Machine Learning of Plasma Proteomics Classifies Diagnosis of Interstitial Lung Disease.

Rationale: Distinguishing connective tissue disease-associated interstitial lung disease (CTD-ILD) from idiopathic pulmonary fibrosis (IPF) can be clinically challenging. Objectives: To identify proteins that separate and classify patients with CTD-ILD and those with IPF. Methods: Four registries with 1,247 patients with IPF and 352 patients with CTD-ILD were included in analyses. Plasma samples were subjected to high-throughput proteomics assays. Protein features were prioritized using recursive feature elimination to construct a proteomic classifier. Multiple machine learning models, including support vector machine, LASSO (least absolute shrinkage and selection operator) regression, random forest, and imbalanced Random Forest, were trained and tested in independent cohorts. The validated models were used to classify each case iteratively in external datasets. Measurements and Main Results: A classifier with 37 proteins (proteomic classifier 37 [PC37]) was enriched in the biological process of bronchiole development and smooth muscle proliferation and immune responses. Four machine learning models used PC37 with sex and age score to generate continuous classification values. Receiver operating characteristic curve analyses of these scores demonstrated consistent areas under the curve of 0.85-0.90 in the test cohort and 0.94-0.96 in the single-sample dataset. Binary classification demonstrated 78.6-80.4% sensitivity and 76-84.4% specificity in the test cohort and 93.5-96.1% sensitivity and 69.5-77.6% specificity in the single-sample classification dataset. Composite analysis of all machine learning models confirmed 78.2% (194 of 248) accuracy in the test cohort and 82.9% (208 of 251) in the single-sample classification dataset. Conclusions: Multiple machine learning models trained with large cohort proteomic datasets consistently distinguished CTD-ILD from IPF. Many of the identified proteins are involved in immune pathways. We further developed a novel approach for single-sample classification, which could facilitate honing the differential diagnosis of ILD in challenging cases and improve clinical decision making.

Molecular Endotypes of Idiopathic Pulmonary Fibrosis: A Latent Class Analysis of Two Multicenter Observational Cohorts.

Rationale: Idiopathic pulmonary fibrosis (IPF) causes irreversible fibrosis of the lung parenchyma. Although antifibrotic therapy can slow IPF progression, treatment response is variable. There exists a critical need to develop a precision medicine approach to IPF. Objectives: To identify and validate biologically driven molecular endotypes of IPF. Methods: Latent class analysis (LCA) was independently performed in prospectively recruited discovery (n = 875) and validation (n = 347) cohorts. Twenty-five plasma biomarkers associated with fibrogenesis served as class-defining variables. The association between molecular endotype and 4-year transplant-free survival was tested using multivariable Cox regression adjusted for baseline confounders. Endotype-dependent differential treatment response to future antifibrotic exposure was then assessed in a pooled cohort of patients naive to antifibrotic therapy at the time of biomarker measurement (n = 555). Measurements and Main Results: LCA independently identified two latent classes in both cohorts (P < 0.0001). WFDC2 (WAP four-disulfide core domain protein 2) was the most important determinant of class membership across cohorts. Membership in class 2 was characterized by higher biomarker concentrations and a higher risk of death or transplant (discovery, hazard ratio [HR], 2.02; 95% confidence interval [CI], 1.64-2.48; P < 0.001; validation, HR, 1.95; 95% CI, 1.34-2.82; P < 0.001). In pooled analysis, significant heterogeneity in treatment effect was observed between endotypes (P = 0.030 for interaction), with a favorable antifibrotic response in class 2 (HR, 0.64; 95% CI, 0.45-0.93; P = 0.018) but not in class 1 (HR, 1.19; 95% CI, 0.77-1.84; P = 0.422). Conclusions: In this multicohort study, we identified two novel molecular endotypes of IPF with divergent clinical outcomes and responses to antifibrotic therapy. Pending further validation, these endotypes could enable a precision medicine approach for future IPF clinical trials.

Biological Age, Chronological Age, and Survival in Pulmonary Fibrosis: A Causal Mediation Analysis.

Rationale: Accelerated biological aging has been implicated in the development of interstitial lung disease (ILD) and other diseases of aging but remains poorly understood. Objectives: To identify plasma proteins that mediate the relationship between chronological age and survival association in patients with ILD. Methods: Causal mediation analysis was performed to identify plasma proteins that mediated the chronological age-survival relationship in an idiopathic pulmonary fibrosis discovery cohort. Proteins mediating this relationship after adjustment for false discovery were advanced for testing in an independent ILD validation cohort and explored in a chronic obstructive pulmonary disease cohort. A proteomic-based measure of biological age was constructed and survival analysis performed, assessing the impact of biological age and peripheral blood telomere length on the chronological age-survival relationship. Measurements and Main Results: Twenty-two proteins mediated the chronological age-survival relationship after adjustment for false discovery in the idiopathic pulmonary fibrosis discovery cohort (n = 874), with 19 remaining significant mediators of this relationship in the ILD validation cohort (n = 983) and one mediating this relationship in the chronic obstructive pulmonary disease cohort. Latent transforming growth factor-ß binding protein 2 and ectodysplasin A2 receptor showed the strongest mediation across cohorts. A proteomic measure of biological age completely attenuated the chronological age-survival association and better discriminated survival than chronological age. Results were robust to adjustment for peripheral blood telomere length, which did not mediate the chronological age-survival relationship. Conclusions: Molecular measures of aging completely mediate the relationship between chronological age and survival, suggesting that chronological age has no direct effect on ILD survival.

Genetic-informed proteome-wide scan reveals potential causal plasma proteins for idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a lethal lung disease for which there are no reliable biomarkers or disease-modifying drugs. Here, we integrated human genomics and proteomics to investigate the causal associations between 2769 plasma proteins and IPF. Our Mendelian randomisation analysis identified nine proteins associated with IPF, of which three (FUT3, ADAM15 and USP28) were colocalised. ADAM15 emerged as the top candidate, supported by expression quantitative trait locus analysis in both blood and lung tissue. These findings provide novel insights into the aetiology of IPF and offer translational opportunities in response to the clinical challenges of this devastating disease.

Fibroblast activation protein and disease severity, progression, and survival in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is characterized by progressive fibrosis in the lungs. Activated fibroblasts play a central role in fibrogenesis and express fibroblast activation protein α. A truncated, soluble form (sFAP) can be measured in blood and is a potential novel biomarker of disease activity. The aim was to study the association between sFAP and clinical, radiological, and histopathological measures of disease severity, progression, and survival in a prospective, multicentre, real-world cohort of patients with IPF. Patients with IPF were recruited from the tertiary interstitial lung disease centres in Denmark and followed for up to 3 years. Baseline serum levels of sFAP were measured by ELISA in patients with IPF and compared to healthy controls. Pulmonary function tests, 6-minute walk test and quality of life measures were performed at baseline and during follow-up. The study included 149 patients with IPF. Median sFAP in IPF was 49.6 ng/mL (IQR: 43.1-61.6 ng/mL) and in healthy controls 73.8 ng/mL (IQR: 62.1-92.0 ng/mL). Continuous sFAP was not associated with disease severity, progression or survival (p > 0.05). After dichotomization of sFAP below or above mean sFAP + 2 SD for healthy controls, higher levels of sFAP were associated with lower FVC % predicted during follow-up (p < 0.01). Higher than normal serum levels of sFAP were associated with longitudinal changes in FVC % predicted, but sFAP did not show clear associations with other baseline or longitudinal parameters. As such, sFAP has limited use as a biomarker of disease progression or survival in patients with IPF.

CXCL10 predicts autoimmune features and a favorable clinical course in patients with IIP: post hoc analysis of a prospective and multicenter cohort study.

BACKGROUND: Interstitial pneumonia with autoimmune features (IPAF), which does not meet any of the criteria for connective tissue diseases (CTD), has been attracting an attention in patients with idiopathic interstitial pneumonia (IIP). However, the biomarkers that reflect the clinical course of these patients have not been fully elucidated. OBJECTIVE: To identify useful serum biomarkers reflecting CTD-related features and favorable prognoses in patients with IIP. METHODS: This was a post hoc analysis of a prospective and multicenter cohort study between 2015 and 2020. Newly diagnosed patients with IIP were consecutively enrolled, and 74 autoimmune features and autoantibodies were comprehensively checked during IIP diagnosis. Serum levels of CXCL10, CXCL1, CCL2, BAFF, angiopoietin-2, and leptin were evaluated at the time of IIP diagnosis. RESULTS: Two hundred twenty-two patients (159 men and 63 women) with IIP were enrolled. The median observation duration was 36 months. The median age was 71 years old, and median %forced vital capacity (FVC) was 84.1% at the time of IIP diagnosis. The proportion of patients who met the classification criteria for IPAF was 11.7%. In patients with high serum CXCL10, changes in both %FVC and %diffusion lung capacity for carbon monoxide at one year were significantly higher than those in patients with low CXCL10 (p = 0.014 and p = 0.009, respectively), whereas these changes were not significant for other chemokines and cytokines. High CXCL10 levels were associated with acute/subacute onset (p < 0.001) and the diagnosis of nonspecific interstitial pneumonia with organizing pneumonia overlap (p = 0.003). High CXCL10 levels were related to a higher classification of IPAF (relative risk for IPAF was 3.320, 95%CI: 1.571-7.019, p = 0.003) and lower classification of progressive pulmonary fibrosis (PPF; relative risk for PPF was 0.309, 95%CI: 0.100-0.953, p = 0.027) compared to those with low CXCL10. Finally, survival was higher in patients with IPF and high CXCL10 (p = 0.044), and high CXCL10 was a significant prognostic factor in multivariate Cox proportional hazards models (hazard ratio 0.368, p = 0.005). CONCLUSIONS: High serum levels of CXCL10 are associated with CTD-related features, the favorable clinical course, and survival in patients with IIP, especially IPF. CLINICAL TRIAL NUMBER: Not applicable.

Blood lipid profiles as a prognostic biomarker in idiopathic pulmonary fibrosis.

BACKGROUND: Dysregulation of lipid metabolism is implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). However, the association between the blood lipid profiles and the prognosis of IPF is not well defined. We aimed to identify the impacts of lipid profiles on prognosis in patients with IPF. METHODS: Clinical data of 371 patients with IPF (145 and 226 in the derivation and validation cohorts, respectively), including serum lipid profiles (total cholesterol, triglyceride, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, apolipoprotein A-I [Apo A-I], and apolipoprotein B), were retrospectively collected. The association with mortality was analyzed using the Cox proportional hazard model. RESULTS: In the derivation cohort, the mean age was 67.5 years, 86.2% were men, and 30.3% died during the follow-up (median: 18.0 months). Non-survivors showed lower lung function and greater gender-age-physiology scores than survivors. Among the serum lipid profiles, the levels of triglyceride and Apo A-I were significantly lower in non-survivors than in survivors. In the multivariate Cox analysis, low Apo A-I levels (< 140 mg/dL) were independently associated with the risk of mortality (hazard ratio 3.910, 95% confidence interval 1.170-13.069; P = 0.027), when adjusted for smoking history, body mass index, GAP score, and antifibrotic agent use. In both derivation and validation cohorts, patients with low Apo A-I levels (< 140 mg/dL) had worse survival (median survival: [derivation] 34.0 months vs. not reached, P = 0.003; [validation] 40.0 vs. 53.0 months, P = 0.027) than those with high Apo A-I levels in the Kaplan-Meier survival analysis. CONCLUSIONS: Our results indicate that low serum Apo A-1 levels are an independent predictor of mortality in patients with IPF, suggesting the utility of serum Apo A-I as a prognostic biomarker in IPF.

Potential role of SIRT-1 and SIRT-3 as biomarkers for the diagnosis and prognosis of idiopathic pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a debilitating and progressive lung disease of unknown aetiology, characterized by the relentless deposition of fibrotic tissue. Biomarkers may play a pivotal role as indicators of disease presence, progression, and treatment response. Sirtuins, a family of enzymes with ADP ribosyltransferase or deacetylase activity, have been implicated in several diseases, including pulmonary fibrosis. METHODS: A cross-sectional, prospective, observational single-center study was conducted to investigate the potential role of serum SIRTs levels as biomarkers in patients with IPF. Demographic, clinical, and functional data and serological samples were collected from 34 patients with IPF followed at the Interstital Lung and Rare Diseases Outpatient Clinic of the Vanvitelli Pneumology Clinic, Monaldi Hospital, Naples, Italy and from 19 age-matched controls. RESULTS: Serum SIRT-1 levels were significantly reduced in IPF patients compared to controls (median IPF 667 [435-858] pg/mL versus controls 925 [794-1173] pg/mL; p < 0.001 ). In contrast, serum SIRT-3 levels were significantly increased in IPF patients compared to controls (median IPF 338 [230-500] pg/mL versus controls 154 [99.8-246] pg/mL; p < 0.001). There were no statistically significant differences in serum SIRT-6 and SIRT-7 levels between IPF and controls. In addition, we found a significant positive correlation between SIRT-1 and lung function parameters such as FEV1% (ϱ=0.417;p = 0.016), FVC% (ϱ=0.449;p = 0.009) and DLCO% (ϱ=0.393;p = 0.024), while a significant negative correlation was demonstrated between SIR-1 and GAP score, demonstrating a significant reduction in SIRT-1 in advanced Gender-Age-Physiology (GAP) stages 2-3 compared to GAP stage 1 (p = 0.008). CONCLUSIONS: This prospective, cross-sectional study showed that SIRT-1 was associated with lung function and IPF severity and that both SIRT-1 and SIRT-3 could be considered as potential biomarkers of IPF, whereas SIRT-6 and SIRT-7 were not associated with IPF.

Lipocalin-2 as a prognostic marker in patients with acute exacerbation of idiopathic pulmonary fibrosis.

BACKGROUND: Lipocalin-2 (LCN2) is a secretory glycoprotein upregulated by oxidative stress; moreover, patients with idiopathic pulmonary fibrosis (IPF) have shown increased LCN2 levels in bronchoalveolar lavage fluid (BALF). This study aimed to determine whether circulatory LCN2 could be a systemic biomarker in patients with IPF and to investigate the role of LCN2 in a bleomycin-induced lung injury mouse model. METHODS: We measured serum LCN2 levels in 99 patients with stable IPF, 27 patients with acute exacerbation (AE) of IPF, 51 patients with chronic hypersensitivity pneumonitis, and 67 healthy controls. Further, LCN2 expression in lung tissue was evaluated in a bleomycin-induced lung injury mouse model, and the role of LCN2 was investigated using LCN2-knockout (LCN2 -/-) mice. RESULTS: Serum levels of LCN2 were significantly higher in patients with AE-IPF than in the other groups. The multivariate Cox proportional hazards model showed that elevated serum LCN2 level was an independent predictor of poor survival in patients with AE-IPF. In the bleomycin-induced lung injury mouse model, a higher dose of bleomycin resulted in higher LCN2 levels and shorter survival. Bleomycin-treated LCN2 -/- mice exhibited increased BALF cell and protein levels as well as hydroxyproline content. Moreover, compared with wild-type mice, LCN2-/- mice showed higher levels of circulatory 8-isoprostane as well as lower Nrf-2, GCLC, and NQO1 expression levels in lung tissue following bleomycin administration. CONCLUSIONS: Our findings demonstrate that serum LCN2 might be a potential prognostic marker of AE-IPF. Moreover, LCN2 expression levels may reflect the severity of lung injury, and LCN2 may be a protective factor against bleomycin-induced acute lung injury and oxidative stress.

Novel prediction model of early screening lung adenocarcinoma with pulmonary fibrosis based on haematological index.

BACKGROUND: Lung cancer (LC), a paramount global life-threatening condition causing significant mortality, is most commonly characterized by its subtype, lung adenocarcinoma (LUAD). Concomitant with LC, pulmonary fibrosis (PF) and interstitial lung disease (ILD) contribute to an intricate landscape of respiratory diseases. Idiopathic pulmonary fibrosis (IPF) in association with LC has been explored. However, other fibrotic interrelations remain underrepresented, especially for LUAD-PF and LUAD-ILD. METHODS: We analysed data with statistical analysis from 7,137 healthy individuals, 7,762 LUAD patients, 7,955 ILD patients, and 2,124 complex PF patients collected over ten years. Furthermore, to identify blood indicators related to lung disease and its complications and compare the relationships between different indicators and lung diseases, we successfully applied the naive Bayes model for a biomarker-based prediction of diagnosis and development into complex PF. RESULTS: Males predominantly marked their presence in all categories, save for complex PF where females took precedence. Biomarkers, specifically AGR, MLR, NLR, and PLR emerged as pivotal in discerning lung diseases. A machine-learning-driven predictive model underscored the efficacy of these markers in early detection and diagnosis, with NLR exhibiting unparalleled accuracy. CONCLUSIONS: Our study elucidates the gender disparities in lung diseases and illuminates the profound potential of serum biomarkers, including AGR, MLR, NLR, and PLR in early lung cancer detection. With NLR as a standout, therefore, this study advances the exploration of indicator changes and predictions in patients with pulmonary disease and fibrosis, thereby improving early diagnosis, treatment, survival rate, and patient prognosis.

Exploring the causal effect between lipid-modifying drugs and idiopathic pulmonary fibrosis: a drug-target Mendelian randomization study.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a respiratory disorder of obscure etiology and limited treatment options, possibly linked to dysregulation in lipid metabolism. While several observational studies suggest that lipid-lowering agents may decrease the risk of IPF, the evidence is inconsistent. The present Mendelian randomization (MR) study aims to determine the association between circulating lipid traits and IPF and to assess the potential influence of lipid-modifying medications for IPF. METHODS: Summary statistics of 5 lipid traits (high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, triglyceride, apolipoprotein A, and apolipoprotein B) and IPF were sourced from the UK Biobank and FinnGen Project Round 10. The study's focus on lipid-regulatory genes encompassed PCSK9, NPC1L1, ABCG5, ABCG8, HMGCR, APOB, LDLR, CETP, ANGPTL3, APOC3, LPL, and PPARA. The primary effect estimates were determined using the inverse-variance-weighted method, with additional analyses employing the contamination mixture method, robust adjusted profile score, the weighted median, weighted mode methods, and MR-Egger. Summary-data-based Mendelian randomization (SMR) was used to confirm significant lipid-modifying drug targets, leveraging data on expressed quantitative trait loci in relevant tissues. Sensitivity analyses included assessments of heterogeneity, horizontal pleiotropy, and leave-one-out methods. RESULTS: There was no significant effect of blood lipid traits on IPF risk (all P＞0.05). Drug-target MR analysis indicated that genetic mimicry for inhibitor of NPC1L1, PCSK9, ABCG5, ABCG8, and APOC3 were associated with increased IPF risks, with odds ratios (ORs) and 95% confidence intervals (CIs) as follows: 2.74 (1.05-7.12, P = 0.039), 1.36 (1.02-1.82, P = 0.037), 1.66 (1.12-2.45, P = 0.011), 1.68 (1.14-2.48, P = 0.009), and 1.42 (1.20-1.67, P = 3.17×10-5), respectively. The SMR method identified a significant association between PCSK9 gene expression in whole blood and reduced IPF risk (OR = 0.71, 95% CI: 0.50-0.99, P = 0.043). Sensitivity analyses showed no evidence of bias. CONCLUSIONS: Serum lipid traits did not significantly affect the risk of idiopathic pulmonary fibrosis. Drug targets MR studies examining 12 lipid-modifying drugs indicated that PCSK9 inhibitors could dramatically increase IPF risk, a mechanism that may differ from their lipid-lowering actions and thus warrants further investigation.

Effects of Active Chronic Cigarette-Smoke Exposure on Circulating Fibrocytes.

PURPOSE: This study aimed to evaluate the hypothesis that active smoking impacts upon mediators and abundance of circulating fibrocyte cells in smoking-related disease characterised by fibrosis. METHODS: Flow cytometry and enzyme-linked immunosorbent assays were used to investigate blood from five patient groups: healthy never-smokers, healthy current smokers, stable chronic obstructive pulmonary disease (COPD) active smokers, idiopathic pulmonary fibrosis (IPF) never-smokers, and IPF active smokers. RESULTS: A significant inverse dose-response relationship was observed in healthy smokers among cumulative smoking burden (pack-years) and fibrocyte abundance (p = 0.006, r = -0.86). Among serum profibrotic fibrocyte chemokines measured, CCL18 rose significantly alongside fibrocyte numbers in all five subject groups, while having an inverse dose-response relationship with pack-year burden in healthy smokers (p = 0.003, r = -0.89). In IPF, CCL2 rose in direct proportion to fibrocyte abundance irrespective of smoking status but had lower serum levels in those currently smoking (p = < 0.001). For the study population, CXCL12 was decreased in pooled current smokers versus never-smokers (p = 0.03). CONCLUSION: The suppressive effect of current, as distinct from former, chronic smoking on circulating fibrocyte abundance in healthy smokers, and modulation of regulatory chemokine levels by active smoking may have implications for future studies of fibrocytes in smoking-related lung diseases as a potential confounding variable.

Causal relationship between circulating glutamine levels and idiopathic pulmonary fibrosis: a two-sample mendelian randomization study.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a progressive and debilitating respiratory disease with a median survival of less than 5 years. In recent years, glutamine has been reported to be involved in the regulation of collagen deposition and cell proliferation in fibroblasts, thereby influencing the progression of IPF. However, the relationships between glutamine and the incidence, progression, and treatment response of IPF remain unclear. Our study aimed to investigate the relationship between circulating glutamine levels and IPF, as well as its potential as a therapeutic target. METHODS: We performed a comprehensive Mendelian Randomization (MR) analysis using the most recent genome-wide association study summary-level data. A total of 32 single nucleotide polymorphisms significantly correlated to glutamine levels were identified as instrumental variables. Eight MR analysis methods, including inverse variance weighted, MR-Egger, weighted median, weighted mode, constrained maximum likelihood, contamination mixture, robust adjusted profile score, and debiased inverse-variance weighted method, were used to assess the relationship between glutamine levels with IPF. RESULTS: The inverse variance weighted analysis revealed a significant inverse correlation between glutamine levels and IPF risk (Odds Ratio = 0.750; 95% Confidence Interval : 0.592-0.951; P = 0.017). Sensitivity analyses, including MR-Egger regression and MR-PRESSO global test, confirmed the robustness of our findings, with no evidence of horizontal pleiotropy or heterogeneity. CONCLUSION: Our study provides novel evidence for a causal relationship between lower circulating glutamine levels and increased risk of IPF. This finding may contribute to the early identification of high-risk individuals for IPF, disease monitoring, and development of targeted therapeutic strategies.

The association between testosterone, estradiol, estrogen sulfotransferase and idiopathic pulmonary fibrosis: a bidirectional mendelian randomization study.

BACKGROUND: The causal relationships between testosterone, estradiol, estrogen sulfotransferase, and idiopathic pulmonary fibrosis (IPF) are not well understood. This study employs a bidirectional two-sample Mendelian Randomization (MR) approach to explore these associations. METHODS: All genetic data utilized in our study were obtained from the IEU Open GWAS project. For the MR analysis, we employed the inverse variance weighted (IVW), MR-Egger, and weighted median methods to assess the causal relationships. We also conducted a multivariate MR (MVMR) analysis, with adjustments made for smoking. To ensure the robustness of our findings, sensitivity analyses were conducted using Cochran's Q test, MR-Egger regression, the MR-PRESSO global test, and the leave-one-out method. RESULTS: Genetically predicted increases in serum testosterone levels by one standard deviation were associated with a 58.7% decrease in the risk of developing IPF (OR = 0.413, PIVW=0.029, 95% CI = 0.187 ∼ 0.912), while an increase in serum estrogen sulfotransferase by one standard deviation was associated with a 32.4% increase in risk (OR = 1.324, PIVW=0.006, 95% CI = 1.083 ∼ 1.618). No causal relationship was found between estradiol (OR = 1.094, PIVW=0.735, 95% CI = 0.650 ∼ 1.841) and the risk of IPF. Reverse MR analysis did not reveal any causal relationship between IPF and testosterone (OR = 1.001, PIVW=0.51, 95% CI = 0.998 ∼ 1.004), estradiol (OR = 1.001, PIVW=0.958, 95% CI = 0.982 ∼ 1.019), or estrogen sulfotransferase (OR = 0.975, PIVW=0.251, 95% CI = 0.933 ∼ 1.018). The MVMR analysis demonstrated that the association between testosterone (OR = 0.442, P = 0.037, 95% CI = 0.205 ∼ 0.953) and estrogen sulfotransferase (OR = 1.314, P = 0.001, 95% CI = 1.118 ∼ 1.545) and the risk of IPF persisted even after adjusting for smoking. CONCLUSIONS: Increased serum levels of testosterone are associated with a reduced risk of IPF, while increased levels of serum estrogen sulfotransferase are associated with an increased risk. No causal relationship was found between estradiol and the development of IPF. No causal relationship was identified between IPF and testosterone, estradiol, or estrogen sulfotransferase.

Oxygen saturation recovery after 6-minute walk test in patients with idiopathic pulmonary fibrosis.

BACKGROUND: The six-minute walk test (6MWT) is widely used to assess functional capacity and prognosis in patients with idiopathic pulmonary fibrosis (IPF). However, studies on oxygen saturation recovery after the 6MWT in patients with IPF are rare. In our study, we investigated the relationship between oxygen saturation recovery time and dyspnea, fatigue, quality of life, prognostic markers and pulmonary hypertension (PH). METHODS: In this cross-sectional study, IPF patients diagnosed according to current guidelines and followed up in our Interstitial Lung Disease Outpatient Clinic between 2021 and 2022 were included. Demographics, data from spirometry, diffusion capacity measurement, arterial blood gas analysis, transthoracic echocardiography and the 6MWT were recorded. The oxygen saturation recovery time, distance saturation product (DSP), gender-age-physiology (GAP) index and composite physiological index (CPI) scores were calculated. Dyspnea severity was assessed by the modified Medical Research Council (mMRC) and Dyspnoea-12 (D-12) scales, fatigue severity by the Multidimensional Fatigue Inventory (MFI-20) and quality of life by the St George's Respiratory Questionnaire (SGRQ). RESULTS: Fifty IPF patients (34 men, 16 women, age: 66.8 ± 7.3 years) were included in the study. The mean FVC was 77.8 ± 19.3%, the DLCO was 52.9 ± 17.1%, the 6-minute walk distance (6MWD) was 385.7 ± 90.6 m, the GAP index was 3.5 ± 1.5, and the CPI was 43.7 ± 14.1. Oxygen saturation after the 6MWT reached pretest values at an average of 135.6 ± 73.5 s. The oxygen saturation recovery time was longer in patients with higher GAP index scores (Rs = 0.870, p < 0.001), CPI scores (Rs = 0.906, p < 0.001), desaturation (Rs = 0.801, p < 0.001), FVC%/DLCO% (Rs = 0.432, p = 0.002), sPAP (Rs = 0.492, p = 0.001), TRV (Rs = 0.504, p = 0.001), mMRC (Rs = 0.913, p < 0.001), MFI-20 (Rs = 0.944, p < 0.001), D-12 scale (Rs = 0.915, p < 0.001) and SGRQ scores (Rs = 0.927, p < 0.001); lower FVC (%) (Rs=-0.627, p < 0.001), DLCO (%) (Rs=-0.892, p < 0.001), PaO2 (Rs=-0.779, p < 0.001), DSP (Rs=-0.835, p < 0.001), and 6MWD (Rs=-0.763, p < 0.001). A total of twenty patients (40%) exhibited an increased risk of PH. According to our multiple regression analysis, oxygen saturation recovery time was independently associated with the GAP index (p = 0.036), the lowest oxygen saturation occurring during the 6MWT (p = 0.011) and the SGRQ score (p < 0.001). CONCLUSIONS: Our results showed that oxygen saturation recovery time is associated with dyspnea, fatigue, quality of life, increased risk of PH and prognostic markers in IPF. Therefore, we recommend continuous measurement of oxygen saturation after 6MWT until pretest values are reached.

Prognostic performance of Krebs von den Lungen-6, surfactant protein A, surfactant protein D levels in the serum and bronchoalveolar lavage fluid in chronic fibrosing interstitial pneumonia: a retrospective study.

BACKGROUND: The serum markers Krebs von den Lungen-6 (KL-6), surfactant protein A (SP-A), and surfactant protein D (SP-D) have been used for the diagnosis, differential diagnosis, and prognosis prediction of interstitial pneumonia. However, the significance of measuring the serum and bronchoalveolar lavage fluid (BALF) KL-6, SP-D, and SP-A levels in predicting the prognosis of chronic fibrosing interstitial pneumonia (CFIP), idiopathic pulmonary fibrosis, and idiopathic nonspecific interstitial pneumonia remains unclear. We aimed to clarify the significance of measuring the serum and BALF KL-6, SP-A, and SP-D levels in predicting the prognosis of patients with CFIP. METHODS: Among 173 patients who were diagnosed with CFIP between September 2008 and February 2021, 39 who underwent bronchoalveolar lavage were included in this study. Among these, patients experiencing an annual decrease in forced vital capacity (FVC) of ≥10% or those facing challenges in undergoing follow-up pulmonary function tests owing to significant deterioration in pulmonary function were categorized as the rapidly progress group. Conversely, individuals with an annual decrease in the FVC of <10% were classified into the slowly progress group. The serum and BALF KL-6, SP-D, and SP-A levels, as well as BALF/serum SP-D and SP-A ratios were compared between the two groups. RESULTS: Among the patients with CFIP, the BALF SP-D level (p=0.0111), BALF SP-A level (p<0.0010), BALF/serum SP-D ratio (p=0.0051), and BALF/serum SP-A ratio (p<0.0010) were significantly lower in the rapidly than in the slowly progress group (p<0.0010). The receiver operating characteristics analysis results demonstrated excellent performance for diagnosing patients with CFIP, with the BALF SP-D level (area under the curve [AUC], 0.7424), BALF SP-A level (AUC, 0.8842), BALF/serum SP-D ratio (AUC, 0.7673), and BALF/serum SP-A ratio (AUC, 0.8556). Moreover, the BALF SP-A level showed a notably superior CFIP diagnostic capability. Survival analysis using the Kaplan-Meier method revealed that patients with a BALF SP-A level of <1500 ng/mL and BALF/serum SP-A ratio of <15.0 had poor prognoses. CONCLUSIONS: Our results suggest that BALF SP-A measurement may be useful for predicting the prognosis in patients with CFIP.

Endothelial-Specific Loss of Sphingosine-1-Phosphate Receptor 1 Increases Vascular Permeability and Exacerbates Bleomycin-induced Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive disease which leads to significant morbidity and mortality from respiratory failure. The two drugs currently approved for clinical use slow the rate of decline in lung function but have not been shown to halt disease progression or reverse established fibrosis. Thus, new therapeutic targets are needed. Endothelial injury and the resultant vascular permeability are critical components in the response to tissue injury and are present in patients with IPF. However, it remains unclear how vascular permeability affects lung repair and fibrosis following injury. Lipid mediators such as sphingosine-1-phosphate (S1P) are known to regulate multiple homeostatic processes in the lung including vascular permeability. We demonstrate that endothelial cell-(EC) specific deletion of the S1P receptor 1 (S1PR1) in mice (EC-S1pr1-/-) results in increased lung vascular permeability at baseline. Following a low-dose intratracheal bleomycin challenge, EC-S1pr1-/- mice had increased and persistent vascular permeability compared with wild-type mice, which was strongly correlated with the amount and localization of resulting pulmonary fibrosis. EC-S1pr1-/- mice also had increased immune cell infiltration and activation of the coagulation cascade within the lung. However, increased circulating S1P ligand in ApoM-overexpressing mice was insufficient to protect against bleomycin-induced pulmonary fibrosis. Overall, these data demonstrate that endothelial cell S1PR1 controls vascular permeability in the lung, is associated with changes in immune cell infiltration and extravascular coagulation, and modulates the fibrotic response to lung injury.

Monocyte Count as a Prognostic Biomarker in Patients with Idiopathic Pulmonary Fibrosis.

Rationale: There is an urgent need for simple, cost-effective prognostic biomarkers for idiopathic pulmonary fibrosis (IPF); biomarkers that show potential include monocyte count. Objectives: We used pooled data from pirfenidone and IFNγ-1b trials to explore the association between monocyte count and prognosis in patients with IPF. Methods: This retrospective pooled analysis included patients (active and placebo arms) from the following four phase III, randomized, placebo-controlled trials: ASCEND (NCT01366209), CAPACITY (NCT00287729 and NCT00287716), and INSPIRE (NCT00075998). Outcomes included IPF progression (≥10% absolute decline in FVC% predicted, ≥50 m decline in 6-minute-walk distance, or death), all-cause hospitalization, and all-cause mortality over 1 year. The relationship between monocyte count (defined as time-dependent) and outcomes was assessed using bivariate and multivariable models. Measurements and Main Results: This analysis included 2,067 patients stratified by monocyte count (at baseline: <0.60 × 109 cells/L [n = 1,609], 0.60 to <0.95 × 109 cells/L [n = 408], and ≥0.95 × 109 cells/L [n = 50]). In adjusted analyses, a higher proportion of patients with monocyte counts of 0.60 to <0.95 × 109 cells/L or ≥0.95 × 109 cells/L versus <0.60 × 109 cells/L experienced IPF progression (P = 0.016 and P = 0.002, respectively), all-cause hospitalization (P = 0.030 and P = 0.003, respectively), and all-cause mortality (P = 0.005 and P < 0.001, respectively) over 1 year. Change in monocyte count from baseline was not associated with any of the outcomes over 1 year and did not appear to be affected by study treatment. Conclusions: In patients with IPF, elevated monocyte count was associated with increased risks of IPF progression, hospitalization, and mortality. Monocyte count may provide a simple and inexpensive prognostic biomarker in IPF.

Autoantibodies targeting telomere-associated proteins in systemic sclerosis.

OBJECTIVES: Systemic sclerosis (SSc) is an autoimmune fibrotic disease affecting multiple tissues including the lung. A subset of patients with SSc with lung disease exhibit short telomeres in circulating lymphocytes, but the mechanisms underlying this observation are unclear. METHODS: Sera from the Johns Hopkins and University of California, San Francisco (UCSF) Scleroderma Centers were screened for autoantibodies targeting telomerase and the shelterin proteins using immunoprecipitation and ELISA. We determined the relationship between autoantibodies targeting the shelterin protein TERF1 and telomere length in peripheral leucocytes measured by qPCR and flow cytometry and fluorescent in situ hybridisation (Flow-FISH). We also explored clinical associations of these autoantibodies. RESULTS: In a subset of patients with SSc, we identified autoantibodies targeting telomerase and the shelterin proteins that were rarely present in rheumatoid arthritis, myositis and healthy controls. TERF1 autoantibodies were present in 40/442 (9.0%) patients with SSc and were associated with severe lung disease (OR 2.4, p=0.04, Fisher's exact test) and short lymphocyte telomere length. 6/6 (100%) patients with TERF1 autoantibodies in the Hopkins cohort and 14/18 (78%) patients in the UCSF cohort had a shorter telomere length in lymphocytes or leukocytes, respectively, relative to the expected age-adjusted telomere length. TERF1 autoantibodies were present in 11/152 (7.2%) patients with idiopathic pulmonary fibrosis (IPF), a fibrotic lung disease believed to be mediated by telomere dysfunction. CONCLUSIONS: Autoantibodies targeting telomere-associated proteins in a subset of patients with SSc are associated with short lymphocyte telomere length and lung disease. The specificity of these autoantibodies for SSc and IPF suggests that telomere dysfunction may have a distinct role in the pathogenesis of SSc and pulmonary fibrosis.

Cutaneous T-cell-attracting chemokine as a novel biomarker for predicting prognosis of idiopathic pulmonary fibrosis: a prospective observational study.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive fibrotic lung disease that leads to respiratory failure and death. Although there is a greater understanding of the etiology of this disease, accurately predicting the disease course in individual patients is still not possible. This study aimed to evaluate serum cytokines/chemokines as potential biomarkers that can predict outcomes in IPF patients. METHODS: A multi-institutional prospective two-stage discovery and validation design using two independent cohorts was adopted. For the discovery analysis, serum samples from 100 IPF patients and 32 healthy controls were examined using an unbiased, multiplex immunoassay of 48 cytokines/chemokines. The serum cytokine/chemokine values were compared between IPF patients and controls; the association between multiplex measurements and survival time was evaluated in IPF patients. In the validation analysis, the cytokines/chemokines identified in the discovery analysis were examined in serum samples from another 81 IPF patients to verify the ability of these cytokines/chemokines to predict survival. Immunohistochemical assessment of IPF-derived lung samples was also performed to determine where this novel biomarker is expressed. RESULTS: In the discovery cohort, 18 cytokines/chemokines were significantly elevated in sera from IPF patients compared with those from controls. Interleukin-1 receptor alpha (IL-1Rα), interleukin-8 (IL-8), macrophage inflammatory protein 1 alpha (MIP-1α), and cutaneous T-cell-attracting chemokine (CTACK) were associated with survival: IL-1Rα, hazard ratio (HR) = 1.04 per 10 units, 95% confidence interval (95% CI) 1.01-1.07; IL-8, HR = 1.04, 95% CI 1.01-1.08; MIP-1α, HR = 1.19, 95% CI 1.00-1.36; and CTACK, HR = 1.12 per 100 units, 95% CI 1.02-1.21. A replication analysis was performed only for CTACK because others were previously reported to be potential biomarkers of interstitial lung diseases. In the validation cohort, CTACK was associated with survival: HR = 1.14 per 100 units, 95% CI 1.01-1.28. Immunohistochemistry revealed the expression of CTACK and CC chemokine receptor 10 (a ligand of CTACK) in airway and type II alveolar epithelial cells of IPF patients but not in those of controls. CONCLUSIONS: CTACK is a novel prognostic biomarker of IPF. Trial registration None (because of no healthcare intervention).