| Ovarian cancer is one of three malignant tumors in female reproductive system. The long-term survival has remained to be 25% to 30%. Owing to high frenquency of abdominal cavity metastasis in ovarian cancer, the life span and quality of survival of patients have been affected directly, therefore, it is very important for clinic to find an effective intraabdominal chemotherapy method. In the beginning of 1980's, studies in China have shown that As2O3 can induce a high rate of clinical remission in patient with relapsed acute promyelocytic leukemia and it becomes hot spot for antitumor effect of As2O3 in the world. Although the mechanism of antileukemic effect of As2O3 is well understood, there has been few reports regarding the effect of As2O3 in solid tumors, especially in ovarian cancer. In this study, light and electron microscopy, MTT, flow cytometry, immunohistochemistry and RT-PCR were used to explore the molecularmechanisms of As2O3 on the growth and induction of apoptosis of human ovarian cancer cell line. The aim was to provide experimental data for therapy of solid tumor and clinical application of As2O3. The main results are as follows: 1. The growth of SKOV3 and 3AO cells were inhibited with treatment of 2-20umol/L As2O3 and demonstrated a dose-and time-dependent growth inhibition. The growth inhibition of 3AO cells with As2O3 were more weaker than that of SKOV3 cells. The apoptotic morphological changes were oberserved in SKOV3 and 3AO cells incubated for 72 hours with light and electron microscopy. Flow cytometry analysis has shown that 2um As2O3 induced sub-G1 peak , a S+G2/M phase arrest in SKVO3 cells following 3 days exposure, while low concentration As2O3 (2, 5uM) induced G2/M phase arrest without sub-G1 peak in 3AO cells following 3 days exposure, this results suggested that As2O3 inhibited the cellular proliferation of SKOV3 and 3AO cells via arrest of cell cycle, low concentration As2O3 inhibited the cellular proliferation of 3AO cells via G2/M phase arrest of cell cycle without inducing apoptosis, while high concentration As2O3 induced apoptosis of 3AO cells, it suggested that it has differences for As2O3 in inducing apoptisis of ovarian cancer cell lines in dose of drug, type of cell line and the mechanism of apoptosis. 2. In this study mitochondrial transmembrane potentials (Δψm) was detected by the double staining of propidium iodide (PI) and Rhodamine123(Rh123) and the results have shown that As2O3 induced the loss of Δψm in SKOV3 and 3AO cells, and the Δψm was disrupted significantly accompanied the increase of concentration of As2O3. The changes of fluorescence intensity by fluorescent microscopy have decreased in SKOV3 and 3AO cells with As2O3 for 72 hours. The ultrastructural alteration of mitochondria was found as following: at 72h mitochodrial swelling lead to balloon-like apperance and its outer membrane disrupted. This results suggested the disruption of Δψm is the key event of As2O3-induced apoptosis. Also down regulated expression of bcl-2 was found in SKOV3 and 3AO cells with As2O3 by immunohistochemistry and immunoflow cytometry. Since down-regulating expression of Bcl-2 is closely related to reative oxygen species in apoptosis, it suggested deregulation of the expression of Bcl-2 may release the antioxidant function of Bcl-2, increase the amount of reactive oxygen species, injurn the Δψm directly, at last cause cell apoptosis. 3. In this study, we explored the relationship of signal transduction between cPKCα and Ras with As2O3 inducing apoptosis of SKOV3,3AO cells. Results showed that As2O3 inhibited H-Ras gene expression in SKOV3 cells at 72h, while in 3AO cells As2O3 inhibited H-Ras gene expression weaker at 12h, 24h, 72h; The protein expression of cPKCα was increased significantly and was activated continuously by As2O3; In SKOV3 and 3AO cells the mRNA expression of c-Raf was increased signifcantly with As2O3for 24h, and the mRNA expression of c-jun wa...
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