Fecal Neutral Sterol Research Articles

Ursodeoxycholic Acid for Trans Intestinal Cholesterol Excretion Stimulation: A Randomized Placebo Controlled Crossover Study.

The trans intestinal cholesterol excretion (TICE) pathway is a potential therapeutic target to reduce plasma low-density lipoprotein (LDL) cholesterol levels. TICE encompasses the direct excretion of cholesterol by enterocytes into feces. In mice, TICE has been shown to be stimulated by a hydrophilic bile acid pool, resulting in increased fecal neutral sterol loss and reduced plasma cholesterol levels. We investigated whether treatment with a hydrophilic bile acid, ursodeoxycholic acid (UDCA), would increase fecal neutral sterols in humans as a proxy for TICE. We performed a randomized, double-blind, placebo-controlled, cross-over trial in 20 male participants aged >18 years, with plasma LDL cholesterol levels ≥2.6 mmol/L. After a run-in period of ezetimibe 20 mg once daily for 3 weeks, patients were randomized to UDCA 600 mg or placebo orally once daily for 2 weeks. After a 3 week washout, patients underwent the alternate treatment. At baseline, mean (SD) age, body mass index, and plasma LDL cholesterol were 59±11.3 years, 26.4±3.1 kg/m2, and 3.9±0.8 mmol/L, respectively. After UDCA treatment, the plasma bile acid hydrophobicity index was reduced compared with placebo (-118.7% versus +2.3%, P<0.001). The fecal neutral sterols did not change (-5.8% versus +18.8%, P=0.51) and treatment with UDCA increased LDL cholesterol with 0.39 mmol/L (+8.1% versus -3.64%, P=0.002) when compared with placebo. UDCA in combination with ezetimibe increased plasma bile acid hydrophilicity in healthy subjects with LDL cholesterol levels >2.6 mmol/L but did not result in increased fecal neutral sterols or decreased LDL cholesterol. This suggests that TICE is not stimulated by an increase in the hydrophilicity of the bile acid pool in humans.

The profile of cholesterol metabolism does not interfere with the cholesterol-lowering efficacy of phytostanol esters

Increasing evidence suggests that high cholesterol absorption efficiency enhances the risk of atherosclerotic cardiovascular diseases. It is not known whether inhibiting cholesterol absorption has different metabolic effects in high- vs. low cholesterol absorbers. We evaluated the effects of phytostanol esters on serum lipids and cholesterol metabolism in a post hoc study of three randomized, double-blind, controlled trials. The participants were classified into low (n=20) and high (n=21) cholesterol absorbers by median cholesterol absorption efficiency based on the plasma cholesterol absorption marker cholestanol at baseline. The participants consumed mayonnaise or margarine without or with phytostanol esters for six to nine weeks without other changes in the diet or lifestyle. Serum cholesterol, cholestanol, lathosterol, and faecal neutral sterols and bile acids were analysed by gas-liquid chromatography. According to power calculations, the size of the study population (n=41) was appropriate. During the control period, cholesterol synthesis, and faecal neutral sterols and bile acids were lower in high- vs. low absorbers (p<0.05 for all). Phytostanol esters reduced low-density lipoprotein cholesterol by 10-13% in both groups, and directly measured cholesterol absorption efficiency by 41±7% in low- and 47±5% in high absorbers (p<0.001 for all) without side effects. Cholesterol synthesis and faecal neutral sterols (p<0.01) increased in both groups, more markedly in the high vs. low absorbers (p<0.01). Low cholesterol absorption combined with high faecal neutral sterol excretion are components of reverse cholesterol transport. Thus, high- vs. low absorbers had a more disadvantageous metabolic profile at baseline. In both groups, phytostanol esters induced favourable changes in serum, lipoprotein, and metabolic variables known to help in prevention of the development of atherosclerotic cardiovascular diseases.

Changes in bile acid composition are correlated with reduced intestinal cholesterol uptake in intestine-specific WASH-deficient mice

The Wiskott-Aldrich syndrome protein and SCAR homolog (WASH) complex is a pentameric protein complex localized at endosomes, where it facilitates the transport of numerous receptors from endosomes toward the plasma membrane. Recent studies have shown that the WASH complex plays an essential role in cholesterol and glucose homeostasis in humans and mice. To investigate the physiological importance of intestinal WASH, we ablated the WASH component WASHC1 specifically in murine enterocytes. Male and female intestine-specific WASHC1-deficient mice (Washc1IKO) were challenged with either a standard chow diet or a high-cholesterol (1.25 %) diet (HCD). Washc1IKO mice fed a standard diet did not present any apparent phenotype, but when fed an HCD, their hepatic cholesterol levels were ~ 50 % lower compared to those observed in control mice. The intestinal cholesterol absorption was almost 2-fold decreased in Washc1IKO mice, which translated into increased fecal neutral sterol loss. The intestinal expression of cholesterogenic genes, such as Hmgcs1, Hmgcr, and Ldlr, was significantly higher in Washc1IKO mice than in control mice and correlated with increased whole-body de novo cholesterol synthesis, likely to compensate for impaired intestinal cholesterol absorption. Unexpectedly, the ratio of biliary 12α−/non-12α-hydroxylated bile acids (BAs) was decreased in Washc1IKO mice and reversing this reduced ratio by feeding the mice with the HCD supplemented with 0.5 % (w/w) sodium cholate normalized the improvement of hepatic cholesterol levels in Washc1IKO mice. Our data indicate that the intestinal WASH complex plays an important role in intestinal cholesterol absorption, likely by modulating biliary BA composition.

Fisetin, a dietary flavonoid, promotes transintestinal cholesterol excretion through the activation of PPARδ

Fisetin, a dietary polyphenol abundantly found in strawberries, exhibits a broad spectrum of health-promoting activities, including antihyperlipidemic effects. This study aimed to investigate the regulatory effect of fisetin on cholesterol elimination through novel transintestinal cholesterol excretion (TICE) pathway. A hypercholesterolemic mouse model and human colon epithelial cancer cell line Caco-2 were utilized to conduct the study. In hypercholesterolemic mice, fisetin (25 mg/kg) treatment reduced serum total cholesterol by 46.48% and significantly decreased lipid accumulation in the liver. Furthermore, fisetin administration led to a substantial increase in the fecal neutral sterol contents, including coprostanol, coprostanone, dihydrocholesterol, and cholesterol. Specifically, these sterol contents increased by approximately 224.20%, 151.40%, 70.40% and 50.72% respectively. The fluorescence intensity of 22-NBD-cholesterol in intestinal perfusion increased by 95.94% in fisetin group (25 mg/kg), indicating that fisetin stimulated TICE. In high cholesterol-induced Caco-2 cells, fisetin at a concentration of 30 μM reduced total cholesterol and free cholesterol by 37.21% and 45.30% respectively, stimulated cholesterol excretion, and inhibited cholesterol accumulation. Additionally, fisetin upregulated the gene and protein expression of cholesterol efflux transporters ABCG5/G8 and ABCB1, while downregulating the cholesterol uptake regulator NPC1L1. Furthermore, fisetin increased LDLR protein expression and decreased PCSK9 expression. Notably, fisetin significantly activated nuclear receptor PPARδ in Caco-2 cells. PPARδ antagonist pretreatment counteracted the regulatory effects of fisetin on TICE regulators, suggesting fisetin lowered cholesterol through enhancing TICE by activation of intestinal PPARδ. Fisetin could be used as functional dietarysupplement for eliminating cholesterol and reducing the incidence of cardiovascular diseases.

Curcumin protects against high-fat diet-induced nonalcoholic simple fatty liver by inhibiting intestinal and hepatic NPC1L1 expression via down-regulation of SREBP-2/HNF1α pathway in hamsters

Niemann-pick C1-like 1 (NPC1L1) mediates cholesterol absorption and plays a key role in the pathogenesis of nonalcoholic simple fatty liver (NASFL). Our previous study showed that curcumin reduced NPC1L1 expression and cholesterol absorption in Caco-2 cells. This study aimed to investigate whether curcumin could inhibit intestinal and hepatic NPC1L1 expression through suppressing sterol regulatory element binding protein-2 (SREBP-2) / hepatocyte nuclear factor 1α (HNF1α) pathway, then exert anti-NASFL effects. Six-week hamsters were fed high-fat diet (HFD) with or without 0.1% curcumin for 12 weeks. Curcumin supplementation lowered blood total cholesterol (TC), triglycerides (TG) and low-density lipoprotein cholesterol levels (20.2%, 48.7%, and 36.5%), and reduced liver TC and TG contents (26.1% and 26.5%). Oil Red O staining demonstrated that curcumin significantly alleviated HFD-induced liver fat accumulation and hepatic steatosis, which was accompanied by reduced intestinal and hepatic NPC1L1, SREBP-2 and HNF1α expression (P < .05) and increased fecal neutral sterol excretion (114.5%). Furthermore, curcumin decreased cholesterol absorption in Caco-2 cells and HepG2 cells (49.2 % and 52.7 %). The inhibitory effects of curcumin on NPC1L1 expression and cholesterol absorption could be prevented by blockade of the SREBP-2 and HNF1α pathway. These findings indicated that curcumin protected against HFD-induced NASFL by inhibiting intestinal and hepatic NPC1L1 expression via down-regulation of SREBP-2/HNF1α pathway, thus reducing intestinal cholesterol absorption and hepatic biliary cholesterol reabsorption, consequently alleviating liver cholesterol accumulation and steatosis. Our study provides evidence for curcumin as a potential nutritional therapy for NASFL by regulating NPC1L1 and enterohepatic circulation of cholesterol.

Abstract 447: Cholesterol Excretion In Whole-body And Liver-specific ABCG5 ABCG8-deficient Mice

Background: The ABCG5 ABCG8 sterol transporter is the body’s primary defense against the accumulation of dietary sterols. Loss of function mutations in ABCG5 or ABCG8 result in Sitosterolemia, a rare disorder that presents as familial hypercholesterolemia, but is distinguished by recessive genetics and the accumulation of phytosterols in plasma and tissues. Biliary cholesterol is reduced by greater than 80% in ABCG5 ABCG8-deficient mice, but fecal neutral sterols are unaffected, suggesting a G5G8-independent pathway for cholesterol excretion. The objectives of the present study were to determine the ability of whole-body and liver-specific G5G8-deficient mice to maintain fecal neutral sterol output when challenged with cholesterol -enriched diet and determine maximal rates of biliary cholesterol secretion. Methods: Wild-type, whole-body and liver-specific ABCG5 ABCG8-deficient mice were maintained on cereal-based mixed diet, a phytosterol-free purified diet, or a purified diet supplemented with 0.2% cholesterol. Following acclimatization to each diet, fecal neutral sterol output was determined over a three-day period by gas chromatography/mass spectrometry. Maximal biliary cholesterol secretion rates were determined by biliary diversion with bile acid infusion over a range of 0 to 1000 nmol/minute. Biliary lipid secretion was determined by enzymatic colorimetric assays and molecular coupling of cholesterol to bile acid and phospholipid secretion calculated by linear regression. Results &amp; Conclusions: Whole-body G5G8-deficient mice exhibited a 40% reduction in fecal neutral sterol output compared to wild-type mice when maintained on cereal-based or purified, low cholesterol containing diets. Cholesterol feeding resulted in a 10-fold increase in fecal neutral sterols in wild-type mice, an effect not observed in G5G8-deficient mice. These data indicate that ABCG5 ABCG8 is indispensable in opposing the accumulation of dietary cholesterol and that alternative pathways cannot compensate for G5G8 deficiency.

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.

Abstract 439: Abcg5 Abcg8 Are Essential To Maintain Sterol Balance When Challenged With Dietary Cholesterol In Mice

Background: ABCG5/ABCG8 (G5/G8) is the primary biliary cholesterol transporter and accounts for &gt;70% of biliary cholesterol secretion. Mice and humans maintain fecal cholesterol excretion under a variety of conditions that disrupt biliary cholesterol secretion, indicating the presence of a non-biliary route for cholesterol elimination. Cholesterol can be actively secreted from the intestine, a pathway termed transintestinal cholesterol excretion (TICE). The present study investigates rates of intestinal cholesterol secretion in mice lacking G5/G8 to determine if adaptive responses within the gut facilitate cholesterol excretion in the absence of the primary biliary cholesterol transporter. Methods: Wild-type (WT) and whole body Abcg5/Abcg8 deficient mice (G5/G8 KO) were maintained on a plant sterol free (PSF) diet to prevent the development of experimental sitosterolemia and potential confounding effects of biologically active phytosterols. Mice were anesthetized, the gall bladder cannulated, and the intestine perfused with model bile to simultaneously determine biliary and intestinal cholesterol secretion rates. Sterol balance studies were conducted in mice sequentially maintained on PSF diet, a PSF diet supplemented with 0.2% cholesterol, and standard rodent chow. Mice were allowed to acclimate to each diet for 14 days, then placed on wire-bottom cages for fecal collection. At termination of the experiment, basal bile, plasma, and tissues were collected. Fecal neutral sterols (FNS) were analyzed by GC-MS. Results: Despite a substantial reduction in biliary cholesterol secretion in G5/G8 KO mice, differences in intestinal cholesterol secretion rates were only observed in males, with KO mice excreting less cholesterol into the intestinal lumen. FNS did not differ between genotypes when maintained on the PSF diet. On chow diet, FNS increased 2x in both genotypes, but did not differ across genotypes. When PSF diet was supplemented with 0.2% cholesterol, FNS increased by 3x in WT mice, but were unchanged in G5/G8 KO mice, relative to PSF diet levels. Conclusions: Abcg5/Abcg8 is essential to maintain sterol balance when cholesterol is present in the diet. TICE is unable to compensate for Abcg5/Abcg8 deficiency in cholesterol fed mice.

Taurine Alleviates Trimethylamine N-Oxide-Induced Atherosclerosis by Regulating Bile Acid Metabolism in ApoE-/- Mice.

Trimethylamine N-oxide (TMAO) widely exists in seafood and is associated with the atherosclerosis progress, but dietary seafood reduced the cardiovascular risk. This intimates that there may be some ingredients in seafood to offset the cardiovascular risk caused by TMAO. Taurine is a marker ingredient in seafood. Thus, this study determined the influences of taurine on TMAO-induced atherosclerosis in apolipoprotein-E-deficient mice. The results showed that dietary taurine significantly reduced the TMAO-induced atherosclerotic lesion area. Further studies found that taurine increased the hepatic- and serum-conjugated bile acid/unconjugated bile acid ratio via increasing hepatic gene expression of conjugated bile acid synthesis. Meanwhile, taurine changed TMAO-induced abnormal bile acid profiles in the gallbladder. Moreover, taurine increased bile acid deconjugation by enhancing the genera Ruminiclostridium level and increased excretion of fecal neutral sterols. Additionally, taurine attenuated inflammation in the serum and artery. These results indicate that taurine alleviated TMAO-induced atherosclerosis via regulating bile acid metabolism.

Deficiency of coiled-coil domain containing 80 increases plasma cholesterol by decreasing fecal sterols excretion in hypercholesterolemic mice

Disorders in cholesterol and bile acid metabolism have been acknowledged as critical in pathogenesis of hypercholesterolemia. Coiled-coil domain containing 80 (CCDC80) has been closely linked to lipid homeostasis in mice, with its role in cholesterol metabolism yet to be fully elucidated. This study aims to uncover the regulatory mechanisms of CCDC80 in diet-induced hypercholesterolemia. We generated a CCDC80 knockout (CCDC80−/−) model in C57BL/6 mouse. The initial transcriptional and metabolic consequences of removing CCDC80 were accessed at baseline by gene expression microarrays and gas chromatography-mass spectrometry / ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry, respectively. The hepatic cholesterol was investigated in both CCDC80+/+ and CCDC80−/− male mice at baseline and after feeding a high-cholesterol diet for 12 weeks. The regulatory effects of CCDC80 on gene expressions and protein masses were measured by RT-qPCR and western blot, respectively. At baseline, the KEGG pathway enrichment analysis combining metabolomics, lipidomics and transcriptomics, revealed a down-regulation of hepatic bile acid biosynthesis by CCDC80-knockout, especially for primary bile acids. In the hypercholesterolemic models, our results showed that deficiency of CCDC80 increased plasma and liver cholesterol levels, but decreased fecal neutral and acidic sterols excretion in mice. Mechanistically, we found that such effects were partly mediated by attenuating the alternative pathway of bile acid synthesis catalyzed by oxysterol 7-alpha-hydroxylase (CYP7B1). In conclusion, our results suggest CCDC80 as a novel modulator of cholesterol homeostasis in male mice. Deficiency of CCDC80 could further impair fecal sterols excretion in diet-induced hypercholesterolemia.

Chronic Oral Administration of Mineral Oil Compared With Corn Oil: Effects on Gut Permeability and Plasma Inflammatory and Lipid Biomarkers.

We investigated the effects of chronic oral administration of mineral oil, versus corn oil as control, on intestinal permeability, inflammatory markers, and plasma lipids in APOE*3-Leiden.CETP mice. Mice received mineral oil or corn oil 15 or 30 μL/mouse/day for 16 weeks (15 mice/group). Intestinal permeability was increased with mineral versus corn oil 30 µL/day, shown by increased mean plasma FITC-dextran concentrations 2 h post-administration (11 weeks: 1.5 versus 1.1 μg/ml, p = 0.02; 15 weeks: 1.7 versus 1.3 μg/ml, p = 0.08). Mean plasma lipopolysaccharide-binding protein levels were raised with mineral versus corn oil 30 µL/day (12 weeks: 5.8 versus 4.4 μg/ml, p = 0.03; 16 weeks: 5.8 versus 4.5 μg/ml, p = 0.09), indicating increased intestinal bacterial endotoxin absorption and potential pro-inflammatory effects. Plasma cholesterol and triglyceride concentrations were decreased with mineral oil, without affecting liver lipids among treated groups. Fecal neutral sterol measurements indicated increased fecal cholesterol excretion with mineral oil 30 µL/day (+16%; p = 0.04). Chronic oral administration of mineral oil in APOE*3-Leiden.CETP mice increased intestinal permeability, with potential pro-inflammatory effects, and decreased plasma cholesterol and triglyceride levels. Our findings may raise concerns about the use of mineral oil as a placebo in clinical studies.

Resveratrol enhances trans-intestinal cholesterol excretion through selective activation of intestinal liver X receptor alpha

Resveratrol (RSV) is a dietary polyphenol with well-documented cardio-protective activity, but its effects on blood cholesterol levels remain to be established. Due to its poor bioavailability, tissue accumulation of RSV is extremely low except for that in the small intestine. In the present study, we aimed to investigate the dose-dependent effects of RSV on blood cholesterol levels and the involvement of small intestine in the cholesterol-lowering impacts of RSV. Mice were administrated with RSV at various doses with high-fat diet (HFD) or high-fat and high-cholesterol diet (HCD) for 12 weeks. The fecal neutral sterol contents were analyzed, and intestinal perfusion test was performed. An enteric barrier model using Caco-2 cells was established. We observed that RSV reduced blood cholesterol levels in a dose-dependent manner in mice fed with HFD or HCD. Further investigation revealed that RSV administration increased the bile acid pool size but did not affect cholesterol consumption or de novo cholesterol synthesis. Interestingly, RSV promoted trans-intestinal cholesterol excretion (TICE) by 2-fold in the intestinal perfusion test. In addition, RSV upregulated the expressions of ATP-binding cassette sub-family G member 5 or 8 (Abcg5/8) and ATP-binding cassette sub-family B member 1a or 1b (Abcb1a/b) by up to 8 times in the duodenum mucosa but not in the liver. RSV also significantly downregulated the expression of intestinal Niemann-Pick C1-Like 1 (Npc1l1). Knock-down of liver X receptor alpha (LXRα) but not Sirt1 by siRNA significantly blocked RSV-induced cholesterol excretion in Caco-2 cells. In conclusion, RSV could decrease circulating cholesterol levels through enhancing TICE and limiting cholesterol absorption via selective activation of intestinal LXRα.

Xylo-oligosaccharides ameliorate high cholesterol diet induced hypercholesterolemia and modulate sterol excretion and gut microbiota in hamsters

• Xylo-oligosaccharides reduced plasma total cholesterol levels. • Xylo-oligosaccharides increased fecal outputs of neutral and acidic sterols. • Xylo-oligosaccharides increased fecal outputs of short chain fatty acids. • Xylo-oligosaccharides influenced gut microbiota composition. The present study investigated the cholesterol-lowering activity of xylo- oligosaccharides and its associated underlying mechanisms in hamsters. Twenty-four hamsters were randomly divided into three groups and fed one of three diets, namely a low cholesterol diet, a high cholesterol diet (HCD), and an HCD diet with supplementation of 5% xylo -oligosaccharides for 6 weeks. The changes in gut microbiota, fecal neutral and acidic sterols were examined. Results exhibited that xylo- oligosaccharides could significantly reduce plasma total cholesterol, non-high-density lipoprotein cholesterol and total triacylglycerol by 11.24%, 24.89% and 38.72%, respectively (p < 0.05). Such benefits were associated with an increase in fecal outputs of neutral and acidic sterols as well as SCFAs. Furthermore, dietary supplementation with xylo -oligosaccharides could change the composition of gut microbiota. It was therefore concluded that xylo -oligosaccharides supplementation could reduce plasma cholesterol levels, enhance the excretion of neutral and acidic sterols, and promote the production of SCFAs via changing the gut microbiota composition.

Effects of dietary oils prepared from the internal organs of the Japanese giant scallop (Patinopecten yessoensis) on cholesterol metabolism in obese type-II diabetic KK-Ay mice.

Our research team has successfully prepared high‐quality scallop oil (SCO), containing high eicosapentaenoic acid (EPA) and phospholipids (PL) from the internal organs of Japanese giant scallop (Patinopecten yessoensis) which is the largest unutilized marine resource in Japan. In this study, we prepared SCOs from scallop internal organs obtained from Mutsu (Aomori) and Uchiura (Hokkaido) bays in Japan, and named them SCO‐M and SCO‐U, respectively. This study aimed to investigate the effects of dietary SCO‐M and SCO‐U on cholesterol metabolism in obese type‐II diabetic KK‐Ay mice. Four‐week‐old male KK‐Ay mice were divided into four groups. The Control group was fed with AIN93G‐modified high‐fat (3 wt% soybean oil + 17 wt% lard) diet, and the other groups were fed with high‐fat diet, in which 7 wt% of the lard contained in the Control diet was replaced with SCO‐M, SCO‐U, or tuna oil (TO). After the mice had been fed with the experimental diet for 49 days, their serum, liver, and fecal lipid contents, as well as their liver messenger ribonucleic acid expression levels, were evaluated. The SCO‐M and SCO‐U groups were significantly decreased liver cholesterol contents compared to those of the Control and TO groups, partially through the enhancement of the fecal neutral sterol excretions and the tendency to increase the cholesterol 7α‐hydroxylase expression level of the liver. These results indicated that dietary SCO‐M and SCO‐U exhibited cholesterol‐lowering functions in the liver that can help prevent the development of lifestyle‐related diseases.

Beneficial effects of plant sterols/stanols-containing milk powder on lipid metabolism in hamsters

The effect of plant sterols/stanols-containing milk powder (phytosterol milk powder, PSMP, containing 2.78% phytosterols mixture) on lipid metabolism in hamsters was investigated. One hundred male 7-week-old Golden Syrian hamsters were given free access to regular rodent chow and water for 1 week to acclimatize. Four extremely (10%) over- and underweight hamsters were eliminated. Sixteen hamsters were killed and examined for plasma and liver lipid compositions to establish the baseline. The remaining hamster were randomly divided into 5 groups, each group of hamsters had statistically similar average body weight, but fed with different experimental diets for 4 weeks. All 5 groups of hamsters were fed with high fat, high cholesterol diet containing different ingredients. Regular rodent chow diet was supplemented with 0.5% (w/w) cholesterol and corn/coconut oil mixture (corn oil/coconut oil = 1:1) to raise the final fat content to 15% (w/w) (Group 1 - control group). Group 2 was the positive control (PC) group, which was fed with diet containing 0.72% (w/w) phytosterols (consisted of 75% β-sitosterol and 10% campesterol). Groups 3-5 were the experimental groups, of which 12.95, 25.90 or 64.75% (w/w) PSMP 1x, 2x and 5x PSMP groups) was added to the diet and the phytosterol mixture contents in these diets were 0.36, 0.72 and 1.8% (w/w), respectively. At the end of 4-week feeding period, hamsters were killed and the plasma and hepatic lipid compositions together with the fecal neutral sterol content were determined. No adverse effects of PSMP on growth and health condition in hamsters were found in this experiment. At the highest feeding dose of PSMP (64.75%, w/w), hamsters had the highest body weight gain and the lowest plasma and liver lipid contents. PSMP showed significant effects on lowering the concentrations of plasma total cholesterol and low density lipoprotein cholesterol. It could also lower the atherogenic index (LDL-C/HDL-C), hepatic lipid levels and relative liver weight while raising the fecal cholesterol and phytosterols excretion in the hyperlipidemic hamsters.

Long-Term β-galacto-oligosaccharides Supplementation Decreases the Development of Obesity and Insulin Resistance in Mice Fed a Western-Type Diet.

ScopeThe gut microbiota might critically modify metabolic disease development. Dietary fibers such as galacto‐oligosaccharides (GOS) presumably stimulate bacteria beneficial for metabolic health. This study assesses the impact of GOS on obesity, glucose, and lipid metabolism.Methods and resultsFollowing Western‐type diet feeding (C57BL/6 mice) with or without β‐GOS (7% w/w, 15 weeks), body composition, glucose and insulin tolerance, lipid profiles, fat kinetics and microbiota composition are analyzed. GOS reduces body weight gain (p < 0.01), accumulation of epididymal (p < 0.05), perirenal (p < 0.01) fat, and insulin resistance (p < 0.01). GOS‐fed mice have lower plasma cholesterol (p < 0.05), mainly within low‐density lipoproteins, lower intestinal fat absorption (p < 0.01), more fecal neutral sterol excretion (p < 0.05) and higher intestinal GLP‐1 expression (p < 0.01). Fecal bile acid excretion is lower (p < 0.01) in GOS‐fed mice with significant compositional differences, namely decreased cholic, α‐muricholic, and deoxycholic acid excretion, whereas hyodeoxycholic acid increased. Substantial changes in microbiota composition, conceivably beneficial for metabolic health, occurred upon GOS feeding.ConclusionGOS supplementation to a Western‐type diet improves body weight gain, dyslipidemia, and insulin sensitivity, supporting a therapeutic potential of GOS for individuals at risk of developing metabolic syndrome.

Small molecule inhibition of gut microbial choline trimethylamine lyase activity alters host cholesterol and bile acid metabolism.

The gut microbe-derived metabolite trimethylamine-N-oxide (TMAO) has recently been linked to cardiovascular disease (CVD) pathogenesis, prompting the development of therapeutic strategies to reduce TMAO. Previous work has shown that experimental alteration of circulating TMAO levels via dietary alterations or inhibition of the host TMAO producing enzyme flavin containing monooxygenase 3 (FMO3) is associated with reorganization of host cholesterol and bile acid metabolism in mice. In this work, we set out to understand whether recently developed nonlethal gut microbe-targeting small molecule choline trimethylamine (TMA) lyase inhibitors also alter host cholesterol and bile acid metabolism. Treatment of mice with the mechanism-based choline TMA lyase inhibitor, iodomethylcholine (IMC), increased fecal neutral sterol loss in the form of coprostanol, a bacteria metabolite of cholesterol. In parallel, IMC treatment resulted in marked reductions in the intestinal sterol transporter Niemann-pick C1-like 1 (NPC1L1) and reorganization of the gut microbial community, primarily reversing choline supplemented diet-induced changes. IMC also prevented diet-driven hepatic cholesterol accumulation, causing both upregulation of the host hepatic bile acid synthetic enzyme CYP7A1 and altering the expression of hepatic genes critical for bile acid feedback regulation. These studies suggest that the gut microbiota-driven TMAO pathway is closely linked to both microbe and host sterol and bile acid metabolism. Collectively, as gut microbe-targeting choline TMA lyase inhibitors move through the drug discovery pipeline from preclinical models to human studies, it will be important to understand how these drugs impact both microbe and host cholesterol and bile acid metabolism.NEW & NOTEWORTHY The gut microbe-dependent metabolite trimethylamine-N-oxide (TMAO) has been strongly associated with cardiovascular mortality, prompting drug discovery efforts to identify points of therapeutic intervention within the microbe host TMAO pathway. Recently, mechanism-based small molecule inhibitors of the major bacterial trimethylamine (TMA) lyase enzymes have been developed, and these drugs show efficacy as anti-atherothrombotic agents. The novel findings of this study are that small molecule TMA lyase inhibition results in beneficial reorganization of host cholesterol and bile acid metabolism. This study confirms previous observations that the gut microbial TMAO pathway is intimately linked to host cholesterol and bile acid metabolism and provides further rationale for the development of small molecule choline TMA lyase inhibitors for the treatment of cardiometabolic disorders.

Plasma Cholesterol-Lowering Activity of Soybean Germ Phytosterols.

Soybean germ phytosterols (SGP) largely exist in soybean germ oil. Our previous study demonstrated that soybean germ oil was effective in reducing plasma cholesterol. However, it remains unknown if its phytosterols are the active ingredients responsible for the plasma cholesterol-lowering activity. The present study aimed to test the effect of SGP on plasma cholesterol and to investigate its associated underlying mechanisms using hamsters as animal model. Male hamsters (n = 40) were randomly divided into five groups (n = 8/group) and fed one of the five diets: a non-cholesterol diet (NCD), a high cholesterol diet (HCD), a HCD diet containing 0.5% cholestyramine (PC), and two HCD diets containing 0.1% (LP) and 0.2% (HP) SGP, respectively, for six weeks. Results showed that SPG reduced plasma cholesterol level in a dose-dependent manner, whereas it dose-dependently increased the excretion of both fecal neutral and acidic sterols. SGP was also effective in displacing cholesterol from micelles. It was concluded that SGP possessed hypocholesterolemic activity, likely by inhibiting cholesterol absorption in the intestine and promoting fecal sterol excretion.

Efficient reabsorption of transintestinally excreted cholesterol is a strong determinant for cholesterol disposal in mice

Transintestinal cholesterol excretion (TICE) is a major route for eliminating cholesterol from the body and a potential therapeutic target for hypercholesterolemia. The underlying mechanism, however, is largely unclear, and its contribution to cholesterol disposal from the body is obscured by the counteracting process of intestinal cholesterol reabsorption. To determine the quantity of TICE independent from its reabsorption, we studied two models of decreased intestinal cholesterol absorption. Cholesterol absorption was inhibited either by ezetimibe or, indirectly, by the genetic inactivation of the intestinal apical sodium-dependent bile acid transporter (ASBT; SLC10A2). Both ezetimibe treatment and Asbt inactivation virtually abrogated fractional cholesterol absorption (from 46% to 4% and 6%, respectively). In both models, fecal neutral sterol excretion and net intestinal cholesterol balance were considerably higher than in control mice (5- and 7-fold, respectively), suggesting that, under physiological conditions, TICE is largely reabsorbed. In addition, the net intestinal cholesterol balance was increased to a similar extent but was not further increased when the models were combined, suggesting that the effect on cholesterol reabsorption was already maximal under either condition alone. On the basis of these findings, we hypothesize that the inhibition of cholesterol (re)absorption combined with stimulating TICE will be most effective in increasing cholesterol disposal.

Glucose-6-Phosphate Regulates Hepatic Bile Acid Synthesis in Mice.

It is well established that, besides facilitating lipid absorption, bile acids act as signaling molecules that modulate glucose and lipid metabolism. Bile acid metabolism, in turn, is controlled by several nutrient‐sensitive transcription factors. Altered intrahepatic glucose signaling in type 2 diabetes associates with perturbed bile acid synthesis. We aimed to characterize the regulatory role of the primary intracellular metabolite of glucose, glucose‐6‐phosphate (G6P), on bile acid metabolism. Hepatic gene expression patterns and bile acid composition were analyzed in mice that accumulate G6P in the liver, that is, liver‐specific glucose‐6‐phosphatase knockout (L‐G6pc−/−) mice, and mice treated with a pharmacological inhibitor of the G6P transporter. Hepatic G6P accumulation induces sterol 12α‐hydroxylase (Cyp8b1) expression, which is mediated by the major glucose‐sensitive transcription factor, carbohydrate response element‐binding protein (ChREBP). Activation of the G6P‐ChREBP‐CYP8B1 axis increases the relative abundance of cholic‐acid–derived bile acids and induces physiologically relevant shifts in bile composition. The G6P‐ChREBP–dependent change in bile acid hydrophobicity associates with elevated plasma campesterol/cholesterol ratio and reduced fecal neutral sterol loss, compatible with enhanced intestinal cholesterol absorption. Conclusion: We report that G6P, the primary intracellular metabolite of glucose, controls hepatic bile acid synthesis. Our work identifies hepatic G6P‐ChREBP‐CYP8B1 signaling as a regulatory axis in control of bile acid and cholesterol metabolism.