Th1 and Th2 cells have critical roles in the development of cell-mediated and humoral immune responses, respectively. This division of function predicts that Th1 cells mediate inflammatory diseases and Th2 cells promote antibody-mediated autoimmunity. Our earlier studies using HEK-293 cells expressing extracellular domain of the TSHR showed that Stat4^-/- mice, that lack Th1 cells, are susceptible while Stat6^-/- mice, that lack Th2 cells, are resistant to the induction of Graves' hyperthyroidism. To investigate the role of Stat4 and Stat6 genes in other murine models of hyperthyroidism, we injected wild-type BALB/c, Stat4^-/- and Stat6^-/- mice with an adenovirus expressing amino acid residues 1-289 of TSHR (TSHR-289-ad or 289-ad). The viral system induces a much stronger immune response with much more rapid onset of disease. Our results showed that 56% of WT, 75% of Stat4^-/-, and 39% Stat6^-/- mice developed hyperthyroidism. Hyperthyroid mice exhibited thyroid stimulatory antibodies. The Stat4^-/- mice developed higher incidence and greater severity of hyperthyroidism compared with WT and Stat6^-/- mice. BALB/c and Stat4^-/- mice showed significantly higher TSHR antibodies of IgG1 subclass, and IL-4, compared with Stat6^-/- mice. In contrast, Stat6^-/- mice had predominantly IgG2a subclass of TSHR antibodies and produced significantly higher amount of IFN- than BALB/c and Stat4^-/- mice. All hyperthyroid mice showed enlarged thyroid glands with hyperactivity. These results suggest that in the TSHR-289-ad model the Th2 cells are more efficient in mediating disease, but in the absence of Th2 cells the Th1 cells may still initiate a reduced incidence of Graves' hyperthyroidism.