Levels Of Delayed-type Hypersensitivity Research Articles

Immunomodulatory Effects of the Somatic Antigens of Fasciola Hepatica and Teladorgasia Circumcincta in Mice Immunized with Sheep Red Blood Cells

Background & Objective: Infestation of humans with helminth parasites can reduce the severity of some auto-inflammatory symptoms in humans. This study was done to evaluate the impact of the somatic antigens extracted from Fasciola hepatica (as an indicator Trematode) ,and Teladorgasia circumcincta (as an indicator nematode) on the immune responses of NMRI mice challenged with sheep red blood cells (SRBCs).
 Materials & Methods: The mice in treatment groups were intraperitoneally immunized with 1×109 SRBCs twice with 14 days intervals. Concurrent with the immunization, the mice received the extract of each of the parasites (50,100. And 150 µg of protein) or placebo, throughout the study on a daily basis. The specific cellular immune responses and the anti-SRBC antibody titers were detected by footpad thickness and, microhemagglutination test, respectively. Splenocytes were also monitored for cytokine production, proliferation rate, and respiratory burst.
 Results: The extracts of F. hepatica and T. circumcincta had an opposite effect on the change of the Anti-sRBC antibody level. The extract of F. hepatica caused a significant decrease in the antibody level whereas, extract T. circumcincta did not show any significant changes in the anti-SRBC antibody. Both extracts caused a significant decrease in the level of delayed-type hypersensitivity. However, the extract of F. hepatica caused a more profound reduction in the severity of delayed-type hypersensitivity. The level of IFN-γ in the splenocytes of immunized mice receiving the F. hepatica showed a more pronounced decrease than the immunized mice receiving the extract of T. circumcincta extract. IL-10 levels were only increased in the immunized mice that received the extract of F. hepatica.
 Conclusion: The extract of F. hepatica may have an immunosuppressive property, while the extract of T. circumcincta may have immunomodulatory properties.

The Immunopathobiology of Syphilis: The Manifestations and Course of Syphilis Are Determined by the Level of Delayed-Type Hypersensitivity

Syphilis has plagued mankind for centuries and is currently resurgent in the Western hemisphere. Although there has been a significant reduction of tertiary disease and recognition of facilitative interactions with human immunodeficiency virus infection, the natural history of syphilis has remained largely unchanged; thus, new strategies are required to more effectively combat this pathogen. The immunopathologic features of experimental syphilis in the rabbit; the course, stages, and pathology of human syphilis; and a comparison of human syphilis with leprosy suggest that the clinical course of syphilis and its tissue manifestations are determined by the balance between delayed-type hypersensitivity (DTH) and humoral immunity to the causative agent, Treponema pallidum. A strong DTH response is associated with clearance of the infecting organisms in a well-developed chancre, whereas a cytotoxic T-cell response or strong humoral antibody response is associated with prolonged infection and progression to tertiary disease. Many of the protean symptoms/appearances of secondary and tertiary human syphilis are manifestations of immune reactions that fail to clear the organism, due to a lack of recruitment and, more importantly, activation of macrophages by sensitized CD4 T cells. The Bacillus Calmette-Guerin vaccination can enhance DTH and has been shown to produce a low, but measurable, beneficial effect in the prevention of leprosy, a disease that shows a disease spectrum with characteristics in common with syphilis. In the prevention of syphilis, a potential vaccine protective against syphilis should be designed to augment the DTH response.

Differentiation and functions of T cell subsets.

The Tc1 and Tc2 subsets of CD8+ T effector cells secrete different patterns of cytokines, but have similar functions, including perforin- and Fas-dependent cytotoxicity, and induction of delayed type hypersensitivity (DTH) reactions involving oedema and granulocytic infiltration. The characteristic cytokines of Tc1 (gamma-interferon) and Tc2 (interleukins 4 and 5) are expressed in vivo during the DTH reaction. Tc1 cells that are deficient in cytokine synthesis also induce similar levels of DTH, supporting the lack of correlation between CD8+ T cell cytokine patterns and DTH. CD8+ T cells often produce lower cytokine levels than CD4 cells because the CD8 cells kill their antigen-presenting cells before full stimulation can occur. This effect can be counteracted by increasing the frequency of stimulation, or using perforin-deficient T cells. A multiparameter analysis of cytokine effects on CD8+ T cell differentiation has been initiated, on the basis of the principle that normal immune responses involve complex cytokine mixtures. All combinations of seven cytokines were tested. In some combinations, the combined effect could not have been predicted from individual cytokine functions. Conditions were identified in which each of interleukins 4, 10 and 12 could have opposite effects on CD8+ T cell differentiation.

Components with potential immunosuppressive activity in lipopolysaccharide of laboratory Pseudomonas aeruginosa strains

The ability of culture filtrate (CF) of two laboratory Pseudomonas aeruginosa strains to inhibit delayed type hypersensitivity (DTH) to non-bacterial antigen in CBA mice has been studied. It was shown that intraperitoneal injection of native CF of the strains did not affect the level of DTH. However, redox treated CF expressed the immunosuppressive activity. Gel filtration of redox activated CF through Sephadex G-200 showed that CF contains three immunosuppressive components differing by their molecular weight and specificity. All components contained lipid group and O-polysaccharide chains that indicated their lipopolysaccharide (LPS) nature. These experiments show that laboratory P. aeruginosa strains have three LPS components but not all of them display the immunosuppressive activity.

The immunosuppressive activity of chemically modified lipopolysaccharide of Shigella sonnei

In the present investigations we aimed to study the effect of Shigella sonnei lipopolysaccharide (LPS) on delayed type hypersensitivity (DTH) to non-bacterial antigen in CBA mice. These experiments showed that intraperitoneal injection of phenol–water extracted LPS and avirulent S. sonnei did not affect the level of DTH. However, an injection of avirulent bacteria and LPS treated with 2-mercaptoethanol reduced significantly the levels of DTH. Gel filtration of redox-reactivated LPS through Sephadex G-200 shows that LPS contains three immunosuppressive components: ∼800 kDa and higher, 150–200 and 50–70 kDa. These components differed by their specificity and heat-sensitivity.

Induction of partial protection against Leishmania donovani by promastigote antigens in negatively charged liposomes.

Negatively charged liposomes, proposed as potential vaccine adjuvants, have been extensively studied in association with various antigens. In the present study, we investigated the adjuvanicity of negatively charged liposomes to enhance the protective immunity of membrane antigens of Leishmania donovani promastigotes (LAg). In comparison to the control mice immunized with phosphate-buffered saline and empty liposomes, immunization with free LAg led to significant levels of protection against infection with virulent promastigotes. Encapsulation of LAg in liposomes also induced effective protection. However, the level of protection by LAg-liposome was not significantly different from that induced by free LAg. Investigation of the immune responses showed, in contrast to free LAg, that immunization with LAg-liposome elicited strong antibody responses. IgG isotype analysis revealed the presence of all 4 isotypes. However, the titer of IgG1 was significantly higher than IgG2a, IgG2b, and IgG3. Following infection, stimulation of IgG and IgG isotypes did not differ in the different immunization groups. Delayed-type hypersensitivity (DTH) analysis after immunization showed significant induction by LAg and LAg-liposomes, in comparison to controls. With infection, again, the level of DTH in all the groups became almost comparable. Stimulation of insufficient cellular response, as reflected by DTH and potentiation of IgG1 over IgG2a, IgG2b, and IgG3 suggest a dominance of Th2 response with this liposome-antigen formulation, resulting in weak protection against visceral leishmaniasis.

Differential Regulation of Epidermal Cell Tumor-Antigen Presentation by IL-1α and IL-1β

IL-1 exists in two forms, termed IL-1alpha and IL-1beta, which exert similar effects in a number of biologic models. Recently, there have been reports of some differences in the activities of these two species in some systems. To address this issue with regard to Langerhans cells, Langerhans cell-enriched preparations of epidermal cells were treated with either IL-1alpha or IL-1beta before pulsing with S1509a tumor-associated antigens and subsequent use for immunization of naive mice to S1509a. While epidermal cells treated with 100 U IL-1beta per ml were able to induce protective tumor immunity (as indicated by the rejection of a subsequent tumor challenge with viable S1509a tumor cells), epidermal cells treated with 100 U IL-1alpha per ml failed to confer protective immunity. At 1000 U per ml, IL-1beta also inhibited the ability of epidermal cells to induce tumor immunity. To investigate the effects of the two IL-1 forms on elicitation of tumor immunity, naive mice were immunized against the S1509a tumor by s.c. injection of dead S1509a cells. Epidermal cells enriched for Langerhans cells were treated with either 100 U IL-1alpha or IL-1beta per ml before tumor-associated antigens-pulsing. Epidermal cells were then washed and injected into a hind footpad of tumor immune mice and 24 h footpad swelling was assessed as a measure of delayed-type hypersensitivity. Exposure to IL-1alpha led to suppressed elicitation of delayed-type hypersensitivity, whereas IL-1beta treated epidermal cells elicited a normal (100 U per ml) or enhanced (1000 U per ml) level of delayed-type hypersensitivity. Previous experiments indicated that the suppressive effects of IL-1alpha on induction of immunity may be mediated by TNF alpha. Therefore, the ability of IL-1alpha or IL-1beta to induce epidermal cell production of TNF alpha was assessed. IL-1alpha induced epidermal cells to secrete significantly higher amounts of TNF alpha protein compared with stimulation with IL-1beta. IL-1alpha and IL-1beta appear to differentially regulate epidermal cell antigen presenting capability.

Acceleration of influenza virus clearance by Th1 cells in the nasal site of mice immunized intranasally with adjuvant-combined recombinant nucleoprotein.

The protective roles of influenza viral nucleoprotein (NP), together with the cellular mechanism of the protection in the nasal site, were examined in BALB/c mice immunized intranasally with an adjuvant (cholera toxin B subunit containing 0.2% of the whole toxin)-combined A or B virus recombinant NP. The NP-immune mice, when challenged intranasally with a sublethal dose of the virus 3 wk after immunization, had accelerated virus clearance from the nasal site in both an influenza type-specific and a nonspecific manner, as shown by the protection from high morbidity from the second day after challenge. Both type-specific and nonspecific acceleration of recovery was confirmed by the increased survival rate after challenge with a lethal dose of virus in mice immunized and boosted with adjuvant-combined NP. The acceleration of nasal virus clearance was accompanied with acceleration of type-specific systemic delayed-type hypersensitivity (DTH) and with IFN-gamma production by nasal lymphocytes. The nasal lymphocytes from the immunized and challenged mice generated a significantly high level of DTH when transferred locally, but no class I MHC-restricted CTL response. Moreover, nasal CD4+ T cells, induced by NP immunization and increased in number by the subsequent challenge, were involved in the accelerated IFN-gamma production. These results suggest that nasal Th1 cells, capable of producing IFN-gamma and mediating DTH, are involved in the type-specific acceleration of recovery from influenza after challenge in mice immunized intranasally with adjuvant-combined NP, although the nonspecific mechanism of accelerated recovery remains to be solved.

A mannoprotein constituent of Candida albicans that elicits different levels of delayed-type hypersensitivity, cytokine production, and anticandidal protection in mice.

To identify major immunogenic constituents of Candida albicans, the effect of a mannoprotein fraction (MP-F2) on the elicitation of a delayed-type hypersensitivity (DTH) reaction, cytokine production, and protection from a virulent Candida challenge in a mouse candidiasis model was studied. In mice immunized with whole cells of a low-virulence strain of C. albicans and thus protected against a challenge with a highly virulent strain of this fungus, MP-F2 was able to elicit a strong DTH response that was accompanied by splenocyte proliferation in vitro in the presence of Candida antigen. The supernatants of MP-F2-stimulated splenocyte cultures contained gamma interferon (IFN-gamma, a typical CD4+ T helper-1 (Th1) cytokine, but no interleukin-4, (IL-4), a typical CD4+ Th2 cytokine. IFN-gamma was produced by CD4+ cells, and its level could be greatly increased by the addition of anti-IL-4 or, mostly, anti-IL-10 antibodies to the CD4+ cell cultures. Upon a suitable schedule of immunization, MP-F2 was also able to induce a vigorous DTH response in Candida-uninfected mice, a response that could be efficiently transferred into naive recipients by CD4+ cells from the spleens of MP-F2-immunized mice. The immunization described above also conferred to mice a low degree of protection against a virulent Candida challenge, both in terms of median survival time and in the number of Candida cells in the kidney. However, while DTH induction by MP-F2 was as strong as that induced by whole cells, MP-F2-induced protection was significantly weaker than that conferred by Candida whole-cell immunization. Mice immunized with either MP-F2 or Candida whole cells had an inverted ratio between the number of CD4+ splenocytes producing IFN-gamma and that of cells producing IL-4, compared with nonimmunized animals. However, the number of IL-4-producing CD4+ cells was significantly higher in MP-F2-vaccinated, weakly protected mice than in Candida whole-cell-vaccinated, highly protected animals. Overall, our data suggest that the MP-F2 fraction contains one or more major immunogens of C. albicans which are capable of interfering with the balance of CD4+ Th1 and Th2 responses that is so critical in the outcome of host-Candida relationship and are thus potentially relevant in the mechanisms of Candida-specific DTH regulation and protection.

Specificity of Salmonella Porin as an Eliciting Antigen for Cell‐Mediated Immunity (CMI) Reaction in Murine Salmonellosis

The specificities of Salmonella porin on elicitations of delayed-type hypersensitivity (DTH) reaction and interleukin-2 (IL-2) production in BALB/c mice immunized with Salmonella typhimurium were examined. Only porin from S. typhimurium was capable of eliciting significant levels of DTH and IL-2 production in S. typhimurium-immunized mice, whereas no significant DTH and IL-2 production were induced by porin from Salmonella enteritidis or Escherichia coli. Our observations suggested that Salmonella porin was a serovar-specific antigen for the elicitation of cell-mediated immunity (CMI) in salmonellosis.

Delayed-type hypersensitivity to weak histocompatibility antigens and its genetic restriction

The analysis of skin allograft survival time and the level of delayed-type hypersensitivity (DTH) reaction to major and minor histocompatibility antigens revealed the correlation between these parameters of the transplantation immunity. The data obtained have shown that histocompatibility in several non-H-2 antigens induces DTH reaction comparable with the reaction caused by H-2 antigens. The effector phase of DTH to non-H-2 antigens is H-2 restricted. No restriction of the afferent phase is revealed. The application of these results to the analysis of the mechanisms of the recognition of minor histocompatibility antigens in DTH is discussed.

Immune responses to yeast and mycelial forms of Candida albicans in intraperitoneally infected mice.

Candida albicans E-139 produced pure mycelial and yeast cultures in a low sulphate medium at different temperatures. The influence of the morphological phase, dose and viability of the fungi on the kinetic of delayed-type hypersensitivity (DTH) and anti-mycelial and anti-yeast antibodies have been studied in mice injected intraperitoneally. The mycelial form elicited higher DTH levels than the yeast phase. This effect seems to be related to its antigenic properties. The effect of dose on the immune response depends on the viability of the fungus. The mycelial cytoplasmic antigens were more effective than the yeast ones in detecting antibodies induced during the experiments, particularly during the later stages of the observation periods, suggesting that such antigens may be useful in the serodiagnosis of Candida infections.

Cellular and humoral immune responses to Candida albicans in subcutaneously infected mice.

Pure mycelial and yeast cultures of Candida albicans were produced in a low sulphate medium. Groups of mice were injected subcutaneously with increasing doses of viable or heat-killed mycelial or yeast cells and the kinetics of delayed-type hypersensitivity (DTH), anti-mycelial and anti-yeast antibodies were studied. Both the dose and the morphological phase of C. albicans showed an influence on the development of the DTH, but the viability is the factor which showed the highest influence on this reaction, since on the one hand mice infected with viable yeast or mycelial cells developed higher DTH levels than mice injected with heat killed cells, and on the other hand this factor seems to play an important role in the kinetics of DTH response. The enzyme-linked immunosorbent assay has been adapted to detect antibodies to yeast and mycelial phase cytoplasmic antigens of C. albicans. In contrast with the DTH reactions, neither dose, morphological phase nor viability played an important role on the antibody titer developed. However, the use of mycelial cytoplasmic antigens seems to be better than the yeasts to detect anti-Candida antibodies over the last days studied.

Depression of macrophage functions and T-cell-mediated immunity to listeria infection in tumor-bearing mice and its prevention by PSK.

The effect of PSK on the depressed bactericidal activity of macrophages and delayed-type hypersensitivity (DTH) to Listeria monocytogenes in BALB/c mice bearing transplantable Meth A fibrosarcoma was studied. In tumor-bearing mice pretreated with PSK, L. monocytogenes was cleared rapidly from the circulating blood and bacterial growth in the liver was inhibited effectively in the early phase of infection. This resistance to the infection could be transferred with adherent peritoneal exudate cells (PEC) but not with nonadherent or adherent spleen cells of PSK-treated mice. In the early phase of infection, tumor-bearing mice developed a lower level of DTH to L. monocytogenes than nongrafted control mice. However, the control levels of DTH could be obtained by pretreatment with PSK in tumor-bearing mice. These results suggest that the restoration of DTH to L. monocytogenes by pretreatment with PSK may be attributable to the restoration of the depressed immunological responsiveness to the normal levels in tumor-bearing mice.

Cutaneous unresponsiveness to Mycobacterium bovis BCG in intravenously infected mice.

Mice injected with 1 mg (about 1 X 10(7) CFU) of Mycobacterium bovis BCG in the footpad showed high levels of delayed-type hypersensitivity (DTH) to BCG antigens and a continuous increase in circulating specific anti-immunoglobulin G (IgG) antibodies throughout a 6-week observation period. In contrast, mice injected intravenously with a 1-mg dose failed to mount a DTH and showed a depression in antibody production at week 5 postinfection. A dose-response study revealed that an optimum dose of BCG, when injected intravenously, can induce a small but significant DTH response. The delayed cutaneous unresponsiveness in intravenously infected mice lasted at least 6 weeks and was not antigenically specific, in that it depressed the DTH response to Corynebacterium parvum. No simple relationship existed between levels of DTH and the amount of circulating anti-IgG antibodies. Splenectomy and treatment with a high dose of cyclophosphamide before infection, although greatly reducing the humoral response, did not reverse the BCG-induced unresponsiveness nor enhance levels of DTH in mice infected in the footpad. It is concluded that the intravenous infection of mice with BCG exerts a nonspecific inhibitory effect on delayed-type immune reactions by the induction of some type of suppressor mechanisms that are resistant to cyclophosphamide.

Effect of specific alloantiserum against T suppressors on resistance of mice to tuberculosis

The writers showed previously that mice of the inbred line I/St are highly susceptible to infection with tuberculosis, whereas those of inbred line A2G are resistant [2]. Parameters of immunity to tuberculosis, such as the level of the delayed-type hypersensitivity (DTH) reaction and of antibody synthesis to tuberculin, were opposite in mice of these lines at the same times after infection: a high level of DTH and a low level of antibodies in A2G mice and the opposite picture in I/St mice [i]. These data confirm the earlier concept [3] of immunological causes of sensitivity and resistance to tuberculosis and the protective role of cellular immunity in this disease.

Augmentation of delayed-type hypersensitivity to serum proteins by vesicular stomatitis virus infection in mice: virus-suppressor cell interactions.

The effects of infection with vesicular stomatitis virus (VSV) on delayed-type hypersensitivity (DTH) to heterologous serum proteins were investigated in mice. DTH was induced by a subcutaneous injection of antigen in complete Freund's adjuvant. Infection with VSV at the time of immunization did not affect the level of DTH elicited 3 wk later. Marked augmentation of DTH was observed only when previously immunized mice were infected with VSV simultaneously with restimulation by soluble antigen; either soluble antigen or the virus infection alone was ineffective. The augmentation was specific to the antigen used for the restimulation; in the mouse previously immunized with both bovine serum albumin (BSA) and human alpha-globulin (HGG), DTH to BSA but not to HGG was augmented by injecting soluble BSA and VSV, and vice versa. These results strongly suggest that cells involved in the suppression of DTH manifestation became susceptible to the virus after specific antigenic restimulation and were then eliminated.

Augmentation of delayed-type hypersensitivity by vesicular stomatitis virus infection in mice.

Effects of the infection with vesicular stomatitis virus (VSV) on delayed-type hypersensitivity (DTH) to heterologous erythrocytes were investigated in mice. Infection at the time of immunization resulted in production of high levels of DTH that were specific to the antigen used for immunization. The high level of DTH produced in VSV-infected mice could not be attributed to the nonspecific enhancement of the footpad swelling with the infection. Augmentation of DTH was observed in all strains of the mouse (CBA, BALB/c, C3H/He, and C57BL/6) used. The augmenting effect of VSV infection was not as apparent in adult thymectomized mice in which the level of VSV-replicating T cells was reduced. These results strongly suggest that DTH-mediating T cells are resistant to infection by VSV, and also that there are VSV-sensitive cells that may be engaged in the suppression of DTH. It seems improbable, however, that the cells sensitive to VSV infection represent the suppressor cells themselves, since the enhancing effect was not observed in mice in which the suppressor cells were induced by the administration of high doses of the antigen.

Immunologic features of mice with streptozotocin-induced diabetes: depression of their immune responses to sheep red blood cells.

The immune responsiveness in streptozotocin (SZ)-induced diabetic mice was studied using the immune responses to sheep red blood cells (SRBC) as an indicator system. In SZ-diabetic mice, the weights of such lymphoid organs as the thymus and spleen were significantly decreased with time after SZ administration, whereas the weight of liver was markedly increased. In SZ-diabetic mice, the level of delayed type hypersensitivity (DTH) to SRBC was not lower than that in normal controls in most cases, although the level of DTH was significantly depressed, on occasion, in SZ-diabetic mice. In contrast, antibody-forming activity, measured as the number of plaque-forming cells (PFC), was markedly decreased in SZ-diabetic mice. It seems that antibody production is more profoundly depressed than is DTH in SZ-diabetic mice. The transfer of normal thymus and bone marrow cells into SZ-diabetic mice caused only a partial restoration of PFC activity. When normal spleen cells were transferred into diabetic irradiated mice, proliferation of spleen cells and production of splenic PFC was greatly reduced as compared with normal irradiated mice. Treatment with insulin completely reversed such depression in the transfer system. These findings suggest that the chronic insulin-deficient diabetic state caused a depression and delay in the proliferation and differentiation of lymphoid cells.

In vivo detection of specific cell-mediated immunity in street rabies virus infection in mice.

In street rabies-infected mice, in vivo expression of delayed type hypersensitivity (DTH) measured by the footpad test was revealed by challenge with inactivated fixed rabies virus (RV). The use of BCG as an adjuvant cell-mediated immunity (CMI) was necessary for the production of significant DTH levels. Typical DTH kinetics were obtained, with a maximum at 24 h after the challenge. DTH was also found to be at highest levels 4 days after infection with street rabies virus. DTH could also be revealed with street rabies virus in RV immunized mice. Adoptive transfer of lymphoid cells from a street rabies infected donor to normal recipient mice was performed and DTH was tested with RV. Susceptibility of DTH to immunosuppression by cyclophosphamide treatment was also assayed in street rabies virus-infected mice and in adoptively-sensitized recipient mice. These results and the relationship between DTH and CMI in rabies infection and immunization are discussed.