莪术油微球释放莪术油对人肝癌SMMC-7721细胞的作用

Effect of CAO released from MS-CAO on human hepatoma cell line SMMC-7721

目的:探讨莪术油微球(CAO-MS)释放出莪术油(CAO)对人肝癌SMMC-7721细胞的生长抑制作用及机制.方法:用MTT法测定CAO及CAO-MS对体外培养的人肝癌细胞系SMMC-7721的生长抑制作用.用FCM测定CAO及CAO-MS对人肝癌细胞周期的影响.用DNA含量测定、AnnexinⅤ标记、电镜观察细胞形态检测CAO及CAO-MS对人肝癌细胞的诱导凋亡作用.用免疫细胞化学(ICC)及RT-PCR方法分别检测p21WAF1/CIP1蛋白与mRNA表达水平.结果:CAO与CAO-MS对肝癌细胞的生长抑制作用与药物浓度及作用时间呈一定的依赖关系(P<0.01).药物作用72h时,CAO与CAO-MS对SMMC-7721的半数抑制浓度(IC50)分别约为50mg/L和100mg/L.50mg/LCAO和100mg/LCAO-MS作用于肝癌细胞72h后,使G0+G1比例增加,S期及G2+M期比例相对下降,细胞周期阻滞于G0+G1期.DNA含量测定、AnnexinⅤ标记、形态学研究结果提示,CAO与CAO-MS均能诱导肝癌细胞的凋亡,CAO的作用优于CAO-MS.ICC和RT-PCR检测结果表明,50mg/LCAO和100mg/LCAO-MS作用于肝癌细胞72h后,均能提高CAO-MSp21WAF1/CIP1蛋白及mRNA表达水平,相同剂量的CAO的作用优于CAO-MS.结论:CAO及CAO-MS对人肝癌细胞SMMC-7721均有显著的抑制增生作用,作用呈剂量依赖性;CAO及CAO-MS能诱导对人肝癌细胞SMMC-7721凋亡,并能使细胞周期阻滞于G0+G1期;CAO及CAO-MS抑制肝癌细胞增生及诱导其凋亡的机制可能与其上调p21WAF1/CIP1基因表达有关.

AIM:To explore the inhibitory effects and mechanisms of CAO released from CAO-MS on the growth of human hepatoma cell line SMMC-7721. METHODS:Inhibitory effects of CAO and CAO-MS on the growth of human hepatoma cell line SMMC-7721 in vitro were determined with MTT assay. The cell cycles of SMMC- 7721 treated with CAO and CAO-MS were analyzed with flow cytometry. The apoptosis of human hepatoma cell line was tested with cellular DNA quantitative analysis, Annexin Ⅴ labeling and morphological study. The expressions of p21WAF1/CIP1 protein and mRNA were detected, respectively,with immunocytochemistry staining (ICC) and reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS:The inhibitory effects of CAO and CAO-MS on the growth of human hepatoma cell line were dose- and time- dependent (P <0.01). The IC50s of CAO and CAO-MS on SMMC-7721 were 50 mg/L and 100 mg/L, respectively. The propoertion of SMMC-7721 in G0/G1 phase increased and that in S and G2+M phase decreased after exposure with 50mg/L CAO and 100mg/L CAO-MS for 72 hours. ICC and RT-PCR analysis showed that both CAO and CAO-MS incresaed the level of p21WAF1/CIP1 protein and mRNA.The effect of CAO was superior to that of CAO-MS with the same dosage of CAO and CAO-MS. CONCLUSION:Dose-dependent inhibitory effects of CAO and CAO-MS on the growth of human hepatoma cell line are remarkable. Both CAO and CAO-MS could induce apoptosis of human hepatoma cell and keep the cell cycle at G0/G1 phase from up-regulating expression of p21WAF1/CIP1 protein and mRNA.