Enhancement of antibody responses by IgD-binding factors induced by anti-IgD treatment of spleen cells.

Treatment of normal BALB/c splenocytes with anti-mouse IgD antibodies at 0 degrees C followed by incubation of the cells at 37 degrees C resulted in the formation of soluble factors that selectively inhibit rosette formation of lymphocytes bearing Fc delta receptors with IgD-coated erythrocytes (i.e., IgD-binding factors).

The low-affinity receptor for IgE (CD23) on B lymphocytes is spatially associated with HLA-DR antigens.

Two hybridomas that produce the mAbs 135 and 449 B4 were obtained that inhibited the binding of IgE to the Fc epsilon RL/CD23 on the EBV-transformed B cell line RPMI 8866. mAb 135 was obtained from a mouse immunized with RPMI 8866 cells, whereas mAb 449B4 was obtained from a mouse immunized with a partially purified preparation of Fc epsilon RL/CD23 obtained as the eluate of an IgE immunoabsorbent loaded with a soluble extract of RPMI 8866 cells. These two mAbs bound to Fc epsilon RL/CD23- cell lines and precipitated two polypeptides with 36,000 Mr and 28,000 Mr, which were the HLA-DR alpha and beta chains, respectively.

Monoclonal antibody specific for T cell-derived human IgE binding factors.

A B cell hybridoma secreting monoclonal antibody against human IgE binding factors was obtained by immunization of BALB/c mice with partially purified IgE binding factors, and fusion of their spleen cells with SP-2/0-AG14 cells. The monoclonal antibody bound all of the 60,000, 30,000, and 15,000 dalton IgE binding factors from two T cell hybridomas and those from activated T cells of a normal individual. The antibody bound both IgE-potentiating factors, which had affinity for lentil lectin, and IgE-suppressive factors, which had affinity for peanut agglutinin.

Augmentation of the antibody response by antigen-specific glycosylation-enhancing factor.

BDF1 mice were immunized with a protein antigen, such as ovalbumin (OA) or keyhole limpet hemocyanin (KLH), absorbed to aluminum hydroxide gel, and their spleen cells were stimulated by homologous antigen for the formation of glycosylation-enhancing factor (GEF). It was found that GEF obtained from OA-primed spleen cells had affinity for OA, whereas those derived from KLH-primed spleen cells had affinity for KLH. Nonspecific GEF, which was obtained by stimulation of normal spleen cells with pertussis toxin, failed to bind OA or KLH.

In vitro modulation of antigen-primed T cells by a glycosylation-inhibiting factor that regulates the formation of antigen-specific suppressive factors.

BDF1 [(C57BL/6 X DBA/2)F1] mice were primed with alum-absorbed ovalbumin, and their spleen cells were cultured with ovalbumin to activate antigen-primed T cells. The activated T cells were then propagated in interleukin 2-containing medium in the presence or absence of affinity-purified glycosylation-inhibiting factor (GIF). Upon incubation with ovalbumin-pulsed macrophages, T cells propagated in the absence of GIF produced IgE-potentiating factor and glycosylation-enhancing factor.

Carrier-specific suppression of antibody responses by antigen-specific glycosylation-inhibiting factors.

We previously established an ovalbumin (OA)-specific T cell clone from spleen cells of BDF1 mice, which had been treated by i.v. injections of OA, and constructed antigen-specific T cell hybridomas from the T cell clone.

Potentiating and suppressive IgE-binding factors are expressed by a single cloned gene.

We have investigated expression of an IgE-binding factor (IgE-BF) cDNA in both COS-7 monkey kidney cells and Chinese hamster ovary cells. Transient expression of the IgE-BF clone in either cell type yielded IgE-BF, which potentiated an in vitro IgE response and had an affinity for lentil lectin. In contrast, when the transient expression experiments were carried out in the presence of tunicamycin, the factors no longer bound to lentil lectin.

Effects of cyclophosphamide treatment and gamma irradiation of SJL mice on the IgE antibody response and the nature of IgE-binding factors.

Immunization with alum-absorbed ovalbumin (OA) failed to induce IgE antibody responses in SJL mice, while mice pretreated with either cyclophosphamide (CY) or gamma-irradiation prior to immunization transiently formed IgE antibodies. Spleen cells of OA-primed SJL mice formed IgE-suppressive factors upon incubation with OA. In contrast, spleen cells of the CY-treated or irradiated mice formed IgE-potentiating factors.

T cell factors involved in the regulation of the IgE synthesis.

The IgE-potentiating and IgE-suppressive factors share a common structural gene and therefore a common polypeptide chain, and their biologic activities are decided by a post-translational glycosylation process. Under physiological conditions, this process is controlled by two T cell factors, i.e.

Biological properties of a recombinant human immunoglobulin epsilon-chain fragment.

A recombinant human immunoglobulin epsilon-chain gene expression product (rFc epsilon) was compared with a human E myeloma protein in the affinity for epsilon-chain Fc fragment receptors (Fc epsilon R) on cultured human basophils. The association-dissociation kinetics of the rFc epsilon-Fc epsilon R interaction are indistinguishable from that of E myeloma protein, indicating that rFc epsilon and IgE have identical affinity for the receptors. The recombinant gene product sensitizes cultured basophils for anti-IgE-induced histamine release.

IgD-binding factors from mouse T lymphocytes.

Incubation of normal mouse splenic lymphocytes with dimeric mouse IgD results in the formation of T-cell factors that selectively inhibit rosette formation of Fc delta receptor-positive lymphocytes with IgD-coated erythrocytes. The rosette-inhibiting factors bound to IgD-Sepharose and were recovered by elution at acid pH. The results indicate that the rosette-inhibiting factors are IgD-binding factors.

Rodent IgE-binding factor genes are members of an endogenous, retrovirus-like gene family.

Synthesis of IgE by B lymphocytes can be regulated by soluble lymphocyte factors which have affinity for the Fc region of IgE (IgE-binding factors). In previous studies, we identified cDNA clones encoding rodent IgE-binding factors by direct expression in transfected mammalian cells. Here we show that IgE-binding factor cDNA clone 8.

Immunosuppressive effects of glycosylation inhibiting factor on the IgE and IgG antibody response.

Glycosylation inhibiting factor (GIF) was purified from culture filtrates of a T cell hybridoma, 23A4, by affinity chromatography on anti-lipomodulin Sepharose. The factor exhibited phospholipase inhibitory activity upon dephosphorylation. Immunization of BDF1 mice with aluminum hydroxide gel (alum)-absorbed dinitrophenyl derivatives of ovalbumin (DNP-OA) resulted in persistent IgE and IgG antibody formation.

Induction of Fc epsilon receptors on mouse macrophages and lymphocytes by homologous IgE.

Normal mouse peritoneal macrophages express Fc gamma 2aR, Fc gamma 1/2bR, and Fc epsilon R, whereas B lymphocytes in mesenteric lymph nodes (MLN) bear Fc gamma 2bR, Fc gamma 1R, and Fc epsilon R. Rosette formation of Fc epsilon R+ macrophages and lymphocytes with IgE-coated ox erythrocytes was inhibited by rodent IgE but not by any other isotype of mouse immunoglobulins. In contrast, IgG1 rosettes and IgG2b rosettes of both macrophages and lymphocytes were inhibited not only by homologous isotype but also by IgE, suggesting that IgE has some affinity for Fc gamma 1/2bR on macrophages and for both Fc gamma 1R and Fc gamma 2bR on lymphocytes.

Regulatory effects of human IgE-binding factors on the IgE response of rat lymphocytes.

Normal human peripheral blood T cells were propagated in the presence of human interleukin 2, and activated cells were incubated with human IgE-dimer to induce IgE binding factor formation. The cells were then fused with a mutant of the human T cell line CEM. Five of the T cell hybridomas formed IgE binding factors upon incubation with human IgE-dimer.

Association of I-J determinants with lipomodulin/macrocortin.

Stimulation with glucocorticoids of mouse splenic lymphocytes and peritoneal adherent cells resulted in the formation of a soluble factor--i.e., glycosylation inhibiting factor (GIF)--that inhibits the assembly of N-linked oligosaccharide(s) to IgE-binding factors during their biosynthesis.

Antigen-induced increase in protein kinase C activity in plasma membrane of mast cells.

Bridging of cell-bound IgE antibody molecules on colony-stimulating factor-dependent mouse mast cell line (PT-18) cells by multivalent antigen induces phospholipid methylation, a transient rise in intracellular cAMP, intracellular mobilization and uptake of Ca2+, and formation of diacylglycerol followed by histamine release. Exposure of the sensitized cells to antigen also induces a substantial increase in protein kinase C activity in the plasma membrane, which is accompanied by a slight decrease in the enzyme in cytosol. Protein kinase C activity in the membrane fraction reached maximum within 30 sec after antigen challenge and then gradually declined.

Relationship among IgE-binding factors with various molecular weights.

The rat-mouse T cell hybridoma 23B6 forms IgE-binding factors on incubation with IgE. The hybridoma cells incubated with IgE contained intracellular IgE-binding factors of 60K, 30K, 14K, and 10K daltons, and secreted the 60K, 30K, and 14K IgE-binding factors. Both intracellular and extracellular IgE-binding factors of 60K and 14K daltons selectively suppressed the IgE response, whereas the 30K and 10K factors failed to do so.

Presence of an antigen-specific T cell subset that forms IgE-suppressive factor and IgG-suppressive factor on antigenic stimulation.

B6D2F1 mice were given three i.v. injections of ovalbumin (OA), and antigen-specific T cell clones were established from their spleen cells.

Biochemical analysis of desensitization of mouse mast cells.

Biochemical mechanisms of desensitization were explored by using peritoneal mouse mast cells saturated with monoclonal mouse IgE anti-DNP antibody. It was found that a 1-min incubation of the sensitized cells with 0.01 micrograms/ml DNP-HSA in the absence of Ca2+ was sufficient to desensitize the cells completely.