A key event in the pathogenesis of allergies is the production of antibodies of the immunoglobulin (Ig)E class. In normal individuals the levels of IgE are tightly regulated, as illustrated by the low serum IgE concentration. In addition, multiple immunizations are usually required to generate detectable IgE responses in normal experimental animals. To define the parameters that regulate IgE production in vivo, we generated mice bearing monoclonal populations of B and T lymphocytes specific for influenza virus hemagglutinin (HA) and chicken ovalbumin (OVA), respectively. A single immunization of the monoclonal mice with the cross-linked OVA-HA antigen led to serum IgE levels that reached 30-200 g/ml. This unusually high IgE response was prevented by the infusion of regulatory ␣ / ␤ CD4 ϩ T cells belonging to both CD25 ϩ and CD25 Ϫ subpopulations. The regulation by the infused T cells impeded the development of fully competent OVA-specific effector/memory Th2 lymphocytes without inhibiting the initial proliferative response of T cells or promoting activation-induced cell death. Our results indicate that hyper IgE responses do not occur in normal individuals due to the presence of regulatory T cells, and imply that the induction of regulatory CD4 ϩ T cells could be used for the prevention of atopy.

Fetal-pig fibroblasts homozygous for the knockout of the alpha1-3 galactosyltransferase gene appear to express low but detectable levels of the gal antigen.

We have previously produced a series of antigalactose (anti-Gal) hybridomas and characterized their heavy chain gene usage. Here we have quantified the affinity of these Abs for the α-Gal epitope and characterized their in vitro effects on endothelial cell activation and apoptosis. We report that anti-Gal mAbs derived from Gal−/− mice show a range of affinity for the α-Gal epitope, and that affinity was generally increased as the VH gene usage transitioned from germline sequences to sequences exhibiting somatic maturation. Despite an 85-fold range in affinity, all the anti-Gal mAbs examined induced α-Gal-specific endothelial cell activation, and after prolonged exposure induced endothelial cell apoptosis in a complement-independent manner. Only murine anti-Gal mAbs of the IgM or IgG3 subclass, but not IgG1, were effective at initiating complement-dependent cell lysis. Using a novel rat to mouse xenograft model, we examined the in vivo ability of these mAbs to induce xenograft rejection and characterized the rejection using histology and immunohistochemistry. Infusion of complement-fixing IgG3 mAbs resulted in either hyperacute rejection or acute vascular rejection of the xenograft. Surprisingly, infusion of an equal amount of a high affinity anti-Gal IgG1 mAb, that fixed complement poorly also induced a rapid xenograft rejection, which we have labeled very acute rejection. These studies emphasize the importance of in vivo assays, in addition to in vitro assays, in understanding the role of anti-Gal IgG-mediated tissue injury and xenograft rejection.

These observations suggest that this model can be used to study the regulation of anti-Gal B cells and can establish a reliable source of functional anti-Gal B cells, which could be used to test the effectiveness of alpha-Gal-specific immunosuppressive reagents.

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