Synthesis and secretion of lipids by long-term cultures of female rat hepatocytes

Abstract

The objective of this work was to characterize lipid metabolism in long-term cultures of adult rat hepatocytes from female rats and explore the potential use of this culture system to study the effect of hormones, drugs and toxic chemicals on it. Hepatocytes, seeded on a feeder layer of 3T3 cells, maintained for 2 weeks their typical morphology. The cultures were able to take up [ 14C]acetic and [ 14C]oleic acid from the culture medium and incorporate them into lipids. The synthesis and secretion of lipids by [ 14C]acetic acid-labeled cultures had a maximum value after 11 and 13 days in culture. Triacylglycerols were the main lipidic species synthesized and secreted by hepatocytes (up to 67% of the total lipids); they also synthesized and secreted phospholipids, cholesterol and cholesterol esters from [ 14C]acetic acid. Similarly, [ 14C]oleic acid-labeled cultures synthesized and secreted mostly triacylglycerols (up to 60–70% of the total lipids), but they were also able to incorporate the labeled precursor into both cellular and secreted phospholipids and cholesterol esters. The activity of glycerol-phosphate-dehydrogenase, marker enzyme of glycerolipid synthesis, decreased slightly during the culture time whereas the activity of malic enzyme, marker of fatty acid synthesis, increased. Our results show that long-term cultures of female rat hepatocytes are able to synthesize and secrete several lipids, specially triacylglycerols, from both [ 14C]acetic and [ 14C]oleic acid for at least 2 weeks and that they maintain enzyme activities related with the synthetic pathways of glycerolipids and fatty acids. They also suggest that these cultures could be a useful model system to study lipid metabolism and the effect of hormones, drugs and toxic chemicals on it.