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Chronic inflammation and proatherogenic lipids are important risk factors of cardiovascular disease (CVD). Specific dietary constituents such as polyphenols and fish oils may improve cardiovascular risk factors and may have a beneficial effect on disease outcomes. We hypothesized that the intake of an antiinflammatory dietary mixture (AIDM) containing resveratrol, lycopene, catechin, vitamins E and C, and fish oil would reduce inflammatory risk factors, proatherogenic lipids, and endpoint atherosclerosis. AIDM was evaluated in an inflammation model, male human C-reactive protein (CRP) transgenic mice, and an atherosclerosis model, female ApoE*3Leiden transgenic mice. Two groups of male human-CRP transgenic mice were fed AIDM [0.567% (wt:wt) powder and 0.933% (wt:wt oil)] or placebo for 6 wk. The effects of AIDM on basal and IL-1β-stimulated CRP expression were investigated. AIDM reduced cytokine-induced human CRP and fibrinogen expression in human-CRP transgenic mice. In the atherosclerosis study, 2 groups of female ApoE*3Leiden transgenic mice were fed an atherogenic diet supplemented with AIDM [0.567% (wt:wt) powder and 0.933% (wt:wt oil)] or placebo for 16 wk. AIDM strongly reduced plasma cholesterol, TG, and serum amyloid A concentrations compared with placebo. Importantly, long-term treatment of ApoE*3Leiden mice with AIDM markedly reduced the development of atherosclerosis by 96% compared with placebo. The effect on atherosclerosis was paralleled by a reduced expression of the vascular inflammation markers and adhesion molecules inter-cellular adhesion molecule-1 and E-selectin. Dietary supplementation of AIDM improves lipid and inflammatory risk factors of CVD and strongly reduces atherosclerotic lesion development in female transgenic mice. [ABSTRACT FROM AUTHOR]

It is unclear whether early metabolic and inflammatory aberrations in the liver are associated with detrimental changes in brain structure and cognitive function. This cross-sectional study examines putative associations between metabolic dysfunction-associated steatotic liver disease (MASLD) and brain health in 36-55 year-old participants with obesity (n = 70) from the BARICO study (BAriatric surgery Rijnstate and Radboudumc neuroImaging and Cognition in Obesity). The participants underwent brain magnetic resonance imaging to study brain volumes and cortical thickness (3T MRI including T1-weighted magnetization-prepared rapid gradient-echo sequence), cerebral blood perfusion (arterial spin labeling) and white matter integrity (diffusion weighted imaging to assess mean-skeletonized mean diffusivity and fluid-attenuated inversion recovery to detect the presence of white matter hyperintensities (WMH)). The participants additionally performed neuropsychological tests to assess global cognition, working and episodic memory, verbal fluency and the ability to shift attention. Liver biopsies were collected and liver dysfunction was examined with histopathological, biochemical, and gene expression analyses. Linear regression analyses were performed between liver and brain parameters and the influence of body-mass index, diabetes and hypertension was explored. Early stages of liver disease were not associated with cognitive status but with cerebrovascular changes independently of age, sex, BMI, diabetes and hypertension: hepatic fibrosis development was associated with higher spatial coefficient of variation (sCoV) in the nucleus accumbens (NAcc), reflecting greater variations in cerebral perfusion and reduced vascular efficiency. Elevated hepatic levels of free cholesterol and cholesteryl esters were associated with increased WMH, indicating cerebral small vessel disease. RNA-seq and pathway analyses identified associations between sCoV in NAcc and WMH and the expression of hepatic genes involved in inflammation and cellular stress. Additionally, sCoV in NAcc correlated with plasma IL-6 levels suggesting that systemic-low grade inflammation may, at least partly, mediate this relationship. In conclusion, this study demonstrates that specific features of liver dysfunction (e.g. free cholesterol, onset of fibrosis) are associated with subtle cerebrovascular impairments, when changes in cognitive performance are not yet noticeable. These findings highlight the need for future research on therapeutic strategies that normalize metabolic-inflammatory aberrations in the liver to reduce the risk of cognitive decline., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors. Published by Elsevier Ltd.)

Background: NAFLD progression, from steatosis to inflammation and fibrosis, results from an interplay of intra- and extrahepatic mechanisms. Disease drivers likely include signals from white adipose tissue (WAT) and gut. However, the temporal dynamics of disease development remain poorly understood., Methods: High-fat-diet (HFD)-fed Ldlr-/-.Leiden mice were compared to chow-fed controls. At t = 0, 8, 16, 28 and 38w mice were euthanized, and liver, WAT depots and gut were analyzed biochemically, histologically and by lipidomics and transcriptomics together with circulating factors to investigate the sequence of pathogenic events and organ cross-talk during NAFLD development., Results: HFD-induced obesity was associated with an increase in visceral fat, plasma lipids and hyperinsulinemia at t = 8w, along with increased liver steatosis and circulating liver damage biomarkers. In parallel, upstream regulator analysis predicted that lipid catabolism regulators were deactivated and lipid synthesis regulators were activated. Subsequently, hepatocyte hypertrophy, oxidative stress and hepatic inflammation developed. Hepatic collagen accumulated from t = 16 w and became pronounced at t = 28-38 w. Epididymal WAT was maximally hypertrophic from t = 8 w, which coincided with inflammation development. Mesenteric and subcutaneous WAT hypertrophy developed slower and did not appear to reach a maximum, with minimal inflammation. In gut, HFD significantly increased permeability, induced a shift in microbiota composition from t = 8 w and changed circulating gut-derived metabolites., Conclusion: HFD-fed Ldlr-/-.Leiden mice develop obesity, dyslipidemia and insulin resistance, essentially as observed in obese NAFLD patients, underlining their translational value. We demonstrate that marked epididymal-WAT inflammation, and gut permeability and dysbiosis precede the development of NAFLD stressing the importance of a multiple-organ approach in the prevention and treatment of NAFLD., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier Ltd.)

Background: Chronic inflammation is an important driver in the progression of non-alcoholic steatohepatitis (NASH) and atherosclerosis. The complement system, one of the first lines of defense in innate immunity, has been implicated in both diseases. However, the potential therapeutic value of complement inhibition in the ongoing disease remains unclear., Methods: After 20 weeks of high-fat diet (HFD) feeding, obese Ldlr-/-.Leiden mice were treated twice a week with an established anti-C5 antibody (BB5.1) or vehicle control. A separate group of mice was kept on a chow diet as a healthy reference. After 12 weeks of treatment, NASH was analyzed histopathologically, and genome-wide hepatic gene expression was analyzed by next-generation sequencing and pathway analysis. Atherosclerotic lesion area and severity were quantified histopathologically in the aortic roots., Results: Anti-C5 treatment considerably reduced complement system activity in plasma and MAC deposition in the liver but did not affect NASH. Anti-C5 did, however, reduce the development of atherosclerosis, limiting the total lesion size and severity independently of an effect on plasma cholesterol but with reductions in oxidized LDL (oxLDL) and macrophage migration inhibitory factor (MIF)., Conclusion: We show, for the first time, that treatment with an anti-C5 antibody in advanced stages of NASH is not sufficient to reduce the disease, while therapeutic intervention against established atherosclerosis is beneficial to limit further progression.

Non-alcoholic steatohepatitis (NASH) is associated with a disturbed metabolism in liver, insulin resistance, and excessive accumulation of ectopic fat. Branched-chain amino acids (BCAAs) may beneficially modulate hepatic lipids, however, it remains unclear whether individual BCAAs can attenuate already established NASH and associated oxidative-inflammatory stress. After a 26 weeks run-in on fast food diet (FFD), obese Ldlr-/-.Leiden mice were treated for another 12 weeks with either valine or isoleucine (3% of FFD) and then compared to FFD controls. Valine and isoleucine did not affect obesity, dyslipidemia, gut permeability, or fecal fatty acid excretion, but significantly reduced hyperinsulinemia. Valine and isoleucine reduced ALT, CK18-M30, and liver steatosis with a particularly pronounced suppression of the microvesicular component (-61% by valine and -71% by isoleucine). Both BCAAs decreased intrahepatic diacylglycerols and 4-hydroxynonenal immunoreactivity, a marker for oxidative stress-induced lipid peroxidation. Functional genomics analysis demonstrated that valine and isoleucine affected BCAA metabolism genes, deactivated master regulators of anabolic pathways related to steatosis (e.g., SREBPF1), and activated master regulators of mitochondrial biogenesis (e.g., PPARGC1A) and lipid catabolism (e.g., ACOX1, AMPK). This correction of critical metabolic pathways on gene expression level was accompanied by a significant decrease in histological liver inflammation, and suppression of FFD-stimulated cytokine and chemokine proteins KC/CXCL1, MCP-1/CCL2, and MIP-2/CXCL2 and their pathways. In conclusion, dietary intervention with either valine or isoleucine corrected liver diacylglycerols, gene expression of multiple metabolic processes, and reduced NASH histology with profound hepatoprotective effects on oxidative stress and inflammatory proteins., (© 2022 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

In obesity-associated non-alcoholic steatohepatitis (NASH), persistent hepatocellular damage and inflammation are key drivers of fibrosis, which is the main determinant of NASH-associated mortality. The short-chain fatty acid butyrate can exert metabolic improvements and anti-inflammatory activities in NASH. However, its effects on NASH-associated liver fibrosis remain unclear. Putative antifibrotic effects of butyrate were studied in Ldlr-/-.Leiden mice fed an obesogenic diet (HFD) containing 2.5% ( w / w ) butyrate for 38 weeks and compared with a HFD-control group. Antifibrotic mechanisms of butyrate were further investigated in TGF-β-stimulated primary human hepatic stellate cells (HSC). HFD-fed mice developed obesity, insulin resistance, increased plasma leptin levels, adipose tissue inflammation, gut permeability, dysbiosis, and NASH-associated fibrosis. Butyrate corrected hyperinsulinemia, lowered plasma leptin levels, and attenuated adipose tissue inflammation, without affecting gut permeability or microbiota composition. Butyrate lowered plasma ALT and CK-18M30 levels and attenuated hepatic steatosis and inflammation. Butyrate inhibited fibrosis development as demonstrated by decreased hepatic collagen content and Sirius-red-positive area. In TGF-β-stimulated HSC, butyrate dose-dependently reduced collagen deposition and decreased procollagen1α1 and PAI1 protein expression. Transcriptomic analysis and subsequent pathway and upstream regulator analysis revealed deactivation of specific non-canonical TGF-β signaling pathways Rho-like GTPases and PI3K/AKT and other important pro-fibrotic regulators (e.g., YAP/TAZ, MYC) by butyrate, providing a potential rationale for its antifibrotic effects. In conclusion, butyrate protects against obesity development, insulin resistance-associated NASH, and liver fibrosis. These antifibrotic effects are at least partly attributable to a direct effect of butyrate on collagen production in hepatic stellate cells, involving inhibition of non-canonical TGF-β signaling pathways.

The development of obesity is characterized by the metabolic overload of tissues and subsequent organ inflammation. The health effects of krill oil (KrO) on obesity-associated inflammation remain largely elusive, because long-term treatments with KrO have not been performed to date. Therefore, we examined the putative health effects of 28 weeks of 3% ( w / w ) KrO supplementation to an obesogenic diet (HFD) with fat derived mostly from lard. The HFD with KrO was compared to an HFD control group to evaluate the effects on fatty acid composition and associated inflammation in epididymal white adipose tissue (eWAT) and the liver during obesity development. KrO treatment increased the concentrations of EPA and DHA and associated oxylipins, including 18-HEPE, RvE 2 and 14-HDHA in eWAT and the liver. Simultaneously, KrO decreased arachidonic acid concentrations and arachidonic-acid-derived oxylipins (e.g., HETEs, PGD 2 , PGE 2 , PGF 2 α, TXB 2 ). In eWAT, KrO activated regulators of adipogenesis (e.g., PPARγ, CEBPα, KLF15, STAT5A), induced a shift towards smaller adipocytes and increased the total adipocyte numbers indicative for hyperplasia. KrO reduced crown-like structures in eWAT, and suppressed HFD-stimulated inflammatory pathways including TNFα and CCL2/MCP-1 signaling. The observed eWAT changes were accompanied by reduced plasma leptin and increased plasma adiponectin levels over time, and improved insulin resistance (HOMA-IR). In the liver, KrO suppressed inflammatory signaling pathways, including those controlled by IL-1β and M-CSF, without affecting liver histology. Furthermore, KrO deactivated hepatic REL-A/p65-NF-κB signaling, consistent with increased PPARα protein expression and a trend towards an increase in IkBα. In conclusion, long-term KrO treatment increased several anti-inflammatory PUFAs and oxylipins in WAT and the liver. These changes were accompanied by beneficial effects on general metabolism and inflammatory tone at the tissue level. The stimulation of adipogenesis by KrO allows for safe fat storage and may, together with more direct PPAR-mediated anti-inflammatory mechanisms, attenuate inflammation.

Background: Non-alcoholic fatty liver disease (NAFLD) is a complex multifactorial disorder that is characterised by dysfunctional lipid metabolism and cholesterol homeostasis, and a related chronic inflammatory response. NAFLD has become the most common cause of chronic liver disease in many countries, and its prevalence continues to rise in parallel with increasing rates of obesity. Here, we evaluated the putative NAFLD-attenuating effects of a multicomponent medicine consisting of 24 natural ingredients: Hepar compositum (HC-24)., Methods: Ldlr-/-.Leiden mice were fed a high-fat diet (HFD) with a macronutrient composition and cholesterol content comparable to human diets for 24 weeks to induce obesity-associated metabolic dysfunction, including hepatic steatosis and inflammation. HC-24 or vehicle control was administered intraperitoneally 3 times/week (1.5 ml/kg) for the last 18 weeks of the study. Histological analyses of liver and adipose tissue were combined with extensive hepatic transcriptomics analysis. Transcriptomics results were further substantiated with ELISA, immunohistochemical and liver lipid analyses., Results: HFD feeding induced obesity and metabolic dysfunction including adipose tissue inflammation and increased gut permeability. In the liver, HFD-feeding resulted in a disturbance of cholesterol homeostasis and an associated inflammatory response. HC-24 did not affect body weight, metabolic risk factors, adipose tissue inflammation or gut permeability. While HC-24 did not alter total liver steatosis, there was a pronounced reduction in lobular inflammation in HC-24-treated animals, which was associated with modulation of genes and proteins involved in inflammation (e.g., neutrophil chemokine Cxcl1) and cholesterol homeostasis (i.e., predicted effect on 'cholesterol' as an upstream regulator, based on gene expression changes associated with cholesterol handling). These effects were confirmed by CXCL1 ELISA, immunohistochemical staining of neutrophils and biochemical analysis of hepatic free cholesterol content. Intrahepatic free cholesterol levels were found to correlate significantly with the number of inflammatory aggregates in the liver, thereby providing a potential rationale for the observed anti-inflammatory effects of HC-24., Conclusions: Free cholesterol accumulates in the liver of Ldlr-/-.Leiden mice under physiologically translational dietary conditions, and this is associated with the development of hepatic inflammation. The multicomponent medicine HC-24 reduces accumulation of free cholesterol and has molecular and cellular anti-inflammatory effects in the liver., Competing Interests: AMu and BS are employees of Heel GmbH. NK and YB are former employees of Heel GmbH. Heel GmbH was involved in the design of the study and the preparation of the manuscript. Heel GmbH was not involved in data acquisition or data analysis. The publication of this study was a requirement of the funding received from Health~Holland, Top Sector Life Sciences & Health. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Mueller, Kleemann, Gart, van Duyvenvoorde, Verschuren, Caspers, Menke, Krömmelbein, Salic, Burmeister, Seilheimer and Morrison.)

The obesity epidemic increases the interest to elucidate impact of short-chain fatty acids on metabolism, obesity, and the brain. We investigated the effects of propionic acid (PA) and caproic acid (CA) on metabolic risk factors, liver and adipose tissue pathology, brain function, structure (by MRI), and gene expression, during obesity development in Ldlr -/- .Leiden mice. Ldlr -/- .Leiden mice received 16 weeks either a high-fat diet (HFD) to induce obesity, or chow as reference group. Next, obese HFD-fed mice were treated 12 weeks with (a) HFD + CA (CA), (b) HFD + PA (PA), or (c) a HFD-control group. PA reduced the body weight and systolic blood pressure, lowered fasting insulin levels, and reduced HFD-induced liver macrovesicular steatosis, hypertrophy, inflammation, and collagen content. PA increased the amount of glucose transporter type 1-positive cerebral blood vessels, reverted cerebral vasoreactivity, and HFD-induced effects in microstructural gray and white matter integrity of optic tract, and somatosensory and visual cortex. PA and CA also reverted HFD-induced effects in functional connectivity between visual and auditory cortex. However, PA mice were more anxious in open field, and showed reduced activity of synaptogenesis and glutamate regulators in hippocampus. Therefore, PA treatment should be used with caution even though positive metabolic, (cerebro) vascular, and brain structural and functional effects were observed., (© 2020 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Obesity characterized by adiposity and ectopic fat accumulation is associated with the development of non-alcoholic fatty liver disease (NAFLD). Treatments that stimulate lipid utilization may prevent the development of obesity and comorbidities. This study evaluated the potential anti-obesogenic hepatoprotective effects of combined treatment with L-carnitine and nicotinamide riboside, i.e., components that can enhance fatty acid transfer across the inner mitochondrial membrane and increase nicotinamide adenine nucleotide (NAD+) levels, which are necessary for β-oxidation and the TCA cycle, respectively. Ldlr -/-.Leiden mice were treated with high-fat diet (HFD) supplemented with L-carnitine (LC; 0.4% w / w ), nicotinamide riboside (NR; 0.3% w / w ) or both (COMBI) for 21 weeks. L-carnitine plasma levels were reduced by HFD and normalized by LC. NR supplementation raised its plasma metabolite levels demonstrating effective delivery. Although food intake and ambulatory activity were comparable in all groups, COMBI treatment significantly attenuated HFD-induced body weight gain, fat mass gain (-17%) and hepatic steatosis (-22%). Also, NR and COMBI reduced hepatic 4-hydroxynonenal adducts. Upstream-regulator gene analysis demonstrated that COMBI reversed detrimental effects of HFD on liver metabolism pathways and associated regulators, e.g., ACOX, SCAP, SREBF, PPARGC1B, and INSR. Combination treatment with LC and NR exerts protective effects on metabolic pathways and constitutes a new approach to attenuate HFD-induced obesity and NAFLD.

Background and Aims: As dietary saturated fatty acids are associated with metabolic and cardiovascular disease, a potentially interesting strategy to reduce disease risk is modification of the quality of fat consumed. Vegetable oils represent an attractive target for intervention, as they largely determine the intake of dietary fats. Furthermore, besides potential health effects conferred by the type of fatty acids in a vegetable oil, other minor components (e.g. phytochemicals) may also have health benefits. Here, we investigated the potential long-term health effects of isocaloric substitution of dietary fat (i.e. partial replacement of saturated by unsaturated fats), as well as putative additional effects of phytochemicals present in unrefined (virgin) oil on development of non-alcoholic fatty liver disease (NAFLD) and associated atherosclerosis. For this, we used pumpkin seed oil, because it is high in unsaturated fatty acids and a rich source of phytochemicals., Methods: ApoE*3Leiden mice were fed a Western-type diet (CON) containing cocoa butter (15% w/w) and cholesterol (1% w/w) for 20 weeks to induce risk factors and disease endpoints. In separate groups, cocoa butter was replaced by refined (REF) or virgin (VIR) pumpkin seed oil (comparable in fatty acid composition, but different in phytochemical content)., Results: Both oils improved dyslipidaemia, with decreased (V)LDL-cholesterol and triglyceride levels in comparison with CON, and additional cholesterol-lowering effects of VIR over REF. While REF did not affect plasma inflammatory markers, VIR reduced circulating serum amyloid A and soluble vascular adhesion molecule-1. NAFLD and atherosclerosis development was modestly reduced in REF, and VIR strongly decreased liver steatosis and inflammation as well as atherosclerotic lesion area and severity., Conclusions: Overall, we show that an isocaloric switch from a diet rich in saturated fat to a diet rich in unsaturated fat can attenuate NAFLD and atherosclerosis development. Phytochemical-rich virgin pumpkin seed oil exerts additional anti-inflammatory effects resulting in more pronounced health effects.

A high-fat diet disturbs the composition and function of the gut microbiota and generates local gut-associated and also systemic responses. Intestinal mast cells, for their part, secrete mediators which play a role in the orchestration of physiological and immunological functions of the intestine. Probiotic bacteria, again, help to maintain the homeostasis of the gut microbiota by protecting the gut epithelium and regulating the local immune system. In the present study, we explored the effects of two probiotic bacteria, Lactobacillus rhamnosus GG (GG) and Propionibacterium freudenreichii spp. shermanii JS (PJS), on high fat-fed ApoE*3Leiden mice by estimating the mast cell numbers and the immunoreactivity of TNF-α and IL-10 in the intestine, as well as plasma levels of several markers of inflammation and parameters of lipid metabolism. We found that mice that received GG and PJS exhibited significantly lower numbers of intestinal mast cells compared with control mice. PJS lowered intestinal immunoreactivity of TNF-α, while GG increased intestinal IL-10. PJS was also observed to lower the plasma levels of markers of inflammation including vascular cell adhesion molecule 1, and also the amount of gonadal adipose tissue. GG lowered alanine aminotransferase, a marker of hepatocellular activation. Collectively, these data demonstrate that probiotic GG and PJS tend to down-regulate both intestinal and systemic pro-inflammatory changes induced by a high-fat diet in this humanised mouse model.

Excess caloric intake leads to metabolic overload and is associated with development of type 2 diabetes (T2DM). Current disease management concentrates on risk factors of the disease such as blood glucose, however with limited success. We hypothesize that normalizing blood glucose levels by itself is insufficient to reduce the development of T2DM and complications, and that removal of the metabolic overload with dietary interventions may be more efficacious. We explored the efficacy and systems effects of pharmaceutical interventions versus dietary lifestyle intervention (DLI) in developing T2DM and complications. To mimic the situation in humans, high fat diet (HFD)-fed LDLr-/- mice with already established disease phenotype were treated with ten different drugs mixed into HFD or subjected to DLI (switch to low-fat chow), for 7 weeks. Interventions were compared to untreated reference mice kept on HFD or chow only. Although most of the drugs improved HFD-induced hyperglycemia, drugs only partially affected other risk factors and also had limited effect on disease progression towards microalbuminuria, hepatosteatosis and atherosclerosis. By contrast, DLI normalized T2DM risk factors, fully reversed hepatosteatosis and microalbuminuria, and tended to attenuate atherogenesis. The comprehensive beneficial effect of DLI was reflected by normalized metabolite profiles in plasma and liver. Analysis of disease pathways in liver confirmed reversion of the metabolic distortions with DLI. This study demonstrates that the pathogenesis of T2DM towards complications is reversible with DLI and highlights the differential effects of current pharmacotherapies and their limitation to resolve the disease.

Background: The prevalence of dyslipidemia and obesity resulting from excess energy intake and physical inactivity is increasing. The liver plays a pivotal role in systemic lipid homeostasis. Effective, natural dietary interventions that lower plasma lipids and promote liver health are needed., Objective: Our goal was to determine the effect of dietary sphingolipids on plasma lipids and liver steatosis., Design: APOE*3Leiden mice were fed a Western-type diet supplemented with different sphingolipids. Body cholesterol and triacylglycerol metabolism as well as hepatic lipid concentrations and lipid-related gene expression were determined., Results: Dietary sphingolipids dose-dependently lowered both plasma cholesterol and triacylglycerol in APOE*3Leiden mice; 1% phytosphingosine (PS) reduced plasma cholesterol and triacylglycerol by 57% and 58%, respectively. PS decreased the absorption of dietary cholesterol and free fatty acids by 50% and 40%, respectively, whereas intestinal triacylglycerol lipolysis was not affected. PS increased hepatic VLDL-triacylglycerol production by 20%, whereas plasma lipolysis was not affected. PS increased the hepatic uptake of VLDL remnants by 60%. Hepatic messenger RNA concentrations indicated enhanced hepatic lipid synthesis and VLDL and LDL uptake. The net result of these changes was a strong decrease in plasma cholesterol and triacylglycerol. The livers of 1% PS-fed mice were less pale, 22% lighter, and contained 61% less cholesteryl ester and 56% less triacylglycerol than livers of control mice. Furthermore, markers of liver inflammation (serum amyloid A) and liver damage (alanine aminotransferase) decreased by 74% and 79%, respectively, in PS-fed mice., Conclusion: Sphingolipids lower plasma cholesterol and triacylglycerol and protect the liver from fat- and cholesterol-induced steatosis.

The present study was designed to investigate the lipid-lowering properties and mechanisms of action of a new HMG-CoA reductase inhibitor, rosuvastatin, in female ApoE*3-Leiden transgenic mice. Mice received a high fat/cholesterol (HFC) diet containing either rosuvastatin (0 [control], 0.00125%, 0.0025%, or 0.005% [w/w]) or 0.05% (w/w) lovastatin. The highest dose of rosuvastatin reduced plasma cholesterol and triglyceride levels by 39% and 42%, respectively, compared with the HFC control. Lovastatin had no effect on plasma cholesterol and triglyceride levels. In ApoE*3-Leiden mice on a chow diet, rosuvastatin (0.005% [w/w]) decreased plasma cholesterol levels by 35% without having an effect on triglyceride levels. On a chow diet, expression of genes involved in cholesterol biosynthesis and uptake in the liver was increased by rosuvastatin. Further mechanistic studies in HFC-fed mice showed that rosuvastatin treatment resulted in decreased hepatic VLDL-triglyceride and VLDL-apolipoprotein B production. VLDL lipid composition remained unchanged, indicating a reduction in the number of VLDL particles secreted. Lipolytic activity and expression of genes involved in cholesterol and triglyceride synthesis and beta-oxidation of fatty acids in the liver were not affected by rosuvastatin treatment, and hepatic lipid content did not change. However, activity of hepatic diacylglycerol acyltransferase was significantly decreased by 25% after rosuvastatin treatment. Moreover, biliary excretion of cholesterol, phospholipids, and bile acids was increased during treatment. The results indicate that rosuvastatin treatment in ApoE*3-Leiden mice on a HFC diet leads to redistribution of cholesterol and triglycerides in the body, both by reduced hepatic VLDL production and triglyceride synthesis and by enhanced hepatobiliary removal of cholesterol, bile acids, and phospholipids, resulting in substantial reductions in plasma cholesterol and triglyceride levels.

Numerous animal studies have reported that garlic can protect against atherosclerosis. However, a comparable number of studies do not support this observation. This contradiction may result from differences in study design, use of different animal models, and use of different garlic formulations and preparations. Here, we investigated the effect of the chemically well-characterized and production-controlled garlic powder printanor on atherosclerosis in the APOE*3-Leiden transgenic mouse, a mouse model well suited for evaluating anti-atherosclerotic properties of drugs and food components under human-like conditions. APOE*3-Leiden mice were fed a Western diet supplemented with either 5 or 50 g kg(-1) printanor. As a reference, the commercially available fermented garlic kyolic was included (1.6 g kg(-1) diet). Treatment with printanor demonstrated reduced body weight, coinciding with increased feces production and fecal fatty acids excretion. Printanor and kyolic treatment did not affect plasma lipids, markers of inflammation (serum amyloid A, serum-soluble intercellular adhesion molecule-1, and blood-leukocytes tumor necrosis factor-alpha (TNFalpha) production) and vascular activation (plasma von Willebrand factor (vWF)). As analyzed after 28 weeks of treatment, printanor and kyolic did not affect atherosclerotic lesion type, area or composition. Under conditions relevant to the human situation, the well-characterized and production-controlled garlic powder printanor does not display hypolipidemic, anti-inflammatory or anti-atherosclerotic properties.

Garlic is reported to have beneficial effects on risk factors associated with cardiovascular disease, including normalization of plasma lipid levels. However, numerous studies do not support this beneficial effect of garlic on plasma lipids. This contradiction may result from the use of different garlic-derived materials, experimental designs, and/or animal models. The present study investigated the hypolipidemic effect of garlic-derived materials in APOE*3-Leiden mice, a model well suited for drug and dietary intervention studies of hyperlipidemia. APOE*3-Leiden mice were fed a garlic-derived sulfur-rich compound, either allicin (0.29 g.L drinking water(-1)) or diallyldisulfide (0.27 g.kg diet(-1)), or powdered garlic, of either the kwai (42 g.kg diet(-1)) or morado (42 g.kg diet(-1)) variety. The amounts of garlic-derived materials supplied allowed free intake of allicin or allicin equivalents (diallyldisulfide, kwai, or morado) at 44 mg.kg body wt(-1).d(-1). Mice were fed a nonpurified diet for 4 wk, followed by a Western diet for 8 wk, both supplemented with the garlic-derived materials. These diets had no consistent effect on plasma lipids and did not affect lipoprotein profiles, which are markers for whole-body cholesterol synthesis and intestinal sterol absorption. The current data indicate that the postulated effects of garlic on cardiovascular disease are not caused via modulation of plasma lipid levels.

We describe 2 patients with glycogen storage disease type 1a and severe hyperlipidemia without premature atherosclerosis. Susceptibility of low-density lipoproteins to oxidation was decreased, possibly related to the ~40-fold increase in palmitate synthesis altering lipoprotein saturated fatty acid contents. These findings are potentially relevant for antihyperlipidemic treatment in patients with glycogen storage disease type 1a.

Dietary plant stanols lower serum cholesterol levels in humans and in hyperlipidemic rodents, mainly by inhibition of the intestinal cholesterol absorption. We used female apolipoprotein E*3-Leiden transgenic mice to investigate the consequences of this effect on serum lipid levels and hepatic lipid metabolism. Five groups of 6 or 7 mice received for 9 weeks a diet containing 0.25% cholesterol and 0.0%, 0.25%, 0.5%, 0.75%, or 1.0% (wt/wt) plant stanols (sitostanol 88% [wt/wt], campestanol 10% [wt/wt]) esterified to fatty acids. Compared with the control diet, plant stanol ester treatment dose-dependently reduced serum cholesterol levels by 10% to 33% (P<0.05), mainly in very low density lipoproteins (VLDLs), intermediate density lipoproteins, and low density lipoproteins. Furthermore, 1.0% of the dietary plant stanols significantly decreased the liver contents of cholesteryl esters (-62%), free cholesterol (-31%), and triglycerides (-38%) but did not change the hepatic VLDL-triglyceride and VLDL-apolipoprotein B production rates. However, plant stanol ester feeding significantly decreased the amounts of cholesteryl esters and free cholesterol incorporated in nascent VLDLs by 72% and 30%, respectively, resulting in a net 2-fold decreased VLDL cholesterol output. Liver mRNA levels of low density lipoprotein receptors, 3-hydroxy-3-methylglutaryl coenzyme A synthase, cholesterol 7alpha-hydroxylase, and sterol 27-hydroxylase were not changed by plant stanol ester feeding. Nevertheless, the serum lathosterol-to-cholesterol ratio was significantly increased by 23%, indicating that dietary plant stanol esters increased whole-body cholesterol synthesis. Plant stanol esters also significantly decreased the cholesterol saturation index in bile by 55%. In conclusion, in apolipoprotein E*3-Leiden transgenic mice, plant stanol ester feeding dose-dependently lowered serum cholesterol levels as a result of a reduced secretion of VLDL cholesterol. This was caused by a decreased hepatic cholesterol content that also resulted in a lowered biliary cholesterol output, indicative of a reduced lithogenicity of bile in these mice.

Although there is evidence that hyperlipidemia and predominance of small dense low density lipoproteins (LDLs) are associated with increased oxidative stress, the oxidation status in patients with hypertriglyceridemia (HTG) has not been studied in detail. Therefore, we studied urinary levels of F(2)-isoprostanes (8-isoprostaglandin F(2alpha) and 2,3-dinor-5,6-dihydro-8-isoprostaglandin F(2alpha)) and susceptibility of very low density lipoproteins (VLDLs) and LDLs to oxidation ex vivo in 18 patients with endogenous HTG and 20 matched control subjects. In addition, the effects of 6 weeks of bezafibrate therapy were assessed in a double-blind, placebo-controlled, crossover trial. Urinary levels of F(2)-isoprostanes were similar in the HTG and normolipidemic group. Bezafibrate caused an increase in 8-isoprostaglandin F(2alpha) (762+/-313 versus 552+/-245 ng/24 h for bezafibrate and placebo therapy, respectively; P=0.03), whereas 2,3-dinor-5, 6-dihydro-8-isoprostaglandin F(2alpha) levels tended to be increased (1714+/-761 versus 1475+/-606 ng/24 h for bezafibrate and placebo therapy, respectively; P=0.11). VLDLs and LDLs were more resistant to copper-induced oxidation in patients with HTG than in control subjects. Bezafibrate reversed the oxidation resistance to the normal range. In conclusion, these results indicate the following: (1) HTG is associated with normal in vivo oxidative stress and enhanced ex vivo resistance of lipoproteins to oxidation. (2) Bezafibrate reduces the resistance of lipoproteins to copper-induced oxidation and enhances oxidative stress in HTG patients.

Evidence has accumulated for oxidative modification of low-density lipoproteins (LDL) to play an important role in the atherogenic process. Therefore, we investigated the relation between susceptibility of LDL to oxidation and risk of peripheral atherosclerosis among 249 men between 45 and 80 years of age. The ankle-arm index was calculated for both legs as the ratio of systolic blood pressure in the leg divided by the arm systolic blood pressure. The lowest of both ankle-arm indices was used to categorize subjects. Thirty-nine men with an ankle-arm index < 1.00 (20% cut-off point of distribution) were classified as subjects with peripheral atherosclerosis. Subjects with peripheral atherosclerosis reported more often the use of a special diet and the use of antihypertensive medication, aspirin and coumarin derivatives. No significant differences in total, LDL and HDL cholesterol and triglycerides were present between groups. Resistance time and maximum rate of oxidation were measured ex vivo using copper-induced LDL oxidation. Subjects with peripheral atherosclerosis had a significantly lower resistance time, whereas the maximum rate of oxidation tended to be increased in subjects with peripheral atherosclerosis. Odds ratios (ORs, and 95% confidence interval) for the successive tertiles of resistance time were 1.00 (reference), 0.37 (0.15-0.89) and 0.37 (0.16-0.86) (p(trend) < 0.01). ORs for the successive tertiles of maximum rate of oxidation were 1.00 (reference), 1.34 (0.47-3.82) and 1.50 (0.55-4.15). This inverse association was borderline significant (p(trend) = 0.07). These results support an association between LDL oxidation and the development of peripheral atherosclerosis.

We investigated the effect of incorporating n-3 polyunsaturated fatty acids (PUFAs) into the diet on the lipid-class composition of LDLs, their size, and their susceptibility to oxidation. Forty-seven healthy volunteers incorporated 30 g sunflower-oil (SO) margarine/d into their habitual diet during a 3-wk run-in period and then used either SO or a fish-oil-enriched sunflower oil (FO) margarine for the following 4 wk. Plasma concentrations of total cholesterol, triacylglycerols, HDL cholesterol, LDL cholesterol, and apolipoproteins A-I and B did not differ significantly between the groups during intervention. The FO margarine increased the concentration of n-3 very-long-chain PUFAs in the LDL particles, showing 93% (P < or = 0.0001), 8% (P = 0.05), and 35% (P = < 0.0001) increases in eicosapentaenoic acid, docosapentaenoic acid, and docosahexaenoic acid, respectively, in the FO group compared with 3%, 7%, and 7%, respectively, in the SO group during the intervention. The cholesterol content of the LDL particles increased in the FO group [total cholesterol: 6% (P = 0.008); cholesterol ester: 12% (P = 0.014)], although it was not significantly different from that in the control group, whereas the other lipid classes and the size of the LDL particles remained unchanged in both groups. A reduction in the alpha-tocopherol content in LDL (6%, P = 0.005) was observed in the FO group. Ex vivo oxidation of LDL induced with Cu2+ showed a significantly reduced lag time (from 91 to 86 min, P = 0.003) and lower maximum rate of oxidation (from 10.5 to 10.2 nmol x mg(-1) x min(-1), P = 0.003) after intake of the FO margarine. The results indicate that consumption of the FO compared with the SO margarine had no effect on LDL size and lipid composition and led to minor changes in LDL a-tocopherol content and oxidation resistance.

Intake of flavonoids is associated with a reduced cardiovascular risk. Oxidation of LDL is a major step in atherogenesis, and antioxidants may protect LDL from oxidation. Because tea is an important source of flavonoids, which are strong antioxidants, we have assessed in a randomized, placebo-controlled study the effect of consumption of black and green tea and of intake of isolated green tea polyphenols on LDL oxidation ex vivo and on plasma levels of antioxidants and lipids. Healthy male and female smokers (aged 34+/-12 years, 13 to 16 per group) consumed during a 4-week period 6 cups (900 mL) of black or green tea or water per day, or they received as a supplement 3.6 grams of green tea polyphenols per day (equivalent to the consumption of 18 cups of green tea per day). Consumption of black or green tea had no effect on plasma cholesterol and triglycerides, HDL and LDL cholesterol, plasma vitamins C and E, beta-carotene, and uric acid. No differences were found in parameters of LDL oxidation. Intake of green tea polyphenols decreased plasma vitamin E significantly in that group compared with the control group (-11% P=.016) but had no effect on LDL oxidation ex vivo. We conclude that consumption of black or green tea (6 cups per day) has no effect on plasma lipids and no sparing effect on plasma antioxidant vitamins and that intake of a high dose of isolated green tea polyphenols decreases plasma vitamin E. Although tea polyphenols had a potent antioxidant activity on LDL oxidation in vitro, no effect was found on LDL oxidation ex vivo after consumption of green or black tea or intake of a green tea polyphenol isolate.

Accumulated evidence indicates that oxidative modification of LDL plays an important role in the atherogenic process. Therefore, we investigated the relation between coronary atherosclerosis and susceptibility of LDL to oxidation in a case-control study in men between 45 and 80 years of age. Case subjects and hospital control subjects were selected from subjects undergoing a first coronary angiography. Subjects with severe coronary stenosis (> or = 85% stenosis in one and > or = 50% stenosis in a second major coronary vessel) were classified as case subjects (n=91). Hospital control subjects with no or minor stenosis (< or = 50% stenosis in no more than two of the three major coronary vessels, n=94) and population control subjects free of plaques in the carotid artery (n=85) were pooled for the statistical analysis into one control category. Enrollment procedures allowed for similar distributions in age and smoking habits. Case subjects had higher levels of total and LDL cholesterol and triglycerides and lower levels of HDL cholesterol. Resistance time, maximum rate of oxidation, and maximum diene production were measured ex vivo using copper-induced LDL oxidation. A borderline significant inverse trend was observed for coronary atherosclerosis risk at increasing resistance time. Odds ratios (95% confidence interval) for the successive quartiles were 1.0 (reference), 0.77 (0.39 to 1.53), 0.67 (0.33 to 1.34), and 0.55 (0.27 to 1.15) (ptrend=0.07). No relation with maximum rate of oxidation was found, and higher maximum diene levels were found in control subjects (P<.01). The main determinant of oxidation was the fatty acid composition of LDL. No effect of smoking or use of medication was observed. We conclude that although LDL resistance to oxidation may be a factor in atherogenesis, the ex vivo measure is not a strong predictor of severity of coronary atherosclerosis.

Accumulating evidence indicates that oxidative modification of low-density lipoproteins is atherogenic and that antioxidants may play a role in protection of LDL against oxidation. Several studies have reported a seasonal fluctuation in antioxidant levels, but to date nothing is known about seasonal fluctuations in parameters of oxidizability. We collected blood from 10 volunteers at four different periods over one year (February, May, September and December), and measured the amount of plasma lipids, plasma antioxidants, lipid and fatty acid composition of the LDL particle, LDL antioxidant content, LDL particle size and oxidation parameters (lag time and propagation rate). No seasonal fluctuation for lag time and propagation rate of copper ion-induced LDL oxidation was found. Small seasonal fluctuations were observed for some determinants of LDL oxidation, e.g. plasma and LDL vitamin E and LDL particle size, and for plasma lipids, plasma and LDL lutein and LDL beta-carotene. Fatty acid composition of LDL did not change during the year. The main determinant of oxidation susceptibility was the fatty acid composition of LDL. We conclude that LDL oxidation parameters do not change over the year.

In type II diabetes mellitus the altered hormonal state after menopause may represent an additional cardiovascular risk factor. Estrogen replacement therapy (ERT) is associated with a decreased cardiovascular risk, at least in nondiabetic postmenopausal women. We studied the effect of ERT on plasma lipids and lipoproteins and on LDL oxidation in 40 postmenopausal women with type II diabetes but with minimal vascular complications in a randomized placebo-controlled trial. Twenty patients were treated orally with 2 mg/d micronized 17 beta-estradiol and 20 patients with placebo for 6 weeks. Plasma total cholesterol (-6%, P = .04), LDL cholesterol (-16%, P = .0001), and apoB (-11%, P = .001) levels decreased and HDL cholesterol (20%, P = .0001) and apoA-I (14%, P = .0001) levels increased after ERT compared with placebo. Glycated hemoglobin (HbA1c) decreased significantly after ERT (-3%, P = .03), the cholesterol content of the LDL particles decreased (-5%, P = .006), triglyceride content increased (16%, P = .01), and LDL particle size did not change significantly. ERT had no effect on parameters of LDL oxidation. We conclude that plasma levels of HDL cholesterol, apoA-I, LDL cholesterol, apoB, and glycated hemoglobin are improved in postmenopausal women with type II diabetes mellitus after treatment with 17 beta-estradiol, indicative of a better metabolic control, and that ERT has no effect on LDL oxidizability.

In hypertriglyceridemic (HTG) patients the addition of fish oil to the diet causes a marked reduction in the concentration of triglyceride-rich lipoproteins in the serum. To investigate the effects of fish oil on the oxidation resistance of VLDL and LDL in HTG patients, nine male patients received 1 g/d fish oil (containing 55.7% n-3 polyunsaturated fatty acids [PUFAS] and l U alpha-tocopherol/g fish oil) for 6 weeks followed by 5 g/d fish oil for an additional 6 weeks. Cu(2+)-induced oxidation of VLDL and LDL was measured by continuous monitoring of conjugated dienes. Supplementation with 1 g/d fish oil caused hardly any changes in the n-3 PUFA content of lipoproteins or lipoprotein concentrations in serum. However, supplementation with 5 g/d fish oil resulted in a significant increase of n-3 PUFA content in VLDL (from 2.5% to 6.4% of total fatty acids) and LDL (from 3.2% to 6.4% of total fatty acids), decreases in serum triglyceride, VLDL triglyceride, and VLDL cholesterol concentrations of 54%, 56%, and 40%, respectively, and an increase in LDL cholesterol of 23%. The lag times of VLDL and LDL oxidation decreased from 197 to 140 minutes (-29%) and 101 to 86 minutes (-15%), respectively. At the end of the 5 g/d fish oil supplementation the lag times of VLDL and LDL oxidation were correlated with their respective n-3 PUFA content (r = -.67; P < .05 and r = -.79; P < .02, respectively). Before and at the end of supplementation with 5 g/d fish oil the lag times and propagation rates of VLDL oxidation also correlated with the total number of double bonds in all PUFAs of VLDL. We conclude that fish oil supplementation strongly reduces serum concentrations of total triglycerides, VLDL triglycerides, and VLDL cholesterol. However, in HTG patients fish oil supplementation increased the serum LDL cholesterol concentration and the susceptibility of VLDL and LDL to oxidation.

There is accumulating evidence that oxidative modification of LDL is an important step in the process of atherogenesis and that antioxidants may protect LDL from oxidation. We and others have previously shown that ingestion of pharmacological doses of the antioxidant D,L-alpha-tocopherol (vitamin E), far above the recommended daily intake (ie, 12 to 15 IU/d for adults), increases the oxidation resistance of LDL. In this study, we ascertained the minimal supplementary dose of vitamin E necessary to protect LDL against oxidation in vitro. Twenty healthy volunteers (10 men and 10 women, aged 21 to 31 years) ingested consecutively 25, 50, 100, 200, 400, and 800 IU/d, D,L-alpha-tocopherol acetate during six 2-week periods. No changes were observed in LDL triglyceride content, fatty acid composition of LDL, or LDL size during the intervention. Concentrations of alpha-tocopherol in plasma and LDL were both 1.2 times the baseline values after the first period (25 IU/d) and 2.6 and 2.2 times, respectively, after the last period (800 IU/d). There was a linear increase in LDL alpha-tocopherol levels up to an intake of 800 IU/d (r = .79, P < .0001) and a good correlation between alpha-tocopherol in plasma and LDL (r = .66, P < .0001). Simultaneously, the resistance of LDL to oxidation was elevated dose-dependently (+28% after the last period) and differed significantly from the baseline resistance time even after ingestion of only 25 IU/d.(ABSTRACT TRUNCATED AT 250 WORDS)