Secondary IgE Responses Research Articles

Potentiation of the reaginic (IgE) antibody response to ovalbumin in the guinea pig with a soluble metabolic product from Ascaris suum.

A soluble antigen, the ACF antigen, obtained from cultures of larvae of Ascaris suum was able to potentiate the reaginic response to an unrelated antigen was administered at the same time or 4 days after ovalbumin, there was a weak primary response and a strong statistically significant secondary IgE response, compared with animals immunized only with ovalbumin. This potentiated response was not observed when the ACF antigen was presented 10 days before or 8 or 12 days after the ovalbumin.

Macrophage regulation of the IgE response. I. Studies on the immunogenicity and antigenicity of macrophage-associated antigen.

The ability of macrophage-associated antigen to both prime for, and subsequently trigger, IgE responses in inbred rats and mice was investigated. Peritoneal exudate cells briefly pulsed <i>in vitro</i> with ovalbumin (PEC-OVA) served as the immunogen, and both primed and unprimed animals as recipients. The experiments revealed that the intraperitoneal administration of relatively small numbers of PEC-OVA, while generally ineffective in inducing primary IgE responses in immunologically virgin mice, successfully primed the latter for secondary IgE responses following later challenge. Furthermore, PEC-OVA was highly effective in triggering vigorous secondary IgE responses in primed mice, producing PCA titres up to 10,000 in both moderate and high IgE-responder strains. The data further indicate that the IgE response induced in mice by PEC-OVA is transient, it exhibits a markedly lower threshold (in both the primary and secondary response) than corresponding haemagglutinating antibody responses, and is MHC-restricted. In the rat strain employed (low IgE-responder WAG), PEC-OVA administration evoked vigorous haemagglutinating antibody responses in pre-immunized rats, and primed immunologically virgin animals for similar secondary responses. However, PEC-OVA was only weakly immunogenic/antigenic for IgE responses in WAG rats.

Anti-timothy IgE formation: suppression with antigen D-dGl conjugates.

Antigen D fragments (AgD1 and AgD2) and chemically synthesized quercetin-glutathione were conjugated to the synthetic polypeptide copolymer of D-glutamic acid and D-lysine (dGL). These conjugates were tested at varying epitope densities to determine their ability to suppress a secondary anti-antigen B IgE response. The data showed that all of the conjugates used with epitope densities of 5-20 groups per dGL produced significant dose-dependent suppression of a secondary IgE response. The duration of the observed suppression was short (about 30 days), but could be extended by additional treatment with the conjugate prior to the loss of unresponsiveness.

The effect of hapten-specific suppression of IgE on antigen-induced histamine release from mouse peritoneal mast cells.

Subcutaneous injections of a mixture of dinitrophenylated ovalbumin (DNP3-OA) and dextran sulfate into Swiss-Webster mice elicited short-lived primary and long-lasting secondary IgE antibody responses to both DNP and OA. Histamine was released on in vitro challenge with antigen (OA or DNP22-BSA) of washed peritoneal mast cells (PMC) obtained from mice during a primary or a secondary IgE response. Administration of an intravenous injection of a tolerogenic conjugate of DNP8-mouse gamma-globulin, either prior to immunizationor during an ongoing IgE response, resulted in almost complete disappearance of circulating anti-DNP IgE antibody and in a very marked decrease in histamine release from PMC on challenge with DNP22-BSA. However, the IgE response to OA of these mice and the histamine release from their PMC on challenge with OA were not affected. Moreover, the PMC of mice, which had been tolerized to DNP, could be passively sensitized with serum containing DNP-specific IgE antibody for the release of histamine on DNP22-BSA challenge. The most significant finding of this study is the observation that the time course for the loss of reactivity of PMC to DNP22-BSA, after administration of the tolerogen during an ongoing secondary response, paralleled the decrease in circulating anti-GNP IgE antibody.

Regulation of IgE Antibody Production by Serum Molecules

Abstract IgE antibody production in mice of high and low IgE responder phenotypes, respectively, can be appreciably enhanced in magnitude after low dose whole body X-irradiation. Such enhanced responses, as well as adoptive secondary IgE responses, can be markedly suppressed by passive transfer of CFA-immune serum in low responder strains, but not in high responder strains. The studies presented here demonstrate that the suppressive activity of CFA-immune serum on IgE antibody production is strain specific. This is true even in reciprocal combinations of low IgE responder SJL and C57BL/6 mice, in which it was shown that serum capable of suppressing mice of the isologous strain was ineffective in diminishing IgE antibody production in the other low responder strain. Absence of suppressive activity in CFA-immune sera obtained from H-2 congenic donors which differed in their H-2 haplotypes while sharing many similarities in the background genome and, conversely, effective suppressive activity of H-2 congenic donor sera when H-2-identities between donor and recipient mice existed, strongly suggested a role, at least in part, of H-2 genes in dictating the strain specificity of such suppressive activity. Additional experiments provided evidence for a possible role of macrophages in catabolism of the active molecules in CFA-immune sera. These observations, together with those presented in the preceding paper, may provide valuable insight toward successful development of appropriate manipulations that could ultimately convert high IgE responder individuals into low responders.

Effect of antigen dose on the secondary IgE response in BN rats.

Inbred Brown Norway rats were immunized at day 0 with a single dose of ovalbumin and 1 mg A1(OH)3. Various doses of ovalbumin without adjuvant were given at day 28. The secondary IgE antibody was antigen dose-dependent. Total serum IgE levels decreased after immunization and during the IgE antibody response.

Suppression of reaginic antibodies with modified allergens. II. Abrogation of reaginic antibodies with allergens conjugated to polyethylene glycol.

The two immunogenic preparations, ovalbumin (OA) and the nondialysable constituents of the aqueous extract of ragweed pollen (RAG), were conjugated with polyethylene glycols of molecular weights of 6,000 and 20,000 (PEG6 and PEG20) with the aid of cyanuric chloride. The i.v. administration of OA-PEG6 or OA-PEG20 into normal mice or into mice sensitized to a state of immediate hypersensitivity to dinitrophenylated OA (DNP3-OA) resulted, respectively, in the suppression of the primary or secondary IgE responses to DNP and OA. Similarly, the administration of RAG-PEG6 abrogated the primary as well as the ongoing anti-RAG reaginic responses in mice sensitized to RAG. The unresponsiveness of spleen cells from animals which had received a tolerogenic dose of OA-PEG6 was also maintained after transfer into X-irradiated (550 rad) mice. The suppressive effects of the tolerogenic PEG conjugates of OA and RAG were shown to be immunologically specific. It is, therefore, suggested that PEG-modified allergens may prove useful for the abrogation of the IgE response to a variety of allergens responsible for conditions of common hypersensitivity in man.

Suppression of reaginic antibodies with modified allergens. I. Reduction in allergenicity of protein allergens by conjugation to polyethylene glycol.

The conjugates of ovalbumin (OA) and of the nondialysable constitutents of the aqueous extract of ragweed pollen (RAG) with polyethylene glycols of molecular weights of 6,000 or 20,000 (PEG6 or PEG20) were shown to be nonantigenic, nonallergenic and nonimmunogenic. Thus, the i.v. administration of OA-PEG and RAG-PEG conjugates into mice did not elicit antibodies to OA and RAG, respectively, and these conjugates were shown to suppress in an immunologically specific manner the capacity of these animals to mount primary as well as secondary IgE responses to sensitizing doses of dinitrophenylated OA or of RAG. Moreover, the Peg-modified antigens did not combine either in vitro or in vivo with IgE antibodies directed against the natural antigens. Hence, OA-PEG and RAG-PEG conjugates were incapable of triggering allergic reactions in animals possessing IgE antibodies to the unmodified antigens. These PEG-modified antigens were also shown to be tolerogenic.

Suppression of Reaginic Antibody Formation

Abstract Reaginic antibodies to the benzylpenicilloyl determinant (BPO) and ovalbumin (OA) were induced readily in B6D2F1 mice by a single i.p. injection of either 1 or 10 µg of BPO4-OA suspended with 1 mg of Al(OH)3 in 0.5 ml of saline. Administration of conjugates consisting of the hapten coupled to the isologous, nonimmunogenic murine γ-globulins (MγG), i.e., BPO9-MγG, BPO11-MγG, or BPO12-MγG, resulted in complete and specific suppression of the induction of the anti-BPO reaginic antibody response without affecting, however, the level of reaginic antibodies to OA. Further study of the effect of epitope density on the immunologic properties of BPOx-MγG revealed that a) the lightly haptenated conjugates, BPO1-MγG and BPO2.9-MγG, were not immunosuppressive, b) the conjugates, BPO4.3-MγG and BPO19-MγG, were partially tolerogenic, and c) the heavily haptenated conjugate, BPO31-MγG, was nontolerogenic. Moreover, most importantly, the ongoing anti-BPO response in sensitized mice was readily abrogated by either four daily or four weekly injections of BPO9-MγG. The immunosuppressive effect of BPO12-MγG conjugates was dose dependent, complete suppression being achieved with 200 µg of the tolerogen. The unresponsiveness to BPO of spleen cells from immunosuppressed donors was also maintained in adoptive cell transfer experiments in spite of the additional administrationof the immunizing antigen under conditions expected to yield a secondary IgE response. Hence, it is suggested that, with special precautions to prevent unleashing an anaphylactic shock, treatment of penicillin-sensitive individuals with polyvalent conjugates of an appropriate number of BPO groups per human γ-globulin molecule would constitute a rational immunotherapeutic procedure for the abrogation of the allergic response to BPO.

HAPTEN-SPECIFIC IgE ANTIBODY RESPONSES IN MICE

The present studies have established conditions for the demonstration of cooperative interactions between specific T and B lymphocyte populations in the development of IgE antibody responses in vivo in mice. This has been accomplished by utilizing a system which permits the successful adoptive transfer to irradiated recipients of DNP-specific secondary IgE responses with spleen cells from suitably primed syngeneic donor mice. Thus, adoptively transferred DNP-KLH or DNP-ASC-primed spleen cells produced high levels of anti-DNP antibodies of both IgE and IgG antibody classes in response to challenge with the appropriate homologous priming conjugate but failed to develop more than meager responses to the reciprocal heterologous conjugate. However, when spleen cells from donors primed to the second carrier were concomitantly transferred with hapten-primed lymphocytes, secondary IgE ant-DNP responses were consistently obtained upon challenge with the heterologous conjugate. Moreover, we have been able to elicit augmented primary IgE anti-DNP antibody responses to either DNP-ASC or DNP-KLH after adoptive transfer of spleen cells from donors primed only to the carrier, ASC or KLH, respectively. This adoptive transfer system has enabled us to provide direct proof for the participation of theta-bearing T lymphocytes in antibody responses of the IgE class. Thus, the capacity of ASC-primed spleen cells to effectively cooperate with the DNP-KLH-primed lymphocytes in the adoptive secondary response to DNP-ASC could be abolished by in vitro treatment of such cells with anti-theta serum plus complement. This was true not only for the anti-DNP response of the IgG antibody class, but for the IgE antibody class as well. These studies have, furthermore, demonstrated the capacity to stimulate secondary anti-DNP antibody production in vivo by the concomitant administration of the DNP and relevant carrier determinants on separate molecules. This was more readily seen in the IgE than in the IgG antibody class. Thus, DNP-ASC-primed cells developed significant IgE, but more variable IgG, anti-DNP responses upon challenge with DNP-KLH plus unconjugated ASC. Antibody responses of both classes elicited in this manner were appreciably improved by the transfer of additional carrier (ASC)-primed cells. These and other results presented herein suggest that IgE B lymphocyte precursors may be inherently more sensitive than IgG B cells to at least certain of the functions of T lymphocytes concerned with regulatory mechanisms involved in antibody production.

Hapten-specific IgE antibody responses in mice. I. Secondary IgE responses in irradiated recipients of syngeneic primed spleen cells.

The present studies have established conditions for the demonstration of cooperative interactions between specific T and B lymphocyte populations in the development of IgE antibody responses in vivo in mice. This has been accomplished by utilizing a system which permits the successful adoptive transfer to irradiated recipients of DNP-specific secondary IgE responses with spleen cells from suitably primed syngeneic donor mice. Thus, adoptively transferred DNP-KLH or DNP-ASC-primed spleen cells produced high levels of anti-DNP antibodies of both IgE and IgG antibody classes in response to challenge with the appropriate homologous priming conjugate but failed to develop more than meager responses to the reciprocal heterologous conjugate. However, when spleen cells from donors primed to the second carrier were concomitantly transferred with hapten-primed lymphocytes, secondary IgE ant-DNP responses were consistently obtained upon challenge with the heterologous conjugate. Moreover, we have been able to elicit augmented primary IgE anti-DNP antibody responses to either DNP-ASC or DNP-KLH after adoptive transfer of spleen cells from donors primed only to the carrier, ASC or KLH, respectively. This adoptive transfer system has enabled us to provide direct proof for the participation of θ-bearing T lymphocytes in antibody responses of the IgE class. Thus, the capacity of ASC-primed spleen cells to effectively cooperate with the DNP-KLH-primed lymphocytes in the adoptive secondary response to DNP-ASC could be abolished by in vitro treatment of such cells with anti-θ serum plus complement. This was true not only for the anti-DNP response of the IgG antibody class, but for the IgE antibody class as well. These studies have, furthermore, demonstrated the capacity to stimulate secondary anti-DNP antibody production in vivo by the concomitant administration of the DNP and relevant carrier determinants on separate molecules. This was more readily seen in the IgE than in the IgG antibody class. Thus, DNP-ASC-primed cells developed significant IgE, but more variable IgG, anti-DNP responses upon challenge with DNP-KLH plus unconjugated ASC. Antibody responses of both classes elicited in this manner were appreciably improved by the transfer of additional carrier (ASC)-primed cells. These and other results presented herein suggest that IgE B lymphocyte precursors may be inherently more sensitive than IgG B cells to at least certain of the functions of T lymphocytes concerned with regulatory mechanisms involved in antibody production.