Distinct VH repertoires in primary and secondary B cell lymphocyte subsets in the preimmune repertoire of A/J mice: the CRI-A idiotype is preferentially associated with the HSA(low) B cell subset.

The anti-arsonate immune response of A/J mice is characterized by the occurrence of several recurrent idiotypes with a different temporal pattern of expression. The CRI-A idiotype is typically a memory idiotype since it appears late in the primary and dominates the secondary as well as subsequent immune responses. The CRI-C idiotype is present throughout the responses, including the primary one. Naive adult A/J mice treated repeatedly with anti-mu or anti-delta monoclonal antibodies exhibit a completely different balance of HSA(low) and HSA(high) B cell subsets and an opposite idiotype profile after immunization with p-azophenylarsonate coupled to hemocyanin. Anti-mu treatment leads to a striking enhancement of the HSA(low) cell subset associated with an earlier important synthesis of CRI-A(+) antibodies, while anti-delta treatment enhances significantly the HSA(high) compartment with a strong decrease of CRI-A and persistence of CRI-C1 antibodies. Semiquantitative PCR analysis reveals that the presence of CRI-A transcripts is associated with the HSA(low) compartment, while CRI-C transcripts are mainly associated with HSA(high) B cell subsets. This has been demonstrated with spleen cells of adult A/J mice treated with anti-mu or anti-delta antibodies and also with purified B cell subsets of unimmunized adult A/J mice and on neonatal spleen cells. It appears that the memory (CRI-A) idiotype is selected into the HSA(low) B cell subset before antigen arrival.

The perinatal presence of antigen (p-azophenylarsonate) or anti-mu antibodies lead to the loss of the recurrent idiotype (CRIA) in A/J mice.

The immune response of A/J mice against p-azophenylarsonate (Ars)-keyhole limpet hemocyanin (KLH) is characterized by the dominance, late in primary and during the secondary, of a recurrent idiotype called CRIA, encoded by a canonical combination of Ig gene segments. In this study, A/J mice were given Ars coupled to deaggregated human gamma globulins (dHGG) within 24 h after delivery. The offsprings from these mice were then exposed as adults to Ars-KLH. These animals developed an unusual immune response. The level of anti-Ars antibodies was nearly normal but a dramatic shift in repertoire was observed: the cross-reactive idiotype which is the hallmark of the anti-Ars response in A/J mice was completely absent. The idiotype could be recovered by injection of anti-idiotypic antibodies alone, with no need of lipopolysaccharide coupling. Therefore the presence of antigen at birth can lead to a strong perturbation of idiotype selection. Similar results were obtained with neonatal treatment using anti-IgM antibodies. After recovery of suppression, A/J mice can mount an anti-arsonate response of normal level but devoid of the dominant idiotype.

Lack of T cell tolerance in mice exposed to a protein antigen through lactation.

Offspring of mother mice treated immediately after delivery with deaggregated human gamma-globulins (dHGG) are unable to produce HGG-specific antibodies when challenged with immunogenic forms of HGG (HGG/CFA) in adulthood. Despite a defective antibody response, animals rendered tolerant to HGG as neonates retain tolerogen-specific T cells able to proliferate and secrete lymphokines. The pattern of IL-2 and IL-4 secretion by T cells isolated from tolerant animals could not be distinguished from the corresponding cells in control mice, suggesting that neonatal exposure to dHGG did not affect T cell reactivity or Th1/Th2 in vivo balance. Moreover, immunization of tolerant animals with haptenated HGG confirmed the presence of tolerogen-specific helper T cells in vivo. Functional T cell depletion by anti-CD3 mAbs during lactation failed to modify induction of B cell tolerance, suggesting that T cells are neither affected nor required to induce the selective tolerance status observed in this model. Based on the finding that antigen-presenting cell functions in secondary organs (spleen, peritoneal cavity) are a late acquisition during ontogeny and reach adult-like levels at weaning, we propose that most soluble proteins elude T cell recognition during lactation due to defective antigen presentation.

Induction of Th2 responses to soluble proteins is independent of B cell tolerance status.

Injection of high doses of monomeric human gamma globulins (dHGG) in naive, adult mice causes antigen-specific tolerance of B cell and Th1 lymphocytes, while inducing the selective expansion of antigen-specific Th2 cells. Several parameters of tolerance induction were analyzed in this work, in order to establish whether B cell tolerance and Th1 unresponsiveness were functionally related in this in vivo model. By varying the antigen form and site of injection, we demonstrate in this work that Th1 unresponsiveness to HGG is not a consequence of peripheral B cell tolerance. In particular, mice pretreated with heat-aggregated antigen (HAHGG) or F(ab')2 HGG were found to develop a strong humoral response while displaying a defective Th1 response. In fact, these animals developed a strong Th2 response in vivo, demonstrating that selective expansion of antigen-specific Th2 cells in this model is not a consequence of B cell tolerance or antigen capture by Fc receptor-expressing cells. We conclude that while B cell tolerance in this model is only observed in response to deaggregated antigen, injection of all forms of adjuvant-free, protein antigens induces T helper precursor cells to differentiate into Th2-type helper cells in vivo irrespectively of the B cell tolerance status.

The injection of deaggregated gamma globulins in adult mice induces antigen-specific unresponsiveness of T helper type 1 but not type 2 lymphocytes.

Injection of adult mice with high doses of monomeric human gamma globulins (dHGG) has been previously shown to produce a state of peripheral tolerance in both B and T cells. To gain insight into the mechanism of induction and maintenance of adult tolerance in this model, we have analyzed the pattern of lymphokines produced by control and tolerant animals in response to the tolerogen. The data presented indicate that HGG-specific, interleukin 2 (IL-2)- and interferon gamma (IFN-gamma)-producing T cells (thus referred to as T helper type 1 [Th1] cells) are rendered unresponsive after in vivo administration of soluble HGG. In contrast, antigenic stimulation of T cells isolated from tolerant adult mice leads to increased production of IL-4 in vitro. In vivo challenge of dHGG-treated adult animals with hapten-coupled HGG (p-azophenylarsonate [ARS]-HGG) induced a significant ARS-specific antibody response, suggesting that tolerance induction in this model does not completely abrogate tolerogen-specific Th activity in vivo. In agreement with the in vitro data, hapten-specific antibody response of tolerant animals is characterized by a selective deficiency in the IFN-gamma-dependent IgG2a subclass. Injection of immunogenic forms of HGG into tolerant animals also produced an IL-4-dependent increase in total serum IgE levels, indicative of an increased activity of HGG-specific Th2 cells in these animals. The finding that tolerance induction differentially affects Th subpopulations suggests that crossregulation among lymphocyte subsets may play a role in the induction and/or maintenance of acquired tolerance in adults.