系统性红斑狼疮患者免疫细胞代谢通路转录组分析

doi:10.3969/j.issn.1674-8115.2022.09.006

上海交通大学学报（医学版） ›› 2022, Vol. 42 ›› Issue (9): 1197-1207.doi: 10.3969/j.issn.1674-8115.2022.09.006

• 创新团队成果专栏 • 上一篇

系统性红斑狼疮患者免疫细胞代谢通路转录组分析

韩夏夏(), 蒋扬, 顾霜霜, 戴黛, 沈南()

上海交通大学医学院附属仁济医院风湿科，上海风湿病学研究所，上海 200127

收稿日期:2022-03-28 接受日期:2022-06-16 出版日期:2022-09-28 发布日期:2022-09-28
通讯作者: 沈南 E-mail:xiaxiahan66@163.com;nanshensibs@gmail.com
作者简介:韩夏夏（1995—），女，硕士生；电子信箱：xiaxiahan66@163.com。
基金资助:
国家自然科学基金(31630021);上海市高水平地方高校创新团队(SSMU-ZDCX 20180100)

Transcriptomic analysis of metabolic characteristics of the immune cells in systemic lupus erythematosus patients

HAN Xiaxia(), JIANG Yang, GU Shuangshuang, DAI Dai, SHEN Nan()

Department of Rheumatology, Renji Hospital, Shanghai Jiao Tong University School of Medicine; Shanghai Institute of Rheumatology, Shanghai 200127, China

Received:2022-03-28 Accepted:2022-06-16 Online:2022-09-28 Published:2022-09-28
Contact: SHEN Nan E-mail:xiaxiahan66@163.com;nanshensibs@gmail.com
Supported by:
National Natural Science Foundation of China(31630021);Innovative Research Team of High Level Local Universities in Shanghai(SSMU-ZDCX20180100)

摘要/Abstract

摘要：

目的·利用生物信息学方法研究系统性红斑狼疮（systemic lupus erythematosus，SLE）患者免疫细胞亚群的代谢通路活化水平。方法·从基因表达综合数据库（Gene Expression Omnibus，GEO）中下载SLE患者和正常人外周血单个核细胞（peripheral blood mononuclear cell，PBMC）的基因表达矩阵，以及SLE患者和正常人外周血中T细胞和B细胞亚群相应的基因表达矩阵，对测序数据进行标准化后筛选差异表达基因（differentially-expressed gene，DEG）。应用Enrichr在线富集工具对差异表达基因进行通路分析，并通过比对分析确定共同上调通路。借助基因集富集分析（gene set enrichment analysis， GSEA）软件对全基因表达矩阵进行通路富集分析。对SLE和正常人PBMC的转录测序数据进行免疫细胞的构成比例分析。对目的通路进行基因表达注释。运用基于转座酶和高通量测序的染色质分析（assay for transposase-accessible chromatin using sequencing，ATAC-seq）技术检测SLE患者和正常人B细胞中糖酵解相关基因的染色质开放性情况。结果·①维恩图展示SLE患者PBMC基因表达谱（GSE169080、GSE144390以及GSE139350）富集了139个共同上调的通路， GSEA分析结果显示其中包括糖酵解、氧化磷酸化（oxidative phosphorylation， OXPHOS）以及脂肪酸氧化（fatty acid oxidation，FAO）在内的多条代谢通路在SLE患者中上调。②免疫细胞比例构成分析发现，与正常人相比，SLE患者外周血中T细胞、B细胞以及NK细胞的比例较高；SLE患者T细胞和B细胞中多个代谢通路关键酶基因的表达水平均较正常人高。③SLE患者和正常人外周血B细胞的ATAC-seq结果显示，糖酵解关键酶基因SLC2A3、PKM以及LDHA的开放性在SLE患者中更高。④SLE患者B细胞亚群的代谢通路GSEA分析结果和代谢通路可视化分析显示，记忆B细胞和浆母细胞的多个代谢通路水平均较初始B细胞高。结论·SLE患者转录水平的多条代谢通路均发生改变，效应B细胞的代谢通路活化水平较初始B细胞高。

关键词: 系统性红斑狼疮, 代谢通路, 基因表达综合数据库, 生物信息学, B细胞

Abstract:

Objective ·To study the metabolic pathway activity level of the immune cell subsets in the patients with systemic lupus erythematosus (SLE) by bioinformatics analysis. Methods ·The matrix expression data of PBMCs collected from SLE patients and healthy controls were downloaded from Gene Expression Omnibus (GEO) Datasets, as well as the transcriptome data of T cell and B cell subsets from SLE patients and healthy controls. The differentially-expressed genes (DEGs) were identified in the standardized sequence data. Pathway enrichment analysis of DEGs was performed by online Enrichr tools, and the common up-regulated pathways were determined by comparative analysis. Gene set enrichment analysis (GSEA) was used to identify pathways that were enriched in the experiment processed with the whole gene expression matrix. RNA-seq data from PBMCs samples of SLE patients and healthy controls were used to characterize the immune cell composition. The targeted pathway was annotated with gene expression. Assay for transposase-accessible chromatin using sequencing (ATAC-seq) technique was performed to detect the chromatin accessibility of glycolysis-related genes in SLE patients and healthy controls. Results ·① Venn diagram depicted 139 common upregulated pathways in GSE169080, GSE144390 and GSE139350 data sets, and GSEA results showed that multiple classical metabolic pathways, including glycolysis, oxidative phosphorylation (OXPHOS) and fatty acid oxidation (FAO), were up-regulated in SLE patients. ② Immune cell composition analysis of PBMCs showed that the proportions of T cells, B cells and NK cells were higher in SLE patients, and the expression of genes encoding multiple enzymes of metabolic pathway in T cells and B cells were higher than those in healthy controls. ③ Compared to healthy controls, the intensity of ATAC-seq signal was significantly enhanced at transcriptional regulatory sites of SLC2A3, PKM and LDHA in peripheral B cells from SLE patients. ④ GSEA results and visualization analysis of metabolic pathways of SLE B cell subsets showed that the memory B cells and plasmablasts displayed a higher metabolic state than na?ve B cells. Conclusion ·Multiple metabolic pathways are altered in SLE patients and the metabolic level of effector B cells is higher than na?ve B cells in SLE patients.

Key words: systemic lupus erythematous (SLE), metabolic pathway, Gene Expression Omnibus (GEO), bioinformatics, B cell

中图分类号:

R593.24⁺1

韩夏夏, 蒋扬, 顾霜霜, 戴黛, 沈南. 系统性红斑狼疮患者免疫细胞代谢通路转录组分析[J]. 上海交通大学学报（医学版）, 2022, 42(9): 1197-1207.

HAN Xiaxia, JIANG Yang, GU Shuangshuang, DAI Dai, SHEN Nan. Transcriptomic analysis of metabolic characteristics of the immune cells in systemic lupus erythematosus patients[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2022, 42(9): 1197-1207.

图/表 7

图1 研究流程图

Fig 1 Flow chart of the study

图2 SLE患者PBMC的代谢通路分析Note： A. The overlap analysis of the upregulated gene sets in GSE169080, GSE139350 and GSE144390. B. Expression levels of the pathways commonly upregulated in GSE169080. C. Gene set enrichment analysis (GSEA) of GSE169080 showed that the KEGG_GLYCOLYSIS_GLUCONEOGENESIS, GO_OXIDATIVE_PHOSPHORYLATION and WP_FATTY_ACID_BETA_OXIDATION were upregulated in SLE patients compared to healthy controls (HCs). D. Heatmap of GSE169080 showed the expression level of key genes encoding the glycolytic and OXPHOS enzymes of SLE patients and HCs.

Fig 2 Metabolic pathways analysis of PBMC in SLE patients

图3 SLE患者免疫细胞亚群通路分析Note： A. The boxplot showed the fraction of multiple immune cell subsets in GSE162828. B/C. Hallmark sets analysis of upregulated DEGs in T cells (B) and B cells (C) of SLE patients compared with healthy controls.

Fig 3 Pathway analysis of immune cell subsets in SLE patients

图4 SLE患者免疫细胞亚群的代谢通路活化水平Note：A/B. Comparison of the expression of key genes encoding the glycolysis, OXPHOS and FAO enzymes in T cells (A) and B cells (B) between SLE patients and healthy controls.

Fig 4 Metabolic pathway activity levels of immune cell subsets in SLE patients

图5 SLE患者和健康者外周血B细胞中糖酵解通路关键酶基因 SLC2A3 、 PKM 和 LDHA 的ATAC-seq可视化信号

Fig 5 Normalized ATAC-seq signal of SLC2A3, PKM and LDHA, encoding the glycolytic enzyme in peripheral B cells from SLE patients and healthy controls

图6 SLE患者B细胞亚群的代谢通路水平Note：A/B. Gene set enrichment analysis (GSEA) of GSE156751 showed that the KEGG_GLYCOLYSIS_GLUCONEOGENESIS, GO_OXIDATIVE_ PHOSPHORYLATION and REACTOME_MITOCHONDRIAL_FATTY_ACID_BETA_OXIDATION were upregulated in memory B cells (A) and plasmablasts (B) compared with na?ve B cells.

Fig 6 Metabolic pathway activity levels of B cell subsets in SLE patients

图7 SLE患者浆母细胞代谢通路水平Note： A. Interactive network analysis of upregulated pathways of plasmablasts compared to memory B cells. B. The gene expression level of OXPHOS pathway were upregulated in plasmablasts compared to memory B cells. Red and green represent up- and down-regulated genes, respectively.

Fig 7 Metabolic pathway activity level of plasmablasts in SLE patients

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系统性红斑狼疮患者免疫细胞代谢通路转录组分析

Transcriptomic analysis of metabolic characteristics of the immune cells in systemic lupus erythematosus patients

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