Iron plays an important role in cell growth and metabolism. In preneoplastic liver nodules, a rise in the number of transferrin receptors (Tf-R) is associated with decreased endocytosis of the Fe2-Tf/Tf-R complex. Because nodules are precursors of hepatocellular carcinoma (HCC), the question arises whether changes in iron uptake by nodules persist in HCC. Current work showed up-regulation of Tf messenger RNA (mRNA) production in preneoplastic nodules, 12 to 37 weeks after initiation, and down-regulation in atypical nodules (at 45 and 50 weeks) and HCCs, induced in rats by the "resistant hepatocyte" model. Tf-R gene expression increased in nodules and HCCs. Tf-R numbers increased, without changes in affinity constant, in HCC. Iron uptake was higher in HCC than in normal liver, 5 to 40 minutes after injection of 59Fe2-Tf, with preferential accumulation in cytosol of tumor cells and in microsomes of normal liver. Purification through Percoll gradient of mitochondria plus lysosomes allowed the identification in liver and HCC of an endosomal compartment sequestering injected 125I-Tf. This subfraction was not seen when 59Fe2-Tf was injected into rats, and 59Fe was found in particulate material of both tissues. Liver and HCC exhibited comparable basal activities of plasma membrane NADH oxidase, an enzyme involved in iron uptake and cell growth. Stimulation of this activity by Fe2-Tf was higher in HCC than in normal liver. These results indicate that Tf expression may be a marker of preneoplastic liver progression to malignancy. Differently from nodules, HCC may sequester relatively high iron amounts, necessary for fast growth, both through the endocytic pathway and the reduced form of nicotinamide adenine dinucleotide (NADH) oxidase system.