Citrus depressa Hayata peel ameliorates nonalcoholic fatty liver and modulates the activity of hepatic drug-metabolizing enzymes and transporters in rats fed a high-fat diet.

Citrus depressa Hayata is a small, green citrus fruit native to Taiwan and Japan. Citrus peel contains polymethoxylated flavones, including nobiletin and tangeretin, and may exhibit strong antioxidant and anti-inflammatory activities. A preliminary study revealed that Citrus depressa Hayata peel (CDHP) ethanolic extract reduced fat accumulation and the concentration of reactive oxygen species in human HepG2 cells exposed to oleic acid. The effects of CDHP on the activity of hepatic drug-metabolizing enzymes and membrane transporters in high-fat (HF) diet-induced fatty liver were investigated. Male rats were fed a low-fat diet, a HF diet, and a HF diet containing 4% CDHP for 11 weeks. The low-fat and HF diet respectively contained 13.5% and 38.1% of daily total calories from dietary fat. CDHP supplementation reduced the HF diet-induced accumulation of triglycerides in the liver and lowered hepatic fatty acid synthase activity. Higher faecal excretions of cholesterol, triglycerides, and total bile acids were observed after CDHP treatment. CDHP lowered the HF diet-induced increase in the mRNA expressions of nuclear factor erythroid 2-related factor 2, aryl hydrocarbon receptor, pregnane X receptor, and peroxisome proliferator-activated receptor-α and the activities of cytochrome P-450 (CYP)1A1, 1A2, 2B, and 2E1. However, increased hepatic CYP3A activity was observed in rats fed the HF diet containing CDHP. A higher hepatic multidrug resistance-associated protein 2 level was observed after CDHP treatment. After CDHP administration (1 g per kg body weight) for 1 h, nobiletin was found in plasma and various tissues and was abundant in the liver. An in vitro study revealed that the activity of various CYP enzymes in liver microsomes was inhibited by CDHP ethanolic extract and nobiletin, with IC50 values ranging from 18.5 to 54.4 µg ml-1 and from 13.0 to 33.2 µM, respectively. The results of this study suggest that CDHP might reduce hepatic steatosis and alter drug-metabolizing enzymes and transporters in HF diet-induced nonalcoholic fatty liver diseases.

Artichoke leaf extract supplementation lowers hepatic oxidative stress and inflammation and increases multidrug resistance-associated protein 2 in mice fed a high-fat and high-cholesterol diet.

Artichoke (Cynara scolymus) leaf extract (ALE) contains many phytonutrients that may have antioxidant and anti-inflammation activities against many diseases including liver damage. To investigate the protective effects of ALE on high-fat and high-cholesterol (HFHC) diet-induced steatohepatitis and liver damage in mice, twenty-four female mice were fed an HFHC diet without or with 0.5% and 1% ALE supplementation for 6 weeks. The antioxidant and anti-inflammation activities and histological changes in the liver after ALE treatment were evaluated. The results show that ALE treatment reduced the HFHC diet-induced elevation of liver damage, as indicated by an increased alanine aminotransferase activity in plasma and perivenular inflammatory infiltrates in the liver. In addition, ALE ameliorated HFHC diet-induced depletion of hepatic glutathione (GSH) and elevations of plasma total cholesterol, triglyceride and hepatic triglyceride. ALE suppressed HFHC diet-induced accumulation of cholesterol precursors, including squalene and desmosterol in the liver. Higher hepatic GSH contents and activities of GSH-related enzymes were observed after ALE treatment. Higher expressions of nuclear factor erythroid 2-related factor 2 and heme oxygenase-1 (HO-1) were induced by the HFHC diet; however, ALE treatment reduced HO-1 expression. The NOD-like receptor protein 3, caspase-1, and interleukin-1ß protein and mRNA levels were reduced in the liver by ALE. A higher multidrug resistance-associated protein 2 expression in the liver was found after ALE treatment. These results suggest that ALE may ameliorate oxidative stress, inflammation and lipid metabolism disorder in HFHC diet-induced steatohepatitis and liver damage.

Qing-Yu-Mu, an Herbal Formula, Reduces Hepatic Oxidative Stress in Rats Fed a High-Frying Oil Diet and Ameliorates Carbon Tetrachloride-Induced Liver Injury.

Qing-Yu-Mu (QYM) is an herbal formula used to prevent and treat liver disease in Taiwan. In this study, the hepatoprotective effects of QYM were evaluated in two experimental models. First, rats were fed a high-frying oil (FO) diet containing 1.25% QYM for 5 weeks to investigate effects of QYM on hepatic oxidative stress and antioxidant enzyme activities. Then, protective effects of QYM on carbon tetrachloride (CCl4)-induced chronic liver injury were evaluated. Results show that QYM treatment reduced FO diet-induced hepatic lipid peroxidation and reactive oxygen species levels and increased glutathione (GSH) S-transferase activity. A higher reduced GSH/oxidized GSH (GSSG) ratio was observed after QYM treatment. Furthermore, QYM ameliorated CCl4-induced liver injury by reducing the activity of plasma alanine aminotransferase and histological lesions in the liver. QYM also increased the level of hepatic GSH and activities of GSH peroxidase and superoxide dismutase. Finally, chlorogenic acid, chrysophanol, and apigenin were found to be present in relative abundance in QYM. Results show that QYM may exhibit a hepatoprotective effect by reducing oxidative stress and increasing antioxidant activity in the liver.

A Diterpenoid, 14-Deoxy-11, 12-Didehydroandrographolide, in Andrographis paniculata Reduces Steatohepatitis and Liver Injury in Mice Fed a High-Fat and High-Cholesterol Diet.

14-Deoxy-11,12-didehydroandrographolide (deAND), a diterpenoid in Andrographis paniculata (Burm. f.) Nees, acts as a bioactive phytonutrient that can treat many diseases. To investigate the protective effects of deAND on reducing fatty liver disease, male mice were fed a high-fat and high-cholesterol (HFHC) diet without or with 0.05% and 0.1% deAND supplementation. Cholesterol accumulation, antioxidant, and anti-inflammatory activities in liver and liver injury were evaluated after deAND treatment. The results show that deAND treatment for seven weeks reduced plasma alanine aminotransferase activity and lowered hepatic cholesterol accumulation, tumor nuclear factor-α, and histological lesions. The 0.1% deAND treatment reduced HFHC diet-induced apoptosis by lowering the caspase 3/pro-caspase 3 ratio. After 11 weeks of deAND treatment, increased NOD-like receptor protein 3 (NLRP3), capase-1, and interleukin-1ß protein levels in liver were suppressed by deAND treatment. In addition, nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA expression, heme oxygenase-1 protein expression, and the activities of glutathione peroxidase and glutathione reductase were increased in mice fed the HFHC diet. However, those activities of antioxidant enzymes or proteins were also upregulated by 0.1% deAND treatment. Furthermore, deAND treatment tended to lower hepatic lipid peroxides. Finally, deAND treatment reversed the depletion of hepatic glutamate level induced by the HFHC diet. These results indicate that deAND may ameliorate HFHC diet-induced steatohepatitis and liver injury by increasing antioxidant and anti-inflammatory activities.

Food-Drug Interaction between the Adlay Bran Oil and Drugs in Rats.

Adlay (Coix lachryma-jobi L. var. ma-yuen Stapf) contains various phytonutrients for treating many diseases in Asia. To investigate whether orally administered adlay bran oil (ABO) can cause drug interactions, the effects of ABO on the pharmacokinetics of five cytochrome P450 (CYP) probe drugs were evaluated. Rats were given a single oral dose (2.5 mL/kg BW) of ABO 1 h before administration of a drug cocktail either orally or intravenously, and blood was collected at various time points. A single oral dose of ABO administration did not affect the pharmacokinetics of five probe drugs when given as a drug cocktail intravenously. However, ABO increased plasma theophylline (+28.4%), dextromethorphan (+48.7%), and diltiazem (+46.7%) when co-administered an oral drug cocktail. After 7 days of feeding with an ABO-containing diet, plasma concentrations of theophylline (+45.4%) and chlorzoxazone (+53.6%) were increased after the oral administration of the drug cocktail. The major CYP enzyme activities in the liver and intestinal tract were not affected by ABO treatment. Results from this study indicate that a single oral dose or short-term administration of ABO may increase plasma drug concentrations when ABO is given concomitantly with drugs. ABO is likely to enhance intestinal drug absorption. Therefore, caution is needed to avoid food-drug interactions between ABO and co-administered drugs.

Epigallocatechin-3-Gallate Reduces Hepatic Oxidative Stress and Lowers CYP-Mediated Bioactivation and Toxicity of Acetaminophen in Rats.

Epigallocatechin-3-gallate (EGCG) is the most abundant polyphenol in green tea. To investigate the effects of dietary EGCG on oxidative stress and the metabolism and toxicity of acetaminophen in the liver, rats were fed diets with (0.54%) or without EGCG supplementation for four weeks and were then injected intraperitoneally with acetaminophen (1 g/kg). The results showed that EGCG lowered hepatic oxidative stress and cytochrome P450 (CYP) 1A2, 2E1, and 3A, and UDP-glucurosyltransferase activities prior to acetaminophen injection. After acetaminophen challenge, the elevations in plasma alanine aminotransferase activity and histological changes in the liver were ameliorated by EGCG treatment. EGCG reduced acetaminophen-induced apoptosis by lowering the Bax/Bcl2 ratio in the liver. EGCG mildly increased autophagy by increasing the LC3B II/I ratio. Lower hepatic acetaminophen-glutathione and acetaminophen-protein adducts contents were observed after EGCG treatment. EGCG increased glutathione peroxidase and NAD(P)H quinone 1 oxidoreductase activities and reduced organic anion-transporting polypeptides 1a1 expression in the liver after acetaminophen treatment. Our results indicate that EGCG may reduce oxidative stress and lower the metabolism and toxicity of acetaminophen. The reductions in CYP-mediated acetaminophen bioactivation and uptake transporter, as well as enhanced antioxidant enzyme activity, may limit the accumulation of toxic products in the liver and thus lower hepatotoxicity.

Bioavailability of the diterpenoid 14-deoxy-11,12-didehydroandrographolide in rats and up-regulation of hepatic drug-metabolizing enzyme and drug transporter expression.

BACKGROUND: 14-Deoxy-11,12-didehydroandrographolide (deAND) is the second most abundant diterpenoid in Andrographis paniculata (Burm. f.) Nees, a traditional medicine used in Asia. To date, the biological activity of deAND has not been clearly investigated. PURPOSE: In this study, we intended to examine the modulatory effect of deAND on hepatic drug metabolism as well as its bioavailability. STUDY DESIGN: deAND prepared from A. paniculata was orally given to Sprague-Dawley rats and changes in plasma deNAD were determined by HPLC-MS. Modulation of deAND on drug-metabolizing enzyme and drug transporter expression as well as the possible mechanism involved was examined in primary rat hepatocytes. RESULTS: After a single oral administration of 50 mg/kg deAND to rats, the maximum plasma concentration (Cmax), time to reach the Cmax, area under the curve (AUC0-24h), mean retention time, and half-life (t1/2) of deAND were 2.65 ± 0.68 µg/ml, 0.29 ± 0.15 h, 6.30 ± 1.66 µg/ml•h, 5.55 ± 2.52 h, and 3.56 ± 1.05 h, respectively. The oral bioavailability was 3.42%. In primary rat hepatocytes treated with up to 10 µM deAND, a dose-dependent increase was noted in the expression of cytochrome P450 (CYP) 1A1/2, CYP2C6, and CYP3A1/2; UDP-glucuronosyltransferase (UGT) 1A1, NAD(P)H:quinone oxidoreductase (NQO1), π form of GSH S-transferase (GSTP), multidrug resistance-associated protein 2, p-glycoprotein, and organic anion transporter protein 2B1. Immunoblotting assay and EMSA revealed that deAND increases the nuclear translocation and DNA binding activity of aryl hydrocarbon receptor (AhR), pregnane X receptor (PXR), and nuclear factor erythroid-derived 2-related factor 2 (Nrf2). Knockdown of AhR and Nrf2 expression abolished deAND induction of CYP isozymes and UGT1A1, NQO1, and GSTP expression, respectively. CONCLUSION: These results indicate that deAND quickly passes through enterocytes in rats and effectively up-regulates hepatic drug-metabolizing enzyme and drug transporter expression in an AhR-, PXR-, and Nrf2-dependent manner.

Freshwater clam extract reduces liver injury by lowering cholesterol accumulation, improving dysregulated cholesterol synthesis and alleviating inflammation in high-fat, high-cholesterol and cholic acid diet-induced steatohepatitis in mice.

Freshwater clam (Corbicula fluminea) is a traditional liver-protective food in Asia. Recent studies have renewed attention on high cholesterol accumulation and dysregulated cholesterol synthesis in the liver as a critical factor in the progression of nonalcoholic fatty liver disease (NAFLD) to nonalcoholic steatohepatitis (NASH). In this study, we investigated the protective effects of freshwater clam extract (FCE) and its fat fraction (FCE oil) on high-fat, high-cholesterol and cholic acid (HFHC) diet-induced lean steatohepatitis in mice. Mice were fed a HFHC diet containing FCE or FCE oil for 6 weeks. FCE, but not FCE oil, feeding reduced liver injury as indicated by decreased plasma alanine aminotransferase activity. Liver total cholesterol accumulation was reduced after FCE and FCE oil treatment. Accumulation of squalene and desmosterol, the precursors of cholesterol, in the liver was reduced by FCE but not by FCE oil. The caspase-1 (p10) and interleukin (IL)-1ß (p17) protein expressions in the liver were suppressed by both FCE and FCE oil. Therefore, FCE may act as functional food that can reduce steatohepatitis and liver injury by reducing cholesterol accumulation, improving dysregulated cholesterol synthesis and attenuating inflammation.

Effects of lemongrass oil and citral on hepatic drug-metabolizing enzymes, oxidative stress, and acetaminophen toxicity in rats.

The essential oil from a lemongrass variety of Cymbopogon flexuosus [lemongrass oil (LO)] is used in various food and aroma industry products and exhibits biological activities, such as anticancer and antimicrobial activities. To investigate the effects of 200 LO (200 mg/kg) and 400 LO (400 mg/kg) and its major component, citral (240 mg/kg), on drug-metabolizing enzymes, oxidative stress, and acetaminophen toxicity in the liver, male Sprague-Dawley rats were fed a pelleted diet and administered LO or citral by gavage for 2 weeks. After 2 weeks of feeding, the effects of LO and citral on the metabolism and toxicity of acetaminophen were determined. The results showed that rats treated with 400 LO or citral had significantly reduced hepatic testosterone 6ß-hydroxylation and ethoxyresorufin O-deethylation activities. In addition, NAD(P)H:quinone oxidoreductase 1 activity was significantly increased by citral, and Uridine 5'-diphospho (UDP) glucurosyltransferase activity was significantly increased by 400 LO in the rat liver. Treatment with 400 LO or citral reduced lipid peroxidation and reactive oxygen species levels in the liver. After acetaminophen treatment, however, LO and citral treatment resulted in little or no change in plasma alanine aminotransferase activity and acetaminophen-protein adducts content in the liver. Our results indicate that LO and citral may change the activities of drug-metabolizing enzymes and reduce oxidative stress in the liver. However, LO and citral may not affect the detoxification of acetaminophen.

Red algae (Gelidium amansii) hot-water extract ameliorates lipid metabolism in hamsters fed a high-fat diet.

The purpose of this study was to investigate the effects of Gelidium amansii (GA) hot-water extracts (GHE) on lipid metabolism in hamsters. Six-week-old male Syrian hamsters were used as the experimental animals. Hamsters were divided into four groups: (1) control diet group (CON); (2) high-fat diet group (HF); (3) HF with GHE diet group (HF + GHE); (4) HF with probucol diet group (HF + PO). All groups were fed the experimental diets and drinking water ad libitum for 6 weeks. The results showed that GHE significantly decreased body weight, liver weight, and adipose tissue (perirenal and paraepididymal) weight. The HF diet induced an increase in plasma triacylglycerol (TG), total cholesterol (TC), low-density lipoprotein cholesterol and very-low-density lipoprotein cholesterol levels. However, GHE supplementation reversed the increase of plasma lipids caused by the HF diet. In addition, GHE increased fecal cholesterol, TG and bile acid excretion. Lower hepatic TC and TG levels were found with GHE treatment. GHE reduced hepatic sterol regulatory element-binding proteins (SREBP) including SREBP 1 and SREBP 2 protein expressions. The phosphorylation of adenosine monophosphate (AMP)-activated protein kinase (AMPK) protein expression in hamsters was decreased by the HF diet; however, GHE supplementation increased the phosphorylation of AMPK protein expression. Our results suggest that GHE may ameliorate lipid metabolism in hamsters fed a HF diet.

Platycodon grandiflorum (PG) reverses angiotensin II-induced apoptosis by repressing IGF-IIR expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Platycodon grandiflorum (PG) is a Chinese medical plant used for decades as a traditional prescription to eliminate phlegm, relieve cough, reduce inflammation and lower blood pressure. PG also has a significant effect on the cardiovascular systems. MATERIALS AND METHODS: The aqueous extract of Platycodon grandiflorum (JACQ.) A. DC. root was screened for inhibiting Ang II-induced IGF-IIR activation and apoptosis pathway in H9c2 cardiomyocytes. The effects were also studied in spontaneously hypertensive rats (five groups, n=5) using low and high doses of PG for 50 days. The Ang II-induced IGF-IIR activation was analyzed by luciferase reporter, RT-PCR, western blot and surface IGF-IIR expression assay. Furthermore, the major active constituent of PG was carried out by high performance liquid chromatography-mass spectrometry (HPLC-MS). RESULTS: Our results indicate that a crude extract of PG significantly suppresses the Ang II-induced IGF-IIR signaling pathway to prevent cardiomyocyte apoptosis. PG extract inhibits Ang II-mediated JNK activation and SIRT1 degradation to reduce IGF-IIR activity. Moreover, PG maintains SIRT1 stability to enhance HSF1-mediated IGF-IIR suppression, which prevents cardiomyocyte apoptosis. In animal models, the administration of PG markedly reduced this apoptotic pathway in the heart of SHRs. CONCLUSION: Taken together, PG may be considered as an effective treatment for cardiac diseases in hypertensive patients.

Docosahexaenoic Acid Downregulates EGF-Induced Urokinase Plasminogen Activator and Matrix Metalloproteinase 9 Expression by Inactivating EGFR/ErbB2 Signaling in SK-BR3 Breast Cancer Cells.

Urokinase plasminogen activator (uPA) and matrix metalloproteinase 9 (MMP-9) play crucial roles in tumor metastasis. Despite the well-known anticancer role of docosa-hexaenoic acid (DHA), its specific effect on ErbB2-mediated breast cancer metastasis is not fully clarified. In this study, we investigated the effect of DHA on epidermal growth factor (EGF)-induced uPA and MMP-9 activity, expression and cell invasion in SK-BR3 breast cancer cells and the possible mechanisms involved. The results showed that EGF (40 ng/ml) induced uPA and MMP-9 mRNA and protein expression, enzyme activity, and 100 µM DHA significantly inhibited EGF-induced uPA and MMP-9 mRNA, protein expression, enzyme activity, cell migration, and cell invasion. EGF increased protein expression and phosphorylation of EGF receptor (EGFR) and ErbB2 as well as of JNK2, ERK1/2, and Akt, and these changes were attenuated by DHA pretreatment. AG1478, an inhibitor of EGFR, also attenuated EGF-induced activation of EGFR, JNK2, ERK1/2, and Akt. Knocked down ErbB2 expression resulted in a similar inhibition of uPA and MMP-9 expression as noted by DHA and AG1478. Taken together, these results suggest that suppression of EGF-induced metastasis by DHA is likely through an inhibition of EGFR and ErbB2 protein expression and the downstream target uPA and MMP-9 activation in SK-BR3 human breast cancer cells.

Effect of commercially available green and black tea beverages on drug-metabolizing enzymes and oxidative stress in Wistar rats.

The effect of commercially available green tea (GT) and black tea (BT) drinks on drug metabolizing enzymes (DME) and oxidative stress in rats was investigated. Male Wistar rats were fed a laboratory chow diet and GT or BT drink for 5 weeks. Control rats received de-ionized water instead of the tea drinks. Rats received the GT and BT drinks treatment for 5 weeks showed a significant increase in hepatic microsomal cytochrome P450 (CYP) 1A1 and CYP1A2, and a significant decrease in CYP2C, CYP2E1 and CYP3A enzyme activities. Results of immunoblot analyses of enzyme protein contents showed the same trend with enzyme activity. Significant increase in UDP-glucuronosyltransferase activity and reduced glutathione content in liver and lungs were observed in rats treated with both tea drinks. A lower lipid peroxide level in lungs was observed in rats treated with GT drink. Electrophoretic mobility shift assay revealed that both tea drinks decreased pregnane X receptor binding to DNA and increased nuclear factor-erythroid 2 p45-related factor 2 binding to DNA. These results suggest that feeding of both tea drinks to rats modulated DME activities and reduced oxidative stress in liver and lungs. GT drink is more effective on reducing oxidative stress than BT drink.

Andrographis paniculata Extract and Andrographolide Modulate the Hepatic Drug Metabolism System and Plasma Tolbutamide Concentrations in Rats.

Andrographolide is the most abundant terpenoid of A. paniculata which is used in the treatment of diabetes. In this study, we investigated the effects of A. paniculata extract (APE) and andrographolide on the expression of drug-metabolizing enzymes in rat liver and determined whether modulation of these enzymes changed the pharmacokinetics of tolbutamide. Rats were intragastrically dosed with 2 g/kg/day APE or 50 mg/kg/day andrographolide for 5 days before a dose of 20 mg/kg tolbutamide was given. APE and andrographolide reduced the AUC0-12 h of tolbutamide by 37% and 18%, respectively, compared with that in controls. The protein and mRNA levels and enzyme activities of CYP2C6/11, CYP1A1/2, and CYP3A1/2 were increased by APE and andrographolide. To evaluate whether APE or andrographolide affected the hypoglycemic action of tolbutamide, high-fat diet-induced obese mice were used and treated in the same manner as the rats. APE and andrographolide increased CYP2C6/11 expression and decreased plasma tolbutamide levels. In a glucose tolerance test, however, the hypoglycemic effect of tolbutamide was not changed by APE or andrographolide. These results suggest that APE and andrographolide accelerate the metabolism rate of tolbutamide through increased expression and activity of drug-metabolizing enzymes. APE and andrographolide, however, do not impair the hypoglycemic effect of tolbutamide.

Indigofera suffruticosa Mill extracts up-regulate the expression of the π class of glutathione S-transferase and NAD(P)H: quinone oxidoreductase 1 in rat Clone 9 liver cells.

Because induction of phase II detoxification enzyme is important for chemoprevention, we study the effects of Indigofera suffruticosa Mill, a medicinal herb, on the expression of π class of glutathione S-transferase (GSTP) and NAD(P)H: quinone oxidoreductase 1 (NQO1) in rat Clone 9 liver cells. Both water and ethanolic extracts of I. suffruticosa significantly increased the expression and enzyme activities of GSTP and NQO1. I. suffruticosa extracts up-regulated GSTP promoter activity and the binding affinity of nuclear factor erythroid 2-related factor 2 (Nrf2) with the GSTP enhancer I oligonucleotide. Moreover, I. suffruticosa extracts increased nuclear Nrf2 accumulation as well as ARE transcriptional activity. The level of phospho-ERK was augmented by I. suffruticosa extracts, and the ERK inhibitor PD98059 abolished the I. suffruticosa extract-induced ERK activation and GSTP and NQO-1 expression. Moreover, I. suffruticosa extracts, especially the ethanolic extract increased the glutathione level in mouse liver and red blood cells as well as Clone 9 liver cells. The efficacy of I. suffruticosa extracts in induction of phase II detoxification enzymes and glutathione content implies that I. suffruticosa could be considered as a potential chemopreventive agent.

Suppressive effects of Indigofera suffruticosa Mill extracts on lipopolysaccharide-induced inflammatory responses in murine RAW 264.7 macrophages.

Indigofera suffruticosa Mill is used as an herbal medicine for the treatment of inflammation. The aim of this study is to assess the anti-inflammatory potency of I. suffruticosa and its likely molecular mechanisms of action in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. Both water and ethanolic extracts of I. suffruticosa significantly decreased LPS-induced nitric oxide (NO) as well as the expression of inducible nitric oxide synthase (iNOS), tumor necrosis factor-α, and pro-interleukin-1ß. Moreover, LPS-induced inhibitory factor-κB-α phosphorylation, nuclear factor-κB (NF-κB) nuclear protein-DNA binding affinity, and NF-κB reporter gene activity were dramatically inhibited by I. suffruticosa extracts. Exogenous addition of I. suffruticosa significantly induced heme oxygenase-1 (HO-1) expression, and the presence of HO-1 small interfering RNA partly reversed the inhibitory effects of I. suffruticosa on LPS-induced NO production and iNOS expression. Furthermore, I. suffruticosa induced HO-1 expression may be through activation of the ERK/nuclear factor E2-related factor 2 pathway. Eight phenolic compounds were found in the I. suffruticosa extracts, but salicylic acid was the only one detected in the plasma of mice fed with I. suffruticosa extracts. In summary, I. suffruticosa have a strong anti-inflammatory property that diminishes pro-inflammatory mediator expressions by lessening LPS-induced NF-κB activation and inducing HO-1 expression in macrophages.

Effects of Bu-Zhong-Yi-Qi-Tang on hepatic drug-metabolizing enzymes and plasma tolbutamide concentration in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Bu-Zhong-Yi-Qi-Tang (BT) is the dry powder derived from the aqueous extract of a mixture of 10 medicinal herbs. It is a traditional Chinese medicine being used for the treatment of various immune-related diseases. AIM OF THE STUDY: To investigate the effect of BT on hepatic drug-metabolizing enzymes and its effect on plasma concentrations of tolbutamide, a substrate of CYP2C, in rats. MATERIALS AND METHODS: EXP 1: Thirty-two male Wistar rats were divided into four groups. Rats were fed a control diet and a control diet containing 1, 2.5 and 5% (w/w) of BT, respectively, for eight weeks. The activities of the major CYP and Phase II conjugating enzymes in rat liver microsomes as well as the antioxidant system in rat liver were assessed. Exp 2: Male Wistar rats were fed a control diet or a control diet containing 2.5% of BT, respectively, for eight weeks. A single 20-mg/kg oral dose of tolbutamide was then administered to each rat. Plasma samples were collected from each rat at 0.5, 1, 2, 4 and 8h after dosing. The concentrations of tolbutamide and glucose level in plasma were determined by high-performance liquid chromatography-mass spectrometer (HPLC/MS) and enzymatic method, respectively. RESULTS: Significant decrease in microsomal CYP2C-catalyzed diclofenac 4-hydroxylation in the liver of rats fed the BT diet was observed. Increased UDP-glucuronosyltransferase (UGT) and glutathione S-transferase (GST) activities were also observed in the liver of rats fed the diet containing 2.5 and 5% of BT. Immunoblot analyses also showed decreases of CYP2C11 proteins in the liver of BT fed rats. In addition, rats fed the 2.5% BT diet for eight weeks had no effects on the disposition of tolbutamide and reduction of glucose level in plasma after orally administered of tolbutamide. CONCLUSIONS: Rats fed the BT diet for eight weeks may decrease CYP2C enzyme activity and protein expression and increase Phase II conjugating enzyme activities in liver. However, BT may not affect the disposition and efficacy of tolbutamide.

Chitosan reduces plasma adipocytokines and lipid accumulation in liver and adipose tissues and ameliorates insulin resistance in diabetic rats.

Chitosan is a natural product derived from chitin. To investigate the hypoglycemic and anti-obesity effects of chitosan, male Sprague-Dawley rats were divided into four groups: normal control, diabetic, and diabetic fed 5% or 7% chitosan. Diabetes was induced in rats by injecting streptozotocin/nicotinamide. After 10 weeks of feeding, the elevated plasma glucose, tumor necrosis factor-α, and interleukin-6 and lower adiponetin levels caused by diabetes were effectively reversed by chitosan treatment. In addition, 7% chitosan feeding also elevated plasma glucagon-like peptide-1 levels and lowered the insulin resistance index (homeostasis model assessment) in diabetic rats. Lower adipocyte granular intensities and higher lipolysis rates in adipose tissues were noted in the 7% chitosan group. Moreover, chitosan feeding reduced hepatic triglyceride and cholesterol contents and increased hepatic peroxisomal proliferator-activated receptor α expression in diabetic rats. Our results indicate that long-term administration of chitosan may reduce insulin resistance through suppression of lipid accumulation in liver and adipose tissues and amelioration of chronic inflammation in diabetic rats.

Induction of Nrf2-dependent Antioxidation and Protection Against Carbon Tetrachloride-induced Liver Damage by Andrographis Herba (chuan xin lián) Ethanolic Extract.

Andrographis paniculata is a traditional Chinese herb and displays diverse biological activities including antioxidation, anti-tumorigenesis, anti-virus, and anti-atherogenesis. In this study, we investigated the up-regulation of ethanolic extract of A. paniculata (APE) on the antioxidant defense in rat livers and whether this enhancement protected against carbon tetrachloride (CCl4)-induced liver damage. Male Sprague-Dawley rats were orally administered (i.g.) 0, 0.75, or 2 g/kg/d APE for 5 d. At d 6, rats were sacrificed and liver tissues were removed. Some animals (n=8) were intraperitoneally injected CCl4 (1 mL/kg, 50% in olive oil) and blood was drawn 24 h after CCl4 treatment. The results showed that APE increased hepatic glutathione (GSH) content and superoxide dismutase, GSH peroxidase, and GSH S-transferase activities in a dose-dependent manner (p < 0.05). Results of immunoblotting and RT-PCR revealed that rats treated with APE had higher glutamate cysteine ligase catalytic and modifier subunits, heme oxygenase 1, superoxide dismutase 1, and GSH S-transferase Ya and Yb protein and mRNA expression than those of control rats. Moreover, APE increased Nrf2 nuclear translocation and Nrf2 binding to DNA in rat liver. In the presence of CCl4, APE decreased hepatic thiobarbituric acid-reactive substances production and plasma aspartate aminotransferase and alanine aminotransferase activities. These results suggest that APE protection against CCl4 insult is attributed, at least in part, to its up-regulation of antioxidant defense in rat liver.

Development of a whole-cell screening system for evaluation of the human CYP1A2-mediated metabolism.

Cytochrome P450 1A2 (CYP1A2) is an important member of cytochrome P450 involved in drug metabolism. In this study, a cell line, Huh7-1A2-I-E, with high expression level of CYP1A2 is established based on Huh7 cells. To achieve this, we constructed a recombinant lentiviral vector, pLenti-1A2-I-E, containing a single promoter encoding CYP1A2 followed by an internal ribosome entry site (IRES) to permit the translation of enhanced green fluorescence protein (EGFP). Such a design has greatly facilitated the selection of stable cell lines because the translations of CYP1A2 and EGFP proteins would be based on a single bi-cistronic mRNA. The Huh7-1A2-I-E cells were evaluated as a cell-based model for identification of CYP1A2 inhibitors and for studies of cytotoxicity resulted from CYP-mediated drug metabolism. Treatment of Huh7-1A2-I-E cells and the Huh7-E control cells with aflatoxin B1 showed that cells with CYP1A2 expression are much more sensitive to aflatoxin B1 and the cellular toxicity of aflatoxin B1 in Huh7-1A2-I-E cells could be prevented by furafylline, a CYP1A2 inhibitor. A collection of approximately 200 drugs were screened using this system and results indicate that for most drugs the metabolism by CYP1A2 is unlikely to have made a major contribution to the in vitro cytotoxicity except for thimerosal and evoxine. Several previously unidentified CYP1A2 inhibitors such as evoxine and berberine were also identified in this study.