反义Bmi-1对Jurkat细胞的抑制作用  

Inhibition effect of antisense Bmi-1 on Jurkat cells

作　　者：刘卫红[1] 孟秀香[1] 刘丹丹[2] 单路娟[3] 赵心宇[1] 

机构地区：[1]大连医科大学检验医学院,116027 [2]大连医科大学诊断学实验中心 [3]大连医科大学附属一院中心实验室

出　　处：《中华血液学杂志》2005年第9期554-556,共3页Chinese Journal of Hematology

摘　　要：目的观察反义Bmi-1表达质粒在体外对T淋巴细胞白血病Jurkat细胞的生长抑制作用。方法选择Bmi-1基因起始密码子及与锌指结构对应的171bp核苷酸片段,反向连接于pLNCX2质粒上,得到反义Bmi-1表达质粒;通过脂质体转染将质粒导入Jurkat细胞中,以G418进行筛选得到阳性克隆;以MTT法和体外集落形成实验检测反义Bmi-1表达质粒对Jurkat细胞增殖的抑制作用;用流式细胞术分析细胞周期;采用免疫荧光组化法检测反义Bmi-1对Jurkat细胞P16蛋白的上调作用。结果转染反义Bmi-1表达质粒组与对照组(空白对照及空质粒组)相比较,生长速度明显变慢,克隆形成能力明显下降,空白对照组Jurkat细胞集落数为(90.70±9.07)/103细胞,空质粒组Jurkat细胞集落数为(83.30±6.11)/103细胞,转染反义质粒组Jurkat细胞集落数为(56.00±5.56)/103细胞(P<0.01);P16蛋白表达也明显增强。结论反义Bmi-1对Jurkat细胞的体外生长具有明显的抑制作用,并上调了P16蛋白的表达。Objectives To investigate whether antisense Bmi-1 plasmid could inhibit the proliferation of Jurkat cells. Methods The antisense plasmid was constructed by PCR amplification of a 171 bp segment spanning Bmi-1 start eodon and zinc finger structure and the PCR product was subsequently inserted reversedly to plasmid pLNCX2. The final construct was eonfirnled through restriction enzyme digestion. G418 was added into the medium after the planlid was successfully introduced into Jurkat cells by using lipofeetinmediated DNA transfection. The proliferation of Jurkat cells were determined by MTT and colony formation assays. Cell cycle was determined by flow cytometry. The p16 expression of Jurkat cells was studied by immunofluorescent histochemistry. Results The growth rate of antisense Bmi-1 transfected Jurkat cells was significantly lower than that of the controls, and the colony forming capacity of the transfected cells decreased significantly （ P 〈 0.01 ） , the colony numbers being（ 90.7 ± 9.07 ）/10^3 cells, （ 83.3 ± 6.11 ）/10^3 eells and（ 56.0 ± 5.56）/10^3 cells for control cells, empty plasmid transfected Jurkat cells and antisense Bmi-1 transfected Jurkat cells, respectively. The percentage of G1 phase cells was increased and the p16 expression of antisense Bmi-1 transfected cells was significantly upregulated than that of control cells. Conclusion Antisense Bmi-1 can inhibit the growth and upregulate the expression of p16 of Jurkat cells in vitro.

关 键 词：细胞系 Jurkat 寡核苷酸 反义 基因 Bmi-1 JURKAT细胞 生长抑制作用 转染反义 P16蛋白表达 T淋巴细胞白血病 体外集落形成 t细胞集落 

分 类 号：R733.7[医药卫生—肿瘤]