Perfluorooctanoic Acid Effects on Steroid Hormone and Growth Factor Levels Mediate Stimulation of Peripubertal Mammary Gland Development in C57Bl/6 Mice

Abstract

Perfluorooctanoic acid (PFOA) is a synthetic, widely used perfluorinated carboxylic acid and a persistent environmental pollutant. It is an agonist of peroxisome proliferator-activated receptor alpha (PPARalpha). Studies have shown that PFOA causes hepatocellular hypertrophy, tumorigenesis, and developmental toxicity in rodents, and some of its toxicity depends on the expression of PPARalpha. Our recent study revealed a stimulatory effect of peripubertal PFOA treatment (5 mg/kg) on mammary gland development in C57Bl/6 mice. The present study was designed to examine the underlying mechanism(s). It was found that mammary gland stimulation by PFOA was similarly observed in PPARalpha knockout and wild-type C57Bl/6 mice. The presence of ovaries was required for PFOA treatment (5 mg/kg) to stimulate mammary gland development with significant increases in the levels of enzymes involved in steroid hormone synthesis in both PFOA-treated wild-type and PPARalpha knockout mouse ovaries. PFOA treatment significantly increased serum progesterone (P) levels in ovary-intact mice and also enhanced mouse mammary gland responses to exogenous estradiol (E), P, and E + P. In addition, PFOA treatment resulted in elevated mammary gland levels of epidermal growth factor receptor (EGFR), estrogen receptor alpha, amphiregulin (Areg, a ligand of EGFR), hepatocyte growth factor, cyclin D1, and proliferating cell nuclear antigen (PCNA) in both wild-type and PPARalpha knockout mouse mammary glands. These results indicate that PFOA stimulates mammary gland development in C57Bl/6 mice by promoting steroid hormone production in ovaries and increasing the levels of a number of growth factors in mammary glands, which is independent of the expression of PPARalpha.

Progesterone Decreases Levels of the Adhesion Protein E-Cadherin and Promotes Invasiveness of Steroid Receptor Positive Breast Cancers

“…These replacement regimens produced systemic hormone levels within the physiological range (Online Resource 4). Plasma hormone levels were measured as previously described [34]. Normal mammary tissues and tumors were fixed in 10 % buffered formalin and paraffin embedded for immunohistochemical analysis or flash-frozen in liquid nitrogen and stored at −80°C for mRNA or protein analysis.…”

“…Immunoblot, Cell Cycle Analysis, and RT-PCR-Based Microarray Analysis Analyses were performed as previously described [34,39]. All rat and human primers for RT-PCR were from Qiagen (Frederick, MD).…”

Impact of diethylhexyl phthalate on gene expression and development of mammary glands of pregnant mouse

“…The fourth inguinal mammary glands of pregnant female mice (3.5 dpc) were isolated with the aid of a stereomicroscope (Nikon, SMZ1500, Japan) as described previously (Zhao et al 2010). The glands were spread onto porous specimen collection bags and then placed in vials containing RNAlater (RNA-EZ Reagents RNA-Be-Locker A, Sangon, B644171-0025, China).…”

Progesterone Stimulates Proliferation and Promotes Cytoplasmic Localization of the Cell Cycle Inhibitor p27 in Steroid Receptor Positive Breast Cancers

“…These replacement regimens produced systemic hormone levels within the physiological range (Online Resources 6). Plasma hormone levels were measured as previously described [41]. For proliferation studies, animals received i.p.…”

“…Immunoblot, Cell Cycle Analysis, and RT-PCR-Based Microarray Analysis Analyses (Rat Cancer Pathway Finder PCR Array, Qiagen, Frederick, MD) were performed as previously described [41,44]. Protein loading in immunoblots was normalized by intensity of β-actin bands.…”