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.     

Antigen-induced T suppressor cells regulate the autoreactive T helper-B cell interaction

The T suppressor (Ts) cell population that functions to regulate antigen-specific MHC-restricted T helper (Th)-B cell interactions also regulates the activation of B cells by cloned autoreactive Th cells. Activated Ts cells were generated by in vivo priming and restimulation in vitro with high concentrations of the specific priming antigen. Once generated, this Ts population inhibits the Th-dependent activation of primed B cells by both antigen-specific and autoreactive T cells in an antigen-nonspecific manner. This suppression requires the participation of both Lyt-1+2- and Lyt-1-2+ T cells. It was also demonstrated that accessory cells were required for the induction of Ts cells. Moreover, the generation of suppression was MHC-restricted and required the recognition by T cells of Ia antigens on accessory cells. These studies demonstrate that the same or a very similar Ts cell population can function to inhibit the activation of B cells by antigen-specific as well as autoreactive T cells.     

Prevention of granuloma development in the mouse by using T cell hybridoma products

The ability of an azobenzenearsonate (ABA)-specific suppressor T cell factor, a soluble extract from first order suppressor T cells (Ts1), and suppressor molecules produced by a long-term T cell hybridoma to regulate ABA-specific granuloma formation was studied. ABA-derivatized syngeneic spleen cells (ABA-SC) administered subcutaneously induced persistent delayed-type hypersensitivity (DTH) responses, detected by footpad swelling and hapten-specific granuloma formation by 72 and 96 hr after challenge with ABA-bovine serum albumin coupled to polyacrylamide beads (ABA-BSA-PAB). Soluble factors from ABA-specific Ts1 prevented DTH and granulomatous development after subcutaneous administration of ABA-SC. Moreover, the in vivo administration of a factor that is derived from a Ts1 functioning hybrid cell line induced a second set of suppressor cells (Ts2) that upon transfer to syngeneic ABA-primed mice were able to inhibit granuloma formation in the footpad, as well as in the gastrointestinal tract after challenge with ABA-BSA-PAB. These experiments demonstrate the dependence of the granulomatous reaction on T cell-mediated events, as well as the potential therapeutic efficacy of an antigen-specific suppressor T cell factor and a hybridoma T cell product in limiting antigen-specific granuloma formation in vivo.     

Characterization of suppressor T cell clones derived from a mouse tolerized with conjugates of ovalbumin and monomethoxypolyethylene glycol.

The induction of antigen-specific tolerance in mice by conjugates of ovalbumin (OVA) and monomethoxypolyethylene glycol (mPEG) previously had been shown to be associated with the generation of antigen-specific suppressor T (Ts) cells. For the elucidation of the nature of these Ts cells, five nonhybridized OVA-specific Ts cell clones were generated from the spleen cells of a BDF1 mouse which had been immunosuppressed by the tolerogenic conjugate, OVA(mPEG)12. The cloned Ts cells were maintained in vitro by periodic stimulation with OVA and feeder cells and were able to suppress the in vitro antibody production in an OVA-specific and MHC class I (H-2Kd or H-2Dd)-restricted manner. All these Ts cell clones were shown to be Thy1.2+, CD4-, CD5-, CD8+, and to express CD3 and the alpha beta heterodimer of the T cell receptor. The cell-free extracts of these cells contained soluble suppressor factors which could mimic in vitro the suppressive activity of the intact cells. In contrast to cytotoxic T lymphocytes (CTL), none of the cloned Ts cells were endowed with cytolytic activity as revealed in the perforin-mediated microhemolysis and in the 18-hr51Cr release assays. These results demonstrate that (i) OVA-specific Ts cell clones can be generated from mice pretreated with OVA(mPEG)12 by employing conventional T cell culture techniques, and (ii) these Ts cells are functionally different from conventional CD8+ CTL.     

Lyt-2+ suppressor T cells are resistant to anti-Lyt-2 antibody blocking

Lyt-2 molecules play a role in antigen recognition by cytotoxic T lymphocytes (CTL). In an attempt to determine whether Lyt-2 molecules play a similar role in suppressor T cell (Ts) functions, the effect of anti-Lyt-2 antibodies on Ts generation and effector activity was studied. Allospecific Ts were induced in allogeneic mixed lymphocyte cultures (MLC). Anti-Lyt-2 antibodies added to MLC in the absence of complement abolished CTL generation, but had no effect on concomitant induction of Ts. In a different experimental system, allospecific Ts were induced in cultures treated with pyrilamine, which blocks generation of CTL but allows differentiation of Ts. The addition of anti-Lyt-2 antibodies to pyrilamine-treated MLC resulted in unaffected induction of Ts. It was further demonstrated that the effector activity of Ts was as resistant to anti-Lyt-2 antibodies as their induction, in contrast to the cytolytic activity of CTL, which was inhibited by the same antibodies. Ts in the present experimental system were Lyt-2+ antigen-specific cells. It therefore appears that Lyt-2 molecules, although expressed on both CTL and Ts, are involved in CTL activity, but do not play an essential role in Ts function.     

The role of suppressor factors in the regulation of immune responses by ultraviolet radiation-induced suppressor T lymphocytes. II. Activity of suppressor cell culture sonicates

The purpose of this study was to determine whether multiple types of suppressor factors play a role in the regulation of immune responses by ultraviolet radiation-induced suppressor T lymphocytes (UV Ts). The UV Ts were induced by applying contact allergens to the ventral, unirradiated skin of mice exposed 5 days earlier to UVB radiation. Previous studies indicated that supernatants from cultures containing UV Ts, normal lymphocytes, and hapten-modified cells suppressed contact hypersensitivity (CHS) in vivo and cytotoxic T lymphocyte (CTL) generation in vitro in a hapten-specific manner. In this report, cell-free lysates from sonically disrupted UV Ts were examined for their ability to suppress these responses. When lysates were injected into normal animals at the time of sensitization, they inhibited CHS in a hapten-nonspecific manner. In addition, the lysates suppressed not only the induction but also the elicitation of CHS, and they suppressed the generation of CTL. Lysates prepared from spleen cells obtained from non-UV-irradiated mice or UV-irradiated, unsensitized mice failed to inhibit either response. Moreover, in contrast to the lysates, the hapten-specific UV Ts culture supernatants inhibited the induction but not the elicitation of CHS. These results suggest that both hapten-specific and nonspecific inhibitory factors may participate in the regulation of immune responses by UV Ts.     

Characterization of two suppressor cells that together prevent in vivo development of cytolytic T cells to hapten-altered self

Two suppressor cell populations that interact to down-regulate in vivo development of the cytolytic T-cell (CTL) response to trinitrophenyl-modified syngeneic spleen cells (TNP-SC) have been further characterized. Suppressor cells induced by the iv injection of trinitrophenyl-modified syngeneic spleen cells possess Thy 1.2 antigen. Their precursors are insensitive to pretreatment of host animals with cyclophosphamide (CY). Suppressor cells that arise after dermal sensitization with trinitrochlorobenzene are also Thy 1.2 antigen positive but their precursors are sensitive to pretreatment with CY. These characteristics of the two suppressor T cells (Ts) are identical to those of the two Ts that are generated by similar methodologies and that together suppress contact sensitivity (CS) to picryl chloride. Neither the CS nor CTL response was suppressed when host animals possessed only one set of Ts. In contrast to suppression of CS at the efferent phase, development of CTL was suppressed only when the two Ts were present early during sensitization (afferent phase). Since the results point to several similarities between the two sets of Ts that are active in the down-regulation of the CS and CTL responses, it is suggested that the two dissimilar immune responses directed to the same hapten, namely CS and CTL, may be controlled by the same suppressor cells. Since it appears that the two sets of Ts interact to affect different phases of the CS and CTL responses, down-regulation of each must be accomplished through different mechanisms.     

Antigen-specific suppression of cytotoxic T cell responses: an idiotype-bearing factor regulates the cytotoxic T cell response to azobenzenearsonate-coupled cells

When A/J mice are injected subcutaneously with azobenzenearsonate- (ABA) coupled spleen cells, their splenocytes contain primed ABA-specific cytotoxic T lymphocyte (CTL) precursors. Animals that are not primed in vivo do not develop vigorous CTL activity when assessed after in vitro culture with ABA-coupled stimulators. Suppressor molecules derived from ligand-induced first-order ABA-specific suppressor T cells were evaluated for their ability to limit cytolytic T cell development. We have shown that an idiotype-bearing, hapten-specific suppressor factor suppresses priming for CTL in an H-2-unrestricted but allotype-restricted manner. The implication of these studies to regulatory networks is discussed.     

T cells in B cell chronic lymphocytic leukemia. II. Lack of antigen-specific T suppressor cells and their progenitors

Nylon wool-purified T cells (Tn) of two patients with chronic lymphocytic leukemia of the B cell type were phenotyped and tested in various assays for antigen-specific T helper (Th), T suppressor effector (Tse), T suppressor precursor (Tsp), and T suppressor inducer (Tsi) function. Antigen-specific Th as well as Tsi activity could be effectively generated. Although phenotypically CD8+ T cells, carrying the receptor for the Fc part of IgG, were present in mononuclear blood cells and Tn fractions, no antigen-specific Tse cell activity could be induced. In addition, Tsp cells were found to be functionally absent. These findings are discussed in relation to a tumor-induced limited heterogeneity within the T suppressor (Ts) cell compartment.     

T cell regulation of B cell activation. An antigen-mediated tripartite interaction of Ts cells, Th cells, and B cells is required for suppression

To determine the requirements underlying the antigen specificity observed in T cell-mediated immune response suppression, cloned major histocompatibility complex (MHC)-restricted T suppressor (Ts) cells specific for keyhole limpet hemocyanin (KLH) and cloned MHC-restricted T helper (Th) cells specific for fowl gamma-globulin (FGG) were employed to study the regulation of trinitrophenyl (TNP)-specific B cell responses. Neither antigen bridging between Ts cells and Th cells (FGG=KLH) nor bridging between Ts cells and B cells (TNP-KLH) was sufficient to allow suppression; a mixture of FGG=KLH and TNP-KLH was also insufficient for suppression. In contrast, suppression was induced by KLH-specific Ts cells only when suppressor determinants (KLH), helper determinants (FGG), and B cell determinants (TNP) were covalently linked on the same molecule (TMP-FGG)=(TNP-KLH) or TNP-(FGG=KLH)). These findings imply that a tripartite antigen-mediated interaction of Ts cells, Th cells, and responding B cells is necessary for the mediation of this antigen-specific suppression.     

Regulatory mechanisms in cytotoxic T lymphocyte development. II. Dissociation of in vitro cytotoxic T-lymphocyte and suppressor T-cell activities with L-ornithine

L-ornithine was found to differentially affect the induction of allospecific cytotoxic T lymphocytes (CTL) and suppressor T cells (Ts). At a concentration of 10 mM, ornithine inhibited the development of CTL in a mixed-leukocyte culture (MLC). This same population of cells suppressed the generation of CTL when irradiated and cocultured with fresh syngeneic lymphocytes and alloantigen. Suppression was mediated by Lyt-1-2+ cells and was antigen specific. Suppression was abrogated when IL-2 (10 U/ml) was added to the cocultures, but could not be reversed by increasing the antigen dose. Ornithine was not toxic to CTL precursors but rather arrested their development. Cells from MLC plus ornithine developed CTL activity within 2 days of transfer to secondary cultures in the absence of ornithine. Development of CTL effector cells (CTLe) was augmented by but did not require exogenous IL-2. Generation of CTLe from the MLC plus ornithine population was radiation sensitive and could be inhibited by reexposure to ornithine, even in the presence of IL-2. Thus, Lyt-1-2+ T cells allostimulated in vitro in MLC plus ornithine and lacking CTL activity convey radiation-resistant, antigen-specific suppression.     

The role of suppressor factors in the regulation of immune responses by ultraviolet radiation-induced suppressor T lymphocytes. I. Activity of suppressor cell culture supernatants

The purpose of this study was to determine whether soluble suppressor factors are involved in the regulation of immune responses by ultraviolet radiation-induced suppressor T lymphocytes (UV Ts). The UV Ts were induced by applying contact allergens to the ventral, unirradiated skin of mice that had been exposed 5 days earlier to UVB radiation. Supernatants from cultures that contained a mixture of UV Ts, normal responder lymphocytes, and hapten-modified stimulator cells were injected iv into normal recipients at the time of sensitization; they inhibited the induction of contact hypersensitivity (CHS) in vivo in an hapten-specific manner. The supernatants similarly suppressed the generation of specific cytotoxic T lymphocytes (CTL) in vitro. Moreover, supernatants from cultures that contained either UV Ts alone or UV Ts in combination with either the responder or the stimulator cells failed to suppress the CHS and CTL responses. These results suggest that hapten-specific inhibitory factors may participate in the regulation of immune responses by suppressor cells generated by epicutaneous sensitization of UV-irradiated mice.     

Down-regulation of cytotoxic T lymphocyte development by a minor stimulating locus-induced suppressor cascade that involves Lyt-1+ suppressor T cells, IA- macrophages, and their factors

Down-regulation of the development of CTL has been studied in mice both in vivo and in vitro. To generate CTL to hapten-altered self Ag in vivo, an immunization protocol has been used in which the host's Th cells are stimulated by a minor locus histocompatibility Ag (Mlsd) and its precursor CTL are activated by trinitrophenylated syngeneic spleen cells. Injecting the H-2 compatible Mls-disparate spleen cells along with the TNP-coupled self cells into the hind paws causes TNP-self specific CTL to appear in popliteal lymph nodes within 5 days. We have previously reported that inducing Ts cells by i.v. injecting Mlsd-bearing cells prevents in vivo generation of TNP-self specific CTL after immunization in this way. Here the induced Ts cell as well as the mechanism by which it functions have been further examined. The suppression was seen to extend to allogeneic as well as TNP-self Ag, provided the Mlsd-tolerized animal was reexposed to Mlsd-bearing cells at the time of immunization for CTL. By transferring the Mlsd-induced suppression adoptively we have learned that the splenic suppressor cell bears Thy-1.2 as well as Lyt-1.1 Ag and inhibits the generation of CTL at the afferent limb. In addition, Mlsd-induced PEC of Mlsd-tolerized mice, but not of normal mice, mediated suppression of development of CTL in vivo. The active cells within the tolerized PEC have been identified as T cells and macrophages (M phi). Furthermore, PEC from mice tolerized to Mlsd suppressed generation of CTL directed toward TNP-self targets in vitro. T cells and M phi separated from PEC of Mlsd-tolerized mice achieved suppression best in culture when present together. In addition, Lyt-1+ splenic cells from tolerized but not normal mice cooperated to down-regulate CTL generation in vitro with peritoneal M phi from either tolerized or normal mice. Supernatants of 24- to 72-h cultures of PEC from tolerized mice were suppressive of CTL generation when incorporated at 40 to 50% of culture volume. Supernatants of T cells from tolerized PEC or spleen were suppressive in culture only when M phi from normal mice were also present. To achieve suppression dialyzed supernatants of M phi from tolerized mice could replace the M phi.(ABSTRACT TRUNCATED AT 400 WORDS)     

Two adjacent epitopes on a synthetic dodecapeptide induce lactate dehydrogenase B-specific helper and suppressor T cells

The outcome of an immune response to the enzyme lactate dehydrogenase B (LDH-B) is determined by the interplay between two types of regulatory T lymphocytes, T helper (Th) and T suppressor (Ts) cells. Most mouse strains are capable of generating Th but not Ts cells, and are therefore high responders to LDH-B in terms of both antibody production and antigen-specific T-cell proliferation. However, in strains expressing the b or k allele at the E beta locus of the major histocompatibility complex (Mhc), Ts cells are induced that partly or totally abrogate the proliferative response of Th cells to LDH-B. As a result, these strains are phenotypically medium (E beta b expressors) or low (E beta k expressors) responders. Because the suppression in the LDH-B system is antigen-specific (i.e. it only affects LDH-B-specific Th cells), it is conceivable that the Th and Ts cells use the antigen itself to communicate with each other. To investigate this possibility, we set out to determine which epitopes of the LDH-B molecule are recognized by Th and Ts cells. On the basis of previous studies, a loop structure extending from residue 211 to residue 224 of pig LDH-B appeared to be preferentially recognized by most Th-type (class II Mhc-restricted, proliferating) clones. By using a synthetic peptide, we demonstrate here that both Th and Ts cells are induced by the 211-222 stretch of LDH-B sequence. The use of two further dodecapeptides, each with a single amino-acid substitution in comparison with the pig 211-222 sequence, has revealed that Th and Ts cells have different fine specificities. Thus the loop appears to have two closely linked, if not overlapping, epitopes, one recognized by Th and the other by Ts cells. This finding is consistent with two possible mechanisms of suppression, namely bridging of Th and Ts cells by antigen and subsequent transmission of a suppressive signal, and competition for antigen between Th and Ts cells.     

Regulation of the activation of cytotoxic T lymphocytes by prostaglandins and antigens

The present study demonstrated the presence of two suppressor circuits in the regulation of the in vitro activation and differentiation of cytotoxic T lymphocytes (CTL); these suppressor circuits were mediated by prostaglandins (PG) and antigens, respectively. In intrinsic suppression, the activation of cytotoxic precursor cells was regulated by the host endogenous production of PG. When the regulation by PG was removed (e.g., using indomethacin), lymphokine-induced cytotoxic cells (LICC) were generated. This activation process can be induced in the absence of antigen or mitogen stimulation. In extrinsic suppression, the presence of antigen induced the generation of antigen-nonspecific suppressor T cells to restrict the expansion of antigen-unrelated cytotoxic lymphocyte clones, whereas the antigen-specific CTL clones were spared. The generation of antigen-specific helper cells further augmented the antigen-specific CTL response. These findings indicate that both antigen specific suppressor T cells and antigen nonspecific suppressor T cells are involved in the regulation of CTL responses. These suppressor circuits not only play an active role in monitoring the activation of CTL clones, they also help to determine the specificity and magnitude of the CTL response.     

Tolerance to Mls-disparate cells induces suppressor T cells that act at the helper level to prevent in vivo generation of cytolytic lymphocytes to hapten-altered self

We have been examining the mechanisms that control in vivo development and down regulation of cytolytic T lymphocytes (CTL) to trinitrophenyl (TNP)-altered self antigens. In vivo generation of hapten-specific CTL requires an auxiliary antigenic stimulus, which can be provided by H-2 compatible but Mls-disparate cells. These experiments were designed to study the effect of tolerization with such Mls-disparate cells on CTL development. C3H/HeN (H-2k, Mlsc ) mice sensitized in the footpads with C3H-TNP spleen cells plus CBA/J (H-2k, Mlsd ) spleen cells develop CTL in the draining lymph nodes that will lyse 51Cr-labeled TNP-modified C3H targets. However, we have found that if C3H/HeN mice are given tolerizing doses of CBA/J spleen cells 5 to 7 days before sensitization, a splenic suppressor T cell (Ts) appears. This Ts will suppress CTL development in its tolerant host, and can be transferred adoptively to function in naive mice. Ts and its precursor are cyclophosphamide insensitive and therefore different from the naturally existing suppressor cell present in mice. When triggered by cells with Mlsd , the Ts produces a factor (TsF) that hinders helper factors from functioning in an in vitro CTL assay. Furthermore, TsF acts to prevent utilization of IL 2 by an IL 2-dependent cell line. Thus, evidence has been provided that the in vivo generation of CTL toward hapten-altered self can be down regulated at the level of helper signals by a Ts. The latter is inducible by the Mls-disparate cells that are needed at a different site to trigger the helper factors in this CTL system.     

Regulation of myelin basic protein-specific helper T cells in multiple sclerosis: generation of suppressor T cell lines.

Suppressor T cell (Ts) lines specific for myelin basic protein (MBP)-reactive helper T cell (Th) clones were generated from two patients with multiple sclerosis (MS) following a primary culture of peripheral blood mononuclear cells (PBMC) with MBP and cyclosporine A (CsA). These suppressor T cell lines were maintained in culture by alternate stimulation with MBP and antigen-presenting cells (APC). The Ts lines expressed preferentially the CD4 phenotype (5/6 Ts lines tested) and exhibited potent antigen-specific suppressor activity on the proliferation of MBP-specific Th clones and not on the T cell lines with other antigen specificity. For some Ts lines, a Ts-to-Th ratio of 1 was sufficient to inhibit the proliferation of MBP-specific T cells by 90%. The suppressor T cells obtained were weakly responsive to MBP and required the presence of the autologous PBMC for proliferation. Furthermore, proliferation of these suppressor T cell lines was restricted by HLA-DR molecules (for CD4+ Ts lines) and HLA class I (for a CD8+ Ts line). The suppressor T cell lines generated and the techniques described in this study may be helpful in our understanding of the events involved in the immune regulation in MS and other autoimmune diseases.     

Downregulation of helper T cells by an antigen-specific monoclonal Ts factor.

The findings of previous studies in this laboratory demonstrating that conjugates of human monoclonal (myeloma) IgG (HIgG) and monomethoxypolyethylene glycol (mPEG) were able to induce in mice antigen-specific tolerance and CD8+ suppressor T (Ts) cells were confirmed in the present study. An extract (TsF) of a nonhybridized clone of Ts cells (viz., clone 23.32), which had been derived from spleen cells of mice tolerized with HIgG(mPEG)26, was shown to possess antigen-specific suppressive activity. This monoclonal TsF was able to specifically suppress in vitro antibody formation only if it was present from the beginning of the culture. From the results of the cellular dissection of the system used it was concluded that (i) the TsF had no effect on fully differentiated primed B cells or plasma cells, and (ii) the TsF inactivated carrier-primed Th cells when the culture contained concomitantly naive CD8+ T cells, accessory cells, and antigen. These data support the view that the monoclonal TsF exerted its downregulating effect on Th cells only if it could first interact with a CD8+ T cell, in the presence of accessory cells and antigen.     

Suppression of cytotoxic response to histoincompatible cells. I. Evidence for two types of T lymphocyte-derived suppressors acting at different stages in the induction of a cytotoxic response.

Spleen and thymus cell populations from normal or allograft tolerant mice have been cultured for 5 days with specific alloantigens and examined for their reactivity in three assay systems. No consistent correlation was observed between the production of cytotoxic T cells (CTL) in these cultures and the ability of such cultured cells to inhibit specifically a CML response from fresh normal spleen cells directed to the priming alloantigens. Furthermore, suppressor cells measured in this latter assay were apparently distinct from those able to inhibit the production of cytotoxic lymphocyte precursors (CTLp) from bone marrow stem cells in lethally irradiated bone marrow protected mice. Velocity sedimentation experiments confirmed that both the precursor and effector cells for the two suppressor systems were physically separable, and were distinct from CTLp or CTL, respectively. Precursor cells for the two suppressor systems investigated belong to the short-lived cortical thymus cell population.     

Regulation of transplantation immunity in vivo by monoclonal antibodies recognizing host class II restriction elements. II. Effects of anti-Ia immunotherapy on host T cell responses to graft alloantigens

Results of the preceding report demonstrated that in vivo treatment with monoclonal anti-I-A antibodies provided an effective means of prolonging the survival of murine tail skin allografts. The mechanism of antibody action was shown to include the activation of alloantigen-specific suppressor T cells (Ts), although the relationship between Ts expression and graft survival was not determined. This issue was addressed in the current studies through a kinetic analysis of suppressor and effector T cell responses in control and treated allograft recipients. Donor-specific delayed-type hypersensitivity (DTH) and cytotoxic T lymphocyte (CTL) responses were detectable in untreated A/J recipients of B10.A allografts 8 days after transplantation, rising to near maximum levels by day 12. Rejection in these animals occurred by day 11. In contrast, the predominant cellular response of anti-I-A treated animals for 12 days after transplantation was that of transferable suppression, DTH and CTL reactivity not being evident until day 15, coincident with the decay of Ts activity. Rejection in these animals was observed approximately 19 days post-transplant. CTL responsiveness in the latter group could not be reconstituted by the addition of antigen-presenting cells to the secondary in vitro culture system, nor was the CTL deficit due to antibody carry-over. It is considered that the altered expression of effector cell responses to graft alloantigens is due at least in part to the in vivo inhibition of helper T cell activity by anti-I-A-induced Ts, and that rejection in the treated host results from an eventual decline in the functional expression of this regulatory T cell subset.