Serum-induced basophil CD63 expression by means of a tricolour flow cytometric method for the in vitro diagnosis of chronic urticaria

BACKGROUND: Functional autoantibodies against the alpha-chain of the high-affinity IgE receptor (FcepsilonRIalpha) identify a subset of patients with chronic urticaria (CU) due to autoreactivity, as assessed by an in vivo positive response to autologous serum skin test (ASST). We performed a study to standardize the serum-induced basophil activation assay by flow cytometry (FCM) using a new tricolour method, assessing the diagnostic performance of this test in discriminating between ASST+ and ASST- CU patients. METHODS: Sera of 64 CU patients (22 ASST+ CU and 42 ASST- CU) and 10 healthy subjects were tested for their ability to induce basophil CD63 expression when incubated with whole blood of both atopic (DA) and non-atopic donors (DNA). Using a triple-labelled strategy with anti-CD123, anti-HLA-DR and anti-CD63 antibodies, CD63+ basophils were identified on a selected population of CD123+ HLA-DR- cells. In 3 ASST+ CU patients who underwent cyclosporine therapy, the assay was performed before and after treatment. RESULTS: The ASST+ CU sera resulted in a significant higher induction of basophil CD63 expression compared with ASST- CU and healthy donors sera; when whole blood from DA was used, sensitivity and specificity of the assay were 95.5% and 90.5% respectively. ASST+ CU serum activity was significantly decreased during cyclosporine A treatment, in parallel with clinical remission. CONCLUSIONS: Chronic urticaria serum-induced CD63 expression assay performed on DA whole blood by means of our tricolour FCM method could be the most useful tool for identification of a subset of patients with autoimmune CU and may become a promising tool also for monitoring treatment efficacy.     

Circulating level of the platelet-derived CXC chemokine platelet factor 4 in chronic urticaria patients with or without coexistent euthyroid Hashimoto's thyroiditis

The concept of autoimmune aetiology of some cases of chronic urticaria (CU) has been supported by several observation including wheal-and-flare reaction induced by intradermal injection of autologous serum as well as association with other autoimmune diseases, in particular Hashimoto's thyroiditis (HT). It is known that activated platelets may actively participate in immune-inflammatory processes. Therefore, we assessed whether autoimmune phenomenon associated with CU influence the systemic platelet activity measured by circulating level of platelet factor 4 (PF-4). Plasma level of PF-4 was analysed using enzyme-linked immunosorbent assay in twelve women with strong positive response to autologous serum skin test (ASST) suffering from CU, twelve female patients with strong positive ASST suffering from both CU and untreated, HT as well as sixteen healthy women. All the subjects were clinically and biochemically euthyroid. There were no statistically significant differences between the CU patients with or without euthyroid HT and plasma PF-4 level in healthy controls. In patients with both CU and thyroiditis, plasma level of PF-4 did not correlate significantly with the level of antibodies against thyroperoxidase. It seems that circulating level of the platelet-derived chemokine is not increased in CU patients with positive response to ASST, regardless the occurrence of euthyroid HT.     

High prevalence of autoimmune urticaria in children with chronic urticaria

BACKGROUND: The etiology of chronic urticaria (CU) in childhood often remains unrecognized. Recently, in adults it has been shown that approximately 40% of patients with CU have autoimmune urticaria (AU); however, no data are available in children. OBJECTIVE: To determine the prevalence and possible risk factors for AU in children with CU. METHODS: Ninety-three consecutive children (52 male; median age, 7.8 years) with CU were evaluated for AU by means of autologous serum skin test (ASST) in all and serum-induced basophil histamine release (HR-urticaria test) in 52. All other known causes of CU were excluded as appropriate. RESULTS: A cause for CU was identified in 44 children (47%), whereas 49 (53%) remained idiopathic. ASST and HR-urticaria test had positive results in 22 of 49 (45%) and in 16 of 31 (52%) children with idiopathic CU compared with 1 of 44 (2%) and 5 of 21 (24%) with CU of a known cause, respectively ( P <.00001; P=.09). Sensitivity, specificity, and positive and negative predictive values of the ASST for diagnosing AU are 78%, 85%, 74%, and 88%. The prevalence of AU in childhood is 31% (15/52; 95% CI, 24%-51%). None of the variables studied were predictive for development of AU. CONCLUSION: Our results demonstrate for the first time that children have the same ability as adults to produce functionally active autoantibodies directed against IgE or IgE receptor and that AU occurs in children in as many as 30% of cases. The addition of screening for AU dramatically decreases the rate of the idiopathic form from 52% to 20%.     

Plasma of patients with chronic urticaria shows signs of thrombin generation, and its intradermal injection causes wheal-and-flare reactions much more frequently than autologous serum

BACKGROUND: Several aspects of the pathogenesis of chronic urticaria (CU) remain contradictory. Autologous serum skin tests (ASSTs) and in vitro histamine release assays seem to look into distinct aspects of the disease, and the specificity of ASST has been questioned. OBJECTIVE: We compared the autologous plasma skin test (APST) with ASST to detect autoreactivity in patients with CU. The clotting process was investigated as well by measuring in vivo thrombin generation. METHODS: A total of 96 adults with CU underwent ASST; 71 of them underwent APST with Na citrate-anticoagulated plasma. Prothrombin fragment 1+2 plasma levels were measured by a sandwich ELISA in Na citrate-anticoagulated plasmas from 28 patients and 27 controls. RESULTS: Fifty-one of 96 (53%) patients scored positive on ASST, whereas 61 of 71 (86%) patients scored positive on APST (21/30 [70%] ASST-negative and 40/41 [98%] ASST-positive). Plasma prothrombin fragment 1+2 was higher in patients than controls (3.06 [SD 3.36] vs 0.80 [0.34]; P < .001) and in ASST-positive/APST-positive than in ASST-negative/APST-positive patients (3.89 [SD 3.68] vs 1.33 [1.64]; P = 0.058) and was directly related to urticaria severity (r = 0.37; P < .05). CONCLUSION: Most patients with CU are positive on APST-Na citrate. CU is associated with the generation of thrombin, a serine protease able to activate mast cells and to cause relevant increase in permeability of endothelium. APST and ASST only partially depend on the presence of circulating antibodies to FcepsilonRI or to IgE. CLINICAL IMPLICATIONS: These findings provide new insights into the pathogenesis of CU and suggest new therapeutic opportunities for treating this disease.     

Elevated serum B-cell activating factor in patients with chronic urticaria

The B and T cells abnormalities that have been described among CU patients, lend support to its regard as an autoimmune disease. In this study we compared serum B-cell activation factor (BAFF) levels in 46 CU patients to 24 healthy controls and evaluated a possible association between elevated serum BAFF in CU patients and the presence of autologous serum skin test, anti-thyroid antibodies, antinuclear antibodies, high total IGE levels, and urticaria disease severity. We found that serum BAFF levels is elevated statistically significant in CU patients compared to healthy control (1228±342 pg/ml vs. 758±313 pg/ml, P<0.0001). CU patients with severe disease activity had significantly higher serum BAFF levels compared to patients with mild CU (1394.6±299.6 vs. 1097.6±221.3, P=0.0008). No association was found between the presence of positive autologous serum skin test, anti-thyroid antibodies or antinuclear antibodies or high levels of total IgE and serum BAFF levels in CU Patients. We conclude that CU patients have higher levels of serum BAFF which associate with disease severity.     

Autologous serum skin test in chronic idiopathic urticaria: prevalence, correlation and clinical implications

Some cases of chronic idiopathic urticaria (CIU) have histamine-releasing IgG autoantibodies in their blood. This disease subgroup is called "autoimmune urticaria". To date, the autologous serum skin test (ASST) is the best in vivo clinical test for the detection of basophil histamine-releasing activity in vitro. This study aimed to find the prevalence of ASST positive cases in Thai patients with CIU, to identify factors related to the positivity of ASST and to find the clinical implications of ASST in CIU. A retrospective study was performed among 85 CIU patients who attended the Urticaria Clinic at the Department of Dermatology, Siriraj Hospital and were willing to perform ASST, from January 2002 to December 2003. Twenty-one (24.7%) patients had a positive ASST. There was no significant difference between patients with positive ASST and negative ASST as to the severity of the disease (wheal numbers, wheal size, itching scores and the extent of body involvement) as well as the duration of the disease.     

The autologous serum skin test in the follow-up of patients with chronic urticaria

BACKGROUND: The presence of anti-FcepsilonRI and anti-IgE autoantibodies in a subset of patients with chronic urticaria suggests their aetiopathogenetic role. In clinical practice, the presence of these antibodies is usually considered when the autologous serum skin test (ASST) is positive. AIMS: To evaluate if the positive ASST follows up the activity of chronic urticaria. METHODS: Autologous serum skin test and thyroid autoantibody detection were performed in 82 patients with chronic urticaria and repeated 1 year later, when the vast majority of patients were symptom-free. Twenty patients with Hashimoto thyroiditis (HT), who had never suffered from urticaria, represented the control group. RESULTS: At the start of the study, the prevalence of positive ASST was 46.6%. The association of HT-urticaria was 29.3%. ASST was positive in 62 and 39% of patients with and without HT, respectively (P > 0.05 ns). One year later, 28 of 34 patients with a positive ASST were symptom-free, but 50% of them were positive for ASST. The ASST was positive in 86.7 and 8% of patients with and without HT, respectively (P < 0.001). In the control group, ASST was always negative. CONCLUSIONS: The co-existence of autoimmune thyroiditis with chronic urticaria seems to induce a significant difference in the persistence of a positive ASST. Consistent with previous reports, a positive ASST correlates with disease exacerbation in chronic urticaria patients without thyroiditis. In patients with thyroiditis and urticaria, positive ASST persists even after the urticaria has disappeared, thus questioning whether a positive ASST to be a surrogate marker of the functional role of anti-FcepsilonRI and anti-IgE autoantibodies.     

Validation of basophil histamine release against the autologous serum skin test and outcome of serum-induced basophil histamine release studies in a large population of chronic urticaria patients

BACKGROUND: Endogenous histamine-releasing factors (HRFs) are involved in 30-60% of patients with chronic urticaria (CU). Evidence for their existence comes from in vivo studies of autoreactivity with the autologous serum skin test (ASST), in vitro immunoassays demonstrating autoantibodies against the immunoglobulin E (IgE) or the high affinity IgE receptor (FcepsilonRI) and serum-induced histamine release (HR) from basophils and mast cells. We have examined the correlation between the ASST and a new basophil histamine-releasing assay (the HR-Urtikaria test) in a group of well-characterized CU patients and subsequently determined the frequency of HR-Urticaria-positive sera from a larger population of CU patients. SUBJECTS: Group 1 consisted of 28 patients with CU (16 were ASST-positive) 20 patients with atopic dermatitis, 24 patients with allergy to birch and nine healthy controls. Group 2 consisted of 873 unselected CU patients. METHODS: White blood cells containing 1-2% basophils from a healthy nonatopic donor were incubated with patients sera in the presence of interleukin (IL)-3. Histamine was measured by the glass fibre method. RESULTS: Using the ASST as the true outcome, the HR-Urticaria test showed a sensitivity and specificity of 75% in group 1 using a cut-off value for HR of >16.5%. None of the controls was positive in the HR-Urticaria test. In group 2, we found no difference in the frequency of positives between male (34.6%, n = 254) and female adults (35.1%, n = 576) but twice as many females as males were tested. CONCLUSIONS: Our studies have shown that the HR-Urticaria test has a good sensitivity and specificity for endogenous HRFs demonstrated by the ASST in patients with CU and that about one-third of unselected patients with CU have a positive result.     

Helicobacter pylori infection: prevalence in chronic urticaria patients and incidence of autoimmune urticaria (study in Dr. Cipto Mangunkusumo Hospital, Jakarta)

AIM: To determine the prevalence of Hp infection in patients with chronic urticaria (CU) and to evaluate the result of autologous serum skin test (ASST) in CU patients with Hp infections. METHODS: In this cross-sectional study, 16 patients with chronic urticaria and 16 non-urticaria volunteers were investigated (matched for age and sex). All subjects were examined for Hp infection with the 13C-urea breath test. Autologous serum skin test was performed in patients with proven Hp infection. RESULTS: Helicobacter pylori was detected in 12.5% of patients and 0% of the control group. There was no significant difference between the two groups (p = 0.484 using Fisher exact test). Autologous serum skin test was positive in 1 of 2 CU patients with Hp infection. CONCLUSION: In this study, there was no significant difference in the seroprevalence of Hp infection between CU patients and controls. Autologous serum skin test was positive in 1 of 2 CU patients with Hp infection.     

Correlation between T-cell and mast cell activity in patients with chronic urticaria

BACKGROUND: The presence of autoantibodies reacting with the high affinity IgE receptor (FcepsilonRIalpha) usually indicates a more severe form of chronic urticaria (CU). Previously, we showed an increased lymphocyte reactivity in CU patients; however, the relation between enhanced cellular immunity and the presence of anti-FcepsilonRIalpha-specific autoantibodies has not been investigated. METHODS: Cellular and humoral immune reactivity of 50 CU patients and 28 healthy controls was studied.Serum sIL-2R, neopterin, and tryptase levels were measured to assess T-cell, monocyte/macrophage and mast cell activity, respectively. Helicobacter pylori (HP)-specific IgG antibody, and IgE levels were also tested. Anti-FcepsilonRIalpha-specific autoantibody was determined by Western blotting. In vivo histamine-releasing activity of patients' sera was assessed by the autologous serum skin test (AST). RESULTS: 17/50 CU patients, who both had IgG-type anti-FcepsilonRIalpha-antibodies by Western blotting and a positive AST response, were classified as autoimmune CU. All patients with CU had significantly higher serum sIL-2R and tryptase levels than healthy controls (p = 0.000257, p = 0.000166, respectively), indicating T-cell and mast cell activation. Patients with higher sIL-2R levels also had higher tryptase levels; the strongest correlation was shown in the autoimmune subgroup of patients (rho = 0.688, p = 0.002). There was a tendency towards higher tryptase levels in the autoimmune subgroup, as compared to the nonautoimmune CU patients. While the serum IgE was significantly lower in autoimmune than in nonautoimmune CU (p = 0.000836), there was no significant difference in their sIL-2R, neopterin and HP-specific IgG antibody levels. CU patients with a positive AST response (38/50) had significantly higher tryptase levels (p = 0.0107) when compared to the negative skin test group. CONCLUSIONS: The significant correlation between sIL-2R and tryptase levels in patients with CU indicates that T cell activation is proportional to mast cell degranulation in these patients. The increased level of tryptase in autoimmune CU may suggest more severe disease.     

EAACI taskforce position paper: evidence for autoimmune urticaria and proposal for defining diagnostic criteria

An autoimmune subset of chronic spontaneous urticaria is increasingly being recognized internationally, based on laboratory and clinical evidence that has accrued over the last 20 years. This evidence has been reviewed by a taskforce of the Dermatology section of the European Academy of Allergy and Clinical Immunology. Functional autoantibodies in chronic urticaria (CU) patient sera have been demonstrated against IgE and FcεRIα by basophil and mast cell histamine release assays and by basophil activation assays. Antibody specificity has been confirmed by immunoassay, but there is a poor correlation between functionality and immunoreactivity. Approximately 25% of CU patients have a positive basophil histamine release assay and show autoreactivity (a positive autologous serum skin test), whereas 50% are negative regarding both. Functionality of CU sera appears to be complement dependent on mast cells but not exclusively on basophils. Basophil activation by CU sera is predominantly restricted to IgG1 and IgG3 subclasses. Circumstantial evidence for CU being an autoimmune disease comes from an observed association with other autoimmune diseases, a strong association between serum functionality and HLA‐DR4 haplotype and the good response of CU patients to immunotherapies. It was proposed that a study should be undertaken to prospectively validate potentially relevant clinical criteria (from the history, examination and routinely available clinical investigations) against a new ‘gold standard’ for the diagnosis of ACU (positive autoreactivity, functional bioassay and immunoassay) to define preliminary criteria sets for the diagnosis of ACU based on clinical and laboratory features with highest individual sensitivity and specificity.     

Assessment of histamine-releasing activity of sera from patients with chronic urticaria showing positive autologous skin test on human basophils and mast cells

BACKGROUND: All previous studies agree that only a proportion of sera from patients with chronic urticaria (CU) positive on the autologous serum skin test (ASST) are able to induce histamine release in vitro. A non-specific release of bradykinins during clotting of blood samples has been suggested; however, ASST seems rather specific and some data point to the existence of a mast cell-specific histamine-releasing factor. OBJECTIVE: To assess whether, and to what extent, the use of both human basophils and mast cells increases the sensitivity of in vitro histamine release assays (HRAs) in ASST-positive patients with CU. METHODS: The histamine-releasing activity of sera from 93 patients with CU selected on the basis of strong skin reactivity on ASST was assessed in vitro on basophils from 1 (n=86), 2 (n=31), or 3 (n=20) normal donors, and on mast cells from 1 (n=3), 2 (n=3), or 3 (n=87) normal donors. RESULTS: Sera from 88/93 (95%) patients induced significant histamine release from mast cells or basophils on at least one HRA. 76/93 (82%), 45/90 (50%), 22/80 (28%), and 6/12 (50%) sera were able to induce significant histamine release from cells of 2/5, 3/5, 4/5 and 5/5 donors, respectively. CONCLUSION: Sera from nearly all ASST-positive patients with CU are able to induce histamine release in vitro. However, the serum from each single patient seems to show its maximal activity on autologous mast cells in vivo, and functional in vitro tests show much variability and seem less sensitive than ASST in the detection of patients with histamine-releasing factors in their blood.     

The addition of zafirlukast to cetirizine improves the treatment of chronic urticaria in patients with positive autologous serum skin test results

BACKGROUND: Because leukotrienes have potent local effects on cutaneous vasculature, leukotriene antagonists might be effective in the treatment of chronic urticaria. OBJECTIVE: A double-blinded, placebo-controlled trial comparing cetirizine 10 mg daily in combination with zafirlukast 20 mg twice a day versus cetirizine 10 mg daily and placebo was conducted to determine whether subjects with chronic urticaria benefit from add-on therapy with a leukotriene-modifying agent. METHODS: Patients 12 years or older with a history of chronic urticaria (more than 6 weeks in duration) required diary documentation of 6 or more hives on at least 2 days/week and a suboptimal response to H(1)-antagonist therapy for enrollment. At baseline, all subjects were skin tested to autologous serum to assess for the potential presence of FcepsilonRI or IgE autoantibodies. Subjects meeting the initial entry criteria were treated with cetirizine 10 mg a day and placebo twice daily for 1 week. Those patients with persistent hives were randomized to receive cetirizine 10 mg daily and zafirlukast 20 mg twice a day or cetirizine 10 mg daily and placebo. At each successive weekly visit, physician and patient treatment effectiveness score (TES) and visual analog scale (VAS) ratings were recorded. Statistical analysis used generalizing estimating equations to compare the effect of combination therapy versus monotherapy on TES and VAS ratings. Results were adjusted for baseline rating, recruiting center, and autologous serum skin test (ASST). A separate analysis evaluated patients with positive ASST results receiving combination therapy versus monotherapy. RESULTS: Combination therapy with zafirlukast demonstrated a modest but significantly greater improvement compared with cetirizine monotherapy in physician and patient recorded VAS ratings at visit 4 and across treatment visits 4 through 6 (P <.05 unless stated otherwise). Subjects with ASST positive results receiving combination therapy as compared with subjects with negative ASST results exhibited a significant improvement in patient recorded VAS ratings across visits 4 through 6. Subgroup analysis of subjects with ASST positive results receiving combination therapy versus monotherapy showed improvement in physician recorded TES at visit 5, physician recorded VAS at visits 4 and 5 and across visits 4 through 6, as well as for patient recorded VAS at visit 5. There were no significant results for patients with ASST negative results. CONCLUSION: The results of this study indicate that only patients with autoimmune (ASST positive) chronic urticaria refractory to H(1)-antagonist monotherapy might benefit from the addition of the leukotriene D(4)-receptor antagonist zafirlukast to their treatment regimen. These results also suggest that routine screening of patients with chronic urticaria with the ASST might be useful in formulating therapeutic algorithms in the management of chronic urticaria.     

Clinical and laboratory parameters in predicting chronic urticaria duration: a prospective study of 139 patients

BACKGROUND: Despite the disabling nature of chronic urticaria (CU), little is known about the disease's duration or the efficacy of adopting aggressive therapeutic regimens such as cyclosporine A. OBJECTIVES: The aim of this study was to evaluate whether parameters such as angioedema, autologous serum test, anti-thyroid antibodies, and total IgE could predict both CU duration and severity. PATIENTS AND METHODS: One hundred and thirty-nine patients suffering from CU were prospectively followed over a 5-year period for disease duration, severity and the presence of angioedema. Also investigated was the association between these clinical parameters and the subsequent detection of autologous serum test, anti-thyroid antibodies, and total IgE. RESULTS: CU lasted over 1 year in more than 70% of cases and in 14% it still existed after 5 years. Angioedema co-existed or appeared during the course of CU in 40% of patients and was associated with disease duration. Autologous serum test and anti-thyroid antibodies were found positive in 28 and 12% of patients, respectively, compared to none of normal individuals, P = 0.001. CU duration was associated with the presence of both autologous serum test and anti-thyroid antibodies; however, autologous serum test and not anti-thyroid antibodies was found in association with CU severity. CONCLUSION: We demonstrate for the first time that CU duration is associated with clinical parameters such as severity and angioedema, and with laboratory parameters such as autologous serum test and anti-thyroid antibodies. The ability to predict CU duration may facilitate decisions regarding the possible early initiation of cyclosporine A as a means by which to reduce disease severity and duration.     

Natural anti-FcepsilonRIalpha autoantibodies may interfere with diagnostic tests for autoimmune urticaria

IgG autoantibodies against the alpha-chain of the high affinity IgE receptor are claimed to play a pathogenetic role in autoimmune urticaria. The best methods for detection of functional autoantibodies are currently the autologous serum skin test and the basophil histamine release assay. A simplified and feasible screening test would facilitate the diagnosis of autoimmune urticaria.Here we offer an explanation for the difficulties in establishing a screening test for autoantibodies directed against the alpha-chain of the high affinity IgE receptor in autoimmune urticaria. Identical autoantibodies in chronic urticaria patients and healthy donors belonging to the natural autoantibody repertoire were found by sequence analysis of anti-alpha-chain autoantibodies isolated by repertoire cloning from antibody libraries. These natural autoantibodies bound to the receptor and triggered histamine release but only if IgE was previously removed from the receptor. Diagnostic assays used for detection of antibodies directed against the IgE receptor may require signal comparison with and without the artificial removal of IgE, immune complexes, and complement in order to avoid false positive or negative results. After IgE removal diagnostic tests will detect natural autoantibodies against the high affinity IgE receptor regardless of whether they are pathogenic or not.     

Assessment of circulating FCεRIa in Chronic Spontaneous Urticaria patients and its correlation with clinical and immunological variables

Chronic spontaneous urticaria (CSU) is a chronic type characterized by episodes of wheals with or without angioedema. Autoantibody against the alpha subunit of Fc epsilon receptor (FcεRIa) was detected in CSU patients' sera. The study aims to evaluate the clinical utility of skin tests in CSU patients. In addition, it assesses the presence of circulating FcεRIa in CSU patients and their correlation with other clinical and immunological variables. The study includes 40 healthy controls and 40 CSU patients who had urticaria symptoms for at least 8 weeks. All subjects underwent the following tests: autologous serum skin test (ASST), autologous plasma skin test (APST), immunoglobulin E (IgE), antinuclear antibodies (ANA), antithyroid antibodies (ATA). An in-house enzyme-linked immunosorbent assay was used for FcεRIa detection. The prevalence of ANA and ATA in CSU was 7.5% and 20% respectively. Total IgE was significantly higher in CSU than in controls (p?<?0.0001). The study detected circulating antibody to FcεRIα in 2.5% of controls and 52.5% of CSU patients (p?<?0.0001). The prevalence of antibody to FcεRIa was 27.3% and 83.3% of ASST negative and positive patients respectively (p?=?0.0004). But the prevalence was 17.6% and 78.3% of APST negative and positive patients respectively (p?=?0.0002). In conclusion, Circulating antibody to FcεRIa has a role in the pathogenic mechanisms of CSU.     

Chronic urticaria serum induces histamine release, leukotriene production, and basophil CD63 surface expression--inhibitory effects ofanti-inflammatory drugs

BACKGROUND: A role of potential histamine-releasing autoantibodies against the high-affinity IgE receptor on the surface of basophils and mast cells is discussed in the pathogenesis of chronic urticaria. This so-called autoimmune urticaria may be diagnosed by a positive intracutaneous autologous serum skin test, which is found in about 30% of patients with chronic urticaria. OBJECTIVE: Our purpose was, first, to compare the effect of complement-inactivated sera of 20 patients with chronic urticaria and positive autologous serum skin tests, 20 patients with chronic urticaria and negative skin tests, and 20 control subjects without chronic urticaria (10 atopic and 10 nonatopic subjects) and, second, to analyze the effect of anti-inflammatory drugs on the serum activity. METHODS: The following assay systems were used: release of histamine in whole blood samples, surface expression of the activation marker CD63 on basophils, and sulfidoleukotriene de novo production in leukocyte suspensions. Whole blood, basophils, and leukocyte suspensions were obtained from a nonatopic and an atopic donor. RESULTS: Sera of patients with autologous serum skin test positive chronic urticaria resulted not only in significantly increased histamine release compared with skin test-negative chronic urticaria sera but also in a significant higher induction of basophil CD63 surface expression and sulfidoleukotriene de novo production. However, serum activity was neither characteristic for chronic urticaria nor for chronic urticaria with a positive autologous serum skin test. Preincubation with dapsone, chloroquine, and lidocaine dose dependently resulted in a significant reduction of all histamine release, CD63 expression, and sulfidoleukotriene production. In addition, mizolastine was able to inhibit serum-induced sulfidoleukotriene production. CONCLUSION: Further studies investigating the in vivo effect of these drugs will have to clarify their role in the management of the subset of patients with chronic urticaria demonstrating serum-induced inflammatory effects.     

Chronic urticaria sera increase basophil CD203c expression

BACKGROUND: Approximately 40% of patients with chronic idiopathic urticaria have antibodies to the alpha subunit of the high-affinity IgE receptor. CD203c is a basophil activation marker known to be upregulated by cross-linking of the FcepsilonRIalpha receptor and may serve as a useful marker to identify these patients. OBJECTIVE: The primary objective was to assess the affect of sera from patients with chronic idiopathic urticaria on basophil CD203c expression. Secondary objectives were to correlate CD203c expression with basophil histamine release and size of the autologous serum skin test and to determine whether the mechanism is mediated by an IgG antibody. METHODS: Sera were obtained from patients with chronic idiopathic urticaria and positive autologous serum skin test or negative autologous serum skin test and normal controls. Sera were incubated with donor whole blood. Activated basophils from whole blood were identified by flow cytometry on the basis of the presence of CD203c on high-expressing IgE positive cells. RESULTS: Incubation of donor basophils with sera from patients with chronic idiopathic urticaria and positive autologous serum skin test demonstrated significant upregulation of CD203c. IgG depletion of representative sera from patients with chronic idiopathic urticaria resulted in significant decrease in CD203c expression on donor basophils. CD203c expression correlated with basophil histamine release and the size of the autologous serum skin test. CONCLUSION: Sera from patients with chronic idiopathic urticaria and positive autologous serum skin test significantly upregulate basophil CD203c and correlate with basophil histamine release. CLINICAL IMPLICATIONS: This article describes an activation marker on basophils whose expression is increased by sera from patients with chronic idiopathic urticaria.     

Allergy-like asthma and rhinitis. A cross-sectional survey of a respiratory cohort and a diagnostic approach using the autologous serum skin test

BACKGROUND: Chronic idiopathic urticaria is considered an allergy-like skin disorder, and in some cases an auto-reactivity caused by mast cell degranulating factors has been demonstrated. A positive reaction to the autologous serum skin test (ASST), reflecting the presence of factors capable to degranulate the mast cells, is regarded as a reliable in vivo diagnostic test in chronic urticaria patients. About one out of three patients complaining for rhinitis or asthma does not show sensitization to any allergenic source. No data are available on the application of ASST to respiratory patients. METHODS: A cohort of respiratory patients, aged 1 to 80 years, complaining about suspected respiratory symptoms was screened using a panel of inhalant allergens by means of SPT and IgE. Current and past information on skin and respiratory symptoms were recorded for each patient. Patients were divided in 'allergy' and 'allergy-like' whether or not they react to at least one allergenic source. Age and gender distribution were analyzed. A control group fulfilling criteria for chronic idiopathic urticaria (CIU) was selected as well. The ASST was applied to the control CIU group, to the allergy-like and allergy subsets, and to a group of 58 healthy adult subjects. RESULTS: Allergy and allergy-like patients were differently distributed in relation to age and gender. Allergy-like children were prevalent before age six, whereas the prevalence of allergics steadily increased with age representing the large majority of patients in late childhood. The ratio of allergy/allergy-like subjects continues to increase in adult males, whereas adult females represent the large majority of the adult patients mainly within the allergy-like subset. Fifty-eight percent of CIU control patients reacted to the ASST. Among allergy-like patients ASST was positive in 47% of the adults (male 26%, female 54%), and in 84% of the children (no gender differences). The percentage of reactive subjects increased in the adult allergy-like subset if a CIU was associated (65%). Eighty-six percent of the pediatric patients and 61% of the adults having an allergy respiratory disease associated with an allergy-like condition had a positive ASST reactivity. Pure allergy patients reacted in 73% and 40% of the cases in the children and adults subgroups, respectively. Forty-five percent of healthy controls reacted to the ASST as well, and statistically significant gender differences were still recorded. CONCLUSIONS: An ASST reactivity is reported for the first time in allergy-like respiratory patients. A higher prevalence of reactive subjects has been recorded in all the pediatric subsets without gender differences, whereas female reactivity is prevalent among adults. The ASST reactivity seems to parallel the patient distribution within the entire respiratory cohort. Further studies are needed to demonstrate that the serum factors causing the ASST reactivity in respiratory patients are the same as for CIU affected subjects. In the light of recent in vitro findings, the ASST reactivity of healthy subjects could lead to a new interpretation of the autologous serum reactivity.     

Autoimmune Diseases Are Linked to Type IIb Autoimmune Chronic Spontaneous Urticaria

PurposePatients with chronic spontaneous urticaria (CSU) have an increased risk for comorbid autoimmune diseases. In this retrospective multicenter study of CSU patients, we evaluated clinical and laboratory features of CSU associated with a higher risk of comorbid autoimmune diseases.MethodsWe analyzed records of CSU patients (n = 1,199) for a history or presence of autoimmune diseases. Patients were diagnosed with type IIb autoimmune CSU (aiCSU) if all 3 tests were positive: autologous serum skin test (ASST), basophil histamine release assay (BHRA) and/or basophil activation test (BAT), and IgG autoantibodies against FcεRIα/IgE detected by immunoassay.ResultsTwenty-eight percent of CSU patients had at least 1 autoimmune disease. The most prevalent autoimmune diseases were Hashimoto''s thyroiditis (HT) (≥ 21%) and vitiligo (2%). Two percent of CSU patients had ≥ 2 autoimmune diseases, most frequently HT plus vitiligo. Comorbid autoimmune diseases, in patients with CSU, were associated with female sex, a family history of autoimmune diseases, and higher rates of hypothyroidism and hyperthyroidism (P < 0.001). Presence of autoimmune diseases was linked to aiCSU (P = 0.02). The risks of having autoimmune diseases were 1.7, 2.9 and 3.3 times higher for CSU patients with a positive ASST, BHRA and BAT, respectively. In CSU patients, markers for autoimmune diseases, antinuclear antibodies and/or IgG anti-thyroid antibodies were associated with non-response to omalizumab treatment (P = 0.013).ConclusionsIn CSU, autoimmune diseases are common and linked to type IIb autoimmune CSU. Our results suggest that physicians assess and monitor all adult patients with CSU for signs and symptoms of common autoimmune diseases, especially HT and vitiligo.