【作者】 刘四明；

【导师】 佟明；

【作者基本信息】 辽宁医学院 ， 外科学， 2011， 硕士

【摘要】 目的研究在激素非依赖性前列腺癌细胞中E2和AR-miRNA-21通路的内在关联性及其重要意义。分析非依赖前列腺癌中miRNA-21调控的下游分子机制。方法通过RT-PCR检查有无E2培养下非依赖性细胞系PC3在转染miRNA-21 inhibitor前后miRNA-21及AR的表达情况。通过CCK-8和FCM检测转染前后细胞的增殖能力。通过western blot检测有无E2培养下PC3细胞AR蛋白的表达情况。通过western blot检测在转染miRNA-21 inhibitor前后PTEN、p-Akt和MMP-2的表达情况。结果RT-PCR检测显示:E2环境培养下转染前miRNA-21的表达明显高于不含E2的培养基中的表达;在无E2的培养基中转染后miRNA-21的表达明显低于转染前的表达;在含有E2的培养基中转染后的miRNA-21的表达明显低于转染前得细胞组。Cck-8方法测转染前PC3细胞系在不同环境下和转染miRNA-21 inhibitor前后的增殖能力,显示在含有E2的环境下PC3细胞增殖能力明显增强;无论有没有E2,转染后PC3的增值能力均明显比未转染组弱。FCM检测转染miRNA-21抑制剂后,对应与转染前PC3细胞的增殖都明显的减慢。RT-PCR检测AR表达发现,转染前后AR的表达变化不具有显著性(P>0.05),但E2能促进AR的表达增加。western blot测定PTEN、p-Akt和MMP-2在转染前后的PC3细胞中表达水平。转染后PTEN的表达明显高于转染前;转染后p-Akt和MMP-2的表达明显低于转染前。结论1. E2通过诱导AR表达在非依赖性前列腺癌的进展中起重要作用。2. E2可能通过AR或其他途径诱导miRNA-21的表达。3. miRNA-21在前列腺癌的非依赖性进程中起重要作用,转染miRNA-21 inhibitor前列腺癌细胞的增值能力明显降低提示miRNA-21 inhibitor有希望成为非依赖性前列腺癌的一个潜在治疗之路。4. MiR-21负面调节PTEN的表达同时上调p-Akt和MMP-2的表达,提示miRNA-21可能通过PTEN-pAkt/MMP-2通路调控前列腺癌的进展。更多还原

【Abstract】 ObjectiveTo study the relationship and the significance of E2 with AR-miRNA-21 pathway, Analyze the downstream regulation mechanisms of miRNA-21 in hormone-independence prostate cancer.MethodsRespectively, in the presence of E2 and E2 in the special medium does not contain cultured PC3 and then were transected with miRNA-21 inhibitor. Detect cell proliferation with CCK-8 and FCM, while the expression of gene and protein with RT-PCR and western blot.ResultsRT-PCR analysis showed that before transfection the expression of miRNA-21 in E2 environment cultured was significantly higher than in the medium without E2; In the presence of E2 the expression of miRNA-21 after transfection was significantly lower than untransfected cell group.Cck-8:in the presence of E2 the PC3 cell proliferation was significantly enhanced ; With or without E2, PC3 after transfection, the capacity of amplification was significantly weaker than the untransfected group.FCM: make sure that the miRNA-21 inhibitor is useful to mitotic cycle in PC3.AR is not different between before transfection and behind transfection (p>0.05), EGF can increase expression of AR by RT-PCR,but E2 can promote increased expression of AR.Western blot show that the expression of PTEN behind transfection is higher than before,but p-Act and MMP-2 behind transfection is lower than before.Conclusions1. E2 through inducting of AR expression in prostate cancer progression plays an important role.2. E2 may induce the expression of miRNA-21 by AR or other means.3. miRNA-21 plays an important role in the process of transfection in hormone-independence prostate cancer. Transfected with miRNA-21 inhibitor of prostate cancer cells significantly reduced the amplification capacity and prompted miRNA-21-inhibitor with prostate cancer will be a potential way of biological treatment.4. miRNA-21 negatively regulate the expression of PTEN also increased p-Akt and the expression of MMP-2, suggesting that miRNA-21 pathway may be through PTEN-pAkt/MMP-2 control the progress of prostate cancer.更多还原