Peripheral blood based T cell-containing and T cell-depleted culture systems for human IgE synthesis: the role of T cells

Background Comparable T cell-containing and T cell-depleted culture systems for human IgE synthesis are currently not available.
Objective This has prompted us to develop peripheral blood mononuclear cell (PBMC) based culture systems for human IgE synthesis in the presence and absence of T cells.
Methods In this paper we describe simplified conditions for in vitro synthesis of high levels of IgE by human peripheral blood B cells, both in T cell-containing cultures and in anti-CD40 stimulated T cell-depleted cultures.
Results T cell-depieted cultures released approximately 20 times more IgE [range 410–2220 ng/mL (mean 1270 ng/mL); based on six experiments using cells from three donors] than did T cell-containing cultures [range 23–105 ng/mL (mean 58 ng/mL); based on 15 experiments using cells from three donors]. Reconstitution experiments were performed to investigate the role of T eells on IgE synthesis. Adding T eells back to the anti-CD40 stimulated T cell-depleted cultures resulted in a dose-dependent inhibition of IgE production. In the absence of anti-CIMO low numbers of T cells stimulated, while high numbers suppressed, IgE production: the optimal ratio of T cells to non-T cells for maximal IgE production was found to be 1:1. At this ratio, irradiated (non-replicating) T cells supported a much greater IgE synthesis than did non-irradiated T cells.
Conclusion The development of these systems provides directly comparable T cell-containing and T cell-depleted cultures for human IgE synthesis from peripheral blood, allowing further study of the role of T cells in IgE regulation. These systems will also be of use for determining whether potential modulators of IgE synthesis act on the T cells or on other cell types.     

Expression of receptors for IgA on mitogen-stimulated human T lymphocytes

This study demonstrates that activation of human peripheral blood mononuclear cells (PBMC) by the T cell mitogens phytohemagglutinin (PHA) and concanavalin A (Con A) induces the expression of receptors for IgA without addition of IgA to the culture medium. Cells bearing receptors for IgA were determined by indirect immunofluorescence using human secretory IgA and fluoresceinated goat anti-human IgA or goat anti-secretory component antibodies. Among freshly isolated PBMC, 4.7 +/- 1.7% of T cells, 12.7 +/- 12.5% of B cells and 14.4 +/- 7.6% of monocytes were found to be IgA receptor positive. In unstimulated PBMC cultures the percentage of IgA receptor-positive cells slightly increased at 48 h and was more elevated after 7 days. In Con A-stimulated cultures 24.3 +/- 18.5% of the cells expressed receptors for IgA after 48 h. Then, the number decreased and rose again thereafter. PHA stimulation induced an increase of smaller magnitude with similar kinetics. Induction of receptor for IgA on activated T cells was demonstrated by double-labelling experiments showing more CD8+ than CD4+ cells with receptors for IgA among Con A-activated PBMC. Furthermore, PHA or Con A stimulation of B cell-depleted PBMC suspensions resulted in a marked increase of cells bearing receptors for IgA. Expression of these receptors was down-regulated by recombinant interferon-gamma (250 units/ml) and by prostaglandin PGE2 (100 nM) both on unstimulated and mitogen-activated PBMC. The receptor for IgA was distinct from the asialoglycoprotein receptor and did not cross-react with the poly-Ig receptor of epithelial cells. It was concluded that, in the absence of inducing exogenous IgA, T cell mitogens trigger the expression of receptors for IgA. Therefore, T cell activation is associated with the down-regulation of receptors for IgM and the increased expression of receptors for IgG, IgA and IgE.     

The effect of cyclosporine on expression of class II major histocompatibility complex antigens on bronchoalveolar cells and peripheral blood mononuclear cells

To evaluate the effect of cyclosporine (CsA) on the expression of class II major histocompatibility complex (MHC) antigens on bronchoalveolar cells (BAC) and peripheral blood mononuclear cells (PBMC), BAC and PBMC were obtained from mongrel dogs before and during CsA treatment. Expression of MHC class II antigens on BAC and PBMC were detected by monoclonal antibodies (Mabs) B1F6, 7.5.10.1 and Q5/13, which recognized canine MHC class II antigens, using cytofluorometry. Total cell counts and cell differentials of canine BAC showed no significant difference before or during CsA treatment (P greater than 0.05). Anti-MHC class II Mabs used in this study reacted with 21-51% of canine BAC and with 31-69% of PBMC. After stimulation with phytohaemagglutinin (PHA) the percentages of MHC class II positive BAC and PBMC were significantly increased (P less than 0.001). Whole blood levels of CsA were 315 +/- 76 (mean +/- SD) ng/ml and 343 +/- 57 ng/ml on days 7 and 14 during CsA treatment at an oral dose of 20 mg/kg/day. During CsA treatment there was no significant difference in the percentages of MHC class II positive BAC and PBMC compared with data obtained before CsA treatment (P greater than 0.05). We likewise failed to observe a suppressive effect of CsA on the increased expression of MHC class II antigens on BAC and PBMC induced by PHA (P greater than 0.05). In summary, at an oral dose of 20 mg/kg/day for a period of two weeks, our results show that: (1) CsA does not affect the total cell counts and cell differentials of canine BAC; (2) CsA does not reduce the basal expression of MHC class II antigens on canine BAC and PBMC; (3) CsA does not suppress the increased expression of MHC class II antigens on canine BAC and PBMC induced by PHA.     

IL-4-producing murine T helper cell line provides help for in vitro production of IgE.

Studies of the regulation of the IgE synthesis have been hampered by the difficulties in making lymphocytes produce IgE in vitro. Synthesis of IgE, IgG1 and IgM were investigated in a murine coculture system in which a conalbumin-specific T helper cell line of Th2 type was cultured together with nonadherent splenocytes from normal MHC-matched mice. Help provided by the antigen-stimulated T cell line induced significant IgE production (20 ng/ml), along with IgG1 (5 micrograms/ml) and IgM (250 micrograms/ml). Immunoglobulin synthesis in cultures was detectable at day 3-4 and culminated at day 7-8. IL-4 production was observed within the first 2 days of culture. The kinetics of the synthesis of the three isotypes were parallel. Stimulation of the Th2 cell line with Con A produced dose-response curves resembling those after antigen stimulation, whereas supernatant from these cells was unable to induce synthesis of IgE. The presence of interferon-gamma in the cultures, inhibited synthesis of IgE, IgM and IgG1, but this inhibition was not due to interference with the IL-4 production, which increased in the presence of high doses of interferon-gamma.     

Effects of retinoids on in vitro and in vivo IgE production

BACKGROUND: Retinoids modulate the growth and number of different cell types, including B cells. We could previously show that retinoic acid (RA) strongly inhibits CD40 + IL-4-mediated IgE production in vitro. The aim of the present study was to extend these findings regarding the potential use of retinoids for the treatment of allergic diseases. METHODS: In vitro IgE production was studied in anti-CD40 + IL-4-stimulated peripheral blood mononuclear cells (PBMC) from allergic donors in the presence of 10(-15)-10(-5) M all-trans and 13-cis RA and in ovalbumin (OVA)-sensitized BALB/c mice treated with RA (20 mg/kg) before and during sensitization. IgE and IgG1 levels were determined in the sera of the mice at day 21 after 2 injections (days 1 and 8) of aluminum hydroxide-absorbed OVA. RESULTS: All-trans and 13-cis RA inhibited in vitro IgE production from PBMC in a dose-dependent manner, but were more efficient in atopic dermatitis patients with low total serum IgE levels (< 400 kU/ml), maximal inhibition for all-trans RA at 10(-7) M (87%) and for 13-cis RA at 10(-5) M (96%) compared to patients with high serum IgE levels (>2,000 kU/ml), maximal inhibition for both all-trans and 13-cis RA at 10(-5) M (53 and 39%, respectively). In contrast, the in vivo data from OVA-sensitized mice revealed comparable total IgE and IgG1 levels in control versus all-trans RA or CD336-treated groups, specific IgE was even higher in the CD336-treated group (n = 10, 2,814 ng/ml), and was comparable in mice treated with OVA alone or with additional all-trans RA (n = 10, 1,447 and 1,354 ng/ml, respectively). CONCLUSIONS: These results indicate that the efficacy of retinoids to inhibit IgE production in vitro depends on the frequency of switched cells in the peripheral blood and that in an in vivo model using OVA-sensitized mice, retinoids fail to inhibit IgE production.     

Induction of a proliferative response of T cells by a high-molecular antigen in Dermatophagoides farinae feces

BACKGROUND: We have previously demonstrated that high-molecular mite antigen (HM1) from Dermatophagoides farinae feces is an allergen which binds to mite-allergic patients IgE. HM1 also induced a proliferative response in lymph node cells from mite-immunized mice as well as nonimmunized mice. In the present study, we demonstrated that HM1 induced T cell proliferation and investigated the HM1-stimulated T cell proliferative pathways using nonallergic human peripheral blood mononuclear cells (PMBC). METHODS: Blood samples were obtained from 10 healthy donors. Using primary culture, T cell response stimulated with HM1 was performed on purified T cells, CD19+ cell-depleted PBMC and CD11b+ cell-depleted PBMC. In addition, interleukin (IL)-5 and interferon (IFN)-gamma produced by mite-allergic and healthy donors stimulated with HM1 were estimated by enzyme immunoassay. RESULTS: T cell proliferation was detected only in CD19+ cell-depleted PBMC. When T cells were cocultured with CD11b+ cells they recovered their proliferative response to HM1. In addition, the pathway of HM1-stimulated T cell proliferation did not involve HLA class II restriction. Both activated CD11b+ cells and their conditioned media were needed to induce HM1-stimulated T cell proliferation. Furthermore, HM1 induced IFN-gamma production in both healthy and allergic donors. CONCLUSION: The high-molecular mite antigen, HM1, induced a proliferative response of T cells in healthy as well as allergic donors, without HLA class II restriction. Our results suggest that further investigation of HM1 could constitute a valuable avenue of research into complex allergic diseases.     

IgE production in vitro by peripheral blood mononuclear cells of patients with parasitic helminth infections.

Helminth parasites induce production of high levels of IgE antibodies but the immunoregulatory mechanisms determining this IgE biosynthesis are poorly understood. To investigate these mechanisms, peripheral blood mononuclear cells were obtained from six normal controls, six atopic patients and eight patients with parasitic helminth infections (three with schistosomiasis, two with loiasis, three with onchocerciasis). Cells were cultured at 1 X 10(6) cells/ml for 8 days in the presence of media alone or media supplemented with pokeweed mitogen (PWM) or cycloheximide; the supernatant fluids from these cultures were then assayed quantitatively for total and parasite specific IgE and IgG using an avidin-biotin amplified (for IgE) or standard (for IgG) microelisa assay. The geometric mean spontaneous IgE production was markedly elevated in peripheral blood mononuclear cells from parasitized individuals (2,487 pg/ml) when compared to those from atopics (358 pg/ml) or normals (152 pg/ml). Spontaneous IgG synthesis was equivalent in all three groups (range 140-420 ng/ml). PWM did not induce IgE production in any group and in the parasitized group even caused significant suppression of total IgE synthesis. Antigen specific antibody production (both IgE and IgG) paralleled total immunoglobulin synthesis. These findings demonstrate for the first time spontaneously enhanced IgE production in vitro in patients with helminth infections and provide a model system for studying the suppressive and regulatory mechanisms controlling IgE secretion.     

Omalizumab decreased IgE-release and induced changes in cellular immunity in patients with allergic asthma

BACKGROUND: Omalizumab, a recombinant monoclonal anti-immunoglobulin E (IgE) antibody, shows proven efficacy in the treatment of allergic diseases. A little is known about the immunological pathways affected by the decrease of circulating free IgE during omalizumab treatment. AIM OF THE STUDY: To investigate the immunological consequence of IgE withdrawal, we studied the influence of omalizumab on stimulated IgE-release of cultured peripheral blood mononuclear cells (PBMC) and on the relative number of lymphocytes in the peripheral blood (cellular immune status) in patients with allergic asthma. METHODS: Nineteen patients were enrolled and received omalizumab at a dose of at least 0.016 mg/kg/IgE (IU/ml) every 4 weeks. PBMC were isolated from peripheral blood. Cells were cultured and stimulated with IL-4 (5 ng/ml) and CD40 ligand (1 microg/ml) for 10 days. IgE release was detected in cell culture supernatants by enzyme-linked immunosorbent assay (ELISA). Cellular immune status was investigated by fluorescence-activated cell sorting. RESULTS: Omalizumab treatment induced significant inhibition of stimulated IgE release (median 1.38-0 ng/ml vs. 1.64-2.0 ng/ml in placebo group, P<0.05). B-lymphocyte counts were also significantly lower in the omalizumab group compared with placebo after 12 weeks of treatment (median 18.2-15.6% lymphocytes vs 12.7-13.7% lymphocytes after placebo, P<0.01). There were no significant differences in the other lymphocyte subpopulations between the groups. CONCLUSIONS: These findings provide evidence of immunological influences of omalizumab treatment, leading to a downregulation of IgE secretion and decrease of lymphocyte subpopulations (B-cells) indicating their anti-inflammatory potency.     

The modulation of B7.2 and B7.1 on B cells by immunosuppressive agents.

Several recent studies demonstrate that B7.2, but not B7.1, play an important role in allergic inflammation and IgE production. Agents that down-regulate B7.2 may therefore be of benefit for the treatment of Th2-driven allergic diseases. Our current study was carried out to investigate the effect of immunosuppressive agents, cyclosporin A (CsA) and dexamethasone, on B7.2 and B7.1 expression on B cells stimulated with the superantigen, toxic shock syndrome toxin-1 (TSST-1). The analysis of B7.2 and B7.1 on the same cells by flow cytometry demonstrated that TSST-1 up-regulated B7.2+B7.1- but not B7.1+B7.2- on B cells in a dose-dependent fashion. CsA and dexamethasone significantly down-regulated B7.2+B7.1- but up-regulated B7.2-B7.1+ B cells in the presence or absence of TSST-1 (100 ng/ml). Interestingly, the combination of CsA and dexamethasone was much more potent in the inhibition of B7.2 expression than either of these agents alone. As CD40 is known to up-regulate B7.2 expression on B cells, the mechanism of B7.2 down-regulation by CsA and dexamethasone was further studied by investigating the effect of these agents on CD40 expression on B cells. TSST-1 significantly increased CD40 expression on B cells. However, the addition of CsA or dexamethasone significantly down-regulated CD40 expression. Anti-CD40 MoAb significantly reversed the effects of CsA or dexamethasone on B7.2 and B7.1 expression, suggesting that T cell engagement of CD40 plays a role in the mechanisms by which CsA and dexamethasone acts on B cells. These data demonstrate the modulatory effect of CsA and dexamethasone on B7.2 and B7.1 expression on B cells and the potential role of CD40 in mediating this effect.     

Suppression of IgE synthesis in vitro by allogeneic T cells from atopic and non-atopic subjects.

The role of T cells in the regulation of IgE synthesis by human PBMC was studied. PBMC or separated and recombined populations of T and B cells from both normal and atopic donors were cultured for 10 days with and without cycloheximide. IgE and IgG synthesis were determined by specific RIA. IgE synthesis was detected in 0/30 non-atopic, 6/34 mildly atopic and 25/31 severely atopic subjects. Autologous T cells from 10/26 atopic donors, whose B cells synthesised IgE, significantly suppressed this IgE synthesis. The addition of allogeneic T cells from atopic or non-atopic subjects to atopic B cells resulted in greater suppression of IgE synthesis than the addition of autologous T cells. These data support the notion that atopic subjects have naturally occurring IgE isotype-specific suppressor T cells as well as suppressor T cells which can be activated during incubation with alloantigen.     

Inhibition of human IgE synthesis by anti-IgE antibodies requires divalent recognition

We used a selection of well-characterized murine monoclonal anti-IgE antibodies to investigate their effect on human in vitro IgE synthesis. We found anti-IgE antibodies that either inhibited or enhanced interleukin-4 plus anti-CD40-induced in vitro IgE synthesis in peripheral blood mononuclear cells (PBMC). This differential activity was isotype specific as neither IgM nor IgG synthesis were affected. Interestingly, only coding IgE mRNA was down-regulated, whereas germ-line ε RNA expression was not influenced by anti-IgE monoclonal antibody (mAb). On purified B cells all anti-IgE mAb inhibited interleukin-4 plus anti-CD40-induced IgE synthesis, implying a role of non-B cells for the enhancing activity observed in PBMC. Using Fab and F(ab')₂ of an inhibitory anti-IgE mAb we could show that divalent recognition was required for inhibition of IgE synthesis.     

Influence of cytokines and cellular interactions on the glucocorticoid-induced Ig (E, G, A, M) synthesis of peripheral blood mononuclear cells.

We studied the mechanisms of action leading to the glucocorticoid (GC)-induced synthesis of the immunoglobulins (IgE, G, A, M) by human peripheral blood mononuclear cells (PBMC). It is shown that the enhanced Ig synthesis of GC-stimulated PBMC is dependent on the presence of T cells and monocytes. After stimulation of purified B cells with GC only a slight enhancement of IgM could be detected. Inhibition studies with neutralizing anti-interleukin-4 (IL-4) and anti-IL-6 antibodies revealed that the GC-induced IgE synthesis of PBMC is not dependent on the presence of IL-4 or IL-6. Stimulation of membrane-separated and co-cultured cell fractions revealed that the GC-induced enhancement of IgA and IgM synthesis is mediated by T-cell derived soluble mediators. The GC-induced IgG and IgE synthesis is dependent upon contact of B cells with monocytes. Antibodies against LFA-1 and ICAM-1 are capable to suppress the GC-induced IgE and IgG synthesis of PBMC. Furthermore, the monocyte expression of lymphocyte function antigen-1 (LFA-1), intercellular adhesion molecule-1 (ICAM-1) and HLA-DR is modulated by GC stimulation.     

High-level IL-10 production by monoclonal antibody-stimulated human T cells.

We investigated interleukin-10 (IL-10) production in freshly isolated mononuclear cells and purified T cells in response to stimulation with monoclonal antibodies (mAb) recognizing CD3, CD2 and CD28, or with the bacterial products Staphylococcus aureus cells (SAC), staphylococcal enterotoxin (SEA) and lipopolysaccharide (LPS). IL-10 production was compared with that of IL-2, IL-4 and interferon-gamma (IFN-gamma). Similar to the other cytokines, in peripheral blood mononuclear cells (PBMC) from adult donors the highest IL-10 levels were produced in response to CD2 plus CD28 stimulation, within 72-96 hr of stimulation. Levels of IL-10 in response to CD2 plus CD28 stimulation (1.9 +/- 1 ng/ml) exceeded those in response to SEA (0.25 +/- 0.16 ng/ml), SAC (0.43 +/- 0.42 ng/ml), or LPS (0.19 +/- 0.14 ng/ml) stimulation. With adult purified T cells, high levels of IL-10 and IL-4 were measured following CD3 plus CD28 stimulation, and the amounts of both T-helper type-2 (Th2) cytokines decreased following the addition of phorbol myristate acetate (PMA), whereas the synthesis of the Th1 cytokines IL-2 and IFN-gamma was enhanced. When PBMC were stimulated with a CD3 mAb and different other cytokines were added, strong enhancement of IL-10 production was seen upon the addition of IL-2, IL-4, IL-7, IL-12 and IFN-gamma, whereas inhibition was found with transforming growth factor-beta 1 (TGF-beta 1). These data illustrate that in freshly isolated PBMC large amounts of IL-10 can be induced rapidly by appropriate mAb stimulation, and that even in freshly isolated cells IL-4 and IL-10 show signs of parallel regulation.     

Improved immunoglobulin production in dialysis patients treated with recombinant erythropoietin.

Improvements in B lymphocyte function have been reported in hemodialysis patients receiving erythropoietin. The present investigation studied whether erythropoietin interferes with B cell function and the mechanisms of this effect. Antibody production by cultured peripheral blood mononuclear cells (PBMC) (7 days) from 15 dialysis patients before and during erythropoietin treatment and from 14 healthy controls was followed. IgG and IgA were formed less in the uremic group than in healthy subjects. After 8 weeks of erythropoietin (hematocrit rose from 19 to 31%) basal IgG formation by PBMC rose from 304 +/- 83 to 566 +/- 49 ng/ml (p less than 0.02), while IgA production rose from 380 +/- 121 to 563 +/- 362 ng/ml (p less than 0.01). IgM production, which appeared to be normal in uremia, remained unchanged during erythropoietin treatment. Production of IgG and IgA stimulated by pokeweed-mitogen was subnormal in uremia, but improved under erythropoietin therapy. To establish whether erythropoietin acted by itself or through correction of the renal anemia, healthy PBMC were directly incubated with 2 U/ml of erythropoietin. Under these conditions production of IgG (+19%), IgA (+28%), and IgM (+32%) was enhanced. Taken together these data indicate a direct stimulant effect of erythropoietin on B lymphocytes in end-stage renal failure.     

The role of T cells and the effect of hydrocortisone on interleukin-4-induced IgE synthesis by non-T cells.

The role of T cells for IL-4-induced IgE synthesis by peripheral blood mononuclear cells (PBMC) was investigated. The removal of monocytes from PBMC abolished IL-4-induced IgE synthesis. When PBMC were separated into T and non-T cells, non-T cells alone were not able to secrete significant amounts of IgE in the presence of IL-4. Depending on the separation procedure, the reconstitution of non-T cells with T cells prepared by rosetting did not restore IgE secretion, whereas T cells obtained by the use of anti-CD3 antibodies could co-induce IgE formation. However, when the T cells were first irradiated, large amounts of IgE were produced, which strongly exceeded those found in unseparated PBMC cultures. IL-4-induced IgE synthesis was also obtained in co-cultures of formaldehyde-fixed T cells with non-T cells. Furthermore, not only autologous but also allogeneic T cells, which have been irradiated or fixed, could provide the costimulatory effect on IgE formation by non-T cells in the presence of IL-4. Mitogenically pre-activated T cells, however, were not able to support IgE synthesis. Hydrocortisone (HC) potentiated the IL-4-induced IgE synthesis by PBMC and enabled non-T cells to secrete IgE in the presence of IL-4. Adding both HC and T cells led to a marked synergistic effect on IL-4-induced IgE production. We conclude that monocytes are required for the induction of IgE synthesis in PBMC in addition to T cells and IL-4. Our results support the view that the T cell signal is delivered via cognate and non-cognate T/B cell membrane interaction. Furthermore, active and proliferating T cells rather suppress IgE synthesis. Finally, HC appears to be a potent alternative stimulus, which bypasses the necessity for T cells in IL-4-induced IgE formation.     

Green tea (Camelia sinensis) mediated suppression of IgE production by peripheral blood mononuclear cells of allergic asthmatic humans

Recent studies in our laboratory demonstrated the suppression of immunoglobulin E (IgE) production by green tea extract (GTE) in U266 cells. However, the effects of GTE or one of its components (EGCG) on IgE production by human peripheral blood mononuclear cells (PBMC) are unknown. PBMC (1.5 × 10⁶) obtained from serum IgE+, allergic asthmatic patients, were cultured ± GTE (1–100 ng/ml) or purified EGCG (0.5–50 ng/ml), and IgE levels were determined on day 10 by enzyme‐linked immunosorbent assay (ELISA). High levels of IgE were detected in supernatants of the PBMC cultures on day 10. When GTE was included in vitro, IgE production by PBMC was suppressed on day 10, compared with control. Purified EGCG included in vitro also suppressed IgE production, but at lower levels, compared with control. This study demonstrates that GTE and its major catechin, EGCG, have immunoregulatory effects on human IgE responses.     

Role of CD8 T cells in rat IgE responses

The role of the cytokines interleukin-4 and interferon-gamma in the regulation of IgE responses in the mouse and man have focused on the role of CD4 T cells. In the rat, antigen-specific CD8 T cells, generated following inhalation of antigen, have been shown to be capable of suppressing IgE responses. Repeated intraperitoneal injections of 1 ng ricin and 1 microgram antigen established a long-lived IgE response in both low- and high-IgE responder rat strains (Wistar and Brown Norway). The duration of the IgE antibody response was 204 and 248 days, respectively. Total IgE levels rose from 30 +/- 20 to 39,000 +/- 7,500 ng/ml in the Wistar rat and from 120 +/- 100 to 47,000 +/- 8,000 ng/ml in the Brown Norway rat. An even greater (10(4)-fold) increase was seen in antigen-specific IgE antibody levels. Ricin alone had no effect and concomitant or prior stimulation with antigen was required. The proportion of CD4+ and CD8+ cells present in the spleen at the peak of the IgE response was markedly increased compared with animals given ricin or antigen alone. Furthermore, CD8 T cells were approximately 100 times more sensitive to ricin than CD4 T cells. These data suggest that enhancement of IgE responses in ricin-treated animals results from the selective deletion of T cells which suppress IgE and are of the CD8 phenotype.     

In vitro production of IgE-binding factors by human mononuclear cells.

This study documents the production of IgE-binding factors (IgE-BFs) by unstimulated and by mitogen-activated human mononuclear cells. IgE-BFs were detected by a sensitive radioimmunoassay employing monoclonal antibodies to lymphocyte Fc epsilon R (MabER). IgE-BFs were found in the 24-hr CSN of unfractionated tonsillar lymphocytes and of their B-cell but not of their T-cell enriched fractions. When cultured for 1 week, PBMC spontaneously synthesized and released IgE-BFs in the CSN; this was significantly reduced by IgE (10 micrograms/ml). PWM, PHA and Con A significantly increased the production of IgE-BFs by PBMC, and this was not influenced by IgE. The production of IgE-BFs in response to mitogens required interactions between T and non-T cells, and IgE-BFs seemed to be derived mainly from non-T cells. However, low levels of IgE-BFs could be detected in the CSN of highly purified T cells cultured for 1 week in the presence of PHA. The production of IgE-BFs by non-T cells was T-cell dependent and it was mediated by soluble factors released from mitogen-activated T cells. T-cell factors increased the secretion of IgE-BFs by: the macrophage cell line U937, adherent cells, and adherent cell-depleted B-cell preparations. It is concluded that the majority of IgE-BFs produced by cultured human mononuclear cells are derived from B cells and monocytes, and that their production is regulated by T lymphocytes.