上调miR-449a表达对肝癌细胞生长和侵袭能力的影响
| 作者: |
1邓雪松,
2潘红艳,
1成俊,
1卢贵余,
1詹勇强,
1倪勇,
1王成友
1 深圳大学第一附属医院/深圳市第二人民医院 肝胆外科，广东 深圳 518035 2 深圳市南山区人民医院 消化内科，广东 深圳 518052 |
| 通讯: |
倪勇
Email: szniyong@qq.com |
| DOI: | 10.3978/.2018.01.011 |
| 基金: | 广东省医学科研基金资助项目（A2017338）；广东省科学技术厅科技计划科研基金资助项目（2014A020212701）；深圳市卫生计生系统科研项目（201401015）；深圳市第二人民医院临床研究项目（20173357201806）。 |
摘要
目的：探讨miR-449a在肝癌（HCC）细胞中的表达及其对HCC细胞生物学行为的影响。
方法：用qRT-PCR检测miR-449a在正常肝细胞L02和4种HCC细胞株（HepG2、Hep3B、SMMC-7721和Bel-7402）的表达。用脂质体法将miR-449a模拟物或阴性对照序列转染HCC细胞后，分别用CCK-8法、流式细胞术、Transwell小室实验检测细胞增殖、细胞周期、侵袭能力的变化，并观察以上两种不同转染的HCC细胞在裸鼠体内的成瘤情况。
结果：miR-449a在4种HCC细胞株的表达水平均明显低于L02细胞（均P<0.05），其中在Bel-7402细胞表达的水平最低。与转染阴性对照序列的Bel-7402细胞比较，转染miR-449a模拟物的Bel-7402细胞增殖活性明显降低、G1/S期阻滞明显增加、穿室细胞数明显减少（均P<0.05）；与转染阴性对照序列的Bel-7402细胞比较，转染miR-449a模拟物的Bel-7402细胞在裸鼠体内成瘤后的移植瘤质量与体积均明显减小（0.748 g vs. 1.234 g；33.667 mm3 vs. 1 400.500 mm3，均P<0.05）。
结论：HCC细胞中miR-449a表达降低，上调miR-449a表达可以抑制HCC细胞在体内外的生长。
关键词：
癌，肝细胞；微RNAs；细胞增殖；肿瘤移植；小鼠，裸
方法：用qRT-PCR检测miR-449a在正常肝细胞L02和4种HCC细胞株（HepG2、Hep3B、SMMC-7721和Bel-7402）的表达。用脂质体法将miR-449a模拟物或阴性对照序列转染HCC细胞后，分别用CCK-8法、流式细胞术、Transwell小室实验检测细胞增殖、细胞周期、侵袭能力的变化，并观察以上两种不同转染的HCC细胞在裸鼠体内的成瘤情况。
结果：miR-449a在4种HCC细胞株的表达水平均明显低于L02细胞（均P<0.05），其中在Bel-7402细胞表达的水平最低。与转染阴性对照序列的Bel-7402细胞比较，转染miR-449a模拟物的Bel-7402细胞增殖活性明显降低、G1/S期阻滞明显增加、穿室细胞数明显减少（均P<0.05）；与转染阴性对照序列的Bel-7402细胞比较，转染miR-449a模拟物的Bel-7402细胞在裸鼠体内成瘤后的移植瘤质量与体积均明显减小（0.748 g vs. 1.234 g；33.667 mm3 vs. 1 400.500 mm3，均P<0.05）。
结论：HCC细胞中miR-449a表达降低，上调miR-449a表达可以抑制HCC细胞在体内外的生长。
Expression and action of miR-449a in hepatocellular carcinoma cells
CorrespondingAuthor:NI Yong Email: szniyong@qq.com
Abstract
Objective: To investigate the miR-449a expression in hepatocellular carcinoma (HCC) cells and its effect on biological behaviors of HCC cells.
Methods: The miR-449a expressions in normal hepatic L02 cells and 4 types of HCC lines (HepG2, Hep3B, SMMC-7721 and Bel-7402) were determined by qRT-PCR method. HCC cells were transfected with miR-449a mimics or scrambled sequences via lipofection, and then, the changes in cell proliferation, cell cycle and invasion ability were detected by CCK-8 assay, flow cytometry and Transwell assay respectively; moreover, the tumor formations of the HCC cells with above transfections in nude mice were observed.
Results: The miR-449a expression levels in the 4 HCC cells were all significantly lower than those in L02 cells (all P<0.05), and showed the lowest level in Bel-7402 cells. Compared with Bel-7402 cells transfected with scrambled sequences, the proliferative activity was significantly inhibited, G1/S phase arrest was significantly increased, and the number of migrating cells was significantly decreased in Bel-7402 cells transfected with miR-449a mimics (all P<0.05). The weight and volume of xenografts derived from Bel-7402 cells transfected with miR-449a mimics in nude mice were all decreased compared with those derived from Bel-7402 cells transfected with scrambled sequences (0.748 g vs. 1.234 g; 33.667 mm3vs. 1 400.500 mm3, both P<0.05).
Conclusion: The miR-449a expression is decreased in HCC cells, and up-regulating miR-449a expression can suppress the growth of HCC cells in vivo and in vitro.
Keywords：
Carcinoma
Hepatocellular; MicroRNAs; Cell Proliferation; Neoplasm Transplantation; Mice
Nude
Methods: The miR-449a expressions in normal hepatic L02 cells and 4 types of HCC lines (HepG2, Hep3B, SMMC-7721 and Bel-7402) were determined by qRT-PCR method. HCC cells were transfected with miR-449a mimics or scrambled sequences via lipofection, and then, the changes in cell proliferation, cell cycle and invasion ability were detected by CCK-8 assay, flow cytometry and Transwell assay respectively; moreover, the tumor formations of the HCC cells with above transfections in nude mice were observed.
Results: The miR-449a expression levels in the 4 HCC cells were all significantly lower than those in L02 cells (all P<0.05), and showed the lowest level in Bel-7402 cells. Compared with Bel-7402 cells transfected with scrambled sequences, the proliferative activity was significantly inhibited, G1/S phase arrest was significantly increased, and the number of migrating cells was significantly decreased in Bel-7402 cells transfected with miR-449a mimics (all P<0.05). The weight and volume of xenografts derived from Bel-7402 cells transfected with miR-449a mimics in nude mice were all decreased compared with those derived from Bel-7402 cells transfected with scrambled sequences (0.748 g vs. 1.234 g; 33.667 mm3vs. 1 400.500 mm3, both P<0.05).
Conclusion: The miR-449a expression is decreased in HCC cells, and up-regulating miR-449a expression can suppress the growth of HCC cells in vivo and in vitro.