Conversion of leukotriene A4 by neutrophils and platelets from patients with atopic dermatitis.

The generation of arachidonic acid-derived inflammatory mediators from unstimulated and stimulated neutrophils (PMN) and platelets in the presence of exogenous LTA4 has been studied in patients with atopic dermatitis (AD) as well as in healthy volunteers. PMN were stimulated with the interleukins IL-3, IL-8, C5a, and the Ca-ionophore A23187. In addition, NaF and thrombin were used to stimulate platelets. The release of leukotriene (LT)B4, 20-COOH- and 20-OH-LTB4, cysteinyl-leukotrienes and 12-HETE was measured. The proinflammatory mediator release from PMN and platelets of patients with AD was significantly higher as compared to the control group. The spontaneous conversion of LTA4 by PMN and platelets was markedly enhanced in patients with AD. Different results with receptor-specific and non-specific stimuli (Ca-ionophore A23187) in the presence of exogenous LTA4 were obtained. The results indicate a higher state of activation for enzymes involved in leukotriene formation. Furthermore, the production of 12-HETE by platelets from patients with AD was enhanced in unstimulated and stimulated cells. Our data emphasize that neutrophils and platelets may play an important role in the pathogenesis of AD by an increased responsiveness to receptor-specific stimuli and cell-cell interaction via LTA4.     

New yeast species, Malassezia dermatis, isolated from patients with atopic dermatitis

Malassezia species are considered to be one of the exacerbating factors in atopic dermatitis (AD). During examination of the cutaneous colonization of Malassezia species in AD patients, we found a new species on the surface of the patients' skin. Analysis of ribosomal DNA sequences suggested that the isolates belonged to the genus MALASSEZIA: They did not grow in Sabouraud dextrose agar but utilized specific concentrations of Tween 20, 40, 60, and 80 as a lipid source. Thus, we concluded that our isolates were new members of the genus Malassezia and propose the name Malassezia dermatis sp. nov. for these isolates.     

Decreased release of leukotriene B4 from monocytes and polymorphonuclear leukocytes in patients with systemic lupus erythematosus

The leukotriene B4 (LTB4) releasing capacities of monocytes and polymorphonuclear leukocytes (PMN) were studied using a specific radioimmunoassay in 28 patients with systemic lupus erythematosus (SLE) and 9 normal controls. LTB4 release from both monocytes (7.3 +/- 3.3 ng) and PMN (6.6 +/- 3.4 ng) in SLE patients was decreased compared with that (15.2 +/- 4.3 ng for monocytes, 20.7 +/- 4.5 ng for PMN) in normal controls. There were no differences in the releasing capacities of monocytes and PMN between patients with active and inactive disease. LTB4 suppressed lymphocyte blastogenesis by PHA-P and induced suppressor cells. The significance of the decreased release of LTB4 from monocytes and PMN in SLE patients was discussed.     

Blood eosinophils,eosinophil-derived proteins,and leukotriene C4 generation in relation to bronchial hyperreactivity in children with atopic dermatitis

To assess the relation among eosinophil-related variables in the peripheral blood, bronchial hyperreactivity, and the presence of atopic dermatitis in children aged 5–14 years, we studied 11 patients with atopic dermatitis alone, six with asthma and atopic dermatitis, 12 with asthma alone, and 12 healthy controls. Eosinophil counts, levels of eosinophil cationic protein, and the capacity of eosinophils to generate leukotriene (LT) C₄, as well as bronchial hyperreactivity and a severity score for atopic dermatitis, were determined. Eosinophil variables were significantly higher in both patient groups with atopic dermatitis than in normal controls. In particular, ionophore A 23187 LTC₄ generation was higher in patients with atopic dermatitis alone (median 82, range 25–273 ng/10⁶ cells) and patients with combined asthma and atopic dermatitis (median 68, range 32–583 ng/10⁶ cells) than in normal controls (median 9, range 1–67 ng/10⁶ cells). However, there was no difference between the group of atopic dermatitis patients with asthma and without asthma. We conclude that eosinophil variables in the peripheral blood are mainly influenced by the presence of atopic dermatitis, and not the presence and the severity of asthma in patients with both asthma and atopic dermatitis.     

Increased leukotriene production by food additives in patients with atopic dermatitis and proven food intolerance

Recently, we identified a subgroup of patients with atopic dermatitis (AD) with a clinical relevant food intolerance proven by double blind placebo controlled challenge. In search of possible pathomechanisms involved in this food intolerance, which leads to aggravation of the disease, the aim of the present study was to determine sulfidoleukotriene production in these patients using isolated leucocytes from the peripheral blood after stimulation with different food additives. Leukotriene production of peripheral leucocytes was detected by incubation of isolated cells with the food additives at different concentrations ranging from 0.2 to 200 microg/mL after pre-stimulation with IL-3. Ten non-atopic donors (A), nine AD patients of the diet responder group with negative oral provocation test against food additives (B) and nine patients of the responder group with positive reactions after the oral provocation test (C) were investigated. In the non-atopic group (A), no increased sulfidoleukotriene (sLT) release was observed for all food additives tested. In group B, increased sLT production was determined using tartrazine in one patient (1/9) and using nitrite in two patients (2/9), whereas sLT production remained below the cut-off range in all patients of group B (9/9) using benzoate, metabisulfite and salicylate. By contrast, in group C increased sLT production was observed with food colour mix in 1/9, with tartrazine in 3/9, with benzoate in 4/9, with nitrite in 5/9, with salicylate in 2/9 and with metabisulfite in 1/9. However, no increased sLT concentration was determined in the presence of the tested food additives in two patients of group C. Increased sLT production by peripheral leucocytes in the presence of single food additives was observed in the majority of patients with a proven food intolerance towards food additives proven by double-blind-placebo-controlled challenges. These food additives were particulary tartrazine, benzoate and nitrite. These findings indicate that single food additives as aggravating factors in AD patients may trigger the disease through increased sLT production as a pathophysiological mechanism.     

Identification of Malassezia species isolated from patients with seborrhoeic dermatitis, atopic dermatitis, pityriasis versicolor and normal subjects.

We identified Malassezia species isolated from 42 patients with seborrhoeic dermatitis, 17 patients with atopic dermatitis, 22 patients with pityriasis versicolor, 35 normal subjects and 73 healthy medical students. Regarding the prevalence of Malassezia species in the 35 normal subjects, the frequency of isolation of Malassezia globosa was 22%, M. sympodialis 10% and M. furfur 3%. M. slooffiae, M. pachydermatis, M. restricta and M. obtusa were infrequently isolated from normal skin. Two different species were isolated coincidentally from seven samples. In the patients with atopic dermatitis, M. furfur was isolated more frequently from lesional skin (21%) than non-lesional skin (11%). However, there was no statistical significance. Therefore, this result, by itself, is insufficient to prove that M. furfur should be considered to be an exacerbating factor of atopic dermatitis. In seborrhoeic dermatitis, M. furfur (35%) and M. globosa (22%) were isolated from lesional skin on the face at significantly high rates in comparison with the normal subjects. Therefore, M. furfur and/or M. globosa may be pathogens of seborrhoeic dermatitis. M. globosa was isolated at a frequency of 55% from lesional skin of pityriasis versicolor, while all other species were below 10%. These data suggest that the pathogenic species of pityriasis versicolor is M. globosa.     

Selective impairment of Toll-like receptor 2-mediated proinflammatory cytokine production by monocytes from patients with atopic dermatitis

BACKGROUND: The skin of patients with atopic dermatitis (AD) exhibits a striking susceptibility to infection with gram-positive bacteria and herpes simplex virus (HSV), which are known to stimulate Toll-like receptor (TLR) 2. OBJECTIVE: We investigated whether TLR2-mediated proinflammatory cytokine production by monocytes is selectively impaired in patients with AD and, if so, whether high FcvarepsilonRI levels on the monocytes could be related to the impairment. METHODS: The 2 subpopulations of monocytes, CD14(dim) proinflammatory and CD14(bright) classical monocytes, from patients with AD and healthy control subjects were stimulated to produce IL-1beta and TNF-alpha with phorbol 12-myristate 13-acetate/ionomycin, LPS (TLR4 ligand), or Pam3Cys (TLR2 ligand) for 4 hours, and simultaneous flow cytometric assessment of surface phenotype and intracellular cytokine synthesis was performed. Surface expression of TLR2, TLR4, and FcvarepsilonRI on the monocyte subpopulations was also assessed by means of flow cytometry. RESULTS: TLR2-mediated IL-1beta and TNF-alpha production by either the CD14(dim) or CD14(bright) monocytes was found to be selectively impaired in patients with AD. The most remarkable reduction in TLR2-mediated proinflammatory cytokine production was observed in CD14(dim) monocytes expressing high FcvarepsilonRI levels from patients with AD. This reduction was restored by means of downregulation of their FcvarepsilonRI expression after preculture in the absence of IgE. CONCLUSION: Monocytes, particularly the proinflammatory monocytes, from patients with AD are functionally defective in their capacity to produce proinflammatory cytokines on TLR2 stimuli in part because of the high levels of their FcvarepsilonRI expression. CLINICAL IMPLICATIONS: This selective impairment of monocytes would explain why patients with AD are specifically susceptible to cutaneous staphylococcal and streptococcal and HSV infections.     

Clinical evaluation of urinary leukotriene E4 levels in children with atopic dermatitis]

To evaluate the significance of peptide leukotrienes in children with atopic dermatitis, we measured urinary LTE4 levels which are thought to reflect in vivo production of peptide LTs. Urine was collected in the early morning. There was no significant difference in urinary LTE4 levels among atopic children with different severities. On the other hand, urinary LTE4 levels were significantly elevated in the patients who had severe nocturnal itches compared with the patients who had mild nocturnal itches and normal controls. These results suggested that increased production of peptide LTs may be relevant to nocturnal itch of atopic dermatitis.     

Interleukin-3, interleukin-8, FMLP and C5a enhance the release of leukotrienes from neutrophils of patients with atopic dermatitis.

The influence of the receptor-specific stimuli interleukin-3 (IL-3), interleukin-8 (IL-8), C5a and formyl-methionyl-leucyl-phenylalanine (FMLP) on the generation of arachidonic acid-derived inflammatory mediators from neutrophils (PMN) has been studied in patients with atopic dermatitis (AD) as well as in healthy, non-atopic volunteers. The release of leukotriene (LT)B4, the omega-oxidation products 20-COOH- and 20-OH-LTB4 and the cysteinyleukotriene LTC4 were measured by reverse-phase HPLC and radioimmunoassay. The incubation of neutrophils with these stimuli led to a significantly higher release of LTB4 and LTC4 in the AD group. The spontaneous leukotriene generation of PMN from patients with AD was on average threefold higher compared to the control group. C5a stimulated the release of LTB4 and its metabolites from atopic cells up to 9 ng in contrast to low amounts from non-atopic cells. Furthermore, FMLP distinctly enhanced the leukotriene release of neutrophils from patients with AD compared to unstimulated cells and to cells of normal donors. IL-3 and IL-8 also significantly stimulated the generation of LTB4 and LTC4 of PMN from atopic patients. Our data emphasize that neutrophils may play an important role in the pathogenesis of AD by an increased responsiveness to receptor-specific stimuli and further suggest that IL-3 and IL-8 influence the acute and chronic inflammatory reactions in patients with AD.     

Interleukin-4 induced IgE and IgG4 secretion by B cells from atopic dermatitis patients

Peripheral blood mononuclear cells from 8 normals and 8 patients with atopic dermatitis (AD) were cultured with recombinant interleukin-4 (IL-4) and the IgE and IgG subclass levels in the culture supernatants measured by radioimmunoassays. IL-4 induced IgE and IgG4 secretion by B cells from both normals and AD patients whereas it has no consistent effect on IgG1, IgG2 and IgG3 secretion. The IL-4 dose response was similar for IgE and IgG4 secretion by cells from both normals and AD patients. On the average, the patients' cells secreted more IgE and less IgG4 than the cells from normals, but because of a large variation, the differences were not significant. However, the ratio of IgG4:IgE secretion was significantly greater for normals than AD patients (mean +/- SEM 7.1 +/- 1.6:1 vs. 1.5 +/- 0.4:1; p less than 0.01). The data demonstrate that IL-4 induces IgE and IgG4 secretion by B cells from both normals and AD patients and suggest that the IL-4 induced switch from IgM to IgG4 or IgE secretion may proceed preferentially to IgE in AD patients as compared to normals.     

Chemical mediators in atopic dermatitis: involvement of leukotriene B4 released by a type I allergic reaction in the pathogenesis of atopic dermatitis

BACKGROUND: The mediators produced from a type I allergic reaction have not yet been able to explain the pathogenesis of atopic dermatitis (AD). OBJECTIVE: The purpose of this study was to elucidate the involvement of leukotriene (LT) B4 produced from a type I allergic reaction in the pathogenesis of AD. METHOD: The release of LTB4 was measured both in vitro, in passively sensitized and antigen-challenged human skin slices, as well as in vivo, in skin chambers on patients with AD. RESULTS: LTB4 was released from in vitro human skin by stimulation of the antigen (54.9 +/- 14.6 pg/g wet weight of skin by antigen challenge and 28.0 +/- 11.1 pg/g in control skin, P <.002). Antigen-specific release of LTB4 and histamine was also observed in vivo in nonlesional skin from the patients with AD by using the skin chamber technique. CONCLUSION: LTB4 release during type I allergic reaction in human skin has been determined in vitro. The released LTB4 possibly contributes to cellular response at the acute inflammatory lesion of AD.     

Expression of IL-18 mRNA and secretion of IL-18 are reduced in monocytes from patients with atopic dermatitis

BACKGROUND: Nasal polyps (NPs) are characterized by eosinophilic inflammation and often coexist with asthma. However, the role of atopy and IgE in NP pathogenesis is unclear. OBJECTIVE: We sought to determine whether there is an association between total and specific IgE to a variety of allergens in polyp and nonpolyp tissue and markers of eosinophilic inflammation or skin test results. METHODS: Homogenates were prepared from nasal tissue of 20 patients with NPs and 20 patients without NPs and analyzed for concentrations of IL-5, IL-4, eotaxin, leukotriene (LT) C4/D4/E4, sCD23, and histamine (ELISA). Eosinophil cationic protein (ECP), tryptase, and total and specific IgE for inhalant allergens and Staphylococcus aureus enterotoxins were measured (ImmunoCAP). RESULTS: The concentrations of total IgE, IL-5, eotaxin, ECP, LTC4/D4/E4, and sCD23 were significantly higher in NP tissue compared with nonpolyp tissue. Total IgE was significantly correlated to IL-5, ECP, LTC4/D4/E4, and sCD23 and to the number of eosinophils in NPs. On the basis of the presence of specific IgE antibodies in tissue, 3 NP groups were defined. NP group 1 demonstrated no measurable specific IgE, and NP group 2 selected specific IgE. The third group demonstrated a multiclonal specific IgE, including IgE to S aureus enterotoxins, a high total IgE level, and a high prevalence of asthma. CONCLUSIONS: These studies suggest that there is an association between increased levels of total IgE, specific IgE, and eosinophilic inflammation in NPs, which may be of relevance in the pathophysiology of nasal polyposis. Similarly, the presence of specific IgE to staphylococcal enterotoxins A and B also points to a possible role of bacterial superantigens.     

Spontaneous release of histamine from basophils and histamine-releasing factor in patients with atopic dermatitis and food hypersensitivity

Patients with hypersensitivity to food documented by a double-blind, placebo-controlled oral food challenge have been reported to have a high rate of release of histamine from basophils in vitro. To determine whether patients with atopic dermatitis and food hypersensitivity had similar high rates of spontaneous histamine release in vitro, whether dietary elimination of relevant food antigens affected this release, and whether a cytokine, histamine-releasing factor, could account for it, we evaluated 63 patients with atopic dermatitis and food hypersensitivity (38 of whom had eliminated the offending foods from their diets), 20 patients with atopic dermatitis without food hypersensitivity, and 18 normal volunteers. Patients with atopic dermatitis and food hypersensitivity were found to have higher rates of spontaneous release of histamine from basophils than controls (mean +/- SE, 35.1 +/- 3.9 percent vs. 2.3 +/- 0.2 percent; P less than 0.001). Those who had eliminated the offending food allergen from the diet for an extended period had a significantly lower rate of histamine release (3.7 +/- 0.5 percent; P less than 0.001). In patients with atopic dermatitis without food hypersensitivity, the rate (1.8 +/- 0.2 percent) did not differ from that in normal controls. Mononuclear cells from persons with food allergies spontaneously produced a histamine-releasing factor in vitro that provoked the release of histamine from the basophils of other food-sensitive persons, but not from those of normal controls. Patients who adhered to a restricted diet had a decline in the rate of spontaneous generation of the factor by their mononuclear cells. The histamine-releasing factor was found to activate basophils through surface-bound IgE. We conclude that in patients with food hypersensitivity, exposure to the relevant antigens produces a cytokine (histamine-releasing factor) that interacts with IgE bound to the surface of basophils, causing them to release histamine.     

Increased levels of urinary leukotriene E4 in children with severe atopic eczema/dermatitis syndrome

BACKGROUND: Leukotrienes are thought to play a role in the pathogenesis of atopic eczema/dermatitis syndrome (AEDS). Urinary leukotriene E4 (U-LTE4) is a marker of whole-body cysteinyl-leukotriene production. AIMS OF THE STUDY: To evaluate the role of leukotrienes in children with AEDS by measuring levels of U-LTE4, and to evaluate whether levels of U-LTE4 may reflect disease activity and allergic sensitization in AEDS. METHODS: U-LTE4 was measured by enzyme-linked immunosorbent assay in 87 children with mild (n=32), moderate (n=34) and severe (n=21) AEDS, as well as in 72 nonatopic healthy controls. Fifty-eight of the children with AEDS were sensitized to common allergens, and 29 were not. RESULTS: Levels of U-LTE4 were higher in children with severe AEDS (140; 66-166 microg/mmol creatinine, median; quartiles) than in controls (52; 30-90, P <0.05), whereas levels of U-LTE4 in moderate and mild disease were similar to controls. U-LTE4 levels were similar in children with or without sensitization to common allergens, but severe AEDS children with sensitization had higher levels of U-LTE4 than those without sensitization. CONCLUSION: The results suggest a role for leukotrienes in the pathogenesis of severe AEDS, and may support a role for leukotriene-antagonists in the treatment of this disorder. Levels of U-LTE4 may reflect the disease severity and sensitization to allergens in AEDS.     

Interleukin-18 is elevated in the sera from patients with atopic dermatitis and from atopic dermatitis model mice, NC/Nga

BACKGROUND: Several lines of in vitro and in vivo studies have demonstrated that interleukin-18 (IL-18) shows both antiallergic and allergy-promoting activities. But its expression in allergic diseases remains unknown. METHODS: Serum IL-18 levels from atopic dermatitis (AD) model mice, NC/Nga and control mice and from patients with AD and healthy volunteers were measured by ELISA. The relationship between IL-18 levels and serum IgE levels or clinical severity was also examined. RESULTS: Serum IL-18 levels from NC/Nga mice were significantly increased compared to those from control mice. The elevation of IL-18 in the sera was observed prior to the onset and during the development of dermatitis in NC/Nga mice. In addition, IL-18 levels in the sera from patients with AD were significantly (p < 0.05) elevated compared to those from healthy volunteers. However, serum IL-18 levels tended to correlate negatively with serum IgE levels in patients with AD and NC/Nga mice. CONCLUSION: IL-18 is overexpressed in AD.     

Platelet-activating factor- and leukotriene B4-induced release of lactoferrin from blood neutrophils of atopic and nonatopic individuals

We found increased accumulation of neutrophils and their components, lactoferrin (Lf) and elastase, as well as platelet-activating factor (PAF) and leukotriene B4 (LTB4) at sites of ongoing human allergic reactions. To determine whether PAF or LTB4, could be the stimulus for in vivo Lf release, blood neutrophils of 17 subjects were incubated with PAF, LTB4, or the phorbol ester, phorbol myristate acetate (PMA), and the released Lf (ELISA assay) was compared with spontaneous release. Significantly increased Lf release was induced by PAF, 10(-5) to 10(-8) mol/L (p less than 0.002); LTB4, 10(-7) to 10(-8) mol/L (p less than 0.004); and PMA (0.05 micrograms/ml) in a dose-dependent reaction. Cytochalasin was not required for Lf secretion but did enhance such responses. PAF-induced Lf secretion was inhibited by the specific PAF antagonist, BN 52063. More Lf was released from neutrophils of atopic than from nonatopic subjects in response to PAF, 10(-6) mol/L (4.2 micrograms/ml +/- 0.2 versus 2.6 micrograms/ml +/- 0.2; p less than 0.001) but not to LTB4, PMA, or buffer (p, not significant). We conclude that (1) PAF and LTB4 released in vivo could stimulate local neutrophils to release Lf with possible pathogenic effects and (2) neutrophils of atopic subjects are more responsive to PAF than neutrophils of nonatopic subjects in this regard.