目的探讨微小RNA(microRNA,miR)-646对EGFR/Akt通路的作用及对肺癌A549细胞增殖扩散的影响及相关的作用机制。方法利用Lipo2000将miR-646转染到肺癌A549细胞中,采用实时荧光定量PCR(real time PCR,RTPCR)法检测转染后各组细胞中miR-646...目的探讨微小RNA(microRNA,miR)-646对EGFR/Akt通路的作用及对肺癌A549细胞增殖扩散的影响及相关的作用机制。方法利用Lipo2000将miR-646转染到肺癌A549细胞中,采用实时荧光定量PCR(real time PCR,RTPCR)法检测转染后各组细胞中miR-646的表达;CCK-8法检测细胞增殖;划痕和Transwell小室分别检测细胞的迁移和侵袭能力;Western blotting法检测EGFR/Akt通路中各蛋白的变化水平。结果 RT-PCR结果显示,转染miR-646组其miR-646表达水平显著高于阴性对照组和空白对照组(P<0.05),A549细胞转染miR-646后,细胞增殖和扩散能力均显著降低(P<0.05);Western blotting结果显示,转染miR-646组中p-EGFR和p-Akt的蛋白表达水平均明显降低(P<0.05)。结论 miR-646可显著降低肺癌A549细胞中p-EGFR、p-Akt的蛋白水平,进而抑制细胞的增殖和扩散能力。展开更多
Objective: The aim of the study was to explore the effect of demethylating agent 5-Aza-2'-deoxycytidine (5-ADC) on expression of Fanconi anemia complementation group F (FANCF) gene and the proliferation of cervica...Objective: The aim of the study was to explore the effect of demethylating agent 5-Aza-2'-deoxycytidine (5-ADC) on expression of Fanconi anemia complementation group F (FANCF) gene and the proliferation of cervical cancer cells, to observe cell's sensitivity to chemotherapeutic drug taxol, and to explore the antitumor effect of 5-ADC as well as the new treatment of cervical cancer. Methods: Cervical cancer cell lines SiHa (FANCF gene full-methylated) and Hela (unmethylated) were treated with 5-ADC. We used the methylation-specific PCR (MSP), reverse transcription-polymerase chain reaction (RT-PCR) and Western blot to detect the FANCF methylation, mRNA and protein respectively. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect the proliferation of cells. The cytotoxicity of taxol was measured by flow cytometer. The nude mice bearing SiHa was used to observe the effect of 5-ADC in vivo. Results: Inhibition of DNA promoter methylation by 5-ADC reactivated the expression of FANCF mRNA and protein in SiHa cells, consistent with decreased growth speed and increased taxol resistance. These results were proven in experiments in vivo. Conclusion: The 5-ADC probably become a potential treatment drug through inhibiting the proliferation of cervical cancer cells in taxol-resistant patients.展开更多
基金Supported by the grant from the National Science Foundation of Chongqing (No. cstc2011jjA10081)
文摘Objective: The aim of the study was to explore the effect of demethylating agent 5-Aza-2'-deoxycytidine (5-ADC) on expression of Fanconi anemia complementation group F (FANCF) gene and the proliferation of cervical cancer cells, to observe cell's sensitivity to chemotherapeutic drug taxol, and to explore the antitumor effect of 5-ADC as well as the new treatment of cervical cancer. Methods: Cervical cancer cell lines SiHa (FANCF gene full-methylated) and Hela (unmethylated) were treated with 5-ADC. We used the methylation-specific PCR (MSP), reverse transcription-polymerase chain reaction (RT-PCR) and Western blot to detect the FANCF methylation, mRNA and protein respectively. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect the proliferation of cells. The cytotoxicity of taxol was measured by flow cytometer. The nude mice bearing SiHa was used to observe the effect of 5-ADC in vivo. Results: Inhibition of DNA promoter methylation by 5-ADC reactivated the expression of FANCF mRNA and protein in SiHa cells, consistent with decreased growth speed and increased taxol resistance. These results were proven in experiments in vivo. Conclusion: The 5-ADC probably become a potential treatment drug through inhibiting the proliferation of cervical cancer cells in taxol-resistant patients.
