辛伐他汀联合干扰miR-21对唾液腺腺样囊性癌细胞迁移和侵袭的影响

doi:10.19438/j.cjoms.2018.02.007

中国口腔颌面外科杂志 ›› 2018, Vol. 16 ›› Issue (2): 132-137.doi: 10.19438/j.cjoms.2018.02.007

辛伐他汀联合干扰miR-21对唾液腺腺样囊性癌细胞迁移和侵袭的影响

王超¹, 黎婷¹, 严斐¹, 蔡文燕², 蒋星宇¹, 孙晋虎¹

1.徐州医科大学口腔医学院,江苏徐州 221000;
2.南京医科大学附属儿童医院口腔科,江苏南京 210000;

收稿日期:2017-10-30 修回日期:2017-12-26 出版日期:2018-03-20 发布日期:2018-04-08
通讯作者: 孙晋虎,E-mail: kqxsjh@163.com
作者简介:王超（1992-）,男,硕士研究生,E-mail: wc452788757@163.com
基金资助:
江苏省高校"青蓝工程"中青年学术带头人资助项目（51934）; 徐州医学院优秀人才基金资助项目（2013012）

Effect of simvastatin combined with simiR-21 on migration and invasion of human salivary adenoid cystic carcinoma

WANG Chao¹, LI Ting¹, YAN Fei¹, CAI Wen-yan², JIANG Xing-yu¹, SUN Jin-hu¹

1.Department of Oral Medicine, School of Stomatology, Xuzhou Medical University. Xuzhou 221000;
2.Department of Stomatology, Children's Hospital Affiliated to Nanjing Medical University. Nanjing 210000, Jiangsu Province, China;

Received:2017-10-30 Revised:2017-12-26 Online:2018-03-20 Published:2018-04-08

摘要/Abstract

摘要： 目的: 探讨辛伐他汀（simvastatin, SIM）联合干扰miR-21对唾液腺腺样囊性癌细胞株（SACC-LM）迁移、侵袭的影响。 方法: 体外培养SACC-LM细胞,随机分为阴性对照组（negative control,NC）、IC₅₀辛伐他汀组（SIM）、miR-21抑制剂转染组（simiR-21）和联合处理组（simiR-21+SIM）。利用qRT-PCR检测4组细胞中 miR-21的表达水平,CCK8法检测不同浓度辛伐他汀（0、10、20、30、40、50、60 μmol/L）作用48 h后的细胞活性,得出48 h的IC₅₀;观察干扰miR-21后SACC-LM对辛伐他汀的耐药性变化。Transwell法观察细胞迁移（不加基质胶）、侵袭能力（加入基质胶）的变化。Western免疫印迹检测EMT途径相关蛋白E-Cadherin、Snail1的表达。应用SPSS 22.0软件包对所得数据进行t检验或单因素方差分析。 结果: 与阴性对照组相比,转染组及联合作用组中miR-21表达显著下调（P<0.01）,单独药物组中miR-21表达无显著变化（P>0.05）。辛伐他汀使SACC-LM细胞的生长受到明显抑制,与药物浓度呈正相关关系。干扰miR-21后,SACC-LM对辛伐他汀的耐药性显著降低（P<0.05）。联合作用组中细胞的迁移、侵袭活性较其他处理组相比均受到显著抑制 (P<0.01),Western免疫印迹结果显示,联合作用组中E-Cadherin蛋白的表达较其他对照组显著升高（P<0.01）,Snail1蛋白表达显著降低（P<0.001）。 结论: 干扰miR-21能有效降低SACC-LM对辛伐他汀的耐药性,通过与辛伐他汀的联合作用,SACC-LM细胞的迁移、侵袭能力受到明显抑制,其机制可能与Snail1表达下调、E-Cadherin表达上调有关。

关键词: 辛伐他汀, microRNA-21, 唾液腺腺样囊性癌, E-Cadherin, Snail1

Abstract: PURPOSE: To investigate the effect of simvastatin combined with miR-21 on migration and invasion of human salivary adenoid cystic carcinoma cell line (SACC-LM) and its underlying mechanism. METHODS: SACC-LM cells were divided into 4 groups randomly. In NC group, unrelated nucleotides sequence was transfected and a blank reagent was added; in SIM group, unrelated nucleotides sequence was transfected and simvastatin was added; in simiR-21 group, miR-21 inhibitor was transfected and a blank reagent that did not contain simvastatin was added; in simiR-21+SIM group, miR-21 inhibitor was transfected and simvastatin was added. qRT-PCR was used to detect the expression of miR-21. CCK-8 method was used to measure the IC₅₀ of SACC-LM while simvastatin (10,20,30,40,50,60 μmol/L) was added for 48 h. Transwell chamber method was used to observe the migration (without matrigel) and invasion (adding matrigel) ability. Western blot was used to detect the expression of EMT relative proteins (E-Cadherin, Snail1). Student's t test or one-way ANOVA were performed to analyze the data using SPSS 22.0 software package. RESULTS: The expression of miR-21 was remarkably down-regulated in the groups with transfected inhibitor(P<0.01). Simvastatin had no effect on the expression of miR-21(P>0.05). Simvastatin could inhibit the proliferation of SACC-LM cells in a dose-dependent manner. miR-21 was effective in reducing the resistance of SACC-LM to simvastatin. The abilities of migration and invasion on SACC-LM cells in simiR-21+SIM group were significantly inhibited compared with other groups (P<0.01). The expression of E-cadherin in the combined group was significantly higher than other groups (P<0.01) with decreased Snail1 expression(P<0.001). CONCLUSIONS: MiR-21 inhibitor transfection can effectively reduce the resistance of SACC-LM to SIM. In simiR-21+SIM group, the abilities of migration and invasion on SACC-LM cells were more greatly inhibited than other groups, indicating the mechanism may be related to down-regulation of Snail1 expression and up-regulation of E-Cadherin expression.

Key words: Simvastatin, MicroRNA-21, Salivary adenoid cystic carcinoma, E-cadherin, Snail1

中图分类号:

R739.8

王超, 黎婷, 严斐, 蔡文燕, 蒋星宇, 孙晋虎. 辛伐他汀联合干扰miR-21对唾液腺腺样囊性癌细胞迁移和侵袭的影响[J]. 中国口腔颌面外科杂志, 2018, 16(2): 132-137.

WANG Chao, LI Ting, YAN Fei, CAI Wen-yan, JIANG Xing-yu, SUN Jin-hu. Effect of simvastatin combined with simiR-21 on migration and invasion of human salivary adenoid cystic carcinoma[J]. China J Oral Maxillofac Surg, 2018, 16(2): 132-137.

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辛伐他汀联合干扰miR-21对唾液腺腺样囊性癌细胞迁移和侵袭的影响

Effect of simvastatin combined with simiR-21 on migration and invasion of human salivary adenoid cystic carcinoma

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