| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Journal of Lipid Research, Vol. 43, 665-670, May 2002
Copyright © 2002 by Lipid Research, Inc.
Review |
Department of Pediatrics and Medicine, NYU School of Medicine, New York, NY 10016
1 e-mail norman.javitt{at}med.nyu.edu
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONOverview of the cholesterol...REFERENCES |
|---|
-hydroxylase activity in human macrophages. The same enzyme determines the proportion of mono-, di-, and tri-hydroxy bile acids synthesized in the liver. Oxysterol 7-hydroxylase deficiency is a molecular basis for cholestatic liver disease. Chenodeoxycholic acid, the major normal end product, downregulates expression of cholesterol 7-hydroxylase via the FXR/short heterodimer protein nuclear receptor and thus limits total bile acid production. The challenge is to quantify in a physiologic setting the magnitude of the pathway in different tissues and to further evaluate the biologic roles of all the intermediates that may function as ligands for orphan nuclear receptors or via other regulatory mechanisms.—Javitt, N. B. 25R,26-Hydroxycholesterol revisited: synthesis, metabolism, and biologic roles. J. Lipid Res. 2002. 43: 665–670.
Abbreviations: StAR, steroid acute response
Supplementary key words CYP27 • CYP7A • CYP7B • ABC transporter • monohydroxy bile acids • StAR • cholestasis • atheroma
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONOverview of the cholesterol...REFERENCES |
|---|
As far as I have been able to discover, 25R,26-hydroxycholesterol was identified by Fredrickson (4) with the help of Scheer, Thompson, and Mosettig (5) at the National Institutes of Health. It represents a classic example of how cooperation between the biological sciences and physical sciences can bring to fruition observations that otherwise would have remained ill-defined.
Fredrickson incubated radioactive cholesterol with mouse liver mitochondria and obtained some products that were difficult to characterize. He was quite certain based on chromatographic criteria that one of the products was 25-hydroxycholesterol but another had slightly different properties. He suspected that it might be 26-hydroxycholesterol but obtaining complete characterization of the miniscule amount of radioactive metabolite was not possible. Enlisting the interest of a group of synthetic organic chemists led to a chemical synthesis of 25R,26-hydroxycholesterol (5), and thus to providing abundant crystalline material for definitive identification of the radioactive metabolite by reverse isotope dilution and other parameters.
Although no one disputes the correctness of the designation of the compound derived from kryptogenin or diosgenin as 25R,26-hydroxycholesterol based on the rules governing the priority of numbering carbon atoms attached to the pro-chiral C25 carbon (6), some ambiguity exists if the designation is shortened to either 26-hydroxycholesterol or 27-hydroxycholesterol. In medical parlance, the designation 26-hydroxycholesterol was changed to 27-hydroxycholesterol in 1991 (7, 8). With the designation of the gene as CYP27 it is sensible to refer to the product as 27-hydroxycholesterol. However, Chemical Abstracts continues to index the sterol as 26-hydroxycholesterol when the stereospecificity is not known or as 25R,26- or 25S,26-hydroxycholesterol, respectively, when the stereospecificities are known.
Actually, the compound derived from natural sources contains both isomers after crystallization. The two enantiomorphs can be separated by chromatographic procedures (9) into mostly 25R,26-hydroxycholesterol with a small amount of 25S,26-hydroxycholesterol. Chemical synthesis, which essentially consists of attaching a stereospecific side chain to the steroid nucleus, can produce only the naturally occurring 25R-isomer (10).
Fredrickson and Ono (11) went on to show that 25R,26-hydroxycholesterol was metabolized to chenodeoxycholic acid in the rat, an exception to the then current paradigm (12). Danielsson (13) repeated these studies and confirmed the correctness of the precursor/product relationship for chenodeoxycholic acid but excluded 25-hydroxycholesterol as an endogenous metabolite, since its occurrence could be accounted for entirely by auto-oxidation.
Using the same approach to the synthesis of 25R,26-hydroxycholesterol as Scheer, Thompson, and Mosettig, Sidney Emerman, and I prepared a radioactive compound of much higher specific activity (14) and found that in the rat it was metabolized to both chenodeoxycholic and cholic acid, although the latter was present in much smaller amounts.
Thus, it was established in both rodents and humans (15) that bile acids could be derived from cholesterol beginning with the oxidation of the terminal carbon atom. However, the existence in both rats and mice of an alternate pathway of bile acid synthesis that yields muricholic rather than cholic acid (16) skewed the precursor/product relationships. These species differences in the genes that govern bile acid production continue to limit the usefulness of knockout mouse models for relating to the changes that occur to the phenotypic expressions as they occur in humans.
|
Overview of the cholesterol 27-hydroxylase metabolic pathway |
|---|
|
TOPABSTRACTINTRODUCTIONOverview of the cholesterol...REFERENCES |
|---|
-hydroxylase expressed in the endoplasmic reticulum (Fig. 1)
. The possibility of a reverse sequence, initial mitochondrial 27-hydroxylation of cholesterol followed by 7-hydroxylation was excluded because cholic acid was not identified as a metabolite of 3ß-hydroxy-5-cholenoic acid in the rat (12, 17). Also, the purified 27-hydroxylase enzyme oxidized the 7-hydroxylated sterol intermediates at rates more than 100-fold greater than that of cholesterol, implying that the latter was not a physiologic substrate (18).
|
-hydroxylase (20–22) and cholesterol 27-hydroxylase (23). Thus, the more recent data could support the view that both metabolic routes are normally expressed in hepatocytes.
However, in the last several years we have learned that steroid acute response (StAR) protein normally expressed in the adrenal gland but not the liver is essential for the normal rate of delivery of cholesterol to the inner mitochondrial membrane for 24S-hydroxylation (24). A similar role for StAR protein can be demonstrated in cell culture for 27-hydroxylation of cholesterol but not 7-hydroxylated intermediates in bile acid synthesis. (25). Thus, because the liver normally lacks StaR protein it is possible that little if any 27-hydroxylation of cholesterol occurs in hepatocytes in vivo.
The subcellular routing that occurs following the generation of mitochondrial 27-hydroxycholesterol is less clear. Because the enzyme is multifunctional and also generates the C27 acid, 3ß-hydroxy-5-cholestenoic acid (8), it might be assumed that all the generated 27-hydroxycholesterol undergoes successive oxidation to the C27 acid before leaving the mitochondria, referred to as the "acidic" pathway (26). The monohydroxy C27 bile acid is then metabolized to 3ß-hydroxy-5-cholenoic acid, a precursor for chenodeoxycholic acid in peroxisomes. However, when the C27 bile acid is administered to rabbits, less than 5% is metabolized to cholic acid (27). By contrast, 27-hydroxycholesterol administered to rabbits (27) and humans (15) yields approximately 90% and 40% cholic acid, respectively. Thus it seems likely that not all the 27-hydroxycholesterol that may occur in hepatic mitochondria is further oxidized to the C27 acid but that some undergoes 7-hydroxylation prior to further side chain oxidation. This view is consistent with cell culture studies in which 27-hydroxycholesterol appears in the medium before the C27 acid (28), indicating that it is released from the enzyme site before complete oxidation.
Another subcellular route depicted in Fig. 1 derives from the finding that the 27-hydroxylase is widely expressed in tissues (7), including vascular endothelium (28) and macrophages (29). Most of the 27-hydroxycholesterol that is generated is transported in plasma esterified with fatty acids and together with esterified cholesterol is part of LDL and HDL (30). Following receptor mediated uptake and the generation of the free sterols by well-defined mechanisms, 7-hydroxylation occurs in the endoplasmic reticulum. Further transformation to 5ß-cholestane-3,7,27-triol and 3,7,12-27-tetrol occurs prior to side chain oxidation in the mitochondria to form C27 bile acids. Further oxidation and shortening of the side chain occurs in peroxisomes to form chenodeoxycholic and cholic acids, respectively.
Recognition that a different 7-hydroxylase (20) determines the production of mostly chenodeoxycholic acid from 27-hydroxycholesterol established the existence of two different but interdependent pathways (31). Thus, chenodeoxycholic acid is a potent ligand for farnesoid X activated receptors (FXRs), which modulates cholesterol 7-hydroxylase activity (32), and therefore the amount of bile acid generated from cholesterol in the liver.
Initial interest in the biological effect of 27-hydroxycholesterol stemmed from the knowledge that it can downregulate cholesterol synthesis (33) at concentrations that could occur intracellularly. Recent studies of the mechanism for downregulation indicate that it is mediated by interruption of the transport of sterol regulatory binding protein by blocking sterol activating cleavage protein-mediated transport (34). Because 27-hydroxycholesterol is rapidly 7-hydroxylated in the liver and the triol no longer downregulates cholesterol synthesis (35), it is difficult to know if 27-hydroxycholesterol has a physiologic role in regulating cholesterol synthesis in the liver. It is known that the activities of both HMG-CoA reductase and cholesterol 7-hydroxylase are greater than normal in the livers of patients who do not generate 27-hydroxycholesterol because of a mutant CYP27 gene. However, feeding chenodeoxycholic acid to these patients returns the activities toward normal, an event that presumably is not mediated by 27-hydroxycholesterol. The finding implies that the higher rate of cholesterol synthesis in the liver than that in most other tissues is related to bile acid production and that this interrelationship is governed by determinants that are independent of oxysterol regulation.
To fully evaluate the role of the cholesterol 27-hydroxylase metabolic pathway, quantitative data are needed with respect to total production of 27-hydroxycholesterol, the tissues of origin, the biologic roles of the intermediates that are generated in these tissues, and the biologic effects of the bile acid end products that are generated in the liver. How to achieve these goals remains an unsolved problem. Some in vivo kinetic data have been obtained (36) using techniques initially developed for determining steroid hormone production (37), but further validation is needed. In these limited studies in older adults (36), the cholesterol 27-hydroxylation pathway accounts for approximately 10% of the daily production of chenodeoxycholic and cholic acids.
Monohydroxy bile acid synthesis
Synthesis of the C27 bile acid, 3ß-hydroxy-5-cholestenoic acid is unique to the cholesterol 27-hydroxylase metabolic pathway and it is probably the the sole source of the C24 bile acid, 3ß-hydroxy-5-cholenoic acid, although one cannot exclude a very minor contribution from 24S-hydroxycholesterol (38). Both monohydroxy bile acids normally circulate in plasma in nanomolar amounts, but only the latter has been reported in amniotic fluid, meconium, bile, and urine (39–42). Presumably the C27 acid undergoes further oxidation to the C24 acid in peroxisomes, perhaps exclusively in the liver, since the C24 acid was not identified as a metabolite when 27-hydroxycholesterol was incubated with nonhepatic tissues (28).
Other metabolic fates are 7-hydroxylation to 3ß,7-dihydroxy-5-cholenoic acid, which is further transformed to chenodeoxycholic acid (43, 44), or direct ring transformation to the monohydroxy bile acid 3-hydroxy-5ß-cholanoic acid (lithocholic acid). The latter bile acid, which is a primary bile acid in the newborn period (45), also derives later in life by bacterial 7-dehydroxylation of chenodeoxycholic acid in the intestines. In urine monohydroxy, bile acids are found almost exclusively as ester sulfates (41).
The interrelationships of the two metabolic pathways in neonatal life are illustrated by the CYP7 knockout mouse (46). A transient neonatal cholestatic syndrome occurs because oxysterol 7-hydroxylase activity is not normally expressed in neonatal mice. Absence of both oxysterol and cholesterol 7-hydroxylase activities yields only monohydroxy bile acids. That these events in the mouse do not regularly occur in human neonates must be attributed to the expression of CYP7B in fetal life since in humans, in contrast to mice, the CYP7A gene is not expressed (47).
Bile acid synthesis in fetal and neonatal life
Relatively little data are available in humans to generate a complete picture of bile acid production during fetal and neonatal life. One needs to consider three sources: placental transfer, initial 7-hydroxylation, and initial 27-hydroxylation.
Although bile acids normally circulate in maternal plasma mostly bound to albumin, exchange of the unbound species across the maternal/fetal capillary circulation can occur and probably accounts for the small amounts of deoxycholic acid found in sterile meconium at birth (48). However, the high proportion of chenodeoxycholic acid in fetal gallbladder bile in early pregnancy does not mimic the composition of maternal bile, and therefore the accumulation of a fetal bile acid pool cannot be attributed entirely to placental exchange.
The knowledge that amniotic fluid in early pregnancy contains relatively large amounts of monohydroxy bile acids (39) indicates the existence of a metabolic pathway that begins with C27 hydroxylation. Expression of the CYP27 gene during fetal life can also be inferred from the finding of monohydroxy bile acids in neonatal urine (41, 42). Oxysterol 7-hydroxylase but not cholesterol 7-hydroxylase is also normally expressed in fetal liver (49). Thus, one can conclude that the neonatal pool of bile acids is derived both via placental transfer and the cholesterol 27-hydroxlation metabolic pathway. Further evidence for this conclusion can also be derived from the statement by Setchell and coworkers that they cannot detect expression of CYP7 in normal neonates for at least the initial few months of life (47). When in the course of normal development the cholesterol 7-hydroxylase pathway begins to contribute to bile acid production and then dominate total bile acid production is not known.
Since the cholesterol 27-hydroxylation metabolic pathway will generate only monohydroxy bile acids unless 7-hydroxylation occurs, the status of expression of CYP7B becomes critical to the proportion of monohydroxy bile acids that are formed. Indeed, two instances of absent oxysterol 7-hydroxylase activity have been associated with severe persistent cholestatic liver disease (47, 50). The severity of the cholestasis depends not only on oxysterol 7-hydroxylase activity, but also on that of cholesterol 7-hydroxylase, since the production of chenodeoxycholic and cholic acids via this latter pathway moderates the cholestatic effect of monohydroxy bile acids (51).
Postnatal bile acid synthesis
Because of the high extraction efficiency of hepatocytes for bile acids returning via the portal vein, the circulating pool of bile acids is confined for the most part to the enterohepatic circulation and supports three major functions. A) Bile acid dependent bile flow is essential for the excretion of endogenous metabolites such as bilirubin glucuronide and many xenobiotics, as well as to maintain in solution the relatively high concentration of cholesterol characteristic of human bile (52); B) Bile acids are essential for the absorption of adequate amounts of vitamin E and other fat-soluble vitamins as well as for the hydrolysis of fat and absorption of long-chain fatty acids; and C) Perhaps less essential but nevertheless important is the role of bile acids in regulating water reabsorption in the ascending colon (53).
Role of the cholesterol 7-hydroxylase metabolic pathway
These biologic roles are maintained by an auto-regulatory process that focuses on the regulation of the activity of cholesterol 7-hydroxylase. Thus, interruption of the enterohepatic circulation by ileal disease, bile fistula, or bile acid sequestrant administration, all of which threaten to reduce bile acid pool size, result in increased enzyme activity to maintain homeostasis. Artifical expansion of pool size by chenodeoxycholic acid administration is a potent mechanism for decreasing enzyme activity (54). This knowledge coupled with the newer information concerning FXR/short heterodimer protein (SHP) nuclear receptor regulation (55) and its relationship to chenodeoxycholic and deoxycholic acids (56) focuses on transcriptional regulation of the CYP7A gene as the dominant but not exclusive mechanism.
Regulation of bile acid pool size and synthesis by this mechanism is essentially independent of dietary cholesterol and depends mostly on the upregulation of cholesterol synthesis in the liver to satisfy the demand for increased bile acid synthesis. However, pharmacologic manipulation of the pathway can adapt it as a mechanism for lowering LDL cholesterol (LDL-C). Thus, suppressing HMG-CoA reductase activity in the liver by the administration of allosteric inhibitors makes it possible to enhance receptor-mediated LDL-C delivery to the enzyme as a source for bile acid production in lieu of de novo synthesis.
Roles of the cholesterol 27-hydroxylase metabolic pathway
Because adult cyp7A knockout mice appear phenotypically normal (46, 57), it could be argued that the cholesterol 27-hydroxylase metabolic pathway can replace all the essential functions, particularly the maintenance of bile acid dependent bile flow. However, until a human counterpart of the knockout cyp7A mouse is identified, one should reserve judgment with respect to the importance of this pathway in human physiology.
Because of species differences in bile acid synthesis (16), knocking out the cyp27 gene in the mouse does not result in the phenotypic expression of cerebrotendinous xanthomas characteristic of the human inherited disease.
The markedly accelerated atherosclerosis in some patients with a mutant CYP27 gene can now be explained at least in part by the loss of 27-hydroxycholesterol/LXR-mediated cholesterol efflux from macrophages as part of reverse cholesterol transport (58). The major question that now requires resolution is how this inherited defect, which affects all cells normally expressing the gene, relates to acquired defects that could occur in selective tissues such as macrophages.
Reverse cholesterol transport
ABC A1 transporter activity
Since the recognition that the molecular basis of Tangier disease is a mutant ABC A1 transporter (59), progress has been very rapid (60, 61). 27-hydroxycholesterol generated in macrophages has been identified as an important endogenous ligand for up-regulating cholesterol efflux via this ABC transporter pathway (58).
Metabolism of cholesterol to 27-hydroxycholesterol in macrophages The rate of conversion of cholesterol to 27-hydroxycholesterol followed by its further metabolism to other intermediates in the bile acid biosynthetic pathway represents a mechanism for returning cholesterol to the liver that is independent of its role of 27-hydroxycholesterol as a ligand for LXR-mediated cholesterol efflux via the ABC A1 transporter.
It is of particular interest, therefore, that the 27-hydroxycholesterol concentration in atheroma obtained from the carotid and coronary arteries and the aorta is much greater than that in plasma (62). Since the major cellular components of atheroma are macrophages that express the CYP27 gene (63), it is reasonable to think that they are the source of this hydroxysterol and account for the much greater ratio of 27-hydroxycholesterol/cholesterol in atheroma than in plasma. The concentration gradient between atheroma and plasma indicates a block in reverse cholesterol transport that is puzzling and may affect the natural history of atheroma. Thus, the local concentration of 27-hydroxycholesterol when expressed per weight of tissue is greater than that needed to initiate apoptosis in cell culture (64) and sufficient to inhibit smooth muscle cell migration (65). On the other hand we do not know what proportion of the 27-hydroxycholesterol may be esterified with fatty acid and/or sulfated and what the local biologic effects of these different naturally occurring derivatives are. It could be argued that the local activity of oxysterol 7-hydroxylase should be equivalent to that of cholesterol 27-hydroxylase. The more water-soluble triol would then enter plasma. Currently, it is not possible to evaluate either the potentially beneficial or the potentially deleterious effects of 27-hydroxycholesterol in atheroma. A major challenge is to design critical studies in an experimental model that might begin to give insights on the biologic significance of 27-hydroxycholesterol and the other steroid intermediates that form locally.
Other ligands regulating gene expression Thus far only the biologic role of 27-hydroxycholesterol as a ligand has been studied in detail. One study indicates that 3ß-hydroxy-5-cholenstenoic acid also may function as a ligand (66). Table 1 indicates the number of intermediates exclusive of fatty acid esters that can be generated by and are unique to the cholesterol 27-hydroxylase metabolic pathway. With the exception of 3ß-hydroxy-5-cholestenoic acid, their possible role as ligands for nuclear receptors have not been evaluated.
|
|
ACKNOWLEDGMENTS |
|---|
Manuscript received December 5, 2001
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONOverview of the cholesterol...REFERENCES |
|---|
,7,12-triol 27-hydroxylase from female rat liver mitochondria. J. Biol. Chem. 263: 18138–18142.-hydroxylation of 27-hydroxycholesterol: biologic role in the regulation of cholesterol synthesis. J. Lipid Res. 37: 1053–1058.
-hydroxylase activity. Gastroenterology. 97: 1498–1505.[Medline]
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  H. Fuda, N. B. Javitt, K. Mitamura, S. Ikegawa, and C. A. Strott Oxysterols are substrates for cholesterol sulfotransferase J. Lipid Res., June 1, 2007; 48(6): 1343 - 1352. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Ren, P. Hylemon, Z.-P. Zhang, D. Rodriguez-Agudo, D. Marques, X. Li, H. Zhou, G. Gil, and W. M. Pandak Identification of a novel sulfonated oxysterol, 5-cholesten-3{beta},25-diol 3-sulfonate, in hepatocyte nuclei and mitochondria J. Lipid Res., May 1, 2006; 47(5): 1081 - 1090. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. A. Pikuleva CHOLESTEROL-METABOLIZING CYTOCHROMES P450 Drug Metab. Dispos., April 1, 2006; 34(4): 513 - 520. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. F. Oram and J. W. Heinecke ATP-Binding Cassette Transporter A1: A Cell Cholesterol Exporter That Protects Against Cardiovascular Disease Physiol Rev, October 1, 2005; 85(4): 1343 - 1372. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Meaney Is C-26 hydroxylation an evolutionarily conserved steroid inactivation mechanism? FASEB J, August 1, 2005; 19(10): 1220 - 1224. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Dubrac, S. R. Lear, M. Ananthanarayanan, N. Balasubramaniyan, J. Bollineni, S. Shefer, H. Hyogo, D. E. Cohen, P. J. Blanche, R. M. Krauss, et al. Role of CYP27A in cholesterol and bile acid metabolism J. Lipid Res., January 1, 2005; 46(1): 76 - 85. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Nishimaki-Mogami, M. Une, T. Fujino, Y. Sato, N. Tamehiro, Y. Kawahara, K. Shudo, and K. Inoue Identification of intermediates in the bile acid synthetic pathway as ligands for the farnesoid X receptor J. Lipid Res., August 1, 2004; 45(8): 1538 - 1545. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. F. Oram HDL Apolipoproteins and ABCA1: Partners in the Removal of Excess Cellular Cholesterol Arterioscler. Thromb. Vasc. Biol., May 1, 2003; 23(5): 720 - 727. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Journal of Biological Chemistry |
| Molecular and Cellular Proteomics | ASBMB Today |
