Table 2_Combined single-cell transcriptome and Mendelian randomization to identify and validate prognostic genes associated with endoplasmic reticulum stress and butyrate metabolism in lung adenocarcinoma.xlsx

BackgroundLung adenocarcinoma (LUAD) is a prevalent and aggressive subtype of lung cancer, with a 5-year survival rate below 20% due to late-stage diagnosis and drug resistance. Endoplasmic reticulum stress (ERS) and butyrate metabolism (BM) play critical roles in tumor progression, but their co-regulatory features in LUAD remain unclear.
MethodsThis study integrated single-cell transcriptome analysis and Mendelian randomization (MR) to identify prognostic genes associated with ERS and BM in LUAD. Public datasets were analyzed using weighted gene co-expression network analysis, differential expression analysis, and MR. A risk model and nomogram were constructed, and immune microenvironment, gene set enrichment, and single-cell analyses were performed to validate findings. Moreover, the expression of prognostic genes was validated in different Non-small cell lung cancer (NSCLC) cell lines through reverse transcription quantitative polymerase chain reaction (RT-qPCR).
ResultsSeven prognostic genes (VDAC1, TXNRD1, GDF15, TRIB3, LPL, KCNQ1, PKP2) were identified, RT-qPCR assays confirmed that these genes exhibited significant expression differences in different NSCLC cell lines. The risk model demonstrated that low-risk patients had significantly better survival outcomes. The nomogram exhibited strong predictive accuracy for 1-, 3-, and 5-year survival. Enriched pathways in high-risk patients included olfactory transduction, while low-risk patients showed enrichment in ribosome and complement-coagulation cascades. Immune profiling revealed 13 differentially abundant immune cell types, including M1 macrophages. Single-cell analysis identified macrophages as key players in LUAD. Notably, VDAC1, TXNRD1, and LPL were highly expressed during early macrophage differentiation.
ConclusionThis study identifies seven ERS- and BM-related prognostic genes and highlights macrophages as pivotal in LUAD progression, the expression differences of candidate genes were verified by RT-qPCR assay. These findings provide novel insights into LUAD diagnosis, prognosis, and potential therapeutic targets, offering a foundation for precision medicine strategies. Further validation in clinical cohorts and functional studies is warranted to translate these discoveries into clinical applications.

背景：肺腺癌（Lung adenocarcinoma, LUAD）是一种高发且侵袭性强的肺癌亚型，因确诊时多处于晚期且易产生耐药性，其5年生存率不足20%。内质网应激（Endoplasmic reticulum stress, ERS）与丁酸代谢（Butyrate metabolism, BM）在肿瘤进展中发挥关键作用，但二者在LUAD中的协同调控特征仍不明确。
方法：本研究整合单细胞转录组分析与孟德尔随机化（Mendelian randomization, MR）技术，筛选与LUAD中ERS及BM相关的预后基因。研究通过加权基因共表达网络分析、差异表达分析及孟德尔随机化方法对公共数据集进行分析；构建了风险预测模型与列线图（nomogram），并通过免疫微环境分析、基因集富集分析及单细胞分析对研究结果进行验证。此外，本研究通过逆转录定量聚合酶链反应（Reverse transcription quantitative polymerase chain reaction, RT-qPCR），在不同非小细胞肺癌（Non-small cell lung cancer, NSCLC）细胞系中验证了预后基因的表达水平。
结果：本研究共鉴定出7个预后基因（VDAC1、TXNRD1、GDF15、TRIB3、LPL、KCNQ1、PKP2），RT-qPCR实验证实上述基因在不同NSCLC细胞系中均存在显著表达差异。风险模型分析显示，低风险组患者的生存预后显著优于高风险组。列线图对LUAD患者1年、3年及5年生存率均具有良好的预测准确性。高风险组患者富集的通路包括嗅觉传导通路，而低风险组则显著富集核糖体通路与补体-凝血级联通路。免疫细胞浸润分析共鉴定出13种差异丰度免疫细胞类型，其中包括M1型巨噬细胞。单细胞分析揭示巨噬细胞是LUAD进展中的关键细胞群。值得注意的是，VDAC1、TXNRD1及LPL在巨噬细胞早期分化阶段呈现高表达。
结论：本研究筛选得到7个与ERS及BM相关的LUAD预后基因，并强调巨噬细胞在LUAD进展中的核心作用，候选基因的表达差异已通过RT-qPCR实验得到验证。本研究结果为LUAD的诊断、预后评估及潜在治疗靶点开发提供了全新视角，为精准医疗策略的制定奠定了基础。后续需在临床队列中开展进一步验证及功能学研究，以推动上述研究成果向临床应用转化。