Dietary sugar kelp (Saccharina latissima) consumption did not attenuate atherosclerosis in low-density lipoprotein receptor knockout mice.

We previously demonstrated the beneficial effects of U.S.-grown sugar kelp (Saccharina latissima), a brown seaweed, on reducing serum triglycerides (TG) and total cholesterol (TC) and protecting against inflammation and fibrosis in the adipose tissue of diet-induced obesity mice. In this current study, we aimed to explore whether the dietary consumption of sugar kelp can prevent atherosclerosis using low-density lipoprotein receptor knockout (Ldlr KO) mice fed an atherogenic diet. Eight-week-old male Ldlr KO mice were fed either an atherogenic high-fat/high-cholesterol control (HF/HC) diet or a HF/HC diet supplemented with 6% (w/w) sugar kelp (HF/HC-SK) for 16 weeks. Consumption of sugar kelp significantly increased the body weight gain without altering fat mass and lean mass. Also, there were no significant differences in energy expenditure and physical activities between the groups. The two groups did not show significant differences in serum and hepatic TG and TC levels or the hepatic expression of genes involved in cholesterol and lipid metabolism. Although serum alanine aminotransferase (ALT) activity did not differ significantly between the two groups, there were significant increases in the expression of macrophage markers, including adhesion G protein-coupled receptor E1 and cluster of differentiation 68, as well as tumor necrosis factor alpha in the HF/HC-SK group compared to the HF/HC mice. The consumption of sugar kelp did not elicit a significant effect on the development of aortic lesions. Moreover, lipopolysaccharide-stimulated splenocytes isolated from HF/HC-SK-fed mice showed no significant changes in the mRNA levels of pro-inflammatory genes compared with those from the HF/HC mice. In summary, the consumption of dietary sugar kelp did not elicit anti-atherogenic and hepatoprotective effects in Ldlr KO mice.

Potential Antiviral Effect of Korean Forest Wild Mushrooms against Feline Coronavirus (FCoV).

Coronaviruses (CoV) are among the major viruses that cause common cold in humans. Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is a high-risk human pathogen that derived from bat coronaviruses, although several other animals serve as CoV hosts, contributing to human infection. As the human activity area expanded, viruses previously prevalent only in animals mutated and became threats to humans as well, leading to worldwide epidemics. Therefore, controlling CoV infections in animals is essential to prevent CoV-related human infections. Feline coronavirus (FCoV) could be reportedly used as an alternative model for SARS-CoV-2. Traditionally, mushrooms are not only foods but are also consumed to prevent diseases. Importantly, certain edible and medicinal mushrooms display antibacterial and antiviral effects against respiratory pathogens; therefore, they could be tested as potential coronavirus treatment agents. In this study, we investigated if wild forest mushrooms with various reported physiological activities could exhibit an antiviral activity against CoV, using FCoV as a SARS-CoV-2 model infecting Crandell Rees feline kidney cells. We measured the antiviral activity of 11 wild mushrooms overall and our results demonstrated that Pleurotus ostreatus and Phallus luteus displayed the highest antiviral efficacy of 55.33%, followed by Tricholoma bakamatsutake at 43.77%. Grifola frondosa, Morchella esculenta, and Sarcodon imbricatus exhibited mild efficacy of 29.21%. We also tested Amanita caesareoides, Marasmius siccus, Pachyma hoelen, Phallus rubrovolvata, and Sparassis latifolia but could not detect any antiviral activity in their case. Our study confirms that wild forest mushrooms could be used as potential functional foods or pharmacological materials against coronavirus.

Roles of Histone Deacetylase 4 in the Inflammatory and Metabolic Processes.

Histone deacetylase 4 (HDAC4), a class IIa HDAC, has gained attention as a potential therapeutic target in treating inflammatory and metabolic processes based on its essential role in various biological pathways by deacetylating non-histone proteins, including transcription factors. The activity of HDAC4 is regulated at the transcriptional, post-transcriptional, and post-translational levels. The functions of HDAC4 are tissue-dependent in response to endogenous and exogenous factors and their substrates. In particular, the association of HDAC4 with non-histone targets, including transcription factors, such as myocyte enhancer factor 2, hypoxia-inducible factor, signal transducer and activator of transcription 1, and forkhead box proteins, play a crucial role in regulating inflammatory and metabolic processes. This review summarizes the regulatory modes of HDAC4 activity and its functions in inflammation, insulin signaling and glucose metabolism, and cardiac muscle development.

Multivariate Analysis among Marker Compounds, Environmental Factors, and Fruit Quality of Schisandra chinensis at Different Locations in South Korea.

This study aimed to investigate the correlation among the contents of marker compounds, growth characteristics, and environmental factors of Schisandra chinensis fruits across South Korea. The fruits were collected from 36 cultivation sites in 28 regions across the country. We investigated nine growth characteristics, twelve soil physicochemical properties, eight meteorological data, and three marker compounds in this study. We optimized and validated an optimized method for quantifying marker compounds using UPLC and performed correlation analysis among the contents of marker compounds, growth characteristics, and environmental factors. The UPLC-UV method for analyzing marker compounds was validated by measuring linearity, LOD, LOQ, precision, and accuracy. The marker compounds were negatively correlated with the fruit size and sugar contents, and growth characteristics were negatively correlated with some physicochemical properties of the soil. The results of this study can be used as basic data for the standard cultural practices and quality control of S. chinensis fruits.

Consumption of Low Dose Fucoxanthin Does Not Prevent Hepatic and Adipose Inflammation and Fibrosis in Mouse Models of Diet-Induced Obesity.

Fucoxanthin (FCX) is a xanthophyll carotenoid present in brown seaweed. The goal of this study was to examine whether FCX supplementation could attenuate obesity-associated metabolic abnormalities, fibrosis, and inflammation in two diet-induced obesity (DIO) mouse models. C57BL/6J mice were fed either a high-fat/high-sucrose/high-cholesterol (HFC) diet or a high-fat/high-sucrose (HFS) diet. The former induces more severe liver injury than the latter model. In the first study, male C57BL/6J mice were fed an HFC diet, or an HFC diet containing 0.015% or 0.03% (w/w) FCX powder for 12 weeks to develop obesity-induced nonalcoholic steatohepatitis (NASH). In the second study, mice were fed an HFS diet or an HFS diet containing 0.01% FCX powder for 8 weeks. FCX did not change body weight gain and serum lipid profiles compared to the HFC or HFS controls. No significant differences were present in liver triglyceride and total cholesterol, hepatic fat accumulation, and serum alanine aminotransferase levels between control and FCX-fed mice regardless of whether they were on an HFC or HFS diet. FCX did not mitigate mRNA abundance of genes involved in lipid synthesis, cholesterol metabolism, inflammation, and fibrosis in the liver and white adipose tissue, while hepatic fatty acid ß-oxidation genes were significantly elevated by FCX in both HFC and HFS feeding studies. Additionally, in the soleus muscle, FCX supplementation significantly elevated genes that regulate mitochondrial biogenesis and fatty acid ß-oxidation, concomitantly increasing mitochondrial DNA copy number, compared with HFC. In summary, FCX supplementation had minor effects on hepatic and white adipose inflammation and fibrosis in two different DIO mouse models.

Nicotinamide riboside supplementation exerts an anti-obesity effect and prevents inflammation and fibrosis in white adipose tissue of female diet-induced obesity mice.

Nicotinamide riboside (NR) is a nicotinamide adenine dinucleotide (NAD+) precursor. We previously reported that NR supplementation prevented the development of liver fibrosis in male mice. However, whether NR exerts a similar effect in females is unknown. Therefore, we determined whether NR supplementation can prevent obesity-induced inflammation and fibrosis in the liver and white adipose tissue (WAT) by providing NAD+ in obese female mice. Female C57BL/6J mice at the age of 8 weeks (young) and 16 weeks (old) were fed a high-fat/high-sucrose/high-cholesterol diet (HF) or HF diet supplemented with NR at 400 mg/kg/d for 20 weeks. While NR had minor effects in young female mice, it significantly reduced body weight gain, fat mass, glucose intolerance, and serum cholesterol levels compared to the HF group in old females. Hepatic NAD+ level tended toward an increase in the NR group (P=.054), but NR did not attenuate serum alanine aminotransferase levels, steatosis, and liver fibrosis in old female mice. However, NR decreased weight and adipocyte size in gonadal WAT (gWAT) of old females. NR also reduced the number of crown-like structures and the expression of inflammatory genes, along with decreases in fibrogenic gene expression and collagen accumulation in gWAT compared with the HF group. Also, old mice fed NR showed increased metabolic rates, physical activity, and energy expenditure compared with the HF. Thus, our results indicated that NR supplementation exerted an anti-obesity effect and prevented the development of inflammation and fibrosis in the WAT of old, but not young, female mice with diet-induced obesity.

MicroRNA-200c coordinates HNF1 homeobox B and apolipoprotein O functions to modulate lipid homeostasis in alcoholic fatty liver disease.

Hepatic steatosis is an initial manifestation of alcoholic liver disease. An imbalance of hepatic lipid processes including fatty acid uptake, esterification, oxidation, and triglyceride secretion leads to alcoholic fatty liver (AFL). However, the precise molecular mechanisms underlying the pathogenesis of AFL remain elusive. Here, we show that mice deficient in microRNAs (miRs)-141 and -200c display resistance to the development of AFL. We found that miR-200c directly targets HNF1 homeobox B (Hnf1b), a transcriptional activator for microsomal triglyceride transfer protein (Mttp), as well as apolipoprotein O (ApoO), an integral component of the mitochondrial contact site and cristae organizing system complex. We show that expression of these miRs is significantly induced by chronic ethanol exposure, which is accompanied by reduced HNF1B and APOO levels. Furthermore, miR-141/200c deficiency normalizes ethanol-mediated impairment of triglyceride secretion, which can be attributed to the restored levels of HNF1B and MTTP, as well as phosphatidylcholine abundance. Moreover, we demonstrate that miR-141/200c deficiency restores ethanol-mediated inhibition of APOO expression and mitochondrial dysfunction, improving mitochondrial antioxidant defense capacity and fatty acid oxidation. Taken together, these results suggest that miR-200c contributes to the modulation of lipid homeostasis in AFL disease by cooperatively regulating Hnf1b and ApoO functions.

A mouse model of the regression of alcoholic hepatitis: Monitoring the regression of hepatic steatosis, inflammation, oxidative stress, and NAD+ metabolism upon alcohol withdrawal.

This study aimed to develop a well-characterized mouse model of alcoholic hepatitis (AH) regression. Male C57BL/6J mice were fed a Lieber-DeCarli (LD) control diet or LD containing 5% ethanol for ten days followed by one binge, which is the chronic-binge model of AH developed by the National Institute on Alcohol Abuse and Alcoholism. To determine AH regression, mice previously exposed to ethanol were put on LD control diet and metabolic and inflammatory features were monitored weekly for three weeks. Serum alcohol, total cholesterol, and alanine transaminase levels were increased in ethanol-fed mice, which declined to those of no ethanol controls within one and three weeks after ethanol withdrawal, respectively. Serum malondialdehyde was increased with ethanol feeding, but it was restored to no ethanol control levels within one week. Ethanol-induced changes in the hepatic expression of genes involved in lipogenesis, fatty acid oxidation, ethanol metabolism, and antioxidant response were restored to those of no ethanol controls after 3 weeks of ethanol withdrawal. Also, ethanol-induced hepatic inflammation was gradually decreased during the 3 weeks of ethanol withdrawal. Hepatic nicotinamide adenine dinucleotide (NAD+) levels and the expression of enzymes involved in the NAD+ salvage pathway were decreased by ethanol feeding, which was mitigated after ethanol withdrawal. Ethanol significantly lowered hepatic sirtuin 1 expression, but its levels were restored with ethanol cessation. This study established a mouse model of AH regression, which can be used as a preclinical model to study the potential of dietary bioactives or therapeutic agents on AH regression.

The loss of histone deacetylase 4 in macrophages exacerbates hepatic and adipose tissue inflammation in male but not in female mice with diet-induced non-alcoholic steatohepatitis.

Epigenetic regulation in macrophages plays a crucial role in the inflammatory response of cells. We investigated the role of macrophage histone deacetylase 4 (HDAC4) in diet-induced obesity and non-alcoholic steatohepatitis using macrophage-specific Hdac4 knockout mice (Hdac4MKO ). Hdac4 floxed control (Hdac4fl/fl ) and Hdac4MKO mice were fed a regular chow diet or an obesogenic high-fat/high-sucrose/high-cholesterol (HF/HS/HC) diet for 12 weeks. The loss of macrophage Hdac4, compared with Hdac4fl/fl control, aggravated the diet-induced inflammation in the liver and white adipose tissue only in male mice. Splenic monocytes isolated from male mice fed the HF/HS/HC diet showed increased lipopolysaccharide (LPS) sensitivity and decreased Ly6C-/Ly6C+ ratios in male Hdac4MKO mice, but not in females. Bone marrow-derived macrophages (BMMs) from male Hdac4MKO mice had a lesser efferocytotic capacity but higher proinflammatory gene expression upon LPS stimulation than male Hdac4fl/fl mice. However, female Hdac4MKO BMMs exhibited the opposite responses. The induction of estrogen receptor α (ERα, Esr1) expression by LPS was less in male but more in female Hdac4MKO BMMs than Hdac4fl/fl BMMs. Moreover, overexpression of human HDAC4 decreased basal expression of Esr1 and abolished its induction by LPS. Inhibition of ERα increased Hdac4 with induction of inflammatory genes, whereas activation of ERα decreased Hdac4 with reduction of inflammatory genes in male and female Hdac4fl/fl BMMs treated with LPS. However, regardless of the inhibition or activation of ERα, proinflammatory genes were induced by LPS more in male Hdac4MKO BMMs than Hdac4fl/fl cells, whereas cells in females showed opposite responses. In conclusion, this study suggests that the lack of macrophage Hdac4 aggravates hepatic and white adipose inflammation in male mice with diet-induced obesity and non-alcoholic steatohepatitis, and not in female mice. HDAC4 and ERα appear to counteract each other, but ERα may not be a major player in sex-dependent inflammatory responses in macrophages deficient in HDAC4. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Sugar kelp (Saccharina latissima) inhibits hepatic inflammation and fibrosis in a mouse model of diet-induced nonalcoholic steatohepatitis.

Nonalcoholic steatohepatitis (NASH), closely associated with obesity, is a health concern worldwide. We investigated whether the consumption of U.S.-grown sugar kelp (Saccharina latissima), an edible brown alga, can prevent obesity-associated metabolic disturbances and NASH in a mouse model of diet-induced NASH. Male C57BL/6J mice were fed a low-fat diet, a high-fat/high-sucrose/high-cholesterol diet (HF), or a HF diet containing sugar kelp (HF-Kelp) for 14 weeks. HF-Kelp group showed lower body weight with increased O2 consumption, CO2 production, physical activity, and energy expenditure compared with the HF. In the liver, there were significant decreases in weight, triglycerides, total cholesterol, and steatosis with HF-Kelp. The HF-Kelp group decreased hepatic expression of a macrophage marker adhesion G protein-coupled receptor E1 (Adgre1) and an M1 macrophage marker integrin alpha x (Itgax). HF-Kelp group also exhibited decreased liver fibrosis, as evidenced by less expression of fibrogenic genes and collagen accumulation than those of HF group. In epididymal white adipose tissue (eWAT), HF-Kelp group exhibited decreases in eWAT weight and adipocyte size compared with those of the HF. HF-Kelp group showed decreased expression of collagen type VI alpha 1 chain, Adgre1, Itgax, and tumor necrosis factor α in eWAT. We demonstrated, for the first time, that the consumption of U.S-grown sugar kelp prevented the development of obesity and its associated metabolic disturbances, steatosis, inflammation, and fibrosis in the liver and eWAT of a diet-induced NASH mouse model.

Nicotinamide riboside, an NAD+ precursor, attenuates inflammation and oxidative stress by activating sirtuin 1 in alcohol-stimulated macrophages.

Macrophages play an essential role in alcohol-induced inflammation and oxidative stress. We investigated the effects of nicotinamide riboside (NR), a natural nicotinamide adenine dinucleotide (NAD+) precursor, on alcohol-induced inflammation and oxidative stress in macrophages. NR significantly decreased ethanol-induced inflammatory gene expression, with a concomitant decrease in nuclear translocation of nuclear factor κB p65 in RAW 264.7 macrophages and mouse bone marrow-derived macrophages (BMDMs). In macrophages incubated with ethanol or acetaldehyde, NR abolished the accumulation of cellular reactive oxygen species. Ethanol decreased sirtuin 1 (SIRT1) expression and activity, and cellular NAD+ level while inducing pro-inflammatory gene expression. However, NR markedly attenuated the changes. SIRT1 inhibition augmented ethanol-induced inflammatory gene expression, but its activation elicited opposing effects. Also, ethanol did not alter glycolysis but increased glycolytic capacity, glycolytic reserve, and non-glycolytic acidification, with concomitant increases in hypoxia-induced factor 1α expression and activity, phosphorylation of pyruvate dehydrogenase, and extracellular lactate levels. Interestingly, ethanol increased mitochondrial respiration and ATP production but decreased maximal respiration and spare respiration capacity. The latter was linked to decreases in mitochondrial copy numbers. NR abolished the ethanol-induced metabolic changes in the glycolytic and oxidative phosphorylation pathways in RAW 264.7 macrophages. In conclusion, NR exerts anti-inflammatory and antioxidant properties by abrogating the inhibitory effects of ethanol on the SIRT1 pathway by increasing Sirt1 expression and its activator, NAD+. Also, SIRT1 activation and normalization of ethanol-induced changes in NAD+/NADH ratios by NR are likely crucial to counteract the changes in energy phenotypes of macrophages exposed to ethanol.

Anthocyanin-Rich Aronia Berry Extract Mitigates High-Fat and High-Sucrose Diet-Induced Adipose Tissue Inflammation by Inhibiting Nuclear Factor-κB Activation.

Obesity-induced inflammation in adipose tissue (AT) promotes the development of metabolic dysregulations by increasing macrophage recruitment in the stromal vascular fraction (SVF). The activation of nuclear factor-κB (NF-κB) signaling in macrophages serves as a pivotal mediator of AT inflammatory responses by increasing the expression of proinflammatory genes in obesity. Given the purported anti-inflammatory effects of berry consumption in humans, we evaluated if anthocyanin-rich aronia berry extract (ARN) can prevent obesity-induced AT inflammation in vivo. We also examined whether ARN suppresses lipopolysaccharide (LPS)-induced NF-κB activation in RAW 264.7 macrophages and mouse bone marrow-derived macrophages (BMDMs). Male C57BL/6J mice were fed a low-fat diet, a high-fat (HF), and high-sucrose (HS) diet or HF/HS diet supplemented with 0.2% ARN (HF/HS + ARN) for 14 weeks. Compared to HF-/HS-fed mice, ARN supplementation tended to decrease fasting serum glucose (P = .07). Furthermore, ARN supplementation significantly inhibited the phosphorylation of NF-κB p65 in epididymal AT with a concomitant decrease in the expression of Cd11b and Tnfα mRNAs in epididymal SVF isolated, compared with those from HF-/HS-fed mice. Consistent with these in vivo findings, ARN treatment significantly decreased the phosphorylation of p65 in LPS-stimulated RAW 264.7 macrophages and BMDMs. Moreover, ARN suppressed LPS-induced mRNA expression of inflammation mediators (iNos, Cox-2, Tnfα, Mcp-1, and Il-6) and glycolysis markers (Glut1, G6pdh, and Hk1) in both cell types. Taken together, our in vivo and in vitro results suggest that ARN supplementation may attenuate obesity-induced AT inflammation by inhibiting NF-κB signaling and glycolytic pathway in macrophages.

Fucoxanthin inhibits lipopolysaccharide-induced inflammation and oxidative stress by activating nuclear factor E2-related factor 2 via the phosphatidylinositol 3-kinase/AKT pathway in macrophages.

PURPOSE: Anti-inflammatory and antioxidant effects of fucoxanthin (FCX), a xanthophyll carotenoid, have been suggested. However, underlying mechanisms are elusive. The objective of this study was to elucidate the mechanisms by which FCX and its metabolites inhibit lipopolysaccharide (LPS)-induced inflammation and oxidative stress in macrophages. METHODS: The effects of the FCX on mRNA and protein expression of pro-inflammatory cytokines and antioxidant genes, and reactive oxygen species (ROS) accumulation were determined in RAW 264.7 macrophages. A potential role of FCX in the modulation of phosphatidylinositol 3-kinase (PI3K)/AKT/nuclear E2-related factor 2 (NRF2) axis was evaluated. RESULTS: FCX significantly decreased LPS-induced interleukin (Il)6, Il1b, and tumor necrosis factor α (Tnf) mRNA abundance and TNFα secretion. FCX attenuated LPS or tert-butyl-hydroperoxide-induced ROS accumulation with concomitant increases in the expression of antioxidant enzymes. Also, trolox equivalent antioxidant capacity assay demonstrated that FCX had a potent free radical scavenging property. FCX markedly increased nuclear translocation of NRF2 in LPS-treated macrophages, consequently inducing its target gene expression. Interestingly, the effect of FCX on NRF2 nuclear translocation was noticeably diminished by LY294002, an inhibitor of PI3K, but not by inhibitors of mitogen-activated protein kinases. Phosphorylation of AKT, a downstream element of PI3K, was also markedly increased by FCX. FCX metabolites, such as fucoxanthinol and amarouciaxanthin A, significantly attenuated LPS-induced ROS accumulation and pro-inflammatory cytokine expression. CONCLUSION: FCX exerts anti-inflammatory and antioxidant effects by the activation of NRF2 in the macrophages activated by LPS, which is mediated, at least in part, through the PI3K/AKT pathway.

Diterpenoids and phenolic analogues from the roots of Aralia continentalis.

Two new compounds, including a nor-pimarane diterpenoid (continentanol, 1) and a phenolic derivative (aralianic acid, 2), along with the known diterpenoids (3-11), polyacetylenes (12-15), phenolic components (16-28), and phytosterols (29 and 30), were isolated from roots of Aralia continentalis. The structures of the new compounds were established by spectroscopic data interpretation, particularly HRESIMS, 1 D and 2 D NMR data including HSQC and HMBC. Also, those of the known compounds were identified by spectral comparison with those of the reported values.[Formula: see text].

Inhibition of alcohol-induced inflammation and oxidative stress by astaxanthin is mediated by its opposite actions in the regulation of sirtuin 1 and histone deacetylase 4 in macrophages.

We previously demonstrated that astaxanthin (ASTX), a xanthophyll carotenoid, repressed ethanol-induced inflammation and oxidative stress in macrophages. We explored the role of sirtuin 1 (SIRT1) and histone deacetylase 4 (HDAC4) in the inhibitory effect of ASTX on inflammation and oxidative stress in macrophages exposed to ethanol. Ethanol decreased mRNA and protein of SIRT1 while increasing those of HDAC4, which was attenuated by ASTX in RAW 264.7 macrophages and mouse bone marrow-derived macrophages (BMDMs). Inhibition of SIRT1 expression or activity augmented ethanol-induced Hdac4 expression, but SIRT1 activation elicited the opposite effect. Consistently, Hdac4 knockdown increased Sirt1 expression with decreases in ethanol-induced inflammatory gene expression, but its overexpression resulted in the opposite effects. Furthermore, BMDMs from mice with macrophage specific-deletion of Hdac4 (Hdac4MKO) showed significant decreases in ethanol-induced inflammatory genes and ROS accumulation but an increase in Sirt1 expression. Macrophage specific deletion of Hdac4 or ASTX abolished the changes in genes for mitochondrial biogenesis and glycolysis by ethanol. Ethanol increased mitochondrial respiration, ATP production, and proton leak, but decreased maximal respiration and spare respiratory capacity, all of which were abolished by ASTX in RAW 264.7 macrophages. The ethanol-induced alterations in mitochondrial respiration were abrogated in Hdac4MKO BMDMs. In conclusion, the anti-inflammatory and antioxidant properties of ASTX in ethanol-treated macrophages may be mediated, at least partly, by its opposite effect on SIRT1 and HDAC4 to empower SIRT1 to counteract ethanol-induced activation of HDAC4.

The effect of cranberry consumption on lipid metabolism and inflammation in human apo A-I transgenic mice fed a high-fat and high-cholesterol diet.

Lipid metabolism and inflammation contribute to CVD development. This study investigated whether the consumption of cranberries (CR; Vaccinium macrocarpon) can alter HDL metabolism and prevent inflammation in mice expressing human apo A-I transgene (hApoAITg), which have similar HDL profiles to those of humans. Male hApoAITg mice were fed a modified American Institute of Nutrition-93M high-fat/high-cholesterol diet (16 % fat, 0·25 % cholesterol, w/w; n 15) or the high-fat/high-cholesterol diet containing CR (5 % dried CR powder, w/w, n 16) for 8 weeks. There were no significant differences in body weight between the groups. Serum total cholesterol, non-HDL-cholesterol and TAG concentrations were significantly lower in the control than CR group with no significant differences in serum HDL-cholesterol and apoA-I. Mice fed CR showed significantly lower serum lecithin-cholesterol acyltransferase activity than the control. Liver weight and steatosis were not significantly different between the groups, but hepatic expression of genes involved in cholesterol metabolism was significantly lower in the CR group. In the epididymal white adipose tissue (eWAT), the CR group showed higher weights with decreased expression of genes for lipogenesis and fatty acid oxidation. The mRNA abundance of F4/80, a macrophage marker and the numbers of crown-like structures were less in the CR group. In the soleus muscle, the CR group also demonstrated higher expression of genes for fatty acid ß-oxidation and mitochondrial biogenesis than those of the control. In conclusion, although CR consumption elicited minor effects on HDL metabolism, it prevented obesity-induced inflammation in eWAT with concomitant alterations in soleus muscle energy metabolism.

Comprehensive characterization of metabolic, inflammatory and fibrotic changes in a mouse model of diet-derived nonalcoholic steatohepatitis.

The objective of this study was to develop a well-characterized mouse model of nonalcoholic steatohepatitis (NASH) with a strong manifestation of liver fibrosis. The progression of metabolic, inflammatory and fibrotic features of this mouse model was monitored by performing in vivo time-course study. Male C57BL/6J mice were fed a high-fat/high-sucrose/high-cholesterol diet (34% fat, 34% sucrose and 2.0% cholesterol, by weight) for 2, 4, 6, 8, 10, 12, 14 or 16 weeks to induce obesity-associated metabolic dysfunctions, inflammation and fibrosis in the liver and white adipose tissue (WAT). Body and liver weights were gradually increased with significant hepatic triglyceride accumulation, i.e., liver steatosis, and marked elevation of serum alanine transaminase levels at week 10. While hepatic inflammation was displayed with the highest expression of macrophage markers and M1 markers at week 6, liver fibrosis determined by collagen accumulation was continuously increased to week 16. In epididymal WAT, weights and adipocyte size peaked at week 6-8. The increased expression of fibrogenic genes preceded inflammatory features (week 2 to 6 vs. week 6 to 16), suggesting that early fibrosis may trigger inflammatory events in the WAT. This study established a mouse model of diet-induced NASH with a strong manifestation of liver fibrosis. This mouse model will be a valuable in vivo tool in studying the pathophysiology of NASH and also in testing preventive and therapeutic potentials of dietary components and drugs against NASH with liver fibrosis.

n-3 PUFAs inhibit TGFß1-induced profibrogenic gene expression by ameliorating the repression of PPARγ in hepatic stellate cells.

Activated hepatic stellate cells (HSCs) are primarily responsible for the accumulation of extracellular matrix substances during the development of liver fibrosis. It has been shown that n-3 polyunsaturated fatty acids (PUFAs) can prevent liver fibrosis development. However, the underlying mechanisms of action need further investigation. The objective of this study was to determine the regulatory roles of fatty acids (FAs) on the expression of profibrogenic genes in HSCs with the elucidation of mechanisms. LX-2 cells and primary human and mouse HSCs were treated with palmitic acid, oleic acid, linoleic acid, α-linolenic acid, eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) to determine their effect on profibrogenic gene expression upon the activation by transforming growth factor ß1 (TGFß1). PUFAs significantly suppressed TGFß1-induced expression of profibrogenic genes in LX-2 and primary human HSCs with n-3 being more potent than n-6 PUFAs. However, PUFAs did not inhibit the phosphorylation and nuclear translocation of SMA- and MAD-related protein in primary human HSCs. Furthermore, PUFAs did not alter the profibrogenic gene expression in primary mouse HSCs. The inhibitory effect of EPA and DHA on TGFß1-induced profibrogenic gene expression was diminished by peroxisome proliferator-activated receptor gamma (PPARG) knockdown, although chemical inhibition of PPARγ did not elicit a similar result. The results suggest that n-3 PUFAs possess the most potent protective effects against TGFß1-induced profibrogenic gene expression, which is, at least in part, PPARγ-dependent in HSCs.

Astaxanthin inhibits alcohol-induced inflammation and oxidative stress in macrophages in a sirtuin 1-dependent manner.

OBJECTIVES: Alcohol induces inflammation and oxidative stress, causing cell damages. We previously demonstrated that astaxanthin (ASTX), a xanthophyll carotenoid, exerts anti-inflammatory and antioxidant properties in macrophages exposed to inflammatory insults. In this study, we investigated whether ASTX can inhibit alcohol-induced inflammation and oxidative stress in macrophages with the elucidation of mechanisms. METHODS: RAW 264.7 macrophages and mouse bone marrow-derived macrophages were treated with 80 mM ethanol in the presence or absence of 25 µM of ASTX for 72 h. Subsequently, the expression of genes related to inflammation and oxidative stress, cellular reactive oxygen species accumulation, cellular NAD+ level and sirtuin 1 (SIRT1) activity were measured. In addition, RAW 264.7 macrophages were treated with sirtinol or resveratrol, which are known inhibitors or activators of SIRT1 activity, respectively, to determine the contribution of SIRT1 to the inhibitory effect of ASTX on inflammation and oxidative stress in macrophages exposed to ethanol. RESULTS: Ethanol increased mRNA expression of interleukin (Il)-6, Il-1b and tumor necrosis factor α with a concomitant increase in nuclear translocation of nuclear factor κB, which was abolished by ASTX. Importantly, ethanol significantly decreased SIRT1 activity and cellular NAD+ level, but ASTX markedly attenuated the decreases in RAW 264.7 macrophages. Sirtinol increased the expression of proinflammatory genes in ethanol-induced RAW 264.7 macrophages. In contrast, resveratrol decreased proinflammatory gene expression. CONCLUSIONS: ASTX showed anti-inflammatory and antioxidant properties by inhibiting decreases in SIRT1 expression and cellular NAD+ level in ethanol-treated macrophages. Therefore, ASTX may be used for the prevention of alcohol-induced cell damages.