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.

ACAT inhibitory activity of exudates from Calocedrus macrolepis var. formosana.

Cholesterol acyltransferase (ACAT) is an enzyme controlling cholesterol esterification in cells. Large amounts of cholesterol esters accumulate in macrophages and smooth muscle cells of blood vessel walls resulting in the initial stages of atherosclerosis. Thus, atherosclerosis might be inhibited through inhibition of the activity of ACAT. In the present study, we identified by spectral analysis and chromatographic quantification that ferruginol was the most abundant component of exudates secreted from the wounding site of Calocedrus macrolepis Kurz var. formosana. Results obtained from the cholesterol absorption assay revealed that ferruginol exhibited a significant inhibitory activity on cholesterol absorption in mice macrophages (RAW 264.7 cell). Based on the results from analyzing the ratio of cholesterol esterification, ferruginol dose-dependently suppressed cholesterol esterification and the IC50 value was 2.0 microg/mL. In conclusion, ferruginol revealed strong inhibitory activities that retarded the absorption and esterification of cholesterol in cells. Our finding indicates that ferruginol might possess a potential for development as a pharmaceutical product for preventing arteriosclerosis.

Anti-inflammatory activity of Flavokawain B from Alpinia pricei Hayata.

Alpinia pricei (Zingiberaceae) is a spicy herb indigenous to Taiwan. A potent anti-inflammatory compound, flavokawain B (FKB), was obtained from A. pricei. FKB significantly inhibited production of NO and PGE(2) in LPS-induced RAW 264.7 cells. Moreover, it also notably decreased the secretion of TNF-alpha. Expression of iNOS and COX-2 proteins was also inhibited by FKB in a dose-dependent manner. FKB blocked the nuclear translocation of NF-kappaB induced by LPS, which was associated with prevention IkappaB degradation, and subsequently decreased NF-kappaB protein levels in the nucleus. Similar anti-inflammatory activities of FKB were observed in an animal assay. NO concentrations in mouse serum rose dramatically from 3.2 to 28.8 microM after mice were challenged with LPS; however, preadministration of 200 mg/kg FKB reduced the NO concentration to 3.8 microM after challenge with LPS. Moreover, FKB strongly suppressed LPS-induced iNOS, COX-2, and NF-kappaB proteins expression in mouse liver.

Antiinflammatory activity of Lindera erythrocarpa fruits.

In this study, in vitro and in vivo antiinflammatory activities of fruits from Lindera erythrocarpa Makino were evaluated. The ethyl acetate soluble fraction derived from the ethanol extract of L. erythrocarpa fruits inhibited significantly nitric oxide (NO) production in lipopolysaccharide (LPS) induced NO in the murine macrophage cell line (RAW264.7) assay, the EC(50) being 16.35 microg/mL. Four compounds, including lucidone (1), cis/trans-methylludicone (2), methyl linderone (3) and linderone (4) were identified from the active fraction based on the bioactivity-guided fractionation procedure. Of these lucidone possessed the strongest NO inhibitory activity with an EC(50) value of 4.22 microg/mL. Furthermore, results from the protein expression assay demonstrated that lucidone suppressed iNOS and COX-2 protein expression in a dose-dependent manner. Lucidone also provided antiinflammatory activity in the croton oil-induced ear edema assay. When it was applied topically at a dosage of 0.5 and 1 mg per ear, the percent edema reduction in treated mice was 44% and 25%, respectively. The results obtained in this study indicated that lucidone has a good potential to be developed as an antiinflammation agent.