Han Zhang ¹ Jiaxing Dai ^2* Qiuqiao Mu ^3* Xiaojiang Zhao ¹ Kai Wang ^1* Meng Wang ^1* Daqiang Sun ^1*

• ¹ Tianjin University Chest Hospital, Tiaijin, China
• ² Tianjin Medical College, Tianjin, China
• ³ Tianjin Medical University, Tianjin, Tianjin Municipality, China

Macrophages play a dual role in the tumor microenvironment(TME), capable of secreting proinflammatory factors to combat tumors while also promoting tumor growth through angiogenesis and immune suppression. This study aims to explore the characteristics of macrophages in lung adenocarcinoma (LUAD) and establish a prognostic model based on macrophage-related genes.We performed scRNA-seq analysis to investigate macrophage heterogeneity and their potential pseudotime evolutionary processes. Specifically, we used scRNA-seq data processing, intercellular communication analysis, pseudotime trajectory analysis, and transcription factor regulatory analysis to reveal the complexity of macrophage subpopulations. Data from The Cancer Genome Atlas (TCGA) was used to assess the impact of various macrophage subtypes on LUAD prognosis.Univariate Cox regression was applied to select prognostic-related genes from macrophage markers.We constructed a prognostic model using Lasso regression and multivariate Cox regression, categorizing LUAD patients into high and low-risk groups based on the median risk score. The model's performance was validated across multiple external datasets. We also examined differences between high and low-risk groups in terms of pathway enrichment, mutation information, tumor microenvironment(TME), and immunotherapy efficacy. Finally, RT-PCR confirmed the expression of model genes in LUAD, and cellular experiments explored the carcinogenic mechanism of COL5A1.We found that signals such as SPP1 and MIF were more active in tumor tissues, indicating potential oncogenic roles of macrophages. Using macrophage marker genes, we developed a robust prognostic model for LUAD that effectively predicts prognosis and immunotherapy efficacy. A nomogram was constructed to predict LUAD prognosis based on the model's risk score and other clinical features.Differences between high and low-risk groups in terms of TME, enrichment analysis, mutational landscape, and immunotherapy efficacy were systematically analyzed. RT-PCR and cellular experiments supported the oncogenic role of COL5A1.Our study identified potential oncogenic mechanisms of macrophages and their impact on LUAD prognosis. We developed a prognostic model based on macrophage marker genes, demonstrating strong performance in predicting prognosis and immunotherapy efficacy. Finally, cellular experiments suggested COL5A1 as a potential therapeutic target for LUAD.