Diagnosing occupational asthma: use of induced sputum.

The diagnosis of occupational asthma (OA) needs to be made with as much objective evidence as possible. If there is airway inflammation, measurement of this should be an asset. The objective of this study was to investigate whether there is an increase in induced sputum and blood eosinophils and eosinophil cationic protein (ECP) in OA after work exposure. Patients were assessed after a 2-4 week period at work and away from work with cell counts and ECP assays performed blind to the clinical data. They were considered to have OA if symptoms were worse at work and there was a fall in forced expiratory volume in one second (FEV1) > or =20% or in the provocative concentration of methacholine causing a 20% fall in FEV1 (PC20) of four-fold or more compared with away from work. Patients whose symptoms were worse at work but had a change in FEV1 of <20% and in methacholine PC20 of less than four-fold were considered as controls. Sixteen patients were studied. Ten had OA and six were controls. Patients with OA had a significant increase in median (interquartile range) sputum eosinophils and ECP when at work compared with the periods out of work, 10.0 (17.05) versus 0.8 (1.6)% (p=0.007) and 3,840 (6,076) versus 116 (180) microg x L(-1) (p=0.01). They also had a higher blood eosinophil count, 0.3 (0.5) x 10(9) versus 0.2 (0.1) x 10(9) x L(-1) (p=0.013), and a trend towards higher serum ECP levels, 44.0 (20.0) versus 32.0 (18.5) microg x L(-1) (p=0.07). In conclusion, the proportion of eosinophils and levels of eosinophil cationic protein in sputum are particularly high at work in patients with occupational asthma, suggesting that the measurement of these factors can supplement other physiological outcomes in establishing the diagnosis of occupational asthma.     

诱导痰技术及其在支气管哮喘、慢性阻塞性肺疾病鉴别中的应用

支气管哮喘、慢性阻塞性肺疾病均为气道炎症性疾病,它们的共同病理特征为气道炎症,由多种细胞及细胞组分参与,因此对参与产生这些疾病的炎症细胞及其细胞因子进行研究,具有重要的价值,本文主要总结了目前使用的诱导痰技术的基本原理及其安全性和可靠性,并综述了运用此技术分析哮喘与慢性阻塞性肺疾病患者痰中细胞成分与细胞因子的异同,对检测指标进行了系统的总结,展望了该技术的应用前景,并分析其不足之处。     

Prednisone-dependent asthma: inflammatory indices in induced sputum.

The kinetics of changes in inflammatory indices in induced sputum from eight prednisone dependent asthmatics whose minimum clinical maintenance and exacerbation doses were known were investigated. The study began on the last day of a course of 30 mg prednisone daily for one week. Thereafter, the daily prednisone was reduced in a structured way to below the maintenance dose. This treatment was continued until a clinical exacerbation occurred. Prednisone 30 mg daily was then given again for one week. The mean duration of prednisone reduction was 7.4 weeks and the median dose was 7.5 mg x day(-1). Increases in sputum eosinophils preceded increases in blood eosinophils by 4 weeks and worsening of symptoms and forced expiratory volume in one second by 6 weeks. The clinical exacerbation was also accompanied by sputum neutrophilia and increases in sputum eosinophil cationic protein (ECP), fibrinogen and interleukin (IL)-5. Treatment with prednisone suppressed median sputum eosinophilia (from 16.3 to 0%, p<0.001), decreased sputum ECP (from 7,480 to 700 microg x L(-1), p = 0.01), but did not improve neutrophil numbers, fibrinogen or IL-5. The results show that the reduction of prednisone treatment in prednisone-dependent asthmatics evokes a severe airway eosinophilic inflammatory response. Clinical and blood indices deteriorate later than those in sputum suggesting that sputum examination may be useful to identify the minimum regular dose of prednisone required in these patients.     

Reproducibility in induced sputum in children with asthma

In asthmatic children sputum-induction with hypertonic saline is useful to quantify the eosinophilic inflammation. However, only few data are available about feasibility and safety of the procedure in children. Therefore, taking 9 non-atopic healthy control children (mean age 11.8 years) and 34 asthmatic children (mean age 11.4 years), inhaling n = 25 Budesonid (400-1200 micrograms/die) and n = 9 DNCG (60 mg/die), sputum induction was performed twice within 6 weeks. Briefly, 10 minutes after inhalation of 200 micrograms salbutamol subjects inhaled hypertonic saline (3, 4 and 5%) for in all 30 minutes, while all 5 minutes lung function was checked and expectoration of sputum was supported. Adequate sputum plugs were separated from contaminating saliva and processed immediately employing native chamber and cytospin cell count as well as measurement of eosinophilic cationic protein (ECP). Sputum-induction could be performed in 84 out of 86 planed tests (97.7%) without any objective clinical adverse effects. The mean fall in FEV1 was 3.0%, the maximum 11.0%. The reproducibility of eosinophil, neutrophil and lymphocyte differential cell count (5-95%-values Test1: 0.0-4.2%, 0.8-11.4%, and 3.2-35.1%, respectively) was moderate for eosinophils and neutrophils (Intraclass-Correlation-Coefficient (ICC) 0.41) as well as for lymphocytes (ICC = 0.49). For ECP 5-95%-values Test1: 39.8-8000.0 micrograms/l) only a fair reproducibility (ICC = 0.24) was found. The ICC levels for total cell count (ICC = 0.31) and for weight of the sputum plug (ICC = 0.30) were also fair. Based on the procedure presented induced sputum is a feasible and safe method in childhood. The differential sputum cell count of eosinophils, neutrophils and lymphocytes can be recommended as parameters with moderate reproducibility.     

An investigation of airway acidification in asthma using induced sputum: a study of feasibility and correlation

RATIONALE: Acidification of the airways seems to be involved in asthma pathophysiology, but its assessment might be difficult. OBJECTIVES: The aim of our study is to assess the feasibility and validity of airway acidification measurement by induced sputum and its clinical significance in asthma. METHODS: Induced-sputum samples were obtained in 57 outpatients with asthma. The between-sample repeatability after 48 hours was measured in an independent population of 14 patients with asthma. pH was measured using a pH meter. The control of asthma was established by the Asthma Control Questionnaire. MEASUREMENTS AND MAIN RESULTS: The pH measurement was feasible in all samples and repeatable both within (intraclass correlation coefficient [ICC], 0.96) and between samples (ICC, 0.621). The mean pH was significantly different between healthy subjects and patients with asthma, including in those with controlled (mean pH: 7.54 in healthy subjects vs. 7.28 in subjects with controlled asthma; p = 0.0105) and uncontrolled disease (mean pH: 7.54 in healthy subjects vs. 7.06 in subjects with uncontrolled disease; p < 0.0001), and between patients with stable asthma and those with poorly controlled asthma (7.28 vs. 7.06, respectively; p = 0.0134). The validity of the method was assessed with the receiver operating characteristic curves and induced-sputum lower pH values (with a cutoff value of 7.3; sensitivity, 72.1%; specificity, 100%). CONCLUSIONS: Patients with asthma show lower pH than healthy subjects. Patients with poorly controlled asthma seem to have the lowest induced-sputum pH, independent of the GINA (Global Initiative for Asthma) severity level. In conclusion, induced sputum is a feasible, repeatable, noninvasive method to measure airway pH. The pH in induced sputum may reflect a different aspect of asthma from sputum eosinophils and be related to different pathophysiologic factors.     

Inflammatory subtypes in asthma: assessment and identification using induced sputum

OBJECTIVE: The authors sought to investigate the detection of non-eosinophilic asthma using induced sputum. Although this is an important subtype of clinical asthma, its recognition is not standardized. METHODS: Adult non-smokers with asthma and healthy controls underwent sputum induction and hypertonic saline challenge. Non-eosinophilic asthma was defined as symptomatic asthma with normal sputum eosinophil counts. The normal range for sputum eosinophil count was determined using the 95th percentile from the healthy control group as a cut-off point. RESULTS: The recognition of non-eosinophilic asthma using eosinophil proportion was in agreement with a definition based on absolute eosinophil count (kappa 0.67). Non-eosinophilic asthma was a stable subtype over both the short term (4 weeks) and longer term (5 years, kappa 0.77). Airway inflammation in asthma could be categorized into four inflammatory subtypes based on sputum eosinophil and neutrophil proportions. These subtypes were neutrophilic asthma, eosinophilic asthma, mixed granulocytic asthma and paucigranulocytic asthma. Subjects with increased neutrophils (neutrophilic asthma and mixed granulocytic asthma) were older and had an increased total cell count and cell viability compared with other subtypes. CONCLUSION: Induced sputum eosinophil proportion is a good discriminator for eosinophilic asthma, providing a reproducible definition of a homogenous group. The remaining non-eosinophilic subjects are heterogeneous and can be further classified based on the presence of neutrophils. These inflammatory subtypes have important implications for the investigation and characterization of airway inflammation in asthma.     

Is the clinical history a satisfactory means of diagnosing occupational asthma?

In some countries a diagnosis of occupational asthma for medicolegal purposes is made when a questionnaire is suggestive and the individual is exposed to a product known to be a sensitizer. The value of an open questionnaire administered by physicians with experience in occupational asthma is unknown, however. We prospectively assessed all subjects (162) referred to our clinic because their physicians thought their asthma might be work related. The medical questionnaire included questions about the nature of the symptoms (dyspnea, wheezing, cough, and chest tightness) and the timing (worse at work or after a shift at work or improved during weekends or holidays). An initial clinical assessment was set by the physician on a scale that ranged from highly probable to probable, uncertain, unlikely, or absent. Subjects then underwent objective assessment with specific inhalation challenges (n = 72), serial monitoring of peak expiratory flow rates for periods at work and away from work (n = 29), or both (n = 61), to confirm or rule out the diagnosis of occupational asthma. A total of 75 subjects (46%) were shown to have occupational asthma. Symptoms alone (type and timing) did not provide a satisfactory differentiation between those subjects with and those without occupational asthma. For example, 66 of 75 (88%) subjects with occupational asthma said that their symptoms improved during holidays, but 66 of 87 (76%) subjects without occupational asthma experienced a similar improvement.(ABSTRACT TRUNCATED AT 250 WORDS)     

Nitric oxide metabolites in patients with asthma: induced sputum versus blood

Nitric oxide (NO) plays an important role in physiological regulation of the airways. The monitoring of airway inflammation has being observed in bronchial asthma directly, by sputum examination, and indirectly, by measurements in peripheral blood. To investigate the diagnostic value of these two methods, we compared NO metabolites in induced sputum and serum obtained in patients with asthma and control subjects. Hypertonic saline induced sputum and serum were obtained in 13 patients with asthma and 10 control subjects. NO metabolite level was assayed by using modified Griess reaction. Eosinophil cationic protein (ECP) was measured by fluoroimmunoassay, and detected interleukin (IL)-5 by a sandwich ELISA. The accuracy of the tests was measured by plotting the data in receiver operating characteristic (ROC) curves and comparing the area under the curve for NO metabolites. Asthmatic patients, compared with control subjects, had significantly higher NO metabolites in induced sputum (1252.5+/-203.3 mol l(-1) vs. 557.2+/-101.5 mol l(-1), P<0.01) but not in serum. IL-5 in induced sputum was detected more frequently in patients with asthma than in control subjects [11/13 (84.6%) vs. 1/10 (10%), P<0.01]. Asthmatic patients, compared with control subjects, had significantly higher ECP concentration in induced sputum (1270.0+/-197.9 g l vs. 154.6+/-47.4 g l(-1), P<0.01). There were significant positive correlations between NO metabolites in induced sputum and eosinophils, ECP in induced sputum (r=0.58 P<0.05; r=0.64, P<0.01) in patients with asthma but not in serum. The area under the ROC curve showed that NO metabolites in induced sputum (0.78) are more accurate marker than NO metabolites in serum (0.53) (P<0.05). These findings suggest that NO metabolites in induced sputum is a more valuable indicator to monitor asthmatic airway inflammation than those in serum.     

Eosinophilic inflammation assessed by induced sputum in corticosteroid-dependent asthma.

Sputum induction can be used to study airway inflammation in asthmatics. However, it has not been used in patients with corticosteroid-dependent asthma requiring long-term oral corticosteroids. The aim of the study was to assess the number of eosinophils and the levels of eosinophil cationic protein (ECP) in sputum of 17 corticosteroid-dependent asthmatics by comparison with nine mild untreated asthmatics, 10 moderate asthmatics receiving inhaled steroids (ICS) and 11 healthy subjects. In the 17 corticosteroid-dependent asthmatics, we examined sputum eosinophil markers on two occasions and correlated with the control of asthma. Eosinophils were undetectable in controls and were detected in 63.8% of asthmatics. There were no significant differences between the three groups of asthmatics. ECP levels were significantly increased in ICS or corticosteroid-dependent asthmatics by comparison to controls and mild asthmatics. There was no significant difference between ICS and corticosteroid-dependent asthmatics. During follow-up, corticosteroid-dependent asthmatics with a controlled disease had no significant change in eosinophil numbers or ECP levels. On the other hand, corticosteroid-dependent asthmatics with recent exacerbations had a non-significant increase in eosinophil numbers and a significant increase in ECP levels. This study shows that ECP levels may be more accurate than eosinophil numbers in assessing exacerbations in corticosteroid-dependent asthmatics.     

诱导痰嗜酸粒细胞与慢性持续期支气管哮喘病情严重程度的关系

目的观察初诊不同病情严重程度慢性持续期支气管哮喘（简称哮喘）患者的诱导痰嗜酸粒细胞（eosinophil,EOS）比例变化,探讨二者之间的关系,并分析诱导痰EOS比例与肺功能的相关性。方法收集专科门诊就诊的63例初诊慢性持续期哮喘患者,根据症状分为轻度持续、中度持续、重度持续3组,分别予诱导痰和肺功能检查。观察不同病情严重程度的患者气道炎症状况。对所得数据用SPSS 15．0软件分析,各组间总体分析采用Kruskal—wallis法,两组间分析采用Mann-Whitney U test法。结果①慢性持续期患者诱导痰EOS比例随病情严重程度增加呈增高趋势,重度持续患者诱导痰EOS比例显著高于轻度持续患者（41．8％vs17．8％,P=0．033）,但轻度持续与中度持续、中度持续与重度持续患者之间比较诱导痰EOS比例差异无统计学意义（P〉0．05）;②诱导痰EOS比例与第1秒用力呼气容积差异无统计学意义（r=-0．111,P〉0．05）,与第1秒用力呼气容积／用力肺活量（％）差异无统计学意义（r=-0．154,P〉0．05）。结论慢性持续期哮喘患者病情严重程度与诱导痰EOS比例有关,但症状不能完全反映气道炎症程度。评价哮喘患者的严重程度时应结合临床症状和气道炎症程度综合考虑。     

Aldosterone antagonism: An emerging strategy for effective blood pressure lowering

Aldosterone antagonists have been available for many decades for the treatment of hypertension, but their use has been mostly limited to patients with classic primary aldosteronism or to combination products with hydrochlorothiazide to minimize risk for hypokalemia. Recently, indications for aldosterone antagonists have been expanded to include congestive heart failure and first-line treatment of mild-to-moderate hypertension. In addition, we have reported that spironolactone has significant antihypertensive benefit when added to existing regimens in patients with resistant hypertension. This benefit was present in patients with and without hyperaldosteronism and was additive to chronic renin-angiotensin blockade with angiotensin-converting enzyme (ACE) inhibitors or angiotensinreceptor blockers (ARBs). Eplerenone, a selective aldosterone antagonist, avoids the androgen and progesterone receptor-related adverse events that sometimes occur with spironolactone, such as breast tenderness, gynecomastia, sexual dysfunction, and menstrual irregularities. In clinical trials, eplerenone has been shown to have antihypertensive benefit in treating mild-to-moderate hypertension similar to other widely used classes of agents. With recent demonstrations of benefit in multiple segments of the hypertensive population, aldosterone antagonists represent emerging opportunity for controlling high blood pressure.     

Comparison of induced sputum inflammatory profiles between childhood and adult-onset asthma

The aim of this study was to investigate differences in airway inflammation between childhood and adult-onset asthma. A total of 47 asthmatic subjects were recruited from patients attending outpatient clinic. A group of 32 adults, mean age 42.8 years (yrs) and a group of 15 children, mean age 11.7 yrs were included. The two groups did not differ in respect to gender, dose of inhaled corticosteroids, atopy status or duration of asthma (mean duration 7.75 yr). Lung function tests, and sputum induction were performed. Flowcytometry was used to study cell population and interleukin-8, eosinophilic cationic protein (ECP) and granulocyte-macrophage colony stimulating factor were measured by enzyme-linked immunosorbent assay (ELISA). Three out of 15 (20%) of the children and 6 out of 32 (19%) of the adult patients were unable to produce a sufficient sputum sample. However, all individuals tolerated the procedure well. The viability of induced sputum cells did not differ among adult-onset asthmatics and children with asthma. Children had greater number of total cells in induced sputum compared with adult subjects (P=0.02). No statistical difference in T-lymphocytes subsets was found between the two groups, except for CD25 (P=0.04). A negative correlation was found between forced expiratory volume (FEV1) values and ECP levels (r=0.338, P=0.04) in the whole population (children and adults). Our study showed that the immunopathology of pediatric and adult asthma is similar and sputum induction provides opportunities for comparison of airway inflammation in childhood and adult asthma safely.     

诱导痰嗜酸粒细胞与慢性持续期支气管哮喘病情严重程度的关系

目的 观察初诊不同病情严重程度慢性持续期支气管哮喘(简称哮喘)患者的诱导痰嗜酸粒细胞(eosinophil,EOS)比例变化,探讨二者之间的关系,并分析诱导痰EOS比例与肺功能的相关性.方法 收集专科门诊就诊的63例初诊慢性持续期哮喘患者,根据症状分为轻度持续、中度持续、重度持续3组,分别予诱导痰和肺功能检查.观察不同病情严重程度的患者气道炎症状况.对所得数据用SPSS 15.0软件分析,各组间总体分析采用Kruskal-Wallis法,两组间分析采用Mann-Whitney U test法.结果 ①慢性持续期患者诱导痰EOS比例随病情严重程度增加呈增高趋势,重度持续患者诱导痰EOS比例显著高于轻度持续患者(41.8%vs 17.8%,P=0.033),但轻度持续与中度持续、中度持续与重度持续患者之间比较诱导痰EOS比例差异无统计学意义(P>0.05);②诱导痰EOS比例与第1秒用力呼气容积差异无统计学意义(r=-0.111,P>0.05),与第1秒用力呼气容积/用力肺活量(%)差异无统计学意义(r=-0.154,P>0.05).结论 慢性持续期哮喘患者病情严重程度与诱导痰EOS比例有关,但症状不能完全反映气道炎症程度.评价哮喘患者的严重程度时应结合临床症状和气道炎症程度综合考虑.     

偏苯三酸酐诱导职业性哮喘大鼠模型的研究

目的 以低分子量化工原料偏苯三酸酐(tri-mellitic-anhydride,TMA)作为致敏剂复制职业性哮喘大鼠模型.方法 采用大鼠背部皮内注射致敏剂致敏,超声雾化器雾化吸入致敏剂激发的方法复制职业性哮喘大鼠模型,以大鼠引喘潜伏期,血液和支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)中细胞学检查、BALF中IL-4和INF-γ检测、血清IgE检测、大鼠离体气管条对乙酰胆碱的反应性、肺组织病理学检查为评判指标,考察TMA复制职业性哮喘大鼠模型的可行性.结果 TMA模型组引喘潜伏期较短(与正常对照组比较P＜0.05),血液及BALF中嗜酸粒细胞(eosinophil,EOS)百分比升高(与正常对照组比较P ＜0.05),血清中IgE升高(与正常对照组比较P＜0.05),BALF中IL-4升高、INF-γ降低(与正常对照组比较P＜0.05～0.01),模型组离体气管条对乙酰胆碱的反应性提高(与正常对照组比较P＜0.01),肺组织病理检查见肺支气管上皮细胞变性坏死脱落,支气管腔内可见大量EOS、单核细胞、淋巴细胞等炎性渗出物,以及管壁增厚、管腔狭窄等炎症表现.结论 通过行为学观察、血液和BALF中细胞学检查、血清IgE检测、BALF中细胞因子检测和肺组织病理检测可见TMA致大鼠哮喘模型具有职业性哮喘的主要特征,表明采用TMA作为造模剂,适当的剂量和致敏方法复制类似职业性哮喘动物模型是可行的.     

偏苯三酸酐诱导职业性哮喘大鼠模型的研究

目的以低分子量化工原料偏苯三酸酐(tri-mellitic-anhydride,TMA)作为致敏剂复制职业性哮喘大鼠模型。方法采用大鼠背部皮内注射致敏剂致敏,超声雾化器雾化吸入致敏剂激发的方法复制职业性哮喘大鼠模型,以大鼠引喘潜伏期,血液和支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)中细胞学检查、BALF中IL-4和INF-γ检测、血清IgE检测、大鼠离体气管条对乙酰胆碱的反应性、肺组织病理学检查为评判指标,考察TMA复制职业性哮喘大鼠模型的可行性。结果 TMA模型组引喘潜伏期较短(与正常对照组比较P<0.05),血液及BALF中嗜酸粒细胞(eosinophil,EOS)百分比升高(与正常对照组比较P<0.05),血清中IgE升高(与正常对照组比较P<0.05),BALF中IL-4升高、INF-γ降低(与正常对照组比较P<0.05～0.01),模型组离体气管条对乙酰胆碱的反应性提高(与正常对照组比较P<0.01),肺组织病理检查见肺支气管上皮细胞变性坏死脱落,支气管腔内可见大量EOS、单核细胞、淋巴细胞等炎性渗出物,以及管壁增厚、管腔狭窄等炎症表现。结论通过行为学观察、血液和BALF中细胞学检查、血清IgE检测、BALF中细胞因子检测和肺组织病理检测可见TMA致大鼠哮喘模型具有职业性哮喘的主要特征,表明采用TMA作为造模剂,适当的剂量和致敏方法复制类似职业性哮喘动物模型是可行的。