Effects And Mechanisms Of Lovastatin On Human Varian Cancer Cell Line SKOV3

Objective: To study the effects of Lovastatin on prolif- eration, apoptosis of human varian cancer cell line SKOV3. Meanwhile, To observe the effects of the combination of lovastatin with cisplatin on proliferation of human varian cancer cell line SKOV3. Furthermore,to investigate the possible mechanism of inhibiting cell proliferation and inducing apoptosis, and feasibility therapy and chemoprevention of varian cancer.Methods:1 SKOV3 cells were incubated in culture medium in vitro. Effect of lovastatin on the proliferation of SKOV3 cells was measured by MTT colorimetric method. Morphological changes of SKOV3 cells were observed by light microscopy and transmission electron microscopy. Apoptosis and distribution of cell cycle were examined by flow cytometry.2 The expression of Survivin, PTEN protein was qualitationly studied by immunocytochemistry staining and was semi-quantitately examined by flow cytometry after treated with lovastatin for 48h.3 MTT colorimetric method was performed to evaluate the potential cytostatic effect of combining lovastatin with cisplatin on human varian cancer cell line SKOV3. To determine whether the combination of lovvastatin and cisplatin results in a synergistic cytostatic effect, Jin's formula was performed;lovastatin and cisplatin were sequentially aplied on SKOV3 cell,the potential cytostatic effect was measured by MTT colorimetric method.Results:1 MTT colorimetric method showed that lovastatin (1.25,2.5,5,10,20,40μmol/L) could inhibit the proliferation of SKOV3 cells. After treated with lovastatin for 24h to 72h, compared with control group, the OD values of lovastatin -treated groups decreased, and there was significant difference between control group and every treatment group (p<0.05). Among every treatment group, there was also significant difference(p<0.05). Furthermore, with the increasing concentration of lovastatin and prolonging of treatment time, the OD values decreased gradually, that was, lovastatin inhibited the proliferation of SKOV3 cells significantly in a dose-and time-dependent manner. When treated with lovastatin, SKOV3 cells had significant morphological changes which were observed by light microscopy: The cells untreated with lovastatin were fusiform or diamond, and they were satiation. But the cells treated with lovastatin were shrinked and broken,and became irregular in shape, spreading a thin and long pseudopodium in both end. Vacuolus could be observed in the kytoplasm of some cells. The cells cast-off increased. Morphological changes of skov3 cells of Giemsa staining were further observed by light microscopy, SKOV3 cells untreated with lovastatin were diamond, and they were satiation, furthermore, the cell nucleus staining were uniform and clear, no floating cells. But SKOV3 cells treated with lovastatin became rounding, the cell kytoplasm was concentrated, the staining cell nucleus was uneven, there were also caryocinesis phenomenon, showing the typical apoptotic morphological changes. The cell ultramicrostructure changes were observed by transmission electron microscope: nucelus overshoot in acute angle outwards, chromatin concentratede highly, electron density raised and they were enriched as crescent below the nuclear membrane. We also observed karyopycnosis.When cells were harvested for the analysis on distribution of cell cycle and apoptosis by flow cytometry, the results were: After treated with 5,10,20μmol/L lovastatin for 48h, the number of cells in G0/G1 phase increased gradually, while the number of cells in S phase decreased grudually. That was, lovastatin could induce an arrest of cell cycle in G1 phase by a dose-dependent manner. In addition, after treated with with 5,10,20μmol/L lovastatin for 48h, the typical apoptotic peak which enhanced gradually with the increasing concentration of lovastatin was observed and the apoptotic percentage was 7.90%,13.07%,22.55%, seperately.2 Immunocytochemistry staining results:In the control group, the staining of Survivin was positive in SKOV3 cells and the buffy grains could be seen in endochylema, and the buffy grains was very thick;lovastatin weakened the stain degree of survivin in the SKOV3 cells,the stain and the buffy grains became thinner and the positive cell population was less than control group. In the control group, the staining of PTEN was weakly positive in SKOV3 cells ,the buffy grains was very thin;lovastatin strengthened the stain degree of PTEN in the SKOV3 cells,the stain and the buffy grains became thicker and the positive cell population was more than control group. FCM assay results:The expression of Survivin and PTEN protein had been examined in SKOV3 cells being treated with 5,10,20μmol/L lovastatin for 48h, the FI-value of Survivin in SKOV3 was 0.907,0.759,0.329, and the FI-value of PTEN was 1.186,1.347,1.491, respectively. The difference was statistically significant(p<0.05).3 Lovastatin combined with cisplatin could enhance proliferation inhibition of human varian cancer SKOV3 cells ,and lovastatin pretreatment was better than cisplatin.Conclusions:1 Lovastatin could have the effect of anti-varian cancer cell line SKOV3 in vitro in a dose-and time-dependent way.2 Lovastatin could induce an arrest of cell cycle in G0/G1 phase as well as apoptosis in SKOV3 cells.3 Within a certain drug concentration, Lovastatin could increase the expression of PTEN and decrease the expression of Survivin in SKOV3 cells. We infered that Lovastatin's effects of inhibiting proliferation and inducing apoptosis in SKOV3 cells seemed to due to up-regulation of the expression of PTEN protein and down-regulation of the expression of Survivin.4 Lovastatin combined with cisplatin could enhance proliferation inhibition of human varian cancer SKOV3 cells, and lovastatin pretreatment was better than cisplatin.