OBJECTIVE: To evaluate the synergic effects of a novel oral supplement formulation, containing prebiotics, yeast β-glucans, minerals and silymarin (Silybum marianum), on lipid and glycidic metabolism, inflammatory and mitochondrial proteins of the liver, in control and high-fat diet-induced obese mice. METHODS: After an acclimation period, 32 male C57BL/6 mice were divided into the following groups: nonfat diet (NFD) vehicle, NFD supplemented, high-fat diet (HFD) vehicle and HFD supplemented. The vehicle and experimental formulation were administered orally by gavage once a day during the last four weeks of the diet (28 consecutive days). We then evaluated energy homeostasis, inflammation, and mitochondrial protein expression in these groups of mice. RESULTS: After four weeks of supplementation, study groups experienced reduced glycemia, dyslipidemia, fat, and hepatic fibrosis levels. Additionally, proliferator-activated receptor-α, AMP-activated protein kinase-1α, peroxisome proliferator-activated receptor γ co-activator-1α, and mitochondrial transcription factor A expression levels were augmented; however, levels of inhibitor of nuclear factor-κB kinase subunit α and p65 nuclear factor-κB expression, and oxidative markers were reduced. Notably, the cortisol/C-reactive protein ratio, a well-characterized marker of the hypothalamic-pituitary-adrenal axis immune interface status, was found to be modulated by the supplement. CONCLUSION We discovered that the novel supplement was able to modify different antioxidant, metabolic and inflammatory pathways, improving the energy homeostasis and inflammatory status, and consequently alleviated hepatic steatosis.
Animals ; Antioxidants ; Dietary Supplements ; Glucans ; Hypothalamo-Hypophyseal System ; Liver ; Mice ; Mice, Inbred C57BL ; Mice, Obese ; Milk Thistle ; Minerals ; Pituitary-Adrenal System ; Prebiotics ; Saccharomyces cerevisiae

(2S)-taxifolin is an important flavonoid that has anti-inflammatory and anti-oxidation effects. It is widely used in pharmaceutical and nutraceutical industries. Flavone 3-hydroxylase (F3H) can catalyze the synthesis of (2S)-taxifolin and other 3-hydroxylated flavonoids from (2S)-eriodictyol. Due to the low catalytic efficiency of F3H, the titer of many 3-hydroxyflavones, such as taxifolin, synthesized by microbial method is relatively low. In this study, a SmF3H was identified from the transcriptome of Silybum marianum (L.) Gaertn. The results of fermentation showed that SmF3H can catalyze the flavone 3-hydroxylation reaction, and its catalytic efficiency was significantly higher than that of commonly used SlF3H from Solanum lycopersicum. Six promoters with different transcription strength were selected to optimize the synthesis pathway from the flavonoid precursor (2S)-naringenin to (2S)-taxifolin. The results showed that the highest titer of (2S)-taxifolin (695.90 mg/L in shake flask) could be obtained when the P(GAL7) promoter was used to control the expression of SmF3H. The titer of (2S)-taxifolin was further improved to 3.54 g/L in a 5-L fermenter, which is the highest titer according to current available literatures.
Antioxidants ; Flavonoids ; Milk Thistle ; Quercetin/analogs & derivatives*

Silymarin is the standardized extract from Silybum marianum which consists mainly of flavonoids and polyphenols. It is highly regarded for its hepatoprotective ability. Silybin B is a flavonolignan and one of the active components of silymarin. The content of silybin B in various parts of S. marianum was analyzed by HPLC-UV. Results show that the extract of seeds contain the highest amount of silybin B (7.434 mg/g DW). The petioles of S. marianum showed a low content of silybin B. This study revealed that seeds of S. marianum contain high amount of silybin B and could be a good source of the compound.
Flavonoids ; Milk Thistle ; Polyphenols ; Silymarin

BACKGROUND/AIMS: Hepatic steatosis is caused by an imbalance between free fatty acids (FFAs) uptake, utilization, storage, and disposal. Understanding the molecular mechanisms involved in FFAs accumulation and its modulation could drive the development of potential therapies for Nonalcoholic fatty liver disease. The aim of the current study was to explore the effects of picroside II, a phytoactive found in Picrorhiza kurroa, on fatty acid accumulation vis-à-vis silibinin, a known hepatoprotective phytoactive from Silybum marianum. METHODS: HepG2 cells were loaded with FFAs (oleic acid:palmitic acid/2:1) for 20 hours to mimic hepatic steatosis. The FFAs concentration achieving maximum fat accumulation and minimal cytotoxicity (500 μM) was standardized. HepG2 cells were exposed to the standardized FFAs concentration with and without picroside II pretreatment. RESULTS: Picroside II pretreatment inhibited FFAs-induced lipid accumulation by attenuating the expression of fatty acid transport protein 5, sterol regulatory element binding protein 1 and stearoyl CoA desaturase. Preatreatment with picroside II was also found to decrease the expression of forkhead box protein O1 and phosphoenolpyruvate carboxykinase. CONCLUSIONS: These findings suggest that picroside II effectively attenuated fatty acid accumulation by decreasing FFAs uptake and lipogenesis. Picroside II also decreased the expression of gluconeogenic genes.
Fatty Acid Transport Proteins ; Fatty Acids, Nonesterified ; Hep G2 Cells* ; Lipogenesis ; Milk Thistle ; Non-alcoholic Fatty Liver Disease ; Phosphoenolpyruvate ; Picrorhiza ; Stearoyl-CoA Desaturase ; Sterol Regulatory Element Binding Protein 1

Fructus Arctii is a traditional Chinese medicine. The main counterfeit species are the seeds of Arctium tomentosum, Onopordum acanthium, Silybum marianum, Saussurea costus, Amorpha fruticosa. Traditional identification methods or molecular barcoding techniques can identify Fructus Arctii and its counterfeit species. However, the identification of the mixture of it and its spurious species is rarely reported. In this paper, we sequenced the ITS2 sequences of Fructus Arctii and 5 kinds of spurious species mix powder by high-throughput sequencing to identify the mixed powder species and providing new ideas for the identification of Fructus Arctii mix powder. The total DNA in mixed powder was extracted, and the ITS2 sequences in total DNA was amplified. Paired-end sequencing was performed on the DNA fragment of the community using the Illumina MiSeq platform. The sequence was analyzed by the software FLASH, QIIME and GraPhlAn etc. The results showed that the high quality ITS2 sequences of 39910 mix samples were obtained from the mixed samples, of which the total ITS2 sequence of the samples genus was 34 935. Phylogenetic analysis showed that the samples contained Fructus Arctii, A. tomentosum, O. acanthium, S. marianum, S. costus and A. fruticosa. Using ITS2 sequences as DNA barcodes, high-throughput sequencing technology can be used to detect the Fructus Arctii and its spurious specie in mixed powder, which can provide reference for the quality control, safe use of medicinal materials of Fructus Arctii and the identification of mixed powder of traditional Chinese medicine.
Arctium ; chemistry ; classification ; DNA Barcoding, Taxonomic ; DNA, Plant ; genetics ; DNA, Ribosomal Spacer ; genetics ; Drug Contamination ; Drugs, Chinese Herbal ; standards ; Fabaceae ; Fruit ; High-Throughput Nucleotide Sequencing ; Milk Thistle ; Onopordum ; Phylogeny ; Saussurea

Silibinin, an active constituent of silymarin extracted from milk thistle, has been previously reported to confer protection to the adult brain against neurodegeneration. However, its effects against epileptic seizures have not been examined yet. In order to investigate the effects of silibinin against epileptic seizures, we used a relevant mouse model in which seizures are manifested as status epilepticus, induced by kainic acid (KA) treatment. Silibinin was injected intraperitoneally, starting 1 day before an intrahippocampal KA injection and continued daily until analysis of each experiment. Our results indicated that silibinin-treatment could reduce seizure susceptibility and frequency of spontaneous recurrent seizures (SRS) induced by KA administration, and attenuate granule cell dispersion (GCD), a morphological alteration characteristic of the dentate gyrus (DG) in temporal lobe epilepsy (TLE). Moreover, its treatment significantly reduced the aberrant levels of apoptotic, autophagic and pro-inflammatory molecules induced by KA administration, resulting in neuroprotection in the hippocampus. Thus, these results suggest that silibinin may be a beneficial natural compound for preventing epileptic events.
Adult ; Animals ; Brain ; Dentate Gyrus ; Epilepsy ; Epilepsy, Temporal Lobe ; Hippocampus* ; Humans ; Kainic Acid ; Mice ; Milk Thistle ; Neuroprotection ; Seizures ; Silymarin ; Status Epilepticus

Luteolin 5-O-glucoside is the major flavonoid from Korean thistle, Cirsium maackii. We previously reported the anti-inflammatory activities of luteolin 5-O-glucoside in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. In this study, we determined the anti-inflammatory mechanisms of luteolin 5-O-glucoside through the inhibition of nitric oxide (NO) production in vitro and in vivo. Results revealed that luteolin 5-O-glucoside dose-dependently inhibited NO production and expression of iNOS and COX-2 in LPS-induced RAW 264.7 cells. Luteolin 5-O-glucoside also significantly inhibited the translocation of NF-κB, the activation of MAPKs, and ROS generation in LPS-induced RAW 264.7 cells. In addition, protein expressions of Nrf-2 and HO-1 were also upregulated by luteolin 5-O-glucoside treatment. Moreover, luteolin 5-O-glucoside inhibited λ-carrageenan-induced mouse paw edema by 65.34% and 48.31% at doses of 50 and 100 mg/kg body weight, respectively. These findings indicate potential anti-inflammatory effect of luteolin 5-O-glucoside particularly by downregulating NF-κB and upregulating HO-1/Nrf-2 pathway.
Animals ; Body Weight ; Cirsium* ; Edema ; In Vitro Techniques ; Luteolin* ; Mice ; Milk Thistle* ; Milk* ; Nitric Oxide ; RAW 264.7 Cells

OBJECTIVEIn the present study, we investigated the antioxidant and anti-aging effects of Silybum marianum protein hydrolysate (SMPH) in D-galactose-treated mice.

METHODSD-galactose (500 mg/kg body weight) was intraperitoneally injected daily for 7 weeks to accelerate aging, and SMPH (400, 800, 1,200 mg/kg body weight, respectively) was simultaneously administered orally. The antioxidant and anti-aging effects of SMPH in the liver and brain were measured by biochemical assays. Transmission electron microscopy (TEM) was performed to study the ultrastructure of liver mitochondri.

RESULTSSMPH decreased triglyceride and cholesterol levels in the D-galactose-treated mice. It significantly elevated the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and total antioxidant capacity (T-AOC), which were suppressed by D-galactose. Monoamine oxidase (MAO) and malondialdehyde (MDA) levels as well as the concentrations of caspase-3 and 8-OHdG in the liver and brain were significantly reduced by SMPH. Moreover, it increased Bcl-2 levels in the liver and brain. Furthermore, SMPH significantly attenuated D-galactose-induced liver mitochondrial dysfunction by improving the activities of Na+-K+-ATPase and Ca2+-Mg2+-ATPase as well as mitochondrial membrane potential (ΔΨm) and fluidity. TEM showed that the degree of liver mitochondrial damage was significantly decreased by SMPH.

CONCLUSIONThe results indicated that SMPH protects against D-galactose-induced accelerated aging in mice through its antioxidant and anti-aging activities.

Aging ; drug effects ; Animals ; Antioxidants ; pharmacology ; Brain ; drug effects ; Caspase 3 ; metabolism ; Galactose ; toxicity ; Gene Expression Regulation, Enzymologic ; drug effects ; Glutathione Peroxidase ; metabolism ; Male ; Malondialdehyde ; metabolism ; Maze Learning ; drug effects ; Mice ; Milk Thistle ; chemistry ; Mitochondria, Liver ; drug effects ; Oxidative Stress ; drug effects ; Plant Proteins ; chemistry ; pharmacology ; Protective Agents ; pharmacology ; Protein Hydrolysates ; chemistry ; pharmacology ; Superoxide Dismutase ; metabolism

The present study aimed to show that pro-inflammatory cytokines [tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and interleukin (IL)-1β] synergistically induce the production of nitric oxide (NO) production in mouse mesangial cells, which play an important role in inflammatory glomerular injury. We also found that co-treatment with cytokines at low doses (TNF-α; 5 ng/ml, IFN-γ; 5 ng/ml, and IL-1β; 1.25 U/ml) synergistically induced NO production, whereas treatment with each cytokine alone did not increase NO production at doses up to 100 ng/ml or 50 U/ml. Silymarin, a polyphenolic flavonoid isolated from milk thistle (Silybum marianum), attenuates cytokine mixture (TNF-α, IFN-γ, and IL-1β)-induced NO production. Western blot and RT-PCR analyses showed that silymarin inhibits inducible nitric oxide synthase (iNOS) expression in a dose-dependent manner. Silymarin also inhibited extracellular signal-regulated protein kinase-1 and -2 (ERK1/2) phosphorylation. Collectively, we have demonstrated that silymarin inhibits NO production in mouse mesangial cells, and may act as a useful anti-inflammatory agent.
Animals ; Blotting, Western ; Cytokines ; Interferons ; Interleukins ; Mesangial Cells* ; Mice* ; Milk Thistle ; Necrosis ; Nitric Oxide ; Nitric Oxide Synthase Type II ; Phosphorylation ; Silymarin*

Silymarin from milk thistle (Silybum marianum) has been reported to show an anti-cancer activity. In previous study, we reported that silymarin induces cyclin D1 proteasomal degradation through NF-κB-mediated threonine-286 phosphorylation. However, mechanism for the inhibition of Wnt signaling by silymarin still remains unanswered. Thus, we investigated whether silymarin affects Wnt signaling in human colorectal cancer cells to elucidate the additional anti-cancer mechanism of silymarin. Transient transfection with a TOP and FOP FLASH luciferase construct indicated that silymarin suppressed the transcriptional activity of β-catenin/TCF. Silymarin treatment resulted in a decrease of intracellular β-catenin protein but not mRNA. The inhibition of proteasome by MG132 and GSK3β inhibition by SB216763 blocked silymarin-mediated downregulation of β-catenin. In addition, silymarin increased phosphorylation of β-catenin and a point mutation of S33Y attenuated silymarin-mediated β-catenin downregulation. In addition, silymarin decreased TCF4 and increased Axin expression in both protein and mRNA level. From these results, we suggest that silymarin-mediated downregulation of β-catenin and TCF4 may result in the inhibition of Wnt signaling in human colorectal cancer cells.
Colorectal Neoplasms* ; Cyclin D1 ; Down-Regulation ; Humans* ; Luciferases ; Milk Thistle ; Phosphorylation ; Point Mutation ; Proteasome Endopeptidase Complex ; RNA, Messenger ; Silymarin* ; Transfection