Animal models of allergic bronchopulmonary aspergillosis

Among the allergic fungi, Aspergillus fumigatus, a saprophytic mold, distributed widely in the environment is a frequently recognized etiologic agent in a number of allergic conditions. Among the different allergic diseases caused by this fungus, allergic bronchopulmonary aspergillosis (ABPA) is by far the most significant one. The immunopathogenesis of this disease is not fully understood. Although several immunomodulatory treatments are available for allergic disease, none of them are applicable or relevant or useful in fungal induced allergy. It is essential to understand the pathogenesis of the disease including the antigen induced immunoregulation and the resulting factors, such as cytokine, chemokines, pathways activating factors, inflammatory and airway remodeling factors need to be understood for intervening with appropriate treatment. Animal models are essential in understanding these features of the disease. Several models of allergic aspergillosis have been developed in recent years in various animals. However, murine models have been studied more carefully and extensively. The exposure to antigen in mice leads to allergy very similar to ABPA with high IgE, elevated peripheral blood and lung eosinophils, pulmonary inflammation, and airway hyperreactivity. The role of various cytokines and chemokines and their receptors were also studied. In addition, immunotherapy and vaccination have been attempted in recent years using the murine model of ABPA. This review covers the murine model of Aspergillus induced allergy and asthma and presented critically our current understanding of the subject and the potential application of such a model in future for developing treatment modalities. This revised version was published online in June 2006 with corrections to the Cover Date.     

变态反应性支气管肺曲霉病临床特点探讨

目的分析变态反应性支气管肺曲霉病(ABPA)的临床特征,以期早期诊断,合理治疗及预防。方法回顾性分析浙江大学医学院附属第一医院2009年6月至2011年7月收治的13例ABPA患者的临床资料,所有患者均依照目前通用的ABPA诊断标准进行诊断,分析其临床表现,影像学特征,治疗方案及疗效等。结果 ABPA好发于中老年患者,且都好发支气管扩张及哮喘患者。13例ABPA患者均有咳嗽,咳痰,8例气喘,4例发热,9例肺部可闻及哮鸣音。13例ABPA患者中有12例CT检查发现囊状或静脉曲张样中心性支气管扩张和支气管粘液栓形成及树芽征为主要特点的小叶中心结节形成。总IgE升高12例,嗜酸粒细胞绝对值升高6例,烟曲菌特异性IgE升高10例。12例接受激素及抗真菌药物治疗,均达到临床症状好转。结论 ABPA临床表现缺乏特异性,为提高对ABPA的认识,对可疑患者早期进行外周血嗜酸性粒细胞数、血清总IgE、烟曲霉菌特异性IgE、肺部CT等检查以确诊,糖皮质激素及伊曲康唑治疗ABPA能取得良好的疗效。     

Diagnosis and treatment of allergic bronchopulmonary aspergillosis

Allergic bronchopulmonary aspergillosis (ABPA) is a rare form of asthma that is most common in corticosteroid-dependent asthmatic patients and cystic fibrosis patients. It is caused by an abnormal T-helper class 2 response of the host to Aspergillus antigens. Although signs and symptoms of typical asthma are usually present with ABPA, unusual features such as fever, expectoration of brown plugs, and central bronchiectasis with or without mucoid impaction on chest radiographs may be present. ABPA is important to diagnose because inadequate therapy may lead to permanent lung destruction. Corticosteroids are the drug of choice for ABPA; however, the doses required are often greater than for routine asthma and corticosteroid dependence is not unusual. Azole therapy appears to have an adjunctive role in treatment in terms of improving signs and symptoms of the disease and demonstrating a corticosteroid-sparing effect.     

变应性支气管肺曲霉病的CT表现——附17例分析

目的探讨变应性支气管肺曲霉病(ABPA)的CT特点,进一步提高对该病的认识。方法回顾性分析17例AB-PA的CT表现,所有的患者均按目前的ABPA诊断标准进行诊断。结果 17例患者均有中心性支气管扩张征象,13例并支气管黏液栓形成,为分支状或挤牙膏状或长管状"Y"形,7例以树芽征为主要表现的小叶中心结节形成,9例有斑片状浸润影,2例随访见中心性支气管扩张有游走性。结论变应性支气管肺曲霉病(ABPA)的CT表现相对有特征性,以中心性支气管扩张为主,常伴有较高密度的支气管腔内黏液栓形成,结合临床一般能做出诊断。     

A murine model of allergic bronchopulmonary aspergillosis with elevated eosinophils and IgE.

A model of allergic bronchopulmonary aspergillosis was developed by exposing BALB/c mice to Aspergillus fumigatus (AF) Ag. Animals immunized intranasally (i.n.) with soluble AF Ag produced low levels of serum IgE compared to animals given alum precipitated AF Ag i.p. The latter treatment also produced higher levels of serum IgG1 and AF-specific IgG1 than soluble AF given i.p. or i.n.. Blood and lung eosinophilia was detected in mice repeatedly exposed to AF by i.n. but not in the groups injected i.p. Particulate AF Ag-induced striking blood and lung eosinophilia and elevated levels of serum IgE in mice preexposed to AF Ag. The results indicate that route of inoculation and physical nature of Ag determine the immune response and can be manipulated to obtain enhanced IgE, eosinophils, or both in the animal model.     

Identification of 45 kD antigen in immune complexes of patients of allergic bronchopulmonary aspergillosis

A glycoprotein antigen with an apparent mw of 45 kD was observed to be predominant in the circulating immune complexes isolated from patients of allergic bronchopulmonary aspergillosis as well as in the immune complexes prepared in vitro. Further characterisation of 45 kD antigen with trypsin showed four immunologically active peptides with mw 43, 36, 33 and 16 kD. Carbohydrate characterization using various lectins (Maackia amurensis agglutinin, Sambucus nigra agglutinin, Peanut agglutinin, Galanthus nivalis agglutinin and Datura stramonium agglutinin) showed presence of both N-linked and O-linked sugar moieties such as mannose, glucose, galactose and N-acetyl glucosamine. Predominant presence of 45 kD antigen in immune complexes, recognition of 45 kD by monoclonal antibodies raised against glycoprotein rich fraction of A. fumigatus and presence of elastinolytic protease activity indicate that 45 kD antigen is probably a potent virulence factor and may be contributing to the pathogenesis of ABPA by its biological as well as immunological activities. (Mol Cell Biochem 166: 111-116, 1997)     

Immunomodulatory role of C10 chemokine in a murine model of allergic bronchopulmonary aspergillosis

The immunomodulatory role of the chemokine C10 was explored in allergic airway responses during experimental allergic bronchopulmonary aspergillosis (ABPA). The intratracheal delivery of Asperigillus fumigatus Ag into A. fumigatus-sensitized mice resulted in significantly increased levels of C10 within the bronchoalveolar lavage, and these levels peaked at 48 h after A. fumigatus challenge. In addition, C10 levels in BAL samples were greater than 5-fold higher than levels of other chemokines such as monocyte-chemoattractant protein-1, eotaxin, and macrophage-inflammatory protein-1alpha. From in vitro studies, it was evident that major pulmonary sources of C10 may have included alveolar macrophages, lung fibroblasts, and vascular smooth muscle cells. Experimental ABPA was associated with severe peribronchial eosinophilia, bronchial hyperresponsiveness, and augmented IL-13 and IgE levels. The immunoneutralization of C10 with polyclonal anti-C10 antiserum 2 h before the intratracheal A. fumigatus challenge significantly reduced the airway inflammation and hyperresponsiveness in this model of ABPA, but had no effect on IL-10 nor IgE levels. Taken together, these data suggest that C10 has a unique role in the progression of experimental ABPA.     

Cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations in allergic bronchopulmonary aspergillosis.

The etiology of allergic bronchopulmonary aspergillosis (ABPA) is not well understood. A clinical phenotype resembling the pulmonary disease seen in cystic fibrosis (CF) patients can occur in some individuals with ABPA. Reports of familial occurrence of ABPA and increased incidence in CF patients suggest a possible genetic basis for the disease. To test this possibility, the entire coding region of the cystic fibrosis transmembrane regulator (CFTR) gene was analyzed in 11 individuals who met strict criteria for the diagnosis of ABPA and had normal sweat electrolytes (< or = 40 mmol/liter). One patient carried two CF mutations (deltaF508/R347H), and five were found to carry one CF mutation (four deltaF508; one R117H). The frequency of the deltaF508 mutation in patients with ABPA was significantly higher than in 53 Caucasian patients with chronic bronchitis (P < .0003) and the general population (P < .003). These results suggest that CFTR plays an etiologic role in a subset of ABPA patients.     

Management of allergic aspergillosis

Treatment of allergic bronchopulmonary aspergillosis depends on accurate diagnosis, which is difficult because of multiple clinical, radiographic, and immunologic criteria; unstandardized immunoassays; and its similarity to cystic fibrosis. Advances in chest CT interpretation, measuring serum Aspergillus fumigatus-specific antibodies and the T_H2 chemokine thymusactivated-and-regulated chemokine, recognizing risk factors, should improve diagnostic accuracy. Oral glucocorticosteroids remain the mainstay of treatment. Dose regimen and duration are not standardized, so expert recommendations are used. Inhaled steroids have not been proven to be effective. Due to relapse and toxicity, itraconazole has been added as second-tier therapy, supported by randomized, placebo-controlled trials in asthma and open-label trials in cystic fibrosis. Itraconazole is steroid sparing with anti-inflammatory aspects and poor bioavailability. Despite combination steroid-itraconazole therapy, relapses and steroid toxicity have led to open-label use of voriconazole, nebulized amphotericin B, monthly high-dose intravenous methylprednisolone, and omalizumab. Controlled trials are needed to evaluate these and other antifungal or immunomodulatory interventions.     

An alternate method of descenting skunks

Striped skunk (Mephitis mephitis) scent glands were ligated closed with waxed dental floss to allow them to be handled during toxicological studies without fear of scenting. This descenting technique was more rapid and less traumatic than scent gland removal. Thirty-four skunks were kept for less than or equal to 127 days and did not display behavioral or physical abnormalities due to this procedure.     

Recurrence of allergic bronchopulmonary aspergillosis after adjunctive surgery for aspergilloma: a case report with long-term follow-up

Background

Coexistence of aspergilloma and allergic bronchopulmonary aspergillosis (ABPA) has rarely been reported. Although the treatment for ABPA includes administration of corticosteroids and antifungal agents, little is known about the treatment for coexisting aspergilloma and ABPA. Furthermore, the impact of surgical resection for aspergilloma on ABPA is not fully understood. Here, we present an interesting case of recurrent ABPA with long-term follow-up after surgical resection of aspergilloma.

Case presentation

A 53-year-old man with a medical history of tuberculosis was referred to our hospital with cough and dyspnea. Imaging revealed multiple cavitary lesions in the right upper lobe of the lung, with a fungus ball and mucoid impaction. The eosinophil count, total serum immunoglobulin E (IgE), and Aspergillus-specific IgE levels were elevated. Specimens collected on bronchoscopy revealed fungal filaments compatible with Aspergillus species. Based on these findings, a diagnosis of ABPA with concomitant aspergilloma was made. Although treatment with corticosteroids and antifungal agents was administered, the patient’s respiratory symptoms persisted. Therefore, he underwent lobectomy of the right upper lobe, which resulted in a stable condition without the need for medication. Twenty-three months after discontinuation of medical treatment, his respiratory symptoms gradually worsened with a recurrence of elevated eosinophil count and total serum IgE. Imaging revealed recurrent bronchiectasis and cavities with mucoid impaction in the right lower lobe, suggesting relapse of aspergilloma and ABPA. Corticosteroids and antifungal agents were re-administered; aspergilloma improved slightly over a 5-year period, and ABPA remained well controlled with low-dose prednisolone (5?mg/day).

Conclusions

We describe the long-term follow-up outcomes of a patient with concomitant ABPA and aspergilloma, who underwent surgical resection for aspergilloma. Physicians should carefully monitor patients with coexisting ABPA and aspergilloma, as the condition may relapse after remission, even despite surgical resection for aspergilloma. Additionally, surgical resection for aspergilloma could result in resolution of ABPA.

     

An alternate method for demonstration of erythrocytic aminotransferases on starch gels

A two-stage procedure using MTT tetrazolium for the demonstration of aminotransferases (GPT and GOT) either singly or together was developed. Identification of phenotypes was unequivocal in over 99% of the individuals studied.     

An alternate method for synthesis of double-stranded DNA segments

Recent progress in the chemical synthesis of DNA has now made it possible to rapidly synthesize single-stranded DNAs over 40 bases in length. We have taken advantage of these longer DNAs in assembling and cloning a 132-base pair gene segment coding for amino acids 126 through the stop codon of human leukocyte interferon alpha 2. The method used involves DNA polymerase I-mediated repair synthesis of synthetic oligonucleotide substrates having short stretches of complementary sequence at their 3' termini. In the presence of DNA polymerase I and the four deoxyribonucleoside triphosphates, those primer-templates are converted to full length double-stranded DNAs. The economy in chemical synthesis using this approach is substantial with a greater than 40% reduction in the amount of chemical synthesis required as compared with the conventional approach. We describe in detail this methodology for the biochemical assembly of long gene segments from synthetic oligodeoxyribonucleotides.     

基于线性判别分析的基因表达数据分类方法研究

由于基因表达数据高属性维、低样本维的特点,Fisher分类器对该种数据分类性能不是很高。本文提出了Fisher的改进算法Fisher-List。该算法独特之处在于为每个类别确定一个决策阀值,每个阀值既包含总体样本信息,又含有某些对分类至关重要的个体样本信息。本文用实验证明新算法在基因表达数据分类方面比Fisher、LogitBoost、AdaBoost、k-近邻法、决策树和支持向量机具有更高的性能。     

An alternate and reversible method for flight restraint of cranes

Flight restraint is important for zoos, safaris, and breeding centers for large birds. Currently used techniques for flight restraint include both surgical and non-surgical approaches. Surgical approaches usually cause permanent change to or removal of tendon, patagial membrane, or wing bones, and can cause pain and inflammation. Non-surgical approaches such as clipping or trimming feathers often alter the bird's appearance, and can damage growing blood feathers in fledglings or cause joint stiffness. We observed microstructure of primary feathers of the red-crowned crane (Grus japonensis) and found that the width of barbs is a determinative factor influencing vane stiffness and geometric parameters. We hypothesized that partial longitudinal excision of barbs on the ventral surface of the primary feathers would reduce the stiffness of the vane and render the feathers unable to support the crane's body weight during flight. Furthermore, we hypothesized that this modification of barbs would also change the aerodynamic performance of feathers such that they could not generate sufficient lift and thrust during flapping to enable the bird to fly. We tested this hypothesis on a red-crowned crane that had normal flight capability by excising the ventral margin of barbs on all 10 primaries on the left wing. The bird was unable to take off until the modified feathers were replaced by new ones. Removal of barbs proved to be a simple, non-invasive, low-cost and reversible method for flight restraint. It is potentially applicable to other large birds with similar structural characteristics of primary feathers.