靶向survivin的RNAi联合X线照射对肺腺癌A549细胞周期、凋亡及增殖的影响

The effect of adenocarcinoma of lung A549 cell cycles,apoptosis and proliferation induced by RNAi targeting survivin gene combined X-rays radiation

目的构建靶向生存素(survivin)基因的RNA干扰(RNAi)载体,观察其联合X射线照射对肺腺癌A549细胞周期、凋亡及增殖影响。方法根据GenBank中survivin的cDNA序列设计干扰序列,构建干扰survivin的重组干扰质粒pGenesil2-survivin。酶切、测序鉴定正确后,干扰质粒经脂质体介导转染肺腺癌A549细胞,联合5Gy射线照射,应用PI单染、流式细胞仪检测细胞周期与凋亡变化,MTT检测细胞增殖的变化。结果酶切和测序结果显示,载体构建正确;转染空载体pGenesil2对细胞G0/G1期、S期及凋亡,增殖能力影响不大,降低了G2/M期(P<0.05);而干扰质粒pGene-sil2-survivin能延长G0/G1期,缩短S期,诱导细胞凋亡增加,降低细胞增殖能力(P<0.05);5Gy照射对细胞周期影响不大,增加细胞凋亡,降低增殖能力(P<0.05);转染pGenesil2-survivin后,进行5GyX射线照射,延长G0/G1期,缩短S期,诱导细胞凋亡增加,降低细胞增殖能力(P<0.05,P<0.01)。结论靶向survivin的RNAi可延长G0/G1期,缩短S期,增加细胞凋亡,降低细胞增殖能力,X射线照射能够增强该质粒的作用。

Objective To construct the vector that RNA interference （RNAi） targeting survivin gene,to observe its effect combined X-rays radiation on adenoearcinoma of lung A549 cell cycle, apoptosis and proliferation. Methods One pair RNAi sequence targeting survivin gene was designed according to cDNA sequence showed in GenBank to construct the recombinant RNAi plasmid pGenesil2-survivin. After identified by enzyme digestion and sequencing, the pGenesil2-survivin plasmid was transfected into A549 cells, the cell cycle and apoptosis of cells were measured by flow eytometry with PI dyed ; the cell proliferation was measured by MTT. Results The identification of pGenesil2-survivin vector by enzyme digestion and sequencing was right, after pGenesil2 was transfected into A549 cells, G0/G1 , S phage, apoptosis and cell proliferation changes were little, G2/M phage was decreased;but after pGenesil2 transfected into A549 cells, G0/G1 phage was prolonged ,S phage was shortened ,apoptosis was enhanced and proliferation was decreased ;5 Gy X rays irradiation enhanced apotosis and decreased proliferation ;after pGenesil2 transfected into A549 cells, the cells was irradiated with 5 Gy X rays, G0/G1 phage was prolonged, S phage was shortened, apoptosis was enhanced and proliferation was decreased. Conclusions The RNAi targeting survivin could prolong G0/ G1 phage,shorten S phage,enhance apoptosis and decrease proliferation,and X rays radiation could enhance these changes.