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Effect of Culture Supernatant of Toxoplasma Gondii on Human Ovarian Cancer Cell Line KOV-3

Author: GeZuo
Tutor: YeBin
School: Chongqing Medical University
Course: Pathogen Biology
Keywords: Toxoplasma gondii supernatant human ovarian cancer cell line SKOV-3 proliferation apoptosis
CLC: R737.31
Type: Master's thesis
Year: 2010
Downloads: 27
Quote: 1
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Abstract

Background and Objective: Ovarian cancer remains the most lethal gynaecological cancer. Ovarian cancer causes non-specific symptoms. Early diagnosis would result in better survival. However, lack of symptoms or vague symptoms in the early stages of ovarian cancer is why the illness is known as the ’silent killer’. The crude death rate of ovarian cancer was 4.8 per 100 000 female population. The treatment methods of this cancer including surgery,chemotherapy, Radiation therapy.But all the methods have side effects, long-term use of patients with immune system and high-dose hematopoietic system, cause a lot of damage.The medicine with less side effects and high selectivity seems to be could improve the clinical efficacy.Toxoplasma gondii is a species of parasitic protozoa in the genus.The focus of recent reserch is on the factors of invasion of host cell by Toxoplasma and virulence. Now, it is proved that Toxoplasma gondii can inhibite host cell proliferation, affect the cell apoptosis and cell cycle.In this study, we try to investigate the inhibitive effects of culture supernatants of T. gondii as a biological agent on the proliferation, apoptosis and cell cycle of human ovarian cancer cell line SKOV-3.METHODS:1. Growth inhibition rates of treated groups were measured with the MTT method.2. FCM was used to test the rates of apoptosis of treated groups by culture supernatants of T. gondii with the fluorescent dye of PI.3. FCM was used to test Cell cycle of treated groups by culture supernatants of T. gondii with the fluorescent dye of Annexin-v-FITC/PI.4. After the treatment of SKOV-3 cells by culture supernatants of T. gondii, morphological change of apoptotic cells was investigated by Annexin-v-FITC/PI fluorescent staining under fluorescent microscope.5. The agarose electrophoresis was used to detect the DNA changes of treated groups.6.Western blot was used to detect the levels of cyclinB1、cdc2、bcl-2 and bax of different groups.RESULTS:1. The culture supernatants of T. gondii could inhibit the proliferation of SKOV-3 cells in a time-dose dependent manner.2. In the flow cytometric analysis, cell cycle was significantly stopped at G2/M phase by the culture supernatants and a maximum apoptosis rate of 18.23% was detected at 48h of treatment with the supernatants, but the apoptosis rates decreased and dead rate increased at the treatment time beyond 48h.3. The classic cellular morphology of apoptosis in the treated trophoblastic cells was observed under the fluorescence microscope.4. The culture supernatant of T. gondii reduced the expressions of gene cyclinB1, cdc2 and bcl-2 of SKOV-3 cells and increased the expressions of gene bax. After 48 hours, contrast the levels of cyclinB1 in negative control group. The integrated density Value of the culture supernatants of T. gondii groups were 0.86+0.04, 0.73+0.06 and 0.60+0.05 respectively, the results of cdc2 proteins were 0.98+0.02, 0.84+0.05 and 0.65+0.06 and the results of cyclinB1 proteins were 0.97+0.05, 0.81+0.02 and 0.78+0.05, nd the results of cdc2 were 1.23+0.03, 1.54+0.04 and 1.75+0.02 respectively. The difference was statistically significant when compared with the negative control group (p<0.05).CONCLUSION:The culture supernatant of T. gondii may inhibit SKOV-3 cells and arrest the cell cycle mainly by regulating the expression of gene cyclinB1 and cdc2 ,While can promote apoptosis of SKOV-3 cells in vitro through up-regulating Bax expressions and down-regulating Bcl-2 expressions.

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