Radio-sensitivity enhancement effect of piplartine on human breast adenoma MDA-MB-231 cells line in vitro

Objective: To investigate the radio-sensitivity enhancement effect of piplartine on human breast cancer MDA-MB-231 cells line and to discuss the possible mechanism of radio-sensitivity enhancement effect. Methods: Human breast cancer cell line MDA-MB-231 was cultured in vitro and the cells in logarithmic growth phase were selected for experiments. MDA-MB-231 cells were divided into four groups: control, X-ray expose, piplartine, and piplartine combined with X-rays. The clonogenic assay was performed to determine the radio-sensitivity enhancement effect of piplartine on MDA-MB-231 cells. The cell survival curve was fitted by single-hit multi-target model and then the survival fraction (SF), average lethal dose (D₀), quasi-threshold dose (D_q) and sensitizing enhance rate (SER) were calculated. Cell apoptosis was analyzed by flow cytometry (FCM). Western blot assay was employed to measure the expression of apoptosis-related protein (Bc1-2 and Bax proteins) after treatment with piplartine and/or X-ray radiation. The intracellular reactive oxygen species (ROS) level was detected by FCM with the probe of DCFH-DA. Results: The cloning formation capacity was decreased in the group of piplartine plus radiation, which showed that the values of SF₂, D₀, D_q were significantly lower than those of radiation alone group(P < 0.05), and the sensitizing enhancement rate (SER) of D₀ was 1.22 and 1.29. The cell apoptosis rate was increased by the combination treatment of piplartine and radiation. Piplartine increased the radiation-induced intracellular levels of ROS. Compared with the control group and single piplartine or radiation group, the combination group decreased the expression of Bcl-2 significantly, while the expression of Bax was increased. Conclusion: Piplartine at non-cytotoxic concentration can enhance the radio-sensitivity of MDA-MB-231 cells, which may be related to its regulation of apoptosis-related protein expression and the increase of intracellular ROS level, thus increasing the radiation-induced apoptosis. (authors)