RETINOIC ACID TREATMENT INDUCES APOPTOSIS OR EXPRESSION OF A MORE DIFFERENTIATED PHENOTYPE ON DIFFERENT FRACTIONS OF CULTURED FETAL-RAT HEPATOCYTES

The present study reports the effects of retinoic acid (RA) on culture d fetal rat hepatocytes. We show that RA treatment induces both differ entiation and apoptosis, Hepatocytes cultured for 48 hours in the pres ence of 5 mu mol/L RA form junctional complexes in the areas of contac t between neighboring cells and develop bile canaliculi, typical featu res of mature and well-differentiated cells. At the same time, about 2 0% of cells are induced to die by apoptosis, and the percentage of apo ptotic cells increases according to the concentration of RA used and t he duration of treatment. The induction of apoptosis, studied at the m orphological and biochemical levels, revealed that, in our system, the classical compaction of chromatin occurs only during the final stages of the process; instead of the common marker of apoptosis, i.e., the ''DNA ladder'' pattern of fragmentation, megabase-sized fragments were found. These observations provide further evidence of the existence o f fundamental differences in the mechanisms of apoptosis among cell ty pes. To investigate the molecular mechanism of the effects of RA, we e valuated the expression of two proteins, c-myc and p53, which are know n to be involved in both cell differentiation and apoptosis, The data obtained show that the amount of p53 remained unchanged after RA treat ment. On the contrary, a dose-dependent reduction in c-myc levels was found, suggesting that RA action may be mediated by modulation of this oncogene. Our findings regarding the apoptosis-inducing effect of RA, which was not found in adult hepatocytes, suggest a possible relation ship between this phenomenon and the proliferative capacity and/or dif ferentiation state of hepatocytes.