目的 探讨表达c FLIPs重组腺病毒诱导淋巴细胞抗凋亡作用。方法 采用RT PCR方法,从人T淋巴细胞株的总RNA中克隆c FLIPs基因;通过pAdeasy系统,构建表达c FLIPs的重组腺病毒Ad c FLIPs;Hochest染色法及PI染色流式细胞仪检测anti Apo 1诱导感染Ad c FLIPs后的H9细胞凋亡。结果 采用RT PCR方法从人T淋巴细胞株中扩增出c FLIPs基因;构建重组腺病毒Ad c FLIPs ;经Hoechst染色,荧光显微镜下观察细胞核的形态,结果发现,经anti Apo 1诱导凋亡处理2 4h后,未经Ad c FLIPs感染的H9细胞有大量的细胞核呈浓染致密的固缩形态或颗粒状荧光的凋亡细胞;Ad c FLIPs感染的H9细胞中细胞核呈浓染致密的固缩形态或颗粒状荧光的细胞数明显减少,大部分细胞染色质呈弥漫均匀低强度荧光;流式细胞仪分析经PI染色后的H9细胞,未经Ad c FLIPs预处理的H9细胞在anti Apo 1作用2 4h后,细胞的凋亡率分别为4 8 33%±7 4 1% ;经Ad c FLIPs预处理1d的H9细胞,其细胞凋亡率分别下降到3 6 0 %±0 2 1%。结论 重组腺病毒Ad c FLIPs可有效地诱导淋巴细胞的抗凋亡作用。
Objective： To investigate the therapeutic efficacy of adenovirus-mediated herpes simplex virus thymidine kinase （HSV-tk） gene transfer under the driving of KDR promoter （AdKDR-tk） in combination of ganciclovir （GCV） against human hepatocellular carcinoma in nude mice. Methods： HepG2 cell line was implanted subcutaneously into 32 nude mice, which were subsequently divided into 4 groups （n=8 each group）： Ganciclovir group （Ⅰ）, Ad group （Ⅱ）, AdCMV-tk/GCV group （under the driving of CMV promoter） （Ⅲ） and AdKDR-tk/GCV group （Ⅳ）. Then intratumoral injection of recombinant adenovirus or Ad was performed in all nude mice, and repeated 24 h later. For the following 10 d GCV was given at a dose of 100 mg/（kg· d）, ip. All the treated animals were killed to evaluate the tumor weight and the histopathological changes and the microvessel density of tumors after the treatment was determined. Results Compared with group Ⅰ, the tumor inhibitory rate was 12.3% in group Ⅲ and 24.5% in group Ⅳ; the inhibition rates were significantly different between group Ⅲand Ⅳ （P〈0.05）. The mean MVDs in group Ⅰ, Ⅱ, Ⅲ and Ⅳ were 37.4±8.6, 30.6±7.8, 27.6±7.1, and 10.7±4.1 （microvessels/mm^2）, respectively. Significant differences were found between group Ⅲ and Ⅱ （P〈0.05）, Ⅳ and Ⅱ （P〈0.01）, and Ⅳ and Ⅲ （P〈0.01）. Conclusion： Intratumoral injection of AdKDR-tk results in marked inhibition of HCC growth through inhibition angiogenesis in nude mice. It may be a new treatment approach for human HCC,
