{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,12]],"date-time":"2026-10-12T17:50:43Z","timestamp":1791827443119,"version":"4.3.4"},"reference-count":42,"publisher":"Public Library of Science (PLoS)","issue":"10","license":[{"start":{"date-parts":[[2026,10,12]],"date-time":"2026-10-12T00:00:00Z","timestamp":1791763200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    Non-obstructive azoospermia (NOA) is a severe male infertility disorder. This study aims to identify immune-related biomarkers for explore immune-related molecular signatures associated with NOA. Here, we integrated transcriptomic data from four independent cohorts (GSE145467, GSE45885, GSE45887, GSE9210) using an integrative pipeline combining differential expression analysis, weighted gene co-expression network analysis (WGCNA), and consensus-based multi-machine learning to identify an immune-related gene signature associated with NOA. We identified 16 immune-related differentially expressed genes through WGCNA, which were refined into a 12-gene panel\u2014\n                    <jats:italic>Hub NOA.Sig<\/jats:italic>\n                    (\n                    <jats:italic>CD14, CD19, CMTM2, CXCL16, HSPA1L, HSPA2, IGF1, IL13, POMC, RBP1, STAB2, TYROBP<\/jats:italic>\n                    )\u2014using five feature selection algorithms (Bagged Trees, Bayesian, Boruta, LASSO, LVQ). A Random Forest classifier based on\n                    <jats:italic>Hub NOA.Sig<\/jats:italic>\n                    yielded good classification performance with an AUC of 0.87. This signature offers a transcriptomic reference for characterizing NOA heterogeneity and exploring immune-related molecular features. Single-sample gene set enrichment analysis (ssGSEA) revealed significant alterations in six immune cell populations\u2014including effector memory CD4\u2009\u207a\u2009\/CD8\u2009\u207a\u2009T cells, regulatory T cells, and plasmacytoid dendritic cells\u2014and demonstrated strong correlations between hub genes and specific immune infiltrates. Furthermore, drug-gene interaction screening and molecular docking predicted loperamide-\n                    <jats:italic>POMC<\/jats:italic>\n                    , acitretin-\n                    <jats:italic>RBP1<\/jats:italic>\n                    , and retinol-\n                    <jats:italic>RBP1<\/jats:italic>\n                    as candidate drug-gene pairs for subsequent experimental validation. This work provides transcriptomic clues and candidate molecular targets for future mechanistic investigations into NOA-related male infertility.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0360305","type":"journal-article","created":{"date-parts":[[2026,10,12]],"date-time":"2026-10-12T17:25:31Z","timestamp":1791825931000},"page":"e0360305","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["Integrated bioinformatics analysis identifies an immune-related gene signature associated with non-obstructive azoospermia"],"prefix":"10.1371","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-1006-226X","authenticated-orcid":true,"given":"Yucheng","family":"Zhong","sequence":"first","affiliation":[{"name":"The Affiliated Guangdong Second Provincial General Hospital of Jinan University","place":["Guangzhou, China"],"department":["Department of Urology"]},{"name":"Assisted Reproductive Technology Center, The Affiliated Foshan Women and Children Hospital, Guangdong Medical University","place":["Foshan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Songqing","family":"Ye","sequence":"additional","affiliation":[{"name":"Affiliated Hospital Group of Guangdong Medical University Shenzhen Baoan Central Hospital","place":["Shenzhen, Guangdong, China"],"department":["Department of Urology"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hongshen","family":"Lai","sequence":"additional","affiliation":[{"name":"The Affiliated Guangdong Second Provincial General Hospital of Jinan University","place":["Guangzhou, China"],"department":["Department of Urology"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiangyu","family":"Zeng","sequence":"additional","affiliation":[{"name":"The Affiliated Guangdong Second Provincial General Hospital of Jinan University","place":["Guangzhou, China"],"department":["Department of Urology"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ge","family":"Song","sequence":"additional","affiliation":[{"name":"Assisted Reproductive Technology Center, The Affiliated Foshan Women and Children Hospital, Guangdong Medical University","place":["Foshan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaofu","family":"Qiu","sequence":"additional","affiliation":[{"name":"The Affiliated Guangdong Second Provincial General Hospital of Jinan University","place":["Guangzhou, China"],"department":["Department of Urology"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2026,10,12]]},"reference":[{"key":"pone.0360305.ref001","doi-asserted-by":"crossref","first-page":"161","DOI":"10.1007\/978-3-030-77779-1_8","article-title":"Male infertility in humans: An update on non-obstructive azoospermia (NOA) and obstructive azoospermia (OA)","volume":"1288","author":"X 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membrane CD14","volume":"6","author":"C Moreno","year":"2004","journal-title":"Microbes Infect"},{"issue":"6","key":"pone.0360305.ref027","doi-asserted-by":"crossref","first-page":"1670","DOI":"10.1002\/eji.200425932","article-title":"KARAP\/DAP12\/TYROBP: Three names and a multiplicity of biological functions","volume":"35","author":"E Tomasello","year":"2005","journal-title":"Eur J Immunol"},{"issue":"2","key":"pone.0360305.ref028","doi-asserted-by":"crossref","first-page":"267","DOI":"10.1189\/jlb.1003465","article-title":"Cell surface-anchored SR-PSOX\/CXC chemokine ligand 16 mediates firm adhesion of CXC chemokine receptor 6-expressing cells","volume":"75","author":"T Shimaoka","year":"2004","journal-title":"J Leukoc Biol"},{"issue":"12","key":"pone.0360305.ref029","doi-asserted-by":"crossref","first-page":"8172","DOI":"10.4049\/jimmunol.179.12.8172","article-title":"Critical role for CXC chemokine ligand 16 (SR-PSOX) in Th1 response mediated by NKT cells","volume":"179","author":"T Shimaoka","year":"2007","journal-title":"J Immunol"},{"issue":"10","key":"pone.0360305.ref030","doi-asserted-by":"crossref","first-page":"531","DOI":"10.1111\/pim.12127","article-title":"Fasciola hepatica tegumental antigens indirectly induce an M2 macrophage-like phenotype in vivo","volume":"36","author":"PN Adams","year":"2014","journal-title":"Parasite Immunol"},{"issue":"2","key":"pone.0360305.ref031","doi-asserted-by":"crossref","first-page":"713","DOI":"10.1093\/micmic\/ozac054","article-title":"Heat shock proteins are differentially expressed in the domestic cat (Felis catus) testis, epididymis, and vas deferens","volume":"29","author":"N Liman","year":"2023","journal-title":"Microsc Microanal"},{"issue":"2","key":"pone.0360305.ref032","doi-asserted-by":"crossref","first-page":"131","DOI":"10.21037\/atm-20-7616","article-title":"The association of CMTM6 expression with prognosis and PD-L1 expression in triple-negative breast cancer","volume":"9","author":"Y Tian","year":"2021","journal-title":"Ann Transl Med"},{"issue":"6","key":"pone.0360305.ref033","article-title":"Zika virus dysregulates human Sertoli cell proteins involved in spermatogenesis with little effect on tight junctions","volume":"14","author":"M-U Rashid","year":"2020","journal-title":"PLoS Negl Trop Dis"},{"issue":"11","key":"pone.0360305.ref034","doi-asserted-by":"crossref","first-page":"1263","DOI":"10.1038\/s41423-022-00925-7","article-title":"Serine metabolism orchestrates macrophage polarization by regulating the IGF1-p38 axis","volume":"19","author":"X Shan","year":"2022","journal-title":"Cell Mol Immunol"},{"issue":"2","key":"pone.0360305.ref035","doi-asserted-by":"crossref","first-page":"1885","DOI":"10.1007\/s11033-022-08179-2","article-title":"Cellular retinol-binding protein 1: A therapeutic and diagnostic tumor marker","volume":"50","author":"Z Guo","year":"2023","journal-title":"Mol Biol Rep"},{"issue":"5","key":"pone.0360305.ref036","doi-asserted-by":"crossref","first-page":"757","DOI":"10.1016\/j.clinthera.2020.03.008","article-title":"Stress-induced interaction of skin immune cells, hormones, and neurotransmitters","volume":"42","author":"N Pondeljak","year":"2020","journal-title":"Clin Ther"},{"issue":"11","key":"pone.0360305.ref037","doi-asserted-by":"crossref","first-page":"1029","DOI":"10.1038\/gt.2013.34","article-title":"Immunotherapy with gene-modified T cells: Limiting side effects provides new challenges","volume":"20","author":"HJ Stauss","year":"2013","journal-title":"Gene Ther"},{"key":"pone.0360305.ref038","doi-asserted-by":"crossref","first-page":"118173","DOI":"10.1016\/j.ecoenv.2025.118173","article-title":"Environmental toxicant 2,3,7,8-tetrachlorodibenzo-p-dioxin induces non-obstructive azoospermia: New insights from network toxicology, integrated machine learning, and biomolecular modeling","volume":"295","author":"L Xu","year":"2025","journal-title":"Ecotoxicol Environ Saf"},{"key":"pone.0360305.ref039","doi-asserted-by":"crossref","DOI":"10.7554\/eLife.97958.3","article-title":"scRNA-seq and scATAC-seq reveal that Sertoli cell mediates spermatogenesis disorders through stage-specific communications in non-obstructive azoospermia","volume":"13","author":"S Wang","year":"2025","journal-title":"Elife"},{"issue":"1","key":"pone.0360305.ref040","first-page":"2","article-title":"Testicular biopsy score count--a method for registration of spermatogenesis in human testes: Normal values and results in 335 hypogonadal males","volume":"1","author":"SG Johnsen","year":"1970","journal-title":"Hormones"},{"issue":"2","key":"pone.0360305.ref041","doi-asserted-by":"crossref","first-page":"67","DOI":"10.1038\/s41585-023-00837-9","article-title":"Molecular mechanisms of cellular dysfunction in testes from men with non-obstructive azoospermia","volume":"21","author":"A Piechka","year":"2024","journal-title":"Nat 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