Evaluation of triclosan in the Hershberger and H295R steroidogenesis assays

Abstract

Triclosan (TCS) is an antibacterial widely used in personal care products that exhibits endocrine disrupting activity in several species, with reports of altered thyroid, estrogen and androgen signaling pathways. To evaluate the androgenic mode of action, TCS was evaluated for androgen receptor mediated effects in the Hershberger assay and for altered androgen synthesis in the H295R steroidogenesis assay. In the Hershberger assay, castrated males were dosed by oral gavage for 10 days with corn oil (vehicle) or TCS (50 or 200 mg/kg/day) in the presence or absence of testosterone proprionate (TP, 0.2 mg/kg/day) prior to assessing accessory sex tissues (ASTs) weights. TCS alone or in combination with TP did not alter androgen dependent AST weights. Assessment of serum thyroxine (T4) demonstrated a significant dose-dependent decrease by TCS (50 or 200 mg/kg/day) co-administered with TP and TCS (200 mg/kg) without TP, but no differences in liver or thyroid weights. In the H295R assay, TCS from 0.01 to 10 μM had no effect on testosterone production but TCS at 3 μM and above did induce a significant increase in estrogen production. At 10 μM, TCS produced significant cytotoxicity which confounded the interpretation of the estrogenic effect at that concentration. Thus, TCS had no effect on androgen synthesis or activity in the models used, but did enhance estrogen production and suppress serum T4.

Keywords: Endocrine disruptor; H295R; Hershberger; Steroidogenesis; Triclosan.

Fig. 1.

Total serum T4 (μg/dl) concentrations in castrated male rats treated with TCS in the presence or absence of testosterone proprionate (TP; 0.2 mg/kg). Bars represent mean ± SEM. *P < 0.01 as compared to corresponding control (white or black bar).

Fig. 2.

Effect of Triclosan on production of estradiol in H295R cells in vitro. Black bar indicates cells treated with solvent control only (DMSO 0.1%). Forskolin (FKN), positive control for induced estradiol production. Prochloraz (PCA), positive control for inhibition of steroid production. Gray bars indicate cells treated with Triclosan in μM concentration. *P < 0.05 compared to solvent control.

Fig. 3.

Effect of Triclosan on production of testosterone in H295R cells in vitro. Black Bar indicates cells treated with only DMSO 0.1%. Forskolin (FKN), positive control for induced testosterone production. Prochloraz (PCZ), positive control for inhibition of T production. Gray bars indicate cells treated with Triclosan in μM concentration. *P < 0.05 compared to solvent control.

Fig. 4.

Cell viability of H295R cells after 48 h exposure to TCS from 0.01 to 30 μM (n=6). * P < 0.05 as compared to control (DMSO 0.1%).

Fig. 5.

The effect of Triclosan on human recombinant aromatase activity. White bar indicates control (Aromatase alone) (n=4), Black bar indicates Aromatase plus 5 μM Letrozole (n=3). Gray bars indicate Aromatase plus Triclosan at 1, 3, 10, or 30 μM (n=4, 4, 5, or 5 respectively). * P < 0.05 as compared to control.