A comparison of mediators released or generated by IFN-gamma-treated human mast cells following aggregation of Fc gamma RI or Fc epsilon RI

The high affinity receptor for IgG (Fc gamma RI, CD64) is expressed on human mast cells, where it is up-regulated by IFN-gamma and, thus, may allow mast cells to be recruited through IgG-dependent mechanisms in IFN-gamma-rich tissue inflammation. However, the mediators produced by human mast cells after aggregation of Fc gamma RI are incompletely described, and it is unknown whether these mediators are distinct from those produced after activation of human mast cells via Fc epsilon RI. Thus, we investigated the release of histamine and arachidonic acid metabolites and examined the chemokine and cytokine mRNA profiles of IFN-gamma-treated cultured human mast cells after Fc gamma RI or Fc epsilon RI aggregation. Aggregation of Fc gamma RI resulted in histamine release and PGD(2) and LTC(4) generation. These responses were qualitatively indistinguishable from responses stimulated via Fc epsilon RI. Aggregation of Fc epsilon RI or Fc gamma RI led to an induction or accumulation of 22 cytokine and chemokine mRNAs. Among them, seven cytokines (TNF-alpha, IL-1beta, IL-5, IL-6, IL-13, IL-1R antagonist, and GM-CSF) were significantly up-regulated via aggregation of Fc gamma RI compared with Fc epsilon RI. TNF-alpha mRNA data were confirmed by quantitative RT-PCR and ELISA. Furthermore, we confirmed histamine and TNF-alpha data using IFN-gamma-treated purified human lung mast cells. Thus, aggregation of Fc gamma RI on mast cells led to up-regulation and/or release of three important classes of mediators: biogenic amines, lipid mediators, and cytokines. Some cytokines, such as TNF-alpha, were released and generated to a greater degree after Fc gamma RI aggregation, suggesting that selected biologic responses of mast cells may be preferentially generated through Fc gamma RI in an IFN-gamma-rich environment.     

Cytokine production by skin-derived mast cells: endogenous proteases are responsible for degradation of cytokines

The current study characterizes the cytokine protein (ELISA) and mRNA (gene array and RT-PCR) profiles of skin-derived mast cells cultured under serum-free conditions when activated by cross-linking of Fc epsilonRI. Prior to mast cell activation, mRNA only for TNF-alpha was detected, while after activation mRNA for IL-5, IL-6, IL-13, TNF-alpha, and GM-CSF substantially increased, and for IL-4 it minimally increased. However, at the protein level certain recombinant cytokines, as measured by ELISAs, were degraded by proteases released by these skin-derived mast cells. IL-6 and IL-13 were most susceptible, followed by IL-5 and TNF-alpha; GM-CSF was completely resistant. These observations also held for the endogenous cytokines produced by activated mast cells. By using protease inhibitors, chymase and cathepsin G, not tryptase, were identified in the mast cell releasates as the likely culprits that digest these cytokines. Their cytokine-degrading capabilities were confirmed with purified chymase and cathepsin G. Soy bean trypsin inhibitor, when added to mast cell releasates, prevented the degradation of exogenously added cytokines and, when added to mast cells prior to their activation, prevented degradation of susceptible endogenous cytokines without affecting either degranulation or GM-CSF production. Consequently, substantial levels of IL-5, IL-6, IL-13, TNF-alpha, and GM-CSF were detected 24-48 h after mast cells had been activated, while none were detected 15 min after activation, by which time preformed granule mediators had been released. IL-4 was not detected at any time point. Thus, unless cytokines are protected from degradation by endogenous proteases, cytokine production by human mast cells with chymase and cathepsin G cells may be grossly underestimated.     

Connective tissue mast cells in contact with fibroblasts express IL-3 mRNA. Analysis of single cells by polymerase chain reaction.

Mast cell-fibroblast interactions have been extensively investigated in the last few years. Fibroblasts support the in vitro survival but not proliferation of mouse connective-tissue type mast cells. However, the factor(s) that allow their survival on fibroblast monolayers has not been identified. We have investigated the presence of mRNA for IL-3 and granulocyte-macrophage-CSF in single mouse mast cells, before and after co-culture with 3T3 fibroblasts, using the polymerase chain reaction technique. The system was calibrated first by using in vitro generated population of mouse bone-marrow derived mast cells (BMMC). Significant differences in the amplification of IL-3 cDNA were observed in each of the BMMC cells examined, whereas the amplification of cDNA for the alpha-subunit of the Fc epsilon RI were similar. Inasmuch as murine cultured IL-3-dependent mast cells differentiate into connective tissue-like mast cells when co-cultured with 3T3 fibroblasts without any exogenous supply of growth factors, it was of interest to determine whether these connective tissue-like mast cells produce IL-3 message. Separation of the differentiated BMMC from the fibroblast monolayer, by either trypsinization or by single cell manipulation revealed the synthesis of a detectable amount of IL-3 mRNA in these mast cells. Whether this IL-3 mRNA was induced by fibroblasts was further investigated using connective tissue mast cells freshly purified from the mouse peritoneal cavity. Only about 20% of these connective tissue mast cells produced detectable amount of granulocyte-macrophage-CSF mRNA whereas in less than 10% of the cells IL-3 mRNA was detected. However, when these connective tissue mast cells were co-cultured with 3T3 fibroblasts for 18 hours and then separated, IL-3 mRNA were detected in most of the cells whereas no such mRNA was detected in tissue mast cells incubated for 18 h with medium derived from 3T3 fibroblasts. Therefore we conclude that fibroblasts induce the accumulation of IL-3 mRNA in connective tissue mast cells. The production of IL-3 may play a role in the survival of this type of mast cells on the fibroblast monolayer.     

Structural requirements of SLP-76 in signaling via the high-affinity immunoglobulin E receptor (Fc epsilon RI) in mast cells

The adapter SLP-76 plays an essential role in Fc epsilon RI signaling, since SLP-76(-/-) bone marrow-derived mast cells (BMMC) fail to degranulate and release interleukin-6 (IL-6) following Fc epsilon RI ligation. To define the role of SLP-76 domains and motifs in Fc epsilon RI signaling, SLP-76(-/-) BMMC were retrovirally transduced with SLP-76 and SLP-76 mutants. The SLP-76 N-terminal and Gads binding domains, but not the SH2 domain, were critical for Fc epsilon RI-mediated degranulation and IL-6 secretion, whereas all three domains are essential for T-cell proliferation following T-cell receptor (TCR) ligation. Unexpectedly, the three tyrosine residues in SLP-76 critical for TCR signaling, Y112, Y128, and Y145, were not essential for IL-6 secretion, but were required for degranulation and mitogen-activated protein kinase activation. Furthermore, a Y112/128F SLP-76 mutant, but not a Y145F mutant, strongly reconstituted mast cell degranulation, suggesting a critical role for Y145 in Fc epsilon RI-mediated exocytosis. These results point to important differences in the function of SLP-76 between T cells and mast cells.     

Expression of a functional high-affinity IgG receptor, Fc gamma RI, on human mast cells: Up-regulation by IFN-gamma

Biologically relevant activation of human mast cells through Fc receptors is believed to occur primarily through the high-affinity IgE receptor Fc epsilon RI. However, the demonstration in animal models that allergic reactions do not necessarily require Ag-specific IgE, nor the presence of a functional IgE receptor, and the clinical occurrence of some allergic reactions in situations where Ag-specific IgE appears to be lacking, led us to examine the hypothesis that human mast cells might express the high-affinity IgG receptor Fc gamma RI and in turn be activated through aggregation of this receptor. We thus first determined by RT-PCR that resting human mast cells exhibit minimal message for Fc gamma RI. We next found that IFN-gamma up-regulated the expression of Fc gamma RI. This was confirmed by flow cytometry, where Fc gamma RI expression on human mast cells was increased from approximately 2 to 44% by IFN-gamma exposure. Fc epsilon RI, Fc gamma RII, and Fc gamma RIII expression was not affected. Scatchard plots were consisted with these data where the average binding sites for monomeric IgG1 (Ka = 4-5 x 108 M-1) increased from approximately 2,400 to 12,100-17,300 per cell. Aggregation of Fc gamma RI on human mast cells, and only after IFN-gamma exposure, led to significant degranulation as evidenced by histamine release (24.5 +/- 4.4%): and up-regulation of mRNA expression for specific cytokines including TNF-alpha, GM-CSF, IL-3 and IL-13. These findings thus suggest another mechanism by which human mast cells may be recruited into the inflammatory processes associated with some immunologic and infectious diseases.     

Secretory granule mediator release and generation of oxidative metabolites of arachidonic acid via Fc-IgG receptor bridging in mouse mast cells.

The releases of beta-hexosaminidase, LTC4, LTB4, and PGD2 after the bridging of Fc gamma R3 were assessed in mouse IL-3-dependent bone marrow-derived progenitor mast cells (BMMC), BMMC maintained in coculture with 3T3 fibroblasts separated by a filter to achieve maturation of the granules toward those of a serosal mast cell (SMC), and SMC that are the prototype of a mouse connective tissue mast cell. Bridging of Fc gamma R on BMMC with the 2.4G2 rat anti-Fc gamma RII/III mAb and anti-rat IgG elicited only 4% net release of beta-hexosaminidase and 4, 2, and 1 ng/10(6) cells of immunoreactive LTC4, LTB4, and PGD2, respectively. Bridging of Fc-IgE receptors (Fc epsilon R) on BMMC yielded 35% net release of beta-hexosaminidase and 9, 4, and 3 ng/10(6) cells of immunoreactive LTC4, LTB4, and PGD2, respectively. BMMC maintained in coculture responded to the bridging of Fc gamma R with statistically significant increases in the net percent release of beta-hexosaminidase to 16% and in the generation of immunoreactive LTC4 to 11 ng/10(6) cells, but without a significant change in the production of either LTB4 or PGD2. Bridging of Fc epsilon R on cocultured mast cells yielded a net percent release of beta-hexosaminidase and lipid mediator amounts and profile similar to those for BMMC. Bridging of Fc gamma R on purified mouse SMC resulted in a maximal net percent release of beta-hexosaminidase of 10% and the generation of 4, 1, and 17 ng/10(6) cells of immunoreactive LTC4, LTB4, and PGD2, respectively; the net percent release of beta-hexosaminidase and PGD2 generation were significantly greater than those obtained from BMMC. The Fc epsilon R-mediated net percent release of beta-hexosaminidase from purified SMC was 34%, with PGD2 being the predominant metabolite of arachidonic acid. That the predominant lipid mediator generated with activation by either Fc gamma R or Fc epsilon R is LTC4 for cocultured mast cells and PGD2 for SMC suggests that the mast cell phenotype rather than the receptor class being bridged determines the lipid mediator profile. The responsiveness to Fc gamma R bridging elicited by coculture of BMMC with fibroblasts in vitro and present in SMC derived in vivo relative to BMMC may relate to the previously measured increases in receptor number per cell, but may also involve the acquisition of an enhanced signal transduction capability, possibly through the increased expression of Fc gamma RIII.     

Carboxypeptidase A in mouse mast cells. Identification, characterization, and use as a differentiation marker

By using a conventional spectrophotometric assay with hippuryl-L-phenylalanine as the substrate, 10(6) BALB/c mouse serosal mast cells possessed 1.5 +/- 0.43 U (mean +/- SE, n = 5, range = 0.48 to 2.5) of carboxypeptidase A activity, while T cell factor-dependent, mouse bone marrow-derived mast cells (BMMC) had barely detectable levels of 0.01 +/- 0.001 U/10(6) cells (mean +/- SE, n = 3). In order to characterize the carboxypeptidase A present in the BMMC, a sensitive assay was developed that used angiotensin I as the substrate and reverse phase-high performance liquid chromatography to separate and quantify production of the cleavage product des-leu-angiotensin I. Using this assay, mouse BMMC carboxypeptidase A had a neutral to basic pH optimum and hydrolyzed angiotensin I with a Km of 0.78 mM. The antigen-induced net percent release of carboxypeptidase A from IgE-sensitized BMMC was proportional to that of the secretory granule component beta-hexosaminidase which indicates a secretory granule location for the exopeptidase. As defined by exclusion during Sepharose CL-2B chromatography, carboxypeptidase A was exocytosed as a greater than 1 X 10(7) m.w. complex bound to proteoglycans. Because BMMC cocultured with mouse skin-derived 3T3 fibroblasts are known to undergo an increase in histamine content and biosynthesis of 35S-labeled heparin proteoglycans, carboxypeptidase A activity was measured during BMMC/fibroblast coculture for 0 to 28 days. The carboxypeptidase A activity increased progressively during 28 days of co-culture from 0.004 +/- 0.002 U/10(6) starting BMMC (mean +/- SE, n = 3) to 0.36 +/- 0.10 U/10(6) co-cultured mast cells. These findings indicate that carboxypeptidase A, a neutral protease, is exocytosed from the secretory granules of mouse mast cells bound to proteoglycan and is increased during the in vitro differentiation of mouse BMMC from mucosal-like mast cells to serosal-like mast cells.     

TLR3-, TLR7-, and TLR9-mediated production of proinflammatory cytokines and chemokines from murine connective tissue type skin-derived mast cells but not from bone marrow-derived mast cells

Recent studies have revealed that murine bone marrow-derived cultured mast cells (BMMC), which are phenotypically immature mast cells, express functional TLR2 and TLR4 that recognize distinct pathogen-associated molecules. However, it remains relatively uncertain whether mast cells express other TLR. We recently established a method to obtain large numbers of murine fetal skin-derived cultured mast cells (FSMC); these cells exhibit important features of connective tissue type mast cells. Working with FSMC and BMMC, the TLR mRNA expression profiles were compared between both cell types. Although TLR2 and TLR4 mRNA were detected in both cells at comparable levels, TLR3, TLR7, and TLR9 mRNA were expressed by FSMC at higher levels than by BMMC, suggesting distinct TLR expression profiles among different mast cell populations. With respect to their functional aspects, FSMC, but not BMMC, dose dependently produced proinflammatory cytokines (TNF-alpha and IL-6) and chemokines (RANTES, MIP-1alpha, and MIP-2) in response to poly(I:C), R-848, and CpG oligodeoxynucleotide, which are TLR3, TLR7, and TLR9 activators, respectively. Interestingly, these TLR activators failed to induce degranulation and IL-13 production by both mast cells, although peptidoglycan and LPS (TLR2 and TLR4 activators, respectively) induced IL-13 production by both cells. Mast cells, thus, may have potential to recruit other immune cells to the infected sites by responding to various bacterial and viral components through TLR signaling pathways, presumably being involved in initiating innate immunity and subsequently linking innate and acquired immune responses.     

Phospholipase C gamma 2 is essential for specific functions of Fc epsilon R and Fc gamma R

Phospholipase Cgamma2 (PLCgamma2) plays a critical role in the functions of the B cell receptor in B cells and of the FcRgamma chain-containing collagen receptor in platelets. Here we report that PLCgamma2 is also expressed in mast cells and monocytes/macrophages and is activated by cross-linking of Fc(epsilon)R and Fc(gamma)R. Although PLCgamma2-deficient mice have normal development and numbers of mast cells and monocytes/macrophages, we demonstrate that PLCgamma2 is essential for specific functions of Fc(epsilon)R and Fc(gamma)R. While PLCgamma2-deficient mast cells have normal mitogen-activated protein kinase activation and cytokine production at mRNA levels, the mutant cells have impaired Fc(epsilon)R-mediated Ca(2+) flux and inositol 1,4,5-trisphosphate production, degranulation, and cytokine secretion. As a physiological consequence of the effect of PLCgamma2 deficiency, the mutant mice are resistant to IgE-mediated cutaneous inflammatory skin reaction. Macrophages from PLCgamma2-deficient mice have no detectable Fc(gamma)R-mediated Ca(2+) flux; however, the mutant cells have normal Fc(gamma)R-mediated phagocytosis. Moreover, PLCgamma2 plays a nonredundant role in Fc(gamma)R-mediated inflammatory skin reaction.     

Molecular cloning of the mouse mast cell protease-5 gene. A novel secretory granule protease expressed early in the differentiation of serosal mast cells

cDNAs were isolated that encode mouse mast cell protease-5 (MMCP-5), an approximately 30,000 Mr serine protease stored in the secretory granules of serosal mast cells (SMC) and Kirsten sarcoma virus-immortalized mast cells. Based on the deduced amino acid sequences of these cDNAs, MMCP-5 is synthesized as a 247-amino acid preproenzyme composed of a novel 19-residue hydrophobic signal peptide, a Gly-Glu activation peptide not present in other mast cell chymases, and a 226-amino acid protein that represents the mature enzyme. MMCP-5 possesses a unique Asn residue in the substrate binding cleft at residue 176 and is highly basically charged. The MMCP-5 gene was isolated, sequenced, and found to belong to a distinct subset of chymase genes. Allelic variations of the MMCP-5 gene were also detected. MMCP-5 is expressed in bone marrow-derived mast cells (BMMC), Kirsten sarcoma virus-immortalized mast cells, and SMC, but not in gastrointestinal mucosal mast cells of helminth-infected mice. The abundant levels of MMCP-5 mRNA in immature BMMC indicate that this chymase is expressed relatively early during the differentiation of mast cells. MMCP-5 is the first chymase to be molecularly cloned from progenitor mast cells and is also the first chymase shown to be expressed preferentially in the SMC subclass.     

Peroxisome proliferator-activated receptor ligands negatively regulate the expression of the high-affinity IgE receptor Fc epsilon RI in human basophilic KU812 cells

The high-affinity IgE receptor Fc epsilon RI is expressed on the cell surface of mast cells and basophils, and plays a central role in IgE-mediated inflammatory reactions. Recently, peroxisome proliferator-activated receptors (PPARs) have been implicated in the anti-inflammatory response. To investigate a possible role for PPAR in human basophils, the effect of PPAR ligands on Fc epsilon RI expression in human basophilic KU812 cells was studied. The PPARalpha ligand, leukotriene B(4), did not affect the cell surface expression of Fc epsilon RI. However, prostaglandin (PG) A(1) and 15-deoxy-Delta(12,14) PGJ(2) (15d-PGJ(2)), which are PPARbeta and gamma ligands, respectively, were both able to decrease Fc epsilon RI expression. Treatment with PGA(1) or 15d-PGJ(2) separately also reduced histamine release from KU812 cells in response to cross-linkage of Fc epsilon RI. In addition, RT-PCR analysis showed that KU812 cells expressed the mRNA for PPARalpha, beta, and gamma, indicating that PPARbeta or gamma may negatively regulate the cell activation via Fc epsilon RI. Cells treated with 15d-PGJ(2) expressed lower levels of Fc epsilon RI alpha and gamma mRNA, and PGA(1) treatment decreased the level of Fc epsilon RI gamma mRNA. These results suggest that the suppression of Fc epsilon RI expression by PPARs may be due to the down-regulation of Fc epsilon RI alpha or gamma mRNA.     

Maturation-related changes in the expression of Fc gamma RII and Fc gamma RIII on mouse mast cells derived in vitro and in vivo.

The expression and function of Fc gamma RII and Fc gamma RIII on three mouse mast cell populations that differ in maturity as assessed by secretory granule constituents were analyzed by cellular and immunochemical approaches. As quantified by flow cytometric analysis of the binding of the rat 2.4G2 anti-Fc gamma RII/III mAb, mouse serosal mast cells (SMC) purified from the peritoneal cavity expressed more receptors per cell than did mouse IL-3-dependent, bone marrow culture-derived mast cells (BMMC), which are progenitors of SMC. Coculture of BMMC with mouse 3T3 fibroblasts for 2 wk, which alters the secretory granule composition toward that of SMC, also increased receptor epitope expression to a level equivalent to that of SMC. As assessed by rosette assays with mouse mAb to SRBC, all three mast cell populations bound IgG1, IgG2a, and IgG2b, essentially all binding was inhibited by 2.4G2 antibody, and greater quantities of the antibody were required to block immune adherence by cocultured mast cells and SMC as compared with BMMC. Immunoprecipitation and SDS-PAGE analysis of Fc gamma RII and Fc gamma RIII from BMMC, cocultured mast cells, and SMC that were surface radiolabeled with Na125I revealed predominant native forms of 62, 57, and 56 kDa, respectively, and an additional surface form of 43 kDa in SMC. Removal of N-linked carbohydrate from immunoprecipitates demonstrated that BMMC expressed peptide cores of 38 kDa (Fc gamma RII-1 gene product) and 31 kDa (Fc gamma RII-2 gene product), and barely detectable amounts of a 28-kDa (Fc gamma RIII gene product) core. The expression of all three was increased by coculture with 3T3 fibroblasts, consistent with the increased expression of their common epitope by cytofluorographic analysis. SMC expressed primarily the Fc gamma RII-1 and some Fc gamma RIII gene product. Thus, the three populations of mast cells express different amounts and ratios of the Fc gamma RII and Fc gamma RIII gene products, and maturation of BMMC during coculture with fibroblasts in vitro and in the peritoneal cavity in vivo augments cell-surface expression of the receptors and immune adherence function.     

曲古菌素A对FcεRI介导的肥大细胞活化的影响

曲古菌素A（TSA）是一种组蛋白去乙酰化酶抑制剂,在变态反应等免疫性疾病和抗肿瘤方面显示了潜在的治疗效应。主要观察了TSA对变态反应的效应细胞—肥大细胞活化的影响。TSA可诱导小鼠骨髓来源的肥大细胞（BMMC）凋亡,并抑制表面FcεRI的表达。TSA处理的BMMC经anti-IgE/IgE刺激之后,脱颗粒及分泌TNF-α、IL-6和IL-13生物活性介质的能力显著受到抑制。由此提示,TSA可能通过诱导BMMC凋亡和/或下调FcεRI表达,进而抑制FcεRI介导的BMMC活化,这为TSA进一步在肥大细胞相关的变态反应性疾病中的应用提供了实验数据。     

Prostaglandin E2 selectively enhances the IgE-mediated production of IL-6 and granulocyte-macrophage colony-stimulating factor by mast cells through an EP1/EP3-dependent mechanism

PGE(2) is an endogenously synthesized inflammatory mediator that is over-produced in chronic inflammatory disorders such as allergic asthma. In this study, we investigated the regulatory effects of PGE(2) on mast cell degranulation and the production of cytokines relevant to allergic disease. Murine bone marrow-derived mast cells (BMMC) were treated with PGE(2) alone or in the context of IgE-mediated activation. PGE(2) treatment alone specifically enhanced IL-6 production, and neither induced nor inhibited degranulation and the release of other mast cell cytokines, including IL-4, IL-10, IFN-gamma, and GM-CSF. IgE/Ag-mediated activation of BMMC induced the secretion of IL-4, IL-6, and GM-CSF, and concurrent PGE(2) stimulation synergistically increased mast cell degranulation and IL-6 and GM-CSF, but not IL-4, production. A similar potentiation of degranulation and IL-6 production by PGE(2), in the context of IgE-directed activation, was observed in the well-established IL-3-dependent murine mast cell line, MC/9. RT-PCR analysis of unstimulated MC/9 cells revealed the expression of EP(1), EP(3), and EP(4) PGE receptor subtypes, including a novel splice variant of the EP(1) receptor. Pharmacological studies using PGE receptor subtype-selective analogs showed that the potentiation of IgE/Ag-induced degranulation and IL-6 production by PGE(2) is mediated through EP(1) and/or EP(3) receptors. Our results suggest that PGE(2) may profoundly alter the nature of the mast cell degranulation and cytokine responses at sites of allergic inflammation through an EP(1)/EP(3)-dependent mechanism.     

IgE enhances Fc epsilon receptor I expression and IgE-dependent release of histamine and lipid mediators from human umbilical cord blood-derived mast cells: synergistic effect of IL-4 and IgE on human mast cell Fc epsilon receptor I expression and mediator release

We investigated the effects of IgE versus IL-4 on Fc epsilon RI surface expression in differentiated human mast cells derived in vitro from umbilical cord blood mononuclear cells. We found that IgE (at 5 micrograms/ml) much more strikingly enhanced surface expression of Fc epsilon RI than did IL-4 (at 0.1-100 ng/ml); similar results were also obtained with differentiated mouse mast cells. However, IL-4 acted synergistically with IgE to enhance Fc epsilon RI expression in these umbilical cord blood-derived human mast cells, as well as in mouse peritoneal mast cells derived from IL-4-/- or IL-4+/+ mice. We also found that: 1) IgE-dependent enhancement of Fc epsilon RI expression was associated with a significantly enhanced ability of these human mast cells to secrete histamine, PGD2, and leukotriene C4 upon subsequent passive sensitization with IgE and challenge with anti-IgE; 2) preincubation with IL-4 enhanced IgE-dependent mediator secretion in these cells even in the absence of significant effects on Fc epsilon RI surface expression; 3) when used together with IgE, IL-4 enhanced IgE-dependent mediator secretion in human mast cells to levels greater than those observed in cells that had been preincubated with IgE alone; and 4) batches of human mast cells generated in vitro from umbilical cord blood cells derived from different donors exhibited differences in the magnitude and pattern of histamine and lipid mediator release in response to anti-IgE challenge, both under baseline conditions and after preincubation with IgE and/or IL-4.