Decreased intracellular degradation and increased secretion of apo B-100 in Hep G2 cells after inhibition of cholesteryl ester synthesis.
Control of apolipoprotein B (apo B) secretion in hepatocytes occurs partly at the post-translational level. The key step in this process appears to be intracellular degradation of newly synthesized apo B. The aim of this paper was to investigate the mechanisms that regulate apo B secretion by Hep G2 cells, in response to the inhibition of Acyl-CoA Acyltransferase (ACAT) by the compound Sandoz 58035 (S-58035). S-58035 (20 microM) reduced cholesteryl ester synthesis from [14C]oleate by 95%, and increased significantly, in a dose-dependent manner, (2-100 microM) apo B secretion, either in control conditions (from 78 +/- 4.3 to 126 +/- 6.1 ng apo B-100/mg cell protein/4 h) or upon stimulation of apo B secretion by oleate (from 134 +/- 4.23 to 177 +/- 4.3 ng apo B/mg cell protein/4 h). This increased secretion of newly synthesized apo B-100 was confirmed by pulse experiments and by gradient ultracentrifugation of the media. Moreover pulse-chase experiments showed that the addition of S-58035 reduced intracellular degradation of apo B-100, both in control conditions and in the presence of oleate. S-58035 (20 microM) did not affect total cellular cholesterol content, but free cholesterol increased with a concomitant decrease of cholesteryl ester (-20%). S-58035 increased cellular triglyceride mass, which was observed in basal conditions (from 12.8 +/- 1.09 to 22.7 +/- 2.7 micrograms TG/mg cellular protein) and also in presence of oleate (from 48 +/- 0.53 to 59 +/- 6.3 micrograms TG/mg cellular protein). This effect is due to a stimulation of triglyceride synthesis, as determined by incorporation of [3H]glycerol into cellular triglycerides. From these data we conclude that, under our experimental conditions, triglyceride synthesis and/or availability is likely to control intracellular degradation of apo B.
Ooyen
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Institute of Pharmacological Sciences, Milano, Italy.
Zecca
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Zanelli
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Catapano
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D000577:Amides / Q000494:pharmacology; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000378:metabolism*; D002788:Cholesterol Esters / Q000037:antagonists & inhibitors* / Q000096:biosynthesis*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D050356:Lipid Metabolism; D008099:Liver / Q000378:metabolism*; D019301:Oleic Acid / Q000494:pharmacology; D017646:Organosilicon Compounds; D002785:Sterol O-Acyltransferase / Q000037:antagonists & inhibitors; D014280:Triglycerides / Q000096:biosynthesis; D014407:Tumor Cells, Cultured
D000577:Amides; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D002788:Cholesterol Esters; D004791:Enzyme Inhibitors; D017646:Organosilicon Compounds; D014280:Triglycerides; D019301:Oleic Acid; C040256:SAN 58035; D002785:Sterol O-Acyltransferase