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目的 探讨Y-盒结合蛋白1(YB1)介导多药耐药基因1(MDR1)表达，从而调控肝内胆管细胞癌（ICC）顺铂诱导耐药性产生的机制。方法 在不同浓度的顺铂处理人ICC细胞HCCC-9810后，采用Western blot和免疫荧光技术检测YB1和磷酸化YB1(pYB1)的表达及其细胞定位的变化，并通过qPCR和Western blot方法分析耐药相关蛋白MDR1、多药耐药相关蛋白2(MRP2)和主要穹隆体蛋白（MVP）的表达变化。使用siRNA敲低HCCC-9810中YB1表达后，通过MTT检测其对顺铂和吉西他滨药物敏感性变化，并使用Western blot检测MDR1、MRP2和MVP表达变化。染色质免疫沉淀分析检测顺铂干预前后HCCC-9810细胞中YB1和MDR1启动子结合情况。通过亚硫酸氢盐测序检测顺铂干预前后HCCC-9810细胞MDR1启动子胞嘧啶-磷酸-鸟嘌呤（CpG）岛甲基化情况。结果 HCCC-9810细胞经顺铂干预后，胞内YB1蛋白总量并没有改变，但pYB1水平逐渐升高，同时伴随YB1和pYB1的核迁移。HCCC-9810细胞经顺铂诱导后，MDR1表达量显著提高，细胞耐药性增加；若预先抑制MDR1表达，则能够逆转细胞耐药性增加。同样，敲低HCCC-9810中YB1表达后，YB1和pYB1蛋白核迁移量显著减少，且MDR1表达量显著下降，对顺铂化疗敏感性显著增加。HCCC-9810细胞未受外界刺激时，MDR1启动子区域CpG岛呈高甲基化状态，抑制包括YB1在内的多种转录因子结合。当HCCC-9810细胞受顺铂干预后，MDR1启动子区域CpG岛发生去甲基化，继而胞核内YB1与MDR1启动子结合力增强，激活MDR1基因表达。结论 YB1高表达于ICC细胞质中，顺铂化疗诱导YB1核转位，同时伴随MDR1启动子CpG岛去甲基化，核YB1结合MDR1启动子并激活转录，最终导致化疗耐药性产生。
Abstract：Objective To explore the mechanism by which Y-box binding protein 1(YB1) mediates the expression of multidrug resistance gene 1(MDR1) and regulates cisplatin-induced resistance in intrahepatic cholangiocarcinoma(ICC).Methods After treating human ICC HCCC-9810 cells with different concentrations of cisplatin, Western blot and immunofluorescence techniques were used to detect the expression of YB1 and phosphorylated YB1(pYB1) and changes in their cellular localization. In addition, qPCR and Western blot were used to analyze the expression changes of drug resistance-related proteins MDR1, multidrug resistance-associated protein 2(MRP2), and major vault protein(MVP). Chromatin immunoprecipitation(ChIP) assay was performed to examine the binding of YB1 to the MDR1 promoter in HCCC-9810 cells with or without cisplatin treatment. Additionally, bisulfite sequencing(BS-seq) was used to determine the methylation status of cytosine-phosphate-guanine(CpG) islands within the MDR1 promoter in HCCC-9810 cells, before and after cisplatin treatment.Results After HCCC-9810 cells were treated with cisplatin, the total amount of intracellular YB1 protein did not change, but the level of pYB1 gradually increased, accompanied by the nuclear migration of YB1 and pYB1 protein. After HCCC-9810 cells were induced with cisplatin, the expression of MDR1 significantly increased, and the drug resistance of the cells also increased. If the expression of MDR1 was inhibited beforehand, the increase in drug resistance of the cells can be reversed. Similarly, after knocking down the expression of YB1 in HCCC-9810 cells, the nuclear migration of YB1 and pYB1 significantly decreased, and the expression of MDR1 significantly decreased, resulting in a significant increase in sensitivity to cisplatin chemotherapy. When HCCC-9810 cells were not stimulated by external factors, the CpG island in the promoter region of MDR1 showed a high methylation state, and multiple transcription factors, including YB1, could not bind to it. When HCCC-9810 cells were treated with cisplatin, the CpG island in the promoter region of MDR1 underwent demethylation, and the binding force between YB1 and the MDR1 promoter in the nucleus increased, activating MDR1 expression.Conclusion YB1 is highly expressed in the cytoplasm of ICC cells. Cisplatin chemotherapy induces nuclear translocation of YB1, accompanied by demethylation of the CpG island in the MDR1 promoter. Nuclear YB1 binds to the MDR1 promoter and activates transcription, ultimately leading to chemotherapy resistance.
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基本信息:
DOI：10.19405/j.cnki.issn1000-1492.2026.08.007
中图分类号:R735.7
引用信息:
[1]王思琦,程淼,罗欣语,等.YB1介导MDR1调控肝内胆管细胞癌顺铂诱导耐药机制研究[J].安徽医科大学学报,2026,61(08):1383-1391.DOI:10.19405/j.cnki.issn1000-1492.2026.08.007.
基金信息:
安徽高校自然科学研究重点项目（编号：KJ2021A0280）~~
2026-07-20
2026-07-20
2026-07-20