Breast cancer (BrCa) as one of the major malignancies threatening women's health worldwide occurs due to the genetic and environmental interactions. Epidemiological studies have suggested that exposure to endocrine disrupting chemicals (EDCs) can elevate the risk of breast cancer. Di-(2-ethylhexyl)-phthalate (DEHP) and bisphenol A (BPA) are known as two typical EDCs. Although several studies have implied that there appear to have adverse effects of exposure to BPA or DEHP alone on breast development, no study to date has demonstrated the exact toxic effect of combined exposure to DEHP and BPA on breast tumorigenesis. In the present study, we performed an in vivo experiment including 160 female Sprague-Dawley (SD) rats, in which 80 rats were randomly allocated to 4 groups including control group given to normal diet, DEHP (150 mg/kg body weight/day), BPA (20 mg/kg body weight/day), and DEHP (150 mg/kg body weight/day) combined with BPA (20 mg/kg body weight/day) by gavage for 30 weeks. Additionally, a DEN/MNU/DHPN (DMD)-induced carcinogenesis animal model was also established to assess their effect on tumor promotion. Namely, the other 80 SD rats were separated into another 4 groups: in addition to DMD initiation each group treated with vehicle, DEHP, BPA and the combination of BPA and DEHP respectively. Our data demonstrated that BPA alone or in combination with DEHP may induce hyperplasia of mammary glands, including the proliferation of ductal epithelial cells and an increase in the number of lobules and acinus after a 30-week exposure. Notably, co-exposure to DEHP and BPA increased the incidence and reduced the latency of mammary tumor, which seemed to enhance the susceptibility of carcinogens-induced tumor. Mechanistically, our results supported the hypothesis that exposure to BPA and DEHP might promote breast cancer dependent on Esr1 and HDAC6 as pivotal factors, and further lead to the activation of oncogene c-Myc. Our study suggested that BPA combined with DEHP facilitate the occurrence of mammary tumors, which contributed to advance our understanding in the complex effects of compound exposure to endocrine disrupting chemicals.

Bisphenol A (BPA) is a widespread endocrine disruptor that induces the impairment of immune cells, but the mechanism remains unknown. Macrophages are one of the most important immune cells in innate and adaptive immunity. In this study, we aimed to probe the effects of BPA on the damage of RAW264.7 cells and its mechanisms of action, especially focusing on the relationship between autophagy and apoptosis. Cells were pretreated with 10 mg/L LPS, or added autophagy activator RAPA, autophagy inhibitor 3-MA or Bcl-2 inhibitor ABT-737, then treated with BPA (0, 10, 100 and 200 μmol/L) for 12 h. Results have shown that BPA decreased the cell viability and disrupted secretory function by promoting pro-inflammatory cytokines TNF-α and IL-6 and reducing anti-inflammatory cytokines IL-10 TGF-β, as well as phagocytic ability. Moreover, autophagy was inhibited by BPA through decreasing p-AMPK/AMPK and increasing p-mTOR/mTOR, and further down-regulating autophagy proteins ATG6, LC3II/I ratio, and up-regulating autophagy flux protein p62. Additionally, BPA significantly increased Bax/Bcl-2 ratio, Caspase-3 expression and apoptosis rate. We found that RAPA ameliorated the cell viability, Bax/Bcl-2 ratio, and macrophage function damage induced by BPA. Intriguingly, ABT-737 might promote ATG6 expression. In summary, our study demonstrated that the effects of BPA on macrophages seemed to be mediated by inhibiting AMPK/mTOR-dependent autophagy and inducing apoptosis via endogenous mitochondrial pathway. Both Bcl-2 and ATG6 were involved in the regulation of apoptosis and autophagy by BPA. These findings provide a broader perspective for understanding the interaction between autophagy and apoptosis in BPA-induced immune cell injury.