脂质体介导bcl-xL反义寡核苷酸对鼻咽癌CNE-2Z细胞株凋亡的影响

背景与目的:实验证明 bcl-xL在鼻咽癌 CNE-2Z细胞中存在高表达 , 它可能在鼻咽癌的发生发展中发挥着重要的作用 . 本研究以脂质体 (lipofectin)为载体 , 将人工合成的 bcl-xL反义寡核苷酸 (antisense oligodeoxynucleotide,ASODN)片段导入 CNE-2Z细胞 , 拟探讨 ASODN对 CNE-2Z细胞的影响 . 方法:以 bcl-xL的编码区为靶点 , 人工合成 20个碱基全硫代修饰的反义寡核苷酸 , 脂质体转染反义寡核苷酸进入 CNE-2Z细胞后 , 用 MTT法检测细胞成活率 ; 用琼脂糖凝胶电泳、荧光染色、流式细胞术等方法检测细胞凋亡 . 结果:MTT结果发现 , ASODN在 Lip介导下即可显著抑制 CNE-2Z细胞株细胞增殖 (P< 0.01), ASODN对细胞增殖的抑制作用随其浓度增加而增强 ; ASODN作用细胞 36 h后 , 经 Hoechst 33258/PI双染在荧光显微镜下可见细胞缩小、染色质固缩、核断裂 ; 流式细胞仪分析发现在 G1峰前出现一个亚二倍体峰即凋亡峰 ; 琼脂糖凝胶电泳分析可见 ASODN/Lip处理组出现明显的 " 梯状 " 外观等凋亡特征性改变 . 结论:ASODN脂质体转染 CNE-2Z细胞后可促进 CNE-2Z的细胞凋亡 ; 在肿瘤研究中 , bcl-xL可作为一个有用的靶基因 , 为开发鼻咽癌基因治疗药物提供实验依据 .

Apoptotic effect of bcl-XL antisense oligodeoxynucleotide mediated by lipofectin on cell strain CNE-2Z

BACKGROUND &OBJECTIVE:Recent studies have shown that overexpression of bcl-x L was detected in human nasopharyng eal carcinoma (NPC)cell strain CNE-2Z,sug g esting it may play a pivotal role in tumorig enesis of NPC.The current study was desig ne d to explore the effect of bcl-x L antisense olig odeoxynucleotide (ASODN)on CNE-2Z.METHODS :A 20-mer g apmer ASODN with a full phosphorothioate b ackbone targ eting a sequence unique of the bcl-x L coding reg ion was artificially synt hesized.Bcl-x L ASODN was transfected into CNE-2Z cells throu g h lipofectin.The survival rate was assessed by MTT assay and internucleosomal fr ag mentation of g enomic DNA was detec ted by ag arose g el electrophoresis.Apo ptotic chang es after treatment with ASODN were observed by fluorescence micro scopy and flow cytometry.RESULTS :MTT assay showed that the proliferation of CNE-2Z cells decreased sig nificantly after treatment with ASODN /Lip as compare d with control (P<0.01).ASODN /Lip reduced the proliferation of CNE-2Z in a dose-dependent manner.After tr eatment with ASODN /Lip for 36hours ,most cel ls stained with Hoechst 33258/PI exhibited apoptotic cell morpholog y such as cell shrinkag e,nuclear condensation,and nuclear frag ment ation under fluorescence microscop e;a apoptotic peak appeared on flow cyto metry;a ladder-like pattern of DNA frag mentation appeared on ag arose g el electrophoresis.CONCLUSION:ASODN can inhibit proliferation of CNE-2Z cells and induce apoptosis of CNE-2Z cells.The results sug g est that bcl-x_L is a promising targ et for g ene therap y of NPC.