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PubMed2003

Serum ferritin concentration is associated with plasma levels of cholesterol oxidation products in man.

Cholesterol oxidation products, oxysterols, are thought to play a part in the initiation and development of human atherosclerotic lesions. Excessive body iron has been suggested to promote atherosclerosis and coronary heart disease through its pro-oxidative properties. In the present study, the associations between serum ferritin and plasma oxysterol concentrations were examined in 669 eastern Finnish men. Serum ferritin concentration had statistically significant (p <.05) direct correlations with most of the measured oxysterols. In multivariate adjusted regression models, serum ferritin concentration predicted significantly the levels of 27-hydroxycholesterol (beta = 0.13, p <.001), 7alpha-hydroxycholesterol (beta = 0.11, p =.005), 25-hydroxycholesterol (beta = 0.10, p =.007), 7-ketocholesterol (beta = 0.10, p =.009), and 7beta-hydroxycholesterol (beta = 0.10, p =.02). In conclusion, excess body iron, as assessed by serum ferritin, is associated with increased levels of circulating oxysterols, both of enzymatic and nonenzymatic origin, in man.

Adult↗
PubMed1996

Recurrent G-to-A substitution in a single codon of SREBP cleavage-activating protein causes sterol resistance in three mutant Chinese hamster ovary cell lines.

Oxygenated sterols such as 25-hydroxycholesterol kill Chinese hamster ovary cells because they inhibit the proteolytic processing of sterol regulatory element binding proteins (SREBPs), a pair of membrane-bound transcription factors that activate genes controlling cholesterol synthesis and uptake from lipoproteins. The unprocessed SREBPs remain membrane-bound, they cannot activate the cholesterol biosynthetic pathway, and the cells die of cholesterol deprivation. Several sterol-resistant hamster cell lines have been isolated previously by chemical mutagenesis and selection for resistance to killing by 25-hydroxycholesterol. We recently identified the defect in one such cell line (25-RA cells) as a point mutation in a newly discovered membrane protein of 1276 amino acids, designated SREBP cleavage-activating protein (SCAP). The mutation in the 25-RA cells resulted from a G-to-A transition in codon 443 of the SCAP gene, changing aspartic acid to asparagine. Wild-type SCAP, when overexpressed by transfection, stimulates the proteolytic processing of both SREBPs. The D443N substitution is an activating mutation that increases the activity of SCAP and renders it resistant to inhibition by 25-hydroxycholesterol. We here report the identical G-to-A transition in two additional lines of Chinese hamster ovary cells that were mutagenized and isolated by a similar protocol. The three mutations occurred independently as indicated by haplotype analysis of the mutant genes using two intragenic sequence polymorphisms. All three cell lines were mutagenized with alkylating agents (nitrosoethylurea or ethylmethane sulfonate) that favor G-to-A transitions. Nevertheless, the finding of the same nucleotide substitution at the same location in all three cell lines indicates that SCAP may be unique in its ability to stimulate SREBP cleavage, and residue 443 is a crucial determinant of the protein's ability to be inhibited by 25-hydroxycholesterol.

Adenine↗
PubMed1995

Three different rearrangements in a single intron truncate sterol regulatory element binding protein-2 and produce sterol-resistant phenotype in three cell lines. Role of introns in protein evolution.

The cholesterol analogue 25-hydroxycholesterol kills animal cells by blocking the proteolytic activation of two sterol-regulated transcription factors designated sterol regulatory element binding protein-1 and -2 (SREBP-1 and SREBP-2). These proteins, each approximately 1150 amino acids in length, are embedded in the membranes of the nucleus and endoplasmic reticulum by virtue of hydrophobic COOH-terminal segments. In cholesterol-depleted cells the proteins are cleaved to release soluble NH2-terminal fragments of approximately 480 amino acids that enter the nucleus and activate genes encoding the low density lipoprotein receptor and enzymes of cholesterol synthesis. 25-Hydroxycholesterol blocks this cleavage, and cells die of cholesterol deprivation. We previously described a mutant 25-hydroxycholesterol-resistant hamster cell line (SRD-1 cells) in which the SREBP-2 gene had undergone a recombination between the intron following codon 460 and an intron in an unrelated gene. The SREBP-2 sequence terminated at residue 460, eliminating the membrane attachment domain and producing a constitutively active factor that no longer required proteolysis and thus was not inhibited by 25-hydroxycholesterol. Here, we report that two additional sterol-resistant cell lines (SRD-2 and SRD-3) have also undergone genomic rearrangements in the intron following codon 460 of the SREBP-2 gene. Although the molecular rearrangements differ in the three mutant lines, each leads to the production of a constitutively active transcription factor whose SREBP-2 sequence terminates at residue 460. These findings provide a dramatic illustration of the advantage that introns provide in allowing proteins to gain new functions in response to new environmental challenges.

Amino Acid Sequence↗
PubMed2002

Protection of superoxide-induced cholesterol oxidation by antioxidants in protic conditions.

It was found in previous experiments that superoxide in the presence of added hydrogen peroxide in protic conditions produces oxysterols. The oxysterols formed under these conditions were 7beta-ketocholesterol, 7alpha-hydroxycholesterol and 7beta-hydroxycholesterol. In the present experiments, the inhibitory effects of three antioxidants, alpha-tocopherol (alpha-T), butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT), on the oxidation of cholesterol in the presence of superoxide anion, water and hydrogen peroxide were investigated. It was found that BHA had the highest antioxidant activity on cholesterol oxidation, followed by alpha-T and BHT. The presence of antioxidants markedly retarded the formation of 7-ketocholesterol. The formation of 7beta-hydroxycholesterol or 7alpha-hydroxycholesterol was also reduced, but to a lesser degree.

Antioxidants↗
PubMed2001

Selective distribution of oxysterols in atherosclerotic lesions and human plasma lipoproteins.

The presence of oxidized sterols (oxysterols) in human serum and lesions has been linked to the initiation and progression of atherosclerosis. Data concerning the origin, identity and quantity of oxysterols in biological samples are controversial and inconsistent. This inconsistency may arise from different analytical methods or handling conditions used by different investigators. In the present study, oxysterol levels and distribution were analyzed by an optimized GC-MS method, in human atherosclerotic coronary and carotid lesions, in atherosclerotic apolipoprotein E deficient mice (E degrees mice) and in native and in vitro oxidized human low and high density lipoproteins. Oxysterol levels were analyzed with a limit of detection of 0.06 - 0.24 ng, with 25-hydroxycholesterol (25-OH) being the least sensitive. In human coronary and carotid lesions, obtained from endatherectomic samples, 27-hydroxycholesterol (27-OH) was the major oxysterol, with about 85% as sterols esterified to fatty acids. While total cholesterol and oxysterols levels were similar in both kinds of human lesions, oxysterol distribution was significantly different. In coronary lesions the mean levels of 27-OH and 7beta-hydroxycholesterol (7beta-OH) were 38% and 20% of total oxysterols, whereas in carotid lesions their mean levels were 66% and 5%, respectively. Unlike in human aortic lesions, 27-OH was entirely absent in E degrees mice, whereas the level of 7alpha-hydroxycholesterol (7alpha-OH) was 28% of the total oxysterols, vs. 5% in human coronary lesions. As 27-OH is an enzymatic product of cholesterol oxidation, this finding may indicate that such an enzymatic process does not take place in E degrees mice.

Animals↗
PubMed2004

The inhibition of the human cholesterol 7alpha-hydroxylase gene (CYP7A1) promoter by fibrates in cultured cells is mediated via the liver x receptor alpha and peroxisome proliferator-activated receptor alpha heterodimer.

In previous work, we showed that the binding of the liver x receptor alpha:peroxisome proliferator-activated receptor alpha (LXRalpha:PPARalpha) heterodimer to the murine Cyp7a1 gene promoter antagonizes the stimulatory effect of their respective ligands. In this study, we determined if LXRalpha:PPARalpha can also regulate human CYP7A1 gene promoter activity. Co-expression of LXRalpha and PPARalpha in McArdle RH7777 hepatoma cells decreased the activity of the human CYP7A1 gene promoter in response to fibrates and 25-hydroxycholesterol. In vitro, the human CYP7A1 Site I bound LXRalpha:PPARalpha, although with substantially less affinity compared with the murine Cyp7a1 Site I. The binding of LXRalpha:PPARalpha to human CYP7A1 Site I was increased in the presence of either LXRalpha or PPARalpha ligands. In HepG2 hepatoblastoma cells, fibrates and 25-hydroxycholesterol inhibited the expression of the endogenous CYP7A1 gene as well as the human CYP7A1 gene promoter when co-transfected with plasmids encoding LXRalpha and PPARalpha. However, a derivative of the human CYP7A1 gene promoter that contains a mutant form of Site I that does not bind LXRalpha:PPARalpha was not inhibited by WY 14,643 or 25-hydroxycholesterol in both McArdle RH7777 and HepG2 cells. The ligand-dependent recruitment of LXRalpha:PPARalpha heterodimer onto the human CYP7A1 Site I can explain the inhibition of the human CYP7A1 gene promoter in response to fibrates and 25-hydroxycholesterol.

Acids, Acyclic↗
PubMed2006

Synergistic effect of peroxisome proliferator activated receptor-gamma and liver X receptor-alpha in the regulation of inflammation in macrophages.

Peroxisome proliferator-activated receptor-gamma (PPARgamma) and liver X receptor-alpha (LXRalpha) are nuclear ligand-activated transcription factors, which regulate lipid metabolism and inflammation. Murine J774.2 macrophages were stimulated with Escherichia coli lipopolysaccharide (concentration, 10 microg/mL) with or without the PPARgamma ligand, 15-deoxy-Delta prostaglandin J2 (15d-PGJ2), or the LXRalpha ligands, 22(R)-hydroxycholesterol and T0901317 (concentration range, 0.01-10 micromol/L), alone or in combination. Nitric oxide (NO) metabolites and tumor necrosis factor alpha production, inducible NO synthase expression, and mitochondrial respiration were measured. When added to the cells as single agents, 15d-PGJ2, 22(R)-hydroxycholesterol, or T0901317 reduced the lipopolysaccharide-induced NO and tumor necrosis factor alpha production and the inducible NO synthase expression, and partially maintained mitochondrial respiration in a concentration-dependent manner. When added to the cells in combination at suboptimal concentrations, 15d-PGJ2 with 22(R)-hydroxycholesterol, or 15d-PGJ2 with T0901317, exerted anti-inflammatory effects similar to much higher concentrations (10,000-fold to 100,000-fold) of each ligand alone. The anti-inflammatory effects of these ligands, alone or in combination, were associated with reduction of nuclear factor-kappaB activation and with enhancement of PPARgamma DNA binding. LXRalpha expression was upregulated in response to 15d-PGJ2 and to the LXRalpha ligands when added alone or in combination. Immunoprecipitation experiments revealed that PPARgamma interacted with LXRalpha. Our data demonstrate that the PPARgamma ligand, 15d-PGJ2, and the LXRalpha ligands, 22(R)-hydroxycholesterol and T0901317, although binding to different nuclear receptors (i.e., PPARgamma and LXRalpha, respectively), affect mediator production through common cell signaling events and exert a synergistic potentiation in a combined treatment at suboptimal concentrations. Thus, our data suggest that PPARgamma and LXRalpha may interact in controlling the inflammatory response in macrophages.

Animals↗
PubMed1993

Oxysterols inhibit gap junctional communication between rat hepatocytes in primary culture.

Several oxysterols were examined for their effect on gap junctional communication between rat hepatocytes in primary culture. 25-Hydroxycholesterol, 22(S)-hydroxycholesterol and 7 beta-hydroxycholesterol, in decreasing order of potency, markedly inhibited gap junctional communication. In contrast, 7-ketocholesterol showed no inhibitory effect. The inhibition of gap junctional communication by oxysterols was not a consequence of changes in cell viability, as measured by lactate dehydrogenase leakage and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction activity. The addition of exogenous cholesterol to the culture medium did not abolish the effect of 25-hydroxycholesterol, suggesting that the capacity of oxysterols to inhibit gap junctional communication is independent of their inhibitory effect on cholesterol synthesis. We suppose that inhibition of gap junctional communication may be an early sign of oxysterols-induced toxicity on hepatocytes.

Animals↗
PubMed1998

Sterol 27-hydroxylase- and apoAI/phospholipid-mediated efflux of cholesterol from cholesterol-laden macrophages: evidence for an inverse relation between the two mechanisms.

Cholesterol-laden, human monocyte-derived macrophages were found to contain 27-hydroxycholesterol in proportion to their content of cholesterol ester. In accordance with previous work with human lung alveolar macrophages, there was a significant efflux of 27-hydroxycholesterol and 3beta-hydroxy-5-cholestenoic acid from the cultured cells. The efflux of 27-hydroxycholesterol was proportional to the cellular content of this steroid. Incubation of cholesterol-laden macrophages with reconstituted discoidal complexes made from apolipoprotein A-I and phospholipids resulted in a decrease in total cellular cholesterol, an increase in the efflux of free cholesterol, and a concomitant decrease in the total production and efflux of 27-oxygenated steroids, in particular, 3beta-hydroxy-5-cholestenoic acid. Reconstituted discoidal complexes with the Milano variant of apolipoprotein A-I gave virtually identical results, whereas high density lipoprotein was less efficient. These results suggest that cultured cholesterol-laden cells can export some of their excess cholesterol in the form of 27-hydroxycholesterol, 3beta-hydroxy-5-cholestenoic acid, and free cholesterol. In the presence of exogenous cholesterol acceptors, export of free cholesterol becomes more effective, resulting in less cholesterol exported via the 27-hydroxylase pathway. The balance between the two mechanisms for removal of cholesterol from macrophages may be of importance for formation of foam cells and development of atherosclerosis.

Apolipoprotein A-I↗
PubMed1998

Inhibition by cholesterol oxides of NO release from human vascular endothelial cells.

Recent studies have demonstrated that, unlike cholesterol, cholesterol oxidized at position 7 can reduce the maximal endothelium-dependent relaxation of isolated rabbit aortas (Circulation. 1997;95:723-731). The aim of the current study was to determine whether cholesterol oxides reduce the release of nitric oxide (NO) from human umbilical vein endothelial cells (HUVECs). The amount of NO released by histamine-stimulated HUVECs was determined by differential pulse amperometry using a nickel porphyrin- and Nafion-coated carbon microfiber electrode. The effects of cholesterol (preserved from oxidation by butylated hydroxytoluene), 7-ketocholesterol, 7beta-hydroxycholesterol, 5alpha,6alpha-epoxycholesterol, 19-hydroxycholesterol (60 microg/mL), and alpha-lysophosphatidylcholine (10 microg/mL) were compared. Pretreatment of HUVECs with cholesterol, 5alpha,6alpha-epoxycholesterol, or 19-hydroxycholesterol did not alter histamine-activated NO production. In contrast, pretreatment with 7-ketocholesterol or 7beta-hydroxycholesterol significantly decreased NO release. The inhibitory effect of 7-ketocholesterol was time and dose dependent and was maintained in the presence of L-arginine. In the absence of serum, lysophosphatidylcholine also reduced NO production. In ionomycin-stimulated cells, pretreatment with 7-ketocholesterol did not inhibit NO release. These results demonstrate that cholesterol derivatives oxidized at the 7 position, the main products of low density lipoprotein oxidation, reduce histamine-activated NO release in HUVECs. Such an inhibitory effect of cholesterol oxides may account, at least in part, for the ability of oxidized low density lipoprotein to reduce the endothelium-dependent relaxation of arteries.

Arginine↗
PubMed1997

Cytotoxicity of some oxysterols on human vascular smooth muscle cells was mediated by apoptosis.

A decrease in smooth muscle cells is observed in advanced atherosclerotic lesion. To understand this mechanism, we selected oxysterols as candidates for toxic lipid, and examined their cytotoxicity on human cultured vascular smooth muscle cells, together with the manner of cell death. In the presence of 7-ketocholesterol or 7 beta-hydroxycholesterol (50 mumol/L), the percentage of detached cells increased significantly with dose dependency, and an increase in detached cell number and DNA nick detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling study (TUNEL) preceded an increase in lactate dehydrogenase released into the medium. DNA extracted from smooth muscle cells incubated with 7-ketocholesterol or 7 beta-hydroxycholesterol showed a laddering pattern on agarose electrophoresis. In the presence of 7-ketocholesterol or 7 beta-hydroxycholesterol, fragmented DNA quantified by the quantitative sandwich enzyme immunoassay was significantly increased. From these results, it is proposed that 7-ketocholesterol and 7 beta-hydroxycholesterol are toxic to smooth muscle cells, and that this cytotoxicity is mediated by apoptosis.

Apoptosis↗
PubMed1976

Inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in hepatoma tissue culture cells by pure cholesterol and several cholesterol derivatives. Evidence supporting two distinct mechanisms.20l.

Pure cholesterol associated in complexes with lipoproteins (whole serum and human low density lipoproteins) or esterified with succinic acid (cholesteryl succinate) and bound to albumin effectively suppresses 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity in hepatoma tissue culture (HTC) cells grown in lipoprotein-poor serum medium during short 4-hour) incubation periods. Simultaneous measurments of the kinetics of uptake of radioactive unesterified cholesterol of whole serum and cholesteryl succinate, their conversion to lipid products, and the decay in enzyme activity, suggest that the cholesterol-induced suppression is mediated by the sterol itself rather than by inhibitory lipid products derived from its metabolism. Several cholesterol derivatives such as cholestenone, 7-ketocholesterol, and 7alpha-and 25-hydroxycholesterol also suppress reductase activiy in HTC cells and are significantly more inhibitory than the pure cholesterol preparations. The decrease in enzyme activity produced by cholesterol and its derivatives is concentration-dependent and specific. [1-14C]Oleate incorporation experiments indicate that cholesterol ester formation in HTC cells is not increased at inhibitory concentrations of the steroids. These data suggest that sterol ester formation is not an obligatory process in the feedback control of HMG-CoA reductase activity. The half-life of the reductase (3 to 4 hours) is not significantly changed by cycloheximide, plus or minus whole serum, and cholesteryl succinate. In contrast, the half-life is strongly reduced when HTC cells are incubated with cycloheximide plus maximal concentrations of 25-hydroxycholesterol, 7-ketocholesterol, or cholestenone, resulting in t1/2 values of 24, 36, and 60 min, respectively. Increasing concentrations of whole serum and cholesteryl succinate have no significant effect on the apparent rate constant of inactivation of the enzyme, whereas its apparent rate of synthesis is decreased 3- and 10-fold, respectively. These results are reversed with oxygenated steroid inhibitors. The rate of synthesis of reductase is essentially unchanged as the concentrations of 25-hydroxycholesterol, 7-ketocholesterol, and cholestenone are increased in the culture medium, whereas the apparent rate constant for degradation is increased 9-, 7-, and 3-fold, respectively. HMG-CoA reductase activity in HTC cells thus appears to be modulated by two different mechanisms in which steroid structure is important. Whole serum and cholesteryl succinate specifically decrease the rate of enzyme synthesis, while 25-hydroxycholesterol, 7-ketocholesterol, and cholestenone increase the rate of inactivation of the reductase.

Alcohol Oxidoreductases↗
PubMed1990

Oxygenation of desmosterol and cholesterol in cell cultures.

In order to determine whether hydration of the delta 24 bond of desmosterol contributes to the formation of the regulatory oxysterol, 25-hydroxycholesterol, [3H]desmosterol was incubated with two cultured cell lines and the labeled products were analyzed. Small amounts of 25-hydroxycholesterol were formed with Chinese hamster lung (Dede) cell cultures, but not with mouse fibroblast (L) cell cultures. Apparently, desmosterol was converted into cholesterol, a process that does not occur in L cells, before 25-hydroxycholesterol takes place. No reliable evidence could be obtained for hydration of the delta 24 bond or for the reverse reaction upon incubation of [3H]25-hydroxycholesterol. Oxygenation of desmosterol occurred in both Dede and L cell cultures to give a mixture of 24(R)- and 24(S)-25-epoxy-cholesterol. This reaction, along with the production of 7-oxygenated sterols, may account for low levels of HMG-CoA reductase repressor activity previously found to be associated with delta 24 sterols.

Animals↗
PubMed1989

Loss of transcriptional repression of three sterol-regulated genes in mutant hamster cells.

Two genes that encode enzymes in cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase and HMG-CoA synthase, and the gene encoding the low density lipoprotein (LDL) receptor are repressed when sterols accumulate in animal cells. Their 5'-flanking regions contain a common element, designated sterol regulatory element-1 (SRE-1). In the HMG-CoA synthase and LDL receptor promoters, the SRE-1 enhances transcription in the absence of sterols and is inactivated in the presence of sterols. In the HMG-CoA reductase promoter, the region containing the SRE-1 represses transcription when sterols are present. In the current studies, we show that the SRE-1 retains enhancer function but loses sterol sensitivity in mutant Chinese hamster ovary cells that are resistant to the repressor, 25-hydroxycholesterol. In the absence of sterols, the mutant cells produced high levels of all three sterol-regulated mRNAs, and there was no repression by 25-hydroxycholesterol. When transfected with plasmids containing each of the regulated promoters fused to a bacterial reporter gene, the mutant cells showed high levels of transcription in the absence of sterols and no significant repression by sterols. When the SRE-1 in the LDL receptor and HMG-CoA synthase promoters was mutated prior to transfection into the mutant cells, transcription was markedly reduced. Thus, the 25-hydroxycholesterol-resistant cells retain a protein that enhances transcription by binding to the SRE-1 in the absence of sterols, but they have lost the function of a protein that abolishes this enhancement in the presence of sterols. Mutation of a 30-base pair segment of the HMG-CoA reductase promoter that contains the SRE-1 did not reduce transcription in the mutant cells, indicating that this promoter is driven by elements other than the SRE-1. Nevertheless, this promoter failed to be repressed by sterols in the mutant cells. These data suggest that a common factor mediates the effects of sterols on the SRE-1 in all three promoters and that this factor has been functionally lost in the 25-hydroxycholesterol-resistant cells.

Amphotericin B↗
PubMed1989

Control of low density lipoprotein receptor gene promoter activity. Ketoconazole inhibits serum lipoprotein but not oxysterol suppression of gene transcription.

We have examined the effects of ketoconazole, a drug which inhibits enzymes involved in cholesterol biosynthesis and metabolism, on the suppressive effects of serum lipoproteins and 25-hydroxycholesterol on low density lipoprotein (LDL) receptor gene promoter activity. A LDL receptor promoter-chloramphenicol acetyltransferase (CAT) fusion gene construct (pLDLR-CAT 6500) was transfected into JEG-3 choriocarcinoma cells, and the transfected cells were cultured in the absence or presence of serum, LDL, or serum and 25-hydroxycholesterol. Serum, LDL, and serum + 25-hydroxycholesterol reduced chloramphenicol acetyltransferase activity in cells transfected with pLDLR-CAT 6500, whereas these treatments had no effect upon enzyme activity in cells transfected with a control construct (pSV2CAT). Ketoconazole (50 microM) overcame the effects of serum and LDL on suppression of pLDLR-CAT 6500 expression, but could not override the combination of serum + 25-hydroxycholesterol. Ketoconazole had no significant effect on expression of pSV2CAT. The drug inhibited cholesterol side chain cleavage enzyme in the cells, but appeared to have no impact on the ability of cells to take up LDL-carried lipids. Our observations are consistent with the idea that serum lipoprotein cholesterol is metabolized to an effector substance which acts to suppress LDL receptor gene transcription. The generation of this effector seems to be sensitive to ketoconazole.

Cell Line↗
PubMed1987

Cytotoxicity of cholesterol oxides and their effects on cholesterol metabolism in cultured human aortic smooth muscle cells.

Incubation of monolayers of cultured human aortic smooth muscle cells with oxygenated sterols (25-hydroxycholesterol, 7-ketocholesterol, or cholesterol 5,6-epoxide) markedly inhibited growth though the viability of the culture was not affected. The effects on growth was concentration dependent, and 25-hydroxycholesterol was the most potent inhibitor of cellular growth as measured by decreased incorporation of thymidine into DNA and suppression of HMG-CoA reductase activity. The inhibitory effect of 25-hydroxycholesterol on cellular growth was not reversible if the cultures were grown in medium with normal fetal calf serum. However, in medium with delipidated serum, addition of purified cholesterol partially prevented growth inhibition induced by 25-hydroxycholesterol. Purified cholesterol, independently or in combination with tocopherol had no toxic effect on cellular growth. Addition of cholesterol oxides to the incubation medium stimulated lysosomal activation and release of acid phosphatase into the culture medium. The effect was concentration dependent and inversely related to cellular growth.

Acid Phosphatase↗
PubMed1985

Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase synthesis by a non-sterol mevalonate-derived product in Mev-1 cells. Apparent translational control.

A somatic cell mutant (Mev-1) auxotrophic for mevalonate by virtue of a complete lack of detectable 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase activity has been shown to demonstrate a requirement for a non-sterol mevalonate-derived product for regulation of synthesis of HMG-CoA reductase. A comparison of the effects of 25-hydroxycholesterol and the combination of 25-hydroxycholesterol and mevalonate on HMG-CoA reductase activity, synthesis, and mRNA levels in Mev-1 is presented in this report. The results show a close correlation between activity, rate of synthesis, and mRNA levels for Mev-1 cells treated with 25-hydroxycholesterol alone. Under the conditions of these experiments these effects are relatively small (approximately a 4-fold decrease). A much larger inhibition of HMG-CoA reductase activity and rate of synthesis (approximately 50-fold) is observed upon treatment of Mev-1 cells with a combination of 25-hydroxycholesterol and mevalonate. Yet, under these conditions mRNA levels are still reduced by only a factor of 4. These results are interpreted to suggest that the non-sterol mevalonate-derived regulatory product of HMG-CoA reductase acts by a translational control mechanism.

Animals↗
PubMed1984

Rapid modulation of platelet aggregation in plasma by oxygenated sterols.

The effect of oxygenated sterols on platelet aggregation induced by thrombin and ADP has been studied. All oxygenated sterols tested with a hydroxyl group on the side chain enhanced thrombin-induced aggregation at 25 microM. In the case of ADP-induced aggregation, however, only 22S-hydroxycholesterol[(22S)-5-cholestene-3 beta,22-diol] enhanced the aggregation, and 22R-hydroxycholesterol[(22R)-5-cholestene-3 beta,22-diol], 24S-hydroxycholesterol[(24S)-5-cholestene-3 beta,24-diol], and 25-hydroxycholesterol[5-cholestene-3 beta,25-diol] inhibited it. These effects were observed within 10 min after the addition of oxygenated sterols to platelet suspensions in plasma.

Adenosine Diphosphate↗