Date Published: April 4, 2018
Publisher: Public Library of Science
Author(s): Qi Han, Rui Chen, Fangjie Wang, Sha Chen, Xiongshan Sun, Xiao Guan, Yao Yang, Bingjie Peng, Xiaodong Pan, Jinfang Li, Weijing Yi, Peng Li, Hongwei Zhang, Dongfang Feng, An Chen, Xiaohui Li, Shuhui Li, Zuoming Yin, Maria Rosaria Scarfi.
Resistance to 5-fluorouracil (5-FU) and its induced immune suppression have prevented its extensive application in the clinical treatment of breast cancer. In this study, the combined effect of 50 Hz-EMFs and 5-FU in the treatment of breast cancer was explored. MCF-7 and MCF10A cells were pre-exposed to 50 Hz-EMFs for 0, 2, 4, 8 and 12 h and then treated with different concentrations of 5-FU for 24 h; cell viability was analyzed by MTT assay and flow cytometry. After pre-exposure to 50 Hz-EMFs for 12 h, apoptosis and cell cycle distribution in MCF-7 and MCF10A cells were detected via flow cytometry and DNA synthesis was measured by EdU incorporation assay. Apoptosis-related and cell cycle-related gene and protein expression levels were monitored by qPCR and western blotting. Pre-exposure to 50 Hz-EMFs for 12 h enhanced the antiproliferative effect of 5-FU in breast cancer cell line MCF-7 in a dose-dependent manner but not in normal human breast epithelial cell line MCF10A. Exposure to 50 Hz-EMFs had no effect on apoptosis and P53 expression of MCF-7 and MCF10A cells, whereas it promoted DNA synthesis, induced entry of MCF-7 cells into the S phase of cell cycle, and upregulated the expression levels of cell cycle-related proteins Cyclin D1 and Cyclin E. Considering the pharmacological mechanisms of 5-FU in specifically disrupting DNA synthesis, this enhanced inhibitory effect might have resulted from the specific sensitivity of MCF7 cells in active S phase to 5-FU. Our findings demonstrate the enhanced cytotoxic activity of 5-FU on MCF7 cells through promoting entry into the S phase of the cell cycle via exposure to 50 Hz-EMFs, which provides a novel method of cancer treatment based on the combinatorial use of 50 Hz-EMFs and chemotherapy.
Breast cancer is a deadly disease due to immense difficulties in prevention and treatment. Multidrug resistance of tumor cells is the main reason for the failure of anticancer drugs. Finding novel therapeutic strategies is therefore of great significance in the treatment of highly malignant breast cancer.
The combination of different antitumor treatment strategies has greatly improved therapeutic efficacy in clinics. For example, combined chemotherapy can not only improve the therapeutic outcome by overcoming multidrug resistance and disrupting multiple cell survival pathways but also improve patient compliance due to reduced dosage of each agent[20–22]. However, the potential effects of combination of electromagnetic fields (EMFs) and chemotherapeutic drugs are usually controversial. It has been shown that stimulation with pulsing electromagnetic fields can enhance the antiproliferative effect of doxorubicin on mouse osteosarcoma cells , while other report demonstrated that pre-exposure to electromagnetic fields appear to protect HL-60 cells from the toxic effects of subsequent treatment with doxorubicin . First, the inconsistent effects can be explained by the heterogeneity of various cancer cells. On the other hand, the exposure conditions, such as magnetic intensity, time and frequency, are the key factors influencing the biological effects of EMFs on cancer cells. In this study, we checked the antitumor effect of 5-FU on MCF7 cells with different 50 Hz-EMF pre-exposure durations. We found that the cytotoxic effect of 5-FU was significantly increased by a 12 h pre-exposure time. However, this pre-exposure condition did not enhance the effect of 5-FU on the normal breast epithelial cell line MCF10A. The potential effect of exposure to ELF EMF for human health have been investigated for many years. Several studies showed that ELF EMF can increase incidence of certain types of cancer[23–24]. While some available evidence showed ELF EMF has no effect on cell cycle distribution and apoptosis and cell growth. Furthermore, the anticarcinogenic ability of low-energy electromagnetic waves has also been demonstrated in several in vitro studies[11,27–31]. Some studies verified the anticancer effect of EMFs in vivo[32–34] or even in patients. These conflicting data might be also due to the differences in frequency, intensity, duration and cell types. However, the effect of 50 Hz-EMF exposure on the physiology of breast cancer MCF7 cells had not been reported. Initially, we also thought that exposure to 50 Hz-EMFs may enhance the antiproliferative effect of 5-FU by inducing cancer cell apoptosis. Surprisingly, we found that 50 Hz-EMF exposure for 12 h had no effect on cell apoptosis but could promote the entry of MCF7 cells into S phase and increase DNA synthesis, thus rendering MCF7 cells in the active phase of S phase more sensitive to the cell cycle-specific drug 5-FU. The mechanism of cytotoxicity of 5-FU is specifically inhibiting DNA synthesis, and 50 Hz-EMF exposure promotes DNA synthesis of MCF7 cells. In our study, 5-FU and ELF-EMF acts in a synergistic manner, this needs to find the right balance point between the ELF-EMF exposure condition and 5-FU concentration. If we use ELF-EMF to induce breast cancer cells into S period, and 5-FU concentration is strong enough to kill cancer cells, the combination of two component would achieve better antiproliferative effects. Crocetti et al. reported low intensity and frequency of pulsed electromagnetic fields selectively impair viability of breast cancer cells, and this difference may result from the different intensity and exposure time used in our study.