HMGCR Protein Expression Research Articles

Oleanolic acid inhibits cholesterol synthesis by suppressing the expression and enzymatic activity of HMGCR

ABSTRACT This study investigated the impact of Oleanolic Acid (OA) on cholesterol metabolism and. MTT assays demonstrated that OA had no cytotoxic effects on HepG2 cells at concentrations below 20 μM. Subsequent experiments focused on 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCR), a key enzyme in cholesterol biosynthesis. OA significantly reduced HMGCR protein expression without affecting other enzymes. Further investigations revealed that OA reduced the protein and mRNA expression of HMGCR and its upstream regulator Sterol regulatory element binding protein-2 (SREBP-2) in HepG2 cells. OA activated Adenosine 5’-monophosphate (AMP)-activated protein kinase (AMPK), enhancing HMGCR phosphorylation and inhibiting its activity. Molecular docking experiments suggested that OA could suppress HMGCR enzymatic activity through hydrophobic interactions, reducing catalytic efficiency. Overall, OA downregulated HMGCR expression and activity, leading to decreased cholesterol synthesis. These findings highlight OA’s regulatory role in cholesterol metabolism and its potential for improving atherosclerosis (AS), supporting its application in functional food development.

Ginsenoside F1 inhibits cholesterol overload in oxidative-damaged cells through SREBP2/HMGCR pathway

To investigate the inhibitory mechanisms of ginsenoside F1 on hydrogen peroxide induced cholesterol metabolism disorder and oxidative stress in HepG2 cells. 1, 1-diphenyl-2-picrylhydrazyl(DPPH) and oxygen radical absorbance capacity(ORAC) tests were used to detect the scavenging effect of ginsenoside F1 on nitrogen and oxygen free radicals. HepG2 cells were treated with 400 μmol/L hydrogen peroxide and pretreated with 10, 20 and 40 μmol/L ginsenoside F1. Mitochondrial membrane potential(MMP) and total cholesterol levels were detected by JC-1 method and cholesterol kit, respectively. The protein expression levels of sterol-regulatory element binding proteins(SREBP2)and 3-hydroxy-3-methylglutaryl coenzyme A reductase(HMGCR) in cholesterol synthesis pathway were detected by Western blot. The DPPH clearance rate of ginsenoside F1 was much lower than that of 6-hydroxy-2, 5, 7, 8-tetramethylchroman-2-carboxylic acid(Trolox), but the ORAC capability of ginsenoside F1 was stronger, which was comparable to Trolox. The MMP and protein expression of SREBP2 were significantly decreased in injured group(P<0.05). The cholesterol and protein expression of HMGCR were significantly increased(P<0.05). Whereas, compared with the injured group, the MMP and protein expression of SREBP2 were significantly increased after 10, 20 and 40 μmol/L ginsenoside F1 pretreatment of injured cells(P<0.05). The cholesterol level and protein expression of HMGCR were significantly lower than injured group with concentration-dependent decreases(P<0.05). Ginsenoside F1 can protect against hydrogen peroxide induced oxidative stress in HepG2 cells by inhibiting oxygen free radicals and protecting mitochondria. And its mechanism may be related to the intervention of SREBP2/HMGCR pathway in regulating cellular cholesterol anabolism.

Paternal cadmium exposure affects estradiol synthesis by impairing intracellular cholesterol homeostasis and mitochondrial function in offspring female mice

Cadmium (Cd) is a toxic heavy metal commonly found in nature and an endocrine disrupting chemical (EDC). Previous studies found that Cd can damage several organs, including the kidneys, bones, cardiovascular system and reproductive system. However, the effect of paternal Cd exposure on the offspring is unclear. In this study, 1 mg/kg of cadmium chloride (CdCl2) was injected intraperitoneally every other day in 8-week-old C57BL/6 J male mice to study the effects on their female offspring. Our results showed an increase in body weight, water intake and food intake in F1 female mice from the Cd-exposed group. The development of secondary follicles and antral follicles in the ovaries of Cd-treated was inhibited. Serum estradiol (E2) was found to be decreased. Further analysis revealed significant downregulation of StAR, P450scc, 17β-HSD, CYP17A1 and CYP19A1, which are related to E2 synthesis. Serum total cholesterol was increased and free cholesterol was reduced. Total cholesterol in ovarian tissue was decreased. qRT-PCR and Western blot analysis revealed a decrease in the mRNA and protein expression of HMGCR, LDLR, and ABCA1, which are associated with cholesterol homeostasis. Oil red O staining indicated that lipid droplets (LDs) were accumulated in ovarian tissues, while the expression of ATGL and HSL proteins associated with lipid droplet degradation was significantly downregulated. In juvenile female mice, ultrastructural alterations of mitochondria in the ovaries were observed by transmission electron microscopy (TEM). In adult female mice, the expression of proteins associated with mitochondrial dynamics (DRP1 and MFN2) was significantly reduced in the ovaries. Overall, our study suggests that paternal Cd exposure inhibits follicular development, and affects serum E2 synthesis by impairing cholesterol homeostasis and affecting mitochondrial function.

Amomum villosum Lour. Fruit extract mitigates hyperlipidemia through SREBP-2/LDLR/HMGCR signaling in high-cholesterol diet-fed mice

Amomum villosum Lour. (Zingiberaceae) is an herbal medicine used in Asian countries for various ailments. In the current experiment, the effect of AV water extract (AVE) on cholesterol-lowering capabilities along with its essential bio-molecular mechanisms have been examined. The efficiency of AVE against high-cholesterol diet (HCD)-generated hyperlipidemia was investigated using C57BL/6 mice. Mice have been divided to six categories: control group (normal diet), high-cholesterol diet (HCD), and HCD treated with AVE at 100, 200, and 500 mg/kg, or simvastatin 40 mg/kg for four weeks. AVE treated animal groups (100, 200, 500 mg/kg) had markedly lessened body mass increase (p < 0.01) and also had reduced liver and epididymal fat weights (p < 0.05) related to HCD alone batch. Hepatic TG levels in AVE 100 mg/kg, 200 mg/kg, and 500 mg/kg treated groups were meaningfully reduced by 13.32%, 14.61%, and 21.78%, respectively; and serum TG decreased by 25.65%, 31.97%, and 32.22%, respectively, compared to the HCD alone group. Serum TC was reduced by 11.17%, 12.48%, and 26.85% with AVE at 100, 200, and 500 mg/kg, respectively; and serum LDL-C declined by 10.08%, 18.50%, and 30.90% (p < 0.05) with AVE at 100, 200, and 500 mg/kg, respectively. Levels of mRNA expression including LDL receptor (LDLR), sterol regulatory element-binding protein 2 (SREBP2), and HMG-CoA reductase (HMGCR) were noticeably amplified by AVE (500 mg/kg) treatment. Similarly, AVE treatment significantly increased Hmgcr protein expression levels in a dose related fashion. These outcomes show that AVE displays a hypolipidemic effect and might function as a novel hypolipidemic therapy, further; they suggest a mechanism of action for this effect.

Amyloid beta increases ABCA1 and HMGCR protein expression, and cholesterol synthesis and accumulation in mice neurons and astrocytes

IntroductionImbalanced cholesterol metabolism in the brain is one of the main pathophysiological mechanisms involved in Alzheimer's disease. We investigated the effect of amyloid-beta (Aβ) on the main proteins involved in regulation of cholesterol metabolism along with cholesterol content in astrocytes and neurons. MethodsAstrocytes and neurons were cultured and treated with Aβ. Apolipoprotein E (apoE) level in the cells and conditioned media, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGCR), ATP-binding cassette transporter A1 (ABCA1), and cytochrome P450 46A1 (CYP46A1) in cell lysates were determined using immunoblotting. Astrocyte media was added to the Aβ-pretreated neurons then, HMGCR was assessed. Cholesterol was measured in both cells and media. ResultsAβ caused a significant increase in HMGCR and ABCA1 protein levels and cholesterol content in both cells without increasing cholesterol efflux. A similar increase was seen for cellular apoE level in astrocytes with no changes in media with a significant reduction of cholesterol efflux. HMGCR level was restored to near control level when Aβ-pretreated neurons were exposed to media from culture astrocytes. ConclusionAlmost all events related to cholesterol homeostasis in neurons and astrocytes, are somehow affected by Aβ. However, because ABCA1 has the most important role(s) in brain cholesterol homeostasis, all subsequent events associated with astrocytes-cholesterol synthesis and its shuttling to neurons are influenced by the effects of Aβ on ABCA1 which could likely be responsible for altered brain cholesterol metabolism in Alzheimer's disease.

Effects of DHA dietary intervention on hepatic lipid metabolism in apolipoprotein E-deficient and C57BL/6J wild-type mice

Lipid metabolic disorder occurs when ApoE gene is deficient. However, the role of Docosahexaenoic acid (DHA) in relieving hepatic lipid metabolic disorder in apolipoprotein E-deficient (ApoE -/-) mice remains unknown. We fed 3-month-old C57BL/6J wild-type (C57 wt) and ApoE -/- mice respectively with normal or DHA fortified diet for 5 months. We found ApoE gene deficiency caused hepatic lipid deposition and increased lipid levels in plasma and liver. Hepatic gene expression of SRB1, CD36 and FABP5 in ApoE -/- mice, protein expression of HMGCR, LRP1 in C57 wt mice and protein expression of LRP1 in ApoE -/- mice increased after DHA intervention. In DHA-fed ApoE -/- mice, LXRα/β and PPARα protein expression down-regulated in cytoplasm, but LXRα/β protein expression up-regulated in nucleus. DHA treatment decreased RXRα and RXRβ expression in C57 wt and ApoE -/- female mice. Deletion of ApoE gene caused lipid metabolism disorder in liver of mice. DHA treatment efficiently meliorated lipid metabolism caused by ApoE deficient through the regulation of gene and protein expressions of molecules involved in liver fatty acids transport and lipid metabolism.

Protective effect of atorvastatin meditated by HMGCR gene on diabetic rats with atherosclerosis: An in vivo and in vitro study

BackgroundAccelerated atherosclerosis in patients suffering from diabetes represents a major cause of morbidity and mortality. The aim of present study was to investigate the protective effects conferred by atorvastatin (AVT) meditated by the HMGCR gene in diabetic rats with atherosclerosis. MethodsSerum triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), very-low-density lipoprotein cholesterol (VLDL-C), fasting blood glucose (FBG) and serum insulin (INS) were all determined by means of in vivo experiments. Following the establishment of the diabetic model of atherosclerosis, the expressions of HMGCR, low density lipoprotein receptor (LDLR), fatty acid synthase (FASN) were detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis in the vitro experiments. Flow cytometry was adopted in order to detect cell cycle and apoptosis. ResultsThe in vivo experiments results indicated that FBG and INS among the diabetic arteriosclerosis rats exhibited markedly higher levels; after injected with AVT and HMGCR, decreased contents of TC, TG, LDL-C and VLDL-C, while increased contents of HDL-C as well as an increased positive rate of HMGCR protein expression were observed. In vitro experiment, the mRNA and protein expression of LDLR were increased and FASN were decreased in cells transfected with HMGCR and AVT; with a greater number of cells arrested at the S phase and less in the G0/G1 phase, as well as data indicating the rate of apoptosis was inhibited after HMGCR and AVT transfection processes. ConclusionThe key findings of the present study suggested that the protective effect conferred by AVT in diabetic rats with atherosclerosis was associated with the overexpression of the HMGCR gene, thus presenting a novel target for atherosclerosis treatment.

Eicosapentaenoic acid inhibits formation of cholesterol gallstone by suppressing gene HMGCR and ABCG5/8 in mice

Objective To observe the effect of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on cholesterol gallstones formation in C57BL/6 mice with diet-induced cholesterol gallstone, and then explore the potential mechanism. Methods Fifty C57BL/6 mice were randomly divided into 5 groups (10 mice in each group), referring to control group, experimental group, experimental plus DHA group, experimental plus EPA group, as well as experimental plus DHA and EPA group. The mice in control group were fed with regular diet, and the rest of the mice with lithogenic diet (LD). Subsequent to feeding the mice with separate diets for two weeks, EPA and/or DHA (70 mg·kg-1·d-1) were orally administered for eight weeks, while the LD feeding was continued during this period. After a total of 10 weeks, the mice were dissected to observe the gallstone formation. The levels of serum lipids, total cholesterol (TC) and phospholipids (PL) in bile, and TC in the liver were tested, and the protein expression of HMGCR, SRBI, ABCG5/ABCG8, CYP7A1 and ABCB11genes in the liver of mice was measured. Results Compared with the experimental group, the experimental plus EPA group had significantly lower TC in liver (0.033±0.008 mmolo/g) and bile (1.807±0.381 mmolo/L), and lower relative protein expression levels of HMGCR (0.545±0.098), ABCG5 (0.418±0.089) and ABCG8 (0.501±0.151)in liver (P 0.05). Conclusions EPA could prevent the cholesterol gallstone formation in mice by decreasing the expression of HMGCR and ABCG5/8 genes in liver, therefore reducing cholesterol synthesis and blocking cholesterol transport from liver to bile as well as diminishing cholesterol content in the bile. However, the inhibition effect of DHA on cholesterol gallstone formation was not obvious. Key words: Eicosapentaenoic acid; Mice; Cholesterol gallstone; Gene

Exogenous administration of chronic corticosterone affects hepatic cholesterol metabolism in broiler chickens showing long or short tonic immobility

Tonic immobility (TI) is an innate characteristic of animals related to fear or stress response. Animals can be classified into long TI (LTI) and short TI (STI) phenotypes based on TI test duration. In this study, effect of TI phenotype, chronic corticosterone administration (CORT), and their interaction on cholesterol metabolism in liver was evaluated in broilers. LTI broilers showed higher level of cholesterol in liver compared to STI chickens (p<0.05), and CORT significantly increased hepatic cholesterol content (p<0.01). Real-time PCR results showed that both TI and CORT potentially altered ABCA1 and CYP7A1 gene expressions (0.05<p<0.1), while there was no significant interaction of CORT and TI on both gene expressions. CORT treatment significantly increased the level of SREBP2 (p=0.00), LDLR (p<0.05), GR (p<0.05) and 11β-HSD2 (p<0.05) protein abundance in liver. However, TI phenotype only affected hepatic HMGCR protein expression, and LTI broilers showed higher level of HMGCR protein expression in liver than STI (p<0.05). These results indicate that chronic CORT administration causes hepatic cholesterol accumulation in broiler chickens mainly by enhancing cholesterol synthesis and uptake into liver. LTI chickens had higher amount of total cholesterol in liver, which might be associated with an increase of hepatic HMGCR protein expression. However, there is no interaction between TI and CORT on cholesterol metabolism in liver of broilers.

고콜레스테롤혈증 ApoE Knockout Mice에서 기능성 수정과의 간지질 축적 억제 효과

In this study, the hepatic lipid-lowering effects and related mechanism of action of sujeonggwa were examined in hypercholesterolemia-induced apoprotein E knockout (apo E ko) mice. Sujeonggwa drink was prepared with cinnamon, ginger, and sugar by modifying the traditional recipe of sujeonggwa. Sugar was partially substituted with either stevia or short chain fructooligosaccharide (scFOS) in order to reduce the calorie content of sujeonggwa, which was measured by descriptive analysis. Apo E ko mice (n=42) were induced to have hypercholesterolemia (plasma total cholesterol concentration >1,000 mg/dL) by administration of a high cholesterol diet for 4 weeks, followed by division into six groups. Experimental groups were orally administered water as a vehicle (normal group), sugar solution (control group), commercially available 'V' sujeonggwa drink (positive control group), or three different types of sujeonggwa drinks (S-sugar, S-stevia, and S-scFOS group) for 6 weeks while high cholesterol diet was provided to all animals. Compared to the control group, concentrations of hepatic triglycerides, total cholesterol, thiobarbituric acid reactive substances, and reactive oxygen species in S-sugar, S-stevia, S-scFOS were significantly reduced (P<0.05), indicating that sujeonggwa had inhibitory effects on hepatic lipid accumulation. Protein expression levels of fatty acid synthase (FAS) and its transcription factor, sterol regulatory element-binding protein (SREBP)-1 responsible for trigly- ceride synthesis, as well as 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCR) and its transcription factor, SREBP-2 responsible for cholesterol synthesis, were also reduced in S-sugar, S-stevia, and S-scFOS groups (P<0.05). These benefits of sujeonggwa were even greater in S-stevia and S-scFOS compared to S-sugar. The beneficial effects of S-stevia on regulation of hepatic lipid metabolism were slightly greater than those of S-scFOS although the differences were not significant. In conclusion, sujeonggwa drinks, especially functional sujeonggwa drinks in which sugar was partially substituted with stevia or scFOS, inhibited hepatic lipid accumulation via suppressing FAS and HMGCR protein expression through down-regulation of SREBP-1 and 2.

Cholesterol deregulation induced by chronic corticosterone (CORT) stress in pectoralis major of broiler chickens

Chronic endogenous glucocorticoid (GC) excess in mammals is associated with metabolic dysfunction and dyslipidemia that are characterized by increased plasma triglyceride and total cholesterol (Tch) levels. However, the effects of chronic GC administration on cholesterol metabolism, particularly in muscle tissues of broiler chickens, are unknown. In this study, broiler chickens were treated chronically with vehicle (CON) or corticosterone (CORT) for 2weeks. Chronic CORT treatment significantly increased Tch levels in pectoralis major muscle (PMC) (p<0.001) as well as in leg muscle (p<0.01), and CORT enhanced triglyceride levels in the PMC (p<0.001). Real-time PCR results showed that HMGCR (p<0.05) mRNA expression was up-regulated by CORT in PMC, and 11β-HSD1 gene transcription (p=0.08) was not significantly downregulated, whereas glucocorticoid receptor (GR) mRNA expression, 11β-HSD2, CYP7A1, CYP27A1, ApoB and LDLR were unchanged by CORT (p>0.05). Western blot results showed that the levels of total GR (p=0.08) tended to be increased and nuclear GR protein (p<0.05) was increased in PMC by CORT administration. Parallel to an increase in gene expression, HMGCR protein expression in PMC was significantly increased (p<0.05) by CORT. Moreover, LDLR (p<0.05), ApoA1 (p=0.06) and 11β-HSD2 (p=0.07) protein expression in PMC tended to be increased by CORT compared to control. These results indicate that chronic CORT administration causes cholesterol accumulation in PMC tissues of broiler chickens by increasing cholesterol synthesis and uptake.