Abstract-Although an immense knowledge has accumulated concerning regulation of cholesterol homeostasis in the body, this does not include the brain, where details are just emerging. Approximately 25% of the total amount of the cholesterol present in humans is localized to this organ, most of it present in myelin. Almost all brain cholesterol is a product of local synthesis, with the blood-brain barrier efficiently protecting it from exchange with lipoprotein cholesterol in the circulation. Thus, there is a highly efficient apolipoprotein-dependent recycling of cholesterol in the brain, with minimal losses to the circulation. Under steady-state conditions, most of the de novo synthesis of cholesterol in the brain appears to be balanced by excretion of the cytochrome P-450 -generated oxysterol 24S-hydroxycholesterol. This oxysterol is capable of escaping the recycling mechanism and traversing the blood-brain barrier. Cholesterol levels and cholesterol turnover are affected in neurodegenerating disorders, and the capacity for cholesterol transport and recycling in the brain seems to be of importance for the development of such diseases. The possibility has been discussed that administration of inhibitors of cholesterol synthesis may reduce the prevalence of Alzheimer disease. No firm conclusions can, however, be drawn from the studies presented thus far.In the present review, the most recent advances in our understanding of cholesterol turnover in the brain is discussed. Key Words: brain cholesterol Ⅲ blood-brain barrier Ⅲ cholesterol 24S-hydroxylase Ⅲ Alzheimer disease Ⅲ statins H ighly sophisticated regulatory systems have evolved for the maintenance of cholesterol homeostasis in the body. There is a distinct difference between the situation in the central nervous system and that in most other extrahepatic tissues. Outside the brain, the cellular needs for cholesterol is covered by de novo synthesis and by cellular uptake of lipoprotein cholesterol from the circulation. In the brain, the blood-brain barrier effectively prevents uptake from the circulation, and de novo synthesis is responsible for practically all cholesterol present in this organ. The independence of the isolated pool of cholesterol in the brain is likely to reflect a high need for constancy in the cholesterol content of membrane and myelin, a constancy that would be difficult to keep if brain cholesterol had been exchangeable with lipoprotein cholesterol.The importance of cholesterol in the nervous system was recognized as early as 1834, when Couerbe's observations lead him to regard cholesterol as un element principal of the nervous system. 1 Despite concerted efforts in the interim, it is only during the past few decades that the brain has begun to surrender the secrets of the behavior of one of its most abundant lipids.In the present brief review, we summarize the basal characteristics of brain cholesterol and some recent findings with respect to homeostasis of this compound in the brain. Emphasis is put on the newly described relation betwee...

Side chain oxidized oxysterols have a unique ability to traverse lipophilic membranes. We tested the hypothesis that there is a net flux of 27-hydroxycholesterol from the circulation into the brain using plasma samples collected from the internal jugular vein and an artery of healthy male volunteers. Two independent studies were performed, one in which total levels of 27-hydroxycholesterol were measured and one in which the free fraction of 27-hydroxycholesterol was measured. In the majority of subjects studied, the level of 27-hydroxycholesterol was higher in the artery than in the vein, and uptake from the circulation was calculated to be about 5 mg/24 h. The distribution of 27-hydroxycholesterol in human brain was found to be consistent with an extracerebral origin, with a concentration gradient from the white to the gray matter-a situation opposite that of 24S-hydroxycholesterol, which is exclusively formed in brain. In view of the fact that the blood-brain barrier is impermeable to cholesterol and that 27-hydroxycholesterol is a potent regulator of several cholesterol-sensitive genes, the flux of 27-hydroxycholesterol into the brain may be an important link between intraand extracerebral cholesterol homeostasis.

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Mammalian CNS contains a disproportionally large and remarkably stable pool of cholesterol. Despite an efficient recycling there is some requirement for elimination of brain cholesterol. Conversion of cholesterol into 24S-hydroxycholesterol by the cholesterol 24-hydroxylase (CYP46A1) is the quantitatively most important mechanism. Based on the protein expression and plasma levels of 24S-hydroxycholesterol, CYP46A1 activity appears to be highly stable in adults. Here we have made a structural and functional characterization of the promoter of the human CYP46A1 gene. No canonical TATA or CAAT boxes were found in the promoter region. Moreover this region had a high GC content, a feature often found in genes considered to have a largely housekeeping function. A broad spectrum of regulatory axes using a variety of promoter constructs did not result in a significant transcriptional regulation. Oxidative stress caused a significant increase in transcriptional activity. The possibility of a substrate-dependent transcriptional regulation was explored in vivo in a sterol-deficient mouse model (Dhcr24 null) in which almost all cholesterol had been replaced with desmosterol, which is not a substrate for CYP46A1. Compared with heterozygous littermates there was no statistically significant difference in the mRNA levels of Cyp46a1. During the first 2 weeks of life in the wild-type mouse, however, a significant increase of Cyp46a1 mRNA levels was found, in parallel with an increase in 24S-hydroxycholesterol level and a reduction of cholesterol synthesis. The failure to demonstrate a significant transcriptional regulation under most conditions is discussed in relation to the turnover of brain and neuronal cholesterol.Although the brain is the most cholesterol-rich organ in the body, relatively little is known about the mechanisms by which it maintains steady-state cholesterol levels (1, 2). This is in marked contrast to the situation in virtually every other tissue or organ. One finding that has been consistently confirmed is that, due to the efficiency of the bloodbrain barrier, the brain is unable to take up cholesterol from the circulation and relies on de novo synthesis to meet its substantial cholesterol requirements. However, the rate of cholesterol synthesis in the adult brain is very low, and the bulk of brain cholesterol has a half-life that is at least 100 times longer than that of cholesterol in most other organs (3).One consequence of this "uncoupling" of brain and whole body cholesterol homeostasis has been the evolution of specific mechanisms for maintenance of cerebral cholesterol levels. Two mechanisms for removal of brain cholesterol are currently recognized (1). The first is analogous to classic "reverse cholesterol transport" and is mediated by a flux of cholesterol present in apolipoprotein E containing lipoproteins through cerebrospinal fluid into the circulation (4, 5). In adults, this mechanism is believed to be responsible for elimination of 1-2 mg of cholesterol per 24 h. The details of this particular ...

Recently, we demonstrated a net blood-to-brain passage of the oxysterol 27-hydroxycholesterol corresponding to 4-5 mg/day. As the steady-state levels of this sterol are only 1-2 mg/g brain tissue, we hypothesized that it is metabolized and subsequently eliminated from the brain. To explore this concept, we first measured the capacity of in vitro systems representing the major cell populations found in the brain to metabolize 27-hydroxycholesterol. We show here that 27-hydroxycholesterol is metabolized into the known C 27 steroidal acid 7a-hydroxy-3-oxo-4-cholestenoic acid by neuronal cell models only. Using an in vitro model of the blood-brain barrier, we demonstrate that 7a-hydroxy-3-oxo-4-cholestenoic acid is efficiently transferred across monolayers of primary brain microvascular endothelial cells. Finally, we measured the concentration of 7a-hydroxy-3-oxo-4-cholestenoic acid in plasma from the internal jugular vein and brachial artery of healthy volunteers. Calculation of the arteriovenous concentration difference revealed a significant in vivo flux of this steroid from the brain into the circulation in human.Together, these studies identify a novel metabolic route for the elimination of 27-hydroxylated sterols from the brain. Given the emerging connections between cholesterol and neurodegeneration, this pathway may be of importance for the development of these conditions.

The side chain oxidized oxysterol 24 S -hydroxycholesterol (24-OH-chol) is formed almost exclusively in the brain, and there is a continuous passage of this oxysterol through the circulation to the liver. 27-Hydroxycholesterol (27-OH-chol) is produced in most organs and is also taken up by the liver. The 27-OH-chol-24-OH-chol ratio is about 0.1 in the brain and about 2 in the circulation. This ratio was found to be about 0.4 in cerebrospinal fluid (CSF) of asymptomatic patients, consistent with a major contribution from the circulation in the case of 27-OH-chol. In accordance with this, we demonstrated a significant flux of deuterium labeled 27-OH-chol from plasma to the CSF in a healthy volunteer. Patients with a defective blood-brain barrier were found to have markedly increased absolute levels (up to 10-fold) of both 27-OH-chol and 24-OH-chol in CSF, with a ratio between the two sterols reaching up to 2. There was a significant positive correlation between the levels of both oxysterols in CSF and the albumin CSF -albumin plasma ratio. The 27-OH-chol CSF -24-OH-chol CSF ratio was found to be about normal in patients with active multiple sclerosis and significantly increased in patients with meningitis, polyneuropathy, or hemorrhages. 24 S -Hydroxycholesterol (24-OH-chol) is almost exclusively formed in the brain, where it is present in greater amounts than in any other organ (8.6-15.1 ng/mg wet weight)(1). There is a daily flux of about 7 mg of this oxysterol from the brain to the circulation, with the majority of this efflux apparently occurring as direct transport across the blood-brain barrier (BBB) (2). It has been estimated that less than 1% of the 24-OH-chol produced by the brain is transported to the circulation via passage through the cerebrospinal fluid (CSF) (1). The enzyme responsible for the 24 S -hydroxylation of cholesterol is a member of the cytochrome P450 superfamily (designated CYP46), and has been mainly localized to neurons (3). Assuming that the expression of this enzyme across the neuronal population is relatively stable, any loss of these cells would result in a decreased production of this oxysterol. In accordance with this, reduced levels of plasma 24-OHchol have been observed in connection with several chronic neurological conditions known to affect the number of neurons (4).In contrast to the above oxysterol, 27-hydroxycholesterol (27-OH-chol) is formed in most cells, and there is a constant flux of this oxysterol from extrahaepatic tissues to the liver (5, 6). A noteworthy exception is the brain (no net flux has been observed from this organ) and the levels of 27-OH-chol are about 10-fold lower than those of 24-OH-chol (1). However, the plasma levels of 27-OH-chol are about twice those of 24-OH-chol.The term BBB is commonly used to describe a whole range of mechanisms that regulate and protect the internal environment in the brain (7-11). More specifically, this term is used to indicate a relative restriction of the entry of the plasma proteins into the brain. This barrier a...

Oxysterols possess powerful biological activities. Some of their effects on the regulation of key enzymes are similar to those of cholesterol, but are much more potent. One of the critical properties of oxysterols is their ability to pass lipophilic membranes at a high rate. Transfer of unesterified 25-hydroxycholesterol from red blood cells to plasma has been reported to occur more than 1,000 times faster than cholesterol. Here we have measured the relative rate of such translocation of the three major oxysterols in human circulation: 27-hydroxycholesterol, 24S-hydroxycholesterol, and 4 ␤ -hydroxycholesterol. The distance from the 3 ␤ -hydroxyl group to the additional hydroxyl group is the greatest possible in 27-hydroxycholesterol and the least possible in 4 ␤ -hydroxycholesterol. The rate of exchange between erythrocytes and plasma was found to be high for 27-hydroxycholesterol and 24S-hydroxycholesterol, and hardly possible to measure for 4 ␤ -hydroxycholesterol and cholesterol. When injected intravenously into humans, deuterium labeled 24-and 27-hydroxycholesterol caused an immediate high enrichment of the corresponding plasma sterols followed by a decay. After injection of labeled 4 ␤ -hydroxycholesterol, the maximum deuterium enrichment occurred after 2-3 h, when secretion of the oxysterol from the liver is likely to be the limiting factor. When radiolabeled cholesterol was injected under the same conditions, maximum appearance of label occurred after about 2 days. The results illustrate the importance of the position of the additional oxygen in oxysterols and are discussed in relation to the rate of metabolism and biological effects of these oxysterols.

There is a clear link between cholesterol turnover and neurodegenerative diseases and hypercholesterolemia is an established risk factor for Alzheimer's disease (AD). The failure to demonstrate a transfer of cholesterol from the circulation into the brain in humans and experimental animals makes it difficult to explain the link between hypercholesterolemia and AD. In contrast to cholesterol itself, side-chain oxidized cholesterol metabolites such as 24S-hydroxycholesterol and 27-hydroxycholesterol are able to pass the blood-brain barrier (BBB). Formation of 24S-hydroxycholesterol is the quantitatively most important mechanism for elimination of cholesterol from the brain and we recently demonstrated a significant net uptake of 27-hydroxycholesterol by the brain from the circulation. We have also shown that patients with AD have increased brain levels of 27-hydroxycholesterol, which may affect the production of beta-amyloid in the brain. The levels of 27-hydroxycholesterol in the circulation are correlated with the levels of cholesterol and the possibility must be considered that the flux of 27-hydroxycholesterol into the brain is the missing link between hypercholesterolemia and Alzheimer's disease. Current knowledge about the role of the two oxysterols for cholesterol homeostasis in the brain as well as their diagnostic potential are reviewed.