Background: Thyrotropin (TSH) and thyroid-stimulating antibodies (TSAbs) activate TSH receptor (TSHR) signaling by binding to its extracellular domain. TSHR signaling has been studied extensively in animal thyrocytes and in engineered cell lines, and differences in signaling have been observed in different cell systems. We, therefore, decided to characterize and compare TSHR signaling mediated by TSH and monoclonal TSAbs in human thyrocytes in primary culture. Methods: We used quantitative reverse transcription-polymerase chain reaction to measure mRNA levels of thyroid-specific genes thyroglobulin (TG), thyroperoxidase (TPO), iodothyronine deiodinase type 2 (DIO2), sodium-iodide symporter (NIS), and TSHR after stimulation by TSH or two monoclonal TSAbs, KSAb1 and M22. We also compared secreted TG protein after TSHR activation by TSH and TSAbs using an enzyme-linked immunosorbent assay. TSHR cell surface expression was determined using fluorescence activated cell sorting (FACS). Results: We found that TSH at low doses increases and at high doses (>1 mU/mL) decreases levels of gene expression for TSHR, TG, TPO, NIS, and DIO2. The biphasic effect of TSH on signaling was not caused by downregulation of cell surface TSHRs. This bell-shaped biphasic dose-response curve has been termed an inverted U-shaped dose-response curve (IUDRC). An IUDRC was also found for TSH-induced regulation of TG secretion. In contrast, KSAb1- and M22-induced regulation of TSHR, TG, TPO, NIS, and DIO2 gene expression, and secreted TG followed a monotonic dose-response curve that plateaus at high doses of activating antibody. Conclusions: Our data demonstrate that the physiological activation of TSHRs by TSH in primary cultures of human thyrocytes is characterized by a regulatory mechanism that may inhibit thyrocyte overstimulation. In contrast, TSAbs do not exhibit biphasic regulation. Although KSAb1 and M22 may not be representative of all TSAbs found in patients with Graves' disease, we suggest that persistent robust stimulation of TSHRs by TSAbs, unrelieved by a decrease at high TSAb levels, fosters chronic stimulation of thyrocytes in Graves' hyperthyroidism.
Keywords: TSH receptor; biphasic dose response; gene expression; thyroglobulin.