Ethanol-Induced Hepatotoxicity and Alcohol Metabolism Regulation by GABA-Enriched Fermented Smilax china Root Extract in Rats.

Chronic alcohol consumption can cause hepatic injury and alcohol-induced toxicities. Extracts from Smilax china root have been widely used in traditional medicine and for their potential pharmacological benefits. We aimed to determine if fermented Smilax china extract (FSC) regulates alcoholic fatty liver and liver injury using two in vivo experiments. Sprague-Dawley rats were administered ethanol (3 g/kg b.w.; po) with or without FSC pretreatment to induce an acute hangover. In another experiment, rats were fed either a normal or Lieber-DeCarli ethanol (6.7%) diet with or without FSC pretreatment (125, 250, and 500 mg/kg b.w.; po) for 28 days. Serum biomarkers, liver histopathology, and the mRNA levels of anti-inflammatory, antioxidant, lipogenic, and lipolytic genes were analyzed. FSC pretreatment significantly reduced blood alcohol and acetaldehyde concentrations, upregulated the mRNA expression of alcohol dehydrogenase, aldehyde dehydrogenase, and superoxide dismutase, and decreased the activities of liver enzymes in a dose-dependent manner. It also downregulated SERBP-1c and upregulated PPAR-α and reduced the gene expression of the anti-inflammatory cytokine IL-6 in the liver. The final extract after fermentation had increased GABA content. Furthermore, FSC was found to be safe with no acute oral toxicity in female rats. Thus, FSC increases alcohol metabolism and exhibits antioxidant and anti-inflammatory effects to induce hepatoprotection against alcohol-induced damage. It may be used as a functional food ingredient after excess alcohol consumption.

Hepatoprotective effects of gamma-aminobutyric acid-enriched fermented Hovenia dulcis extract on ethanol-induced liver injury in mice.

BACKGROUND: Various extracts of Hovenia dulcis have been commonly used in Asia for cases of alcohol-related disorders. Fermentation is reported to enhance the level and biological activities of various bio-constituents of plant extracts. Therefore, this study was undertaken to evaluate the effects of fermented H. dulcis extract (FHDE) on ethanol-induced liver injury in mice. METHODS: FHDE was prepared using Bacillus subtilis and Lactobacillus plantarum. The effects of FHDE on ethanol-induced liver injury were evaluated in C57BL/6 N CrSlc mice. A mixed feed preparation containing the fermented extract with and without ethanol was given to mice for 29 days, according to its group. At the end of the experiment, blood and liver samples were collected from all mice in the group. Plasma biochemical analysis and histopathological investigation were performed to evaluate the impacts of treatment on the biomarkers of hepatic damage and inflammatory changes. Besides, the expression of genes that regulate the activities of enzymes associated with alcohol metabolism, antioxidant activity, and fatty acid oxidation was assessed using a quantitative real-time polymerase chain reaction. Moreover, the amino acid contents and the active ingredients of the extract were evaluated before and after fermentation. RESULTS: Fermentation resulted in a marked increase and decrease in the amount of Gamma-Amino-n-butyric acid (GABA) and glutamic acid, respectively. FHDE enhanced the body weight gain of mice compared to ethanol. Besides, plasma levels of triglyceride, low-density lipoprotein, the activities of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were significantly (P < 0.05) reduced in the FHDE-treated groups relative to the ethanol-treated control. FHDE upregulated the expression of genes associated with enzymes involved in alcohol dehydrogenation (Adh1 and Aldh2), antioxidant activity (SOD and CAT), and fatty acid oxidation (PPAR-α and PGC-1α). However, the expressions of Cytochrome peroxidase Cyp2E1 and genes related to lipogenesis (SREBP-1c, FAS, SCD-1, and ACC) were significantly (P < 0.05) downregulated following treatment with the FHDE. Histopathological investigation demonstrated a slight degree of inflammatory cell infiltration and occasional fatty changes in the FHDE-treated groups. CONCLUSION: The GABA-enriched fermented H. dulcis extract prevented ethanol-induced hepatic damage by enhancing the antioxidant defense system, fatty acid oxidation, and reducing lipogenesis.

An in vivo immunomodulatory and anti-inflammatory study of fermented Dendropanax morbifera Léveille leaf extract.

BACKGROUND: Medicinal plants represent a source of new drugs for the prevention and treatment of infectious diseases. Dendropanax morbifera Léveille is an economically and medicinally important subtropical tree that has various biological activities. However, its ability to affect immune responses in vivo is unknown. Hence, this study was designed to examine the immunomodulatory activity of fermented D. morbifera extract in BALB/c mice. METHODS: five-week-old female BALB/c mice were arranged in six groups and kept under a standard laboratory condition. Splenocyte counts were determined using the trypan blue dye exclusion method, and splenic lymphocyte proliferation was determined using concanavalin A and lipopolysaccharide (LPS). Flow cytometric analysis was performed to phenotype T-lymphocytes. Next, cytokine and immunoglobulin quantitation was performed using sandwich ELISA. RESULTS: The results showed an increase in spleen cells by 71 and 67% in mice treated with 125 and 250 mg/kg of D. morbifera, respectively. In addition, splenocyte proliferation was increased 58.7% in response to concanavalin A treatment, while LPS treatment induced a 73.3% increase in mice treated with 125 mg/kg. T-cell phenotypic analysis indicated that D. morbifera-treated groups showed higher CD8a+, CD11b and CD3+ T-cell expression. However, the treatment groups showed suppression of IL-1α, Il-1ß and IL-4. In addition, the IgG super-family was downregulated in a dose-dependent manner by 4.5% up to 43.7%. CONCLUSIONS: Taken together, we show that D. morbifera increases the number and proliferation of T- and B-lymphocytes. Moreover, these effects may play a role in boosting non-specific immunity, while suppressing proinflammatory cytokines and immunoglobulins after a single antigen exposure.

Enhancement of Lipid Metabolism and Hepatic Stability in Fat-Induced Obese Mice by Fermented Cucurbita moschata Extract.

The aim of this study was to evaluate the potentials of fermented Cucurbita moschata extract (FCME) in the treatment of obesity and nonalcoholic fatty liver disease (NAFLD). Five-week-old male C57BL/6 mice were assigned to 6 groups and treated for 8 weeks by feeding the normal diet (ND) and high fat diet (HFD) with and without FCME. Changes in body weight gain and consumption of feed and water were recorded. Major organs, adipose tissues, and blood samples were collected after the experimental period. The serum lipid profile, histological features of liver and adipose tissues, and mRNA expression of different adipogenic/lipogenic genes from liver tissue were evaluated. The supplementation of FCME in HFD significantly prevented HFD-induced increment of bodyweight. The adipose tissue mass, liver enzymes, and plasma lipids were also reduced significantly (p < 0.05) by the consumption of FCME. The mRNA expressions of adipogenic/lipogenic genes (PPARγ, C/EBPα, C/EBP ß , C/EBPγ, and SREBP-1C) in FCME-treated obese mice were considerably (p < 0.05) suppressed. FCME showed its antiobesity potential by suppressing the body weight gain and by modulating the plasma lipids and liver enzymes through the regulation of adipogenic/lipogenic transcriptional factors. Fermented Cucurbita moschata could be an opportunistic agent in controlling obesity and fatty liver changes.

The protective effect of fermented Curcuma longa L. on memory dysfunction in oxidative stress-induced C6 gliomal cells, proinflammatory-activated BV2 microglial cells, and scopolamine-induced amnesia model in mice.

BACKGROUND: Curcuma longa L. is a well-known medicinal plant that has been used for its anti-cancer, neuroprotective, and hepatoprotective effects. However, the neuroprotective effect of fermented C. longa (FCL) has not been reported. Therefore, in this study, the effectiveness of FCL for the regulation of memory dysfunction was investigated in two brain cell lines (rat glioma C6 and murine microglia BV2) and scopolamine-treated mice. METHODS: C. longa powder was fermented by 5% Lactobacillus plantarum K154 containing 2% (w/v) yeast extract at 30 °C for 72 h followed by sterilization at 121 °C for 15 min. The protective effects of fermented C. longa (FCL) on oxidative stress induced cell death were analyzed by MTT assay in C6 cells. The anti-inflammatory effects of FCL were investigated by measuring the production of nitric oxide (NO) and prostaglandin E2 (PGE2) as well as the expression levels of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated BV2 cells. The step-through passive avoidance test, Morris water maze test, acetylcholinesterase (AChE) activity, and expression of cAMP response element-binding protein (CREB) and brain-derived neurotropic factor (BDNF) were employed to determine the effects of FCL on scopolamine-induced memory deficit in mice. The contents of curcuminoids were analyzed through LC/MS. RESULTS: Pretreatment with FCL effectively prevented the cell death induced by oxidative stress in C6 cells. Moreover, FCL inhibited the production NO and PGE2 via the inhibition of iNOS and COX-2 expression in BV2 cells. FCL significantly attenuated scopolamine-induced memory impairment in mice and prevented scopolamine-induced AChE activity in the hippocampus. Additionally, FCL reversed the reduction of CREB and BDNF expression. The curcuminoids content in FCL was 1.44%. CONCLUSION: FCL pretreatment could alleviate scopolamine-induced memory impairment in mice, as well as oxidative stress and inflammation in C6 and BV2 cells, respectively. Thus, FCL might be a useful material for preventing impairment of learning and memory.

Defatted safflower seed extract inhibits adipogenesis in 3T3-L1 preadipocytes and improves lipid profiles in C57BL/6J ob/ob mice fed a high-fat diet.

In the present study, we hypothesized that defatted safflower seed which is known to be rich in polyphenols might influence adipogenesis and obesity-related disorders, and therefore the anti-adipogenic and hypolipidemic effects of ethanol extract from defatted safflower (Cathamus tinctorius L.) seeds (CSE) were investigated both in cultured 3T3-L1 preadipocytes and in C57BL/6J ob/ob mice fed a high-fat diet. CSE inhibited adipocyte differentiation of 3T3-L1 preadipocytes and decreased expression of the adipogenic transcription factors, SREBP1c and PPARγ, and their target genes. Six-week-old obese (ob/ob) mice were fed a high-fat diet and treated with CSE (50 or 100 mg/kg/day) by oral gavage for 6 weeks. Body fat mass (epididymal and perirenal white adipose tissues) in the CSE-treated groups was significantly lower than that in the high-fat diet control (HFD) group, whereas average daily food intake was not significantly different among the groups. Plasma and hepatic triglyceride levels and plasma low-density lipoprotein cholesterol level were also significantly lower in the CSE groups compared to the HFD group. These results suggest that CSE which decreases body fat mass and improves lipid profiles in plasma and liver, represents a potential treatment option for obesity and associated metabolic disorders, including hyperlipidemia.

Effect of Ganoderma applanatum mycelium extract on the inhibition of adipogenesis in 3T3-L1 adipocytes.

Ganoderma applanatum (GA) and related fungal species have been used for over 2000 years in China to prevent and treat various human diseases. However, there is no critical research evaluating the functionality of GA grown using submerged culture technology. This study aimed to evaluate the effects of submerged culture GA mycelium (GAM) and its active components (protocatechualdehyde [PCA]) on preadipocyte differentiation of 3T3-L1 cells. Mouse-derived preadipocyte 3T3-L1 cells were treated with differentiation inducers in the presence or absence of GAM extracts. We determined triglyceride accumulations, glycerol-3-phosphate dehydrogenase (GPDH) activities, and differentiation makers. PCA, the active component of GAM extract, was also used to treat 3T3-L1 cells. The MTT assay showed that the GAM extract (0.01-1 mg/mL) was not toxic to 3T3-L1 preadipocyte. Treatment of cells with GAM extracts and its active components significantly decreased the GPDH activity and lipid accumulation, a marker of adipogenesis, in a dose-dependent manner. Western blot analysis results showed that the protein expression levels of peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein 1 (SREBP1) were inhibited by the GAM extract. In addition, adipogenic-specific genes such as perilipin, fatty acid synthase (FAS), fatty acid transport protein 1 (FATP1), and fatty acid-binding protein 4 (FABP4) decreased in a dose-dependent manner. Quantitative high-performance liquid chromatography analysis showed that the GAM extract contained 1.14 mg/g PCA. GAM extracts suppressed differentiation of 3T3-L1 preadipocytes, in part, through altered regulation of PPARγ, C/EBPα, and SREBP1. These results suggest that GAM extracts and PCA may suppress adipogenesis by inhibiting differentiation of preadipocytes.

Hepatoprotective effects of fermented field water-dropwort (Oenanthe javanica) extract and its major constituents.

Dropwort (Oenanthe javanica) has been used for many years for the treatment of inflammatory conditions, including hepatitis. We investigated the protective effects of fermented field water-dropwort extract (FDE) on tert-butyl hydroperoxide (t-BHP)-induced hepatotoxicity in HepG2 cells and carbon tetrachloride (CCl4)-induced liver damage in rats. Pretreatment with FDE prior to the t-BHP treatment of HepG2 cells inhibited cell death and lactate dehydrogenase (LDH) leakage in a dose-dependent manner. In addition FDE significantly prevented the increase of hepatic enzyme markers (ALT, AST) in vivo. Moreover, FDE administration for 7 days significantly affected CYP2E1, CYP4A2, and PPARγ gene expressions. CYP2E1 and CYP4A2 gene expression in the liver, increased 2 and 22-fold by CCl4 administration, respectively, was attenuated to normal levels by pretreatment with FDE. PPARγ gene expression, completely blocked by CCl4 treatment, was increased by FDE pretreatment compared to normal control group. Histopathological examination of the livers also revealed that FDE reduced the incidence of liver lesions. Caffeic acid and chlorogenic acid were identified as major constituents of FDE. These results demonstrate the protective effects of FDE against hepatocytotoxicity induced by CCl4 and t-BHP in rats and HepG2 cells, thus indicating the potential of FDE as a therapeutic for acute liver diseases.

Inhibitory effects of actinidiamide from Actinidia polygama on allergy and inflammation.

Actinidia polygama Max. was subjected to supercritical fluid extraction (SFE), and the resulting ethanol extract of marc (SFEM) was subjected to sequential fractionation with various solvents. Each extract and fraction was assayed for anti-inflammatory effect. The ethyl acetate fraction (EtOAc) contained the highest level (70.8% inhibition) of anti-inflammatory activity. In order to identify the active constituents, the EtOAc fraction was further fractionated by silica gel and ODS column chromatography. By activity-guided fractionation, an active ceramide was identified as the anti-inflammatory component, and its structure was determined by NMR and MS analysis. The novel ceramide was named actinidiamide, and was found significantly to inhibit nitric oxide (NO) production (30.6% inhibition at 1 µg/mL) in lipopolysaccaride (LPS)-stimulated RAW 264.7 cells and ß-hexosaminidase release (91.8% inhibition at 1 µg/mL) in IgE-sensitized RBL-2H3 cells. Thus the presence of actinidiamide conveys allergy and inflammation treatment ability to A. polygama.

Antimicrobial effect and resistant regulation of Glycyrrhiza uralensis on methicillin-resistant Staphylococcus aureus.

In the present study, we investigated antimicrobial activity of Glycyrrhiza uralensis against various strains of methicillin-resistant Staphylococcus aureus (MRSA) (KCCM 11812, 40510, 40512). Glycyrrhiza uralensis was extracted by 80% MeOH and fractionated by organic solutions. The extract and fractions showed antimicrobial activity against standard S. aureus as well as MRSA. In the minimum inhibitory concentration test, G. uralensis showed 0.25 mg mL(-1) in hexane fraction and 0.10-0.12 mg mL(-1) in chloroform fraction. Especially, chloroform fraction showed 2.5 times higher antimicrobial activity than penicillin. Furthermore, chloroform fraction correlated with MRSA gene expression (MecA, MecI, MecRI, FemA). These results suggest that G. uralensis may have potent antimicrobial activity and thus, this medicinal herb can be a suitable phytotherapeutic agent for treating MRSA infections.

Comparative immunomodulating activities of polysaccharides isolated from Phellinus spp. on cell-mediated immunity.

The comparison of potent immunomodulating activities of hot-aqueous polysaccharides isolated from Phellinus spp., P. linteus (PL), P. baumii (PB) and P. gilvus (PG), on cellular immunity were investigated. The results showed that all three kinds of Phellinus spp. could stimulate the proliferation of murine splenocytes, and PG has more powerful stimulating activity than other Phellinus spp. Furthermore, PL and PB significantly increased the proliferation of the mixed splenocytes in a concentration-response manner, and the stimulating effect of PL was significantly higher than that of PB at all concentrations used in the present study, but no stimulating effect was found with the addition of PG. The phagocytosis of both peritoneal macrophage and RAW264.7 cells was also increased in the presence of PG, PB or PL, and the stimulating activity of PG was higher than that of PB or PL at all concentrations tested.

Production of fibrinolytic enzyme from soybean grits fermented by Bacillus firmus NA-1.

Bacillus firmus NA-1 producing fibrinolytic enzyme was isolated from Japanese traditional fermented soybean (Natto). Seed starter was cultured in 5% soy milk prepared with micronized soybean powder. To optimize the production of fibrinolytic enzyme, the soybean grits were mixed with 1 volume of water and sterilized at 121 degrees C for 5 minutes, and then used as the medium for solid-state fermentation at 42 degrees C. The fibrinolytic enzyme activity of the fermented grits was 1,120 plasmin units/100 g (wet weight) after fermentation for 24 hours. The water extract of the fermented grits showed the highest viscosity after fermentation for 12 hours. However, the tyrosine content was the highest (962 mg%) after fermentation for 60 hours. The color of raw soybean grit was affected by heat treatment. The activity of fibrinolytic enzyme was stable after freezing-drying, but was completely destroyed by heating at 70 degrees C for 10 minutes. The color of soybean grit was greatly darkened by increasing fermentation time. Soybean grits were completely converted into valuable functional ingredients containing fibrinolytic enzyme, peptide, and mucilage by the solid-state fermentation.

A new in vitro assay of cholesterol adsorption by food and microbial polysaccharides.

The adsorption of total cholesterol by polysaccharides was measured in vitro by enzymatic reactions, including the polysaccharide precipitation procedure. Total cholesterol adsorption capacities, in a mixture of polysaccharide and total cholesterol, were compared for apple pectin, gelrite gellan gum, xanthan gum, high-methoxyl pectin, citrus pectin, high-viscous alginate, low-viscous alginate, dextran, and zooglan. Acidic polysaccharides such as pectins, alginate, and xanthan gum at concentrations of 0.1% (wt/vol) were able to adsorb over 90% of the total cholesterol when dissolved in distilled water, sodium acetate buffer (pH 4.6), or sodium phosphate buffer (pH 7.0). However, total cholesterol adsorptions by gellan and zooglan were dependent upon the salt concentration and pH value, which decreased cholesterol adsorption in the following order by degree: distilled water, acidic pH, and alkaline pH. In particular, total cholesterol adsorption of zooglan was greatly decreased by the addition of sodium chloride. With 0.1% (wt/vol) polysaccharide dissolved in distilled water, the adsorption capacities of alginate, pectins, gellan gum, xanthan gum, and zooglan were 2.9, 2.88, 2.5, 2.9, and 2.4 mg/dL, respectively. However, 0.2% of zooglan was able to completely adsorb the cholesterol (3 mg/dL), whereas dextran did not adsorb cholesterol at all, producing no precipitate with hexadecyltrimethylammonium bromide.