南方医科大学学报

南方医科大学学报 ›› 2023, Vol. 43 ›› Issue (2): 232-241.doi: 10.12122/j.issn.1673-4254.2023.02.11

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载脂蛋白E通过激活ERK/MMP9信号通路促进子宫内膜癌细胞的迁移

吴超英，程文俊   

  1. 南京医科大学常州医学中心，常州市妇幼保健院妇科，江苏 常州 213000；南京医科大学第一附属医院妇科，江苏 南京 210029
  • 出版日期:2023-02-20 发布日期:2023-03-16

Apolipoprotein E enhances migration of endometrial cancer cells byactivating the ERK/MMP9 signaling pathway

WU Chaoying, CHENG Wenjun   

  1. Department of Gynecology, Changzhou Maternity and Child Health Care Hospital, Changzhou Medical Center, Nanjing Medical University, Changzhou 213000, China; Department of Gynecology,First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China
  • Online:2023-02-20 Published:2023-03-16

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摘要： 目的 研究载脂蛋白E（APOE）在子宫内膜癌转移中的具体作用及分子调控机制。方法 子宫内膜癌HEC-1B细胞常规培养，空白对照组siRNA以及APOE靶向的siRNA转染子宫内膜癌HEC-1B细胞，以及采用空载质粒和APOE过表达质粒转染子宫内膜癌HEC-1B细胞，分为空白对照组siCtrl、siAPOE组、空载质粒组、APOE过表达质粒组。利用划痕实验和Transwell实验检测子宫内膜癌细胞的迁移能力，通过MTT实验及流式细胞术验证APOE对子宫内膜癌细胞增殖能力的影响，使用Hoechst染色法检测干扰APOE后HEC-1B细胞的凋亡情况，采用qPCR、Western blotting观察ERK/MMP9信号通路的变化；通过免疫组化的方法分析APOE的表达与子宫内膜癌组织分化程度的相关性。结果 划痕实验和Transwell小室实验结果显示，与空白对照组相比，转染了siAPOE的HEC-1B细胞的迁移能力均明显下降（P<0.05）。与空载质粒组相比，过表达APOE能显著促进子宫内膜癌细胞HEC-1B的迁移能力（P<0.05）。MTT实验及流式细胞术结果显示APOE对细胞增殖无影响。Hoechst染色实验结果表明干扰APOE后HEC-1B细胞的凋亡能力没有受到影响。Western blotting结果显示敲低APOE能够降低子宫内膜癌细胞中MMP9的蛋白表达水平（P<0.05），而过表达APOE 能使细胞内MMP9的蛋白水平升高（P<0.05）；U0126能够抑制过表达APOE对MMP9的促进作用（P<0.05）。免疫组化结果显示，APOE在子宫内膜癌组织的表达水平比癌旁组织高。结论 APOE在子宫内膜癌组织中呈高表达，APOE可通过影响ERK的磷酸化，促进子宫内膜癌细胞中MMP9的表达，促进子宫内膜癌细胞的迁移。

关键词: APOE；ERK；子宫内膜癌；细胞迁移

Abstract: Objective To study the role of apolipoprotein E (APOE) in regulating endometrial cancer metastasis and explore the signaling pathway in the regulatory mechanism. Methods Human endometrial cancer cell line HEC-1B was transfected with a control siRNA (siCtrl) or a specific siRNA targeting APOE (siAPOE) or with either pEGFP-N1 plasmid or an APOE-overexpressing plasmid. The changes in migration, proliferation, apoptosis and cell cycle of the transfected cells were examined using wound healing assay, Transwell migration assay, MTT assay, flow cytometry, and Hoechst staining. The activity of the ERK/MMP9 signaling pathway in the transfected cells was assessed using RT-qPCR and Western blotting. The expression level of APOE in clinical specimens of endometrial cancer tissues were detected using immunohistochemistry and its correlation with differentiation of endometrial cancer tissues was analyzed. Results Wound healing assay and Transwell migration assay showed that compared with those in siCtrl group, HEC-1B cells transfected with siAPOE showed significantly reduced migration ability (P<0.05), whereas APOE overexpression significantly promoted the migration of the cells (P<0.05). Neither APOE knockdown nor overexpression produced significant effects on HEC-1B cell proliferation as shown by MTT assay and flow cytometry. Hoechst staining revealed that transfection with siAPOE did not significantly affect apoptosis of HEC- 1B cells. APOE knockdown obviously reduced and APOE overexpression enhanced ERK phosphorylation and MMP9 expression in HEC-1B cells (P<0.05). Treatment with U0126 partially reversed the effects of APOE overexpression on ERK phosphorylation, migration and MMP9 expression in HEC-1B cells (P<0.05). APOE is highly expressed in clinical samples of endometrial cancer tissues as compared with the adjacent tissues. Conclusion APOE is highly expressed in endometrial cancer tissues to promote cancer cell migration by enhancing ERK phosphorylation and MMP9 expression.

Key words: apolipoprotein E; extracellular signal-regulated kinases; endometrial carcinoma; cell migration