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A crude aqueous acetone extract of Osbeckia aspera Blume (Melastomataceae), a plant from Sri Lanka used traditionally to treat liver disease, was fractionated by column and preparative paper chromatography, and the fractions were analyzed by high-performance liquid chromatography (HPLC) using diode array and mass spectrometric detection. Phenolic acids (gallic, protocatechuic, and ellagic acid), flavonol glycosides [quercetin 3-O-β-galactopyranoside, quercetin 3-O-β-glucopyranoside, kaempferol 3-O-β-glucopyranoside, and kaempferol 3-O-(6''-O-p-coumaroyl-β-glucopyranoside) (tiliroside)] and flavonol aglycones (quercetin and kaempferol) were identified by comparison of their retention times, UV and MS spectra with those of authentic standards. Five compounds from a methanol extract were identified by NMR spectroscopy as the flavonol glycosides, quercetin 3-O-(3''-O-acetyl-β-galactopyranoside) and kaempferol 3-O-[2'',6''-di-O-(E,E)-p-coumaroyl-β-glucopyranoside], and the norsesquiterpenoids 6,9-dihydroxy-4,7-megastig-madien-3-one, 9-hydroxy-4,7-megastigmadien-3-one and 9-hydroxy-4-megastigmen-3-one. A crude water extract, 50% acetone extract and fractions from this extract, a 100% methanol extract, and three of the phenolic acids in the fractions were tested for in vitro hepatoprotective activity against bromobenzene and 2,6-dimethyl-N-acetyl p-quinoneimine toxicity to HepG2 liver-derived cells. The crude water extract showed protective activity against both liver toxins, whereas the fractions and compounds were more protective against 2,6-dimethyl-N-acetyl p-quinoneimine than bromobenzene. Of the three phenolic acids present in the extracts that were tested, gallic and protocatechuic acids were more active at protecting the liver cells from the two toxic compounds than ellagic acid. [ABSTRACT FROM AUTHOR]

5-Caffeoylquinic acid (5-CQA) is one of the most abundant compounds found in natural foods including coffee. We investigated whether 5-CQA had a cytoprotective effect through the NF-E2-related factor 2 (Nrf2)-antioxidant response element (ARE) signalling pathway. Nrf2 activation in response to 5-CQA treatment at the concentration of 10–100 μM is evaluated by Western blotting of Nrf2 and ARE reporter gene assay as well as its target gene expression in HepG2 cells. Intracellular reactive oxygen species (ROS) and glutathione (GSH) levels were measured in the tert-butyl hydroperoxide-induced hepatocytes to examined cytoprotective effect of 5-CQA (10–100 μM). The specific role of 5-CQA on Nrf2 activation was examined using Nrf2 knockout cells or Nrf2 specific inhibitor, ML-385. Nuclear translocation of Nrf2 is increased by 5-CQA in HepG2 cells which peaked at 6 h. Consequently, 5-CQA significantly increases the ARE reporter gene activity and downstream antioxidant proteins, including glutamate cysteine ligase (GCL), hemeoxygenase-1 (HO-1), NAD(P)H quinone oxidoreductase 1, and Sestrin2. Nrf2 deficiency or inhibition completely antagonized ability of 5-CQA to induce HO-1 and GCL expression. Cells pre-treated with 5-CQA were rescued from tert-butyl hydroperoxide-induced ROS production and GSH depletion. Nrf2 activation by 5-CQA was due to increased phosphorylation of MAPKs, AMPK and PKCδ. Taken together, our results demonstrate that as a novel Nrf2 activator, 5-CQA, may be a promising candidate against oxidative stress-mediated liver injury. Additional efforts are needed to assess 5-CQA, as a potential therapeutic in liver diseases in vivo and in humans. [ABSTRACT FROM AUTHOR]

The potential hepatotoxicity of Polygoni Multiflori Radix (PMR) has attracted much attention, but the specific mechanism of inducing hepatotoxicity is still unclear due to the complexity of its components. This study investigated the specific mechanism by which 2,3,5,4′-tetrahydroxy-stilbene-2-O-β-d-glucoside (TSG) regulates hepatotoxicity. The toxic effects of TSG (10, 100, 1000 μg/mL) on WRL-68 cells were examined using MTT, flow cytometry, and LDH assay after 24 h of incubation. Untreated cells served as the control. Gene and protein expression levels were determined by quantitative real-time PCR and Western blot, respectively. Immunofluorescence analysis was conducted to investigate the expression of light chain 3 (LC3). Luciferase activity assay was used to assess the targeted regulation of RUNX1 by miR-122. The half maximal inhibitory concentration (IC50) of TSG in WRL-68 cells was calculated as 1198.62 μg/mL. TSG (1000 μg/mL) inhibited cell viability and LDH activity and promoted WRL-68 cell apoptosis by inducing autophagy. Subsequent findings showed that TSG induced autophagy and promoted apoptosis in WRL-68 cells by downregulating the levels of p-PI3K, p-Akt, and p-mTOR proteins, while RUNX1 overexpression rescued this inhibition. Additionally, the effect of TSG on hepatocyte apoptosis was reversed by miR-122 knockdown. Furthermore, bioinformatics and dual luciferase reporter assay results indicated that miR-122 targeted RUNX1. Our data demonstrate for the first time that TSG regulates hepatotoxicity, possibly by upregulating miR-122 and inhibiting the RUNX1-mediated PI3K/Akt/mTOR pathway to promote autophagy and induce hepatocyte apoptosis. Further in vivo research is necessary to verify our conclusion. [ABSTRACT FROM AUTHOR]

Context:Eruca sativaMill. (Brassicaceae), commonly known as rocket salad, is a popular leafy-green vegetable with many health benefits. Objective:To evaluate the antidiabetic activities of this plant in major insulin-responsive tissues. Materials and methods:FiveE. sativaleaf extracts of varying polarity were prepared (aqueous extract, 70% and 95% ethanol extracts, then-hexane-soluble fraction of the 95% ethanol extract (ES3) and the defatted 95% ethanol extract).Eruca sativaextracts were investigated through a variety of cell-basedin vitrobioassays for antidiabetic activities in C2C12 skeletal muscle cells, H4IIE hepatocytes and 3T3-L1 adipocytes. Guided by the results of these bioassays, ES3 was fractionated into the saponifiable (SM) and the unspaonifiable (USM) fractions. Glucose uptake was measured using [3H]-deoxy-glucose, while the effects on hepatic glucose-6-phosphatase (G6Pase) and adipogenesis were assessed using Wako AutoKit Glucose and AdipoRed assays, respectively. Results:ES3 and its SM fraction significantly stimulated glucose uptake with EC50values of 8.0 and 5.8 μg/mL, respectively. Both extracts significantly inhibited G6Pase activity (IC50values of 4.8 and 9.3 μg/mL, respectively). Moreover, ES3 and SM showed significant adipogenic activities with EC50of 4.3 and 6.1 μg/mL, respectively. Fatty acid content of SM was identified by GC-MS.trans-Vaccenic and palmitoleic acids were the major unsaturated fatty acids, while palmitic and azelaic acids were the main saturated fatty acids. Discussion and conclusion:These findings indicate that ES3 and its fatty acid-rich fraction exhibit antidiabetic activities in insulin-responsive cell lines and may hence prove useful for the treatment of type 2 diabetes. [ABSTRACT FROM PUBLISHER]

Context:Zea maysL. (Poaceae)Stigma maydisis an underutilized product of corn cultivation finding therapeutic applications in oxidative stress-related disorders. Objectives:This study investigated its aqueous extract against acetaminophen (APAP)-perturbed oxidative insults in rat hepatocytes. Materials and methods:Hepatotoxic rats were orally pre- and post-treated with the extract (at 200 and 400 mg/kg body weight) and vitamin C (200 mg/kg body weight), respectively, for 14 days. Liver function, antioxidative and histological analyses were thereafter evaluated. Results:The APAP-induced marked (p < 0.05) increases in the activities of alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma glutamyl transferase and the concentrations of bilirubin, oxidized glutathione, protein carbonyls, malondialdehyde, conjugated dienes, lipid hydroperoxides and fragmented DNA were dose-dependently extenuated in the extract-treated animals. The extract also significantly (p < 0.05) improved the reduced activities of superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase as well as total protein, albumin and glutathione concentrations in the hepatotoxic rats. These improvements may be attributed to the bioactive constituents as revealed by the gas chromatography–mass spectrometric chromatogram of the extract. The observed effects compared favourably with vitamin C and are informative of hepatoprotective and antioxidative attributes of the extract and were further supported by the histological analysis. Conclusion:The data from the present findings suggest thatStigma maydisaqueous extract is capable of preventing and ameliorating APAP-mediated oxidative hepatic damage via enhancement of antioxidant defence systems. [ABSTRACT FROM AUTHOR]

Context: Natural products are good sources of natural dietary antioxidants that are believed to protect the body against hepatotoxic effect induced by oxidative stress.Hedyotis diffusaWilld (Rubiaceae) (HDW) is a traditional Chinese medicinal herb that has been shown to possess a variety of antioxidant properties. Objective: The present study examines and explains the cell protective property of HDW water extract (WEHDW). Materials and methods: 2,2-Diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl (DPPH) assay was used to measure the free radical scavenging property of WEHDW (0.001–10 mg/mL). The protective effect of WEHDW (0.3–10 mg/mL 2 h pretreatment) against hydrogen peroxide (H2O2,200 μM for 6 h) induced cytotoxicity in human hepatic cells, LO2, was evaluated using cell viability assay and nuclear staining. The molecular pathway of WEHDW's effect was investigated by using Western blot assay. Results: WEHDW had a 50% scavenging concentration (SC50) at 0.153 mg/mL in the DPPH assay. Exposure of LO2cells to H2O2resulted in apoptosis which could be markedly attenuated by pre-treating WEHDW in a concentration-dependent manner (0.5, 1, 3, 5, or 10 mg/mL) (all withp < 0.001, versus control). Moreover, Hoechst (nuclear) staining showed that 1 mg/mL WEHDW could protect LO2cells by attenuating apoptotic cell death mediated by H2O2. It was found that WEHDW reversed H2O2-induced activation of MEK/ERK pathway and H2O2-induced inhibition of P13-K/AKT/GSK3β pathway in LO2cells. Discussion and conclusion: WEHDW may help to improve the antioxidant defense system, resulting in prevention of oxidative stress-related fatty liver diseases. [ABSTRACT FROM AUTHOR]

Context: Fumonisin B1 (FB1) is a mycotoxin produced by Fusarium verticillioides (Sacc.) Nirenberg (Nectriaceae) mold that contaminates maize and other agricultural products. Although the effects of FB1 on sphingolipid metabolism are clear, little is known about early molecular changes associated with FB1 carcinogenicity. Objective: Alteration on DNA methylation, as an early event in non-genotoxic carcinogenesis, may play an important role in the mechanism of FB1 toxiciy. Materials and methods: Dose-related effects of FB1 (1-50 µM for 24 h) on global DNA methylation by using high-performance liquid chromatography with UV-diode array detection (HPLC-UV/DAD) and CpG promoter methylation by methylation-specific PCR (MSP) were performed in rat liver (Clone 9) and rat kidney (NRK-52E) epithelial cells. Results: Cell viability reduction is 39% and 34% by the XTT test and LDH release in the growth medium is 32% and 26% at 200 µM of FB1 treatment in Clone 9 and NRK-52E cells, respectively. No significant dose-related effects of FB1 on global DNA methylation which ranged from 4 to 5% were observed in both cells compared with controls. Promoter regions of c-myc gene were methylated (>33%) at 10 and 50 µM of FB1 treatment in Clone 9 cells while it was unmethylated in NRK-52E cells. Promoter regions of p15 gene were unmethylated while VHL gene were found to be methylated (>33%) at 10, 25, and 50 µM and 10 and 50 µM of FB1 treatment in Clone 9 and NRK-52E cells, respectively. Discussion and conclusion: Alteration in DNA methylation might play an important role in the toxicity of FB1 in risk assessment process. [ABSTRACT FROM AUTHOR]

Context: Indian mustard [ Brassica juncea (L.) Czern. & Coss. (Brassicaceae)] is reported to possess diverse pharmacological properties. However, limited information is available concerning its hepatoprotective activity and mechanism of action. Objective: To study the protective mechanism of mustard seed extract against acetaminophen (APAP) toxicity in a hepatocellular carcinoma (HepG2) cell line. Materials and methods: Hepatotoxicity models were established using APAP (2.5-22.5 mM) based on the cytotoxicity profile. An antioxidant-rich fraction from mustard seeds was extracted and evaluated for its hepatoprotective potential. The mechanism of action was elucidated using various in vitro antioxidant assays, the detection of intracellular generation of reactive oxygen species (ROS), and cell cycle analysis. The phytoconstituents isolated via HPLC-DAD were also evaluated for hepatoprotective activity. Results: Hydromethanolic seed extract exhibited hepatoprotective activity in post- and pre-treatment models of 20 mM APAP toxicity and restored the elevated levels of liver indices to normal values ( p < 0.05). Post-treatment suppressed the generation of ROS by 58.37% and pre-treatment effectively prevented the generation of ROS by 90.5%. The mechanism of ROS suppression was further supported by antioxidant activity (IC50) data from DPPH (103.37 ± 4.2 µg AAE/mg), FRAP (83.26 ± 1.1 µg AAE/mg), ORAC (1115 µM GAE/ml), ABTS (83.05 µg GAE/ml), and superoxide (345.22 ± 5.15 µg AAE/mg) scavenging assays and by the restoration of cell cycle alterations. HPLC-DAD analysis revealed the presence quercetin, vitamin E, and catechin, which exhibited hepatoprotective activity. Discussion and conclusions: A phytoextract of mustard seeds acts by suppressing the generation of ROS in response to APAP toxicity. [ABSTRACT FROM AUTHOR]

Context: Rubus aleaefolius Poir. (Rosaceae) is used as a folk medicine to treat various types of hepatitis with significant effects in Fujian Province of China. The ethyl acetate fraction of R. aleaefolius root ethanol extract proved effective after our testing in vivo animal experiments. Objective: The protective effects of a major constituent, 1β-hydroxyeuscaphic acid isolated from R. aleaefolius was first investigated against carbon tetrachloride (CCl4)-induced injury in liver cells from hepatocytes cell line (BRL-3A). Materials and methods: Treatment of BRL-3A with CCl4 led to generation of free radicals detected after a 2 h incubation and produced cell injury demonstrated by increased leakage of alanine aminotransaminase (ALT) and aspartate aminotransaminase (AST) in the media. Exposure to CCl4 caused apoptosis to cells but did not induce lipid peroxidation. Following treatment with 1β-hydroxyeuscaphic acid at doses ranging from 1 to 100 µg/mL for 24 h, cellular morphology, cell growth function (MTT assay), ALT, AST, malondialdehyde (MDA) and superoxide dismutase (SOD) were assessed and evaluated under control and exposed conditions. Results: The IC50 of 1β-hydroxyeuscaphic acid was 15 μg/mL. Exposure of injured BRL-3A at 20 μg/mL changed abnormal size, cellular shrinkage, and enhanced regulation. ALT, AST, MDA enzyme levels were reduced and SOD activity was increased. Discussion and conclusion: Treatment with 1β-hydroxyeuscaphic acid has significant hepatoprotective activity by lowering the leakage of intracellular enzymes, reducing the oxidation of proteins and decreasing the incidence of apoptosis. These results provide a basis for confirming the traditional uses of R. aleaefolius in treating hepatic diseases. [ABSTRACT FROM AUTHOR]

Context: Ursolic acid is a pentacyclic triterpenoid which has hepatoprotective and antihepatotoxic activities. Objective: This study investigated whether ursolic acid is able to stimulate liver regeneration in partially hepatectomized mice. Materials and methods: Ursolic acid or the vehicle solution was orally administered to the experimental, sham-operated and vehicle-treated group mice for 7 days, positive control animal (mice) was treated with recombinant human hepatocyte growth factor (rhHGF), and then the 70% liver partial hepatectomy was performed. The liver mass recovery rate was estimated by measuring the ratios of mice liver weight to body weight. The liver cells undergoing DNA synthesis were identified by immunohistochemistry analysis using monoclonal anti-BrdU antibodies. The expression levels of cyclin D1, cyclin E and C/EBP proteins (C/EBPα and C/EBPβ) were detected by the Western blotting technique. Results: Our results showed administration of ursolic acid significantly increased the ratio of the liver to body weight and BrdU labeling index at 36 and 48 h after partial hepatectomy, and the potency of UA is similar to rhHGF treated positive control mice. In addition, ursolic acid treatment significantly increased cyclin D1, cyclin E and C/EBPβ protein expression levels at 36 h after liver PHx compared with the vehicle-treated control mice. Discussion and conclusion: All these results suggest that ursolic acid stimulates liver proliferation after partial hepatectomy, and this effect may be associated with the stimulation of C/EBPβ expression. [ABSTRACT FROM AUTHOR]

Objective: Punarnavashtak kwath (PNK) is a classical Ayurvedic formulation, mentioned in Ayurvedic literature Bhaishajya Ratnavali, for hepatic disorders and asthma. This study investigated the hepatoprotective activity of PNK to validate the traditional use of this formulation. Materials and methods: PNK was prepared in the laboratory according to the method given in Ayurvedic literature. Phytochemical screening was performed to determine the presence of phytoconstituents. Hepatoprotective activity was evaluated against CCl4-induced hepatotoxicity in rats and by its effect on the HepG2 cell line. Results: Preliminary phytochemical screening revealed the presence of alkaloids, tannins, flavonoids, saponins, and a bitter principle in PNK. Administration of PNK produced significant hepatoprotective effect as demonstrated by decreased levels of serum liver marker enzymes such as aspartate transaminase, serum alanine transaminase, serum alkaline phosphatase, and serum bilirubin and an increase in protein level. Thiopentone-induced sleeping time was also decreased in the PNK-treated animals compared with the CCl4-treated group. It also showed antioxidant activity by increase in activity of glutathione, superoxide dismutase, and catalase and by a decrease in thiobarbituric acid reactive substance level compared with the CCl4-treated group. Results of a histopathological study also support the hepatoprotective activity of PNK. Investigation carried out on the HepG2 cell line depicted significant increase in viability of cells exposed to PNK as compared with CCl4-treated cells. Discussion and Conclusion: It can be concluded that PNK protects hepatocytes from CCl4-induced liver damages due to its antioxidant effect on hepatocytes. An in vitro study on HepG2 cell lines also supports its protective effect. [ABSTRACT FROM AUTHOR]

Morinda citrifolia Linn. (Rubiaceae), common name noni, has been used as a herbal medicine for over 2000 years. The consumption of noni, and especially the fruit, stresses the importance, urgency, and possibility of the examination of drug interaction when concomitantly administered with a drug. The objectives of this study were to determine the effects of noni juice (NJ) on aminopyrine N-demethylase (APND), uridine diphosphoglucuronosyl-transferase (UGT), and cytosolic glutathione S-transferase (GST) drug metabolizing enzymes and the molecular mechanism elucidation of NJ on APND using different inhibitors and stimulators. The in vitro results for APND showed that different concentrations of NJ significantly increased the activity in isolated hepatocytes at 1.0 ng/mL, 10 ng/mL, 10 μg/mL, 20 μg/mL, 50 μg/mL, and 100 μg/mL. The ex vivo results demonstrated that NJ (210 mg/kg) produced a statistically significant increase in APND activity following 1 day of NJ treatment. The results for UGT and GST showed a decrease in the activity of UGT at a dose of 21 mg/kg following 1 day of treatment, and at 2.1 and 21 mg/kg following 14 days of treatment. GST enzyme demonstrated an increase in activity by 100% for all doses following 1 day of treatment. Molecular mechanism elucidation of the ex vivo effect of NJ on phase I APND showed that KT5720 significantly reduced the activity as compared to control. A change in activity of APND, UGT, and GST following 1 day and 14 days of treatment suggests that all three metabolic pathways may play a role in herb–drug interaction by modulation of metabolic enzymes. [ABSTRACT FROM AUTHOR]

Glutathione S-transferase (GST) is a family of enzymes involved in the detoxification of electrophilic compounds. Different classes of GST are expressed in various organs, such as liver, lungs, stomach and others. Expression of GST can be modulated by diet components and plant-derived compounds. The importance of controlling GST expression is twofold: increasing levels of GST are beneficial to prevent deleterious effects of toxic and carcinogenic compounds, while inhibition of GST in tumor cells may help overcoming tumor resistance to chemotherapy. A screening of 16 plants used in the Brazilian pharmacopoeia tested their effects on GST expression in hepatocytes and Jurkat (leukemia) T-cells. The methanol extracts of five plants inhibited GST expression in hepatocytes. Three plants significantly inhibited and four others induced GST expression in Jurkat cells. Among these, the extracts of Bauhinia forficata Link. (Leguminosae) and Cecropia pachystachya Tréc. (Urticaceae) inhibited GST expression at relatively low concentrations. With the exception of B. forficata, all plants were cytotoxic when administered to Jurkat cells at high doses (1 mg/mL) and some extracts were considerably cytotoxic even at lower concentrations. [ABSTRACT FROM AUTHOR]

Diabetes mellitus is a chronic disease which affects millions of people worldwide. The prevalence of this disease is increasing annually and the number of diabetics is projected to rise above 300 million before 2025. The growing number of diabetics, coupled with the harsh side effects of some synthetic drugs has led to the increasing search for more natural products of plant origin. Vernonia amygdalina Del. (Asteraceae) is one of the plants commonly used for the treatment of diabetes in Africa. This study evaluated the effect of leaf extracts of this plant on glucose utilization in 3T3-L1, C2C12 muscle, and Chang-liver cells. Treatment of the cells with the acetone, methanol, water, and n-hexane/isopropanol extracts of V. amygdalina leaves significantly increased glucose utilization in the C2C12 muscle and Chang-liver cells but showed no effect on the 3T3-L1 cells. The water and n-hexane/isopropanol extracts were the most active in the C2C12 cells with a response of 78.3 and 95.6% above the control, respectively, while in the Chang-liver cells, water and acetone extracts had a response of 65.8 and 59.6% above the control, respectively. The results, especially of the water extract, strongly corroborate the ethnomedical uses of V. amygdalina as an antidiabetic plant. [ABSTRACT FROM AUTHOR]

A purified saponin mixture (PSM) isolated from Astragalus corniculatus Bieb. (Fabaceae) was investigated for its protective effect in two models of toxicity, carbon tetrachloride (CCl4) and tert-butyl hydroperoxide (t-BuOOH), using isolated rat hepatocytes. CCl4 undergoes dehalogenation in the liver endoplasmic reticulum. This process leads to trichlormethyl radical (CCl3) formation, initiation of lipid peroxidation, and measurable toxic effects on the hepatocytes. Oxidative damage is widely recognized as being involved in the development of many pathological conditions. In our experiment, t-BuOOH was used as a model of oxidative stress. The hepatocytes were incubated with the PSM alone (0.01-100 μ M) and along with CCl4 (86 μ M) and t-BuOOH (75 μ M). As a sign of cytotoxicity, cell viability was used. CCl4 and t-BuOOH significantly decreased hepatocyte viability. Our data indicate that PSM showed lower toxic effects compared to CCl4 and t-BuOOH and in combination exerted statistically significant protection of cell viability against the toxic agents. [ABSTRACT FROM AUTHOR]