| Objective:To evaluate the effect of salidroside on the proliferation of a human cervical cancer cell line SiHa, and further investigating its possible molecular mechanisms.Methods:SiHa cells, a human cervical cancer cell line, were cultured in vitro. MTT cell viability assay was used to evaluate the cell viability of SiHa cells with varying concentrations of salidroside for12h,48h and72h, then IC50was calculated. Morphological alterations were observed in SiHa cells pretreated with varying concentrations of salidroside for48h by inverted phase contrast microscope. Flow cytometry technology analyzed the changes of cell cycle distribution and apoptotic rate induced by varying concentrations of salidroside for48h exposure. After48h treatment of salidroside at varying concentrations, SiHa cells were stained by Hoechst33258to observe the morphological changes through fluorescence microscopy. Western blot further detected the expression changes of cell cycle related proteins Cdc2, cyclinBl, CDK2and cyclinA as well as apoptosis related proteins (Bax, Bcl-2, Fas, FasL and cleaved caspase-3) in SiHa cells, which were treated with varying concentrations of salidroside.Results:The results determined by MTT assay showed that the viability of human cervical cancer SiHa cells with salidroside was markedly attenuated in a concentration and time dependent manner (P<0.05). The IC50values were calculated as (107.42±1.44)μg/ml,(86.87±2.90)μg/ml and (76.98±1.74)μg/ml in24h,48h and72h, respectively. The morphological feature of SiHa cells changed significantly with the increase of the concentration of salidroside after treated for48h. Flow cytometry analyzed that salidroside could affect cell cycle distribution of SiHa cells. With increasing concentrations of salidroside, the percentages of G0/G1phase cells were gradually decreased while the percentages of G2/M phase were increased (P<0.05). In addition, as compared with the control (Oμg/ml), SiHa cells were markedly accumulated at the S phase induced by varying concentrations of salidroside (P<0.05), except a concentration of100μg/ml. Besides, the apoptotic rates in salidroside treated SiHa cells at the concentrations of20μg/ml,40μg/ml,60μg/ml and80μg/ml were(8.55±0.54)%,(19.44±0.51)%,(42.52±0.61)%and (58.87±1.08)%, respectively, which were significantly higher than the contro(1.67±0.51)%(P<0.05). Microscopic observation by Hoechst33258staining determined the cell apoptosis in salidroside-treated SiHa cells and cells with typical apoptosis characteristics were observed. Western blot confirmed that20μg/ml,40μg/ml and80μg/ml salidroside could induce a significant decrease of Cdc2, cyclinB1, CDK2and cyclinA (P<0.05). In addition, salidroside (20μg/ml,40μg/ml,60μg/ml and80μg/ml) significantly up-regulated the expression levels of Bax, Fas and cleaved caspases-3as compared to the control (P<0.05), while down-regulated the expression levels of FasL and Bcl-2, the differences between control and groups with varying concentrations of salidroside were statistically significant (P<0.05) except the level of Bcl-2at a concentration of20μg/ml salidroside (P>0.05). Moreover, the ratio of Bax to Bcl-2were increased with the concentration of salidroside raised.60μg/ml and80μg/ml salidroside significantly increased the ratio of Bax to Bcl-2(P<0.05).Conclusion:salidroside inhibited the proliferation of human cervical cancer SiHa cells, which was probably associated with the induction of cell apoptosis and cell cycle arrest at G2/M and S phase. |
PDF Full Text Request