Flavopiridol对尤文肉瘤SK-N-MC细胞增殖的抑制作用  

Inhibitory effects of flavopiridol on the proliferation of Ewing sarcoma SK-N-MC cell line in vitro and in vivo

作　　者：李旭[1] 范姝丽[2] 李岩 朱悦[1] 屠冠军[1] 范广宇[1] 

机构地区：[1]中国医科大学附属第一医院骨科,沈阳110001 [2]中国医科大学附属第一医院干诊科,沈阳110001

出　　处：《中华骨科杂志》2007年第7期534-538,共5页Chinese Journal of Orthopaedics

基　　金：国家自然科学基金专项基金资助课题（30640001）

摘　　要：目的探讨细胞周期蛋白激酶抑制剂（flavopiridol）在体内外抑制尤文肉瘤细胞增殖的作用及其机制。方法以不同浓度的flavopiridol作用于培养的SK-N-MC细胞,用噻唑蓝法（MTT）检测细胞增殖,流式细胞分析仪测定细胞周期。采用DNA凝胶电泳检测法,观察flavopiridol诱导细胞凋亡的情况;蛋白质电泳观察Bcl-2及Mcl-1表达变化;建立SK-N-MC细胞裸小鼠皮下移植瘤模型,腹腔内给药,观察肿瘤生长情况;肿瘤标本石蜡包埋切片,免疫组织化学原位末端标记法（TUNEL）检测flavopiridol在体内对肿瘤细胞的凋亡诱导情况。结果（1）Flavopiridol对人尤文肉瘤细胞增殖有抑制作用,时间越长、浓度越高,抑制作用越强。（2）Flavopiridol可改变尤文肉瘤细胞周期,随药物浓度增加Sub-G1期百分比逐渐上升。尤文肉瘤细胞经flavopiridol中、高浓度给药后,DNA凝胶电泳可见典型的梯状带。（3）Flavopiridol给药前后,Bcl-2蛋白表达无变化,而Mcl-1蛋白表达明显下降。（4）TUNEL染色结果显示实验组阳性细胞比例上升,与对照组比较差异有统计学意义。结论Flavopiridol能在体内外抑制尤文肉瘤细胞的增殖,其机制主要是阻滞细胞周期的G1/S期,诱导细胞凋亡。Objective To investigate the effects of cyclin-dependent kinase inhibitor （flavopiridol） on the proliferation of human Ewing sarcoma in vitro and in vivo. Methods SK-N-MC cell lines were treated with flavopiridol in different concentration. Methabenzthiazuron test （MTT） was performed to measure the proliferation of the cells; cell cycle distribution was determined with flow cytometre; apoptosis of SK-N- MC cells was detected by DNA ladder; Bcl-2 and Mcl-1 gene expression related to apoptosis were examined by immunoblotting. The subcutaneous tumors of SK-N-MC cells were established in nude mice to study the antitumor effects of flavopiridol; TUNEL staining was examined to study weather flavopiridol induces apeptosis in vivo. Results 1） Flavopiridol inhibited cell growth in vitro in dose-dependent and time-dependent manner. The ICs0 was 105 nmol/L. 2） The flavopiridol changed cell cycle progression in human Ewing sarcoma cell lines, and the cell population in the Sub-G1 DNA content in human Ewing sarcoma cells dose-dependently increased with treatment of flavopiridol. When human Ewing sarcoma cell lines were treated by flavopiridol in middle and high dose, the formation of DNA ladders was observed in the cells. 3） The expression of Bcl-2 protein was not affected by flavopiridol, whereas the protein expression of Mcl-1 decreased in flavopiridol-treated cell lines. 4） TUNEL staining suggested that flavopiridol could induce apoptosis of Ewing sarcoma in vivo, there was statistical significant difference compared with control group. Conclusion Flavopiridol can inhibit the proliferation of SK-N-MC cells not only in vitro but also in vivo mainly by induction of apoptosis.

关 键 词：肉瘤 Ewing 细胞周期 细胞凋亡 

分 类 号：R686[医药卫生—骨科学]