Establishment of Rat Models of Different Pathological Types of Hypersensitivity Pneumonitis Using Pigeon Droppings.

BACKGROUND: The pathogenesis and pulmonary histopathological characteristics of hypersensitivity pneumonitis (HP) are not yet fully understood. Therefore, we established animal models of HP of different stages, aiming to provide support for research on this disease. METHODS: We established rat models of pigeon breeder's lung of different pathological types by creating freeze-dried allergen powder from fresh pigeon feathers, dander, and other droppings. Freeze-dried allergen powder suspensions of pigeon droppings were used to establish 2 rat models of HP, one by aerosol inhalation and one by airway instillation, and the rats were sacrificed after different lengths of time to observe the pathological changes in their lung tissues. RESULTS: By the 40th week after allergen inhalation, granulomas were the main changes in the model, without fibrotic changes. When using airway instillation to establish the model, at the 20th week, group 1 (low dose + twice/week) and group 2 (medium dose + twice/week) showed granuloma changes, but no fibrosis; group 3 (high dose + once/week) and group 4 (high dose + twice/week) both showed obvious pulmonary fibrotic changes, but the death rate of rats in group 4 was greater. CONCLUSIONS: Both aerosol inhalation and airway instillation of freeze-dried pigeon allergen powder can successfully establish an HP model. The airway instillation method can cause pulmonary fibrotic changes in a short time, and the pulmonary pathological changes of animal models manifest with an obvious time-dose effect.

Frequency of goose and duck down causation of hypersensitivity pneumonitis within an 80-patient cohort.

BACKGROUND: A variety of antigens have been identified as causative of hypersensitivity pneumonitis (HP), which is characterized by inflammation to the lung parenchyma that is induced by exposure. Goose and duck down (GDD) bedding is often overlooked by physicians as a potential cause, yet the use of GDD has markedly increased in recent years, paralleling an increased frequency of reports of GDD-induced HP. OBJECTIVE: To determine the frequency of GDD as the causative antigen in patients with HP who use bedding that contains GDD. METHODS: Patients referred with a working diagnosis of HP underwent a detailed environmental history. Those who were using GDD were asked to remove it as an avoidance procedure. Signs, symptoms, spirometry, and inflammatory markers were followed up at weekly intervals for up to 1 month to determine the effect of remediation. RESULTS: Eighty patients with HP were seen during an 8-year period. Thirty-two patients (40%) were using GDD bedding. Of these 32 patients, 12 (37.5% of those exposed and 15% of the total HP population experienced remission (or nonprogression) of disease by simply avoiding GDD bedding. Eleven (92%) of these 12 patients were female. In patients with GDD-induced HP, lung biopsy patterns were varied. CONCLUSION: Approximately one-third of patients with HP, who slept with GDD, had persistent improvement or remission with simple avoidance. The higher incidence of GDD-induced HP in females may be hormonal and/or sociocultural related. Lung biopsy findings were across the spectrum of histopathologic patterns. Avoidance-challenge techniques were effective in confirming diagnoses and causation and mitigating the need for additional remediation.

T lymphocytes dominate local leukocyte infiltration in response to intradermal injection of functionalized graphene-based nanomaterial.

Graphene-based nanomaterials (GBN) have many potential biomedical applications. However, information regarding their biological properties and interactions with cells and/or soluble factors within a complex tissue is limited. The objective of this study was to use the growing feather (GF) of chickens as a minimally invasive cutaneous test-site to assess and monitor leukocyte recruitment in response to intradermal GBN injection. Specifically, the dermis of 20 GFs per chicken was injected with 10 µl of phosphate-buffered saline (PBS)-vehicle or 10 µl of 300 µg ml-1 oxygen-functionalized (f) GBN (6 chickens/treatment). GFs were collected before- (0) and at 0.25, 1, 2, 3, 4, 5, and 7 days post-injection and used for leukocyte-population analysis of immunofluorescently stained pulp cell suspensions or histological examination. Based on flow-cytometric cell population analysis, lymphocytes and macrophages were the major leukocyte-populations infiltrating GFs in response to f-GBN presence. Compared with PBS-controls, levels of T cells (γδ-, αß-, CD4- and CD8-T cells) were greatly elevated in f-GBN-injected GFs within 6 h and remained elevated throughout the 7-day examination period. f-GBN's effects on local tissue leukocyte recruitment were not reflected in the blood, except for a higher percentage of lymphocytes on 7 days. These observations together with a visual examination of f-GBN-injected GF tissue-sections suggest a delayed-type hypersensitivity-like, inflammatory cell-mediated response to the non-biodegradable f-GBN. The GF 'in vivo test-tube'system together with blood sampling provided unique insight into the time-course, qualitative, and quantitative aspects of immune system activities initiated by the presence of f-GBN in a complex tissue of a living animal. Copyright © 2017 John Wiley & Sons, Ltd.

[Improved IgG Antibody Diagnostics of Feather Duvet Lung by an Antibody Screening Test]. / Optimierte IgG-Antikörperdiagnostik der Bettfedern-Alveolitis mittels EAA-Suchtest.

The underdiagnosed feather duvet lung, an extrinsic allergic alveolitis (hypersensitivity pneumonitis) caused by duck and goose feathers, can be more frequently diagnosed, if duck and goose feather antibodies are included in the panel of the routinely applied IgG antibody screening test. This does not necessarily require extending the screening test to include duck and goose feather antigens. By analysing 100 sera with duck and goose antibodies we found that the commonly used pigeon and budgerigar antibodies can also screen for feather duvet antibodies. All examined sera lacking pigeon and budgerigar antibodies also lacked clear-cut duck and goose feather antibodies. The examined sera with strong pigeon or budgerigar antibodies always also contained feather duvet antibodies. However, sera with medium or low concentrated pigeon or budgerigar antibodies are not always associated with feather duvet antibodies. In the light of these observations, we find that 71% of the duck and goose antibody analyses would be dispensable without essential loss of quality, if the results of screening for pigeon and budgerigar antibodies were incorporated into the procedure of a step-by- step diagnostics.

Paediatric feather duvet hypersensitivity pneumonitis.

A previously well 12-year-old boy was admitted with a second insidious episode of dyspnoea, dry cough, anorexia, weight loss and chest pain. At admission, he had an oxygen requirement, significantly impaired lung function and reduced exercise tolerance. Initial forced expiratory volume in 1 s was 26%; a 3 min exercise test stopped at 1 min 50 when saturations dropped to 85%. CT scan showed ground-glass nodularity with lymphadenopathy. Bronchoalveolar lavage (BAL) for Pneumocystis carinii pneumonia and viruses were negative, and microbiology results for the BAL were reported in the absence of histology. This is because at the time the BAL samples were collected, a lung biopsy was performed. The biopsy was consistent with hypersensitivity pneumonitis. Echo was normal and CT pulmonary angiography negative. After taking a thorough history, exposure to feather duvets prior to each episode was elicited. IgG of avian precipitants was raised at 10.6 mgA/L (normal <10 mgA/L). Clinical improvement began with avoidance of exposure, while the boy was an inpatient. Antigen avoidance continued on discharge. He continues to improve since discharge. The condition was diagnosed as hypersensitivity pneumonitis secondary to exposure to antigens from feather duvets.

Enzymatic hydrolyzed feather peptide, a welcoming drug for multiple-antibiotic-resistant Staphylococcus aureus: structural analysis and characterization.

This study aimed to explore the bactericidal activity of a feather-degraded active peptide against multiple-antibiotic-resistant (MAR) Staphylococcus aureus. An antibacterial peptide (ABP) was isolated from the chicken feathers containing fermented media of Paenibacillus woosongensis TKB2, a keratinolytic soil isolate. It was purified by HPLC, and its mass was found to be 4666.87 Da using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) spectroscopy. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of this peptide were 22.5 and 90 µg/ml, respectively. SEM study revealed the distorted cell wall of the test strain along with pore formation. The possible reason for bactericidal activity of the peptide is due to generation of reactive oxygen species (ROS), resulting in membrane damage and leakage of intracellular protein. Complete sequence of the peptide was predicted and retrieved from the sequence database of chicken feather keratin after in silico trypsin digestion using ExPASy tools. Further, net charge, hydrophobicity (77.7 %) and molecular modelling of the peptide were evaluated for better understanding of its mode of action. The hydrophobic region (17 to 27) of the peptide may facilitate for initial attachment on the bacterial membrane. The ABP exhibited no adverse effects on RBC membrane and HT-29 human cell line. This cytosafe peptide can be exploited as an effective therapeutic agent to combat Staphylococcal infections.

Presenting features of feather duvet lung.

BACKGROUND: Feather duvet lung (FDL) is a rare subgroup of bird fancier's lung. It is caused by inhalation of organic dust due to goose or duck feathers in duvets or pillows. METHODS: A retrospective review of the medical records of 13 patients with FDL was performed to assess the specific history and review clinical characteristics of patients with this disease. RESULTS: All patients were female with a mean age of 53 years (26-71). They were recently exposed to feather duvets (6), pillows (1) or both (6). Specific histories were duvets or pillows filled with raw goose feathers from their own farms (4), intensive contact with goose feathers in youth (3), and bird exposure prior to symptom onset (5). In all patients specific IgG antibodies to goose and/or duck feathers were detected. Pulmonary function tests revealed a moderate to severe reduced diffusion capacity and a mild restrictive pattern. High-resolution computed tomography was performed in 11 patients and demonstrated predominantly ground-glass opacities (10) and fibrosis (6). In bronchoalveolar lavage fluid, lymphocytic alveolitis was demonstrated in all patients. Lung biopsies were obtained in 9 patients and demonstrated lymphocytic alveolitis (8), granulomas (3), bronchiolitis obliterans organizing pneumonia pattern (2), and usual interstitial pneumonia pattern (1). CONCLUSIONS: The clinical findings of FDL are typical of extrinsic allergic alveolitis. Primary sensitization could be due to former exposure to bird antigens at home or goose/duck feather exposure in youth. In view of the increasing popularity of feather duvets, FDL should be considered in the differential diagnosis of patients with extrinsic allergic alveolitis.

Increased DNA damage and oxidative stress in chickens with natural Marek's disease.

Oxidative stress contributes to the accumulation of genomic abnormalities, prevents cellular apoptosis, and also mediates immunosuppression resulting in tumor formation. Marek's Disease provides excellent opportunities for the study of herpesvirus-induced tumors both in experimental- and natural conditions. The aim of this study was to examine the effects of Marek's Disease (MD) on basal levels of DNA strand breaks and on the oxidative-antioxidative status of chickens with MD. White-Lohmann hens-fifteen infected with Marek's Disease Virus (MDV) and fifteen healthy-of same age and sex were included in this study. MD infection was diagnosed via clinical signs, gross- and micro-pathological findings and also by detection of viral antigens in feather follicle epithelium by the indirect immunoperoxidase method. Compared with healthy controls, DNA damage was greater and levels of malondialdehyde (MDA) and plasma protein carbonyl (PCO), and plasma concentration of nitric oxide metabolites (NOx) higher in the MD group. Furthermore, total antioxidant activities (AOAs) were found lowered and glutathione (GSH) levels reduced in the MD group compared to the control group. Significantly positive correlation was found between DNA damage, MDA, PCO, and NOx in the MD group. DNA strand breaks were found negatively associated with AOA and GSH concentrations in the MD group. Our results demonstrated that oxidative stress markers and DNA damage substantially increased in chickens with MD, which indicated that increased DNA damage levels might be related to the increased oxidative stress and reduced antioxidant activity.

Marek's disease virus-induced expression of cytokine genes in feathers of genetically defined chickens.

Marek's disease (MD) vaccines, although effective in reducing lymphoproliferation, cannot control infectious virus production in the feather follicle epithelium (FFE) which is the site of virus shedding. Therefore, we investigated Marek's disease virus (MDV) replication as well as the expression of cytokine genes in feathers of MDV-infected chickens belonging to genetically defined lines (N2a or B(21)/B(21) haplotype-resistant and P2a or B(19)/B(19) haplotype-susceptible). Though there was not a difference in MDV genome load and transcripts between feathers of these chicken lines at 4 and 10 days post-infection (d.p.i.), feathers of resistant chickens carried significantly lower viral genome load and transcripts at 21 d.p.i. Irrespective of genetic background of the chickens examined, MDV replication showed a significant positive correlation with the expression of IFN-gamma gene. The results imply the usefulness of genetic control approach in reducing virulent MDV transmission.

Host responses are induced in feathers of chickens infected with Marek's disease virus.

Control measures are ineffective in curtailing Marek's disease virus (MDV) infection and replication in the feather follicle epithelium (FFE). Therefore, vaccinated birds which subsequently become infected with MDV, shed the virulent virus although they remain protected against disease. The present study investigated host responses generated against MDV infection in the feather. We observed that in parallel with an increase in viral genome load and viral replication in the feather, there was a gradual but progressive increase in infiltration of CD4+ and CD8+ T cells into the feather pulp of MDV-infected chickens, starting on day 4 and peaking by day 10 post-infection. Concomitant with infiltration of T cells, the expression of interleukin (IL)-18, IL-6, interferon (IFN)-gamma and major histocompatibility complex class I genes was significantly enhanced in the feather pulp of MDV-infected chickens. The finding that host responses are generated in the feather may be exploited for developing strategies to control MDV infection in the FFE, thus preventing horizontal virus transmission.

Are cutaneous hypersensitivity tests to inhalant allergens a severity marker for vernal keratoconjunctivitis?

PURPOSE: The purpose of this study was to analyze the cutaneous sensitivity to a variety of allergens in patients with vernal keratoconjunctivitis (VKC) and to demonstrate the relation between skin response and clinical aspects of the disease. METHODS: Twenty patients with vernal keratoconjunctivitis were randomly chosen from the External Disease and Cornea Sector. They were clinically evaluated, and a score ranging from 0 to 20 was applied based on signs and symptoms on ophthalmic examination. All subjects underwent a skin prick test against standardized allergens, such as house dust mites D. pteronyssinus, D. farinae, and Blomia tropicalis, as well as allergens from cat, dog, fungi and feather. RESULTS: Seventy-five per cent of patients were positive for at least one of the allergens tested. House dust mites were responsible for the majority of the cases (75 percent). There was a poor correlation between the clinical score and sensitivity to allergens (r= - 0.119 for fungi; r= - 0.174 for dog; r= - 0.243 for house dust mites; r= - 0.090 for feather). A significant correlation was found only for cat allergen extract (r = - 0.510; p=0.024). CONCLUSIONS: Our study demonstrated poor correlation between cutaneous hypersensitivity tests and clinical findings in patients with vernal keratoconjuntivitis. We concluded that skin response to inhalant allergens is not a useful test to identify clinical severity and chronicity of inflammatory process in this disease.

OBJETIVO: Avaliar o papel da sensibilização cutânea a diferentes aeroalérgenos em pacientes com ceratoconjuntivite vernal e a correlação entre esta e os aspectos clínicos da doença. MÉTODOS: Vinte pacientes do setor de doenças externas e córnea foram aleatoriamente convidados para participar deste estudo. Os pacientes foram avaliados e a eles foi atribuído um escore clínico variando de 0 a 20 de acordo com sinais e sintomas presentes no exame oftalmológico. Todos os pacientes foram submetidos a testes cutâneos de hipersensibilidade imediata contra aeroalérgenos padronizados como os ácaros domiciliares D. pteronyssinus, D. farinae e Blomia tropicalis, assim como também a alérgenos de epitélio de gato, epitélio de cão, mistura de fungos e mistura de penas. RESULTADOS: Setenta e cinco por cento dos pacientes tiveram teste de hipersensibilidade imediata positivo contra pelo menos um dos antígenos testados. Os ácaros domiciliares foram responsáveis pela maioria destes casos (75 por cento). Houve uma pobre correlação entre o escore clínico e a hipersensibilidade cutânea aos alérgenos (r= -0,119 para fungos; r= -0,174 para epitélio de cão; r= -0,243 para ácaros domiciliares; r= -0,090 para mistura de penas). Houve correlação significativa apenas contra epitélio de gatos (r= -0,510; p=0,024). CONCLUSÃO: O estudo demonstrou uma pobre correlação entre os testes cutâneos de hipersensibilidade imediata e os achados clínicos nos pacientes com ceratoconjuntivite vernal. Os testes cutâneos de hipersensibilidade imediata contra aeroalérgenos não foi parâmetro eficaz na identificação dos casos de maior gravidade e cronicidade de ceratoconjuntivite vernal.

A clinical study of hypersensitivity pneumonitis presumably caused by feather duvets.

BACKGROUND: Bird fancier's lung (BFL) is a type of hypersensitivity pneumonitis induced by the inhalation of bird-related antigens. The BFL induced by feathers is difficult to diagnose because feathers are generally unrecognized as a causative antigen. OBJECTIVE: To determine the clinical features of BFL presumably induced by feather duvets (feather duvet lung) to provide clues for diagnosis. METHODS: We performed a retrospective review of the medical records of patients with feather duvet lung evaluated between April 1, 2000, and June 30, 2003, at the Tokyo Medical and Dental University Hospital in Japan. RESULTS: Seven patients with feather duvet lung were included in this study; 4 patients had acute disease and 3 had chronic BFL. Duration of contact with feather duvets was 1 month to 10 years. Serum KL-6 and surfactant protein D levels were elevated in all the patients. Specific antibodies against avian antigens were positive in acute BFL but negative in chronic BFL. Antigen-induced lymphocyte proliferation in peripheral blood or bronchoalveolar lavage cells was positive in all the patients. The diagnosis was confirmed by an environmental or inhalation provocation test. CONCLUSIONS: Feather duvets can induce acute and chronic BFL. Physicians should be aware of feather duvets as a cause of BFL because feather duvets are becoming more prevalent.

[A case of acute bird fancier's lung caused by feather duvet].

A 57-year-old woman was admitted to our hospital because of cough and low-grade fever for 2 months and shortness of breath for 2 weeks. She had raised two budgerigars for the last 15 years and had been using a feather duvet for one year. A chest radiograph showed diffuse ground-glass opacities in both lung fields, and a chest CT scan showed centrilobular micronodules and ground-glass opacities. Bronchoalveolar lavage (BAL) revealed a marked increase in lymphocytes, and a transbronchial lung biopsy (TBLB) specimen showed alveolitis due to the infiltration of mononuclear cells. Since she had specific antibodies against pigeon and budgerigar dropping extracts and her peripheral blood lymphocytes proliferated on addition of pigeon serum, she was diagnosed as having bird fancier's lung (BFL). She was treated with steroids, which brought about a marked improvement. After she visited her husband who had been hospitalized where a feather duvet was provided for each patient, both subjective and objective findings deteriorated. This deterioration was preventable when she wore a protective mask for micro-dust while visiting her husband. The feather duvets seemed to induce acute BFL in this case, though raising budgerigars may well be related to her sensitization with bird-related antigens.

Feather duvet lung.

BACKGROUND: Bird fancier's lung (BFL) is a type of hypersensitivity pneumonitis, which is induced by inhalation of bird related antigens. The diagnosis of BFL induced by feathers is difficult because feathers are generally not recognized as a causative antigen of BFL. We report a female case of chronic BFL presumably due to a feather duvet, which presents as pulmonary fibrosis. CASE REPORT: A 73 year-old woman presented with exertional dyspnea for the last three years. She had raised two pigeons for three years (1971-1973) in her forties and had been using a feather duvet for the last eight years (1992-2000). A chest X-ray showed reticular infiltrates in the both peripheral lung field and an HRCT scan showed scattered consolidation, micronodules, and peribronchial ground-glass opacities. Lymphocyte proliferation to the feather antigen was positive and inhalation provocation test using a bird antigen was also positive. Thoracoscopic biopsy specimens showed organization, cholesterol clefts, alveolitis around terminal and respiratory bronchioles--all of which are consistent with chronic hypersensitivity pneumonitis. Clinical findings have spontaneously improved after she stopped using her feather duvet. CONCLUSIONS: Feather beds including duvets, pillows, and cushions are now popular all over the world. Physicians should be aware of feathers as a cause of BFL since this induction seems to be more prevalent.

Profiles of pulp infiltrating lymphocytes at various times throughout feather regeneration in Smyth line chickens with vitiligo.

Smyth line (SL) chickens develop a spontaneous, autoimmune, posthatch loss of pigment cells (vitiligo) in regenerating feather tissue. Smyth line vitiligo (SLV) is associated with lymphocyte infiltrations prior to and throughout the development of the disorder. It was the purpose of this study to determine the type, relative amounts, and proportions of pulp-infiltrating lymphocytes at various times throughout the growth of regenerating feathers. Feathers were plucked from 8-week-old chickens with and without SLV. Feather pulp cell suspensions were prepared when the regenerating feathers were 2, 3, 4, and 6 weeks of age. Cells were fluorescently labeled using a panel of mouse monoclonal antibodies specific for chicken lymphocytes. Both T and B cells infiltrated the feather pulp of chickens with SLV. T cell levels remained elevated throughout the 6 weeks of feather growth, while B cell levels steadily declined to control levels over the same time. The pulp-infiltrating cells were primarily T cells with an alphabeta T cell receptor expressing the Vbeta1 gene (TCR2+). The ratio between CD4+ and CD8+ cells was 1.42 and 0.75 in 2- and 6-week-old regenerating feathers from chickens with autoimmune SLV, respectively. In non-vitiliginous chickens this ratio was always near 1. These data suggest that TCR2+ T cells play an important role in SLV. CD4+ cells may play a recruiting/activating role, whereas CD8+ cells may have cytotoxic activity specifically directed against melanocytes. Additionally, this is the first report demonstrating the infiltration of B cells into the feather pulp of vitiliginous chickens. These B cells may directly/indirectly contribute to melanocyte destruction in SLV.

Synergism between the endogenous viral loci ev6 and ev9 in inducing immunological tolerance to avian leukosis virus.

1. The course of infection by exogenous avian leukosis virus was followed in a commercial strain of White Leghorn domestic fowls by measuring viral antigen in feather pulp and egg albumin. Ten days after hatching, 2 out of 360 birds tested positive and at 286 days of age about 60% of the birds had been antigen positive at least once. 2. Among the antigen positive birds, two groups could be distinguished: those which permanently and those which transiently expressed viral antigen. Permanent antigen expression was associated with low antibody titres, while transient antigen expression was associated with high antibody titres. 3. The strain segregated for the two endogenous viral genes ev6 and ev9, both of which express endogenous viral envelope protein, and have been implicated in affecting immune-responsiveness. The antibody titre in individuals positive for both ev6 and ev9, was significantly lower than in those which had none or only one of the two ev-genes. In addition, individuals positive for both ev-genes occurred more frequently in the group permanently positive for viral antigen than in the group transiently antigen positive. 4. The results indicate that there was a strong synergism between ev6 and ev9 in reducing the antibody response to exogenous avian leukosis virus infection, perhaps by inducing immune tolerance or interfering with antibody formation.

Semen traits and fertility of White Leghorn males shown to be positive or negative for lymphoid leukosis virus in semen and feather pulp.

1. Males from strains selected for high egg production (and other economic traits) and from unselected control strains were used to determine the frequency of shedding of lymphoid leukosis virus (LLV) into semen. The effect of the male's LLV status on semen production, fertility and hatchability was also examined in males of the unselected control strains. 2. The frequency of detection of exogenous LLV in semen by the phenotypic mixing test, and high concentrations of the viral group specific antigen in feather pulp by the complement fixation test, were both higher in control strains than in strains selected for high egg production. 3. Semen production was not reduced in LLV-shedding males. 4. Significant associations of LLV shedding with higher incidence of abnormal spermatozoa and reduced fertility were found in some populations but not in others. No significant effect of LLV shedding on hatchability was detected. 5. Tests for group specific antigen in feather pulp proved useful in identifying males that shed LLV in semen.

Effect of the expression of an endogenous viral gene on the growth of tumours induced by Rous sarcoma virus in chickens.

The endogenous group-specific antigen of avian retroviruses has been detected in feather pulp of chickens of the inbred CB line (WL breed). Expression of the gs antigen has been demonstrated by ELISA, whereas the classical methods have classified the CB line as gs-. The number of gs+ chickens decreased with increasing age after hatching, and all chickens were gs- already at 20 weeks of age. The longer duration of gs antigen expression in line CB chickens had an adverse effect on their ability to regress Rous sarcomas. Chickens from a closed flock of the BL breed that segregated to gs+ and gs- also significantly differed in the ability to regress Rous sarcomas.