摘要
目的：探讨微小RNA-10a（microRNA-10a,miR-10a）对肝癌细胞增殖的影响及其作用机制。方法：收集广西医科大学附属肿瘤医院肿瘤科2001年10月至2005年7月144例肝癌患者手术切除的肝癌组织和癌旁组织（距癌灶组织边缘2~5 cm）标本,Real-time PCR法分析144例肝癌组织及癌旁组织中miR-10a的表达量。在肝癌细胞（QGY-7701、Huh7、PCL/PRF/5）中转染miR-10a模拟物,Real-time PCR法检测转染后细胞miR-10a的表达水平;CCK-8法检测过表达miR-10a的肝癌细胞的增殖水平,流式细胞术检测过表达miR-10a的肝癌细胞的凋亡和细胞周期;生物信息学预测并以Western blotting检测过表达miR-10a的肝癌细胞中转录因子E2F3的表达量。结果：与癌旁组织相比,肝癌组织中的miR-10a显著低表达[（-9.89±1.68）vs（-7.84±1.97）,P=0.000]。转染miR-10a模拟物后肝癌细胞系中miR-10a的表达量是转染对照小RNA组或空白组细胞的16倍左右。过表达miR-10a可显著抑制7种肝癌细胞（QGY-7701、QGY-7703、Huh7、PCL/PRF/5、HepG2、BeL-7402、SMMC-7721）的增殖（均P〈0.05）,并引起肝癌细胞细胞周期G1/S期阻滞,但并不能诱导肝癌细胞发生凋亡。生物信息学预测显示E2F3是miR-10a可能的靶分子,Western blotting检测显示过表达miR-10a可明显抑制肝癌细胞中E2F3的表达[（0.50±0.12）vs（0.79±0.21）,P〈0.05]。结论：人肝癌组织中低表达miR-10a,转染miR-10a模拟物后多种肝癌细胞的增殖均受到明显抑制,其机制可能与miR-10a靶向作用转录因子E2F3并阻滞肝癌细胞细胞周期于G1/S期有关。
Objective： To investigate the role of microRNA-10a（ miR-10a） in hepatocellular carcinoma（ HCC）growth. Methods： Paired HCC and adjacent non-tumor tissue specimens were surgically collected from 144 patients who were diagnosed with primary HCC in Guangxi Medical University-Affiliated Tumor Hospital between October 2001 and July2005. HCC QGY-7701,Huh7,and PCL /PRF /5 cells were transfected with miR-10 a mimics or scramble control miRNA.The abundance of miR-10 a in both tissue specimens and transfected cells was quantified by real-time PCR and E2F3 protein in transfected cells was assessed by Western blotting. Proliferation of the transfectants was assessed by a colorimetric cell counting assay. Cell cycle progression and apoptosis of the transfectants were assessed by FACS. Results： The abundance of miR-10 a mRNA was significantly lower in HCC tissue specimens than in normal tissue specimens（- 9. 89 ±1. 68 vs- 7. 84 ± 1. 97,P = 0. 0001）. HCC cells transfected with miR-10 a mimics had miR-10 a abundance 16 times higher than both wild-type HCC cells and HCC cells transfected with the control miRNA with scrambled sequences. Over-expression of miR-10 a resulted in significant increases in suppression of HCC cell proliferation（ P 0. 05） and G1 phase arrest. In contrast,overexpression of miR-10 a had no influence on apoptosis of HCC cells. Bioinformatics suggested that transcription factor E2F3 might be a downstream target of miR-10 a and the expression of E2F3 in HCC cells transfected with miR-10 a was significantly lower than in wild-type HCC cells and HCC cells transfected with the control miRNA（ 0. 50 ±0.12 vs 0.79 ±0.21,P 0.05）. Conclusion： MiR-10 a may suppress HCC cell proliferation through G1 phase arrest in an E2F3-dependent mechanism.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2014年第4期383-388,共6页
Chinese Journal of Cancer Biotherapy
基金
国家高技术研究发展计划(863计划)资助项目~~
关键词
肝癌
微小RNA-10a
增殖
转录因子
E2F3
G1
S期阻滞
hepatocellular carcinoma
microRNA-10a（miR-10a）
transcription factor
E2F3
proliferation
G1/S phase arrest
