Apoptin引起肿瘤细胞G2-M阻滞

目的:研究凋亡素(apoptin)特异诱导肿瘤细胞凋亡的作用,探讨其用于肿瘤治疗的可能性.方法:用PCR技术克隆了apoptin基因,将其插入预先装有FLAG标签的真核表达质粒pcDNA3.1中,并构建了带有apoptin基因的腺病毒载体,观察转染pcDNA3.1/FLAG/apoptin和/或Ad/apoptin重组腺病毒感染后apoptin在几种细胞中的表达及细胞形态的改变,用流式细胞术分析PI染色后Ad/apoptin感染细胞的DNA含量变化.结果:apoptin在Hela、Bcap37、A549、HepG2、SKOV3等肿瘤细胞核中表达,apoptin重组腺病毒感染后细胞核染色质呈颗粒状,最后凝聚成不规则大块并出现核固缩.正向和反向插入apoptin的重组腺病毒均引起G2-M细胞增多,但前者引起G2-M阻滞更为显著,需要的感染复数(MOI)也较低.结论:apoptin可引起肿瘤细胞周期G2-M阻滞和染色质凝聚.

Apoptin induces G2-M arrest in cancer cells

Objective: To further study the effect of apoptin in inducing cancer cell specific apoptosis and the possible applications in cancer therapy. Methods: Apoptin gene was amplified by PCR and inserted into pcDNA3.1( ) with a FLAG tag in front of the multi-cloning-site. Apoptin gene with the FLAG tag was sub-cloned into an adenovirus vector. Several cancer cell lines were transfected with pcDNA3. 1/FLAG/apoptin or infected with apoptin containing recombinant adenoviruses to study the morphologic changes. Ad/apoptin infected cells were also analyzed by flowcytometry after staining with PI. Results: Expressed apoptin was localized in the nucleus of cancer cells. Chromatin condensation occurred 2 or 3 days after Ad/apoptin( ) infection. Cell number in G 2-M phase increased dramatically after Ad/apoptin( ) infection. Conclusion: Apoptin can induce cell cycle G 2-M arrest and chromatin condensation in cancer cells.