In rats fed a high fat diet (HFD), long-term administration of 3,5-diiodo-L-thyronine (T2), a naturally occurring iodothyronine, was shown to reduce body-weight gain, fat mass, and hepatic lipid accumulation. This work was aimed at investigating the mechanisms of T2 action in the liver of HFD rats. The results show that HFD induces liver lipid peroxidation and stimulates the activity of enzymes involved in hydrogen peroxide (H2O2) metabolism, catalase in particular. Moreover, quantitative RT-PCR revealed HFD-induced upregulation of the transcription factor PPAR alpha, as well as of metallothionein isoforms (MT-1 and MT-2). T2 administration prevented the HDF-induced lipid peroxidation, as well as the increase in H2O2 metabolism, and reduced the upregulation of both PPAR alpha and MT-2. These data demonstrate that in the liver of HFD rats, T2 prevents both lipid accumulation and oxidative stress associated with increased fat metabolism.