基础医学与临床 ›› 2007, Vol. 27 ›› Issue (6): 616-620.

• 研究论文 • 上一篇    下一篇

mCD99L2基因对小鼠B淋巴瘤细胞株A20细胞转化H/RS样

何滢 沈丽佳 李祖国 谢卫兵 谢思明 刘芳 刘鹏飞 蒋会勇 赵彤   

  1. 南方医科大学附属南方医院病理科 南方医科大学附属南方医院病理科 南方医科大学附属南方医院病理科 南方医科大学附属南方医院病理科 南方医科大学附属南方医院病理科 青海医学院生物化学教研室 南方医科大学附属南方医院病理科 南方医科大学附属南方医院病理科 军事医学科学院基础医学研究所 北京
  • 收稿日期:2006-07-17 修回日期:2007-01-05 出版日期:2007-06-25 发布日期:2007-06-25
  • 通讯作者: 何滢

Effect of mCD99L2 gene on the cell biological characteristic of mice lymphoma cell A20

  

  • Received:2006-07-17 Revised:2007-01-05 Online:2007-06-25 Published:2007-06-25

摘要: 目的 探究mCD99L2基因沉默对小鼠B淋巴瘤细胞系A20细胞转化为H/RS样细胞的影响。 方法 重组SiRNA表达质粒LV-mCD99L2,体外转染内源性mCD99L2表达阳性的A20细胞,筛选出稳定表达LV质粒的细胞株并扩增培养;采用免疫荧光技术和流式细胞仪检测转化前后两组细胞鼠源CD30表达;透射电镜观察转化后细胞超微结构的形态特点; 细胞计数方法动态观测培养细胞干扰组A20-LV-mCD99L2和未经干扰组A20细胞的H/RS样细胞(直径≥25μm)转型率,以人霍奇金淋巴瘤细胞系L428作为对照;采用流式细胞仪检测细胞周期变化。 结果 获得了稳定表达LV质粒的单克隆细胞株A20-LV- mCD99L2;免疫荧光标记显示转化细胞CD30(+);流式细胞仪检测A20-LV-mCD99L2细胞 CD30阳性率为54.4%;透射电镜观察转化后细胞核增大,可见单核、双核及多核,核仁明显的H/RS样细胞;干扰组H/RS样细胞的转型率明显高于未经干扰组(P<0.01)。两组处于S期的细胞无明显差异,两组细胞均未见凋亡峰。结论 mCD99L2基因沉默可诱导小鼠B淋巴瘤细胞系A20细胞转化为H/RS样细胞。

Abstract: Objective To investigate the effects of mCD99L2 gene silencing on the mice B lymphoma cell line A20 transferred to H/RS-like cells. Method Recombinant plasmid LV-mCD99L2 was transfected into A20 cells which endogenetically express mCD99L2. The stably expressed cell line was selected by Blasticidin. The expression of CD30 in transferred cells was detected by immunofluorescense and FCM method. Observation of A20-LV-mCD99L2 cells was applied by transmission electron microscope. The proliferation of the tumor cells (the diameter of H/RS-like ≥25μm) of A20- LV- mCD99L2 and A20 in vitro and in nude mice was examined by continuously cell counting, and taken the human Hodgkin lymphoma cell line L428 as control. The cell cycle was detected by FCM method. Result Stable expression cell clone A20-LV- mCD99L2 was obtained after recombinant plasmid transfection. The expression of CD30 in transferred cells was positive, and the rate of expression of CD30 in A20-LV- mCD99L2 was 54.4%. The observation of transferred cells showed that the cells nuclear were larger and H/RS-like cells with obvious nucleoli which were uninucleated, binucleated and multinucleated cells. The result of cell counting showed that the rate of H/RS-like cells of A20-LV-mCD99L2 was obviously higher than that of A20(P<0.01)in vitro and in vivo. Cell cycle analysis showed that no obvious differences between A20-LV- mCD99L2 and A20 in S stage and no obvious apoptosis was detected in A20-LV- mCD99L2 and A20. Conclusion mCD99L2 gene silencing may transfer mice B lymphoma cell line A20 into H/RS-like cells in vitro and in vivo.