中文摘要：

目的研究肿瘤RNA转染树突状细胞（DCs）与同源细胞因子诱导的杀伤细胞（CIKs）共培养对前列腺癌细胞PC3内Akt/NF-κB生存信号通道的影响,探讨这种免疫治疗引起肿瘤细胞凋亡的分子机制。方法体外培养CIKs、DC-CIKs和RNA转染的共培养的DC-CIKs作为效应细胞,以传代培养的PC3细胞作为靶细胞,分别加到TRANSWELL 6孔细胞培养板上、下室。收集靶细胞,分别提取细胞可溶性总蛋白和核蛋白,利用Western Blotting检测Akt,phospho-Akt,phospho-IKKα/β以及NF-κBp65的表达量,以未干预的PC3细胞总蛋白和核蛋白作为阴性对照,以PI3K抑制剂LY294002作为实验的阳性对照。以β-actin作为内参照,利用Quantity One软件定量分析。结果以未干预的PC3细胞作为阴性对照,CIKs、DC-CIKs和RNA-DC-CIKs各免疫效应细胞处理组均显著抑制细胞浆中磷酸化Akt和磷酸化IKKα/β的表达。转录因子NF-κBp65的核定位减少,表达量降低。但胞浆中总Akt表达在实验组和对照组之间无显著性差异。各实验组间表达量比较,结果显示在RNA-DC-CIKs组phospho-Akt和phospho-IKKα/β和转录因子NF-κBp65表达量比CIK组更低,但无显著性差异。结论肿瘤RNA负载DC-CIKs、DC-CIKs和CIKs均能显著抑制PC3细胞内Akt的活化,降低活化的IKKα/β的表达,导致转录因子NF-κBp65核内定位和表达降低,从而抑制细胞生存信号通道,诱导肿瘤细胞凋亡。

英文摘要：

【Objectives】To test the effcets of tumor RNA-pulsed DCs cocultured with autologous CIKs on Akt/ NF-κB survival signaling pathway of HRPC cell lines PC3 and explore the molecule mechanisms of cancer cells apoptosis induced by these immune cells.【Methods】PC3 cells were seeded in TRANSWELL plate and interacted with different effector cells including CIKs,DC-CIKs（CIKs cocultured with nude DCs） and RNA-DC-CIKs（CIKs cocultured with autologous tumor RNA-transfected DCs）.The target PC3 cells in defferent experiment group were collected respectively and extracted cell total protein and nuclear extracts respectively.Akt,phospho-Akt,phospho-IKKα/β and transcription factor NF-κBp65 were checked by western blotting assay,using untreated PC3 as negative control and PC3 treated by PI3K inhibitor LY294002 as positive control.The housekeeping gene-product β-actin is used as an internal loading control.Western blot assays were analyzed with densitometry software（Quantity One）.【Results】Compared to untreated PC3,the activations of Akt and IKKα/β were significantly inhibited by CIKs,DC-CIKs and RNA-DC-CIKs,and the expression and nucleic localization of transcription factor NF-κBp65 were re-duced in all experiment groups.However,the expression of total Akt was no difference in all experiment groups andcontrol groups.Furthermore,the results showed the expressions of phospho-Akt,phospho-IKKα/β and NF-κBp65 inRNA-DC-CIKs group were lower than the ones in CIKs group,but this difference was no statistical significance.【Conclusion】CIKs,DC-CIKs and RNA-DC-CIKs could inhibit the activation of Akt and IKKα/β in PC3 cells,andreduce the expression and nucleic localization of transcription factor NF-κBp65,therefore they can regulate and con-trol the Akt/NF-κB survival signaling pathway,resulting in cancer cells apoptosis.