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中国癌症防治杂志

中国癌症防治杂志 ›› 2024, Vol. 16 ›› Issue (4): 433-.doi: 10.3969/j.issn.1674-5671.2024.04.08

• 临床研究 • 上一篇    下一篇

基于单细胞RNA测序技术的肝癌手术前后外周血B细胞基因表达谱研究

  

  1. 广西医科大学公共卫生学院流行病与卫生统计学教研室；广西医科大学公共卫生学院儿少卫生与妇幼保健医学教研室；广西医科大学护理学院临床内科护理学教研室；广西医科大学附属肿瘤医院内分泌代谢肾病内科
  • 出版日期:2024-08-25 发布日期:2024-08-23
  • 通讯作者: 曾小云 E-mail：zengxiaoyun@gxmu.edu.cn
  • 基金资助:
    国家自然科学基金项目（82060616）

Gene expression profiling of peripheral blood B cells before and after surgery for hepatocellular carcinoma based on single-cell RNA sequencing technology

  • Online:2024-08-25 Published:2024-08-23

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摘要： 目的 探索肝细胞癌（hepatocellular carcinoma，HCC）患者手术前后外周血B细胞基因表达谱的变化。 方法 收集2022年9月在广西医科大学附属肿瘤医院行肝切除术治疗的3例HCC患者手术前及手术后的外周血样本，提取外周血单个核细胞（peripheral blood mononuclear cell，PBMC）并进行单细胞RNA测序（single⁃cell RNA sequencing，scRNA⁃seq）。根据测序数据进行B细胞数量分析、上下调基因差异分析、KEGG分析、GSEA⁃KEGG及GSEA⁃GO分析和细胞通讯分析，比较手术前后B细胞的动态改变。结果 PBMC经scRNA⁃seq分析识别出了标记基因为MS4A1、CD79A的细胞簇，即B细胞群。B细胞群重新聚类分析后可分为记忆B（memory B）细胞、初始B（naive B）细胞和浆母细胞（plasmablasts）共3个细胞亚群。与术前组相比，术后组naive B和plasmablasts细胞占比升高，memory B细胞占比降低。手术前后B细胞中共筛选出285个显著差异表达基因（differentially expressed genes，DEGs），其中S100A8、S100A9、IL1R2、MALAT1等152个基因在手术后显著上调（P<0.05），RPS27A、RPS3A、RPS12、RPS3等133个基因在手术后显著下调（P<0.05）。KEGG分析发现手术后B细胞中显著下调的DEGs主要富集到核糖体、抗原处理和提呈通路。GSEA⁃KEGG分析发现手术后B细胞中表达上调的基因集显著富集于癌症中的转录失调、P53信号通路，表达下调的基因集显著富集于核糖体、抗原处理和提呈通路；GSEA⁃GO分析发现手术后B细胞中表达上调的基因集显著富集于环氧化酶P450途径、信号模式识别受体活性，表达下调的基因集显著富集于免疫球蛋白复合物、T细胞受体复合物。手术后B细胞与T细胞之间的通讯增强，且手术后配体⁃受体对CD22⁃PTPRC在B细胞与B细胞、B细胞与Mono细胞之间的通讯概率上调（P<0.01）。无论是手术前还是手术后，CD22信号通路对B细胞与T细胞之间的通讯均具有较强调控能力。结论 HCC患者外周血B细胞基因表达谱在手术前后发生动态变化且与外周血B细胞免疫功能相关，可为研究外周血B细胞的抗肿瘤免疫作用提供新的参考依据。

关键词: 单细胞RNA测序, B细胞, 肝癌, 基因表达谱

Abstract: Objective To investigate the changes of gene expression profiles of peripheral blood B cells in hepatocellular carcinoma (HCC) patients before and after surgery. Methods Peripheral blood samples of 3 HCC patients who underwent hepatectomy for treatment at the Guangxi Medical University Cancer Hospital in September 2022 were collected before and after surgery, peripheral blood monoculear cell (PBMC) were extracted and single⁃cell RNA sequencing (scRNA⁃seq) was performed. B cell number analysis, up⁃ and down⁃regulated gene difference analysis, KEGG analysis, GSEA⁃KEGG and GSEA⁃GO analysis, and cell communication analysis were performed based on the sequencing data to compare the dynamic changes of B cells before and after surgery. Results PBMC was analyzed by scRNA⁃seq to identify clusters of cells with marker genes MS4A1 and CD79A, namely, B⁃cell clusters. After regrouping analysis, the B⁃cell clusters were classified into three cell subpopulations, namely, memory B cells, naive B cells, and plasmablasts. Compared with pre⁃operative group, the percentage of naive B cells and plasmablasts increased, and the percentage of memory B cells decreased after operation. A total of 285 significantly differentially expressed genes (DEGs) were screened in B cells before and after surgery, of which 152 genes including S100A8, S100A9, IL1R2 and MALAT1 were significantly up⁃regulated after surgery (P<0.05), and 133 genes including RPS27A, RPS3A, RPS12 and RPS3 were significantly down⁃regulated after surgery(P<0.05). KEGG analysis revealed that significantly down⁃regulated DEGs in post⁃operative B cells were mainly enriched in ribosomal, antigen processing and presentation pathways. GSEA⁃KEGG analysis revealed that the gene set up⁃regulated in post⁃operative B cells was significantly enriched in transcriptional dysregulation and P53 signaling pathway in cancer, while the gene set down⁃regulated was significantly enriched in ribosomal, antigen processing and presentation pathways. GSEA⁃GO analysis revealed that the gene set up⁃regulated in post⁃operative B cells was significantly enriched in cyclooxygenase P450 pathway, signaling pattern recognition receptor activity, and the down⁃regulated gene set was significantly enriched in immunoglobulin complex and T⁃cell receptor complex. Communication between B cells and T cells was enhanced after surgery, and the communication probability of post⁃operative ligand and receptor pairs of CD22⁃PTPRC between B cells and B cells, and between B cells and Mono cells was up⁃regulated (P<0.01). Either before or after surgery, the CD22 signaling pathway had a strongly ability to regulate the communication between B cells and T cells. Conclusions The gene expression profile of peripheral blood B cells in HCC patients changes dynamically before and after surgery, and is correlated with the immune function of peripheral blood B cells. This result provides a new reference for the study of the anti⁃tumor immune function of peripheral blood B cells.

Key words: Single-cell RNA sequencing, B cells, Hepatocellular carcinoma, Gene expression profiling

中图分类号: 

  • R735.7