Normal Antigen-presenting Cells Research Articles

B3a2 BCR-ABL fusion peptides as targets for cytotoxic T cells in chronic myeloid leukaemia.

Peptide sequences spanning the BCR-ABL protein junction potentially constitute novel leukaemia-specific antigens. 9-mer b3a2 fusion peptides have been reported to bind with high affinity to HLA-A3, -A11 and -B8. We have studied the effect of b3a2 BCR-ABL junctional peptides on the cytotoxic T-cell (CTL) response against normal and chronic myeloid leukaemia (CML) cells. Antigen-presenting cells (APCs) were prepared from HLA-A3- or -B8-positive peripheral blood mononuclear cells (PBMCs) by incubation with phytohaemagglutinin (PHA) and interleukin (IL)-2 for 7 d. These APCs were pulsed with the respective b3a2 junctional peptide in the presence of beta2-microglobulin and were then used to challenge autologous PBMCs at 7-d intervals in the presence of IL-2, IL-6, IL-7 and IL-12. On subsequent exposure to target cells (either further pulsed normal APCs or unpulsed CML cells), specific HLA-restricted CTL responses were observed against all HLA-A3/-B8 matched normal target cells tested, but not to targets that were HLA mismatched. Cytotoxicity was also induced against HLA-A3/-B8 unpulsed CML cells, but not against unmatched CML cells. These data indicate (i) that endogenous BCR-ABL junctional peptides may be presented by CML cells and (ii) that exogenous peptides are potential stimulators of autologous antileukaemic CTLs.

Early endosomal maturation of MHC class II molecules independently of cysteine proteases and H-2DM.

Major histocompatibility complex (MHC) class II molecules bind and present to CD4(+) T cells peptides derived from endocytosed antigens. Class II molecules associate in the endoplasmic reticulum with invariant chain (Ii), which (i) mediates the delivery of the class II-Ii complexes into the endocytic compartments where the antigenic peptides are generated; and (ii) blocks the peptide-binding site of the class II molecules until they reach their destination. Once there, Ii must be removed to allow peptide binding. The bulk of Ii-class II complexes reach late endocytic compartments where Ii is eliminated in a reaction in which the cysteine protease cathepsin S and the accessory molecule H-2DM play an essential role. Here, we here show that Ii is also eliminated in early endosomal compartments without the intervention of cysteine proteases or H-2DM. The Ii-free class II molecules generated by this alternative mechanism first bind high molecular weight polypeptides and then mature into peptide-loaded complexes.

The majority of H2-M3 is retained intracellularly in a peptide-receptive state and traffics to the cell surface in the presence of N-formylated peptides.

We used a new monoclonal antibody (mAb 130) to analyze the intracellular trafficking and surface expression of H2-M3, the major histocompatibility complex class Ib molecule that presents N-formylated peptides to cytotoxic T cells. M3 surface expression is undetectable in most cell types due to the paucity of endogenous antigen. M3 is induced on the cell surface by addition of high-affinity N-formylated peptides from mitochondria and listeria. Peptide-induced M3 expression is most efficient on antigen presenting cells. Basal and inducible expression of M3 is transporter associated with antigen processing (TAP)-dependent, distinguishing M3 from the class Ib molecules TL and CD1. Unlike the expression of class Ia molecules and a previously described M3/L(d) chimera, surface expression of M3 cannot be rescued by lowered temperature, suggesting that the alpha3 domain and transmembrane region of M3 may control trafficking. Pulse-chase analysis and use of trafficking inhibitors revealed a pool of empty M3 in the endoplasmic reticulum or early Golgi apparatus. Addition of exogenous peptide allows maturation with kinetics matching those of D(d). The lack of endogenous N-formylated peptide allows discovery of novel pathogen-derived peptides in normal antigen presenting cells. The nonpolymorphic nature of M3 and its ability to present bacterial antigens rapidly and dominantly make it an attractive target for peptide vaccination strategies.

Effects of UV on the migration and function of epidermal antigen presenting cells

Depletion of epidermal Langerhans cells (LC) and the concomitant depression of the skin immune system after excessive exposure to ultraviolet B light (UVB) has been established in the international literature for some time. Our investigations were intended to determine whether or not these phenomena occurred as a direct result of increased LC migration being triggered by the UVB exposure. To test this hypothesis, a sheep model was established in which the lymphatic vessels draining a defined region of skin were cannulated and the cells migrating towards the regional lymph node continuously collected. Cell populations in these collections were identified and enumerated by indirect immunofluorescence and flow cytometry. These experiments showed there was a significant, dose-dependent increase in the rate of LC migration from sheep skin after exposure to doses of UVB light exceeding 1 minimal erythemal dose (MED). In a series of parallel experiments, the functional characteristics of dendritic cells (DC) migrating from normal or UVB irradiated sheep were studied. To assay them, enriched preparations of DC were collected via cannulated afferent lymphatic vessels and pulsed with antigen prior to incubation with autologous peripheral blood lymphocytes. The relative efficiency of antigen presentation was determined by the ability of DC to induce T cell proliferation. Our data clearly demonstrate that there is a profound loss of normal antigen-presenting cell function after exposure to UVB light. Various experiments were undertaken to determine the mechanism(s) associated with these changes in migration kinetics and cellular function. Electron microscopic examinations of LC migrating from normal or UVB irradiated skin have demonstrated a profound loss of dendritic processes after UVB exposure. This provides a possible explanation for the changes in skin immunity after UVB exposure.

Cutaneous delayed type hypersensitivity in SCID mice adoptively transferred with lymphocytes is B cell independent and H-2 restricted.

Severe combined immunodeficiency (SCID) mice are largely devoid of functional T and B lymphocytes but have normal antigen presenting cell (APC) capacity. The authors have examined the requirements for cutaneous delayed type hypersensitivity (DTH) in SCID recipients by i.p. transfer of freshly isolated naive mature. T cells or non-fractionated spleen cells of H-2 compatible or incompatible origin. Recipient SCID mice were epicutaneously sensitized with oxazolone (OXA) within 24 h after cell transfer. SCID mice injected with as few as 10(5) H-2 compatible BALB/c (H-2d) spleen cells were able to mount DTH ear swelling reaction upon sensitization and challenge with OXA. Non-fractionated spleen cells from H-2 incompatible B6 or NZW mice were also able to restore DTH capacity in SCID recipients. However, when thymocytes were transferred, only donor mice expressing H-2d, but not H-2b and H-2z, haplotype restored DTH reactivity. Serum levels of IgM and IgM anti-OXA antibodies in SCID mice 27 days post-transfer with 10(7) BALB/c spleen cells were similar to that of intact donor mice. In contrast, specific antibodies of IgG isotype were approximately only one-tenth of that found in BALB/c controls. The results of this study show that for the development of cutaneous DTH, in SCID mice transferred with T cells, H-2 restricted APC-T cell interaction is required, whereas B cells are not mandatory. Also, SCID mice transferred with splenocytes show signs of defect immunoglobulin switch function. The authors believe that this model of DTH will be useful in delineating the effects of immunomodulatory substances in vivo on distinct host versus donor cell populations.

T cells are unresponsive to transgenic MHC class II molecules expressed on pancreatic beta cells.

It has been proposed that the autoimmune attack on the pancreatic beta cells leading to insulin-dependent diabetes mellitus can be caused by the expression of MHC class II molecules on the beta cells. Transgenic mice expressing normal levels of allogeneic MHC class II Ak on the beta-cell surface (IP-Ak) do not develop either insulitis or diabetes, yet these mice are not tolerant to Ak when expressed on normal antigen-presenting cells. The authors have stimulated T cells from IP-Ak mice in vitro with Ak-expressing beta cells. Mice were also primed in vivo in order to facilitate the antiallogeneic response. The authors found that neither IP-Ak positive nor IP-Ak negative mice were able to respond to Ak-expressing beta cells, and that in vivo priming does not overcome this inability. They suggest that beta cells do not act as antigen-presenting cells, probably due to inability of delivering costimulatory signals. This strengthens the notion that MHC class II expression per se is not sufficient to induce an autoimmune attack on the beta cells.

MHC class I presentation of live and heat-inactivated Sendai virus antigen in T2Kb cells depends on an intracellular compartment with endosomal characteristics.

T2Kb cells, which do not express TAP1/2 peptide transporters or the low molecular weight protein 2/7 (LMP2/7) proteasomal subunits, can still process and present both live and heat-inactivated Sendai virus (SV). As this operation may also reflect the existence of an alternative processing pathway in normal antigen-presenting cells (APC), the authors have characterized it using intracellular inhibitors and anti-Kb monoclonal antibodies (MoAbs). From the results with lipophilic amines (ammonium chloride, methylamine and chloroquine), cytoskeletal inhibitors (cytochalasin B and vinblastine), and an endoprotease inhibitor (phenylmethylsulfonyl fluoride, PMSF), the authors conclude that the processing of SV antigen in T2Kb cells has endosomal characteristics depending on cellular activities such as uptake, vesicular transport and intracellular-vesicular proteolysis. In addition, internalized 'empty' Kb molecules derived from the T2Kb cell surface appeared to be involved in the presentation of SV antigen, as demonstrated by a protocol using a combination of the Golgi inhibitor brefeldin A(BFA) and anti-Kb antibodies. The results thus indicate that T2Kb cells process SV antigen in an endosomal-like compartment which contain recycling 'empty' Kb molecules.

A Novel HLA Class II-Independent TCR-Mediated T Cell Activation Mechanism Is Distinguished by the Vβ Specificity of the Proliferating Oligoclones and Their Capacity to Generate Interleukin-2

Superantigens can induce proliferative T cell responses after complex formation with major histocompatibility complex (MHC) class II antigens. In this study, highly purified variant T cells from a histocompatibility leukocyte antigen (HLA) (human major histocompatibility complex) class II-deficient patient were stimulated with toxic shock syndrome toxin 1 (TSST-1) in the presence of either HLA class II-negative or normal antigen-presenting cells (APC). The proliferative responses were similar to those of normal T cells even when HLA class II structures were absent on both responding T cells and costimulatory APC. However, the Vβ specificity of responding T cells differed depending on the presence or absence of HLA class II antigens on the APC. In the presence of HLA class II-negative costimulatory APC, TSST-1 induced primarily proliferation of Vβ2-expressing T cells, whereas in the presence of normal APC virtually all responding T cells expressed Vβ elements different from Vβ2. In addition, in the presence of normal APC, T cells responding to TSST-1 produced much higher amounts of interleukin-2 than T cells proliferating in the presence of HLA class II-negative APC. These findings suggest that T cells bearing selected Vβ elements can directly interact with and respond to superantigen without involvement of HLA class II structures. Thus, our findings introduce a novel and distinct T cell receptor-mediated activation mechanism by which specific subpopulations of T cells respond to superantigen in the absence of HLA class II structures on APC.

(n-3) Polyunsaturated Fatty Acids Modulate the Expression of Functionally Associated Molecules on Human Monocytes in Vitro

Diets rich in (n-3) polyunsaturated fatty acids (PUFA) are associated with suppression of the immune system, but the mechanisms are unclear. Specific immune responses are initiated by antigen-presenting cells. This study examines the in vitro effect of the (n-3) PUFA eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on the expression of cell surface molecules required for normal antigen-presenting cell function on human blood monocytes. Monocytes were incubated with or without EPA or DHA for 48 h at 37°C. Following incubation, expression of major histocompatibility complex (MHC) class II molecules (HLA-DR, -DP and -DQ) and adhesion molecules [intercellular adhesion molecule-1 (ICAM-1) and leucocyte function associated antigen-1] was quantified by flow cytometry. In the presence of EPA alone there was a significantly lower median intensity of expression of HLA-DR and ICAM-1 relative to incubations without EPA. In contrast, significantly greater median intensities of expression of HLA-DR and -DP were observed following incubation with DHA. In parallel experiments, where monocytes were simultaneously activated by the addition of interferon-gamma to the cultures, median expression intensities of HLA-DR, -DP and ICAM-1 were significantly lower in the presence of either EPA or DHA compared with incubations without the (n-3) PUFA. These findings support previous animal studies that suggest that (n-3) PUFA can influence immune reactivity by modulating antigen-presenting cell function.

Poor loading of major histocompatibility complex class II molecules with endogenously synthesized short peptides in the absence of invariant chain.

In normal antigen-presenting cells, newly synthesized major histocompatibility complex (MHC) class II molecules associate with the invariant chain (Ii) glycoprotein in the endoplasmic reticulum (ER). They are loaded with peptides only after proteolytic removal of the Ii in post-Golgi endocytic vesicles. Since the Ii inhibits peptide binding to MHC class II molecules, this association could protect MHC class II molecules from being loaded with endogenous peptides early after biosynthesis. If this were an important function of the Ii in vivo, MHC class II molecules synthesized in cells lacking the Ii should be loaded efficiently with short endogenous peptides in the ER; such peptides are known to be present there due to TAP-mediated import from the cytosol. To examine this possibility, we have studied peptide loading in HeLa transfectants expressing murine H-2Ak MHC class II molecules either alone or together with an excess of Ii. Endogenous peptides could readily be extracted from conformationally intact Ak alpha beta dimers of biosynthetically labeled Ii+ cells, whereas peptide loading was greatly (> 95%) diminished in the absence of Ii. Significant amounts of sodium dodecyl sulfate-(SDS) stable 55-kDa peptide: Ak complexes were only found in the Ii+ transfectants. In the absence of Ii, the MHC class II molecules instead formed stable complexes with long (20 and 50 kDa) polypeptides. Known Ak-binding peptides bound stably to Ak molecules on Ii- cells, could be co-purified with them, and were resistant to release in SDS, suggesting that poor recovery of endogenous peptides was not due to decreased stability of Ak:peptide complexes in the absence of Ii. We conclude that protection of MHC class II molecules from endogenous short peptides does not appear to be a quantitatively important function of the Ii molecule, because peptide loading is inefficient in its absence.

Genetically engineered vaccines. Comparison of active versus passive immunotherapy against solid tumors.

Late tumor bearers (LTB), that is, mice that had tumors 3 weeks or longer, can have a selective immune dysfunction and fail to respond to target antigens expressed by the cancer cells. Such mice, however, do respond to nonmalignant cells engineered to express the rejection antigen, and they can be vaccinated with such cells to reject growing tumors. In this study, we compared the efficacy of passive immunization with that of active immunization using the engineered vaccine. An allogeneic MHC class I molecule was used as model tumor antigen. We found that active immunotherapy was only effective for small tumors in early stages of growth. In a later stage of tumor growth, active immunotherapy did not cure any mice, whereas passive immunotherapy was successful in all animals. Reasons for the failure of these LTB to respond to active vaccination with the engineered vaccine may be related to the decreased primary or secondary response we observed in these mice after active immunization. It is suggested that normal antigen-presenting cells expressing the tumor rejection antigen can elicit, in the presence of IL-2, antigen-specific T-cell responses by LTB, and that such T cells may be curative when used in adoptive therapy. We also suggest that the stage of tumor growth and the immune status of LTB more closely simulate the conditions observed in cancer patients.

Strong influence of the processing of the antigen on negative effects on T cell activation by regions outside the determinant area of bovine alpha s1-casein.

alpha s1-Casein can elicit a proliferative response in responding T cell clone 3D20 cells (specific for I-Ab plus fragment 136-151), even when using fixed splenic antigen-presenting cells (APC) not carrying antigen processing ability. The order of potency of each tested antigen for fixed APC was the determinant peptide (136-151) > the long peptide (136-195) > the intact protein (199 residues), indicating that regions outside the determinant area negatively affected the stimulatory potency of the antigens. On the other hand, the order for normal splenic APC was the short peptide > the intact protein > the long peptide. This shows that negative effects by regions outside the determinant area were strongly influenced by the antigen processing.

Direct binding of a synthetic multichain polypeptide to class II major histocompatibility complex molecules on antigen-presenting cells and stimulation of a specific T-cell line require processing of the polypeptide.

T-cell activation involves the recognition of foreign antigens as a complex with self-major histocompatibility complex (MHC) proteins on the surface of antigen-presenting cells (APC). Protein antigens usually require uptake by the APC and processing that results in the generation of peptide fragments. The branched synthetic polypeptide (Tyr, Glu)-Ala--Lys was chosen as a model antigen to follow the processing requirements, leading to T-cell activation. It has been demonstrated, by using fixed APC and various inhibitors of proteases, that (Tyr, Glu)-Ala--Lys has to be processed to stimulate a (Tyr, Glu)-Ala--Lys-specific T-cell line of C3H.SW (H-2b) origin to proliferate. To determine whether processing of (Tyr,Glu)-Ala--Lys is required to allow its association with the MHC class II molecules, biotin was covalently attached to it. Binding of the biotinylated (Tyr,Glu)-Ala--Lys to MHC class II gene products on the surface of intact normal APC was directly detected by phycoerythrin-streptavidin. The specificity of the binding was confirmed by its inhibition with anti-I-Ab antibodies as well as with excess of nonlabeled (Tyr,Glu)-Ala--Lys. Furthermore, introducing several inhibitors of proteases to the binding assay, we could substantiate that the proteolysis of (Tyr,Glu)-Ala--Lys is required to allow association of the resulting peptidyl T-cell epitopes with the MHC class II molecules themselves. The presence of the biotin moiety in the resulting peptides suggests that the T-cell epitopes of (Tyr,Glu)-Ala--Lys contain the N-terminal portion of the side chains of the branched polypeptide. An apparent Kd of 8.05 x 10(-8) M was determined, and optimal binding was detected after 10 hr of incubation with the antigen. The latter phenomenon is not due to slow uptake, since uptake of (Tyr,Glu)-Ala--Lys occurs mainly during the first 30 min of incubation, but rather reflects the events of processing that precede MHC interaction.

T-cell Clonal Anergy

Chemical fixation of splenic antigen-presenting cells (APC) with paraformaldehyde or 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (ECDI) destroys their ability to stimulate a proliferative response from interleukin-2 (IL-2)-producing T-cell clones (Jenkins and Schwartz 1987). In addition, exposure of the clones to antigen in the presence of the chemically fixed APC induces in them a hyporesponsive state to subsequent stimulation by normal APC and antigen (Fig. 1A). This induction of proliferative nonresponsiveness requires the correct allelic form of the major histocompatibility complex (MHC) class II molecule and the precise peptide recognized by the antigen receptor of the T-cell clone. These observations suggest that antigen-receptor occupancy is qualitatively normal and that the delivery of some other signal required for T-cell activation has been damaged by the fixation.

Cis-Urocanic Acid, a Product formed by Ultraviolet B Irradiation of the Skin, Initiates and Antigen Presentation Defect in Splenic Dendritic Cells In Vivo

Urocanic acid (UCA, deaminated histidine) is a major ultraviolet-absorbing component of the stratum corneum. On UV irradiation, the naturally occurring trans form converts to the cis isomer. We have previously postulated that UV-induced systemic suppression is initiated by cis-UCA by way of an antigen-presenting cell defect. To test this hypothesis further, we have investigated the antigen-presenting cell (APC) function of splenic dendritic cells (DC). Splenic DC were prepared from mice 7 days after 1 h UV irradiation (27 kJ/m2) or i.v. administration of 50-200 micrograms/mouse of cis- or trans-UCA. Dendritic cells from UV-irradiated or cis-UCA-treated mice had a significantly impaired (APC) ability, assessed by the proliferative response of purified T cells from mice immune to DNP6 OVA to DC pulsed with this antigen. Dendritic cells from mice given trans-UCA had normal APC ability. The number of FcR+ cells was the same in DCs from all four treatment groups, and the number of IAd+ cells and the intensity of IAd expression were not decreased in DCs from UV-irradiated or cis-UCA-treated mice. Mixture of DCs from UV- or cis-UCA-treated mice with DCs from normal mice did not suppress APC activity. Dendritic cells taken 3 days after UV or cis-UCA treatment, in contrast to DC taken 7 days after treatment, had normal APC ability, indicating a time delay in the generation of the APC defect. In contrast, addition of cis-UCA or trans-UCA (66 micrograms/ml) directly to an in vitro proliferation assay had no effect, suggesting that cis-UCA may be activated in vivo. These results support our original hypothesis that cis-UCA has a natural role as a modulator of immune function.

Molecular events in the induction of a nonresponsive state in interleukin 2-producing helper T-lymphocyte clones.

Exposure of normal interleukin 2 (IL-2)-producing helper T-cell clones to antigen and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide-treated antigen-presenting cells results in proliferative unresponsiveness to subsequent stimulation with antigen and normal antigen-presenting cells. In the present study, we have examined the molecular events that accompany the induction of this unresponsive state. T cells stimulated in this manner failed to produce IL-2, but interleukin 3, interferon-gamma, and IL-2 receptors were partially induced and T-cell receptor beta mRNA was fully induced. Although T-cell unresponsiveness correlated with an IL-2 production defect, addition of IL-2 during the induction phase failed to prevent development of the unresponsive state. The critical biochemical event appeared to be an increase in intracellular calcium. Removal of calcium from the medium prevented induction of the unresponsive state, whereas addition of the calcium ionophore ionomycin induced unresponsiveness as well as all of the related partial activation events. Thus, an increase in intracellular calcium under nonmitogenic conditions appears to initiate an alternative activation program that prevents the T cell from producing IL-2 in response to subsequent normal activation signals. The significance of this in vitro model for tolerance induction in vivo is discussed.

Quantitative analysis of the T cell response to antigen and planar membranes containing purified Ia molecules.

Planar lipid membranes containing the purified Ia molecule E beta k:E alpha k can present a peptide antigen derived from cytochrome c to the T cell hybridoma 2B4.11. The incorporation of E beta k:E alpha k into planar membranes was linear over a 120-fold range of Ia molecule concentrations, permitting the dependence of the T cell response on the Ia molecule concentration to be examined. As the Ia molecule concentration was increased in the planar membranes, two parameters changed: less antigen was needed to stimulate the T cells, and the plateau response seen at functionally saturating concentrations of antigen increased. The antigen sensitivity was analyzed by plotting the antigen concentration (log2) required to stimulate the release of 10 U of IL 2 from the T cells as a function of the Ia molecule concentration (log2). If the T cell recognized a simple unit of one antigen molecule and one Ia molecule, this plot should have generated a straight line with a slope of -1. Surprisingly, a line with a slope of -2.04 X/divided by 1.12 was observed, suggesting that the T cell might recognize one antigen molecule and two Ia molecules. This complexity, however, resulted from changes in the maximal response achieved at different Ia molecule concentrations. A similar phenomenon was observed when the Ia molecule concentration was decreased in cultures containing splenic antigen-presenting cells (APC) by the addition of an anti-E beta k:E alpha k monoclonal antibody, or the use of [B10.A(4R) X B10.PL]F1APC. The Ia molecule concentration can therefore be limiting for T cell hybridomas in cultures containing normal APC and functionally saturating amounts of antigen. When the planar membrane data were normalized to the maximal response to eliminate the effect of the changing plateau response, the resulting plot generated a line with a slope of -1.17 X/divided by 1.11. These results suggest that the sole stimulatory signal for this T cell hybridoma consisted of a 1:1 ratio of antigen and Ia molecules.

The effect of in vitro UV irradiation on the production of IL 1 by murine macrophages and P388D1 cells.

Ultraviolet irradiation (UV) exposure may lead to the development of multiple immunologic defects. One such defect is a dysfunction of normal antigen-presenting cell (APC) activation of T lymphocytes after whole body or in vitro UV. Although the mechanism of this interaction is not clearly defined, several possibilities have been suggested. One proposal is that UV may inhibit or abrogate the APC IL 1 signal and thus prevent normal T cell activation. To investigate this possibility further, we examined the functional consequences of UV on murine peritoneal adherent cell (PAC) activation of a cloned antigen-specific T cell hybridoma (A2.2.E10). In agreement with previous reports, we found a marked UV-induced inhibition of PAC activation of A2.2.E10 after sublethal UV. To correlate this UV-APC dysfunction with UV alterations of IL 1 production, both the IL 1-producing murine macrophage cell line P388D1 and normal murine PAC were exposed to various amounts of in vitro UV and the 24-hr post-UV IL 1 activity production of these cells was determined. The results surprisingly indicated that certain amounts of sublethal UV may actually augment the production of IL 1 activity, by using a dose range that clearly inhibits antigen presentation. This UV-induced activity was cycloheximide-sensitive, suggesting that de novo protein synthesis rather than release from cells was responsible for the increased IL 1 activity. In addition, the UV-induced IL 1 activity had a m.w. of 14K, consistent with previous reports, and demonstrated pyrogen activity when tested in the rabbit pyrogen assay. Thus UV clearly inhibits normal APC function; however, this may not be due to abrogation of IL 1 production, but rather the result of UV toxicity for other complex events involved in antigen presentation.

Adherent Ia + murine cell lines with characteristics of dendritic cells : II. Characteristics of I region-restricted antigen presentation

The ability of an adherent Ia +, interleukin 1 + (IL-1) tumor cell line (P388AD) to present turkey γ-globulin (TGG) to primed T lymphocytes was demonstrated and compared with normal antigen-presenting cells (APC) found in mouse spleen. P388AD tumor cells presented TGG to long-term cultures of TGG-reactive T cells (LTTC) and to lymph node-derived T cells which were enriched on nylon wool columns and subsequently depleted of endogenous antigen-presenting cells with anti-Ia antisera and complement. MHC-restricted antigen presentation by P388AD was observed when long-term cultures of TGG-reactive T cells were used as the responding T-cell population. Furthermore, antisera directed against I-region determinants expressed on the P388AD tumor cells inhibited TGG-specific T-cell proliferation in a dose-related fashion, suggesting a functional role for the tumor cell-associated Ia molecules. The kinetics of antigen presentation to LTTC by P388AD were similar to the kinetics observed for splenic APC, although the magnitude of the proliferative response to LTTC to TGG was generally lower when antigen (Ag) was presented by the tumor cells compared to splenic antigen-presenting cells (APC). However, the magnitude of T-cell proliferation of immune lymph node (LN) T cells was comparable when Ag was presented on tumor cells or splenic APC. Several experiments suggested that Ag uptake and/or processing may be less effective in P388AD tumor cells as compared to normal splenic APC. A nonadherent Ia +, IL-1 − tumor cell line (P388NA), which was isolated from the same parental tumor as P388AD, was also tested for the ability to present Ag to primed T lymphocytes and Ag-reactive LTTC. In contrast, to P388AD, the nonadherent tumor cell failed to present TGG under identical culture conditions even though Ia molecules were expressed on the tumor cells and Ag uptake had occurred. However, the defect in Ag presentation by P388NA could be corrected if an exogenous source of purified interleukin 1 was supplied to the cultures. A unique opportunity thus exists with both the P388AD and P388NA tumor cell lines to decipher some of the molecular interactions leading to T-cell proliferation during antigen presentation.