| Objective:To study the effects of dihydroartemisinin(DHA) on the proliferation, cell cycle distribution, apoptosis of PC-3 cells and its possible mechanism.Methods:PC-3 cells were divided into three groups: Control, DHA and DHA+ transferrin (TF). The control group included HamF12, TF (25μg/ml) and 0.1%DMSO subgroups. The DHA group and the DHA+TF group were both subdivided into 5 subgroups according to the concentration of DHA while that of TF was 25μg/ml in each DHA+TF subgroups. Proliferation assay was tested by MTT method on PC-3 cells in different groups as described above and all treated for 24, 48, 72 hours. Cell cycle distribution, apoptosis and necrosis of PC-3 cells treated with HamF12, DHA and DHA+TF (DHA concentration: 12.5, 50, 200μmol/L) were determined by using flow cytometry (FCM) analysis after 48, 72 hours, and 48 hours groups among them were examined for Bcl-2, Bax and P53 by immunocytochemical staining and the results were undergone an image analysis for average optical. Ultrastructural changes of PC-3 cells treated with DHA of different concentration for 48 hours were observed by SEM and TEM.Results:Comparison with inhibition ratios of PC-3 cells treated with DHA and DHA+TF, both of them had significant time and concentration-dependent relation (P<0.001) while no significant deviation between the two groups (P>0.05), inhibition ratios were 13.66±20.46%, 78.40±6.79% when treated with DHA 12.5μmol/ml for 24h and 200μmol/ml for 72h, respectively, and that was up to 82.64±8.31% when treated with DHA+TF (DHA 200μmol/ml, 72h). FCM analysis showed DHA concentration had an inverse correlation with the portion of PC-3 cells in G0/G1 cell cycle phase and a positive correlation with which in G2/M phase after treated with DHA and DHA+TF for 48 hours, and there was no significant deviation between the two groups; Compared to the control group, the rate of cell apoptosis and necrosis raised both in DHA and DHA+TF groups, especially in high DHA concentration (200μmol/ml) for 48, 72 hours. Immunocytochimical staining showed low-expression of Bcl-2 and over-expression of Bax which had correlation with DHA concentration after treated with DHA and DHA+TF for 48 hours, but negative expression of P53 was shown in all groups. EM showed the decrease and loss of microvilli on the cellular surface in both apoptotic and necrotic cells. in apoptotic cells, drumstick-shaped and globe-like membrane blebbing on the cellular surface occured, cellular nucleus chromatin condensed, margination, light mitochondrial swelling, nuclear fragmentations and apoptosis body with intact membrane were also observed; in necrotic cells, Cellular volume increased, electron density of cytoplasm decreased obviously, mitochondrial swelling, karyolysis, broken plasma membranes and cytoplasma outflow were observed in necrotic cells.Conclusion:DHA can effectively inhibit PC-3 cells proliferation, display strong cytotoxic effects and accumulate PC-3 cells in the G2/M cell cycle phases. It can low express Bcl-2 and over express Bax to induce apoptosis in PC-3 cells, and its effects on P53 can't be indicated in this test. 25μg/ml TF added has not significant influence on the effects of DHA.
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