Journal of Lipid Research, Vol 35, 2241-2253, Copyright © 1994 by Lipid Research, Inc.

ARTICLES

In vivo formation of 25-hydroxycholesterol from endogenous cholesterol after a single meal, dietary cholesterol challenge

KA Johnson, CJ Morrow, GD Knight and TJ Scallen
Department of Biochemistry, University of New Mexico, Albuquerque 87131.

The role of oxysterols as regulatory molecules in the suppression of 3- hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity was investigated in the intact rat in response to an acute dietary cholesterol challenge. When rats were fed highly purified cholesterol as a single meal at a level of 5% of the diet, maximal inhibition of enzyme activity (66%) occurred 120 min after the completion of the meal. Furthermore, when nonsaponifiable liver extracts were chromatographically resolved and analyzed by high performance liquid chromatography (HPLC) and capillary gas chromatography-mass spectrometry (GC-MS), 25-hydroxycholesterol was identified in the livers of rats 120 min after the completion of the single cholesterol meal. Significantly, only barely detectable amounts of 25- hydroxycholesterol were observed in the livers from control rats fed a sterol-free diet. The biosynthetic origin of 25-hydroxycholesterol was investigated with the use of deuterated water. Rats were fed deuterium oxide (33%) ad libitum for 3 days and then killed 120 min after the completion of a single cholesterol meal. As before, 25- hydroxycholesterol was detected in the livers from cholesterol-fed rats, but not to a significant extent in livers from control-fed rats receiving a sterol-free diet. Isotope ratio mass spectrometry revealed that the fractional incorporation of deuterium into 25- hydroxycholesterol (21%) was less than that observed for cholesterol (24%) isolated from the same livers, indicating that 25- hydroxycholesterol was produced endogenously from exogenous cholesterol and not from autoxidation of cholesterol. In a separate experiment it was also shown that [3H]mevalonate was incorporated into 25- hydroxycholesterol after a single meal cholesterol challenge, but was barely detected in the livers of control rats. The evidence obtained in the present article supports the hypothesis that 25-hydroxycholesterol is endogenously produced from cholesterol at early time intervals after an acute dietary cholesterol challenge. In addition, rat liver HMG-CoA reductase was inhibited by the administration of a single intragastric dose (1 microgram/kg) of an aqueous solution of 25-hydroxycholesterol. Thus, the results provide strong support for the conclusion that 25- hydroxycholesterol plays a significant role in the in vivo regulation of rat liver cholesterol biosynthesis after an acute dietary cholesterol challenge.
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