**Author details**

378 Thyroid Hormone

assumed *in vivo* [102].

were reproduced.

**6. Conclusion** 

synthesis.

cell membrane, due to loss of polarity [109].

demonstrated with rat or pig isolated and closed follicle cultures.

iodide dose [105]. This was perhaps presented by reduced AM2Pc levels at 48 h, as excess Iinhibited an increase in AMPc stimulated by TSH in hypophysectomised rats [106] and mice [34], which could have explained the low NIS level during Wolff-Chaikoff effect and in

production and increased Ca2+ flow induced by TSH, which could have led to reduced peroxide production during Wolff-Chaikoff effect [107,108]. The organification observed in the presence of 10E-5 M (Table 8) did not completely inhibit TPO and had no effect on NIS in the presence of 10E-3 M NaI (Figures 13E and 13H), thereby demonstrating that organification depends on TPO and not on NIS as in cells transfected with the TPO gene [34]. TSH did not stimulate the organification of iodide captured in 10E-3 M but did so in 10E-5 M NaI (Tables 8 and 10), as it has been described that the effect of TSH on thyroid physiology becomes reduced in the presence of excess I- , meaning that antagonic roles are

TSH modulated relative NIS expression and its subcellular localisation in the thyrocytes of isolated and closed follicles *in vitro*. These results were similar to those found in FRTL5cells, where it has been demonstrated that *de novo* synthesis [32], half-life time, NIS targeting and/or retention regarding cytoplasmatic membrane requires TSH to be located throughout

Thyrocyte disposition in follicles has not been necessary for iodide accumulation, since it has been present in foetal thyroids before follicular lumen formation [110] and also in primary cultures from normal thyrocytes [111] or goitre patients [112] and in the FRTL cell line [113]; however, these cultures have required TSH, hormones and other molecules for maintaining them. Nevertheless, isolating the colloidal cavity from the exterior must be ensured for iodide accumulation and incorporation in Tg, T3 and T4 hormone synthesis, as

Rat and pig follicles thus inhibited iodide organification in the presence of strong concentrations of iodide, i.e. performed the Wolff-Chaikoff effect. Neither thyrocytes' follicular architecture nor ultra-structure was modified and no sign of cell death was presented. The TSH and iodide effects observed *in vivo* during the Wolff-Chaikoff effect

Loss of follicular structure during the first 24 h of culture has been the main drawback of *in vitro* thyroid studies and, therefore, hormone synthesis. It is not enough to conserve thyrocytes' apical-basal polarity in culture in this specific tissue for maintaining colloid's extracellular functions for the enzymes implicated in iodide fixation on Tg and hormone

We have shown that follicular architecture must be conserved in culture, especially the follicular cavity isolated from extracellular medium as this is indispensable for maintaining ultra-structure and the polarity of thyrocytes around the follicular cavity; such premise

inhibited IP3

follicles at 48 h in the presence of excess iodide (Figure s13G and 13H). Excess I-

Clara Spinel1,3, Magnolia Herrera3 and and Jhon Rivera2,3 *Membrane Biophysics and Biology Group, 1Biology Department, Science Faculty, Universidad Nacional de Colombia, Colombia 2Chemistry Department, Science Faculty, Universidad Nacional de Colombia, Colombia 3International Physics Center, Bogotá, Colombia* 
