梁挺, 王培军. miR-512-3p在乳腺癌中的表达及其功能研究[J]. 中国肿瘤临床, 2013, 40(19): 1145-1149. DOI: 10.3969/j.issn.1000-8179.20121921
引用本文: 梁挺, 王培军. miR-512-3p在乳腺癌中的表达及其功能研究[J]. 中国肿瘤临床, 2013, 40(19): 1145-1149. DOI: 10.3969/j.issn.1000-8179.20121921
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Ting LIANG, Peijun WANG. MiR-512-3p expression pattern and function in breast cancer[J]. CHINESE JOURNAL OF CLINICAL ONCOLOGY, 2013, 40(19): 1145-1149. DOI: 10.3969/j.issn.1000-8179.20121921
Citation: Ting LIANG, Peijun WANG. MiR-512-3p expression pattern and function in breast cancer[J]. CHINESE JOURNAL OF CLINICAL ONCOLOGY, 2013, 40(19): 1145-1149. DOI: 10.3969/j.issn.1000-8179.20121921
shu

miR-512-3p在乳腺癌中的表达及其功能研究

MiR-512-3p expression pattern and function in breast cancer

    摘要
  • 摘要:
      目的  观察miR-512-3p在浸润性乳腺癌和癌旁组织中的表达，以及对乳腺癌细胞系MDA-MB-231增殖、凋亡、周期和克隆形成能力的影响，并分析其可能机制。
      方法  运用荧光定量PCR检测浸润性乳腺癌和癌旁组织中的miR-512-3p的表达量。运用MTT法、流式细胞仪和平板克隆形成实验分析miR-512-3p对细胞增殖、凋亡、周期和克隆形成能力的影响。通过TargetScan、PicTar和miRanda软件寻找miR-512-3p可能靶基因，应用RT-PCR和Western blot技术进行验证。
      结果  乳腺癌组织中miR-512-3p表达明显低于正常组织(P < 0.05)。MTT试验中，72 h浓度为100 nmol/L时，miR-512-3p对乳腺癌细胞增殖的抑制作用最明显，抑制率为45.38%。miR-512-3能明显诱导细胞发生早期凋亡(9.32±0.41)%，与空白对照组(3.1±0.54)%，负对照组(2.9±0.39)% 比较，差异有统计学意义(P < 0.05)。与空白对照组比较，G0/G1期细胞明显增多，G2/M期细胞明显减少(P < 0.05)。平板克隆试验显示过表达miR-512-3p可显著抑制细胞的克隆形成能力，并与对照组差异性显著。荧光定量PCR和Western bolt实验结果显示100 nmol/L miR-512-3p显著抑制c-FLIP mRNA和蛋白表达水平。
      结论  miR-512-3p在乳腺癌组织中表达明显低于正常组织，其可能通过作用于下游基因c-FLIP抑制乳腺癌细胞株MDA-MB-231的增殖和克隆形成能力，诱导凋亡，其在肿瘤发生发展中可能担任着抑癌基因的角色，可能成为未来乳腺癌诊断和治疗的一个新的靶点。

     

    Abstract:
      Objective  This study aims to investigate the expression pattern of miR-512-3p in breast cancer and noncancerous paired specimens as well as the effects of miR-512-3p on the proliferation, apoptosis, cell cycle, and cloning of MD-MBA-231 breast cancer cells. The study also aims to identify the miR-512-3p target gene.
      Methods  Reverse transcription-polymerase chain reaction (RT-PCR) was conducted to quantify the miR-512-3p expression in breast cancer and noncancerous paired specimens. Methylthiazol tetrazolium (MTT) assay, flow cytometry, and clone formation assay were used to characterize the function of miR-512-3p in breast cancer. Target prediction was performed using the TargetScan, PicTar, and miRanda software. The results were validated using RT-PCR and western blot target validation.
      Results  The relative expression of miR-512-3p in breast cancer specimens was significantly lower than those in normal breast specimens (P < 0.05). MTT assay revealed that 48 h after transfection, miR-512-3p significantly repressed the proliferation of MD-MBA-231 cells at a suppression rate of 45.38% and at a concentration of 100 nmol/L. MiR-512-3p increased the percentage of early apoptotic cells in the treatment groups (9.32 ± 0.41)% compared with those in the blank controls (3.1 ± 0.54)% and in the negative controls (2.9 ± 0.39)%(P < 0.05). Significant differences were found in the percentages of the G0/G1- and G2/M-phase cells after miR-512-3p transfection compared with those in the controls (P < 0.05). In the cloning assay, clone formation was inhibited in the miR-512-3p-transfected groups compared with those in the control groups. RT-PCR and western blot results indicate that miR-512-3p significantly inhibited the c-FLIP mRNA and protein expression.
      Conclusion  MiR-512-3p expression is relatively decreased in breast cancer specimens compared with those in the normal samples. The negative effect of miR-512-3p on cell proliferation and clone formation and its positive effect on early apoptosis through c-FLIP targeting suggest that miR-512-3p acts as a tumor suppressor gene in breast cancer. Therefore, miR-512-3p may be a new target for the diagnosis and treatment of breast cancer in the future.

     

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