HMG-CoA Reductase Protein Expression Research Articles

Amyloid Beta sharply increases HMG-CoA reductase protein levels in astrocytes isolated from C57BL/6 mice

IntroductionIt is believed that the accumulation of Amyloid Beta (Aβ) and brain cholesterol are involved in Alzheimer's pathophysiology. Consequently, regulation of brain cholesterol is greatly important. In astrocytes, like other cells, HMG-CoA reductase plays a key role in the maintenance of normal cholesterol level. In this study expression and protein levels of HMG-CoA reductase is assessed in the presence and absence of Aβ in mouse astrocytes. MethodsMouse astrocytes were cultured and treated with Aβ for 24 h. Protein expression of HMG-COA reductase was detected using western blotting. Moreover, real-time PCR was executed to determine the alterations in the HMG-COA reductase expression. ResultsStatistical analysis of results showed that the protein levels of HMG-COA reductase in treated group with Aβ has increased by 4-fold, while no significant changes was observed in the mRNA of the HMG-COA reductase gene compared to the control group. ConclusionOur findings showed that Aβ can sharply increase protein level of HMG-COA reductase which can potentially increase cholesterol synthesis. Therefore, Aβ may somehow increase HMG-COA reductase stability which could be resulted in excessive cholesterol build-up and in the other side increasing of cholesterol leads to an increase in Aβ deposition. We showed a new insights on the interplay between Aβ and HMG-COA reductase which may be a scientific and reasonable explanation for over accumulation of cholesterol in Alzheimer's disease.

NPY stimulates cholesterol synthesis acutely by activating the SREBP2-HMGCR pathway through the Y1 and Y5 receptors in murine hepatocytes

AimsThe development of non-alcoholic fatty liver disease (NAFLD) is partially attributed to disturbance in cholesterol metabolism and sympathetic overactivity. Excessive levels of the sympathetic neurotransmitter neuropeptide Y (NPY) positively correlated with both NAFLD and cholesterol accumulation. We wanted to determine, for the first time, whether NPY promotes cholesterol accumulation directly in hepatocytes and elucidate the underlying mechanism. Main methodsIn vivo, NPY was injected through the hepatic portal vein into SD rats. One hour later, serum and liver tissues were collected. In vitro, BRL-3A hepatocytes were treated with NPY, and with Y1, Y2, Y5, receptor antagonists as well as with extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) antagonist, respectively. Cholesterol content was measured by coupled enzyme method. Precursor sterol-regulatory element binding protein 2 (pSREBP2), mature SREBP2 (mSREBP2), HMG-CoA reductase (HMGCR), ERK1/2, pERK1/2, cAMP-dependent protein kinase (PKA), and pPKA protein expression levels were examined by western blotting. Key findingsIn rats, intraportal vein injection of NPY activates pSREBP2, mSREBP2, and HMGCR protein expression, and induces hepatic cholesterol accumulation. In BRL-3A cells, we observed that NPY increases cholesterogenic protein expression and cholesterol synthesis through Y1 and Y5 receptors. This effect is mediated by the activation of the ERK1/2 signaling pathway. SignificanceWe demonstrated, for the first time, that NPY can activate the cholesterogenic pathway and elucidated the underlying mechanism. Thus, NPY and NPY receptors might be new targets for the treatment of NAFLD and dyslipidemia.

Lactoferrin Alleviates the Progression of Atherosclerosis in ApoE-/- Mice Fed with High-Fat/Cholesterol Diet Through Cholesterol Homeostasis.

Lactoferrin (LF) is a multifunctional glycoprotein and has beneficial effects on the regulation of lipid metabolism. However, whether LF supplementation alleviates the development of atherosclerosis (AS) remains unclear. In the present study, all of 48 male Apolipoprotein E-/- mice were fed with high-fat diet with 1.25% added cholesterol and divided to four treatment groups with either distilled water (HFCD), LF solutions at 2 mg/mL (low LF), 10 mg/mL (middle LF or MLF), or 20 mg/mL (high LF or HLF) for 12 weeks. Oral glucose tolerance tests (OGTT) were performed at weeks 0, 4, 8, and 12. At the end of the experiment, lipids in serum, liver, and feces were determined. The livers, whole aortas, and aortic sinuses were pathologically examined. The protein expression of factors related to cholesterol synthesis, absorption, and excretion were detected through western blot. No significant difference in body weight, food intake, and OGTT was observed among the four groups. Compared with the HFCD group, the MLF and HLF groups had significantly decreased serum and hepatic cholesterol levels and significantly increased fecal cholesterol contents. LF alleviated the hepatic steatosis and lipid droplet, especially in the MLF group. LF also significantly decreased the average lesion areas in the whole aorta, especially in the MLF group. On the other hand, LF downregulated hepatic protein expression of HMG-CoA reductase (the rate-limiting enzyme in cholesterol synthesis) and upregulated cholesterol 7-alpha hydroxylase (the rate-limiting enzyme in bile acid synthesis from cholesterol). LF also downregulated the intestinal expression of Niemann-Pick C1-like 1 protein, which is known to bind to a critical mediator of cholesterol absorption. In conclusion, LF supplementation alleviates the AS in mice on HFCD likely by reducing the synthesis and absorption of cholesterol and increasing cholesterol excretion.

Chlorogenic acid-enriched extract from Eucommia ulmoides leaves inhibits hepatic lipid accumulation through regulation of cholesterol metabolism in HepG2 cells

Context: Eucommia ulmoides Oliver (Eucommiaceae) leaf exhibits beneficial lipid-lowering and anti-obesity effects. However, the mechanisms remain unknown.Objective: The objective of this study is to investigate the lipid-lowering effects of chlorogenic acid (CGA)-enriched extract from this plant (CAEF) in human hepatoma HepG2 cells, focusing on cholesterol metabolism.Materials and methods: HepG2 cells were treated with CAEF (10, 20, 25, 40, 60, and 80 mg/L), CGA (0.3, 3, 30, 300, and 600 μmol/L), and simvastatin (0.1, 1, 10, 50, and 100 μmol/L) for 24 or 48 h. The cytotoxicity, Oil red O staining, total cholesterol, and triacylglycerol in supernatants were determined. The mRNA expression of genes involved in cholesterol metabolism was determined with RT-PCR. The protein expression of HMG-CoA reductase (HMGCR) was examined by immunocytochemistry and western-blot.Results: The IC50 values were 59.2 mg/L for CAEF, 335.9 μmol/L for CGA, and 10.5 μmol/L for simvastatin. By treating cells with CAEF (25 mg/L), CGA (30 μmol/L), or simvastatin (10 μmol/L) for 48 h, the efflux of total cholesterol and triacylglycerol was increased (CAEF, 4.06- and 31.00-folds; CGA, 2.94- and 2.17-folds; and simvastatin, 3.94- and 24.67-folds), and the cellular lipid droplets were reduced in Oil red O staining. CAEF and CGA increased mRNA expression of ABCA1, CYP7A1, and AMPKα2, while CAEF and simvastatin decreased SREBP2. However, their effects on LXRα mRNA expression were variable. Importantly, all drugs significantly inhibited protein expression of HMGCR at mRNA and protein levels.Discussion and conclusion: CAEF is a promising dietary supplement to prevent obesity and dyslipidemia and the effects appear to be due, at least in part, to regulating cholesterol metabolism through inhibition of HMGCR in HepG2 cells.

The target for statins, HMG-CoA reductase, is expressed in ductal carcinoma-in situ and may predict patient response to radiotherapy.

Patients with ductal carcinoma-in-situ (DCIS) are currently not prescribed adjuvant systemic treatment after surgery and radiotherapy. Prediction of DCIS patients who would benefit from radiotherapy is warranted. Statins have been suggested to exert radio-sensitizing effects. The target for cholesterol-lowering statins is HMG-CoA reductase (HMGCR), the rate-limiting enzyme in the mevalonate pathway. The aim of this study was to examine HMGCR expression in DCIS and study its treatment predictive value. A population-based cohort including 458 women diagnosed with primary DCIS between 1986 and 2004 were followed until November 2011 to study long-term survival. Tumor tissue microarrays were constructed, and immunohistochemical analyses were performed to detect cytoplasmic protein expression of HMGCR. The association between DCIS HMGCR expression and invasive breast cancer recurrence-free survival (RFSinv) and overall survival (OS) was analyzed by Kaplan-Meier curves, log rank test, and Cox proportional hazard analysis. HMGCR was strongly expressed in 24 % of the assessed DCIS samples, moderately expressed in 46 %, and weakly expressed in 23 %; no expression was detected in 7 % of the samples. During the follow-up time (median 13.8 years), 61 patients were diagnosed with an invasive breast cancer recurrence, and 80 patients died. A crude analysis showed no survival benefit from radiotherapy. However, patients with strong HMGCR expression showed an improved RFSinv (log rank, p = 0.03) and OS (log rank, p = 0.04) after radiotherapy. No statistically significant interaction was observed for HMGCR and radiotherapy (RFSinv p = 0.69 and OS p = 0.29). This study demonstrates HMGCR expression in DCIS and suggests HMGCR as a predictive marker of response to postoperative radiotherapy in DCIS, although the test for interaction was nonsignificant. Future DCIS studies addressing the potential of statin treatment targeting HMGCR are warranted.

Single dose testosterone increases total cholesterol levels and induces the expression of HMG CoA Reductase

BackgroundCholesterol is mainly synthesised in liver and the rate-limiting step is the reduction of 3-hydroxy-3methylglutaryl coenzyme A (HMG-CoA) to mevalonate, a reaction catalysed by HMG-CoA reductase (HMGCR). There is a comprehensive body of evidence documenting that anabolic-androgenic steroids are associated with deleterious alterations of lipid profile. In this study we investigated whether a single dose of testosterone enanthate affects the cholesterol biosynthesis and the expression of HMGCR.Methods39 healthy male volunteers were given 500 mg testosterone enanthate as single intramuscular dose of Testoviron®--Depot. The total cholesterol levels prior to and two days after testosterone administration were analysed. Protein expression of HMGCR in whole blood was investigated by Western blotting. In order to study whether testosterone regulates the mRNA expression of HMGCR, in vitro studies were performed in a human liver cell-line (HepG2).ResultsThe total cholesterol level was significantly increased 15% two days after the testosterone injection (p = 0.007). This is the first time a perturbation in the lipoprotein profile is observed after only a single dose of testosterone. Moreover, the HMGCR mRNA and protein expression was induced by testosterone in vitro and in vivo, respectively.ConclusionHere we provide a molecular explanation how anabolic androgenic steroids may impact on the cholesterol homeostasis, i.e. via an increase of the HMGCR expression. Increasing knowledge and understanding of AAS induced side-effects is important in order to find measures for treatment and care of these abusers.

Linalool reduces the expression of 3-hydroxy-3-methylglutaryl CoA reductase via sterol regulatory element binding protein-2- and ubiquitin-dependent mechanisms

We investigated hypocholesterolemic mechanisms of linalool, an aromatic anti-oxidative monoterpene, which is abundant in teas and essential oils. Oral administration of linalool to mice for 6weeks significantly lowered total and low-density lipoprotein cholesterol concentrations, and HMG-CoA reductase protein expression (−46%; P<0.05) by both transcriptional and posttranscriptional mechanisms. Linalool suppressed the gene expression of HMG-CoA reductase by reducing the binding of SREBP-2 to its promoter, as assessed by qPCR and chromatin immunoprecipitation, and by inducing ubiquitin-dependent proteolysis of the HMG-CoA reductase. These findings suggest that food molecules with a pleasant scent could exert beneficial metabolic effects through multiple mechanisms.

Nutrigenomic analysis of hypolipidemic effects of Agastache rugosa essential oils in HepG2 cells and C57BL/6 mice

Hypolipidemic effects of Agastache rugosa essential oil (AREO) were investigated. Gas chromatographymass spectrometry (GC-MS) analysis showed that the major compound in AREO is limonene (47%, w/w). AREO (1 mg/mL) markedly reduced low density lipoprotein (LDL) oxidation (−93%). After 3-week feeding of AREO, plasma cholesterol (−28%) and triglyceride levels (−26%) were significantly decreased in C57BL/6J mice. Mouse hepatic transcriptome profiling with oligonucleotide microarray revealed that AREO altered the expression of 2,524 genes. Notably, significant reductions in sterol regulatory element binding factor (SREBF)-1 and SREBF-2 mRNA levels were detected. Protein expression of HMG-CoA reductase, a major target for SREBP-2, was reduced in HepG2 cells (−36%) and in mouse liver (−35 %). AREO also significantly increased mRNA (+40%) and protein (+83%) expression of the LDL receptor in HepG2 cells and mouse liver, respectively. Our findings suggest that AREO may prevent atherosclerosis via the inhibition of LDL oxidation, down-regulation of SREBF-2 and HMG-CoA reductase expression, and up-regulation of LDL receptor expression.

Novel hypoglycemic effects of Ganoderma lucidum water-extract in obese/diabetic (+db/+db) mice

In this study, we evaluated the pharmacological effects of Ganoderma lucidum (G. lucidum) (water-extract) (0.003, 0.03 and 0.3g/kg, 4-week oral gavage) consumption using the lean (+db/+m) and the obese/diabetic (+db/+db) mice. Different physiological parameters (plasma glucose and insulin levels, lipoproteins-cholesterol levels, phosphoenolpyruvate carboxykinase (PEPCK), 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG CoA reductase) and isolated aorta relaxation of both species were measured and compared. G. lucidum (0.03 and 0.3g/kg) lowered the serum glucose level in +db/+db mice after the first week of treatment whereas a reduction was observed in +db/+m mice only fed with 0.3g/kg of G. lucidum at the fourth week. A higher hepatic PEPCK gene expression was found in +db/+db mice. G. lucidum (0.03 and 0.3g/kg) markedly reduced the PEPCK expression in +db/+db mice whereas the expression of PEPCK was attenuated in +db/+m mice (0.3g/kg G. lucidum). HMG CoA reductase protein expression (in both hepatic and extra-hepatic organs) and the serum insulin level were not altered by G. lucidum. These data demonstrate that G. lucidum consumption can provide beneficial effects in treating type 2 diabetes mellitus (T2DM) by lowering the serum glucose levels through the suppression of the hepatic PEPCK gene expression.

Acute P-407 Administration to Mice Causes Hypercholesterolemia by Inducing Cholesterolgenesis and Down-Regulating Low-Density Lipoprotein Receptor Expression

Poloxamer 407 (P-407) is a chemical that induces a dose-controlled dyslipidemia in mice. Our aim was to determine the acute effects of P-407 treatment on the mechanisms that influence hepatic cholesterol homeostasis. We measured lipid levels in plasma and liver samples from control and P-407-treated mice (24 h post-i.p. injection of 0.5 g kg(-1) of P-407 or saline for the control mice). We measured acyl-coenzyme A:cholesterol acyltransferase (ACAT) and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activities in liver microsomes. The protein expression of ACAT2, scavenger receptor class B, type I (SR-BI), ATP-binding cassette transporter A1 (ABCA1), ATP-binding cassette transporter G8 (ABCG8), low-density lipoprotein receptor (LDLr), and actin was measured by immunoblot. We found an increase in plasma cholesterol and triglyceride levels as well as increased hepatic cholesteryl esters (CE) in P-407-treated mice. The hepatic ACAT microsomal activity and ACAT2 protein expression were not altered by P-407. The protein expression of the LDLr was decreased in the livers of P-407-treated mice. This decrease was specific, because the expression of the SR-BI was unchanged. The P-407-induced hypercholesterolemia was accounted for by increased activity and protein expression of HMG-CoA reductase. ATP-binding cassette transporters A1 and G8 protein expression were not significantly different in P-407-treated mice compared to controls. The increased hepatic CE levels, following P-407 treatment, was neither related to an up-regulation of ACAT2 nor enhanced SR-BI expression. Hypercholesterolemia was associated with an up-regulation of both the protein expression and activity of HMG-CoA reductase and decreased LDLr expression.