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PPARγ基因静默抑制肝癌HCCLM3细胞增殖并诱导其凋亡
         
Silencing PPARγ gene inhibits proliferation and inducs apoptosis of hepatoma HCCLM3 cells

摘    要
目的:观察过氧化物酶体增殖物活化受体γ(peroxisome proliferators-activated receptorγ,PPARγ)基因静默对肝癌细胞增殖与凋亡的影响。方法:构建靶向PPARγ的短发夹状RNA真核表达载体并转染HCCLM3细胞,采用RT-PCR、Western印迹法检测HCCLM3细胞中PPARγ的表达;MTT法检测细胞增殖;TUNEL法及FCM法检测细胞的凋亡;免疫细胞化学法检测PCNA、野生型p53的表达。结果:shPPARγ转染组PPARγmRNA表达下调80.5%。转染后48 h,pshPPARγ组细胞增殖抑制率为71.5%;40 h时,PCNA阳性率为(23.8±7.2)%;TUNEL法检测凋亡率为(24.2±4.7)%,FCM法检测凋亡率为(23.2±4.2)%,2种方法的检测结果一致;shPPARγ转染组细胞被阻滞于G0/G1期,G2/M期细胞减少,与阴性对照和空白对照组比较(P < 0.01)差异有统计学意义,且野生型p53表达增加。结论:PPARγ基因静默能诱导HCCLM3细胞凋亡,抑制增殖;与促进野生型p53表达相关。
标    签 肝肿瘤,实验性   RNA,小分子干扰   PPARγ   细胞增殖   细胞凋亡   Live neoplasms   experimental   RNA small interfering PPAR gamma   Cell proliferation   Apoptosis  
 
Abstract
Objective: To observe the effects of knocking down the expression of peroxisome proliferators-activated receptor gamma(PPARγ) with RNA interference techniques on the proliferation and apoptosis of hepatocellular carcinoma HCCLM3 cells.Methods: A short-hairpin RNA(shRNA) eukaryotic expression vector against PPARγ was constructed and transfected into HCCLM3 cells.The changes of PPARγ expression were detected by semi-quantitative RT-PCR and Western blotting analysis.The proliferation of HCCLM3 cells was tested by MTT assay.Apoptosis ratio of HCCLM3 cells was detected by TUNEL method and flow cytometry(FCM).Expressions of PCNA and wide-type p53 protein were analyzed by immuocytochemistry(ICC) methods.Results: The sequence-specific shRNA(pshPPARγ) efficiently blocked the expression of PPARγ mRNA by 80.5%.At 48 h after transfection of pshPPARγ, proliferation of HCCLM3 cells was significantly suppressed by 71.5%.The positive rate of PCNA expression was(23.8±7.2)% at 40 h transfection.The apoptotic rates were(24.2±4.7)% as detected by TUNEL assay and(23.2±4.2)% of cells as measured by FCM test, respectively.The detection results of the two methods were consistent.In pshPPARγ transfection group, cell cycle of HCCLM3 cells was arrested in G0/G1 phase and the proportion of cells in G2/M phase decreased.Moreover, expression of wide-type p53 protein increased significantly.Conclusion: Knockdown PPARγ expression with RNA interference technology can significantly suppress proliferation and induce apoptosis of HCCLM3 cells.It is related with up-regulation of wide-type p53 protein expression.

中图分类号 R735.7   DOI 10.3781/.jissn.1000-7431.2008.08.010

 
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所属栏目 基础研究

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收稿日期 2007/9/21

修改稿日期 2007/10/18

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备注吴旭东(1970-),男(汉族),博士,主治医师

引用该论文: WU Xu-dong,CHEN Wei-chang,HUANG Xiao-ping. Silencing PPARγ gene inhibits proliferation and inducs apoptosis of hepatoma HCCLM3 cells[J]. Tumor, 2008, 28(8): 676~680
吴旭东,陈卫昌,黄小平. PPARγ基因静默抑制肝癌HCCLM3细胞增殖并诱导其凋亡[J]. 肿瘤, 2008, 28(8): 676~680


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参考文献
【1】SAEZE,ROSENFELDJ,LIVOLSIA,et al. PPAR gamma signaling exacerbates mammary gland tumor development[J].Genes Dev,2004,18(5):528-540.
 
【2】郭鸣雷,李锦军,李宏年,等. 过氧化物酶体增殖激活性受体γ在肝癌组织中的差异性表达[J].肿瘤,2005,25(5):413-415.
 
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