Induction Of Ia Antigen Research Articles

Mononuclear cells in salivary glands of normal and isoproterenol-treated rats

The purpose of this study was to analyse the phenotypical distribution of resident cells of the mononuclear phagocyte system in rat salivary glands, and to determine whether isoproterenol induces alterations in macrophage and lymphocyte surface-marker expression. Frozen sections of gland tissues were prepared from five normal rats, and from six rats treated with 20 mg/kg isoproterenol/day for 10 days. A panel of six monoclonal antibodies was used to identify membrane markers associated primarily with monocytes (ED1), mature tissue macrophages (ED2), lymphoid macrophages (ED3), MHC class II (Ia) antigens (OX6), CD5-positive T lymphocytes (OX19), and rat B lymphocytes (OX33). Double-labelling techniques were used to detect the coexpression of ED1/ED2 and OX6/ED2 mononuclear cell markers in the major salivary glands. ED2- positive macrophages were predominant in all three major glands, ranging from 96 cells/0.87 mm 2 field in the parotid gland to 165 cells/0.87 mm 2 in the submandibular. OX19-positive T lymphocytes were rarely observed in submandibular and parotid glands but represented a distinguishing feature of the sublingual. Moderate numbers of ED3-positive macrophages also were detected in sublingual tissues. In the submandibular and parotid glands, isoproterenol resulted in a decrease in ED2-positive cells, but ED2-positive macrophages increased in sublingual glands with isoproterenol. Isoproterenol resulted in a decrease in MHC class II antigen expression on submandibular and sublingual mononuclear cells but an induction of Ia antigen in the parotid gland. Double labelling revealed that isoproterenol induced coexpression of ED1/ED2 markers on mononuclear cells in the submandibular glands, but ED1/ED2-positive cells were absent from other glands. However, coexpression of MHC class II markers on ED2-positive cells in the sublingual and parotid glands of normal rats was frequently observed, with isoproterenol decreasing coexpression in the sublingual gland and increasing it in the parotid. B lymphocytes were not detected in any of the glands examined. These findings indicate that important differences exist in normal resident mononuclear cell subsets among the major salivary glands of the rat. The differential effects of isoproterenol on inflammatory cells may reflect important differences in local salivary gland immunoregulation. Although salivary gland inflammation induced by isoproterenol does not appear to result from immune mechanisms, the rich population of T lymphocytes and ED3-positive macrophages, and presence of MHC class II antigens, suggest that the sublingual gland may function as an immune organ and have a role in mucosal immunity.

Antigen presentation of herpes simplex virus by corneal epithelium--an in vitro and in vivo study.

Ia antigen (class II antigen) is a histocompatibility antigen that foreign peptides associate with, before antigen presentation to T cells and subsequent triggering of the CD4 T cells. Although corneal epithelium is normally Ia negative it may become Ia positive under abnormal circumstances but the functional significance of this is uncertain. In this study the expression of Ia antigen on corneal epithelium of mice during in vivo primary and secondary herpes simplex keratitis and the in vitro accessory function of corneal epithelium in the presentation of herpes simplex virus (HSV) antigen to in vivo HSV primed T cells were evaluated. Whole mount preparations of corneal epithelium were found to express Ia antigen on days 3, 5, and 7 following corneal inoculation with live HSV. The intensity of the Ia expression was greater in non-immune mice on day 7 after corneal inoculation compared with immune mice. A cellular suspension of corneal epithelium induced HSV primed T cells to proliferate in the presence of HSV antigen. Induction of Ia antigen on corneal epithelium during herpes simplex keratitis may functionally expand the population of antigen presenting cells in the cornea and contribute to T cell activation.

Tetraethylammonium, a K + channel blocker, inhibits interferon-γ-induced major histocompatibility class II antigen (Ia) expression and DNA synthesis in rat astrocytes

Astrocytes play an important role in antigen presentation to T lymphocytes by their ability to express major histocompatibility class II (Ia) antigen upon exposure to a number of agents, including interferon-γ (IFN-γ). Astrocytes have been shown to express a variety of voltage-sensitive ion channels including voltage-sensitive K + channels. The function(s) of these channels in astrocyte functions is not clearly understood. In this investigation, we examined: (1) the comparative effects of mouse, rat, and human recombinant IFN-γ (rIFN-γ) on the induction of Ia antigen and DNA synthesis in rat astrocytes; (2) the effect of tetraethylammonium (TEA), a K + channel blocker, on rat IFN-γ-induced Ia expression and DNA synthesis in rat astrocytes. Our data show that all INF-γ induce DNA synthesis in rat astrocytes but only rat and mouse and not the human IFN-γ will induce Ia expression. TEA in a dose-dependent manner inhibited both Ia expression and DNA synthesis in rat astrocytes. The concentration kinetics of TEA with regard to maximum inhibition of Ia and DNA synthesis were different. Furthermore, these inhibitory effects were not a result of toxic or nonspecific effect of TEA on astrocytes as demonstrated by viability data and lack of any effect of tetramethylammonium, an analogue of TEA, that does not block K + ion channels. These data suggest a role of K + channels in Ia expression and DNA synthesis, therefore in immunological functions of astrocytes.

Lymphocyte subsets differentially induce class II human leukocyte antigens on allogeneic microvascular endothelial cells.

Increased expression of major histocompatibility complex class II (Ia) antigens on vascular endothelium is a common observation in allografts undergoing acute rejection. This phenomenon is generally ascribed to the host immune response directed against graft alloantigens, but its cellular and molecular basis are incompletely understood. In the present study we show that constitutively Ia-negative human microvascular endothelial cells (EC) can be induced to express surface class II human leukocyte antigens shortly after exposure to allogeneic lymphocytes in vitro. CD16+ (natural killer) and CD8+ (cytotoxic/suppressor) lymphocytes were efficient in triggering Ia antigen expression by EC, whereas CD4+ (helper/inducer) lymphocytes induced EC Ia expression only if cultured in the presence of autologous monocytes. Binding of lymphocytes to EC was shown to be essential for the subsequent induction of EC Ia, and anti-CD18 (LFA-1) antibody, which blocks lymphocyte-EC adhesion, was the only antibody of a panel of antilymphocyte antibodies that completely blocked the induction of EC Ia. Antibodies to interferon-gamma, which is a potent inducer of EC Ia, and to the CD3 T cell-surface antigen partly inhibited the induction of EC Ia by T cells, but neither antibody had any effect on Ia induction mediated by CD16+ cells, suggesting that T cells and natural killer cells utilize different mechanisms to induce Ia on EC. When combined with data from other laboratories indicating that Ia+ but not Ia- EC stimulate allogeneic T cell proliferation and cytotoxicity, our results suggest that the binding of EC by lymphocyte subpopulations followed by the induction of Ia antigen may represent the initial stage of incompatible allograft rejection.

Induction of macrophage Ia antigen expression by rIFN-gamma and down-regulation by IFN-alpha/beta and dexamethasone are mediated by changes in steady-state levels of Ia mRNA.

In previous studies, the induction of Ia antigens on murine peritoneal exudate macrophages by recombinant IFN-gamma (rIFN-gamma) and the antagonism of rIFN-gamma-induced Ia expression by the inhibitors IFN-alpha/beta and glucocorticoids have been examined. In this report, these findings have been extended to an analysis of total or cytoplasmic mRNA from macrophage cultures treated with rIFN-gamma in the absence or presence of these two inhibitors. Recombinant IFN-gamma induced a 5.7- to 6.5-fold increase in steady-state levels of Ia (A alpha-specific) mRNA. Coordinate increases in steady-state mRNA for A beta, and E alpha were observed in response to rIFN-gamma. Maximum induction occurred 24 hr post-treatment and required the continued presence of rIFN-gamma. Induction of A alpha-specific mRNA was sensitive to the protein synthesis inhibitor cycloheximide. Simultaneous treatment of macrophage cultures with rIFN-gamma and IFN-alpha/beta or the glucocorticoid dexamethasone (DEX) resulted in a significant decrease in steady-state, A alpha-specific mRNA levels compared with treatment with rIFN-gamma alone. This analysis suggests that both the induction of Ia expression by rIFN-gamma, and the antagonism of rIFN-gamma-induced Ia gene expression by IFN-alpha/beta and DEX, are regulated by cognate changes in Ia mRNA.

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.

Changes in cell surface antigen expression on human articular chondrocytes induced by gamma-interferon. Induction of Ia antigens.

Ia antigens (class II HLA molecules) have been detected on cells eluted from affected human cartilage in certain disease states, but not on normal cartilage cells. Because the presence of Ia antigens on chondrocytes may play an important role in rheumatic diseases, we investigated the induction of these molecules by gamma-interferon (gamma-IFN), a potent Ia-inducing lymphokine. Human articular chondrocytes were incubated with recombinant gamma-IFN, and the expression of Ia antigens was studied by cell sorter analysis, using a panel of reagents that detect monomorphic and polymorphic specificities of the DR and DQ Ia antigen families. While the induction of DR antigens, including polymorphic DR specificities, was readily obtained with gamma-IFN (50-95% positive cells), DQ antigens were negative or were displayed only on a lower percentage of chondrocytes (5-60%). In addition, incubation with gamma-IFN led to an increased expression of HLA class I antigens. The expression of various other surface markers either remained unchanged (as in 4F2 and BA-2) or showed tendencies toward decreased percentages (as in 83c2) or increased percentages (as in M phi R-17). No apparent change in cell morphology or growth pattern was observed.

Factors in AMLR culture supernatant which mediate the cytotoxic T-cell response to hapten-altered self: Interaction with macrophages

Supernatants from autologous mixed lymphocyte reaction (AMLR) cultures mediated the cytotoxic T-cell response in a system containing T cells and mitomycin C-treated, TNP-modified syngeneic thymocytes. No cytotoxic activity developed in the absence of the AMLR supernatants. The removal of thymic adherent cells abrogated the effect of the AMLR supernatants in the generation of cytotoxic cells. AMLR helper activity was restored following the addition of small numbers of splenic adherent cells. These results led to speculation that the AMLR supernatant interacted with accessory cells. Examination of the supernatant revealed significant levels of colony stimulating factor (CSF), and interferon-γ (IFN-γ). Functionally, incubation of the AMLR supernatant with P388D 1, a macrophage tumor line, resulted in the production of interleukin 1 (IL-1). CSF is a major inducer of IL-1 synthesis. In addition, the AMLR supernatant induced the expression of Ia antigens on the P388D 1 cells. IFN-γ has been reported to mediate this activity. Although both lymphokines were present and appeared to have an effect on macrophages, it was unknown if both were required for cytotoxic T-cell development in this system. Samples of AMLR supernatant were incubated for 10 min at various temperatures. The ability of the AMLR supernatants to mediate cytotoxic T-cell activation was sensitive to incubation at 80 or 90°C. Interestingly, at these temperatures CSF activity in the bone marrow colony-forming cell assay was enhanced. In separate experiments, AMLR culture supernatant was dialyzed against a pH 2 buffer. The induction of IA antigens by these supernatants was sensitive to dialysis at this pH. The similarity with IFN-γ provides further evidence that this activity was mediated by IFN-γ in the AMLR supernatants. The ability of the AMLR supernatants to mediate the cytotoxic T-cell response to altered self was also pH 2 sensitive. In contrast, IL-1 inducing capability and bone marrow colony growth, both CSF activities, were not reduced by pH 2 dialysis. Taken together, these data demonstrate a primary role for an IFN-like molecule, present in the AMLR supernatant, on cytotoxic T-cell activation. CSF involvement in this response is suggested by its IL-1 inducing activity, but cannot be definitively proven in the present study.

Expression of class II antigen in endotoxin induced uveitis.

Uveitis can be induced by systemic or intravitreal administration of endotoxin (lipopolysaccharide, LPS). In this study we correlated the expression of class II antigens (la in rat) of the Major Histocompatibility Complex (MHC), with this experimental model of uveitis. Ia antigen was detected by immunohistochemistry using the Avidin-Biotin-Peroxidase Complex (ABC) method and the monoclonal antibody OX6. Ia antigen was not expressed in normal eyes. However, Ia was expressed in the anterior uvea epithelial cells in all eyes with LPS induced uveitis. This study demonstrates that the ocular Ia expression is a localized process in the anterior uvea in response to systemic or intravitreal LPS. This response appears to be distinct from the action of LPS on macrophage Ia expression, where LPS has been shown to inhibit the induction of Ia antigen in macrophages by gamma interferon.

Induction of Ia antigens on thyroid cultures by syngeneic T lymphocytes: A model of autoimmune disease

We previously reported that the expression of Ia antigens on cultured monolayers of murine thyroid epithelial cells (TEC) occurred with a particular distribution exclusively on the basal part of the cultured thyroid cells, while class I antigens of the major histocompatibility complex (MHC) are only detected on the apical surface. It appears that deposition of syngeneic lymphocytes induces, 24 h later, Ia expression on the apical side of cultured TEC, the surface that is in direct contact with the responder lymphocytes during syngeneic sensitization of T lymphocytes. We hypothesized that this phenomenon could represent, in syngeneic situations, the restriction process in antigen recognition by T cells, as demonstrated by Ia restricted primary syngeneic sensitization (PSS) on murine TEC.

Induction of IA antigens in murine renal transplants.

Induction of IA antigens in murine renal transplants.

Soluble factors produced during an immune response regulate Ia antigen expression by murine adenocarcinoma and fibrosarcoma cells.

LT-85 is an alveologenic adenocarcinoma of C3Hf/HeN mice. Comparisons of the in vitro and in vivo surface properties of these cells revealed that under normal conditions, they expressed I-A and I-E antigens iv vivo only. By using clonally derived cells, it was established that this phenomenon was not due to the selection of an Ia antigen-positive tumor cell subpopulation, but resulted from phenotypic conversion of Ia antigen-negative tumor cells. These tumor cells and 1053 cells (a fibrosarcoma of C3H/HeN MTV- mice) could, however, be induced to express I-A, I-E, and much higher levels of H-2 antigens in vitro by co-culturing them with spleen cells from LT-85 tumor-bearing C3H/HeN MTV- mice. In vitro induction of Ia and H-2 antigens did not result from contaminating splenocytes or from antigen transfer, because splenocytes from BALB/c (H-2d) mice immunized with A/J (H-2k/d) cells were able to induce the expression of Iak antigens by both tumor cell lines. It was found that this phenomenon was neither H-2-restricted nor antigen-specific. The results clearly indicated, however, that an immune response was required to generate phenotypic conversion of the tumor cells, both in vivo and in vitro. It was further found that soluble, rather than cellular, factors produced during an immune response induced the expression of Ia antigens by LT-85 and 1053 tumor cells. In contrast to what has been reported about the induction of Ia antigens on macrophages and normal epithelial and endothelial cells, the induction of Ia antigens on LT-85 and 1053 cells did not appear to require T cells, and did not involve gamma-interferon. These findings demonstrate that some tumor cells are capable of altering their MHC antigen phenotype in response to factors produced during an immune response in vivo or in vitro. Because of the involvement of Ia antigens in several aspects of immune phenomena, the ability of tumor cells to differentially express Ia antigens in response to environmental factors may have profound effects on host-tumor interactions. Furthermore, the differences seen in the phenotypes of tumor cells grown in vitro and in vivo suggest that in vitro methodologies of tumor cell characterization may not present a complete picture of the natural state of the tumor cell surface.

Induction of Ia antigens on murine epidermal cells during the rejection of skin allografts.

Induction of Ia antigens on murine epidermal cells during the rejection of skin allografts.

Induction of Ia antigen in rat epidermal cells and gut epithelium by immunological stimuli.

The expression of Ia antigen in rat keratinocytes and gut epithelium was found to be inducible by a variety of immunological stimuli. Graft-vs.-host disease (GvHD) was accompanied by the appearance of Ia antigen in both sites, whereas local immunological stimuli, such as a contact-sensitizing agent applied to the skin and Trichinella spiralis infection of the gut, caused the expression of Ia antigen confined to the sites of contact of these stimuli with the tissues involved. Both T helper and T cytotoxic/suppressor subsets of parental lymphocytes, used to produce GvHD in F1 hybrid recipients, induce Ia expression in the skin and gut of these hosts, but simultaneous removal of both subsets from the donor inocula prevented induction. The Ia antigen expression associated with GvHD was shown to be of host origin but was not acquired from bone marrow-derived cells. Attempts to detect Ia antigen in serum or lymph of rats with GvHD gave negative results, and it was shown that Ia+ cells in the lamina propria of the small intestine did not take up detectable amounts of Ia antigen from the Ia+ intestinal epithelium. It appears that the local recognition of antigen by T lymphocytes can result in the induction of Ia antigen in keratinocytes and in the epithelial cells of the intestine. This antigen is synthesized by the cells in which it is found, and the observation that immunological stimuli are responsible for its appearance suggests that its role is an immunological one. Failure to find evidence that the gut epithelium Ia antigen was transferred to lymph or taken up by other Ia+ cells in the intestinal villi supports the view that this Ia (and, by analogy, that found in keratinocytes) serves a local function, and the possibility that it is involved in antigen presentation to T cells is discussed.

Differentiation of B Lymphocytes in C3H/HeJ Mice: The Induction of Ia Antigens by Lipopolysaccharide

The lipid A moiety of bacterial lipopolysaccharide (LPS) elicits several types of responses in murine B lymphocytes. First, lipid A induces the nonproliferative expression of cell surface antigens in more immature cell types. Second, lipid A induces a mitogenic response in more mature B cell types. Lipid A induces the expression of Ia antigens on bone marrow cells from C3H/DiSn but not C3H/HeJ mice. The Ia-inducible cells possess surface immunoglobulin. Agents that elevate intracellular levels of adenosine 3',5'-cyclic monophosphate (cyclic AMP) induce the appearance of Ia antigens on B lymphocytes from both C3H/HeJ and C3H/DiSn mice, suggesting that lipid A exerts its inductive effects by increasing cyclic AMP levels in cells. In contrast to what is observed by using other strains of mice, mature B lymphocytes from C3H/HeJ mice do not support a mitogenic response to lipid A. The subpopulation of B lymphocytes in C3H/HeJ mice that normally respond mitogenically to LPS not only appear to lack an LPS-response mechanism utilized in the mitogenic pathway, but they lack the LPS-response pathway of the immature B cell types. A lipid A-bound protein (LAP) induces both the expression of Ia and a mitogenic response in the different subpopulations of B lymphocytes from C3H/HeJ and C3H/DiSn mice. The genetic defect in C3H/HeJ mice that limits responses to lipid A may be associated with a receptor that is normally expressed on many different cell types.