Alkylphenol Xenoestrogens with Varying Carbon Chain Lengths Differentially and Potently Activate Signaling and Functional Responses in GH3/B6/F10 Somatomammotropes

BACKGROUND Alkylphenols varying in their side-chain lengths [ethyl-, propyl-, octyl-, and nonylphenol (EP, PP, OP, and NP, respectively)] and bisphenol A (BPA) represent a large group of structurally related xenoestrogens that have endocrine-disruptive effects. Their rapid nongenomic effects that depend on structure for cell signaling and resulting functions are unknown. OBJECTIVES We compared nongenomic estrogenic activities of alkylphenols with BPA and 17beta-estradiol (E(2)) in membrane estrogen receptor-alpha-enriched GH3/B6/F10 pituitary tumor cells. These actions included calcium (Ca) signaling, prolactin (PRL) release, extracellular-regulated kinase (ERK) phosphorylation, and cell proliferation. METHODS We imaged Ca using fura-2, measured PRL release via radioimmunoassay, detected ERK phosphorylation by fixed cell immunoassay, and estimated cell number using the crystal violet assay. RESULTS All compounds caused increases in Ca oscillation frequency and intracellular Ca volume at 100 fM to 1 nM concentrations, although long-chain alkylphenols were most effective. All estrogens caused rapid PRL release at concentrations as low as 1 fM to 10 pM; the potency of EP, PP, and NP exceeded that of E(2). All compounds at 1 nM produced similar increases in ERK phosphorylation, causing rapid peaks at 2.5-5 min, followed by inactivation and additional 60-min peaks (except for BPA). Dose-response patterns of ERK activation at 5 min were similar for E2, BPA, and PP, whereas EP caused larger effects. Only E2 and NP increased cell number. Some rapid estrogenic responses showed correlations with the hydrophobicity of estrogenic molecules; the more hydrophobic OP and NP were superior at Ca and cell proliferation responses, whereas the less hydrophobic EP and PP were better at ERK activations. CONCLUSIONS Alkylphenols are potent estrogens in evoking these nongenomic responses contributing to complex functions; their hydrophobicity can largely predict these behaviors.