全氟辛酸促进鼻咽癌细胞HK1增殖和迁移的机制研究

Study on the role of perfluorooctanoic acid in promoting the proliferation and migration of nasopharyngeal carcinoma cell line HK1

摘要:

目的 探究全氟辛酸(PFOA)对鼻咽癌细胞 HK1 生物学行为的影响及其相关机制。方法 采用50 μM浓度PFOA连续处理HK1细胞10 d，通过细胞计数试剂盒-8(CCK-8)法检测0、24、48、72 h细胞增殖能力，并利用Transwell实验评估细胞迁移能力；随后对PFOA处理组及对照组细胞进行RNA测序(RNA-seq)分析，筛选差异表达基因(DEGs)，采用实时荧光定量聚合酶链反应(qPCR)对关键DEGs进行验证；对DEGs进行京都基因与基因组百科全书(KEGG)、基因本体(GO)功能富集分析。结果 随培养时间延长，对照组和PFOA处理组的细胞活力均呈上升趋势，至72 h时，PFOA处理组的细胞活力显著高于对照组(P<0.05)。Transwell实验显示，PFOA组的迁移细胞数显著多于对照组(P<0.05)。RNA-seq共筛选出891个DEGs，其中上调279个，下调612个；qPCR验证结果显示，HBEGF、EPHA2、SPRY4、IL1B等关键上调DEGs的mRNA及蛋白表达水平与测序结果一致(P<0.05)。功能富集分析表明，DEGs主要富集于Toll样受体信号通路、细胞因子-细胞因子受体相互作用、白细胞介素(IL)-17信号通路及相关自身免疫性疾病通路等。结论 在HK1细胞模型中，PFOA可促进鼻咽癌细胞的增殖和迁移，其作用可能与调控HBEGF、EPHA2等基因及炎症相关信号通路有关，为后续探索鼻咽癌环境风险因素提供了初步实验线索。

Abstract:

Objective To investigate the effect of perfluorooctanoic acid (PFOA) on the biological behavior of nasopharyngeal carcinoma (NPC) cell line HK1 and its potential mechanism of action. Methods HK1 cells were continuously treated with 50 μM concentration PFOA for 10 days. The cell proliferation ability at 0, 24, 48 and 72 hours was detected by the cell counting Kit-8 (CCK-8) method, and the cell migration ability was evaluated by Transwell assay. Subsequently, RNA sequencing (RNA-seq) analysis was performed on the cells of the PFOA-treated group and the control group to screen for differentially expressed genes (DEGs). The key DEGs were verified by real-time fluorescence quantitative polymerase chain reaction (qPCR). Functional enrichment analyses of the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) were conducted on DEGs. Results With the extension of culture time, the cell viability of both the control group and the PFOA treatment group showed an upward trend. By 72 hours, the cell viability of the PFOA treatment group was significantly higher than that of the control group (P<0.05). The Transwell assay showed that the number of migrating cells in the PFOA group was significantly higher than that in the control group (P<0.05). A total of 891 DEGs were screened out by RNA-seq, among which 279 were upregulated and 612 were downregulated. The results of qPCR and WB verification showed that the expression levels of mRNA and protein of key up-regulating DEGs such as HBEGF, EPHA2, SPRY4, and IL1B were consistent with the sequencing results (P<0.05). Functional enrichment analysis indicated that DEGs were mainly enriched in Toll-like receptor signaling pathways, cytokine-cytokine receptor interactions, IL-17 signaling pathways, and related autoimmune diseases pathways. Conclusions In the HK1 cell model, PFOA can promote the proliferation and migration of NPC cells. Its effect may be related to the regulation of DEGs such as HBEGF and EPHA2 and inflammation-related signaling pathways, providing preliminary experimental clues for further exploration of environmental risk factors for NPC.