The Exploration Of Protein Chip In The Classification And Evaluation Of Chronic Obstructive Pulmonary Disease

BackgroundChronic obstructive pulmonary disease(COPD),or Chronic obstructive pulmonary disease(COPD),is a common and frequent,preventable and treatable respiratory disease worldwide and the third leading cause of death globally by far,imposes a huge burden on society and the economy.At present,the diagnosis of COPD is mainly depends on pulmonary function examination,which plays a core role in the diagnosis of COPD and grading of airflow restriction.However,the use of lung function indicators alone in disease reflection and prognosis inference is not comprehensive,nor can it well reflect the heterogeneity of COPD,and requires patients to have a high degree of cooperation Therefore,seeking more biomarkers to reflect the severity of COPD disease and evaluate the treatment response horizontally,reflect the disease progression and predict the clinical outcome vertically,and further distinguish the specific phenotype is the premise of realizing the personalized medicine of COPD.In recent years,more and more studies have been conducted to explore the role of protein microarray in the pathogenesis,disease evaluation,phenotypic biomarker characteristics,differential diagnosis and drug therapy of COPD.However,However,the existing studies on protein microarray in COPD have some problems,such as small sample size and less number of screened protein markers;For the studies on the characteristics of different subtypes of COPD,the biomarker characteristics of airflow restriction,acute exacerbation risk and other phenotypes have not been comprehensively analyzed.The present study has some limitations and cannot fully interpret the dynamic changes of COPD progression.ObjectiveThe comprehensive value of discovery protein microarray in identifying biomarkers,reflecting phenotypic characteristics,assessing disease conditions and predicting clinical deterioration in COPD provides a research basis for personalized medicine of COPD.Subjects and MethodsA prospective observational cohort study was used in this study.Object of study:Normal subjects and patients with chronic obstructive pulmonary disease who met the GOLD2017 diagnostic criteria were included in the cohort between January 2016 and May 2022.According to inclusion and exclusion criteria,a total of 51 normal subjects and 148 COPD subjects were included as healthy controls.Clinical data:baseline data of subjects were collected:gender,age,body mass index(BMI),number of smoking pack years,current smoking status,respiratory distress questionnaire score(mMRC),CAT score,number of acute exacerbations in the past 1 year,risk of acute exacerbations(low or high risk),lung function and chest HRCT were examined,sputum cytological classification and emphysema index were collected Subjects were followed up for 1 year after baseline,including mMRC and CAT scores for lung function after 1 year of follow-up for acute exacerbations.Protein chip assay for biomarkers:Sputum supernatant and serum from each subject were collected at baseline.After pretreatment of sputum and serum,280 protein biomarkers,including inflammatory factors,chemokines,growth factor receptor cytokines,were detected using protein chip assay.Statistical analysis:Comparative analysis of the demographic characteristics of normal subjects and COPD subjects;The expression of interprotein factors of different phenotypes in COPD subjects was analyzed,and the correlation analysis was conducted with various clinical indicators.ROC curve was used to calculate the AUC,and the sensitivity and specificity of differential proteins among different COPD phenotypes in different populations were evaluated.Multivariate logistics regression was used to analyze the statistical results of biomarkers with independent predictive value for COPD clinical deterioration in the next one year.Results1.Demographic and clinical characteristics of baseline subjects:The mean age of COPD subjects(67.0 ± 8.3)was higher than that of normal subjects(58.6 ±9.9);Most of them were male(93.9%vs 68.6%).The number of pack years of smoking increased significantly compared with normal subjects(P<0.001).Compared with normal subjects,total CAT mMRC score and%LAA-950 were increased in COPD subjects;Lung function indicators:post-diastolic FEV1%pred,post-diastolic PEF,post-diastolic MMEF,post-diastolic MEF50 and MEF25 were decreased(P<0.001).2.Differential protein markers between COPD and normal subjects:42 differential proteins were detected in sputum,and ROC showed an AUC of 0.992.Five differential proteins were detected in serum,and ROC showed an AUC of 0.784,indicating that protein markers in both sputum and serum of COPD patients were significantly different from those of healthy subjects.3.Differential protein markers in sputum may better reflect the difference in protein markers expression of different COPD phenotypes3.1 Difference in phenotype protein expression between severely airflow restricted and non-severely airflow restricted COPD group:Compared with the non-severely airflow restricted group,the levels of inflammatory factor IL-6,IL-10,MIP-1a,growth factor HGF and cytokine LAP(TGFb1)，MMP-1 in sputum of severely airflow restricted group were significantly increased;The chemokine Axl,I-TAC,CTACK,MSP,MCP-4,LIF,IL-18,BPa,IL-31 growth factor HB-EGF,receptor RAGE and the cytokine CD9 7,galectin-7,MMP-10,hCGb,IL-17B,BMP-9,FGF-19 decreased significantly(P<0.001).Both chemokine MCP-4 and cytokine galectin-3 were significantly increased in serum(P<0.001)and the AUC of sputum protein markers was 0.880,and that of serum protein markers was 0.634,suggesting that protein markers in sputum were more able to significantly distinguish the phenotypes of COPD with different degree of airflow restriction.3.2 Protein expression differences between low-risk and high-risk COPD phenotypes in acute exacerbations:IL-10 growth factor HGF was significantly increased in high-risk COPD subjects compared with low-risk subjects in acute exacerbations;Chemokine MCP-3 increased significantly,while MSP,Axl decreased significantly.Both MMP-13 and MMP-1,cytokine LAP(TGFb1)were significantly increased,while CD97 MMP-10 and receptor RAGE were significantly decreased(P<0.001);The AUC of these protein markers was 0.856,suggesting that the sputum protein profile could significantly differentiate the risk of COPD exacerbation.3.3 Protein expression difference between COPD with emphysema and non-emphysema:Compared with non-emphysema,chemokine I-TAC was significantly decreased in sputum of COPD with emphysema,while growth factor HGF and growth factor HGF were significantly increased;In serum,inflammatory factor MIG,chemokine OPN and cytokine galectin-3 were significantly increased(P<0.001).The AUC of protein markers in sputum was 0.703 and that in serum was 0.733,suggesting that there were significant differences in protein expression profiles in sputum and serum of COPD subjects with different imaging phenotypes.3.4 Differential expression of protein markers in COPD subjects with different sputum cytological classifications:There were statistically significant differences in the sputum of COPD subjects with different sputum cytological classifications of chemokine MDC,MSP and receptor VCAM-1(P<0.05).4.Correlation between COPD protein markers and clinical characteristics:Among COPD protein markers in subjects,(1)mMRC score:HGF,MIP-1A,LAP(TGFb1),IL-10,LIGHT in sputum and MCP-4 in serum MIG was positively correlated with mMRC score,while MMP-10,RAGE,Axl,Galectin-7,CD97 was negatively correlated with mMRC score in sputum(P<0.05);(2)Total CAT score:IL-10 in sputum and OPN,MCP-4 in serum were positively correlated with total CAT score,while MMP-10,ANG-2,RAGE,Axl,Galectin-7,CD97,IL-17B in sputum were negatively correlated with total CAT score(P<0.05);(3)In sputum,APRIL,MMP-10,RAGE,MSP,Axl,Galectin-7,FGF-19,CD97,hCGb,IL-17B was positively correlated with post-diastolic FEV1%Pred,while in sputum,HGF,MIP-1A,LAP(TGFb1),IL-10,IL-6,Mmp-1 and serum OPN,MCP-4 were negatively correlated with postdiastolic FEV1%Pred(P<0.05).(4)%LAA-950,HGF,IL-10,MMP-1 in sputum and OPN,MCP-4,MIG in serum were positively correlated with%LAA-950;However,MMP-10,ANG-2,RAGE,MSP,Axl,Galectin-7,VCAM-1,CD97 and serum IL-23 were negatively correlated with%LAA-950(P<0.05);(5)FeNO:HGF,MIP-1A,MMP-1 in sputum was positively correlated with FeNO(P<0.05);(6)Blood Eos absolute count:serum MCP-4,MIG was positively correlated with blood Eos absolute count(P<0.05).5.COPD protein markers predicted that after 1 year of follow-upFEV1 was lower than baseline and CAT total score was higher than baseline(P<0.05),suggesting that progressive exacerbation of lung function and symptoms was associated with significant clinical exacerbation 1 year later.The results of univariate analysis showed that protein biomarkers for severe acute exacerbation 1 year after baseline in COPD subjects included receptor RAGE and cytokine MMP-10,CD97.The protein markers that affected the increase of CAT scores 1 year after baseline were the chemokine MCP-3 in sputum and the cytokine IL-23 in serum.The Multivariate analysis of serum chemokine OPN for significant clinical exacerbations after 1 year showed that baseline postdiastolic FEV1%Pred was an independent risk factor for severe acute exacerbations 1 year after baseline in COPD subjects.CAT baseline scores in the serum IL-23 is 1 year after the baseline COPD subjects independent risk factors for the development of CAT scores increase research.Conclusions1.The expression of protein markers in the sputum and serum of COPD patients was significantly different from that of healthy subjects.There was a greater difference in protein markers in the sputum than in the serum between COPD patients and normal subjects.Airway biomarkers of COPD may be better than serum biomarkers to characterize disease.2.Different COPD phenotypes(severe vs.non-severe airflow restriction,low risk of acute exacerbation vs.There were significant differences in protein marker expression between high risk,emphysema vs.non-emphysema,and sputum cytology.These differences can distinguish clinical phenotypes and suggest differences in biological processes behind clinical phenotypes.3.Protein markers of COPD were closely correlated with mMRC score,total CAT score,post-diastolic FEV1%Pred,number of acute exacerbations,%LAA-950,FeNO and blood Eos absolute count,which could be used to assess disease change.4.In addition to conventional clinical indicators(baseline post-diastolic FEV1%Pred,baseline CAT total score),serum IL-23 expression level is also an independent risk factor for clinical deterioration in COPD subjects,suggesting its potential value in predicting clinical outcome.