Expansion of adipose tissue in the orbits is a key feature of Graves' ophthalmopathy. Recent evidence shows that orbital fibroblasts are committed to differentiate into adipocytes under appropriate stimuli. Rosiglitazone, an agonist of the nuclear hormone receptor, peroxisome proliferator-activated receptor gamma (PPARgamma) is able to induce both differentiation of orbital fibroblasts into mature adipocytes and expression of the TSH receptor (TSHr) gene. Several studies have suggested an important role of the high mobility group AT-hook 2 (HMGA2) gene in adipocytic cell growth and development. To investigate further the association between adipogenesis-related genes and orbital fibroblasts, we treated fibroblasts from Graves' ophthalmopathy (FGOs) and from normal orbital tissues with fenofibrate, a specific agonist for PPARalpha. We then evaluated the expression of the PPARalpha, PPARgamma2, HMGA2, leptin and TSHr genes before and after 24 h of fenofibrate treatment, using semiquantitative and real-time PCR. For up to 96 h after exposure to fenofibrate, FGOs differentiated into adipocytes. PPARalpha and PPARgamma2 were expressed more in FGOs than in normal cultures, whereas TSHr mRNA was detected only in FGOs. Expression of HMGA2 mRNA and protein was significantly increased in FGOs from 6 to 24 h after fenofibrate, confirming its role in the early phase of adipocyte differentiation. Treatment with fenofibrate for 24 h significantly increased the expression of leptin and TSHr genes. Moreover, TSH treatment significantly increased the accumulation of cAMP, demonstrating that FGOs express functional TSHr. The high level of expression of PPARalpha other than PPARgamma2 transcripts and the stimulating effect of fenofibrate on adipogenesis and on HMGA2, leptin and TSHr genes also indicate that the PPARalpha pathway plays an important part in the adipocyte differentiation of FGOs. These findings suggest that novel drugs to antagonize PPARalpha, other than the PPARgamma signalling system, may also need to be considered in the treatment or prevention of Graves' ophthalmopathy.