Heterologous antigenic stimulation in induction of delayed hypersensitivity.

An influence of a delayed hypersensitive reaction to a primary antigen on the induction of delayed hypersensitivity to a second unrelated antigen was observed in guinea pigs immunized with azobenzenearsonate-N-acetyl-L-tyrosine (ABAT), and injected intradermally 3 weeks later with a mixture of ABAT and secondary antigen. Animals so treated developed delayed hypersensitivity to sheep red blood cells (SRBC) or Type II pneumococcal polysaccharide as secondary antigens, as measured by skin test reactivity and inhibition of macrophage migration, whereas ABAT unsensitized control groups did not. However, attempts to induce delayed reactivity to proteins as secondary antigens were unsuccessful. The injection of secondary antigen into a mineral oil-induced inflammatory lesion did not induce delayed hypersensitivity, suggesting that specific reactivity to ABAT is a prerequisite for heterologous induction. Possible mechanisms for the observed phenomenon, including a role for macrophages, are discussed.     

Studies on delayed hypersensitivity to protein antigen. Induction of delayed hypersensitivity by chemically modified antigen.

it was shown in our previous paper that mice primed with chemically modified bacterial alpha-amylase (BaA), which was neither cross-reactive with anti-BaA antibody nor able to induce a humoral anti-BaA response, developed enhanced responses to a subsequent challenge with native BaA and that the magnitude of the immunological memory was closely related to the priming dose of modified BaA. This paper describes the experimental conditions for induction of delayed hypersensitivity (DH) by modified BaA in relation to the development of immunological memory for antibody response to native BaA. Mice primed with either an intraperitoneal (i.p.) or subcutaneous (s.c.) injection of modified BaA in complete Freunds adjuvant (CFA) developed enhanced anti-BaA as the immunogen and modified BaA as the eliciting antigen, the relationship of anti-BaA responses to a subsequent challenge with BaA. In contrast, when mice were immunized with an s.c. injection of the modified BaA only, a significant level of DH to native BaA could be induced, as measured by the footpad reaction (FPR). The highest degree of DH was observed in mice given 50 micrograms of modified BaA. DH was detectable within 5 days and persisted for 25 days after immunization. In the reciprocal combination of native BaA as the immunogen and modified BaA as the eliciting antigen, the relationship of anti-BaA responses to DH was examined. The primary anti-BaA responses induced by an i.p. injection of large doses of BaA was markedly higher than those induced by an s.c. injection, while DH was exhibited only in mice given s.c. injection of BaA in CFA. With respect to DH to native BaA induced by the modified BaA, it was shown that C3H/He mice were high and C57BL/6 mice were low responders.     

Studies on delayed hypersensitivity to protein antigen. Antigen-specific suppression of delayed hypersensitivity by chemically modified antigen.

An extensively modified protein antigen (methylated bacterial α-amylase, M-BαA) which was neither reactive with anti-BαA antibody nor able to induce a humoral anti-BαA response, retained the ability to prime native BαA-specific T cells which were responsible for the enhanced anti-BαA response to subsequent immunization with BαA and delayed hypersensitivity (DH). The splenic T cell-rich fraction from mice primed with M-BαA collaborated with a native BαA-primed B cell-rich fraction to give a good adoptive IgG anti-BαA response in syngeneic irradiated mice, whereas M-BαA-primed B cell fractions failed to cooperate with native BαA-primed T cell fractions. Splenic T cells from mice given a subcutaneous (s.c.) injection of M-BαA in complete Freund's adjuvant (CFA) exhibited DH in syngeneic cyclophosphamide-treated mice. In the present study, native and methylated BαA were tested for their ability to generate suppressor T cells capable of inhibiting the development of DH. An intraperitoneal (i.p.) injection of either native or methylated BαA in incomplete Freund's adjuvant (IFA) interferred with the development of DH to M-BαA by an s.c. injection of the same antigen in CFA. Transfer of spleen cells from mice given an i.p. injection of either of these antigens 5 days previously, suppressed antigen-specifically induction and expression of DH in the syngeneic recipient mice. The suppressive activity was sensitive to treatment with anti-θ antiserum plus complement. These results indicate that the early phase of inhibition of DH after an i.p. injection is in part mediated by suppressor T cells and that M-BαA cross-reacts with native BαA at the suppressor T cell level as well as the level of effector T cells in DH.     

Mechanisms involved in the antibody-mediated suppression of tuberculin-type delayed hypersensitivity: II. The sensitivity of tolerance induction to cyclophosphamide pretreatment and splenectomy,and the demonstration of active suppression

The induction of tuberculin-type delayed hypersensitivity, as measured by skin test, can be specifically inhibited by administration of antibody during sensitization. The cellular mechanisms involved in this tolerance were investigated in CAP₁ mice, using chicken conalbumin as antigen. Tolerance was prevented when mice were treated with Cyclophosphamide 2 days before sensitization and suppression. However, it was not affected by splenectomy 7 or 21 days before sensitization. This tolerance could be transferred to normal CAF₁ mice with spleen cells, but not with thymocytes, when taken from donor mice 21 to 28 days after sensitization and tolerance induction. Production of these cells in the donors required both antibody and antigen. The cells responsible for the transfer were B cells, as shown by their sensitivity to rabbit anti-mouse-immunoglobulin serum and complement. In addition to B cells, serum from tolerant mice also could transfer suppression at 21 to 28 days. We conclude that sensitizing mice, and treating them with specific immunosuppressive antiserum, induces the recipients to make suppressor B cells and suppressive humoral factors, which are involved in arresting the induction of tuberculin-type delayed hypersensitivity.     

Effect of thymectomy on induction of immunologic tolerance in the effectors of delayed hypersensitivity]

The influence of thymectomy (Tx) on induction of tolerance of delayed type hypersensitivity effectors (DHE) was examined. Tx did not interfere with induction of tolerance to sheep red blood cells (SRBC) achieved with combined injections of the massive dose of the antigen and cyclophosphamide (Cy). Tx resulted in prolongation of unresponsiveness. The injection of mice with the massive dose of SRBC alone also resulted in tolerance formation. However, this type of tolghtly depressed formation of DHE in intact but not in Cy treated mice. The results obtained are in agreement with the idea of the existence of diverse mechanisms of tolerance induction (clonic-deficient and suppressor). These data also suggest the existence of two subpopulations differing in susceptibility to Cy and Tx in DHE effectors and their precursors.     

The macrophagal-lymphocytic interaction of heterochronic cells in the induction of delayed hypersensitivity]

The influence of senescence on the functional activity of lymphocytes and macrophages in the induction of sensitivity to tuberculosis has been studied in experiments on 226 CBA mice. The study has revealed that after the injection of BCG old animals exhibit decreased capacity for the formation of delayed hypersensitivity, and their lymphocytes, transplanted to recipients, induce a lower level of hypersensitivity. Joint incubation of lymphocytes and macrophages from animals of different ages has shown that immunological defect appearing with age is localized in lymphocytes, while the antigen-presenting function of macrophages remains basically unchanged.     

Development of delayed hypersensitivity to sheep erythrocytes after separate and combined administration of the antigen and cyclophosphamide]

The ability to the development of delayed hypersensitivity (DHS) to the appropriate antigen was studied in mice treated with large doses of SRBC and cyclophosphamide at varying time prior to the experiment. Suppression of DHS development induced by administering either the cytostatic alone or a large dose of the antigen was examined at the same periods of time. The combined treatment was shown to induce tolerance according to diverse tests for DHS (skin and macrophage migration inhibition tests). At the basis of this tolerance lies genuine deficiency of the appropriate clone of T cells. Administration of cyclophosphamide alone leads to a slight suppression of DHS, while a large dose of the antigen induces a different form of areactivity due to the suppressor cells whose nature is not yet clear.     

Effect of polysaccharides from Ganoderma applanatum on immune responses. I. Enhancing effect on the induction of delayed hypersensitivity in mice.

Prior intraperitoneal (i.p.) or oral administration of the polysaccharide preparation from a kind of mushroom, Ganoderma applanatum (Pers.) Pat. of Basidiomycetes, exerted an enhancing effect on the induction of delayed hypersensitivity (DH) to protein antigen as measured by the footpad reaction (FPR), and expanded the size of T cell memory for the IgG antibody response. One of the active principles was partially purified and found to be associated with a polysaccharide-rich fraction. The induction of DH was enhanced by treatment with an appropriate dose of the mushroom extract, whereas increasing the dose resulted in almost complete loss of the enhancing activity. The mechanism for the enhancing effect of the mushroom extract on the induction of DH was explored by the adoptive cell transfer technique. Although an i.p. injection of methylated bacterial α-amylase (M-BαA) in incomplete Freund's adjuvant (IFA) has been found to generate in the spleen the antigen-specific suppressor T cells capable of inhibiting the induction of DH 5 days after immunization, the same treatment of mice given prior injections of the mushroom extract did not raise the suppressor cell activity, but transfer of these spleen cells (6 × 10⁷) into syngeneic recipient mice which had been primed with a subcutaneous (s.c.) injection of M-BαA in complete Freund's adjuvant (CFA) resulted in substantial amplification of the expression of DH. The absence of effector T cells for DH in the transferred spleen cells was confirmed by the failure to transfer DH into cyclophosphamide (CY)-treated mice with the amplifying cells. The amplifying activity was antigen-nonspecific and mediated by cells sensitive to treatment with anti-θ antiserum plus complement. Therefore, the nonspecific enhancing effect of the mushroom extract could not be explained by the possibility that pretreatment with the extract eliminated the antigen-specific suppressor T cells. Other adoptive cell transfer experiments revealed that nylon wool-passed cells from mice unprimed but treated with the mushroom extract were able to exert an enhancing activity on the expression of effector T cells in DH. The results indicate that the treatment with an appropriate dose of the extract enhances the induction of DH by activation of the nonspecific amplifier T cells.