My research career has focused on the causes of asthma and its treatment. After establishing the key role that mast cells play in the inflammatory response in asthma, attention was turned towards understanding disease chronicity and variability across the lifecourse. Through a combination of studies on airway biopsies and primary cell cultures we have established that asthma is primarily an epithelial disease driven by increased environmental susceptibility to injury and an altered repair response as depicted by sustained activation of the epithelial mesenchymal trophic unit (EMTU) that is invoked in foetal branching morphogenesis. Varied activation of the EMTU connects the origins of asthma to its progression over time with involvement of epithelial susceptibility through impaired barrier and innate immune functions and altered mesenchymal susceptibility as exemplified by polymorphisms of the metalloprotease gene, ADAM33. Taken together these observations have led to a fundamental re-evaluation of asthma pathogenesis. Rather than placing allergic inflammation as the prime abnormality, it is proposed that the airway epithelium lies at the center of asthma pathogenesis, and that in conjunction with the underlying mesenchyme, it is the principle orchestrator of both the induction of asthma and its evolution over the lifecourse. This concept has provided 'the basis for a new preventative and therapeutic approach focused more on increasing the airways resistance to environmental insults rather than suppressing downstream inflammation once it is established.
Asthma; management; prevention; treatment
Sensitization and exposure to the allergenic fungus Alternaria alternata has been associated with increased risk of asthma and asthma exacerbations. The first cells to encounter inhaled allergens are epithelial cells at the airway mucosal surface. Epithelial barrier function has previously been reported to be defective in asthma. This study investigated the contribution of proteases from Alternaria alternata on epithelial barrier function and inflammatory responses and compared responses of in vitro cultures of differentiated bronchial epithelial cells derived from severely asthmatic donors with those from non-asthmatic controls. Polarised 16HBE cells or air-liquid interface (ALI) bronchial epithelial cultures from non-asthmatic or severe asthmatic donors were challenged apically with extracts of Alternaria and changes in inflammatory cytokine release and transepithelial electrical resistance (TER) were measured. Protease activity in Alternaria extracts was characterised and the effect of selectively inhibiting protease activity on epithelial responses was examined using protease inhibitors and heat-treatment. In 16HBE cells, Alternaria extracts stimulated release of IL-8 and TNFα, with concomitant reduction in TER; these effects were prevented by heat-treatment of the extracts. Examination of the effects of protease inhibitors suggested that serine proteases were the predominant class of proteases mediating these effects. ALI cultures from asthmatic donors exhibited a reduced IL-8 response to Alternaria relative to those from healthy controls, while neither responded with increased thymic stromal lymphopoietin (TSLP) release. Only cultures from asthmatic donors were susceptible to the barrier-weakening effects of Alternaria. Therefore, the bronchial epithelium of severely asthmatic individuals may be more susceptible to the deleterious effects of Alternaria.
Biodiversity loss and climate change secondary to human activities are now being associated with various adverse health effects. However, less attention is being paid to the effects of biodiversity loss on environmental and commensal (indigenous) microbiotas. Metagenomic and other studies of healthy and diseased individuals reveal that reduced biodiversity and alterations in the composition of the gut and skin microbiota are associated with various inflammatory conditions, including asthma, allergic and inflammatory bowel diseases (IBD), type1 diabetes, and obesity. Altered indigenous microbiota and the general microbial deprivation characterizing the lifestyle of urban people in affluent countries appear to be risk factors for immune dysregulation and impaired tolerance. The risk is further enhanced by physical inactivity and a western diet poor in fresh fruit and vegetables, which may act in synergy with dysbiosis of the gut flora. Studies of immigrants moving from non-affluent to affluent regions indicate that tolerance mechanisms can rapidly become impaired in microbe-poor environments. The data on microbial deprivation and immune dysfunction as they relate to biodiversity loss are evaluated in this Statement of World Allergy Organization (WAO). We propose that biodiversity, the variability among living organisms from all sources are closely related, at both the macro- and micro-levels. Loss of the macrodiversity is associated with shrinking of the microdiversity, which is associated with alterations of the indigenous microbiota. Data on behavioural means to induce tolerance are outlined and a proposal made for a Global Allergy Plan to prevent and reduce the global allergy burden for affected individuals and the societies in which they live.
Allergy plan; Biodiversity; Civilization disease; Epigenetics; Immune dysfunction; Microbiota; Microbiome; Urbanization
The asthma susceptibility gene, a disintegrin and metalloprotease-33 (ADAM33), is selectively expressed in mesenchymal cells, and the activity of soluble ADAM33 has been linked to angiogenesis and airway remodeling. Transforming growth factor (TGF)-β is a profibrogenic growth factor, the expression of which is increased in asthma, and recent studies show that it enhances shedding of soluble ADAM33. In this study, we hypothesized that TGF-β also affects ADAM33 expression in bronchial fibroblasts in asthma. Primary fibroblasts were grown from bronchial biopsies from donors with and those without asthma, and treated with TGF-β2 to induce myofibroblast differentiation. ADAM33 expression was assessed using quantitative RT-PCR and Western blotting. To examine the mechanisms whereby TGF-β2 affected ADAM33 expression, quantitative methylation-sensitive PCR, chromatin immunoprecipitation, and nuclear accessibility assays were conducted on the ADAM33 promoter. We found that TGF-β2 caused a time- and concentration-dependent reduction in ADAM33 mRNA expression in normal and asthmatic fibroblasts, affecting levels of splice variants similarly. TGF-β2 also induced ADAM33 protein turnover and appearance of a cell-associated C-terminal fragment. TGF-β2 down-regulated ADAM33 mRNA expression by causing chromatin condensation around the ADAM33 promoter with deacetylation of histone H3, demethylation of H3 on lysine-4, and hypermethylation of H3 on lysine-9. However, the methylation status of the ADAM33 promoter did not change. Together, these data suggest that TGF-β2 suppresses expression of ADAM33 mRNA in normal or asthmatic fibroblasts. This occurs by altering chromatin structure, rather than by gene silencing through DNA methylation as in epithelial cells. This may provide a mechanism for fine regulation of levels of ADAM33 expression in fibroblasts, and may self-limit TGF-β2–induced ectodomain shedding of ADAM33.
a disintegrin and metalloprotease-33; myofibroblast; transforming growth factor-β; histone modification
Asthma is a common disease of children with a complex genetic origin. Understanding the genetic basis of asthma susceptibility will allow disease prediction and risk stratification.
We sought to identify asthma susceptibility genes in children.
A nested case-control genetic association study of children of Caucasian European ancestry from a birth cohort was conducted. Single nucleotide polymorphisms (SNPs, n=116,024) were genotyped in pools of DNA samples from cohort children with physician-diagnosed asthma (n=112) and normal controls (n=165). A genomic region containing the ATPAF1 gene was significantly associated with asthma. Additional SNPs within this region were genotyped in individual samples from the same children and in eight independent study populations consisting of Caucasian, African American, Hispanic, or other ancestries. SNPs were also genotyped or imputed in two consortia control populations. ATPAF1 expression was measured in bronchial biopsies from asthmatics and controls.
Asthma was associated with a cluster of SNPs and SNP haplotypes containing the ATPAF1 gene with two SNPs achieving significance at a genome-wide level (p=2.26×10−5 to 2.2×10−8). Asthma severity was also associated with SNPs and haplotypes in the primary population. SNP and/or gene-level associations were confirmed in the four non-Hispanic populations. Haplotype associations were confirmed in the non-Hispanic populations (p=0.045 to 0.0009). ATPAF1 total RNA expression was significantly (p<0.01) higher in bronchial biopsies from asthmatics than controls.
Genetic variation in the ATPAF1 gene predisposes children of different ancestry to asthma.
asthma; ATPAF1; children; gene; genetic; genome-wide association; purinergic; respiratory; single nucleotide polymorphism; SNP
Rhinovirus (RV) infection is a major cause of asthma exacerbations which may be due to a deficient innate immune response in the bronchial epithelium. We hypothesized that the pleiotropic cytokine, TGF-β, influences interferon (IFN) production by primary bronchial epithelial cells (PBECs) following RV infection. Exogenous TGF-β2 increased RV replication and decreased IFN protein secretion in response to RV or double-stranded RNA (dsRNA). Conversely, neutralizing TGF-β antibodies decreased RV replication and increased IFN expression in response to RV or dsRNA. Endogenous TGF-β2 levels were higher in conditioned media of PBECs from asthmatic donors and the suppressive effect of anti-TGF-β on RV replication was significantly greater in these cells. Basal SMAD-2 activation was reduced when asthmatic PBECs were treated with anti-TGF-β and this was accompanied by suppression of SOCS-1 and SOCS-3 expression. Our results suggest that endogenous TGF-β contributes to a suppressed IFN response to RV infection possibly via SOCS-1 and SOCS-3.
Rationale: Airway mucous cell metaplasia and chronic inflammation are pathophysiological features that influence morbidity and mortality associated with asthma and other chronic pulmonary disorders. Elucidation of the molecular mechanisms regulating mucous metaplasia and hypersecretion provides the scientific basis for diagnostic and therapeutic opportunities to improve the care of chronic pulmonary diseases.
Objectives: To determine the role of the airway epithelial–specific transcription factor NK2 homeobox 1 (NKX2-1, also known as thyroid transcription factor-1 [TTF-1]) in mucous cell metaplasia and lung inflammation.
Methods: Expression of NKX2-1 in airway epithelial cells from patients with asthma was analyzed. NKX2-1+/− gene targeted or transgenic mice expressing NKX2-1 in conducting airway epithelial cells were sensitized to the aeroallergen ovalbumin. In vitro studies were used to identify mechanisms by which NKX2-1 regulates mucous cell metaplasia and inflammation.
Measurements and Main Results: NKX2-1 was suppressed in airway epithelial cells from patients with asthma. Reduced expression of NKX2-1 in heterozygous NKX2-1+/− gene targeted mice increased mucous metaplasia in the small airways after pulmonary sensitization to ovalbumin. Conversely, mucous cell metaplasia induced by aeroallergen was inhibited by expression of NKX2-1 in the respiratory epithelium in vivo. Genome-wide mRNA analysis of lung tissue from ovalbumin-treated mice demonstrated that NKX2-1 inhibited mRNAs associated with mucous metaplasia and Th2-regulated inflammation, including Spdef, Ccl17, and Il13. In vitro, NKX2-1 inhibited SPDEF, a critical regulator of airway mucous cell metaplasia, and the Th2 chemokine CCL26.
Conclusions: The present data demonstrate a novel function for NKX2-1 in a gene network regulating mucous cell metaplasia and allergic inflammation in the respiratory epithelium.
asthma; goblet cell; respiratory epithelium; NK2 homeobox 1
Allergist/clinical immunologist maintenance of certification and training program reaccreditation are mandatory in some countries. The World Allergy Organization conducted surveys in 2009 and 2011 to assess where such programs were available and to promote the establishment of such programs on a global level. This was done with the presumption that after such an “inventory,” World Allergy Organization could offer guidance to its Member Societies on the promotion of such programs to assure the highest standards of practice in the field of allergy and clinical immunology. This review draws on the experience of countries where successful programs are in place and makes recommendations for those wishing to implement such programs for the specialty.
allergy and clinical immunology; allergist; physician skills; physician competencies; physician performance measurement
Asthma exacerbations remain a major unmet clinical need. The difficulty in obtaining airway tissue and bronchoalveolar lavage samples during exacerbations has greatly hampered study of naturally occurring exacerbations. This study was conducted to determine if mRNA profiling of peripheral blood mononuclear cells (PBMCs) could provide information on the systemic molecular pathways involved during asthma exacerbations.
Over the course of one year, gene expression levels during stable asthma, exacerbation, and two weeks after an exacerbation were compared using oligonucleotide arrays. For each of 118 subjects who experienced at least one asthma exacerbation, the gene expression patterns in a sample of peripheral blood mononuclear cells collected during an exacerbation episode were compared to patterns observed in multiple samples from the same subject collected during quiescent asthma. Analysis of covariance identified genes whose levels of expression changed during exacerbations and returned to quiescent levels by two weeks. Heterogeneity among visits in expression profiles was examined using K-means clustering. Three distinct exacerbation-associated gene expression signatures were identified. One signature indicated that, even among patients without symptoms of respiratory infection, genes of innate immunity were activated. Antigen-independent T cell activation mediated by IL15 was also indicated by this signature. A second signature revealed strong evidence of lymphocyte activation through antigen receptors and subsequent downstream events of adaptive immunity. The number of genes identified in the third signature was too few to draw conclusions on the mechanisms driving those exacerbations.
This study has shown that analysis of PBMCs reveals systemic changes accompanying asthma exacerbation and has laid the foundation for future comparative studies using PBMCs.
Rationale: Asthma is a chronic inflammatory airway disease that affects more than 300 million individuals worldwide. Asthma is caused by interaction of genetic and environmental factors. Bronchial hyperresponsiveness (BHR) is a hallmark of asthma and results from increased sensitivity of the airways to physical or chemical stimulants. BHR and asthma are linked to chromosome 5q31-q33.
Objectives: To identify a gene for BHR on chromosome 5q31-q33.
Methods: In 200 Dutch families with asthma, linkage analysis and fine mapping were performed, and the Protocadherin 1 gene (PCDH1) was identified. PCDH1 was resequenced in 96 subjects from ethnically diverse populations to identify novel sequence variants. Subsequent replication studies were undertaken in seven populations from The Netherlands, the United Kingdom, and the United States, including two general population samples, two family samples, and three case-control samples. PCDH1 mRNA and protein expression was investigated using polymerase chain reaction, Western blotting, and immunohistochemistry.
Measurements and Main Results: In seven out of eight populations (n = 6,168) from The Netherlands, United Kingdom, and United States, PCHD1 gene variants were significantly associated with BHR (P values, 0.005–0.05) This association was present in both families with asthma and general populations. PCDH1 mRNA and protein were expressed in airway epithelial cells and in macrophages.
Conclusions: PCDH1 is a novel gene for BHR in adults and children. The identification of PCDH1 as a BHR susceptibility gene may suggest that a structural defect in the integrity of the airway epithelium, the first line of defense against inhaled substances, contributes to the development of BHR.
bronchial hyperresponsiveness; asthma genetics; protocadherin-1; cell adhesion; airway epithelium
The original concept of asthma being primarily a disease of airways smooth muscle drove the development of bronchodilator drugs. However when it was realised that airway inflammation underpinned the disordered airway function, this gave way to the development of controller therapies such as inhaled cromones and corticosteroids. More recently the discovery of complex interconnecting cytokine and chemokine networks has stimulated the development of biologics with varying success. With the recognition that airway wall "remodelling" is present early in asthma inception and is in part driven by aberrant epithelial-mesenchymal communication both genetic and environmental factors beyond allergen exposure such as virus infection and air pollution are being seen as being increasingly important not only in asthma exacerbations but in the origins of asthma and its evolution into different sub-phenotypes. This brings us round full circle to once again considering that the origins of asthma lie in defects in the formed elements of the airway; the epithelium, smooth muscle, and vasculature. Over the last 25 years Professor You Young Kim has engaged in the exciting discovery science of allergy and asthma and has made an enormous contribution in bringing Korea to the forefront of disease management and research, a position that both he and his colleagues can justly be proud of.
Asthma; airway inflammation; airway remodeling; infection; epithelial-mesenchymal trophic unit; ADAM33
Respiratory diseases are placing an increasing burden on the UK health system
Asthma is an inflammatory disorder of the conducting airways that has strong association with allergic sensitization. The disease is characterized by a polarized Th-2 (T-helper-2)-type T-cell response, but in general targeting this component of the disease with selective therapies has been disappointing and most therapy still relies on bronchodilators and corticosteroids rather than treating underlying disease mechanisms. With the disappointing outcomes of targeting individual Th-2 cytokines or manipulating T-cells, the time has come to re-evaluate the direction of research in this disease. A case is made that asthma has its origins in the airways themselves involving defective structural and functional behaviour of the epithelium in relation to environmental insults. Specifically, a defect in barrier function and an impaired innate immune response to viral infection may provide the substrate upon which allergic sensitization takes place. Once sensitized, the repeated allergen exposure will lead to disease persistence. These mechanisms could also be used to explain airway wall remodelling and the susceptibility of the asthmatic lung to exacerbations provoked by respiratory viruses, air pollution episodes and exposure to biologically active allergens. Variable activation of this epithelial–mesenchymal trophic unit could also lead to the emergence of different asthma phenotypes and a more targeted approach to the treatment of these. It also raises the possibility of developing treatments that increase the lung's resistance to the inhaled environment rather than concentrating all efforts on trying to suppress inflammation once it has become established.
allergen; asthma; inflammation; remodelling; T-cell; virus infection; BHR, bronchial hyper-responsiveness; CT, computed tomography; DC, dendritic cell; ADC, airway DC; EBUS, endobronchial ultrasound; EMTU, epithelial–mesenchymal trophic unit; ETS, environmental tobacco smoke; IFN, interferon; IL, interleukin; IoW, Isle of Wight; LT, leukotriene; mAb, monoclonal antibody; RV, rhinovirus; TGF-β, transforming growth factor-β; Th-2, T-helper-2; TJ, tight junction; TSLP, thymic stromal lymphopoietin
Exposure to fine ambient particulate matter (PM) has consistently been associated with increased morbidity and mortality. The relationship between exposure to ultrafine particles (UFP) and health effects is less firmly established. If UFP cause health effects independently from coarser fractions, this could affect health impact assessment of air pollution, which would possibly lead to alternative policy options to be considered to reduce the disease burden of PM. Therefore, we organized an expert elicitation workshop to assess the evidence for a causal relationship between exposure to UFP and health endpoints.
An expert elicitation on the health effects of ambient ultrafine particle exposure was carried out, focusing on: 1) the likelihood of causal relationships with key health endpoints, and 2) the likelihood of potential causal pathways for cardiac events. Based on a systematic peer-nomination procedure, fourteen European experts (epidemiologists, toxicologists and clinicians) were selected, of whom twelve attended. They were provided with a briefing book containing key literature. After a group discussion, individual expert judgments in the form of ratings of the likelihood of causal relationships and pathways were obtained using a confidence scheme adapted from the one used by the Intergovernmental Panel on Climate Change.
The likelihood of an independent causal relationship between increased short-term UFP exposure and increased all-cause mortality, hospital admissions for cardiovascular and respiratory diseases, aggravation of asthma symptoms and lung function decrements was rated medium to high by most experts. The likelihood for long-term UFP exposure to be causally related to all cause mortality, cardiovascular and respiratory morbidity and lung cancer was rated slightly lower, mostly medium. The experts rated the likelihood of each of the six identified possible causal pathways separately. Out of these six, the highest likelihood was rated for the pathway involving respiratory inflammation and subsequent thrombotic effects.
The overall medium to high likelihood rating of causality of health effects of UFP exposure and the high likelihood rating of at least one of the proposed causal mechanisms explaining associations between UFP and cardiac events, stresses the importance of considering UFP in future health impact assessments of (transport-related) air pollution, and the need for further research on UFP exposure and health effects.
The aim of this study is to compare asthma controller therapy in children between Europe and Japan.
Materials and Methods
A questionnaire-based survey was conducted at the 2007 annual meeting of the European Respiratory Society held in Stockholm. In total, 120 answers were collected from European doctors. We divided Europe into 5 areas: South, West, North, East, and Central. The same survey was conducted at the 40th annual meeting of the Japanese Society of Pediatric Pulmonology. Forty-three answers were collected from Japanese doctors.
Inhaled corticosteroids were used more frequently in Europe and antileukotrienes were used more frequently in Japan. There were also some differences in treatment in different areas of Europe.
This survey shows differences in the treatment of children with asthma in Europe and in Japan. European doctors prefer using inhaled corticosteroids, and Japanese doctors prefer using oral antileukotrienes. Because the number of the respondents is small and there may be some bias, further study on a large-scale for general clinicians providing medical care to asthma children is desirable.
asthma treatment; children; Europe; Japan; questionnaire-based survey
Exacerbations of Chronic obstructive pulmonary disease (COPD) are an important cause of the morbidity and mortality associated with the disease. Strategies to reduce exacerbation frequency are thus urgently required and depend on an understanding of the inflammatory milieu associated with exacerbation episodes. Bacterial colonisation has been shown to be related to the degree of airflow obstruction and increased exacerbation frequency. The aim of this study was to asses the kinetics of cytokine release from COPD parenchymal explants using an ex vivo model of lipopolysaccharide (LPS) induced acute inflammation.
Lung tissue from 24 patients classified by the GOLD guidelines (7F/17M, age 67.9 ± 2.0 yrs, FEV1 76.3 ± 3.5% of predicted) and 13 subjects with normal lung function (8F,5M, age 55.6 ± 4.1 yrs, FEV1 98.8 ± 4.1% of predicted) was stimulated with 100 ng/ml LPS alone or in combination with either neutralising TNFα or IL-10 antibodies and supernatant collected at 1,2,4,6,24, and 48 hr time points and analysed for IL-1β, IL-5, IL-6, CXCL8, IL-10 and TNFα using ELISA. Following culture, explants were embedded in glycol methacrylate and immunohistochemical staining was conducted to determine the cellular source of TNFα, and numbers of macrophages, neutrophils and mast cells.
In our study TNFα was the initial and predictive cytokine released followed by IL-6, CXCL8 and IL-10 in the cytokine cascade following LPS exposure. The cytokine cascade was inhibited by the neutralisation of the TNFα released in response to LPS and augmented by the neutralisation of the anti-inflammatory cytokine IL-10. Immunohistochemical analysis indicated that TNFα was predominantly expressed in macrophages and mast cells. When patients were stratified by GOLD status, GOLD I (n = 11) and II (n = 13) individuals had an exaggerated TNFα responses but lacked a robust IL-10 response compared to patients with normal lung function (n = 13).
We report on a reliable ex vitro model for the investigation of acute lung inflammation and its resolution using lung parenchymal explants from COPD patients. We propose that differences in the production of both TNFα and IL-10 in COPD lung tissue following exposure to bacterial LPS may have important biological implications for both episodes of exacerbation, disease progression and amelioration.
Epidemiological studies have demonstrated adverse health effects of environmental pollution. Diesel exhaust (DE) is a major contributor to particulate matter pollution. DE exposure has been shown to induce a pronounced inflammatory response in the airways, together with an enhanced epithelial expression of cytokines such as IL-8, Gro-α, IL-13 and activation of redox sensitive transcription factors (NFκB, AP-1), and MAP kinases (p38, JNK). The aim of the present investigation was to elucidate the involvement of the epidermal growth factor receptor (EGFR) signalling pathway in the epithelial response to DE in-vivo.
Immunohistochemical staining was used to quantify the expression of the EGFR, phosphorylated Tyrosine residues, MEK and ERK in the bronchial epithelium of archived biopsies from 15 healthy subjects following exposure to DE (PM10, 300 μg/m3) and air. DE induced a significant increases in the expression of EGFR (p = 0.004) and phosphorylated C-terminal Tyr 1173 (p = 0.02). Other investigated EGFR tyrosine residues, Src related tyrosine (Tyr 416), MEK and ERK pathway were not changed significantly by DE.
Exposure to DE (PM10, 300 μg/m3) caused enhanced EGFR expression and phosphorylation of the tyrosine residue (Tyr 1173) which is in accordance with the previously demonstrated activation of the JNK, AP-1, p38 MAPK and NFkB pathways and associated downstream signalling and cytokine production. No effects were seen on the MEK and ERK pathway suggesting that at the investigated time point (6 hours post exposure) there was no proliferative/differentiation signalling in the bronchial epithelium. The present findings suggest a key role for EGFR in the bronchial response to diesel exhaust.
An understanding of the needs and behaviors of asthma patients is important in developing an asthma-related healthcare policy. The primary goal of the present review was to assess patient perspectives on key issues in asthma and its management, as captured in patient surveys.
Local, national, and multinational asthma surveys were reviewed to assess patient perspectives, and where possible healthcare provider (HCP) perspectives, on key issues, including diagnosis, treatment, control, quality of life, and other patient-centered outcomes. Twenty-four surveys, conducted or published between 1997 and 2003 in Europe and North America, were included in this review. Substantial differences among studies prevented a formal meta-analysis; instead, data were pooled to allow for general comparisons and qualitative analysis.
The results indicate that patients' knowledge of the underlying causes of asthma and treatment options remains inadequate. Moreover, patients often tolerate poor symptom control, possess meager knowledge of correct drug usage, and display insufficient adherence to therapy. Many patients have a low expectation of receiving an appropriate therapy or of having a positive encounter with the HCP. Among HCPs, there is evidence of inadequate understanding of disease etiology and poor or unstructured communication with patients, resulting often in inaccurate assessment of disease severity. Moreover, patients often underreport their symptoms and severity, which in turn could lead to misclassification and undertreatment.
Improving patient education about the importance of achieving optimal asthma control, along with improved communication between patients and HCPs, emphasizing treatment options and optimal treatment of inflammation, may lead to better outcomes and improved asthma management in daily practice.
Fibroblasts are implicated in sub-epithelial fibrosis in remodeled asthmatic airways and contribute to airway inflammation by releasing cytokines and other mediators. Fibroblast activity is influenced by members of the leukotriene family of bronchoconstrictor and inflammatory mediators, but it is not known whether human bronchial fibroblasts can synthesize leukotrienes.
The expression of leukotriene biosynthetic enzymes and receptors was investigated in primary fibroblasts from the bronchi of normal and asthmatic adult subjects using RT-PCR, Western blotting, immunocytochemistry and flow cytometry.
These techniques revealed that human bronchial fibroblasts from both subject groups constitutively express 5-lipoxygenase, its activating protein FLAP, the terminal enzymes leukotriene A4 hydrolase and leukotriene C4 synthase, and receptors for leukotriene B4 (BLT1) and cysteinyl-leukotrienes (CysLT1). Human bronchial fibroblasts generated immunoreactive leukotriene B4 and cysteinyl-leukotrienes spontaneously and in increased amounts after calcium-dependent activation. Flow cytometry showed that human bronchial fibroblasts transformed to a myofibroblast-like phenotype by culture with transforming growth factor-β1 expressed 320–400% more immunofluorescence for leukotriene C4 synthase and CysLT1 receptors, with 60–80% reductions in leukotriene A4 hydrolase and BLT1 receptors.
These results indicate that human bronchial fibroblasts may not only respond to exogenous leukotrienes but also generate leukotrienes implicated in narrowing, inflammation and remodeling of the asthmatic airway.