-TsF. This indicates a requirement for a molecule or molecular complex possessing both I-J determinants and antigen (GT)-binding specificity, together with GT itself, for Ts2 induction. The induced Ts2 are I-J+, since their function can be eliminated by treatment with anti-I-Jk plus C. These I-J determinants are coded for by the precursor of the Ts2 and do not represent passively adsorbed, I-J coded TsF, since anti-Ijk antiserum [(3R X DBA/2)F1 anti-5R] which cannot recognize the BALB/c (I-Jd) TsF used for induction still eliminates the activity of induced A/J (I-Jk) Ts2. These data provide further evidence for and information about the minimum of two T cells involved in antigen-specific suppressor T cell systems.
The GAT-specific suppressor T-cell factor (GAT-TsF) extracted from lymphoid cells from GAT-primed, nonresponder DBA/1 mice has been partially characterized. It is a protein that has affinity for GAT and determinants encoded by the I region of the H-2 complex. On the basis of specificity and avidity, GAT-TsF resembles anti-GAT-MBSA antibodies produced by DBA/1 mice in spite of the fact that it is too small to be classical antibody and has no constant-region determinants of heavy or light chains. Further, GAT or a fragment of GAT is associated with the GAT-TsF. GAT-TsF has been partially purified from the crude extract by absorption to GAT-Sepharose and elution with 0.4 to 0.6 KCl. GAT-TsF purified on the basis of its affinity for GAT bears I-region determinants but not detectable GAT or GAT fragment.
Through the use of specific immunoadsorbent columns, it is shown that Escherichia coli aspartokinase I-homoserine dehydrogenase I, aspartokinase II-homoserine dehydrogenase II, aspartokinase III, and homoserine kinase, enzymes involved in the same complex biosynthetic pathway, share antigenic determinants. This raises the question of a common origin for the four cibtenoirart kinases. (Aspartate kinase or ATP:L aspartate 4-phosphotransferase, EC 2.7.2.4; homoserine dehydrogenase or Lhomoserine:NADP oxidoreductase, EC 1.1.1.3; homoserine kinase or ATP:L-homoserine O-phosphotransferase, EC 2.7.1.39.)
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Two murine T cell lines (C30.1 and Line 1) were used to study the expression of the p55 interleukin-2 receptor gene. C30.1 is an IL-2-dependent T cell line that can be stimulated for a short period of time by IL-4. Line 1 cells are propagated in IL-4 but they also proliferate in response to IL-2. In both cell lines stimulation by IL-2 leads to a strong induction of p55 IL-2 receptor mRNA while stimulation by IL-4 leads only to a very moderate increase in expression of this mRNA. The induction of p55 IL-2 receptor mRNA by IL-4 is comparable to that of beta-actin mRNA. These data confirm that IL-2 upregulates p55 IL-2 receptor gene expression while IL-4, which also activates T cells, does not lead to specific induction of this gene. We have also determined the transcription initiation sites utilized by the p55 IL-2 receptor gene in C30.1 and Line 1 cells. Seven sites were identified, one of which predominates. Resting cells, or cells stimulated with IL-2 or IL-4, display the same pattern of transcription site utilization.
We have investigated the effects of supernatants from human placenta explant cultures on lymphokine dependent T cell proliferation using mainly the CTLL-2 reference murine cell line. A profound, dose dependent, inhibition of both IL-2 and IL-4-dependent cell proliferation was observed. In addition, supernatants from human placental cultures inhibited B cell proliferation, IL-5-driven proliferation of B13 cells, and IL-6-induced proliferation of 7TD.1. Conversely, the supernatants from human placental cultures enhance the growth of human embryonic fibroblasts. Characterization of the material by HPLC yielded a 68-70 kDa peak at pH 7.4, but the peak activity was at a smaller molecular weight (20-25 kDa) if pH 2.9 acetic acid, KC1 buffer was used. The same fractions were inhibitory for IL-2 and IL-4 driven proliferation of CTLL-2 cells. We conclude that the substances tested are acting as a general growth inhibitor for lymphocytes.
The helper activity of GAT (L-glutamic acid 60-L-alanine 30-L-tyrosine 10)-specific T cells from BALB/c mice has been characterized. The isotypic pattern of the PFC response (plaque-forming cell) obtained in vitro is different depending on the origin of the T helper cells. T helper cells from primed spleen induce a very predominant IgG1 PFC response while T helper cells from primed lymph node induce IgG1-IgG2a- and IgG2b-PFC responses. When primed spleen cells are added to primed lymph node cells in vitro, the IgG2a- and IgG2b-PFC responses are abolished. This results suggests the presence of isotype-specific suppressor cells in the spleen of immunized mice.
Cytokines form a complex network of interaction which is crucial in maintaining the homeostasis of the immune and hemopoietic organs and in controlling the balance between different responses of these two systems. The International Congress held in Osaka (17-21 October 1993) was an opportunity to summarize all the information accumulated in the field of cytokines and to evaluate the impact of new approaches and ideas which have invaded laboratories these last two years (the 7th International Congress was held in San Antonio in 1991). In this paper, we will summarize some striking points that were presented in Osaka.