Chronic obstructive pulmonary disease (COPD) is a heterogeneous respiratory disorder characterized by concurrent injury and remodeling of the airways, lung parenchyma, and pulmonary vasculature. These lung impairments lead to progressively worsening airflow limitation, which in turn results in increased dyspnea, functional disability, and premature death (Qian et al., 2023). COPD is a major global health burden, ranking as the fourth leading cause of death worldwide, characterized by high prevalence, high mortality, and significant healthcare resource consumption (Wen et al., 2018; Ferrera et al., 2021). It showed an annual incidence rate of roughly 12%, with a global prevalence ranging from 9% to 10% among people aged 40 and older (Adeloye et al., 2015; Diseases and Injuries, 2020). The development of COPD involves a complex interplay of genetic susceptibility (e.g., α1-antitrypsin deficiency), environmental exposures (e.g., smoking, air pollution), and dysregulated immune responses (e.g., neutrophil/macrophage activation, protease-antiprotease imbalance). Dysregulation of immune responses in COPD leads to the activation of multiple immune cells, such as neutrophils, macrophages, lymphocytes, and eosinophils (Rabe et al., 2023). These cells secrete a large number of inflammatory mediators and cytokines, including interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), and chemokines. They can recruit more immune cells to the lungs, perpetuate the inflammatory process, and contribute to airway remodeling and lung tissue destruction. However, the complex pathogenesis of COPD remains incompletely understood (Zhu et al., 2025). Early diagnosis and timely therapeutic intervention are critical in improving prognosis and survival rates for individuals with COPD (Lin et al., 2023). Current diagnostic methods for COPD mainly include pulmonary function tests, chest X-ray radiography, and computed tomography (CT), etc. Due to the limited sensitivity and specificity of the equipment, there are certain difficulties in distinguishing COPD from other lung diseases. Moreover, these methods have poor visualization of mild lesions and suboptimal performance in the differential diagnosis of early-stage COPD (Kumar et al., 2024; Chen et al., 2025). In addition, pharmacological treatments currently focus on antibiotics and corticosteroids to manage inflammation, yet no clinically effective targeted therapies are available. Therefore, understanding the underlying molecular mechanisms and identifying new biomarkers are essential for establishing early diagnostic standards and discovering precision therapeutic targets for COPD.

Lactylation, a novel post-translational modification that involves the covalent attachment of lactate to lysine residues, has emerged as a critical regulatory mechanism in various cellular processes (Zhang et al., 2019; Gaffney et al., 2020). Recent studies indicate that lactylation plays a pivotal role in diverse physiological and pathological processes, including immunity (Chen et al., 2022), metabolism (Chen et al., 2021; Merkuri et al., 2024), and cancer (Yu et al., 2021), by modifying both histones and non-histone proteins. A recent study demonstrated that lactylation significantly influences the phenotype and functional properties of immune cells, including macrophage polarization (Irizarry-Caro et al., 2020) and T cell reprogramming (Lopez Krol et al., 2022). Pulmonary macrophages, encompassing tissue-resident macrophages (TRMs) and monocyte-derived macrophages (Hashimoto et al., 2013; Byrne et al., 2016), play pivotal roles in COPD pathophysiology. The M1 and M2 phenotypic states of these macrophages mediate distinct functional outcomes (Kim et al., 2024). Notably, lactylation has been identified as a key modulator of macrophage polarization, influencing both inflammatory mediator production and tissue repair capacity (Irizarry-Caro et al., 2020). Moreover, lactylation regulates gene expression related to inflammation, cytokine synthesis, and metabolic pathways in macrophages, ultimately affecting their activation states and functional outcomes (Cui et al., 2021; Wang et al., 2022; Li et al., 2024). Abnormal macrophage function, immune dysregulation, and sustained inflammation are well-established contributors to COPD pathogenesis (Lee et al., 2021). Therefore, investigating the role of lactylation in macrophage activation is essential for advancing our understanding of COPD pathogenesis and for developing novel therapeutic approaches and molecular targets.

This study utilized two transcriptomic datasets comprising blood samples from smokers with COPD and smokers with normal lung function. Differentially expressed genes (DEGs) related to macrophage lactylation between COPD and control samples were identified through various bioinformatics approaches, including differential expression analysis, immune infiltration analysis, and weighted gene co-expression network analysis (WGCNA). The underlying mechanisms of these biomarkers in COPD were further explored using a nomogram model, Gene Set Enrichment Analysis (GSEA), molecular regulatory networks, compound prediction, and molecular docking. Additionally, clinical samples were analyzed to validate the expression levels of the identified biomarkers, confirming the bioinformatics findings. In summary, this study provides a solid foundation for a deeper understanding of the molecular mechanisms driving COPD and offers potential avenues for novel therapeutic strategies.

COPD is a prevalent respiratory disorder and the fourth leading cause of global disease-related mortality, posing a significant economic burden worldwide (Brassington et al., 2022; Safiri et al., 2022). Given its profound impact, early detection is critical to slowing disease progression and alleviating pressures on healthcare systems. Emerging research underscores the close association between immune cell infiltration and COPD pathogenesis. In particular, macrophage polarization and lactylation—a recently identified post-translational modification—play central roles in immune dysregulation during COPD progression (Cruz et al., 2019; Bu et al., 2020). Thus, exploring the immune microenvironment, especially lactylation-related mechanisms, could provide novel diagnostic biomarkers that enhance clinical management and personalized therapies (Huang et al., 2022; MacDonald et al., 2023). In the present study, the infiltration levels of nine immune cell types showed significant differences between COPD and control groups, particularly among macrophages, including both M1 and M2 phenotypes. Additionally, three biomarkers associated with macrophage lactylation—ALDH2, ASGR2, and CYP1B1—were identified through bioinformatics analysis and further validated with clinical samples. GSEA analysis suggested that these biomarkers are primarily associated with metabolic pathways and B-cell receptor signaling. Furthermore, five TFs, including STAT3 and SOX9, were found to be linked to all three biomarkers. This study aims to identify significant novel biomarkers related to macrophage lactylation, which may aid in the early diagnosis and therapeutic management of COPD.

COPD is a chronic airway inflammatory disorder involving the participation of various immune cells and inflammatory mediators. Among them, macrophages, as the first line of defense in innate immunity, are one of the earliest immune cells to respond to inflammation. In COPD, macrophages play a key pathophysiological role via distinct M1 and M2 phenotypes (Lee et al., 2021). M1 macrophages, linked to infections or smoking-induced lung damage, promote Th1-type inflammation through pro-inflammatory cytokines (e.g., IFN-γ, TNF-α), exacerbating inflammation and tissue damage. In contrast, M2 macrophages, active in non-inflammatory states, aid tissue repair/remodeling and maintain homeostasis by secreting anti-inflammatory cytokines and clearing debris. Lactylation is a post-translational modification that adds lactate groups to proteins, thereby altering their function and stability. Recent study indicates that lactylation plays essential roles in regulating the M1/M2 polarization of macrophages (Bao et al., 2025). Research has showed that bone marrow-derived macrophages (BMDMs) stimulated with LPS and IFN-γ exhibited an M1 phenotype and increased lactylation. Following 24–48 h of M1 polarization, they began expressing M2-like genes (e.g., ARG1, VEGFA). In contrast, BMDMs treated solely with lactate displayed an M2-like phenotype and elevated lactylation without an initial M1 phase. These results indicate that lactylation induces an M2-like phenotype and facilitates the return of BMDMs to homeostasis during the late stage of M1 polarization (Zhang et al., 2019). However, the precise regulatory effects of lactylation on macrophage function have not been fully elucidated and need further investigation.

Through the application of machine learning algorithms and subsequent RT-qPCR validation of clinical samples, three diagnostic biomarkers associated with macrophage lactylation in COPD were identified. Aldehyde dehydrogenase 2 (ALDH2), a key enzyme involved in the metabolism of acetaldehyde, a product of alcohol metabolism, was one of these biomarkers (Wang et al., 2020). This tetrameric allosteric enzyme is highly expressed in critical organs such as the heart, brain, liver, and lungs (Ma et al., 2011; Koppaka et al., 2012). Reduced enzyme activity is associated with increased susceptibility to various diseases, including coronary heart disease (Xu et al., 2011), late-onset Alzheimer’s disease (Zhu Z. Y. et al., 2022), and cancer (Zhang and Fu, 2021; Tran et al., 2023). One study demonstrated that upregulating endogenous ALDH2 expression in fibrotic cells using CRISPR activation effectively inhibited the expression of profibrotic genes (Tan et al., 2021). Moreover, the ALDH2 loss-of-function polymorphism is linked to subtle alterations in pulmonary tissues, some of which resemble changes seen in normal pulmonary aging, suggesting a “premature lung aging” effect (Kuroda et al., 2017). A recent study has identified ALDH2 as a critical pathogenic mechanism linked to endogenous lactate accumulation in acute kidney injury and proposes it as a potential therapeutic target. ALDH2 lactylation at lysine 52 promotes PHB2 degradation via the ubiquitin-proteasome system, thereby inhibiting PHB2-mediated mitophagy and exacerbating mitochondrial dysfunction (Li J. et al., 2025). However, ALDH2 has not been studied in COPD. In this study, ALDH2 was significantly upregulated in COPD patients and may be involved in macrophage lactylation in the development of COPD, which might be helpful in the diagnosis and treatment of COPD.

Asialoglycoprotein Receptor 2 (ASGR2), a subunit of the asialoglycoprotein receptor, is a transmembrane protein predominantly expressed in hepatocytes. It specifically recognizes N-acetylgalactosamine and galactose and functions primarily in the internalization and degradation of glycoproteins via desialylation, a process essential for maintaining serum glycoprotein homeostasis (Grewal, 2010). ASGR2 expression correlates significantly with the clinical stage of hepatocellular carcinoma (Zhang et al., 2021). In gastric cancer, ASGR2 contributes to the manifestation of cancer hallmarks upon PS exposure and confers resistance to both chemotherapy and monoclonal antibody-based therapies (Kim et al., 2022). A recent study indicated that serum ASGR2 levels could serve as a biomarker for assessing the therapeutic effects of balloon pulmonary angioplasty (BPA) in patients with chronic thromboembolic pulmonary hypertension (CTEPH). Prior to BPA, ASGR2 levels were associated with HDL-C levels and platelet counts. Post-BPA, ASGR2 levels correlated with LYM%, which may provide insights into the immune and inflammatory states of patients with CTEPH (Xu et al., 2024). However, the specific regulatory mechanism of ASGR2 in COPD remains unclear and requires further in-depth exploration.

Cytochrome P450 Family 1 Subfamily B Member 1 (CYP1B1), a member of the CYP450 enzyme family, is expressed in both hepatic and extrahepatic tissues and plays a critical role in metabolizing a broad range of xenobiotics, including the metabolic activation of polycyclic aromatic hydrocarbons. CYP1B1 has been implicated in processes such as metabolism, inflammation, angiogenesis, and anticancer drug resistance (Li et al., 2017). CYP1B1 contributes to colorectal cancer (CRC) resistance to ferroptosis, with its metabolite, 20-HETE, mediating this resistance (Chen et al., 2023). Furthermore, CYP1B1 significantly influences CRC liver metastasis by regulating tumor cell proliferation through the “CYP1B1-LCFAs-G1/S transition,” suggesting its potential as a therapeutic target for CRC liver metastasis (Jin et al., 2023). It was reported that long-term exposure to incense smoke induces CYP1A1, CYP1A2, and CYP1B1 in rat lung and liver tissues with tissue-specific differences, accompanied by increased oxidative stress (elevated MDA and GSH levels, altered catalase activity in the liver) and inflammation (increased TNF-α and IL-4 levels), thereby potentially promoting carcinogenesis and health complications in chronically exposed individuals (Hussain et al., 2014). And CYP1B1 has been shown to play a protective role in preventing the exacerbation of allergic airway inflammation by ragweed extract and house dust mite, including increased IgE levels, infiltration of inflammatory cells, and especially an increase in Th2 cells (Alessandrini et al., 2022). In summary, these genes—ALDH2, ASGR2, and CYP1B1—are involved in key processes such as metabolism and inflammation. However, their expression and roles in COPD remain unexplored and need further exploration.

Previous studies have highlighted significant changes in the cellular components of the small airways in patients with COPD, with key alterations including epithelial cell senescence (Wu et al., 2022), a notable increase in neutrophils (Kapellos et al., 2023), elevated macrophage numbers accompanied by phenotypic shifts and impaired phagocytic function (Eltboli et al., 2014; Eapen et al., 2017), augmented T and B cell populations, and the proliferation and activation of epithelial dendritic cells (Stoll et al., 2015). In COPD, dysregulated immune cell activation and the release of immune mediators contribute significantly to the exacerbation of pulmonary inflammation. The altered immune microenvironment in COPD has gained increasing attention, and targeting immune cells could offer potential avenues for precision therapy (Tzortzaki et al., 2013; Lan et al., 2025). In the present study, immune infiltration analysis of MRGs revealed significant differences in the infiltration levels of nine immune cell types between the COPD and control groups, particularly monocytes, macrophages, dendritic cells, B cells, and CD8⁺ T cells. Macrophages, as integral components of the innate immune system, displayed significant differences between M1 and M2 phenotypes. Macrophage dysregulation is closely associated with COPD pathology and severity. Thus, strategies aimed at restoring the macrophage phenotype, improving phagocytosis, reducing inflammation, and addressing foamy macrophages may provide promising therapeutic targets for COPD. Furthermore, the role of innate immunity in COPD pathogenesis is complex, with different macrophage subsets contributing to proinflammatory responses, while M2 macrophages are involved in attenuating inflammation, promoting tissue repair, and decreasing the secretion of proinflammatory cytokines (Kim et al., 2024). Further analysis of the core genes identified their primary enrichment in metabolic pathways, B-cell receptor signaling, wound healing, and antibacterial humoral immune responses. Previous research has suggested that B cells, through antibody production, may play a role in airway inflammation in patients with COPD. B lymphocytes infiltrate the adventitia of small airways in patients with COPD, with a rise in lymphocyte fractions and lymphoid aggregates containing germinal centers as the disease progresses (Caramori et al., 2016). Previous research has indicated the antioxidant and wound-healing properties of baru nut extract in lung epithelial cells, suggesting its potential for COPD treatment (Coco et al., 2022). These findings offer valuable insights for future treatment strategies and further mechanistic investigations into COPD.

Database-based predictions identified 154 TFs associated with the biomarkers, with five TFs—STAT3, SOX9, TFAP4, IKZF2, and TP63—targeting all biomarkers. As is well established, the chronic inflammatory response plays a central role in the pathogenesis and progression of COPD. The JAK-STAT signaling pathway is crucial in the activation of cytokines during inflammation, significantly contributing to COPD development (Purohit et al., 2023). SOX9 alleviates cigarette smoke extract (CSE)-induced inflammatory injury in human bronchial epithelial cells by suppressing stromal interaction molecule 1 (STIM1) expression (Zhu X. et al., 2022). TFAP4, predominantly recognized as an oncogene (Wong et al., 2021), has recently been implicated in exacerbating liver fibrosis and tissue inflammation in mice by promoting the activation of the STING signaling pathway (Han et al., 2025). Moreover, the predicted compounds targeting the biomarkers primarily included CGP 52608, benzo(a)pyrene, and bisphenol A. These compounds warrant further exploration and may serve as the basis for the development of new targeted drugs, offering novel treatment options for COPD.

The PCR validation results provide strong evidence that the expression levels of biomarkers such as ALDH2, ASGR2, and CYP1B1 are significantly higher in patients with COPD. These findings, confirmed by clinical samples, align with the bioinformatics analysis and validate the reliability of the results. Consequently, this study offers clinical value for diagnosing COPD and identifying novel potential immune targets for COPD immunotherapy.

However, this study still has some limitations. Firstly, the clinical sample size of this study is relatively small, which may compromise the generalizability of the conclusions. Secondly, the samples validated via RT-qPCR in this study were whole blood, rather than those of purified macrophages or lung tissues, and thus may be confounded by the influence of other blood cell types. Thirdly, alterations in mRNA levels can be affected by multiple factors; relying solely on such changes is insufficient to definitively elucidate the lactylation status. There is a paucity of experimental validation and biological functional verification that directly link these biomarkers (ALDH2, ASGR2, and CYP1B1) to lactylation, coupled with potential batch effects in public databases (e.g., data from acute exacerbation phases, which fail to fully capture the heterogeneous features of chronic obstructive pulmonary disease). In addition, the data acquisition technologies used for the training set and validation set in this study are different (microarray technology and RNA sequencing). Such a difference may cause some DEGs to fail to be validated across platforms, which may further affect the completeness of candidate gene screening. In future studies, we will perform additional experiments to validate the findings. Firstly, we will employ macrophage-specific gene knockout models or organoid technology to mechanistically dissect how ALDH2/ASGR2/CYP1B1 regulates cell polarization through lactylation. Secondly, we will collect a more diverse set of samples (e.g., macrophages or lung tissues) for RT-qPCR or proteomic assays to validate changes in protein levels and their functional implications, thereby mitigating the influence of other blood cells. Thirdly, we will expand the scale of the dataset, attempt to use data from the same technical platform to reduce the impact of technical bias on the results, and strive to evaluate the relationship between biomarkers and disease stages—especially those biomarkers associated with metabolic syndrome in COPD. Additionally, we will incorporate mass spectrometry (MS) analyses to detect whether these proteins undergo lactylation under specific conditions, and validate the lactylation status of relevant enzymes or target proteins via Western blotting using anti-lactylation-specific antibodies, so as to more precisely characterize the modification status of proteins. Furthermore, leveraging the strong interaction between CYP1B1 and benzo[a]pyrene, we will develop targeted inhibitors or environmental exposure interventions. We aim to fill existing gaps through these studies and lay the groundwork for an in-depth understanding of the immunometabolic mechanisms of chronic obstructive pulmonary disease and their translational applications.

In conclusion, this study identified ALDH2, ASGR2, and CYP1B1 as novel biomarkers related to macrophage lactylation in COPD, demonstrating their roles in regulating the immune microenvironment (e.g., macrophage polarization) and metabolic pathways (e.g., oxidative stress, B-cell signaling). These findings provide new targets for early diagnosis and therapy.