24-hydroxycholesterol is a substrate for hepatic cholesterol 7alpha-hydroxylase (CYP7A).

(24S)-Hydroxycholesterol is formed from cholesterol in the brain and is important for cholesterol homeostasis in this organ. Elimination of (24S)-hydroxycholesterol has been suggested to occur in the liver but little is known about the metabolism of this oxysterol. In the present investigation, we report formation of 7alpha, 24-dihydroxycholesterol in pig and human liver. 7alpha-hydroxylase activity toward both isomers of 24-hydroxycholesterol [(24S) and (24R)] was found in a partially purified and reconstituted cholesterol 7alpha-hydroxylase (CYP7A) enzyme fraction from pig liver microsomes. In contrast, a purified enzyme fraction of pig liver oxysterol 7alpha-hydroxylase with high activity toward 27-hydroxycholesterol did not show any detectable activity toward 24-hydroxycholesterol. 7alpha-Hydroxylation of 24-hydroxycholesterol was strongly inhibited by 7-oxocholesterol, a known inhibitor of CYP7A. Human CYP7A, recombinantly expressed in Escherichia coli and in simian COS cells, showed 7alpha-hydroxylase activity toward both cholesterol and the two isomers of 24-hydroxycholesterol, with a preference for the (24S)-isomer. Our results show that 24-hydroxycholesterol is metabolized by CYP7A, an enzyme previously considered to be specific for cholesterol and cholestanol and not active toward oxysterols. Because CYP7A is the rate-limiting enzyme in the major pathway of bile acid biosynthesis, the possibility is discussed that at least part of the 24-hydroxycholesterol is converted into 7alpha-hydroxylated bile acids by the enzymes involved in the normal biosynthesis of bile acids.

[1]  D. Russell,et al.  Expression Cloning of an Oxysterol 7α-Hydroxylase Selective for 24-Hydroxycholesterol* , 2000, The Journal of Biological Chemistry.

[2]  J. van der Linden,et al.  Elimination of cholesterol as cholestenoic acid in human lung by sterol 27-hydroxylase: evidence that most of this steroid in the circulation is of pulmonary origin. , 1999, Journal of lipid research.

[3]  S. Kliewer,et al.  Structural requirements of ligands for the oxysterol liver X receptors LXRalpha and LXRbeta. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[4]  J. Wahren,et al.  Cholesterol homeostasis in human brain: turnover of 24S-hydroxycholesterol and evidence for a cerebral origin of most of this oxysterol in the circulation. , 1998, Journal of lipid research.

[5]  M. Norlin,et al.  Biochemical characterization of the 7alpha-hydroxylase activities towards 27-hydroxycholesterol and dehydroepiandrosterone in pig liver microsomes. , 1998, Biochimica et biophysica acta.

[6]  D. Lütjohann,et al.  Importance of a Novel Oxidative Mechanism for Elimination of Brain Cholesterol , 1997, The Journal of Biological Chemistry.

[7]  I. Björkhem,et al.  Elimination of Cholesterol in Macrophages and Endothelial Cells by the Sterol 27-Hydroxylase Mechanism , 1997, The Journal of Biological Chemistry.

[8]  R. Lathe,et al.  Identification and Characterization of a Mouse Oxysterol 7α-Hydroxylase cDNA* , 1997, The Journal of Biological Chemistry.

[9]  R. Lathe,et al.  Cyp7b, a novel brain cytochrome P450, catalyzes the synthesis of neurosteroids 7α-hydroxy dehydroepiandrosterone and 7α-hydroxy pregnenolone , 1997 .

[10]  E. Baulieu,et al.  Metabolism of 27-, 25- and 24-hydroxycholesterol in rat glial cells and neurons. , 1997, The Biochemical journal.

[11]  D. Lütjohann,et al.  Cholesterol homeostasis in human brain: evidence for an age-dependent flux of 24S-hydroxycholesterol from the brain into the circulation. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[12]  J. Mason,et al.  A Novel Cytochrome P450 Expressed Primarily in Brain (*) , 1995, The Journal of Biological Chemistry.

[13]  E. Adashi,et al.  A novel nonhepatic hydroxycholesterol 7 alpha-hydroxylase that is markedly stimulated by interleukin-1 beta. Characterization in the immature rat ovary. , 1995, The Journal of biological chemistry.

[14]  O. Larsson,et al.  7α-Hydroxylation of 25-hydroxycholesterol and 27-hydroxycholesterol in human fibroblasts , 1995 .

[15]  E. Baulieu,et al.  7 Alpha-hydroxylation of 27-hydroxycholesterol in rat brain microsomes. , 1995, Comptes rendus de l'Academie des sciences. Serie III, Sciences de la vie.

[16]  I. Björkhem,et al.  7α Hydroxylation of 25‐Hydroxycholesterol in Liver Microsomes , 1994 .

[17]  N. Javitt,et al.  Cholesterol and 27-hydroxycholesterol 7 alpha-hydroxylation: evidence for two different enzymes. , 1993, Journal of lipid research.

[18]  D. Clark,et al.  Hepatic uptake and metabolism of ingested 24-hydroxycholesterol and 24(S),25-epoxycholesterol. , 1993, Biochimica et biophysica acta.

[19]  I. Björkhem,et al.  Evidence that 24- and 27-hydroxylation are not involved in the cholesterol-induced down-regulation of hydroxymethylglutaryl-CoA reductase in mouse liver. , 1992, The Journal of biological chemistry.

[20]  K. Einarsson,et al.  7α-Hydroxylation of 27-hydroxycholesterol in human liver microsomes , 1992 .

[21]  J. Sjövall,et al.  7α‐Hydroxylation of 26‐hydroxycholesterol, 3β‐hydroxy‐5‐cholestenoic acid and 3β‐hydroxy‐5‐cholenoic acid by cytochrome P‐450 in pig liver microsomes , 1992 .

[22]  J. Sjövall,et al.  Potential bile acid precursors in plasma--possible indicators of biosynthetic pathways to cholic and chenodeoxycholic acids in man. , 1990, Journal of steroid biochemistry.

[23]  M. Noshiro,et al.  Molecular cloning and sequence analysis of cDNA encoding human cholesterol 7α‐hydroxylase , 1990 .

[24]  K. Boberg,et al.  Apparent lack of conversion of sitosterol into C24-bile acids in humans. , 1990, Journal of lipid research.

[25]  C. Slaughter,et al.  Cloning and regulation of cholesterol 7 alpha-hydroxylase, the rate-limiting enzyme in bile acid biosynthesis. , 1990, The Journal of biological chemistry.

[26]  H. Boström Binding of cholesterol to cytochromes P-450 from rabbit liver microsomes. , 1983, The Journal of biological chemistry.

[27]  T. Boulikas,et al.  Silver staining of proteins in polyacrylamide gels. , 1981, Analytical biochemistry.

[28]  Y. Yasukochi,et al.  Some properties of a detergent-solubilized NADPH-cytochrome c(cytochrome P-450) reductase purified by biospecific affinity chromatography. , 1976, The Journal of biological chemistry.

[29]  P. Eneroth,et al.  Studies on the formation of C7-oxygenated cholesterol and beta-sitosterol metabolites in cell-free preparations of rat liver. , 1973, Journal of lipid research.

[30]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[31]  E. Mosettig,et al.  5-Cholestene-3β,26-diol , 1956 .

[32]  Oliver H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[33]  M. Waterman,et al.  Heterologous expression of mammalian P450 in COS cells. , 1991, Methods in enzymology.

[34]  S. Andersson,et al.  Purification from rabbit and rat liver of cytochromes P-450 involved in bile acid biosynthesis. , 1985, Methods in enzymology.

[35]  S. Margolis,et al.  Effects of drugs and sterols on cholesterol 7 alpha-hydroxylase activity in rat liver microsomes. , 1983, Journal of lipid research.

[36]  L. Stárka Reaktion der steroide mit tert-butylperbenzoat I. Über die 7-acyloxylierung Δ 5 -ungesättigter steroide , 1961 .