色谱

色谱 ›› 2025, Vol. 43 ›› Issue (1): 96-103.DOI: 10.3724/SP.J.1123.2024.05022

• 研究论文 • 上一篇    

多维方法评价全氟辛酸对人结直肠癌细胞的毒性效应

张睿佳1,2, 林颖诗2, 涂蓝尹2, 陈梓潼4, 张维蔚5,*(), 栾天罡2, 陈保卫3,*()   

  1. 1.海南大学生命健康学院, 海南 海口 570228
    2.中山大学生命科学学院, 广东 广州 510275
    3.中山大学海洋科学学院, 广东 珠海 519082
    4.中山大学人工智能学院, 广东 珠海 519082
    5.广州市疾病控制中心, 广东 广州 510440
  • 收稿日期:2024-05-24 出版日期:2025-01-08 发布日期:2024-12-26
  • 通讯作者: E-mail:chenbw5@mail.sysu.edu.cn(陈保卫); E-mail:249726849@qq.com(张维蔚).
  • 基金资助:
    国家自然科学基金(22206207);国家自然科学基金(22127810);国家自然科学基金(22276224);广州市科技计划(2024A04J3793);广州市科技计划(2024A03J0414)

Evaluation of the toxicity of perfluorooctanoic acid toward human colorectal cancer cells using multi-dimensional approaches

ZHANG Ruijia1,2, LIN Yingshi2, TU Lanyin2, CHEN Zitong4, ZHANG Weiwei5,*(), LUAN Tiangang2, CHEN Baowei3,*()   

  1. 1. School of Life and Health Sciences, Hainan University, Haikou 570228, China
    2. School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China
    3. School of Marine Sciences, Sun Yat-sen University, Zhuhai 519082, China
    4. School of Artificial Intelligence, Sun Yat-sen University, Zhuhai 519082, China
    5. Guangzhou Center for Disease Control and Prevention, Guangzhou 510440, China
  • Received:2024-05-24 Online:2025-01-08 Published:2024-12-26
  • Supported by:
    National Natural Science Foundation of China(22206207);National Natural Science Foundation of China(22127810);National Natural Science Foundation of China(22276224);Science Program Foundation of Guangzhou(2024A04J3793);Science Program Foundation of Guangzhou(2024A03J0414)

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摘要：

全氟辛酸(PFOA)的暴露与溃疡性结肠炎的发生密切相关,但目前缺少PFOA暴露对人结直肠癌细胞(HCT116)产生毒性效应的相关分子机理研究。本研究从细胞毒性表型、细胞呼吸和代谢相关基因转录水平3个层次评价了PFOA对HCT116的毒性效应。首先,利用水溶性甲臜化合物(MTS)来评价PFOA暴露对HCT116细胞相对活性的影响;随后,利用细胞外流量分析仪对HCT116的线粒体呼吸活性进行测定;最后,用定量即时聚合酶链式反应(qPCR)对HCT116中代谢相关基因的转录水平进行检测。细胞毒性实验结果表明,与对照组相比,经高浓度(300 μmol/L)PFOA暴露48 h后,HCT116的细胞活性受到显著抑制(p<0.01),并在G0/G1细胞周期受到阻滞,而低浓度(30、50 μmol/L)的PFOA反而提高了细胞相对活性;低浓度(50 μmol/L)的PFOA能够促进HCT116的线粒体呼吸活性。利用自主开发的Metabolic Gene and Pathway Query检索软件和比较毒理基因组学数据库,本研究发现代谢相关基因二肽酶1(DPEP1)和鞘甘氨酸-1激酶(SPHK1)与PFOA引起的溃疡性结肠炎相关。qPCR实验结果表明,高浓度(300 μmol/L)PFOA能够显著诱导DPEP1和SPHK1的转录表达上调(约8~10倍),低浓度(50 μmol/L)PFOA未引起DPEP1和SPHK1转录表达水平的变化。本文发现细胞线粒体呼吸活性是评价低浓度PFOA干扰效应的一个敏感指标,DPEP1和SPHK1介导的细胞代谢过程可能是PFOA引起肠道毒性的潜在机制。

关键词: 全氟辛酸, 人结直肠肠道毒性, 代谢紊乱, 代谢基因和通路检索软件

Abstract:

While human exposure to perfluorooctanoic acid (PFOA) can lead to ulcerative colitis, the molecular mechanisms responsible for PFOA-induced intestinal toxicity are unclear. Herein, we examined the toxicity of PFOA toward human colorectal cancer cells (HCT116) from three dimensions: the cytotoxic phenotype, cell respiration, and transcription levels of metabolism-related genes. Formazan was used to assess how PFOA exposure affects HCT116-cell relative viability, after which the mitochondrial respiratory activities of these cells were determined by analyzing extracellular flux. The quantitative real-time polymerase chain reaction (qPCR) method was used to detect metabolism-related gene expression levels. The cytotoxicity assay revealed that the HCT116 showed significantly inhibited relative activities compared to those of the control when exposed to 300 μmol/L PFOA for 48 h (p<0.01), with most cells retained at the G0/G1 stage. In contrast, the mitochondrial respiratory activities of the HCT116 were promoted by concentrations of PFOA as low as 50 μmol/L. Two genes related to cellular metabolism (dipeptidase 1 (DPEP1) and sphingosine kinase 1 (SPHK1)) were found to be related to the PFOA-promoted formation of ulcerative colitis using our self-developed Metabolic Gene and Pathway Query software and Comparative Toxicogenomics Database (CTD). The qPCR studies revealed that DPEP1 and SPHK1 expression levels were enhanced by 8-10 times in HCT116 exposed to 300 μmol/L PFOA relative to the control, whereas this trend was not observed for HCT116 exposed to 50 μmol/L PFOA. Collectively, these results suggest that the respiratory activity of cellular mitochondria may serve as an index for determining the interference effects associated with PFOA and that metabolic pathways mediated by DPEP1 and SPHK1 may be involved in the development of PFOA-induced ulcerative colitis. Future studies should investigate the relationships between changes in metabolism-related genes (DPEP1 and SPHK1) and the mitochondrial respiratory activities of intestinal cells, and verify the roles played by the DPEP1 and SPHK1 genes in PFOA-induced intestinal inflammation using in-vivo models.

Key words: perfluorooctanoic acid (PFOA), human colorectal enterotoxicity, metabolic disorder, Metabolic Gene and Pathway Query software

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