Hepatoprotective effect of lucidone against alcohol-induced oxidative stress in human hepatic HepG2 cells through the up-regulation of HO-1/Nrf-2 antioxidant genes.

Lucidone was previously reported to exhibit anti-inflammatory activity in vitro and in vivo. In the present study, we characterized the mechanisms underlying the hepatoprotective effect of lucidone against alcohol-induced oxidative stress in vitro. Human hepatoma (HepG2) cells were pretreated with lucidone (1-10μg/mL) and then hepatotoxicity was stimulated by the addition ethanol (100mM). With response to ethanol-challenge, increased amount of alanine aminotransferase (ALT) and aspirate aminotransferase (AST) release were observed, whereas lucidone pretreatment significantly inhibited the leakage of AST and ALT in HepG2 cells without appreciable cytotoxic effects. We also found that lucidone pretreatment significantly decreased ethanol-induced nitric oxide (NO), tumor necrosis factor-α (TNF-α), malondialdehyde (MDA), reactive oxygen species (ROS) and glutathione (GSH) depletion in HepG2 cells. Furthermore, Western blot and quantitative-PCR analyses showed that ethanol-exposure apparently down-regulated endogenous anti-oxidant hemoxygenase-1 (HO-1) expression, whereas pretreatment with lucidone significantly up-regulates HO-1 expression followed by the transcriptional activation of NF-E2 related factor-2 (Nrf-2). Interestingly, the profound up-regulation of HO-1 and Nrf-2 were observed in only ethanol-challenged cells, which evidenced that lucidone-induced induction of HO-/Nrf-2 were specific with oxidative stress. Thus, we concluded that lucidone-mediated up-regulation of phase-II enzymes and HO-1 via Nrf-2 signaling pathway may provide a pivotal mechanism for its hepatoprotective action.

Senthil Kumar KJ ，Liao JW ，Xiao JH ，Gokila Vani M ，Wang SY ... -  《-》

Antroquinonol from ethanolic extract of mycelium of Antrodia cinnamomea protects hepatic cells from ethanol-induced oxidative stress through Nrf-2 activation.

In recent years, the medicinal mushroom Antrodia cinnamomea, known as "niu-chang chih" has received much attention with regard to its possible health benefits; especially its hepatoprotective effects against various drugs, toxins, and alcohol induced liver diseases. However, the molecular mechanism underlying this protective effect of Antrodia cinnamomea and its active compound antroquinonol was poorly understood. In the present study we evaluated to understand the hepatoprotective efficacy of antroquinonol and ethanolic extracts of mycelia of Antrodia cinnamomea (EMAC) in vitro and in vivo. The protective mechanism of antroquinonol and EMAC against ethanol-induced oxidative stress was investigated in cultured human hepatoma HepG2 cells and ICR mice model, respectively. HepG2 cells were pretreated with antroquinonol (1-20μM) and oxidative stress was induced by ethanol (100mM). Meanwhile, male ICR mice were pretreated with EMAC for 10 days and hepatotoxicity was generated by the addition of ethanol (5g/kg). Hepatic enzymes, cytokines and chemokines were determined using commercially available assay kits. Western blotting and real-time PCR were subjected to analyze HO-1 and Nr-2 expression. EMSA was performed to monitor Nrf-2 ARE binding activity. Possible changes in hepatic lesion were observed using histopathological analysis. Antroquinonol pretreatment significantly inhibited ethanol-induced AST, ALT, ROS, NO, MDA production and GSH depletion in HepG2 cells. Western blot and RT-PCR analysis showed that antroquinonol enhanced Nrf-2 activation and its downstream antioxidant gene HO-1 via MAPK pathway. This mechanism was then confirmed in vivo in an acute ethanol intoxicated mouse model: serum ALT and AST production, hepatocellular lipid peroxidation and GSH depletion was prevented by EMAC in a dose-dependent manner. EMAC significantly enhanced HO-1 and Nrf-2 activation via MAPKs consistent with in vitro studies. Ethanol-induced hepatic swelling and hydropic degeneration of hepatocytes was significantly inhibited by EMAC in a dose-dependent manner. These results provide a scientific basis for the hepatoprotective effects of Antrodia cinnamomea. Data also imply that antroquinonol, a potent bioactive compound may be responsible for the hepatoprotective activity of Antrodia cinnamomea. Moreover, the present study highly supported our traditional knowledge that Antrodia cinnamomea as a potential candidate for the treatment of alcoholic liver diseases.

Kumar KJ ，Chu FH ，Hsieh HW ，Liao JW ，Li WH ，Lin JC ，Shaw JF ，Wang SY ... -  《-》

Lucidone protects human skin keratinocytes against free radical-induced oxidative damage and inflammation through the up-regulation of HO-1/Nrf2 antioxidant genes and down-regulation of NF-κB signaling pathway.

We investigated the protective effects of lucidone, a naturally occurring cyclopentenedione isolated from the fruits of Lindera erythrocarpa Makino, against free-radical and inflammation stimulator 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH)-induced oxidative stress in human keratinocyte (HaCaT) cells, with the aim of revealing the possible mechanisms underlying the protective efficacy. Lucidone pretreatment (0.5-10 μg/mL) markedly increased HaCaT cell viability and suppressed AAPH-induced reactive oxygen species (ROS) generation, lipid peroxidation, and DNA damage. Notably, the antioxidant potential of lucidone was directly correlated with the increased expression of an antioxidant gene, heme oxygenase 1 (HO-1), which was followed by the augmentation of the nuclear translocation and transcriptional activation of NF-E2-related factor-2 (Nrf2), with or without AAPH. Nrf2 knockdown diminished the protective effects of lucidone. We also observed that lucidone pretreatment inhibited AAPH-induced inflammatory chemokine prostaglandin E₂ (PGE₂) production and the expression of cyclooxygenase-2 (COX-2) in HaCaT cells. Lucidone treatment also significantly inhibited AAPH-induced nuclear factor-κB (NF-κB) activation and suppressing the degradation of inhibitor-κB (I-κB). Furthermore, lucidone significantly diminished AAPH-induced COX-2 expression through the down-regulation of the extracellular signal-regulated kinase (ERK) and p38 MAPK signaling pathways. Therefore, lucidone may possess antioxidant and anti-inflammatory properties and may be useful for the prevention of free radical-induced skin damage.

Kumar KJ ，Yang HL ，Tsai YC ，Hung PC ，Chang SH ，Lo HW ，Shen PC ，Chen SC ，Wang HM ，Wang SY ，Chou CW ，Hseu YC ... -  《-》

The activation of HO-1/Nrf-2 contributes to the protective effects of diallyl disulfide (DADS) against ethanol-induced oxidative stress.

Diallyl disulfide (DADS) is a garlic-derived organosulfur compound. The current study is designed to evaluate the protective effects of DADS against ethanol-induced oxidative stress, and to explore the underlying mechanisms by examining the HO-1/Nrf-2 pathway. We investigated whether or not DADS could activate the HO-1 in normal human liver cell LO2, and then evaluated the protective effects of DADS against ethanol-induced damage in LO2 cells and in acute ethanol-intoxicated mice. The biochemical parameters were measured using commercial kits. HO-1 mRNA level was determined by RT-PCR. Histopathology and immunofluorescence assay were performed with routine methods. Protein levels were measured by western blot. DADS significantly increased the mRNA and protein levels of HO-1, stimulated the nuclear translocation of Nrf-2 and increased the phosphorylation of MAPK in LO2 cells. The nuclear translocation of Nrf-2 was abrogated by MAPK inhibitors. DADS significantly suppressed ethanol-induced elevation of lactate dehydrogenase (LDH) and aspartate transaminase (AST) activities, decrease of glutathione (GSH) level, increase of malondialdehyde (MDA) levels, and apoptosis of LO2 cells, which were all blocked by ZnPPIX. In mice, DADS effectively suppressed acute ethanol-induced elevation of aminotransferase activities, and improved liver histopathological changes, which might be associated with HO-1 activation. These results demonstrate that DADS could induce the activation of HO-1/Nrf-2 pathway, which may contribute to the protective effects of DADS against ethanol-induced liver injury. DADS may be beneficial for the prevention and treatment of ALD due to significant activation of HO-1/Nrf-2 pathway.

Zeng T ，Zhang CL ，Song FY ，Zhao XL ，Yu LH ，Zhu ZP ，Xie KQ ... -  《-》

In vitro and in vivo hepatoprotective effect of ganodermanontriol against t-BHP-induced oxidative stress.

Ganoderma lucidum (Fr.) Karst. (Ganodermataceae) is a mushroom which is used as a traditional remedy in the treatment of human diseases such as hepatitis, liver disorders, hypercholesterolemia, arthritis, bronchitis and tumorigenic diseases. This study targets the evaluation of hepatoprotective activity of ganodermanontriol, a sterol isolated from Ganoderma lucidum, and the investigation of its mechanism of action in Hepa1c1c7 and murine liver cells upon tert-butyl hydroperoxide (t-BHP)-induced inflammation. t-BHP was utilized to stimulate an anti-inflammatory reaction in the hepatic cell lines and murine hepatic tissue examined. Western blot and reverse transcription-quantitative polymerase chain reaction (RT-PCR) were used to estimate the expression of ganodermanontriol (GDT)-induced proteins, including heme oxidase-1 (HO-1) and mitogen-activated protein kinases (MAPKs) as well as the corresponding mRNA. Luciferase assays were conducted to evaluate the interaction between NF-E2-related factor-2 (Nrf-2), the antioxidant response element (ARE), and the promoter region of the HO-1 gene and subsequent gene expression. Biochemical markers for hepatotoxicity were monitored to assess whether GDT protected the cells from the t-BHP-mediated oxidative stimuli. GDT induced HO-1 expression via the activation of Nrf-2 nuclear translocation and the subsequent transcription of the HO-1 gene in vitro and in vivo, which seemed to be regulated by phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) and p38 signaling pathways. GDT exhibited in vitro and in vivo hepatoprotective activity as determined by the lowered levels of hepatic enzymes and malondialdehydes and the elevated glutathione levels. This study validates the ethnopharmacological application of Ganoderma lucidum as a treatment for hepatic disorders. GDT induced in vitro and in vivo anti-inflammatory activity in t-BHP-damaged hepatic cells through the expression of HO-1, and in which PI3K/Akt and p38 kinases are involved. Our study motivates further research in the exploration of potent hepatoprotective agents from Ganoderma lucidum.

Ha do T ，Oh J ，Khoi NM ，Dao TT ，Dung le V ，Do TN ，Lee SM ，Jang TS ，Jeong GS ，Na M ... -  《-》