Suppression Of Delayed-type Hypersensitivity Response Research Articles

The effect of immune status, age and genetic background on induction of oral tolerance to Actinomyces viscosus in mice

The aim of the present study was to determine the effect of immune status, age and genetic background on the induction of oral tolerance to Actinomyces viscosus. Suppression of delayed type hypersensitivity (DTH) response and antigen-specific serum antibody levels could be induced in DBA/2 mice intragastrically and systemically immunized with A. viscocus, suggesting the induction of oral tolerance. In contrast, this immune suppression could be abrogated if the animals had been systemically immunized prior to the induction of oral tolerance with the same bacterium. Long-term systemic immunization prior to intragastric immunization with A. viscocus suppressed DTH response only. Cell transfer of this group of animals also suppressed DTH response in the donors, indicating the action of suppressor cells for inhibition of DTH response. Furthermore, oral tolerance to A. viscocus failed to occur in mice aged at 3days and 1, 2, 4, 6 and 36weeks old. Mice bearing H-2d haplotype were the most susceptible to oral tolerization, followed by H-2b and H-2k. Therefore, the results of the presence study suggest that the induction of oral tolerance to A. viscosus in mice may be dependence on the immune status and genetic background but not age.

Immunomodulatory effects of Stachytarpheta cayennensis leaf extract and its synergistic effect with artesunate.

BackgroundThe leaves of Stachytarpheta cayennensis C. Rich. (Verbenaceae) have been reported to possess potent anti-inflammatory and anti-malarial activities. Due to close association between inflammatory and immune responses, we evaluated the immunomodulatory activity of leaves extract of S. cayennensis. The combined effects of the leaves extract and artesunate, a standard antimalarial agent with immunomodulatory effects, were also evaluated.MethodsThe immunomodulatory activity of the methanol extract of the leaves of S. cayennensis (MESC) was evaluated in mice using the Delayed-Type hypersensitivity response (DTHR), primary and secondary humoral immune responses and the in vivo leucocyte mobilization tests. The immunomodulatory effect of artesunate and the combined effects of MESC and artesunate were evaluated using the phagocytic activity of polymorphonuclear neutrophils. Acute toxicity and lethality test in addition to the preliminary phytochemical studies of MESC were also performed.ResultsThe MESC exhibited 64.21% inhibition of DTHR at 500 mg/kg dose and evoked 139.64% of phagocytic stimulation at 100 μg/ml dose. Also MESC significantly (p < 0.05) showed dose related stimulation of humoral immunity and a highest percentage leucocyte mobilization of 10.15% at 250 mg/kg dose. Artesunate offered a non-significant (p > 0.05) percentage phagocytic stimulation (PPS) while the combined effect of artesunate and MESC exhibited a significant (p < 0.05) dose dependent PPS with highest PPS of 393.77% at 100 μg/ml. The LD50 of the MESC was estimated to be greater than 5000 mg/kg since there were no lethality and signs of acute intoxication after 48 h observation. Preliminary phytochemical analysis revealed the presence of carbohydrates, glycosides, flavonoids, saponins, alkaloids, terpenoids and steroids.ConclusionsThe results of this study indicated that MESC possesses immunostimulatory action with significant synergistic effects with artesunate, and can therefore, offer immune boosting activities in disorders of immune suppression.

The induction of anterior chamber associated immune deviation with Type II collagen (P1045)

Abstract The administration of antigens into the anterior chamber (AC) of the eye induces a special form of antigen-specific peripheral immune tolerance termed AC-associated immune deviation (ACAID), which prevents delayed-type hypersensitivity (DTH) responses and other inflammatory responses. Type-II collagen (CII) is highly expressed in cartilage tissues and has been linked to Rheumatoid arthritis, aging, and osteoarthritis. To explore the potential for ACAID induction via CII, we checked for different signs of ACAID generation following the AC injection of CII in BALB/c mice. We hypothesized that the mechanism of ACAID induction involves efferent T regulatory cells (Tregs). Both local adoptive transfer (LAT) assays and DTH assays were performed. Results indicated that ACAID induction was driven by the AC injection of CII. Spleen cells of mice injected with CII in the AC significantly suppressed DTH responses. ACAID induction was mediated by efferent Tregs in the spleen. CII-mediated ACAID induction could have multiple therapeutic implications, especially in autoimmunity.

Type II Collagen Induces Peripheral Tolerance in BALB/c Mice via the Generation of CD8+ T Regulatory Cells

Antigens introduced into the anterior chamber (AC) of the eye induce a potent form of antigen-specific peripheral immune tolerance termed AC-associated immune deviation (ACAID), which prevents inflammatory immune responses and is characterized by impaired delayed-type hypersensitivity (DTH) responses. Type-II collagen (CII) is a fibrillar protein expressed exclusively in cartilage tissues. Although of its clinical relevance to Rheumatoid arthritis, aging, and osteoarthritis, there have been no studies to date to test if CII has the ability to induce ACAID. We hypothesized that ACAID could be generated via AC injection of CII in BALB/c mice. Using a DTH assay, the hypothesis was supported and led to another hypothesis that CII is capable of inducing specific immune tolerance via CD8+ T regulatory cells (Tregs). Thus, we performed functional local adoptive transfer (LAT) assays to examine the regulatory roles of spleen cells, T cells, and CD8+ T cells in the specific immune regulation induced by CII injection into the AC. Results indicated that CII induced ACAID when injected into the AC. Spleen cells of mice injected with CII in the AC significantly suppressed DTH responses. The T cell compartment of the spleen was capable of expressing this suppression. CD8+ Tregs could solely express this CII-driven suppression and even exerted more noticeable suppression than spleen cells or splenic T cells. This study suggests a crucial role for CD8+ Tregs in mediating CII-driven ACAID-mediated immune tolerance. This could have therapeutic implications in Rheumatoid arthritis, aging, osteoarthritis, and other diseases in which CII is involved.

Tumor-Derived Exosomes Confer Antigen-Specific Immunosuppression in a Murine Delayed-Type Hypersensitivity Model

Exosomes are endosome-derived small membrane vesicles that are secreted by most cell types including tumor cells. Tumor-derived exosomes usually contain tumor antigens and have been used as a source of tumor antigens to stimulate anti-tumor immune responses. However, many reports also suggest that tumor-derived exosomes can facilitate tumor immune evasion through different mechanisms, most of which are antigen-independent. In the present study we used a mouse model of delayed-type hypersensitivity (DTH) and demonstrated that local administration of tumor-derived exosomes carrying the model antigen chicken ovalbumin (OVA) resulted in the suppression of DTH response in an antigen-specific manner. Analysis of exosome trafficking demonstrated that following local injection, tumor-derived exosomes were internalized by CD11c+ cells and transported to the draining LN. Exosome-mediated DTH suppression is associated with increased mRNA levels of TGF-β1 and IL-4 in the draining LN. The tumor-derived exosomes examined were also found to inhibit DC maturation. Taken together, our results suggest a role for tumor-derived exosomes in inducing tumor antigen-specific immunosuppression, possibly by modulating the function of APCs.

Chronic exposure to dim light at night suppresses immune responses in Siberian hamsters.

Species have been adapted to specific niches optimizing survival and reproduction; however, urbanization by humans has dramatically altered natural habitats. Artificial light at night (LAN), termed 'light pollution', is an often overlooked, yet increasing disruptor of habitats, which perturbs physiological processes that rely on precise light information. For example, LAN alters the timing of reproduction and activity in some species, which decreases the odds of successful breeding and increases the threat of predation for these individuals, leading to reduced fitness. LAN also suppresses immune function, an important proxy for survival. To investigate the impact of LAN in a species naive to light pollution in its native habitat, immune function was examined in Siberian hamsters derived from wild-caught stock. After four weeks exposure to dim LAN, immune responses to three different challenges were assessed: (i) delayed-type hypersensitivity (DTH), (ii) lipopolysaccharide-induced fever, and (iii) bactericide activity of blood. LAN suppressed DTH response and reduced bactericide activity of blood after lipopolysaccharide treatment, in addition to altering daily patterns of locomotor activity, suggesting that human encroachment on habitats via night-time lighting may inadvertently compromise immune function and ultimately fitness.

The generation of a ‘tolerogen’ after the ingestion of ovalbumin is time-dependent and unrelated to serum levels of immunoreactive antigen

In an attempt to investigate the molecular basis of the mechanisms underlying oral tolerance, we have evaluated the molecular and biological features of ovalbumin subjected to intestinal processing. Immunoreactive ovalbumin absorbed by the gut was measured by a sandwich ELISA at different times after feeding 25 mg ovalbumin to adult mice. Ovalbumin was detected as early as 5 min after the feed (36.7 +/- 16 ng/ml; mean +/- 1 s.d.) and reached maximal levels at 1 h (73.3 +/- 20 ng/ml). Pooled mouse serum, collected 5 min or 1 h after the feed, was transferred intraperitoneally into the naive recipients. Suppression of systemic delayed-type hypersensitivity (DTH) was found in mice receiving 0.8 ml of serum obtained 1 h after ovalbumin feeding but not when using serum obtained 5 min after feeding. In order to transfer samples containing similar levels of ovalbumin, an increased amount (1.3 ml) of serum collected 5 min post-feed was used in further experiments but again failed to induce DTH tolerance. Serum samples obtained 5 and 60 min after ovalbumin feeding were analysed by fast-protein liquid chromatography (FPLC) fractionation followed by ELISA. Both the charge characteristics and molecular weight of intestinally absorbed ovalbumin were indistinguishable from native ovalbumin. Although intact native ovalbumin is the only molecular species detected by ELISA, the results suggest that this has no role in the suppression of DTH responses.

Metalloproteinase Inhibition Has Differential Effects on Alloimmunity, Autoimmunity, and Histopathology in the Transplanted Lung

Upregulation of matrix metalloproteinases (MMPs) has been associated with chronic lung allograft rejection known as bronchiolitis obliterans syndrome. It has been suggested that MMP inhibition could prevent the rejection response. However, the effect of MMP inhibition on lung allograft rejection has not been reported. Utilizing a rat model of lung transplantation, tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) were overexpressed by gene therapy in F344 rat lung allografts prior to transplantation into WKY recipient rats. Separately, WKY rats that received F344 lung allografts were treated systemically with COL-3, a global MMP inhibitor. TIMP-1 and TIMP-2 had differential effects on delayed type hypersensitivity (DTH) responses to donor antigens and type V collagen, an autoantigen involved in the rejection response. Neither TIMP-1 or TIMP-2 affected the onset of rejection pathology. COL-3 suppressed DTH responses to donor antigens and type V collagen, abrogated local production of tumor necrosis factor-alpha, and interleukin-1beta. Although it did not prevent rejection pathology, COL-3 (30 mg/kg) induced intragraft B cell hyperplasia suggestive of posttransplant proliferative disorder (PTLD). These data identify a complex role for MMPs and TIMPs in the immunopathogenesis of lung allograft rejection, and indicate their effects are not limited to matrix remodeling.

Immune Function in PTSD

Disturbed regulation of both the hypothalamic-pituitary-adrenal (HPA) axis and sympathoadrenomedullary system in posttraumatic stress disorder (PTSD) suggests that immune function, which is modulated by these systems, may also be dysregulated. Two dermatologic, in vivo measures of immune function, delayed-type hypersensitivity (DTH) and skin barrier function recovery, were examined in female subjects with PTSD and compared to measures in healthy female comparison subjects. In addition, at the time of DTH test placement, circulating numbers of lymphocyte subtypes were assessed. In separate studies, the effects of acute psychological stress on DTH and skin barrier function recovery were examined in healthy volunteer subjects. Both DTH and barrier function recovery were enhanced in women with PTSD. These findings contrast with the effects of acute stress in healthy control subjects, which was associated with suppression of DTH responses and skin barrier function recovery. There was no difference between subjects with PTSD and healthy control subjects in proportions of circulating lymphocyte subsets or in expression of the lymphocyte markers CD62, CD25, and CD45RO/CD45RA. These results suggest that cell-mediated immune function is enhanced in individuals with PTSD, a condition that imposes chronic physiologic and mental stress on sufferers. These findings contrast with suppression of DTH and skin barrier function recovery in healthy volunteers in response to acute psychological stress.

Expression of Tim-3 Is Transiently Increased before Development of Anterior Chamber-Associated Immune Deviation

Purpose: To assess the expression of T-cell immunoglobulin- and mucin-domain-containing molecule 3 (Tim-3) in the spleens of BALB/c mice undergoing anterior chamber-associated immune deviation (ACAID). Methods: ACAID was generated after intracameral (i.c.) injection of ovalbumin (OVA) into BALB/c mice and evaluated by assessing the delayed-type hypersensitivity (DTH) response following a subsequent subcutaneous (s.c.) injection of OVA emulsified in complete Freund's adjuvant (CFA) on Days 0, 3, 7, 14, 21 and 28. Tim-3 mRNA levels were detected using real-time RT-PCR, and the frequency of CD4+Tim-3+ T cells in splenocytes as well as the coexpression of Tim-3 with CD25 on CD4+ T cells were assessed by flow cytometry. Results: A significantly suppressed DTH response was observed on Days 7, 14, 21, and 28, but not on Days 0 and 3 during the development of ACAID. The levels of Tim-3 mRNA and the frequency of CD4+CD25+Tim-3+ T cells in the splenocytes reached a peak on Day 3, declined on Day 7, and returned to basal levels thereafter. Conclusions: A transient upregulation of Tim-3 expression was observed in the early stage of ACAID, suggesting its possible involvement in the development of ACAID.

Topical exposure to exogenous ultraviolet-irradiated urocanic acid enhances Mycobacterium ulcerans infection in a Crl:IAF(HA)-hrBR hairless guinea-pig model of Buruli ulcer disease.

Ultraviolet radiation (UVR) pre-exposure enhances Mycobacterium ulcerans infection in the Crl:IAF(HA)-hrBR hairless guinea-pig, possibly via a photoimmunosuppressive mechanism. The trans-cis photoisomerization of epidermal urocanic acid is an important initiator of the web of events leading to photoimmunosuppression. Thus, the hypothesis tested in this paper was that topical pre-exposure to UVR-irradiated urocanic acid mixture containing cis-urocanic acid (UVR-UCA) enhances the ulcerative form of M. ulcerans infection in the Crl:IAF(HA)-hrBR hairless guinea-pig model of human Buruli ulcer disease. Groups of six animals were subjected to daily topical treatment with either 0 (vehicle only), 0.1, 0.5 or 1 mg of trans (tUCA) or UVR-UCA (contained a cis : trans urocanic acid isomer ratio of 1 : 9) for three consecutive days. A sham treatment group was also included in the experiment. Three days following their final treatment, the guinea-pigs were intradermally infected in the right dorsal flank with 1.5 x 107 CFU of M. ulcerans in 0.1 ml of phosphate-buffered saline (PBS) and sham infected with 0.1 ml of PBS in the left dorsal flank. The resultant skin lesions were then measured over the next 21 days. At day 21 postinfection, the animals were tested for delayed-type hypersensitivity (DTH) reactivity to M. ulcerans cell fragment antigens (MCF). Distinct, well-demarcated, dermally situated skin nodules were present at infected, but not sham-infected, skin sites by day 3 postinfection, and the lesions progressed to frank ulcers by day 5. Between days 5 and 21, the mean lesion diameters of the UVR-UCA-treated animals were significantly (P<0.001) greater than those of the sham, vehicle only or tUCA-treated groups. UVR-UCA-treated guinea-pigs also had significantly (P<0.001) suppressed DTH responses to MCF compared with the other treatment groups. There were no significant (P>0.4) differences between the lesion sizes and DTH responses of the tUCA, vehicle only or sham treatment groups. These results demonstrate that topical exposure to UVR-UCA promotes M. ulcerans infection and suppresses DTH responses to M. uclerans antigens in infected animals. These results lend credence to the hypothesis that UVR-mediated enhancement of Buruli ulcer disease in the Crl:IAF(HA)-hrBR hairless guinea-pig model occurs via modulation of cis-urocanic acid-susceptible immune pathways.

Induction of suppressor cells following mucosal presentation of Actinomyces viscosus in mice.

Mucosal presentation of Actinomyces viscosus results in antigen-specific systemic immune suppression, known as oral tolerance. The aim of the present study was to determine the mechanism by which this oral tolerance is induced. DBA/2 mice were gastrically immunized with A. viscosus. Serum, Peyer's patch (PP) and spleen cells were transferred to syngeneic recipients which were then systemically challenged with the sameiA. viscosus strain. To determine antigen-specificity of cells from gastrically immunized mice, recipients which received immune spleen cells were also challenged with Porphyromonas gingivalis. One week after the last systemic challenge, the delayed type hypersensitivity (DTH) response was determined by footpad swelling and the level of serum IgG, IgA and IgM antibodies to A. viscosus or P. gingivalis measured by an ELISA. No suppression of DTH response or of specific serum antibodies was found in recipients which received serum from gastrically immunized mice. Systemic immune suppression to A. viscosus was observed in recipients which had been transferred with PP cells obtained 2 days but not 4 and 6 days after gastric immunization with A. viscosus. Conversely, suppressed immune response could be seen in recipients transferred with spleen cells obtained 6 days after gastric immunization. The immune response to P. gingivalis remained unaltered in mice transferred with A. viscosus-gastrically immunized cells. The results of the present study suggest that oral tolerance induced by A. viscosus may be mediated by antigen-specific suppressor cells which originate in the PP and then migrate to the spleen.

Regulatory functions of self-restricted MHC class II allopeptide-specific Th2 clones in vivo.

We studied T-cell clones generated from grafts of rejecting and tolerant animals and investigated the regulatory function of Th2 clones in vitro and in vivo. To prevent allograft rejection, we treated LEW strain recipient rats of WF strain kidney grafts with CTLA4Ig to block CD28-B7 costimulation. We then isolated epitope-specific T-cell clones from the engrafted tissue, using a donor-derived immunodominant class II MHC allopeptide presented by recipient antigen-presenting cells. Acutely rejected tissue from untreated animals yielded self-restricted, allopeptide-specific T-cell clones that produced IFN-gamma, whereas clones from tolerant animals produced IL-4 and IL-10. Adoptive transfer into naive recipients of Th1 clones, but not Th2 clones, induced alloantigen-specific delayed-type hypersensitivity (DTH) responses. In addition, Th2 clones suppressed DTH responses mediated by Th1 clones in vivo and blocked Th1 cell proliferation and IFN-gamma production in vitro. A pilot human study showed that HLA-DR allopeptide-specific T-cell clones generated from patients with chronic rejection secrete Th1 cytokines, whereas those from patients with stable graft function produce Th2 cytokines in response to donor-specific HLA-DR allopeptides. We suggest that self-restricted alloantigen-specific Th2 clones may regulate the alloimmune responses and promote long-term allograft survival and tolerance.

ICAM-1 deficiency suppresses host allosensitization and rejection of MHC-disparate corneal transplants.

We used a murine model of orthotopic corneal transplantation to determine whether host deficiency in ICAM-1 promotes survival of corneal grafts with different degrees of allodisparity. ICAM-1-/- and wild-type C57BL/6 (ICAM-1+/+) received corneal grafts from the following strains of mice: BALB/c (fully mismatched), BALB.b (mismatched at multiple minor H only), or B10.D2 [including major histocompatibility complex (MHC) mismatch]. Graft rejection, induction of allospecific delayed-type hypersensitivity (DTH) responses, and leukocytic infiltration of grafts were measured. There were no differences in long-term survival of allografts that were either fully mismatched or had only minor H disparity in ICAM-1+/+ vs. ICAM-1-/-hosts. However, whereas B10.D2 grafts were accepted in only 58% of the ICAM-1+/+ hosts, graft survival in ICAM-1-/- recipients was 100% (P=0.006). Moreover, none of the ICAM-1-/- mice receiving B10.D2 grafts developed allospecific DTH. Prolonged survival seen in MHC-mismatched grafts in ICAM-1-/- mice, along with a suppressed DTH response to donor alloantigens after transplantation, suggest that ICAM-1 is associated with recipient sensitization to MHC alloantigens.

Estrogen mediates the sex difference in post-burn immunosuppression.

Previous studies in our laboratory have demonstrated that cell-mediated immune function was suppressed in female, but not male, mice at 10 days after burn injury and was mediated, in part, by increased production of interleukin-6 (IL-6). Because 17beta-estradiol (E(2)) influences immune function after trauma and the hormone is known to regulate IL-6 production, the effect of E(2) on immune function after thermal injury was examined. Increased circulating concentrations of E(2) corresponded with suppressed delayed-type hypersensitivity (DTH) and splenocyte-proliferative responses, and increased circulating concentrations of IL-6 in female mice after burn. Ovariectomy restored the suppressed DTH response and decreased IL-6 concentrations, and administration of exogenous E(2) to both ovariectomized females and intact male mice resulted in a suppressed DTH response. In addition, in vitro treatment with E(2) suppressed splenocyte proliferation in a macrophage-dependent manner and enhanced macrophage production of IL-6. These results strongly suggest that the sex difference in cell-mediated immunity 10 days after burn injury is mediated by altered concentrations of E(2), which in turn modulate key macrophage-derived immunoregulatory cytokines.

Reduced Ultraviolet-Induced Carcinogenesis in Mice with a Functional Disruption in B7-Mediated Costimulation

Immunosuppression by UV light contributes significantly to the induction of skin cancer by suppressing the cell-mediated immune responses which control the development of carcinogenesis. The B7/CD28-CTLA-4 signaling pathway provides costimulatory signals essential for Ag-specific T cell activation. To investigate the role of this pathway in photocarcinogenesis, we utilized transgenic (Tg) mice which constitutively express CTLA-4Ig, a high-affinity CD28/CTLA-4 antagonist that binds to both B7-1 and B7-2. The transgene is driven by a skin-specific promoter yielding high levels of CTLA-4Ig in the skin and serum. Chronic UV exposure of CTLA-4Ig Tg mice resulted in significantly reduced numbers of skin tumors, when compared to control mice. In addition, Tg mice were resistant to UV-induced suppression of delayed-type hypersensitivity responses to alloantigens. Most importantly, upon stimulation with mitogens and alloantigens, T cells isolated from CTLA-4Ig Tg mice produced significantly less IL-4 but more IFN-gamma compared to control T cells, suggesting an impaired Th2 response and a relative increase of Th1-type immunity. Together, these data show that overall B7 engagement directs immune responses toward the Th2 pathway. Moreover, they point out the crucial role of Th1 immune reactions in the protection against photocarcinogenesis.

A Critical Role for Dermal Mast Cells in Cis‐Urocanic Acid‐induced Systemic Suppression of Contact Hypersensitivity Responses in Mice

Many studies have implicated cis-urocanic acid (cis-UCA) in UVB-induced immunomodulation. The strongest evidence came from studies in mice whereby a cis-UCA antibody blocked UVB-induced suppression of delayed-type hypersensitivity responses. Furthermore, in several studies, the cis-UCA antibody at least partially reversed UVB suppression of contact hypersensitivity responses. Previous reports suggested that cis-UCA was immunomodulatory through its effects on keratinocytes, Langerhans cells, fibroblasts, T lymphocytes, natural killer cells and monocytes/macrophages. As dermal mast cells were recently demonstrated to be critical to UVB-induced systemic suppression of certain delayed-type and contact hypersensitivity responses, we investigated whether they were involved in the processes by which cis-UCA was immunomodulatory. Not only was there a correlation between dermal mast cell prevalence and the degree of susceptibility of different strains of mice to the immunomodulatory effects of cis-UCA, there was also a functional link. Mast cell-depleted Wf/Wf mice were rendered susceptible to immunomodulation by cis-UCA injected subcutaneously only after their dorsal skin had been reconstituted with bone marrow-derived mast cell precursors. These studies suggest that mast cells are critical to the processes by which cis-UCA suppresses systemic contact hypersensitivity responses to the hapten, trinitrochlorobenzene, in mice.

Correolide and Derivatives Are Novel Immunosuppressants Blocking the Lymphocyte Kv1.3 Potassium Channels

The voltage-gated potassium channel, Kv1.3, is specifically expressed on human lymphocytes, where it controls membrane potential and calcium influx. Blockade of Kv1.3 channels by margatoxin was previously shown to prevent T cell activation and attenuate immune responses in vivo. In the present study, a triterpene natural product, correolide, was found to block Kv1.3 channels in human and miniswine T cells by electrophysiological characterization. T cell activation events, such as anti-CD3-induced calcium elevation, IL-2 production, and proliferation were inhibited by correolide in a dose-dependent manner. More potent analogs were evaluated for pharmacokinetic profiles and subsequently tested in a delayed-type hypersensitivity (DTH) response to tuberculin in the miniswine. Two compounds were dosed orally, iv, or im, and both compounds suppressed DTH responses, demonstrating that small molecule blockers of Kv1.3 channels can act as immunosuppressive agents in vivo. These studies establish correolide and its derivatives as novel immunosuppressants.

Persistent suppression of delayed-type hypersensitivity in adult F344 rats after perinatal exposure to 2,3,7,8-tetrachlorodibenzo- p-dioxin

Recently we observed a suppressed delayed-type hypersensitivity (DTH) response to bovine serum albumin (BSA) in the 4–5-month-old offspring of F344 rat dams receiving as little as 1.0 μg 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD)/kg on gestational day (GD) 14. This study was designed to characterize better this suppression of the DTH response. First, the persistence of the DTH suppression was determined by measuring the DTH response to BSA in the offspring of dams dosed orally with 3.0 μg TCDD/kg on GD14 as well as in age-matched controls at 4, 8, 12 and 19 months of age. TCDD significantly suppressed the males’ DTH response through 19 months of age. While the females’ DTH response was reduced at 8, 12 and 19 months, significant suppression was observed only at 4 months of age. Secondly, the lowest maternal dose of TCDD that produced DTH suppression was determined by measuring the DTH response to BSA in the 4- and 14-month-old offspring of dams dosed orally with 0, 0.1, 0.3 or 1.0 μg TCDD/kg on GD14. In the males, suppression was observed at a maternal dose as low as 0.1 μg TCDD/kg at 14 months of age, while a maternal dose of 0.3 μg TCDD/kg was necessary to cause suppression in the 14-month-old females. Both males and females were more sensitive to the suppression at 14 months of age than at 4 months of age. Lastly, the DTH response to a second antigen was examined by measuring the DTH response to either BSA or keyhole limpet hemocyanin (KLH) in the 5- or 4-month-old male offspring, respectively, of dams dosed orally with either 0 or 3.0 μg TCDD/kg on GD14. The DTH response to both antigens was suppressed significantly. Phenotypic analysis was performed on thymus and lymph node suspensions. Significant effects in the thymus included an increased percentage of γδ TCR + cells and a decreased percentage of γδ TCR +/CD4 −CD8 − and MHCI − MHCII − cells. In the popliteal lymph node draining the BSA-injected footpad, there was a decreased percentage of γδ TCR + and MHCI −MHCII − cells and an increased percentage of MHCI + cells. In conclusion, the suppression of the DTH response associated with perinatal TCDD exposure is persistent through late adulthood, occurs at a low dose (i.e. 0.1 μg TCDD/kg) to the dam, and is more pronounced in males than females. While phenotypic analysis identified differences in subsets of thymocytes and lymph node cells between control and TCDD exposed offspring, no clear correlations were established between altered subpopulations and suppressed DTH responses.

Suppression of allograft rejection with anti-alphabeta T cell receptor antibody in rat corneal transplantation.

Anti-alphabeta T cell receptor monoclonal antibody (R73) has been reported to be a potent immunosuppressant. The suppressive effects of this antibody on allograft rejection after corneal transplantation are unknown. Orthotopic rat penetrating keratoplasty was performed using Lewis rats as recipients and Brown Norway and Fisher rats as donors. The treated groups received R73 intraperitoneally until day 12 after the transplantation. In grafted rats with or without R73 treatment, cytokine expression of the aqueous humor, corneal-infiltrating cells, draining lymph nodes, and splenocytes was determined. Delayed-type hypersensitivity (DTH) responses were compared. All allografts in the untreated controls of Fisher-to-Lewis or BN-to-Lewis rat combinations were rejected within 14 days. In contrast, indefinite survival rates of the postoperative R73-treated group increased to 86% in the Fisher-to-Lewis and 23% in the Brown Norway-to-Lewis combinations, respectively. Interferon-y, interleukin (IL)-2 (T helper [Th]1), and IL-10 (Th2), but not IL-4 (Th2), expression of the eye and DTH responses in the control group were suppressed in the R73-treated group. Both IL-2 and IL-10 expression after mixed lymphocyte culture in the R73-treated group were significantly lower than those of the naive and untreated control group. alphabeta T cell receptor-targeted therapy prevents allograft rejection in rat corneal transplantation as evidenced by suppression of DTH responses. The cytokine profile after R73 treatment was characterized by low interferon-gamma, IL-2, and IL-10, and high IL-4 expression.