The 2-acetaminofluorene/partial hepatectomy (AAF/Phx) model is widely used to induce oval/progenitor cell proliferation in the rat liver. We have used this model to study the impact of a primary hepatocyte mitogen, triiodothyronine (T3) on the liver regenerating by the recruitment of oval/progenitor cells. Administration of T3 transiently accelerates the proliferation of the oval cells, which is followed by rapid differentiation into small hepatocytes. The oval cell origin of the small hepatocytes has been proven by tracing retrovirally transduced and BrdU marked oval cells. The differentiating oval cells become positive for hepatocyte nuclear factor-4 and start to express hepatocyte specific connexin 32, alpha1 integrin, Prox1, cytochrom P450s, and form CD 26 positive bile canaliculi. At the same time oval cell specific OV-6 and alpha-fetoprotein expression is lost. The upregulation of hepatocyte specific mRNAs: albumin, tyrosine aminotransferase and tryptophan 2,3-dioxygenase detected by real-time PCR also proves hepatocytic maturation. The hepatocytic conversion of oval cells occurs on the seventh day after the Phx in this model while the first small hepatocytes appear 5 days later without T3 treatment. The administration of the primary hepatocyte mitogen T3 accelerates the differentiation of hepatic progenitor cells into hepatocytes in vivo, and that may have therapeutic potential.