高糖诱导腹膜间皮细胞凋亡的蛋白质组学研究

doi:10.3969/j.issn.1674-8115.2026.10.003

上海交通大学学报（医学版） ›› 2026, Vol. 46 ›› Issue (10): 1331-1338.doi: 10.3969/j.issn.1674-8115.2026.10.003

• 论著 · 基础研究 • 上一篇

高糖诱导腹膜间皮细胞凋亡的蛋白质组学研究

李若琳¹, 李子萌², 宋阿会³, 陶冶¹, 李辰²(), 刘英莉¹()

^1.上海交通大学医学院附属第九人民医院肾脏内科，上海 200011
^2.上海交通大学医学院单细胞组学与疾病研究中心，上海 201318
^3.上海交通大学医学院附属第九人民医院内分泌代谢科，上海 200011

收稿日期:2026-01-14 接受日期:2026-04-13 出版日期:2026-10-19 发布日期:2026-10-19
通讯作者: 李辰，研究员，博士；电子信箱：cli@shsmu.edu.cn。
刘英莉，主任医师，博士；电子信箱：liuyingli@sjtu.edu.cn。
基金资助:
国家自然科学基金(82170760);国家自然科学基金(82373446);国家自然科学基金(82502656);国家自然科学基金(32671158);上海市科学技术委员会基金(23ZR1435800);上海市科学技术委员会基金(21DZ2201700)

Proteomic analysis of high glucose-induced apoptosis in peritoneal mesothelial cells

Li Ruolin¹, Li Zimeng², Song Ahui³, Tao Ye¹, Li Chen²(), Liu Yingli¹()

^1.Department of Nephrology, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011, China
^2.Center for Single-Cell Omics, Shanghai Jiao Tong University School of Medicine, Shanghai 201318, China
^3.Department of Endocrinology and Metabolism, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011, China

Received:2026-01-14 Accepted:2026-04-13 Online:2026-10-19 Published:2026-10-19
Contact: Li Chen, E-mail: cli@shsmu.edu.cn.
Liu Yingli, E-mail: liuyingli@sjtu.edu.cn.
Supported by:
National Natural Science Foundation of China(82170760);Fund of Science and Technology Commission of Shanghai Municipality(23ZR1435800)

摘要/Abstract

摘要：

目的·基于蛋白质组学技术探究高糖诱导腹膜间皮细胞（peritoneal mesothelial cell，PMC）凋亡的生物学机制及分子靶点。方法·采用人PMC系HMrSV5，将其分为正常对照组和高糖组（138 mmol/L葡萄糖处理），刺激24 h后收集细胞蛋白。利用非标记定量蛋白质组学技术，分析高糖刺激下HMrSV5细胞蛋白质表达谱变化，并对差异表达蛋白进行信号通路富集和蛋白质相互作用（protein-protein interaction，PPI）网络构建。采用CCK-8（cell counting kit-8）法检测细胞增殖活力，流式细胞术检测细胞凋亡率，Western blotting验证关键蛋白检查点激酶2（checkpoint kinase 2，CHEK2）的表达及其磷酸化水平。结果·蛋白质组学共鉴定到7 366种蛋白质，主成分分析显示对照组与高糖组间存在显著差异。与对照组相比，高糖组筛选出417个差异表达蛋白（differentially expressed proteins，DEPs），包括237个上调DEPs和180个下调DEPs。亚细胞定位分析显示，上调DEPs主要分布于细胞质、线粒体和核质，下调DEPs主要定位于核质和细胞质。基因本体论（Gene Ontology，GO）功能注释及京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes，KEGG）通路富集分析结果显示，DEPs显著富集于有丝分裂细胞周期、P53信号通路、细胞衰老等细胞周期相关通路。基因集富集分析（gene set enrichment analysis，GSEA）进一步提示P53信号通路在高糖组显著正向富集，而E2F靶基因通路与G₂/M检查点通路显著负向富集。PPI网络分析显示，CHEK2与多种细胞周期相关DEPs存在相互作用，可能是潜在关键蛋白。CCK-8及流式细胞实验结果表明，高糖刺激可降低HMrSV5细胞活力并诱导其凋亡；Western blotting结果提示，高糖刺激可显著上调CHEK2表达并提升其磷酸化水平。结论·高糖刺激可能通过影响PMC的细胞周期调控进而促进其凋亡，其中CHEK2可能是关键调控蛋白。

关键词: 腹膜透析, 蛋白质组学, 高糖, 细胞凋亡, 细胞周期, 检查点激酶2

Abstract:

Objective ·To investigate the biological mechanisms and molecular targets of high glucose-induced apoptosis in peritoneal mesothelial cells (PMCs) using proteomics. Methods ·The human peritoneal mesothelial cell line HMrSV5 was employed in this study. Cells were divided into a control group (untreated) and a high glucose group (treated with 138 mmol/L glucose). Cellular proteins were collected after 24 h of stimulation. Label-free quantitative proteomics was utilized to analyze changes in the protein expression profile of HMrSV5 cells under high glucose stimulation. Differentially expressed proteins (DEPs) were subjected to pathway enrichment analysis and protein-protein interaction (PPI) network construction. Cell proliferation viability was measured using the CCK-8 assay, and the apoptosis rate was detected by flow cytometry. Western blotting was performed to verify the expression and phosphorylation levels of the key protein checkpoint kinase 2 (CHEK2). Results ·A total of 7 366 proteins were identified. Principal component analysis (PCA) revealed significant differences between the control and glucose-treated groups. Compared with the control group, 417 DEPs were identified in the high glucose group, comprising 237 upregulated and 180 downregulated proteins. Subcellular localization analysis showed that upregulated DEPs were primarily distributed in the cytoplasm, nucleoplasm, and mitochondria, while downregulated DEPs were mainly located in the nucleoplasm and cytoplasm. Gene Ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses indicated that DEPs were significantly enriched in cell cycle-related pathways, including the mitotic cell cycle, P53 signaling pathway, and cellular senescence. Gene Set Enrichment Analysis (GSEA) further suggested that the P53 signaling pathway was significantly positively enriched in the high glucose group, whereas the E2F target gene pathway and G₂/M checkpoint pathway were negatively enriched. PPI network analysis revealed that CHEK2 interacted with multiple cell cycle-related DEPs, suggesting it may be a potential key protein. CCK-8 and flow cytometry assays showed that high glucose stimulation reduced HMrSV5 cell viability and induced apoptosis. Western blotting results confirmed that high glucose stimulation significantly upregulated CHEK2 expression and increased its phosphorylation levels. Conclusion ·High glucose stimulation may promote apoptosis in PMCs by affecting cell cycle regulation, and CHEK2 may be a key regulatory protein.

Key words: peritoneal dialysis, proteomics, high glucose, apoptosis, cell cycle, checkpoint kinase 2 (CHEK2)

中图分类号:

R692.5

李若琳, 李子萌, 宋阿会, 陶冶, 李辰, 刘英莉. 高糖诱导腹膜间皮细胞凋亡的蛋白质组学研究[J]. 上海交通大学学报（医学版）, 2026, 46(10): 1331-1338.

Li Ruolin, Li Zimeng, Song Ahui, Tao Ye, Li Chen, Liu Yingli. Proteomic analysis of high glucose-induced apoptosis in peritoneal mesothelial cells[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2026,(10): 1331-1338.

图/表 4

图1 高糖刺激HMrSV5细胞后蛋白质组学表达谱的变化Note： A. Number of identified protein groups (PGs) in each group (n=4). The HMrSV5 cells were treated with 138 mmol/L glucose for 24 h. B. Pearson correlation coefficients between each pair of biological replicates in each group. C. The PCA plot demonstrates global proteomic variations between the control group and the glucose group. D. The volcano plot illustrates the DEPs detected by proteomics. E. Subcellular distribution of DEPs.

Fig 1 Proteomic alterations in high glucose-treated HMrSV5 cells

图2 高糖刺激HMrSV5细胞后DEPs的功能富集分析Note： A. GO enrichment analysis of DEPs, including biological processes, cellular components, and molecular functions. Bubble size represents the number of enriched proteins, and color indicates -log10(P value). B. KEGG pathway enrichment analysis of DEPs. The length of the bar represents gene counts, and the color denotes -log10(P value). C. GSEA results showed that the P53 pathway was positively enriched, whereas E2F targets and the G2/M checkpoint pathways were negatively enriched in the glucose group compared with the control group.

Fig 2 Functional enrichment analysis of DEPs in glucose-treated HMrSV5 cells

图3 高糖刺激HMrSV5细胞后细胞周期相关DEPs的PPI网络分析及表达量变化Note： A. PPI network of cell cycle-related DEPs. Red nodes indicate upregulated proteins, and blue nodes indicate downregulated proteins. B. Heatmap of DEPs enriched in cell cycle pathway. C. Log-transformed intensities of representative proteins across the control and the glucose groups. norm.int.—normalized intensity.

Fig 3 PPI network and expression patterns of cell cycle-related DEPs in glucose-treated HMrSV5 cells

图4 高糖刺激HMrSV5细胞后细胞活性、凋亡水平及CHEK2磷酸化水平的变化Note： A. Cell viability was assessed using the CCK-8 assay. B. Apoptosis was evaluated by Annexin Ⅴ-FITC/PI double staining and analyzed by flow cytometry. C. Protein expressions of p-CHEK2 and CHEK2 were evaluated by Western blotting.

Fig 4 Changes in cell viability, apoptosis level, and CHEK2 phosphorylation level after high-glucose stimulation of HMrSV5 cells

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高糖诱导腹膜间皮细胞凋亡的蛋白质组学研究

Proteomic analysis of high glucose-induced apoptosis in peritoneal mesothelial cells

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