The costimulatory molecules CD80, CD86 and OX40L are up-regulated in Aspergillus fumigatus sensitized mice.

Aspergillus fumigatus (Af) is a fungus associated with allergic bronchopulmonary aspergillosis (ABPA) and other allergic diseases. Immune responses in these diseases are due to T and B cell responses. T cell activation requires both Af-specific engagement of the T-cell-receptor as well as interaction of antigen independent costimulatory molecules including CD28-CD80/CD86 and OX40-OX40L interactions. Since these molecules and their interactions have been suggested to have a potential involvement in the pathogenesis of ABPA, we have investigated their role in a model of experimental allergic aspergillosis. BALB/c mice were primed and sensitized with Af allergens, with or without exogenous IL-4. Results showed up-regulation of both CD86 and CD80 molecules on lung B cells from Af-sensitized mice (79% CD86+ and 24% CD80+) and Af/rIL-4-treated mice (90% CD86+ and 24% CD80+) compared to normal controls (36% and 17%, respectively). Lung macrophages in Af-sensitized mice treated or not with IL-4 showed enhanced expression of these molecules. OX40L expression was also up-regulated on lung B cells and macrophages from both Af-sensitized and Af/rIL-4 exposed mice as compared to normal controls. All Af-sensitized animals showed peripheral blood eosinophilia, enhanced total serum IgE and allergen-specific IgG1 antibodies and characteristic lung inflammation. The up-regulation of CD80, CD86 and OX40L molecules on lung B cells and macrophages from Af-allergen exposed mice suggests a major role for these molecules in the amplification and persistence of immunological and inflammatory responses in ABPA.

Detection of focal cerebral hemisphere lesions using the neurological examination.

OBJECTIVE: To determine the sensitivity and specificity of clinical tests for detecting focal lesions in a prospective blinded study. METHODS: 46 patients with a focal cerebral hemisphere lesion without obvious focal signs and 19 controls with normal imaging were examined using a battery of clinical tests. Examiners were blinded to the diagnosis. The sensitivity, specificity, and positive and negative predictive values of each test were measured. RESULTS: The upper limb tests with the greatest sensitivities for detecting a focal lesion were finger rolling (sensitivity 0.33 (95% confidence interval, 0.21 to 0.47)), assessment of power (0.30 (0.19 to 0.45)), rapid alternating movements (0.30 (0.19 to 0.45)), forearm rolling (0.24 (0.14 to 0.38)), and pronator drift (0.22 (0.12 to 0.36)). All these tests had a specificity of 1.00 (0.83 to 1.00). This combination of tests detected an abnormality in 50% of the patients with a focal lesion. In the lower limbs, assessment of power was the most sensitive test (sensitivity 0.20 (0.11 to 0.33)). Visual field defects were detected in 10 patients with a focal lesion (sensitivity 0.22 (0.12 to 0.36)) and facial weakness in eight (sensitivity 0.17 (0.09 to 0.31)). Overall, the examination detected signs of focal brain disease in 61% of the patients with a focal cerebral lesion. CONCLUSIONS: The neurological examination has a low sensitivity for detecting early cerebral hemisphere lesions in patients without obvious focal signs. The finger and forearm rolling tests, rapid alternating movements of the hands, and pronator drift are simple tests that increase the detection of a focal lesion without greatly increasing the length of the examination.

Cerebral venous sinus thrombosis: a diagnostic challenge.

BACKGROUND: Cerebral venous sinus thrombosis (CVT) is a potentially serious but treatable disorder that has been underdiagnosed in the past. Delay in diagnosis and treatment of this disorder has resulted in the death of one of our patients. AIM: To review the local experience with CVT in order to identify factors that may allow diagnosis and appropriate treatment decisions to be made more readily in the future. METHODS: A retrospective review of all cases of CVT diagnosed or treated at Auckland Hospital between 1990 and 1999. RESULTS: Twenty-five cases of CVT were identified. The number of cases diagnosed increased from less than one per year in 1990-94 to eight in 1999. Clinical signs at presentation included headache (96%), focal neurological deficits (60%), seizures (40%) and papilloedema (43%). Delayed diagnosis after admission to hospital occurred in two young women presenting with neurological symptoms during pregnancy or puerperium, in two cases in whom focal symptoms were not explained by negative computed tomography and in five cases presenting with intracerebral haemorrhage. Twenty patients received anticoagulant therapy and their condition remained stable or improved after treatment. CONCLUSIONS: The diagnosis of CVT should be considered in women with any neurological symptoms during pregnancy or puerperium and in all cases of unexplained intracerebral haemorrhage. CVT should also be considered in cases of recent onset and progressive headache, particularly when associated with focal neurological symptoms or signs, seizures or papilloedema. Magnetic resonance imaging with magnetic resonance venography is the investigation of choice. Anticoagulation with heparin remains the mainstay of treatment, even in the presence of intracerebral haemorrhage.

Anaphylaxis to celecoxib.

BACKGROUND: Adverse reactions such as urticaria, angioedema, asthma, and anaphylaxis are known to be associated with nonsteroidal anti-inflammatory agents (NSAIDs). Celecoxib (Pfizer/Searle, Caguas, PR) is a new NSAID that differs in structure and mechanism of action of other similar drugs of this class. OBJECTIVE: Evaluation of a case of anaphylaxis to celecoxib (Celebrex). METHODS AND RESULTS: This report describes a 55-year-old woman who experienced the acute onset of pruritus, urticaria, respiratory distress, and hypotension minutes after ingesting a celecoxib capsule. She had taken the drug a previous time for tendonitis without difficulty. Treatment with epinephrine, corticosteroids, and intravenous fluids was successful. An IgE mechanism could not be detected. She has avoided the drug and has had no further problems. CONCLUSIONS: This is the first patient report of anaphylaxis attributable to celecoxib, a new NSAID. This suggests that physicians and other health care professionals should be aware of the potential serious side effects of this drug.

Myasthenia gravis with thymoma is more common in the Maori and Pacific Island populations in New Zealand.

BACKGROUND: The association of myasthenia gravis (MG) with thymoma is well recognized. Our clinical impression has been that MG associated with thymoma may be more common in patients of Polynesian descent than in other races. AIM: To determine the influence of ethnicity on the association of MG with thymoma in our population. METHOD: Review of all cases of thymectomy performed at Greenlane Hospital in Auckland for the 20-year period from June 1978 to June 1998. RESULTS: There were 103 thymectomies performed in the study period. Fifty-five thymomas were identified, 15 in subjects of Maori or Pacific Island ethnicity and 40 in subjects of other races, predominantly Caucasian. Ten of 15 Maori or Pacific Island subjects with thymoma had MG (67%), compared with 15 of 40 subjects of other races (37.5%, P = 0.05). The mean age of Maori or Pacific Island subjects with thymoma and MG was 42.5 years, compared with 56.3 years in subjects from other races (P = 0.06). All five Maori and Pacific Island subjects with invasive thymoma had MG, whereas only four of 15 subjects (27%) from other races with invasive tumours had MG (P < 0.01). The overall incidence of thymoma and the proportion of thymomas that were invasive did not differ between the ethnic groups. CONCLUSIONS: Myasthenia gravis with thymoma occurs more frequently among Maori or Pacific Island people than in other racial groups in our population. This is due to an increase in the proportion of cases with thymoma who have MG in this group, while the overall frequency of cases of thymoma is similar between groups. MG with thymoma in the Maori or Pacific Island populations also presents at a younger age and is more often associated with tumour invasion.

T cell proliferation and cytokine secretion to T cell epitopes of Asp f 2 in ABPA patients.

The pathogenesis of allergic bronchopulmonary aspergillosis (ABPA) involves specific cytokines secreted by lymphocytes in response to Aspergillus fumigatus (Af) allergens. To gain information about the lymphoproliferative response and cytokine production against a major Af allergen, Asp f 2, we studied Asp-f-2-specific T cell clones (TCCs) from ABPA patients. TCCs were stimulated with rAsp f 2, its deletion mutants, and synthetic peptides to identify the T cell epitope(s) and to understand cytokine production. PBMCs from four of five ABPA patients showed proliferation in response to Asp f 2. Three TCCs from one patient showed higher IL-5 secretion compared to IL-4 and IFN-gamma. Two TCCs from the second patient showed a mixed Th1/Th2 response, as evidenced by production of IL-4, IL-5, and IFN-gamma. An epitope from the N-terminal region of Asp f 2 induced only IL-5 secretion. High IL-5 secretion might explain the marked eosinophilia observed in ABPA patients.

Cloning and expression of Aspergillus fumigatus allergen Asp f 16 mediating both humoral and cell-mediated immunity in allergic bronchopulmonary aspergillosis (ABPA).

BACKGROUND: Aspergillus fumigatus, a ubiquitous fungus, is responsible for a number of lung disorders in atopic and non-atopic individuals. Standardized, pure, and relevant allergens are desirable for reliable immunodiagnosis of the disease and to understand the structural and functional properties of these allergens and the role they play in causing ABPA. OBJECTIVE: Molecular cloning and characterization of a relevant allergen from A. fumigatus cDNA library. MATERIALS AND METHODS: A cDNA library was constructed from 96 h old mycelium of A. fumigatus using lambda ZAP expression vector. A novel gene encoding an A. fumigatus allergen was identified by screening the library with sera from ABPA patients. The gene was cloned and the allergen over-expressed in Escherichia coli. This recombinant allergen, Asp f 16, was evaluated in ELISA and Western blots using sera from patients and normal subjects and peripheral blood mononuclear cells (PBMC) for antigen-induced stimulation. RESULTS: Seventy percent of the patients with ABPA demonstrated high levels of serum IgE antibodies to Asp f 16, a 43-kDa protein, whereas patients with allergic asthma, Aspergillus skin test-positive asthmatics without clinical evidence of ABPA, and normal controls failed to show Asp f 16-specific IgE binding by ELISA. The deduced amino acid sequences of Asp f 16 showed extensive sequence homology to 30.6-kDa Asp f 9 at the N-terminal region of the protein. PBMC from the majority of patients with ABPA exhibited significant proliferation with the recombinant Asp f 16 allergen. CONCLUSION: Specific humoral and cell-mediated immune responses of Af-sensitized patients against Asp f 16 suggest its usefulness in the immunodiagnosis of hypersensitivity diseases due to Af and understanding the pathophysiology of ABPA.

Hyperthyroidism complicating asthma treatment.

Asthma is one of the most common chronic medical conditions. The usual treatment includes quick relief bronchodilator medications of the sympathomimetic class and controller medications that may include the long-acting inhaled bronchodilator salmeterol. Mild adverse cardiac and central nervous system effects are common with these medications, requiring modifications in dose or occasionally switching to a different medication. Both asthma and thyroid disease are common disorders that occasionally occur together. Hyperthyroidism may exacerbate asthma. Many symptoms of hyperthyroidism are identical to the adverse effects of the commonly used inhaled bronchodilators and include tremor, nervousness, tachycardia, wide pulse pressure, palpitations, emotional lability, agitation, nightmares, aggressive behavior, and diarrhea. In this report we describe a patient with hyperthyroidism whose symptoms initially were thought to be adverse effects of the inhaled bronchodilator medications.

Antibody binding of deletion mutants of Asp f 2, the major Aspergillus fumigatus allergen.

Asp f 2, a 268 amino acid major allergen from Aspergillus fumigatus (Af) demonstrated nine linear IgE binding regions. It is not known whether any of these linear epitopes are also conformatory epitopes. Hence, we constructed deletion mutants of Asp f 2 devoid of one or more epitopes, and the IgE binding of these proteins with sera from patients with ABPA was compared with the full-length Asp f 2 expressed in E. coli and Pichia. The Pichia expressed protein reacted weakly with IgE, but strongly with IgG of ABPA sera compared to E. coli expressed Asp f 2. Weak IgE binding only was seen when the C-terminal or N-terminal was deleted, while depletion of both ends negated all reactivity. The monoclonal antibody IL-B8 and IgE and IgG of ABPA sera bound significantly to the Asp f 2 E-4 fragment indicating that the major B-cell epitope is located at the N-terminal end of Asp f 2.

Detection of immunoglobulin antibodies in the sera of patients using purified latex allergens.

BACKGROUND: Latex allergy is largely an occupational allergy due to sensitization to natural rubber latex allergens present in a number of health care and household products. Although several purified allergens are currently available for study, information on the usefulness of these purified, native or recombinant allergens in the demonstration of specific immunoglobulin (Ig) E in the sera of patients is lacking. OBJECTIVE: To evaluate the purified latex allergens and to demonstrate specific IgE antibody in the sera of health care workers and spina bifida patients with clinical latex allergy. METHODS: Two radioallergosorbent and an enzyme-linked immunosorbent assay (ELISA) using latex proteins Hev b 1, 2, 3, 4, 6 and 7 along with two glove extracts and Malaysian nonammoniated latex (MNA) were evaluated to demonstrate IgE in the sera of health care workers and spina bifida with latex allergy and controls with no history of latex allergy. RESULTS: ELISA using the purified latex allergens demonstrated specific IgE in 32-65% health care workers and 54-100% of spina bifida patients with latex allergy. The corresponding figures for RAST were 13-48 and 23-85 for RAST-1 and 19-61 and 36-57 for RAST-2. These results were comparable with the results obtained with glove extracts and crude rubber latex proteins. CONCLUSIONS: When used simultaneously, latex proteins Hev b 2 and Hev b 7 reacted significantly with specific serum IgE in 80% of health care workers and 92% of spina bifida patients with latex allergy by ELISA technique, while this combination gave lower positivity when the RASTs were used. By the addition of Hev b 3, specific IgE was detected in all spina bifida patients with latex allergy. Both RASTs failed to show specific IgE in the control subjects, while the ELISA showed significant latex-specific IgE in 22% of controls.

Unique and shared IgE epitopes of Hev b 1 and Hev b 3 in latex allergy.

Of the several latex proteins cloned and expressed, the rubber elongation factor, Hev b 1, and the closely related Hev b 3, represent two major allergens associated with latex allergy. Although both allergens demonstrated IgE binding with sera from latex allergic patients, it was not known whether these two molecules shared any epitopes. Hence, in the present study using health care workers (HCW) and spina bifida (SB) patients with latex allergy, we investigated the IgE binding epitopes in Hev b 1 and Hev b 3. Recombinant Hev b 1 and Hev b 3 were expressed in a prokaryotic expression system, while overlapping decapeptides of Hev b 1 and Hev b 3 were synthesized on derivatized cellulose membrane. Eight IgE binding epitopes for Hev b 1 and eleven for Hev b 3 were identified using sera from latex allergic patients with SB. On further analysis of synthetic peptides encompassing these epitopes, similar IgE antibody reactivity was demonstrated with three Hev b 1 epitopes b1E3, b1E5, b1E6 and two Hev b 3 epitopes; b3E10 and b3E 11. For Hev b 1, a unique IgE binding epitope was identified in the region of amino acid residues 16-25. In competitive ELISA, peptides bIE2 and bIE4 together inhibited 58% of IgE binding of Hev b 1, while b3E5 showed 22% inhibition in the IgE binding of Hev b 3. The results of the present study suggest that the understanding of linear and conformational IgE epitopes in the major latex allergens may provide better insight into the structure-function relationship of the allergens, and may lead to the development of better patient care and management strategies in latex allergy.

Therapy of allergic bronchopulmonary aspergillosis.

The management of ABPA depends on the extent and stage of the disease. Underlying asthma should be controlled with environmental changes, pharmaco- and immunotherapy. Baseline examinations and evaluations of pulmonary function, airway and parenchymal anatomy, and serum total IgE levels are important and should be re-evaluated based on the clinical course of the patient. The mainstay of pharmacotherapy for ABPA remains oral corticosteroids. The dose and duration of treatment in the initial stage of the disease depends on when it was diagnosed as well as the patient's clinical course. Anti-fungal agents should be considered as adjunctive. Clinical data suggests that the early institution of treatment is likely to prevent progression of ABPA to end-stage fibrosis.

Molecular characterization of aspergillus fumigatus allergens.

Aspergillus fumigatus (Af) is ubiquitous saprophytic fungus associated with a broad spectrum of diseases in humans. These diseases range from benign colonization of the lung to life threatening diseases such as allergic bronchopulmonary aspergillosis (ABPA) and invasive aspergillosis. Af is the etiologic agent identified in most of the Aspergillus related human diseases and is therefore of particular clinical importance. Af induced obstructive airway diseases may be due to transient exposure to fungal spores resulting in a T helper 2 response. The IgE mediated inflammatory reaction could be due to colonization of bronchial airway epithelium by Af. Early and precise diagnosis of Aspergillus induced respiratory allergy is essential for preventing irreversible lung damages. The major problems in the diagnosis of A. fumigatus induced diseases are due to the lack of standardized and well characterized fungal extracts. The advent of molecular cloning technology and the development of phage surface display technology for cloning genes have facilitated the isolation of more relevant recombinant allergens. Using these techniques, a panel of different Af allergens having distinct IgE binding with various groups of Af sensitized patients have been cloned and characterized. These allergens can be categorized functionally as secreted and cytoplasmic proteins. The distinct IgE binding property of these purified and well characterized recombinant Af allergens may be useful for the differential diagnosis of Af related pulmonary complications.

Molecular biology and immunology of fungal allergens.

Fungi are non-chlorophyllus microorganisms, which constitutes the main source of outdoor and indoor allergens. The antigens present in the spores and fragments of hyphae induce allergic responses in sensitized patients. The frequently recognized fungi associated with asthma include Alternaria, Cladosporium, Aspergillus, and Penicillium. With the advent of molecular biology techniques a number of fungal genes encoding relevant allergens have been cloned and the expressed allergens purified and characterized. In this review, we have presented the recent developments, where recombinant allergens have been used in the precise diagnosis of fungal allergy. We have also discussed the role played by these allergens and the T- and B-cell epitopes in the immune mechanism in fungal allergy.