地塞米松对人肝癌细胞系SMMC-7721凋亡的影响

Effect of Apoptosis on Human Hepatocellular Carcinoma Cell Line SMMC-7721 Influenced by Dexamethasone

观察不同浓度的地塞米松(dexamethasone,Dex)对人肝癌细胞系SMMC-7721凋亡的影响。SMMC-7721细胞培养于DMEM(高糖,含10%胎牛血清)培养液中,待细胞融合度达70%时,分别加入地塞米松,使其终浓度分别达到0.5μg/m L(低浓度组)、1.0μg/m L(中浓度组)和2.0μg/m L(高浓度组),另设一生理盐水对照组。诱导24 h后,Hoechst 33258染色,荧光倒置显微镜观察凋亡细胞形态和比例;发现随着Dex浓度的变化,凋亡的肝癌细胞数量、形态变化明显,Dex浓度越高,凋亡的肝癌细胞越多,并呈现浓度依赖性,即凋亡数与Dex浓度呈正相关。提取细胞总RNA,逆转录成为c DNA,荧光定量PCR法测定凋亡相关基因Bcl-2、Caspase 3、Caspase 5、Caspase 7、Caspase 9、Fas及P53的表达。与对照组比较,3种浓度Dex处理的细胞其凋亡细胞形态和比例均发生显著差异,凋亡细胞数量显著增加,各相关基因表达均显著下降。Dex对SMMC-7721细胞具有诱导凋亡的作用。

To observe the effect of apoptosis on human hepatocellular carcinoma cell line SMMC-7721 influenced by dexamethasine （Dex）. SMMC-7721 cells were cultured in DMEM medium （high glucose, containing 10% fetal bovine serum）. After fusion of cells reached to 70%, Dex added to medium at the concentration of 0.5μg/mL （low concentration group）, 1.0 μg/mL （middle concentration group） and 2.0/xg/mL （high concentration group） respectively, and a saline group was set as a control. After 24 h, cells were stained using Hoechst 33258 and observed the morphology of apoptotic cells and ratio under fluorescent inverted microscope. With the change of Dex concentration, the number and morphology of apoptotic cells changed obviously. As Dex concentration increased, the number of apoptotic cells increased in a concentration-dependent manner, that is to say, the number of apoptotic cells was positively correlated with the Dex concentration. Total RNA was isolated and cDNA was synthesized by reverse transcription reaction. Realtime quantitative PCR （qPCR） was performed to detect the exp- ression of apoptosis related genes Bcl-2, Caspase 3, Caspase 5, Caspase 7, Caspase 9, Fas and P53. Compared to control group, the morphology of apoptotic cells and ratio changed obviously at 3 concentration of Dex, the number of apoptotic cells increased markedly. The expression of apoptosis related genes decreased significantly. Dex could induce apoptosis of SMMC-7721 cell line.