Tri-iodothyronine induces proliferation in cultured bovine thyroid cells: Evidence for the involvement of epidermal growth factor-associated tyrosine kinase activity

Abstract

The effects of the tri-iodothyronine (T3) secreted by thyroid cells on the growth of the thyrocyte are poorly known. In this study we analyzed the effects of T3 on the proliferation of bovine thyroid follicles in primary culture previously depleted of endogenous T3. Cellular deoxiribonucleic acid (DNA) synthesis, determined by [3H]thymidine incorporation, was stimulated by T3 (0.1-5.0 nM) for 24 h in a concentration-dependent fashion with a maximal effect at 1.0 nM T3 (P < 0.01). This T3 action was time-dependent when assayed from 12 to 72h. The induction of mitogenic activity was corroborated by the increase in proliferating cell nuclear antigen (PCNA) measured by Western blot analysis. PCNA increased after treatment with T3 (0.1-5.0 nM) in a concentrationdependent manner. Since T3 modifies the activity of growth factors whose actions are mainly mediated by tyrosine kinase (TK) activation in diverse cellular types, we assayed the effects ofgenistein, a general TK inhibitor, and tyrphostin A25, a specific epidermal growth factor (EGF)-receptor (EGFR)-dependent TK activity inhibitor, on the proliferative effects of T3. The T3-induced [3H]thymidine incorporation was inhibited by both agents in a concentration-dependent manner. A significant increase in the total TK activity measured in cellular protein extracts was induced by 0·5 and 1·0 nM T3 (P < 0.001). Tyrosine phosphorylation of the EGFR was also stimulated by T3 (P < 0.001) with no change in the EGFR expression as determined by Western blot analysis. Both, the T3-stimulated [3H]thymidine incorporation and the TK activity were inhibited by a anti-mouse EGF antibody. These results lead us to propose that T3 could operate as a proliferative agent in bovine thyroid cells through a mechanism involving an autocrine/paracrine EGF/EGFR-dependent regulation.