Yu Qi, Meng Xiu-xiang, Liu Ben, Liu Dan-dan, Zhang Wei, Liu Wei-hong, Yang Guang
College of Laboratory Medicine, Dalian Medical University, Dalian 116044, China.
Zhonghua Bing Li Xue Za Zhi. 2009 Dec;38(12):829-32.
To study the effects of antisense Bmi-1 (B cell-specific moloney murine leukemia virus insertion site 1) RNA on the growth, cell cycle and apoptosis of lung cancer cell line A549.
Recombinant plasmids carrying antisense Bmi-1 RNA were transfected into A549 cells, which expressed a high level of endogenous Bmi-1. The mRNA level of A549 cell was analyzed by real time quantitative RT-PCR and the protein level was determined using Western blot. MTT growth curve and plate colony forming assay were used to measure the effect of antisense Bmi-1 RNA expression on the growth of A549. Flow cytometry was used to analyze cell cycle and apoptosis.
Antisense Bmi-1 RNA reduced the Bmi-1 expression at the protein level, but did not alter the mRNA level in A549 cells. Compared with the control cells, A549 cells transfected with antisense Bmi-1 RNA showed a strong inhibition of the cell growth. The number of plate colony formation of the antisense Bmi-1 transfected cells (0.67 +/- 0.50) was less than those of the control (73.0 +/- 4.1) and cells transfected with empty vector (67.0 +/- 4.0, P < 0.01). Transfection of antisense Bmi-1 RNA arrested the A549 cells at G₀/G₁ phase of the cell cycle and did not increase the apoptosis.
Antisense Bmi-1 RNA expression inhibits A549 cells proliferation, likely through the interference of Bmi-1 leading to an arrest of the proliferating cells at the G₀/G₁ phase.
研究反义Bmi-1（B细胞特异性莫洛尼鼠白血病病毒插入位点1）RNA对肺癌细胞系A549生长、细胞周期及凋亡的影响。
将携带反义Bmi-1 RNA的重组质粒转染至内源性Bmi-1高表达的A549细胞。采用实时定量逆转录聚合酶链反应分析A549细胞的mRNA水平，并用蛋白质免疫印迹法测定蛋白质水平。采用MTT生长曲线及平板集落形成试验检测反义Bmi-1 RNA表达对A549细胞生长的影响。运用流式细胞术分析细胞周期及凋亡情况。
反义Bmi-1 RNA在蛋白质水平降低了Bmi-1的表达，但未改变A549细胞中的mRNA水平。与对照细胞相比，转染反义Bmi-1 RNA的A549细胞的生长受到显著抑制。反义Bmi-1转染细胞的平板集落形成数（0.67±0.50）少于对照细胞（73.0±4.1）及转染空载体的细胞（67.0±4.0，P<0.01）。转染反义Bmi-1 RNA使A549细胞停滞于细胞周期的G₀/G₁期，且未增加细胞凋亡。
反义Bmi-1 RNA表达抑制A549细胞增殖，可能是通过干扰Bmi-1导致增殖细胞停滞于G₀/G₁期。
