Prepubertal exposure to elevated manganese results in estradiol regulated mammary gland ductal differentiation and hyperplasia in female rats

Evidence suggests that environmental substances regulating estrogenic pathways during puberty may be detrimental to the developing mammary gland (MG). Manganese (Mn) is a trace mineral required for normal physiological processes. Prepubertal exposure to Mn induces precocious puberty in rats, an event associated with early elevations in puberty-related hormones, including estradiol (E₂). However, until now the effect of Mn-induced precocious MG development has not been determined. Therefore, we assessed the ability of prepubertal Mn exposure to advance normal MG development and alter E₂ driven pathways involved in tumorigenesis. Sprague Dawley female rats were gavaged daily with either 10 mg/kg manganese chloride (MnCl₂) or saline (control) from postnatal day (PND) 12 through PND 30. Blood and MGs were collected on PNDs 30 and 120. Compared to controls, serum E₂ levels on PND 30 were elevated (p < 0.05) in the Mn-treated group. Mn exposure significantly increased differentiated MG terminal ductal structures and the percentage of MG epithelial cells that stained positive for the proliferative marker, Ki67, at PND 30 (p < 0.001) and PND 120 (p < 0.001). Levels of Mn (ppm) were not elevated in these MGs. Mn-treated animals (40%) exhibited reactive stroma and intra-luminal focal hyperplasia in hemotoxylin and eosin stained MGs at PND 120. Furthermore, Mn exposure resulted in elevated protein expression levels of estrogen receptor α, activator protein 2α, phosphorylated (p)-Akt, and p53 in MGs on PND 120, but not on PND 30. Collectively, these data show that exposure to a supplemental dose of Mn causes accelerated pubertal MG growth which can progress to adult hyperplasia; thus, providing evidence that early life Mn exposure may increase susceptibility to breast cancer.