Pharmacological activities and adverse side effects of ginkgolic acids (GAs), major components in extracts from the leaves and seed coats of Ginkgo biloba L, have been intensively studied. However, there are few reports on their hepatotoxicity. In the present study, the metabolism and hepatotoxicity of GA (17 : 1), one of the most abundant components of GAs, were investigated. Kinetic analysis indicated that human and rat liver microsomes shared similar metabolic characteristics of GA (17 : 1) in phase I and II metabolisms. The drug-metabolizing enzymes involved in GA (17 : 1) metabolism were human CYP1A2, CYP3A4, UGT1A6, UGT1A9, and UGT2B15, which were confirmed with an inhibition study of human liver microsomes and recombinant enzymes. The MTT assays indicated that the cytotoxicity of GA (17 : 1) in HepG2 cells occurred in a time- and dose-dependent manner. Further investigation showed that GA (17 : 1) had less cytotoxicity in primary rat hepatocytes than in HepG2 cells and that the toxicity was enhanced through CYP1A- and CYP3A-mediated metabolism.
Animals ; Cells, Cultured ; Cytochrome P-450 CYP1A2 ; metabolism ; Cytochrome P-450 CYP3A ; metabolism ; Ginkgo biloba ; chemistry ; Glucuronosyltransferase ; metabolism ; Hepatocytes ; chemistry ; drug effects ; enzymology ; metabolism ; Humans ; Kinetics ; Liver ; chemistry ; drug effects ; enzymology ; metabolism ; Microsomes, Liver ; chemistry ; drug effects ; enzymology ; metabolism ; Plant Extracts ; chemistry ; metabolism ; toxicity ; Rats ; Rats, Sprague-Dawley ; Salicylates ; chemistry ; metabolism ; toxicity

To investigate the effect of Jiawei Foshou San and its various combined administration on hepatic P450 enzyme activity and hepatocyte morphology in rats. Rats were orally administered with drugs for four weeks and then sacrificed to prepare liver microsomes. The liver microsomes were incubated with the cocktail method; The metabolites were determined with the rapid liquid chromatography with tandem mass spectrometry (LC-MS/MS) to investigate the hepatocyte P450 enzyme activity. In addition, the hepatic pathological changes were observed by using the hematoxylin and eosin (HE) staining. Compared with the control group, the enzyme activity of CYP1A2 and CYP3A4 in the Jiawei Foshou san group showed a significant rise (P < 0.05); the enzyme activity of CYP1A2, CYP2C9, CYP2D6, CYP2E1 and CYP3A4 in the ferulic acid + ligustrazine group and the ligustrazine + tetrahydropalmatine group showed a significant rise (P < 0.05) ; the enzyme activity of CYP1A2, CYP2D6 and CYP2E1 in the ligustrazine group showed a significant rise (P < 0.05); the enzyme activity of CYP3A4 in the ferulic acid group showed a significant reduction (P < 0.05). After the administration with various drugs, the hepatocyte morphologies in the ferulic acid group and the ligustrazine group were normal. The pathological changes were observed in the tetrahydropalmatine group, such as unclear boundary of hepatic lobules, disordered hepatic cell arrangement, blurred edge, anisokaryosis and infiltration of inflammatory cells. The ferulic acid + tetrahydropalmatine group, the ligustrazine + tetrahydropalmatine group and the Jiawei Foshou San group also showed inflammatory infiltration, but with less pathological changes, particularly the Jiawei Foshou San group. The study result shows that Jiawei Foshou San can induce the enzyme activity of CYP1A2 and CYP3A4, and ligustrazine may be the effective substance for inducing CYP1A2. Its combination with ferulic acid and ligustrazine can significantly reduce the liver toxicity of tetrahydropalmatine.
Animals ; Cytochrome P-450 Enzyme System ; metabolism ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Hepatocytes ; drug effects ; enzymology ; Microsomes, Liver ; drug effects ; enzymology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo observe the effects of different therapeutic methods and the recipes of Chinese medicine (CM) on the activation of c-Jun N-terminal kinase (JNK) in Kupffer cells of rats with fatty liver disease and to explore the mechanisms of these therapeutic methods.

METHODSBy using a random number table, 98 rats were randomly divided into 7 groups: control group, model group, and 5 treatment groups, including soothing Liver (Gan) recipe group, invigorating Spleen (Pi) recipe group, dispelling dampness recipe group, promoting blood recipe group, and complex recipe group. Rats in the control group were fed with normal food and distilled water by gastric perfusion, while rats in the model group were fed with high-fat food and distilled spirits by gastric perfusion. Rats in the 5 treatment groups were fed with high-fat food and corresponding recipes by gastric perfusion. Twelve weeks later, all rats were sacrificed and liver tissues were stained for pathohistological observation. Kupffer cells were isolated from livers of rats to evaluate JNK and phospho-JNK expressions by Western blotting.

RESULTSThe grade of hepatic steatosis was higher in the model group than the control group (P<0.05). Compared with the model group, the grade of fatty degeneration in soothing Liver recipe group and invigorating Spleen recipe group were significantly ameliorated (P<0.05). Expressions of JNK and phospho-JNK in Kupffer cells were significantly higher in the model group than those in the control group (P<0.05, P<0.01). Compared with the model group, expressions of JNK in all treatment groups decreased, especially in invigorating Spleen recipe group and promoting blood recipe group (P<0.05). Compared with the model group, expressions of phospho-JNK in all treatment groups declined significantly (P<0.01), especially in soothing Live recipe group and invigorating Spleen recipe group.

CONCLUSIONSThe high expressions of JNK and phospho-JNK in Kupffer cells might play an important role in the pathogenesis of fatty liver disease in rats. The recipes of CM, especially invigorating Spleen recipe and soothing Liver recipe, might protect liver against injury by reducing the total JNK protein content and inhibiting the activation of JNK protein in Kupffer cells of fatty liver model rats, which showed beneficial effects on fatty liver disease.

Animals ; Cell Survival ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Enzyme Activation ; drug effects ; Fatty Liver ; enzymology ; pathology ; therapy ; Hepatocytes ; drug effects ; enzymology ; pathology ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Kupffer Cells ; drug effects ; enzymology ; pathology ; Liver ; drug effects ; enzymology ; pathology ; Phosphorylation ; drug effects ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the protective effect of Shaoganduogan (SGDG) on serum transaminase, liver pathology and hepatocyte mitochondria in rat with subacute liver injury induced by carbon tetrachloride.

METHODSubacute liver injury of rats were induced by carbon tetrachloride, and cured by different doses of SGDG through intragastric administration. The activity of serum ALT, AST, liver pathology and ultrastructure, activity of ATPase, SOD and content of MDA of hepatocyte mitochondria were observed.

RESULTSGDG can remarkably reduce the transaminase, alleviate the degeneration and necrosis of liver cells ,enhance activity of Na+ -K+ ATPase, Ca2+ ATPase, SOD, reduce content of MDA of mitochondria, alleviate ultrastructure change of mitochondria, reduce section area, perimeter equivalent diameter and average optical density perimeter of liver cells.

CONCLUSIONSGDG has obvious effect of liver protection, the mechanisms are related with alleviating mitochondria injury.

Animals ; Carbon Tetrachloride ; adverse effects ; Chemical and Drug Induced Liver Injury ; pathology ; Drugs, Chinese Herbal ; pharmacology ; Hepatocytes ; drug effects ; pathology ; Male ; Malondialdehyde ; metabolism ; Mitochondria ; drug effects ; enzymology ; metabolism ; ultrastructure ; Rats ; Rats, Sprague-Dawley

This study is to investigate the effects of aqueous extract of Schisandra chinensis Baill (WWZ), kadsurin, schisandrin A, schisandrin B and schisandrol B on rat hepatic CYP3A. Rats received a daily gavage of aqueous extract of WWZ for different times. The livers were harvested after gavage and subjected to microsome preparation. Microsomal CYP3A activity was determined by measuring the amount of the metabolite of testosterone (6 beta-hydroxytestosterone) with HPLC. Aqueous extract of WWZ, kadsurin and schisandrin A were incubated with microsomes obtained from rat. Microsomal CYP3A activity was determined by HPLC. Primary hepatocytes were separated and extracted from rat, then were treated with aqueous extract of WWZ, schisandrin A, schisandrin B and schisandrol B. Then, the expression of CYP3A1 mRNA was analyzed by RT-PCR. As for the in vivo assay, aqueous extract of WWZ significantly inhibited the enzyme activity of CYP3A after 12 h gavage. The inhibitory effect was converted to inductive effect after 3-day gavage. Aqueous extract of WWZ could induce the enzyme activity of CYP3A after 6-day gavage. Aqueous extract of WWZ and kadsurin showed a dose-dependent inhibition of CYP3A (IC50 of 487.8 microg mL(-1) and 6.2 micromol L(-1), separately). In rat primary hepatocytes, aqueous extract of WWZ (2.5 mg mL(-1)), schisandrin A (0.1 micromol L(-1)), schisandrin B (0.1 micromol L(-1)) and schisandrol B (10 micromol L(-1)) increased significantly the expression of CYP3A1 mRNA by 23%, 55%, 42% and 27%, respectively. Aqueous extract of WWZ could show dual effect on the enzyme activity of CYP3A in rat in vivo. Meanwhile, kadsurin showed a dose-dependent inhibition of the enzyme activity of hepatic CYP3A in vitro. And schisandrin A, schisandrin B and schisandrol B showed significant inductive effect on the expression of rat CYP3A1 mRNA.
Animals ; Cyclooctanes ; administration & dosage ; isolation & purification ; pharmacology ; Cytochrome P-450 CYP3A ; genetics ; metabolism ; Dioxoles ; administration & dosage ; isolation & purification ; pharmacology ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; administration & dosage ; isolation & purification ; pharmacology ; Hepatocytes ; drug effects ; enzymology ; Inhibitory Concentration 50 ; Lignans ; administration & dosage ; isolation & purification ; pharmacology ; Male ; Microsomes, Liver ; enzymology ; Plants, Medicinal ; chemistry ; Polycyclic Compounds ; administration & dosage ; isolation & purification ; pharmacology ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Schisandra ; chemistry

OBJECTIVETo optimize the animal model of liver injury that can properly represent the pathological characteristics of dampness-heat jaundice syndrome of traditional Chinese medicine.

METHODSThe liver injury in the model rat was induced by alpha-naphthylisothiocyanate (ANIT) and carbon tetrachloride (CCl(4) ) respectively, and the effects of Yinchenhao Decoction (, YCHD), a proved effective Chinese medical formula for treating the dampness-heat jaundice syndrome in clinic, on the two liver injury models were evaluated by analyzing the serum level of alanine aminotransferase (ALT), asparate aminotransferase (AST), alkaline phosphatase (ALP), malondialchehyche (MDA), total bilirubin (T-BIL), superoxide dismutase (SOD), glutathione peroxidase (GSH-PX) as well as the ratio of liver weight to body weight. The experimental data were analyzed by principal component analytical method of pattern recognition.

RESULTSThe ratio of liver weight to body weight was significantly elevated in the ANIT and CCl(4) groups when compared with that in the normal control (P<0.01). The contents of ALT and T-BIL were significantly higher in the ANIT group than in the normal control (P<0.05,P<0.01), and the levels of AST, ALT and ALP were significantly elevated in CCl(4) group relative to those in the normal control P<0.01). In the YCHD group, the increase in AST, ALT and ALP levels was significantly reduced (P<0.05, P<0.01), but with no significant increase in serum T-BIL. In the CCl(4) intoxicated group, the MDA content was significantly increased and SOD, GSH-PX activities decreased significantly compared with those in the normal control group, respectively (P<0.01). The increase in MDA induced by CCl(4) was significantly reduced by YCHD P<0.05).

CONCLUSIONYCHD showed significant effects on preventing liver injury progression induced by CCl(4), and the closest or most suitable animal model for damp-heat jaundice syndrome may be the one induced by CCl(4).

1-Naphthylisothiocyanate ; toxicity ; Alanine Transaminase ; blood ; Alkaline Phosphatase ; blood ; Animals ; Annonaceae ; Aspartate Aminotransferases ; blood ; Bilirubin ; blood ; Body Weight ; Carbon Tetrachloride ; toxicity ; Chemical and Drug Induced Liver Injury ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Glutathione ; metabolism ; Hepatocytes ; drug effects ; enzymology ; pathology ; Jaundice ; chemically induced ; drug therapy ; pathology ; Liver ; drug effects ; enzymology ; pathology ; Liver Diseases ; drug therapy ; pathology ; Male ; Malondialdehyde ; metabolism ; Organ Size ; Rats ; Rats, Wistar ; Superoxide Dismutase ; metabolism

OBJECTIVETo investigate the effects of hydroquinone (HQ) on expression of ubiquitin-ligating enzyme Rad18 in human hepatic cells (L-02), and to explore the role and possible mechanism of Rad18 involved in toxicity of HQ to hepatic cells.

METHODSAfter L-02 hepatic cells were exposed to HQ with various concentrations (0, 5, 10, 20, 40, 80 and 160 micromol/L) for 24 h, cell survival rate was measured by MTT assay; DNA impairment was evaluated by single cell gel electrophoresis (SCGE); The expression levels of Rad18 mRNA and protein were detected by Real-time fluorescent quantitative polymerase chain reaction (QPCR) technique and Western blot method respectively.

RESULTSHQ with concentration from 0 to 80 micromol/L had little effect on survival rate of L-02 (P > 0.05); Whereas the survival rate in the group of 160 micromol/L was significantly lower than in the control with the significant difference (P < 0.01) after treated with HQ for 24 h; The higher dose of HQ presented, the more degrees of olive tail moment (OTM) were produced and a dose-dependent relationship was shown. HQ in a low concentration (0 to approximately 40 micromol/L) could induce increase in the expression of Rad18 mRNA and protein which was in proportion to the increment of HQ concentration; the expression of Rad18 mRNA was enhanced increasingly, while the expression of Rad18 protein unchanged basically once the concentration of HQ exceeded 40 micromol/L; Besides, there was a positive correlation between OTM and the expression level of Rad18 mRNA (r = 0.919, P < 0.01).

CONCLUSIONHQ could regulate up the expression of Rad18 in L-02 hepatic cells.

Cell Survival ; drug effects ; Cells, Cultured ; DNA Damage ; drug effects ; DNA-Binding Proteins ; metabolism ; Hepatocytes ; drug effects ; enzymology ; Humans ; Hydroquinones ; toxicity ; Ubiquitin-Protein Ligases

OBJECTIVETo investigate the effects of sodium selenite on telomerase activity, apoptosis and expression of TERT, c-myc and p53 in rat hepatocytes.

METHODSSelenium at doses of 2.5, 5.0, and 10 micromol/kg was given to SD rats by gavage. In rat hepatocytes, telomerase activity was measured by the telomeric repeat amplification protocol (TRAP), apoptosis was detected by flow cytometry, and expressions of telomerase reverse transcriptase (TERT), c-myc and p53 were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). c-Myc and P53 proteins were detected by immunochemistry.

RESULTSSelenium at doses of 2.5, 5.0, and 10 micromol/kg significantly increased hepatocellular telomerase activity and induced apoptosis in a dose-dependent manner. Although selenium at doses of 2.5, 5.0, and 10 micromol/kg displayed no obvious enhancing effect on the TERT mRNA expression in rat hepatocytes (P > 0.05), it significantly increased the c-myc mRNA and p53 mRNA expression at the dose of 10 micromol/kg (P < 0.05). Selenium at doses of 5.0 and 10 micromol/kg obviously increased the content of P53 protein in rat hepatocytes, but only at the dose of 10 micromol/kg, it significantly promoted the value of c-Myc protein in them.

CONCLUSIONSelenium can slightly increase telomerase activity and TERT expression, and significantly induce apoptosis and over-expression of c-myc and p53 at relatively high doses. The beneficial effects of selenium on senescence and aging may be mediated by telomerase activation and expression of TERT, c-myc, and p53 in rat hepatocytes.

Animals ; Apoptosis ; drug effects ; Gene Expression Regulation ; drug effects ; Hepatocytes ; cytology ; drug effects ; enzymology ; Male ; Proto-Oncogene Proteins c-myc ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Selenium ; pharmacology ; Telomerase ; genetics ; metabolism ; Tumor Suppressor Protein p53 ; genetics ; metabolism

OBJECTIVETo explore how arylamine N-acetyltransferases (NATs) is related to cell apoptosis.

METHODSNAT activity in apoptotic HepG2 cells was measured using high performance liquid chromatography (HPLC); the apoptosis rate of HepG2 cells acted upon by an NAT inhibitor was measured using flow cytometry.

RESULTSNAT activity was lowered in apoptotic HepG2 cells; apoptosis rate induced by camptothecin (CAM) increased after inhibition of NAT activity in HepG2 cells.

CONCLUSIONNAT can inhibit apoptosis in HepG2 cells.

Apoptosis ; drug effects ; Arylamine N-Acetyltransferase ; antagonists & inhibitors ; metabolism ; Camptothecin ; administration & dosage ; Cells, Cultured ; Dose-Response Relationship, Drug ; Hepatocytes ; cytology ; drug effects ; enzymology ; Humans

OBJECTIVETo study the effect of salvianolic acid B (SAB) and curcumin, the extracts of Salvia Miltiorrhiza and Curcuma Longa, on the proliferation and activation of hepatic stellate cell (HSC), and the extracellular signal regulated kinase (ERK) expression in it.

METHODSRat's HSC-T6 were cultured and treated by SAB or curcumin. The inhibitory effect on cell proliferation was determined by 3-(4, 5-dimthyl-2-2thiazoly)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) colorimetry, and the expression levels of alpha smooth actin (alpha-SMA), collagen type I, and ERK were determined by Western blot.

RESULTSSAB and curcumin inhibited the proliferation and activation of rat's HSC-T6 in dose-dependent fashion and significantly reduced the expression level of alpha-SMA (P < 0.01). Curcumin significantly reduced the expression of collagen type I (P < 0.05). Both SAB and curcumin showed insignificant effect on the ERK expression level, but they could significantly reduce the level of phosphorylated-ERK expression, showing significant difference as compared with that in the control group (P < 0.01 and P < 0.05 respectively).

CONCLUSIONSAB and curcumin could significantly inhibit the proliferation, activation of HSC, and the production of type I collagen in HSC, the mechanism may be associated with their inhibition on ERK phosphorylation.

Animals ; Cell Division ; drug effects ; Cell Line ; Curcuma ; Drugs, Chinese Herbal ; pharmacology ; Extracellular Matrix ; drug effects ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Hepatocytes ; drug effects ; enzymology ; Liver Cirrhosis ; drug therapy ; metabolism ; MAP Kinase Signaling System ; drug effects ; Phosphorylation ; drug effects ; Plant Extracts ; Rats ; Salvia miltiorrhiza ; Vasodilator Agents ; pharmacology