Effects of IL-4 and IL-13 on total and allergen specific IgE production by cultured PBMC from allergic patients determined with recombinant pollen allergens

BACKGROUND: Interleukin (IL)-4 and IL-13 have been shown to be potent switch factors for IgE synthesis in human B cells. OBJECTIVE: In this study we investigated the effects of recombinant human IL-4 and IL-13 on total and allergen specific IgE synthesis by peripheral blood mononuclear cells (PBMC) from pollen allergic patients and healthy control individuals. METHODS: Peripheral blood mononuclear cells (PBMC) from allergic patients were investigated for their capacity to produce allergen specific IgE in vitro. Total protein extracts from birch pollen and timothy grass pollen as well as purified recombinant birch pollen allergens, Bet v I, birch profilin (Bet v II) and recombinant timothy grass pollen allergens, Phl p I, Phl p II, and Phl p V were used to measure specific IgE-antibody synthesis in cell culture supernatants by IgE-immunoblot and ELISA. RESULTS: PBMC obtained from allergic patients spontaneously secreted allergen specific IgE in the culture supernatants. Addition of Interleukin 4, Interleukin 13 and anti-CD40 antibody to the cultures alone or in combinations significantly induced total IgE production whereas allergen specific IgE production was not affected. CONCLUSION: Our results indicate that the peripheral blood of allergic individuals contains long lived allergen specific B cells which have already switched to IgE production and which are not sensitive to IL-4 and IL-13 treatment. These results may have implications on attempts to use cytokines or cytokine antagonists in therapy of Type I allergy.     

Risk factors for sensitisation to methyltetrahydrophthalic anhydride

OBJECTIVES: To examine an association between specific IgE to methyltetrahydrophthalic anhydride (MTHPA) and exposure time, atopic history, smoking habits, and total IgE concentrations. METHODS: A cross sectional survey was carried out on a population of 148 workers from two condenser plants using epoxy resin with MTHPA, an acid anhydride curing agent known to cause allergy. RESULTS: Using a Pharmacia CAP system with a MTHPA human serum albumin conjugate, specific IgE antibody was detected in serum from 97 (66%) out of the 148 workers exposed to MTHPA. Stepwise multiple linear regression analysis showed a striking relation between log concentrations of specific and total IgE (P < 0.0001). Furthermore, when the workers were divided into two groups according to a cut-off point (100 IU/ml) between low and high total IgE, current smoking was significantly (P = 0.025) associated with specific IgE production only in the group with low total IgE (< 100 IU/ml). CONCLUSIONS: Smoking is the most significant risk factor for raising specific IgE to MTHPA in the group with low total IgE concentrations.     

Interleukin 1 production by peripheral blood mononuclear cells of children with bronchial asthma in remission

To analyse the change of the immunological response in the remission state of children with bronchial asthma, we studied the interleukin 1 (IL-1) production of peripheral blood mononuclear cells (PBMC) obtained from children with bronchial asthma sensitized by mite antigen. After PBMC were cultured for 24 hours with Dermatophagoides farinae (Df) or lipopolysaccharide (LPS), the PBMC-derived culture supernatant was estimated for IL-1 alpha and beta by enzyme-linked immunosorbent assay (ELISA). PBMC from some of subjects with active asthma produced IL-1 alpha and beta without any stimulation, but not those from controls or subjects in remission. IL-1 alpha and beta production of PBMC stimulated with Df was observed in all three groups, but IL-1 produced by subjects with active asthma was higher than that produced by subjects in the other two groups. Moreover, when PBMC were incubated with LPS, the secretion of both IL-1 alpha and beta was enhanced. PBMC from patients with active asthma produced both IL-1 alpha and beta in amounts comparable to those produced by PBMC from control subjects, but IL-1 production of PBMC from patients in remission was lower than in the other two groups. IL-1 beta production was about ten times as much as IL-1 alpha. Df-induced IL-1 production of PBMC from asthmatic patients sensitized by mite antigen, which was increased in the active state, was down-regulated in the remission state. Moreover, nonspecific stimuli such as LPS may induce the suppressive factors which down-regulate IL-1 production.(ABSTRACT TRUNCATED AT 250 WORDS)     

Central administration of the neurotensin receptor antagonist, SR48692, modulates diurnal and stress-related hypothalamic-pituitary-adrenal activity

BACKGROUND: The mechanism of bronchoobstruction in asthmatics following inhalation of lipopolysaccharide (LPS) is unclear. OBJECTIVE: In this study we have investigated the effects of the LPS on eosinophil survival rate in stimulating peripheral blood mononuclear cells (PBMCs) from patients with asthma. METHODS: PBMC supernatants from 10 asthmatic patients and 4 healthy subjects were compared for eosinophil survival rate using the same purified eosinophils. Two x 10(6) cells/ml of PBMCs were cultured for 24 hours in RPMI 1640 containing 10% fetal bovine serum along with 10 micrograms/ml of LPS. 10(6) cells/ml of eosinophils were incubated for 96 hours in the presence of PBMC-derived culture supernatants at 75% in volume. RESULTS: The viability of eosinophils incubated with PBMC supernatants from asthmatic patients stimulated with LPS was higher, 62.5% +/- 10.6% (mean +/- SD), than that without LPS, 26.9% +/- 10.8%, and also higher than that of PBMCs from healthy subjects with LPS, 40.0% +/- 15.4%. This increasing activity of asthmatic PBMC stimulated with LPS was markedly inhibited from 72.2% +/- 9.7% to 38.5% +/- 6.8% by addition of mouse anti-human GM-CSF antibody to the PBMC supernatant but not mouse anti-human IL-5 antibody. CONCLUSION: These results suggest that LPS enhances eosinophil survival in asthmatics by increasing the production of GM-CSF from PBMCs.     

A case of spontaneous remission of paragonimiasis miyazakii

A 52-year-old man was admitted with fever and chest pain. Chest X-ray showed a soft infiltration in the right lung and bilateral pleural effusions. A strong tuberculin reaction was elicited. Significant laboratory findings included eosinophilia (37% in peripheral blood and 78% in pleural fluid) and elevated IgE levels (577 IU/ml in sera and 6700 IU/ml in pleural fluid). Adenosine deaminase activity in the pleural fluid was high. No helminth eggs were detected after repeated examination of the pleural fluid and sputum. No definitive diagnosis was made. Three months of chemotherapy with INH and rifampicin resulted in little improvement. Corticosteroid was then administered orally under a tentative diagnosis of idiopathic eosinophilic pleurisy, which proved to be a successful treatment and resulted in a marked reduction of pleural fluid volume. Two years after discharge, the patient's chest X-ray was normal and laboratory findings were normal including the eosinophil count and IgE level. The pleural fluid obtained at the first admission and kept frozen was subjected to immunological analysis for anti-parasite antibody activity. The pleural fluid showed an unexpectedly high titer of antibody activity (x6400 dilution) against Paragonimus miyazakii antigen assayed by double diffusion Ouchterlony method. Examination of the sera obtained from the patient two years after discharge, however, revealed no detectable antibody activity against the parasite antigens assayed either by Ouchterlony or ELISA method. We concluded from the clinical as well as laboratory findings that the patient had recovered from Paragonimiasis miyazakii without specific intervention for the disease.     

A study of CD23 expression and regulation in the patients with asthma

To study the pathogenesis of asthma, we examined the CD23 expression on peripheral blood mononuclear cells (PBMC) from 25 healthy donors and 28 asthmatic patients with APAAP test. The results demonstrated that CD23 antigen expressed strongly on PBMC from patients and their PBMC correlated positively with their IgE level (r = 0.78 P < 0.01). We investigated the regulation of CD23 expression by IL-4 and IFN-gamma in vitro, the results demonstrated that IFN-gamma could inhibit spontaneous and IL-4 induced expression of CD23. It was suggested that the analysis of the percentage of positive CD23 expression is useful for making diagnosis and assessing severity of asthma and is of significance in elucidating the pathogenesis of the asthma.     

Immunomodulation by TYB-2285 of Dermatophagoides farinae (Df) antigen-induced IFN-gamma and IL-4 production in lymphocytes from children with bronchial asthma

Effect of TYB-2285 and its metabolites on immune responses by peripheral blood mononuclear cells (PBMC) from children with bronchial asthma was investigated. TYB-2285 and its metabolites have immunosuppressive activity for the proliferation by Df-stimulated patients' lymphocytes. Concanavalin A (Con A)-activated peripheral blood mononuclear cells (PBMC) from the patients were not affected by the same treatment. The results indicate that TYB-2285 and its metabolites are capable of suppressing antigen-induced 3H-thymidine (3H-TdR) uptake but not the response induced by Con A. Interferon-gamma (IFN-gamma) production by Df-stimulated PBMC from patients with active asthma, which was lower than that of normal lymphocytes, were reversed beyond the levels of that in normal subjects. Thus reduced production of IFN-gamma by Df-stimulated patients' lymphocytes was increased by TYB-2285 and its metabolites in a dose-dependent manner. This phenomenon was not observed in lymphocytes from normal subjects. Furthermore, TYB-2285 inhibited IL-4 production induced by Df antigen in asthmatic patients' lymphocytes. Taken, together, TYB-2285 could work as a weak immunosuppressant to modify lymphocytes' responses to allergen in patients with bronchial asthma. These data underscore the potential benefit for the patients with bronchial asthma.     

Influence of HLA-DR phenotype on tumor necrosis factor-alpha production in renal-transplant recipients

In healthy subjects, previous studies have demonstrated a great interindividual variability in the ability for tumor necrosis factor-alpha (TNF-alpha) production. The gene for TNF-alpha is closely linked to and located in the major histocompatibility complex (MHC) and it has been suggested that these interindividual differences may be HLA related. Since TNF-alpha is likely to be an important mediator in renal allograft rejection, we investigated the role of HLA antigens on TNF-alpha production rates by peripheral blood mononuclear cells (PBMC) from renal transplant recipients during stable graft function. HLA-DR2-positive recipients showed significantly lower spontaneous TNF-alpha production than DR2-negative patients (p < 0.001). Upon stimulation with OKT3, HLA-DR2-positive patients also showed significantly lower TNF-alpha production than DR2-negative subjects (p < 0.001). HLA-DR3-positive recipients, however, showed significantly higher spontaneous TNF-alpha production than DR3-negative individuals (p < 0.05). These results suggest that differences in TNF-alpha production, both spontaneous and induced, may be due to the expression of certain DR allotypes.     

The effect of T cell depletion with Campath-1M on immune reconstitution after chemotherapy and allogeneic bone marrow transplant as treatment for leukaemia

Thirty asthmatic children were examined allergic reaction against egg white, gelatin and vaccine solution before and after vaccination using skin prick test. We also measured the levels of specific IgE and IgG antibody against gelatin. The changes in clinical symptoms before and after vaccination were investigated in 25 asthmatic children by evaluating symptom and treatment score. The results were as follows; 1. In one subject who had delayed type of skin reaction to gelatin, the adverse reaction was also recognized at the skin site around 24 hrs after vaccination. In this subject, the levels of serum specific IgE and IgG to gelatin became positive after 5 months. 2. Specific IgE antibodies to gelatin were not detected in all subjects before and after vaccination. 3. The mean values of asthma symptom score before and after vaccination were 3.3 +/- 4.2 and 1.5 +/- 3.3 respectively. Those of treatment score before and after vaccination were 75.6 +/- 35.2 and 76.0 +/- 35.0 respectively. These results suggest that skin testing with gelatin and vaccine solution is useful as a screening method for predicting adverse reactions in asthmatic children and that influenza vaccination can be performed safely in skin test negative children.     

Frequencies of T cells expressing interleukin-4 and interleukin-5 in atopic asthmatic children. Comparison with atopic asthmatic adults

T-cell-derived cytokines have been implicated in the pathogenesis of asthma and it has been suggested that Th2-type cytokines (interleukin-4 [IL-4], interleukin-5 [IL-5]) are pivotal in the allergic inflammation. However, there are little data on human cytokine production by individual T cells at the protein level, in particular in asthmatic children. In this study we analyzed the cytokine production at the single cell level in peripheral blood from mild atopic asthmatic (AA) children and adults and age-matched atopic nonasthmatic (AN) and nonatopic nonasthmatic (NN) control subjects (n = 9 in each group) using the technique of intracellular cytokine detection by flow cytometry. Comparing asthmatic children with atopic and nonatopic control subjects, an increased percentage of IL-5-producing T cells (AA: median 4.9% [range 1.1 to 8.9%]; AN: 0.3% [0.2 to 0.9%], p = 0.003; NN: 0.4% [0.1 to 3.8%], p = 0.001) was detectable, with a positive correlation to the number of peripheral eosinophils and to bronchial hyperresponsiveness. The frequency of IL-4-producing T cells was increased in both atopic groups compared with nonatopic controls (AA: 1.2% [0.2 to 2.6%], p = 0.011; AN: 0.8% [0.4 to 3.7%], p = 0.007; NN: 0.4% [0.2 to 0.9%]) with a positive correlation to total IgE concentration. In adults there were no differences in IL-5- or IL-4-producing T cells between all three groups. A substantial proportion of T cells coproducing IL-4 and IL-5 was not detectable in children and adults. These findings indicate that in asthmatic children the frequencies of Th2-type-producing T cells are increased and that expression of IL-4 and IL-5 is regulated independently.     

Effects of Russell''s viper venom on blood coagulation, platelets and the fibrinolytic enzyme system

Abnormalities of second-wave platelet aggregation were demonstrated in 17 of 33 asthmatic patients in whom drug and diet intake were controlled in the hospital. Mean abnormal responses were significantly greater after epinephrine- (p less than 0.001), adenosine diphosphate-(less than 0.001), collagen- (p = 0.01), and thrombin- (p less than 0.001) induced platelet aggregation in patients with immunologically mediated asthma and serum IgE levels greater than 250 U/ml as compared to patients without immunologic factors and/or normal controls. Mean pollen-specific radioallergosorbent (RAST) binding was also significantly higher in patients with abnormal aggregation as compared to normal platelet responders (p = 0.02). Release of serotonin generally reflected abnormal aggregation patterns in asthmatic patients. Platelet factor 4 release was significantly decreased in the same groups of patients. These results suggest that the allergic state may affect platelet membrane responsiveness to multiple aggregating agents.     

Anti-fungal and cytokine producing activities of CD8 + T lymphocytes from HIV-1 infected individuals

Lymphokine activated killer (LAK) cells are capable of killing not only malignant cells but also hyphal form of Candida albicans in vitro. When peripheral blood mononuclear cells (PBMC) from normal healthy donors were cultured for 72-96 hrs with 1,500 international unit (IU)/ml interleukin-2 (IL-2), marked LAK activity was induced. However, even prior to IL-2 activation, PBMC isolated from some normal subjects and those from almost all individuals who are infected by human immunodeficiency virus type 1 (HIV-1) exhibited significant levels of anti-fungal activity. Such pre-activation ("in situ") antifungal activity of PBMC decreased during the initial 48 hrs of IL-2 activation. PBMC from HIV-1 seropositive subjects showed higher levels of "in situ" anti-fungal activity than normal PBMC did. After a decline of "in situ" activity during the initial 48 hours, LAK activity gradually increased and reached near maximal levels by day 4 and remained more or less constant until day 6. No significant difference was observed between the LAK activity of normal and HIV-1(+) PBMCs on days 4-6. In IL-2 activated normal and HIV-1(+) PBMC cultures, both CD4 and CD8 T cells produced IL-2, INF-gamma as well as TNF-alpha. Production of IL-2 by both CD4 and CD8 T cells was suppressed in HIV-1(+) PBMC cultures, but no significant suppression of INF-gamma production was noted. Meanwhile, TNF-alpha production by CD4 was very much suppressed but no significant changes in TNF-alpha production by CD8 T cells was noted in HIV-1(+) PBMC cultures.     

Factors influencing immediate responses to intradermal allergen administration in patients with respiratory allergy

BACKGROUND: Immediate skin reactions to allergens are influenced by several factors, such as the amount of administered allergen, the level of specific IgE, releasability of mast cells and hyperresponsiveness of the target organ. METHODS: For the evaluation of factors influencing immediate skin response to intradermal allergen administration, we measured the wheal size 15 min after intradermal injection of 0.01-0.02 ml of the following agents: whole-body extract of Dermatophagoides farinae, 1,000 allergy units/ml; histamine, 0.1 mg/ml, and codeine sulfate, 0.09% in saline, and determined total IgE level, specific IgE and IgG subclass antibodies to D. farinae in 53 patients with respiratory allergy. RESULTS: Multiple regression analysis for factors influencing wheal size after intradermal injection of D. farinae, specific IgE antibody level to D. farinae and wheal size after intradermal administration of histamine showed statistically significant results (R2 = 0.42739, p = 0.0000; R2 = 0.50243, p = 0.0185, respectively). Multiple regression analysis for factors influencing wheal size after intradermal administration in the group with high levels of specific IgE to D. farinae (RAST class 3 or more) showed that wheal size after intradermal administration of codeine was the only factor exerting a statistically significant influence (p = 0.0119). CONCLUSION: Based on the above results, we can state that immediate responses to intradermal allergen administration were influenced by the level of specific IgE and hyperresponsiveness of the target organ to histamine, but that the immediate skin allergic responses in the presence of high levels of specific IgE were partially but significantly influenced by the releasability of skin mast cells.     

Anti-IgE autoantibodies in asthma: a diagnostic artefact or an explanation for non-allergic asthma?

Autoantibodies to IgE can be detected in sera of individuals with atopic disease, but occasionally elevated levels are also found in sera of normal individuals. During the last years we studied therefore the functional properties of such autoantibodies. Depending on the studied in vitro system one can always detect minimally two different antibody types. Antibodies have been found that either trigger or inhibit mediator release from basophils, that enhance or inhibit binding of IgE to the low IgE receptor and either stimulate or inhibit human IgE synthesis. Based on such in vitro experiments one may conclude that also in vivo autoantibodies exist that either neutralize IgE or have no effect on IgE mediated clinical events. Thus, anti-IgE autoantibodies may hide IgE as for example in those bee sting allergic individuals where we could not detect specific IgE but found IgE hidden within immune complexes, suggesting that the biological activity of IgE was not neutralized. A similar phenomenon may exist in asthmatic individuals. In a recent study we found that non-atopic asthmatic children had indeed low levels of serum IgE, but showed the same levels of autoantibodies to IgE, against suggesting that IgE was hidden within immune complexes. Thus, our ongoing research addresses the question whether in diseases of unclear atopic origin IgE may nevertheless play a critical role but based on possible artefacts in the IgE detection assays some of the clinically relevant IgE may escape.     

Norepinephrine accelerates HIV replication via protein kinase A-dependent effects on cytokine production

In vivo, IgE production is related to bronchial hyperresponsiveness and, in vitro, passive sensitization of human airways with asthmatic serum containing a high concentration of IgE enhances the contractile response to a variety of agonists. However, cell types implicated in this IgE sensitization are not fully determined. The aim of this study was to determine IgE-bearing cells during passive sensitization with special reference to mast cells. Peripheral bronchi were dissected out from 10 lung specimens obtained at thoracotomy and processed into glycolmethacrylate resin. Sections, each 2 microm thick, were passively sensitized by incubation for 2 h at 37 degrees C in either buffer supplemented with monoclonal IgE or asthmatic serum with a high concentration of IgE (> or = 1,000 IU/ml). Immunohistochemistry was performed using monoclonal antibodies directed against the epsilon chain, and markers of the various IgE-bearing cells (e.g., AA1, antichymase). The number of IgE-bearing cells was significantly higher in passively sensitized specimens as compared with nonsensitized specimens (6.63 +/- 1.71 versus 4.29 +/- 1.35/mm2; p = 0.013, n = 10). Mast cells represented 65% of IgE-bearing cells, 41.6 and 23.4% for TC and T subtypes, respectively. These results indicate that mast cell is the main cell type involved in IgE-induced passive sensitization. The involvement of mast cell-derived tryptase in the mechanisms of IgE-related hyperresponsiveness should be further examined.     

Abortion in guinea pigs by Chlamydia psittaci isolates from natural sheep abortion

The effect of a novel anti-allergic drug, betotastine besilate (betotastine) on interleukin (IL)-5 production by human peripheral blood mononuclear cells (PBMC) was investigated. PBMC of Dermatophagoides farinae extract (Df)-sensitive donors produced IL-5 and showed a proliferative response upon stimulation with relevant antigen (10 microg/ml). Df-induced IL-5 production by PBMC was significantly inhibited by betotastine at 10 and 100 microM. Betotastine also suppressed proliferation of PBMC with less potency. The effect of betotastine on IL-5 production was enhanced and significant even at 0.1 microM when the drug was added 120 min before antigen stimulation. Ketotifen and cetirizine also inhibited IL-5 production, but the effects of these drugs were significant only at 100 microM. These findings indicate that the suppression of IL-5 production may be involved in the anti-allergic effect of betotastine.     

Antibody response to outer membrane protein of nontypeable Haemophilus influenzae in otitis-prone children

One of the major outer membrane proteins of nontypeable Haemophilus influenzae, P6, is highly conserved among strains, serves as a target for bactericidal antibody, and has been proposed as a possible vaccine candidate. The serum antibody response to P6 was studied in otitis-prone and normal children by an enzyme-linked immunosorbent assay. Of 20 otitis-prone children, 12 (60%) had a serum IgG antibody response to P6 after otitis media; however, the mean acute antibody level for the group, 4.6 micrograms/ml, was not significantly different from the convalescent level, 5.4 micrograms/ml. Anti-P6 antibody levels were also measured longitudinally for 10 to 25 months in 30 otitis-prone and 13 healthy children. Antibody levels increased sevenfold in the normal group compared with less than three-fold for the otitis-prone group and were significantly higher in the normal children after the age of 18 months (p < 0.05). Finally, otitis-prone children who had two or more episodes of otitis media with nontypeable H. influenzae did not have an anamnestic antibody response to P6. The failure to recognize P6 as a specific immunogen may account for recurrent infections. Moreover, the data suggest that otitis-prone children may not respond adequately to a vaccine containing P6.     

Keeping people well despite life change crises

IgE was found in urine from healthy adult volunteers at very low levels (approximately 0.003-0.010 IU/ml), corresponding to a 24-h excrection rate of 3-16 IU. IgE was not detected in 36 out of 47 samples of milk and colostrum. In samples from six women the protein was present at low concentration 1-2 days postpartum but was not detected in later samples (usually day 5 or 6). Mammary secretions from four allergic donors were studied, and IgE was detected at low concentrations in samples from the two most severely affected individuals. The levels of IgE observed in both urine and milk suggest that there is no significant synthesis of the protein in either the urinary tract or in mammary tissue.     

Lactate metabolism in the dog during shock from hemorrhage, cardiac tamponade or endotoxin

Serum IgE concentrations of patients with paragonimiasis were determined by a radioimmunosorbent test. The mean concentration was 3,462.3 IU/ml in a group of 13 cases of paragonimiasis miyazakii in which patients showed clinical symptoms and/or positive immunological diagnostic tests, and 1,026.6 IU/ml in a control group of 13 individuals who had eaten uncooked freshwater crabs, Potamon dehaani, but had been found to be free from the infection. Moreover, the IgE level of the pleural exudates obtained from four patients with paragonimiasis miyazakii on the day of bleeding or within several days after was significantly higher than that of their sera, ranging between 4,200 IU/ml and 10,000 IU/ml. This was true also in a case of paragonimiasis westermani. Sera and pleural exudates of patients with both forms of paragonimiasis were applied to immunosorbent columns of Sepharose 4B beads coupled with saline extracts of Paragonimus miyazakii, P. weetermani, or P. ohirai. IgE eluted from the corresponding column was considered to be specific, being around 5% to 10% of the total IgE.