Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin, 12-O-tetradecanoylphorbol-13-acetate and 17β-estradiol on estrogen receptor regulation in MCF-7 human breast cancer cells

Abstract

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) exhibits remarkably potent antiestrogenic activity. To further elucidate the role of estrogen receptor (ER) regulation in this response, we examined the effects of exposure to TCDD in MCF-7 human breast cancer cells on ER mRNA levels by using an RNase protection assay, on ER accumulation by using an ER immunocytochemical essay (ER-ICA), and on ER function by competitive binding assays under conditions of saturating 17 beta-estradiol (E2). Comparative studies were conducted with E2 and 12-O-tetradecanoylphorbol-13-acetate (TPA), as both compounds are known to suppress ER expression. Our results indicate that 1 nM E2 and 100 nM TPA both suppress ER mRNA levels as early as 4 h after exposure and to 33.6% and 16.5% of control levels, respectively, after 72 h. In contrast, no significant effect on ER mRNA levels was attributed to exposure to 10 nM TCDD. A greater than 50% reduction in positive staining was observed by ER-ICA after 72 h exposure to 1 nM E2 and to 100 nM TPA, while only an 11% reduction in positive staining was observed with 10 nM TCDD. Specific binding of [3H]E2 under saturating conditions (10 nM E2) in whole cells was reduced by 50% in cultures exposed to 100 nM TPA, although no effect on binding was observed with exposure to 10 nM TCDD. In contrast, specific binding using subsaturating 1 nM [3H]E2 was depressed by 49% in MCF-7 cells exposed to 10 nM TCDD for 72 h. This depression was inhibited by a 1-h treatment with 5 microM alpha-naphthoflavone, which inhibits TCDD-induced, P450-mediated, E2 metabolism, and subsequent E2 depletion. In conclusion, while TPA and E2 effectively down-regulate ER expression, TCDD, under antiestrogenic conditions, has little if any effect on total ER levels in MCF-7 cells, and thus ER modulation is probably not necessary for the suppression of estrogenic activity in MCF-7 cells by TCDD.

Antiestrogenicity of Clarified Slurry Oil and Two Crude Oils in a Human Breast-Cancer Cell Assay

“…The underlying details for the increase in metabolism have been well documented previously (Gierthy et al, 1996;Pang et al, 1999;Spink et al, 1992Spink et al, , 1994. Briefly, it is assumed that compounds within the oil samples, particularly the PACs, bind the aryl hydrocarbon receptor (AhR), inducing cytochrome P-450 (CYP450) enzymes that catalyze the metabolism of E 2 .…”

Diosgenin Induces Hypoxia-Inducible Factor-1 Activation and Angiogenesis through Estrogen Receptor-Related Phosphatidylinositol 3-kinase/Akt and p38 Mitogen-Activated Protein Kinase Pathways in Osteoblasts

“…The whole-cell estrogen receptor binding assay was carried out as described previously (Gierthy et al, 1996). MC3T3-E1 cells were suspended in serum-free medium and seeded into 24-well plates at a density of 2 ϫ 10 5 cells per well.…”

Induction of CYP1A1 and CYP1B1 in T-47D Human Breast Cancer Cells by Benzo[a]pyrene Is Diminished by Arsenite

“…T-47D cells were obtained from the American Type Culture Collection (Rockville, MD) and were maintained in a 37°C incubator with a humidified atmosphere containing 5% CO 2 . For experimental protocols, the medium was DC 5 (Gierthy et al, 1996) consisting of Dulbecco's modified Eagle's medium, without phenol red, and with 5% bovine calf serum (Cosmic calf serum; Hyclone Laboratories, Logan, UT), 10 mM nonessential amino acids, 2 mM L-glutamine, and 10 g/l insulin. BAP was added to the cultures alone or in combination with varying concentrations of NaAsO 2 .…”