Thyroglobulin (Tg) induces thyroid cell growth in a concentration-specific manner by a mechanism other than thyrotropin/cAMP stimulation Yoshihiko Noguchi a,b,c , Norikazu Harii c , Cesidio Giuliani c , Ichiro Tatsuno b , Koichi Suzuki d, * , Leonard D. Kohn c, * a Department of Internal Medicine, Ito Hospital, Tokyo 150-8308, Japan b Department of Clinical Cell Biology, Graduate School of Medicine, Chiba University, Chiba 260-8670, Japan c Edison Biotechnology Institute and Department of Biomedical Sciences, Building 25, The Ridges, College of Osteopathic Medicine, Ohio University, Athens, OH 45701, USA d Department of Bioregulation, Leprosy Research Center, National Institute of Infectious Diseases, 4-2-1 Aoba-cho, Higashimurayama-shi, Tokyo 189-0002, Japan article info Article history: Received 24 November 2009 Available online 29 November 2009 Keywords: Thyroglobulin Thyroid FRTL-5 cells Cell growth abstract Thyroglobulin (Tg), a major product of the thyroid gland, serves as a macromolecular precursor of thyroid hormone biosynthesis. In addition, Tg stored in the thyroid follicles is a potent regulator of thyroid-specific gene expression. In conjunction with thyroid stimulating hormone (TSH) and iodide, Tg regulates thyroid follicle function, which is the minimal functional unit of the thyroid gland. In the present study, we show that Tg stimulates growth of FRTL-5 thyroid cells in the absence of TSH, insulin and serum. Unlike TSH, Tg did not increase cellular cyclic AMP (cAMP) levels; rather, the TSH signal counteracted Tg-induced cell growth. A specific inhibitor of A-kinase, H-89, did not modulate the effect of Tg. Tg increased kinase activ- ity of Akt to the same level as TSH, insulin and 5% serum, while LY294002 abolished Tg-induced growth. Interestingly, low Tg concentrations maximized growth-promotion activity and induction of the apical iodide transporter (PDS; SLC26A4), whereas high Tg concentrations suppressed both cell growth and the expression of thyroid-specific genes. These results suggest that a low levels of Tg in the follicular lumen might stimulates cell growth and iodide transport to accelerate the iodide organification process; however, elevated Tg levels in the follicle might then shut down all of these functions. Ó 2009 Elsevier Inc. All rights reserved. Introduction Thyroglobulin (Tg) is a large glycoprotein produced and stored in the follicular lumen of the thyroid gland. Its primary function is to serve as a macromolecular substrate for the coupling of iodide to its tyrosine residues during thyroid hormone biosynthesis [1]. In addition to this conventional role, follicular Tg acts as a potent autocrine regulator of follicular function and mediates the tran- scriptional suppression of genes essential for iodide transport and hormone synthesis [2,3]. The addition of follicular Tg to the culture medium of FRTL-5 thyroid cells dramatically decreased the RNA levels of thyroid-specific genes such as Tg, thyroid perox- idase (TPO), and the sodium/iodide symporter (NIS; SLC5A5) [2,3]. Tg suppressed iodide uptake in a dose-dependent manner in FRTL- 5 cells in vitro, and the in vivo accumulation of Tg in the follicular lumen correlates with low iodide uptake in the rat thyroid [3]. Tg concentration used in these studies is in the lower range of follicular Tg in different follicles of the normal thyroid, which var- ies from 0.1 mg/ml up to 250 mg/ml as measured by aspiration biopsy or micropuncture of a single follicle [4–6]. The relatively large range in estimates of follicular Tg concentration is the result of significant functional heterogeneity among follicles. Follicular Tg not only exhibits suppressive activity, but also increases mRNA levels and promoter activity of major histocom- patibility complex (MHC) class I genes, and significantly induces the Pendred syndrome gene (PDS; SLC26A4), an apical iodide transporter in the thyroid [2,7,8]. NIS and PDS, basal and apical io- dide transporters, respectively, are differentially regulated by Tg concentration and exposure time. One of the interesting differ- ences between the suppressive and activating effects of Tg is that the former seems to occur in a dose-dependent manner, while the latter maximizes at a lower concentration of Tg [2,7,8]. It was therefore proposed that accumulated follicular Tg within the col- loid is a major regulator of follicular function [7,9–12]. The physi- ological turnover of follicular function seems to be dynamically and coordinately regulated by Tg and TSH. In addition to the effects of Tg on modulation of thyroid gene expression and function, it is evident that Tg also regulates cell growth and gene transcription in various cell types. In mink lung epithelial cells, Tg is activated by disuccinimidyl suberate (DDS) to act as an agonist of transforming growth factor (TGF)-b and 0006-291X/$ - see front matter Ó 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2009.11.158 * Corresponding authors. Fax: +81 42 394 9092 (K. Suzuki); fax: +425 885 0205 (L.D. Kohn). E-mail addresses: koichis@nih.go.jp (K. Suzuki), leonardkohn@gmail.com (L.D. Kohn). Biochemical and Biophysical Research Communications 391 (2010) 890–894 Contents lists available at ScienceDirect Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc