Effect of combined use of a low-carbohydrate, high-protein diet with omega-3 polyunsaturated fatty acid supplementation on glycemic control in newly diagnosed type 2 diabetes: a randomized, double-blind, parallel-controlled trial.

Background: The combined effect of a low-carbohydrate, high-protein (LCHP) diet and omega-3 (n-3) polyunsaturated fatty acid (PUFA) supplementation on patients with type 2 diabetes (T2D) is not known. Objective: The aim of this study was to evaluate the effect of an LCHP diet combined with ω-3 (LCHP+ω-3) on glycemic control in patients with T2D. Design: In this randomized, double-blind, parallel-controlled trial, 122 newly diagnosed participants with T2D were randomly assigned to receive a high-carbohydrate, low-protein diet with low ω-3 PUFAs [control (CON)], an LCHP, ω-3, or LCHP+ω-3 diet for 12 wk. The ratio of carbohydrate to protein was 42:28 in the LCHP and LCHP+ω-3 diet and 54:17 in the CON and ω-3 diet. The participants were given 6 g fish oil/d (containing 3.65 g docosahexaenoic acid, eicosapentaenoic acid, and docosapentaenoic acid/d) in the ω-3 and LCHP+ω-3 diet groups or 6 g corn oil/d (placebo) in the CON and LCHP diet groups. Results: Compared with the CON diet group, greater decreases in glycated hemoglobin (HbA1c) and fasting glucose were observed in all of the other 3 diet groups at 12 wk. Of note, HbA1c reduction in the LCHP+ω-3 diet group (-0.51%; 95% CI: -0.64%, -0.37%) was greater than that in the LCHP (P = 0.03) and ω-3 (P = 0.01) diet groups at 12 wk. In terms of fasting glucose, only the LCHP+ω-3 diet group showed a significant decrease at 4 wk (P = 0.03 compared with CON). Moreover, the reduction in fasting glucose in the LCHP+ω-3 diet group (-1.32 mmol/L; 95% CI: -1.72, -0.93 mmol/L) was greater than that in the LCHP (P = 0.04) and ω-3 (P = 0.03) diet groups at 12 wk. Conclusions: The LCHP+ω-3 diet provided greater effects on HbA1c and fasting glucose and faster effects on fasting glucose than both the LCHP and ω-3 diets, indicating the potential necessity of combining an LCHP diet with ω-3 PUFAs in T2D control. This trial was registered at chictr.org.cn/ as ChiCTR-TRC-14004704.

Dietary Factors Modulate Colonic Tumorigenesis Through the Interaction of Gut Microbiota and Host Chloride Channels.

SCOPE: In recent decades, the association among diet, gut microbiota, and the risk of colorectal cancer (CRC) has been established. Gut microbiota and associated metabolites, such as bile acids and butyrate, are now known to play a key role in CRC development. The aim of this study is to identify that the progression to CRC is influenced by cholic acid, sodium butyrate, a high-fat diet, or different dose of dihydromyricetin (DMY) interacted with gut microbiota. METHODS AND RESULTS: An AOM/DSS (azoxymethan/dextran sodium sulfate) model is established to study the gut microbiota compsition before and after tumor formation during colitis-induced tumorigenesis. All above dietary factors profoundly influence the composition of gut microbiota and host colonic tumorigenesis. In addition, mice with DMY-modified initial microbiota display different degrees of chemically induced tumorigenesis. Mechanism analysis reveals that gut microbiota-associated chloride channels participated in colon tumorigenesis. CONCLUSION: Gut microbiota changes occur in the hyperproliferative stage before tumor formation. Gut microbiota and host chloride channels, both of which are regulated by dietary factors, are associated with CRC development.

Resveratrol Attenuates Trimethylamine-N-Oxide (TMAO)-Induced Atherosclerosis by Regulating TMAO Synthesis and Bile Acid Metabolism via Remodeling of the Gut Microbiota.

UNLABELLED: The gut microbiota is found to be strongly associated with atherosclerosis (AS). Resveratrol (RSV) is a natural phytoalexin with anti-AS effects; however, its mechanisms of action remain unclear. Therefore, we sought to determine whether the anti-AS effects of RSV were related to changes in the gut microbiota. We found that RSV attenuated trimethylamine-N-oxide (TMAO)-induced AS in ApoE(-/-) mice. Meanwhile, RSV decreased TMAO levels by inhibiting commensal microbial trimethylamine (TMA) production via gut microbiota remodeling in mice. Moreover, RSV increased levels of the genera Lactobacillus and Bifidobacterium, which increased the bile salt hydrolase activity, thereby enhancing bile acid (BA) deconjugation and fecal excretion in C57BL/6J and ApoE(-/-) mice. This was associated with a decrease in ileal BA content, repression of the enterohepatic farnesoid X receptor (FXR)-fibroblast growth factor 15 (FGF15) axis, and increased cholesterol 7a-hydroxylase (CYP7A1) expression and hepatic BA neosynthesis. An FXR antagonist had the same effect on FGF15 and CYP7A1 expression as RSV, while an FXR agonist abolished RSV-induced alterations in FGF15 and CYP7A1 expression. In mice treated with antibiotics, RSV neither decreased TMAO levels nor increased hepatic BA synthesis. Additionally, RSV-induced inhibition of TMAO-caused AS was also markedly abolished by antibiotics. In conclusion, RSV attenuated TMAO-induced AS by decreasing TMAO levels and increasing hepatic BA neosynthesis via gut microbiota remodeling, and the BA neosynthesis was partially mediated through the enterohepatic FXR-FGF15 axis. IMPORTANCE: Recently, trimethylamine-N-oxide (TMAO) has been identified as a novel and independent risk factor for promoting atherosclerosis (AS) partially through inhibiting hepatic bile acid (BA) synthesis. The gut microbiota plays a key role in the pathophysiology of TMAO-induced AS. Resveratrol (RSV) is a natural phytoalexin with prebiotic benefits. A growing body of evidence supports the hypothesis that phenolic phytochemicals with poor bioavailability are possibly acting primarily through remodeling of the gut microbiota. The current study showed that RSV attenuated TMAO-induced AS by decreasing TMAO levels and increasing hepatic BA neosynthesis via gut microbiota remodeling. And RSV-induced hepatic BA neosynthesis was partially mediated through downregulating the enterohepatic farnesoid X receptor-fibroblast growth factor 15 axis. These results offer new insights into the mechanisms responsible for RSV's anti-AS effects and indicate that the gut microbiota may become an interesting target for pharmacological or dietary interventions to decrease the risk of developing cardiovascular diseases.

Capsaicin-containing chili improved postprandial hyperglycemia, hyperinsulinemia, and fasting lipid disorders in women with gestational diabetes mellitus and lowered the incidence of large-for-gestational-age newborns.

BACKGROUND & AIMS: Gestational diabetes mellitus (GDM) may increase the future health risks of women and their offspring. The aim of this study was to determine the effect of capsaicin supplementation on blood glucose, lipid metabolism and pregnancy outcomes in women with GDM. METHODS: Forty-four pregnant women with GDM at 22-33 gestational weeks were randomly assigned to the capsaicin group (5 mg/d of capsaicin) or to the placebo group (0 mg/d of capsaicin) for 4 weeks in a randomized, double-blind, placebo-controlled trial. The concentrations of fasting plasma glucose and serum insulin, 2-h postprandial plasma glucose (2-h PG) and serum insulin (2-h INS), and fasting serum lipids, liver and kidney function parameters, and calcitonin gene-related peptide (CGRP) were measured at 0 and 4 weeks. The maternal and neonatal outcomes were also recorded. RESULTS: Forty-two women completed the trial. Compared to the placebo group, 2-h PG and 2-h INS concentrations and 2-h postprandial HOMA-IR (2-h HOMA-IR) levels, and the fasting serum total cholesterol and triglycerides concentrations significantly decreased in the capsaicin group after treatment (P < 0.05). Moreover, the fasting serum apolipoprotein B and CGRP concentrations significantly increased in the capsaicin group (P < 0.05). The changes in the 2-h PG and 2-h INS concentrations and in the 2-h HOMA-IR were negatively correlated with the change in the serum CGRP concentration (P < 0.05). Furthermore, the incidence of large-for-gestational-age (LGA) newborns was significantly lower in the capsaicin group than in the placebo group (P = 0.022). CONCLUSIONS: Capsaicin-containing chili supplementation regularly improved postprandial hyperglycemia and hyperinsulinemia as well as fasting lipid metabolic disorders in women with GDM, and it decreased the incidence of LGA newborns.

Fish Oil Supplements Lower Serum Lipids and Glucose in Correlation with a Reduction in Plasma Fibroblast Growth Factor 21 and Prostaglandin E2 in Nonalcoholic Fatty Liver Disease Associated with Hyperlipidemia: A Randomized Clinical Trial.

UNLABELLED: Fish oil has been used effectively in the treatment of cardiovascular disease via triglyceride reduction and inflammation modulation. This study aimed to assess the effects of fish oil on patients with nonalcoholic fatty liver disease (NAFLD) associated with hyperlipidemia. Eighty participants with NAFLD associated with hyperlipidemia were randomly assigned to consume fish oil (n=40, 4 g/d) or corn oil capsules (n=40, 4 g/d) for 3 months in a double-blind, randomized clinical trial. Blood levels of lipids, glucose and insulin, liver enzymes, kidney parameters and cytokines at baseline and the end of the study were measured. Seventy people finished the trial. Plasma concentrations of eicosapentaenoic acid and docosahexaenoic acid significantly increased in the fish oil group after intervention. After adjustment for age, gender and BMI, fish oil significantly decreased fasting serum concentrations of total cholesterol, triglyceride, apolipoprotein B and glucose (by (mean±SD) 0.49±0.43 mmol/L, 0.58±0.89 mmol/L, 0.28±0.33 g/L and 0.76±0.56 mmol/L, respectively, P<0.05), as well as alanine aminotransferase and γ-glutamyl transpeptidase levels (by (median (interquartile)) 9.0(0.5, 21.5) and 7.0(2.2, 20.0) IU/L, respectively, P<0.05), significantly increased serum adiponectin levels (by 1.29±0.62 µg/mL, P<0.001), and reduced serum levels of tumor necrosis factor α, leukotrienes B4, fibroblast growth factor 21 (FGF21), cytokeratin 18 fragment M30 and prostaglandin E2 (by 1.70±1.18 pg/mL, 0.59±0.28 ng/mL, 121±31 pg/mL, 83±60 IU/L and 10.9±2.3 pg/mL, respectively, P<0.001). Corn oil had no effect except for increasing serum creatinine concentrations by 7.7±8.9 µmol/L (P=0.008). The effects of fish oil on lipids, glucose and γ-glutamyl transpeptidase were positively correlated with the reductions of serum FGF21 and prostaglandin E2 concentrations after adjustment for age, gender and BMI (r = 0.275 to 0.360 and 0.261 to 0.375, respectively, P<0.05). In conclusion, our findings suggest that fish oil can benefit metabolic abnormalities associated with NAFLD treatment. TRIAL REGISTRATION: ChiCTR-TRC-12002380.

Resveratrol improves hepatic steatosis by inducing autophagy through the cAMP signaling pathway.

SCOPE: Resveratrol (RSV), a natural polyphenol, has been reported to attenuate nonalcoholic fatty liver disease (NAFLD); however, its underlying mechanism is unclear. Autophagy was recently identified as a critical protective mechanism during NAFLD development. Therefore, we investigated the role of autophagy in the beneficial effects of RSV on hepatic steatosis. METHODS AND RESULTS: Via Oil red O staining, triglyceride, and ß-hydroxybutyrate detection, we found that RSV decreased palmitate-induced lipid accumulation and stimulated fatty acid ß-oxidation in hepatocytes. Based on Western blot assay, confocal microscopy and transmission electron microscopy, we found that RSV induced autophagy in hepatocytes, whereas autophagy inhibition markedly abolished RSV-mediated hepatic steatosis improvement. Moreover, RSV increased cAMP levels and the levels of SIRT1 (sirtuin 1), pPRKA (phosphorylated protein kinase A), and pAMPK (phosphorylated AMP-activated protein kinase), as well as SIRT1 activity in HepG2 cells. Incubation with inhibitors of AC (adenylyl cyclase), PRKA, AMPK, SIRT1, or with AC, PRKA, AMPK, or SIRT1 siRNA abolished RSV-mediated autophagy. Similar results were obtained in mice with hepatic steatosis. CONCLUSION: RSV improved hepatic steatosis partially by inducing autophagy via the cAMP-PRKA-AMPK-SIRT1 signaling pathway, which provides new evidence regarding RSV's effects on NAFLD treatment.

Genistein inhibits DNA methylation and increases expression of tumor suppressor genes in human breast cancer cells.

It has been previously demonstrated that genistein exhibits anticancer activity against breast cancer. However, the precise mechanisms underlying the anticancer effect of genistein, in particular the epigenetic basis, remain unclear. In this study, we investigated whether genistein could modulate the DNA methylation status and expression of cancer-related genes in breast cancer cells. We treated MCF-7 and MDA-MB-231 human breast cancer cells with genistein in vitro. We found that genistein decreased the levels of global DNA methylation, DNA methyltransferase (DNMT) activity and expression of DNMT1. Yet, the expression of DNMT3A and DNMT3B showed no significant change. Using molecular modeling, we observed that genistein might directly interact with the catalytic domain of DNMT1, thus competitively inhibiting the binding of hemimethylated DNA to the catalytic domain of DNMT1. Furthermore, genistein decreased DNA methylation in the promoter region of multiple tumor suppressor genes (TSGs) such as ataxia telangiectasia mutated (ATM), adenomatous polyposis coli (APC), phosphatase and tensin homolog (PTEN), mammary serpin peptidase inhibitor (SERPINB5), and increased the mRNA expression of these genes. However, we detected no significant changes in the DNA methylation status or mRNA expression of stratifin (SFN). These results suggest that the anticancer effect of genistein on breast cancer may be partly due to its ability to demethylate and reactivate methylation-silenced TSGs through direct interaction with the DNMT1 catalytic domain and inhibition of DNMT1 expression.

Resveratrol attenuates vascular endothelial inflammation by inducing autophagy through the cAMP signaling pathway.

Inflammation participates centrally in all stages of atherosclerosis (AS), which begins with inflammatory changes in the endothelium, characterized by expression of the adhesion molecules. Resveratrol (RSV) is a naturally occurring phytoalexin that can attenuate endothelial inflammation; however, the exact mechanisms have not been thoroughly elucidated. Autophagy refers to the normal process of cell degradation of proteins and organelles, and is protective against certain inflammatory injuries. Thus, we intended to determine the role of autophagy in the antiinflammatory effects of RSV in human umbilical vein endothelial cells (HUVECs). We found that RSV pretreatment reduced tumor necrosis factor ? (TNF/TNF?)-induced inflammation and increased MAP1LC3B2 (microtubule-associated protein 1 light chain 3 ? 2) expression and SQSTM1/p62 (sequestosome 1) degradation in a concentration-dependent manner. A bafilomycin A 1 (BafA1) challenge resulted in further accumulation of MAP1LC3B2 in HUVECs. Furthermore, autophagy inhibitors 3-methyladenine (3-MA), chloroquine as well as ATG5 and BECN1 siRNA significantly attenuated RSV-induced autophagy, which, subsequently, suppressed the downregulation of RSV-induced inflammatory factors expression. RSV also increased cAMP (cyclic adenosine monophosphate) content, the expression of PRKA (protein kinase A) and SIRT1 (sirtuin 1), as well as the activity of AMPK (AMP-activated protein kinase). RSV-induced autophagy in HUVECs was abolished in the presence of inhibitors of ADCY (adenylyl cyclase, KH7), PRKA (H-89), AMPK (compound C), or SIRT1 (nicotinamide and EX-527), as well as ADCY, PRKA, AMPK, and SIRT1 siRNA transfection, indicating that the effects of RSV on autophagy induction were dependent on cAMP, PRKA, AMPK and SIRT1. In conclusion, RSV attenuates endothelial inflammation by inducing autophagy, and the autophagy in part was mediated through the activation of the cAMP-PRKA-AMPK-SIRT1 signaling pathway.

Absorption of resveratrol by vascular endothelial cells through passive diffusion and an SGLT1-mediated pathway.

Resveratrol is a natural polyphenol that exerts potent effects to suppress atherosclerosis. However, its low concentration in plasma has placed this role in doubt. Thus, resveratrol effects might be dependent on its transport into vascular endothelium, a question not previously addressed in spite of its obvious and fundamental importance. Via high-performance liquid chromatography and liquid chromatography/mass spectrometry, we found that resveratrol was absorbed by human umbilical vein endothelial cells in a temperature-, concentration- and time-dependent manner, suggesting the involvement of passive diffusion and active transport. As determined by confocal laser scanning microscopy, resveratrol primarily distributed throughout the cytoplasm. Furthermore, resveratrol absorption was modulated by serum proteins and sodium-dependent glucose transporter 1 (SGLT1) yet inhibited by glucose (an SGLT1 substrate) and phlorizin (an SGLT1 selective inhibitor), as well as SGLT1 siRNA transfection. Additionally, Sprague-Dawley rats were intragastrically administrated with 100mg/kg of resveratrol and the concentration of resveratrol in blood vessels declined more slowly up to 24h compared to that in the blood. Our results suggested that resveratrol uptake by vascular endothelial cells involved both passive diffusion and an SGLT1-mediated process, at least partially. Moreover, the intracellular resveratrol pool may be more important than the serum level in vivo. These provide new insights into the cardiovascular benefits of resveratrol.

Delphinidin-3-glucoside protects against oxidized low-density lipoprotein-induced mitochondrial dysfunction in vascular endothelial cells via the sodium-dependent glucose transporter SGLT1.

Delphinidin-3-glucoside (Dp) is a member of a family of bioactive compounds known as anthocyanins that occur naturally in pigmented plants and are known to ameliorate oxidative stress. Previous studies have showed that Dp decreased oxidative stress in vascular endothelial cells, however, the underlying mechanisms remain largely unknown. In the present study, we showed that pretreatment with Dp significantly suppressed oxidized low-density lipoprotein (oxLDL)-induced cell proliferation inhibition and apoptosis in primary human umbilical vein endothelial cells (HUVECs). Also, Dp pretreatment attenuated oxLDL-induced mitochondrial dysfunction via decreased reactive oxygen species (ROS) and superoxide anion generation, thereby repressing mitochondrial membrane potential and closing mitochondrial permeability transition pore. Furthermore, in vitro and in vivo data showed that Dp was transported into endothelial cells in a temperature, concentration, and time-dependent manner via the sodium-dependent glucose transporter (SGLT1). Suppression of SGLT1 by its substrate glucose, its inhibitor phlorizin or SGLT1 siRNA blocked Dp transportation. Repression of SGLT1 significantly inhibited Dp function of ameliorating mitochondrial dysfunction induced by pro-apoptotic factors (Apoptosis-inducing factor, Cytochrome c, Caspase-3 and Bax/Bcl-2 ratio). Taken together, our data indicate that Dp protects VECs via the SGLT1-ROS-mitochodria pathway. This new insight may help to elucidate the molecular mechanisms underlying the vascular protection afforded by Dp, and anthocyanins in general, in the context of prevention of endothelial dysfunction and atherosclerosis.

Isoflavones for hypercholesterolaemia in adults.

BACKGROUND: Hypercholesterolaemia is a significant risk factor for cardiovascular diseases. Isoflavones may be effective in improving hypercholesterolaemia. OBJECTIVES: To assess the effects of isoflavones for hypercholesterolaemia. SEARCH METHODS: We searched the following databases: The Cochrane Library (Issue 9, 2012), MEDLINE, EMBASE, Chinese BioMedical Database and China National Knowledge Infrastructure (all to September 2012). SELECTION CRITERIA: We considered randomized controlled clinical trials in hypercholesterolaemic participants comparing isoflavones versus placebo, or soy isolated protein added with isoflavones versus soy isolated protein alone. DATA COLLECTION AND ANALYSIS: Two review authors independently abstracted relevant population and intervention characteristics. We resolved any disagreements through discussion, or if required by a third party. We assessed the risk of bias of trials against key criteria: random sequence generation, allocation concealment, blinding of participants and personnel, blinding of outcome assessment, incomplete outcome data, selective reporting and other sources of bias. MAIN RESULTS: We included five randomized trials (208 participants, 104 in the intervention group and 104 in the control group). Interventions ranged from three to six months. Four trials reported results in non-Asian populations published in English. One trial reported results in Chinese people published in Chinese. Overall, the risk of bias of included trials was high or unclear. There were no outcome data on death from any cause, morbidity, complications, health-related quality of life and costs. Two trials reported adverse effects, including gastrointestinal discomfort (bloating and constipation) and an increased number of hot flushes. None of the trials found serious adverse events. There was a slight significant effect on triglycerides in favour of isoflavones when compared with placebo (mean difference (MD) -0.46 mmol/L (95% confidence interval (CI) -0.84 to -0.09; P = 0.02; 52 participants; 2 trials). No statistically significant effects on total cholesterol, low-density lipoprotein cholesterol and high-density lipoprotein cholesterol were shown in favour of isoflavones. AUTHORS' CONCLUSIONS: We found no evidence for effects of isoflavones on patient-important outcomes or lowering of cholesterol levels in people with hypercholesterolaemia. Our findings have to be interpreted with caution due to high or unclear risk of bias in several risk of bias domains, and low number of participants in trials.

Isoflavone consumption and risk of breast cancer: a dose-response meta-analysis of observational studies.

Epidemiologic studies that examine whether isoflavone consumption protects against breast cancer have yielded inconsistent results. The controversy focuses on the effects of the menopausal status and exposure dose of isoflavone. We aim to conduct a meta-analysis on the association between isoflavone intake and breast cancer risk by comprehensively assessing isoflavone exposure in the targeted populations. We searched PUBMED and EMBASE databases for case-control and cohort studies that assess the association between isoflavone intake and breast cancer risk. We extracted relative risks (RR) and odds ratios (OR) of different reported categories of isoflavone intake from each study. Fixed- or random-effects models were used to summarize dose-response data. Twenty-two studies were selected for the meta-analysis. Overall, the results showed that isoflavone reduced the breast cancer risk (a combined RR/OR of 0.68, 95% CI: 0.52-0.89) in Asian populations rather than Western populations (a combined RR/OR of 0.98, 95% CI: 0.87, 1.11) for the high-dose category. Further analysis showed that the intake of isoflavone in postmenopausal Asian women 0.46 (95% CI: 0.28-0.78) was better than premenopausal 0.63 (95% CI: 0.50-0.80) but similar in postmenopausal Western women 1.00 (95% CI: 0.98-1.02) and premenopausal 0.99 (95% CI: 0.87-1.12). Exposure to high isoflavone may be associated with a reduced breast cancer risk in Asian populations, especially in postmenopausal women. However, no significant difference in the studies of Western populations may be due to the low intake of isoflavone levels.

Activation of nuclear factor erythroid 2-related factor 2 and PPARγ plays a role in the genistein-mediated attenuation of oxidative stress-induced endothelial cell injury.

We investigate the cytoprotective effects and the molecular mechanism of genistein in oxidative stress-induced injury using an endothelial cell line (EA.hy926). An oxidative stress model was established by incubating endothelial cells with H2O2. According to the present results, genistein pretreatment protected endothelial cells against H2O2-induced decreases in cell viability and increases in apoptosis. Genistein also prevented the inhibition of B-cell lymphoma 2 and the activation of caspase-3 induced by H2O2. Genistein increased superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) levels and attenuated the decrease in these antioxidants during oxidative stress. We also found that genistein induced the promoter activity of both nuclear factor erythroid 2-related factor 2 (Nrf2) and PPARγ. Additionally, genistein induced the nuclear translocation of Nrf2 and PPARγ. While genistein caused the up-regulation of both Nrf2 and PPARγ, it also activated and up-regulated the protein expression and transcription of a downstream protein, haem oxygenase-1 (HO-1). Moreover, the use of Nrf2 small interfering RNA transfection and HO-1- or PPARγ-specific antagonists (Znpp and GW9662, respectively) blocked the protective effects of genistein on endothelial cell viability during oxidative stress. Therefore, we conclude that oxidative stress-induced endothelial cell injury can be attenuated by treatment with genistein, which functions via the regulation of the Nrf2 and PPARγ signalling pathway. Additionally, the endogenous antioxidants SOD, CAT and GSH appear to play a role in the antioxidant activity of genistein. The present findings suggest that the beneficial effects of genistein involving the activation of cytoprotective antioxidant genes may represent a novel strategy in the prevention and treatment of cardiovascular endothelial damage.

Differential suppression of intracellular reactive oxygen species-mediated signaling pathway in vascular endothelial cells by several subclasses of flavonoids.

Increased intracellular reactive oxygen species (ROS) is crucial for vascular endothelial dysfunction, a key step in the initiating of atherosclerosis (AS). The antioxidant activity of flavonoids has been suggested to contribute to AS prevention. However, The association of the structure characteristics to antioxidant capacities in relation to the inhibitory effects on endothelial dysfunction has not been well established. In this study, four subclasses of flavonoids with similar structures, including two anthocyanins (delphinidin and cyanidin), two flavonols (myricetin and quercetin), two flavones (luteolin and apigenin) and two isoflavones (genistein and daidzein) were examined for their inhibitory effects on intracellular ROS-mediated signaling pathway in the human umbilical vein endothelial cell EA.hy926. Cells were pretreated with different flavonoids for 2 h and then exposed to oxLDL of 100 µg/ml for another 24 h. It was found that treatment with different flavonoids alone had no notable effects on cell viability. However, the oxLDL-induced decrease of cell viability, generation of O(2)(·-) and ROS, p38MAPK activation, NF-κB nuclear translocation, NF-κB-modulated transcriptional activity as well as the mRNA expression of genes including ICAM-1, VCAM-1, E-selectin, MMP-1, MMP-2 and MMP-9 were notably inhibited by the pretreatment of different flavonoids through blunting ROS-triggered signaling pathway, in spite of apparent differences. And the number of hydroxyl groups in total, 3',4'-ortho-dihydroxyl in B-ring and 3-hydroxyl group in C-ring of flavonoids were important structure characteristics for the inhibitory effects. Thus, anthocyanins and flavonols such as delphinidin and myricetin exert higher ROS scavenging activities and more significant endothelium-protective effects compared to the other compounds. Our results provide evidence for AS prevention and a basis for designing the potent anti-atherosclerotic agents.

Chemical structures of 4-oxo-flavonoids in relation to inhibition of oxidized low-density lipoprotein (LDL)-induced vascular endothelial dysfunction.

Vascular endothelial dysfunction induced by oxidative stress has been demonstrated to be the initiation step of atherosclerosis (AS), and flavonoids may play an important role in AS prevention and therapy. Twenty-three flavonoids categorized into flavones, flavonols, isoflavones, and flavanones, all with 4-oxo-pyronenucleus, were examined for what structural characteristics are required for the inhibitory effects on endothelial dysfunction induced by oxidized low-density lipoprotein (oxLDL). Human vascular endothelial cells EA.hy926 were pretreated with different 4-oxo-flavonoids for 2 hs, and then exposed to oxLDL for another 24 hs. Cell viability and the level of malondialdehyde (MDA), nitric oxide (NO) and soluble intercellular adhesion molecule-1 (sICAM-1) were measured, respectively. Then, correlation analysis and paired comparison were used to analyze the structure-activity relationships. Significant correlations were observed between the number of -OH moieties in total or in B-ring and the inhibitory effectson endothelial dysfunction. Furthermore, 3',4'-ortho-dihydroxyl on B-ring, 3-hydroxyl on C-ring and 2,3-double bondwere correlated closely to the inhibitory effects of flavonolson cell viability decrease and lipid peroxidation. 5,7-meta-dihydroxyl group on A-ring was crucial for the anti-inflammatory effects of flavones and isoflavones in endothelial cells. Moreover, the substituted position of B-ring on C3 rather than C2 was important for NO release. Additionally, hydroxylation at C6 position significantly attenuated the inhibitory effects of 4-oxo-flavonoids on endothelial dysfunction. Our findings indicated that the effective agents in inhibiting endothelial dysfunction include myricetin, quercetin, luteolin, apigenin, genistein and daidzein. Our work might provide some evidence for AS prevention and a strategy for the design of novel AS preventive agents.

Inhibitory effect of delphinidin on monocyte-endothelial cell adhesion induced by oxidized low-density lipoprotein via ROS/p38MAPK/NF-κB pathway.

Monocyte adhesion to the vascular endothelium and their subsequent trans-endothelial migration are pivotal early events in atherogenesis. In this study, the effect of delphinidin, belonging to the group of anthocyanin, on adhesion of monocytes to endothelial cells induced by ox-LDL was investigated. The results showed that the pre-treatment with delphinidin (50, 100, or 200 µM) dose-dependently decreased the ox-LDL-induced up-regulation of the expression of ICAM-1 and P-selectin, and the enhanced adhesion and transmigration of monocytes. To determine the role of ROS/p38MAPK/NF-κB pathway, intracellular ROS level, p38MAPK protein expression, NF-κB transcription activity and protein expression, IκB-α degradation, NADPH oxidase subunit (Nox2 and p22phox) protein, and mRNA expression were measured. The results showed that delphinidin attenuated ox-LDL-induced generation of ROS, p38MAPK protein expression, NF-κB transcription activity and protein expression, IκB-α degradation, NADPH oxidase subunit (Nox2 and p22phox) protein and mRNA expression in endothelial cells in a dose-dependent manner. These results suggest that delphinidin attenuates ox-LDL induced expression of adhesion molecules (P-selectin and ICAM-1) and the adhesion of monocytes to endothelial cells by inhibiting ROS/p38MAPK/NF-κB pathway. These findings provide a basis for the design of potent antiatherosclerotic agents that will have therapeutic potential in the prevention of AS.

Effects of different dietary fatty acids on the fatty acid compositions and the expression of lipid metabolic-related genes in mammary tumor tissues of rats.

In this study, the effects of dietary fatty acids on the fatty acid compositions and lipid metabolic-related genes expression in N-methyl-N-nitrosourea (MNU)-induced rat mammary carcinogenesis were evaluated. The 50-day-old female Sprague-Dawley rats were intervened by different dietary fats (15% wt/wt), including saturated fatty acid (SFA), monounsaturated fatty acid (MUFA), n-6 polyunsaturated fatty acid (PUFA), n-3 PUFA, 1:1 n-6/n-3, 5:1 n-6/n-3, 10:1 n-6/n-3, and 1:2:1 S/M/P (1:1 n-6/n-3), alone or in combination with MNU. There was no mammary tumor occurrence in the control and MNU-treated n-3 PUFA groups after 18 wk. n-3 PUFA diet retarded the weight growth of rats. 1:1 n-6/n-3 diet significantly reduced the MNU-induced tumor incidence and tumor multiplicity compared with SFA, MUFA, n-6 PUFA, 5:1 n-6/n-3, 10:1 n-6/n-3 and 1:2:1 S/M/P diets (42.86% vs. 83.33%-92.31%, 0.79 vs. 2.62-2.85, P < 0.01). Additionally, 1:1 n-6/n-3 diet substantially increased cis-5,8,11,14,17-eicosapentaenoic acid and cis-4,7,10,13,16,19-docosahexaenoic acid levels, whereas it decreased C20:4 level and the mRNA expressions of fatty acid synthase, Cyclooxygenase-2 (COX-2), and 5-lipoxygenase (5-LOX) in mammary tissues (P < 0.05). These results suggest that 1:1 n-6/n-3 in the diet is effective in the prevention of mammary tumor development by increasing the n-3 PUFA content and reducing the expression of lipid metabolic-related genes.

[Role of Rho GTPase in inhibiting metastatic ability of human prostate cancer cell line PC-3 by omega-3 polyunsaturated fatty acid].

BACKGROUND & OBJECTIVE: Recently, researches refer to the influence of polyunsaturated fatty acid (PUFA) on cancer initiation and progression had been highly concerned. This study was to investigate the effects of 2 kinds of omega-3 PUFA, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on the metastatic ability of human prostate cancer cell line PC-3, and explore the role of Rho GTPase in inhibiting cancer metastasis by omega-3 PUFA. METHODS: MTT assay was used to determine the effects of omega-3 PUFA on the proliferation of PC-3 cells. Adhesion assay, invasion assay, and migration assay were used to observe the effects of omega-3 PUFA on the metastatic ability of PC-3 cells. Western blot was used to observe the effects of omega-3 PUFA on the expression of RhoA, Rac1, Rac2, and Cdc42 proteins in PC-3 cells. Laser confocal microscopy was used to investigate the effect of omega-3 PUFA on the reorganization of the microfilaments and microtubules marked by immunofluorescent cytochemistry technology. RESULTS: Both EPA and DHA inhibited the proliferation of PC-3 cells, and the proliferation inhibition rate increased along with the increase of the concentration and treatment time. When treated with 60 mumol/L EPA or DHA for 48 h, the abilities of adhesion, invasion and migration of PC-3 cells were inhibited (P<0.05). omega-3 PUFA significantly suppressed the expression of Rac1, Rac2 and Cdc42 proteins (P<0.05), influenced the distribution and structure of sytoskeletons. CONCLUSION: omega-3 PUFA could inhibit the metastatic ability of PC-3 cells through down-regulating the expression of Rho GTPase and inhibiting the cytoskeleton reorganization.