Ji Sun Lim

Dehydroglyasperin C isolated from licorice caused Nrf2-mediated induction of detoxifying enzymes.

Our preliminary experiment demonstrated that a n-hexane/EtOH (9:1, volume) extract of Glycyrrhiza uralensis (licorice) caused a significant induction of NAD(P)H:oxidoquinone reductase (NQO1), one of the well-known phase 2 detoxifying enzymes. We isolated dehydroglyasperin C (DGC) as a potent phase 2 enzyme inducer from licorice. DGC induced NQO1 both in wild-type murine hepatoma Hepa1c1c7 and ARNT-lacking BPRc1 cells, indicating that the compound is a monofunctional inducer. The compound induced not only NQO1 but also some other phase 2 detoxifying/antioxidant enzymes, such as glutathione S-transferase, gamma-glutamylcysteine synthase, glutathione reductase, and heme oxygenase 1. Similar to most monofunctional inducers, DGC caused the accumulation of Nrf2 in the nucleus in dose- and time-dependent manners and thereby activated expression of phase 2 detoxifying enzymes. It also resulted in a dose-dependent increase in the luciferase activity in the reporter assay, in which HepG2-C8 cells transfected with antioxidant response element (ARE)-luciferase construct were used, suggesting that the induction of phase 2 detoxifying and antioxidant enzymes could be achieved through the interaction of Nrf2 with the ARE sequence in the promoter region of their genes.

Neuroprotective Effects of Dehydroglyasperin C through Activation of Heme Oxygenase-1 in Mouse Hippocampal Cells

Licorice, the root of the Glycyrrhiza species ( Glycyrrhiza uralensis Fisher), is known to have antioxidant, anti-inflammatory, antiviral, and antitumor properties. The objective of this study is to explore the neuroprotective effect of dehydroglyasperin C (DGC) against glutamate-induced oxidative stress in mouse hippocampal HT22 cells. DGC significantly reduced cytotoxicity and reactive oxygen species (ROS) generation induced by glutamate in HT22 cells, whereas DGC did not restore glutathione depletion caused by glutamate. In addition, it was further investigated whether DGC affected the expression of heme oxygenase (HO)-1, one of the major cellular antioxidant defense systems, and it was found that DGC dose-dependently increased HO-1 expression. DGC-mediated cytoprotection of HT22 neuronal cells from glutamate insult was abrogated by either HO-1 inhibitor (Tin protoporphyrin, SnPP) or AKT inhibitor (LY294002). In conclusion, the present results demonstrate for the first time that DGC protects neuronal cells against glutamate-induced oxidative injury through the induction of HO-1 expression, which is, in turn, activated maybe through Nrf2-Keap1 and PI3K/AKT signaling pathways.

Protection by Chrysanthemum zawadskii extract from liver damage of mice caused by carbon tetrachloride is maybe mediated by modulation of QR activity

Our previous study demonstrated that methanolic extract of Chrysanthemum zawadskii Herbich var. latilobum Kitamura (Compositae) has the potential to induce detoxifying enzymes such as NAD(P)H:(quinone acceptor) oxidoreductase 1 (EC 1.6.99.2) (NQO1, QR) and glutathione S-transferase (GST). In this study we further fractionated methanolic extract of Chrysanthemum zawadskii and investigated the detoxifying enzyme-inducing potential of each fraction. The fraction (CZ-6) shown the highest QR-inducing activity was found to contain (+)-(3S,4S,5R,8S)-(E)-8-acetoxy-4-hydroxy-3-isovaleroyloxy-2-(hexa-2,4-diynyliden)-1,6-dioxaspiro [4,5] decane and increased QR enzyme activity in a dose-dependent manner. Furthermore, CZ-6 fraction caused a dose-dependent enhancement of luciferase activity in HepG2-C8 cells generated by stably transfecting antioxidant response element-luciferase gene construct, suggesting that it induces antioxidant/detoxifying enzymes through antioxidant response element (ARE)-mediated transcriptional activation of the relevant genes. Although CZ-6 fraction failed to induce hepatic QR in mice over the control, it restored QR activity suppressed by CCl4 treatment to the control level. Hepatic injury induced by CCl4 was also slightly protected by pretreatment with CZ-6. In conclusion, although CZ-6 fractionated from methanolic extract of Chrysanthemum zawadskii did not cause a significant QR induction in mice organs such as liver, kidney, and stomach, it showed protective effect from liver damage caused by CCl4.

Dehydroglyasperin C, a component of liquorice, attenuates proliferation and migration induced by platelet-derived growth factor in human arterial smooth muscle cells

Liquorice is one of the botanicals used frequently as a traditional medicine in the West and in the East. Platelet-derived growth factor (PDGF)-BB is involved in the development of CVD by inducing abnormal proliferation and migration of vascular smooth muscle cells. In our preliminary study, dehydroglyasperin C (DGC), an active compound of liquorice, showed strong antioxidant activity. Since phytochemicals with antioxidant activities showed beneficial effects on chronic inflammatory diseases, the present study aimed to investigate the effects of DGC on PDGF-induced proliferation and migration of human aortic smooth muscle cells (HASMC). Treatment of HASMC with DGC for 24 h significantly decreased PDGF-induced cell number and DNA synthesis in a dose-dependent manner without any cytotoxicity, as demonstrated by the 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide test and thymidine incorporation. Upon cell cycle analysis, DGC blocked the PDGF-induced progression through the G0/G1 to S phase of the cell cycle, and down-regulated the expression of cyclin-dependent kinase (CDK); 2, cyclin E, CDK4 and cyclin D1. Furthermore, DGC significantly attenuated PDGF-stimulated phosphorylation of PDGF receptor-b, phospholipase C-g1, AKT and extracellular-regulated kinase 1/2, and DGC inhibited cell migration and the dissociation of actin filaments by PDGF. In a rat vascular balloon injury model, DGC suppressed an excessive reduction in luminal diameters and neointimal formation compared with the control group. These results demonstrate the mechanistic basis for the prevention of CVD and the potential therapeutic properties of DGC.

Abstract 4243: Potential protective role of phytoalexins derived from soybean by biotic elicitor on inflammatory mechanism

Given the preventive effect of soy intake against several chronic diseases, this study was conducted to investigate the inhibitory activity of phytoalexins glyceollins derived from soybean isoflavones by treating with biotic elicitor against LPS-induced inflammation in RAW 264.7 cell culture system. Our data showed that glyceollins effectively inhibited NO production, IL-6 release, and COX-2 expression induced by LPS. In particular, glyceollins suppressed the LPS-induced phosphorylation of NF-B p65, suggesting that the compounds inhibit the production of NO and transcriptional activation of COX-2 by regulating NF-B activity. In another experiment we found that glyceollins enhanced the expression of heme oxygenase 1 and repressed the phosphorylation of ERK1/2 and p38 but not JNK in LPS-treated RAW 264.7 cells. Glyceollins also reduced TPA-induced skin inflammation in mouse model, confirming the anti-inflammatory activity of glyceollins in vivo system as well as cell culture system. Taken together, glyceollins merit further study as potential therapeutic agents for inflammatory disorders. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4243. doi:10.1158/1538-7445.AM2011-4243

Abstract 5666: Nrf2-mediated induction of phase 2 detoxifying enzymes by an active compound isolated from Chrysanthemum zawadskii

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Previous study demonstrated that methanolic extract of Chrysanthemum has the potential to induce detoxifying enzymes such as NAD(P)H:(quinone acceptor) oxidoreductase 1 (EC [1.6.99.2][1]) (NQO1, QR). In this study we further fractionated methanolic extract of Chrysanthemum and investigated the detoxifying enzyme-inducing potential of each fraction. The fraction shown the highest NQO1-inducing activity was found to contain (+)-(3S,4S,5R,8S)-(E)-8-acetoxy-4-hydroxy-3-isovaleroyloxy-2-(hexa-2,4-diynyliden)-1,6-dioxaspiro[4,5]decane (CZ-5) which increased NQO1 enzyme activity in a dose-dependent manner. And the compound (CZ-5), like most phase 2 enzyme inducers, caused a increasing expression pattern of phase 2 detoxifying enzymes in a dose-dependent manner in mouse hepatoma Hepa1c1c7 and BPRc1 cells. Moreover, CZ-5 caused a dose-dependent enhancement of luciferase activity in HepG2-C8 cells generated by stably transfecting antioxidant response element-luciferase gene construct, suggesting that it induce antioxidant/detoxifying enzymes through antioxidant response element (ARE)-mediated transcriptional activation of the relevant genes. CZ-5 also stimulated nuclear accumulation of Nrf2 as evaluated by western blot analysis. In conclusion, our data showed that CZ-5 acted as a potent phase 2 detoxifying enzyme inducer by stimulating the nuclear accumulation of Nrf2 and had potential as chemopreventive agent Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5666. [1]: pending:yes

Abstract 227: Antitumor activity of glyceollins through induction of apoptosis

Glyceollins, which are synthesized from daidzein in soybean infected with fungi, have been shown to have anti-fungal effects and cancer preventive properties. However, the effects of glyceollins on apoptosis of cancer cell are not well known. We have investigated the antitumor effects of glyceollins on murine hepatoma cell lines. Cell viability was tested with a MTT assay. In Hepa1C1C7, glyceollins markedly reduced cell viability which was due to apoptosis. Glyceollins were observed to induce apoptosis of Hepa1C1C7 cells as evidenced by cell-morphological changes, an early redistribution of plasma membrane phosphatidylserine, increases in membrane leakage (propidium iodide) staining), Annexin-V binding, sub G1 phase in the cell cycle, and DNA laddering. Western blot analysis showed that upon treatment of tumor cells with the glyceollins, cleavage of pro-caspases-3 and significant release of mitochondrial cytochrome c into the cytosol were readily observed. This study demonstrated that glyceollins caused cell death via an apoptotic pathway, which might be exploited for prevention of tumor development. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 227.