奇异南星挥发油通过miR-762/NF2轴对甲状腺癌细胞恶性生物学行为的影响

doi:10.11904/j.issn.1002-3070.2023.03.003

实用肿瘤学杂志 ›› 2023, Vol. 37 ›› Issue (3): 207-214.doi: 10.11904/j.issn.1002-3070.2023.03.003

奇异南星挥发油通过miR-762/NF2轴对甲状腺癌细胞恶性生物学行为的影响

张慧¹, 闫旺², 孙明华³

1.廊坊市人民医院检验科(廊坊 065000);
2.张北县医院外科;
3.华北医疗集团峰峰总医院外科

收稿日期:2022-03-15 修回日期:2023-01-05 出版日期:2023-06-28 发布日期:2023-08-07
通讯作者: 张慧,E-mail:yangyangz2@163.com
作者简介:张慧,女,(1977-),本科,副主任检验师,从事检验管理方面的研究。
基金资助:
河北省卫健委项目(编号:20201301)

Effects of arisaema decipiens volatile oil on the malignant biological behavior of thyroid cancer cells through a miR-762/NF2 axis

ZHANG Hui¹, YAN Wang², SUN Minghua³

1. Department of Laboratory,Langfang People's Hospital,Langfang 065000,China;
2. Department of Surgery,Zhangbei County Hospital;
3. Department of Surgery,Fengfeng General Hospital of North China Medical Group

Received:2022-03-15 Revised:2023-01-05 Online:2023-06-28 Published:2023-08-07

摘要/Abstract

摘要： 目的探讨奇异南星挥发油(ADVO)对甲状腺癌细胞恶性生物学行为的影响及可能作用机制。方法用含不同浓度ADVO培养液培养甲状腺癌TPC-1细胞48 h,CCK-8法检测细胞存活率,筛选最适作用浓度;TPC-1细胞分为空白组(常规培养)、ADVO组(60 μg/mL ADVO)、miR-762 mimics组(转染miR-762 mimics)、miR-762 mimics NC组(转染miR-762 mimics NC)、ADVO+miR-762 mimics组(60 μg/mL ADVO+转染miR-762 mimics)、ADVO+miR-762 mimics NC组(60 μg/mL ADVO+转染miR-762 mimics NC)并培养,行CCK-8法、流式细胞术、Transwell小室实验;双荧光素酶报告基因实验验证miR-762与2型神经纤维瘤(Neurofibromatosis type 2,NF2)基因的靶向关系;RT-qPCR法检测miR-762及NF2 mRNA相对表达水平;Western blot实验检测Merlin、Yes相关蛋白1(YAP1)、p-YAP1、裂解半胱氨酸蛋白酶-3(C-caspase-3)、E钙粘附蛋白(E-cadherin)、波形蛋白(Vimentin)相对表达水平。结果 TPC-1细胞存活率随ADVO作用浓度增大而降低(P＜0.05),60 μg/mL ADVO浓度作用下的细胞存活率接近50%;与空白组比较,ADVO组细胞存活率、miR-762及YAP1、Vimentin蛋白相对表达水平降低,细胞凋亡率、NF2 mRNA与Merlin、p-YAP1、C-caspase-3和E-cadherin蛋白相对表达水平升高、细胞迁移及侵袭数减少(P＜0.05),miR-762 mimics组细胞存活率、miR-762及YAP1、Vimentin蛋白相对表达水平升高,细胞凋亡率、NF2 mRNA与Merlin、p-YAP1、C-caspase-3和E-cadherin蛋白相对表达水平降低、细胞迁移及侵袭数增加(P＜0.05);ADVO可减弱miR-762过表达的作用效果,抑制TPC-1细胞恶性生物学行为(P＜0.05);经生物信息学预测和双荧光素酶报告基因实验验证,NF2基因可能为miR-762的潜在靶基因。结论 ADVO可抑制TPC-1细胞增殖、侵袭及迁移并促进其凋亡,其作用机制可能与抑制miR-762表达并提高其靶基因NF2编码Merlin蛋白水平,激活相关抑癌信号有关。

关键词: 奇异南星挥发油, 甲状腺癌, 微小RNA-762, 2型神经纤维瘤基因

Abstract: Objective The aim of this study was to investigate the effects and possible mechanism of arisaema decipiens volatile oil(ADVO)on the malignant biological behavior of thyroid cancer cells. Methods Human thyroid cancer TPC-1 cells were cultured in medium containing different concentrations of ADVO for 48 hours,and the cell viability was detected by CCK-8 method to screen for the optimal concentration.TPC-1 cells were divided into the blank group(conventional culture)and ADVO group(60 μg/mL),miR-762 mimics group(transfected with miR-762 mimics),miR-762 mimics NC group(transfected with miR-762 mimics NC),ADVO+miR-762 mimics group(60 μg/mL+transfected with miR-762 mimics),and ADVO+miR-762 mimics NC group(60 μg/mL+transfected with miR-762 mimics NC).The CCK-8 assay,flow cytometry and Transwell assays were performed in this study.The dual luciferase reporter gene assay was used to verify the targeting relationship between the miR-762 and neurofibromatosis type 2(NF2)gene.The relative expression of miR-762 and NF2 mRNA was detected by RT-qPCR.The expression of merlin,Yes-related protein 1(YAP1),p-YAP1,cleavage cysteine protease-3(C-caspase-3),E-cadherin and vimentin proteins was detected by Western blot. Results The viability of TPC-1 cells decreased with the increase of ADVO concentration(P＜0.05),and the viability of TPC-1 cells at the concentration of 60 μg/mL of ADVO was close to 50%.Compared with the blank group,the cell viability,the expression of miR-762,YAP1 and vimentin protein decreased,the apoptotic rate,the expression of NF2 mRNA and Merlin,p-YAP1,C-caspase-3 and E-cadherin proteins increased,and the number of cell migration and invasion decreased in the ADVO group(P＜0.05).ADVO could weaken the overexpression of miR-762 and inhibit the malignant biological behavior of TPC-1 cells(P＜0.05).According to bioinformatics prediction and the dual luciferase reporter gene assay,NF2 gene might be a potential target gene of miR-762. Conclusion ADVO can inhibit the proliferation,invasion and migration of TPC-1 cells and promote their apoptosis.The mechanism of action may be related to inhibiting the expression of miR-762,increasing the level of Merlin protein encoded by its target gene NF2,and activating related tumor suppressor signals.

Key words: Arisaema decipiens volatile oil, Thyroid cancer, microRNA 762, Neurofibromatosis type 2 gene

中图分类号:

R739.4

张慧, 闫旺, 孙明华. 奇异南星挥发油通过miR-762/NF2轴对甲状腺癌细胞恶性生物学行为的影响[J]. 实用肿瘤学杂志, 2023, 37(3): 207-214.

ZHANG Hui, YAN Wang, SUN Minghua. Effects of arisaema decipiens volatile oil on the malignant biological behavior of thyroid cancer cells through a miR-762/NF2 axis[J]. Journal of Practical Oncology, 2023, 37(3): 207-214.

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奇异南星挥发油通过miR-762/NF2轴对甲状腺癌细胞恶性生物学行为的影响

Effects of arisaema decipiens volatile oil on the malignant biological behavior of thyroid cancer cells through a miR-762/NF2 axis

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