Neurodegeneration in the Niemann-Pick C mouse: glial involvement.

A mouse model of Niemann-Pick type C disease has been found that exhibits neuropathology similar to the human condition. There is an age-related neurodegeneration in several brain regions and a lack of myelin in the corpus callosum in these mice. The purpose of the present study was to examine the Niemann-Pick mouse and determine whether: (1) microglia and astrocytes exhibit ultrastructural pathology similar to that found in neurons; (2) nerve fiber number is reduced when the myelin sheath is absent; and (3) the lysosomal hydrolase, cathepsin-D, is involved in the neurodegenerative process. Using light and electron microscopic methods, and immunocytochemistry, Niemann-Pick and control animals were examined at several ages. Cathepsin-D content was semi-quantitatively measured in neurons and glial cells in brain regions known to exhibit neurodegeneration, as was the density of glial fibrillary acidic protein-labeled astrocytes. The Niemann-Pick mouse exhibited: (1) an age-related increase in inclusion bodies in microglia and astrocytes, similar to that observed within neurons; (2) an almost complete absence of myelin in the corpus callosum by 7-8 weeks of age, along with a 30% reduction in the number of corpus callosum axons; (3) a mild age-related increase in cathepsin-D content within nerve cells in many brain regions. However, the cathepsin-D elevation was greatest in microglial cells; (4) an age-related increase in the number of microglial cells containing intense cathepsin-D immunoreactivity in both the thalamus and cerebellum. Both of these brain regions have been shown previously to exhibit an age-related loss of neurons; and (5) an increase in the number of reactive astrocytes immunostained for glial fibrillary acidic protein, especially in the thalamus and cerebellum. These data indicate that glial cells are a major target for pathology in the Niemann-Pick mouse. The lack of myelin within the corpus callosum may be related to the loss of nerve fibers in this structure. The increase in cathepsin-D-laden microglial cells, in brain regions previously shown to undergo neurodegeneration, is consistent with a role for microglia in the phagocytosis of dead neurons and in actively contributing to the neurodegenerative process. The activation of astrocytes in regions that undergo neurodegeneration is also consistent with a role for these glial cells in the neurodegenerative process.

Alternate pathways of bile acid synthesis in the cholesterol 7alpha-hydroxylase knockout mouse are not upregulated by either cholesterol or cholestyramine feeding.

Bile acids are synthesized via the classic pathway initiated by cholesterol 7alpha-hydroxylase (CYP7A1), and via alternate pathways, one of which is initiated by sterol 27-hydroxylase (CYP27). These studies used mice lacking cholesterol 7alpha-hydroxylase (Cyp7a1(-/-)) to establish whether the loss of the classic pathway affected cholesterol homeostasis differently in males and females, and to determine if the rate of bile acid synthesis via alternate pathways was responsive to changes in the enterohepatic flux of cholesterol and bile acids. In both the Cyp7a1(-/-) males and females, the basal rate of bile acid synthesis was only half of that in matching Cyp7a1(+/+) animals. Although bile acid pool size contracted markedly in all the Cyp7a1(-/-) mice, the female Cyp7a1(-/-) mice maintained a larger, more cholic acid-rich pool than their male counterparts. Intestinal cholesterol absorption in the Cyp7a1(-/-) males fell from 46% to 3%, and in the matching females from 58% to 17%. Bile acid synthesis in Cyp7a1(+/+) males and females was increased 2-fold by cholesterol feeding, and 4-fold by cholestyramine treatment, but was not changed in matching Cyp7a1(-/-) mice by either of these manipulations. In the Cyp7a1(-/-) mice fed cholesterol, hepatic cholesterol concentrations increased only marginally in the males, but rose almost 3-fold in the females. CYP7A1 activity and mRNA levels were greater in females than in males, and were increased by cholesterol feeding in both sexes. CYP27 activity and mRNA levels did not vary as a function of CYP7A1 genotype, gender, or dietary cholesterol intake. We conclude that in the mouse the rate of bile acid synthesis via alternative pathways is unresponsive to changes in the enterohepatic flux of cholesterol and bile acid, and that factors governing gender-related differences in bile acid synthesis, pool size, and pool composition play an important role in determining the impact of CYP7A1 deficiency on cholesterol homeostasis in this species.

Degeneration of neurons and glia in the Niemann-Pick C mouse is unrelated to the low-density lipoprotein receptor.

The BALB/c mouse model of Niemann-Pick type C disease exhibits similar neuropathological features to the human condition, including cerebral atrophy, demyelination of the corpus callosum, and degeneration of cerebellar Purkinje cells. The gene defect in Niemann-Pick C disease causes cholesterol to accumulate within the lysosomal compartment of neurons and glial cells. In order to determine whether cholesterol accumulation through the low-density lipoprotein receptor pathway plays an important role in the degenerative process, Niemann-Pick C mice were crossed with low-density lipoprotein receptor knockout mice. The purpose of the present study was to determine whether degeneration of neurons and glial cells is reduced in Niemann-Pick C animals lacking the low-density lipoprotein receptor. Using stereological counting methods, Purkinje cells were counted in the cerebellum and glial cell bodies were counted in the corpus callosum in mice at 3, 7.5 and 11 weeks of age. In the Niemann-Pick C animals, compared to wild-type control mice, there were 48% fewer glial cells at 3 weeks of age, and by 11 weeks of age there were 63% fewer glial cells. Purkinje cells were decreased in number by 13% at 3 weeks of age, and by 11 weeks of age there was a 96% loss. In the Niemann-Pick C animals lacking low-density lipoprotein receptors, there was no difference in the magnitude of glial cell or Purkinje cell loss compared to the Niemann-Pick C animals. These data indicate that both neurons and glia are vulnerable to degeneration in the Niemann-Pick C mouse, but that blocking the accumulation of cholesterol through the low-density lipoprotein receptor pathway does not alter the degenerative phenotype of Niemann-Pick C disease.

Selective neurodegeneration, without neurofibrillary tangles, in a mouse model of Niemann-Pick C disease.

The BALB/c mouse model of Niemann-Pick type C (NPC) disease exhibits neuropathological similarities to the human condition. There is an age-related cerebral atrophy, demyelination of the corpus callosum, and degeneration of cerebellar Purkinje cells in the NPC mouse. In human NPC, many cortical and subcortical neurons contain neurofibrillary tangles, which are thought by some investigators to play an important role in the neurodegenerative process. The purpose of the present study was to determine whether neurodegeneration occurs in the NPC mouse, in brain regions other than the cerebellum and whether the degeneration is related to the presence of neurofibrillary tangles. Using light microscopic methods with immunohistochemistry, electron microscopy, and cell counting methods, 11-week-old NPC(+/+) and NPC(-/-) animals were examined. In the NPC(-/-) mice, there were 96% fewer Purkinje cells, 28% fewer neurons in the prefrontal cortex, 20% fewer neurons in the thalamus, and 63% fewer glial cells in the corpus callosum. On the other hand, previous studies indicate normal numbers of neurons and glial cells in these same neuroanatomical regions in young NPC(-/-) mice. There were normal numbers of cholinergic neurons in sections assessed in the striatum and basal forebrain in the 11-week-old animals and no evidence of neurofibrillary tangles within cells. The present data indicate that both neurons and glial cells die in the NPC mouse but that all cells are not equally vulnerable. There was no evidence for neurofibrillary tangles in the NPC mouse, and therefore the degenerative process in the mouse is unrelated to the neurofibrillary tangle.

Cholesterol metabolism in the brain.

The central nervous system accounts for only 2% of the whole body mass but contains almost a quarter of the unesterified cholesterol present in the whole individual. This sterol is largely present in two pools comprised of the cholesterol in the plasma membranes of glial cells and neurons and the cholesterol present in the specialized membranes of myelin. From 0.02% (human) to 0.4% (mouse) of the cholesterol in these pools turns over each day so that the absolute flux of sterol across the brain is only approximately 0.9% as rapid as the turnover of cholesterol in the whole body of these respective species. The input of cholesterol into the central nervous system comes almost entirely from in situ synthesis, and there is currently little evidence for the net transfer of sterol from the plasma into the brain of the fetus, newborn or adult. In the steady state in the adult, an equivalent amount of cholesterol must move out of the brain and this output is partly accounted for by the formation and excretion of 24S-hydroxycholesterol. This cholesterol turnover across the brain is increased in neurodegenerative disorders such as Alzheimer's disease and Niemann-Pick type C disease. Indirect evidence suggests that large amounts of cholesterol also turn over among the glial cells and neurons within the central nervous system during brain growth and neuron repair and remodelling. This internal recycling of sterol may involve ligands such as apolipoproteins E and AI, and one or more membrane transport proteins such as members of the low density lipoprotein receptor family. Changes in cholesterol balance across the whole body may, in some way, cause alterations in sterol recycling and apolipoprotein E expression within the central nervous system, which, in turn, may affect neuron and myelin integrity. Further elucidation of the processes controlling these events is very important to understand a variety of neurodegenerative disorders.

Induction of bile acid synthesis by cholesterol and cholestyramine feeding is unimpaired in mice deficient in apolipoprotein AI.

High density lipoprotein (HDL) cholesterol is believed to be preferentially utilized for bile acid synthesis and biliary secretion. In mice, the deletion of apolipoprotein AI (apo AI), the major apolipoprotein in HDL, results in very low plasma HDL-cholesterol levels. This article describes bile acid metabolism in apo AI-deficient (Apo AI(-/-)) mice and their C57BL/6 (Apo AI(+/+)) controls fed either a basal rodent diet alone or containing cholesterol or cholestyramine. Basal plasma HDL-cholesterol levels in the (-/-) mice (<10 mg/dL) were less than 20% of those in their (+/+) controls, but there were no phenotypic differences in either the relative cholesterol content of gallbladder bile, bile acid pool size and composition, fecal bile acid excretion or the activity of, or mRNA level for, cholesterol 7alpha-hydroxylase. However, compared with their (+/+) controls, the (-/-) mice absorbed more cholesterol (33 vs. 24%) and manifested lower rates of hepatic sterol synthesis (534 vs. 1,019 nmol/h per g). Cholesterol feeding increased hepatic cholesterol levels in the (+/+) animals from 2.7 to 4.4 mg/g and in the (-/-) mice from 2.6 to 8.1 mg/g. Bile acid synthesis increased 70% in both genotypes. Cholestyramine feeding stimulated bile acid synthesis 3.7 fold in both (-/-) and (+/+) mice. We conclude that the virtual loss of HDL-cholesterol from the circulation in apo AI deficiency has no impact on the ability of the hepatocyte to adapt its rate of bile acid synthesis in concert with the amount of cholesterol and bile acid returning to the liver from the small intestine.

Disruption of the sterol 27-hydroxylase gene in mice results in hepatomegaly and hypertriglyceridemia. Reversal by cholic acid feeding.

Sterol 27-hydroxylase (CYP27) participates in the conversion of cholesterol to bile acids. We examined lipid metabolism in mice lacking the Cyp27 gene. On normal rodent chow, Cyp27(-/-) mice have 40% larger livers, 45% larger adrenals, 2-fold higher hepatic and plasma triacylglycerol concentrations, a 70% higher rate of hepatic fatty acid synthesis, and a 70% increase in the ratio of oleic to stearic acid in the liver versus Cyp27(+/+) controls. In Cyp27(-/-) mice, cholesterol 7alpha-hydroxylase activity is increased 5-fold, but bile acid synthesis and pool size are 47 and 27%, respectively, of those in Cyp27(+/+) mice. Intestinal cholesterol absorption decreases from 54 to 4% in knockout mice, while fecal neutral sterol excretion increases 2.5-fold. A compensatory 2.5-fold increase in whole body cholesterol synthesis occurs in Cyp27(-/-) mice, principally in liver, adrenal, small intestine, lung, and spleen. The mRNA for the cholesterogenic transcription factor sterol regulatory element-binding protein-2 (SREBP-2) and mRNAs for SREBP-2-regulated cholesterol biosynthetic genes are elevated in livers of mutant mice. In addition, the mRNAs encoding the lipogenic transcription factor SREBP-1 and SREBP-1-regulated monounsaturated fatty acid biosynthetic enzymes are also increased. Hepatic synthesis of fatty acids and accumulation of triacylglycerols increases in Cyp27(-/-) mice and is associated with hypertriglyceridemia. Cholic acid feeding reverses hepatomegaly and hypertriglyceridemia but not adrenomegaly in Cyp27(-/-) mice. These studies confirm the importance of CYP27 in bile acid synthesis and they reveal an unexpected function of the enzyme in triacylglycerol metabolism.

Regulation of absorption and ABC1-mediated efflux of cholesterol by RXR heterodimers.

Several nuclear hormone receptors involved in lipid metabolism form obligate heterodimers with retinoid X receptors (RXRs) and are activated by RXR agonists such as rexinoids. Animals treated with rexinoids exhibited marked changes in cholesterol balance, including inhibition of cholesterol absorption and repressed bile acid synthesis. Studies with receptor-selective agonists revealed that oxysterol receptors (LXRs) and the bile acid receptor (FXR) are the RXR heterodimeric partners that mediate these effects by regulating expression of the reverse cholesterol transporter, ABC1, and the rate-limiting enzyme of bile acid synthesis, CYP7A1, respectively. Thus, these RXR heterodimers serve as key regulators of cholesterol homeostasis by governing reverse cholesterol transport from peripheral tissues, bile acid synthesis in liver, and cholesterol absorption in intestine.

Centripetal cholesterol flow from the extrahepatic organs through the liver is normal in mice with mutated Niemann-Pick type C protein (NPC1).

Niemann-Pick type C (NPC) protein functions to move unesterified cholesterol from the lysosomal compartment to other intracellular sites for further metabolism and/or excretion. This cholesterol is brought into the cell through the coated-pit pathway and accumulates in the lysosomes when NPC protein is mutated. The present study quantitated the alternative uptake process that brings cholesterol into the cell through the scavenger receptor, class B, type I (SR-BI) pathway in animals with this mutation. In homozygous NPC mice, the tissues of the extrahepatic compartment accumulated an excess of 14 mg of cholesterol each day per kg body weight, and synthesis increased by a similar amount (to 111 mg/day per kg) to compensate for this functional loss of sterol through lysosomal sequestration. An amount of cholesterol (108 mg/day per kg) nearly equal to that synthesized in the extrahepatic compartment was carried through the circulation by high density lipoprotein (HDL) and taken up by the liver. The rate of hepatic cholesterol excretion from the NPC mice as fecal acidic (65 mg/day per kg) and neutral (85 mg/day per kg) sterols was elevated 61% above control values and was accounted for by the total amount of cholesterol brought to the liver in HDL and synthesized in the hepatocytes. These studies demonstrated that while cholesterol entering tissues of the NPC animals through the coated-pit pathway became sequestered in the lysosomal compartment and was metabolically inactive, cholesterol that was newly synthesized or that entered cells through the SR-BI pathway was metabolized and excreted normally.

Cholesterol is sequestered in the brains of mice with Niemann-Pick type C disease but turnover is increased.

In Niemann-Pick Type C (NPC) disease, the concentration of cholesterol increases with age in every tissue except the brain. This study investigates whether accumulation of cholesterol might also occur within the cells of the central nervous system (CNS), but be obscured by the simultaneous loss of sterol from myelin as neurodegeneration proceeds. At birth, when there is little myelin in the CNS, the concentration of cholesterol is significantly elevated in every region of the brain in the homozygous NPC mouse. At 7 wk of age, myelination is nearly complete. In the NPC mouse, however, there is striking neurodegeneration and a reduction in both myelin protein and myelin cholesterol. Furthermore, net loss of cholesterol from the CNS is much higher in the NPC mouse than in the control animal (2.23 versus 1.37 mg/day per kg) so that the concentration of sterol in most regions of the brain is reduced. This neurodegeneration and loss of myelin cholesterol is not prevented by deletion of either the low-density lipoprotein receptor or apolipoprotein E in the NPC animal. Thus, the cholesterol sequestration seen in every organ in NPC disease also occurs in cells of the CNS and may be etiologically related to the neurodegeneration.

Cholesterol accumulation in tissues of the Niemann-pick type C mouse is determined by the rate of lipoprotein-cholesterol uptake through the coated-pit pathway in each organ.

Niemann-Pick type C (NPC) disease is associated with the accumulation of unesterified cholesterol in nearly all tissues and with progressive neurodegeneration. A murine model of this disease, the NPC mouse, was used to determine whether this sequestered cholesterol represented sterol carried in low density lipoprotein (LDL) and chylomicrons (CMs) taken up into the tissues through the coated-pit pathway. By 7 weeks of age, the sterol pool in the NPC mice had increased from 2,165 to 5,669 mg/kg body weight because of the daily sequestration of 67 mg of cholesterol per kg in the various organs. This was 7-fold greater than the rate of accumulation in control mice. The rate of LDL clearance in the NPC mouse was normal (523 ml/day per kg) and accounted for the uptake of 78 mg/day per kg of cholesterol in LDL whereas 8 mg/day per kg was taken up from CMs. Deletion of the LDL receptor in NPC mice altered the concentration of unesterified cholesterol in every organ in a manner consistent with the changes also observed in the rate of LDL cholesterol uptake in those tissues. Similarly, altering the flow of cholesterol to the liver through the CM pathway changed the concentration of unesterified cholesterol in that organ. Together, these observations strongly support the conclusion that, in NPC disease, it is cholesterol carried in LDL and CMs that is sequestered in the tissues and not sterol that is newly synthesized and carried in high density lipoprotein.

Genetic differences in cholesterol absorption in 129/Sv and C57BL/6 mice: effect on cholesterol responsiveness.

This study compared the cholesterolemic response of two strains of mice with genetically determined differences in cholesterol absorption. When fed a basal low-cholesterol diet, 129/Sv mice absorbed cholesterol twice as efficiently as did C57BL/6 mice (44% vs. 20%). Total lipid absorption, in contrast, averaged 80-82% in both strains. The higher level of cholesterol absorption in the 129/Sv animals was reflected in an adaptive reduction in hepatic and intestinal sterol synthesis. When fed lipid-enriched diets, the 129/Sv mice became significantly more hypercholesterolemic and had twofold higher hepatic cholesterol concentrations than did the C57BL/6 animals even though the conversion of cholesterol to bile acids was stimulated equally in both strains. The difference in cholesterol absorption between these mouse strains was not the result of physicochemical factors relating to the size and composition of the intestinal bile acid pool but more likely reflects an inherited difference in one or more of the biochemical steps that facilitate the translocation of sterol across the epithelial cell.

Cholesterol balance and metabolism in mice with loss of function of Niemann-Pick C protein.

Type C Niemann-Pick disease is due to a mutation in Niemann-Pick C (NPC) protein, a putative determinant of intracellular cholesterol transport. This study quantifies cholesterol balance in vivo across all tissues in mice with this defect. Cholesterol balance in the heterozygous animal is normal, but in the homozygous mouse the whole animal cholesterol pool expands continuously from birth, reaching 5, 442 mg/kg at 7 wk. The size of this pool in each organ is proportional to the rate at which each tissue clears low-density lipoprotein-cholesterol. Despite this expansion, however, cholesterol synthesis is increased so that whole animal synthesis equals 180 mg. day-1. kg-1. Forcing additional cholesterol into the liver through the clathrin-coated pit pathway increases the hepatic cholesterol pool in control mice, all of which is esterified, while there is a much greater increase in this pool in mutant mice, all of which is unesterified. These findings are consistent with the view that there is a block in sterol movement from the lysosome to the sites of regulation in NPC disease and have important implications for understanding the function of the NPC protein in intracellular cholesterol metabolism, in general, and in the brain, in particular.

Centripetal cholesterol flux to the liver is dictated by events in the peripheral organs and not by the plasma high density lipoprotein or apolipoprotein A-I concentration.

The major net flux of cholesterol in the intact animal or human is from the peripheral organs to the liver. This flux is made up of cholesterol that is either synthesized in these peripheral tissues or taken up as lipoprotein cholesterol. This study investigates whether it is the concentration of apolipoprotein (apo) A-I or high density lipoprotein in the plasma that determines the magnitude of this flux or, alternatively, whether events within the peripheral cells themselves regulate this important process. In mice that lack apoA-I and have very low concentrations of circulating high density lipoprotein, it was found that there was no accumulation of cholesterol in any peripheral organ so that the mean sterol concentration in these tissues was the same (2208 +/- 29 mg/kg body weight) as in control mice (2176 +/- 50 mg/kg). Furthermore, by measuring the rates of net cholesterol acquisition in the peripheral organs from de novo synthesis and uptake of low density lipoprotein, it was demonstrated that the magnitude of centripetal sterol movement from the peripheral organs to the liver was virtually identical in control animals (78 +/- 5 mg/day per kg) and in those lacking apoA-I (72 +/- 4 mg/day per kg). These studies indicate that the magnitude of net sterol flux through the body is not related to the concentration of high density lipoprotein or apolipoprotein A-I in the plasma, but is probably determined by intracellular processes in the peripheral organs that dictate the rate of movement of cholesterol from the endoplasmic reticulum to the plasma membrane.

Gender-related differences in bile acid and sterol metabolism in outbred CD-1 mice fed low- and high-cholesterol diets.

These studies were undertaken to determine whether in young adult outbred CD-1 mice there were any gender-related differences in basal bile acid metabolism that might be important in determining how males and females in this species responded to a dietary cholesterol challenge. When fed a plain cereal-based rodent diet without added cholesterol, 3-month-old females, compared with age-matched males, manifested a significantly larger bile acid pool (89.1 vs. 54.1 micromol/100 g body weight), a higher rate of fecal bile acid excretion (13.6 vs. 8.5 micromol/d/100 g body weight), a more efficient level of intestinal cholesterol absorption (41.1% vs. 25. 3%), and a lower rate of hepatic sterol synthesis (338 vs. 847 nmol/h/g). Similar results were found in C57BL/6 and 129Sv inbred mice. In matching groups of CD-1 mice fed a diet containing 1% cholesterol for 21 days, hepatic cholesterol levels increased much more in the females (from 2.4 to 9.1 mg/g) than in the males (from 2. 1 to 5.2 mg/g). This occurred even though the level of stimulation of cholesterol 7-hydroxylase activity in the females (79%) exceeded that in the males (55%), as did the magnitude of the increase in fecal bile acid excretion (females: 262% vs. males: 218%). However, in both sexes, bile acid pool size expanded only modestly and by a comparable degree (females: 19% vs. males: 26%) so that in the cholesterol-fed groups, the pool remained substantially larger in the females than in the males (102.3 vs. 67.6 micromol/100 g body weight). Together, these data demonstrate that while male and female CD-1 mice do not differ qualitatively in the way cholesterol feeding changes their bile acid metabolism, the inherently larger bile acid pool in the female likely facilitates the delivery of significantly more dietary cholesterol to the liver than is the case in males, thereby resulting in higher steady-state hepatic cholesterol levels.

Marked reduction in bile acid synthesis in cholesterol 7alpha-hydroxylase-deficient mice does not lead to diminished tissue cholesterol turnover or to hypercholesterolemia.

These studies used mice that were deficient in cholesterol 7alpha-hydroxylase to determine the effects of reduced bile acid synthesis on cholesterol homeostasis. In mice lacking this enzyme, bile acid synthesis was reduced from 8.3 to 3.4 micromol/day per 100 g body weight, the intestinal bile acid pool was decreased from 62.5 to 13.2 micromol/100 g bw, and the proportion of hyodeoxycholate, relative to cholate, in this pool was significantly increased. Associated with these changes, intestinal cholesterol absorption decreased from 37% to <1% while triacylglycerol absorption and animal weight gain remained essentially unaffected. The very low rate of cholesterol absorption could be corrected by feeding the mutant mice cholate, but not hyodeoxycholate. The reduction in sterol uptake across the intestine was associated with a 2-fold increase in cholesterol synthesis in the small bowel and liver and an increase in fecal neutral sterol excretion from 15.2 to 35.7 micromol/day per 100 g bw. The size of the cholesterol pools in the plasma, various organs and whole animal remained constant. Thus, under circumstances where the excretion of sterol as bile acids was markedly reduced, total cholesterol turnover actually increased from 164 to 239 mg/day per kg bw. This study demonstrates the complex interactions between bile acid and cholesterol metabolism and the dramatic effects of eliminating a single gene product; however, even though a major catabolic pathway was deleted, cholesterol balance across the animal was maintained.

Preferential utilization of newly synthesized cholesterol for brain growth in neonatal lambs.

These studies used the suckling lamb as a model to determine the sources of cholesterol that are utilized for development of the central nervous system in the neonate. Lambs were studied at 1.3 and 16.4 days after birth. Over this 15-day interval, 14 g of new brain tissue were formed. About 9-10 mg of cholesterol were utilized daily for this new tissue growth. To determine the source of this cholesterol, the absolute rates of low-density lipoprotein clearance and cholesterol synthesis were measured in vivo in nine separate regions of the central nervous system. Low-density lipoprotein clearance throughout the brain was very low and at most could have contributed only 0.3-0.4 mg cholesterol daily for new brain growth. In contrast, the brain synthesized 7-8 mg of cholesterol/day. There were pronounced regional differences in the concentration of cholesterol throughout the brain, and these correlated closely with the rate of sterol synthesis (r = 0.95) in these same regions. We conclude that the principal source of sterol for brain growth in suckling lambs is de novo synthesis.

Dietary fatty acids and the regulation of plasma low density lipoprotein cholesterol concentrations.

Epidemiologic studies over the past 25 years have shown that the level of dietary fat intake is positively correlated with the average serum cholesterol value and mortality from coronary heart disease (CHD). A number of different investigators demonstrated that in addition to total fat, the fatty acid composition of diets influenced serum total cholesterol (TC) in humans. In general, saturated fatty acids were found to elevate the serum cholesterol concentration, and unsaturated fatty acids were found to decrease this value. The lipoprotein fraction most affected was the level of cholesterol carried in low density lipoprotein (LDL-C). It has now been demonstrated that the steady-state level of LDL-C is predominantly dictated by metabolic events in the liver. As the amount of dietary cholesterol entering the body is increased, there is expansion of the sterol pool in the liver cell and down regulation of LDL receptors (LDLR) that are primarily responsible for clearing LDL-C from the blood stream. When dietary cholesterol intake is kept constant, however, long-chain saturated fatty acids further suppress hepatic LDLR activity, whereas several unsaturated fatty acids have the opposite effect. These regulatory events depend upon the availability of the various fatty acids to shift intracellular cholesterol between a regulatory and storage pool of cholesterol, and this effect is mediated by the enzyme acyl-CoA:cholesterol acyltransferase (ACAT).

Identification of a metabolic difference accounting for the hyper- and hyporesponder phenotypes of cynomolgus monkey.

These studies were done to determine whether an underlying metabolic difference could account for the higher concentration of cholesterol carried in low density lipoproteins (LDL-C) in male hyperresponder (HR) cynomolgus monkeys than in their hyporesponder (HO) counterparts during dietary cholesterol challenge. All animals were fed to steady state at 5 months a diet that had a constant concentration of cholesterol (0.19 mg/g), triacylglycerol (175 mg/g), and soluble fiber. There were no differences in these two phenotypes with respect to the profile of fatty acids in the liver and bile acids in the gallbladder, or in the relationship of cholesterol synthesis to cholesteryl ester formation in the liver. The rate of cholesterol synthesis in all extrahepatic tissues was also the same in the HO and HR animals but was 2.1 mg/day per kg body weight less in the liver of the HR monkeys. When challenged with a greater dietary cholesterol load, therefore, the HR animal could not readily further down-regulate synthesis and so shifted more cholesterol into the ester pool (9.4 mg/g) than did the HO animal (3.9 mg/g). Also the LDL-C concentration was more markedly elevated (412 mg/dl) compared to the hyporesponder monkey (188 mg/dl). Thus, this single metabolic alteration apparently accounted for the HO and HR phenotypes. As this difference was not due to variation in the delivery of sterol from the extrahepatic organs to the liver, it must reflect a difference in either net intestinal sterol absorption or net hepatic sterol excretion in the two phenotypes.