2nd International Conference on Biomedicine and Pharmaceutics (ICBP) 2014

EFFECTS OF RADIATION ON APOPTOSIS, ROS AND CELL CYCLE IN BREAST CANCER STME CELLS Li Yan, WANG Zhi-cheng, Qi Yali*. Department of Epidemiology, School of Public Health, Beihua University, Jilin, China. Key Laboratory of Radiobiology, Ministry of Health, School of Public Health, Jilin University, Changchun, China. *Department of Epidemiology, School of Public Health, Beihua University, Jilin, China. 95614837@qq.com; 20496586@qq.com. *Corresponding author: E-mail:lhbqyl26@sina.com Introduction: Cancer stem cells occupy a small portion of whole cancer cells, it has stem cell characteristics, and can immeasurably proliferate, differentiate, and initiate tumorigenesis. In addition, it can cause tumor recurrence, metastasis, and radiotherapy and chemotherapy resistance, its basic mechanism involves some regulation pathways of DNA damage and repair, cell cycle disturbance, autophagy, apoptosis, and cell hypoxia (1-6). Zielske and his colleagues found that after breast cancer radiotherapy, CD44 CD24 lin phenotype breast stem cells enrich in tumor tissue sample of breast cancer patients, and can cause radioresistance (7). Philips and his colleagues found that oxygen-free radical in breast cancer stem cells was lower than that in breast cancer cells, and 10 Gy irradiation could increase cancer cell oxygen-free radical level, but had no obvious change in cancer stem cells, it indicated that ionizing radiation had more effects on cancer cells than cancer stem cells (8). In order to explore the exact mechanism of radioresistance, in our study, changes of apoptosis, reactive oxygen species (ROS) level, and cell cycle were measured in breast cancer cells and breast cancer stem cells, and provided a new perspective and direction for breast cancer radiotherapy. Methods: Breast cancer stem cells (MCF-7-S) were sorted by flow cytometry (FCM) with CD44 and CD24 antibodies in breast cancer MCF-7 cells. MCF-7 and MCF-7-S were irradiated with 8 Gy X-rays, after irradiation 4 to 24 hours, apoptotic rate and ROS level were measured by FCM with Annexin V/PI and 2¶,7¶-dichlorodihydrofluorescin diacetate (DCFH-DA) staining, respectively. After irradiation for 12 hours, cell percentage of each phage was measured by FCM with PI staining. Statistical evaluations are presented as meanTSD. Data were analyzed using the Student t test for statistical significance using SPSS v17.0 software. P values were considered significant if P was less than 0.05. Results: CD44CD24 phenotype breast stem cells were sorted successfully. With time prolongation, apoptotic rate and ROS level in MCF-7 and MCF-7-S showed increasing tendency and reached for maximum value at 12 hours and 24 hours. ROS level in MCF-7-S was significantly lower than that in MCF-7 at 4, 8,12, and 24 hours (P G 0.05 or P G 0.001), and apoptotic rate in MCF-7-S was significantly higher than those in MCF-7 cells only at 8 hours (P G 0.05). After 8 Gy irradiation, ROS level (4, 8,12, and 24 hours) and apoptotic rate (12 hours) significantly increased in MCF-7 (P G 0.05), and apoptotic rate (4, 8,12, and 24 hours) significantly decreased in MCF-7-S (P G 0.05 or P G 0.001), but had no obvious effects on ROS level in MCF-7-S. At 12 hours, as compared with MCF-7 cells, cell percentage of G0-G1 phage and G2-M phage in MCF-7-S significantly decreased, and cell percentage in S phage significantly increased (P G 0.05 ); and after 8 Gy irradiation, cell percentage of G2-M phage in MCF-7 significantly increased (P G 0.05 ), but it had no obvious changes in MCF-7-S. Conclusion: First, CD44CD24 phenotype breast stem cells were obtained successfully by FCM. Second, ionizing irradiation caused apoptosis and ROS level increasing, and G2-M phage arrest in breast cancer cells, but had no obvious effects on breast cancer stem cells, even caused apoptosis to decrease, it indicated that radioresistance of breast cancer might be related to apoptosis, ROS level, and G2-M phage arrest. Acknowledgment: This research was supported by Jilin province education department science and technology research projects(2014-192), the National Natural Science Foundation of China (30970681), Young Scholars Program of Norman Bethune Health Science Center of Jilin University (2013202017), Jilin University innovative training program (2013A72261).