Fecal Cholesterol Excretion Research Articles

Quantitative analysis of sterol balance in a mouse model of hepatic lipid accumulation induced by cholesterol and cholic acid supplementation.

The cholesterol balance and bile acid metabolism in a mouse model of hepatic lipid accumulation induced by a diet supplemented with cholesterol and cholic acid (CA) were quantitatively evaluated. The mice were fed diets supplemented with different levels of cholesterol (0, 3, or 6 g/kg diet) and CA (0.5 g/kg diet) for six weeks. Cholesterol supplementation doubled the hepatic triglyceride concentration, regardless of the supplementation level, without inflammation or gallstone formation. Both cholesterol supplementations enhanced fecal excretion of muricholic acid. Additionally, the higher cholesterol supplementation led to an increase in fecal cholesterol excretion, accompanied by elevated expression of hepatic cholesterol exporters and a reduction in fecal bile acid excretion. In this mouse study, supplementation with 3 g cholesterol/kg diet and 0.5 g CA/kg diet was sufficient to induce hepatic lipid accumulation.

Evaluation of the Decreased Cholesterol Potential of Levilactobacillus brevis M-10 Isolated from Spontaneously Fermented Sour Porridge in Mice with High-Cholesterol Levels.

Excessive cholesterol levels can lead to hypercholesterolemia, which is related to cardiovascular diseases (CVDs), and CVDs are a serious threat to human health. Therefore, lowering cholesterol levels is necessary, and diet intervention is safer than drugs are. The cholesterol-lowering effect of Levilactobacillus brevis M-10 isolated from spontaneously fermented millet sour porridge was investigated in fifty C57BL/6N male mice. After a 4-week intervention, the food intake, weight gains and organ indices were calculated; the lipid contents in the serum, liver, and feces were determined; the histopathology of the liver tissues was observed; the expression of metabolism-related genes was determined; and short-chain fatty acid (SCFA) levels in the droppings were monitored. The results showed that administration of a high dose of L. brevis M-10 (1 × 1010CFU/mL) significantly reduced food intake, suppressed weight gain; prevented excessive liver growth; and reduced the total serum cholesterol, triglycerides, low-density lipoproteins; and total hepatic cholesterol and triglyceride contents (P < 0.05) in high-cholesterol mice. Moreover, a high dose of L. brevis M-10 significantly promoted the fecal excretion of cholesterol and triglycerides (P < 0.05) and alleviated liver damage induced by a high-cholesterol diet. Furthermore, a high dose of L. brevis M-10 significantly downregulated the cholesterol metabolism-related gene expression of NPC1L1, ACAT2, HMG-CoA, and SREBP2 but upregulated the gene expression of ABCG5, CYP7A1, and LXR-α (P < 0.05). Additionally, a high dose of L. brevis M-10 significantly increased SCFA contents, including those of acetic acid, propionic acid and n-butyric acid (P < 0.05). These findings could provide support for the use of L. brevis M-10 in the application of functional foods to alleviate hypercholesterolemia.

Influence of Varied Dietary Cholesterol Levels on Lipid Metabolism in Hamsters.

Syrian hamsters are valuable models for studying lipid metabolism due to their sensitivity to dietary cholesterol, yet the precise impact of varying cholesterol levels has not been comprehensively assessed. This study examined the impact of varying dietary cholesterol levels on lipid metabolism in Syrian hamsters. Diets ranging from 0% to 1% cholesterol were administered to assess lipid profiles and oxidative stress markers. Key findings indicate specific cholesterol thresholds for inducing distinct lipid profiles: below 0.13% for normal lipids, 0.97% for elevated LDL-C, 0.43% for increased VLDL-C, and above 0.85% for heightened hepatic lipid accumulation. A cholesterol supplementation of 0.43% induced hypercholesterolemia without adverse liver effects or abnormal lipoprotein expression. Furthermore, cholesterol supplementation significantly increased liver weight, plasma total cholesterol, LDL-C, and VLDL-C levels while reducing the HDL-C/LDL-C ratio. Fecal cholesterol excretion increased, with stable bile acid levels. High cholesterol diets correlated with elevated plasma ALT activities, reduced hepatic lipid peroxidation, and altered leptin and CETP levels. These findings underscore Syrian hamsters as robust models for hyperlipidemia research, offering insights into experimental methodologies. The identified cholesterol thresholds facilitate precise lipid profile manipulation, enhancing the hamster's utility in lipid metabolism studies and potentially informing clinical approaches to managing lipid disorders.

Apolipoprotein O modulates cholesterol metabolism via NRF2/CYB5R3 independent of LDL receptor

Apolipoprotein O (APOO) plays a critical intracellular role in regulating lipid metabolism. Here, we investigated the roles of APOO in metabolism and atherogenesis in mice. Hepatic APOO expression was increased in response to hyperlipidemia but was inhibited after simvastatin treatment. Using a novel APOO global knockout (Apoo−/−) model, it was found that APOO depletion aggravated diet-induced obesity and elevated plasma cholesterol levels. Upon crossing with low-density lipoprotein receptor (LDLR) and apolipoprotein E (APOE) knockout hyperlipidemic mouse models, Apoo−/−Apoe−/− and Apoo−/−Ldlr−/− mice exhibited elevated plasma cholesterol levels, with more severe atherosclerotic lesions than littermate controls. This indicated the effects of APOO on cholesterol metabolism independent of LDLR and APOE. Moreover, APOO deficiency reduced cholesterol excretion through bile and feces while decreasing phospholipid unsaturation by inhibiting NRF2 and CYB5R3. Restoration of CYB5R3 expression in vivo by adeno-associated virus (AAV) injection reversed the reduced degree of phospholipid unsaturation while decreasing blood cholesterol levels. This represents the first in vivo experimental validation of the role of APOO in plasma cholesterol metabolism independent of LDLR and elucidates a previously unrecognized cholesterol metabolism pathway involving NRF2/CYB5R3. APOO may be a metabolic regulator of total-body cholesterol homeostasis and a target for atherosclerosis management.Apolipoprotein O (APOO) regulates plasma cholesterol levels and atherosclerosis through a pathway involving CYB5R3 that regulates biliary and fecal cholesterol excretion, independently of the LDL receptor. In addition, down-regulation of APOO may lead to impaired mitochondrial function, which in turn aggravates diet-induced obesity and fat accumulation.

Novel Synthesis of Phytosterol Ferulate Using Acidic Ionic Liquids as a Catalyst and Its Hypolipidemic Activity.

Phytosterol ferulate (PF) is quantitively low in rice, corn, wheat, oats, barley, and millet, but it is potentially effective in reducing plasma lipids. In this study, PF was synthesized for the first time using acidic ionic liquids as a catalyst. The product was purified, characterized using Fourier transform infrared, mass spectroscopy, and nuclear magnetic resonance, and ultimately confirmed as the desired PF compound. The conversion of phytosterol surpassed an impressive 99% within just 2 h, with a selectivity for PF exceeding 83%. Plasma lipid-lowering activity of PF was further investigated by using C57BL/6J mice fed a high-fat diet as a model. Supplementation of 0.5% PF into diet resulted in significant reductions in plasma total cholesterol, triacylglycerols, and nonhigh-density lipoprotein cholesterol by 13.7, 16.9, and 46.3%, respectively. This was accompanied by 55.8 and 36.3% reductions in hepatic cholesterol and total lipids, respectively, and a 22.9% increase in fecal cholesterol excretion. Interestingly, PF demonstrated a higher lipid-lowering activity than that of its substrates, a physical mixture of phytosterols and ferulic acid. In conclusion, an efficient synthesis of PF was achieved for the first time, and PF had the great potential to be developed as a lipid-lowering dietary supplement.

Plasma cholesterol level reduction in albino rats by β-d glucan (pleuran) from Plurotus ostreatus

Abstract Background: The progression of atherosclerotic lesions and coronary artery disease is known to be accelerated by hypercholesterolemia. Objective: This study investigated at the effects of adding an extract of oyster mushroom fruiting bodies to rats with high and normal cholesterol levels on their biochemical and histological changes. This study was aimed to assess the effect of Pleurotus ostreatus extraction on hypercholesterolemia in albino rats and show the revelation of the relationship of these with some biochemical parameters. Materials and Methods: Three sets of ten albino rats each were formed from 6-week-old rats. To lower plasma total cholesterol, triglycerides, low-density lipoprotein (LDL), total lipid, phospholipids, and LDL/high-density lipoprotein, extracts of P. ostreatus fruiting bodies were fed to hypercholesterolemic rats. Results: In rats with high cholesterol, feeding oyster mushrooms drastically lowered body weight. The excretion of total lipids and cholesterol in feces was enhanced by feeding mushrooms. Conclusion: These findings imply that P. ostreatus food supplement improved health by altering hypercholesterolemic rats’ atherogenic lipid profiles.

Laminarin Reduces Cholesterol Uptake and NPC1L1 Protein Expression in High-Fat Diet (HFD)-Fed Mice.

Aberrantly high dietary cholesterol intake and intestinal cholesterol uptake lead to dyslipidemia, one of the risk factors for cardiovascular diseases (CVDs). Based on previous studies, laminarin, a polysaccharide found in brown algae, has hypolipidemic activity, but its underlying mechanism has not been elucidated. In this study, we investigated the effect of laminarin on intestinal cholesterol uptake in vitro, as well as the lipid and morphological parameters in an in vivo model of high-fat diet (HFD)-fed mice, and addressed the question of whether Niemann-Pick C1-like 1 protein (NPC1L1), a key transporter mediating dietary cholesterol uptake, is involved in the mechanistic action of laminarin. In in vitro studies, BODIPY-cholesterol-labeled Caco-2 cells were examined using confocal microscopy and a fluorescence reader. The results demonstrated that laminarin inhibited cholesterol uptake into Caco-2 cells in a concentration-dependent manner (EC50 = 20.69 μM). In HFD-fed C57BL/6J mice, laminarin significantly reduced the serum levels of total cholesterol (TC), total triglycerides (TG), and low-density lipoprotein cholesterol (LDL-C). It also decreased hepatic levels of TC, TG, and total bile acids (TBA) while promoting the excretion of fecal cholesterol. Furthermore, laminarin significantly reduced local villous damage in the jejunum of HFD mice. Mechanistic studies revealed that laminarin significantly downregulated NPC1L1 protein expression in the jejunum of HFD-fed mice. The siRNA-mediated knockdown of NPC1L1 attenuated the laminarin-mediated inhibition of cholesterol uptake in Caco-2 cells. This study suggests that laminarin significantly improves dyslipidemia in HFD-fed mice, likely by reducing cholesterol uptake through a mechanism that involves the downregulation of NPC1L1 expression.

Genetic Deletion of Hepatic NCOR1 regulates the bile acid metabolism and leads to reduction in atherosclerosis

Abstract Background The nuclear receptor corepressor 1 (NCOR1) plays an important role in the regulation of gene expression in immunometabolic conditions by connecting chromatin-modifying enzymes, coregulators and transcription factors. NCOR1 has been shown to be involved in cardiometabolic diseases. Since NCOR1 transrepresses the function of several nuclear receptors involved in hepatic lipid transport and synthesis, we hypothesized that genetic NCOR1 deletion in the liver alters lipid metabolism and atherosclerosis development. Methods To test this hypothesis, we generated hepatocyte-specific Ncor1 knockout mice on a Ldlr-/- background. Besides assessing the progression of the disease in thoraco-abdominal aortae en face, we analyzed hepatic cholesterol and bile acid metabolism at expression and functional level. Results This study findings demonstrated that specific Ncor1 knockout mice on an atherosclerosis-prone background develop less atherosclerotic lesions than controls. Interestingly, under chow diet plasma cholesterol levels of liver-specific Ncor1 knockout mice were slightly higher compared to control, but strongly reduced compared to control mice after feeding them an atherogenic diet for 12 weeks. Moreover, hepatic cholesterol and triglyceride contents were decreased in liver-specific Ncor1 knockout compared to control mice. Our mechanistic data revealed that NCOR1 reprograms the synthesis of bile acids towards the alternative pathway, which in turn reduce bile hydrophobicity and enhances fecal cholesterol excretion. Conclusions Our data suggest that hepatic Ncor1 deletion in mice decreases atherosclerosis development by reprograming bile acid metabolism and enhancing fecal cholesterol excretion. Hepatocyte NCOR1 may therefore provide a novel target for development of anti-atherosclerotic drugs.

Regulation of Cholesterol Transporters by Nuclear Receptors

Atherosclerosis is a pathological condition characterized by the accumulation of plaques in the arteries, leading to cardiovascular diseases. The deposition of cholesterol in peripheral cells increases the risk of atherosclerosis. Reverse cholesterol transport (RCT) is essential to reduce the risk of atherosclerosis because it removes excessive cholesterol from the peripheral tissues. ATP-binding cassette transporters such as ABCA1, ABCG1, ABCG5, and ABCG8 are involved in the efflux of cholesterol. The upregulation of these ABC transporters enhances RCT, thereby promoting the removal of excess cholesterol from the body. The expression and activity of ABC transporters are regulated by transcriptional and post-transcriptional mechanisms, as well as by post-translational modifications. In this review, the regulation of ABC transporters by nuclear receptors such as farnesoid X receptor, liver X receptor, retinoid X receptor, retinoic acid receptor, and peroxisome proliferator-activated receptors is discussed. Pharmacological and natural compounds serving as agonists for the nuclear receptors have been identified to elevate the mRNA levels of the transporters. Consequently, it is anticipated that these compounds will attenuate the development of atherosclerosis through stimulation of the ABC transporters, thereby enhancing RCT and fecal cholesterol excretion. Understanding these regulatory processes can aid in the development of therapeutic approaches to prevent atherosclerosis.

Mei-Gin Formula Ameliorates Obesity through Lipolysis, Fatty Oxidation, and Thermogenesis in High-Fat Diet-Induced Obese Rats.

Obesity is a metabolic dysfunction characterized by excessive body fat deposition as a consequence of an energy imbalance. Novel therapeutic strategies have emerged that are safe and have comparatively low side effects for obesity treatment. Functional foods and nutraceuticals have recently received a great deal of attention because of their components with the properties of antimetabolic syndrome. Based on our previous in vitro and in vivo investigations on anti-adipogenesis activity and improved body fat accumulation in serials, the combination of three ingredients (including bainiku-ekisu, black garlic, and Mesona procumbens Hemsl), comprising the Mei-Gin formula (MGF), was eventually selected as a novel inhibitor that exhibited preventive effects against obesity. Herein, we verify the anti-obesity effects of MGF in obese rats induced by a high-fat diet and discuss the potential molecular mechanisms underlying obesity development. Oral administration of MGF significantly suppressed the final body weight, weight change, energy and water intake, subcutaneous and visceral fat mass, liver weight, hepatic total lipids and triglycerides (TG), and serum levels of TG, triglycerides (TC), low-density lipoprotein cholesterol (LDL-C), alanine transaminase (AST), uric acid, and ketone bodies and augmented fecal total lipids, TG, and cholesterol excretion in the high-dose MGF-supplemented groups. Furthermore, the corresponding lipid metabolic pathways revealed that MGF supplementation effectively increased lipolysis and fatty acid oxidation gene expression and attenuated fatty acid synthesis gene expression in the white adipose tissue (WAT) and liver and it also increased mitochondrial activation and thermogenic gene expression in the brown adipose tissue (BAT) of rats with obesity induced by a high-fat diet (HFD). These results demonstrate that the intake of MGF can be beneficial for the suppression of HFD-induced obesity in rats through the lipolysis, fatty oxidation, and thermogenesis pathway. In conclusion, these results demonstrate the anti-obesity efficacy of MGF in vivo and suggest that MGF may act as a potential therapeutic agent against obesity.

Genetic deletion of hepatic NCOR1 protects from atherosclerosis by promoting alternative bile acid-metabolism and sterol excretion

BackgroundThe nuclear receptor corepressor 1 (NCOR1) plays an important role in the regulation of gene expression in immunometabolic conditions by connecting chromatin-modifying enzymes, coregulators and transcription factors. NCOR1 has been shown to be involved in cardiometabolic diseases. Recently, we demonstrated that the deletion of macrophage NCOR1 aggravates atherosclerosis by promoting CD36-triggered foam cell formation via PPARG derepression.PurposeSince NCOR1 modulates the function of several key regulators involved in hepatic lipid and bile acid metabolism, we hypothesized that its deletion in hepatocytes alters lipid metabolism and atherogenesis.MethodsTo test this hypothesis, we generated hepatocyte-specific Ncor1 knockout mice on a Ldlr-/- background. Besides assessing the progression of the disease in thoracoabdominal aortae en face, we analyzed hepatic cholesterol and bile acid metabolism at expression and functional levels.ResultsOur data demonstrate that liver-specific Ncor1 knockout mice on an atherosclerosis-prone background develop less atherosclerotic lesions than controls. Interestingly, under chow diet, plasma cholesterol levels of liver-specific Ncor1 knockout mice were slightly higher compared to control, but strongly reduced compared to control mice after feeding them an atherogenic diet for 12 weeks. Moreover, the hepatic cholesterol content was decreased in liver-specific Ncor1 knockout compared to control mice. Our mechanistic data revealed that NCOR1 reprograms the synthesis of bile acids towards the alternative pathway, which in turn reduce bile hydrophobicity and enhances fecal cholesterol excretion.ConclusionsOur data suggest that hepatic Ncor1 deletion in mice decreases atherosclerosis development by reprograming bile acid metabolism and enhancing fecal cholesterol excretion.

Plasma Total and Lipoprotein Cholesterol, Liver Cholesterol and Fecal Cholesterol Excretion in Hamsters Fed Fiber Diets

The effect of dietary fibers on plasma lipids, liver cholesterol and fecal cholesterol excretion was investigated in hamsters. Male 9- to 11-wk-old Golden-Syrian hamsters (n = 155) were fed a purified hypercholesterolemic diet (0.1% cholesterol, 10% fat) for 5 wk to elevate plasma lipid concentrations. Sixteen animals with elevated plasma total cholesterol were randomly assigned to each treatment group: control, oat bran, guar gum, cellulose, xylan and terminal groups. After 4 wk of fiber diet consumption, the plasma total cholesterol concentrations were significantly decreased in the oat bran, guar gum and xylan groups (16, 12 and 15%, respectively) (P < 0.05). Plasma HDL cholesterol concentration was significantly decreased only in the guar gum group (12%) (P < 0.05). The combined plasma VLDL+LDL cholesterol concentrations were significantly lowered by the oat bran, xylan and cellulose diets (38, 34 and 40%, respectively) (P < 0.05). After 4 wk of control diet consumption, the liver cholesterol concentration quadrupled to 10.6 mumol cholesterol/g liver (P < 0.05). After 4 wk of consumption of the treatment diets, liver cholesterol was further increased (P < 0.05) only in the cellulose group, to 14.5 mumol cholesterol/g liver. The total fecal cholesterol excretion was the highest (P < 0.05) in the oat bran group. In the present study, oat bran, guar gum and xylan were effective hypocholesterolemic agents in hamsters.

Dietary Grape Seed Tannins Affect Lipoproteins, Lipoprotein Lipases and Tissue Lipids in Rats Fed Hypercholesterolemic Diets , ,

The effects of monomeric and polymeric grape seed tannins on rat plasma lipoproteins, lipoprotein lipase, hepatic lipase and aortic and hepatic lipid concentration were studied. Male Sprague-Dawley rats received either a normal diet (with no added cholesterol and no tannins), a control diet (hypercholesterolemic diet) or hypercholesterolemic diets supplemented with 2% tannin monomers or 2% polymers 3 or 9 wk. Plasma total cholesterol, triacylglycerol, LDL cholesterol and VLDL concentrations were significantly higher and the HDL cholesterol concentration lower in controls and in rats fed the diet supplemented with monomers compared with rats fed polymeric tannins at both time points. Lipoprotein lipase and hepatic lipase activities were significantly higher in control and in monomer-fed groups than in the polymer-fed group. Hepatic and aortic cholesterol and triacylglycerol concentrations were significantly higher in control rats and those fed monomers than in the polymer-fed group. Moreover, plasma HDL cholesterol and hepatic lipase activity were closely associated with low aortic cholesterol and triacylglycerol in rats fed polymeric tannins. These rats also exhibited greater fecal excretion of cholesterol and especially bile acids than the control or monomer-fed groups. Thus dietary grape seed tannins have a pronounced anti-hypercholesterolemic effect by enhancing reverse cholesterol transport and also by reducing intestinal cholesterol absorption and increasing bile acid excretion.

Supplementation of Weizmannia coagulans BC2000 and Ellagic Acid Inhibits High-Fat-Induced Hypercholesterolemia by Promoting Liver Primary Bile Acid Biosynthesis and Intestinal Cholesterol Excretion in Mice

The probiotic Weizmannia coagulans (W. coagulans) BC2000 can increase the abundance of intestinal transforming ellagic acid (EA) bacteria and inhibit metabolic disorders caused by hyperlipidemia by activating liver autophagy. This study aimed to investigate the inhibitory effects of W. coagulans BC2000 and EA on hyperlipidemia-induced cholesterol metabolism disorders. C57BL/6J mice (n = 10 in each group) were fed a low-fat diet, high-fat diet (HFD), HFD supplemented with EA, HFD supplemented with EA and W. coagulans BC77, HFD supplemented with EA, and W. coagulans BC2000. EA and W. coagulans BC2000 supplementation prevented HFD-induced hypercholesterolemia and promoted fecal cholesterol excretion. Transcriptome analysis showed that primary bile acid biosynthesis in the liver was significantly activated by EA and W. coagulans BC2000 treatments. EA and W. coagulans BC2000 treatment also significantly increased the intestinal Eggerthellaceae abundance and the liver EA metabolites, iso-urolithin A, Urolithin A, and Urolithin B. Therefore, W. coagulans BC2000 supplementation promoted the intestinal transformation of EA, which led to the upregulation of liver bile synthesis, thus preventing hypercholesterolemia.

Ergosterol fraction from Agaricus bisporus modulates adipogenesis and skeletal glucose uptake in high fat diet induced obese C57BL/6 mice

AimThe study aimed to optimize a method of extracting ergosterol rich concentrate (ECF) and to evaluate its significant impact on adipogenesis and associated complications in high-fat diet (HFD) induced obese mice. MethodsA comparative analysis (soxhlet and ultra sound assisted extraction) was done to obtain the highest yield of ergosterol from Agaricus bisporus. The ECF was evaluated for the biological effect on 3T3-L1 pre-adipocytes in-vitro and in male C57BL/6 mice model in-vivo. Key findingsUltra sound assisted extraction method using the solvent n-hexane resulted in highest ergosterol yield. ECF treatment significantly reduced the differentiation and lipid accumulation in pre-adipocyte cells without any cytotoxicity. In-vivo study illustrated beneficial impact on cholesterol metabolism by down regulating the hepatic gene expression of LXR-α, HMG-CoR and up-regulating LDL-R expression. Significant increase in fecal excretion of cholesterol and bile acids have also been observed among the ECF treated animals compared to high fat diet (HFD) fed mice. ECF had an anti-adipogenic activity in-vivo mainly by inhibiting the activity of PPAR-γ, C/EBP-α and SREBP-1c. The results also depicted the improvement of obesity associated insulin resistance by ECF treatment manly via decrease in plasma resistin and up-regulation in skeletal GLUT4 protein expression. SignificanceOur study illustrated diverse activity of ECF in the therapeutic management of obesity associated metabolic complications mainly by reducing adipogenesis and improving glucose uptake in skeletal muscle in conjunction with improved cholesterol metabolism.

Hypotriglyceridemic and hepatoprotective effect of pumpkin (Cucurbita moschata) seed flour in an experimental model of dyslipidemia

This study evaluated the effect of pumpkin ( Cucurbita moschata ) seed flour on the lipid profile and biomarkers of liver function in an experimental dyslipidemia. The hamsters were divided into three groups: normal control group (n = 8) fed commercial feed; dyslipidemia control group (n = 8) with dyslipidemia induced by consumption of feed enriched with saturated fat and cholesterol (dyslipidemic diet with 13.5% coconut fat and 0.1% cholesterol); pumpkin seed group (n = 10), maintained on a dyslipidemic diet plus pumpkin seed flour. After 28 days of treatment, the treatment with a diet enriched with pumpkin seeds significantly reduced the serum triglycerides, increased the faecal excretion of cholesterol, and reduced the accumulation of lipids in the hepatocytes, thus decreasing the degree of hepatic steatosis. In the experimental model used in this study, pumpkin seed flour had a hypotriglyceridemic property and a hepatoprotective effect in relation to hepatic steatosis. • Feeding enriched with pumpkin seeds reduced the serum triglycerides level. • Creole pumpkin seed flour increased the excretion of cholesterol in the faeces. • Pumpkin seeds reduced the degree of hepatic steatosis.

NUDT7 regulates total hepatic CoA levels and the composition of the intestinal bile acid pool in male mice fed a Western diet

Nudix hydrolase 7 (NUDT7) is an enzyme that hydrolyzes CoA species, is highly expressed in the liver, and resides in the peroxisomes. Peroxisomes are organelles where the preferential oxidation of dicarboxylic fatty acids occurs and where the hepatic synthesis of the primary bile acids cholic acid and chenodeoxycholic acid is completed. We previously showed that liver-specific overexpression of NUDT7 affects peroxisomal lipid metabolism but does not prevent the increase in total liver CoA levels that occurs during fasting. We generated Nudt7-/- mice to further characterize the role that peroxisomal (acyl-)CoA degradation plays in the modulation of the size and composition of the acyl-CoA pool and in the regulation of hepatic lipid metabolism. Here, we show that deletion of Nudt7 alters the composition of the hepatic acyl-CoA pool in mice fed a low-fat diet, but only in males fed a Western diet does the lack of NUDT7 activity increase total liver CoA levels. This effect is driven by the male-specific accumulation of medium-chain dicarboxylic acyl-CoAs, which are produced from the β-oxidation of dicarboxylic fatty acids. We also show that, under conditions of elevated synthesis of chenodeoxycholic acid derivatives, Nudt7 deletion promotes the production of tauromuricholic acid, decreasing the hydrophobicity index of the intestinal bile acid pool and increasing fecal cholesterol excretion in male mice. These findings reveal that NUDT7-mediated hydrolysis of acyl-CoA pathway intermediates in liver peroxisomes contributes to the regulation of dicarboxylic fatty acid metabolism and the composition of the bile acid pool.

Abstract 9700: Totum-070 Reduces Hypercholesterolemia in Western Diet Fed Mice and Increases Cholesterol Fecal Excretion

Lowering blood cholesterol level reduces the incidence of cardiovascular diseases. Totum-070 (T070) is a polyphenol-rich active substance composed by the association of 5 plant extracts. We tested the effect of supplementation of T070 in dyslipidemic mice and characterized the mechanisms of action. C57Bl/6 mice were fed with either a normal diet (ND) or a western diet (WD) for 6 weeks. Mice fed WD were split in different groups (vehicle, T070 1g/kg and T070 3g/kg) depending on the treatment received daily by gavage (n=14 per group). Compared to WD-fed mice, the concentration of total cholesterol was decreased after 3 weeks of supplementation by -9% (p&lt;0.05) and -12.7% (p&lt;0.01) with 1 g/kg T070 and 3 g/kg T070, respectively. At the end of the study, total cholesterol was reduced by -7.3% (n.s) and -11% (p&lt;0.01) with 1 g/kg and 3 g/kg T070 compared to WD-fed mice. Mice receiving 1g/kg T070 exhibited similar body weight gain than mice from the WD group. On the contrary, WD-fed mice receiving 3 g/kg T070 had reduced body weight gain by 29.5% compared to WD-fed mice (p&lt;0.01). The supplementation of T070 reduced hepatic steatosis in a dose-dependent manner with a 17.4% drop at 1 g/kg T070 (p&lt;0.05) and a 40% reduction at 3g/kg T070 (p&lt;0.0001) compared to WD-fed mice. Hepatic and intestinal gene expression were altered by WD feeding compared to ND, while supplementation with T070 tended to restore normal levels of expression on several genes implicated in lipid and lipoprotein metabolism. Fecal neutral sterols were quantified in feces collected at the end of the study. Sterol quantity was increased by 16% in feces from the 3g/kg T070 group compared to the WD group. Furthermore, a significant 13% increase in feces production was observed in mice supplemented with T070 3 g/kg compared to the mice fed with WD alone (p&lt;0.01). Fresh feces were harvested of the mice and composition of microbiota was assessed by 16S rDNA sequencing. WD induced a decrease in richness (Chao) and diversity (Shannon) indexes, compared to ND. T070 supplementation was able to partially restore diversity index, compared to WD. In conclusion, T070 supplementation is a promising strategy to prevent hypercholesterolemia though increased of fecal cholesterol excretion and amelioration of gut microbial imbalance.

A Novel, Orally Bioavailable, Small-Molecule Inhibitor of PCSK9 With Significant Cholesterol-Lowering Properties In Vivo

Proprotein convertase subtilisin kexin type 9 (PCSK9) inhibits the clearance of low-density lipoprotein (LDL) cholesterol (LDL-C) from plasma by directly binding with the LDL receptor (LDLR) and sending the receptor for lysosomal degradation. As the interaction promotes elevated plasma LDL-C levels, and therefore a predisposition to cardiovascular disease, PCSK9 has attracted intense interest as a therapeutic target. Despite this interest, an orally bioavailable small-molecule inhibitor of PCSK9 with extensive lipid-lowering activity is yet to enter the clinic. We report herein the discovery of NYX-PCSK9i, an orally bioavailable small-molecule inhibitor of PCSK9 with significant cholesterol-lowering activity in hyperlipidemic APOE∗3-Leiden.CETP mice. NYX-PCSK9i emerged from a medicinal chemistry campaign demonstrating potent disruption of the PCSK9-LDLR interaction in vitro and functional protection of the LDLR of human lymphocytes from PCSK9-directed degradation ex vivo. APOE∗3-Leiden.CETP mice orally treated with NYX-PCSK9i demonstrated a dose-dependent decrease in plasma total cholesterol of up to 57%, while its combination with atorvastatin additively suppressed plasma total cholesterol levels. Importantly, the majority of cholesterol lowering by NYX-PCSK9i was in non-HDL fractions. A concomitant increase in total plasma PCSK9 levels and significant increase in hepatic LDLR protein expression strongly indicated on-target function by NYX-PCSK9i. Determinations of hepatic lipid and fecal cholesterol content demonstrated depletion of liver cholesteryl esters and promotion of fecal cholesterol elimination with NYX-PCSK9i treatment. All measured in vivo biomarkers of health indicate that NYX-PCSK9i has a good safety profile. NYX-PCSK9i is a potential new therapy for hypercholesterolemia with the capacity to further enhance the lipid-lowering activities of statins.

Totum-070 prevents diet-induced hypercholesterolemia in western diet fed mice and inhibits cholesterol uptake in human enterocyte caco2 cell line

Abstract Introduction It is well known that hypercholesterolemia is an important modifiable risk factor in the development of cardiovascular diseases. Lowering blood cholesterol level reduces the incidence of atherosclerotic disease. Totum-070 (T070) is a clinical stage polyphenol-rich active substance composed by the association of 5 plant extracts. Purpose We assessed the hypothesis that administration of T070 prevents hypercholesterolemia in western diet fed mice and explored the potential mechanisms involved in vitro in human enterocytes. Methods C57BL/6 mice were fed either a normal diet (ND) or western diet (WD) for 6 weeks and received either vehicle or T070 (1mg/kg or 3mg/kg) daily by gavage (n=14 mice per group). Food intake, body weight and plasma lipid were monitored along study duration. Liver and feces lipid content were quantified at the end of the study. For in vitro studies, human Caco2 cells were differentiated in transwells for 21 days. To perform cholesterol uptake assay, [1,2-3H(N)]-cholesterol was incubated in apical medium for 1 hour in presence or not of T070 prior to quantification of radioactivity in the cells. Results In mice, supplementation with T070 had no effect on food intake during the study. At the end of the in vivo study, body weight in WD-fed mice was increased to 28.3±0.8 g compared to the ND group (24.5±0.4 g, p&amp;lt;0.001). Interestingly, a significant (p&amp;lt;0.01) 29% reduction in body weight gain was observed in the T070 3mg/kg group compared to the WD group at the end of the study. Compared to WD group, total-cholesterol in mice supplemented by T070 was reduced in a dose dependent manner (298±8 mg/dl WD vs 276±9 mg/dl T070 1mg/kg, p=0.19, and 265±8 mg/dl T070 3mg/kg, p&amp;lt;0.05). Furthermore, hepatic steatosis induced by the WD was reduced by 73% (p&amp;lt;0.001) in the T070 3mg/kg group. Hepatic and intestinal gene expression were drastically altered by WD feeding compared to ND, while supplementation with T070 tended to restore normal expression levels for many genes implicated in lipid and lipoprotein metabolism. A significant increase in fecal cholesterol excretion was observed in mice supplemented with T070 3 mg/g compared to the mice fed with WD alone (442.3±151.7 nmol/day WD + T070 3mg/g vs 75.4±5.9 nmol/day WD, p&amp;lt;0.05). Studies in human enterocytes demonstrated that 1 g/l T070 reduced cholesterol uptake by 35% (p&amp;lt;0.001), suggesting that T070 inhibits intestinal cholesterol absorption. Conclusion Our results show the beneficial effect of supplementation with Totum-070 to prevent induction of hypercholesterolemia in a nutritional mouse model. Investigation of mechanisms of action in enterocytes demonstrate the role of Totum-070 to inhibit intestinal cholesterol absorption. These results highlight the interest of using Totum-070 for the management of hypercholesterolemia. Funding Acknowledgement Type of funding sources: None.