Sputum Biomarkers Research Articles

Airway inflammation in mild cystic fibrosis

BackgroundAirway infection and inflammation play major roles in the progression of cystic fibrosis (CF) lung disease. In patients with mild disease, airway inflammation is a clinically relevant and often underdiagnosed feature. Lung function, sputum cell counts, and cytokine profiles in CF with mild disease might be different in patients with and without involvement of small airway disease (SAD). MethodsPatients with mild CF (n=32) and 22 healthy controls were enrolled in this study. Patients with CF were assigned to two groups: (1) patients without SAD (n=19, median age 12.3years, MEF25>50% predicted), and (2) patients with SAD (n=13 median age, 13.2years, MEF25<50% predicted). Lung function parameters were measured, cells in induced sputum were counted, and cytokines/chemokines (IL-1β, IL-6, IL-8, TNF-α) were analyzed by real-time quantitative PCR (qRT-PCR) and cytometric bead array (CBA). ResultsPatients with CF had significant elevated levels of pro-inflammatory genes in qRT-PCR and secreted gene products in CBA compared to controls. Patients with CF and SAD had significantly increased trapped air (RV/TLC) and pronounced airway inflammation compared to controls as indicated by elevated levels of sputum biomarkers like total cells, neutrophils, and IL6. ConclusionsOur study demonstrated that patients with CF with mild disease defined by lung function might be further endotyped according to their involvement of SAD. In patients with CF and SAD, airway neutrophilic inflammation is more pronounced and is in part distinct from that seen in patients without SAD.

Do we really need asthma–chronic obstructive pulmonary disease overlap syndrome?

The association of asthma and chronic obstructive pulmonary disease (COPD) in the same patient, which is designated as mixed asthma-COPD phenotype or overlap syndrome (ACOS), remains a controversial issue. This is primarily because many conflicting aspects in the definition of ACOS remain, and it is extremely difficult to summarize the distinctive features of this syndrome. Furthermore, we are realizing that asthma, COPD, and ACOS are not single diseases but rather syndromes consisting of several endotypes and phenotypes and, consequently, comprising a spectrum of diseases. The umbrella term ACOS blurs the lines between asthma and COPD and allows an approach that simplifies therapy. However, this approach contradicts the modern concept according to which we must move toward more targeted and personalized therapies to treat patients with these diseases. Therefore we argue that the term ACOS must be abandoned and ultimately replaced when new phenotypes and underlying endotypes are identified and a new taxonomy of airway diseases is generated.

Diagnosing Lung Cancers through Examination of Micro-RNA Biomarkers in Blood, Plasma, Serum and Sputum: A Review and Summary of Current Literature

Lung cancer is the leading cause of cancer related morbidity and mortality worldwide. Currently, the vast majority of lung cancers are diagnosed at a late stage, when patients become symptomatic leading to dismal, less than 15% five-year survival rates. Evidence has demonstrated that screening computed tomography scans can be used to detect lung cancer, but these scans have high false positive rates. Therefore, there is a continued need for the development of minimally-invasive methods to screen the high risk population and diagnose lung cancer at an earlier, curable stage. One such promising area is the use micro-RNAs. These are short, non-coding RNA molecules that have been shown in previous research to be dysregulated in cancers. This review will focus on the potential use of miRNA levels in various biological fluids (whole blood, plasma, serum, and sputum) and demonstrate their potential utility as screening and diagnostic biomarkers for lung cancer. Current research will be analyzed and compared, and future directions in establishing the use of miRNAs for detecting lung cancer will be discussed.

Lung microbiome dynamics in COPD exacerbations.

Increasing evidence suggests that the lung microbiome plays an important role in chronic obstructive pulmonary disease (COPD) severity. However, the dynamics of the lung microbiome during COPD exacerbations and its potential role in disease aetiology remain poorly understood.We completed a longitudinal 16S ribosomal RNA survey of the lung microbiome on 476 sputum samples collected from 87 subjects with COPD at four visits defined as stable state, exacerbation, 2 weeks post-therapy and 6 weeks recovery.Our analysis revealed a dynamic lung microbiota where changes appeared to be associated with exacerbation events and indicative of specific exacerbation phenotypes. Antibiotic and steroid treatments appear to have differential effects on the lung microbiome. We depict a microbial interaction network for the lung microbiome and suggest that perturbation of a few bacterial operational taxonomic units, in particular Haemophilus spp., could greatly impact the overall microbial community structure. Furthermore, several serum and sputum biomarkers, in particular sputum interleukin-8, appear to be highly correlated with the structure and diversity of the microbiome.Our study furthers the understanding of lung microbiome dynamics in COPD patients and highlights its potential as a biomarker, and possibly a target, for future respiratory therapeutics.

An update on sputum MicroRNAs in lung cancer diagnosis.

Lung cancer is one of the leading cause of cancer mortality in the world. It is well known that genetic damages could result in lung tumor genesis. Despite years of research, the survival rate of the patients has not been markedly improved. According to lack of high sensitivity and specificity in diagnostic tests, just about 15-20% of lung cancer cases are discovered prior to progression of the disease. In last decade, sputum biomarkers have been developed for early detection/diagnosis of lung cancer. MicroRNAs are a class of small endogenous noncoding RNAs, which act as post-transcriptional regulators. Some specific miRNAs can have multifunctions in lung development and their aberrant expression could induce lung tumor genesis. The differences in miRNAs between the normal and cancerous lung lead to emerging of a novel type of biomarkers, which can be helpful in screening of high risk individuals, diagnosis of lung cancer as well as its therapy.

The metabolomic detection of lung cancer biomarkers in sputum

ObjectivesDeveloping screening and diagnosis methodologies based on novel biomarkers should allow for the detection of the lung cancer (LC) and possibly at an earlier stage and thereby increase the effectiveness of clinical interventions. Here, our primary objective was to evaluate the potential of spontaneous sputum as a source of non-invasive metabolomic biomarkers for LC status. Materials and methodsSpontaneous sputum was collected and processed from 34 patients with suspected LC, alongside 33 healthy controls. Of the 34 patients, 23 were subsequently diagnosed with LC (LC+, 16 NSCLC, six SCLC, and one radiological diagnosis), at various stages of disease progression. The 67 samples were analysed using flow infusion electrospray ion mass spectrometry (FIE-MS) and gas-chromatography mass spectrometry (GC–MS). ResultsPrincipal component analysis identified negative mode FIE-MS as having the main separating power between samples from healthy and LC. Discriminatory metabolites were identified using ANOVA and Random Forest. Indications of potential diagnostic accuracy involved the use of receiver operating characteristic/area under the curve (ROC/AUC) analyses. This approach identified metabolites changes that were only observed with LC. Metabolites with AUC values of greater than 0.8 which distinguished between LC+/LC− binary classifications where identified and included Ganglioside GM1 which has previously been linked to LC. ConclusionThis study indicates that metabolomics based on sputum can yield metabolites that can be used as a diagnostic and/or discriminator tool. These could aid clinical intervention and targeted diagnosis of LC within an ‘at risk’ LC− population group. The use of sputum as a non-invasive source of metabolite biomarkers may aid in the development of an at-risk population screening programme for lung cancer or enhanced clinical diagnostic pathways.

Tolerability in man following inhalation dosing of the selective TLR7 agonist, AZD8848

BackgroundMany patients with asthma have a T-helper type 2 (Th2) driven inflammation of the lung, whereas toll-like receptor 7 (TLR7) agonists, by inducing type I interferons, inhibit Th2 responses. In...

Patients with chronic obstructive pulmonary disease and chronically colonized with Haemophilus influenzae during stable disease phase have increased airway inflammation.

BackgroundSome patients with chronic obstructive pulmonary disease (COPD) show increased airway inflammation and bacterial colonization during stable phase. The aim of this study was to follow COPD patients and investigate chronic colonization with pathogenic bacteria during stable disease phase, and relate these findings to clinical parameters, inflammatory pattern, lung function, and exacerbations.MethodsForty-three patients with COPD were included while in a stable state and followed up monthly until exacerbation or for a maximum of 6 months. The patients completed the Clinical COPD Questionnaire and Medical Research Council dyspnea scale questionnaires, and exhaled breath condensate was collected, followed by spirometry, impulse oscillometry, and sputum induction.ResultsTen patients were chronically colonized (ie, colonized at all visits) with Haemophilus influenzae during stable phase. These patients had higher sputum levels of leukotriene B4 (P<0.001), 8-isoprostane (P=0.002), myeloperoxidase activity (P=0.028), and interleukin-8 (P=0.02) during stable phase when compared with other patients. In addition, they had lower forced vital capacity (P=0.035) and reactance at 5 Hz (P=0.034), but there was no difference in forced expiratory volume in 1 second (FEV1), FEV1 % predicted, forced vital capacity % predicted, exhaled breath condensate biomarkers, C-reactive protein, or Clinical COPD Questionnaire and Medical Research Council dyspnea scale results. Three patients had intermittent colonization (colonized at only some visits) of H. influenzae during stable phase, and had lower levels of inflammatory biomarkers in sputum when compared with the chronically colonized patients. The difference in airway inflammation seen during stable phase in patients chronically colonized with H. influenzae was not observed during exacerbations.ConclusionSome COPD patients who were chronically colonized with H. influenzae during stable phase showed increased airway inflammation and reduced lung volumes when compared with non-chronically colonized patients.

Characterization of the inflammatory response to inhaled lipopolysaccharide in mild to moderate chronic obstructive pulmonary disease.

Lipopolysaccharide (LPS) inhalation causes increased airway and systemic inflammation. We investigated LPS inhalation in patients with chronic obstructive pulmonary disease (COPD) as a model of bacterial exacerbations. We studied safety, changes in sputum and systemic biomarkers. We have also investigated interleukin (IL)-17 concentrations in this model. Twelve COPD patients inhaled 5 μg LPS. Safety was monitored over 24 h. Sputum was induced at baseline, 6 and 24 h for cells and IL-8, IL-17, neutrophil elastase, monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1β (MIP-1β) in supernatants. Serum was collected at baseline, 4, 8 and 24 h for IL-6, C-reactive protein (CRP) and Clara cell protein (CC-16) concentrations. Peripheral blood mononuclear cells (PBMCs) were isolated at baseline and 4 h for systemic IL-17 analysis. LPS 5 μg was well tolerated. The greatest FEV1 change was 11.7% (mean) at 1 h (95% CI 5.1-18.2%). There was a large range in maximal fall (2.5-37.7%). Total sputum cell count and neutrophil count significantly increased 6 and 24 h post-LPS. There was no change in sputum supernatant mediators. IL-6, CRP and CC-16 increased post-inhalation, with different temporal patterns. CD4+ and CD8+ cell associated IL-17 significantly increased at 4 h. Inhaled LPS in COPD patients safely causes increased airway and systemic inflammation. This may be a model for studying COPD exacerbations.

Kinetics of TH2 biomarkers in sputum of asthmatics following inhaled allergen

BackgroundAllergen-induced late airway response offers important pharmacodynamic targets, including T helper 2 (TH2) biomarkers. However, detection of inflammatory markers has been limited in dithiothreitol-processed sputum.ObjectivesTo test whether allergen-induced TH2 inflammatory markers can be reproducibly quantified by sensitive detection techniques in ultracentrifuged sputum and the effect of fluticasone (FP) on these endpoints.MethodsThirteen allergic asthmatics with dual allergen-induced airway responses, documented during a single-blind placebo run-in period, participated in a double-blind, two-period crossover study. Each period consisted of three consecutive days, separated by ≥3 weeks. Following randomization, subjects inhaled FP (500 µg bid, five doses total) or placebo. On Day 2 in each study period, allergen challenge was performed and airway response measured by forced expiratory volume in 1 sec (FEV1) until 7 h post-challenge. Sputum was induced 24 h pre-allergen and 7 and 24 h post-allergen. Sputum samples were split into two portions: TH2 biomarkers were quantified by Meso Scale multiplex platform following ultracentrifugation, and cell differentials were counted on Giemsa–May-Grünwald-stained cytospins. Allergen-induced changes in inflammatory endpoints were compared between FP and placebo using a mixed model ANCOVA.ResultsInhaled allergen induced dual airway responses in all subjects during both placebo periods with reproducible late asthmatic response (LAR) and increased sputum inflammatory biomarkers (IL-2, IL-4, IL-13, and eotaxin-1) and eosinophil counts. FP effectively blunted both the LAR and the inflammatory biomarkers.ConclusionsCombining novel, sensitive quantification methods with ultracentrifugation allows reproducible quantification of sputum biomarkers following allergen challenge, reversed by FP. This approach allows non-invasive identification of pharmacodynamic targets for anti-asthma therapies.

Airway inflammation in Japanese COPD patients compared with smoking and nonsmoking controls.

PurposeTo assess the importance of inflammation in chronic obstructive pulmonary disease (COPD) by measuring airway and systemic inflammatory biomarkers in Japanese patients with the disease and relevant control groups.Patients and methodsThis was the first study of its type in Japanese COPD patients. It was a non-treatment study in which 100 participants were enrolled into one of three groups: nonsmoking controls, current or ex-smoking controls, and COPD patients. All participants underwent standard lung function assessments and provided sputum and blood samples from which the numbers of inflammatory cells and concentrations of biomarkers were measured, using standard procedures.ResultsThe overall trends observed in levels of inflammatory cells and biomarkers in sputum and blood in COPD were consistent with previous reports in Western studies. Increasing levels of neutrophils, interleukin 8 (IL-8), surfactant protein D (SP-D), and Krebs von den Lungen 6 (KL-6) in sputum and clara cell 16 (CC-16), high-sensitivity C-reactive protein (hs-CRP), and KL-6 in serum and plasma fibrinogen were seen in the Japanese COPD patients compared with the non-COPD control participants. In sputum, significant correlations were seen between total cell count and matrix metalloproteinase 9 (MMP-9; P<0.001), neutrophils and MMP-9 (P<0.001), macrophages and KL-6 (P<0.01), total cell count and IL-8 (P<0.05), neutrophils and IL-8 (P<0.05), and macrophages and MMP-9 (P<0.05). Significant correlations were also observed between some inflammatory cells in sputum and biomarkers in serum, with the most significant between serum CC-16 and both total cell count (P<0.005) and neutrophils (P<0.005) in sputum.ConclusionThese results provide evidence for the first time that COPD in Japanese patients is a multicomponent disease, involving both airway and systemic inflammation, in addition to airway obstruction. Therefore, intervention with anti-inflammatory therapy may provide additional benefit in disease management of COPD in Japan.

Neutrophilic Bronchial Inflammation Correlates with Clinical and Functional Findings in Patients with Noncystic Fibrosis Bronchiectasis.

Background. Neutrophilic bronchial inflammation is a main feature of bronchiectasis, but not much is known about its relationship with other disease features. Aim. To compare airway inflammatory markers with clinical and functional findings in subjects with stable noncystic fibrosis bronchiectasis (NCFB). Methods. 152 NFCB patients (62.6 years; females: 57.2%) underwent clinical and functional cross-sectional evaluation, including microbiologic and inflammatory cell profile in sputum, and exhaled breath condensate malondialdehyde (EBC-MDA). NFCB severity was assessed using BSI and FACED criteria. Results. Sputum neutrophil percentages inversely correlated with FEV1 (P < 0.0001; rho = −0.428), weakly with Leicester Cough Questionnaire score (P = 0.068; rho = −0.58), and directly with duration of the disease (P = 0.004; rho = 0.3) and BSI severity score (P = 0.005; rho = 0.37), but not with FACED. Sputum neutrophilia was higher in colonized subjects, P. aeruginosa colonized subjects showing greater sputum neutrophilia and lower FEV1. Patients with ≥3 exacerbations in the last year showed a significantly greater EBC-MDA than the remaining patients. Conclusions. Sputum neutrophilic inflammation and biomarkers of oxidative stress in EBC can be considered good biomarkers of disease severity in NCFB patients, as confirmed by pulmonary function, disease duration, bacterial colonization, BSI score, and exacerbation rate.

Pharmacokinetics and tolerability of oral sildenafil in adults with cystic fibrosis lung disease

RationaleAirway inflammation is central to cystic fibrosis (CF) pathophysiology. Pre-clinical models have shown that phosphodiesterase inhibitors (PDEi) like sildenafil have anti-inflammatory activity. PDEi have not been studied in CF subjects. ObjectivesWe evaluated the pharmacokinetics, tolerability, and safety of sildenafil in subjects with CF. Sputum biomarkers were used to explore efficacy. MethodsAn open-label pilot study of oral sildenafil administration was conducted in adults with mild to moderate CF lung disease. Subjects received oral sildenafil 20 or 40mg p.o. t.i.d. for 6weeks. Measurements and main resultsTwenty subjects completed the study. Estimated elimination rate constants were statistically different in subjects with CF compared to previously published non-CF subjects. Side effects were generally mild. There were no drug-related serious adverse events. Sputum neutrophil elastase activity decreased. ConclusionsSubjects with CF may eliminate sildenafil at a faster rate than non-CF subjects. Sildenafil administration was safe in subjects with CF and decreased sputum elastase activity. Sildenafil warrants further study as an anti-inflammatory in CF.

S132 Sputum And Bronchial Biopsy Expression Of 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxodg) In Asthma Is Related To Neutrophilic Inflammation And Poor Asthma Control

Introduction and objectives Oxidative stress has been implicated in the pathogenesis of asthma. Validated sputum biomarkers are required to assess this and its relationship to other clinical variables. We sought to compare sputum and bronchial 8-oxodG expression in asthma and health; assess the sputum repeatability; and explore its relationship with induced sputum inflammatory cells counts and exacerbations. Methods Asthmatics and healthy controls were recruited from a single centre and underwent clinical characterisation including sputum induction (asthma n = 58, health n = 27) and bronchial biopsy (asthma n = 16, health n = 10). Sputum and epithelial 8-oxodG expression was measured by ELISA and Immunohistochemistry respectively. Sputum asthmatics were assessed at a repeat stable visit at 6 months. Results Between health and asthma, there were no significant differences in the median (IQR) sputum 8-oxodG levels [12 (16) ngml -1 vs. 11 (15) ngml -1 , p = 0.36] or the mean (SEM) percentage area of epithelium reaching threshold intensity for 8-oxodG [2.0 (0.7)% vs. 4.4 (1.0) %, p = 0.12]. Asthma sputum 8-oxodG correlated with the sputum total cell count ( r s =0.53, p r s =0.27, p = 0.04), sputum macrophages ( r s =-0.31, p = 0.02) and serum IgE ( r s =-0.27, p = 0.04). Epithelial 8-oxodG correlated to the number of exacerbations in the previous year (r s =0.70,p s =-0.52,p = 0.04). The upper 95th confidence interval of sputum 8-oxodG and epithelium 8-oxodG reaching threshold in healthy controls was used to split asthma patients into 8-oxodG high and low groups. The sputum 8-oxodG high group (n = 13) had significantly higher sputum total cells 8.08 [8.41] x10 6 g -1 vs. 2.25 [2.91] x10 6 g -1 , p -1 vs. 157 [212.90] KUL -1 , p high group (n = 8) had significantly more exacerbations 3.9 (0.3) vs. 0.5 (0.3) p In the asthmatic group, the intra-class correlation coefficient of sputum 8-oxodG between the 2 visits was 0.51 (p Conclusions 8-oxodG expression in sputum and bronchial biopsies was not different between asthma and health, although we did identify an 8-oxodG high group in asthma. Interestingly, expression in asthma was associated with neutrophilic inflammation and poor asthma control.

Investigating the spill-over hypothesis: Analysis of the association between local inflammatory markers in sputum and systemic inflammatory mediators in plasma

Exposure to air pollutants represents a risk factor not only for respiratory diseases and lung cancer, but also for cardiometabolic diseases. It has been hypothesised that local inflammation in the lung and systemic subclinical inflammation are linked by impaired lung function and the spill-over of proinflammatory factors from the lung into the circulation which could act as intermediaries between environmental exposures and disease risk. We wanted to investigate whether local and systemic inflammatory markers are associated, which would support the spill-over hypothesis.Sputum and plasma samples were obtained from 257 women of the German SALIA cohort. We performed immunoassays to measure multiple biomarkers of airway inflammation in sputum as well as cytokines, chemokines and soluble adhesion molecules in plasma. Correlations were calculated and adjusted for potentially confounding variables.Even though several significant associations were detected between inflammatory mediators in sputum and plasma, correlation coefficients were rather low ranging from r≥−0.20 to r≤0.20. Comparatively stronger associations were observed between nitrite, eosinophil cationic protein, leukotrienes C/D/E4 and interleukin-8 in sputum. Notably, correlations were positive with all proinflammatory biomarkers and interleukin-1 receptor antagonist in plasma, whereas negative correlations were observed with the anti-inflammatory adipokine adiponectin.In conclusion, local inflammation in the lung and systemic subclinical inflammation appear mainly independently regulated in elderly women from the general population. Although we found multiple significant correlations between inflammatory biomarkers in sputum and plasma, our results do not provide clear support for the spill-over hypothesis.

The effect of inhaled IFN-β on worsening of asthma symptoms caused by viral infections. A randomized trial.

Ex vivo, bronchial epithelial cells from people with asthma are more susceptible to rhinovirus infection caused by deficient induction of the antiviral protein, IFN-β. Exogenous IFN-β restores antiviral activity. To compare the efficacy and safety of inhaled IFN-β with placebo administered to people with asthma after onset of cold symptoms to prevent or attenuate asthma symptoms caused by respiratory viruses. A total of 147 people with asthma on inhaled corticosteroids (British Thoracic Society Steps 2-5), with a history of virus-associated exacerbations, were randomized to 14-day treatment with inhaled IFN-β (n = 72) or placebo (n = 75) within 24 hours of developing cold symptoms and were assessed clinically, with relevant samples collected to assess virus infection and antiviral responses. A total of 91% of randomized patients developed a defined cold. In this modified intention-to-treat population, asthma symptoms did not get clinically significantly worse (mean change in six-item Asthma Control Questionnaire <0.5) and IFN-β treatment had no significant effect on this primary endpoint, although it enhanced morning peak expiratory flow recovery (P = 0.033), reduced the need for additional treatment, and boosted innate immunity as assessed by blood and sputum biomarkers. In an exploratory analysis of the subset of more difficult-to-treat, Step 4-5 people with asthma (n = 27 IFN-β; n = 31 placebo), Asthma Control Questionnaire-6 increased significantly on placebo; this was prevented by IFN-β (P = 0.004). Although the trial did not meet its primary endpoint, it suggests that inhaled IFN-β is a potential treatment for virus-induced deteriorations of asthma in difficult-to-treat people with asthma and supports the need for further, adequately powered, trials in this population. Clinical trial registered with www.clinicaltrials.gov (NCT 01126177).

Identification of ENO1 As a Potential Sputum Biomarker for Early-Stage Lung Cancer by Shotgun Proteomics

BackgroundLung cancer is the leading cancer killer. Early detection will reduce the related deaths. The objective of this study was to identify potential biomarkers for early-stage lung cancer in sputum supernatant. Materials and MethodsUsing shotgun proteomics, we detected changes in protein profiles that were associated with lung cancer by analyzing sputum supernatants from 6 patients with early-stage lung cancer and 5 cancer-free controls. Using western blotting, we validated the proteomic results in 22 lung cancer cases and 22 controls. Using enzyme-linked immunosorbent assay (ELISA), we evaluated the diagnostic performance of the biomarker candidates in an independent set of 35 cases and 36 controls. ResultsProteomics identified 8 biomarker candidates for lung cancer. Western blotting validation of the candidates showed that enolase 1 (ENO1) displayed a higher expression level in patients with cancer than in cancer-free individuals (P = .015). ELISA revealed that the assessment of ENO1 expression in sputum supernatant had 58.33% sensitivity and 80.00% specificity in distinguishing patients with stage I lung cancer from cancer-free individuals. ConclusionThe analysis of protein biomarkers in sputum may provide a potential approach for the early detection of lung cancer. Future validation of all the candidates defined by shotgun proteomics in a large cohort study may help develop additional biomarkers that can be added to ENO1 to provide more diagnostic efficacy for lung cancer.

227 Detecting early markers of CF-related pulmonary exacerbations using home telemetry and sputum biomarkers

227 Detecting early markers of CF-related pulmonary exacerbations using home telemetry and sputum biomarkers

Association of pro-inflammatory cytokines, cortisol and depression in patients with chronic obstructive pulmonary disease.

Evidence suggests that pro-inflammatory cytokines and cortisol play a crucial role in the etiology of chronic obstructive pulmonary disease (COPD) and depression. Depression occurs commonly among COPD patients and an earlier diagnosis would be beneficial. This study investigated the associations between depression, sputum cytokines and salivary cortisol in COPD patients. The diurnal rhythms of sputum IL-1, IL-6, TNF-α and salivary cortisol were measured in COPD patients with depression compared to those only with depression, or COPD and healthy controls. The area under the diurnal variation curves (AUC) over the 24h time course and relative diurnal variation (VAR) were calculated while correlation and regression analysis were performed. Patients with co-morbid depression and COPD showed an increasing sputum IL-1, sputum TNF-α AUC and a decreasing salivary cortisol VAR (P<0.001). The combination of sputum TNF-α AUC, sputum IL-1 AUC, sputum IL-6 AUC and salivary cortisol VAR performed best as a potential biomarker in the diagnosis of depression in COPD patients, with a sensitivity of 94.74% and a specificity of 96.67%. Positive correlations were found between sputum IL-1 AUC and sputum TNF-α AUC versus depressive symptoms, respectively a negative correlation was found between salivary cortisol VAR and depression. They were independently associated with depression in logistic regression models. Depression in COPD is associated with higher 24-h overall levels of sputum IL-1, TNF-α and flattened diurnal salivary cortisol. These non-invasive sputum and salivary biomarkers may serve as a simple clinical tool for the early diagnosis of depression in COPD patients.

Rapid diagnosis of tuberculosis using ex vivo host biomarkers in sputum

To the Editor: Tuberculosis continues to be a major public health problem in developing countries [1]. One of the roadblocks in reducing tuberculosis transmission is the lack of accurate laboratory-free diagnostic tests for use at the point of care. If tuberculosis is to be eliminated, we need a robust, low-cost and safe point-of-care diagnostic test, which in turn requires identification of appropriate biomarkers [2]. Rapid tests based on microfluidics (lateral flow tests) hold great promise for tuberculosis diagnostics. They are easy to use, cheap, provide an answer within minutes, do not require specialised equipment and are stable at room temperature, making them ideal for use in high tuberculosis burden, resource-poor settings. To date, however, no such test has been developed for tuberculosis due to lack of sensitivity related to the markers and/or sample type. Development of tests based on host biomarkers requires evaluation of different sample types [3]–[5] and markers other than interferon (IFN)-γ [5] to provide differential diagnosis of active tuberculosis, latent infection and other respiratory disorders. We have previously shown that a combination of three host factors in pleural fluid resulted in 96% correct classification of tuberculosis among other respiratory diseases (ORD) (including bacterial pneumonia) regardless of HIV status [6]. However, this sample type is not easy to obtain and we therefore wanted to determine if we could use ex vivo sputum, which is noninvasive and easy to obtain in adult pulmonary tuberculosis patients. Subjects were consecutively recruited from the outpatient clinic and ward at the Medical Research Council Unit, Fajara, the Gambia. All subjects were adults (≥18 years of age) with symptoms suggestive of tuberculosis. Subjects were subsequently classified into two groups: those with culture-confirmed tuberculosis and those with ORD. 75% of the tuberculosis and …