Blood transcriptomic signature in type-2 biomarker-low severe asthma and asthma control.

BACKGROUND: Patients with type-2 (T2) cytokine-low severe asthma often have persistent symptoms despite suppression of T2 inflammation with corticosteroids. OBJECTIVES: We sought to analyze whole blood transcriptome from 738 samples in T2-biomarker-high/-low patients with severe asthma to relate transcriptomic signatures to T2 biomarkers and asthma symptom scores. METHODS: Bulk RNA-seq data were generated for blood samples (baseline, week 24, week 48) from 301 participants recruited to a randomized clinical trial of corticosteroid optimization in severe asthma. Unsupervised clustering, differential gene expression analysis, and pathway analysis were performed. Patients were grouped by T2-biomarker status and symptoms. Associations between clinical characteristics and differentially expressed genes (DEGs) associated with biomarker and symptom levels were investigated. RESULTS: Unsupervised clustering identified 2 clusters; cluster 2 patients were blood eosinophil-low/symptom-high and more likely to be receiving oral corticosteroids (OCSs). Differential gene expression analysis of these clusters, with and without stratification for OCSs, identified 2960 and 4162 DEGs, respectively. Six hundred twenty-seven of 2960 genes remained after adjusting for OCSs by subtracting OCS signature genes. Pathway analysis identified dolichyl-diphosphooligosaccharide biosynthesis and assembly of RNA polymerase I complex as significantly enriched pathways. No stable DEGs were associated with high symptoms in T2-biomarker-low patients, but numerous associated with elevated T2 biomarkers, including 15 that were upregulated at all time points irrespective of symptom level. CONCLUSIONS: OCSs have a considerable effect on whole blood transcriptome. Differential gene expression analysis demonstrates a clear T2-biomarker transcriptomic signature, but no signature was found in association with T2-biomarker-low patients, including those with a high symptom burden.

Inhaled nebulised unfractionated heparin (UFH) for the treatment of hospitalised patients with COVID-19: A randomised controlled pilot study.

There is a strong scientific rationale to use nebulised unfractionated heparin (UFH) in treating patients with COVID-19. This pilot study investigated whether nebulised UFH was safe and had any impact on mortality, length of hospitalisation and clinical progression, in the treatment of hospitalised patients with COVID-19. This parallel group, open label, randomised trial included adult patients with confirmed SARS-CoV-2 infection admitted to two hospitals in Brazil. One hundred patients were planned to be randomised to either "standard of care" (SOC) or SOC plus nebulized UFH. The trial was stopped after randomisation of 75 patients due to falling COVID-19 hospitalisation rates. Significance tests were 1-sided test (10% significance level). The key analysis populations were intention to treat (ITT) and modified ITT (mITT) which excluded (from both arms) subjects admitted to ITU or who died within 24 h of randomisation. In the ITT population (n = 75), mortality was numerically lower for nebulised UFH (6 out of 38 patients; 15.8%) versus SOC (10 out of 37 patients; 27.0%), but not statistically significant; odds ratio (OR) 0.51, p = 0.24. However, in the mITT population, nebulised UFH reduced mortality (OR 0.2, p = 0.035). Length of hospital stay was similar between groups, but at day 29, there was a greater improvement in ordinal score following treatment with UFH in the ITT and mITT populations (p = 0.076 and p = 0.012 respectively), while mechanical ventilation rates were lower with UFH in the mITT population (OR 0.31; p = 0.08). Nebulised UFH did not cause any significant adverse events. In conclusion, nebulised UFH added to SOC in hospitalised patients with COVID-19 was well tolerated and showed clinical benefit, particularly in patients who received at least 6 doses of heparin. This trial was funded by The J.R. Moulton Charity Trust and registered under REBEC RBR-8r9hy8f (UTN code: U1111-1263-3136).

Exacerbation Profile and Risk Factors in a Type-2-Low Enriched Severe Asthma Cohort: A Clinical Trial to Assess Asthma Exacerbation Phenotypes.

Rationale: The past 25 years have seen huge progress in understanding of the pathobiology of type-2 (T2) asthma, identification of measurable biomarkers, and the emergence of novel monoclonal antibody treatments. Although present in a minority of patients with severe asthma, very little is known about the mechanisms underlying T2-low asthma, making it a significant unmet need in asthma research. Objectives: The objective of this study was to explore the differences between study exacerbators and nonexacerbators, to describe physiological changes at exacerbation in those who are T2HIGH and T2LOW at the time of exacerbation, and to evaluate the stability of inflammatory phenotypes when stable and at exacerbation. Methods: Exacerbation assessment was a prespecified secondary analysis of data from a 48-week, multicenter, randomized controlled clinical study comparing the use of biomarkers and symptoms to adjust steroid treatment in a T2-low severe asthma-enriched cohort. Participants were phenotyped as T2LOW (fractional exhaled nitric oxide ⩽ 20 ppb and blood eosinophil count ⩽ 150 cells/µl) or T2HIGH (fractional exhaled nitric oxide > 20 or blood eosinophil count > 150) at study enrollment and at each exacerbation. Here, we report the findings of the exacerbation analyses, including comparison of exacerbators and nonexacerbators, the physiological changes at exacerbation in those who had evidence of T2 biology at exacerbation versus those that did not, and the stability of inflammatory phenotypes when stable and at exacerbation. Measurements and Main Results: Of the 301 participants, 60.8% (183) had one or more self-reported exacerbations (total of 390). Exacerbators were more likely to be female, have a higher body mass index, and have more exacerbations requiring oral corticosteroid and unscheduled primary care attendances for exacerbations. At enrollment, 23.6% (71) were T2LOW and 76.4% (230) T2HIGH. The T2LOW group had more asthma primary care attendances, were more likely to have a previous admission to HDU (high dependency unit)/ICU and to be receiving maintenance oral corticosteroids. At exacerbation, the T2LOW events were indistinguishable from T2HIGH exacerbations in terms of lung function (mean fall in T2LOW FEV1, 200 [400] ml vs. T2HIGH 200 [300] ml; P = 0.93) and symptom increase (ACQ5: T2LOW, 1.4 [0.8] vs. T2HIGH, 1.3 [0.8]; P = 0.72), with no increase in T2 biomarkers from stable to exacerbation state in the T2LOW exacerbations. The inflammatory phenotype within individual patients was dynamic; inflammatory phenotype at study entry did not have a significant association with exacerbation phenotype. Conclusions: Asthma exacerbations demonstrating a T2LOW phenotype were physiologically and symptomatically similar to T2HIGH exacerbations. T2LOW asthma was an unstable phenotype, suggesting that exacerbation phenotyping should occur at the time of exacerbation. The clinically significant exacerbations in participants without evidence of T2 biology at the time of exacerbation highlight the unmet and pressing need to further understand the mechanisms at play in non-T2 asthma. Clinical trial registered with www.clinicaltrials.gov (NCT02717689).

Effects of switching from a metered dose inhaler to a dry powder inhaler on climate emissions and asthma control: post-hoc analysis.

OBJECTIVE: To compare the effects of switching from a pressurised metered dose inhaler (pMDI)-based to a dry powder inhaler (DPI)-based maintenance therapy versus continued usual care on greenhouse gas emissions (carbon dioxide equivalents, CO2e) and asthma control. METHODS: This post-hoc analysis was based on a subset of 2236 (53%) patients from the Salford Lung Study in Asthma who at baseline were using a pMDI-based controller therapy. During the study patients were randomised to fluticasone furoate/vilanterol (FF/VI) via the ELLIPTA DPI (switched from pMDI to DPI) (n=1081) or continued their usual care treatment (n=1155), and were managed in conditions close to everyday clinical practice. Annual CO2e (kg) was calculated for the total number of maintenance and rescue inhalers prescribed. Asthma control was assessed by the proportion of ACT responders (composite of ACT total score ≥20 and/or increase from baseline ≥3). RESULTS: The groups were well matched for demographic characteristics and baseline Asthma Control Test (ACT) total score (mean age: 49 years; mean ACT score: usual care, 16.6; FF/VI, 16.5). Annual CO2e kg per patient (maintenance plus rescue therapy) was significantly lower with FF/VI DPI treatment ('switch' group) than usual care (least squares geometric mean 108 kg (95% CI 102 to 114) vs 240 kg (95% CI 229 to 252), p<0.001). Asthma control was consistently superior over the 12 months in the FF/VI DPI group compared with usual care. CONCLUSIONS: Patients switching from a pMDI-based to a DPI-based maintenance therapy more than halved their inhaler carbon footprint without loss of asthma control. The remaining inhaler carbon footprint could be reduced through switches from pMDI to DPI rescue medications or alternative lower-carbon footprint rescue inhalers if available. Asthma control improved in both groups, with greater control demonstrated in those initiated on FF/VI DPI. TRIAL REGISTRATION NUMBER: NCT01706198.

The environmental impact of inhaled therapy: making informed treatment choices.

When selecting the best inhaler and drug combination for a patient with respiratory disease, a number of factors should be considered. While efficacy and safety of medical treatments are always a priority, in recent years the environmental impacts of all aspects of life have become an increasingly necessary consideration and inhaled therapies are no exception. The carbon footprint of an item, individual or organisation is one of the most important and quantifiable environmental impacts, assessed by the amount of greenhouse gases (often expressed in terms of carbon dioxide equivalents) generated throughout the life cycle. The two most commonly prescribed and manufactured inhaler types worldwide are pressurised metered-dose inhalers (pMDIs) containing hydrofluorocarbon (HFC) propellants and dry powder inhalers (DPIs). Most of the carbon footprint of current pMDIs is a result of the propellants that they contain (HFC-134a and HFC-227ea, which are potent greenhouse gases). In comparison, the powder in DPIs is dispersed by the patient's own inhalation, meaning DPIs do not contain a propellant and have a lower carbon footprint than most pMDIs currently available. Soft mist inhalers are another propellant-free option: the device contains a spring, which provides the energy to disperse the aqueous medication. In this review, we examine the published data on carbon footprint data for inhalers, providing an analysis of potential implications for treatment decision making and industry initiatives.

Factors affecting adherence with treatment advice in a clinical trial of patients with severe asthma.

BACKGROUND: Understanding why patients with severe asthma do not follow healthcare provider (HCP) advice to adjust treatment is critical to achieving personalised disease management. METHODS: We reviewed patient choice to follow HCP advice to adjust asthma treatment in a UK-based randomised, controlled, single-blind (study participant), multicentre, parallel group 48-week clinical study comparing biomarker-directed treatment adjustment with standard care in severe asthma. RESULTS: Of 1572 treatment advisories (291 participants), instructions were followed in 1377 cases (87.6%). Patients were more likely to follow advice to remain on treatment (96.7%) than to either reduce (70.3%) or increase (67.1%) their treatment, with 64% of patients following all treatment advice. Multivariate analysis associated belonging to an ethnic minority group (OR 3.10, 95% CI 1.68-5.73) and prior study medication changes (two or more changes: OR 2.77, 95% CI 1.51-5.10) with failure to follow treatment advice. In contrast, emergency room attendance in the prior year (OR 0.54, 95% CI 0.32-0.92) was associated with following treatment advice. The largest effect was seen with transition onto or off oral corticosteroids (OR 29.28, 95% CI 16.07-53.36) when compared with those requested to maintain treatment. Centre was also an important determinant regarding the likelihood of patients to follow treatment advice. CONCLUSIONS: Belonging to an ethnic minority group and multiple prior treatment adjustments were associated with not following HCP treatment advice. Patients also responded differently to HCP advice across UK specialist centres. These findings have implications for the generalisability of models of care in severe asthma and require further focused studies.

The environmental impact of inhalers for asthma: A green challenge and a golden opportunity.

The propellants in metered-dose inhalers (MDIs) are powerful greenhouse gases, which account for approximately 13% of the NHS's carbon footprint related to the delivery of care. Most MDI use is in salbutamol relievers in patients with poorly controlled disease. The UK lags behind Europe in this regard, with greater reliance on salbutamol MDI and correspondingly greater greenhouse gas emissions, roughly treble that of our European neighbours. There has been a broad switch towards MDIs in asthma treatment in the UK over the last 20 years to reduce financial costs, such that the treatment for two-thirds of asthma patients in the UK is dominated by salbutamol MDI. Strategies that replace overuse of reliever MDIs with regimes emphasising inhaled corticosteroids have the potential to improve asthma control alongside significant reductions in greenhouse gas emissions. Real-world evidence shows that once-daily long-acting combination dry-powder inhalers (DPIs) can improve compliance and asthma control, and reduce the carbon footprint of care. Similarly, maintenance and reliever therapy (MART), which uses combination reliever and inhaled steroids in one device (usually a DPI), can simplify therapy, improve asthma control and reduce greenhouse gas emissions. Both treatment strategies are popular with patients, most of whom would be willing to change treatment to reduce their carbon footprint. By focussing on patients who are currently using high amounts of salbutamol MDI and prioritising inhaled steroids via DPIs, there are golden opportunities to make asthma care in the UK more effective, safer and greener.

The Effect of Pain Conditioning on Experimentally Evoked Cough: Evidence of Impaired Endogenous Inhibitory Control Mechanisms in Refractory Chronic Cough.

The pathophysiology of refractory chronic cough (RCC) is unclear. We hypothesised that endogenous inhibitory control mechanisms, such as those activated by noxiousstimuli inducing pain (conditioned pain modulation (CPM)), may be capable of inhibiting coughing and urge to cough evoked by inhaled capsaicin. Furthermore, these mechanisms may be impaired in patients with RCC.The objective was to investigate the effects of pain on cough and urge to cough (UTC) in healthy volunteers (HV) and RCC. HV and RCC patients underwent a randomised, controlled, 4-way cross-over study comparing the effect of 4 interventions on capsaicin evoked coughing and UTC. The interventions comprised immersing a hand in i) noxious cold-water, ii) warm water, iii) warm water but instructed to voluntarily supress coughing and iv) no intervention. The co-primary outcomes were numbers of evoked coughs and UTC scores.Twenty HV (mean age 50.1(±SD14.2), M:F 10:10) and 20 RCC (age 60.1(±7.9), M:F 9:11) participated. Overall, noxious cold water reduced capsaicin evoked UTC scores and cough numbers compared with warm water (1.6 (95% C.I. 1.3-2.0) versus 2.2 (1.8-2.6) p<0.001; 4.8 coughs (3.7-6.2) versus 7.9 coughs (6.7-9.5) p<0.001, respectively). HV and RCC demonstrated similar reductions in the UTC during noxious cold-water immersion, but noxious cold water and voluntary suppression interventions were less effective in RCC than HV in reducing capsaicin evoked cough (p=0.041).Endogenous inhibitory control mechanisms, specifically those activated by pain, can reduce both coughing and the UTC. Impairment of endogenous inhibitory control mechanisms may contribute to excessive coughing in RCC.

Association of bacterial load in drinking water and allergic diseases in childhood.

BACKGROUND: Treatment of drinking water may decrease microbial exposure. OBJECTIVE: To investigate whether bacterial load in drinking water is associated with altered risk of allergic diseases. METHODS: We recruited 1,110 schoolchildren aged 6-16 years between 2011 and 2013 in Pozega-Slavonia County in Croatia, where we capitalized on a natural experiment whereby individuals receive drinking water through public mains supply or individual wells. We obtained data on microbial content of drinking water for all participants; 585 children were randomly selected for more detailed assessments, including skin prick testing. Since water supply was highly correlated with rural residence, we compared clinical outcomes across four groups (Rural/Individual, Rural/Public, Urban/Individual and Urban/Public). For each child, we derived quantitative index of microbial exposure (bacterial load in the drinking water measured during the child's first year of life). RESULTS: Cumulative bacterial load in drinking water was higher (median [IQR]: 6390 [4190-9550] vs 0 [0-0]; P < .0001), and lifetime prevalence of allergic diseases was significantly lower among children with individual supply (5.5% vs 2.3%, P = .01; 14.4% vs 6.7%, P < .001; 25.2% vs 15.1%, P < .001; asthma, atopic dermatitis [AD] and rhinitis, respectively). Compared with the reference group (Urban/Public), there was a significant reduction in the risk of ever asthma, AD and rhinitis amongst rural children with individual supply: OR [95% CI]: 0.14 [0.03,0.67], P = .013; 0.20 [0.09,0.43], P < .001; 0.17 [0.10,0.32], P < .001. Protection was also observed in the Rural/Public group, but the effect was consistently highest among Rural/Individual children. In the quantitative analysis, the risk of allergic diseases decreased significantly with increasing bacterial load in drinking water in the first year of life (0.79 [0.70,0.88], P < .001; 0.90 [0.83,0.99], P = .025; 0.80 [0.74,0.86], P < .001; current wheeze, AD and rhinitis). CONCLUSIONS AND CLINICAL RELEVANCE: High commensal bacterial content in drinking water may protect against allergic diseases.

Effectiveness of Fluticasone Furoate-Vilanterol for COPD in Clinical Practice.

BACKGROUND: Evidence for the management of chronic obstructive pulmonary disease (COPD) comes from closely monitored efficacy trials involving groups of patients who were selected on the basis of restricted entry criteria. There is a need for randomized trials to be conducted in conditions that are closer to usual clinical practice. METHODS: In a controlled effectiveness trial conducted in 75 general practices, we randomly assigned 2799 patients with COPD to a once-daily inhaled combination of fluticasone furoate at a dose of 100 µg and vilanterol at a dose of 25 µg (the fluticasone furoate-vilanterol group) or to usual care (the usual-care group). The primary outcome was the rate of moderate or severe exacerbations among patients who had had an exacerbation within 1 year before the trial. Secondary outcomes were the rates of primary care contact (contact with a general practitioner, nurse, or other health care professional) and secondary care contact (inpatient admission, outpatient visit with a specialist, or visit to the emergency department), modification of the initial trial treatment for COPD, and the rate of exacerbations among patients who had had an exacerbation within 3 years before the trial, as assessed in a time-to-event analysis. RESULTS: The rate of moderate or severe exacerbations was significantly lower, by 8.4% (95% confidence interval, 1.1 to 15.2), with fluticasone furoate-vilanterol therapy than with usual care (P=0.02). There was no significant difference in the annual rate of COPD-related contacts to primary or secondary care. There were no significant between-group differences in the rates of the first moderate or severe exacerbation and the first severe exacerbation in the time-to-event analyses. There were no excess serious adverse events of pneumonia in the fluticasone furoate-vilanterol group. The numbers of other serious adverse events were similar in the two groups. CONCLUSIONS: In patients with COPD and a history of exacerbations, a once-daily treatment regimen of combined fluticasone furoate and vilanterol was associated with a lower rate of exacerbations than usual care, without a greater risk of serious adverse events. (Funded by GlaxoSmithKline; Salford Lung Study ClinicalTrials.gov number, NCT01551758 .).

Effectiveness of fluticasone furoate plus vilanterol on asthma control in clinical practice: an open-label, parallel group, randomised controlled trial.

BACKGROUND: Evidence for management of asthma comes from closely monitored efficacy trials done in highly selected patient groups. There is a need for randomised trials that are closer to usual clinical practice. METHODS: We did an open-label, randomised, controlled, two-arm effectiveness trial at 74 general practice clinics in Salford and South Manchester, UK. Patients aged 18 years or older with a general practitioner's diagnosis of symptomatic asthma and on maintenance inhaler therapy were randomly assigned to initiate treatment with a once-daily inhaled combination of either 100 µg or 200 µg fluticasone furoate with 25 µg vilanterol or optimised usual care and followed up for 12 months. The primary endpoint was the percentage of patients who achieved an asthma control test (ACT) score of 20 or greater or an increase in ACT score from baseline of 3 or greater at 24 weeks (termed responders), in patients with a baseline ACT score less than 20 (the primary effectiveness analysis population). All effectiveness analyses were done according to the intention-to-treat principle. This study is registered with ClinicalTrials.gov, number NCT01706198. FINDINGS: Between Nov 12, 2012, and Dec 16, 2016, 4725 patients were enrolled and 4233 randomly assigned to initiate treatment with fluticasone furoate and vilanterol (n=2114) or usual care (n=2119). 1207 patients (605 assigned to usual care, 602 to fluticasone furoate and vilanterol) had a baseline ACT score greater than or equal to 20 and were thus excluded from the primary effectiveness analysis population. At week 24, the odds of being a responder were higher for patients who initiated treatment with fluticasone furoate and vilanterol than for those on usual care (977 [71%] of 1373 in the fluticasone furoate and vilanterol group vs 784 [56%] of 1399 in the usual care group; odds ratio [OR] 2·00 [95% CI 1·70-2·34], p<0·0001). At week 24, the adjusted mean ACT score increased by 4·4 points from baseline in patients initiated with fluticasone furoate and vilanterol, compared with 2·8 points in the usual care group (difference 1·6 [95% CI 1·3-2·0], p<0·0001). This result was consistent for the duration of the study. Pneumonia was uncommon, with no differences between groups; there was no difference in other serious adverse events between the groups. INTERPRETATION: In patients with a general practitioner's diagnosis of symptomatic asthma and on maintenance inhaler therapy, initiation of a once-daily treatment regimen of combined fluticasone furoate and vilanterol improved asthma control without increasing the risk of serious adverse events when compared with optimised usual care. FUNDING: GlaxoSmithKline.

Effectiveness versus efficacy trials in COPD: how study design influences outcomes and applicability.

Guidelines for chronic obstructive pulmonary disease (COPD) management are based largely on results from double-blind randomised controlled trials (RCTs) of efficacy. These trials have high internal validity and test whether a drug is efficacious, but they are conducted in highly selected populations that may differ significantly from patients with COPD seen in routine practice.We compared the baseline characteristics, healthcare use and outcomes between the Salford Lung Study (SLS), an open-label effectiveness RCT, with six recent large-scale efficacy RCTs. We also calculated the proportion of SLS patients who would have been eligible for inclusion in an efficacy RCT by applying the inclusion criteria used in efficacy trials of combination treatments.SLS patients were older, included more females and more current smokers, had more comorbidities (including asthma), and had more often experienced exacerbations prior to inclusion. In the SLS, rates of moderate or severe exacerbations, incidence of overall serious adverse events (SAEs), and SAEs of pneumonia were more frequent. A maximum of 30% of patients enrolled in the SLS would have been eligible for a phase IIIa regulatory exacerbation study.Patients in large COPD efficacy RCTs have limited representativeness compared with an effectiveness trial. This should be considered when interpreting efficacy RCT outcomes and their inclusion into guidelines.

Neurophenotypes in Airway Diseases. Insights from Translational Cough Studies.

RATIONALE: Most airway diseases, including chronic obstructive pulmonary disease (COPD), are associated with excessive coughing. The extent to which this may be a consequence of increased activation of vagal afferents by pathology in the airways (e.g., inflammatory mediators, excessive mucus) or an altered neuronal phenotype is unknown. Understanding whether respiratory diseases are associated with dysfunction of airway sensory nerves has the potential to identify novel therapeutic targets. OBJECTIVES: To assess the changes in cough responses to a range of inhaled irritants in COPD and model these in animals to investigate the underlying mechanisms. METHODS: Cough responses to inhaled stimuli in patients with COPD, healthy smokers, refractory chronic cough, asthma, and healthy volunteers were assessed and compared with vagus/airway nerve and cough responses in a cigarette smoke (CS) exposure guinea pig model. MEASUREMENTS AND MAIN RESULTS: Patients with COPD had heightened cough responses to capsaicin but reduced responses to prostaglandin E2 compared with healthy volunteers. Furthermore, the different patient groups all exhibited different patterns of modulation of cough responses. Consistent with these findings, capsaicin caused a greater number of coughs in CS-exposed guinea pigs than in control animals; similar increased responses were observed in ex vivo vagus nerve and neuron cell bodies in the vagal ganglia. However, responses to prostaglandin E2 were decreased by CS exposure. CONCLUSIONS: CS exposure is capable of inducing responses consistent with phenotypic switching in airway sensory nerves comparable with the cough responses observed in patients with COPD. Moreover, the differing profiles of cough responses support the concept of disease-specific neurophenotypes in airway disease. Clinical trial registered with www.clinicaltrials.gov (NCT 01297790).

Research in progress: Medical Research Council United Kingdom Refractory Asthma Stratification Programme (RASP-UK).

The UK Refractory Asthma Stratification Programme (RASP-UK) will explore novel biomarker stratification strategies in severe asthma to improve clinical management and accelerate development of new therapies. Prior asthma mechanistic studies have not stratified on inflammatory phenotype and the understanding of pathophysiological mechanisms in asthma without Type 2 cytokine inflammation is limited. RASP-UK will objectively assess adherence to corticosteroids (CS) and examine a novel composite biomarker strategy to optimise CS dose; this will also address what proportion of patients with severe asthma have persistent symptoms without eosinophilic airways inflammation after progressive CS withdrawal. There will be interactive partnership with the pharmaceutical industry to facilitate access to stratified populations for novel therapeutic studies.

Morphologic Characterization of Nerves in Whole-Mount Airway Biopsies.

RATIONALE: Neuroplasticity of bronchopulmonary afferent neurons that respond to mechanical and chemical stimuli may sensitize the cough reflex. Afferent drive in cough is carried by the vagus nerve, and vagal afferent nerve terminals have been well defined in animals. Yet, both unmyelinated C fibers and particularly the morphologically distinct, myelinated, nodose-derived mechanoreceptors described in animals are poorly characterized in humans. To date there are no distinctive molecular markers or detailed morphologies available for human bronchopulmonary afferent nerves. OBJECTIVES: Morphologic and neuromolecular characterization of the afferent nerves that are potentially involved in cough in humans. METHODS: A whole-mount immunofluorescence approach, rarely used in human lung tissue, was used with antibodies specific to protein gene product 9.5 (PGP9.5) and, for the first time in human lung tissue, 200-kD neurofilament subunit. MEASUREMENTS AND MAIN RESULTS: We have developed a robust technique to visualize fibers consistent with autonomic and C fibers and pulmonary neuroendocrine cells. A group of morphologically distinct, 200-kD neurofilament-immunopositive myelinated afferent fibers, a subpopulation of which did not express PGP9.5, was also identified. CONCLUSIONS: PGP9.5-immunonegative nerves are strikingly similar to myelinated airway afferents, the cough receptor, and smooth muscle-associated airway receptors described in rodents. These have never been described in humans. Full description of human airway nerves is critical to the translation of animal studies to the clinical setting.