Nasal Nitric Oxide Is Dependent on Sinus Obstruction in Allergic Rhinitis

Transcription

1 The Laryngoscope VC 2014 The American Laryngological, Rhinological and Otological Society, Inc. Nasal Nitric Oxide Is Dependent on Sinus in Allergic Rhinitis Hille Suojalehto, MD; Tapio Vehmas, MD, PhD; Irmeli Lindstr om, MD, PhD; David W. Kennedy, MD; Maritta Kilpel ainen, MD, PhD; Tuomas Plosila, MD; Sauli Savukoski, BA; Jukka Sipil a, MD, PhD; Matti Varpula, MD, PhD; Henrik Wolff, MD, PhD; Harri Alenius, PhD; Elina Toskala, MD, PhD Objectives/Hypothesis: The aim of this study was to evaluate the associations between nasal nitric oxide and nasal symptoms, sinus opacification, and markers of allergic inflammation in allergic and in nonallergic rhinitis while taking into account the effect of sinus obstruction. Study Design: We studied 175 young adult subjects divided into three groups: 1) allergic rhinitis, 2) nonallergic rhinitis, and 3) controls. Methods: We measured nasal nitric oxide using the breath-holding method and exhaled nitric oxide and scored semiquantitatively nasal computed tomography scans for opacification and obstruction. We also assessed the visual analogue scores of nasal symptoms, eosinophil count, and interleukin-13 mrna levels in nasal biopsies. Results: The level of nasal nitric oxide correlated with exhaled nitric oxide (r , P <.001). In allergic rhinitis, nasal nitric oxide was elevated when compared to the controls, and an inverse correlation existed between the nasal nitric oxide level and sinus ostial obstruction (r , P 5.013). In nonallergic rhinitis, the level of nasal nitric oxide was similar to that of the controls. In allergic rhinitis, nasal nitric oxide correlated positively with the opacification score (r , P 5.033) and the nasal eosinophil count (r , P 5.030) of patients without a marked sinus ostial obstruction. Conclusions: Sinus ostial obstruction lowers the level of nasal nitric oxide and reduces its value as an indicator of allergic mucosal inflammation. A high nasal nitric oxide level may be a useful marker of eosinophilic inflammation in the nasal cavity and indicate the absence of marked sinus ostial obstruction. Key Words: Nasal nitric oxide, exhaled nitric oxide, computed tomography, eosinophils, interleukin-13, rhinitis. Level of Evidence: 3b. Laryngoscope, 124:E213 E218, 2014 INTRODUCTION Allergic rhinitis (AR) is the most common form of noninfectious rhinitis, affecting approximately 30% of the population. 1 It is associated with immunoglobulin E (IgE)-mediated immune response against allergens and characterized by an infiltration of eosinophils and an increase in Cd41 lymphocytes and Th-2 type cytokines. 2 Interleukin-13 (IL-13) is a Th-2 type cytokine involved Additional Supporting Information may be found in the online version of this article. From the Control of Hypersensitivity Diseases Team (H.S., I.L.); and Occupational Medicine Team (T.V.); and Unit of Systems Toxicology (S.S., H.W., H.A.), Finnish Institute of Occupational Health, Helsinki, Finland; and Department of Otolaryngology Head and Neck Surgery (D.W.K.), University of Pennsylvania, Philadelphia, Pennsylvania, U.S.A; Department of Pulmonary Diseases and Allergology (M.K.); and Department of Otorhinolaryngology Head and Neck Surgery (T.P., J.S.), University of Turku, Finland; Imaging Center (M.V.), Central University Hospital of Turku, Turku, Finland; and the Department of Otolaryngology Head and Neck Surgery (E.T.), Temple University, Philadelphia, Pennsylvania, U.S.A Editor s Note: This Manuscript was accepted for publication January 7, The authors have no funding, financial relationships, or conflicts of interest to disclose. Send correspondence to Hille Suojalehto, MD, Finnish Institute of Occupational Health, Topeliuksenkatu 41 a A, Helsinki, Finland. hille.suojalehto@ttl.fi DOI: /lary in B-cell maturation, differentiation, and isotype switching. Substantial evidence supports the important role of IL-13 in AR. 3 5 Nonallergic rhinitis (NAR) is characterized by chronic nasal symptoms, predominantly nasal congestion and sneezing, without allergic sensitization relevant to symptoms and without signs of infection. It is a heterogeneous condition currently not completely understood. 6 Nitric oxide (NO) is a gas produced naturally in the airways. Exhaled NO (feno) is used as a surrogate of eosinophilic airway inflammation in asthma. 7 Most of the NO production in the airways originates from the nasal region. 8,9 The main production of nasal nitric oxide (nno) is located within the mucosal epithelium of the paranasal sinuses, where it is synthesized by calciumindependent inducible nitric oxide synthase (inos). 10,11 The level of nno is influenced by several airway diseases such as AR, rhinosinusitis and nasal polyps, cystic fibrosis and primary ciliary dyskinesia. 12 Increased levels of inos have been evident in the nasal mucosa of AR patients. 13 Some studies have shown increased levels of nno in AR, 14,15 whereas others have not. 16,17 In a study by Maniscalco et al., subjects with AR showed higher NO levels than controls when sinus diseases were excluded. 18 The NO released from the inflamed nasal mucosa might be increased in AR, but Suojalehto et al.: Nasal NO and Sinus in Rhinitis E213

2 swelling of the nasal mucosa may lead to the obstruction of the sinus ostia, thus reducing the influx of NO from the sinuses to the nasal cavity. 19 Similarly, in nasal polyposis paradoxically low nno levels have been found. 20,21 In the present study we assessed the effect of sinus ostial obstruction on nno in subjects with AR and NAR by using a semiquantitative computed tomography (CT) scoring system. In addition, we evaluated the association between the level of nno and 1) feno, 2) opacification of sinuses, 3) the mucosal eosinophilia, and 4) IL-13 messenger RNA (mrna) level in the nasal mucosa when marked ostial obstruction was excluded. MATERIALS AND METHODS Study Design The study population was selected from a cohort of firstyear university students followed up for 12 years (Supporting Fig. S1). 22 The examinations were performed in two centers, Helsinki and Turku. The study groups were: 1) AR, 2) NAR, and 3) control. The inclusion criteria for the AR group were at least one positive skin prick test (SPT) and relevant rhinitis symptoms to that allergen. In the case of dermographism, sensitization was confirmed with the specific IgE. The NAR group consisted of subjects with periodic or perennial rhinitis symptoms unrelated to allergens. Subjects in the control group had no diagnosed chronic respiratory diseases and no recurrent or persistent respiratory symptoms. Subjects using nasal steroids within 14 days before the examination were excluded from the study. The study was approved by the ethical committee of Turku University and Turku University Central Hospital. All the participants signed informed consent. Clinical Examination Subjects were asked to withhold leukotriene antagonists for 3 days and antihistamines for 7 days before the examination. Participants filled in a questionnaire covering their medical history, respiratory symptoms, medication, smoking history, and a 10-cm visual analogue scale (VAS) of rhinorrhea, nasal congestion, and nasal itchiness within a week. 23 A rhinologist interviewed and examined all subjects. Nasal and Fractional Exhaled NO nno and feno were measured using an online chemiluminescence analyzer equipped with nno software (NIOX; Aerocrine AB, Solna, Sweden) in compliance with American Thoracic Society/ European Respiratory Society recommendations. 24 nno was measured from the nostril while breath was held for 40 seconds with an aspiration flow of 5 ml/s. Nasal Biopsies Nasal biopsies obtained from the anteriosuperior part of the inferior turbinate were fixed in 10% buffered formalin and embedded in paraffin. Sections of 2.5 lm were cut and stained with hematoxylin and eosin and examined under light microscopy (Leica DM LB; Leica Microsystems, Wetzlar, Germany). Eosinophils were counted in three high-power fields at 4003 magnification. Quick-frozen nasal biopsies were kept at 270 C before the extraction of RNA. Total RNA was extracted by using Trisure Reagent (Bioline, Taunton, MA) according to the manufacturer s protocol. The total amount of RNA was measured using the NanoDrop ND-1000 Spectrophotometer (NanoDrop Technologies Inc., Wilmington, DE). mrna expressions of IL-13 were measured as described earlier. 25 CT Scoring On the day of the NO measurements, we obtained coronal CT scans (Helsinki patients) or both coronal and axial scans (Turku patients). They were scored by using the Zinreich methodology, a modification of the Lund-Mackay scoring system. 26 The extent of sinus opacification was the bilateral sum score of the five major sinuses (frontal, maxillary, anterior and posterior ethmoid, and sphenoid), each scored on a six-point scale as follows: 0 for 0%, 1 for 1% to 25%, 2 for 26% to 50%, 3 for 51% to 75%, 4 for 76% to 99%, and 5 for 100% opacification. The maximum sum score was 50. Sinonasal obstructions that could interfere with the nno measurements (the frontal recess, middle meatus, infundibulum, and sphenoethmoid recess) were scored independently. No obstruction was 0, partial/suspected obstruction was 0.5, and total/definitive obstruction was 1. The individual scores were added together to form the sum score, the maximum being 8. In addition to the main drainage pathway from the maxillary sinus (the ostiomeatal complex), we documented the additional (natural or man made) ostia (being present in five subjects) and took this into account while scoring. All of the CT images were scored by a single radiologist (T.V.) blinded to the patient information. To work out the intra- and inter-reader consistencies of scorings, two radiologists (T.V., M.V.) separately scored a subset of CT images (n 5 28). IgE Measurements and SPTs Serum total and specific IgE was measured using the Phadia UniCAP System (Phadia, Uppsala, Sweden). Total IgE <110 ku/l and specific IgE <0.35 ku/l were regarded as normal. The SPT panel included a negative control; a positive control (histamine); and standardized antigens of birch, alder, timothy grass, and mugwort pollen; Alternaria alternate; Aspergillus fumigatus; cat, dog and horse epithelium, and house dust mite Dermatophagoides pteronyssimus (ALK Abello, Nieuwegein, the Netherlands). Results were regarded as positive if the mean wheal diameter was at least 3 mm without a reaction to a negative control (dermographism). E214 Statistical Analysis Continuous variables were expressed as means (6standard deviation) and categorical values as percentages. The differences between the groups were analyzed by using Kruskal-Wallis, analysis of variance, v 2 test, Student t test, or Mann-Whitney U test depending on their distribution. For multiple comparisons, Bonferroni correction was used. The analyses for the studied variables were also rerun while adjusting for sex and smoking. In this case, these covariates and the particular item to be studied were set as independent variables, and group membership (AR, NAR, control) was the dependent variable in a multinomial logistic regression model. The effect of smoking, gender, obstruction, opacification, and nasal polyposis on nno was studied using the general linear model. We also studied the interaction between opacification and obstruction. Spearman correlation between continuous variables was computed. The intra- and inter-reader agreements were computed by using the single measures intraclass correlation (weighted kappa). A P value of <.05 was considered statistically significant. IBM SPSS Suojalehto et al.: Nasal NO and Sinus in Rhinitis

3 TABLE I. Characteristics of Study Subjects. Control, n 5 42 Allergic Rhinitis, n 5 89 Nonallegic Rhinitis, n 5 44 P Value Female, n (%) 25 (59.5) 52 (58.4) 36 (81.8).022 Age, yr, mean (SD) 33.5 (1.8) 32.9 (1.3) 33.2 (1.5) NS,.104 Body mass index, kg/m 2, mean (SD) 23.8 (3.9) 24.3 (3.8) 23.4 (3.6) NS,.443 Smokers, n (%) 5 (11.9) 6 (6.7) 3 (6.8) NS,.560 No. of positive SPTs, n (%), n <.001* 1 3 positive SPTs 2 (4.9) 30 (34.9) 8 (19.0) At least 4 positive SPTs 0 (0.0) 56 (65.1) 0 (0.0) At least 1 positive SPT to 2 (4.9) 67 (77.9) 2 (4.8) <.001 perennial allergen, n Total IgE, ku/l, mean (SD) 31.2 (39.6) (346.0) 42.0 (65.9) <.001 *P value between the groups in subjects having one or more positive SPTs. IgE 5 immunoglobulin E; NS 5 nonsignificant; SD 5 standard deviation; SPT 5skin prick test. Statistics for Windows version 20.0 (IBM Corp., Armonk, NY) software was used. RESULTS Study Population A total of 175 subjects were enrolled in the study groups in 2008 to 2009; 95 subjects were examined in Helsinki and 80 in Turku (Supporting Fig. S1). Altogether, 19 subjects reported having symptoms suggestive of respiratory infection (mainly flu-like symptoms) within the preceding 2 weeks. The characteristics of the study subjects are presented in Table I. The mean age of the participants was 33 years; 64% of them were women. Altogether, 38 subjects in the AR group and six subjects in NAR group had concomitant asthma. Symptoms, Eosinophils, and IL-13 mrna in the Nasal Mucosa The VAS score of nasal itching was significantly higher in the AR group than among the controls (Table II). In the NAR group, the rhinorrhea and nasal congestion scores were also elevated. The three groups did not TABLE II. Nasal Symptoms and Markers of Allergic Inflammation, Computed Tomography Findings, and Nitric Oxide Levels Among Subjects With Allergic and Nonallergic Rhinitis and Among Controls. Control, n 5 42 Allergic Rhinitis, n 5 89 Nonallergic Rhinitis, n 5 44 P Value Crude P 1 Value* P 2 Value* P 3 Value* P Value Adjusted For Sex and Smoking VAS rhinorrhea, mm, n VAS nasal congestion, mm, n VAS nasal itch, mm, n Nasal eosinophil count, n Nasal IL-13 mrna, RU, n CT opacification score, n CT obstruction score, n Nasal nitric oxide, ppb, n Exhaled nitric oxide, ppb, n (20.2) 26.4 (25.1) 31.7 (25.7).029 NS (.176).026 NS (.729) (20.3) 22.8 (24.1) 33.2 (28.8).003 NS (.261).002 NS (.076) (7.8) 25.0 (25.5) 19.9 (23.0) <.001 < NS (.642) < (4.0) 2.0 (3.7) 1.5 (2.9) NS (.773) NS (1.000) NS (1.000) NS (1.000) NS (.876) 43.1 (46.6) 95.9 (157.0) (412.4).035 NS (.144) NS (1.000) NS (1.000) NS (.123) 3.9 (5.0) 2.7 (3.6) 2.9 (4.5) NS (.253) NS (.270) NS (.987) NS (1.000) NS (.209) 0.8 (1.6) 0.6 (1.3) 0.7 (1.6) NS (.598) NS (1.000) NS (1.000) NS (1.000) NS (.493) (191.6) (276.0) (288.2) NS (1.000) NS (.122) (16.6) 19.5 (12.8) 13.2 (6.1).012 NS (.129) NS (1.000) Data are presented as mean (standard deviation). P value crude and P value adjusted present a comparison among all of the study groups. P 1 value is the crude P value between control and allergic rhinitis groups. P 2 value is the crude P value between control and nonallergic rhinitis groups. P 3 value is the crude P value between allergic rhinitis and nonallergic rhinitis groups. *Bonferroni correction. CT 5computed tomography; IL-13 5 interleukin-13; mrna 5 messenger RNA; NS 5 statistically nonsignificant (P.05); ppb 5 parts per billion; RU 5 relative unit; VAS 5 visual analogue scale. Suojalehto et al.: Nasal NO and Sinus in Rhinitis E215

4 Association Between nno and CT Findings In Figure 1, nno is plotted on the total obstruction score for subjects with AR. nno seems to decrease when the obstruction score exceeds 2. A significant negative correlation between nno and total obstruction was seen in the AR group (r , P 5.013), but not among the controls (r , P 5.534) or subjects with NAR (r , P 5.989). Fig. 1. Nasal nitric oxide (nno) plotted on total obstruction score for subjects with allergic rhinitis. ppb 5 parts per billion. differ in terms of nasal eosinophils or IL-13 mrna. We did not detect significant differences in symptom scores, eosinophils, or IL-13 mrna between the subgroups of asthmatics and nonasthmatics in the AR group (data not shown). CT Findings Partial or total obstruction of frontal recesses was detected in 27 subjects (15.3%), within the middle meatus in 11 (6.3%), within the infundibula in 29 (22.2%), and within the sphenoethmoidal recess in 20 (11.4%). The mean obstruction score was 0.7, and the mean opacification score was 3.0. The scores did not differ between the study groups (Table II) or between the subgroups of asthmatics and nonasthmatics in the AR group (data not shown). We detected a significant positive correlation between the opacification and obstruction (r , P <.001). Visible fluid in at least one nasal cavity was detected in five subjects (12.5%) in the control group, in one (1.1%) in the AR group, and in one (2.3%) in the NAR group. The intra-reader agreement (weighted kappa) for sinus opacification was and for sinus obstruction 0.897, whereas the respective interreader values were and 0.846, all being excellent. Nasal Nitric Oxide Compared to the controls, the level of nno and feno was increased in AR, but not in NAR (Table II). When the subgroups of asthmatics and nonasthmatics in AR group were compared, we did not find a difference in the level of nno (P 5.826). The level of feno was significantly higher in asthmatics (P 5.031). The level of nno correlated with feno (r , P <.001) in the study population. In addition, the level of nno was negatively associated with obstruction (P 5.005). The effects of smoking (P 5.343), sex (P 5.075), fluid in the sinuses (P 5.830), or opacification (P 5.467) on the level of nno were not significant. E216 nno in Subjects Without Marked Ostial In Figure 1, we selected the obstruction score 2 as a cutoff for marked ostial obstruction. In subjects with an obstruction score of <2, significant differences between the study groups were detected in nno (P 5.003), in exno (P 5.018), and in IL-13 mrna (P 5.043) (Table III). In AR subjects with an obstruction score of <2, a significant positive correlation was seen between nno and feno, between nno and opacification, and between nno and nasal eosinophils (Table IV). In addition, a positive correlation between nno and feno was detected in NAR and control groups with an obstruction score <2. On the other hand, no significant positive correlation was detected between nno and IL-13 mrna or between nno and VAS scores of nasal symptoms (data not shown). We did not detect a significant difference in any of the presented variables on the basis of reported respiratory infection symptoms (data not shown). The subjects with fluid in nasal sinuses (n 5 8) had an elevated opacification (P 5.003) and obstruction (P <.001) score, but did not differ significantly in terms of other evaluated parameters from those without nasal fluid (data not presented). DISCUSSION We found that the level of nno was increased in subjects with AR; in NAR no difference was detected from controls. We detected no difference in the CT scoring of nasal obstruction or opacification between the AR, NAR, and control groups. nno inversely correlated with sinus ostial obstruction in AR. Analysis of AR patients without marked ostial obstruction revealed positive correlations between nno and sinus opacification and nasal eosinophil count. The Zinreich classification was used, because it is more sensitive to changes in the sinus mucosa than the Lund-Mackay score. 26,27 In this system, both obstruction and opacification are separately classified, and we found a strong correlation between these two scores. Also, partial or suspected ostial obstruction in CT scans was noted to achieve a precise scoring. Inaccuracies in the obstruction score results would most likely cause random error and dilute the effects found, but not bias the study results. Early mucosal swelling causes radiological opacification and ostial obstruction, which leads to the accumulation of fluid, thus causing further opacification. By using the Zinreich classification, the levels of opacification or obstruction scores were low, and they did not differ significantly Suojalehto et al.: Nasal NO and Sinus in Rhinitis

5 TABLE III. Nasal and Exhaled Nitric Oxide, Computed Tomography-Based Opacification Score, and Eosinophil Count and Interleukin-13 mrna Levels in Nasal Biopsies in Subjects With Allergic Rhinitis, Nonallergic Rhinitis, and in the Control Group. Control Allergic Rhinitis Nonallergic Rhinitis (n 5 32) (n 5 9) (n 5 79) (n 5 10) (n 5 37) (n 5 6) Nasal nitric oxide, ppb (187.7) (214.7) (269.9) (277.7) (241.7) (576.5) Exhaled nitric oxide, ppb 14.1 (18.3) 16.5 (10.0) 19.9 (13.4) 17.1 (6.3) 13.0 (6.4) 12.9 (4.6) Opacification score 1.6 (1.7) 12.1 (4.4) 2.0 (2.1) 8.1 (6.0) 1.4 (1.9) 12.2 (4.6) Nasal eosinophil count 1.5 (4.3) 3.2 (3.6) 1.9 (3.8) 3.0 (2.7) 1.4 (2.9) 2.4 (2.7) Nasal IL-13 mrna, RU 40.7 (39.6) 54.4 (76.2) 92.1 (161.8) (99.7) 40.8 (52.6) 39.9 (44.7) Data are presented as mean (standard deviation). Subjects are subdivided into two groups based on the sinus ostial obstruction score. IL-13 5 interleukin-13; ppb 5 parts per billion; mrna 5 messenger RNA, RU 5 relative unit. between the groups. Previous studies also found no difference in opacification between AR and NAR 28 or in Lund- Mackay score of opacification between intermittent and perennial AR and controls. 29 Limited mucosal changes detected in CT scans should therefore not be regarded as indicative of allergy, even though mucosal thickening is found among some AR patients. 30,31 nno was slightly elevated in AR, and this effect was relatively robust for covariate (sex and smoking) adjustment. This finding is in line with some former studies, 14,15 but not all. 16,17 The mean nno levels of control and NAR groups were within the suggested normal range ( parts per billion), whereas the level of the AR group was slightly above it. 32 It has been proposed that the breath-holding method 33 we used collects NO mainly from the mucosa of the nasal cavity, whereas the humming method mostly reflects the sinus contribution. 18,34 By using the humming method we could have detected more distinct reduction of the nno level in subjects with ostial obstruction. The study of Colantonio et al. showed significantly lower NO levels in subjects with nasal polyposis than those with uncomplicated AR when the breath-holding method was used. 20 In other studies, nasal polyposis or sinus obstruction did not affect the nno levels during a quiet exhalation, but the humming increase in nno was abolished. 18,35 Also, the ostiomeatal complex patency correlated with the increased nno level in quiet exhalation with normal controls but not in AR. 29 In our study, sinus ostial obstruction was negatively associated with the level of nno. Nevertheless, in subjects with marked ostial obstruction, the mean level of nno (862.5 parts per billion) was within suggested normal range. This finding supports the view that extremely low levels of nno found in patients with primary ciliary dyskinesia are not solely explained by sinus ostial obstruction. 9 In the present study, the nno level positively correlated with the Zinreich opacification score in subjects with AR who did not have marked sinus obstruction. In the former studies, the extent of opacification scored with the Lund-Mackay system inversely correlated with nno in subjects with polyposis 36 and also in a group of AR and nonrhinitic subjects without severe rhinosinusitis. 29 This discrepancy may be due to the interaction between the obstruction and opacification score in these studies. In addition, the Zinreich scoring is more sensitive to mild changes in mucosal opacification. 27 We found no increase in nasal eosinophils or in IL- 13 mrna in AR, which may be related to low allergen exposure and symptom scores. In the absence of allergen exposure, no increase in inflammatory cells or markers of eosinophilic activation has been detected in seasonal TABLE IV. Correlation of Nasal Nitric Oxide With Exhaled Nitric Oxide, Opacification Score, and Eosinophil Count and IL-13 mrna Level in Nasal Biopsies. Control Allergic Rhinitis Nonallergic Rhinitis n 5 32 n 5 9 n 5 79 n 5 10 n 5 37 n 5 6 Exhaled nitric oxide 0.489* Opacification score Nasal eosinophil count Nasal IL-13 mrna Results are shown as correlation coefficient (r). *P <.01. P <.05. IL-13 5 interleukin-13; mrna 5 messenger RNA. Suojalehto et al.: Nasal NO and Sinus in Rhinitis E217

6 AR. 37,38 We found a correlation with nno and feno as reported earlier. 39 In asthma, an association exists between sputum eosinophils and feno. 40 However, to our knowledge, there is little information available regarding the correlation of nno and nasal mucosal eosinophils and Th2 cytokines. In the present study, a correlation between nasal eosinophils and nno was detected in AR when subjects with sinus obstruction were excluded. In a former study, nno and nasal smear eosinophils decreased simultaneously in AR when levocetirizine was introduced. 41 In our sample, nno and IL-13mRNA level did not correlate. This is in line with the study of Truyen et al., who found no correlation between feno and IL-13 in induced sputum among asthmatics and healthy controls. 42 CONCLUSION We found a slight elevation nno, a clear correlation between feno and nno, and an inverse correlation between the nno and the CT score of sinus ostial obstruction in AR. A significant positive correlation was revealed between nno and opacification and the nasal eosinophil count in AR patients without marked ostial obstruction. Our findings support the view that sinus ostial obstruction affects the level of nno in AR when the breath-holding method is used. This phenomenon reduces the value of nno as an indicator of allergic inflammation. However, a high nno level may be a useful marker of eosinophilic inflammation in the nasal cavity and indicate the absence of marked sinus ostial obstruction. BIBLIOGRAPHY 1. Jarvis D, Newson R, Lotvall J. Asthma in adults and its association with chronic rhinosinusitis: the GA2LEN survey in Europe. Allergy 2012;67: Bousquet J, Khaltaev N, Cruz AA, et al. Allergic Rhinitis and its Impact on Asthma (ARIA) 2008 update (in collaboration with the World Health Organization, GA(2)LEN and AllerGen). Allergy 2008;63(suppl 86): Ghaffar O, Laberge S, Jacobson MR, et al. IL-13 mrna and immunoreactivity in allergen-induced rhinitis: comparison with IL-4 expression and modulation by topical glucocorticoid therapy. Am J Respir Cell Mol Biol 1997;17: Erin EM, Leaker BR, Zacharasiewicz AS, et al. Single dose topical corticosteroid inhibits IL-5 and IL-13 in nasal lavage following grass pollen challenge. Allergy 2005;60: Miyahara S, Miyahara N, Matsubara S, Takeda K, Koya T, Gelfand EW. IL-13 is essential to the late-phase response in allergic rhinitis. J Allergy Clin Immunol 2006;118: Bousquet J, Fokkens W, Burney P, et al. Important research questions in allergy and related diseases: nonallergic rhinitis: a GA2LEN paper. Allergy 2008;63: Dweik RA, Boggs PB, Erzurum SC, 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: Alving K, Weitzberg E, Lundberg JM. Increased amount of nitric oxide in exhaled air of asthmatics. Eur Respir J 1993;6: Lundberg JO, Weitzberg E, Nordvall SL, et al. Primarily nasal origin of exhaled nitric oxide and absence in Kartagener s syndrome. Eur Respir J 1994;7: Lundberg JO, Rinder J, Weitzberg E, et al. Nasally exhaled nitric oxide in humans originates mainly in the paranasal sinuses. Acta Physiol Scand 1994;152: Lundberg JO, Farkas-Szallasi T, Weitzberg E. High nitric oxide production in human paranasal sinuses. Nat Med 1995;1: Sahin G, Klimek L, Mullol J, et al. Nitric oxide: a promising methodological approach in airway diseases. Int Arch Allergy Immunol 2011;156: Kawamoto H, Takeno S, Yajin K. Increased expression of inducible nitric oxide synthase in nasal epithelial cells in patients with allergic rhinitis. Laryngoscope 1999;109: Kharitonov SA, Rajakulasingam K, O Connor B, Durham SR, Barnes PJ. Nasal nitric oxide is increased in patients with asthma and allergic rhinitis and may be modulated by nasal glucocorticoids. J Allergy Clin Immunol 1997;99: Arnal JF, Didier A, Rami J, et al. Nasal nitric oxide is increased in allergic rhinitis. Clin Exp Allergy 1997;27: Palm JP, Alving K, Lundberg JO. Characterization of airway nitric oxide in allergic rhinitis: the effect of intranasal administration of L-NAME. Allergy 2003;58: Henriksen AH, Sue-Chu M, Holmen TL, Langhammer A, Bjermer L. Exhaled and nasal NO levels in allergic rhinitis: relation to sensitization, pollen season and bronchial hyperresponsiveness. Eur Respir J 1999;13: Maniscalco M, Sofia M, Weitzberg E, et al. Humming-induced release of nasal nitric oxide for assessment of sinus obstruction in allergic rhinitis: pilot study. Eur J Clin Invest 2004;34: Maniscalco M, Lundberg JO. Nasal nitric oxide. In: Horvath I, de Jongste JC. European Respiratory Monograph. Vol. 49. Lausanne, Switzerland: European Respiratory Society; 2010: Colantonio D, Brouillette L, Parikh A, Scadding GK. Paradoxical low nasal nitric oxide in nasal polyposis. Clin Exp Allergy 2002;32: Alobid I, Benitez P, Valero A, Munoz R, Langdon C, Mullol J. Oral and intranasal steroid treatments improve nasal patency and paradoxically increase nasal nitric oxide in patients with severe nasal polyposis. Rhinology 2012;50: Suojalehto H, Toskala E, Kilpelainen M, et al. MicroRNA profiles in nasal mucosa of patients with allergic and nonallergic rhinitis and asthma. Int Forum Allergy Rhinol 2013;3: Spector SL, Nicklas RA, Chapman JA, et al. Symptom severity assessment of allergic rhinitis: part 1. Ann Allergy Asthma Immunol 2003;91: ATS/ERS recommendations for standardized procedures for the online and offline measurement of exhaled lower respiratory nitric oxide and nasal nitric oxide, Am J Respir Crit Care Med 2005;171: Kankkunen P, Rintahaka J, Aalto A, et al. Trichothecene mycotoxins activate inflammatory response in human macrophages. J Immunol 2009; 182: Kennedy DW, Kuhn FA, Hamilos DL, et al. Treatment of chronic rhinosinusitis with high-dose oral terbinafine: a double blind, placebocontrolled study. Laryngoscope 2005;115: Meltzer EO, Hamilos DL, Hadley JA, et al. Rhinosinusitis: developing guidance for clinical trials. J Allergy Clin Immunol 2006;118: Ono N, Kase K, Homma H, Kusunoki T, Ikeda K. Maxillary sinus infundibulum narrowing influences sinus abnormalities in spite of the presence or absence of allergy. Acta Otolaryngol 2011;131: Shusterman DJ, Weaver EM, Goldberg AN, Schick SF, Wong HH, Balmes JR. Pilot evaluation of the nasal nitric oxide response to humming as an index of osteomeatal patency. Am J Rhinol Allergy 2012;26: Naclerio RM, detineo ML, Baroody FM. Ragweed allergic rhinitis and the paranasal sinuses. A computed tomographic study. Arch Otolaryngol Head Neck Surg 1997;123: Piette V, Bousquet C, Kvedariene V, et al. Sinus CT scans and mediator release in nasal secretions after nasal challenge with cypress pollens. Allergy 2004;59: Scadding G, Scadding GK. Update on the use of nitric oxide as a noninvasive measure of airways inflammation. Rhinology 2009;47: Struben VM, Wieringa MH, Mantingh CJ, de Jongste JC, Feenstra L. Nasal NO measurement by direct sampling from the nose during breathhold: aspiration flow, nasal resistance and reproducibility. Eur Arch Otorhinolaryngol 2006;263: Weitzberg E, Lundberg JO. Humming greatly increases nasal nitric oxide. Am J Respir Crit Care Med 2002;166: Lundberg JO, Maniscalco M, Sofia M, Lundblad L, Weitzberg E. Humming, nitric oxide, and paranasal sinus obstruction. JAMA 2003;289: Delclaux C, Malinvaud D, Chevalier-Bidaud B, Callens E, Mahut B, Bonfils P. Nitric oxide evaluation in upper and lower respiratory tracts in nasal polyposis. Clin Exp Allergy 2008;38: Juliusson S, Pipkorn U, Karlsson G, Enerback L. Mast cells and eosinophils in the allergic mucosal response to allergen challenge: changes in distribution and signs of activation in relation to symptoms. J Allergy Clin Immunol 1992;90: Milanese M, Ricca V, Canonica GW, Ciprandi G. Eosinophils, specific hyperreactivity and occurrence of late phase reaction in allergic rhinitis. Eur Ann Allergy Clin Immunol 2005;37: Williamson PA, Vaidyanathan S, Clearie K, Stewart M, Lipworth BJ. Relationship between fractional exhaled nitric oxide and nasal nitric oxide in airways disease. Ann Allergy Asthma Immunol 2010;105: Berry MA, Shaw DE, Green RH, Brightling CE, Wardlaw AJ, Pavord ID. The use of exhaled nitric oxide concentration to identify eosinophilic airway inflammation: an observational study in adults with asthma. Clin Exp Allergy 2005;35: Bautista AP, Eisenlohr CP, Lanz MJ. Nasal nitric oxide and nasal eosinophils decrease with levocetirizine in subjects with perennial allergic rhinitis. Am J Rhinol Allergy 2011;25: Truyen E, Coteur L, Dilissen E, et al. Evaluation of airway inflammation by quantitative Th1/Th2 cytokine mrna measurement in sputum of asthma patients. Thorax 2006;61: E218 Suojalehto et al.: Nasal NO and Sinus in Rhinitis