Arsenite alters heme synthesis in long-term cultures of adult rat hepatocytes

Arsenite (As[III]) effects on the intermediate steps of heme biosynthesis w ere studied in adult rat hepatocytes seeded on a feeder layer of 3T3 cells (3T3-hepatocytes) and maintained for 2 weeks with culture medium non-supple mented or supplemented with 150 mu M 5-aminolevulinic acid (ALA), The activ ities of the intracellular enzymes porphobilinogen deaminase (PBG-D), uropo rphyrinogen III synthase (UROIII-S), and uroporphyrinogen In decarboxylase (URO-D), and the intermediary uroporphyrins (URO), coproporphyrins (COPRO) and protoporphyrin IX (PROTO) were determined in these cultures. The 3T3-he patocytes maintained the activities of PBG-D, UROIII-S and URO-D during 2 w eeks and ALA addition to the culture medium increased PBG-D (2-3-fold) and UROIII-S (50%) activities and porphyrin production, which accumulated as PR OTO, Exposure to 3.9 mu M As(III) inhibited UROIII-S activity (down to 34%) , and PBG-D and URO-D activities to a lower extent; these effects were magn ified by ALA supplementation. As(III) also produced an intracellular accumu lation and a decreased excretion of PROTO, and a 31% reduction of the COPRO /URO ratio in the culture medium. Additionally, As(III) caused cytoplasmic vacuolization and lipid accumulation. Our results show that As(III) exposur e selectively inhibits several intermediary enzymes of heme metabolism and affects the intra- and extracellular content of porphyrins and their ratio in the culture medium. They also confirm that 3T3-hepatocytes are a suitabl e in vitro model to study hepatic heme metabolism and its alterations by he patotoxic chemicals.