对批量测序和单细胞测序的综合分析揭示TNFSF9是微卫星不稳定型胃腺癌的潜在调节因子。

Integrative analysis of bulk and single-cell sequencing reveals TNFSF9 as a potential regulator in microsatellite instability stomach adenocarcinoma.

作者信息

Zhou Jianlong, Zhang Yucheng, Liu Yongfeng, Li Jiehui, Zhang Wenxing, Wang Junjiang, Yao Xueqing, Feng Huolun, Zheng Jiabin, Li Yong

机构信息

Department of Gastrointestinal Surgery, Department of General Surgery, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, 510080, Guangdong, China.

Department of General Surgery, Guangdong Provincial People's Hospital Ganzhou Hospital (Ganzhou Municipal Hospital), Ganzhou, 341000, China.

出版信息

Eur J Med Res. 2025 Mar 28;30(1):214. doi: 10.1186/s40001-025-02471-0.

DOI:10.1186/s40001-025-02471-0

PMID:40148957

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11951761/

Abstract

BACKGROUND

Stomach adenocarcinoma (STAD) with microsatellite instability (MSI) is associated with a better prognosis compared to Non-MSI. This study aims to elucidate the differences in the tumor microenvironment (TME) of MSI and explore its underlying mechanisms in STAD.

METHODS

TME differences between MSI and Non-MSI were analyzed using single-cell RNA sequencing (MSI = 7, Non-MSI = 19) and bulk RNA sequencing (MSI = 39, Non-MSI = 198). Differentially expressed genes (DEGs) were used to identify enriched pathways and hub genes. TNFSF9 expression was validated by immunohistochemistry (IHC) on 23 STAD sections (MSI = 13, Non-MSI = 10) and confirmed in tumor epithelial cells using SNU-1 (MSI) and AGS (Non-MSI) cell lines through quantitative polymerase chain reaction (qPCR) and Western blot (WB).

RESULTS

The results showed MSI was significantly associated with a better prognosis (P < 0.05). Within the TME, MSI was associated with a higher abundance of antigen-presenting cells, including M1 macrophages (40.1% vs. 27.9%) and activated dendritic cells (22.1% vs. 10.5%), as well as pro-inflammatory Th1-like CD4⁺ T cells (15% vs. 11%). However, MSI also showed an increase in exhausted T cells, indicating a complex immune landscape. Signaling pathway and cell communication analyses revealed an enrichment of cytokine-related pathways in MSI. Hub gene analysis revealed that TNFSF9 was predominantly expressed in stromal cells and partially in tumor epithelial cells in MSI, with its upregulation further confirmed through IHC, qPCR, and WB. Correlation analysis demonstrated a positive relationship between TNFSF9 expression and the abundance of M1 macrophages.

CONCLUSIONS

These findings provide new insights into the TME of MSI in STAD, emphasizing the significant role of TNFSF9 in shaping MSI-specific TME, enhancing immunotherapy efficacy, and improving patient survival.

摘要

背景

与微卫星稳定（Non-MSI）的胃腺癌（STAD）相比，微卫星不稳定（MSI）的胃腺癌预后更好。本研究旨在阐明MSI的肿瘤微环境（TME）差异，并探索其在STAD中的潜在机制。

方法

使用单细胞RNA测序（MSI = 7，Non-MSI = 19）和批量RNA测序（MSI = 39，Non-MSI = 198）分析MSI和Non-MSI之间的TME差异。使用差异表达基因（DEG）来鉴定富集的通路和枢纽基因。通过对23个STAD切片（MSI = 13，Non-MSI = 10）进行免疫组织化学（IHC）验证TNFSF9表达，并通过定量聚合酶链反应（qPCR）和蛋白质免疫印迹（WB）在SNU-1（MSI）和AGS（Non-MSI）细胞系的肿瘤上皮细胞中进行确认。

结果

结果显示MSI与更好的预后显著相关（P < 0.05）。在TME中，MSI与更高丰度的抗原呈递细胞相关，包括M1巨噬细胞（40.1%对27.9%）和活化的树突状细胞（22.1%对10.5%），以及促炎性Th1样CD4⁺ T细胞（15%对11%）。然而，MSI中耗竭性T细胞也有所增加，表明免疫格局复杂。信号通路和细胞通讯分析显示MSI中细胞因子相关通路富集。枢纽基因分析显示TNFSF9主要在MSI的基质细胞中表达，部分在肿瘤上皮细胞中表达，通过IHC、qPCR和WB进一步证实其上调。相关性分析表明TNFSF9表达与M1巨噬细胞丰度之间存在正相关。