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The Study on the Effect and Mechanism of Artesunate on the Radiosensitivity of HeLa and Siha Cells of Human Cervical Cancer

Author: JiRong
Tutor: CaoJianPing
School: Suzhou University
Course: Radiation Medicine
Keywords: Artesunate Radiosensitization Human cervical carcinoma cell Apoptosis Cell Cycle wee1 cyclinB1 cdc2
CLC: R737.33
Type: Master's thesis
Year: 2010
Downloads: 84
Quote: 1
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Objective To study the artesunate on p53 mutant human cervical carcinoma HeLa cells and p53 wild-type human cervical cancer cell radiosensitivity Siha, and by comparing artesunate with different p53 function in human cervical carcinoma cell radiosensitivity effect discrepancies further study the mechanism of radiosensitization. Methods HeLa cells and Siha as the experimental cells. (A) the use of tetrazolium salt colorimetric test (MTT) detection of artesunate on human cervical carcinoma HeLa and Siha cell survival fraction influence radiosensitivity experiments to determine the required concentration of the drug artesunate; (2) Application colony formation assay and flow cytometry to study artesunate on HeLa, Siha cells radiosensitivity; (3) the use of flow cytometry artesunate γ ray irradiation, HeLa and Siha cell cycle changes in the distribution situation; (4) application of Western-blot analysis artesunate γ ray irradiation, cell cycle regulatory protein wee1, cyclinB1, cdc2 expression level changes. Results (1) MTT test analysis showed: artesunate on human cervical cancer HeLa, Siha cells inhibition rate with the increase of drug concentration (P lt; 0.01). Identified sensitizer used in the experiments artesunate drug concentrations: HeLa was 2.0μmol / L, Siha is 2.5μmol / L, reaction time was 24 h, and duration of action in selected points lower drug concentration, artesunate itself Inhibition of cell is small (10%); (2) Experimental results show clonogenic: artesunate same concentration, the cell clone formation rate decreased with increasing radiation dose (P lt; 0.01). For HeLa cells, irradiation group and drug exposure group, the average lethal dose (D 0 ) values ??were 2.95Gy and 2.07Gy, quasi-threshold dose (D q ) values ??were as 2.01Gy and 1.24Gy, radiosensitization ratio (SER) was 1.43; For Siha cells, irradiation group and drug exposure group, the average lethal dose (D 0 ) values ??were 2.68Gy and 2.61Gy , quasi-threshold dose (D q ) values ??were 1.91Gy and 1.86Gy, radiosensitization ratio (SER) was 1.03; (3) Annexin Ⅴ / PI double staining apoptosis detection results showed that: For HeLa cells, pure 2,6 Gy irradiation group apoptosis rate was 12.26% and 40.08%, after artesunate treatment, the same dose of irradiation apoptosis rate increased to 22.71% and 59.92%, the difference was statistically significance (P lt; 0.01); For Siha cells, pure 2,6 Gy irradiation cell apoptosis rates were 12.27% and 32.25%, after artesunate treatment, the same radiation dose rate of apoptosis were 14.35% and 33.67%, the difference was not statistically significant (P gt; 0.05); (4) Flow cytometric analysis of cell cycle showed that: 2,6 Gy dose point, for HeLa cells irradiated with artesunate group and irradiation group Compared, G 0 / G 1 increase in the proportion of cells (P lt; 0.01), G 2 / M phase cells decreased (P lt; 0.01); For Siha cells irradiated group artesunate compared with radiation alone, G 0 / G 1 phase, G 2 / M phase cell levels did not change significantly (P gt; 0.05); (5) Western-blot analysis showed that: HeLa cells, simply by 2,6,12 G y γ-ray irradiation, cell cycle-related protein expression wee1 rising, and with the increase of radiation dose increased expression, cyclinB1 decreased protein expression, and the expression increased with the increase of radiation dose reduction is more obvious, cdc2 protein expression showed no significant change, Under the same dose, drug exposure group wee1 protein expression compared with radiation alone group decreased, cyclinB1 protein compared with radiation alone group increased, cdc2 protein did not change significantly; Siha cells in drug irradiated group wee1 protein, cyclinB1 protein, cdc2 protein expression compared irradiation group did not change significantly. Conclusion (1) of artesunate on human cervical carcinoma HeLa and Siha cells inhibition rate in a dose dependent manner within the study area; (2) of artesunate on the inhibition of HeLa and Siha cells different from the same drug concentration, the stronger inhibitory effect on HeLa cells in Siha cells; (3) of artesunate on p53 mutant human cervical carcinoma HeLa cells significantly radiosensitizing effect, while the p53 wild-type human cervical Siha cells without radiosensitizing effect . Artesunate on HeLa cells and radiosensitizing effect of reducing the radiation caused by the sublethal damage repair capacity of relevant; (4) Artesunate can increase p53 mutant HeLa cells were treated with γ-ray irradiation of cells apoptosis, whereas wild-type p53 Siha cells after irradiation had no effect on the rate of apoptosis; (5) Artesunate inhibits cells from ionizing radiation-induced activation of G 2 / M arrest, so that more cells with radiation injury due to insufficient time to repair, directly into mitosis, cell death occurs; (6) γ-ray irradiation, by increasing G 2 / M phase related to the expression levels of cyclin wee1 reduce cyclinB1 the expression level of p53 mutant induced HeLa cells G 2 / M phase arrest, artesunate combined radiation, can reverse This change makes wee1 expression was reduced, cyclinB1 expression increased, thereby weakening G 2 / M phase arrest effect, play radiosensitizing effect. Siha cells after irradiation occurs mainly G 1 / S phase arrest, so artesunate after irradiation, G 2 / M phase-associated protein expression was not significantly change ; (7) HeLa and Siha cells were treated with artesunate γ-ray irradiation, cdc2 protein expression were no significant changes, which indicated that artesunate is wee1 indirect effects on cdc2 to influence the G 2 / M phase arrest.

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CLC: > Medicine, health > Oncology > Genitourinary tumors > Female genital tumors > Uterine tumors
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