FcR epsilon+ lymphocytes and regulation of the IgE antibody system. V. Preliminary physicochemical characterization of the T cell-selective IgE-induced regulant EIRT

We previously showed that murine lymphoid cells exposed to elevated levels of IgE exhibit the de novo expression of Fc receptors for IgE (FcR epsilon), and the production of soluble mediators, which we have termed IgE-induced regulants (EIR). Described herein is the preliminary physicochemical characterization of one such regulant, that being the EIR responsible for the Lyt-2+ T cell-dependent expression of FcR epsilon and secretion of an IgE-binding factor (IgE-BF) which can potentiate IgE synthesis; the former activity has been denoted EIRT for its selectivity of action on T cells, and the latter activity has been termed enhancing effector molecule (EEM) for its presumed potentiating influence on IgE antibody synthesis. Characterized in parallel was the conventional lymphoid cell-derived cEIRT and a murine monoclonal T cell hybridoma-(MBI-2)-derived mcEIRT. EIRT from either source was shown to exhibit the characteristics of a protein with a molecular mass of 45 to 60 kd. Once enriched by gel filtration, neither cEIRT nor mcEIRT preparations displayed any other EIR-like activity except that of EIRT, as evidenced by the ability of these preparations to act selectively to induce the Lyt-2 T cell-dependent expression of FcR epsilon and the production of EEM, the lack of detectable SFA activity that could induce Lyt-1+ T cells to produce the IgE-BF denoted suppressive effector molecule (SEM), and the lack of detectable levels of the B cell-selective EIRB, as indicated by the incapacity of either preparation to induce B cell FcR epsilon expression. Neither cEIRT nor mcEIRT displayed IgE-binding affinity, in contrast to the EEM produced in response to stimulation with these regulants. The only EIR-like activity detected in the unfractionated supernatant fluid from cultures of the monoclonal T cell hybridoma MBI-2 was that of EIRT. Careful in vitro analysis established that such preparations did not contain enhancing factor of allergy (EFA), SFA, EIRB, or IgE-BF. Thus, the enhancement of IgE synthesis observed in animals given this mcEIRT preparation was most likely due to the activity of EIRT known to be present. During the course of these studies, clues as to the physicochemical nature of other EIR activities was obtained. Thus, upon molecular sieve analysis, two distinct molecular mass species of EIRB (one 15 to 20 kd and the other 30 to 35 kd) were demonstrated to be present in conventional lymphocyte-derived cEIR.(ABSTRACT TRUNCATED AT 400 WORDS)     

Fc receptors for IgE (Fc epsilon R) on human lymphoid cells: inducible expression of Fc epsilon RII (CD23) on lymphocytes and detection by monoclonal anti-Fc epsilon RII antibody

The studies presented herein describe (1) a sensitive, quantitative, and objective assay for detecting cell membrane-bound form of Fc receptors for IgE displayed on human lymphoid cells based on measuring unlabeled Fc epsilon R-bound IgE by a solid-phase RIA of cell lysate fluids; (2) the development and characterization of an IgM monoclonal antibody, termed 7E4, which is specific for human lymphocyte Fc epsilon RII (CD23) molecules; and (3) a system for reproducibly inducing de novo synthesis and expression of Fc epsilon RII proteins on human lymphocytes following exposure to the mitogenic lectin, pokeweed mitogen. The Fc epsilon RII molecules induced by exposure to PWM were proven to be present on lymphocytes, and not on other cell types in several ways, including (1) documenting sensitivity of such proteins to both acid pH and trypsin treatment, the latter manipulation being ineffective in removing Fc epsilon RII molecules on basophils and mast cells; (2) demonstrating specific reactivity of the expressed Fc epsilon RII molecules with the 7E4 monoclonal antibody, which is specific for human lymphocyte Fc epsilon RII molecules and does not react with Fc epsilon R molecules on other cell types; and (3) observing the required concomitant presence of both T and B lymphocytes during the induction process and proving that the induced Fc epsilon R+ cells are indeed B cells of the Leu-12+ phenotype by fluorescence analysis. The ability to induce expression of Fc epsilon RII molecules on human lymphocytes exposed to a mitogen such as PWM requires special technical attention to the method of preparation and isolation of human lymphoid cells from peripheral blood. This in vitro system for up-regulating Fc epsilon RII expression on human lymphocytes should provide us with an important new tool to analyze the participation of such cells in the regulatory mechanisms controlling the human IgE antibody system.     

Thymus-dependent in vivo suppression of IgE synthesis in a murine IgE-secreting hybridoma

In previous studies we demonstrated that BALB/c mice bearing ascitic tumors of the IgE-secreting hybridoma B53 (epsilon, kappa, anti-dinitrophenyl) developed large numbers of Lyt-1-2+ Fc epsilon R(+) T lymphocytes (T cells with membrane Fc receptors) in response to the elevated serum IgE concentration. The development of Fc epsilon R(+) T lymphocytes was followed by a progressive decrease in the levels of serum IgE in spite of continued proliferation of the hybridoma cells. This sequence of events suggested that the IgE-secreting hybridoma triggered a suppressive immunoregulatory circuit of the host that inhibited IgE expression by the hybridoma cells. The present studies were undertaken to investigate the basis for the subsequent decline in serum IgE levels in mice with B53 tumors and to identify host factors that might be involved in this process. We observed that ascitic B53 cells recovered at increasing time points from BALB/c mice exhibited a selective decline in steady state levels and rates of synthesis of epsilon-heavy chain protein and mRNA. The expression of kappa-light chain protein and mRNA appeared relatively unchanged. The decrease in epsilon-heavy chain gene expression did not occur when B53 tumors were passaged in nu+/nu+ mice or in BALB/c mice depleted of Lyt-2+ cells (suppressor/cytotoxic cell lineage), but did occur in nu+/nu+ mice reconstituted with neonatal BALB/c thymus and in BALB/c mice depleted of L3T4+ cells (helper/inducer cell lineage). That Fc epsilon R(+) T lymphocytes were directly involved in the inhibition of IgE expression was supported by the earlier and more pronounced inhibition of B53 IgE in mice infused with Fc epsilon R(+) T lymphocytes. We conclude from these findings that: 1) the decline in serum IgE levels that occurs toward the end of each generation of in vivo passage of the B53 hybridoma is due to decreased production of IgE by the hybridoma cells, 2) the decreased production of IgE is due to a selective loss of epsilon mRNA expression, 3) the decrease production of IgE by B53 cells is dependent on the presence of Lyt-2+ cells, and 4) Fc epsilon R(+) T lymphocytes participate in the mechanism by which IgE production is suppressed.     

In vivo and in vitro regulation of IgE production in murine hybridomas

Normal BALB/c mice injected i.p. with the IgE-secreting hybridomas B53 (epsilon, kappa anti-DNP), SE1.3 (epsilon, kappa, anti-arsonate) or A3B1 (epsilon, kappa, anti-TNP) were monitored for serum IgE concentrations and frequencies of splenic T lymphocytes with surface membrane receptors for the Fc portion of IgE (Fc epsilon R+ T lymphocytes). Mice with B53 or SE1.3 hybridomas initially developed high concentrations of IgE and CD8+ Fc epsilon R+ T lymphocytes, followed by a progressive decline in both serum IgE and expression of cytoplasmic epsilon-chains in the hybridoma cells. Serum IgE concentrations in mice with A3B1 hybridomas progressively increased without development of Fc epsilon R+ T lymphocytes nor a subsequent decline in IgE or change in cytoplasmic epsilon-chain expression in the A3B1 cells. An in vitro system in which the IgE-secreting hybridoma cells were cocultured with spleen cells harvested from mice with established B53 tumors was used to investigate the mechanisms involved in the inhibition of IgE production by the hybridoma cells. The results of these studies indicate that: 1) the induction/upregulation of Fc epsilon R on CD8+ T lymphocytes in vivo requires factors in addition to high serum IgE concentrations; 2) in addition to CD8+ Fc epsilon R+ T lymphocytes and monocytes, another, as yet unidentified, splenic cell component appears to contribute to the process by which epsilon-chain expression in IgE-secreting hybridoma cells is suppressed, and 3) a hybridoma (A3B1) that fails to induce CD8+, Fc epsilon R+ T lymphocytes in vivo and is not inhibited in IgE expression in vivo, nonetheless is inhibited in IgE expression in vitro when cocultured with spleen cells from mice with B53 tumors.     

The expression of B cell surface receptors. I. The ontogeny and distribution of the murine B cell IgE Fc receptor

The ontogenic appearance and lymphoid tissue distribution of the murine B cell IgE FcR (Fc epsilon R) was examined. Flow cytometry was utilized to study the expression of the Fc epsilon R on splenic B cells from mice of increasing age, as well as B cells from various lymphoid organs. A large panel of B cell tumors was also screened for the presence of the Fc epsilon R. The results demonstrate that the Fc epsilon R appears very late in B cell development, and is preceded in appearance even by IgD. In adult animals, the Fc epsilon R was found to be expressed on virtually all mature IgM, IgD bearing B cells, whether taken from the spleen, lymph nodes, or Peyer's patches. Further examination showed that B cells which had switched to express an isotype other than IgD, appeared to no longer display the Fc epsilon R. When surveying a variety of B cell tumors, the Fc epsilon R was found to be present on WEHI 279, an IgM, IgD-bearing lymphoma. The receptor was not found on pre-B cell, immature B cell, switched B cell, or secreting B cell tumors. Taken together, these results indicate that the B cell Fc epsilon R is expressed predominantly on mature, virgin B cells, and is lost after activation and switching.     

Mechanisms of IgE homeostasis. Sequestration of IgE by murine type II IgE Fc receptor-bearing B cell hybridomas.

Among all classes of Ig, IgE exhibits the highest rate of fractional catabolism of which the site and mechanisms is not understood. We construct a panel of murine B cell hybridomas to investigate the catabolism of IgE; one of these hybridomas, 17A11, constitutively expresses high levels of type II IgE FcR (Fc epsilon RII, CD23) (Kd:1.77 nM; B max: 1.65 x 10(5], and is capable of clearing receptor-bound IgE. Receptor-mediated endocytosis of IgE ligand ensues after binding monomeric and DNP-BSA:IgE immune complexes, and the binding is inhibited by treating 17A11 with anti-CD23. IgE ligands are sequestered and are not susceptible to acid stripping from the cell surface. The internalized IgE ligands redistributed into acid hydrolase containing high density lysosomal vesicles and were degraded; metabolic inhibitors such as chloroquine and monensin that elevate intracellular pH of 17A11 also prevent entry of IgE ligand into lysosomes. These observations raise the possibility that normal Fc epsilon RII-bearing mature B cells in the circulation and lymphoid tissues may function in sequestration and catabolic turnover of IgE molecules through IgE or IL-4 up-regulated Fc epsilon RII uptake; B cell Fc epsilon RII may perform an important role in determining the short biological half-life of IgE molecules, and contributes to IgE homeostasis.     

Superinduction of the murine B cell Fc epsilon RII by T helper cell clones. Role of IL-4

Th cell clones are known to induce an IL-4 dependent polyclonal IgE synthesis. Because IL-4 can induce the expression of the low affinity FcR for IgE (Fc epsilon RII) the ability of Th cell clones to induce Fc epsilon RII on purified splenic B cells was analyzed. It was found that a TH2 clone could cause a 50- to 100-fold superinduction of Fc epsilon RII after 2 days in culture; after 3 days, the Fc epsilon RII levels had almost returned to base line. The superinduction was inhibited by an anti-IL-4 antibody, 11B11, indicating its dependence on IL-4. A TH1 clone could cause a modest (four fold) induction of Fc epsilon RII, and this induction was not influenced by 11B11. A similar Fc epsilon RII induction was seen when using the supernatant from activated TH1 cells. The component(s) causing this relatively low level Fc epsilon RII induction is not known; a variety of known lymphokines were tested, and only IL-4 demonstrated any capacity for Fc epsilon RII induction on LPS-activated B cells. Addition of rIL-4 at concentrations of 400 U/ml or greater to the TH1 culture was sufficient to cause a Fc epsilon RII superinduction similar to that seen with the TH2 clone, while 40 U/ml was not. In order to determine a potential role for the Fc epsilon RII or its soluble fragment on the IgE synthesis mediated by TH2, a monoclonal anti-Fc epsilon RII, B3B4, was added to the culture. The addition of B3B4 did not have an influence on IgE levels in this system.     

Human Fc epsilon RIalpha-specific human single-chain Fv (scFv) antibody with antagonistic activity toward IgE/Fc epsilon RIalpha-binding

The alpha-chain of Fc epsilon RI (Fc epsilon RIalpha) plays a critical role in the binding of IgE to Fc epsilon RI. A fully human antibody interfering with this interaction may be useful for the prevention of IgE-mediated allergic diseases. Here, we describe the successful isolation of a human single-chain Fv antibody specific to human Fc epsilon RIalpha using human antibody phage display libraries. Using the non-immune phage antibody libraries constructed from peripheral blood lymphocyte cDNA from 20 healthy subjects, we isolated three phage clones (designated as FcR epsilon 27, FcR epsilon 51, and FcR epsilon 70) through two rounds of biopanning selection. The purified soluble scFv, FcR epsilon 51, inhibited the binding of IgE to recombinant Fc epsilon RIalpha, although both FcR epsilon 27 and FcR epsilon 70 showed fine binding specificity to Fc epsilon RIalpha. Since FcR epsilon 51 was determined to be a monomer by HPLC, BIAcore analysis was performed. The dissociation constant of FcR epsilon 51 to Fc epsilon RIalpha was estimated to be 20 nM, i.e., fortyfold lower than that of IgE binding to Fc epsilon RIalpha (K(d) = 0.5 nM). With these characteristics, FcR epsilon 51 exhibited inhibitory activity on the release of histamine from passively sensitized human peripheral blood mononuclear cells.     

Monoclonal anti-Fc epsilon receptor antibodies with different specificities and studies on the expression of Fc epsilon receptors on human B and T cells

Three monoclonal antibodies, 1-7 (gamma 2b), 3-5 (gamma 1), and 8-30 (mu), specific to Fc epsilon receptors (Fc epsilon R) on human B cells were established. The two monoclonals (1-7 and 8-30) could inhibit the binding of IgE to Fc epsilon R in rosette formation assays, as well as FACS analysis, and were shown to recognize the same epitope of Fc epsilon R. The other monoclonal antibody (3-5) recognized the same molecule but a different epitope, and marginally inhibited the IgE binding. The molecules on RPMI 8866 cells recognized by these monoclonal antibodies had Mr of 46,000 and 25,000 to 30,000 daltons as determined by immunoprecipitation and SDS-PAGE analysis. By employing these monoclonal antibodies, the expression of Fc epsilon R on circulating lymphocytes was studied. Approximately 50% of B cells from normal, nonatopic individuals were found to express Fc epsilon R, and a remarkable increase in the expression of Fc epsilon R was observed in B cells of atopic patients. The expression of Fc epsilon R was not detected in T cells from atopic patients (including hyper IgE syndrome) as well as normal individuals. Incubation of B cells with PHA-conditioned medium plus IgE augmented the expression of Fc epsilon R in the Fc epsilon R+ B cell population but not in Fc epsilon R- population. PHA-conditioned medium plus IgE did not induce Fc epsilon R expression on T cells.     

Possible role of human lymphocyte receptor for IgE (CD23) or its soluble fragments in the in vitro synthesis of human IgE

The present study demonstrates that human rIL-4 is capable of inducing the secretion of IgE by PBMC. At a concentration of 200 U/ml, an IgE response was observed in 11/26 cultures of PBMC from normal donors and in 12/15 cultures from allergic individuals. The same rIL-4-stimulated cells released significant amounts of IgE-binding factors (IgE-BF) in their culture supernatant. These IgE-BF were shown for the first time to bind simultaneously to some mAb against Fc epsilon R II (mAbER) and to soluble IgE. The lack of correlation between the rIL-4-induced secretion of IgE-BF and IgE indicates that the production of IgE-BF or the expression of Fc epsilon R II is not the only factor involved in the induction of IgE synthesis by rIL-4. However, the observation that mAbER suppressed the rIL-4-induced IgE synthesis strongly suggests that either Fc epsilon R II or IgE-BF are necessary for an IgE response. Finally, the spontaneous in vitro synthesis of IgE by enriched B cell preparations isolated from atopic donors was suppressed in an isotype-specific manner by the same mAbER or by their F(ab')2 fragments. These observations suggest that the ongoing IgE synthesis by in vivo pre-activated B cells is also regulated by IgE-BF or Fc epsilon R II. It is concluded that Fc epsilon R II or IgE-BF play an essential role both in the induction of IgE synthesis by normal B cells and in the ongoing IgE synthesis by in vivo activated B cells from allergic donors.     

The binding of IgE to murine Fc epsilon RII is calcium-dependent but not inhibited by carbohydrate

Despite extensive study, little is known about the functions of the moderate affinity IgE receptors (Fc epsilon RII) on B cells. Recent cDNA and genomic cloning studies have demonstrated that, in contrast to other FcR, Fc epsilon RII is not a member of the Ig gene superfamily. Moreover, it uniquely expresses a region that is highly homologous with a membrane-associated, calcium-dependent binding lectin, the asialoglycoprotein receptor. We now report that the interaction between IgE and the Fc epsilon RII of murine B cells and macrophages requires calcium. Furthermore, as might be expected of asialoglycoprotein lectins, this binding was pH-dependent and resulted in ligand internalization. However, although 125I-Fc epsilon RII bound in a calcium-dependent manner to monosaccharide-agarose beads, high concentrations of mono- and disaccharides did not inhibit the interaction between either 125I-IgE and intact B cells or 125I-Fc epsilon RII (from surface-labeled B cells) and IgE-Sepharose. These results suggest that although murine Fc epsilon RII is a lectin, it is not strictly dependent upon IgE oligosaccharides for its binding to IgE.     

IFN-alpha and IFN-gamma have different regulatory effects on IL-4-induced membrane expression of Fc epsilon RIIb and release of soluble Fc epsilon RIIb by human monocytes

We used highly purified human monocytes to study the regulation of cell surface and secretion of the low affinity FcR for IgE (Fc epsilon RIIb). IL-4 induces Fc epsilon RIIb expression and soluble Fc epsilon RIIb release in a dose-dependent manner. Significant levels of Fc epsilon RIIb expression were obtained after 12 h of incubation with IL-4 and maximal expression was observed between 24 to 48 h after which the expression declined. Surface expression was followed by secretion of soluble Fc epsilon RIIb which reached maximal levels after 3 to 4 days of incubation and which remained constant throughout 7 days of culture. Induction of Fc epsilon RIIb expression by IL-4 was completely blocked by anti-IL-4 antibodies. Furthermore, IL-1 alpha, IL-2, IL-5, granulocyte-macrophage-CSF, IFN-alpha, IFN-gamma, low m.w. BCGF and also LPS all failed to induce Fc epsilon RIIb expression, demonstrating the specificity of the induction. Fc epsilon RIIb membrane expression induced by IL-4 was reduced in the presence of IFN-gamma and IFN-alpha. Strong inhibition of IL-4-induced Fc epsilon RIIb expression was observed at IFN-alpha concentrations of 450 U/ml (80%), and 100 U/ml of IFN-gamma reduced IL-4-induced Fc epsilon RIIb expression by 70%. Interestingly, soluble Fc epsilon RIIb release was strongly inhibited by IFN-alpha. In contrast, IFN-gamma did not affect soluble Fc epsilon RIIb release, suggesting that reduced membrane expression of Fc epsilon RIIb observed in the presence of IFN-gamma does not reflect inhibition of Fc epsilon RIIb expression but may represent enhanced cleavage or reduced anchoring in the membrane of Fc epsilon RIIb. Finally, IL-5 that has been shown to enhance IL-4-induced Fc epsilon RII on B cells does not enhance significantly IL-4-induced Fc epsilon RIIb membrane expression or subsequent soluble Fc epsilon RIIb release by monocytes. Taken together these results show that IFN-alpha and IFN-gamma have different regulatory effects on IL-4-induced Fc epsilon RIIb membrane expression and soluble Fc epsilon RIIb release by human monocytes.     

The murine lymphocyte receptor for IgE. IV. The mechanism of ligand-specific receptor upregulation on B cells

Rodent B cells respond to culture with IgE by increasing their IgE-specific Fc receptors (Fc epsilon R). The mechanism of this upregulation was characterized on Fc epsilon R+ murine B cell hybridoma lines. Measurements of [35S]methionine incorporated into the Fc epsilon R over time indicated that IgE did not appreciably increase the rate of Fc epsilon R synthesis. In contrast analysis of Fc epsilon R decay from surface radioiodinated B hybridoma cells demonstrated that IgE acted to slow the rate of Fc epsilon R degradation. Very little endocytosis of monomeric IgE was seen; this, combined with the observation that lysomotropic agents failed to inhibit Fc epsilon R degradation suggested that decay occurs at the cell surface. A soluble receptor immunoassay was developed, using monoclonal anti-Fc epsilon R, and this assay demonstrated that cell-bound IgE inhibited the release into the culture media of soluble immunoreactive Fc epsilon R. Examination of the soluble Fc epsilon R by SDS-PAGE after isolation with monoclonal anti-Fc epsilon R demonstrated that it was 10,000 m.w. smaller than the cell-associated Fc epsilon R. IgE affinity columns failed to bind the Fc epsilon R fragment, indicating that the ligand binding activity was largely lost. Thus this study demonstrated that IgE-dependent Fc epsilon R induction on B cells occurs because IgE upon binding to the B cell surface, inhibits the proteolytic cleavage and release of the Fc epsilon R into the surrounding medium, and it is this inhibition of degradation that causes the higher Fc epsilon R levels.     

B lymphocyte regulation of the immune system. II. Inhibition of Fc receptor expression of lymphocytes by BEF, a lymphokine of B cell origin

Recently, we described a new lymphokine of B cell origin, capable of selectively preventing the differentiation of T suppressor cells from the precursor into the effector stage. As a result, antibody production against various antigens is markedly increased. We termed this lymphokine B cell-derived enhancing factor (BEF). To discern the mechanism(s) by which BEF interferes with the activation of T suppressor cells, experiments were undertaken to explore the effect of BEF on the induction of Fc receptors (FcR). The induction of FcR on T cells has been implicated in the down-regulation of antibody synthesis, and it has been suggested that the expression of FcR for a given immunoglobulin precedes the release of factors with regulatory functions for the corresponding isotype. In the experiments reported here, murine spleen cells were incubated for 24 hr in the presence of IgG1 or IgA monoclonal antibodies, were washed, and the number of FcR gamma 1+ and FcR alpha+ cells were calculated by a rosette assay. The effect of BEF was studied either during the inductive phase or before, i.e., by pretreating the cells with BEF for 18 hr at 37 degrees C before the inductive phase. Our results show that BEF abolishes, in a dose-dependent manner, the expression of isotype-specific FcR in spleen cells when present during the inductive phase, as well as when cells are pretreated with it. In successive experiments, we tested the effect of BEF on the induction of FcR on T cell-enriched or B cell-enriched spleen cells. The results show that BEF is effective in selectively inhibiting FcR expression on T lymphocytes, but not on B lymphocytes, once isolated from the total spleen cell population. These findings provide further insight into the mechanism by which BEF modulates the immune response, and suggest that different mechanisms may be involved in the induction of FcR on T and B lymphocytes, respectively.     

The murine epsilon receptor modulating protein: a novel serine protease which modulates CD23 binding of IgE.

In our recent previous studies, we have identified and purified a murine 17-kDa protein which diminishes the avidity of binding between IgE and CD23 (low-affinity Fc receptor for IgE) without decreasing the quantitative expression of the CD23. The protein was thus designated epsilon receptor modulating protein (epsilon RMP). In this study, we have further characterized this protein and have found that (i) epsilon RMP is inactivated by phenylmethylsulfonyl fluoride and decomposes N,alpha-benzyloxycarbonyl-L-lysine thiobenzyl ester, as well as N-succinyl-Ala-Ala-Pro-Phe p-nitroanilide; (ii) epsilon RMP does not work directly on B cells but requires CD4+ T cells to decrease functional expression of CD23 on B cells; and (iii) the partial internal amino acid sequence of epsilon RMP, obtained by using in situ cyanogen bromide cleavage on polyvinylidene difluoride membrane is unique. These data thus clearly demonstrate that epsilon RMP is a novel serine protease controlling the functional expression of CD23 through the participation of CD4+ T cells. Mechanisms of the involvement of CD4+ T cells are discussed.     

Characterization with monoclonal antibodies of T lymphocytes bearing Fc receptors for IgE (T epsilon cells) and IgG (T gamma cells) in atopic patients

Peripheral blood T lymphocytes from nonatopic control donors, asymptomatic atopic donors, and patients with moderate to severe atopic dermatitis were analyzed for Fc receptors for IgE (T epsilon cells) and IgG (T gamma cells) by rosette assays and were characterized with monoclonal antibodies. The T cells were reacted first with monoclonal antibodies, followed by fluoresceinated F(ab')2 goat antimouse Ig; they were then rosetted, and subsequently the rosetting cells were examined for immunofluorescence. Seven nonatopic control donors had less than 0.1% T epsilon cells and a mean +/- SD of 10.5% +/- 4.1 T gamma cells. Seven asymptomatic atopic donors with low IgE levels (2 to 233 IU/ml) varied from less than 0.1 to 1.3% T epsilon cells and 7.2% +/- 3.7 T gamma cells. Six of seven patients with moderate to severe atopic dermatitis and IgE levels of 1339 to 24,261 IU/ml had less than 0.1% T epsilon cells and significantly fewer T gamma cells (3.1% +/- 2.7, p less than 0.01) than the nonatopic control donors and the atopic donors in remission. Both T epsilon and T gamma cells reacted with the pan-T cell antibody Lyt-3 (anti-sheep red cell receptor) but not with antibodies OKT3, OKT4, or OKT6. Subpopulations of both T epsilon and T gamma cells reacted with antibodies OKT8 and the antimonocyte antibody OKM1. The OKM1+ cells did not appear to be monocytes, however, because the T cells did not react with another antimonocyte antibody, BRL.2, and were negative for nonspecific esterase activity. (ABSTRACT TRUNCATED AT 250 WORDS)     

Increased T gamma and T mu cells in BALB/c mice with IgG and IgM plasmacytomas and hybridomas

The results of previous studies in our laboratory have shown that mice bearing plasmacytomas and hybridomas that secrete IgA or IgE are accompanied by increased frequencies of Lyt-1-2+ T lymphocytes bearing Fc receptors (FcR) for IgA (T alpha) or IgE (T epsilon), respectively. The present study was undertaken to examine whether IgG- or IgM-secreting tumors influenced the frequency of T lymphocytes that express FcR for IgG or IgM. We studied mice bearing IgG- and IgM-secreting plasmacytomas and hybridomas. BALB/c mice injected subcutaneously with the IgG-secreting hybridoma HDP1 (gamma 1 kappa, anti-TNP) were sequentially examined for the frequencies and Lyt phenotypes of splenic lymphocytes bearing FcR for IgG (T gamma), IgM (T mu), and IgA (T alpha). A threefold increase in the frequency of T gamma lymphocytes that were Lyt-1-2+, L3T4- was seen. The frequencies of T mu and T alpha lymphocytes in these mice were not significantly altered. Similarly, mice injected subcutaneously with the IgM-secreting plasmacytoma MOPC 104E (mu lambda, anti-dextran) or the IgM-secreting hybridoma C1D1 (mu kappa, anti-ox RBC) were examined sequentially for the frequencies of T gamma, T mu, and T alpha lymphocytes. Mice with established IgM subcutaneous tumors showed a twofold increase in splenic, nylon wool-nonadherent T mu lymphocytes. This was associated with a relative increase in Lyt-2+ splenic T lymphocytes and a relative decrease in Lyt-1+ splenic T lymphocytes. No changes were observed in the frequencies of either T gamma or T alpha lymphocytes. These studies extend to IgG and IgM the observation that plasmacytomas and hybridomas secreting immunoglobulins of a specific isotype cause an expansion of T lymphocytes bearing FcR specific for the corresponding isotype. The expansion of FcR+ Lyt-1-2+ T lymphocytes likely represents an exaggerated, but otherwise normal, immunoregulatory response of the host. These cells may be an important element in the regulation of isotype expression.     

A B cell-specific differentiation antigen, CD23, is a receptor for IgE (Fc epsilon R) on lymphocytes

Two independent L cell transformants expressing human lymphocyte Fc epsilon R were established by using cellular DNA from RPMI 8866 cells. The surface expression of the receptor was confirmed on the basis of the binding of a panel of anti-Fc epsilon R antibodies and its ability to bind IgE. Anti-CD23 antibodies strongly stained the transformants, indicating possible identity or antigenic relationship between Fc epsilon R and CD23. This interesting observation warrants additional clarification as to the role of CD23 and Fc epsilon R in B cell differentiation.