Cloned murine Ia+ BK-BI-2.6.C6 T cells function as accessory cells presenting protein antigens to long-term-cultured antigen-specific T cell lines

A variant clone, BK-BI-2.6.C6, was derived from the murine bovine insulin-reactive T cell line BK-BI-2.6 with helper/amplifier phenotype. Variant cells have lost reactivity to insulin, but have acquired constitutive IL 2 receptor expression, growing in IL 2-containing medium without feeder cells. In contrast to their ancestor line, variant cells synthesize and express I-A and I-E region-dependent class II molecules as indicated by metabolic radiolabeling, immunoprecipitation with subregion-specific monoclonal antibodies and two-dimensional (2D) gel electrophoresis (1D isoelectric focusing, 2D SDS-PAGE). BK-BI-2.6.C6 cells can act as accessory cells, presenting the protein antigens bovine insulin and ovalbumin to antigen-dependent long-term cultured T cell lines BK-BI-1.2 and BK-OVA-1 in the context of I-A restriction elements. Antigen recognition on presenting BK-BI-2.6.C6 accessory cells resulted in highly efficient IL 2 production. However, in contrast to splenic antigen-presenting cells, BK-BI-2.6.C6 cells did not initiate antigen-specific [3H]thymidine incorporation by the T cell lines tested. Further study of accessory function of Ia+ T cell clones might provide insight into processes regulating T cell responses to antigen.     

Transfer of antigen-presenting capacity to Ia-negative cells upon fusion with Ia-bearing liposomes

The role of Ia in T cell activation was investigated by incorporating affinity-purified I-Ad molecules into synthetic liposomal membranes and by using these as antigen-presenting units. IL 2 production by I-Ad-restricted, chicken ovalbumin-specific T cell hybridomas was measured in a system in which antigen processing by the presenter was not required. I-Ad-bearing liposomes were found to have no antigen-presenting capacity. It was shown, however, that antigen-presenting capacity could be conferred on Ia-negative cells by fusion of these cells with liposomes bearing I-Ad molecules, together with Sendai virus envelope glycoproteins, as fusogenic agents. Both Ia-negative B lymphoma cells and mouse L cells were capable of antigen presentation of predigested ovalbumin after fusion with vesicles formed from phosphatidylserine and phosphatidylethanolamine in a 1:1 w:w ratio. The cell surface expression of the transferred Ia remained stable for at least 7 hr. These results indicate that Ia is the only additional cell surface molecule required, at least by Ia-negative B cell lymphomas and L cells, to convert them into effective antigen-presenting cells. This system should be useful in future studies of the cellular requirements for antigen processing and presentation.     

Influence of accessory cell and T cell surface antigens on mitogen-induced IL 2 receptor expression

We have assessed the inhibitory effects of various monoclonal antibodies on the expression of the IL 2 receptor. Anti-LFA-1, but not anti-Ly-2, markedly inhibited the induction of the IL 2 receptor on the Ly-2+ subset. T-depleted spleen cells, L cells, and B lymphoma cells all functioned as potent accessory cells (AC) for the induction of the IL 2 receptor on L3T4+ T cells. Anti-LFA-1 inhibited the induction of the IL 2 receptor irrespective of the type of AC used. Anti-L3T4 only inhibited the induction of IL 2 receptor expression when L cells were the source of AC. The inhibitory capacity of anti-L3T4 was not related to the expression of Ia on the AC population, because the magnitude of inhibition was comparable in cultures containing either Ia+ or Ia- L cells, whereas no inhibition was seen with either Ia+ or Ia-B lymphoma cells. We conclude from these studies that LFA-1 plays a critical role in mitogen-induced activation of both T cell subsets by promoting both T-AC and T-T interactions. Although anti-L3T4 can inhibit T cell activation in the absence of the recognition of Ia, the mechanism of inhibition and the proposed target molecule for L3T4 on the AC or the T cell have not been determined in our studies. A number of different models for the function of this cell surface antigen are discussed.     

Sodium periodate-induced T cell mitogenesis: an analysis of the requirement for Ia and IL 1

T lymphocytes oxidized with the mitogen sodium periodate undergo a proliferative response when cultured in the presence of Ia+ accessory cells. However, the exact role(s) the accessory cells play in such a response has not been clearly defined. We have evaluated the role of Ia and the requirement for interleukin 1 (IL 1) in periodate mitogenicity by using the Ia+ cloned tumor cell lines P388AD (Ia+, IL 1 inducible) and P388NA (Ia+, IL 1 noninducible) as accessory cells. P388AD but not P388NA was able to supply accessory cell function to periodate-treated T cells, suggesting that Ia expression alone was not sufficient to reconstitute a response. Monoclonal anti-I-Ad and anti-I-Ed antibody blocked the accessory cell function of P388AD. In addition, monoclonal antibody GK 1.5, directed against the T cell determinant L3T4a, blocked the P388AD/periodate-treated T cell interaction, confirming that this interaction was restricted by class II molecules. Although Ia expression was required, the response was not major histocompatibility complex (MHC) restricted, because allogeneic as well as syngeneic macrophages were capable of supplying accessory cell function to periodate-treated T cells. Exogenous IL 1 alone was able to trigger periodate-treated T cells, suggesting that Ia was required for the induction of IL 1 synthesis by the accessory cells. Furthermore, purified IL 2, devoid of IL 1 activity, was able to fully reconstitute the proliferative response of accessory cell-depleted oxidized T cells to a level equal to that of whole spleen accessory cells or P388AD. These data suggest that periodate-treated T cells can proliferate with IL 1 alone and that Ia+ accessory cells in periodate-mediated T cell mitogenicity may function in the release of IL 1 and the induction of IL 2 synthesis by the T cells.     

Molecular signals in antigen presentation. II. Activation of cytolytic cells in vitro after ultraviolet radiation or combined gamma and ultraviolet radiation treatment of antigen-presenting cells

Murine low-density spleen cells have potent antigen-presenting ability in a hapten-specific cytolytic T lymphocyte (CTL) system using the hapten azobenzenearsonate (ABA). Exposure of these cells to 0.33 KJ/m2 of ultraviolet radiation (UVR) after coupling to hapten results in markedly inhibited antigen-presenting function that can be substantially corrected or bypassed by interleukin 1 (IL 1). These results have been interpreted to reflect an inhibition of Lyt-1+ T cell activation by UVR-treated APC. Treatment of these cells sequentially with 1500 rad of gamma-radiation (GR) prior to hapten coupling, followed by 0.33 KJ/m2 of UVR radiation after coupling, results in an antigen-presenting defect only minimally improved by IL 1. However, partially purified interleukin 2 (IL 2) can completely bypass or correct this defect. Thus, combined GR and UVR induces a different or more profound defect in APC function when compared to UVR alone. However, these cells do provide a signal(s) other than hapten necessary for CTL activation because ABA-coupled high density spleen cells do not activate CTL cells, even with the addition of IL 2. Fluorescence-activated cell sorter analysis demonstrates that exposure of these low density spleen cells to GR or UVR results in decreased I-A antigen expression at 24 hr than either alone. The addition of nonhapten-coupled low-density APC partially reconstitutes the ability of combined GR/UVR-treated LD-APC to present antigen, and this effect is enhanced by the administration of exogenous IL 1. This occurs despite a lack of significant accessory cell activity by the LD-APC for the ABA hapten, and indicates that combined GR/UVR-treatment of APC is not functionally equivalent to completely removing them.     

Presentation of antigen by B cells: functional dependence on radiation dose, interleukins, cellular activation, and differential glycosylation

Whether resting B cells can present antigen to T cells is controversial. Several factors can influence the outcome of an assessment of the presenting function of resting B cells: the method of purifying resting B cells and maintaining them in culture without altering their resting state, the sensitivity of resting B cells to gamma-irradiation, the activation state of the T cells used to assess presenting function, and the requirement for exogenous interleukin 1. We have examined all of these variables and find that one adherent antigen-presenting cell is functionally equivalent to four LPS-activated B cells and to 1000 resting B cells. In addition, we have examined the potential functional relevance of the differential glycosylation of Ia molecules on resting B cells compared with adherent antigen-presenting cells. Altering the surface glycosylation of resting B cells by neuraminidase treatment results in a 25-fold increase in B cell antigen presentation without altering their resting state. More important, among antigen-presenting cells the effect of neuraminidase is limited to resting B cells. It also appears to involve a restricting element such as the Ia molecule rather than total cell surface charge, because neuraminidase treatment has no effect on the capacity of resting B cells to serve as accessory cells in the Con A response.     

Cell receptors for the mammalian reovirus: reovirus-specific T-cell hybridomas can become persistently infected and undergo autoimmune stimulation.

We have previously described the development of virus-specific helper T cell hybridomas which recognize structural determinants shared by type 1 and type 3 reoviruses that have been exposed to UV radiation. We have found that T-cell hybridomas become persistently infected with live type 3 reovirus used for the immunization. Persistently infected T-hybridoma cells were found to spontaneously produce interleukin 2 (IL-2). To analyze the mechanism of induction of IL-2 secretion of persistently infected T-cell hybridomas, we exposed T-cell hybridomas specific for UV-treated virus to replicating type 3 reovirus. The T-cell hybridomas became infected but did not produce IL-2 unless simultaneously exposed to syngeneic I-A+ antigen-presenting cells. In this situation, the persistently infected T-cell hybridomas produced IL-2 without being reexposed to virus. This process was not a consequence of nonspecific IL-2 gene activation, which occurs in cells persistently infected with reovirus, because reovirus infection did not activate IL-2 secretion in T-cell hybridomas with other antigenic specificities. Reovirus exposure also resulted in persistent infection of certain antigen-presenting B-cell tumor lines. The persistently infected B-cell tumor lines could stimulate reovirus-specific helper T cells but not T-cell hybridomas of other specificities. The data support the thesis that persistent infection of reovirus-specific T cells creates a mechanism in which the virus released from these cells is processed and then reexpressed by I-A+ antigen-presenting cells. The IA antigen and reovirus structures on the antigen-presenting cells then restimulate the T cells through their specific receptors, resulting in IL-2 synthesis and release. These observations may be relevant to mechanisms of autoimmunity induced by virus.     

The major histocompatibility complex-restricted antigen receptor on T cells. IX. Role of accessory molecules in recognition of antigen plus isolated IA

Antibody inhibition studies were done to determine which molecules on the surface of the T cell hybridomas other than their receptors for antigen plus IAd were involved in interaction with antigen-presenting B cells, with artificial IAd membranes on glass beads, or with anti-receptor antibodies coupled to Sepharose beads. We found that T cell LFA-1 was only involved when B cells were used to present antigen plus IAd, whereas T cell L3T4 was involved in the response of T cells to antigen plus IAd either on cells or in artificial membranes, but not if anti-receptor antibodies were used to stimulate the T cells. From these results we concluded that LFA-1 may be involved in the recognition of a ligand on cells that was not present in artificial membranes, but that L3T4 might interact with a nonpolymorphic portion of class II molecules present in both intact antigen-presenting cells and the antigen-presenting artificial membranes.     

Evidence for effects of interleukin 4 (B cell stimulatory factor 1) on macrophages: enhancement of antigen presenting ability of bone marrow-derived macrophages

We have studied the effects of recombinant mouse interleukin 4 (IL 4) (previously known as B cell stimulatory factor 1) on the antigen-presenting ability of murine splenic B cells and bone marrow macrophages. Our assay is based on the induction of antigen-presenting ability in these cells after incubation with IL 4 for 24 hr. The presenting cells were then used to stimulate IL 2 production by antigen-specific, I-Ad-restricted T cell hybridomas, a response mainly dependent on the induction of Ia antigens. Consistent with our previously published data using partially purified natural IL 4, we show here that recombinant IL 4 (but not interferon-gamma (IFN-gamma) or IL 1) induces antigen-presenting ability in B cells. Recombinant IL 4 was also found to induce antigen-presenting ability in a cloned, bone marrow derived-macrophage cell line (14M1.4), and in normal bone marrow-derived macrophages. These macrophage populations also respond to IFN-gamma showing enhanced antigen-presenting ability (mediated by increased Ia antigen expression). A small but significant increase in Ia antigen expression was also detected in 14M1.4 macrophages induced with IL 4. However, additional analysis suggested that the effect of IL 4 on 14M1.4 is different from that of IFN-gamma, because IL 4 (but not IFN-gamma) is able to maintain the viability and increase the size of and metabolic activity of bone marrow macrophages. However, IL 4 may not affect all macrophages because the macrophage cell line P388D1, which responds to IFN-gamma, failed to show enhanced antigen-presenting function after stimulation with IL 4. These observations indicate that IL 4, a lymphokine previously considered to be B cell lineage specific, has effects on macrophages and may be involved in their activation.     

Antigen presentation in the rat. II. An Ia+ radiosensitive T cell can present antigen to primed Ia- T cells

We demonstrated previously the presence of an Ia+ (OX-6+) antigen-presenting cell within the rat T cell fraction that is capable of presenting antigen to antigen-primed OX-6-T cells. This antigen-presenting cell (T-APC) reacted with the monoclonal antibodies W3/25 and W3/13, which is known to react mainly with rat T cells. Further characterization of the T-APC indicated that the cell also reacted with the monoclonal antibody OX-19, which is highly specific for rat T cells. Moreover, the antigen-presenting function of the T-APC was sensitive to treatment with mitomycin C or gamma-irradiation (2000 rad). Under similar conditions, antigen presentation by partially purified dendritic cells or macrophages was totally resistant to these treatments. The antigen-presenting activity of gamma-irradiated T-APC was not reconstituted by the addition of the lymphokines IL 1, IL 2, or Con A supernatants. Although unirradiated T-APC were able to stimulate an MLR response, this function was also sensitive to gamma-irradiation, whereas the MLR-stimulating ability of macrophages and dendritic cells was resistant to gamma-irradiation. These data indicate that Ia+ T cells from the rat are capable of presenting antigen to antigen-primed T lymphocytes and that, in contrast to antigen presentation by macrophages and dendritic cells, the function of T-APC is gamma-radiation sensitive.     

Modulation of suppressor T cell induction with gamma-interferon

The ability of antigen-coupled splenic adherent cells to induce suppressor T cells (Ts) is dependent on the presence of I-J determinants on antigen-presenting cells. After 4 days of in vitro culture, antigen-coupled adherent cells lose the capacity to induce Ts. Supernatants from Con A-stimulated lymphocyte cultures and purified interferon-gamma can sustain accessory function for the induction of Ts. Furthermore, after in vitro culture of splenic adherent cells, there is an apparent correlation between the loss of I-A determinants and the decrease in I-J-restricted Ts induction. Stimulation of Ia expression with interferon-gamma results in a simultaneous increase in the ability to induce Ts. Finally, elimination of I-A-bearing splenic adherent cells with antibody + C eliminates I-J-restricted Ts induction. The combined data imply a co-regulation of I-A and I-J on the antigen-presenting cells involved in the induction of both the Ts1 and Ts3 suppressor T cell subsets.     

Inability of human alveolar macrophages to stimulate resting T cells correlates with decreased antigen-specific T cell-macrophage binding

Alveolar macrophages (AM) from the majority of human volunteers are defective antigen presenting cells (APC) in T cell proliferation assays despite the display by the cells of HLA-D region antigens. We have confirmed that AM secrete relatively little interleukin 1 (IL 1), but addition of exogenous IL 1 did not improve the capacity of AM to initiate antigen-induced T cell proliferation. Thus, the presence of HLA-D region antigens and IL 1 is not sufficient to enable an accessory cell to act as an APC. We developed a T cell-accessory cell binding assay to investigate early events in T cell activation. AM demonstrated a diminished capacity as compared with monocytes to bind antigen-specific T cell clones. Nevertheless, AM often induced proliferation of T cell clones as effectively as monocytes, indicating that antigen display was intact. The inefficiency of AM in bind T cell clones correlated with their reduced capacity to induce resting T cells to express IL 2 receptors, secrete IL 2, and proliferate in response to antigen. Indirect immunofluorescence established that similar percentages of AM and monocytes expressed LFA molecules, but the density of the molecules was greater on monocytes than AM. A role for LFA antigens in the physical binding of T cells to monocytes was demonstrated by blocking antigen-specific binding with a monoclonal antibody to LFA-1 antigen. LFA-1 antibody also blocked the low levels of specific binding between AM and T cell clones, indicating that LFA-1-ligand interactions were operative between these two cell types. These studies indicate that there are critical cell membrane characteristics that promote binding of T cells to APC in addition to T cell receptor-antigen interactions. This combination of nonspecific and specific interactions leads to avid T cell-APC binding that may be essential for activation of resting T cells. Furthermore, we postulate that the failure to AM to act as effective APC results from an inability to bind T cells efficiently.     

The murine IL 2 receptor. III. Cellular requirements for the induction of IL 2 receptor expression on T cell subpopulations

The accessory cell requirements for the induction of the IL 2 receptor by the lectin Con A on murine T cell subsets were directly assayed with anti-IL 2 receptor monoclonal antibodies. Substantial levels of IL 2 receptor expression were induced on T lymphocytes of the MHC class I-restricted, suppressor/cytotoxic phenotype (L3T4-, Ly-2+) in the presence and absence of accessory cells. In contrast, high levels of IL 2 receptor expression could only be induced on T cells of the MHC class II-restricted, helper/inducer phenotype (L3T4+, LY-2-) in the presence, but not in the absence, of accessory cells. Ia- cells such as the P388D1 macrophage line or cultured fibroblasts (DAP X 3) were as efficient as the Ia+ B cell hybridoma LB in providing accessory cell function for the L3T4+, Ly-2- subset. PMA, but not purified human IL 1, could substitute for accessory cells for both IL 2 receptor expression and IL 2 secretion by the L3T4+, Ly-2- subset. These data suggest that IL 2 receptor induction on the L3T4+, Ly-2- subset is complex, possibly requiring a T cell-accessory cell interaction, whereas the lectin may directly trigger IL 2 receptor expression on L3T4-, Ly-2+ T cells.     

Antigen presentation to human T lymphocytes. II. Requirements for Mac-120+ macrophages and responsiveness to interleukin 2

To investigate mechanisms by which antigen, macrophages, and interleukin 2 (IL2) participate in the induction of secondary T-cell proliferative responses, trinitrophenyl (TNP) was presented in three distinct modes: (i) TNP-modified peripheral blood mononuclear cells (TNP-PBMC), (ii) TNP-PBMC cell sonicates, and (iii) TNP-ovalbumin (TNP-OVA). Stimulators were depleted of Mac-120+ macrophages using Mac-120 monoclonal antibody plus complement. TNP-Mac-120 macrophages stimulated primed T cells nearly as well as TNP-unfractionated macrophages (which were about 40% Mac-120+). In contrast, although greater than 70% DR+, Mac-120- macrophages plus either TNP-OVA or TNP-PBMC sonicate elicited minimal responses compared to unfractionated macrophages plus antigen. After 21-28 days of in vitro priming, macrophage-depleted T cells were not stimulated to proliferate by either IL2 alone or sonicates alone. IL2 plus TNP-PBMC sonicates, however, stimulated significant proliferation. Furthermore, this response was considerably greater than that to IL2 plus either TNP-T cell sonicates or TNP-mouse spleen sonicates. Thus, the Mac-120+ macrophage population may have an important antigen-presenting and/or accessory function in the stimulation of primed T cells by soluble or particulate antigen, although it is unnecessary for responses to intact TNP-Ia+ PBMC. In addition, the data suggest that Ia+ sonicates alone may suffice for induction of IL2 responsiveness, but not for endogenous IL2 production and subsequent proliferation by primed T cells.     

Different antigen-presenting cells differ in their capacity to induce lymphokine production and proliferation of an apo-cytochrome c-specific T cell clone

The activation of an apo-cytochrome c-specific T cell clone was found to differ, depending on the antigen-presenting cell population. Whereas total syngeneic spleen cells and bone marrow macrophages could be shown to trigger proliferation, IL 2, and MAF production by the T cell clone, a B cell lymphoma only induced MAF secretion. Further studies demonstrated that this effect was not due to a different antigen processing by the B lymphoma or to limiting amounts of Ia and antigen molecules on the B lymphoma cell surface. The dissociation of induction of MAF production from IL-2 production/proliferation found with the different antigen-presenting cells indicates strongly that molecules other than Ia and antigen may be required for the complete functional activation of antigen-specific T cell clones.     

Antigen presentation by interferon-gamma-treated endothelial cells and fibroblasts: differential ability to function as antigen-presenting cells despite comparable Ia expression

The effect of interferon-gamma (IFN-gamma) on endothelial cell (EC) and fibroblast (FB) class II major histocompatibility complex (MHC) gene product expression and antigen presenting ability was examined. Control FB did not express class II MHC gene products, whereas a small (less than 1%) population of passaged EC expressed class II gene products. IFN-gamma induced a comparable density of HLA-DR expression on nearly all EC and FB. IFN-gamma-treated EC and FB also expressed HLA-DP but at a lower density, whereas HLA-DQ expression was barely detectable on either cell type. Control FB were not able to stimulate allogeneic T4 cell DNA synthesis or function as antigen-presenting cells (APC). Control EC were also unable to stimulate allogeneic T4 cell DNA synthesis unless large numbers of stimulator cells were used. Small numbers of IFN-gamma-treated EC were able to stimulate allogeneic T4 cell DNA synthesis, whereas larger numbers were markedly more effective than control EC. In contrast, IFN-gamma-treated FB were ineffective stimulators of allogeneic T4 cell DNA synthesis. IFN-gamma-treated FB were able to present the exogenous antigen SKSD to autologous but not allogeneic T4 cells, but they were extremely inefficient APC. The inability of IFN-gamma-treated FB to function as APC could not be explained by FB-mediated immunosuppression, Ia density, or HLA-DQ expression. This limited capacity of IFN-gamma-treated FB to participate in Ia-restricted functional interactions with T4 cells correlated with a similar diminished capacity to support nonspecific mitogen-induced proliferation of T4 cells before IFN-gamma-induced Ia expression. This accessory cell function was not enhanced by IFN-gamma treatment. Monocytes syngeneic to the responding T4 cells but not interleukin 1 (IL 1) permitted IFN-gamma-treated FB but not control FB to stimulate allogeneic T4 cell DNA synthesis, but they remained markedly less effective stimulators than monocytes. Moreover, IFN-gamma-treated FB were effective stimulators of alloprimed T4 cells, in contrast to their inability to stimulate fresh T4 cells. Furthermore, monocytes and IFN-gamma-treated FB were comparably effective stimulators of alloreactive T cell lines. These data suggest that accessory cells perform functions unrelated to Ia and IL 1 that are necessary for mitogen-, alloantigen-, and antigen-induced proliferation of freshly isolated T cells. Monocytes and EC effectively perform this function, but FB do not. This accessory cell function does not seem to be as important for the activation of primed T cells.     

Accessory cells in immune suppression. I. Role of I-A+ accessory cells in effector phase idiotype-specific suppression of myeloma function

The suppression of MOPC 315 myeloma cells by idiotype-specific effector Ts requires the presence of non-immune AC. This requirement was demonstrated in cultures where myeloma targets and Ts were separated by cell-impermeable membranes or were in direct contact. The AC were adherent, radioresistant, and present in peritoneal exudates and in FcR+ as well as FcR- fractions of low density splenocytes; they bore cell surface I-A determinants and did not have to be H-2 compatible with myeloma cells and Ts. These studies demonstrate a novel role for Ia+ AC in immune regulation, and suggest that their accessory function may involve processing of T lymphocyte-derived suppressor factors or presentation of such factors to target cells.     

A subpopulation of adherent accessory cells bearing both I-A and I-E or C subregion antigens is required for antigen-specific murine T lymphocyte proliferation.

A murine T lymphocyte proliferation assay that used antigen-primed lymph node T cells, was antigen specific, and required exogenous accessory cells was used to characterize the accessory cells that supported proliferation. These cells were Thy 1.2 negative, radioresistant, glass-adherent, and were functional only if alive. The accessory cell function of spleen adherent cells was much greater than that of peritoneal cells. Also, the accessory cell function of spleen adherent cells was proportional to the length of time such cells were incubated with antigen and very small numbers of such cells provided accessory cell function. Cytotoxic studies with subregion-restricted anti-Ia antibodies and complement indicated that accessory cell function resided in a subpopulation of spleen adherent cells that bore both I-A and I-E or C subregion antigens. The function of such cells was not related to a selective ability (vs other spleen adherent cells) to take up antigen. These data indicate that antigen-specific stimulation of T lymphocyte proliferation requires at least one specific subpopulation of spleen adherent cells that can be phenotypically identified by its expression of Ia antigens and are consistent with the possibility that Ia antigens may be Ir gene products.     

Mouse splenic macrophage cell lines with different antigen-presenting activities for CD4+ helper T cell subsets and allogeneic CD8+ T cells.

A panel of seven mouse splenic macrophage cell lines, derived from cloned progenitors, was compared for their ability to present antigen to Th1 or Th2 helper T cell lines and hybridomas, as well as to naive T cells, and to provide accessory cell function for the synthesis of antibody from primed B cells. One of the cell lines expressed MHC class II molecules and was the only line with constitutive antigen-presenting activity for Th1 cells. It may represent a subset of splenic macrophages responsible for the activation of naive Th1 helper cells in situ. The remaining six cell lines responded to INF-gamma by up-regulating their class II expression and acquiring Th1 antigen-presenting activity. They may represent cells which, in situ, lack constitutive antigen-presenting activity but are promoted to presenting status by Th1-derived INF-gamma. Five of the cell lines provided accessory cell function to Th2 cells, as indicated by antibody synthesis in suspensions of spleen cells from primed mice depleted of their antigen-presenting cells. One of the cell lines lacking accessory cell activity had constitutive antigen-presenting activity for Th1 cells. This reciprocal expression of antigen-presenting activity supports the idea that Th1 and Th2 helper cells are activated by different antigen-presenting cells. Finally, the cell lines differed in their ability to constitutively induce an allogeneic response; a response that was limited to CD8+ T cells occurred in a CD4+ helper cell-independent manner and was unaffected by the addition of INF-gamma. The alloantigen-presenting macrophage cell lines also possessed the most efficient accessory cell activity for antibody synthesis. These cell lines, which represent a spectrum of antigen-presenting activities in the spleen afford models for defining the roles of macrophages in the induction of immune responses and for resolving issues concerning their development.     

Stimulation of normal inducer T cell clones with antigen presented by purified Ia molecules in planar lipid membranes: specific induction of a long-lived state of proliferative nonresponsiveness

Culture of normal inducer T cell clones with antigen and purified Ek beta:Ek alpha incorporated into planar lipid membranes resulted in specific T cell activation as determined by cell volume increase and IL 3 production. However, in contrast to results obtained with T cell hybridomas, antigen presentation by planar membranes did not induce measurable IL 2 production, and proliferative responses were not detected. Rather, recognition of only Ek beta:Ek alpha and antigen resulted in the specific induction of a long-lived state of proliferative nonresponsiveness to subsequent stimulation by conventional APC and antigen. Induction of nonresponsiveness required protein synthesis, and was not simply due to the absence of IL 2. The antigen-nonresponsive cells could respond to either PMA plus ionomycin or IL 2, and they expressed normal levels of surface antigen-receptor molecules. These results demonstrate that recognition by normal T cell clones of antigen and Ia molecules in the absence of other accessory cell molecules and signals results in a prolonged state of proliferative nonresponsiveness, possibly similar to a state of T cell tolerance in vivo.