摘要
目的:探讨PI3K/Akt通路与肝癌对索拉非尼获得性耐药机制的关系。方法:采用浓度梯度递增法建立肝癌细胞系Hep G2和Huh7耐药细胞株Hep G2-SR和Huh7-SR,并采用CCK-8法检测细胞活力,验证细胞耐药特性。Western blot检测p-Akt、Akt和β-actin蛋白表达水平。以哌立福辛抑制p-Akt,采用CCK-8法检测细胞活力,并采用流式细胞仪检测细胞凋亡,探讨哌立福辛与索拉非尼的协同作用及对耐药的逆转作用。结果:索拉非尼对Huh7及Hep G2细胞抑制细胞活力的作用明显强于Huh7-SR、Hep G2-SR细胞(P<0.05)。耐药细胞株中Akt磷酸化蛋白p-Akt的表达水平明显高于亲本细胞株(P<0.05),而Akt表达在耐药株与亲本细胞株中无明显差异(P>0.05)。哌立福辛可协同索拉非尼抑制Huh7-SR、Hep G2-SR细胞活力,促进凋亡。结论:索拉非尼持续作用导致PI3K/Akt通路的激活导致肝癌对索拉非尼获得性耐药,其机制是通过调节发生在翻译后水平的Akt磷酸化而非Akt转录或翻译。抑制Akt可逆转肝癌对索拉非尼耐药。
Objective： To investigate the role of PI3K/Akt pathway in the acquired resistance to sorafenib in hepatocellular carcinoma（HCC）. Methods： Two sorafenib-resistant cells Hep G2-SR and Huh7-SR were established by incubating human HCC Hep G2 and Huh7 cells at a increasing concentration of sorafenib, and verify resistant cell properties by detecting the cell viability using CCK-8 assay. The protein levels of p-Akt, Akt, and β-actin were detected by Western-blot. Perifosine was used to inhibiting p-Akt, then the cell viability and cell apoptosis were detected by CCK-8and flow cytometry to explore the effect of reversing the drug resistance and synergy in combination with sorafenib. Results：Sorafenib induced cell viability inhibition is significantly in Huh7 and Hep G2 cells than the corresponding Huh7-SR and Hep G2-SR cells（P〈0.05）. The P-Akt expression levels were significantly higher in sorafenib-rsistant cells than the corresponding parental cells（P〈0.05）,while the Akt expression levels were no significant（P〉0.05）. When in combination with sorafenib,perifosine showed the synergistic effect in inhibiting cell viability and inducing cell apoptosis in sorafenib-resistant cells. Conclusion： The activation of PI3K/Akt pathway for sustaining action sorafenib was respondible for the acquired resistance to sorafenib in HCC. The mechanism was due to post-translational Akt phosphorylation, not the transcription or translation of Akt. Inhibiting Akt could reverse the acquired resistance to sorafenib in HCC.
出处
《中国现代普通外科进展》
CAS
2016年第1期6-10,共5页
Chinese Journal of Current Advances in General Surgery
基金
黑龙江省教育厅科学技术研究(面上)项目(12541329)
