Iodide excess regulates its own efflux: a possible involvement of pendrin

Abstract

Adequate iodide supply and metabolism are essential for thyroid hormones synthesis. In thyrocytes, iodide uptake is mediated by the sodium-iodide symporter, but several proteins appear to be involved in iodide efflux. Previous studies demonstrated that pendrin is able to mediate apical efflux of iodide in thyrocytes. Acute iodide excess transiently impairs thyroid hormone synthesis, a phenomenon known as the Wolff-Chaikoff effect. Although the escape from this inhibitory effect is not completely understood, it has been related to the inhibition of sodium-iodide symporter-mediated iodide uptake. However, the effects of iodide excess on iodide efflux have not been characterized. Herein, we investigated the consequences of iodide excess on pendrin abundance, subcellular localization, and iodide efflux in rat thyroid PCCl3 cells. Our results indicate that iodide excess increases pendrin abundance and plasma membrane insertion after 24 h of treatment. Moreover, iodide excess increases pendrin half-life. Finally, iodide exposure also increases iodide efflux from PCCl3 cells. In conclusion, these data suggest that pendrin may have an important role in mediating iodide efflux in thyrocytes, especially under conditions of iodide excess.

Keywords: Wolff-Chaikoff effect escape; iodide efflux; iodide excess; pendrin.

Copyright © 2016 the American Physiological Society.

Figures

Fig. 1.

Pendrin protein abundance in PCCl3 rat thyroid cells is increased by treatment with 10−3 M sodium iodide (NaI; A), but not by 10−5 M (B), 10−7 M (C), or 10−9 M (D). Western blot analysis of total pendrin content was performed in cells treated or not [control (C)] for 30 min, or 1, 24, or 48 h with 10−3, 10−5, 10−7, or 10−9 M NaI. Pendrin levels were normalized to β-actin content. Immunoblots shown are representative of at least 3 independent experiments. Values are means ± SE in arbitrary units (AU). ****P < 0.0001 vs. C (ANOVA, Student-Newman-Keuls).

Fig. 2.

Treatment with iodide excess increases pendrin insertion at the membrane of PCCl3 cells. Thyroid cells were incubated or not (C) with 10−3 M NaI for 2, 12, 24, or 48 h. Cells were fixed and incubated with an antibody directed against the extracellular epitopes of pendrin under nonpermeabilized condition. Thereafter, the cells were incubated with an anti-rabbit IgG-FITC antibody. Pictures are representative of 2 independent experiments. Immunofluorescence was analyzed with a ZEISS Axiovert 100M fluorescence microscope. Green signal, pendrin; blue signal, 4′,6-diamidino-2-phenylindole. Magnification ×20.

Fig. 3.

Iodide excess increases pendrin abundance at the plasma membrane of PCCl3 cells. Relative quantification of fluorescence intensity from flow cytometry data of PCCl3 cells incubated with 10−3 NaI for 12, 24, and 48 h under nonpermeabilized condition is shown. Ten thousand events were evaluated per sample. Values are means ± SE of at least 3 independent experiments. ***P < 0.001 vs. C (unpaired two-tailed Student's t-test).

Fig. 4.

Pendrin half-life is increased in iodide-treated cells. PCCl3 cells were treated for 1 h with cycloheximide and then exposed to NaI (10−3 M) for 0, 3, 6, 8, or 10 h. Solid and shaded lines represent the pendrin decay rate of control and iodide-treated cells, respectively. Pendrin content was normalized to total protein loading. Values are means ± SE in AU. Three independent experiments were performed in triplicate. *P < 0.05 vs. 0 h (two-way ANOVA).

Fig. 5.

Iodide efflux in PCCl3 cells treated for short periods (A) or longer periods (B) of iodide excess exposure. A: cells were incubated for 1, 4, and 6 h with 10−3 M non-radioactive NaI. After that, 20 μCi/μmol I− carrier-free Na125I was added to the medium for 30 min. The medium was then replaced by medium containing perchlorate after 90, 150, and 210 s. B: cells were incubated with 10−3 M nonradioactive NaI and 20 μCi/μmol I− carrier-free Na125I for 12, 24, or 48 h. After that, the medium was replaced by medium containing perchlorate after 90, 150, and 210 s. The radioactivity in each fraction was measured using a gamma-counter. The amount of 125I was normalized by the amount of DNA in each sample. Values are means ± SE. Three independent experiments were performed in triplicate. ****P < 0.0001 vs. control (one-way ANOVA, Student-Newman-Keuls).

Fig. 6.

Iodide excess does not alter Ano1 (anoctamin 1) mRNA expression. Relative Ano1 mRNA expression was evaluated by real-time PCR. PCCl3 cells were incubated with NaI (10−3 M) for 24 h. Results are indicated as fold change relative to the mRNA levels of untreated cells. Values are means ± SE. P > 0.05 vs. control cells (unpaired two-tailed Student's t-test).

Fig. 7.

Hypothetical mechanism involved in the escape of the Wolff-Chaikoff effect. In response to high intracellular iodide concentrations, sodium-iodide symporter (NIS) synthesis and activity is inhibited, while pendrin [Pendred syndrome (PDS)] is inserted more abundantly in the apical membrane, and its mRNA expression is stimulated, which results in increased iodide efflux under conditions of a high intracellular iodide concentration.