Cyclic Adenosine Monophosphate-Mediated Enhancement of Vascular Endothelial Growth Factor Released by Differentiated Human Monocytic Cells: The Role of Protein Kinase A.

OBJECTIVE: Our investigation was designed to examine the signaling pathway involved in the enhancement of vascular endothelial growth factor (VEGF) release by ß-adrenoceptor agonists. MATERIALS AND METHODS: Human U937 cells differentiated into macrophages were primed with lipopolysaccharide (LPS) in the absence or presence of ß-adrenoceptor agonists and antagonists. The VEGF released and the intracellular cyclic adenosine monophosphate (cAMP) generated were assayed by ELISA. Where necessary, differences between mean values were tested for significance using Student's t test. RESULTS: Isoprenaline, procaterol and salbutamol concentration-dependently enhanced the release of VEGF induced by LPS in U937 cells. R*,R*-(±)-4-[2-[(2-(3-chlorophenyl)-2-hydroxyethyl)amino]propyl]phenoxyacetic acid (BRL 37344), a selective ß3-adrenoceptor agonist, did not enhance VEGF release. Using isoprenaline as an agonist, propranolol, ICI 118551 and atenolol produced a parallel rightward shift of the concentration-response curve with no reduction in the maximum response. The -logKB values were 8.12 ± 0.17, 8.03 ± 0.05 and 7.23 ± 0.05 for propranolol, ICI 118551 and atenolol, respectively, indicating the possible involvement of both ß1- and ß2-adrenoceptor subtypes. Isoprenaline and prostaglandin E2 concentration-dependently increased cAMP generation in U937 cells. Isoprenaline, db-cAMP and 6-Bnz-cAMP, a protein kinase A (PKA) activator, all enhanced VEGF release induced by LPS, and this effect was abolished by KT 5720 and Rp-cAMPS, which are both selective PKA inhibitors, suggesting that PKA is the downstream effector of cAMP activity. 8-CPT-cAMP, a selective activator of the Epac system, had no effect on VEGF release induced by LPS, indicating that the Epac pathway played no role in the release process. CONCLUSION: In this study, we established that ß1- and ß2- but not ß3-adrenoceptors mediated cAMP-dependent enhancement of VEGF release induced by LPS in differentiated U937 cells, and that PKA was the downstream effector of cAMP activity.

The role of adenosine A(2) receptors in the regulation of TNF-alpha production and PGE(2) release in mouse peritoneal macrophages.

The adenosine A(2) receptors are known to mediate most of the anti-inflammatory activities of adenosine. In lipopolysaccharides (LPS)-stimulated macrophages adenosine strongly inhibits TNF-alpha release, but may also enhance PGE(2) generation. The aims of this study were to determine the relative contributions of the A(2A) and A(2B) receptor subclasses in these two effects and to determine whether the enhanced release of PGE(2) contributes to the inhibition of TNF-alpha release. In LPS-stimulated mouse macrophages, adenosine potently inhibited TNF-alpha production and also potentiated PGE(2) release, though less potently (IC(50)=250 nM vs EC(50) approximately 8 microM, respectively). The non-selective adenosine receptor agonist NECA, and the selective A(2A) receptor agonist CGS21680 also inhibited TNF-alpha production even more potently (IC(50)=4.8 and 2.3 nM, respectively). NECA, but not CGS21680, also enhanced PGE(2) production. The selective A(2A) receptor antagonist ZM241385 (30 nM), but not the selective A(2B) receptor antagonist MRS1754 (30 nM), blocked the inhibitory effect of NECA and CGS21680 on TNF-alpha release. On the other hand, MRS1754, but not ZM241385, abolished the PGE(2) potentiating effect of NECA. Pre-treatment with indomethacin (1 microM) abolished adenosine-induced PGE(2) release enhancement but did not prevent the inhibition of TNF-alpha release. These results show that in this system, the inhibition of TNF-alpha release by adenosine is mediated by the A(2A) receptors whereas the enhancement of PGE(2) release appears to be mediated by the A(2B) receptors. The results also show that while exogenous PGE(2) is a potent inhibitor of TNF-alpha release, the enhanced PGE(2) release induced by adenosine does not appear to contribute to the inhibition of TNF-alpha release.

Characteristics of asthmatic children in Kuwait.

This article summarizes clinical characteristics and identifies sensitizing allergens in 135 asthmatic children under 13 years of age in Kuwait, a desert environment with scant vegetation and weather conditions least associated with asthma. There were 84 males (M:F 1.65:1). Almost 70% were breast-fed (1-24 months), 59% had eczema, 52% allergic rhinitis, 78% of first-degree relatives had atopy, and 52% of parents were consanguinous. Cough was the presenting symptom in 92% and together with wheezing occurred in 76%. Most (91%) were < or = 5 years of age at diagnosis and 42% < 2 years. Mean duration of symptoms prior to diagnosis was 9.3+/-2 months (1 week-1 year). Viral upper respiratory tract infections, cigarette smoke, and exercise were the commonest triggers of symptoms (79%, 68%, 62%). Fumes of traditional Bokhour (incense) constituted a major indoor hazard. The most common sensitizing allergens were pollens of imported plants, molds, house dust mites, cockroaches, and peanuts. Management showed considerable under-treatment and included alternative medicines. In conclusion, childhood asthma in this desert environment starts at an early age, and is associated with high rate of atopy and high frequency of sensitization to aero- and food allergens. Asthmatic children are disadvantaged by delay in diagnosis, undertreatment, exposure to indoor cigarette smoke, and local traditions.

Effect of wortmannin on human eosinophil responses in vitro and on bronchial inflammation and airway hyperresponsiveness in Guinea pigs in vivo.

Many mediators activate eosinophils via transduction pathways involving the enzyme phosphatidylinositol 3-kinase. The initial investigation of wortmannin, a specific inhibitor of PI3-kinase, was of its effect on human and guinea pig eosinophil superoxide (O(2)(-)) release and degranulation in vitro. Subsequently, the effect on allergen- and Sephadex-induced bronchial inflammation and airway hyperresponsiveness (AHR) in vivo in guinea pigs was investigated. Wortmannin potently inhibited complement C5a-induced O(2)(-) generation and eosinophil peroxidase (EPO) release from human eosinophils, with 50% inhibition produced by a 1-10 nM concentration. Both aerosol allergen challenge of sensitized guinea pigs and intravenous injection of Sephadex beads in normal guinea pigs caused, in 24 h, significant eosinophilia and increased EPO activity in bronchoalveolar lavage fluid (BALF) and AHR to intravenous acetylcholine and histamine. In the allergic model, intranasal pretreatment with wortmannin had no effect on BALF eosinophilia, but dose dependently inhibited BALF EPO activity. At 1 mg/kg, the drug abolished the AHR to histamine, but not acetylcholine. In the Sephadex model, the drug significantly inhibited all three parameters (eosinophilia, increased EPO activity, and AHR to both spasmogens). These results show that wortmannin is a potent inhibitor of human eosinophil degranulation and that when administered intranasally can prevent AHR in allergen-challenged guinea pigs, probably by inhibiting eosinophil degranulation, but not their accumulation in BALF. This may be relevant to the possible clinical utility of wortmannin in conditions involving eosinophilic inflammation and AHR.

Involvement of A(3) receptors in the potentiation by adenosine of the inhibitory effect of theophylline on human eosinophil degranulation: possible novel mechanism of the anti-inflammatory action of theophylline.

The current use of theophylline in asthma is based on both the bronchodilatory and the anti-inflammatory effects. The exact mechanism of these actions is still controversial and may include the inhibition of adenosine 3',5'-monophosphate phosphodiesterase enzyme (PDE) and antagonism of adenosine receptors. In this study, the mechanism of the anti-inflammatory action was investigated by studying the inhibition by theophylline of complement C5a (C5a)-induced degranulation of human eosinophils and its interaction with adenosine. Theophylline (10-1000 microM) inhibited C5a-induced release of eosinophil peroxidase (EPO) in a concentration-dependent manner with an IC(50) of 233.5 microM and a maximal inhibition of 90.3 +/- 3.0%. In contrast, the PDE4 inhibitor rolipram (up to 50 microM) had no effect. The adenosine A(3) receptor agonist N(6)-(3-iodobenzyl)-5'-N-methylcarbamoyladenosine (IB-MECA) also inhibited release (IC(50) = 7.5 microM), but neither adenosine itself nor the selective A(1) and A(2) agonists and antagonists had any significant effect, even at 100 microM. The inhibition produced by clinically relevant concentration of theophylline (50 microM) was potentiated by ineffective concentrations of exogenous adenosine and additive to that produced by IB-MECA. The potent and selective A(3) antagonist MRS 1220, but not the A(1) or A(2) antagonists, significantly reversed the inhibitory effect of theophylline. These results suggest that therapeutic concentrations of theophylline inhibit human eosinophil partly by acting as an A(3) agonist. Together with the potentiation of theophylline action by adenosine, perhaps via the A(3) receptors, these novel actions may, at least in part, contribute to the mechanism of the anti-inflammatory action of this drug in vivo.

Requirement of additional adenylate cyclase activation for the inhibition of human eosinophil degranulation by phosphodiesterase IV inhibitors.

Human eosinophils contain predominantly phosphodiesterase type IV, but selective inhibitors of this isoenzyme fail to inhibit certain eosinophil responses such as degranulation. In this study, the effect of activation of adenylate cyclase on the ability of several highly selective PDE IV inhibitors to inhibit complement C5a-induced O2- release and degranulation of human eosinophils in vitro was investigated. All four selective PDE IV inhibitors, N-(3,5-dichloropyrid-4-yl)-3-cyclopentyl-oxy-4-methoxybenzamide (RP 73401), rolipram, N-(3,5-dichloropyrid-4-yl)-[1-(4-fluorobenzyl)-5-hydroxy-indol-3-yl]glyoxylacidamide (AWD 12-281) and c-4-cyano-4-(3-cyclopentyloxy-4-methoxyphenyl-r-1-cyclohexane carboxylic acid) (SB 207499) potently inhibited C5a-induced O2- generation (IC50 = 0.03, 0.42, 0.55 and 0.86 microM, respectively), but generally failed to inhibit degranulation. The only exception was AWD 12-281, which inhibited degranulation (IC50 = 16.2 microM). In the presence of different AC activators (histamine, salbutamol, prostaglandin E2 and forskolin), the PDE IV inhibitors became potent inhibitors of degranulation. The interaction between the PDE IV inhibitors and the AC activators resulted in a synergistic increase in intracellular levels of adenosine 3', 5'-monophosphate (cAMP). These results show that PDE IV inhibitors generally require an additional cAMP signal to be able to inhibit eosinophil degranulation, and that this signal can be generated via both membrane receptors and direct AC activation. This may be relevant to the in vivo effectiveness of PDE IV inhibitors in eosinophilic inflammation.

Therapy of bronchial asthma with adenosine receptor agonists or antagonists.

Adenosine is an endogenous nucleoside that is released under pathological conditions and interacts with four G-protein-coupled receptor subtypes. These receptors are widely distributed throughout the body. They are involved in many central and peripheral processes, including immunological and inflammatory responses. In inflammatory and asthmatic conditions, the extracellular concentration of adenosine increases in the airway tissue. It enhances mast cell degranulation and bronchoconstriction, but may also inhibit eosinophil or lymphocyte function or modulate reactive oxygen species generation in neutrophils. Despite a large number of studies clearly indicating the effects of adenosine in vitro, many aspects of the mechanisms involved in the adenosine-mediated responses are still unclear, and our knowledge is limited in understanding the complex multifactorial interactions occurring in the whole body. The discovery of adenosine receptor compounds acting with increasing selectivity will bring new approaches to the use of adenosine receptor agonists and antagonists and may clarify some of the current uncertainties. On the basis of our present knowledge, the development of adenosine A(2A)- or (A3)-receptor agonists as antiinflammatory agents or A(2B)-receptor antagonists as inhibitors of mast cell degranulation for the treatment of asthma holds promise.

Histamine H(2) receptors mediate the inhibitory effect of histamine on human eosinophil degranulation.

The effect of histamine on human eosinophil degranulation and the receptor mediating such effect were studied in vitro using the complement C5a-mediated eosinophil peroxidase (EPO) release model. Following pre-treatment with 5 microg ml(-1) cytochalasin B(CB), C5a induced a concentration-dependent release of EPO from eosinophils isolated from healthy donors. Histamine (0.1-50 microM), but not L-histidine, inhibited concentration-dependently C5a-induced EPO release with IC(50) (95% CI) of 0.6 microM (0.3-1.2 microM) and maximal inhibition of approximately 60%. A similar effect was seen with the selective H(2) agonists dimaprit (IC(50) (95% CI)=6.9 microM (3.2-10.6 microM)) and amthamine (IC(50) (95% CI)=0.4 microM (0.2-0.7 microM)). Neither the selective H(1) agonist 6-(2-(4-imidazolyl)ethylamino)-N-(4-trifluoromethylphenyl) heptanecarboxamide(HTMT), nor the selective H(3) agonists imetit (up to 100 microM) had any significant effect. The inhibition by histamine was reversed by cimetidine (0.1-30 microM) and other H(2) antagonists, but not the H(1) antagonist mepyramine (1.0- 100 microM), nor the H(3) antagonist thioperamide (1.0-100 microM). Cimetidine (1-30 microM) shifted to the right the dimaprit log dose-response curve, producing a pA(2) value of 5.9 and Schild's plot slope of 0.98, thus confirming simple competitive antagonism. Histamine (10-100 microM) increased intracellular level of adenosine 3',5'-cyclic monophosphate, which was completely abolished by cimetidine (30 microM), but not mepyramine or thioperamide. The cyclic AMP analogue - dibutyryl cyclic AMP - also inhibited degranulation (IC(50) approximately 300 microM). The cyclic AMP phosphodiesterase(PDE) IV inhibitor rolipram (10 microM) synergistically enhanced the inhibition of EPO release by histamine. These results suggest that histamine, via stimulation of H(2) receptors and a consequent elevation of intracellular levels of cyclic AMP, inhibits human eosinophil degranulation.

The association of skin test reactivity, total serum IgE levels, and peripheral blood eosinophilia with asthma in Kuwait.

There is evidence that elevated serum immunoglobulin E (IgE) and eosinophilia correlate well with allergic skin test reactivity. These parameters have been used as alternative methods to characterize atopic subjects. Skin test reactivity is the only measure used routinely in clinical practice in Kuwait to reflect atopy in asthma patients. This study examines the usefulness of the two other parameters of atopy in patients with asthma, and to determine the most common allergens involved in Kuwait. Between 1998 and 1999, 101 asthma patients and 33 healthy controls were recruited for this study. Skin sensitivity test, serum total and specific IgE, total blood eosinophil count (B-EOS), and eosinophil cationic protein (ECP) tests were performed in patients and controls. Nine allergens known to be prevalent in this environment were selected for the skin test and specific IgE test. Spirometry was also measured. These parameters were repeated after 4 weeks of therapy in the patients only. Skin test reaction was positive in 81% of the patients, while total IgE above 200 kU/L was obtained in 63% of cases. B-EOS above 300 x 10(3)/L was found in 75% of cases. House dust mite reactivity (positivity) was the most frequently encountered skin allergy, occurring in 28% of the patients. IgE correlated positively with B-EOS and ECP. B-EOS similarly correlated positively with ECP. There was a negative correlation between ECP and forced expiratory volume in 1 sec (FEV1) (% predicted) as expected. At least one positive parameter of atopy was found in 95% of the patients. In 48% of the patients, all three parameters of atopy were found to be positive. Skin test reactivity and elevated IgE were found together in 62% of the cases. This study reveals a significant degree of allergy among patients with asthma in this environment. Skin testing was found to be the most effective measure of atopy in this environment, and correlates well with the other more sensitive newer tests.

IgE-mediated sensitization to mould allergens among patients with allergic respiratory diseases in a desert environment.

BACKGROUND: The importance of fungal allergens in the development of allergic diseases in a desert environment is uncertain. This study evaluated the prevalence of IgE sensitization to moulds among patients with allergic respiratory diseases in Kuwait - a desert country. METHODS: A total of 810 patients (male:female ratio 1.4) with a mean age of 32.3 years (range 2-76 years) with extrinsic asthma or allergic rhinitis were studied. Sera from the patients were tested by the CAP-RAST method for specific IgE to 6 fungi (Penicillium, Cladosporium, Aspergillus, Candida, Alternaria and Helminthosporium). For comparison house dust mite and Bermuda grass were also assessed. RESULTS: The overall positivity to at least one mould was 20.9%. Among 120 matched control subjects, the value was 5. 8%. The value was much higher among patients with asthma alone (45. 8%) or both asthma and rhinitis (28.3%) than those with rhinitis alone (11.8%; p < 0.001). Asthmatic children had the highest sensitization rate (66.0% in the 7- to 12-year age group), which declined sharply with age. Among asthmatics, Candida and Aspergillus had the highest sensitization rates (23.1 and 21.3%, respectively), followed by Helminthosporium (18.8%), Cladosporium (15.9%), Alternaria (14.6%) and Penicillium (13.9%). The values for mite and Bermuda grass were 41.2 and 54.6%, respectively. Among asthmatic children, severe asthma was significantly more frequent among mould-positive (51.6%) than mould-negative patients (17.5%; p < 0. 0001). CONCLUSIONS: Even in this desert environment, sensitization to moulds is quite common among patients with allergic respiratory diseases, with a striking preponderance among children with asthma. Mould allergy could also be an important factor determining asthma severity in this environment.

Sensitization to aeroallergens among patients with allergic rhinitis in a desert environment.

BACKGROUND: The causative allergens of allergic rhinitis in desert environments are uncertain. OBJECTIVE: To determine the sensitizing aeroallergens in patients with allergic rhinitis in Kuwait, a desert country. METHODS: A total of 706 patients aged 6 to 64 years (mean 34.3 years) with allergic rhinitis were studied. Sera from the patients were screened for specific IgE to 14 inhalant allergens by the CAP-RAST method. RESULTS: Specific IgE to any allergen was detected in 86.3% of patients. The prevalence rates for allergen groups were: pollens (77.3%), house dust (62.3%), and molds (14.7%). The individual allergens with the highest positive rates were pollens of the weed Chenopodium (64.3%); Bermuda grass (55.0%), and Prosopis tree (50.3%). These plants were all imported and cultivated for the purpose of "greening" the desert. German cockroach (48.2%) and house dust mites (32.4% to 39.2%) were the most prevalent indoor sensitizers. With the exception of the molds, sensitization rates were higher for males than females. The youngest age group (6 to 17 years) had significantly higher sensitization rates than the older ones, particularly with respect to the molds (P < .01 to .001). Severe sensitization was more common with Alternaria than the other allergens and in general mold sensitization was more frequently associated with severe symptoms. Polysensitization was very common, with 81.8% of all sensitized patients positive to more than one allergen. CONCLUSIONS: Pollens of the local horticultural plants are the main sensitizing allergens among patients with allergic rhinitis in this desert environment. The practices that "green" the desert seem to also encourage allergen sensitization.

Allergen-induced bronchial eosinophilia in guinea-pigs is inhibited by both pre- and post-induction cyclosporin-A treatments.

Repeated treatment of sensitized guinea-pigs with cyclosporin-A (CS-A) before aerosol allergen challenge is known to inhibit the subsequent bronchial eosinophilia. It is not known, however, if the drug is also effective on established/on-going bronchial eosinophilia. We have, therefore, studied the effect of CS-A on allergen-induced eosinophil recruitment into the bronchoalveolar lavage (BAL) fluid of guinea-pigs when given before or after induction.Ovalbumin-immunized guinea-pigs were treated with CS-A (20 mg/kg subcutaneously) or vehicle daily for varying periods before a single aerosol allergen challenge. In animals in which bronchial eosinophilia was maintained with repeated aerosol allergen challenge, CS-A or vehicle was given daily for varying periods after the first allergen challenge. BAL and cell count were performed 24 h after the last challenge. In vehicle-treated animals, a single allergen challenge caused a 4-5 fold increase in the number of eosinophils in the BAL fluid after 24 h, declining to baseline by 7 days. In repeatedly-challenged animals, this response was sustained throughout. Eosinophil infiltration was significantly inhibited when CS-A was given daily for 7-14 days, but not for 1 or 3 days, before allergen challenge. When given during an established/on-going eosinophil infiltration, a significant inhibition was seen after administration for 5 or 7 days, but not for 1 or 3 days.These results show that repeated CS-A administration inhibits not only the induction of allergic bronchial eosinophilia but also the maintenance of an established one. This may be relevant in the treatment of allergic diseases, such as asthma, in which drug administration often begins when eosinophilia is already established.

Asthma in the desert: spectrum of the sensitizing aeroallergens.

BACKGROUND: Bronchial asthma is common in Kuwait, a desert country, but the sensitizing allergens are uncertain. This study investigated the sensitizing allergens in Kuwaiti patients with extrinsic asthma. METHODS: A total of 553 asthmatics (male:female ratio: 1.4; mean age: 31.7 years [range 3-76 years]) and 112 matched controls were studied. Sera from all patients/subjects were tested by the CAP-RAST method for specific IgE to 14 locally relevant inhalant allergens. RESULTS: Specific IgE to at least one allergen was detected in 87.2% of the patients compared with 24.1% of controls. Among the confirmed extrinsic asthmatics, the sensitization rates for the allergen groups were as follows: pollens (87.1%), house dust (76.1%), and molds (30.3%). The three most prevalent sensitizing pollens were from Chenopodium (70.7%), Bermuda grass (62.9%), and Prosopis (62.7%), all of which are horticultural plants imported for the purpose of "greening" the desert. For all allergens, except the molds, the prevalence rate was higher in males than females, but age had only a weak effect. Severe asthma occurred significantly more frequently among mold-sensitized patients. CONCLUSIONS: These results show that even in a desert environment, pollens and house-dust allergens may be important sensitizing allergens. They also illustrate how practices that "green" the desert can affect public health.

Adenosine A3 receptors on human eosinophils mediate inhibition of degranulation and superoxide anion release.

The role of adenosine A3 receptors on human eosinophil degranulation and superoxide anion (O2-) release was studied in vitro using the complement fragment C5a as the main stimulus and employing a number of selective agonists and antagonists. In the presence of cytochalasin B (CB), C5a induced a dose-dependent release of the granular eosinophil peroxidase (EPO), but not O2-, whereas in the absence of CB O2- , but not EPO, was released. C5a-induced EPO release was inhibited dose-dependently by the selective A3 agonist N6-(3-iodobenzyl)-5'-N-methylcarbamoyladenosine (IB-MECA) and to a lesser extent by the less-selective N6-2-(4-amino-3-iodophenyl) ethyladenosine (APNEA). The IC50 (95% CI) for IB-MECA was 6.8 microM (3.1-12.0 microM). At concentrations up to 100 microM, neither adenosine nor the selective A1 agonist N-cyclopentyladenosine (CPA) and the selective A2 agonist 2-[[2-[4-(2-carboxyethyl)phenyl]ethyl]amino]-N-ethylcarboxamidoadenosine (CGS 21680) had any significant effect. The inhibitory effect of IB-MECA was almost completely abolished by pre-treatment with 1 microM of the selective A3 antagonist 9-chloro-2-(2-furyl)-5-phenylactylamino[1,2,4]triazolo[1,5-c]quina zoline (MRS 1220), but not the selective A1 antagonist 1,3-dipropyly-8-cyclopentylxanthine (DPCPX) or the selective A2 antagonist 3,7-dimethyl-1-propargylxanthine (DMPX). IB-MECA also significantly inhibited C5a-induced O2- release with IC50 (95% CI) of 9.5 microM (4.6-13.1 microM) whereas adenosine and the A1 agonist CPA potentiated this effect at low concentrations. The potentiation appeared to be a result of their direct O2- release from these cells, probably mediated via A1 receptors. The inhibition by IB-MECA was selectively reversed by MRS 1220. These results show that the A3 receptors on human eosinophils mediate inhibition of both degranulation and O2- release and suggest a therapeutic potential for A3 agonists in diseases such as asthma in which activated eosinophils are involved.

Reference values of total serum IgE and their significance in the diagnosis of allergy among the young adult Kuwaiti population.

BACKGROUND: The reference total serum immunoglobulin (IgE) values and the usefulness of total IgE values in the diagnosis of allergy have not been established for the Kuwaiti population. The literature reference values may not be applicable since such values often vary among ethnic nationalities. OBJECTIVE: The aim of this study was to establish the reference IgE values for the young adult Kuwaiti population and to determine the usefulness of such values in the diagnosis of allergic diseases in the community. METHODS: A total of 1057 randomly selected young adults were screened for atopy using the Pharmacia CAP-PhadiatopR method. Atopy was detected in 423 individuals (40.0%). Total serum IgE was then measured in 542 randomly selected Phadiatop-negative (non-atopic) cases in the age range 18-50 years (mean 28.9 years) and male:female ratio of 1.3. RESULTS: Serum total IgE values in non-atopics covered a very wide range (< 2-1993 kU/L) with a geometric mean (GM) value of 43.7 kU/L. The reference range, calculated as the 95% confidence interval of the log IgE (95% CI) was 3.2-602.5 kU/L. The 90% CI was 11.7-162 kU/L. The GM was significantly higher for males than females, (53.7 vs. 35.5 kU/L, P < 0.001) and for smokers than non-smokers, (64.6 vs. 40.7 kU/L, P < 0.01), but was independent of age. Although the GM for the non-atopics (43.7 kU/L) was significantly lower than those of the asymptomatic atopics (213.8 kU/L) and allergic asthmatics (626.6 kU/L), the 95% CI for the three groups showed considerable overlap. CONCLUSIONS: These results show that the normal total IgE values in the young adult Kuwaiti population are generally high and that the distribution of the values is so wide that the diagnostic value of total serum IgE in this community is likely to be very limited.

Serum eosinophil cationic protein as a predictor of disease activity in acute and chronic asthma.

BACKGROUND: Eosinophils may contribute to airway hyper responsiveness in asthma through the effects of eosinophil derived granular proteins in the bronchial epithelium. Increased concentration of eosinophil cationic protein (ECP) has been reported in patients with acute and chronic asthma. OBJECTIVE: To examine if ECP can serve as a marker of disease activity in acute and chronic asthma patients. DESIGN: Prospective case control study. PATIENTS: Sixteen non smoking asthmatics in exacerbation (group 1); twenty two in relatively stable state (group 2); and sixteen normal control subjects (group 3) were recruited into the study. SETTING: Casuality and outpatients departments, Mubarak hospital, Kuwait between August 1997 and July 1998. MAIN OUTCOME MEASURES: The mean serum ECP, blood eosinophil count and peak expiratory flow rate (PEFR). RESULTS: There was a statistically significant difference between the groups in blood eosinophil count (p < 0.01) and in PEFR (p < 0.0001). At week four, the mean ECP and blood eosinophil count fell as a result of therapy in group 1. The difference in PEFR values between week 0 and 4 in group 1 reached statistical significance (p < 0.05). In group 2 patients, the mean serum ECP, blood eosinophil count and PFER values between week 0 and 4 did not show any significant difference. A correlation was observed between ECP and PEFR in group 1 (p < 0.05) and between ECP and eosinophil count in group 2 (p < 0.01). CONCLUSION: Serum ECP has the potential to serve as a marker for predicting and monitoring the clinical course of asthma. Further studies are required to verify these baseline findings in our environment.

Effects of some anti-asthma drugs on human eosinophil superoxide anions release and degranulation.

BACKGROUND: Eosinophil infiltration of bronchial tissues and subsequent release of inflammatory mediators by them are the hallmarks of bronchial asthma but it has not yet been clarified whether anti-asthma drugs affect these cells directly. In this study, we investigated the direct effects of 8 clinically used anti-asthma drugs [salbutamol, salmeterol, theophylline, denbufylline, disodium cromoglycate (DSCG), azelastine, ketotifen and dexamethasone] on superoxide anions (O2-) and eosinophil peroxidase (EPO) release from human blood eosinophils in vitro. METHODS: Highly purified eosinophils were stimulated for O2- release with platelet-activating factor (PAF) or interleukin-5 (IL-5), while for EPO release complement fragment (C5a) or N-formyl-methionyl-leucyl-phenylalanine (FMLP) was employed. Generated products were assayed by standard techniques. RESULTS: All the drugs, except ketotifen and dexamethasone, inhibited PAF-induced O2- release in a dose-dependent manner. The IC50 values were 0.7, 5.8, 330, 3,500, 4,200 and 6,250 nM for DSCG, denbufylline, salmeterol, azelastine, salbutamol and theophylline, respectively. On IL-5-induced release, the effects were similar except that salbutamol completely failed to inhibit the release induced by this stimulus. In contrast, EPO release was generally poorly inhibited, especially when the release was induced by C5a. Only theophylline and azelastine (both at 10(-4) M or more) were able to inhibit EPO release by both C5a and FMLP. Salbutamol and, to a lesser extent, salmeterol inhibited FMLP-, but not C5a-induced EPO release, while all the other drugs tested were inactive. CONCLUSIONS: The results show that some of the anti-asthma drugs, but not all, do exert direct effects on human blood eosinophils but these effects may be stimulus-dependent and by far more pronounced against O2- release than against degranulation.

Differential effects of salbutamol and salmeterol on human eosinophil responses.

In the treatment of bronchial asthma, salmeterol is believed to have a greater anti-inflammatory activity than salbutamol. To determine whether the comparative effects of these drugs on eosinophil function are the basis of their differential anti-inflammatory properties, we studied the effect of the two drugs on interleukin-5 (IL-5) and 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine (PAF)-induced O2- release and adherence to fibronectin-coated plates, as well as the C5a- and N-formylmethionyl-leucyl-phenylalanine (FMLP)-induced degranulation of purified human blood eosinophils in vitro. Salmeterol significantly inhibited IL-5-induced O2- release in a concentration-dependent manner with an IC50 of 2.2 X 10(-6) M (95% CI, 1.6-2.7 X 10(-6) M) and a maximal inhibition of about 70%. In contrast, salbutamol had no significant effect even at 10(-5) M. Both drugs significantly inhibited PAF-induced O2- generation, but salmeterol was approximately 20 times more potent than salbutamol. Salmeterol also significantly inhibited adherence induced by both IL-5 and PAF, whereas salbutamol had no significant effect on adherence induced by both agents. Both drugs failed to block C5a-induced eosinophil peroxidase release, whereas for FMLP-induced release, salbutamol, but not salmeterol, produced significant inhibition. Unlike salbutamol, all the actions of salmeterol were independent of beta-2 adrenoceptors. These results confirm that human eosinophils can be modulated directly by beta-2 adrenoceptor agonists, but that salmeterol and salbutamol have differential effects which depend on both the stimulus used and the response being measured and that the reportedly greater in vivo anti-inflammatory effect of salmeterol may reflect its superior ability to inhibit eosinophil O2- release and adherence, rather than degranulation.

The effect of salmeterol on human eosinophils is both stimulus- and response-dependent.

Salmeterol, a long-acting beta 2-adrenoceptor agonist, also possesses some anti-inflammatory properties, but whether eosinophils are the target of such action has been equivocal. To clarify the direct effect of salmeterol on eosinophil functions, we have studied the effect of the drug on the various responses of purified human eosinophils. Superoxide anions (O2-) release and adherence to fibronectin-coated plastic plates induced by platelet-activating factor (PAF), interleukin-5 (IL-5), leukotriene B4 (LTB4) and phorbol myristate acetate (PMA), as well as degranulation induced by C5a and formyl methionyl leucyl phenylalanine (FMLP), in the presence of cytochalasin B (CB) were studied. In the concentration range 10(-8)-10(-5) M, the drug inhibited PAF- and IL-5-induced O2- release, with an IC50 values of 3.2 +/- 1.2 x 10(-7) M and 2.2 +/- 0.4 x 10(-6) M, respectively, Superoxide anion release by LTB4 was only modestly inhibited while that due to PMA was completely unaffected. On the other hand, eosinophil adherence induced by all the 4 stimuli were significantly inhibited within the same concentration range. On eosinophil degranulation, the drug failed to significantly inhibit the release of eosinophil peroxidase (EPO) induced by either C5a or FMLP. In contrast, beta-hexoseaminidase (beta-HA) release by the same agents was significantly inhibited, the inhibition being more pronounced for FMLP-induced, than C5a-induced release. None of the effects of the drug was reversed by the selective beta 2-adrenoceptor antagonist ICI 118551 at a concentration of 10(-7) M. These results show that salmeterol may have some direct inhibitory effects on human eosinophil functions but that these effects are both stimulus- and response-dependent, and are unlikely to be mediated via beta 2 adrenoceptors.