INTRODUCTION
Sputum induction has been used for many years in the diagnosis of various respiratory diseases. It is being increasingly used to study pathophysiology of various respiratory diseases.1 Curschmann's Spirals and Charcot Leyden crystals are popularly used to diagnose bronchial asthma. Sputum cytology for presence of malignant cells and various biochemical markers in sputum are also used in patients of suspected lung cancers as well as prognostic marker during follow-up. Isolation of various pathogens e.g., bacteria, fungi, viruses are useful in diagnoses of various respiratory infections.
The inhalation of an aerosol of hypertonic saline to produce sputum was firstly used by Bickerman et al in 1958 in cytology of lung cancer2. Later on, induced sputum was used for clinical purposes to detect numerous pathogenic microorganism.34 More than three decades ago, sputum induction was widely used for the diagnosis of pulmonary tuberculosis as it was superior in yield to gastric lavage. After the invent of fibreoptic bronchoscopy, it was abandoned for few years. But, again revived because of lower risk of TB transmission and low cost.56 Various studies over the past decade favours superior yield of sputum induction over fibreoptic bronchoscopy in diagnosis of pulmonary tuberculosis.7 Moreover patient tolerability and safety are good.5678 Procedure is also feasible in developing countries setting and in young children.9 In resource poor countries, sputum induction may be particularly helpful in diagnosis is because of high prevalence of tuberculosis in these countries. Though, sputum for AFB is still a gold standard to diagnose pulmonary tuberculosis, the chest symptomatics, who have dry cough and/or scanty sputum, pose a major problem in diagnosis of pulmonary tuberculosis. Adequate sputum is essential for quality microscopy diagnosis and is prerequisite for the DOTS therapy. Younger women mostly have either dry cough or scanty sputum and may have only minimal disease radiologically. A study from Vietnam particularly stress the need for quality sputum microscopy in these women.10
In early nineties, first attempt to standardize sputum induction and processing was made. First task force on sputum induction method was sponsored by European Respiratory Society.111213
METHODOLOGY
Various agents like distilled water, normal saline, hypertonic saline, glucose, surfactant active agents like tyloxapril, Tegemist and b2-agonist have been used for sputum induction. Hypertonic saline is most commonly used because of high success rate and safety. It is most helpful in sputum production for studying airway inflammation.14 β2 agonist like terbutaline has been demonstrated to enhance mucocilliary transport in healthy subjects and in patients with chronic bronchitis and mucovisidosis.15 These agents are delivered through ultrasonic nebulizer.
It is advisable that bronchodilatation medication, oxygen supply and resuscitation equipment should be available at procedure place. Although experienced technician can conduct sputum induction, a physician should supervise the procedure. Two methods are commonly employed. One proposed by Ireda et al16 in which the inhalation of the same concentration of hypertonic saline (4.5%) for increasing time interval is used while in second proposed by Pin et al1 inhalation for the same period of increasing concentration of hypertonic saline (3%, 4%, 5%) is used. Study has shown that hypertonic saline 3% is as successful as 3-5% saline.17
Mechanism
The ultrasonic nebuliser produces a mist of hypertonic saline droplets. The smaller droplets are deposited peripherally in the lung. It is suggested that the hypertonicity of the deposited saline draws interstitial fluid into the lower airways by osmosis. The hypertonic fluid also causes bronchial irritation and this stimulates bronchial secretions. After 10-20 minutes of nebulisation the fluid produced mobilize the material in the lower airways. Repeated coughing by the patient help in movement of this material into trachea to facilitate expectoration.7
Standard protocol for sputum induction
Patient is pretreated with 200 mg of albuterol or equivalent β2-agonist by inhalation. Pre-induction spirometry 10 minutes before and 10 minutes after β2-agonist is done. After placing a nasal clip, induction is started with 3% hypertonic saline (5 to 7 ml). Patient is asked to expectorate whenever he feels or at every 5 minutes. FEV1 and PEF is also checked at every 5 minutes. Saline induction is continued for 15 minutes (3 times, 5 minute each). If sputum sample is inadequate, induction can be continued for another 5 minutes. Procedure is stopped after 20 minutes if FEV1 dropped by 20%.7
Processing of the Induced Sputum
Sputum sample is kept in cold place (temp. 4°C) and processed within two hours. It is solubilized with DTT (dithiothreitol) or DTE (ditheoerythritol) followed by Dulbecco's phosphate buffer saline (D-PBS). Then the mixture is filtered and centrifuged. The supernatant is aspirated and frozen at -80°C for further analysis.18 Numerous laboratories prefer to process the entire sputum.19 Other prefer to collect and analyse the more viscid proportion of sputum (plugs).116
Characteristics of the Induced Sputum
It is a complex medium rich in mucin (protein) and also contains degradation products of DNA. Sputum is rich in mediators much more than BAL. With increase in the concentration of the hypersonic saline, there is no difference in the cell composition of the sputum, but differential effect on mediator concentration in sputum fluid phase is seen. Cell composition varies with duration of inhalation. Neutrophils decreases and macrophages increases by increasing the duration of inhalation reflecting that the sample is derived from the distal air base. Contamination of sputum with saliva is major concern and problem. It can be decreased by blowing nose, bearing nose clips or by washing the mouth.19
The quality of induced sputum is better because it is derived from the peripheral airways. This was confirmed by using a radio labeled aerosol bolus delivery technique (99m Tc Sulphur colloid particles). Induced sputum contains higher concentration of fluid phase components such as eosinophil cationic proteins (ECP), mucinlites glycoproteins and albumin as compared with BAL. These are present in airways secretions as compared with alveolar spaces.
Success and Safety of Sputum Induction
Success was obtained by non-squamous total and differential cell counts containing macrophages and safety by the fall in forced expiratory volume in one second (FEV 1). The overall success has been reported as 93%.7 The procedure is safe even amongst patients with an FEV1 less than 60% of predicted value or less than one L. Thus carefully standardized sputum induction can be safe and successful procedure in patients with asthma and COPD in clinical practice, even in presence of moderate to severe airflow limitation. Sputum induction using hypersonic saline is a safe procedure even in children and it is useful to study airway inflammation. Sputum induction is performed with b2-agonist pretreatment using ultrasonic nebulizer with 4.5% hypertonic saline gave 98% procedure completion rate. 4% of patients demonstrated 15% decrease in FEV1. An adequate sputum sample could be obtained in 92% of the children. Distressing cough was noticed in 13%. In 1% mucosal irritation was noticed.2021
Clinical uses of induced sputum
1. Asthma
It is helpful in better understanding of pathology of bronchial asthma, assessment of severity of disease, evaluation of effect of treatment and in confirmation or exclusion of diagnosis.232425 Increased level of eosinophils in sputum allows to forecast the efficiency of glucocorticosteroids in the therapy of asthma. Eosinophilic bronchitis is an important cause of chronic cough and sputum eosinophil, ECP (eosinophilic cationic protein) in induced sputum confirms the diagnosis.2627 Increased eosinophil count above the upper limit of 3% of non-squamous cells in the absence of typical clinical and PFT findings is used as a diagnostic tool. It is also used for better monitoring of drug activity and response to treatment and also to decide minimum dose of inhaled corticosteroids. Induced sputum eosinophil count is used as a diagnostic tool in occupational asthma.28
2. COPD
Sputum analysis after sputum induction gives useful information about airways inflammation in COPD.29 Sputum eosinophilia has been found as characteristic of patients with steroid response and sputum examination might serve as a screening test for continuation of long term steroid treatment in COPD. It can be used to investigate different aspects of airway inflammation. Sputum analysis has given evidence for increased number of macrophages, neutrophils and eosinophils in COPD. Changes in various mediators have been found in sputum supernatant of COPD patients (IL-8, LTB-4 and TNFa). It is also useful in serial monitoring of induced sputum inflammatory markers in COPD2430 It is useful in assessment of the effect of glucocorticosteroids or markers of neutrophilic airway inflammation like activation of neutrophils which signifies clinical improvement in patients of COPD.31 Thus, sputum induction can also guide treatment also.
3. Lung Cancer
In elderly patients of central pulmonary masses, cytology of induced sputum is useful as the first diagnostic procedure because of its safety and high sensitivity. Fernandez et al32 found 74% sensitivity in his study with a Kappa index of 0.66 with high degree of histologic correlation. Hidaka et al33 found its usefulness in diagnosis of bronchoalveolar carcinoma.
4. Tuberculosis
a) Pleural tuberculosis: Pleural tuberculosis may occur in the presence or absence of pulmonary parenchymal lesions on the chest radiograph.34 Pleural effusion associated with tuberculosis contains relatively small number of organisms making diagnosis of pleural tuberculosis difficult and oftenly requiring invasive procedures like bronchoscopy and pleural biopsy. Sputum specimens are not oftenly evaluated because many patients are not able to produce sputum spontaneously. Conde et al35 in their study observed that the yield of sputum cultures obtained by sputum induction is high in patients suspected of having pleural tuberculosis, even when there is no radiographic evidence of pulmonary parenchymal disease.
b) Pulmonary tuberculosis: Diagnosis of pulmonary tuberculosis in smear negative cases has been resting mainly upon high index of clinical suspicion, x-ray findings and tuberculin testing. Recently sputum induction has shown good results as additional yield in diagnosis of smear negative pulmonary tuberculosis.36 Diagnostic yield of single induced sputum is as good as FOB and yield of repeated induction is better favouring this tool over bronchoscopy for diagnosis of smear negative pulmonary tuberculosis. A study from India also favoring it by using β2 agonists for sputum induction.37
5. Interstitial Lung Diseases (ILD)
Induced sputum has been used to study ILD, more specifically pneumoconiosis, sarcoidosis and nongranulomatous ILD. Results have been found comparable to BAL findings for occupational lung disease and were able to distinguish sarcoidosis patients from healthy subjects and from patients with nongranulomatous lung disease. Thus it can be used as good complementary tool to BAL both in research as well as in clinical monitoring of patients with ILD.38
6. Opportunistic Infection in immunocompromised (IC) host
It is widely used for the diagnosis of pneumocystic pneumonia (PCP) with AIDS. It has also clinical utility for diagnosing PCP in IC without HIV infection.39
7. Community acquired pneumonia (CAP)
Sputum induction is a standard diagnostic procedure to identify pathogens in lower respiratory tract secretions in adults with pneumonia. Zare et al40 found it as safe and useful procedure in infants and children also with CAP from a high HIV prevalence area.
8. Others
Cough due to gastroesophageal reflex disease (GRD) has shown increased macrophages laden with lipid in induced sputum.41
Sputum induction is a safe, simple noninvasive available tool which can be used for diagnosis and management of various respiratory illness like bronchial asthma, COPD, tuberculosis, lung cancer, ILD etc. It can be a good research tool for several lung diseases.
REFERENCES
1. Pin I, Gibson PG, Kolendonicz R. Use of induced sputum cell counts to investigate airway inflammation Thorax. 1992;47:25–9
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
2. Bickerman HA, Sproul EE, Barach AL. An aerosol method ofproducing bronchial secretions in human subject a clinical technique for the detection oflung cancer Dis Chest. 1958;33:347–62
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
3. Bandyopadhyay T, Gerardi DA, Melosky ML. A comparison of induced and expectorated sputum for the microbiological diagnosis of communityacquired pneumonia Respiration. 2000;67:173–6
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
4. Larson IL, Ridzon R, Harman MM. Sputum induction versus fibreoptic bronchoscopy in the diagnosis of tuberculosis Am J Respir Crit Care Med. 2001;163:1279–80
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
5. Anderson C, Inbaber M, Menzris RL. Comparison of sputum induction with fibreoptic bronchoscopyin the diagnosis of tuberculosis Am JRespir Crit Care Med. 1995;152:1570
- Cited Here |
- Google Scholar
6. Li IM, Yang HL, Mao CF, Tang RT, Chen YF, Chem SM, et al Sputum induction to improve the diagnostic yield in patients with suspected pulmonary tuberculosis IntJTuberc Lung Dis. 1999;3:1137–9
- Cited Here |
- Google Scholar
7. Al Zahrani K, Al Jahdali H, Poirier L, Rene P, Menzies D. Yield ofsmear, culture and amplification tests from repeated sputum induction for the diagnosis of pulmonary tuberculosis In t J Tubercule Lung Dis. 2001;5:855–60
- Cited Here |
- Google Scholar
8. Conde MB, Soares LM, Mello CQ, Rezende VM, Almeida LL, Reingold AL, et al Comparison of sputum induction with fibreoptic bronchoscopyin diagnosis of tuberculosis Am J Respir Crit Care Med. 2000;162:2238–40
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
9. Zar HJ, Tannenbaum F, Apolles P, Raux P, Hanelo D, Hussey G. Sputum induction for the diagnosis of pulmonary tuberculosis in infants and young children in an urban setting in South Africa Arch Dis Child. 1999;82:305–8
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
10. Long MH, Johansson E, Lonnvoth K, Eriksson B, Winkrist A, Diwan VK. Longerdelaysin tuberculosis diagnosis among women in Vietnam IntJTubercLung Dis. 1999;3:388
- Cited Here |
- PubMed |
- Google Scholar
11. Louis R, Shute 1, Goldring K. The effect of processing on inflammatory markers in induced sputum Eur Respir J. 1999;13:660–7
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
12. Djukanovic R, Sterk PJ, Fahy] V, Hougreave FE. Standardised methodology ofsputum induction and processing Eur Respir J. 2002;37(Suppl):1S–2S
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
13. Vignola AM, Renmar SI, Hargreave FE, Fahy IV, Bonsignore MR. Standardized methodology of sputum induction and processing: Future directions Fur Respir J. 2002;37(Suppl):51S–55S
- Cited Here |
- Google Scholar
14. Pavord ID, Pizzichini MM, Hargreve FE. The use ofinduced sputum to investigate airwayinfiammation Thorax. 1997;52:498–502
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
15. Sardool P, Puchelle E, Zahni IM. Effect of terbutaline on mucociliary transport and sputum properties in chronic bronchitis Chest. 1991;80:388–91
- Cited Here |
- Google Scholar
16. Iredate MI, Wanklyn SA, Philips IP, Krausz T, Ind PW. Noninvasive assessment of bronchial inflammation in asthma No correlation between easinophilia ofinduced sputum and BHR to inhaled hypertonic saline Clin Exp Allergy. 1994;24:940–5
- Cited Here |
- Google Scholar
17. Popav TA, Pizzichini MM, Kolendoncz R, Dolovich J. Some technical factors influencing the induction of sputum for cell analysis Eur Respir J. 1995;8:559–65
- Cited Here |
- PubMed |
- Google Scholar
18. Spanevello A, Beghe B, Bianchi A, Migliori GB, Ambrosetti M, Neri M, et al Comparison of two methods ofprocessing induced sputum: Selected versus entire sputum Am JRespir Crit Care Med. 1998;157:665–8
- Cited Here |
- Google Scholar
19. Alexis NE, Hu SC, Zemen K, Alter T, Bennett WD. Induced sputum derives from the central airways Am JRespirCrit Care Med. 2001;164:1964–70
- Cited Here |
- Google Scholar
20. Hunter CI, Ward R, Woltmann G, Wardlaw AJ, Pavord ID. The safety and success rate of sputum induction using a low output ultrasonic nebuliser Respir Med. 1999;93:345–8
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
21. Jones PD, Hankin R, Simpson 1, Gibson PG, Henry RL. The tolerability, safety and success of sputum induction and combined hypertonic saline challenge in children Am J Respir Crit Care Med. 2001;164:1146–9
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
22. Polosa R, Louis R, Cacciola R. Sputum eosinophilia is more closelyassociated with airway responsiveness to bradykinin than methacholine in asthma EurRespir1. 1998;12:551–6
23. Louis R, Lau LC, Bron AO. The relationship between airways inflammation and asthma severity Am J Respir Crit Care Med. 2000;161:9–16
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
24. Louis R, Shute J, Biagi S. Cell infiltration, ICAM-1 expression, and eosinophil chemotacticactivityin asthmatic sputum Am J Respir Crit Care Med. 1997;155:466–72
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
25. Pavord ID, Brightling CE, Woltmann G. Non-eosinophilic corticosteroid unresponsive asthma [letter] Lancet. 1999;353:2213–14
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
26. Brightling CE, Ward R, Gob KL. Eosinophilic bronchitis is an important cause of chronic cough Am JRespir Crit Care Med. 1999;160:406–10
- Cited Here |
- Google Scholar
27. Siergiejko Z. Bronchoalveolarlavage and induced sputum in asthmatic and COPD patients Pol Merkuriusz Lek. 2003;14):545–7
- Cited Here |
- PubMed |
- Google Scholar
28. Maestrelli P, Calcagni PG, Saetta M. Sputum eosinophilia after asthmatic responses induced by isocyanates in sensitized subjects Clin Exp Allergy. 1994;24:29–34
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
29. Pizzichini E, Pizzichini MM, Gibson P. Sputum eosinophilia predicts benefit from prednisone in smokers with chronic obstructive bronchitis Am J Respir Crit Care Med. 1998;158:1511–17
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
30. Troumakidou M, Tzanakis M, Siafakar NM. Induced sputum in the investigation ofairwayinflammation ofCOPD Respir Med. 2003;97:863–71
- Cited Here |
- Google Scholar
31. Barezyk A, Sozaneka E, Trzaska M. Decreased levels of myeloperoxidase in induced sputum ofpatients with COPD after treatment with oral glucocorticoids Chest. 2004;126:389–93
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
32. Aloneo Fernandez A, Garcia Rio F, Mayoralas Alises S. Usefulness ofinduced sputum cytologyin the studyof central masses in elderly patients JRev Clin Exp. 2001;201:444–7
- Cited Here |
- Google Scholar
33. Hidaka N, Nagao K. Bronchoalveolarcarcinomaaccompanied by severe bronchorrhea Chest. 1996;110:281–2
- Cited Here |
- View Full Text | PubMed | CrossRef |
- Google Scholar
34. Berger HW, Mejia E. Tuberculous pleurisy Chest. 1973;63:88–92
- Cited Here |
- PubMed | CrossRef |
- Google Scholar
35. Conde MB, Loivos AC, Rezende VM, Soares LM, Mello FCO, Reingold AL, et al Yield of sputum induction in the diagnosis of pleural tuberculosis Am I Respir Crit Care Med. 2003;167:723–5
- Cited Here |
- Google Scholar
36. Parry CM, Kamato 0, Harries AD, Wirima JJ, Nyirenda CM, Nyanulu DS, et al The use of sputum induction for establishinga diagnosis in patients with suspected pulmonary tuberculosis in Malawi Tubercle LungDis. 1995;76:72–6
- Cited Here |
- Google Scholar
37. Yazdani A, Kiran L, Murthy KJR. Sputum induction by oral salbutamol Ind JTuber. 2002;49:221–3
- Cited Here |
- Google Scholar
38. Olivieri D, D'Ippolito R, Chetta A. Induced sputum: diagnostic value in interstitial lung disease Curr Opin Pul Med. 2000;6:411–14
- Cited Here |
- Google Scholar
39. Larosgue RC, Katz JT, Perruzzi P, Baden LR. The utility of sputum induction fordiagnosis ofpneumocystitis pneumonia in immunocompromised patients without human immunodeficiency virus CIinInfect Dis. 2003;37:1380–3
- Cited Here |
- Google Scholar
40. Zar HJ, Tannenbaum E, Hanslo D, Hussey G. Sputum induction as a diagnostic tool for community-acquired pneumonia in infants and young children from a high HIV prevalence area PediatrPulmonol. 2003;36:58–62
- Cited Here |
- Google Scholar
41. Parameswaran K, Anvari M, Efthimiadis A. Lipid-laden macrophages in induced sputum are a marker of oropharyngeal reflux and possible gastric aspiration Am J Respir Crit Care Med. 2001;164:1146–9
- Cited Here |
- Google Scholar
Keywords:
Sputum induction; Pulmonary tuberculosis; Fibero bronchoscopy