摘要
目的研究阿霉素诱导骨肉瘤细胞凋亡过程中Mcl-1含量的变化以及Mcl-1对骨肉瘤细胞凋亡的调控作用。方法采用半定量PCR法检测Mcl-1、Bcl-2、β-actin的mRNA水平变化;Western blot法检测Mcl-1、Bcl-2、β-actin蛋白变化;转染pcDNA3-Mcl-1质粒使细胞高表达Mcl-1、Bcl-2;采用Caspase3/7活性试剂盒检测Caspase3/7活性。结果骨肉瘤细胞在阿霉素作用下,Mcl-1和Bcl-2的mRNA水平无变化,而蛋白水平下调;过表达Mcl-1可以明显抑制凋亡标志物PARP的剪切,而Bcl-2不具有抑制效应;同样,过表达Mcl-1可以抑制阿霉素作用下Caspase3/7的活化。结论阿霉素作用下,Mcl-1蛋白水平下降是骨肉瘤细胞发生凋亡的重要原因,并且该凋亡过程与Caspase的活化有关。
Objective To explore the expression level and functional role of Mcl-1 under doxorubicin-induced apoptosis.Methods The mRNA levels of Mcl-1,Bcl-2 and β-actin were detected by semi-quantitative RT-PCR.The protein levels of Mcl-1,Bcl-2 and β-actin were detected by Western blot.Transfections of pcDNA-flag-Mcl-1 and pcDNA-flag-Bcl-2 were used to overexpress Mcl-1 and Bcl-2 in cells.Caspase-3 /7 activities were measured by Caspase-3 /7 assay kit.Results Although the expression of Mcl-1 and Bcl-2 mRNA stayed the same level,Mcl-1 protein level decreased in osteosarcoma under doxorubicin-induced apoptosis.Overexpression of Mcl-1,not Bcl-2,significantly inhibited cleavage of PARP,which was marker for apoptosis.Meanwhile,overexpression of Mcl-1 also inhibited activity of caspase-3 /7 by doxorubicin.Conclusion The downregulation of Mcl-1 protein level leads to an increase of apoptosis in osteosarcoma cells under doxorubicin treatment.In addition,this process is related to Caspase.
出处
《安徽医科大学学报》
CAS
北大核心
2012年第5期492-495,共4页
Acta Universitatis Medicinalis Anhui
基金
安徽省教育厅自然科学基金(编号:KJ2011A167)
