Fractional nitric oxide measurement in exhaled air (FeNO): perspectives in the management of respiratory diseases

Abstract

Exhaled nitric oxide (NO) production, upregulated by inflammatory cytokines and mediators in central and peripheral airways, can be easily and non-invasively detected in exhaled air in asthma and other respiratory conditions as a promising tool for disease monitoring. The American Thoracic Society and European Respiratory Society released recommendations that standardize the measurement of the fractional exhaled NO (FeNO). In asthma, increased FeNO reflects eosinophilic-mediated inflammatory pathways and, as a biomarker of T2 inflammation can be used to identify asthma T2 phenotype. In this setting its measurement has shown to be an important tool especially in the diagnostic process, in the assessment and evaluation of poor adherence or predicting positive response to inhaled corticosteroids treatment, in phenotyping severe asthma patients and as a biomarker to predict the response to biologic treatments. The discovery of the role of NO in the pathogenesis of different diseases affecting the airways and the possibility to estimate the predominant site of increased NO production has provided new insight on its regulatory role in the airways, making it suitable for a potential extended use in clinical practice for different pulmonary diseases, even though its role remains less clear than in asthma. Monitoring FeNO in pulmonary obstructive lung diseases including chronic bronchitis and emphysema, interstitial lung diseases, obstructive sleep apnea and other pulmonary diseases is still under debate but has opened up a window to the role NO may play in the management of these diseases. The use of FeNO is reliable, cost effective and recommendable in both adults and children, and should be implemented in the management of patients with asthma and other respiratory conditions.

Keywords

asthma management central and peripheral airways fractional exhaled nitric oxide respiratory diseases

Introduction

Endogenous nitric oxide (NO) is a gaseous molecule produced by residential and inflammatory cells in both large and peripheral airways playing an important role in regulating airway and vascular function. It is produced by three isoforms of NO synthases: neuronal nitric oxide synthase (NOS) (nNOS, NOS1), endothelial NOS (eNOS, NOS3), and inducible NOS (iNOS, NOS2) with different expression and pathophysiologic roles in the airways. The presence of NO in exhaled air was first described in 1991 by Gustafsson et al.¹ and, in 1992 NO was voted molecule of the year by the scientific journal Science.² Subsequently, researchers from the Karolinska Institute in Sweden (1993) were the first to report increased fractional exhaled NO (FeNO) levels in asthmatics by induced cell types during an inflammatory response.³ This was soon followed by a number of publications reporting high FeNO in subjects with asthma.⁴ Since then, a large amount of evidences has confirmed a role of endogenous NO as responsible for the physiological regulation of airways, thus its possible involvement in many pulmonary conditions. Moreover, the in-depth study of different inflammatory pathways and the development of techniques to detect FeNO using different flows offers the possibility to estimate the predominant site of increased FeNO production, making it suitable for a potential extended use in the clinical management of many conditions even though its role remains less established than in asthma. The identification and monitoring of biomarkers like FeNO in obstructive lung diseases including chronic bronchitis and emphysema, interstitial lung diseases (ILDs), obstructive sleep apnea (OSA) and other respiratory conditions might reflect the disease state and be helpful in understanding the underlying pathology though its role is still controversial and under debate. The aim of this narrative review is to resume the most important highlights on the usefulness of FeNO as a consolidated tool for assessment and management of bronchial asthma and to provide new insight about the role of FeNO in the regulation of the airways in different respiratory diseases including chronic obstructive pulmonary disease (COPD) and emphysema, ILDs, OSA and other pulmonary diseases to open up a window on its potential contribution as a surrogate biomarker of inflammation and as a method to detect disease activity, useful in the clinical management of these diseases. The use of FeNO is reliable, recommendable and cost effective in both adults and children, and should be implemented and encouraged when used in the diagnosis and management of patients with asthma and other respiratory conditions as shown in Figure 1.

Figure 1.

Main respiratory diseases in which fractional exhaled nitric oxide assessment may have a potential role.

Exhaled NO assessment

The gold standard technique for the detection of FeNO uses a chemiluminescence reaction. The reaction between the NO in the breath sample and ozone determines the emission of electromagnetic radiation having wavelength between 600 and 3000 μm which, detected and amplified, provide a proportional signal of the concentration of NO.⁵ The chemiluminescence equipment needs an external, highly sensitive source of NO to generate ozone, with detection thresholds of parts per billion (1:109 ppb) and a fast response time (0.5–0.7 s).⁶ The chemiluminescence FeNO analyzers currently on the market are NOA 280i (Sievers, GE Analytical Instruments), Logan model LR2149 (Logan Research), NIOX (Circassia), and CLD 88 (Eco Medics).An electrochemical sensor is able to convert NO concentration into an electrical signal, in the presence of a buffer system and a catalytic sensor. A subsequent chemical reaction induces a quantifiable physical change in the sample directly proportional to the concentration of NO. Among electrochemical or infrared sensor devices commercially available there are: NIOX VERO (Circassia), Medisoft (Hypair), NObreath (Bedfont Scientific), and Vivatmo-PRO (Bosch). The NIOX VERO is portable, usable in both adults and children with a sensor to be replaced after 100–300 measurements. Patients should exhale for at least 10 s at a pressure ensuring a flow rate of 50 ± 5 ml/s. An electrochemical sensor evaluates the final 3 s of exhalation and subsequently express the result in a range of 5–300 ppb. NObreath requires 12 s of exhalation in adults and 10 in children.⁶ The Medisoft device is semi-portable and can offer either off-line and on-line measurements, in a range of 0–600 ppb. The ‘Off-line’ measurement involves the analysis of a previously collected gas sample, while the ‘on-line’ one collects gas sample in real time. Recent research has shown a good correlation between some of these devices although FeNO absolute values may show clinically relevant differences.⁷ Recently, a comparison between FeNO levels measured by different portable analyzers produced by different manufacturers was published. FeNO levels obtained using the NIOX MINO® and NObreath® were more variable than those measured using the NA623NP®. There were strong positive correlations in FeNO levels measured by the NA623NP®, NIOX MINO®, and NObreath® (p < 0.001). No significant differences were observed in FeNO levels obtained using the NIOX MINO® and NIOX VERO®.⁸ The authors concluded that globally it seems that there may be significant differences in the FeNO measurements carried out with different instruments especially if from different manufacturers; this must be kept in mind in clinical practice and in the evaluation of studies especially in relation to the normal cut-offs considered. Optical sensors consisting of a laser light source that reacts with sample gas, a collection cell, and a sensing system that records signals in the mid-infrared absorption band (5.1–5.7 μm) have been developed.⁹

Exhaled NO measurement

Standardized procedures for measuring FeNO concentration were approved by the American Thoracic Society (ATS) and the European Respiratory Society (ERS).¹⁰ The document was recently integrated by adding the evaluation of nasal NO measurement and introducing recommendations for NO dynamic models as extended NO analysis has been developed.^11,12 According to the guidelines, FeNO measurement in adults occurs instantaneously during an expiratory maneuver. The subject should inhale to total lung capacity with purified air to limit contamination of the sample with potentially high NO levels in the surrounding environment. Then exhalation should be performed for 10 s at a pressure of 5–20 cmH₂O, which ensures the closing of the soft palate to reduce the potential contamination from higher airways and paranasal sinuses.¹³ An exhalation is considered adequate if a stable concentration level (plateau) is reached at a flow rate of 50 ml/s. An expiratory flow of 50 ml/s is recommended, considering that the region of interest of NO production is the lower respiratory tract. The plateau values, corresponding at the expiratory flow rate, vary considerably with the flow rate due to the diffusion of the NO into airways and transit time. For this reason, low speeds (<10 ml/s) result in higher levels. The measurement of FeNO at a single expiratory flow does not allow the identification of the site of NO production in the respiratory system. Therefore, mathematical models have been developed, extensively revised,¹⁴ to calculate the production of pulmonary NO. The fractions of NO produced in the airway walls and alveolar region [concentration of alveolar nitric oxide (CANO)] can be calculated at different high flows. In addition, the association of mathematical calculations with NO measurements at high and low air flows can provide values of the concentration of the gas released by the air conduction system and the capacity of diffusion of NO. There is a possibility to modify expiratory flow rates by resistors in all chemiluminescence analyzers, whereas most electrochemical sensors are not suitable for the analysis of multiple streams except for Medisoft. The two-compartment model allowing the evaluation of NO dynamics in central and in more distal airways with mathematical models to calculate bronchial and alveolar NO parameters measuring NO at different exhalation flows seems to be theoretically plausible, even though, the feasibility to exhale at low flows in patients with obstructive lung diseases such as COPD or severe asthma may be limited. To address this issue, recent research proposed a suitable protocol for extended NO analysis to calculate CANO with a linear model obtained with elevated expiratory high flow rates in patients with asthma of differing severity including severe phenotypes.¹⁵

Interpretation of FeNO measurement results

In clinical practice a FeNO value <25 ppb (<20 ppb in children) is considered normal, while values between 25 and 50 ppb (20 and 35 ppb in children) should be interpreted in the clinical context. Reference values for FeNO have been defined for healthy adults, older aging people and children.^16–19 The upper limit of FeNO₅₀ for healthy individuals, confirmed by a very large survey by the National Health and Nutrition Examination Survey, reported the fifth to 95th percentile values of FeNO being 3.5–39 ppb for subjects 12–80 years of age.²⁰ A recent review reported different studies in which the application of multiple regression modeling reported normal values of FeNO in never-smoking adults, ranging from 24 to 53 ppb. The data showed, in an unselected population, a distribution of FeNO skewed to the right, therefore the authors concluded that it is unlikely that reference values derived from a ‘normal’ population will be as helpful as cut points in patients with airway disease or respiratory symptoms.²¹ FeNO values may be influenced by several conditions related to the patients like genetics, sex, height and weight, use of medications (corticosteroids, ACE-inhibitors), atopy, diet and smoking habits.^22,23 Some foods are known to be capable of influencing FeNO values, with some enhancing, while others lowering FeNO. Nitrate-rich foods, especially leafy vegetables such as lettuce, spinach and kale, can increase FeNO.^24,25 Chronic alcohol consumption and recent consumption of coffee were both related with lowering of FeNO values.^26,27 Moreover, NO is produced by NOS, and alterations in NOS activity or expression may contribute to the pathophysiology of asthma. Genetic variations in NOS genes have been suggested as a potential contributor to altered NO production and airway inflammation in asthma patients. A large meta-analysis in 2023²⁸ concluded that several polymorph NOS gene variants appear to have influence on asthma prevalence or outcomes. However, current evidence is still affected by too many different factors such as ethnicity, study designs and disease variants to drive clear conclusions. The relationship between NOS gene mutations and FeNO levels is complex and may be influenced by other factors such as age, sex, and disease severity. Further research is needed to fully understand the relationship between NOS gene mutations and FeNO levels in patients with respiratory diseases. For these reasons, to avoid any analytic interference, it is strongly recommended to perform FeNO measurement before spirometric maneuvers and before consuming any food, beverage or cigarette smoking. The latter, in particular is one of the most important determinants of FeNO levels. Current smokers show lower levels of FeNO when compared to ex-smokers or non-smokers²⁹ and different mechanisms have been suggested such as a mechanism of downregulation of NO synthetase by NO from cigarette smoke, increased breakdown of NO or lack of tetrahydrobiopterin.³⁰ Also, viral respiratory infections can influence FeNO values³¹ as represented in Table 1.

Table 1.

Factors influencing FeNO production.

Variables	Increasing effect	Decreasing effect
Demographics
Age	∧
Height	∧
Gender: Males Females	∧	∨
Type of inflammation
Eosinophil inflammation	∧
Neutrophil inflammation		∨
Comorbidities
Allergic rhinitis	∧
CRSwNP	∧
Obesity		∨
GERD		∨
Drugs
ICS		∨
Biologics (anti-IL4/IL13-antibody)		∨
Other conditions
Cigarette smoke		∨
Atopy	∧
Viral infections	∧
Acute bronchoconstriction		∨
Asthma exacerbation	∧

CRSwNP, Chronic rhinosinusitis with nasal polyps; FeNO, fractional exhaled nitric oxide; GERD, gastro-esophageal reflux; ICS, inhaled corticosteroids.

Main clinical applications of FeNO: State of art

Asthma

Asthma diagnosis

High FeNO values in patients with relevant symptoms and in combination with other diagnostic options may help in confirming the diagnosis of asthma and, National Institute of Clinical Excellence (NICE) and ERS guidelines recommend supplementing the asthma diagnostic algorithm with FeNO testing in combination with lung function assessment.^32,33 According to current literature, increased FeNO levels (cut-off of 40 ppb in adults and 35 ppb in children) are associated with a higher probability of asthma diagnosis, while in range values do not exclude asthma.³¹ In a study conducted by Karrasch et al.,³⁴ high FeNO levels showed higher specificity than sensitivity and a positive predictive value for asthma, being an important tool in ruling in asthma diagnosis. Although disease prevalence in the population is known to influence the predictive value of the test, the cited study, analyzed populations with widely differing asthma prevalence (9–80%) and found no significant differences in the accuracy of FeNO to diagnosing asthma. FeNO measurement may also be used as a rule-in test for detecting Cough-Variant Asthma (CVA) in adult patients with chronic cough^35,36 and may be useful to predict bronchial hyperactivity in young adults with CVA being an alternative to the use of bronchial provocation tests which can be time-consuming and expensive³⁷ (Table 2).

Table 2.

Reference values and clinical interpretation of FeNO in asthma diagnosis and treatment.

FeNO (ppb)	Low (healthy)	Intermediate	High (asthma)
Adults	<25	25–50	>40
Children (<12 years)	<20	20–35	>35

In clinical practice a FeNO value <25 ppb (<20 ppb in children) is considered normal, values between 25 and 50 ppb (20 and 35 ppb in children) should be interpreted in the clinical context. Values >40 ppb (>35 ppb in children) are likely to reflect a significant T2-mediated inflammation and a good answer to biologics.

FeNO, Fractional exhaled nitric oxide (ppb at flow 50 ml/s).

Asthma control, response to treatments and adherence

Being a T2-mediated biomarker³⁸ related to airway eosinophilic inflammation FeNO was investigated as a promising marker able to predict asthma exacerbations and poor asthma control.^39,40 Moreover, its rapid reduction after anti-inflammatory treatment makes it a suitable tool to predict therapeutic efficacy of treatments. Many studies had previously investigated these aspects but, actually, a recent meta-analysis concluded that the correlation between FeNO levels and asthma control is weak.⁴¹ These results may be partly explained by clinical characteristics of the patients: FeNO levels correlates with asthma control only in patients not regularly treated with ICS;⁴² moreover sino-nasal comorbidities frequently correlated with asthma may influence FeNO levels independently.⁴³ Some asthma phenotypes are characterized by eosinophilic inflammation (T2 phenotypes) which are sensitive to glucocorticoids; in these patients FeNO (directly related to eosinophilic inflammation) has been reported to be an ideal tool in monitoring the clinical response to inhaled or systemic corticosteroid treatments especially in steroid-naïve patients, with high levels of FeNO associated with a good response to treatment.^40,44 Moreover, elevated high FeNO levels before starting ICS treatment may predict improvements in lung function either in adults and children,⁴⁵ reduction of symptoms⁴⁶ and a better quality of life.⁴⁷ FeNO assessment has also been proposed for therapeutic strategies based on personalized treatment regimen according to FeNO levels^44,48,49 (combined impact of exhaled nitric oxide) due to the rapid response to anti-inflammatory treatments together with a rapid increase in case of worsening of asthma control and exacerbations.^39,50 A meta-analysis including data from three Cochrane reviews^51,52 concluded that a strategy based on FeNO levels for tailoring asthma therapy resulted in significant attenuation of exacerbations in adults and children when compared to a guideline-based strategy, without increasing ICS dose.⁴⁴ FeNO has also been examined as a tool for the identification of non-adherence to treatment, helping to distinguish ‘difficult to treat’ versus actual severe asthma patients. Patients not adhering to treatment tend to have high FeNO values^53–58 and therefore like all other asthmatics who do not respond to therapy, would undergo to the so-called ‘FeNO suppression test’.⁵⁹ Globally, recent evidence-based recommendations suggest that FeNO testing is indicated to optimize asthma treatment, in particular to control exacerbations and titrating ICS therapy and should be introduced in the management of asthma, from diagnosis to follow-up⁶⁰ as summarized in Figure 2. Overall, literature evidence underlines FeNO utility in the diagnosis of asthma, prediction of ICS response and evaluation of the risk of exacerbation/loss of control and lung function impairment, although the clinical use of a single biomarker in asthma has to be carefully weighted according to the goals it is demonstrated to satisfy in clinical trials and real-life experiences. Moreover, several factors may influence the measure of FeNO as mentioned before (the variability over time, corticosteroids intake, comorbidities, environmental exposure or habits). Among positive aspects FeNO has been shown to be easy-to-measure, reliable and cost-effective, useful both in adults and children. Therefore, the use of FeNO in combination with other diagnostic tests and biomarkers is highly suggested to increase the specificity of the outcomes and, as an integrated tool, may be useful for routine clinical practice (Figure 3).

Figure 2.

Usefulness of fractional exhaled nitric oxide measurement in the management of asthma and severe asthma.

Figure 3.

Flow diagram of treatment decisions based on fractional exhaled nitric oxide measurement at initial evaluation of patients with suspected asthma.

Severe asthma

Severe asthma is defined by the ATS/ERS guidelines on the basis of the treatment needed to achieve asthma control. Recent studies have shown the association of severe asthma with the degree of eosinophilic inflammation⁶¹ and with an increased expression of iNOS which is the major player responsible for the production of FeNO. iNOS expression is mediated by IL-13, one of the most important inflammatory cytokines involved in TH2 inflammatory response. The utility of FeNO measurement in the management of severe asthma is still debated, especially in light of the ICS inhibitor action on iNOS. However, a recent study reported elevated FeNO levels in 50% of severe steroid-dependent asthmatics. Moreover, high concentrations of FeNO and eosinophils in blood and sputum that characterize severe late-onset asthma, predominantly non-atopic, were found to be associated with the severity of asthma. Further research has shown that, in severe asthmatics, elevated FeNO values correlated with increased airway reactivity, increased sputum eosinophilia, atopy, hyperventilation, reduced perception of symptoms, increased airflow obstruction, more pronounced air-trapping and higher emergency room visits.⁶² All this evidence suggests that FeNO is a reliable biomarker for identifying patients with severe asthma. According to the Global Initiative for Asthma (GINA) guidelines, to evaluate the asthma phenotype a FeNO ⩾ 20 ppb is considered the cut-off for Type 2 inflammation in severe asthma and may be used together with blood (⩾300 cells/μl) and sputum (⩾3%) eosinophil count. Reference values may be used as general guidance, taking into account that they may be subject to change significantly in patients. FeNO values >50 ppb (>35 ppb in children) are associated with eosinophilic inflammation of the airways and may predict response to anti-inflammatory therapy or a loss of control of the disease³⁹ suggesting the opportunity for treatment with ICS. Instead, a FeNO < 25 ppb (<20 ppb in children) is correlated with lower eosinophilic inflammation and corticosteroid response. Measures >30 ppb have been associated with uncontrolled asthma, while a value between 16 and 25 ppb defines the smallest clinically significant difference (an increase of up to 60% in baseline). During an exacerbation, FeNO can increase by more than 50% and up to 150 ppb following exposure to an allergen or acute infection. FeNO in association with patients’ symptoms monitoring may aid clinicians in gradually decreasing oral corticosteroids (OCS) dose in case of good responses to Type-2 targeted therapy, though the ERS/ATS guidelines suggest that FeNO should be avoided in guiding OCS-therapy in adults with severe asthma.⁶³ More recently, FeNO, being a biomarker of T2 inflammation, has been proposed as a predictor of response/efficacy to treatment with biological agents.^64–67 In particular, FeNO ⩾ 25 ppb, together with eosinophilic counts ⩾150 cells/μl, is predictive of therapeutic efficacy of Dupilumab (an IL-4/IL-13 antagonist) in severe asthmatics on high doses of ICS.^68,69 Also, treatment with Tezepelumab, a humanized antibody targeting thymic stromal lymphopoietin, which acts in an upstream position in the inflammatory cascade and modulating both the Th1 and TH2 pathways, has demonstrated persistent reductions in FeNO levels and blood eosinophil count⁷⁰ which promises to be a therapeutic strategy in severe uncontrolled asthma patients irrespective of the asthma T2 phenotype.^71,72 It is important to underline that FeNO levels are primarily controlled by IL-13-induced upregulation of NOS in the airway epithelium; for this reason, FeNO seems not to be a reliable prediction marker for the efficacy of Mepolizumab and Benralizumab.^73,74 In the DREAM study FeNO failed to show a pharmacodynamic response during Mepolizumab treatment and did not demonstrate as a predictive biomarker for reductions in exacerbation rates.⁷⁵ This suggests that FeNO is not responsive to modulation through the IL-5 pathway and is potentially more relevant to different aspects of the T2 inflammatory response (IL-13). However, a small study on Benralizumab in real life found that the responder could be predicted by FeNO at 40 ppb in Type 2 asthma.⁷⁶ Recently, an Italian Consensus paper supported the use of FeNO in the diagnosis and management of asthma and severe asthma and to titrate treatment with ICS and biologics.¹³ Globally, the current evidence suggests FeNO as a reliable biomarker in the identification of asthmatic patients with a more severe behavior, though the heterogeneity of the disease needs an integration of different tools to completely characterize specific phenotypes.

Non-asthmatic eosinophilic bronchitis

Non-asthmatic eosinophilic bronchitis (NAEB) is characterized by chronic dry cough, sputum eosinophilia and good response to glucocorticoids similarly to bronchial asthma. However, in contrast to asthma, NAEB presents no airflow obstruction and airway hyperresponsiveness. It has been hypothesized that the difference of inflammatory cells location, inflammatory mediators and airway remodeling might result in pathogenesis of NAEB and asthma. Compared with asthmatics, in fact, inflammatory cells mainly infiltrate bronchial mucosa in subjects with NAEB thus explaining the absence of airway hyperresponsiveness, as less inflammatory mediators exert their action on smooth muscle.⁷⁷ Some reports indicate that, in patients with NAEB, FeNO levels are significantly elevated when compared to those of patients with other causes of chronic cough.⁷⁸ According to a meta-analysis, FeNO seems to be less precise in predicting NAEB in non-asthmatic patients with chronic cough. However, the same study highlighted the reliability of FeNO values between 30 and 40 ppb in identifying asthmatics with chronic cough responsive to corticosteroids, hence implying not to neglect the clinical utility of FeNO in these patients.³⁵ A recent systematic review of guidelines and expert panel report on the management of chronic cough due to asthma and non-asthmatic eosinophilic bronchitis (NAEB) in adults and adolescents, suggest that non-invasive measurement of airway inflammation may be useful and the presence of eosinophilic airway inflammation is likely to be associated with a more favorable response to ICS especially in patients with chronic cough due to asthma.⁷⁹

COPD and related comorbidities

Fractional exhaled nitric oxide in COPD

The clinical value of FeNO in COPD patients despite its potential benefit as a non-invasive monitoring biomarker of airway inflammation during routine outpatient visits (easy and standardized test, with low intra-subject variability) is still debated with smoking habits, disease severity and ICS use being important factors influencing FeNO levels. According to a previous meta-analysis, COPD induces a slight increase in NO, with higher levels in former smokers than in current smokers; globally no association has been found between the levels of FeNO and COPD flare-ups⁸⁰ even though in a recent large prospective observational study of 226 COPD patients, an association was reported between increased FeNO and the risk of COPD exacerbations.⁸¹ Recent evidence underlines that FeNO at a flow of 50 ml/s (FeNO₅₀) is related to sputum eosinophilia in COPD,^82,83 and may be considered as a marker of T2 inflammation in the central bronchial airways. Moreover, the persistently elevated FeNO₅₀ in stable COPD has been found to be associated with an increased risk of exacerbations⁸¹ and to predict inhaled and systemic steroid responsiveness in stable and disease exacerbations, respectively.^84,85 However, chronic airflow limitation in COPD is the result of either small airway disease and lung parenchyma destruction,⁸⁶ which may not be captured by FeNO₅₀. In this setting the extended NO analysis allows the assessment of pulmonary NO dynamics in the central bronchial and more distal airways (small airways and the alveolar/acinar region).¹¹ A recent, interesting study investigated for the first time the CANO during COPD exacerbations finding an increased CANO in both stable and disease exacerbation, remaining unchanged during convalescence and lung function improvement, while FeNO₅₀ reflecting central airway production was found to be elevated only during exacerbations and suppressed in convalescence. The authors also found that CANO was correlated with circulating eosinophils suggesting a possible upregulation of iNOS/nNOS activity in distal airways, but not correlated with airflow limitation markers of small airway dysfunction/alveolar integrity.⁸⁷ They concluded that CANO and bronchial NO parameters, reflecting nitrative stress at different anatomical sites, can be successfully measured in stable patients with severe airflow limitation and during COPD exacerbation using a feasible extended NO analysis. Moreover, central and peripheral airway NO, is differentially regulated in stable and exacerbated COPD suggesting CANO as a marker for detecting the constantly elevated nitrative stress in distal airways, while bronchial NO to monitor inflammatory activity during an exacerbation. The activity of NOS isoenzymes such as nNOS expression has been previously shown to be linked with the severity of airflow limitation in COPD;⁸⁸ therefore modulation of nitrative stress pathways in peripheral airways may be a potential therapeutic goal even though possible clinical application of CANO should be further explored in certain aspects, including its role in predicting disease progression or exacerbation development. Patients with COPD have a predominantly neutrophilic airway inflammation, although new scientific evidence shows that a proportion of COPD patients present with an eosinophilic airway inflammation. In these patients a significant direct correlation between FeNO and blood eosinophil count in stable disease, especially in GOLD E patients with frequent exacerbations was reported.⁸⁹ FeNO was also shown, in the same phenotype of patients, to predict response to ICS treatment during stable disease⁹⁰ and to systemic corticosteroid treatment in patients suffering an acute exacerbation.⁸⁵ In conclusion, in this specific COPD phenotype, FeNO, as eosinophilic surrogate biomarker, may have an important role, being an easier and useful tool to identify those patients who are most likely to benefit from ICS.

FeNO in alpha-1 antitrypsin deficiency

Alpha-1 antitrypsin (AAT) deficiency is a genetic disorder that, in the homozygotic form of the disease, is considered an important risk factor for developing COPD. Severe AAT deficiency (PiZZ: homozigous genotype) is characterized by lower FeNO levels compared to healthy non-smokers and COPD patients, due to the predictable pulmonary function impairment in these patients.^91,92 On the other hand, patients with a lower reduction of plasma AAT levels (PiMZ: heterozygous genotype) show increased FeNO levels, and an inverse correlation with plasma AAT levels.⁹³ In these patients FeNO measurement may be important to monitor the progression of airway inflammation toward possible COPD development.⁹⁴

FeNO in pulmonary arterial hypertension and cardiovascular diseases

An altered NO signaling in the pulmonary endothelium due to the decreased expression and dysfunction of eNOS contributes to the increased vascular smooth muscle tone and vascular remodeling, leading to the development and progression of pulmonary arterial hypertension (PAH). NO deficiency of the pulmonary vasculature can be assessed by detecting FeNO, a non-invasive biomarker that may be useful to correlate with changes in pulmonary hemodynamics and predicting therapies response. It has been shown that patients with PAH may have lower,^95–98 similar^97,99–101 or elevated FeNO levels compared to health¹⁰² and that patients who respond to therapy replacing NO (phosphodiesterase Type 5 inhibitors, prostacyclin and endothelin receptor antagonists, other pharmacological strategies) show higher FeNO levels compared to those who do not, suggesting a role of FeNO in monitoring response to therapy.^97,103–107 However, this remains controversial as some studies failed to confirm this correlation.^108–111 Several experimental and human studies highlighted the involvement of the reduced endothelial NO signaling in the pulmonary vasculature in PAH patho-mechanisms; however, studies on both bronchial and alveolar NO showed ambiguous results, likely related to unstandardized measurement protocols, low number of patients, different PAH etiologies and confounding analytical and physiological factors.¹¹² Interestingly, a recent study evaluating the clinical value of FeNO in the diagnosis of PAH in patients with acute exacerbation of COPD (AECOPD) demonstrated that FeNO levels at admission can act as an indicator for PAH diagnosis in AECOPD patients.¹¹³ Additionally, FeNO has been assessed in heart failure with ambivalent outcomes.^114–116 In stable ischemic heart disease FeNO was found to be inversely associated with some of risk markers of atherosclerosis (triglycerides and hemoglobin A1c) implying endothelial dysfunction resulting in a decreased production and increased degradation of NO.¹¹⁷

Interstitial lung diseases

The possibility to obtain multiple-flows assessment of FeNO may provide a reliable measurement of bronchial and alveolar production of NO, supporting its potential value as a biomarker in peripheral lung diseases, such as ILDs. Recent research proposed FeNO as a marker of severity of idiopathic pulmonary fibrosis (IPF) and systemic sclerosis-associated ILD. Patients affected by ILD have been reported to have an elevated high flow FeNO and CANO values compared to healthy controls, indicating a potential role of nitrosative stress in lung fibrosis. The significant difference of CANO levels between idiopathic ILDs and connective tissue disease (CTD)-associated ILD is interesting, and may be proposed as a differential diagnosis tool between the two entities, suggesting that NO is also implicated in lung inflammation associated with rheumatological diseases.¹¹⁸ A recent systematic review on the assessment of extended FeNO analysis in the management of patients with ILDs (ILDs associated with systemic sclerosis, idiopathic pulmonary fibrosis, granulomatous ILDs, eosinophilic pneumonia) reported that multiple-flow FeNO analysis, and CANO in particular, demonstrated a suggestive potential in discriminating idiopathic ILDs from connective-ILDs, and in estimating survival and disease progression in terms of forced vital capacity (FVC) deterioration, suggesting its possible implementation in the clinical management of IPF due to its reproducibility, repeatability and non-invasive nature.¹¹⁹

Obstructive sleep apnea

OSA is a sleep disorder characterized by airway and systemic inflammation. Many studies have investigated FeNO patterns in these patients, reporting slightly, but still significantly higher levels of FeNO than healthy controls, especially evident in non-smokers and not consistently related to the severity of the disease.^120–122 Obesity, being a risk factor for OSA and, independently, for airway inflammation has been the objective of studies, although, conflicting results were found comparing FeNO values in obese OSA patients with healthy obese subjects.^123–125 Some studies investigated alveolar NO reporting lower levels in OSA patients compared to healthy subjects with a significant increase after continuous positive airway pressure (CPAP) treatment,^126,127 while other studies found elevated alveolar NO levels in OSA with a positive correlation with the apnea-hypopnea index (AHI), nocturnal desaturation and BMI, linking distal airway inflammation to intermittent hypoxia. No differences were found in bronchial NO.¹²⁸ nNO seems to have a greater value than FeNO reflecting upper airway inflammation in OSA patients.¹²² Interestingly, CPAP treatment has been shown to reduce/normalize high FeNO values in OSA patients, suggesting its potential role as a biomarker of compliance and response to therapy^129,130 although some studies failed to find this correlation.¹²⁷ Overall, even though FeNO seems to have a limited screening potential for sleep disordered breathing,¹³¹ a composite index including BMI, age, carboxy-hemoglobin saturation, neck circumference, FEF₅₀/FIF₅₀ and FeNO has shown to be predictive for OSA,¹³² hence new investigations are needed to overcome methodological limitations and better characterize airway inflammation and oxidative stress in these patients, which may aid in understanding the complex pathophysiology of OSA.¹³³

Conclusions and future perspectives

FeNO is an easy-to-measure, reliable and cost-effective marker of T2 airway inflammation recommended in both adults and children, for asthma diagnosis and monitoring, especially in combination with other diagnostic tests and biomarkers and in the appropriate clinical setting, considering its high variability over time. The use of FeNO may be considered for the identification of precise asthmatic phenotypes and for the consequent development of titrating-therapy strategy aimed at the individual. Of note, FeNO is not completely satisfactory in differentiating Type 2 asthma phenotypes, especially in the context of severe asthma choice of biologic therapies. As there is no currently universal shared algorithm for biomarker application in asthma and severe asthma management, an integrated approach including clinical molecular phenotyping is aimed to see if combination of different markers including FeNO can increase diagnostic accuracy and make decisions more reliable in the whole approach of asthma management and in routinely clinical practice, after the advent of personalized medicine.

The possibility to estimate the predominant site and distinguish the bronchial from the distal airways production of increased fraction of exhaled NO, moreover, opens a window in the monitoring of nitrative stress at different anatomical sites within the airways, making it now suitable for a potential extended use also in the management of other respiratory diseases including COPD and ILDs, though its role is still less clear and hence less established than in asthma. In this field, CANO has demonstrated a potential in discriminating idiopathic ILDs from CTD-ILDs and in estimating survival and disease progression in terms of lung function deterioration, suggesting its possible implementation in the clinical management of IPF, also due to its reproducibility, repeatability and noninvasive nature. These promising findings need to be confirmed by larger, prospective and multicenter studies to better investigate the dynamics of FeNO parameters throughout the clinical course of disease and to evaluate the influence on CANO in related comorbidities and during antifibrotic treatment.

Footnotes

Acknowledgements

None.

Declarations

ORCID iD

Beatrice Ragnoli

References

Gustafsson

Leone

Persson

, et al. Endogenous nitric oxide is present in the exhaled air of rabbits, guinea pigs and humans. Biochem Biophys Res Commun 1991; 181: 852–857.

Koshland

Jr.

The molecule of the year. Science 1992; 258: 1861.

Alving

Weitzberg

Lundberg

. Increased amount of nitric oxide in exhaled air of asthmatics. Eur Respir J 1993; 6: 1368–1370.

Kharitonov

. Exhaled markers of inflammatory lung diseases: ready for routine monitoring? Swiss Med Wkly 2004; 134: 175–192.

Maniscalco

Vitale

Vatrella

, et al. Fractional exhaled nitric oxide-measuring devices: technology update. Med Devices Auckl NZ 2016; 9: 151–160.

Harnan

Tappenden

Essat

, et al. Measurement of exhaled nitric oxide concentration in asthma: a systematic review and economic evaluation of NIOX MINO, NIOX VERO and Nobreath. Health Technol Assess 2015; 19: 1–330.

Molino

Fuschillo

Mosella

, et al. Comparison of three different exhaled nitric oxide analyzers in chronic respiratory disorders. J Breath Res 2019; 13: 021002.

Saito

Kikuchi

Fukuhara

, et al. Comparison of fractional exhaled nitric oxide levels measured by different analyzers produced by different manufacturers. J Asthma 2020; 57: 1216–1226.

Wang

Sahay

. Breath analysis using laser spectroscopic techniques: breath biomarkers, spectral fingerprints, and detection limits. Sensors 2009; 9: 8230–8262.

10.

ATS/ERS recommendations for standardized procedures for the online and offline measurement of exhaled lower respiratory nitric oxide and nasal nitric oxide. American Journal of Respiratory and Critical Care Medicine, https://www.atsjournals.org/doi/10.1164/rccm.200406-710ST (2005, accessed 17 March 2023)

11.

Horváth

Barnes

Loukides

, et al. A european respiratory society technical standard: exhaled biomarkers in lung disease. Eur Respir J 2017; 49: 1600965.

12.

Högman

Malinovschi

Norbäck

, et al. Added value with extended NO analysis in atopy and asthma. Clin Physiol Funct Imaging 2011; 31: 294–299.

13.

Heffler

Carpagnano

Favero

, et al. Fractional exhaled nitric oxide (FENO) in the management of asthma: a position paper of the Italian respiratory society (SIP/IRS) and Italian society of allergy, asthma and clinical immunology (SIAAIC). Multidiscip Respir Med 2020; 15: 36.

14.

George

Hogman

Permutt

, et al. Modeling pulmonary nitric oxide exchange. J Appl Physiol 2004; 96: 831–839.

15.

Lázár

Horváth

Puskás

, et al. A suitable protocol for measuring alveolar nitric oxide in asthma with differing severity to assess peripheral airways inflammation. J Asthma 2019; 56: 584–593.

16.

Kovesi

Kulka

Dales

. Exhaled nitric oxide concentration is affected by age, height, and race in healthy 9- to 12-year-old children. Chest 2008; 133: 169–175.

17.

Olin

A-C

Bake

Torén

. Fraction of exhaled nitric oxide at 50 mL/s: reference values for adult lifelong never-smokers. Chest 2007; 131: 1852–1856.

18.

Olivieri

Talamini

Corradi

, et al. Reference values for exhaled nitric oxide (reveno) study. Respir Res 2006; 7: 94.

19.

Malerba

Damiani

Carpagnano

, et al. Values in elderly people for exhaled nitric oxide study. Rejuvenation Res 2016; 19: 233–238.

20.

See

Christiani

. Normal values and thresholds for the clinical interpretation of exhaled nitric oxide levels in the US general population: results from the national health and nutrition examination survey 2007-2010. Chest 2013; 143: 107–116.

21.

Guida

Bagnasco

Carriero

, et al. Critical evaluation of asthma biomarkers in clinical practice. Front Med 2022; 9: 969243.

22.

Grob

Dweik

. Exhaled nitric oxide in asthma. From diagnosis, to monitoring, to screening: are we there yet? Chest 2008; 133: 837–839.

23.

Ahovuo-Saloranta

Csonka

Lehtimäki

. Basic characteristics and clinical value of FeNO in smoking asthmatics-a systematic review. J Breath Res 2019; 13: 034003.

24.

Kroll

Werchan

Rosenfield

, et al. Acute ingestion of beetroot juice increases exhaled nitric oxide in healthy individuals. PLoS One 2018; 13: e0191030.

25.

Kerley

Kilbride

Greally

, et al. Dietary nitrate acutely and markedly increased exhaled nitric oxide in a cystic fibrosis case. Clin Med Res 2016; 14: 151–155.

26.

Yurach

Davis

Cockcroft

. The effect of caffeinated coffee on airway response to methacholine and exhaled nitric oxide. Respir Med 2011; 105: 1606–1610.

27.

Afshar

Poole

Cao

, et al. Exhaled nitric oxide levels among adults with excessive alcohol consumption. Chest 2016; 150: 196–209.

28.

Fan

Liu

. Association of nitric oxide synthase gene polymorphism with asthma: a systematic review and meta-analysis. Clin Respir J 2023; 17: 516–526.

29.

Malinovschi

Janson

Holmkvist

, et al. Effect of smoking on exhaled nitric oxide and flow-independent nitric oxide exchange parameters. Eur Respir J 2006; 28: 339–345.

30.

Hoyt

Robbins

Habib

, et al. Cigarette smoke decreases inducible nitric oxide synthase in lung epithelial cells. Exp Lung Res 2003; 29: 17–28.

31.

Dweik

Boggs

Erzurum

, et al. An official ATS clinical practice guideline: interpretation of exhaled nitric oxide levels (FENO) for clinical applications. Am J Respir Crit Care Med 2011; 184: 602–615.

32.

Overview|Asthma: diagnosis, monitoring and chronic asthma management|Guidance|NICE, https://www.nice.org.uk/guidance/ng80 (2017, accessed March 17, 2023).

33.

Louis

Satia

Ojanguren

, et al. European respiratory society guidelines for the diagnosis of asthma in adults. Eur Respir J 2022; 60: 2101585.

34.

Karrasch

Linde

Rücker

, et al. Accuracy of FENO for diagnosing asthma: a systematic review. Thorax 2017; 72: 109–116.

35.

Song

W-J

Kim

Shim

J-S

, et al. Diagnostic accuracy of fractional exhaled nitric oxide measurement in predicting cough-variant asthma and eosinophilic bronchitis in adults with chronic cough: a systematic review and meta-analysis. J Allergy Clin Immunol 2017; 140: 701–709.

36.

Zhang

Liu

, et al. Diagnostic value of fractional exhaled nitric oxide in cough-variant asthma: an updated meta-analysis. J Asthma 2020; 57: 335–342.

37.

Malerba

Ragnoli

Azzolina

, et al. Predictive markers of bronchial hyperreactivity in a large cohort of young adults with cough variant asthma. Front Pharmacol 2021; 12: 630334.

38.

Pavlidis

Takahashi

Ng Kee Kwong

, et al. “T2-high” in severe asthma related to blood eosinophil, exhaled nitric oxide and serum periostin. Eur Respir J 2019; 53: 1800938.

39.

Mansur

Srivastava

Sahal

. Disconnect of type 2 biomarkers in severe asthma; dominated by FeNO as a predictor of exacerbations and periostin as predictor of reduced lung function. Respir Med 2018; 143: 31–38.

40.

Lehtimäki

Csonka

Mäkinen

, et al. Predictive value of exhaled nitric oxide in the management of asthma: a systematic review. Eur Respir J 2016; 48: 706–714.

41.

Wang

Pianosi

Keogh

, et al. The clinical utility of fractional exhaled nitric oxide (FeNO) in asthma management. Rockville (MD): Agency for Healthcare Research and Quality (US), http://www.ncbi.nlm.nih.gov/books/NBK487497/ (2017, accessed 17 March 2023).

42.

Lee

Suh

Song

, et al. Asthma control test reflects not only lung function but also airway inflammation in children with stable asthma. J Asthma 2020; 57: 648–653.

43.

Guida

Rolla

Badiu

, et al. Determinants of exhaled nitric oxide in chronic rhinosinusitis. Chest 2010; 137: 658–664.

44.

Petsky

Cates

Kew

, et al. Tailoring asthma treatment on eosinophilic markers (exhaled nitric oxide or sputum eosinophils): a systematic review and meta-analysis. Thorax 2018; 73: 1110–1119.

45.

Szefler

Phillips

Martinez

, et al. Characterization of within-subject responses to fluticasone and montelukast in childhood asthma. J Allergy Clin Immunol 2005; 115: 233–242.

46.

Cowan

Palmay

, et al. Effects of steroid therapy on inflammatory cell subtypes in asthma. Thorax 2010; 65: 384–390.

47.

Price

Buhl

Chan

, et al. Fractional exhaled nitric oxide as a predictor of response to inhaled corticosteroids in patients with non-specific respiratory symptoms and insignificant bronchodilator reversibility: a randomised controlled trial. Lancet Respir Med 2018; 6: 29–39.

48.

Malerba

Ragnoli

Radaeli

, et al. Usefulness of exhaled nitric oxide and sputum eosinophils in the long-term control of eosinophilic asthma. Chest 2008; 134: 733–739.

49.

Malerba

Ragnoli

Radaeli

, et al. Long-term adjustment of stable asthma treatment with fractional exhaled nitric oxide and sputum eosinophils. Eur J Inflammation 2012; 10: 383–392.

50.

Zeiger

Schatz

Zhang

, et al. Elevated exhaled nitric oxide is a clinical indicator of future uncontrolled asthma in asthmatic patients on inhaled corticosteroids. J Allergy Clin Immunol 2011; 128: 412–414.

51.

Petsky

Kew

Turner

, et al. Exhaled nitric oxide levels to guide treatment for adults with asthma. Cochrane Database Syst Rev 2016; 9: CD011440.

52.

Petsky

Kew

Chang

. Exhaled nitric oxide levels to guide treatment for children with asthma. Cochrane Database Syst Rev 2016; 11: CD011439.

53.

Malinovschi

Van Muylem

Michiels

, et al. FeNO as a predictor of asthma control improvement after starting inhaled steroid treatment. Nitric Oxide Biol Chem 2014; 40: 110–116.

54.

Klok

Brand

PLP

. Can exhaled nitric oxide fraction predict adherence to inhaled corticosteroids in atopic and nonatopic children with asthma? J Allergy Clin Immunol Pract 2017; 5: 521–522.

55.

Tsai

Y-G

Sun

H-L

Chien

J-W

, et al. High exhaled nitric oxide levels correlate with nonadherence in acute asthmatic children. Ann Allergy Asthma Immunol 2017; 118: 521–523.e2.

56.

Hunt

Flynn

MacHale

, et al. Reduction in exhaled nitric oxide tracks improved patient inhaler compliance in difficult asthma–a case study. J Asthma 2018; 55: 1373–1375.

57.

Heaney

Busby

Bradding

, et al. Remotely monitored therapy and nitric oxide suppression identifies nonadherence in severe asthma. Am J Respir Crit Care Med 2019; 199: 454–464.

58.

Price

Bosnic-Anticevich

Pavord

, et al. Association of elevated fractional exhaled nitric oxide concentration and blood eosinophil count with severe asthma exacerbations. Clin Transl Allergy 2019; 9: 41.

59.

McNicholl

Stevenson

McGarvey

, et al. The utility of fractional exhaled nitric oxide suppression in the identification of nonadherence in difficult asthma. Am J Respir Crit Care Med 2012; 186: 1102–1108.

60.

Khatri

Iaccarino

Barochia

, et al. Use of fractional exhaled nitric oxide to guide the treatment of asthma: an official American thoracic society clinical practice guideline. Am J Respir Crit Care Med 2021; 204: e97–e109.

61.

Carr

Zeki

Kraft

. Eosinophilic and noneosinophilic asthma. Am J Respir Crit Care Med 2018; 197: 22–37.

62.

Ricciardolo

FLM

Silkoff

. Perspectives on exhaled nitric oxide. J Breath Res 2017; 11: 047104.

63.

Chung

Wenzel

Brozek

, et al. International ERS/ATS guidelines on definition, evaluation and treatment of severe asthma. Eur Respir J 2014; 43: 343–373.

64.

Menzies-Gow

Corren

Bourdin

, et al. Tezepelumab in adults and adolescents with severe, uncontrolled asthma. New Engl J Med 2021; 384: 1800–1809.

65.

Kurokawa

Koya

Takeuchi

, et al. Association of upper and lower airway eosinophilic inflammation with response to omalizumab in patients with severe asthma. J Asthma 2020; 57: 71–78.

66.

Luo

Liu

. The efficacy and safety of antiinterleukin 13, a monoclonal antibody, in adult patients with asthma: a systematic review and meta-analysis. Medicine 2016; 95: e2556.

67.

Russell

Chachi

FitzGerald

, et al. Effect of tralokinumab, an interleukin-13 neutralising monoclonal antibody, on eosinophilic airway inflammation in uncontrolled moderate-to-severe asthma (MESOS): a multicentre, double-blind, randomised, placebo-controlled phase 2 trial. Lancet Respir Med 2018; 6: 499–510.

68.

Castro

Corren

Pavord

, et al. Dupilumab efficacy and safety in moderate-to-severe uncontrolled asthma. N Engl J Med 2018; 378: 2486–2496.

69.

Dupilumab shows rapid and sustained suppression of inflammatory biomarkers in asthma patients in LIBERTY ASTHMA QUEST, https://www.ers-education.org/lr/show-details/?idP=213133 (2020, accessed 17 March 2023).

70.

Corren

Parnes

Wang

, et al. Tezepelumab in adults with uncontrolled asthma. New Engl J Med 2017; 377: 936–946.

71.

Diver

Khalfaoui

Emson

, et al. Effect of tezepelumab on airway inflammatory cells, remodelling, and hyperresponsiveness in patients with moderate-to-severe uncontrolled asthma (CASCADE): a double-blind, randomised, placebo-controlled, phase 2 trial. Lancet Respir Med 2021; 9: 1299–1312.

72.

Ragnoli

Morjaria

Pignatti

, et al. Dupilumab and tezepelumab in severe refractory asthma: new opportunities. Ther Adv Chron Dis 2022; 13: 1–19.

73.

Yancey

Keene

Albers

, et al. Biomarkers for severe eosinophilic asthma. J Allergy Clin Immunol 2017; 140: 1509–1518.

74.

Hearn

Kavanagh

d’Ancona

, et al. The relationship between FeNO and effectiveness of mepolizumab and benralizumab in severe eosinophilic asthma. J Allergy Clin Immunol Pract 2021; 9: 2093–2096.e1.

75.

Pavord

Korn

Howarth

, et al. Mepolizumab for severe eosinophilic asthma (DREAM): a multicentre, double-blind, placebo-controlled trial. J Lancet 2012; 380: 651–659.

76.

Watanabe

Shirai

Hirai

, et al. Blood eosinophil count and FeNO to predict benralizumab effectiveness in real-life severe asthma patients. J Asthma 2022; 59: 1796–1804.

77.

Lai

Chen

Peng

, et al. Non-asthmatic eosinophilic bronchitis and its relationship with asthma. Pulm Pharmacol Ther 2017; 47: 66–71.

78.

Lee

Rhee

Lim

, et al. Fraction of exhaled nitric oxide in patients with acute eosinophilic pneumonia. Chest 2012; 141: 1267–1272.

79.

Côté

Russell

Boulet

. Managing chronic cough due to asthma and NAEB in adults and adolescents: CHEST guideline and expert panel report. Chest 2020; 158: 68–96.

80.

Huang

Wang

, et al. Exhaled nitric oxide in patients with chronic obstructive pulmonary disease: a systematic review and meta-analysis. Int J Chron Obstruct Pulmon Dis 2018; 13: 2695–2705.

81.

Alcázar-Navarrete

Ruiz Rodríguez

Conde Baena

, et al. Persistently elevated exhaled nitric oxide fraction is associated with increased risk of exacerbation in COPD. Eur Respir J 2018; 51: 1701457.

82.

Schleich

Corhay

J-L

Louis

. Blood eosinophil count to predict bronchial eosinophilic inflammation in COPD. Eur Respir J 2016; 47: 1562–1564.

83.

Predicting sputum eosinophilia in exacerbations of COPD using exhaled nitric oxide|SpringerLink, 2013 https://link-springer-com-443.web.bisu.edu.cn/article/10.1007/s10753-013-9653-8 (accessed 21 May 2023).

84.

Kunisaki

Rice

Janoff

, et al. Exhaled nitric oxide, systemic inflammation, and the spirometric response to inhaled fluticasone propionate in severe chronic obstructive pulmonary disease: a prospective study. Ther Adv Respir Dis 2008; 2: 55–64.

85.

Antus

Barta

Horvath

, et al. Relationship between exhaled nitric oxide and treatment response in COPD patients with exacerbations. Respirology 2010; 15: 472–477.

86.

Vogelmeier

Criner

Martinez

, et al. Global strategy for the diagnosis, management, and prevention of chronic obstructive lung disease 2017 report: gold executive summary. Eur Respir J 2017; 49: 1700214.

87.

Lázár

Kelemen Gálffy

, et al. Central and peripheral airway nitric oxide in patients with stable and exacerbated chronic obstructive pulmonary disease. J Breath Res 2018; 12: 036017.

88.

Nitric oxide synthase isoenzyme expression and activity in peripheral lung tissue of patients with chronic obstructive pulmonary disease|American Journal of Respiratory and Critical Care Medicine, https://www.atsjournals.org/doi/full/10.1164/rccm.200904-0493OC (2009, accessed 21 May 2023).

89.

Vincken

Sylvia

Daniel

, et al. The role of FeNO in stable COPD patients with eosinophilic airway inflammation. Respir Med 2021; 181: 106377.

90.

Yamaji

Oishi

Hamada

, et al. Detection of type2 biomarkers for response in COPD. J Breath Res 2020; 14: 026007.

91.

Machado

Stoller

Laskowski

, et al. Low levels of nitric oxide and carbon monoxide in alpha 1-antitrypsin deficiency. J Appl Physiol (1985) 2002; 93: 2038–2043.

92.

Malerba

Clini

Cremona

, et al. Exhaled nitric oxide in patients with PiZZ phenotype-related α 1-anti-trypsin deficiency. Respir Med 2001; 95: 520–525.

93.

Malerba

Ragnoli

Radaeli

. Exhaled nitric oxide levels in alpha-1-antitrypsin PiMZ subjects. J Intern Med 2009; 265: 382–387.

94.

Malerba

Radaeli

Olivini

, et al. Exhaled Nitric Oxide as a Biomarker in COPD and Related Comorbidities. BioMed Res Int 2014;2014: 271918.

95.

Carpagnano

Radaeli

Lacedonia

, et al. Exhaled nitric oxide and exhaled breath temperature as potential biomarkers in patients with pulmonary hypertension. Biomed Res Int 2018; 2018: 7292045.

96.

Malerba

Radaeli

Ragnoli

, et al. Exhaled nitric oxide levels in systemic sclerosis with and without pulmonary involvement. Chest 2007; 132: 575–580.

97.

Girgis

Champion

Diette

, et al. Decreased exhaled nitric oxide in pulmonary arterial hypertension: response to bosentan therapy. Am J Respir Crit Care Med 2005; 172: 352–357.

98.

Kharitonov

Cailes

Black

, et al. Decreased nitric oxide in the exhaled air of patients with systemic sclerosis with pulmonary hypertension. Thorax 1997; 52: 1051–1055.

99.

Girgis

Gugnani

Abrams

, et al. Partitioning of alveolar and conducting airway nitric oxide in scleroderma lung disease. Am J Respir Crit Care Med 2002; 165: 1587–1591.

100.

Riley

Pórszász

Miranda

, et al. Exhaled nitric oxide during exercise in primary pulmonary hypertension and pulmonary fibrosis. Chest 1997; 111: 44–50.

101.

Malinovschi

Henrohn

Eriksson

, et al. Increased plasma and salivary nitrite and decreased bronchial contribution to exhaled NO in pulmonary arterial hypertension. Eur J Clin Invest 2011; 41: 889–897.

102.

Archer

Djaballah

Humbert

, et al. Nitric oxide deficiency in fenfluramine- and dexfenfluramine-induced pulmonary hypertension. Am J Respir Crit Care Med 1998; 158: 1061–1067.

103.

Ozkan

Dweik

Laskowski

, et al. High levels of nitric oxide in individuals with pulmonary hypertension receiving epoprostenol therapy. Lung 2001; 179: 233–243.

104.

Machado

Londhe Nerkar

M-V

Dweik

, et al. Nitric oxide and pulmonary arterial pressures in pulmonary hypertension. Free Radic Biol Med 2004; 37: 1010–1017.

105.

Henrohn

Björkstrand

Lundberg

, et al. Effects of oral supplementation with nitrate-rich beetroot juice in patients with pulmonary arterial hypertension—results from BEET-PAH, an exploratory randomized, double-blind, placebo-controlled, crossover study. J Card Fail 2018; 24: 640–653.

106.

Simon

Vanderpool

Nouraie

, et al. Acute hemodynamic effects of inhaled sodium nitrite in pulmonary hypertension associated with heart failure with preserved ejection fraction. JCI Insight 2016; 1: e89620.

107.

Rix

Vick

Attkins

, et al. Pharmacokinetics, pharmacodynamics, safety, and tolerability of nebulized sodium nitrite (AIR001) following repeat-dose inhalation in healthy subjects. Clin Pharmacokinet 2015; 54: 261–272.

108.

Akbay

Bingol

Kiyan

, et al. Fractional exhaled nitric oxide measurement in pulmonary hypertension: a follow-up study. Clin Appl Thromb Hemost 2018; 24: 483–488.

109.

Malekmohammad

Folkerts

Kashani

, et al. Exhaled nitric oxide is not a biomarker for idiopathic pulmonary arterial hypertension or for treatment efficacy. BMC Pulm Med 2019; 19: 188.

110.

Cao

Mathai

Hummers

, et al. Exhaled nitric oxide in pulmonary arterial hypertension associated with systemic sclerosis. Pulm Circ 2016; 6: 545–550.

111.

Rothe

Karrer

Schindler

. A single dose of sildenafil does not enhance FeNO: a randomised, cross-over and placebo-controlled study. Respir Med 2010; 104: 788–793.

112.

Lázár

Mészáros

Bikov

. The nitric oxide pathway in pulmonary arterial hypertension: pathomechanism, biomarkers and drug targets. Curr Med Chem 2020; 27: 7168–7188.

113.

Guo

Wang

Cui

, et al. Clinical value of FeNO for pulmonary hypertension diagnosis in patients with acute exacerbation of chronic obstructive pulmonary disease. Emerg Med Int 2022; 2022: 9924047.

114.

Hare

Nguyen

Massaro

, et al. Exhaled nitric oxide: a marker of pulmonary hemodynamics in heart failure. J Am Coll Cardiol 2002; 40: 1114–1119.

115.

Agostoni

Bussotti

. Exhaled nitric oxide and exercise performance in heart failure. Arch Physiol Biochem 2003; 111: 293–296.

116.

Schuster

Thakur

Wang

, et al. Increased exhaled nitric oxide levels after exercise in patients with chronic systolic heart failure with pulmonary venous hypertension. J Card Fail 2012; 18: 799–803.

117.

Salonen

Huttunen

Hirvonen

M-R

, et al. Exhaled nitric oxide and atherosclerosis. Eur J Clin Invest 2012; 42: 873–880.

118.

Cameli

Bargagli

Bergantini

, et al. Evaluation of multiple-flows exhaled nitric oxide in idiopathic and non-idiopathic interstitial lung disease. J Breath Res 2019; 13: 026008.

119.

Cameli

Bargagli

Bergantini

, et al. Extended exhaled nitric oxide analysis in interstitial lung diseases: a systematic review. Int J Mol Sci 2020; 21: 6187.

120.

Zhang

Luo

Qiao

, et al. Measurement of exhaled nitric oxide concentration in patients with obstructive sleep apnea: a meta-analysis. Medicine 2017; 96: e6429.

121.

Zhang

Xiao

Luo

, et al. Measurement of fractional exhaled nitric oxide and nasal nitric oxide in male patients with obstructive sleep apnea. Sleep Breath 2019; 23: 785–793.

122.

Duarte

RLM

Rabahi

Oliveira-e-Sá

, et al. Fractional exhaled nitric oxide measurements and screening of obstructive sleep apnea in a sleep-laboratory setting: a cross-sectional study. Lung 2019; 197: 131–137.

123.

Depalo

Carpagnano

Spanevello

, et al. Exhaled NO and iNOS expression in sputum cells of healthy, obese and OSA subjects. J Intern Med 2008; 263: 70–78.

124.

Carpagnano

Spanevello

Sabato

, et al. Exhaled pH, exhaled nitric oxide, and induced sputum cellularity in obese patients with obstructive sleep apnea syndrome. Transl Res J Lab Clin Med 2008; 151: 45–50.

125.

Petrosyan

Perraki

Simoes

, et al. Exhaled breath markers in patients with obstructive sleep apnoea. Sleep Breath 2008; 12: 207–215.

126.

Fortuna

Miralda

Calaf

, et al. Airway and alveolar nitric oxide measurements in obstructive sleep apnea syndrome. Respir Med 2011; 105: 630–636.

127.

Foresi

Leone

Olivieri

, et al. Alveolar-derived exhaled nitric oxide is reduced in obstructive sleep apnea syndrome. Chest 2007; 132: 860–867.

128.

Hua-Huy

Le-Dong

N-N

Duong-Quy

, et al. Increased alveolar nitric oxide concentration is related to nocturnal oxygen desaturation in obstructive sleep apnoea. Nitric Oxide Biol Chem 2015; 45: 27–34.

129.

Chua

A-P

Aboussouan

Minai

, et al. Long-term continuous positive airway pressure therapy normalizes high exhaled nitric oxide levels in obstructive sleep apnea. J Clin Sleep Med 2013; 09: 529–535.

130.

Liu

, et al. Exhaled nitric oxide from the central airway and alveoli in OSAHS patients: the potential correlations and clinical implications. Sleep Breath Schlaf Atm 2016; 20: 145–154.

131.

Cowan

Allardice

Macfarlane

, et al. Predicting sleep disordered breathing in outpatients with suspected OSA. BMJ Open 2014; 4: e004519.

132.

Bucca

Brussino

Maule

, et al. Clinical and functional prediction of moderate to severe obstructive sleep apnoea. Clin Respir J 2011; 5: 219–226.

133.

Bikov

Hull

Kunos

. Exhaled breath analysis, a simple tool to study the pathophysiology of obstructive sleep apnoea. Sleep Med Rev 2016; 27: 1–8.