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Identification of Lipocalin and Apolipoprotein A1 as Biomarkers of Chronic Obstructive Pulmonary Disease
Abstract and Figures
Much effort is being made to discover noninvasive biomarkers of chronic airway disease that might enable better management, predict prognosis, and provide new therapeutic targets.
To undertake a comprehensive, unbiased proteomic analysis of induced sputum and identify novel noninvasive biomarkers for chronic obstructive pulmonary disease (COPD).
Induced sputum was obtained from patients with COPD with a spectrum of disease severity and from control subjects. Two-dimensional gel electrophoresis and mass spectrometric identification of differentially expressed proteins were first applied to induced sputum from patients with GOLD stage 2 COPD and healthy smoker control subjects. Initial results thus obtained were validated by a combination of immunoassays (Western blotting and ELISA) applied to a large subject cohort. The biomarkers were localized to bronchial mucosa by immunohistochemistry.
Of 1,325 individual protein spots identified, 37 were quantitatively and 3 qualitatively different between the two groups (P < 0.05%). Forty protein spots were subjected to tandem mass spectrometry, which identified 15 separate protein species. Seven of these were further quantified in induced sputum from 97 individuals. Using this sequential approach, two of these potential biomarkers (apolipoprotein A1 and lipocalin-1) were found to be significantly reduced in patients with COPD when compared with healthy smokers. Their levels correlated with FEV(1)/FVC, indicating their relationship to disease severity.
A potential role for apolipoprotein A1 and lipocalin-1 in innate defense has been postulated previously; our discovery of their reduction in COPD indicates a deficient innate defense system in airway disease that could explain increased susceptibility to infectious exacerbations.
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... 12 For example, Nicholas et al collected sputum samples from 15 patients with COPD and 18 healthy smokers, and used twodimensional gel electrophoresis and mass spectrometric identification techniques to identify 40 DEPs between the two groups. 13 Such studies analyzing proteomics and peptide profiles in serum samples obtained from patients with COPD may identify biomarkers linked to the occurrence and development of the disease. Liquid chromatography tandem mass spectrometry (LC-MS/MS) is a powerful proteomics methods that can identify many differentially expressed proteins (DEPs) with high sensitivity and specificity. ...
... The expression of APOA-I (apolipoprotein A-I) was greatly decreased in the specimens from patients with COPD. 13 Li et al used enzyme linked immunosorbent assays to determine the levels of ApoM and pentraxin-3 in 110 patients with COPD and 110 controls. In addition, they used automated biochemical analyzers to assess the levels of C-reactive protein, cholesterol, and triglycerides. ...
Purpose:
As a common respiratory disease, chronic obstructive pulmonary disease (COPD) has a high morbidity and mortality. Current clinical therapies are not ideal and do not improve lung function or long-term life quality. It is very important to find new potential pathogenic mechanisms, biomarkers, and targets with therapeutic value in COPD.
Methods:
Serum samples collected from acute exacerbation and stable COPD and healthy participants were analyzed using label-free liquid chromatography tandem mass spectrometry to identify the differentially expressed proteins (DEPs) between two groups. Bioinformatics analyses were performed to determine the biological processes associated with those DEPs. Key proteins were validated by enzyme linked immunosorbent assay (ELISA).
Results:
In total, 661 proteins were detected in serum from patients with COPD and healthy participants. Compared with healthy participants, patients with acute exacerbation of COPD had 45 DEPs, 13 were upregulated and 32 were downregulated; and patients with stable COPD had 79 DEPs, 18 were upregulated and 61 were downregulated. Gene Ontology functional annotation results indicated that the DEPs identified in patients with COPD were associated with the terms cellular anatomical entity, binding, and cellular process. Kyoto Encyclopedia of Genes and Genomes functional annotation analysis and the Clusters of Orthologous Genes database analysis indicated that the functions of these DEPs were primarily in signal transduction mechanisms and amino acid transport and metabolism. The ELISA results for three key proteins of IGFBP2, LRG1 and TAGLN were consistent with the LC-MS/MS results and the area under the receiver operating characteristic of the combined index was 0.893 (95% CI: 0.813, 0.974).
Conclusion:
Our findings suggested pathogenic mechanisms underlying COPD stages and indicated three key proteins that may warrant further study as potential biomarkers for early diagnosis or prognosis of COPD or as therapeutic targets.
... The normal distribution was checked using the Kolmogorov-Smirnov criterion. Due to the nonparametric distribution of the studied indicators, the data are presented for continuous variables in the form of Me [25,75], where Me is the median and 25 and 75 are the first and third quartiles in the case of categorical variables in the form of absolute and relative values: n (%). The nonparametric Mann-Whitney U-test was used to compare two independent samples. ...
... There have been practically no studies of this biomarker in patients with bronchitis. There is evidence of a decrease in the level of lipocalin-2 in patients with COPD compared with healthy smokers [25]. In a small study performed among people with chronic cough (more than 1 month), there were no differences in the level of lipocalin-2 compared to the control group [26]. ...
The pathogenesis of the development of chronic lung diseases assumes the participation of systemic inflammation factors, as well as hormone-like substances produced by adipose tissue. The aim of this study was to evaluate the associations of certain adipokines/cytokines and chronic bronchitis against the background of abdominal obesity in young people. The study included 1415 people aged 25−44. In total, 115 people were selected by the random numbers method, who were divided into two subgroups: those with chronic bronchitis and abdominal obesity and those with chronic bronchitis without abdominal obesity. A control group of patients with comparable gender and age was also selected. In the group of patients with chronic bronchitis, adiponectin, TNFa and GIP levels were 1.4 times higher. The levels of C-peptide, MCP-1 and PP in the group of chronic bronchitis were 1.3 times higher compared to the control. Adipsin, lipocalin-2, IL-6 and resistin were significantly higher in the group with chronic bronchitis. Glucagon, amylin and ghrelin were 2.2, 2.3 and 3.2 times lower, respectively, in the group of patients with chronic bronchitis. Against the background of abdominal obesity, the probability of having chronic bronchitis increased with an increase in the level of lipocalin-2 and GIP and TNFa.
... In this study, except S100A6, there were no differentially expressed transcripts between the former smoker and never smoker groups (Titz et al., 2015). Nicholas et al. (2010) used a proteomic approach to compare novel noninvasive biomarkers using induced sputum in GOLD stage 2 COPD and healthy smoker control subjects. In this study, the authors used a sequential approach of identifying 1,325 individual protein spots in which 37 were quantitatively and 3 qualitatively different between the two groups. ...
... Using this approach, apolipoprotein A1 and lipocalin-1 were potential biomarkers and reduced in patients with COPD when compared with healthy smokers (Table 2). Immunohistochemistry revealed that these two markers were localized to bronchial mucosa (Nicholas et al., 2010). Hoonhorst et al. (2016) assessed the association of AGEs and sRAGE in plasma, sputum, bronchial biopsies, and skin and found no differences in sputum sRAGE between smokers and COPD groups. ...
Biomarkers of potential harm (BoPH) are indicators of biological perturbations which may contribute to the pathophysiology of disease. In this review, we critically assessed the published data on lung-related BoPH in human lung disease for potential use in evaluating the effects of tobacco and nicotine products. A Scopus literature search was conducted on lung disease biomarkers used in a clinical setting over the last 10 years. We identified 1171 papers which were further screened using commercial software (Sciome SWIFT-Active Screener) giving 68 publications that met our inclusion criteria (data on the association of the biomarker with cigarette smoking, the impact of smoking cessation on the biomarker, and differences between smokers and non-smokers), the majority of which investigated chronic obstructive pulmonary disease. Several physiological and biochemical measures were identified that are potentially relevant for evaluating the impact of tobacco products on lung health. Promising new candidates included blood biomarkers, such as surfactant protein D (SP-D), soluble receptor for advanced glycation end products (sRAGE), skin autofluorescence (SAF), and imaging techniques. These biomarkers may provide insights into lung disease development and progression; however, all require further research and validation to confirm their role in the context of tobacco and nicotine exposure, their time course of development and ability to measure or predict disease progression.
... Sputum induction and processing was performed as previously reported (13) at least 4 weeks after a respiratory tract infection. The mucous elements were selected from the expectorated sample and treated with 5 mM dithioerythritol in HEPES-buffered saline (HBS) to liquefy the mucus and obtain sputum cell cytospin slides as previously described (13). Differential cell counts were obtained using the rapid Romanowski method. ...
Asthmatics are more susceptible to viral infections than healthy individuals and are known to have impaired innate anti-viral defences. Influenza A virus causes significant morbidity and mortality in this population. Immuno-modulatory regulators (IMRs) such as PD-1 are activated on T cells following viral infection as part of normal T cell activation responses, and then subside, but remain elevated in cases of chronic exposure to virus, indicative of T cell exhaustion rather than activation. There is evidence that checkpoint inhibition can enhance anti-viral responses during acute exposure to virus through enhancement of CD8+T cell function. Although elevated PD-1 expression has been described in pulmonary tissues in other chronic lung diseases, the role of IMRs in asthma has been relatively unexplored as the basis for immune dysfunction. We first assessed IMR expression in the peripheral circulation and then quantified changes in IMR expression in lung tissue in response to ex-vivo influenza infection. We found that the PD-1 family members are not significantly altered in the peripheral circulation in individuals with severe asthma but are elevated in pulmonary tissues following ex-vivo influenza infection. We then applied PD-1 Mab inhibitor treatment to bronchial biopsy tissues infected with influenza virus and found that PD-1 inhibition was ineffective in asthmatics, but actually increased infection rates in healthy controls. This study, therefore, suggests that PD-1 therapy would not produce harmful side-effects when applied in people with severe asthma, but could have important, as yet undescribed, negative effects on anti-viral responses in healthy individuals that warrant further investigation.
... The relatively high representation of immunomodulators among the candidate agents and the increased centrality of fibrosis-related proteins is consistent with the paradigm of airway remodeling as central to COPD pathology [51]. With additional data, this regulatory circuit could be used as a testbed for computational evaluation of these and other candidate drug effects using network topological methods [52]. ...
Bronchoalveolar lavage of the epithelial lining fluid (BALF) can sample the profound changes in the airway lumen milieu prevalent in chronic obstructive pulmonary disease (COPD). We compared the BALF proteome of ex-smokers with moderate COPD who are not in exacerbation status to non-smoking healthy control subjects and applied proteome-scale translational bioinformatics approaches to identify potential therapeutic protein targets and drugs that modulate these proteins for the treatment of COPD. Proteomic profiles of BALF were obtained from (1) never-smoker control subjects with normal lung function (n = 10) or (2) individuals with stable moderate (GOLD stage 2, FEV1 50–80% predicted, FEV1/FVC < 0.70) COPD who were ex-smokers for at least 1 year (n = 10). After identifying potential crucial hub proteins, drug–proteome interaction signatures were ranked by the computational analysis of novel drug opportunities (CANDO) platform for multiscale therapeutic discovery to identify potentially repurposable drugs. Subsequently, a literature-based knowledge graph was utilized to rank combinations of drugs that most likely ameliorate inflammatory processes. Proteomic network analysis demonstrated that 233 of the >1800 proteins identified in the BALF were significantly differentially expressed in COPD versus control. Functional annotation of the differentially expressed proteins was used to detail canonical pathways containing the differential expressed proteins. Topological network analysis demonstrated that four putative proteins act as central node proteins in COPD. The drugs with the most similar interaction signatures to approved COPD drugs were extracted with the CANDO platform. The drugs identified using CANDO were subsequently analyzed using a knowledge-based technique to determine an optimal two-drug combination that had the most appropriate effect on the central node proteins. Network analysis of the BALF proteome identified critical targets that have critical roles in modulating COPD pathogenesis, for which we identified several drugs that could be repurposed to treat COPD using a multiscale shotgun drug discovery approach.
... As noted above, Apo A1is a structural protein of the HDL and has numerous protective effects on the healthy lung and pulmonary diseases (24). It was observed that ApoA1 and lipocalin-1 present in the sputum of patients with COPD significantly decreased compared with those of the healthy smokers (28). ApoA1 ...
Background
Chronic obstructive pulmonary disease (COPD) is a major health issue worldwide.
Introduction
This study aimed to evaluate the effect of aerobic exercises at steep surfaces on Apo A1 and Apo B serum levels and their ratio in COPD patients.
Methods
This interventional study was undertaken on 16 COPD patients. The sample was selected by using the random sampling method. Patients were randomly divided into 2 groups of 8 members: uphill and downhill. Spirometry was conducted prior to the exercises and pulmonary volumes were measured. Aerobic exercises (stationary bicycle and treadmill) were started 3 times a week for 8 weeks. The duration of exercises in each session was determined based on the patients’ ability (30-40 min). In the first and second group, exercises were performed on sloped surfaces that were downwards and upwards at a 10-degree angle, respectively. Blood samples were taken from the patients before and after the 8 weeks. Apo A1 and Apo B serum levels and Apo A1 to Apo B ratio were measured. Data were analyzed by SPSS software. P<0.05 was considered statistically significant.
Results
ApoA1/Apo B ratio in the uphill group before and after the intervention was 1.43±0.21 and 1.53±0.24 mg/dL, respectively, with P-value=0.36. In the downhill group, this value was 1.27±0.17 and 1.30±0.18, respectively, with a P-value=0.032.
Conclusion
In light of the results of the present study, incorporating downhill exercises in COPD patients' rehabilitation program can help improve pulmonary function and prevent atherosclerotic events.
Aim
Respiratory disease (RD) is one of the most common diseases characterized by lung dysfunction. Many diagnostic mechanisms have been used to identify the pathogenic agents of responsible for RD. Among these, proteomics emerges as a valuable diagnostic method for pinpointing the specific proteins involved in RD pathogenesis. Therefore, in this study, for the first time, we examined the protein markers involved in the pathogenesis of chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF), asthma, bronchiolitis obliterans (BO), and chemical warfare victims exposed to mustard gas, using the proteomics method as a systematic study.
Materials and Methods
A systematic search was performed up to September 2023 on several databases, including PubMed, Scopus, ISI Web of Science, and Cochrane. In total, selected 4246 articles were for evaluation according to the criteria. Finally, 119 studies were selected for this systematic review.
Results
A total of 13,806 proteins were identified, 6471 in COPD, 1603 in Asthma, 5638 in IPF, three in BO, and 91 in mustard gas exposed victims. Alterations in the expression of these proteins were observed in the respective diseases. After evaluation, the results showed that 31 proteins were found to be shared among all five diseases.
Conclusion
Although these 31 proteins regulate different factors and molecular pathways in all five diseases, they ultimately lead to the regulation of inflammatory pathways. In other words, the expression of some proteins in COPD and mustard‐exposed patients increases inflammatory reactions, while in IPF, they cause lung fibrosis. Asthma, causes allergic reactions due to T‐cell differentiation toward Th2.
A wealth of evidence indicates that high-density lipoprotein assumes the unique antiatherosclerosis and other cardioprotective properties. Based on that, HDL-C has been considered as a promising therapy target to reduce the cardiovascular diseases. Recombinant HDL (rHDL) and apolipoprotein mimetic peptides emerge in recent years and have great potential in the future. Here we discussed the pleiotropic therapeutic effect of rHDL based on the effects of atherogenic, angiogenesis, platelet, vascular, and Alzheimer's disease. On the other hand, rHDL not only plays the key role as the major protein component of HDL, it is also used as a nanovector in antiatherosclerotic, antitumor, cardiovascular diagnosing and other therapeutic areas. Synthetic apolipoprotein mimetic peptides like apoA-I and and apoE mimetics have undergone clinical assessment, and we have also reviewed the advances of clinical trials and gave an outlook for the therapy of rHDL and mimetic peptides.
Rationale
Bronchoalveolar lavage of the epithelial lining fluid can sample the profound changes in the airway lumen milieu prevalent in Chronic Obstructive Pulmonary Disease (COPD). Characterizing the proteins in bronchoalveolar lavage fluid in COPD with advanced proteomic methods will identify disease-related changes, provide insight into pathogenetic mechanisms and potential therapeutics that will aid in the discovery of more effective therapeutics for COPD.
Objectives
We compared epithelial lining fluid proteome of ex-smokers with moderate COPD who are not in exacerbation status COPD, to non-smoking healthy control subjects using advanced proteomics methods and applied proteome-scale translational bioinformatics approaches to identify potential therapeutic protein targets and drugs that modulate these proteins towards the treatment of COPD.
Methods
Proteomic profiles of bronchalveolar lavage fluid were obtained from 1) never-smoker control subjects with normal lung function (n=10) or 2) individuals with stable moderate (GOLD stage 2, FEV1 50% – 80% predicted) COPD who were ex-smokers for at least one year (n=10). NIH’s Database for Annotation, Visualization and Integrated Discovery (DAVID) and Ingenuity’s Ingenuity Pathway Analysis (IPA) were the two bioinformatics tools employed for network analysis on the differentially expressed proteins to identify potential crucial hub proteins. The drug-proteome interaction signature comparison and ranking approach implemented in the Computational Analysis of Novel Drug Opportunities (CANDO) platform for multiscale therapeutic discovery was utilized to identify potential repurposable drugs for the treatment of COPD based on the BALF proteome. Subsequently, a literature-based knowledge graph was utilized to rank combinations of drugs that would most likely ameloriate inflammatory processes by inhibition or activation of their functions.
Results
Proteomic network analysis demonstrated that 233 of the >1800 proteins identified in the BALF were differentially expressed in COPD versus control, including proteins associated with inflammation, structural elements, and energy metabolism. Functional annotation of the differentially expressed proteins by their implicated biological processes, cellular localization, and transcription factor interactions was accomplished via DAVID. Canonical pathways containing the differential expressed proteins were detailed via the Ingenuity Pathway Analysis application. Topological network analysis demonstrated that four proteins act as central node proteins in the inflammatory pathways in COPD. The CANDO multiscale drug discovery platform was used to analyze the behavioral similarity between the interaction signatures of all FDA-approved drugs and the identified BALF proteins. The drugs with the signatures most similar interaction signatures to approved COPD drugs were extracted with the CANDO platform. The analysis revealed 189 drugs that putatively target the proteins implicated in COPD. The putative COPD drugs that were identified using CANDO were subsequently analyzed using a knowledge based technique to identify an optimal two drug combination that had the most appropriate effect on the central node proteins.
Conclusion
Analysis of the BALF proteome revealed novel differentially expressed proteins in the epithelial lining fluid that elucidate COPD pathogenesis. Network analyses identified critical targets that have critical roles in modulating COPD pathogenesis, for which we identified several drugs that could be repurposed to treat COPD using a multiscale shotgun drug discovery approach.
Asthma is a disease of complex origin and multiple pathologies. There are currently very few biomarkers of proven utility in its diagnosis, management or response to treatment. Recent studies have identified multiple asthma phenotypes following biofluid analysis; however, such findings may be driven by the well-characterised alterations in immune cell populations in asthma. We present a study designed to identify cell type-specific gene signatures of severe allergic asthma in peripheral blood samples. Using transcriptomic profiling of four magnetically purified peripheral blood cell types, we identify significant gene expression changes in monocytes and NK cells but not T lymphocytes in severe asthmatics. Pathway analysis indicates dysfunction of immune cell regulation and bacterial suppression in the NK cells. These gene expression changes may be useful on their own as prognostic peripheral blood cell markers of severe asthma, but also may indicate novel cell pathways for therapeutic intervention.
Objective.
—To determine whether a program incorporating smoking intervention and use of an inhaled bronchodilator can slow the rate of decline in forced expiratory volume in 1 second (FEV1) in smokers aged 35 to 60 years who have mild obstructive pulmonary disease.
Chronic obstructive pulmonary disease (COPD) is an inflammatory disease characterized by the progressive deterioration of pulmonary function and irreversible airway obstruction. Investigations of the molecular pathogenesis of COPD have not yet provided complete answers to the mechanisms that determine the onset and progression of this illness. Therefore, therapeutic choices are limited and new strategies are needed to prevent, manage and treat this disorder. In particular, the application of complementary approaches, including gel- and liquid chromatography mass spectrometry-based proteomic techniques on sputum and/or bronchoalveolar lavage may provide a better understanding of the proteome differentially expressed by COPD patients in the course of the disease. The identification of appropriate and reliable biomarkers is, thus, an essential step for the diagnostics and treatment of these patients.
A prospective epidemiological study of the early stages of the development of chronic obstructive pulmonary disease was performed on London working men. The findings showed that forced expiratory volume in one second (FEV1) falls gradually over a lifetime, but in most non-smokers and many smokers clinically significant airflow obstruction never develops. In susceptible people, however, smoking causes irreversible obstructive changes. If a susceptible smoker stops smoking he will not recover his lung function, but the average further rates of loss of FEV1 will revert to normal. Therefore, severe or fatal obstructive lung disease could be prevented by screening smokers' lung function in early middle age if those with reduced function could be induced to stop smoking. Infective processes and chronic mucus hyper-secretion do not cause chronic airflow obstruction to progress more rapidly. There are thus two largely unrelated disease processes, chronic airflow obstruction and the hypersecretory disorder (including infective processes).
Immunoglobulin binding factor (IgBF) is known to bind immunoglobulin, to interact with anti-Fc gamma-III antibodies and to be present in the lower respiratory tract of normal healthy subjects. In this study, in order to clarify the role of IgBF in respiratory diseases, we investigated whether IgBF exists in the airway of patients with chronic airway diseases.
IgBF was measured in the sputum of 28 normal subjects and 59 patients with chronic airway diseases including 37 cases of chronic bronchitis, 18 bronchiectasis, and four diffuse panbronchiolitis by enzyme-linked immunosorbent assay.
Immunoglobulin binding factor concentration in the sputum of patients with chronic airway diseases (purulent sputum, 50.2 +/- 8.2 microg/mL; mucoid sputum, 88.6 +/- 12.8 microg/mL) was higher than that in induced sputum of normal subjects (6.3 +/- 5.5 microg/mL; P < 0.001). Immunoglobulin binding factor level in mucoid sputum was significantly higher than that in purulent sputum (P < 0.05). A significant inverse correlation was shown between the IgBF level and the elastase activity in sputum, and the concentration of IgBF purified from seminal plasma was decreased by treatment with neutrophil elastase, indicating that IgBF might be degraded by elastase. In the gel filtration chromatography of both types of sputum, IgBF was eluted in a region corresponding to a molecular weight of 27 kDa as a single peak. Western blot analysis with a monoclonal antibody to IgBF indicated that IgBF in both types of sputum had a molecular weight of 27 kDa under non-reducing conditions and of 16 kDa under reducing conditions.
These results demonstrate that a high level of IgBF is present in the respiratory tract of patients with chronic airway diseases and may be related to the pathogenesis of these diseases.
Lipocalins are carrier proteins for hydrophobic molecules in many biological fluids. In the oral sphere (nasal mucus, saliva, tears), they have an environmental biosensor function and are involved in the detection of odours and pheromones. Herein, we report the first identification of human lipocalins involved in odorant binding. They correspond to a gene family located on human chromosome 9q34 produced by genomic duplications: two new odorant-binding protein genes ( hOBP (IIa) and hOBP (IIb) ), the previously described tear lipocalin LCN1 gene and two new LCN1 pseudogenes. Although 95% similar in sequence, the two hOBP (II) genes were differentially expressed in secretory structures. hOBP (IIa) was strongly expressed in the nasal structures, salivary and lachrymal glands, and lung, therefore having an oral sphere profile. hOBP (IIb) was more strongly expressed in genital sphere organs such as the prostate and mammary glands. Both were expressed in the male deferent ducts and placenta. Surprisingly, alternatively spliced mRNAs resulting in proteins with different C-termini were generated from each of the two genes. The single LCN1 gene in humans generated a putative odorant-binding protein in nasal structures. Finally, based on the proposed successive genomic duplication history, we demonstrated the recruitment of exons within intronic DNA generating diversity. This is consistent with a positive selection pressure in vertebrate evolution in the intron-late hypothesis.
COPD is a heterogeneous collection of conditions that can affect various structures within the lung in a number of different ways. These various processes can all result in limitation of expiratory airflow. If severe enough, this physiologic abnormality defines COPD. The various conditions that can lead to this syndrome are prevalent and often relentlessly progressive. In aggregate, they represent an important public health problem. This supplement outlines diagnostic and therapeutic strategies by which the practitioner can assist patients suffering from this condition.