Airway Inflammation And Hyperresponsiveness In Patients With Rhinitis Without Asthma

Background and ObjectiveAllergic rhinitis (AR) is the chronic inflammatory disease of nasal mucus triggered by IgE antibody-mediated release of inflammatory mediators (mainly histamine) by mast cells under comorbid actions of multiple immuno-active cells and cytokines after contact with allergens in susceptible individuals. Categorized as airway allergic diseases, both allergic rhinitis and asthma share close correlations. In 1997, Grossman et al first proposed the concept of'One airway, one disease', which emphasized the integraty of respiratory inflammatory diseases. In 2001, World Health Organization published a guideline document entitled'allergic rhinitits and its impact on asthma'(ARIA), in which allergic rhinitis was stated as one of the major factor contributing to asthma. Furthermore, a 2008 update of ARIA stated the principle more explicitly that allergic rhinitis and asthma are both manifestation of a syndrome in two separate airway positions, which indicated the significance of integration of both diseases. Both allergic rhinitis and asthma share numerous similarities in immunopathological pathogenesis and are characterized of airway eosinophil infiltration. Induced sputum cytology test is currently a mature objective approach for measurement of lower airway eosinophilic inflammation, while nasal lavage cytology test could well represent upper airway inflammation, which lacks unified standard for evaluation as well as positive threshold. Its limitation in clinical applications has led to the failure of being a routine method for clinical diagnosis of upper airway inflammatory diseases and follow-up visits.With a growing trend in worldwide prevalence rate, allergic rhinitis has exerted its impact on the quality of life, loaded huge social economic burden and has become a global health issue. To date, most international or national epidemiological data were obtained mainly through questionnaire surveys or telephone investigations, judgments were made only according to subject's own perceptions, and there is a lack of objective and comparable parameter for the measurements. From the perspective of symptom, various similarities are present in allergic and non-allergic rhinitis, which made distinction demanding solely on symptoms. The difficulty in determining the proportion of AR and non-allergic rhinitis (NAR) in these self-reported subjects would lead to adverse impact on the accuracy and scientificity of investigation data on prevalence of allergic-rhinitis, for which further studies need to be conducted. Moreover, few studies were available concerning the prevalence of rhinitis and asthma in China.Furthermore, although close correlation between AR and NAR has been suggested by more and more recent studies, there's a lack of large scale comparison on the characters of lower airway inflammation and hyperresponsiveness in subjects with simple AR or NAR. Clinical issues, for instance, whether NAR shares identical risk factors for development of disease with bronchial asthma, or could the risk of lower airway hyperresponsiveness be predicted through assessment of systemic and lower airway allergic inflammation prior to existence of lower airway symptoms in subjects with rhinitis are yet to be solved. No report was available around the world, therefore more objective, scientific and prudent clinical trials are to be carried out. In this study, the distribution character of cell components was analyzed systematically, the approach for assessment of cell differentiation and related normal reference values were established, and clinical significance in assisting the diagnosis of AR was evaluated through collection and processing of nasal lavage fluid in a large scale of normal individuals and subjects with AR. The prevalence of rhinitis in college students of Guangdong could be determined, and the proportion of AR and NAR as well as the correlation with occurrence of asthma in subjects with self-reported rhinitis were assessed thourgh sampling survey on rhinitis and its related disorders in a sampled college in combination with otolaryngopharyngologic examinations as well as measurements of allergens in clinics. In the last section, the inflammatory characters of subejcts without lower airway symptoms who had AR or NAR and its correlation with lower airway inflammation and hyperresponsiveness were determined through blood routine test, nasal lavage test, induced sputum test, measurement of exhaled nitric oxide and bronchial provocation test in subejcts with AR, NAR as well as normal individuals. The intrinsic correlation between upper airway inflammation, lower airway inflammation and hyperreposniveness was also explored. The parameters of peripheral blood or upper/ lower airway allergic inflammation were identified for prediction of the risk of lower airway hyperesponsiveness prior to occurrence of lower airway symptoms in subjects with rhinits, thereby offering scientific proves for clinical monitor, early intervention and control of rhinitis and asthma. ObjectivesTo establish the methodology for nasal lavage cytology differential test and the normal reference range, and assess the clinical significance in assissting diagnosis of allergic rhinitis.MethodsNormal healthy students in a college of Guangzhou and adult volunteers recruited in The Physical Examination Center, The First Affiliated Hospital of Guangzhou Medical College from January 2009 to January 2011 were enrolled in the study as normal subjects. No abnormality in physical examination, blood routine test, chest X-ray roentgenography or spirometric test was revealed, and allergen skin prick test proved negative in normal controls. Subjects with typical symptoms and signs of allergic rhinitis were recruited in allergic rhinitis group according to the diagnostic criteria of ARIA 2008, who should had no use of oral or intranasal corticosteroid or anti-histamine. All subjects had neither history of chronic cough, wheezing or other systemic disease, nor upper respiratory tract infection for the past 8 weeks, and had neither the history of nasal or facial injury nor smoking. No intranasal septum deviation was revealed by nasal speculum examination. All female subjects must not be within pregnancy or lactation period. The protocol was approved by the Ethics Committee of The First Affiliated Hospital of Guangzhou Medical College. The items of the study were interpreted to all subjects, and informed written consent was given prior to the study. Nasal lavage procedures were performed in both groups of subjects. Inflammatory cells and their percentages in the sediments of nasal lavage fluid were calculated under microscopic vision, with the standard of whether epithelial cells or inflammatory cells were visible under 200×microscopic vision for judgment of quality of test. Neutrophils, macrophages, eosinophilss and lymphocytes were counted in 20 non-repeated fields under 200×microscopic vision, with the averages being the cell counts of inflammatory cells. Percentage of neutrophils was calculated using the formula: sum of neutrophils in 20 fields/ sum of inflammatory cells in 20 fields×100%, and percentages of macrophages, eosinophilss and lymphocytes were deduced following the identical approach. Distinction of the two measurements in normal control group and allergic rhinitis group was compared in order to establish the optimal parameter for judgment of positive outcomes of nasal lavage test. The 95% upper reference limit of inflammatory cells was calculated for normal individuals. Receiver operation characteristic curve (ROCC) was depicted adopting non-parametric approach, and the threshold was determined according to the maximal Youden's index. The corresponding sensitive and specificity were calculated for evaluation of the significance of clinical diagnosis.Results1. We enrolled 162 healthy volunteers and 184 subjects with allergic rhinitis. No statistical difference (P>0.05) was revealed in constitutional proportion of gender, age, height, weight and body mass index (BMI), which indicated comparability between two groups.2. According to the existence of epithelial cells or inflammatory cells visible under 200×microscopic vision as successful standard of test, the successful rate of nasal lavage test was 98.77% and 98.91% in normal control group and allergic rhinitis group, with no significant difference (P>0.05) being found between two groups.3. Besides epithelial cells, inflammatory cells were visible in nasal lavage fluid among 53 subjects (33.12%) in normal control group and 155 subjects in AR group (85.16%), with significant statistical difference (P<0.01) between both groups. No statistical difference (P>0.05) was shown in the percentage of each inflammatory cell between the two groups.4. The neutrophil and eosinophil count was 1.74±9.37 /×200 and 0.76±3.46 /×200 (HE stain) in normal control group, with significant statistical diffenrece (P<0.01) compared with that of AR group [13.61±46.93/×200 and 27.88±62.47 /×200 (HE stain)], respectively. The 95% upper reference limit of inflammatory cells in nasal lavage test was 10.03/×200 for neutrophils and 3.47/×200 for eosinophils (HE stain) in healthy volunteers, respectively.5. The clinical diagnostic criteria were set as the golden standard for allergic rhinitis, and ROC curves were depicted based on the positive standard using neutrophil or eosinophil counts in nasal lavage fluid, with the results revealing eosinophil count to be the optimal positive criterion (area under curve: 0.86, P<0.01) . The optimal threshold of eosinophil count in nasal lavage fluid for allergic rhinitis was 0.350/×200 according to ROC curve, with the sensitivity, specificity, Youden's index, positive likelihood ratio and negative likelihood ratio of 0.76, 0.89, 0.66, 7.21 and 0.14, respectively.Conclusions1. Nasal lavage inflammatory cell absolute count is capable of offering clear discrimination between normal individuals and subjects with allergic rhinitis, and may serve as a surrogate assessment toll for clinical diagnosis.2. Eosinophils count is the optimal positive parameter for judgment of the results of nasal lavage test, with the 95% upper reference limit of 3.47 /×200 (HE stain). ObjectivesTo determine the prevalence rate of rhinitis in college students and assess the proportion of allergic rhinitis and non-allergic rhinitis in subjects with self-reported rhinitis as well as its correlation with occurrence of asthma.MethodsA sampling survey was performed in a college of Guangdong using'Questionnaire on rhinitis and its complications in college students of Guangdong region', with the items including epidemiological characters, family history, rhinitis-associated symptoms, asthma-associated symptoms, cough symptoms and its character, ocular region-associated symptoms and dermal symptoms, etc.. A one-to-one questionnaire survey using uniform method was conducted in all subjects enrolled in the study by investigators who underwent uniform training course. Subjects with any symptom of nasal secretion, sneezing, nasal congestion or itchy nose, or had susceptible history of rhinitis as well as the healthy volunteers without discomforts were primarily screened. The clinical diagnoses were made according to the skin prick test result using 11 common inhaled allergens (ALK Co. Ltd, Denmark) and measurement of serum specific IgE (sIgE) of inhaled allergens (Pharmacia kits, Phadiatop Co. Ltd, Switzerland), therefore 4 groups (AR, NAR, atopy and normal control group) were divided. The diagnosis, classification and severity of AR took reference on the guideline of ARIA (version 2008). Data of all questionnaires were defined according to the variables in uniform database structure, and were input into Excel TM electronic tables, with at least 2 personnels performing corss-checks. SPSS 13.0 was adopted for data reorganization and analysis. And chi-square test was applied for comparison of rates. Results1. 2362 questionnaires were delivered to the subjects, with 2339 valid questionnaires and the response rate of 99.03%. The prevalence rate of rhinitis was 19.20% in college students of Guangdong, with no statistical difference between genders. Among all individuals in normal control group, 1357 subejcts (58.02%) had no symptoms of allergic disorders, 534 subjects had atopic dermatitis (22.83%), 201 subjects had allergic conjunctivitis (8.59%), 85 subjects had sinusitis (3.63%), and 35 subejcts had asthma (1.50%).2. The prevalence rate of rhinitis in college students of Guangdong region was 19.20%, with no statistical difference between genders.The average span of rhinitis was (4.48±3.64) years,with the minimum and maximum of 0.5 year and 20 years. There were 75 (16.70%), 250 (55.68%) and 124 (27.62%) subjects without episode, with intermittent episodes and continuous episodes of rhinitis in the last year, respectively. 149 (33.18%) and 300 (66.82%) subjects had mild and moderate-to-severe rhinitis.3. The major symptoms followed the sequence of sneezing [287 cases (63.92%)], running clear secretion [264 cases (58.80%)], nasal congestion [239 cases (53.23%)] and itchy nose [205 cases (45.66%)] in subjects with rhinitis.4. The major pattern of rhinitis episode followed the sequence of seasonal alternative [131 cases (29.18%)], in winter [109 cases (24.28%)], perennial continuous [62 cases (13.68%)], in spring [48 cases (10.69%)], in summer [33 cases (7.35%)], in autumn [24 cases (5.35%)] and irregular [70 cases (15.59%)].5. The major triggering factors of rhinitis followed the sequence of cold air [274 cases (61.02%)], dust contact [180 cases (40.09%)], stimulus gas contact [144 cases (32.07%)], contact with cat and dog or animals with furs [50 cases (11.14%)], flower or grass contact [43 cases (9.58%)], ingestion of food [36 cases (8.02%)], exercise [29 cases (6.46%)], contact with penicillin [4 cases (0.89%)], contact with aspirin [1 case (0.22%)], and other causes [60 cases in total (13.36%)], which included climate changes, air pollution, in the morning or prior to sleeping, contact with fragrance or cosmetics, and second-hand smoking, etc.6. There were 221 subjects (49.22%) with rhinitis reported family history of allergic disorders, with the incidence rates following the sequence of allergic rhinitis [132 cases (29.40%)], food allergy [70 cases (15.59%)], asthma [26 cases (5.79%)], atopic dermatitis [21 cases (4.68%)] and allergic conjunctivitis [14 cases (3.12%)].7. Common complications of rhinitis followed the sequence of atopic dermatitis [138 cases (30.73%)], allergic conjunctivitis [79 cases (17.59%)], sinusitis [43 cases (9.58%)], asthma [32 cases (7.13%)] and chronic cough [8 cases (1.78%)].8. Among 250 subjects with symptoms of rhinitis, allergen skin prick test and measurement of sIgE proved positive in 136 subejcts (54.40%) and was negative in 114 subjects (45.60%). Among 168 normal individuals without symptoms of allergic disorders, allergen skin prick test and measurement of sIgE proved positive in 27 subejcts (16.07%) and was negative in 114 subjects (83.93%).9. The three most common allergens in rhinitis groups were Dermatophagoides pterynyssinus, Dermatophagoides farinae and Dermatophagoides tropicalis, respectively. Significant statistical difference (P<0.05) was shown in the sort of allergens and the extent of reaction in normal individuals compared with that of rhinitis groups.10. The prevalence rate of asthma was 22.52%, 6.54%, 8.00% and 0.71% in AR, NAR, atopy and normal control group, respectively. Significant statistical difference (P<0.01) was revealed in the prevalence rate between AR group and normal control group. Statistical difference (P<0.05) was also shown between NAR group and normal control group.Conclusions1. The prevalence rate of rhinitis reaches to 19.20%, with a similar proportion of non-allergic rhinitis and allergic rhinitis.2. An increase in prevalence rate of asthma in subejcts with AR and NAR has been revealed, both of which are the predisposing factors of onset of asthma. ObjectivesTo determine the inflammatory character of allergic rhinitis and non-allergic rhinitis in subjects without lower airway symptoms and its correlation with lower airway inflammation and airway hyperresponsiveness.MethodsAll subjects were recruited from the outpatient clinics of otolaryngopharyngology department, The First Affiliated Hospital of Guangzhou Medical College from January 2009 to January 2011. Subjects with typical symptoms and signs of allergic rhinitis as well as positive allergen skin prick test result were allocated in allergic rhinitis group (AR) according to the diagnostic criteria of ARIA 2008, while subjects with typical symptoms and signs of allergic rhinitis as well as negative allergen skin prick test result were allocated in non-allergic rhinitis group. All subjects enrolled in the study should have no use of oral or intranasal corticosteroid or anti-histamine for the past 4 weeks. Normal healthy students in a college of Guangzhou and adult volunteers recruited in The Physical Examination Center, The First Affiliated Hospital of Guangzhou Medical College were allocated in the normal control group. No abnormality in physical examination, blood routine test, chest X-ray roentgenography or spirometric test was revealed, and allergen skin prick test proved negative in normal controls. All subjects enrolled had neither history of chronic cough, wheezing or other systemic disease, nor upper respiratory tract infection for the past 8 weeks, and had neither the history of nasal or facial injury nor smoking. No intranasal septum deviation was revealed by nasal speculum examination. All female subjects must not be within pregnancy or lactation period. The protocol was approved by the Ethics Committee of The First Affiliated Hospital of Guangzhou Medical College. The items of the study were interpreted to all subjects, and informed written consent was given prior to the study.Peripheral five-classify blood examinations (eosinophil count >0.30×109/L as positive criterion), measurements of exhaled nitric oxide concentration (FENO> 25ppb as postive criterion), nasal lavage fluid differentiation cytology tests (eosinophil count >3.47 /×200 as positive criterion), spirometry tests, methacholine bronchial provocation tests (a≥15% and≥20% fall in FEV1 as susceptive positive and positive criterion, respectively) and induced sputum differential cytology tests (eosinophil percentage >2.5% as positive criterion) were performed. The character and difference in peripheral blood cells, systemic, upper and lower airway inflammation as well as airway hyperresponsiveness among three groups were compared. T-test, one-way analysis of variance (ANOVA) and chi-square continuous correction test was adopted for comparison between two groups, among multiple groups, and comparison on rates, respectively. Multiple variance Logistic regression was applied to determine the predisposing factors of lower airway hyperresponsiveness and eosinophils-associated inflammation.Results1. We enrolled 184 subjects with allergic rhinitis, 129 subjects with non-allergic rhinitis and 162 normal controls. No significant difference (P>0.05) in the constitutional proportion of gender, height, weight and body mass index (BMI) was revealed, which indicated comparability among the three groups.2. The proportion of eosinophilss in peripheral blood was (3.34±2.26)%, (2.49±1.79)% and (1.81±1.22)% in allergic rhinitis, non-allergic rhinitis and normal control group (P<0.05). Eosinophils count was (0.20±0.16)×109/L, (0.16±0.13)×109/L and (0.10±0.07)×109/L in the three respective groups (P<0.05). The positive rate of peripheral blood eosinophil count was 17.79%, 14.06% and 1.96% (P<0.01), respectively.3. Neutrophil count in nasal lavage fluid of non-allergic rhinitis group was (48.28±141.17)/×200, which had statistical difference (P<0.05) with that of allergic rhinitis group [(13.61±46.93)/×200] and normal control group [(1.74±9.37)/×200]. Eosinophil count in nasal lavage fluid of allergic rhinitis group was (27.88±62.47)/×200, which had statistical difference (P<0.05) with that of non-allergic rhinitis group [(9.04±22.56)/×200] and normal control group [(0.76±3.46)/×200]. The positive rate of peripheral blood eosinophil count in nasal lavage fluid was 59.89%, 28.35% and 5.00% (P<0.01), respectively.4. The proportion of eosinophil in induced sputum of allergic rhinitis group was (4.84±8.63)%, which had statistical difference (P<0.05) with that of non-allergic rhinitis group [(1.29±3.23)%] and normal control group [(0.37±0.91)%]. The positive rate of eosinophil proportion in induced sputum test was 38.32%, 13.93% and 1.42% in AR, NAR and normal control group (P<0.001).5. Fractional exhaled nitric oxide of AR group was (25.82±18.58)ppb, which had statistical difference (P<0.05) with that of NAR group [(17.77±11.93)ppb] and normal control group [(14.15±6.79)ppb]. The positive rate was 34.07%, which had statistical difference (P<0.05) with that of NAR group and normal control group (15.15% and 6.25%, respectively).6. No stastistical difference (P<0.05) was shown in major spirometric parameters among three groups. The sum of susceptive positive and positive rate of bronchial provocation test in AR group was 11.96%, which had significant statistical difference (P<0.01) compared with that of NAR group (2.33%) and normal control group (1.23%).7. Two subgroups were further classified according to existence of upper airway eosinophilic inflammation in AR group, with the positive percentage rate of 48.41% and 7.50% (P<0.01) in subjects with and withour upper airway eosinophilic inflammation, respectively.8. Two subgroups were further classified according to existence of upper airway eosinophilic inflammation in NAR group, with the positive percentage rate of 26.23% and 1.69% (P<0.01) in subjects with and withour upper airway eosinophilic inflammation, respectively.9. A multiple variance Logistic regression equation of Logit(P)= -4.781+1.410X₁+2.274X₂+2.797X₃+1.134X₄ on the impact factors of eosinophils-associated inflammation was obtained, in which X₁ referred to allergen skin prick test result, X₂ was eosinophils count in peripheral blood, X₃ represented eosinophils count in nasal lavage fluid and X4 referred to bronchial provocation test result. The predisposing factors of eosinophils-associated inflammation were positive allergen skin prick test result (OR=4.096), increased eosinophils count in peripheral blood (OR=9.715), increased eosinophils count in nasal lavage fluid (OR=16.398) and susceptible positive bronchial provocation test result (OR=3.107).10. A multiple variance Logistic regression equation of Logit(P)= -4.167 + 1.453X₁ + 1.587X₂ on the impact factors of airway hyperresponsiveness was obtained, in which X₁ referred to allergen skin prick test result and X₂ was the proportion of eosinophilss in induced sputum. The predisposing factors of airway hyperresponsiveness were positive allergen skin prick test result (OR=4.276) and increased eosinophils proportion in induced sputum (OR=4.890).Conclusions1. Lower airway eosinophilsic inflammation and airway hyperresponsiveness have developed despite the absence of lower airway symptoms in a proportion of individuals with allergic rhinitis.2. Lower airway inflammatory changes have also developed despite absence of lower airway symptoms in part of subjects with non-allergic rhinitis.3. Upper airway eosinophilic inflammation was the independent predisposing factor of lower airway eosinophilic inflammation.4. Atopy and lower airway eosinophilic inflammation were independent predisposing factors of airway hyperresponsiveness. General conclusions of the study1. The method for assessment of inflammatory cell differentiation in nasal lavage fluid has been established successfully. Nasal lavage inflammatory cell absolute count is capable of offering clear discrimination between normal individuals and subjects with allergic rhinitis. Eosinophils count is the optimal positive parameter for judgment of the results of nasal lavage test, with the 95% upper reference limit of 3.47 /×200 (HE stain).2. The prevalence rate of rhinitis reaches to 19.20%, with a similar proportion of non-allergic rhinitis and allergic rhinitis. An increase in prevalence rate of asthma in subejcts with AR and NAR has been revealed, both of which are the predisposing factors of onset of asthma.3. Lower airway eosinophilic inflammation or lower airway hyperresponsiveness has evolved in some subjects without lower airway symptoms who have allergic rhinitis or non-allergic rhinitis, which might be the early stage during progression of rhinitis to asthma.