Table3_CD40LG and GZMB were correlated with adipose tissue macrophage infiltration and involved in obstructive sleep apnea related metabolic dysregulation: Evidence from bioinformatics analysis.DOCX

Both obesity and obstructive sleep apnea (OSA) can lead to metabolic dysregulation and systemic inflammation. Similar to obesity, increasing evidence has revealed that immune infiltration in the visceral adipose tissue (VAT) is associated with obstructive sleep apnea-related morbidity. However, the pathological changes and potential molecular mechanisms in visceral adipose tissue of obstructive sleep apnea patients need to be further studied. Herein, by bioinformatics analysis and clinical validation methods, including the immune-related differentially expressed genes (IRDEGs) analysis, protein-protein interaction network (PPI), functional enrichment analysis, a devolution algorithm (CIBERSORT), spearman’s correlation analysis, polymerase chain reaction (PCR), Enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC), we identified and validated 10 hub IRDEGs, the relative mRNA expression of four hub genes (CRP, CD40LG, CCL20, and GZMB), and the protein expression level of two hub genes (CD40LG and GZMB) were consistent with the bioinformatics analysis results. Immune infiltration results further revealed that obstructive sleep apnea patients contained a higher proportion of pro-inflammatory M1 macrophages and a lower proportion of M2 macrophages. Spearman’s correlation analysis showed that CD40LG was positively correlated with M1 macrophages and GZMB was negatively correlated with M2 macrophages. CD40LG and GZMB might play a vital role in the visceral adipose tissue homeostasis of obstructive sleep apnea patients. Their interaction with macrophages and involved pathways not only provides new insights for understanding molecular mechanisms but also be of great significance in discovering novel small molecules or other promising candidates as immunotherapies of OSA-associated metabolic complications.

肥胖与阻塞性睡眠呼吸暂停（obstructive sleep apnea, OSA）均可引发代谢失调与全身炎症。与肥胖类似，越来越多研究表明，内脏脂肪组织（visceral adipose tissue, VAT）中的免疫浸润与阻塞性睡眠呼吸暂停相关发病风险存在关联。然而，阻塞性睡眠呼吸暂停患者内脏脂肪组织的病理变化及潜在分子机制仍有待进一步探究。本研究通过生物信息学分析与临床验证手段，包括免疫相关差异表达基因（immune-related differentially expressed genes, IRDEGs）分析、蛋白质相互作用网络（protein-protein interaction network, PPI）构建、功能富集分析、反卷积算法（CIBERSORT）、斯皮尔曼相关性分析，以及聚合酶链式反应（polymerase chain reaction, PCR）、酶联免疫吸附试验（Enzyme-linked immunosorbent assay, ELISA）与免疫组织化学（immunohistochemistry, IHC）实验，最终筛选并验证了10个核心免疫相关差异表达基因；其中4个核心基因（CRP、CD40LG、CCL20及GZMB）的相对mRNA表达水平，与2个核心基因（CD40LG与GZMB）的蛋白表达水平，均与生物信息学分析结果一致。免疫浸润分析结果进一步显示，阻塞性睡眠呼吸暂停患者体内促炎型M1巨噬细胞比例更高，而M2巨噬细胞比例更低。斯皮尔曼相关性分析表明，CD40LG与M1巨噬细胞呈正相关，GZMB则与M2巨噬细胞呈负相关。CD40LG与GZMB可能在阻塞性睡眠呼吸暂停患者的内脏脂肪组织稳态维持中发挥关键作用。二者与巨噬细胞的相互作用及参与的信号通路，不仅为阐明阻塞性睡眠呼吸暂停相关分子机制提供了新视角，同时为开发针对阻塞性睡眠呼吸暂停相关代谢并发症的新型小分子免疫治疗药物或其他潜在候选疗法具有重要意义。