Identification of allergens from Aspergillus fumigatus-Potential association with lung damage in asthma.

BACKGROUND: Component-resolved diagnosis allows detection of IgE sensitization having the advantage of reproducibility and standardization compared to crude extracts. The main disadvantage of the traditional allergen identification methods, 1- or 2-dimensional western blotting and screening of expression cDNA libraries with patients' IgEs, is that the native structure of the protein is not necessarily maintained. METHODS: We used a novel immunoprecipitation technique in combination with mass spectrometry to identify new allergens of Aspergillus fumigatus. Magnetic Dynabeads coupled with anti-human IgE antibodies were used to purify human serum IgE and subsequently allergens from A. fumigatus protein extract. RESULTS: Of the 184 proteins detected by subsequent mass peptide fingerprinting, a subset of 13 were recombinantly expressed and purified. In a panel of 52 A. fumigatus-sensitized people with asthma, 23 non-fungal-sensitized asthmatics and 18 healthy individuals, only the former showed an IgE reaction by immunoblotting and/or ELISA. We discovered 11 proteins not yet described as A. fumigatus allergens, with fructose-bisphosphate aldolase class II (FBA2) (33%), NAD-dependent malate dehydrogenase (31%) and Cu/Zn superoxide dismutase (27%) being the most prevalent. With respect to these three allergens, native versus denatured protein assays indicated a better recognition of the native proteins. Seven of 11 allergens fulfilled the WHO/IUIS criteria and were accepted as new A. fumigatus allergens. CONCLUSION: In conclusion, we introduce a straightforward method of allergen identification from complex allergenic sources such as A. fumigatus by immunoprecipitation combined with mass spectrometry, which has the advantage over traditional methods of identifying allergens by maintaining the structure of the proteins.

The airway fungal microbiome in asthma.

BACKGROUND: Fungal involvement in asthma is associated with severe disease. The full spectrum of fungal species in asthma is not well described and is derived largely from insensitive culture techniques. OBJECTIVES: To use high-throughput sequencing to describe the airway mycobiota in asthmatics with and without fungal sensitization and healthy controls; to compare samples representing different airway compartments; to determine whether the mycobiota was influenced by the fungal composition of outdoor air; and to compare findings with clinically relevant outcomes. METHODS: We amplified the internal transcribed spacer region 2 of the nuclear ribosomal operon to identify the fungal species present. Ninety-seven subjects were recruited and provided sputum (83 asthmatics; 14 healthy subjects), with 29 also undergoing a bronchoscopy. A subset of airway samples were compared with matched outdoor air and mouthwash samples. RESULTS: Two hundred and six taxa at the species level were identified in sputum, most at low relative abundance. Aspergillus fumigatus, Candida albicans and Mycosphaerella tassiana had the highest relative abundances and were the most prevalent species across all subjects. The airway mycobiota consisted of a complex community with high diversity between individuals. Notable shifts in the balance of fungi detected in the lung were associated with asthma status, asthma duration and biomarkers of inflammation. Aspergillus tubingensis, a member of the Aspergillus niger species complex, was most prevalent from bronchoscopic protected brush samples and significantly associated with a low sputum neutrophilia. Cryptococcus pseudolongus, from the Cryptococcus humicola species complex, was more abundant from bronchoscopy samples than sputum, and differentially more abundant in asthma than health. CONCLUSIONS AND CLINICAL RELEVANCE: The airway mycobiota was dominated by a relatively small number of species, but was distinct from the oropharyngeal mycobiota and air samples. Members of the A. niger and C. humicola species complexes may play unexpected roles in the pathogenesis of asthma.

IRP2 as a potential modulator of cell proliferation, apoptosis and prognosis in nonsmall cell lung cancer.

IREB2 is a gene that produces iron regulatory protein 2 (IRP2), which is critical to intracellular iron homeostasis and which relates to the rate of cellular proliferation. IREB2 lies in a lung cancer susceptibility locus. The aims were to assess 1) the relationship between iron loading, cell proliferation and IRP2 expression in lung cancer; 2) the potential of iron related pathways as therapeutic targets; and 3) the relevance of IRP2 in operated lung cancer patients.Cells of two nonsmall cell cancer (NSCLC) lines and primary bronchial epithelial cells (PBECs) were cultured with and without iron; and proliferation, apoptosis and migration were assessed. Reverse transcriptase PCR and Western blot were used to assess expression of iron homeostasis genes/proteins. Iron chelation and knockdown of IREB2 were used in vitro to explore therapeutics. A cohort of operated NSCLC patients was studied for markers of systemic iron status, tumour IRP2 staining and survival.Iron loading caused cell proliferation in cancer cell lines, which were less able to regulate IREB2 expression than PBECs. Iron chelation resulted in a return of proliferation rates to baseline levels; knockdown of IREB2 had a similar effect. IRP2-positive tumours were larger (p=0.045) and higher percentage staining related to poorer survival (p=0.079).Loss of iron regulation represents a poor prognostic marker in lung cancer.

Eosinophilia in Pulmonary Disorders.

Lung disease associated with marked peripheral blood eosinophilia is unusual and nearly always clinically significant. Once recognized, it is generally easy to manage, albeit with long-term systemic corticosteroids. A failure to respond to oral steroids in the context of good compliance suggests a malignant cause for the eosinophilia. An important development is the introduction of antieosinophil therapies, particularly those directed against the interleukin 5 pathway, which is hoped to provide benefit in the full spectrum of eosinophilic lung disease as well as asthma, reducing the burden of side effects and resultant comorbidities.

Allergic fungal airway disease: pathophysiologic and diagnostic considerations.

PURPOSE OF REVIEW: Fungal spores are ubiquitously present in indoor and outdoor air. A number can act as aeroallergens in Immunoglobulin E (IgE)-sensitized individuals and some thermotolerant fungi germinate in the lung where they can cause a combined allergic and infective stimulus leading to a number of clinical presentations characterized by evidence of lung damage. We discuss which biomarkers are useful in helping to guide diagnosis, prognosis and treatment of allergic fungal airway disease (AFAD). RECENT FINDINGS: Diagnostic biomarkers, such as specific IgEs and fungal culture, for AFAD are limited by sensitivity, although this may be improved with novel agents such as specific IgEs to fungal components and quantitative PCR. Total IgE and hypereosinophilia are nonspecific and do not clearly relate to disease activity. High attenuation mucus and proximal bronchiectasis are specific, albeit insensitive markers of AFAD. Biomarkers that predict prognosis and treatment response are yet to be defined. SUMMARY: This review summarizes the fungi involved and the current debate regarding the diagnostic criteria to define fungal-associated lung disease. We advocate the phasing out of the term allergic bronchopulmonary aspergillosis and the use of a more inclusive term such as AFAD, together with a more liberal set of criteria based largely on IgE sensitization to thermotolerant fungi, which identifies those patients at risk of developing lung damage.

Assessing the risk of attack in the management of asthma: a review and proposal for revision of the current control-centred paradigm.

Asthma guidelines focus on day-to-day control of symptoms. However, asthma attacks remain common. They continue to cause mortality and considerable morbidity, and are a major financial burden to the UK National Health Service (NHS) and the wider community. Asthma attacks have chronic consequences, being associated with loss of lung function and significant psychological morbidity. In this article we argue that addressing daily symptom control is only one aspect of asthma treatment, and that there should be a more explicit focus on reducing the risk of asthma attacks. Management of future risk by general practitioners is already central to other conditions such as ischaemic heart disease and chronic renal impairment. We therefore propose a revised approach that separately considers the related domains of daily control and future risk of asthma attack. We believe this approach will have advantages over the current 'stepwise' approach to asthma management. It should encourage individualised treatment, including non-pharmacological measures, and thus may lead to more efficacious and less harmful management strategies. We speculate that this type of approach has the potential to reduce morbidity and healthcare costs related to asthma attacks.

Acid suppression does not reduce symptoms from vocal cord dysfunction, where gastro-laryngeal reflux is a known trigger.

Vocal cord dysfunction (VCD) is characterized by paradoxical inspiratory abduction of the vocal cords. Gastro-oesophageal reflux disease (GORD) is a known trigger. We studied 77 patients referred to a tertiary VCD clinic. VCD was diagnosed in 62, of which 83.9% had proven GORD. Following 8 weeks of acid suppression, 24.2% reported improvement in the severity and frequency of VCD attacks. This study suggests that empirical treatment of GORD in VCD marginally improves symptom control.

Polycystic liver disease presenting with an exudative pleural effusion: a case report.

INTRODUCTION: Polycystic liver disease is asymptomatic in 95% of patients. In the remaining 5% it causes symptoms due to the local mass effect of the polycystic liver. We describe the case of a patient who presented with symptoms of a pleural effusion and was also found to have polycystic liver disease. The effusion recurred despite repeated efforts at drainage and only resolved following surgical debridement of the cystic liver. CASE PRESENTATION: A 50-year-old Caucasian woman presented with a two-week history of increasing dyspnoea. An examination revealed a large right pleural effusion and gross hepatomegaly. An ultrasound confirmed a large polycystic liver and diagnostic thoracocentesis revealed an exudate, which was sterile to culture. The pleural effusion proved refractory to drainage and our patient underwent surgery to deroof the main hepatic cysts in an attempt to reduce the pressure on her right diaphragm. The histology was compatible with that of polycystic liver disease. No evidence of malignancy was found. After surgery, our patient had no recurrence of her effusion and, to date, has remained asymptomatic from her polycystic liver disease. CONCLUSION: The case in this report illustrates that an exudative pleural effusion is a rare complication of polycystic liver disease. We feel that the mechanical effects of a large polycystic liver, and subsequent disruption of sub-diaphragmatic capillaries, resulted in a persistent exudative pleural effusion. Thus, surgical debulking of the hepatic cysts is required to manage these effusions.