Regulation of hepatic lanosterol 14 alpha-demethylase gene expression by dietary cholesterol and cholesterol-lowering agents.

Binding of sterol response element binding protein 1a to sterol response element-1 (SRE-1) in the promoter region of lanosterol 14 alpha-demethylase (14DM) has been demonstrated previously. Decreased 14DM activity has been shown to result in accumulation of the intermediate, 3 beta-hydroxy-lanost-8-en-32-al, a known translational downregulator of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. Since it has also been demonstrated that feedback regulation of hepatic HMG-CoA reductase occurs primarily at the level of translation, the effects of dietary cholesterol and cholesterol lowering agents on levels of hepatic 14DM mRNA and immunoreactive protein were investigated. Addition of 1% cholesterol to a chow diet markedly decreased hepatic 14DM mRNA and protein levels in Sprague-Dawley rats. The extent and time course of this decrease in 14DM immunoreactive protein closely paralleled that of HMG-CoA reductase. Supplementation of the diet with the HMG-CoA reductase inhibitor, Lovastatin, to a level of 0.02%, raised 14DM mRNA and protein levels 2- to 3-fold. Addition of 2% Colestipol, a bile acid binding resin, to the chow diet caused smaller increases. The highest level of 14DM protein expression was observed in liver, the major site of feedback regulation of HMG-CoA reductase by cholesterol. Taken together, these observations suggest a critical role for 14DM in the feedback regulation of hepatic HMG-CoA reductase.

7-Dehydrocholesterol-dependent proteolysis of HMG-CoA reductase suppresses sterol biosynthesis in a mouse model of Smith-Lemli-Opitz/RSH syndrome.

Smith-Lemli-Opitz/RSH syndrome (SLOS), a relatively common birth-defect mental-retardation syndrome, is caused by mutations in DHCR7, whose product catalyzes an obligate step in cholesterol biosynthesis, the conversion of 7-dehydrocholesterol to cholesterol. A null mutation in the murine Dhcr7 causes an identical biochemical defect to that seen in SLOS, including markedly reduced tissue cholesterol and total sterol levels, and 30- to 40-fold elevated concentrations of 7-dehydrocholesterol. Prenatal lethality was not noted, but newborn homozygotes breathed with difficulty, did not suckle, and died soon after birth with immature lungs, enlarged bladders, and, frequently, cleft palates. Despite reduced sterol concentrations in Dhcr7(-/-) mice, mRNA levels for 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the rate-controlling enzyme for sterol biosynthesis, the LDL receptor, and SREBP-2 appeared neither elevated nor repressed. In contrast to mRNA, protein levels and activities of HMG-CoA reductase were markedly reduced. Consistent with this finding, 7-dehydrocholesterol accelerates proteolysis of HMG-CoA reductase while sparing other key proteins. These results demonstrate that in mice without Dhcr7 activity, accumulated 7-dehydrocholesterol suppresses sterol biosynthesis posttranslationally. This effect might exacerbate abnormal development in SLOS by increasing the fetal cholesterol deficiency.

Feedback and hormonal regulation of hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase: the concept of cholesterol buffering capacity.

Regulation of the expression of hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase by the major end product of the biosynthetic pathway, cholesterol, and by various hormones is critical to maintaining constant serum and tissue cholesterol levels in the face of an ever-changing external environment. The ability to downregulate this enzyme provides a means to buffer the body against the serum cholesterol-raising action of dietary cholesterol. The higher the basal expression of hepatic HMG-CoA reductase, the greater the "cholesterol buffering capacity" and the greater the resistance to dietary cholesterol. This review focuses on the mechanisms of feedback and hormonal regulation of HMG-CoA reductase in intact animals rather than in cultured cells and presents the evidence that leads to the proposal that regulation of hepatic HMG-CoA reductase acts as a cholesterol buffer. Recent studies with animals have shown that feedback regulation of hepatic HMG-CoA reductase occurs at the level of translation in addition to transcription. The translational efficiency of HMG-CoA reductase mRNA is diminished through the action of dietary cholesterol. Oxylanosterols appear to be involved in this translational regulation. Feedback regulation by dietary cholesterol does not appear to involve changes in the state of phosphorylation of hepatic HMG-CoA reductase or in the rate of degradation of this enzyme. Several hormones act to alter the expression of hepatic HMG-CoA reductase in animals. These include insulin, glucagon, glucocorticoids, thyroid hormone and estrogen. Insulin stimulates HMG-CoA reductase activity likely by increasing the rate of transcription, whereas glucagon acts by opposing this effect. Hepatic HMG-CoA reductase activity undergoes a significant diurnal variation due to changes in the level of immunoreactive protein primarily mediated by changes in insulin and glucagon levels. Thyroid hormone increases hepatic HMG-CoA reductase levels by acting to increase both transcription and stability of the mRNA. Glucocorticoids act to decrease hepatic HMG-CoA reductase expression by destabilizing reductase mRNA. Estrogen acts to increase hepatic HMG-CoA reductase activity primarily by stabilizing the mRNA. Deficiencies in those hormones that act to increase hepatic HMG-CoA reductase gene expression lead to elevations in serum cholesterol levels. High basal expression of hepatic HMG-CoA reductase, whether due to genetic or hormonal factors, appears to result in greater cholesterol buffering capacity and thus increased resistance to dietary cholesterol.

Effects of 15-oxa-32-vinyl-lanost-8-ene-3 beta,32 diol on the expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase and low density lipoprotein receptor in rat liver.

The mechanisms by which oxylanosterols regulate expression of hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase and lower serum cholesterol levels were examined by using a novel nonmetabolizable oxylanosterol mimic, 15-oxa-32-vinyl-lanost-8-ene-3 beta, 32 diol (DMP 565). This compound, unlike other nonmetabolizable oxylanosterols, is not a substrate for lanosterol 14 alpha-methyl demethylase. Feeding rats a diet supplemented with 0.02% DMP 565 markedly decreased HMG-CoA reductase immunoreactive protein and enzyme activity levels without affecting mRNA levels. The rate of reductase protein degradation was unaffected. However, the rate of translation was reduced to less than 20% of control. Thus, DMP 565 appears to regulate hepatic HMG-CoA reductase gene expression primarily at the level of translation. The pronounced inhibition of HMG-CoA reductase by DMP 565 resulted in a compensatory increase in the functioning of the hepatic low density lipoprotein (LDL) receptor, possibly by increased cycling, as evidenced by a marked increase in the rate of degradation of the LDL receptor. The half-life of the receptor was decreased from over 7 h to only 1 h in animals receiving DMP 565. This increase in the rate of degradation occurred without a change in the steady state level of the receptor. Addition of dietary cholesterol attenuated the increased turnover of the LDL receptor. These effects on the hepatic LDL receptor have also been observed with HMG-CoA reductase inhibitors (G. C. Ness et al., 1996, Arch. Biochem, Biophys. 325, 242-248). However, the effect of DMP 565 on the rate of degradation of the hepatic LDL receptor was of a greater magnitude when equal doses of the drugs were used. These regulatory actions of DMP 565 provide, in part, an explanation for the observed hypocholesterolemic action of this compound.

Effects of L-triiodothyronine and the thyromimetic L-94901 on serum lipoprotein levels and hepatic low-density lipoprotein receptor, 3-hydroxy-3-methylglutaryl coenzyme A reductase, and apo A-I gene expression.

The mechanisms by which thyroid hormone (triiodothyronine (T3)) and a thyromimetic, 2-amino-3-(3,5-dibromo-4-[4-hydroxy-3-(6-oxo-1,6-dihydro-pyridazin -3-ylmethyl)-phenoxyl]-phenyl)propionic acid (L-94901), lower plasma low density lipoprotein (LDL) cholesterol and raise plasma high density lipoprotein (HDL) cholesterol levels was investigated in thyroidectomized and sham-operated rats. Thyroidectomy resulted in a 77% increase in plasma LDL cholesterol, a 60% decrease in plasma triglycerides, and a modest reduction in HDL cholesterol. Daily oral dosing with T3 (10-170 nmol/kg) or L94901 (100-1000 nmol/kg) for 7 days decreased plasma LDL cholesterol in thyroidectomized rats by 60-80%, respectively. This reduction in LDL cholesterol was accompanied by a dose-dependent increase in HDL cholesterol levels of up to 60%. Thus, the ratio of LDL to HDL was decreased from 1.01 to 0.12 after treatment with L-94901 and to 0.25 after dosing with T3. In sham-operated animals, T3 and L-94901 lowered LDL cholesterol by 61 and 46%, respectively, and increased HDL cholesterol by 25 and 53%, respectively. Immunoblotting analysis of liver membranes prepared from thyroidectomized or sham-operated rats demonstrated that LDL receptor protein levels were increased by up to eight-fold. Northern blotting analysis revealed similar large increases in hepatic LDL receptor mRNA levels that accounted for the increases in LDL receptor protein levels. Hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase mRNA, protein, and activity were increased 2- to 3-fold. The T3- and L-94901-mediated increases in serum HDL levels were associated with 2- to 3-fold increases in apo A-I mRNA levels. In contrast with most other hypocholesterolemic agents, T3 and L-94901 significantly increase HDL cholesterol levels in addition to decreasing LDL cholesterol levels due to induction of hepatic apo A-I and LDL receptor gene expression.

Dietary cholesterol regulates hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase gene expression in rats primarily at the level of translation.

The level of gene expression at which dietary cholesterol exerts feedback regulation on hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase was investigated using young male Sprague-Dawley rats. Previous studies suggested that this regulation might be exerted posttranscriptionally. Thus, possible regulation at the levels of catalytic efficiency, protein turnover, and translation was investigated. To examine possible regulation at the level of catalytic efficiency, rats were placed on chow diets supplemented with 2% cholesterol and the rates of decline in hepatic HMG-CoA reductase activity and immunoreactive protein levels were determined. Both decreased slowly over a 72-h period. The catalytic efficiency did not change. These observations are inconsistent with phosphorylation-dephosphorylation or thiol-disulfide interchange as possible mechanisms. The possibility that dietary cholesterol might act by increasing the rate of turnover of HMG-CoA reductase protein was examined by determining the half-life of the enzyme in livers from rats consuming chow or chow supplemented with 2% cholesterol. The half-life of HMG-CoA reductase protein was not decreased in the animals receiving cholesterol, thus ruling out this possibility. Regulation at the level of translation was investigated by measuring the rate of HMG-CoA reductase protein synthesis in liver slices using [35S]methionine and [35S]cysteine. It was found that the rate of synthesis was reduced by over 80% in liver slices from rats fed a diet supplemented with 2% cholesterol. Similar results were obtained with liver slices from rats given mevalonolactone, which supplies both sterol and nonsterol endproducts. These results indicate that cholesterol regulates hepatic HMG-CoA reductase gene expression in rats primarily at the level of translation.

Compensatory responses to inhibition of hepatic squalene synthase.

The mechanism by which depletion of hepatic cholesterol levels, achieved by inhibition of squalene synthase, alters hepatic LDL receptor, HMG-CoA reductase, and cholesterol 7alpha-hydroxylase gene expression was investigated by measuring transcription rates, mRNA stability, rates of translation, translational efficiency, and levels of sterol response element binding proteins. It was found that the transcription of both hepatic LDL receptor and HMG-CoA reductase were increased about twofold. The increase in LDL receptor transcription occurred within 2 h after giving 2 mg/kg zaragozic acid A, a potent inhibitor of squalene synthase. This preceded the increase in transcription of HMG-CoA reductase that occurred at 4 h. Increases in the stability of both of these mRNAs were also observed. These changes account for the increases in LDL receptor and HMG-CoA reductase mRNA levels previously observed. The rate of transcription of hepatic cholesterol 7alpha-hydroxylase was decreased to about 25% of control within 3 h after administration of zaragozic acid A, which correlates with the decrease in this mRNA. The rates of translation, as determined by pulse labeling, of both hepatic HMG-CoA reductase and LDL receptor were increased two- to threefold. The translational efficiency of these two mRNAs was also increased as judged by polysome profile analysis. There was an increase in mRNA associated with the heaviest polysome fraction and a decrease in that associated with monosomes. No significant change was observed in the levels of sterol response element binding protein 2, the form that mediates induced transcription, in response to zaragozic acid A treatment, indicating that this protein might not be involved in mediating the observed transcriptional changes. An increase in sterol response element binding protein -1 was observed 30 min after giving zaragozic acid A. The results suggest that compensatory responses to depletion of squalene-derived products involve alterations in the rates of transcription, mRNA stability, and translational of key proteins involved in cholesterol homeostasis.

Atorvastatin action involves diminished recovery of hepatic HMG-CoA reductase activity.

The effects of atorvastatin on the expression of the hepatic HMG-CoA reductase and LDL receptor genes were investigated in rats. Like the other statins, atorvastatin increased the rate of degradation and presumably cycling of the hepatic LDL receptor. In atorvastatin-treated rats, the half-life of the receptor was decreased by over 60%. Hepatic HMG-CoA reductase mRNA levels were increased about 3-fold by feeding a diet containing 0.04% atorvastatin while reductase protein levels were increased by as much as 700-fold. Apparent HMG-CoA reductase activity was not increased as much as protein levels. Washing experiments revealed that atorvastatin is more difficult to remove from microsomes than lovastatin. The results support the conclusion that the potent hypocholesterolemic action of atorvastatin involves decreased hepatic VLDL production due to effective inhibition of in vivo cholesterol biosynthesis resulting from diminished recovery of HMG-CoA reductase activity following drug treatment.

Inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase unmask transcriptional regulation of hepatic low-density lipoprotein receptor gene expression by dietary cholesterol.

The mechanism by which dietary cholesterol regulates expression of the hepatic low-density lipoprotein (LDL) receptor was investigated. In a previous study (Arch. Biochem. Biophys. 325, 242-248, 1996), we demonstrated that dietary cholesterol reduces the rate of LDL receptor protein degradation without affecting steady-state levels of receptor protein. In view of these findings, it was expected that dietary cholesterol would decrease the rate of transcription of the hepatic LDL receptor gene, resulting in lower mRNA levels and lower rates of synthesis of LDL receptor protein. Surprisingly, neither the rate of transcription nor the level of LDL receptor mRNA was reduced in response to dietary cholesterol, even though hepatic cholesterol levels were increased twofold. This suggests that under normal conditions, dietary cholesterol does not affect LDL receptor gene expression at the level of transcription. In contrast, feeding 2% cholesterol to rats fed a diet supplemented with 0.04% lovastatin significantly decreased hepatic LDL receptor mRNA levels and transcription rates. These results suggest that lovastatin unmasks transcriptional regulation of the hepatic LDL receptor by dietary cholesterol. The levels of the mature nuclear forms of sterol response element binding proteins-1 and -2 were unaffected despite significant changes in hepatic cholesterol levels, mRNA levels, and transcription rates caused by lovastatin treatment. This suggests that the observed changes in transcription rates may not be mediated by these proteins in rat liver.

3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors unmask cryptic regulatory mechanisms.

The possibility that potent inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase may alter the mechanisms by which dietary cholesterol and farnesol regulate this gene was investigated by comparing the regulatory responses of rats maintained on diets with or without 0.04% Lovastatin supplementation to dietary cholesterol. It was found that the rate of hepatic HMG-CoA reductase transcription was significantly decreased by dietary cholesterol in animals fed Lovastatin-supplemented diets, whereas animals maintained on a normal chow diet showed no decrease in the rate of transcription. The levels of reductase mRNA were decreased to about 10% of controls in Lovastatin-supplemented animals in response to dietary cholesterol but not affected in nonsupplemented animals. Administration of farnesol, reputed to be the nonsterol regulator of reductase, to rats maintained on a diet containing Lovastatin decreased hepatic HMG-CoA reductase protein by 30% and the half-life of reductase immunoreactive protein to 4.0 h, which is close to that observed in chow-fed animals. In contrast, farnesol treatment does not affect the turnover rate of reductase protein in rats fed a normal chow diet. These results suggest that potent inhibitors of HMG-CoA reductase may unmask transcriptional regulation by dietary cholesterol and accelerated degradation of the reductase by the putative nonsterol regulator farnesol.

Translational regulation of hepatic HMG-CoA reductase by dietary cholesterol.

The question of whether dietary cholesterol exerts feedback regulation on hepatic HMG-CoA reductase at the level of translation was examined by performing polysome profile analysis. Liver polysomes from rats fed 2% cholesterol in their diets for 3 days were compared with those isolated from rats fed a normal chow diet. Northern blotting analysis of the individual fractions revealed that cholesterol feeding reduced the portion of HMG-CoA reductase mRNA associated with translationally active polysomes by over 50% and progressively increased the percentage of reductase mRNA present in the monosomal fractions. In the lightest monosomal fraction over 10 times as much reductase mRNA was present in samples from cholesterol animals as compared to controls. These findings indicate that dietary cholesterol exerts significant feedback regulation on hepatic HMG-CoA reductase at the translational level.

Increased expression of low-density lipoprotein receptors in a Smith-Lemli-Opitz infant with elevated bilirubin levels.

We report on an infant girl with severe RSH or Smith-Lemli-Opitz syndrome with hyperbilirubinemia. The infant died at age 2 months. Sterol analysis of liver and brain tissues showed marked elevations of 7-dehydrocholesterol with decreased levels of cholesterol. Immunocytochemical analysis demonstrated remarkable increases in low-density lipoprotein (LDL) receptors in these tissues, indicative of a deficiency in available cholesterol for tissue needs.

Smith-Lemli-Opitz syndrome produced in rats with AY 9944 treated by intravenous injection of lipoprotein cholesterol.

A limitation to treating Smith-Lemli-Opitz infants by giving dietary cholesterol is their impaired ability to absorb cholesterol due to a deficiency of bile acids. Since intravenously administered lipoprotein cholesterol should not require bile acids for uptake into tissues, we tested the effects of this form of cholesterol on tissue cholesterol and 7-dehydrocholesterol levels in an animal model of SLO, created by feeding rats 0.02% AY 9944. Intravenous administration of 15 mg of bovine cholesterol supertrate twice daily increased serum cholesterol levels from 11 to over 250 mg/dl. This treatment increased liver cholesterol levels from 309 to over 900 micrograms/g and lowered hepatic 7-dehydrocholesterol levels from 1546 to 909 micrograms/g. A combination of iv cholesterol and 2% dietary cholesterol was most effective as it raised hepatic cholesterol levels to 1950 micrograms/g, which is 50% above normal. 7-Dehydrocholesterol levels were decreased to 760 micrograms/g. Similar responses were seen for heart, lung, kidney, and testes. Brain sterol levels were not significantly affected. AY 9944 caused a modest increase in hepatic HMG-CoA reductase activity. Administration of dietary cholesterol together with iv cholesterol lowered hepatic HMG-CoA reductase activity to barely detectable levels. The data indicate that the combination of iv and dietary cholesterol was most effective in raising cholesterol levels, lowering 7-dehydrocholesterol levels, and inhibiting de novo cholesterol biosynthesis.

Increasing hepatic cholesterol 7alpha-hydroxylase reduces plasma cholesterol concentrations in normocholesterolemic and hypercholesterolemic rabbits.

The effect of bile acid depletion and replacement with glycodeoxycholic acid on plasma cholesterol concentrations, hepatic low-density lipoprotein (LDL) receptor binding and messenger RNA (mRNA) levels, and hepatic activities and mRNA levels for 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase and cholesterol 7alpha-hydroxylase was investigated in 19 New Zealand white (NZW) and 15 Watanabe heritable hyperlipidemic (WHHL) rabbits. Bile acid depletion was produced by external bile drainage for 5 days, which maximized cholic acid synthesis. Replacement was achieved by infusing glycodeoxycholic acid intraduodenally for 24 hours so that the hepatic bile acid flux reached prefistula levels. Plasma and liver cholesterol concentrations were 13 times and 50% greater, respectively, hepatic LDL receptor-mediated binding was 26% less, and cholesterol 7alpha-hydroxylase activity and mRNA levels were 62% and 86% less in WHHL than NZW rabbits. After bile drainage, plasma cholesterol concentrations decreased 29% in NZW rabbits and 40% in WHHL rabbits and were associated with a 2.1-fold increase in hepatic LDL receptor-mediated binding in the NZW rabbits, but there was no change in the WHHL rabbits. Cholesterol 7alpha-hydroxylase activity and mRNA levels increased three and four times in NZW and WHHL rabbits, respectively, although liver cholesterol levels remained unchanged. Replacement with exogenous glycodeoxycholic acid increased plasma cholesterol concentrations 1.7 times in NZW rabbits and decreased enhanced cholesterol 7alpha-hydroxylase activity 54%, mRNA levels 86%, cholic acid synthesis 38%, and hepatic LDL receptor-mediated binding 57% in NZW rabbits. Bile acid depletion stimulated cholic acid synthesis by up-regulating cholesterol 7alpha-hydroxylase to use cholesterol and reduce plasma concentrations substantially in both NZW and WHHL rabbits, although LDL receptors did not function in WHHL rabbits. Glycodeoxycholic acid replacement inhibited elevated cholesterol 7alpha-hydroxylase, cholic acid synthesis, and hepatic LDL receptor binding to reestablish baseline plasma cholesterol levels in NZW rabbits. Hypercholesterolemia in WHHL rabbits was related to the combination of dysfunctional LDL receptors and inhibited cholesterol 7alpha-hydroxylase. Plasma cholesterol concentrations were reduced significantly when cholesterol 7alpha-hydroxylase was stimulated even in the absence of LDL receptor function.

Farnesol is not the nonsterol regulator mediating degradation of HMG-CoA reductase in rat liver.

A recent report, in which cultured tumor cells were used, identified farnesol as the nonsterol mevalonate-derived metabolite required for the accelerated degradation of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (C. C. Correll, L. Ng, and P. A. Edwards, 1994, J. Biol. Chem. 269, 17390-17393). We examined this proposed linkage in animals by measuring hepatic farnesol levels and rates of HMG-CoA reductase degradation under conditions previously shown to alter the stability of the reductase. In normal rats, the hepatic farnesol level, quantified by high-pressure liquid chromatography, was 0.10 +/- 0.08 microgram/g and the half-life of HMG-CoA reductase was 2.5 h. Administration of mevalonolactone at 1 g/kg body wt to provide all nonsterol metabolites in addition to cholesterol increased farnesol levels 6-fold without significantly affecting the half-life of the reductase. Treatment of rats with zaragozic acid A, an inhibitor of squalene synthase, raised hepatic farnesol levels 10-fold and decreased the half-life of HMG-CoA reductase to 0.25 h. However, feeding lovastatin to rats did not lower hepatic farnesol levels despite a marked stabilization of HMB-CoA reductase protein. Moreover, intubation of rats with 500 mg/kg body wt of farnesol failed to decrease the half-life of HMG-CoA reductase protein, alter the levels of enzyme activity, or change of the levels of immunoreactive protein despite an increase of 1000-fold in hepatic farnesol levels. These observations indicate that farnesol per se does not induce accelerated degradation of HMG-CoA reductase in rat liver.

The diurnal variation of hepatic HMG-CoA reductase activity is due to changes in the level of immunoreactive protein.

The diurnal variation in levels of hepatic HMG-CoA reductase immunoreactive protein and enzyme activity were determined in rats. Immunoreactive protein levels changed together with enzyme activity. Thus the catalytic efficiency of HMG-CoA reductase was not significantly changed. The data suggest that the diurnal variation in hepatic HMG-CoA reductase activity is due to changes in enzyme protein levels rather than changes in phosphorylation state of the enzyme, for example, which would cause changes in catalytic efficiency.

Inhibitors of cholesterol biosynthesis increase hepatic low-density lipoprotein receptor protein degradation.

Inhibitors of cholesterol biosynthesis are believed to lower serum cholesterol levels by enhancing the removal of serum low-density lipoprotein (LDL) by increasing hepatic LDL receptor function. Thus, the effects of several different inhibitors of cholesterol biosynthesis were examined for their effects on the expression of the hepatic LDL receptor in rats. We found that administration of inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A reductase such as lovastatin, pravastatin, fluvastatin, and rivastatin resulted in increased hepatic LDL receptor mRNA levels. Surprisingly, these agents failed to increase levels of immunoreactive LDL receptor protein in rat liver even when the dose and length of treatment were increased. Treatment of rats with zaragozic acid A, an inhibitor of squalene synthase, caused even greater increases in hepatic LDL receptor mRNA levels, but did not increase levels of immunoreactive protein. Further investigation revealed that the rate of degradation of the hepatic LDL receptor was increased in rats given inhibitors of cholesterol biosynthesis. The greatest increase in the rate of degradation was seen in animals treated with zaragozic acid A which caused the largest increase in hepatic LDL receptor mRNA levels. In contrast, hepatic LDL receptor protein was stabilized in cholesterol-fed rats. It appears that increased potential for LDL receptor protein synthesis, reflected in increased mRNA levels, is offset by a corresponding increase in the rate of receptor protein degradation resulting in constant steady-state levels of hepatic LDL receptor protein. These findings are suggestive of increased cycling of the hepatic LDL receptor. This postulated mechanism can provide for enhanced hepatic uptake of lipoproteins without increasing steady-state levels of LDL receptor protein.

Transcriptional regulation of rat hepatic low-density lipoprotein receptor and cholesterol 7 alpha hydroxylase by thyroid hormone.

Utilizing hypophysectomized and thyroidectomized rats, it is demonstrated that the rapid increase in expression of hepatic low-density lipoprotein (LDL) receptor and cholesterol 7 alpha-hydroxylase in response to thyroid hormone is due to increases in the rates of transcription of these genes. An increase in transcription of the hepatic LDL receptor gene was seen within 30 min after administration of triiodothyronine. The increase in transcription fully accounted for the increase in hepatic LDL receptor protein. Stimulation of transcription of these two genes provides for enhanced removal of LDL from serum and elimination from the body.

Treatment of the cholesterol biosynthetic defect in Smith-Lemli-Opitz syndrome reproduced in rats by BM 15.766.

BACKGROUND & AIMS: The Smith-Lemli-Opitz syndrome is a recessive inherited disorder characterized by neurological developmental defects and dysmorphic features with a defect in cholesterol synthesis at the conversion of 7-dehydrocholesterol to cholesterol. BM 15.766 inhibits 7-dehydrocholesterol-delta 7-reductase and reproduces the biochemical defect. The aim of this study was to investigate the effects of cholesterol, cholic acid, and lovastatin feeding on rats fed BM 15.766. METHODS: Plasma cholesterol and 7-dehydrocholesterol concentrations were related to the hepatic 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase. RESULTS: With the inhibitor treatment, plasma cholesterol concentrations decreased 67%; 7-dehydrocholesterol concentrations increased from trace to 17 mg/dL; and hepatic HMG-CoA reductase activity and messenger RNA levels were stimulated 74% and two times, respectively. In inhibitor-treated rats, feeding cholesterol increased plasma cholesterol concentrations 3.7 times, decreased 7-dehydrocholesterol concentrations 88%, and reduced elevated HMG-CoA reductase activity and messenger RNA levels 74% and 49%. Feeding cholic acid increased plasma cholesterol without reducing 7-dehydrocholesterol concentrations. The combination of cholic acid and cholesterol enhanced plasma cholesterol 9.5 times without decreasing 7-dehydrocholesterol levels. Feeding lovastatin depressed plasma cholesterol further without reducing 7-dehydrocholesterol levels. CONCLUSIONS: Cholesterol is essential to correct abnormal cholesterol synthesis induced by BM 15.766 in rats by expanding the pool and inhibiting HMG-CoA reductase. Neither cholic acid nor lovastatin are effective separately, but cholic acid plus cholesterol may offer some additional benefit.