Regulation of the cell cycle and P13K/AKT/mTOR signaling pathway by phthalates in normal human breast cells

Abstract

ObjectiveTo investigate the impact of phthalates, including Butyl benzyl phthalate (BBP), di(n-butyl) phthalate (DBP), and di(2-ethylhexyl) phthalate (DEHP), in breast carcinogenesis. Materials and methodsMCF-10A normal breast cells were treated with phthalates (100 nM) and 17β-estradiol (E2, 10 nM), which were co-cultured with fibroblasts from normal mammary tissue adjacent to estrogen receptor positive primary breast cancers. Cell viability was determined using a 3-(4,5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Cell cycles were analyzed using flow cytometry. The proteins involving cell cycles and P13K/AKT/mTOR signaling pathway were then evaluated by Western blot analysis. ResultsMCF-10A co-cultured cells treated with E2, BBP, DBP, and DEHP exhibited a significant increase in cell viability using MTT assay. The expressions of P13K, p-AKT, and p-mTOR, as well as PDK1 expression, were significantly higher in MCF-10A cells treated with E2 and phthalates. E2, BBP, DBP, and DEHP significantly increased cell percentages in the S and G2/M phases. The significantly higher expression of cyclin D/CDK4, cyclin E/CDK2, cyclin A/CDK2, cyclin A/CDK1, and cyclin B/CDK1 in MCF-10A co-cultured cells were induced by E2 and these three phthalates. ConclusionThese results provide consistent data regarding the potential role of phthalates exposure in the stimulating proliferation of normal breast cells, enhancing cell viability, and driving P13K/AKT/mTOR signaling pathway and cell cycle progression. These findings strongly support the hypothesis that phthalates may play a crucial role in breast tumorigenesis.