凋亡素诱导人成骨肉瘤细胞多药耐药株凋亡的研究  

Apoptin Inducing Apoptosis in the Multidrug-Resistant Model of Human Osteosarcoma Cell Line

作　　者：孙大辉[1] 金宁一[2] 谷贵山 龚伟[2] 段德生[3] 

机构地区：[1]吉林大学第一医院骨科,长春130021 [2]解放军军需大学基因工程重点实验室 [3]吉林大学第三医院骨科

出　　处：《中国肿瘤生物治疗杂志》2001年第3期193-195,共3页Chinese Journal of Cancer Biotherapy

基　　金：国家973项目 (G1990 1190 2 )资助

摘　　要：目的 :研究凋亡素对人成骨肉瘤细胞OS 732多药耐药株R OS 732的作用。方法 :凋亡素基因VP3 经酶切后克隆入质粒 pIRES1,获得重组质粒pIRVP3,通过脂质体法 pIRVP3转染R OS 732细胞 ,RT PCR检测凋亡素在细胞中的表达 ;光镜及电镜下观察瘤细胞 ;提取瘤细胞基因组DNA进行琼脂糖凝胶电泳 ;流式细胞仪检测凋亡率。结果 :凋亡素基因已在转录水平表达 ;光镜及电镜下 2组细胞形态变化明显 ;电泳见转染组DNA呈梯状条带 ,对照组为单一条带 ;转染组凋亡率明显高于空白对照组 (P <0 .0 1)。结论 :凋亡素可有效诱导R OS 732细胞凋亡。Objective: To investigate the curative effects of apoptin on the multidrug-resistant model, of human osteosarcoma cell line (R-OS-732). Methods: 363 bpVP 3 gene was cloned into the vector pIRES1 and the recombinant eukaryon expression vector pIRVP3 was transfected into R-OS-732 cells with liposome. The expression of apoptin gene at transcription level was proved by RT-PCR. The pathological changes of the cells was observed by light microscope and electronmicroscope. The transfected R-OS-732 cells were collected after 48 hours. The genomic DNA extracted from the cells was observed by agarose gel electrophoresis. The apoptosis rate of the cells was analysed by flow cytometry. Results: The expression of apoptin gene at transcription level had been proved by RT-PCR. The construction changes of the two groups were obviously different under light microscope: most of R-OS-732 cells in the transfected group existed in form of being away from the bottom of the culture dish after 24 hours, in form of apoptosis after 48 hours and in form of necrosis after 72 hours; but those cells in the controlled group grow luxuriantly. And under electronmicroscope there was much change of cell nucleus between the two groups. DNA ladder of the transfected cells was observed through agarose gel electrophoresis only one band was observed in the controlled group; but bands of fragmented DNA observed in ositive control group. Flow cytometry analysis showed that the apoptosis rate of the cells in the transfected group was relatively higher than that of the controlls( P <0.01).Conclusion: Apoptin can induce apoptosis in the multidrug-resistant model of human osteosarcoma cell line(R-OS-732) .

关 键 词：凋亡素 成骨肉瘤 多药耐药性 凋亡细胞 

分 类 号：R738.1[医药卫生—肿瘤] R730.53[医药卫生—临床医学]