Autologous rosette-forming T cells regulate responses of T cells. Phenotypic and functional analysis of suppressor cells generated from autologous rosette-forming T cells after autologous mixed lymphocyte reactions.

An average of 5--9% of human peripheral blood of T lymphocytes from rosettes with autologous erythrocytes (ARFT). This population responded only slightly against autologous and allogeneic non-T cells. In contrast, T cells that did not form rosettes with autologous erythrocytes (NRFT) proliferated to a greater degree in auto- and allogeneic mixed lymphocyte reactions (MLR) and also in reactions to trinitrophenyl (TNP) modified autologous non-T cells (TNP-auto-MLR) as compared with ARFT or unfractionated T cells. The ARFT populations could suppress the increased allogeneic (allo)MLR and TNP-auto-MLR of NRFT when the ARFT were added to the NRFT at the beginning of the cultures. Fluorescence-activated cell-sorter (FACS) analysis of these freshly obtained T cell fractions using monoclonal antibodies to subpopulations of T cells did not demonstrate any selective gain or less of T cell subsets in the ARFT and NRFT as compared with unfractionated T cells. But when each T cell fraction was cultured separately for a week in the presence of autologous non-T cells (auto-MLR) and the cells were again analyzed by fluorescence-activated cell sorter, there was an increase in OKT8-positive cells (suppressor/cytotoxic subset) only in the ARFT fraction. The above findings strongly suggest that suppressor T cells are generated from the ARFT fraction during an auto-MLR, these may then regulate the responses on NRFT.     

Regulatory mechanisms in cell-mediated immune responses. IV. Expression of a receptor for mixed lymphocyte reaction suppressor factor on activated T lymphocytes

Suppression of the mixed lymphocyte reaction (MLR) by a soluble factor produced by alloantigen-activated spleen cells requires genetic homology between the factor-producing cells and responder cells in MLR. The ability of lymphocytes used as MLR responder cells to adsorb MLR suppressor factor was tested to investigate the expression of a receptor structure for suppressor molecules. Normal spleen or thymus cells had no effect on suppressor activity. Concanavalin A (Con A)-activated thymocytes, however, effectively removed suppressor activity, suggesting that the receptor is expressed only after activation and is not present or not functional on resting cells. Significantly neither phytohemagglutinin- nor lipopolysaccharide-activated lymphoid cells absorbed the factor. Furthermore, only Con A-activated thymocytes demonstrating genetic homology with the cell producing suppressor factor for H-2 regions to the right of I-E were effective absorbants. Alloantigen-stimulated spleen cells syngeneic to the suppressor cell also removed suppressor activity. These data support an hypothesis that subsequent to stimulation in MLR, T lymphocytes express a receptor, either through synthesis or alteration of an existing molecular structure, which then provides the appropriate site for interaction with suppressor molecules.     

Immunoregulatory T cell circuits in man. Alloantigen-primed inducer T cells activate alloantigen-specific suppressor T cells in the absence of the initial antigenic stimulus

Although alloantigen-specific suppressor T cells are generated in MLR, the cellular signals that lead to activation of suppressor T cells as opposed to cytotoxic T cells are unknown. The current study was undertaken to characterize interactions among T cell subsets involved in the generation of suppressor T cells in MLR. Human peripheral blood Leu-2+ (suppressor/cytotoxic) and Leu-3+ (helper/inducer) T cell subsets were activated with allogeneic non-T cells and then examined for their inductive effects on fresh autologous T cells. Fresh Leu-2+ cells proliferated in response to alloantigen-primed Leu-3+ cells and subsequently suppressed the response of fresh autologous Leu-3+ cells to the original, but not third party, allogeneic stimulator non-T cells. Moreover, only Leu-2+ cells that lacked the 9.3 marker, an antigen present on the majority of T cells including precursors of cytotoxic T cells, differentiated into suppressor cells. The alloantigen-specific suppressive effect of Leu-2+,9.3-cells was not mediated by cytolysis of allogeneic stimulator cells, nor could it be explained by alteration of MLR kinetics. Suppression was observed only when activated Leu-2+ cells were added to fresh MLRs within 24 h of initiation of cultures, suggesting that these cells block an early phase of the activation of Leu-3+ cells in MLR. These results indicate that alloantigen-primed inducer T cells can activate alloantigen- specific suppressor T cells in the absence of allogeneic stimulator cells.     

Autologous Mixed Lymphocyte Reaction in Patients with Hodgkin's Disease: EVIDENCE FOR A T CELL DEFECT

The proliferative response of T lymphocytes cultured with autologous non-T lymphocytes is known as the autologous mixed lymphocyte reaction (MLR). This reaction can be demonstrated reproducibly in healthy individuals and has been shown to generate specific cytotoxic T cells, as well as T cells that regulate antibody synthesis and cell-mediated immunity. In this study, we demonstrate that the autologous MLR is impaired or absent in most patients with Hodgkin's disease regardless of age, sex, pathologic stage, or histologic classification. In 64 patients, the mean autologous MLR was 3,084±1,878 cpm compared to 16,552±6,532 in 29 healthy donors. A defect in autologous MLR was observed in newly diagnosed patients before the initiation of therapy, but was also found in patients without evidence of recurrent disease up to 15 yr after treatment.     

Immunoglobulin secretion in the human autologous mixed leukocyte reaction. Definition of a suppressor-amplifier circuit using monoclonal antibodies

The induction of immunoglobulin (Ig) synthesis in the autologous MLR has an absolute requirement for helper/inducer (Leu-3) T cells, whereas an excess of suppressor/cytotoxic (leu-2) cells suppresses the response. The current study was an effort to assess the immunoregulatory potential to T cells activated in the autologous mixed- leukocyte response (MLR). T cells were cultured with autologous non-T cells for 8-9 d, after which the activated T cells were fractionated into subsets with monoclonal antibodies to T cell markers and HLA-DR antigen. Each population was co-cultured in fresh autologous MLR, and on the 8th day of culture, Ig-secreting cells were measured in a reverse hemolytic plaque assay. The results show that activated Leu-2, DR+ T cells, but neither Leu-2, DR- nor Leu-3 T cells, were at least 50 times more potent as suppressors of IgM and IgG synthesis than fresh Leu-2 cells alone. The activation of this Leu-2, DR+ subpopulation required Leu-3 cells in the primary culture. Furthermore, in the absence of Leu-2 cells in the second culture, little or no suppression was observed, suggesting that the Leu-2, DR+ cells act to amplify or induce suppressor effects of fresh Leu-2 cells. This indicates that at least two distinct subpopulations of Leu-2 cells are required for maximal suppression of an immune response, and that immunoregulatory circuits analogous to those described in the mouse exist in man.     

Suppressor cell activity after concanavalin A treatment of lymphocytes from normal donors

Pretreatment of normal human peripheral blood lymphocytes with the plant lectin, concanavalin A (Con A), results in inhibition of blast transformation and [3H]thymidine incorporation by untreated allogeneic lymphocytes from healthy volunteers donors in one-way mixed leukocyte culture. Similarly, responses to mitogens, certain microbial antigens, and allogeneic lymphocytes are inhibited by Con A-treated allogeneic cells. Con A pretreated autologous lymphocytes can also be induced to manifest suppressor activities. This antimitotic effect occurs without evidence of cytotoxicity and is active on de novo lymphocyte responses and does not require prior sensitization of the cells being tested. Suppression of the lymphocyte response to pokeweed mitogen, a potent B-cell stimulator, by Con A-pretreated suppressor cells was not as consistent as was inhibition of response to other mitogens, including phytohemagglutinin and Con A. Furthermore, suppression of lymphocyte transformation to the microbial antigens, tuberculin purified protein derivative, and Canadida albicans extracts could be similarly induced by Con A pretreatment of either allogeneic or autologous cells. Induction of autologous suppressor activity in lymphocytes from healthy donors is compatible with a model that includes a role for suppressor cells in the modulation of the normal immune response.     

Functional heterogeneities among concanavalin A-activated OKT4+ and OKT8+ cells by using autologous erythrocyte rosette technique.

Normal human peripheral blood T lymphocytes activated by concanavalin A (Con A) were fractionated into OKT4+ and OKT8+ populations by complement-dependent cell lysis using OKT8 and OKT4 antibodies, respectively. By using the preferential ability of some, but not all, Con A-activated T cells to form rosettes with autologous erythrocytes, each population was further divided into autorosetting cells and nonautorosetting cells, and thus Con A-activated OKT4+ autorosetting, OKT4+ nonautorosetting, OKT8+ autorosetting, and OKT8+ nonautorosetting cells were obtained. The immune regulatory function of these populations was then investigated using a pokeweed mitogen-driven B cell plaque-forming cell system. These studies demonstrated that (a) autorosetting cells can exert potent suppressor activity regardless of their phenotypes of OKT4+ and OKT8+ antigens, and fail to help B cell differentiation; suppressor function mediated by these cells is radiosensitive; moreover, receptors for autologous erythrocytes may constitute either the interleukin 2 (IL2) receptors themselves or a component of an IL2 receptor-effector complex involved in modulating the growth signal that IL2 transmits to T cells; (b) OKT4+ nonrosetting cells serve adequately as radioresistant helper cells, but are devoid of suppressor cells; and (c) OKT8+ nonrosetting cells are found to lack either suppressor or helper activity, suggesting that they may belong to a T lymphocyte subset distinct from the subsets related to immune regulation. The results lead us, therefore, to the conclusion that there may exist functional heterogeneities among both the OKT4+ and OKT8+ populations; these heterogeneities can be dissected by virtue of the autologous erythrocyte rosette technique.     

Separation of concanavalin A-induced human suppressor and helper T cells by the autologous erythrocyte rosette technique.

Very few normal human peripheral blood T cells are capable of binding autologous erythrocytes to form rosettes, whereas in the T cell population activated by concanavalin A (Con A) the autorosette levels are markedly enhanced. Fractionation of the Con A-activated T cells with autologous erythrocytes into autorosetting and nonrosetting cells demonstrates that suppressor, but not helper, activity resides in the autorosetting population, whereas the reverse is true of the nonrosetting population. Both these activities are found to be Con A dependent. The Con A-induced human suppressor cells can be identified and separated from the Con A-induced human helper cells by the autorosette technique. Studies on the surface properties of autorosetting and nonrosetting T cells indicate that there is little correlation between the activated suppressor and helper T cell subsets defined by autorosette technique and either those defined by monoclonal antibodies (which are able to distinguish these subsets in the resting but not activated T cells) or those defined by Fc receptors. Since the autorosetting T cell population (which acts as suppressor cells) bears receptors for peanut agglutinin, the nature of Con A-induced human suppressor cells appears to be analogous to that of Con A-induced murine suppressor cells.     

Lymphocyte transformation induced by autologous cells. IV. Human T- lymphocyte proliferation induced by autologous or allogeneic non-T lymphocytes

An autologous mixed lymphocyte reaction was demonstrated between T and non-T lymphocytes. Sheep erythrocyte rosetting was used to separate human lymphocytes into T and non-T lymphoid preparations. Non-T lymphocytes stimulated the proliferation of autologous T lymphocytes. The cell in this preparation that was most stimulatory had the characteristics of a K lymphocyte. The allogeneic mixed lymphocyte reaction was also shown to reflect the proliferation of T lymphocytes stimulated by allogeneic non-T lymphocytes. Proliferation of T lymphocytes in the allogeneic mixed lymphocyte culture probably reflects a response to both foreign histocompatibility determinants and determinants present on non-T lymphocytes. It is suggested that the proliferative response of T lymphocytes to autologous non-T lymphocytes may be a step in the process by which T lymphocytes regulate immunity.     

Impairment of the autologous mixed lymphocyte reaction in atopic dermatitis.

The T cell proliferative response to autologous non-T cells is termed the autologous mixed lymphocyte reaction (AMLR). Recent studies have suggested that the AMLR represents an inducer circuit for the activation of T8+ suppressor/cytotoxic effector cells. Since atopic dermatitis (AD) patients are deficient in T8+ cytolytic T cell function, we investigated the AMLR in AD. When sheep erythrocytes were used to separate T cells from non-T cells, the AMLR was found to be significantly decreased (P less than 0.001) in AD patients (n = 11; delta cpm = 1,550 +/- 393) when compared with normal control subjects (n = 13; delta cpm = 25,819 +/- 4,609). To exclude the possibility that these results were an artifact of the sheep erythrocyte separation, T cells were also separated on a fluorescence-activated cell sorter after treatment of peripheral blood lymphocytes with the OKT3 monoclonal antibody. AD T cells separated by the latter method were also found to have a significantly reduced AMLR response when compared with similarly treated normal T cells. Co-culture studies using cells from AD patients and their HLA identical siblings indicated that the defect resided at the responder T cell level rather than at the stimulator non-T cell level. Co-culture studies revealed no evidence for excessive suppressor cell activity resulting in the decreased AMLR. However, enumeration of T cells reactive with the monoclonal antibody T29, which recognizes a subset of T cells proliferating in the AMLR, demonstrated that AD patients (n = 8; % T29 = 2.5 +/- 0.7) had a significantly decreased (P less than 0.001) number of circulating T29+ T cells when compared with normal controls (n = 8; % T29 = 10.4 +/- 0.8). These studies suggest that a deficiency of T4+ T29+ cells contributes to the deficient AMLR in AD and possibly underlies the abnormalities of T8+ effector cells present in this disease.     

Analysis of the defects responsible for the impaired regulation of Epstein-Barr virus-induced B cell proliferation by rheumatoid arthritis lymphocytes. I. Diminished gamma interferon production in response to autologous stimulation

T cells of patients with rheumatoid arthritis (RA) do not control the rate of B lymphoblast transformation induced by Epstein-Barr virus (EBV) as efficiently as T cells from healthy individuals; thus, lymphoblast cell lines are established more readily in RA lymphocytes in vitro after EBV infection. In the present experiments, we have asked whether this T cell regulation can be reproduced by lymphocytes. We found that normal T cells, activated in allogeneic or autologous mixed leukocyte reactions (MLR), produce lymphokines that inhibit in vitro EBV-induced B cell proliferation. Allogeneic MLR supernatants inhibited EBV-induced DNA synthesis 62 +/- 4% (mean +/- SE) at 10 d post- infection, whereas autologous MLR supernatants suppressed it 50 +/- 3%. RA T cell supernatants produced in an allogeneic MLR suppressed as well as normal T cell supernatants (64 +/- 5% inhibition). In contrast, supernatants from RA autologous MLR had little inhibitory activity. EBV- induced DNA synthesis at 10 d was reduced only 8 +/- 3%, compared with the 50 +/- 3% suppressive activity of normal autologous MLR supernatants. The magnitude of the proliferative responses in the autologous MLR regenerating the lymphokines was similar in the normal and RA populations. After depletion of adherent cells from the RA auto- MLR stimulators, supernatant inhibitory activities increased to normal levels (from 11 +/- 6 [SE] to 52 +/- 6% [SE]). The inhibitory factor involved in the regulation of in vitro EBV infection is a protein with a molecular weight of approximately 50,000. Its activity is eliminated by hearing at 56 degrees C and by exposure to acid at pH 2. The inhibitory activity is blocked by mixing the MLR supernatants with a polyvalent antisera or monoclonal antibodies specific for human gamma interferon. Gamma interferon produced by activating T cells in allo- or auto-MLR can reproduce T cell-mediated regulation of EBV-induced B cell proliferation, and the failure of RA auto-MLR to generate that lymphokine parallels the defective T cell regulation of EBV-induced B cell proliferation characteristic of RA lymphoid cells.     

Effect of BM 12 531 on in vitro induction of suppressor cells by concanavalin A in cancer patients and normal healthy donors

The effects of BM 12 531, a 2-cyanaziridinyl derivative, on in vitro generation of Con A induced suppressor cells as well as generation of spontaneous suppressor cells in peripheral blood lymphocytes of 9 chronic myeloid leukemia (CML) patients in remission, 9 patients with active Hodgkin's disease (HD) and 12 normal healthy donors were studied. The suppressor cells generated spontaneously and with Con A, in the presence and absence of the drug, were tested for their modulating effect on mitogenic (PHA) response of autologous lymphocytes. The results indicate that the addition of the drug at the time of generation of suppressor cells decreased the spontaneous suppressor cell activity in 2/4 healthy donors, 3/7 CML patients and 4/6 HD patients. Con A induced suppressor cell activity generated in presence of the drug was significantly reduced in 7/12 healthy donors, 6/9 CML patients and 3/9 HD patients.     

Effect of oral colchicine on T cell subsets, monocytes and concanavalin A-induced suppressor cell function in asthmatic patients

Asthmatic patients have a deficiency of concanavalin A-(Con A) induced suppressor cell function. We tested whether oral colchicine 0.5 mg twice daily for 7 days could correct this immunoregulatory abnormality. Peripheral blood mononuclear cells were incubated with Con A and then suppression of proliferation was measured by coculture of these cells with healthy volunteers' mononuclear cells and phytohaemagglutinin. Sixteen asthmatic patients had significantly (P less than 0.002) decreased Con A-induced suppressor cell function (17.0 +/- 17.2%, mean +/- s.d.) as compared to 13 healthy volunteers (37.9 +/- 14.9%). Oral colchicine significantly (P less than 0.05) increased, though only partially corrected, these 16 asthmatic patients' Con A-induced suppressor cell function (28.1 +/- 14.3%). Asthmatic patients had an increased number of monocytes (691 +/- 289 vs 388 +/- 271/mm3 for normals, P less than 0.01) and a normal number of lymphocytes, Leu 4+ total T cells, Leu 3+ helper/inducer T cells, and Leu 2+ suppressor/cytotoxic T cells as well as a normal Leu 3/Leu 2 ratio. Oral colchicine significantly (P less than 0.005) decreased the number of monocytes (451 +/- 255/mm3) without significantly affecting the number of lymphocytes, Leu 4+, Leu 3+, or Leu 2+ T cells, or the Leu 3/Leu 2 ratio. These results are consistent with the hypothesis that the deficiency of Con A-induced suppressor cell function in asthmatic patients may be due, in part, to an increased number and/or abnormal activity of monocytes. If so, then oral colchicine may have partially corrected the deficiency of Con A-induced suppressor cell function by decreasing the number and/or modulating the activity of monocytes.     

Suppressor T cell numbers and function in atopic patients with high serum IgE levels

Suppressor T cell function and the sensitivity of lymphocyte transformation to histamine has been studied in 9 atopic patients with high serum IgE levels and in 14 controls. The effect of concanavalin A (Con A) induced suppressor T cells on the proliferative response of fresh lymphocytes to mitogens and antigens was measured; in atopic subjects the median suppression to 5.0 micrograms/ml Con A was 80% to 1.0 microgram/ml Phytohaemagglutinin (PHA) was 73%, to Dermatophagoides pteronyssinus extract was 45% and to streptokinase--streptodornase was 24%. Indomethacin increased lymphocyte proliferation to mitogens, but to a variable degree, in both groups. Histamine suppressed lymphocyte transformation to PHA and Con A (median suppression in normal subjects 38 and 46%, and in atopics 51 and 60%) and to D. Pteronyssinus extract. There was no significant difference between normal subjects and atopics in any of these functional assays. The relative and absolute numbers of total T cells, helper T cells and suppressor T cells measured by monoclonal antibodies and the helper to suppressor T cell ratio were normal in the atopic group. These results show that the activity of Con A induced suppressor T cells and the effect of histamine and indomethacin on lymphocyte proliferation is normal in highly atopic subjects. No suppressor T cell defect has been identified using these assays.     

Human monocyte-derived soluble product(s) has an accessory function in the generation of histamine- and concanavalin A-induced suppressor T cells.

We have analyzed the cellular interactions required for the generation of histamine- and concanavalin A (Con A)-induced suppressor T cells by employing a co-culture assay and techniques for fractionation of human blood mononuclear cells (PBMC). PBMC cultured in the presence of histamine (0.1 mM-1 mM) or Con A (20 micrograms/ml) for 24 h, mitomycin treated and subsequently combined with autologous mitogen-stimulated mononuclear cells, significantly suppressed a subsequent blastogenic response. PBMC fractionated over nylon wool columns and depleted of adherent cells and enriched for T cells (NWNA-T) were unable to generate suppressor activity. However, suppressor cell function by NWNA-T cells was reconstituted by the addition of autologous monocytes. In both the histamine and ConA suppressor systems, the requirement for monocytes in the activation process was enhanced by suspending the NWNA-T population in supernatants derived from allogeneic monocytes stimulated with heat-killed Staphylococcus albus. These crude supernatants contained leukocytic pyrogen (LP) and lymphocyte activating factor (LAF). Sequential purification and separation of the crude supernatants using gel-filtration, immunoadsorption, and isoelectric focusing demonstrated that only those fractions containing LP and LAF were capable to reconstituting NWNA-T cell histamine and Con A-induced suppressor activity. Thus, these studies suggest that the accessory role of supernatants derived from activated monocytes in the generation of suppressor cells may be mediated by LP/LAF. Further studies are in progress to explore the mechanism by which soluble factors stimulate suppressor T cells.     

The cellular basis of the impaired autologous mixed lymphocyte reaction in patients with systemic lupus erythematosus.

The autologous mixed lymphocyte reaction is impaired in patients with systemic lupus erythematosus. This is because nonthymus-derived (T)-lymphocyte preparations from such patients do not stimulate autologous T-lymphocyte proliferation normally. This defect may explain the impaired generation of suppressor activity in systemic lupus erythematosus and thereby the occurrence of autoantibodies in this disease.     

Enhancement of the impaired autologous mixed leukocyte reaction in patients with systemic lupus erythematosus.

The cellular basis of the impaired autologous mixed leukocyte reaction (AMLR) in patients with systemic lupus erythematosus (SLE) was investigated. Non-T cells from normal subjects and from SLE patients were fractionated into low and high density subpopulations. SLE patients were found to have an increased proportion of low density to high density non-T cells when compared to normal subjects. Although normal low-density non-T cells were highly enriched in AMLR stimulatory capacity, SLE low-density non-T cells induced minimal proliferation by autologous T cells. Brief incubation of SLE non-T cells with phorbol myristate acetate or formalin-treated Staphylococcus aureus resulted in marked augmentation of the capacity of those non-T cells to stimulate an AMLR, although the magnitude of the activated non-T cell-induced AMLR did not achieve that observed in normal subjects. No significant alterations in the expression of Ia molecules on the surface of the non-T cells were detected after in vitro activation. These experiments support the hypothesis that the impaired capacity of SLE T lymphocytes to proliferate in response to autologous non-T cells may in part represent a failure of SLE non-T cells to present an appropriate stimulus for the generation of a T cell response.     

Decreased in vitro humoral immune responses in aged humans.

Induction of antigen-specific and non-specific (polyclonal) humoral immune responses in vitro was investigated in peripheral blood mononuclear cells of aged (65-85 yr) and young (20-30 yr) volunteers. In vitro immunization of lymphocytes with antigen (sheep erythrocytes) was performed in a recently described microculture system, and anti-sheep erythrocyte plaque forming cells were quantitated in a direct hemolytic plaque assay. Immunoglobulin secreting cells, induced polyclonally with pokeweed mitogen, were quantitated in a reverse hemolytic plaque assay. Significant depressions of antigen-specific as well as polyclonal responses were noted in relation to advancing age. Antigen-specific responses were more frequently depressed than polyclonal responses. T cell mitogen concanavalin A (Con A) was used to amplify functions of autologous immunoregulatory T cells. Addition of 10 microgram/ml Con A to lymphocytes of young donors at culture initiation resulted in activation of suppressor cells and abrogated antigen-specific responses. Delayed addition of Con A, on the other hand, enhanced responses, presumably because of activation of helper T cells. Similar manipulations of lymphocyte cultures from aged donors showed failure of Con A to suppress antigen-specific responses in approximately half of the responders. In many nonresponders, responses within normal range were elicited by the delayed addition of Con A to their lymphocyte cultures. Deviations beyond the range of expected responses were noted in 32.5% of the co-cultures between pokeweed mitogen stimulated young and aged cells. Our findings suggest that age-related deficiencies of B cell function are frequently associated with dysfunction of immunoregulatory T cells and are only occasionally due to intrinsic defects of B cells.     

Deficient concanavalin-A-induced suppressor cell activity in women with untreated breast cancer

Peripheral blood mononuclear cells from 14 women with untreated breast cancer and 5 patients with non-malignant breast disease were studied for concanavalin-A (Con A) inducible suppressor activity against proliferative response of lymphocytes to phytohemagglutinin (PHA). Six of 14 patients with breast cancer demonstrated deficiency of suppressor cell activity against proliferation of autologous lymphocytes to PHA. This is in contrast to only 1 of 5 patients with benign breast disease who demonstrated deficiency of inducible suppressor activity. No correlation was observed between the deficiency of suppressor cells and the presence or absence of metastasis in the regional lymph nodes. The significance of these observations is discussed.     

A Suppressor T Cell of the Mixed Lymphocyte Reaction Specific for the HLA-D Region in Man

The mixed lymphocyte reaction (MLR) is the proliferative response of one individual's lymphocytes cultured in the presence of another individual's lymphocytes. In man, the MLR is elicited by cell surface antigens coded for by the HLA-D gene locus. This locus is among a cluster of genes which are located on the sixth chromosome and which include genes coding for the major histocompatibility antigens HLA-A, B, and C as well as HLA-D. If the stimulator cell possesses D locus antigens not present in the responder, the lymphocytes of the latter will undergo blast transformation resulting in DNA synthesis which can be measured. A vigorous response in the MLR to allogeneic cells is the rule among healthy individuals.We describe studies of a 23-yr-old man whose lymphocytes respond normally to mitogens and soluble antigens but fail to respond to allogeneic cells in the MLR. His medical history is unremarkable except that he received thymic irradiation as an infant. HLA typing revealed that he is homozygous for HLA-A2, B12, and Cw5 as well as for the D locus antigen Dw4. When his lymphocytes were added to the responder lymphocytes of other persons homozygous for the same HLA antigens, their responses to allogeneic cells but not mitogens were suppressed by 50-95%. Their responses to a soluble antigen, tetanus toxoid, were suppressed to a lesser degree. These inhibitory effects were mediated by a relatively radioresistant thymus-derived (T) lymphocyte.Further studies of the requirements for MLR suppression revealed that only persons heterozygous or homozygous for the Dw4 antigen were inhibited by the suppressor T cell. This effect was not altered by differences in the HLA-A, B, or C antigens between the suppressor and responder. It is concluded that genes in or near the HLA-D locus code not only for antigens (primarily on bone marrow-derived (B) cells), that elicit the MLR, but also for structures on T cells, or possibly macrophages, which are recognized by MLR suppressor T cells.