Lack of significant estrogenic or antiestrogenic activity of pyrethroid insecticides in three in vitro assays based on classic estrogen receptor α-mediated mechanisms

Abstract

Estrogenic and antiestrogenic activity of pyrethroid insecticides (d-trans-allethrin, cypermethrin, empenthrin, fenvalerate, imiprothrin, permethrin, d-phenothrin and prallethrin) was evaluated using a suite of three in vitro assays based on classic human estrogen receptor α (hERα)-mediated mechanisms. A mammalian cell-based luciferase reporter gene assay was developed for examining effects on hERα-mediated gene activation. hERα-independent effects on the gene activation were examined using control cells with constitutive luciferase activation by a herpes simplex virus thymidine kinase (HSV-TK) promoter for determining appropriate dose levels of test chemicals. Moreover, the test chemical-dependent interaction between hERα and a coactivator (transcriptional intermediary factor 2: TIF2) was analyzed by a yeast two-hybrid method, competitive binding to hERα being assayed by a fluorescence polarization method. Significant (p < 0.05) positive effects of estrogenic substances (E2/estradiol, diethylstilbestrol, and p-nonylphenol) were detected in all assays. An antiestrogen, 4-hydroxytamoxifen, significantly inhibited E2-mediated transactivation and interaction between hERα and TIF2 through hERα binding (p < 0.05). However, none of the pyrethroids tested showed significant (p < 0.05) estrogenic or antiestrogenic effects (100 nM-10 μM), indicating that they do not impact on the classic hERα-mediated activation pathway in vitro.