Maturation of peroxisomes in differentiating human hepatoblastoma cells (HepG2): possible involvement of the peroxisome proliferator-activated receptor alpha (PPAR alpha)

We have studied the alterations of peroxisomes in the human hepatoblastoma cell line HepG2, induced to differentiate by long-term cultivation (20 days without passaging) using morphological and biochemical techniques as well as mRNA analysis. Ultrastructural studies revealed alterations in shape and size of peroxisomes, with significant increases in mean diameter and forma tion of small clusters exhibiting heterogeneous staining for catalase after 20 days in culture. These alterations of peroxisomes correspond to the cha nges described during the maturation process from prenatal to adult human h epatocytes. As revealed by Northern and Western blotting there was marked e levation of the mRNA (190%) and protein (180%) of the peroxisomal branched- chain acyl-CoA oxidase. This protein is the key regulatory enzyme for the s ide chain oxidation of cholesterol for bile acid synthesis, a pathway assoc iated with mature hepatocytes. Concomitantly a marked increase of bile cana liculi was noted by light and electron microscopy. This differentiation pro cess was confirmed also by the increase of albumin synthesis (mRNA: 160%; p rotein: 190%) which is generally used as a differentiation marker of hepato cytes in culture. Interestingly, the mRNA for peroxisome proliferator-activ ated receptor a (PPAR alpha) increased drastically by almost 390% and its c orreponding protein by 150%, suggesting its involvement in maturation of th e peroxisomal compartment in differentiating HepG2 cells. In contrast to th e well-known increases during the drug-induced peroxisome proliferation of cytochrome P450 4A, multifunctional enzyme 1, palmitoyl-CoA oxidase and the 70-kDa peroxisomal membrane protein, those proteins were either not altere d or only slightly elevated during the differentiation process, suggesting that peroxisome proliferation and maturation are two distinct and different ially regulated processes.