Fractionation of lymphocyte subpopulations which regulate mixed lymphocyte reactions.

Four days after injection of allogeneic lymphocytes BALB/c splenic T cells suppress proliferation of syngeneic cells in mixed lymphocyte reactions (MLR). Conversely, lymph node cells from the same mice amplify MLR responses. To further characterize these functional subpopulations, alloantigen-primed lymphocyte suspensions from both organs were fractionated by velocity sedimentation at unit-gravity. After fractionation MLR suppressor cells from spleens localized exclusively in rapidlly sedimenting fractions of large cells. MLR suppressor activity of cells from these fractions, as well as that of unfractionated spleen cell suspensions, was abolished by treatment with anti-Thy-1.2 serum and complement. Spleen cell fractions of similar sedimentation velocity also secreted a soluble MLR suppressor into culture supernatants. Although inhibitory of MLR, spleen cells of rapid sedimentation velocity did not suppress responses to T cell mitogens. In marked contrast with the effects of spleen cells, large 4-day-alloantigen-primed lymph node cells had no suppressive activity in MLR. MLR amplifier cells of uncertain derivation were found in fractions of medium sedimentation velocity from both spleens and lymph nodes. Fractionation of alloantigen-primed lymph node cell suspensions did reveal, however, a subpopulation of small cells with MLR suppressor acitivty which was unaffected by treatment with anti-Thy-1 serum and complement. The data thus indicate that large alloantigen-activated lymphocytes are not intrinsically suppressive nor are cells which suppress MLR necessarily large. We consequently conclude that regulation of MLR responses by alloantigen-primed lymphocytes involves a complex interaction between distinct functional subpopulations of cells which are separable both by physical and biologic properties.     

Suppression of mixed lymphocyte reactions by alloantigen-activated spleen-localizing thymocytes.

Heavily irradiated BALB/c and C57BL/6 mice were injected intravenously with BALB/c thymocytes. At varying times thereafter spleen and lymph node cell suspensions from these animals were treated with mitomycin C and added as regulator populations to mixed lymphocyte reactions (MLR) with syngeneic responder cells. Alloantigen-activated spleen-localizing thymocytes suppressed MLR responses 40 to 95%. Suppressor activitity, manifested as a quantitative reduction in peak proliferative responses, was maximal 4 days after cell transfer. Antigenic specificity for the stimulator cell strain in MLR was not demonstrated. The effect of lymph nodelocalizing thymocytes as regulators in MLR was variable, but in most experiments these cells slightly enhanced responses. We conclude that splenic localization is an intrinsic property of the thymocyte subpopulation capable of suppressing MLR responses, and that the suppressor activity of this subpopulation is substantially enhanced by activation in allogeneic hosts.     

Biological expressions of lymphocyte activation. IV. Concanavalin A-activated suppressor cells in mouse mixed lymphocyte reactions.

Thymus-derived lymphocytes (T cells) from mouse spleen, activated in vitro or in vivo with concanavalin A (Con A), suppress proliferative responses of syngenic lymphocytes in mixed lymphocyte reactions (MLR). Replication in vitro was not required for expression of suppressor activity by Con A-activated cells and was blocked in MLR by treating suppressor cells with mitomycin C or irradiation. Kinetics of MLR responses and viability of cultures were not altered by addition of activated suppressor cells. The data are consistent with a direct inhibitory effect of suppressor T cells on antigen-induced DNA replication. These observations extend a model previously described for regulation of antibody synthesis by Con A-activated T cells to control of cell-mediated immune responses. This model should be particularly useful in further definition of regulatory T cell subpopulations, and in investigation of interactions and relationships between such populations.     

Enhancement of the mixed lymphocyte response in the mouse by addition of thymocytes

Synthesis of DNA by mixtures of mouse lymph node and thymic cells was studied in vitro using mitomycin-treated allogeneic spleen cells as stimulator cells. The tests were performed to see whether there occurs a similar cell synergy during this reaction as has been reported during the in vivo graft-vs-host response.It was observed that mixtures of thymocytes and lymph node cells give higher incorporations of isotope-labelled thymidine than can be explained by a pure additive effect of the two cell populations tested separately. This enhancement of the reactivity was more pronounced using combinations of lymph node cells and medullary thymocytes obtained from cortisone-treated donors. Enhancement was also noted between lymph node cells and spleen cells. Blocking of the capacity of lymph node cells to synthesize DNA by treatment with mitomycin abolished this enhanced activity when mixed with thymic cells. On the contrary, mitomycin treatment of thymocytes did not abolish their capacity to increase the reactivity when mixed with normal lymph node cells. Thymocytes, which were unresponsive to the mitomycin-treated cells for genetic reasons, were also found to increase DNA synthesis when combined with lymph node cells. The mechanism by which thymocytes increase DNA synthesis of lymph node cells is not clear, but the results show that they have to be present during the reaction, since culture medium “conditioned” by thymocytes did not exhibit any enhanced capacity to promote a mixed lymphocyte reaction of lymph node cells.The results are thus in agreement with the findings obtained by others showing that mixtures of lymph node cells and thymic cells yield higher immunological reactivities in vivo against foreign transplantation, antigens than can be explained by a pure additive effect of the reactivities by the two cell populations tested separately. However, in contrast to these findings, the thymic cells do not have to be able to synthesize DNA or to react against the foreign cells in vitro to yield an enhanced response when mixed with lymph node cells.     

Regulation by adherent cells of macromolecular synthesis in lymphocyte populations.

The role of adherent cells in the regulation of lymphocyte DNA, RNA, and protein synthesis was investigated. Splenic adherent cells suppressed DNA synthesis of spleen cells. The reverse was true of lymph node adherent cells, i.e., DNA synthesis was less in the absence of adherent cells. Histocompatibility was not required for interaction between adherent and nonadherent cells. The regulatory adherent cell in either case was not θ positive. Removal of splenic adherent cells decreased RNA and protein synthesis of nonadherent spleen cells. RNA synthesis by lymph node cells increased when adherent cells were removed, but protein synthesis was lower. Autoradiographic analysis revealed that removal of adherent spleen cells allowed a greater number of nonadherent cells to respond to the presence of mitogen. Adherent cells were observed to regulate DNA synthesis of B lymphocytes also. Lymph node adherent cells amplified DNA synthesis in both nonadherent spleen and lymph node cells, while splenic adherent cells suppressed splenic nonadherent cells but stimulated nonadherent lymph node cells. We feel the data are compatible with the idea that at least two functionally distinct adherent cell populations exist. A mechanism is suggested to explain the data.     

T and B lymphocyte migration into syngeneic tumors.

The migration of splenic T and B lymphocytes into syngeneic tumors undergoing immunologic rejection was investigates. Spleen cells were obtained from normal BALC/c mice or BALB/c mice bearing tumors induced by murine sarcoma virus (MSV). Either whole spleen cells or immunoabsorbent purified T and B cells were radiolabeled with sodium chromate-51 and injected i.v. into normal or MSV inducted-tumor bearing syngeneic recipients. Twenty-four hours later the recipient mice were sacrificed and radioactivity was assessed for tumor, contralateral normal muscle, the lymph nodes draining the tumor and contralateral draining lymph nodes, peripheral lymph nodes, spleen, and liver. Both T and B lymphocytes from either normal or MSV tumor-bearing animals show greatly increased migration into the tumor when compared with normal muscle. Migration of T cells from both normal and MSV tumor bearers was 30 times that of migration to normal muscle. B cells from tumor-bearing mice, on the other hand, localized in the tumor itself only 50% as frequently as did B cells from normal animals. In addition, T cells from MSV tumor bearers were found in the highest proportion in the lymph node draining the tumor site. We conclude that T and B lymphocytes from either normal or tumor-bearing mice migrate to a syngeneic tumor undergoing immunologic rejection. In contrast, the migration of both T and B cells from tumor-bearing animals was decreased to the peripheral lymph nodes at the time of maximum tumor growth.     

Progression of armed CTL from draining lymph node to spleen shortly after localized infection with herpes simplex virus 1.

We have examined the generation of CTL immunity immediately after localized footpad infection with herpes simplex virus 1 (HSV-1) using three coordinated in vivo T cell tracking methodologies. Tetrameric MHC class I containing the immunodominant peptide from HSV-1 glycoprotein B (gB) showed that after infection the proportion of Ag-specific T cells peaked at day 5 within draining popliteal lymph nodes and 2 days later in the spleen. Preferential expression of the activation marker CD25 by tetramer-positive cells in draining popliteal nodes but not spleen suggested that gB-specific T cells were initially activated within the lymph node. In vivo cytotoxicity assays showed that Ag-specific effector cells were present within the draining lymph nodes as early as day 2 after infection, with a further 2-day lag before detection in the spleen. Consistent with the very early arming of effector CTL in the draining lymph node, adoptive transfer of CFSE-labeled gB-specific transgenic T cells showed that they had undergone one to four rounds of cell division by day 2 after infection. In contrast, proliferating T cells were first detected in appreciable numbers in the spleen on day 4, at which time they had undergone extensive cell division. These data demonstrate that HSV-1-specific T cells are rapidly activated and armed within draining lymph nodes shortly after localized HSV-1 infection. This is followed by their dissemination to other compartments such as the spleen, where they further proliferate in an Ag-independent fashion.     

Lymphocyte function in experimental African trypanosomiasis. VIII. Loss of suppressor T cell function in lymph nodes

The immunosuppression that occurs in mice experimentally infected with African trypanosomiasis has been examined further. In the present study we have examined lymph node cells from Trypanosoma rhodesiense-infected C57Bl/6J mice for the ability to produce mitogen induced antigen-nonspecific suppressor T cells (Ts). Inguinal, mesenteric, and brachial lymph node cells were harvested from uninfected control mice and from mice at different periods of infection. These cells were cultured with or without concanavalin A (Con A) for 48 hr to induce Ts activity. After stimulation, the control and infected lymph node cells were passed over Sephadex G-10 columns to remove suppressor macrophages that arise during the infection from Con A-induced Ts. The column passed cells were then added to normal mouse responder spleen cells in a primary in vitro antibody response culture system with sheep erythrocytes (SRBC) as antigen. The resultant plaque-forming cell responses to SRBC indicated that Ts function was not induced in infected lymph node cell populations. However, early in the infection, a stimulatory signal was provided by both the untreated and Con A-treated infected lymph node cells, which was lost in the terminal stage. Determinations of T cell subpopulations revealed that the infected Lyt 2.2-bearing subpopulation was not significantly altered from normal controls. We conclude that T. rhodesense infected mice fail to mount normal lymph node cell antigen nonspecific Ts responses and that this loss of activity may be due to an intrinsic dysfunction in the suppressor T cell population.     

The antimetastatic function of concomitant antitumor immunity. II. Evidence that the generation of Ly-1+2+ effector T cells temporarily causes the destruction of already disseminated tumor cells

Progressive growth of the P815 mastocytoma in an immunocompetent host evokes the generation of an antitumor immune response that can be measured in terms of the production of cytolytic Ly-1+2+ T cells in the draining lymph node and spleen. This immunity, designated concomitant immunity, is present on day 6 of tumor growth, peaks on day 9, and decays progressively thereafter. It fails to develop in mice made T cell deficient by thymectomy and lethal whole-body gamma-radiation, and reconstituted with syngeneic bone marrow cells (TXB mice). Employment of a mouse survival assay, capable of enumerating metastatic P815 cells in cell suspensions, showed that the P815 tumor metastasizes to the draining lymph node and spleen at the same rate in normal and TXB mice for the first 6 days of growth of an intradermal P815 tumor. By day 6 of tumor growth there were approximately 10(3) P815 cells in the draining lymph node in both types of mice. However, during the generation of concomitant immunity between days 6 and 9, the number of metastatic P815 cells in the draining lymph nodes and spleens of normal tumor-bearing mice declined by nearly 90%. After day 12, however, the number of tumor cells in the nodes and spleens increased concordantly with the decay of concomitant immunity. These findings, together with the demonstration that T cell-deficient mice failed to restrain the number of metastatic P815 cells in the draining lymph node and spleen, suggest that concomitant immunity is an important defense mechanism against the development of systemic disease. Additional evidence consistent with this interpretation was provided by studies which showed that adoptive immunization with spleen cells from concomitant immune donors significantly prolonged the median survival time of TXB tumor-bearing mice by destroying a substantial proportion of P815 tumor cells already seeded in the draining lymph node. Adoptive immunization also delayed the appearance of metastatic tumor cells in the spleen.     

Cross presentation of antigen on MHC class II via the draining lymph node after corneal transplantation in mice

We investigated Ag trafficking from the cornea and T effector cell activation in secondary lymphoid tissue after corneal transplantation. In preliminary experiments, the central cornea was shown to contain a population of CD45(+), CD11b(+), CD11c- cells, with a few MHC class II(+) cells, and F4/80(+) cells. However, MHC class II(+) passenger leukocytes in donor cornea after allografting did not traffic to the draining lymph node. Instead, Ag (plasmid) delivered to the eye via the donor cornea during allograft was detected in host CD11c(+) and F4/80(+) APC in the draining lymph nodes and spleen. The earliest detection of APC-associated Ag was at 6 h in the draining lymph node and 24 h in the spleen. After 48 h Ag was not detected in the draining lymph node but was still present in the spleen. Ag applied to the donor corneal epithelium before allografting induced Ag-specific T cell activation and expansion in the draining lymph node with a peak response at 4-6 days, indicating that cross-presentation of Ag had occurred. We conclude therefore, that Ag is transported from the donor cornea within host APC and that this event occurs within hours after grafting. Ag is cross-presented to host CD4(+) T cells on MHC class II and leads to the activation of Ag-specific effector T cells and clonal expansion in the draining lymph node.     

PHA activation of encephalitogenic T cells: in vitro line selection overcomes splenic suppression

In this study we compared myelin basic protein (MBP) and phytohemagglutinin (PHA) for their ability to induce proliferation and experimental autoimmune encephalomyelitis (EAE) transfer activity in mixed cell cultures obtained from spleen and lymph nodes versus highly selected MBP-specific T cell lines and clones. Established MBP-specific cells derived initially from immune lymph nodes attained both proliferative and EAE-transfer activities after in vitro activation with either MBP or PHA. In contrast, PHA was unable to induce immune spleen cells to transfer EAE, in spite of its potent mitogenic activity. On the basis of these results, we evaluated the in vitro proliferation and differentiation responses of MBP-specific T cells during the line selection process using cells derived from both immune lymph node and immune spleen. During the initial selection process with MBP, proliferation of MBP-specific T cell precursors from immunized spleen populations was reduced relative to lymph node cells. After antigen-dependent selection the encephalitogenic cells from either organ exhibited identical in vitro response characteristics. Freshly isolated immune spleen cells were potent suppressors of MBP-specific T cell proliferation suggesting that the in vitro differences between the two organs was due to splenic suppression of the encephalitogenic cells.     

Correlation of cytotoxicity with experimental allergic encephalomyelitis.

Cytotoxicity of immune lymph node cells in experimental allergic encephalomyelitis (EAE) was maximal 9 days after injection of encephalitogenic emulsion. The ability of these cells to passively transfer EAE was also maximal at this time. Immune spleen cells were more cytotoxic than lymph node cells 9 days after injection; however, these cells did not passively transfer EAE. Twelve days after injection of encephalitogenic emulsion immune spleen cells passively transferred EAE with resulting mild histopathologic lesions. At this time the spleen cells were 50% more cytotoxic than comparable lymph node cells. Cyclophosphamide suppressed the development of clinical EAE and the development of cytotoxic lymphoid cells. It also reduced clinical signs and cytotoxic activity of lymph node cells. Spleen cell cytotoxic activity was enhanced by Cyclophosphamide. It was concluded that cytotoxic activity of lymph node and spleen cells was correlated with the ability of these cells to produce EAE. Lymph node cell populations differed qualitatively and/or quantitatively from immune spleen cell populations in EAE. Capacity to passively transfer EAE coincided with the maximal Cytotoxicity of the lymphoid cells from each tissue.     

The B cell is the initiating antigen-presenting cell in peripheral lymph nodes

We have examined the role of B cells in antigen presentation in lymph nodes in several ways. We found that mice depleted of B lymphocytes via chronic injection of anti-mu-chain antibody do not mount peripheral lymph node T cell proliferative responses to normally immunogenic doses of antigen. Depletion of B cells by passage of immune lymph node cells over anti-immunoglobulin columns early after immunization depletes antigen-presenting function from draining lymph nodes, and this function can be restored by using B cells or splenic adherent cells to allow the remaining T cells to proliferate. Lymph node B cells present antigen very effectively to lines of antigen-specific T cells. However, unfractionated lymph node cells from anti-mu-treated mice present very poorly, if at all, whereas unfractionated spleen cells from the same mice do present antigen. This is in keeping with our previous finding that helper T cell function in the spleen is normal in B cell-deprived mice. Finally, when mice homozygous for the lymphoproliferative gene lpr are treated chronically with anti-mu-chain antibody, lymphadenopathy is greatly retarded, suggesting a role for B cells in the massive proliferation of T cells in this syndrome. From this analysis, it would appear that the initiating antigen-presenting cell in the lymph node is a B lymphocyte, and that B lymphocytes in lymph nodes may be distinct from those in the spleen. It is of interest that these results also suggest that the lymph node lacks an antigen-presenting cell that is found in the spleen, perhaps the dendritic cell.     

Immunologic characterization of jird lymphocyte responsiveness to Brugia pahangi ribosomal protein S13.

Ribosomal protein S13 of the nematode Brugia pahangi is recognized by B and T cells from parasite-infected animals. To identify helper T cell sites on the protein, 15 overlapping synthetic peptides spanning the entire molecule (Bp17.4) were tested for their ability to stimulate lymph node and spleen cells of peptide-immunized and recombinant antigen-immunized jirds. Lymph node cells from animals immunized with peptides 6, 8, 9, 13, and 14, corresponding to Bp17.4 amino acids (AA) 50-70, 70-90, 80-100, 120-140, and 130-150, respectively, showed strong and specific responses to the homologous peptide, while only those lymph node cells from jirds immunized with peptides 8, 9, 13, and 14 proliferated in response to Bp17.4. These results suggest the existence of at least two T cell epitopes. Lymph node cells from infected jirds also responded to these peptides and to Bp17.4 (80,000 cpm). In contrast to the lymph node cells, spleen cells from microfilaria-positive animals failed to mount significant responses to any of the peptides or to Bp17.4. Splenic T cell responsiveness was restored upon removal of nylon wool adherent cells, suggesting active regulation of Bp17.4 reactivity. In liquid-phase competitive inhibition immunoassays, peptides 1 (AA 1-30) and 6 (50-70) blocked antibody binding and, therefore, these regions contain conformational antibody-binding sites. This model system should prove useful for analyzing regulation of epitope-specific responses in experimental filariasis.     

Variable lymphocyte responses in rats after space flight.

Most studies of human blood lymphocyte function following space flight have indicated that microgravity suppresses T cell proliferation. However, several other postflight experiments with animals have shown no decrease in proliferation of lymphocytes from peripheral lymphatic tissues, suggesting that different tissues may be variably affected by microgravity. Therefore, we examined the proliferation of lymphocytes from both spleen and lymph nodes of rats following a 4-day flight aboard the Space Shuttle. The experiments were designed to investigate tissue variability as well as potential mechanisms involved in suppressing proliferation. We found that proliferation of lymph node lymphocytes (LNL) from flight (FLT) animals stimulated with the antigen receptor-dependent T cell mitogen concanavalin A was depressed and could not be restored by supplementing cultures with interleukin 1 or interleukin 2 (IL-2). Response to another receptor-dependent mitogen, phytohemagglutinin, was not decreased. However, proliferation of FLT LNL following stimulation with the receptor-independent, mitogenic combination of phorbol ester and ionomycin was depressed. LNL IL-2 activity, cell surface marker expression, and B cell responses to mitogen were normal. Thus, deficits in antigen receptor/ligand interactions, cell surface marker expression, or IL-2 did not account for the suppressed lymphocyte proliferation observed postflight. In contrast to LNL, FLT splenocyte proliferation was not depressed. Assayable IL-2, IL-2 receptor expression, and cell surface marker expression likewise were unaffected by space flight. The differences between lymph node and splenic responses demonstrate the tissue-specific nature of microgravity effects on individual lymphatic tissues.     

Freezing of rat lymphocytes. I. The effects of dimethyl sulfoxide and freezing on the phytohemagglutinin- and pokeweed mitogen-responding lymphocyte subpopulations.

Rat spleen and lymph node lymphocytes have been frozen with dimethyl sulfoxide (DMSO) at 1 °C/min and stored at ?196 °C for 10 min. The functional recovery of the cell populations was monitored by the mitogenic response (uptake of [³H]thymidine) to phytohemagglutinin (PHA) or pokeweed mitogen (PWM) in culture after thawing. With 5 to 10% DMSO in the freezing medium, frozen-thawed lymph node cells were found to retain about 40% of their response to PHA. In contrast, frozen-thawed spleen cells responded better to PHA than fresh cells. The response to PWM was markedly decreased in both spleen and lymph node cell cultures.A similar effect was observed when DMSO was added to the culture medium of fresh spleen cells, i.e., an augmentation of the response to PHA and a suppression of the response to PWM. However, the concentrations of DMSO needed to induce this effect was more than 10-fold higher than that present in the culture medium after freezing and thawing.Since removal of adherent cells from the spleen cell population also produced an augmentation of the response to PHA, it is suggested that the freezing procedure and DMSO may have an inhibitory effect on suppressor cell functions present in spleen cell populations.     

Lymphocyte function in experimental African trypanosomiasis. III. Loss of lymph node cell responsiveness

The relationship between immunosuppression and suppressor cell activity in the lymphoid organs of animals with experimental African trypanosomiasis has been examined further. In the present study we measure the primary in vitro PFC response to SRBC by spleen and lymph node cells from Trypanosoma rhodesiense infected or drug-cured C57BL/6 mice. Passive transfer experiments with this culture system tested for the presence or absence of suppressor cells. We demonstrate that infected mice exhibit immunosuppression in the spleen cell population several weeks before becoming suppressed at the level of the lymph node cell populations. Although suppressor cells are present in immunosuppressed spleen cell populations, suppression of lymph node cell responsiveness was not attributable to suppressor cells detectable withi, lymph nodes. After Berenil treatment of terminally infected mice immunocompetence was restored gradually, first to the lymph node cells and subsequently to the spleen cell population. Recovery of spleen cell responsiveness was attributable to the loss of detectable suppressor cell activity within spleens. These results demonstrate that there is anatomical restriction of the suppressor cell population to trypanosome-infected mouse spleen and that loss of immunocompetence in the lymph nodes may be due to factors unrelated to suppressor cell effects.     

Identification of profound peripheral T lymphocyte immunodeficiencies in the spontaneously diabetic BB rat

The BB rat is presently the best available animal model for human insulin dependent diabetes (IDD). Because of the extreme susceptibility of the strain to opportunistic infections and because current studies suggest that they have an autoimmune diathesis, of which IDD is but one result, aspects of the immune system of the BB rat were studied. Severe T lymphopenia was observed in all BB rats, irrespective of sex or the presence of IDD, while numbers of B cells and serum immunoglobulin levels were normal. Both the helper T lymphocyte and cytotoxic/suppressor T lymphocyte subsets, defined by reactions with monoclonal antibodies, were depressed, and an inversion of the helper T cell subset to cytotoxic/suppressor T lymphocyte subset ratio occurred in all BB rats with increasing maturity. Concomitantly, severe impairments of T cell-mediated immune responses were noted. BB rats poorly rejected allografts across both major and minor histocompatibility barriers, and BB splenic or peripheral blood lymphocytes had markedly defective proliferative responses to mitogens and to allogeneic cells in MLC. Irradiated and nonirradiated BB spleen cells did not inhibit WF mitogenic or MLC responses, which suggests that the T cell defect in BB rats is not solely due to increased suppressor activity. Because irradiated WF cells and Con A supernatants did not restore BB proliferative responses, and BB lymphocytes were able to produce IL-2 normally, a reduced ability of BB lymphocytes to respond to helper factors such as IL-2 is suggested. In contrast to T lymphocytes from spleen or peripheral blood, BB thymocytes responded as well as did WF thymocytes to Con A or Con A supernatants. Percentages of T lymphocyte subsets and histology of BB thymuses were also normal when compared to WF thymuses. However, spleens and lymph nodes from BB rats were severely depleted of T lymphocytes, and thymocytotoxic autoantibodies were detected in many BB rat sera. The above findings indicate that BB rats have T lymphocyte immunoincompetence, which appears to be a post-thymic or peripherally acquired maturational defect.     

Antigen-specific lymphocyte proliferative responses in inbred mice after Trichinella spiralis infection.

The in vitro antigen-specific lymphoproliferative response of spleen, mesenteric lymph node (MLN), and coeliac lymph node (CLN) cells taken from various strains of inbred mice infected with Trichinella spiralis was assessed. In most experiments cell populations were stimulated with excretory/secretory antigens (ESA) derived from adult and larval worms. Lymphoid cells collected 5-7 days postinfection were usually the most responsive to ESA as measured by [3H]thymidine uptake. Spleen cells were more responsive than either MLN or CLN cells. There was a correlation between in vitro ESA stimulation and worm rejection in strong- and weak-responder strains of mice. Spleen and MLN cells of NFS mice showed higher antigen-specific responsiveness, whereas the same cells from B10.BR (H-2k) and B10.Q (H-2q) strains of mice were less responsive. Among intermediate responder strains 2 patterns were observed. Spleen and MLN cells of BuB and DBA/1 mice responded more strongly than those of C3H mice. Dose-response experiments demonstrated that increasing the infective dose of larvae to the host usually increased subsequent in vitro antigen-specific lymphoproliferation. Furthermore, non-MHC-linked genes appear to be the primary determinant of antigen-specific T-cell-proliferative responses in inbred mice infected with T. spiralis.     

Age-related changes, including synergy and suppression, in the mixed lymphocyte reaction in long-lived mice.

The mixed lymphocyte culture reaction represents the in vitro counterpart of the recognition phase of the graft-versus-host reaction, and of allograft rejection. The mixed lymphocyte culture reactivities of lymph node and spleen cells from all strains show a striking decline with advanced age. Furthermore, studies of "synergy" between subpopulations of T cells in the mixed lymphocyte culture reaction suggest that the cells of the recirculating lymphoid pool (T2 cells) in particular display a functional decline. Finally, spleen cells from old mice of appropriate strains inhibit or suppress the mixed lymphocyte culture reactivity of lymph node or spleen cells from young mice.