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PubMed1991

HMG CoA reductase inhibitors affect Na(+)-H+ antiport activity in human lymphoblasts.

The Na(+)-H+ antiport is a membrane-bound glycoprotein that extrudes intracellular acid loads and regulates cellular volume. Cellular synthesis of the oligosaccharide side chains of glycoproteins is dependent on a supply of mevalonate, itself a product of the rate-limiting enzyme of cholesterol synthesis 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase. The effect of two HMG CoA reductase inhibitors (simvastatin and 25-hydroxycholesterol) on intracellular pH and Na(+)-H+ exchange was therefore studied. Inhibition of the Na(+)-H+ antiport by these agents led to a fall in intracellular pH but did not impair the regulatory volume increase response to a hypertonic stimulus. The inhibitory effect of simvastatin was prevented by mevalonate but not dolichol or squalene. The effect of 25-hydroxycholesterol was more complex and not easily reversed. Thus HMG CoA reductase inhibitors reduced the ability of human lymphoblasts to expel an intracellular acid load via the Na(+)-H+ antiport, although the response of the antiport to an osmotic stimulus was preserved.

Blastomeres↗
PubMed2001

Fas/Fas ligand-mediated death pathway is involved in oxLDL-induced apoptosis in vascular smooth muscle cells.

Oxidized low-density lipoprotein (oxLDL) is a potent inducer of apoptosis for vascular cells. In the present study, we demonstrate that the expression of death mediators, including p53, Fas, and Fas ligand (FasL) was substantially upregulated by oxLDL in cultured vascular smooth muscle cells (SMCs). The induction of these death mediators was time dependent and was accompanied by an increase in apoptotic death of SMCs following oxLDL treatment. Two oxysterols, 7beta-hydroxycholesterol and 25-hydroxycholesterol, were also effective to induce the expression of death mediators and apoptosis. alpha-Tocopherol and deferoxamine significantly attenuated the induction of death mediators and cell death induced by oxLDL and oxysterols, suggesting that reactive oxygen species are involved in triggering the apoptotic event. Incubation of cells with FasL-neutralizing antibody inhibited the oxLDL-induced cell death up to 50%. Furthermore, caspase 8 and caspase 3 activities were induced time dependently in SMCs following oxLDL treatment. Collectively, these data suggest that the Fas/FasL death pathway is activated and responsible for, at least in part, the apoptotic death in vascular SMCs upon exposure to oxLDL.

Animals↗
PubMed2006

Modulation of human Niemann-Pick C1-like 1 gene expression by sterol: Role of sterol regulatory element binding protein 2.

Niemann-Pick C1-like 1 (NPC1L1) is an essential intestinal component of cholesterol absorption. However, little is known about the molecular regulation of intestinal NPC1L1 expression and promoter activity. We demonstrated that human NPC1L1 mRNA expression was significantly decreased by 25-hydroxycholesterol but increased in response to cellular cholesterol depletion achieved by incubation with Mevinolin (an inhibitor of 3-hydroxy-3-methylglutaryl-CoA reductase) in human intestinal Caco-2 cells. We also showed that a -1741/+56 fragment of the NPC1L1 gene demonstrated high promoter activity in Caco-2 cells that was reduced by 25-hydroxycholesterol and stimulated by cholesterol depletion. Interestingly, we showed that the NPC1L1 promoter is remarkably transactivated by the overexpression of sterol regulatory element (SRE) binding protein (SREBP)-2, suggesting its involvement in the sterol-induced alteration in NPC1L1 promoter activity. Finally, we identified two putative SREs in the human NPC1L1 promoter and established their essential roles in mediating the effects of cholesterol on promoter activity. Our study demonstrated the modulation of human NPC1L1 expression and promoter activity by cholesterol in a SREBP-2-dependent mechanism.

Base Sequence↗
PubMed1998

Absorption of cholesterol oxidation products from ordinary foodstuff in humans.

BACKGROUND: Information on the absorption of cholesterol oxidation products (COP) from ordinary foodstuff in humans is scarce. METHODS: Five healthy young men were offered a salami- and Parmesan-containing meal naturally rich in COP. Plasma and lipoprotein COP concentrations were measured over the following 9 h. RESULTS: The mean plasma free (nonesterified) COP concentration showed its maximal increase 3 and 5 h after meal consumption. In contrast, the raise in plasma total COP concentration began 6 h after the meal with a maximum at 8 h and was statistically significant for 7alpha- and 7beta-hydroxycholesterol and 7-ketocholesterol. The increase in plasma total cholesterol concentration was comparable to that of total COP. Comparing the COP composition of the chylomicrons and the test meal, cholestanetriol, 7-ketocholesterol, and to a lesser extent cholesterol-alpha-epoxide were underrepresented in the chylomicrons as was the opposite for 7beta-hydroxycholesterol. In very-low-density lipoprotein, a steady increase in the COP:cholesterol ratio was observed from 6 h on. CONCLUSION: COP from ordinary foodstuff were absorbed in the human intestinal tract but differences in the bioavailability of the single COP compounds were found.

Absorption↗
PubMed2002

ATP-binding cassette transporter 1 participates in LDL oxidation by artery wall cells.

OBJECTIVE: We have previously reported that products of the lipoxygenase pathway, hydroperoxyoctadecadienoic acid and hydroperoxyeicosatetraenoic acid, as well as cholesterol linoleate hydroperoxides, collectively termed seeding molecules, are removed by apolipoprotein A-I (apoA-I) from the artery wall cells and render low density lipoprotein (LDL) resistant to oxidation by human artery wall cells. The mechanisms by which oxidized lipids are transported and/or transferred to lipoproteins and the pathways by which apoA-I facilitates their removal remain unclear. ATP-binding cassette transporter 1 (ABCA1) is known to facilitate the release of cellular phospholipids and cholesterol from the plasma membrane to apoA-I and high density lipoprotein. Therefore, we evaluated whether ABCA1 participates in LDL oxidation. METHODS AND RESULTS: In this report, we show that (1) chemical inhibitors of ABCA1 function, glyburide and DIDS, block artery wall cell-mediated oxidative modification of LDL, (2) inhibition of ABCA1 with the use of antisense (but not sense) oligonucleotides prevents LDL-induced lipid hydroperoxide formation and LDL-induced monocyte chemotactic activity by the artery wall cells, and (3) oxysterols that induce ABCA1 expression, such as 22(R)hydroxycholesterol, enhance cell-mediated LDL oxidation. Furthermore, we also show that 22(R)hydroxycholesterol induces the production of reactive oxygen species in the artery wall cells, which can be removed by incubating the artery wall cells with apoA-I. CONCLUSIONS: Our data suggest that ABCA1 plays an important role in artery wall cell-mediated modification/oxidation of LDL by modulating the release of reactive oxygen species from artery wall cells that are necessary for LDL oxidation.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2004

Cytotoxicity of oxidized low-density lipoprotein in cultured RPE cells is dependent on the formation of 7-ketocholesterol.

PURPOSE: To determine which components present in oxidized LDL are responsible for the cytotoxicity associated with its internalization by culture ARPE19 cells. METHODS: ARPE19 cells were grown in 24-well and 96-well plates. Cell viability was measured by MTT and/or adenosine triphosphate (ATP) content. LDL was oxidized with Cu(+2) and oxysterol content analyzed by a novel HPLC method. RESULTS: OxLDL showed increased cytotoxicity with prolonged oxidation. Analysis of the oxLDL showed a predominance of the 7-oxygenated products, 7 alpha-hydroxycholesterol (7 alpha HCh), 7 beta-hydroxycholesterol (7 beta HCh), and 7-ketocholesterol (7kCh). Addition of these oxysterols to the ARPE19 cell in free form indicated that 7kCh is the most cytotoxic of the oxysterols but at physiologically unrealistic concentrations. Partitioning of individual oxysterols into nonoxidized LDL at concentrations similar to those found in the oxLDL also indicated that 7kCh is the most cytotoxic of the oxysterols. Transition metals are tightly bound by LDL and play an important role in the oxidation of LDL, but do not seem to enhance its cytotoxicity directly. CONCLUSIONS: Prolonged oxidation of LDL increases the levels of 7kCh due to further oxidation of 7 alpha HCh and 7 beta HCh. The formation of 7KCh seems to be responsible for most of the cytotoxicity associated with oxLDL internalization in ARPE19 cells.

Adenosine Triphosphate↗
PubMed1998

Oxysterols, but not cholesterol, inhibit human immunodeficiency virus replication in vitro.

Oxysterols, oxygenated derivatives of cholesterol selected for their cytostatic activity and their inhibitory effect on cholesterol synthesis, have been investigated for their anti-human immunodeficiency virus (HIV) activity in vitro. The three oxysterols tested, 7 beta-hydroxycholesterol (7 beta-OHC), 25-hydroxycholesterol (25-OHC) and 7 beta, 25-dihydroxycholesterol (7,25-OHC), inhibit viral replication at micromolar concentrations. The selectivity indexes for 7 beta-OHC and 25-OHC are quite modest (2 to 8) but reproducible; the dihydroxycholesterol 7,25-OHC exhibited antiviral properties at concentrations 13- to 25-fold lower than the highest concentration tested at which no toxicity was measurable. Oxysterols are naturally occurring compounds, and we speculate on their physiological relevance in HIV-infected individuals.

Anti-HIV Agents↗
PubMed2003

Side chain oxidized oxysterols in cerebrospinal fluid and the integrity of blood-brain and blood-cerebrospinal fluid barriers.

The side chain oxidized oxysterol 24S-hydroxycholesterol (24-OH-chol) is formed almost exclusively in the brain, and there is a continuous passage of this oxysterol through the circulation to the liver. 27-Hydroxycholesterol (27-OH-chol) is produced in most organs and is also taken up by the liver. The 27-OH-chol-24-OH-chol ratio is about 0.1 in the brain and about 2 in the circulation. This ratio was found to be about 0.4 in cerebrospinal fluid (CSF) of asymptomatic patients, consistent with a major contribution from the circulation in the case of 27-OH-chol. In accordance with this, we demonstrated a significant flux of deuterium labeled 27-OH-chol from plasma to the CSF in a healthy volunteer. Patients with a defective blood-brain barrier were found to have markedly increased absolute levels (up to 10-fold) of both 27-OH-chol and 24-OH-chol in CSF, with a ratio between the two sterols reaching up to 2. There was a significant positive correlation between the levels of both oxysterols in CSF and the albuminCSF-albuminplasma ratio. The 27-OH-cholCSF-24-OH-cholCSF ratio was found to be about normal in patients with active multiple sclerosis and significantly increased in patients with meningitis, polyneuropathy, or hemorrhages. Results are discussed in relation to the possible use of 24-OH-cholCSF as a surrogate marker of central nervous system demyelination and/or neuronal death.

Biomarkers↗
PubMed1985

Restoration effects of exogenous luteinizing hormone on the testicular steroidogenesis and Leydig cell ultrastructure.

In the present study, we explored the restoration effects of exogenous LH on Leydig cell ultrastructure and testicular steroidogenesis in rats that were deprived of endogenous LH via treatment with testosterone-17 beta-estradiol-filled Silastic implants for 10 days. Exogenous LH was supplied continuously via Alzet miniosmotic pumps at the rate of 1 microgram/h for 3, 6, 12, and 24 h or 1, 2, 4, 8, and 12 days. Testes were then perfused in vitro with medium containing 1) LH, 2) 20 alpha-hydroxycholesterol, or 3) pregnenolone substrate, which allowed us to assess LH-stimulated testosterone secretion, cholesterol side-chain cleavage activity, or the conversion of pregnenolone to testosterone, respectively. Other testes were perfusion fixed via the testicular artery for morphometric measurement of Leydig cell number and volume per testis and the surface area of Leydig cell cytoplasmic smooth endoplasmic reticulum (SER), inner mitochondrial membrane, and outer mitochondrial membrane. The results verified that Leydig cell smooth endoplasmic reticulum and inner and outer mitochondrial membrane surface areas are drastically diminished (P less than 0.05 vs. intact controls) by LH withdrawal. Also, the results verified that exogenous LH administered in situ restores Leydig cell ultrastructure and capacity to biosynthesize testosterone. However, the recovery of Leydig cell structure and steroidogenic reactions occurred at strikingly different rates upon restoration of LH after 10 days of the treatment with testosterone-17 beta-estradiol implants. For example, the restoration of testicular capacity to synthesize progesterone in response to LH stimulation or 20 alpha-hydroxycholesterol substrate was completed within 24 h. In contrast, the restoration of Leydig cell SER and testicular capacity to synthesize testosterone from pregnenolone was completed only after 8 days of continuous LH treatment (P greater than 0.05 vs. intact controls). Thus, our results show that LH rapidly restores Leydig cell post-LH receptor steroidogenic events up to and including cholesterol side-chain cleavage activity. Interestingly, there is no temporal association between the recovery of cholesterol side-chain cleavage activity and the surface area of inner mitochondrial membrane surface area. In contrast, 8 days are required to coincidentally restore SER surface area and the capacity of Leydig cells to synthesize testosterone from pregnenolone. We conclude that different cellular mechanisms are involved in the LH-dependent restoration of inner mitochondrial cholesterol side-chain cleavage activity and SER-associated conversion of pregnenolone to testosterone.

Animals↗
PubMed1988

Protein malnutrition in the domestic fowl induces alterations in adrenocortical function.

Adrenocortical function was investigated in immature, dietary protein-restricted domestic fowl (Gallus gallus domesticus). White Leghorn cockerels (2 weeks old) were fed isocaloric semipurified diets containing either 8% [low (L)] or 20% [normal (N)] soy protein for 4 weeks ad libitum. Cockerels were quickly killed by decapitation and exsanguination. Trunk plasma corticosterone (B) and ACTH levels were measured by RIA. Maximal B-binding capacity (CBC) of plasma was also measured. In addition, in randomly selected cockerels, a rough index of the B clearance rate was determined. Finally, to determine the influence of protein malnutrition on adrenocortical cell function per se, we measured the acute (2-h) B responses of highly enriched adrenocortical cell populations to various ACTH analogs, 8-bromo-cAMP (8-Br-cAMP), and cellular B production maximally supported with 25-hydroxycholesterol. Plasma B and ACTH concentrations of L cockerels were, respectively, 160% greater and 32% less than those of N cockerels. In addition, plasma B clearance rate of L birds was 85% greater than that of N birds, thus suggesting a greater B secretion rate in L birds. However, maximal plasma CBC of L cockerels was 59% less than that of N cockerels. Thus, the free plasma B concentration of L birds was greater than that of N birds. The increase in the plasma B concentration of L cockerels is explained in part by the relative adrenal weight of these birds which was 88% greater than that of N cockerels. In addition, there were differences at the adrenocortical cell level. On an equal cell concentration basis, basal and maximal B production values (stimulated by ACTH analogs and 8-Br-cAMP, and supported by 25-hydroxycholesterol) of L cockerel adrenocortical cells were, respectively, 73% and 139% greater than those of N cockerel adrenocortical cells. In addition, maximal ACTH-induced aldosterone production of L bird cells was 104% greater than that of N bird cells, such that the ratio of aldosterone production to B production was not altered by protein deprivation. The data suggest that the greater steroidogenic capacity of L cockerel cells was due to an increase in intracellular steroidogenic enzyme content and/or activity and not to an alteration in the composition of adrenocortical cell types within the populations of isolated cells. Furthermore, ACTH analog ED50 values for B, aldosterone, and cAMP production by L bird cells were about one third to one fifth the values for N bird cells, thus indicating that L bird cells were about 3-5 times more sensitive to ACTH than were N bird cells.(ABSTRACT TRUNCATED AT 400 WORDS)

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1992

Sites of interaction between epidermal growth factor and transforming growth factor-beta 1 in the control of steroidogenesis in cultured porcine Leydig cells.

The present study examines how the hormonal action of gonadotropin is modulated by transforming growth factor-beta 1 (TGF beta 1) and epidermal growth factor (EGF) in primary cultures of purified porcine Leydig cells. Although TGF beta 1 (1 ng/ml; 48 h) and EGF (10 ng/ml; 72 h) individually enhanced hCG-stimulated testosterone formation, the effects of EGF were more pronounced than those of TGF beta 1. When studied in combination, the effects of maximal concentrations of TGF beta 1 and EGF were additive on gonadotropin hormonal action. In the present study we demonstrate that their additive effects resulted from a complex interaction occurring at the levels of cholesterol substrate availability in the mitochondria and of 3 beta-hydroxysteroid dehydrogenase/isomerase activity (3 beta HSDI). First, TGF beta 1 (1 ng/ml; 48 h) and EGF (10 ng/ml; 72 h) were, respectively, shown to reduce and enhance dehydroepiandrosterone (DHEA) formation (evaluated in the presence of 10(-5) M WIN 24540, an inhibitor of 3 beta HSDI) in Leydig cells when acutely (3 h) stimulated with hCG (0.01-1 ng/ml), but not when incubated with 22R-hydroxycholesterol (3 micrograms/ml). Such findings indicate that TGF beta 1 and EGF did not affect cholesterol side-chain cleavage cytochrome P450 activity, but, respectively, decreased and increased cholesterol substrate availability for this enzyme in the mitochondria. Furthermore, when Leydig cells were treated with the combined factors, the formation of delta 5-steroid intermediates (such as DHEA) in untreated (control) and EGF-plus TGF beta 1-treated cells was not significantly different whether the cells were acutely stimulated with the gonadotropin or incubated with 22R-hydroxycholesterol. Such findings indicate that the effects of EGF and TGF beta 1 on cholesterol substrate availability in the mitochondria are antagonistic. Second, EGF, TGF beta 1, and EGF plus TGF beta 1 significantly (P less than 0.001) increased delta 5-steroid intermediate (i.e. pregnenolone and DHEA), but not delta 4-steroid intermediate (i.e. progesterone and androstenedione), conversion into testosterone, indicating that the growth factors increased, individually or in combination in an additive manner, 3 beta HSDI activity (respectively, 90.7 +/- 0.6%, 80.6 +/- 2.6%, and 164 +/- 4.5% increase in the presence of EGF, TGF beta 1, and EGF plus TGF beta 1). Together, the reciprocal suppression of the effects of TGF beta 1 and EGF on the mitochondrial cholesterol substrate availability coupled to their stimulatory additive actions on 3 beta HSDI activity provide an explanation of the additive actions of the two growth factors on gonadotropin-induced testicular androgen formation.

Androstenedione↗
PubMed1996

Ouabain production by cultured adrenal cells.

The adrenal cortex releases a sodium pump inhibitor. The present studies tested whether this material was endogenous and identical to ouabain by 1) studying the production of ouabain in long term cultures of adrenocortical cells, 2) seeking evidence that ouabain might be taken up from exogenous sources by adrenocortical cells, 3) examining the release of adrenocortical cells loaded with exogenous ouabain, 4) attempting to stimulate ouabain steroidogenesis in cultured adrenocortical cells, and 5) performing further chemical analysis on ouabain immunoreactivity released by cultured adrenocortical cells. Our results indicate that ouabain immunoreactivity is present in conditioned medium from both murine Y-1 adrenocortical cultures and primary bovine adrenocortical cell (BAC) cultures. We also found that BACs bind and internalize [3H]ouabain. Bound [3H]ouabain is released from BACs by both receptor dissociation and cytoplasmic release of internalized [3H]ouabain. Only one isoform of membrane sodium, potassium-adenosine triphosphatase, alpha 1, was expressed in the adrenal. Authentic ouabain was not metabolized during membrane binding or while present intracellularly. Stimulation of steroidogenesis in Y-1 and BAC with 22R-hydroxycholesterol and 25-hydroxycholesterol was performed and confirmed increased steroidogenesis; however, there was no effect on ouabain immunoreactivity content or release. Comparison of the ouabain binding density in cultured BAC, hepatoma cells, and 3T3 fibroblasts indicated that adrenocortical cells have a high ouabain-binding capacity. HPLC studies of the ouabain immunoreactivity released by bovine adrenocortical cells indicated that essentially no authentic ouabain was secreted. The present studies confirm that both BAC and Y-1 cultures release a ouabain-like material that differs in structure from authentic plant ouabain and is not a product of cholesterol side-chain cleavage.

Adrenal Cortex↗
PubMed2004

Diagnostic use of cerebral and extracerebral oxysterols.

BACKGROUND: 24S-Hydroxycholesterol (24OHC) and 27-hydroxycholesterol (27OHC) are two structurally similar oxysterols of different origins--the former almost exclusively formed in the brain and the latter formed to a lesser extent in the brain than in most other organs. HYPOTHESIS TO BE TESTED: Neuronal damage and/or demyelination causes increased flux of 24OHC from the brain into the cerebrospinal fluid (CSF), whereas a defect blood-brain barrier causes increased flux of 27OHC from the circulation into the CSF. METHODS: Isotope dilution-mass spectrometry was used to assay the two oxysterols in CSF and plasma from more than 250 patients with different neurological and geriatric diseases. RESULTS: The CSF-levels of the two oxysterols were much more affected by the different diseases than the plasma levels. Patients with active demyelinating diseases had increased levels of 24OHC in CSF with a relatively high 24OHC/27OHC ratio. Patients with meningitis in general had high levels of both steroids with a low 24OHC/27OHC ratio. Patients with Alzheimer's disease had slightly increased levels of 24OHC in CSF with less increase in 27OHC. Patients with multiple sclerosis had a tendency to have higher levels of 24OHC during active periods with a high 24OHC/ 27OHC ratio. CONCLUSIONS: Measurements of the two oxysterols in CSF and plasma may add significantly to existing biochemical methods for evaluation of neurological diseases.

Adult↗
PubMed1986

The interaction of hCG, hydroxysteroids and interstitial fluid on rat Leydig cell steroidogenesis in vitro.

Rat testicular interstitial fluid and hydroxycholesterol both stimulated testosterone production by isolated Leydig cells in vitro in a dose-dependent manner, but the dose-response lines were not parallel. The addition of cycloheximide blocked the stimulation by interstitial fluid but not that of hydroxycholesterol. Use of the compounds SU 10603 and cyanoketone (which inhibit 3 beta-hydroxysteroid dehydrogenase and 17 alpha-hydroxylase respectively) or aminoglutethimide (which acts on the cholesterol side-chain cleavage enzyme) showed that the stimulatory factor(s) in interstitial fluid stimulated steroidogenesis at the cholesterol side-chain cleavage enzyme, before the conversion of pregnenolone. This enzyme is rate-limiting in the synthesis of testosterone by Leydig cells and a site of action of LH; therefore, these results support the view that an interstitial fluid factor may be involved in the paracrine regulation of testicular steroidogenesis.

Animals↗
PubMed1992

Maintenance or stimulation of steroidogenic enzymes and testosterone production in rat Leydig cells by continuous and pulsatile infusions of luteinizing hormone during passive immunization against gonadotrophin-releasing hormone.

The importance of the pulsatility of luteinizing hormone (LH) secretion in maintaining key enzymes in the testosterone biosynthetic pathway in Leydig cells was studied using rats in which LH secretion was suppressed by passive immunization against gonadotropin-releasing hormone (GnRH) and replaced by continuous or pulsatile i.v. infusions of exogenous LH, all delivering the same daily dose of the hormone (300 ng per 100 g NIDDK-ovine LH-24). Continuous infusions (12.5 ng per 100 g h-1) were compared with infusions of 1 min pulses every 2 h (25 ng per 100 g) and every 4 h (50 ng per 100 g). After 5 days of treatment in vivo with sheep anti-GnRH serum (or normal sheep serum) and LH (or vehicle), Leydig cells were purified and assayed in vitro for maximum production of testosterone stimulated by human chorionic gonadotrophin (hCG) and supported by 25-hydroxycholesterol and for the activities of cholesterol side-chain cleavage, delta 5-3 beta-hydroxysteroid dehydrogenase-delta 5-4-isomerase (3 beta-HSD-isomerase) and 17 alpha-hydroxylase. Relative contents of cholesterol side-chain cleavage and 17 alpha-hydroxylase were also quantified by western and immunoblotting analysis. Activity of 3 beta-HSD-isomerase was reduced by about 40% by anti-GnRH treatment and was increased by all LH regimens in anti-GnRH-treated animals, with no consistent pattern in the effects of the different LH regimens. Results for testosterone-producing capacity and the other two enzymes differed in several respects. Treatment with anti-GnRH serum markedly reduced basal, hCG-stimulated and 25-hydroxycholesterol-supported testosterone production (by 80-90%) and the activities of cholesterol side-chain cleavage (about 80%) and 17 alpha-hydroxylase (about 65%). Infusion of exogenous LH in any of the regimens tested prevented these changes or increased the activities to values greater than those in normal serum-treated controls. Differences in immunodetectable contents of the two enzymes generally paralleled those in enzyme activities. There was a consistent trend in the effects of LH replacement regimens on these parameters of steroidogenic activity: continuous infusions were more effective than pulses at 2 h intervals and these in turn were more effective than pulses at 4 h intervals, suggesting that the frequency of LH exposure is more important than the amplitude of individual exposures in maintaining Leydig cell steroidogenic function.(ABSTRACT TRUNCATED AT 400 WORDS)

3-Hydroxysteroid Dehydrogenases↗
PubMed1983

The regulating effect of cholesterol derivatives isolated from human sera on lymphocyte response to phytohemagglutinin.

Cholesterol derivatives, including 7 alpha-hydroxycholesterol (7 alpha-HC), 7 beta-hydroxycholesterol (7 beta-HC), and 7-ketocholesterol (7-KC), were isolated from the Cohn fraction IV-I of normal human sera and were shown to be the moieties responsible for the suppression of lymphocyte proliferation. When 5-10 micrograms/ml of 7 alpha-HC, 7 beta-HC or 7-KC were added in vitro to lymphocyte cultures, incorporation of 3H-thymidine (3H-TdR) into the lymphocyte DNA was suppressed by about 50% when compared with the control. When 10-50 mg/kg/day of 7 alpha-HC, 7 beta-HC and 7 KC were administered in vivo to skin grafts on rats, the prolongation of skin grafts was observed. The concentration of 7-KC in cancer patient sera was 2 to 3 times the amount in normal pooled human sera.

Animals↗
PubMed1996

Insulin-like growth factor I enhancement of steroidogenesis by bovine granulosa cells and thecal cells: dependence on de novo cholesterol synthesis.

Studies were conducted to determine the importance of de novo cholesterol synthesis and cholesterol side-chain cleavage enzyme in the action of IGF-I in bovine granulosa and thecal cells. Granulosa and thecal cells from bovine follicles were cultured for 2 days in 10% fetal calf serum and then treated with luteinizing hormone (100 ng/ml) and IGF-I (0 or 100 ng/ml) for an additional 2 days in serum-free medium. During the last 24 h of treatment, cells were concomitantly treated with simvastatin (0, 0.5 or 5 micrograms/ml) or 25-hydroxycholesterol (0 or 10 micrograms/ml). Simvastatin, a potent inhibitor of the key enzyme controlling de novo cholesterol synthesis, 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, completely inhibited (P < 0.05) progesterone production by granulosa cells and progesterone and androstenedione production by thecal cells. Simvastatin also inhibited (P < 0.05) granulosa cell and thecal cell proliferation. Concomitant treatment with mevalonate, an immediate product of HMG-CoA reductase, attenuated the inhibitory effect of simvastatin on progesterone and androstenedione production by thecal cells and blocked the inhibitory effect of simvastatin on cell proliferation. The addition of 25-hydroxycholesterol, a substrate for cholesterol side-chain cleavage enzyme, had no effect (P > 0.10) on IGF-I-stimulated progesterone or androstenedione production by thecal cells and actually inhibited (P < 0.05) IGF-I-stimulated progesterone production by granulosa cells. These results provide indirect evidence indicating that stimulation of HMG-CoA reductase is an important locus of IGF-I action in bovine granulosa and thecal cells, whereas IGF-I has little or no effect on side-chain cleavage enzyme activity in these same cell types under the culture conditions employed.

Androstenedione↗
PubMed1995

Depletion of alpha-tocopherol in human atherosclerotic lesions.

Estimations of alpha-tocopherol content were made on a series of human necropsy samples of normal arterial wall and of atherosclerotic lesions. The results were compared with stage of lesion, shown by histology, and with the amounts of cholesterol and hydroxycholesterols in the same lesions. The ratio of alpha-tocopherol to cholesterol levels varied widely in normal arterial wall but was consistently low in lesions, especially in lesions rich in macrophage foam cells. The results suggested that significant accumulation of hydroxycholesterols, found almost exclusively in lesions, only occurred when alpha-tocopherol levels were low in relation to the cholesterol content. This suggests that oxidative activity in the lesion may lead to significant oxidation of constituents of low-density lipoprotein only after alpha-tocopherol has been depleted.

Adolescent↗