Low-affinity Fc Receptor For IgE Research Articles

FCER2 (CD23) Asthma-Related Single Nucleotide Polymorphisms Yields Increased IgE Binding and Egr-1 Expression in Human B Cells

CD23 is the low-affinity Fc receptor for IgE. When expressed on B cells, CD23 appears to play a role in regulation of IgE synthesis. Polymorphisms within FCER2, the gene encoding CD23, have been associated with atopy, increased risk of exacerbations in patients with asthma, and high serum IgE levels. A single-nucleotide polymorphism (rs2228137) present in exon 4 of FCER2 encodes a nonsynonymous amino acid change (R62W) and is the subject of the present analysis. Human B cell stable transfectants were established to characterize the functional relevance of the R62W SNP. We demonstrate that CD23b-R62W-expressing human B cells bind IgE with greater affinity than wild-type cells and display differences in kinetics of CD23-mediated ERK1/2 activation that may be responsible for the increased levels of Egr-1 mRNA observed after stimulation through CD23. Finally, the R62W SNP seems to alter the tertiary or quaternary structure of CD23 because in the absence of N-glycosylation the CD23b-R62W-expressing cells appear to be less sensitive to endogenous proteases. These observations may have implications in mechanisms responsible for the atopic phenotypes observed in patients with asthma who possess this genotype.

Structural Changes in the Lectin Domain of CD23, the Low-Affinity IgE Receptor, upon Calcium Binding

CD23, the low-affinity receptor for IgE (Fc epsilonRII), regulates IgE synthesis and also mediates IgE-dependent antigen transport and processing. CD23 is a unique Fc receptor belonging to the C-type lectin-like domain superfamily and binds IgE in an unusual, non-lectin-like manner, requiring calcium but not carbohydrate. We have solved the high-resolution crystal structures of the human CD23 lectin domain in the presence and absence of Ca2+. The crystal structures differ significantly from a previously determined NMR structure and show that calcium binding occurs at the principal binding site, but not at an auxiliary site that appears to be absent in human CD23. Conformational differences between the apo and Ca2+ bound structures suggest how IgE-Fc binding can be both calcium-dependent and carbohydrate-independent.

Epidemiology, clinical features, and immunology of the "intrinsic" (non-IgE-mediated) type of atopic dermatitis (constitutional dermatitis).

Departmentof Pharmacology, University of Bern, Bern, SwitzerlandKey words: atopic dermatitis; clinical features;constitutional dermatitis; epidemiology; extrinsictype; immunopathology; intrinsic type.Prof. Dr B. Wu¨thrichAllergy Unit, Department of DermatologyUniversity Hospital8091 ZurichSwitzerlandAccepted for publication 22 March 2001

CD23 expression in activated human T cells is enhanced by interleukin-7.

Despite evidence for the expression of the low-affinity Fc receptor for IgE (Fc epsilon RII/CD23) in several pathological conditions, the role played by CD23 in normal human T cells is still unclear. We studied the effect of a stomal-derived cytokine, interleukin (IL-7), on the expression of CD23 in human T cells stimulated with 10 micrograms/ml phytohemagglutinin (PHA). The results demonstrate that IL-7 did not induce CD23 expression in the resting T cells. However, PHA-induced CD23 expression was enhanced by costimulation with IL-7 (1,000 U/ml). Cytofluorometric analysis revealed that CD23 expression in activated T cells was enhanced by the addition of IL-7 (from 2% to 18%). It was also observed that the effect of IL-7 on CD23 expression is exclusively on CD4+ T cells. The enhanced expression of CD23 was blocked by an anti-IL-7 monoclonal antibody (mAb), but not by IL-2 and IL-4 mAbs. This suggests that IL-7 is a potent regulatory cytokine capable of acting independently of IL-2 and IL-4 in the expression of CD23. Northern blot analysis showed an increase in CD23 mRNA when activated T cells were cultured in the presence of IL-7. A significant increase in receptor numbers on activated T cells was detected by Scatchard analysis when IL-7 was added to the cell cultures. The induction of CD23 expression by IL-1, IL-3, IL-4, IL-5, IL-6, interferon-8 and OKT3 on PHA-activated T cells was not of the same magnitude as observed in the presence of IL-7. These results demonstrate a selective induction of CD23 expression on activated human T cells cultured in the presence of IL-7. These data indicate that the stromal-derived growth factor points to an important role of CD23 in the regulatory network of the immune response.

Plasma cell tumors decrease CD23 mRNA expression in vivo in murine splenic B cells

CD23, the low-affinity Fc receptor for IgE, is constitutively expressed on mature, naive B cells, but is lost following B cell activation. We and others have shown that CD23 expression on B cells decreases in mice bearing plasma cell tumors. In contrast to these findings, we find that IgE-secreting tumors do not cause a loss of CD23 expression on host splenic B cells. Decreased expression of CD23 on B cells induced by plasma cell tumors requires direct contact between tumor cells and B cells; and other host cells are not involved. The loss of surface CD23 expression is associated with a decrease in steady-state CD23 mRNA levels in B cells from plasma cell tumor-bearing mice. Interestingly, loss of CD23 mRNA is observed even in B cells from mice with IgE-secreting tumors, where we find surface CD23 protein expression to be similar to that of normal mice. The maintenance of surface CD23 expression on B cells in mice with IgE-secreting tumors is dependent solely on the presence of IgE, and is not a tumor-specific effect. Therefore, we conclude that in vivo, IgE secreted by plasma cell tumors can result in the maintenance of normal levels of surface CD23 expression by a post-transcriptional mechanism, even when the tumor induces a down-regulation of CD23 mRNA levels.

Functional significance of CD23- on CD23-transfected Th2 clone

CD23, a low-affinity IgE Fc receptor, is not displayed on most resting T cells but its expression has been shown to be transiently induced in vivo and in vitro on some CD4 + T cells [1–4] and in vivo on CD8 + T cells by IgE-secreting hybridoma tumors [5]. To investigate the functional role of CD23 on T cells, we inserted a CD23 construct into an expression vector driven by a CD2 promoter and transfected it into a murine Th2 clone D10.G4.1 (D10). We stimulated the transfected D10 cells (D10.3M.24) with anti-TCR antibody in the presence or absence of IgE, and measured IL-4, IL-5 and IL-6 production in the culture supernatants. Activation of D10.3M.24 cells by anti-TCR antibody induced greater levels of IL-4, IL-5 and IL-6 production, when the TCR and CD23 were co-crosslinked by TNP anti-TCR and IgE anti-TNP antibodies. IgG anti-TNP antibody did not enhance lymphokine production by D10.3M.24 cells. The enhanced lymphokine production by IgE was blocked by monoclonal anti-CD23 antibody. IgE anti-TNP antibody did not enhance lymphokine production by the wild-type D10 cells induced by TNP anti-TCR antibody. These studies show that when co-crosslinked with the TCR, CD23 can modulate the lymphokine production in activated Th2 cells. Since CD23 binds to IgE and also binds to CD21 [6], a complement receptor commonly expressed on B cells, T-cell CD23 could play an immunoregulatory role during cognate T-B cell interaction and during IgE antibody responses.

Identification of novel CD23 transcripts on human T and B lymphocytes and eosinophil cell line

The main aim of the present studies was to investigate the structure of the human low-affinity IgE Fc receptor (CD23) present on T and B lymphoid cells and eosinophil cell line. A novel finding in these studies has been the detection and sequence analysis of CD23 transcripts in human T lymphocytes. These studies have established that some of the human T-cell populations analyzed express CD23 mRNA and that its structure is quite similar to that previously described for human B lymphocytes. A second major finding in these studies is that some human T- and B-cell lines and eosinophil cell line contain multiple forms of CD23 transcripts. These appear to be generated via alternative splicing, resulting in transcripts that may encode a truncated, possibly secretory form of CD23. These findings in human T and B lymphocytes and eosinophils provide new information about the structure of lymphocyte CD23 and suggest that alternative processing of transcripts generates CD23 mRNA that encodes CD23 isoforms. These studies are the first experimental evidence showing that CD23 isoforms may occur in the human and are the first direct evidence for production of CD23 by human T lymphocytes. In addition, these studies provide the first experimental evidence that T and B lymphocytes express CD23 transcripts lacking exon 3-encoded sequences, raising the possibility that a secretory form of CD23 may be synthesized by human T and B lymphocytes, and eosinophils.

Soluble FcϵRII/CD23 in Patients with Autoimmune Diseases and Epstein-Barr Virus-Related Disorders: Analysis by ELISA for Soluble FcϵRII/CD23

The low-affinity Fc receptor for IgE (FcϵRII/CD23) and its soluble form (sCD23, IgE-binding factor) have multiple functions, and enhanced levels of these are associated with various immunological diseases. We established two sensitive ELISA systems using enzyme-conjugated mAb and biotinylated mAb. The detection limits of the ELISA systems were 0.03 and 1.0 ng/ml, which showed good correlation in the range 1.0-10 ng/ml. In the ELISA system using enzyme-conjugated mAb, the average sCD23 concentration in 303 normal healthy volunteers was 1.4 ± 0.3 ng/ml. In the ELISA system using biotinylated mAb, sCD23 levels in normal healthy volunteers showed almost the same values. In patients with autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus, Sjögren syndrome, progressive systemic sclerosis, and mixed connective tissue disease, the sCD23 levels were significantly higher than those in normal individuals. Furthermore, in Epstein-Barr virus-related disorders after liver transplantation with immunosuppression, plasma levels of sCD23 rapidly Increased to more than 12 ng/ml when clinical symptoms were evident. In addition, the sCD23 values remained high, although elevated GOT levels gradually decreased to standard values and EBV hepatitis improved. These data suggest that sCD23 levels are a sensitive marker of autoimmune diseases and EBV-related disorders in addition to allergic disorders. The ELISA system for sCD23 may be an additional diagnostic tool in estimating the clinical courses of these diseases.

Cloning of cDNAs for new subtypes of murine low-affinity Fc receptor for IgE (Fc epsilon RII/CD23).

In humans, two subtypes of the low-affinity Fc receptor for IgE, Fc epsilon RIIa and Fc epsilon RIIb, have been identified. The two forms differ only in a short stretch of amino acids at the cytoplasmic amino terminus. On the other hand, only one Fc epsilon RIIa-like receptor was reported in the mouse, although the existence of another subtype of transcript has been suggested. A new cDNA subtype for murine Fc epsilon RII that has amino acid homology to the human subtype b at the amino terminus was cloned by polymerase chain reaction. When expressed transiently in COS cells, the cDNA is capable of generating a functional cell surface protein that is recognized by an anti-Fc epsilon RII antibody and that has affinity to IgE. In the spleen, although the expression level is considerably lower than that of subtype a, the transcript appears in B cells and in non-B cells upon stimulation with IL-4 and LPS, in contrast to the constitutive expression of the conventional Fc epsilon RII transcript. It is also detectable in several cell lines of B- or T-cell lineages. These results suggest that this gene may be the murine counterpart of human Fc epsilon RII subtype b. Another cDNA clone for the distinct Fc epsilon RII transcript has also been isolated. The cDNA also encodes a surface protein functional on COS cells, but its human counterpart has not been found.

Switching capacity of FcϵRII‐positive and ‐negative murine B cells

In previous studies, our laboratory demonstrated the utility of the low affinity IgE Fc receptor (Fc epsilon RII) in delineating a number of murine B cell subsets. In the spleen, the Fc epsilon RII is expressed on mature conventional B cells but is absent on marginal zone B cells. In the peritoneal cavity, the receptor is present on all conventional B cells, but is not expressed on fresh peritoneal Ly1/sister B cells. The studies in this report compared the ability of these B cell populations to isotype switch. Using a lipopolysaccharide (LPS)- and interleukin (IL)-4-driven system, sort-purified Fc epsilon RII-positive and -negative B cells from peritoneum and spleen were tested for switching to IgG1, IgE, and IgA. The results demonstrated that regardless of their source, Fc epsilon RII+ B cells produced significant levels of IgG1 and IgE. Similar results were obtained with Fc epsilon RII- (marginal zone) B cells obtained from spleen. In contrast, Fc epsilon RII- (Ly1/sister) peritoneal B cells were found to produce IgG1 and IgA, but were incapable of secreting significant levels of IgE. Further studies tested for LPS and IL-4-induced expression of Fc epsilon RII and Thy1 on the various B cell populations. These experiments demonstrated the induction of the Fc epsilon RII on all B cells, regardless of their initial resting levels. Additionally, Thy1 was found to be induced only on those B cell subsets capable of producing IgE. Taken together, the results demonstrate a correlation between IgE secretion and Thy1 expression, and no apparent correlation between the presence of the Fc epsilon RII and isotype commitment.

CD23/Fc epsilon RII expression on phytohemagglutinin-A- or phorbol-12myristate-13acetate-Ca(2+)-activated human tonsil T cells.

The low-affinity Fc receptor for IgE, CD23/Fc epsilon RII, has been expressed in T cell lines and pathologic T cells, but its presence on normal human T cells is still debated. We studied the expression of CD23/Fc epsilon RII on purified T cells from normal peripheral blood mononuclear cells (PBMC) or tonsil T cells stimulated with 10 micrograms/ml phytohemagglutinin A (PHA) or phorbol-12myristate- 13acetate (PMA) Ca2+. Using two-dimensional flow cytometry, the tonsil T-cell-enriched population showed > 10% of CD23/Fc epsilon RII expression when coexpressed with the CD3 antigen. CD4+ T cells appear to be principally involved in the expression of CD23/Fc epsilon RII, although we were unable to detect a clear expression of CD23/Fc epsilon RII in PBMC that were activated with either PHA or PMA Ca2+. PHA stimulation resulted in the release of IgE binding factor (IgEBF). The induction of CD23/Fc epsilon RII expression in PHA- and PMA-Ca(2+)-activated T cells was enhanced by IL-4, but not by IgE or IL-6. IL-4 also augmented the PHA- and PMA-Ca(2+)-induced release of IgEBF. The addition of supernatant from the Epstein-Barr virus (EBV)-infected cell line to PHA- or PMA-Ca(2+)-stimulated tonsil T cells did not increase CD23/Fc epsilon RII expression. The expression of CD23/Fc epsilon RII mRNA was detected in RNA prepared from a tonsil T-cell-enriched population by Northern blot analysis.

CD23 isoforms in murine T and B lymphocytes.

A major objective of the present studies was to seek biochemical evidence for the production of the low affinity IgE Fc receptor (CD23) by murine T cells. These studies have established that most of the murine T cell populations analyzed express CD23 mRNA. Some T cell populations contained a transcript whose nucleotide sequence was identical to that previously reported for the CD23 cDNA cloned from murine B cells. Some T and B cells contained CD23 transcripts lacking exon-3-encoded sequences. If translated, these transcripts would yield a CD23 polypeptide lacking the transmembrane segment and all but six amino acids of the intracytoplasmic tail. The truncated CD23 transcripts detected in T and B cells in these studies appear to be generated via alternative splicing. It is possible that these transcripts encode soluble, secretory forms of CD23, or that these transcripts have regulatory functions that influence CD23 gene expression. These investigations provide new information about the structure of murine lymphocyte CD23, they predict the occurrence of CD23 isoforms in the mouse, and they present direct evidence for the production of CD23 by murine T lymphocytes.

Regulation of the low affinity IgE Fc receptor (CD23) in atopic dermatitis.

Alterations in the production of CD23, the IgE Fc receptor, may play a role in the etiology of atopic dermatitis and other allergic conditions. Interleukin-4 (IL-4), interferon-gamma (IFN-gamma), and interferon-alpha (IFN-alpha) are involved in coordinate regulation of CD23. IL-4 stimulates production of both CD23 and IgE. IFN-gamma also stimulates production of CD23, but suppresses production of IgE and inhibits IL-4-mediated production of CD23. IFN-alpha also suppresses these IL-4-mediated activities and, in addition, suppresses IFN-gamma-mediated stimulation of CD23 production. Changes in this coordinate regulation may be involved in the development of atopic dermatitis. Expression of CD23 by Langerhans cells is stimulated by IL-4 and IFN-gamma. Further, levels of CD23-positive T cells are elevated in atopic dermatitis subjects as compared to non-atopic controls, and observed skin changes appear to be related to changes in CD23-positive mononuclear cell populations. Coordinate regulation of CD23 by IFN-gamma and IL-4 may also be important in other allergic conditions, parasitic infections, and a variety of disease states.

Two forms of the low-affinity Fc receptor for IgE differentially mediate endocytosis and phagocytosis: identification of the critical cytoplasmic domains.

We have previously identified two species of the low-affinity human Fc receptor for IgE, Fc epsilon RIIa and Fc epsilon RIIb, which differ only in a short stretch of amino acids at the N-terminal cytoplasmic end. Their differential expressions on B cells and monocytes suggest that Fc epsilon RIIa and Fc epsilon RIIb are involved in B-cell function and IgE-mediated immunity, respectively. Here we show that Fc epsilon RII-mediated endocytosis is observed only in Fc epsilon RIIa-expressing cells, whereas IgE-dependent phagocytosis is observed only in Fc epsilon RIIb-expressing cells, demonstrating the functional difference between Fc epsilon RIIa and Fc epsilon RIIb. Furthermore, site-directed mutagenesis revealed that the tyrosine residue in the Fc epsilon RIIa-specific region is important for endocytosis, and the Asn-Pro residues in the Fc epsilon RIIb-specific region are required for phagocytosis. These findings suggest that endocytosis and phagocytosis are functionally separable phenomena involving distinct amino acid residues.

The low affinity IgE Fc receptor (CD23) participates in B cell activation.

The low affinity IgE Fc receptor (FcɛRII or CD23) is expressed on all mature B lymphocytes (reviewed in 1 and 2). The FcʵRII is a 45 to 49 KD glycoprotein, and is categorized as a class II transmembrane molecule on account of its inverted membrane orientation. In addition to the transmembrane form of the receptor, a soluble form is readily released from B cells, a result of cleavage by a cell surface protease. cDNA cloning has revealed that the FceRII is a member of the hepatic lectin family, a somewhat unusual finding since all other characterized Fc receptors are known to be members of the immunoglobulin (Ig) gene superfamily.

Fyn tyrosine kinase associated with Fc epsilon RII/CD23: possible multiple roles in lymphocyte activation.

Expression of low-affinity Fc receptor for IgE (Fc epsilon RII), which is identical to the lymphocyte differentiation antigen CD23, is associated with activation of lymphoid cells. The mechanism of signal transduction through Fc epsilon RII/CD23 was dissected by transfection of cDNA coding for Fc epsilon RII to the YT human natural killer-like cell line, activation of which was easily detected by the induction of interleukin 2 receptor/p55(Tac). Cross-linking of Fc epsilon RII/CD23 with H107 anti-Fc epsilon RII monoclonal antibody markedly enhanced interleukin 2 receptor/p55 expression on the YT cells transfected with Fc epsilon RII cDNA (YTSER cells). Similar induction of interleukin 2 receptor/p55 by the cross-linking of Fc epsilon RII was observed on an Epstein-Barr virus-transformed B-cell line, 3B6, and fresh leukemic cells isolated from a patient with B-cell chronic lymphoblastic leukemia. Thus Fc epsilon RII/CD23 provides activation signals not only in YTSER cells but also in activated B cells. A possible involvement of protein-tyrosine kinase in the Fc epsilon RII-mediated signal transduction was studied. Fc epsilon RII was physically associated with a src family tyrosine kinase p59fyn and not with p56lck, which was also found in YT cells. Recently it was reported that p59fyn was associated with T-cell antigen receptor. Our results collectively suggest the multiple functions of p59fyn that may be implicated in Fc epsilon RII-mediated activation signal in YT cells.

The murine ϵ 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 ϵ receptor modulating protein (ϵRMP). In this study, we have further characterized this protein and have found that (i) ϵRMP is inactivated by phenylmethylsulfonyl fluoride and decomposes N,α-benzyloxycarbonyl- l-lysine thiobenzyl ester, as well as N-succinyl-Ala-Ala-Pro-Phe p-nitroanilide; (ii) ϵRMP 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 (ϵRMP, obtained by using in situ cyanogen bromide cleavage on polyvinylidene difluoride membrane is unique. These data thus clearly demonstrate that ϵRMP 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.

High Frequency of IL-4–Producing CD4+ Allergen-Specific T Lymphocytes in Atopic Dermatitis Lesional Skin

In atopic dermatitis (AD) hypersensitivity reactions to allergens are commonly observed and are assumed to make a major contribution in the pathomechanism of the disease. It may be expected that allergen-reactive Th cells play a central role in these reactions. In the present study the occurrence and function of allergen-specific T lymphocytes in dermal inflammatory lesions were studied. To this aim panels of randomly cloned CD4+ T cells from lesional skin biopsies of two housedust mite Dermatophagoides pteronyssinus (Dp)-allergic AD patients were screened for reactivity with Dp allergens. The results were compared with similar tests for Dp reactivity of T-lymphocyte clones (TLC) from the peripheral blood of these patients. In the panels of TLC generated from lesional skin (S-TLC), a considerable number of TLC appeared to be Dp-specific, 47% (n = 17) and 10% (n = 29), respectively. In the panels from the peripheral blood, the percentages of Dp-specific TLC were only 0% (n = 22) and 3% (n = 34), suggesting accumulation or expansion of these T cells in lesional skin. The function of these TLC was studied by assaying the secretion of IL-4 and IFN-gamma, which have been shown to be produced in aberrant ratios by Dp-specific TLC from the peripheral blood of AD patients (Wierenga et al: J Immunol 144:4651, 1990). All Dp-specific S-TLC produced IL-4 in combination with no or low levels of IFN-gamma, whereas many of the non-Dp-specific S-TLC and blood-derived TLC (B-TLC) were observed to produce high levels of IFN-gamma without significant amounts of IL-4. A functional consequence of these cytokine profiles was demonstrated by the finding that TLC producing substantial amounts of IL-4 enhanced expression of the low-affinity Fc receptor for IgE (CD23) on antigen-presenting cells to a greater extent than did IFN-gamma-producing TLC.

Purification of murine suppressive factor of allergy into distinct CD23-modulating and IgE-suppressive proteins.

The murine suppressive factor of allergy (SFA) has been purified from a T-cell hybridoma and found to consist of two functionally and biochemically distinct protein molecules. One protein (17 kDa) modulates the low-affinity Fc receptor for IgE on lymphocytes (i.e., CD23); it decreases the binding avidity of IgE to CD23-bearing B cells without affecting quantitative expression of CD23 and is thus designated epsilon-receptor-modulating protein. The second protein (30 kDa) suppresses IgE biosynthesis (i.e., SFA). This purified SFA suppresses interleukin 4-induced IgE and IgG1 synthesis by lipopolysaccharide-activated spleen cells but has no effect on other antibody isotypes; since the activity of SFA is not blocked by anti-interferon gamma monoclonal antibody, it is thus distinct from interferon gamma. The data presented indicate that epsilon-receptor-modulating protein and SFA are protein molecules that are involved in modulating the CD23 molecule and IgE antibody synthesis, respectively.

CD77: an antigen of germinal center B cells entering apoptosis

We have previously reported that a neutral glycolipid (globotriosylceramide; Gb3) was specifically expressed on Burkitt's lymphoma cells and on a subset of germinal center tonsillar B lymphocytes. Recently the Gb3 molecule was recognized as a new B cell differentiation antigen and now defines the CD77 cluster. Here we report an extensive phenotypic and functional characterization of the tonsillar CD77+ B lymphocytes. These cells have a low buoyant density and are thus purified using a Percoll gradient. They express various B cell antigens such as CD19, CD20, CD21, CD22 and CD40, as well as the adhesion molecules LFA-1, LFA-3 and CD44. They are positive for surface IgM and negative for surface IgD. Although these results suggest a phenotype of activated B cells, the CD77+ cells are negative for the classical activation antigens: CD23 (the low-affinity Fc receptor for IgE), CD25 [the interleukin (IL) 2 receptor alpha chain] and CD71 (the transferrin receptor). Proliferation and protein synthesis of CD77+ cells was measured after stimulation with a range of mitogens and IL. None of the agents tested are able to induce proliferation and protein synthesis with the exception of a combination of recombinant IL 4 plus anti-CD40 antibody. When examined by electron microscopy, CD77+ B lymphocytes present a morphology similar to that of cells undergoing programmed cell death, also called apoptosis (i.e. chromatin condensation, nuclear fragmentation, membrane blebbing). As shown by direct examination of DNA, these CD77+ cells are indeed in the process of apoptosis. Treatment of the CD77+ cells by recombinant IL 4 and anti-CD40 antibody prevents apoptosis. All these results suggest that the CD77 molecule defines a B lymphocyte maturation pathway, specific for germinal center, where the cells undergo programmed cell death.