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Biomedical subjects

J T Schroeder

Publications and source records attributed to J T Schroeder.

At least 19 recordsLinked to original sources
PubMed2001

Selective inhibition of interleukin-4 gene expression in human T cells by aspirin.

Previous studies indicated that aspirin (acetylsalicylic acid [ASA]) can have profound immunomodulatory effects by regulating cytokine gene expression in several types of cells. This study is the first in which concentrations of ASA in the therapeutic range were found to significantly reduce interleukin (IL)-4 secretion and RNA expression in freshly isolated and mitogen-primed human CD4+ T cells. In contrast, ASA did not affect IL-13, interferon-gamma, and IL-2 expression. ASA inhibited IL-4, but not IL-2, promoter-driven chloramphenicol acetyltransferase expression in transiently transfected Jurkat T cells. The structurally unrelated nonsteroidal anti-inflammatory drugs indomethacin and flurbiprofen did not affect cytokine gene expression in T cells, whereas the weak cyclo-oxygenase inhibitor salicylic acid was at least as effective as ASA in inhibiting IL-4 expression and promoter activity. The inhibitory effect of ASA on IL-4 transcription was not mediated by decreased nuclear expression of the known salicylate target nuclear factor (NF)-kappaB and was accompanied by reduced binding of an inducible factor to an IL-4 promoter region upstream of, but not overlapping, the NF of activated T cells- and NF-kappaB-binding P1 element. It is concluded that anti-inflammatory salicylates, by means of a previously unrecognized mechanism of action, can influence the nature of adaptive immune responses by selectively inhibiting the expression of IL-4, a critical effector of these responses, in CD4+ T cells.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed2001

Inhibition of cytokine generation and mediator release by human basophils treated with desloratadine.

BACKGROUND: Desloratadine is a non-sedating, clinically effective, anti-allergic therapy that has been shown to exhibit anti-inflammatory properties that extend beyond its ability to antagonize histamine at H(1)-receptor sites. This latter effect has been shown in vitro to be both IgE-dependent and -independent. OBJECTIVE: In this study, we addressed the ability of desloratadine to inhibit the in vitro generation of interleukin (IL)-4 and IL-13 from human basophils while concurrently comparing its efficacy in preventing mediator release by these cells. METHODS: Basophil-enriched suspensions were treated with various concentrations of desloratadine for 15 min before stimulating with either anti-IgE antibody, calcium ionophore, IL-3 or phorbol ester. Histamine (fluorimetry), LTC(4) (RIA) and IL-4 (ELISA) were all assayed using the same 4-h culture supernatants. IL-13 (ELISA) was measured in supernatants harvested after 20 h incubation. IL-4 mRNA expression (dilutional RT-PCR) was also examined. RESULTS: Desloratadine was found to be nearly six-seven times more potent in preventing the secretion of IL-4 and IL-13 induced by anti-IgE than it was at inhibiting the release of histamine and LTC(4). These cytokines were equally inhibited by desloratadine following activation with ionomycin despite the lack of an effect on the histamine induced with ionomycin. Desloratadine had a lesser effect regarding inhibition of the IL-13 secreted in response to IL-3 and PMA. There was no evidence that desloratadine mediated its inhibitory effects by causing decreased cell viability. Finally, IL-4 mRNA accumulation was remarkably inhibited, by as much as 80%, following pretreatment with desloratadine. CONCLUSION: While capable of inhibiting histamine and LTC(4) release by human basophils, desloratadine is more effective at targeting the signals regulating IL-4 and IL-13 generation in these cells. This inhibitory effect on cytokine generation provides additional evidence that this antihistamine exerts anti-inflammatory properties.

Basophils↗
PubMed2001

IL-4 production by human basophils found in the lung following segmental allergen challenge.

BACKGROUND: Human blood basophils secrete high levels of IL-4 following activation with specific allergen, yet their role as cytokine-producing cells in allergic lesions has not been described. OBJECTIVE: Our objective was to investigate whether and under what conditions basophils infiltrating allergic lesions in the lung secrete IL-4 in vitro. METHODS: Bronchoalveolar lavage (BAL) cells were recovered 20 hours after segmental allergen challenge. Basophils were enriched with Percoll using a protocol commonly used for blood basophils. IL-4 and histamine were measured in culture supernatants following activation with a variety of stimuli. Two-color flow cytometry was performed to detect intracellular IL-4. RESULTS: IL-4 protein was detected in all basophil culture supernatants following a 4- to 5-hour incubation in medium alone; the levels obtained did not significantly increase with the addition of anti-IgE. BAL basophils failed to release histamine in response to specific allergen but showed nearly 60% histamine release with N-formyl-methionyl-leucyl-phenylalanine, suggesting that they were desensitized to IgE-mediated stimuli as a result of their activation in vivo. Using these same conditions, IL-4 was not detected in BAL cell fractions enriched for lymphocytes and eosinophils. Ionomycin induced IL-4 secretion by BAL basophils, and this response was reduced with the addition of phorbol myristate acetate. In contrast, phorbol myristate acetate promoted the secretion of IL-4 by BAL cells enriched for lymphocytes; both findings are identical to those reported for basophils and lymphocytes purified from blood. Flow cytometry confirmed the secretion of IL-4 by BAL basophils. CONCLUSIONS: These data suggest that basophils migrating to the lung following allergen challenge represent a major source of IL-4.

Allergens↗
PubMed2001

Effects of prednisone on the cellular responses and release of cytokines and mediators after segmental allergen challenge of asthmatic subjects.

BACKGROUND: Systemic glucocorticoids are a major therapy for the management of allergic inflammation and asthma; however, information about their effects in vivo are limited. OBJECTIVE: This study was performed to examine the effects of prednisone on inflammatory mediators, cytokines, and cellular responses in the model of segmental allergen challenge (SAC) of allergic asthmatic subjects. METHODS: The effects of a 3-day pretreatment with oral prednisone (30 mg twice daily) on the physiologic and inflammatory responses to SAC were studied in 10 allergic asthmatic subjects in a double-blind, placebo-controlled, crossover protocol. RESULTS: Prednisone improved baseline FEV(1) by 10% and modestly inhibited the SAC-induced fall in FEV(1) at 30 minutes and at 6 to 8 hours. Five minutes after challenge, levels of histamine, PGD(2), 9alpha,11beta-PGF(2), and thromboxane B(2) increased in bronchoalveolar lavage fluid (median increase, 5- to 14-fold); prednisone did not inhibit these responses. Prednisone inhibited (median decrease, 66%-97%) the total influx of inflammatory cells, specifically eosinophils, basophils, and some subsets of T lymphocytes (CD4, CD45RA, and CD45RO cells) assessed 19 hours after SAC, but it did not inhibit the influx of neutrophils. Increases in soluble E-selectin, kinins, and albumin were also inhibited by the glucocorticoid (median decrease, 36%-74%). Prednisone treatment inhibited the appearance of mRNA, protein, or both for T(H)2 cytokines (IL-4 and IL-5), as well as for IL-2 and transforming growth factor alpha, but did not inhibit increases of immunoreactive GM-CSF in bronchoalveolar lavage fluid. CONCLUSION: These studies indicate that prednisone suppresses multiple components of allergic airway inflammation, including cell recruitment, adhesion molecule expression or release, airway permeability, and production of cytokines potentially involved in airway immunity or remodeling.

Adult↗
PubMed2001

Nerve growth factor or IL-3 induces more IL-13 production from basophils of allergic subjects than from basophils of nonallergic subjects.

BACKGROUND: Studies show that nerve growth factor (NGF) exhibits immunomodulatory activity. This neurotrophin is found at high levels in the serum of asthmatic individuals, is released during allergic reactions, and is reported to augment in vitro histamine and leukotriene C4 release by human basophils. OBJECTIVE: Because basophils represent a substantial source of IL-4 and IL-13, we tested the effects of NGF on the secretion of these cytokines by cells prepared from allergic subjects and cells prepared from nonallergic subjects. METHODS: Cytokine and histamine were measured in culture supernatants by ELISA and fluorimetry, respectively. Both real-time RT-PCR and conventional RT-PCR were used to measure IL-13 mRNA expression. NGF receptor expression was determined by 2-color flow cytometry. RESULTS: Basophil suspensions from allergic subjects secreted some 2.5-fold greater levels of IL-13 when cultured with NGF than did cells prepared from normal control subjects. Flow cytometry revealed no significant differences in TrkA receptors on basophils to explain these findings. The levels of IL-13 secreted by the 2 groups of donors also differed when cells were activated with IL-3 but not when they were activated with anti-IgE antibody. Both NGF and IL-3 failed to induce IL-13 in cell cultures depleted of basophils, suggesting that the measurable IL-13 was indeed basophil-derived. Real-time RT-PCR showed an average induction of IL-13 message above medium control that was 4.3 (+/- 1.7)-fold with NGF and 8.9 (+/- 3.7)-fold with IL-3. Finally, NGF priming resulted in a remarkable enhancement of IL-13 induced by anti-IgE. This was significantly greater than the priming observed for either the IL-4 or histamine when this stimulus was used. CONCLUSION: NGF (like IL-3) can both directly stimulate IL-13 secretion and modulate IgE-mediated responses in basophils. Its enhanced effect on cells from allergic individuals raises the importance of this cytokine in the pathogenesis of allergic disease.

Adjuvants, Immunologic↗
PubMed2000

Functional consequences of FcepsilonRIalpha up-regulation by IgE in human basophils.

These studies examine the functional changes that occur after up-regulation of FcepsilonRIalpha by immunoglobulin E (IgE) for human basophils. Basophils were cultured with and without IgE antibody (PS myeloma IgE or anti-gp120-specific IgE) for 1 week and challenged with anti-IgE, anti-FcepsilonRIalpha, or antigen for histamine and IL-4 secretion. There were no statistically significant changes in their response to anti-IgE or anti-receptor antibodies, as compared with controls incubated for the same period, whereas receptor expression increased an average of 4-fold. There was increased responsiveness to antigenic challenge, most notably at suboptimal concentrations of antigen (gp120 peptide-ovalbumin conjugate). For a 6-fold difference in cell surface density of gp120-specific IgE, there was a 2.2-fold change in antigen potency or 3-fold increases in histamine release at lower antigen concentrations. Similar results were found for secretion of IL-4. Basophil sensitivity, which is a measure of the density of antigen-specific IgE required for 50% of maximal secretion, was used to determine whether up-regulation of FcepsilonRIalpha was coordinated with up-regulation of other components of the IgE-signaling pathway. The results indicated up-regulation of FcepsilonRI is not always accompanied by changes that allow sensitivity to be maintained. These results indicate that functional up-regulation does occur but that its magnitude may be modulated because not all components of the signaling pathway are up-regulated in a balanced manner.

Antibodies, Anti-Idiotypic↗
PubMed1999

Extracellular signal-regulated kinases regulate leukotriene C4 generation, but not histamine release or IL-4 production from human basophils.

Human basophils secrete histamine and leukotriene C4 (LTC4) in response to various stimuli, such as Ag and the bacterial product, FMLP. IgE-mediated stimulation also results in IL-4 secretion. However, the mechanisms of these three classes of secretion are unknown in human basophils. The activation of extracellular signal-regulated kinases (ERKs; ERK-1 and ERK-2) during IgE- and FMLP-mediated stimulation of human basophils was examined. Following FMLP stimulation, histamine release preceded phosphorylation of ERKs, whereas phosphorylation of cytosolic phospholipase A2 (cPLA2), and arachidonic acid (AA) and LTC4 release followed phosphorylation of ERKs. The phosphorylation of ERKs was transient, decreasing to baseline levels after 15 min. PD98059 (MEK inhibitor) inhibited the phosphorylation of ERKs and cPLA2 without inhibition of several other tyrosine phosphorylation events, including phosphorylation of p38 MAPK. PD98059 also inhibited LTC4 generation (IC50 = approximately 2 microM), but not histamine release. Stimulation with anti-IgE Ab resulted in the phosphorylation of ERKs, which was kinetically similar to both histamine and LTC4 release and decreased toward resting levels by 30 min. Similar to FMLP, PD98059 inhibited anti-IgE-mediated LTC4 release (IC50, approximately 2 microM), with only a modest effect on histamine release and IL-4 production at higher concentrations. Taken together, these results suggest that ERKs might selectively regulate the pathway leading to LTC4 generation by phosphorylating cPLA2, but not histamine release or IL-4 production, in human basophils.

Antibodies, Anti-Idiotypic↗
PubMed1998

In vitro regulation of FcepsilonRIalpha expression on human basophils by IgE antibody.

In vivo studies suggested the possibility of an IgE-dependent regulation of high-affinity (FcepsilonRI) IgE receptor expression on basophils. The current studies extend these observations to in vitro cultures of human basophils. Incubation of basophils for 3 to 4 weeks resulted in a slow dissociation of IgE antibody, during which time FcepsilonRI expression decreased, as measured by flow cytometry using the anti-FcepsilonRIalpha monoclonal antibody, 22E7, or by measuring FcRIalpha mass by Western blotting of whole-cell lysates. Culture of basophils with IgE resulted in upregulation of FcepsilonRIalpha expression by both flow cytometry and Western blotting of whole-cell lysates. Upregulation followed a linear time course during 2 weeks of culture. The relative increase in FcepsilonRIalpha density depended on the starting density; with starting densities of FcepsilonRIalpha of 10,000 to 170,000 per basophil, the upregulation varied 20- to 1.1-fold, respectively. Upregulation occurred in high-purity basophils, was not influenced by IgG at concentrations up to 1 mg/mL, and was inhibited by dimeric IgE. Heat-inactivated IgE was less effective and the monoclonal antibody CGP51901 that prevents IgE binding to FcepsilonRIalpha blocked the ability of IgE to induce upregulation. The dose-response curve for IgE-induced upregulation had an effective concentration50 of 230 ng/mL. Although the receptor through which IgE induces this upregulation is not yet known, several characteristics suggest that the upregulation is mediated by IgE interacting through FcepsilonRIalpha itself.

Antibodies, Monoclonal↗
PubMed1998

Differential regulation of IL-4 and IL-13 secretion by human basophils: their relationship to histamine release in mixed leukocyte cultures.

Human basophils are an important source of IL-4, a cytokine that is central to the pathogenesis of allergic inflammation. Recent reports have indicated that these cells also generate IL-13, which shares a number of biologic properties with IL-4. We found basophils to be the major source of IL-13 produced in mixed leukocyte cultures following 20-h activation with a variety of stimuli. While the magnitude of IL-4 protein generated correlated with the percent histamine secreted (r = 0.8; p = 0.007), there was no relationship between the levels of IL-13 detected and the amount of either IL-4 or histamine in cultures activated with IL-3/anti-IgE. The induction of IL-13 secretion also occurred in response to IL-3 alone, without concomitant secretion of either IL-4 or histamine. Although previously shown to inhibit IL-4 secretion, the phorbol ester PMA was a potent stimulus for IL-13 generation from basophils, and this secretion was sensitive to the protein kinase C inhibitor, bisindolylmaleimide. In contrast, bisindolylmaleimide did not prevent cytokine secretion induced by either anti-IgE or IL-3. The immunosuppressant, FK506, while strikingly inhibiting the accumulation of IL-4 mRNA and the secretion of protein in response to IL-3/anti-IgE, had no effect on the generation of IL-13 in these cultures; the resistance was attributed to the IL-3-dependent signaling. Similarly, FK506 had no effect on the secretion of IL-13 in basophil cultures stimulated with PMA. This study suggests that multiple intracellular mechanisms control the generation of IL-13 in basophils, some of which are distinct from those regulating IL-4.

Adult↗
PubMed1998

IL-4 secretion and histamine release by human basophils are differentially regulated by protein kinase C activation.

The role of protein kinase C (PKC) activation was investigated in the secretion of interleukin-4 (IL-4) protein by human basophils. Phorbol myristate acetate (PMA) induced little to no detectable IL-4 protein in culture supernatants, despite being a potent secretagogue of histamine release by basophils. In fact, the secretion of IL-4 by basophils stimulated with ionomycin alone was down-regulated (30-70%) with the simultaneous addition of PMA. In peripheral blood lymphocytes (PBL), however, the combination of ionomycin and PMA were highly synergistic, resulting in maximum IL-4 release but at a slower rate. PKC inhibitors reversed these effects on IL-4 secretion. In sharp contrast to its inhibitory effect on IL-4 protein secretion, PMA did not block the accumulation of IL-4 mRNA in basophils activated by ionomycin. These data suggest that there are marked differences in the regulatory processes for IL-4 transcription, translation, or secretion between basophils and lymphocytes.

Basophils↗
PubMed1998

HrHRF: function and regulation.

Since basophils appear to play a fundamental role in the maintenance of allergic inflammation, the factors involved in basophil activation have become a focus of investigation in many laboratories. From this interest, the field of histamine-releasing factors (HRFs) has evolved. Our laboratory reported that a factor was present in late-phase skin blister fluids that caused basophil histamine release. It was hypothesized that basophil degranulation in these donors was mediated by the interaction of these HRFs with a certain kind of IgE, and upon experimentation a functional heterogeneity of the IgE molecule was uncovered. We designated the IgE from HRF responders as IgE+; the remaining IgE molecules designated were IgE-. Initially, it was thought that HrHRF might exert its activity by directly interacting with IgE+; however, a number of recent experiments have questioned this hypothesis. Observations suggest, in part, that HrHRF mediates biological activities on inflammatory cells by binding to a specific receptor rather than to the IgE molecule and/or the FcepsilonRI. In parallel with the search for an HrHRF receptor, a number of experiments have been performed to determine whether mediator release by this protein proceeds via a signal transduction pathway other than the one triggered by classic IgE-dependent stimuli such as anti-IgE antibody or antigen. Although further characterization is ongoing, the evidence thus far is consistent with the concept that HRF may be an important regulator of the cellular inflammation involved in the pathophysiology of allergy.

Journal Article↗
PubMed1997

Recombinant histamine-releasing factor enhances IgE-dependent IL-4 and IL-13 secretion by human basophils.

Human recombinant histamine-releasing factor (HrHRF) is known to directly stimulate histamine release and IL-4 secretion from basophils of selected atopic donors in a reaction requiring the expression of a particular type of IgE, referred to as IgE+. In this study, HrHRF is shown to affect the IgE-mediated release of IL-4, IL-13, and histamine from basophils not normally releasing to this protein (i.e, those expressing IgE-). Priming with several different concentrations of HrHRF for 15 min enhanced basophil secretion of IL-4 and histamine after 4 h in a dose-dependent fashion following activation with anti-IgE Ab (10 ng/ml). This effect of HrHRF priming also occurred in cultures activated with 1 or 100 ng/ml of anti-IgE Ab. The secretion of IL-13 protein was enhanced similarly by HrHRF priming in cultures stimulated for 16 to 20 h with anti-IgE Ab. There were, however, no apparent changes in the secretion of histamine or cytokine by basophils primed with HrHRF and activated with the IgE-independent secretogogue, FMLP. These findings suggest that HrHRF modifies the response of basophils for IgE-dependent secretion by binding to a specific receptor, broadening the possible role of this protein in chronic allergic inflammation.

Basophils↗
PubMed1997

Regulation of IgE-dependent IL-4 generation by human basophils treated with glucocorticoids.

Glucocorticoids are widely used in the therapeutic intervention of allergic diseases, affecting the function of a variety of proinflammatory cell types that participate in these disorders. These drugs have been shown to inhibit the release of histamine by human basophils, but not by mast cells, in a reaction requiring 8 to 24 h. To address whether the generation of IL-4 by basophils is similarly affected, we investigated the actions of glucocorticoids on the in vitro release of this cytokine induced by anti-IgE Ab or by the human recombinant histamine-releasing factor. The ability of basophils to generate IL-4 was immediately affected, with secretion of this protein being inhibited >50% with <1-h preexposure to steroid. However, the release of histamine in these cultures was inhibited only after 24-h preincubation, suggesting that the mechanisms controlling the release of this mediator differ significantly from those regulating cytokine secretion. A rank order of potency of the steroids tested for inhibition of IL-4 protein was as follows: triamcinolone > dexamethasone > betamethasone > hydrocortisone. The sex steroids, testosterone and estrogen, showed no effect on basophil secretion. Experiments using reverse transcription-PCR indicate that glucocorticoids inhibit IL-4 generation in basophils on the level of transcription. These studies suggest that the success of glucocorticoids in the treatment of allergic conditions is due in part to their ability to inhibit both mediator release and cytokine secretion by basophils.

Antibodies, Monoclonal↗
PubMed1996

An immunoglobulin E-dependent recombinant histamine-releasing factor induces interleukin-4 secretion from human basophils.

A novel recombinant histamine-releasing factor (rHFR), which stimulates secretion from a subpopulation of human basophils that express a particular type of immunoglobulin E (IgE) or IgE+, was found to induce interleukin-4 (IL-4) production from cells isolated from these same donors. The secretion of IL-4 protein induced by rHRF significantly correlated with histamine release and the amount of protein generated, and the kinetics were identical to those caused by anti-IgE activation. Furthermore, the ability of rHRF to induce IL-4 protein production from cells not normally responsive to this protein was transferred by passive sensitization with plasma containing IgE+ antibody. That this novel protein stimulates both mediator release and the secretion of IL-4 protein from human basophils suggests a prominent role for this molecule in allergic disease.

Basophils↗
PubMed1996

Comparative ultrastructural morphology of human basophils stimulated to release histamine by anti-IgE, recombinant IgE-dependent histamine-releasing factor, or monocyte chemotactic protein-1.

An ultrastructural analysis of human basophils stimulated with anti-IgE, recombinant histamine-releasing factor (rHRF), or monocyte chemotactic protein-1 (MCP-1) (compared with unstimulated cells) was performed. Partially purified peripheral blood basophils were prepared for electron microscopy at time points known to precede histamine release and at half-maximum histamine release times for each secretagogue. Activation morphologies associated with stimulation included granule-vesicle attachments, piecemeal degranulation, anaphylactic degranulation, and uropod formation. These features were qualitatively similar in the stimulated samples. Quantitative differences were evident, however, when stimulated samples were compared with controls or at different time points after stimulation with a single agent or when individual secretagogues were compared. All stimulated samples differed quantitatively from the control samples. Rank orders for morphologic activation events revealed that the most effective trigger for anaphylactic degranulation was anti-IgE > MCP-1 > rHRF, whereas the most effective trigger for uropod formation was rHRF > anti-IgE > MCP-1. Rank orders for piecemeal degranulation and granule-vesicle attachments were the same: MCP-1 > anti-IgE > rHRF. Important relationships among these anatomic events reveal that the development of motile configurations is not associated with the development of secretion morphologies and that piecemeal degranulation precedes and is inversely related to anaphylactic degranulation in stimulated samples.

Antibodies, Anti-Idiotypic↗