anti-Ia Antibodies Research Articles

The Medawar Prize Acceptance Speech 2022.

The Medawar Prize Acceptance Speech 2022.

Autoimmunity, Insulin Resistance and Beta Cell Function in Subjects with Low Body Weight Type 2 Diabetes Mellitus

In developing countries like India, the majority of Type 2 diabetics are non-obese, and many are "lean" with a body mass index (BMI) below 18.5. This type is referred to as "Low Body Weight Type 2 DM" (LB Type 2 DM). LB Type 2 DM are confused with Type 1 DM or late autoimmune diabetes in adults (LADA) due to their high blood glucose levels and early insulin-requiring state. We assessed pancreatic islet cell autoimmunity by using both glutamic acid decarboxylase 65 antibody (GADA) and anti-IA(2) antibody estimation in 23 patients with LB Type 2 DM and 10 age-matched normal weight (NW) Type 2 DM. Fasting blood glucose (FBG) and 2 hr postchallenge blood glucose (PGBG2) were significantly higher in LB (p < 0.05), while mean values of fasting insulin (24.47 +/- 73.15 muIU/mL vs. 13.4 +/- 16.54 muIU/mL, p > 0.7) and fasting C-peptide (180.81 +/- 357.08 pM/mL vs. 279.83 +/- 281.38 pM/mL, p > 0.5) in LB and NW respectively were not statistically different. All 23 LB and 10 NW subjects were GADA negative while IA(2) positivity was found in 1/23 and 1/10 cases, respectively. LB Type 2 DM revealed good beta cell function with homeostasis model assessment beta cells (HOMAB) values of 57.41 +/- 153.18 as compared to 44.74 +/- 56.24 (p > -0.2) in NW Type 2 DM. Insulin resistance as assessed by homeostasis model assessment insulin resistant (HOMA IR) was 13.50 +/- 42.83 and 5.68 +/- 6.90 (p > 0.6) in LB and NW Type 2 DM, respectively, suggesting that LB Type 2 DM are a phenotypic variant of Type 2 DM.

Conjunctival inflammation induces langerhans cell migration into the cornea

The virtual absence of Langerhans cells (LC) in donor or recipient corneal epithelium is known to be an important factor in the acceptance of orthotopic corneal allografts. Though it is well known that various types of stimulation to the cornea induce LC migration into the corneal epithelium, resulting in poor graft survival, the influence of conjunctival inflammation on LC migration into the cornea has not been elucidated. Therefore we examined whether LCs migrate into the cornea in the presence of conjunctival inflammation. Sixteen BALB/c mice were divided into four groups. Group A: 4 mice with corneal inflammation induced by two 9-0 silk interrupted sutures in the central cornea (positive control); Group B: 4 mice with conjunctival inflammation induced by two 9-0 silk interrupted sutures in the temporal and nasal bulbar conjunctiva 1 mm from the limbus; Group C: 4 mice with conjunctival inflammation by two 10-0 nylon interrupted sutures in the temporal and nasal bulbar conjunctiva 1 mm from the limbus; and Group N: 4 mice with no inflammation (untreated, naive control). Fourteen days after suturing, the mice were sacrificed and LCs migrated into the corneal epithelium were counted by immunofluorescence assay using anti-Ia antibody. In Group A, Ia(+) cells in the cornea totaled 29.4 +/- 3.8 cells/mm(2); in Group B, 7.9 +/- 1.2 cells/mm(2); in Group C, 7.8 +/- 0.7 cells/mm(2); and in Group N, 1. 6 +/- 0.5 cells/mm(2). Significantly greater numbers of Ia(+) cells were detected in Groups A, B and C than in Group N (p < 0.005). Conjunctival inflammation caused by sutures in the bulbar conjunctiva induced LC migration into the cornea. These results indicate that conjunctival inflammation influences the corneal immunological environment, and may affect the fate of orthotopic corneal allografts.

Conjunctival inflammation induces langerhans cell migration into the cornea

Purpose. The virtual absence of Langerhans cells (LC) in donor or recipient corneal epithelium is known to be an important factor in the acceptance of orthotopic corneal allografts. Though it is well known that various types of stimulation to the cornea induce LC migration into the corneal epithelium, resulting in poor graft survival, the influence of conjunctival inflammation on LC migration into the cornea has not been elucidated. Therefore we examined whether LCs migrate into the cornea in the presence of conjunctival inflammation. Methods. Sixteen BALB/c mice were divided into four groups. Group A: 4 mice with corneal inflammation induced by two 9–0 silk interrupted sutures in the central cornea (positive control); Group B: 4 mice with conjunctival inflammation induced by two 9–0 silk interrupted sutures in the temporal and nasal bulbar conjunctiva 1 mm from the limbus; Group C: 4 mice with conjunctival inflammation by two 10–0 nylon interrupted sutures in the temporal and nasal bulbar conjunctiva 1 mm from the limbus; and Group N: 4 mice with no inflammation (untreated, naive control). Fourteen days after suturing, the mice were sacrificed and LCs migrated into the corneal epithelium were counted by immunofluorescence assay using anti-Ia antibody. Results. In Group A, Ia + cells in the cornea totaled 29.4 ± 3.8 cells/mm 2; in Group B, 7.9 ± 1.2 cells/mm 2; in Group C, 7.8 ± 0.7 cells/mm 2; and in Group N, 1.6 ± 0.5 cells/mm 2. Significantly greater numbers of Ia + cells were detected in Groups A, B and C than in Group N (p < 0.005). Conclusions. Conjunctival inflammation caused by sutures in the bulbar conjunctiva induced LC migration into the cornea. These results indicate that conjunctival inflammation influences the corneal immunological environment, and may affect the fate of orthotopic corneal allografts.

Selective Elimination of Antigen-Specific Line T Cells andex VivoAntigen-Primed Lymph Node Cells by Antigen-Targeted Drug-Labeled Antigen-Presenting Cell Membranes

Since antigen-specific autoaggressive T cells have been found in association with many autoimmune diseases, a treatment to eliminate such antigen-specific T cell clones was developed. The complex of peptide antigen and class II MHC protein is used to target a cytotoxic drug to antigen-specific T cells. The drug is bound covalently to antigen-presenting cells (APC) and protein antigens (Ag) are added to the cells for processing and presentation of peptides. The APC contain class II MHC (Ia) protein to present the peptide Ag to the T cell receptor and adhesion proteins for optimal interaction with the T cell. Either the Ag-bearing intact APC or Ia+membranes shed or released spontaneously from them were used as drug carriers to target the drug to the T cells. The drugs being used are phototoxic compounds. When irradiated with light of an appropriate wavelength, they give off toxic free radicals and singlet oxygen. These toxic by-products are short-lived and damage cells only in their immediate vicinity, cutting down on nonspecific side effects. APC from thymus cells, or their shed membranes bearing Ia–Ag peptide complexes, were able to target the phototoxic drug specifically to Ag-specific T line cells andex vivoAg-specific lymph node cells. Proliferation of the target T cells was inhibited at a three to four times lower drug concentration than required to affect control T cells. The Ag-specific effect was inhibited by anti-Ia antibody and by drug-free membranes carrying the Ag–Ia complex. This indicated that the antigen-specific phototoxic effect was mediated by interaction of the Ag–Ia complex with the T cell receptor.

MHC Class II-Mediated T Cell Response to DNA

Anti-DNA antibodies are elevated in several autoimmune diseases, including systemic lupus erythematosus. Yet, DNA was not shown to be presented by molecules of the major histocompatibility complex (MHC), nor was it reported to specifically stimulate T cellsin vivo,although these steps are essential for antibody production. We now demonstrate DNA-specific T cell activation, which involves presentation of DNA by MHC class II molecules. T cells, isolated from lymph nodes of mice immunized with a murine monoclonal anti-anti-DNA antibody, proliferated in response to DNA. Moreover, presentation of DNA by murine antigen-presenting cells could be inhibited with an isotype-specific anti-Ia antibody, and with peptides restricted by the same H-2 haplotype, suggesting that it is MHC class II-mediated. These results indicate that DNA can play a direct role in the regulation of T cells and in autoimmune processes.

The lymphoid substance of the chicken's harderian gland is organized in two histologically distinct compartments.

Light and electron microscopical investigations revealed that the lymphoid structure of the chicken Harderian gland is organized in different histological frameworks. In the head the surface epithelium of the central canal can be classified as a lymphoepithelial tissue which covers the dense lymphoid substance. It consists of small and medium-sized lymphocytes, dendritic-like cells, and occasional macrophages. High endothelial venules are associated with intense lymphocyte migration and homing that gives circumstantial evidence for a T-dependent region, as found in a secondary lymphoid organ. The B-dependent germinal centers are also common structural units of the head region's lymphoid substance. The body of the gland is loaded with plasma cells of different maturation stages. They immigrate into the epithelium of the central canal and produce IgM and IgA. Only a few scattered IgG producing plasma cells can be found in the gland of Harder. This plasmocytic region accounts for the immunosurveillance on the conjunctiva and in the upper respiratory tract through antibody production against bacterial or parasitic infections. In both the head and body regions of the gland, anti-B-L (anti-Ia) antibody recognized scattered elongated cells which might represent dendritic cells. The immunological relationship between the two histologically different parts of the Harderian gland is unknown, but we speculate that the dense lymphoid tissue with high endothelial venule receives the blood-borne, immunologically mature, but uncommitted B cells. By the influence of local antigen stimulus, these B cells transform to plasma cells which gradually appear in the body of the gland. The lymphoid structures of the head and the body fulfill the function of secondary and tertiary lymphoid organs, respectively.

Trinitrophenol Reactive T-Cell Hybridomas Recognize Antigens That Require Antigen Processing

Protein antigens must be taken up, processed, and displayed on the surface of antigen-presenting cells in association with major histocompatibility complex molecules before they can be recognized by T cells. Whether recognition of the haptens used to study allergic contact hypersensitivity in murine models similarly requires processing has not been determined. We analyzed whether presentation of trinitrophenol to trinitrophenol reactive T-cell hybridomas requires antigen processing by studying the effects of inhibitors of antigen processing and presentation on the ability of a syngeneic B-cell tumor (A20) to present trinitrophenol to a series of interleukin-2 producing, trinitrophenol specific, major histocompatibility complex class II-restricted T-cell hybridomas. The ability of trinitrophenol modified A20 cells to stimulate the hybridomas was completely inhibited by monoclonal, anti-trinitrophenol, or anti-Ia antibodies and was significantly reduced by paraformaldehyde fixation immediately after trinitrophenol modification. Trinitrophenol-modified A20 cultured at 37 degrees C for 2 h prior to fixation was significantly more effective at stimulating the hybridomas than trinitrophenol-modified A20 fixed immediately. The ability of A20 to present trinitrophenol was inhibited by chloroquine. Paraformaldehyde fixation and chloroquine treatment had similar effects on the ability of trinitrophenol modified lymph node dendritic cells to stimulate the trinitrophenol specific hybridomas. Paraformaldehyde fixation and chloroquine treatment had similar effects on the ability of A20 cells to present ovalbumin to ovalbumin-specific hybridomas as they had on the ability of trinitrophenol modified A20 cells to present trinitrophenol to the trinitrophenol specific hybridomas. One of seven T-cell hybridomas responded to trinitrophenol modified ovalbumin but not other trinitrophenol modified proteins. These results suggest that, at least in part, T cells in the contact hypersensitivity response to trinitrophenol recognize antigens that require processing and that trinitrophenol modified proteins can be recognized.

ALTERATION OF MONONUCLEAR CELL IMMUNE-ASSOCIATED ANTIGEN EXPRESSION, INTERLEUKIN-1 EXPRESSION, AND ANTIGEN PRESENTATION DURING INTRA-ABDOMINAL INFECTION

Intact peritoneal macrophage (M phi) function is critical to successful localization of intra-abdominal infection. Peritoneal macrophage antigen presentation capacity (APC), interleukin-1 (IL-1) expression, and immune-associated (Ia) antigen expression and abscess formation were determined following cecal ligation and puncture. APC and IL-1 expression were measured by coculture with a T-helper cell clone and by measuring subsequent proliferation. Ia expression was determined in blood, peritoneal M phi, and splenocytes using anti-Ia monoclonal antibody stain and flow cytometric analysis. Significant reductions in both Ia expression and APC were found 1 and 4 days after CLP with no change in IL-1 expression. Muramyl dipeptide, which enhances M phi phagocytosis, partially abrogated the depression in antigen presentation but did not affect Ia expression. Peritoneal M phi Ia expression and APC, but not IL-1 expression, were depressed after experimental peritonitis. The recovery of M phi function by day 14 coincides with clinical recovery and abscess formation, and restoration of early M phi depression may improve outcome.

Unexpected lack of reactivity of allogeneic anti-Ia monoclonal antibodies with MHC class II molecules expressed by mouse intestinal epithelial cells.

We have examined MHC class II molecules expression by murine gut epithelial cells using a large panel of anti-Ia antibodies. In contrast to conventional APC (i.e., B cells and macrophages), only two anti-Ia antibodies reacted with enterocytes: a xenogeneic rat anti-Ia mAb (CD311) directed against a monomorphic class II determinant, and a polyclonal antiserum directed against both I-A and I-E heterodimers. In contrast, allogeneic anti-Ia mAb were either unreactive (17 of 20) or reacted weakly (3 of 20) with enterocytes, even after in vivo treatment with IFN-gamma. This pattern of Ia reactivity of epithelial cells was tissue specific (restricted to gut mucosa) and cell specific (restricted to gut epithelial cells). Biochemical and molecular studies confirmed that enterocytes expressed I-A and I-E isotypes on their cell surface and contained mRNA of both subregion loci. Interestingly, enterocytes appeared deficient in expression of the MHC class II-associated invariant chain, and are not able to stimulate allogeneic T cells. These data suggest that gut epithelial cells express a conformation of class II molecules, antigenically distinct from that expressed on conventional APC.

Molecular Events in B Lymphocyte Activation: Consequences of Signals Transduced through MHC Class II Molecules

Evidence is presented to demonstrate that murine B lymphocytes receive growth stimulatory signals from their surface class II molecules. Monoclonal anti-la antibodies enhanced anti-μ-induced B cell proliferative response. The signals through surface immunoglobulin (Ig) and Ia appeared to act sequentially since preexposure to anti-μ was required to observe anti-la-induced increase in B cell proliferation. Anti-Ia antibodies did not increase the number of B cells entering the G 1 phase of cell cycle but always enhanced transition from G 1 into the S phase in response to stimulation with anti-μ. Analysis of early gene expression revealed that signaling through class II molecules led to an increase in anti-μ-induced expression of c-myc, a proto-oncogene, and of ornithine decarboxylase, a key enzyme in polyamine biosynthesis that has been shown to be intimately related to increased cell proliferation.

Deleterious effects of cis-urocanic acid and UVB radiation on Langerhans cells and on induction of contact hypersensitivity are mediated by tumor necrosis factor-alpha.

Ultraviolet B (UVB) light disrupts epidermal Langerhans cells (LC) universally and impairs the induction of contact hypersensitivity (CH) to epicutaneously applied haptens in certain strains of mice. Similar effects are observed when tumor necrosis factor-alpha (TNF alpha) is injected intradermally (ID) in mice. Trans-urocanic acid (UCA), a photoreceptor for UVB radiation, is known to be immunosuppressive. To determine whether cis-UCA is important in the process by which UVB and/or TNF alpha act in the skin, cis-UCA was injected ID into C57BL/6, C3H/HeN, BALB/c, and C3H/HeJ mice. Whole mounts of epidermis were removed 5 h later and stained immunochemically with anti-Ia antibodies. Microscopy revealed that Ia-bearing LC had lost their dendrites, had rounded up, and were reduced in number in all strains examined. Moreover, when dinitrofluorobenzene (DNFB) was applied epicutaneously to the injected site, induction of CH was grossly impaired. When neutralizing anti-TNF alpha antibodies were administered intraperitoneally 2 h prior to ID injection of cis-UCA, the deleterious effects on LC and CH induction were largely reversed. These results indicate that the actions of cis-UCA on LC and on CH induction are very similar to those achieved by ID injections of TNF alpha and by cutaneous exposure to low-dose UVB. Because the effects of UVB radiation and cis-UCA are reversed by anti-TNF alpha antibodies, we propose that UVB radiation impairs the induction of CH in mice by converting trans-UCA to cis-UCA within the epidermis; cis-UCA in turn causes the local release of TNF alpha, which thwarts sensitization by its ability to alter the functional program of epidermal Langerhans cells, thereby preventing the induction of CH.

Stimulation or tolerization of an anti-myelin basic protein T lymphocyte line with membrane fragments from antigen presenting cells

We have investigated the abilities of a cell-free supernatant of splenocytes or thymocytes, which have been incubated with myelin basic protein (MBP), and of membranes prepared by lysing these cells, to stimulate proliferation of a Lewis rat anti-MBP T lymphocyte line in vitro. The supernatant fraction, obtained by low-speed centrifugation, is thought to contain shed membrane fragments bearing class II MHC protein (Ia) and processed antigen. Almost all of 67 preparations of supernatant fraction and about a third ( 26 70 ) of the membrane preparations stimulate proliferation of the line cells in the absence of other antigen-presenting cells and antigen. Some membrane preparations bearing the synthetic peptide S69 (residues 69–89 of MBP), containing the immunodominant encephalitogenic determinant for the Lewis rat, instead of processed MBP could also stimulate proliferation. Those membrane preparations bearing either processed MBP or synthetic S69, which do not stimulate proliferation, induce a state of unresponsiveness in which the cells do not proliferate but produce inositol phosphate. Stimulation of proliferation and induction of unresponsiveness were both inhibited by anti-Ia antibody. Addition of cyclosporin A prevents induction of unresponsiveness. Addition of allogeneic splenocytes or the cell-free supernatant fraction of syngeneic or allogeneic splenocytes or thymocytes, prevents induction of unresponsiveness by providing a necessary costimulatory signal. Further fractionation of the cell-free supernatant by high-speed ultracentrifugation showed that the costimulatory signal resided in a particulate fraction which sedimented and not in the supernatant. These results indicate that the encephalitogenic peptide can induce anergy in T cells when presented on class II MHC in the absence of the costimulatory signal. Tolerizing forms of the membrane preparations which lack the costimulatory signal may be useful for in vivo treatment of autoimmune response.

Generation of helper T cells that recognize a cross-reactive idiotype through a network mechanism.

T cells that recognize the cross-reactive idiotype expressed on the heavy (H) chain of M104E (IgM, lambda 1) were induced in BALB/c mice by immunization with Dextran B-1355. T cells derived from mice immunized with 1 mg of Dextran B-1355 showed a marked proliferative response against M104E, whereas T cells from mice immunized with Ficoll or lesser amounts of Dextran B-1355 did not. BCL1Id, which had an immunoglobulin isotype identical to M104E, did not induce proliferation of the T cells. These T cells also proliferated against J558 (IgA, lambda 1) which shared the cross-reactive idiotype of the anti-alpha (1----3) glucosidic linkage antibody with M104E on the H chain. The T cells proliferated more efficiently against F(ab')2-104E, Fab-104E and H104E, the H chain of M104E, than against intact M104E. The T cell proliferative response against the idiotype on M104E or even H104E required macrophages as antigen-presenting cells (APC) and the response was inhibited when APC were treated with NH4Cl or chloroquine, inhibitors of antigen processing. Moreover, anti-CD4 antibody or anti-Ia antibody inhibited the proliferative response. These results indicated that anti-idiotypic T cells of the helper type, which recognized a cross-reactive idiotype associated with Ia molecules in processed form, could be induced physiologically through a network mechanism.

One-step double immunolabeling of mouse interdigitating reticular cells: simultaneous application of pre-formed complexes of monoclonal rat antibody M1-8 with horseradish peroxidase-linked anti-rat immunoglobulins and of monoclonal mouse anti-Ia antibody with alkaline phosphatase-coupled anti-mouse immunoglobulins.

A novel one-step double immunolabeling method was elaborated on the basis of the simultaneous application of preformed molecular complexes of two primary antibodies with their specific secondary antibodies labeled with different enzymes. Treatment with a rat monoclonal antibody (MAb), M1-8, pre-coupled with horseradish peroxidase-linked sheep anti-rat immunoglobulins, and enzyme reaction revealed by the 3-amino-9-ethylcarbazole/hydrogen peroxide reaction, resulted in red-brown intracytoplasmic staining of interdigitating reticular cells in the lymph nodes of Balb/c mice. Another molecular complex, made of mouse anti-Ia MAb with alkaline phosphatase-linked rabbit anti-mouse immunoglobulins, applied at the same time and then developed with naphthol AS-BI-phosphate/fast blue BB as substrate, yielded blue surface staining of this cell type in addition to labeling of B-lymphocytes. The method described provides the possibility of relatively rapid double antigen detection where the binding sites of the secondary antibodies are saturated by the specific primary immunoglobulins. This approach seems to avoid nonspecific binding of primary antibodies to Fc receptors, and the unwanted binding of secondary antibodies with cell surface immunoglobulins on B-lymphocytes or with crossreactive primary antibodies used in the other sequence, if the primary antibodies and the tissue are the same or crossreactive animal species.

Examination of Tetrachlorosalicylanilide (TCSA) Photoallergy Using In Vitro Photohapten-Modified Langerhans Cell-Enriched Epidermal Cells

Lymphocytes from BALB/c mice photosensitized in vivo to tetrachlorosalicylanilide (TCSA) were investigated to determine whether they could be stimulated to proliferate when cultured with Langerhans cell-enriched cultured epidermal cells (LC-EC) photohapten-modified in vitro with TCSA + UVA radiation. Cultured LC-EC were photohapten-modified in vitro by irradiation in TCSA-containing medium using a 1000-watt solar simulator equipped with filters to deliver primarily UVA radiation (320-400 nm). Lymphocytes from TCSA-photosensitized mice were incubated with LC-EC that had been treated in vitro with 0.1 mM TCSA and 2 J/cm2 UVA radiation (TCSA + UVA). Responder lymphocytes demonstrated a significant increase in their blastogenesis response compared to lymphocytes that were incubated with LC-EC irradiated with UVA prior to treatment with TCSA (UVA/TCSA) or with LC-EC that had received no treatment. Lymphocytes from naive mice or mice photosensitized with musk ambrette (MA) demonstrated a significantly lower response to LC-EC modified with TCSA + UVA, indicating the specificity of the response. Maximum blastogenesis response was achieved when LC-EC were treated with 0.1 mM TCSA and a UVA radiation dose of at least 0.5 J/cm2. Epidermal cells depleted of LC by treatment with anti-Ia antibody plus complement or by an adherence procedure were unable to stimulate this blastogenesis response. Epidermal cells treated in vitro with TCSA + UVA demonstrated enhanced fluorescence compared to control cells. The fluorescence observed was not restricted to any specific epidermal cell type; however, fluorescence microscopy studies revealed that dendritic Ia-positive cells, presumably LC, were also TCSA fluorescent. Flow cytometry showed that Ia-positive epidermal cells demonstrated the greatest UV fluorescence when treated with TCSA + UVA compared to both cells irradiated with UVA and subsequently treated with TCSA and untreated cells. This is consistent with the enhanced antigen presentation capability of TCSA + UVA treated LC-EC, which leads to the conclusion that LC photohapten-modified in vitro with TCSA + UVA demonstrate enhanced TCSA fluorescence and are capable of stimulating lymphocytes from TCSA photosensitized mice in an antigen-specific manner.

Haplotype-specific inhibition of homing of radiolabeled lymphocytes in experimental allergic encephalomyelitis following treatment with anti-IA antibodies

In vivo treatment with anti-IA antibodies has been shown to induce a haplotype-specific inhibition of EAE when the disease was following passive transfer of MBP-sensitized T cells. In order to determine the mechanism by which anti-IA antibody prevents passively transferred EAE, the homing of radiolabeled cells to the brain following anti-IA therapy was studied. Administration of anti-IA antibodies at the earliest onset of clinical signs of EAE prevented the homing of radiolabeled cells to the brain. In Fl (Balb/c X SJL/J) mice that developed EAE and received anti-IA s antibody there was a decreased homing of radiolabeled cells when compared to animals that received anti-IA d antibody. In addition, there was preferential expression of IA s antigen, over IA d antigen on capillary endothelium of the brain. The differential expression of IA antigens and the homing of radiolabeled cells in Fl (SJL X Balb/c) mice could in part explain the haplotypespecific suppression of disease following treatment with anti-IA antibodies.

L3T4 (CD4 +) cells that mediate contact sensitivity to trinitrochlorobenzene express I-A determinants

The present study has further characterized the T cell-mediated inflammatory response of contact sensitivity (CS) to the hapten trinitrochlorobenzene (TNCB) in mice. A discernible CS response was found to be induced as early as 2 days after epicutaneous application of TNCB. The response peaked on Days 4 to 5 and it then declined to a nearly undetectable level by Days 10 to 11. Examination of the draining lymph nodes demonstrated that development of CS coincided with an increase in cellular proliferation and in the total number of cells present. Despite a severalfold increase in the cellular contents of the draining lymph nodes of sensitized mice, the relative percentages of most subsets of T cells remained unchanged. Flow cytometric studies revealed that the subpopulation of T cells characterized as Thy 1.2 + L3T4 + I-A + increased substantially in comparison to its presence in unsensitized mice. Whether the Thy 1.2 + L3T4 + I-A + cells that increased following sensitization represented the effector population that mediates CS was then examined. Four-day immune lymph node T cells or L3T4 cells positively selected from them were capable of adoptively transferring CS to normal mice. However, these cells, after treatment with anti-Ia antibody or anti-I-A monoclonal antibody and complement, were unable to transfer CS. These findings imply that expression of I-A determinants may indicate antigeninduced T cell activation in vivo and that L3T4 cells that mediate CS are I-A positive.

Effect of high ligand concentration on West Nile virus‐specific T cell proliferation

In this paper the phenomenon of suppression of proliferation in vitro of 14 day primed, West Nile Virus (WNV)-specific, murine CD4+ T cells by large numbers of antigen-presenting macrophages and B cells has been investigated. Suppression was apparently not mediated by prostaglandins, as the use of indomethacin in cultures at four times the usual concentration did not reverse suppression. Experiments were designed to evaluate the contribution of major histocompatibility complex (MHC) Class II and nominal WNV antigens in causing suppression of T cell proliferation. Listeria- or thioglycollate-induced macrophages from CBA/H (H-2k) mice, when treated with heat-killed Listeria in vitro for 1 h to maintain or increase, respectively, MHC Class II levels before the addition of alloreactive Iak-specific T cells caused inverse dose-responses; the highest T cell proliferation occurred at a stimulator to responder (S : R) ratio of 0.25 and profound suppression at a S : R ratio of 1 or 2. In contrast, untreated thioglycollate-induced macrophages, which express low MHC Class II levels, gave a direct dose-response with increasing T cell proliferation as antigen-presenting cell (APC) numbers increased. Addition of anti-Ia antibodies (or their Fab fragments) to cultures caused a significant reversal of suppression of anti-WNV T cells imposed by high numbers of Listeria-induced macrophages or 14 day WNV-primed B cell APC. Suppression was also reversed by reducing the concentration of WNV antigen. These observations support the notion that the suppression of T cell proliferation observed at high S : R ratios was due to high concentrations of ligand (WNV-derived peptide complexed with Class II MHC) on APC.

Ontogeny of macrophage subpopulations and Ia-positive dendritic cells in pulmonary tissue of the rat

The development of macrophage subpopulations dendritic cells in the rat lung was studied from day 15 of gestation until day 21 after birth by means of immunohistochemical techniques combined with acid phosphatase staining. To characterize these cell populations, monoclonal antibodies raised against rat macrophage subpopulations were used (ED1, ED2, ED7, and ED8) in addition to anti-Ia antibodies. Ia-positive cells with a dendritic morphology were found on day 16 of gestation. During ontogeny, the number of these cells gradually increased. They were always found in mesenchymal lung tissue between the epithelial tubules of future alveoli, and in perivascular or peribronchial areas. ED1-positive macrophages were found on day 17 of gestation, with a distribution different from that of Ia-positive dendritic cells. The distribution of ED1-positive cells changed during ontogeny: before birth, ED1-positive cells were present in mesenchymal areas of lung tissue, whereas after the first week of postnatal life ED1 recognized all free alveolar macrophages. No Ia-expression was found on free alveolar macrophages. This developmental pattern resembles the ontogeny of Ia-positive dendritic cells and ED1-positive macrophages in gut-associated tissue. The comparable development of these cell populations in gut and lung tissue indicates a common ontogeny in the mucosal immune system.