Original Article
  

Cytokine Expression and Promoter Methylation Signatures Underlying Immune Dysregulation in Mild and Severe COPD

Abstract

Chronic obstructive pulmonary disease (COPD) is a progressive inflammatory lung condition and a leading cause of morbidity and mortality worldwide. Despite well-established links to smoking, emerging evidence highlights the involvement of complex immune and epigenetic mechanisms in its pathogenesis. This study aimed to investigate the expression, secretion, and promoter methylation status of key pro- and anti-inflammatory cytokines in peripheral blood mononuclear cells (PBMCs) of patients with mild and severe COPD, compared with healthy individuals.
PBMCs were isolated from 90 participants divided into three groups: severe COPD, mild COPD, and healthy controls. Quantitative polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and methylation-specific PCR were employed to evaluate gene expression, protein secretion, and promoter methylation of inflammatory cytokines.
Patients with COPD exhibited significant upregulation of pro-inflammatory cytokines (Interleukin1β (IL-1β), IL-6, IL-18, IFN-γ, and TNF-α) at both transcript and protein levels, with more pronounced alterations in the severe group. Conversely, anti-inflammatory mediators (IL-10 and TGF-β) were significantly downregulated. Promoter methylation analysis revealed hypomethylation in pro-inflammatory cytokine genes and hypermethylation in anti-inflammatory ones, correlating with disease severity.
The findings demonstrate that COPD progression is associated with a shift toward a hyper-inflammatory, hypo-regulatory immune phenotype sustained by epigenetic modifications. These results support the potential for integrating cytokine-methylation signatures into clinical staging and for targeting epigenetic and immune pathways in future therapeutic strategies.

1. Verma A, Behera A, Kumar R, et al. Mapping of digital health interventions for the self-management of COPD: a systematic review. Clinical Epidemiology and Global Health. 2023;24:101427. doi:10.1016/ j.cegh. 2023.101427.
2. Pham HQ, Pham KHT, Ha GH, et al. Economic burden of chronic obstructive pulmonary disease: a systematic review. Tuberculosis and Respiratory Diseases. 2024;87(3):234-45. doi:10.4046/trd.2023.0118.
3. Choi JY, Milne S, Yunus F, et al. Current chronic obstructive pulmonary disease treatment status in Asia: a position statement of the Asian Pacific Society of Respirology. Tuberculosis and Respiratory Diseases. 2022;85(3):279-91. doi:10.4046/trd.2022.0025.
4. Lange P, Ahmed E, Lahmar ZM, et al. Natural history and mechanisms of COPD. Respirology. 2021;26(4):298-321. doi:10.1111/resp.14007.
5. Patel N. An update on COPD prevention, diagnosis, and management: the 2024 GOLD Report. The Nurse Practitioner. 2024;49(6):29-36. doi:10.1097/01.NPR. 0000000000000185.
6. Kim SH, Moon J-Y, Min KH, Lee H. Proposed etiotypes for chronic obstructive pulmonary disease: controversial issues. Tuberculosis and Respiratory Diseases. 2024;87(3):221-33. doi:10.4046/trd.2023.0125.
7. Choi JY, Kim JW, Kim YH, et al. Clinical characteristics of non-smoking chronic obstructive pulmonary disease patients: findings from the KOCOSS cohort. COPD: Journal of Chronic Obstructive Pulmonary Disease. 2022;19(1):174-81.doi:10.1080/15412555.2022.2045263.
8. Lahouty M, Fadaee M, Aghaei R, et al. Gut microbiome and colorectal cancer: from pathogenesis to treatment. Pathology - Research and Practice. 2025;265:156034. doi:10.1016/j.prp.2024.156034.
9. Abdi M, Karimzadeh H, Jourabchi A, et al. Protective effects of Tribulus terrestris extract on cisplatin-induced ovarian damage: antioxidants and anti-inflammatory insights. Human & Experimental Toxicology. 2025;44:09603271251353492.doi:10.1177/09603271251353492
10. Fadaee M, Lahouty M, Abdi M, et al. The effects of n-3 PUFAs on colorectal cancer carcinogenesis: comprehensive review. Food and Agricultural Immunology. 2025;36(1):2501967. doi:10.1080/ 09540105.2024. 2501967.
11. Lahouty M, Soleymanzadeh A, Kazemi S, et al. Cell-based immunotherapy in oesophageal cancer. Journal of Drug Targeting. 2025:1-11. doi:10.1080/1061186X.2024.2384192.
12. Orooji N, Babaei S, Fadaee M, et al. Novel therapeutic approaches for non-small cell lung cancer: an updated view. Journal of Drug Targeting. 2025:1-16. doi:10.1080/1061186X.2024.2378910.
13. Parsania S, Salehpoor S, Ameli MS, et al. Investigating the therapeutic potential of clove extract in mitigating testicular injury associated with testicular hypoxia in male rats. Crescent Journal of Medical & Biological Sciences. 2024;11(3):145-52.
14. Rovina N, Koutsoukou A, Koulouris NG. Inflammation and immune response in COPD: where do we stand? Mediators of Inflammation. 2013;2013:413735. doi:10.1155/2013/413735.
15. Di Stefano A, Coccini T, Roda E, et al. Blood MCP-1 levels are increased in chronic obstructive pulmonary disease patients with prevalent emphysema. International Journal of Chronic Obstructive Pulmonary Disease. 2018;13:1691-700. doi:10.2147/COPD.S164775.
16. El-Gazzar AG, Kamel MH, Elbahnasy OKM, El-Naggar ME-S. Prognostic value of platelet and neutrophil to lymphocyte ratio in COPD patients. Expert Review of Respiratory Medicine. 2020;14(1):111-6. doi:10.1080/17476348.2020.1685387.
17. Shyam Prasad Shetty B, Chaya SK, Kumar V S, et al. Inflammatory biomarkers interleukin 1 beta (IL-1β) and tumour necrosis factor alpha (TNF-α) are differentially elevated in tobacco smoke associated COPD and biomass smoke associated COPD. Toxics. 2021;9(4):72. doi:10.3390/toxics9040072.
18. Wang Z, Locantore N, Haldar K, et al. Inflammatory endotype–associated airway microbiome in chronic obstructive pulmonary disease clinical stability and exacerbations: a multicohort longitudinal analysis. American Journal of Respiratory and Critical Care Medicine. 2021;203(12):1488-502. doi:10.1164/rccm.202009-3448OC.
19. Wu C-W, Wang L, Bolton S, et al. Progressive immunologic dysregulation with exposure to tobacco smoke in a model of chronic obstructive pulmonary disease (COPD). The Journal of Immunology. 2020;204(1_Supplement):74.17.
20. Mu Q, Wang Q, Yang Y, et al. HMGB1 promotes M1 polarization of macrophages and induces COPD inflammation. Cell Biology International. 2025;49(1):79-91. doi:10.1002/cbin.12247.
21. Feng X, Dong H, Li B, et al. Integrative analysis of the expression profiles of whole coding and non-coding RNA transcriptomes and construction of the competing endogenous RNA networks for chronic obstructive pulmonary disease. Frontiers in Genetics. 2023;14:1165243. doi:10.3389/fgene.2023.1165243.
22. Hernandez Cordero AI, Yang CX, Li X, et al. The blood DNA methylation clock GrimAge is a robust surrogate for airway epithelia aging. Biomedicines. 2022;10(12):3094. doi:10.3390/biomedicines10123094.
23. Armstrong DA, Chen Y, Dessaint JA, et al. DNA methylation changes in regional lung macrophages are associated with metabolic differences. ImmunoHorizons. 2019;3(7):274-81. doi:10.4049/immunohorizons.1900038
24. Li P, Peng J, Chen G, et al. DNA methylation profiling in a cigarette smoke-exposed mouse model of airway inflammation. International Journal of Chronic Obstructive Pulmonary Disease. 2022;17:2443-50. doi:10.2147/COPD.S374240.
25. Qiu J, Liu X, Yang G, et al. MiR-29b level-mediated regulation of Klotho methylation via DNMT3A targeting in chronic obstructive pulmonary disease. Cells & Development. 2023;174:203827. doi:10.1016/j.cdev. 2023.203827.
26. Qiu W, Baccarelli A, Carey VJ, et al. Variable DNA methylation is associated with chronic obstructive pulmonary disease and lung function. American Journal of Respiratory and Critical Care Medicine. 2012;185(4):373-81. doi:10.1164/rccm.201108-1382OC.
27. Nakahira K, Hisata S, Choi AM. The roles of mitochondrial damage-associated molecular patterns in diseases. Antioxidants & Redox Signaling. 2015;23(17):1329-50. doi:10.1089/ars.2015.6407.
28. Remels AH, Schrauwen P, Broekhuizen R, et al. Peroxisome proliferator-activated receptor expression is reduced in skeletal muscle in COPD. Euro Respir J. 2007;30(2):245-52. doi:10.1183/ 09031936.00115006.
29. Kaur G, Batra S. Regulation of DNA methylation signatures on NF-κB and STAT3 pathway genes and TET activity in cigarette smoke extract–challenged cells/COPD exacerbation model in vitro. Cell Biology and Toxicology. 2020;36(5):459-80. doi:10.1007/s10565-020-09520-w.
30. Jacobs M, Verschraegen S, Salhi B, et al. IL-10 producing regulatory B cells are decreased in blood from smokers and COPD patients. Respiratory Research. 2022;23(1):287. doi:10.1186/s12931-022-02213-y.
31. Masjedy A, Salesi M, Ahmadi A, et al. Association between single-nucleotide polymorphism of cytokines genes and chronic obstructive pulmonary disease: a systematic review and meta-analysis. Cytokine. 2023;171:156352. doi:10.1016/j.cyto.2023.156352.
32. Asgharzadeh M, Jigheh ZA, Kafil HS, et al. Association of interleukin-1 and interleukin-1 receptor antagonist gene polymorphisms with multiple sclerosis in Azeri population of Iran. Endocrine, Metabolic & Immune Disorders - Drug Targets. 2020;20(7):1110-6. doi:10.2147/COPD.S164775.
33. Asgharzadeh M, Najafi-Ghalehlou N, Poor BM, et al. IFN-γ and TNF-α gene polymorphisms in multiple sclerosis patients in northwest Iran. Endocrine, Metabolic & Immune Disorders - Drug Targets. 2021;21(3):520-5. doi:10.2174/1871530320666200610141630.
34. Asgharzadeh M, Fadaee M, Leylabadlo HE, et al. TNF-α gene polymorphism in Iranian Azeri population. Gene Reports. 2020;19:100651. doi:10.1016/j.genrep. 2020.100651.
35. Asgharzadeh M, Fadaee M, Mahdavipoor B, et al. Polymorphism of the IFN-γ gene in the Azeri population of Iran. Gene Reports. 2020;19:100596. doi:10.1016/j.genrep.2020.100596.
36. Obling N, Backer V, Hurst JR, Bodtger U. Nasal and systemic inflammation in chronic obstructive pulmonary disease (COPD). Respiratory Medicine. 2022;195:106774. doi:10.1016/j.rmed.2022.106774.
37. Zhang T, Wang G, Li Q, et al. Relationship between serum TH1/TH2 imbalance and depression in elderly patients with COPD and its clinical implications. Technology and Health Care. 2023;31(6):2047-58. doi:10.3233/THC-230009.
38. Jiang E, Fu Y, Wang Y, et al. The role and clinical significance of myeloperoxidase (MPO) and TNF-α in prognostic evaluation of T-COPD. BMC Pulmonary Medicine. 2025;25(1):192. doi:10.1186/s12890-025-03190-7.
39. Małujło-Balcerska E, Sipowicz K, Pietras T. Comparing chronic obstructive pulmonary disease and depressive disorder in terms of inflammation-related biomarkers. Archives of Medical Science: AMS. 2023;19(3):814-23. doi:10.5114/aoms/161494.
40. Amirkhosravi A, Salari E, Hashemi-Bajgani S-M, et al. Association of polymorphisms in IL-10, TGF-β1, IFN-γ, and TNF-α genes with the susceptibility to chronic obstructive pulmonary disease in Kerman, Iran. Journal of Kerman University of Medical Sciences. 2023;30(3):128-35. doi:10.22062/jkmu.2023.92193.
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Keywords
Chronic obstructive pulmonary disease Cytokines Epigenetics Inflammation Methylation
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1.
Faghih Ojaroodi A, Gholamzadeh N, Pouya K, Abbaspour-Aghdam S, Mikaeili H, Sadeghi A, Ahmadi M, Nadiri M. Cytokine Expression and Promoter Methylation Signatures Underlying Immune Dysregulation in Mild and Severe COPD. Iran J Allergy Asthma Immunol. 2026;:1-10.