长链非编码RNA PVT1沉默对K562细胞增殖的影响

目的 筛选靶向抑制长链非编码RNA(lncRNA)浆细胞瘤转化迁移基因1(PVT1)的小干扰RNA(siRNA),并探讨其对人白血病细胞株K562细胞增殖的影响.方法 针对PVT1设计合成PVT1-siRNA1、2、3、4序列及无关对照siRNA(SC-siRNA)序列.利用HiperFect转染试剂将各条siRNA转入K562细胞,实验依此分为siRNA1组、siRNA2组、siRNA3组、siRNA4组和SC-siRNA组,并设空转组和细胞组.转染后用实时定量RT-PCR检测细胞中PVT1 RNA的相对表达情况,用CCK8法检测细胞增殖情况,用流式细胞仪检测细胞周期.结果 转染24、48 h后siRNA4组PVT1 RNA水平下降,与SC-siRNA组、空转组和细胞组相比差异有统计学意义(P<0.01), 而siRNA1组、siRNA2组和siRNA3组PVT1 RNA水平与细胞组相比差异无统计学意义(P>0.05);转染24、48、72 h后siRNA4组细胞增殖抑制率都增加,与SC-siRNA组和空转组相比差异均有统计学意义(P<0.01);转染48、72 h后siRNA4组出现了G1期阻滞,G1期细胞比例增加,与SC-siRNA组、空转组和细胞组相比差异有统计学意义(P<0.01).结论 PVT1-siRNA可通过阻滞细胞周期的进展而抑制K562细胞的增殖.

The effect of long non-coding RNA PVT1 silencing on K562 cell proliferation

Objective To screen the effective small interfering RNAs (siRNAs) targeting against long non-coding RNA (lncRNA) PVT1 (plasmacytoma variant translocation 1), and to observe its influence on the proliferation of human leukemic cell K562.Methods Four siRNA sequences targeting against PVT1 (PVT1-siRNA1, 2, 3, 4) and scramble control sequence (SC-siRNA) were designed and synthesized.Using HiperFect transfection reagent, all of the siRNAs were transfected into K562 cells.The experiment was divided into siRNA1 group, siRNA2 group, siRNA3 group, siRNA4 group, SC-siRNA group, idling group and untransfected group.The PVT1 RNA expression level was assessed by real-time RT-PCR.The cell proliferation was determined by CCK8 assay and the cell cycle was detected by Flow cytometry.Results The expression level of PVT1 RNA of K562 cells in siRNA4 group was significantly reduced compared with SC-siRNA group, idling group and untransfected group 24 and 48 hours after transfection (P<0.01), while there was no significant difference among siRNA1 group, siRNA2 group, siRNA3 group and untransfected group (P>0.05).The cell proliferation of K562 cells in the siRNA4 group was significantly inhibited as compared with SC-siRNA group and idling group 24, 48 and 72 hours after transfection (P<0.01).The cells in the siRNA4 group were arrested at G1 phase 48 and 72 hours after transfection (P<0.01).Conclusion PVT1-siRNA can inhibit the proliferation of K562 cells through arresting the cell cycle at G1 phase.