摘要
PI3K/AKT/m TOR信号通路的激活能引发人体细胞发生癌变,哺乳动物靶标m TOR作为PI3K/AKT信号通路下游的一个效应分子,被视为针对肿瘤发生和形成的关键治疗靶点,因此阻断该信号通路的相关靶点可以作为恶性肿瘤治疗的一个策略。白细胞介素24(interleukin 24,IL24)是一个选择性诱导肿瘤细胞凋亡的重要抑癌基因。该研究分别利用MTT法和实时无标记细胞功能分析仪检测携带IL24基因的溶瘤腺病毒ZD55-IL24与m TOR抑制剂雷帕霉素(rapamycin)单独或联合作用对多种肝癌细胞的体外杀伤效果;倒置显微镜分别观察ZD55-IL24、雷帕霉素单独作用及两者联合作用引起的细胞形态学变化;Hoechst 33342、流式细胞术和TUNEL染色检测各处理组细胞的凋亡情况;Western blot检测IL24、AKT以及凋亡相关蛋白Bax和Bcl-2的蛋白质水平。结果显示,溶瘤腺病毒ZD55-IL24与雷帕霉素联合作用较两者单独作用更显著地抑制了肝癌细胞Hep3B的生长并诱导了肝癌细胞的凋亡;此外,两者联合作用更有效地上调了肝癌细胞Hep3B中的细胞因子IL24和促凋亡蛋白Bax的表达,同时下调了PI3K/AKT/m TOR信号通路关键蛋白AKT和抗凋亡蛋白Bcl-2的表达。该研究结果表明,雷帕霉素很可能在一定程度上促进了ZD55-IL24病毒介导的IL24表达而增强对肝癌细胞的杀伤作用,进而为肝癌治疗研究提供了一条新型有效的方案。
Activation of PI3K/AKT/mTOR signal pathway can trigger the formation of human solid tu- mors. mTOR, as a downstream effector of the PI3K/AKT pathway, is a key therapeutic target involved in onco- genesis and progression of cancer. Therefore, blocking relevant targets of this pathway could be a novel treatment strategy for human malignancies. IL24 （interleukin 24）, which is able to selectively induce tumor cell apoptosis, is an important antitumor gene. In this study, we explored the killing effects of either oncolytic adenovirus mediatedIL24 gene （ZD55-IL24） alone or in combination with novel immunosuppressor, rapamycin, against hepatoma cells in vitro by MTT detecting method and the iCELLigence real time cellular analysis system. The morphological anal- ysis was assessed by inverted microscope in hepatoma cells treated by single ZD55-IL24 or rapamycin, and their combination. Hoechst 33342 staining, flow cytometry assay and TUNEL were performed to determine the apoptosis effects of single and combinational therapy on hepatoma cells, respectively. The protein levels of IL24, AKT, and apoptosis related protein Bax and Bcl-2 were determined by Western blot. The results indicated that the combina- tion treatment of oncolytic adenovirus ZD55-IL24 and rapamycin remarkably inhibited Hep3B cell proliferation and induced the cell apoptosis compared with ZD55-IL24 alone or rapamycin alone. Furthermore, the combination treatment significantly up-regulated cytokines IL24 and apoptosis-induction protein Bax expression, while down- regulated the key protein AKT expression of PI3K/AKT/mTOR signal pathway and anti-apoptosis protein Bcl-2 expression. In conclusion, the results showed that rapamycin could promote the expression mediated by ZD55-IL24 replication and strengthen its antitumor effects in hepatoma cells in vitro, which provides a novel and promising therapeutic approach for targeting treatment of hepatoma cells.
出处
《中国细胞生物学学报》
CAS
CSCD
2015年第10期1353-1361,共9页
Chinese Journal of Cell Biology
基金
国家自然科学基金(批准号:81272687)
浙江省自然科学基金(批准号:LZ13H160004)
浙江省公益技术研究计划项目(批准号:2014C33275)资助的课题~~
