Evaluating the anticancer potential of Polygonum multiflorum root-derived stilbenes against H2452 malignant pleural mesothelioma cells.

A naturally occurring stilbene, resveratrol, shows promising effects in the treatment of malignant pleural mesothelioma (MPM) both as a single agent and in combination with chemotherapeutic drugs. To discover new anticancer agents targeting MPM, stilbene-targeted isolation was performed on the roots of Polygonum multiflorum Thunb., an herbal medicine rich in stilbene compounds. In this study, seven stilbene glycosides (1-7) were isolated, along with four non-stilbenes (8-11), of which compounds 4 and 9-11 have not previously been isolated from this species. Stiquinoside A (1) is a previously undescribed stilbene glycoside, and its structure was elucidated as (E)-2,3,5,4'-tetrahydroxystilbene 2-O-ß-d-quinovopyranoside based on 1D and 2D-NMR, HR-ESI-MS, and acid hydrolysis experiments. Compounds 1, 4, 6, and 8 significantly inhibit the growth of MPM cancer cells H2452. These results demonstrate the potential utility of stilbenes in new strategies for the treatment of MPM.

Diterpene and biflavone derivatives from Thuja koraiensis and their cytotoxicities against A549 cells.

During the screening of the cytotoxicity of rare Korean endemic plants, the extract of Thuja koraiensis Nakai displayed potent cytotoxicity against the adenocarcinomic human alveolar basal epithelial A549 cell line. Through a series of separations via column chromatography, three undescribed abietanes, an undescribed labdane along with a labdane, and a biflavonoid were purified from methylene chloride (CH2Cl2) fraction possessing a potent cytotoxic effect. Extensive 1D and 2D NMR spectroscopic data analyses, in combination with quantum chemical calculations were conducted to establish the planar and absolute configurations of thujakoraienes A-C. The chemical structure of thujakoraiene D was elucidated by spectroscopic data analysis and competing enantioselective acylation. Thujakoraienes A and C along with 7,7″-di-O-methylamentoflavone, showed cytotoxic effects on A549 cells, with IC50 values of 64.86, 47.97, and 16.14 µM, respectively. Finally, thujakoraiene C and 7,7″-di-O-methylamentoflavone were identified as potent cytotoxic compounds in A549 cells, followed by an additional cytotoxicity test in the normal human lung fibroblast MRC-5 cell line. This is the first study on the non-volatile chemicals in the extract of T. koraiensis and comparison of chemical profiles of T. orientalis and T. koraiensis.

Inhibitory Effects of Euscaphic Acid in the Atopic Dermatitis Model by Reducing Skin Inflammation and Intense Pruritus.

Atopic dermatitis (AD) is a complex and multifactorial skin disease characterized by skin inflammation and intense pruritus. There are many commercially available treatments such as topical corticosteroids and immunosuppressants to treat of AD, but their effectiveness is limited, and frequent use of these treatments can cause serious side effects. Therefore, the development of new therapeutic agents is necessary for the treatment of AD. Hence, an alternative agent that was derived from natural products that are effective and safe for AD treatment was investigated using experimental models. The biological activity of euscaphic acid has anti-inflammatory, anticoagulant, and antioxidant effects. Despite the various biomedical properties of euscaphic acid, its therapeutic effects on AD have not been well studied. In this study, we investigated the effects of euscaphic acid on skin inflammation and pruritus in AD mouse model. The effects of euscaphic acid were investigated in activated human epidermal keratinocytes and leukemia T lymphoblast cell lines, and Dermatophagoides farina extract and 2,4-dinitrochlorobenzene-induced AD mouse model. Euscaphic acid ameliorated AD properties, such as the expression of inflammatory cytokines and activation of transcription factors. In addition, euscaphic acid reduced critical factors for pruritus such as immunoglobulin E hyperproduction, mast cell invasion, and interleukin-33 expression. Taken together, euscaphic acid could be a potent therapeutic agent for the treatment of AD.

Polyozellin alleviates atopic dermatitis-like inflammatory and pruritic responses in activated keratinocytes and mast cells.

Polyozellus multiplex is an edible mushroom that offers beneficial pharmacological effects against intestinal inflammation and cancer. Previous studies have demonstrated that polyozellin, a major component of P. multiplex, has therapeutic activities against inflammation, cancer, and oxidative stress-related disorders. This study aimed to determine the pharmacological effects of polyozellin on inflammatory and pruritic responses, the major symptoms of atopic dermatitis (AD), and to define its underlying mechanism of action. Our results showed that polyozellin inhibited the expression of inflammatory cytokines and chemokines through blockade of signal transducer and activator of transcription 1 and nuclear factor-κB in activated keratinocytes, the major cells involved in AD progression. Based on the histological and immunological analyses, oral treatment with polyozellin attenuated the Dermatophagoides farinae extract (DFE)/2,4-dinitrochlorobenzene (DNCB)-induced atopic inflammatory symptoms in the skin. Pruritus is an unpleasant sensation for AD patients that causes scratching behavior and ultimately exacerbates the severity of AD. To find a possible explanation for the anti-pruritic effects of polyozellin, we investigated its effects on mast cells and mast cell-derived histamines. Oral treatment with polyozellin reduced the DFE/DNCB-induced tissue infiltration of mast cells, the serum histamine levels, and the histaminergic scratching behaviors. Additionally, polyozellin decreased the immunoglobulin E-stimulated degranulation of mast cells. Taken together, the findings of this study provide us with novel insights into the potential pharmacological targets of polyozellin for treating AD by inhibiting the inflammatory and pruritic responses.

1,7-Bis(4-hydroxyphenyl)-4-hepten-3-one from Betula platyphylla induces apoptosis by suppressing autophagy flux and activating the p38 pathway in lung cancer cells.

Betula platyphylla (BP) is frequently administered in the treatment of various human diseases, including cancers. This study was undertaken to investigate the pharmacological function of the active components in BP and the underlying mechanism of its chemotherapeutic effects in human lung cancer cells. We observed that BP extracts and 1,7-bis(4-hydroxyphenyl)-4-hepten-3-one (BE1), one of the components of BP, effectively decreased the cell viability of several lung cancer cell lines. BE1-treated cells exhibited apoptosis induction and cell cycle arrest at the G2/M phase. Further examination demonstrated that BE1 treatment resulted in suppression of autophagy, as evidenced by increased protein expression levels of both LC3 II and p62/SQSTM1. Interestingly, the pharmacological induction of autophagy with rapamycin remarkably reduced the BE1-induced apoptosis, indicating that apoptosis induced by BE1 was associated with autophagy inhibition. Our data also demonstrated that BE1 exposure activated the p38 pathway resulting in regulation of the pro-apoptotic activity. Taken together, we believe that BE1 is a potential anticancer agent for human lung cancer, which exerts its effect by enhancing apoptosis via regulating autophagy and the p38 pathway.

Avenanthramide C from germinated oats exhibits anti-allergic inflammatory effects in mast cells.

Mast cells play a crucial role in allergic diseases via the release of inflammatory mediators, particularly histamine and pro-inflammatory cytokines. Avenanthramide (Avn) C, a polyphenol found mainly in oats, is known to exhibit various biological properties. In this study, we aimed to evaluate the effectiveness of Avn C from germinated oats against mast cell-mediated allergic inflammation. For the in vitro study, RBL-2H3, mouse bone marrow-derived mast cells and rat peritoneal mast cells were used. Avn C (1-100 nM) inhibited the immunoglobulin (Ig)E-stimulated mast cells degranulation by suppressing phosphorylation of phosphoinositide 3-kinase and phospholipase Cγ1 and decreasing intracellular calcium levels. It inhibited IgE-stimulated secretion of inflammatory cytokines via suppression of FcεRI-mediated signaling proteins Lyn, Syk, Akt, and nuclear factor-κB. To verify the effects of Avn C in vivo, ovalbumin-induced active systemic anaphylaxis (ASA) and IgE-mediated passive cutaneous anaphylaxis (PCA) models were used. Oral administration of Avn C dose-dependently attenuated the ASA reactions, as evidenced by the inhibition of hypothermia and reduction of elevated serum histamine, IgE, and interleukin-4 levels. Avn C also inhibited the PCA reactions, such as ear swelling and plasma extravasation. Our results suggested that Avn C from germinated oats might be a possible therapeutic candidate for mast cell-mediated allergic inflammation.

Ethanol extract of Glycyrrhizae Radix modulates the responses of antigen-specific splenocytes in experimental autoimmune encephalomyelitis.

BACKGROUND: Multiple sclerosis (MS) is an autoimmune disorder resulting in paralysis, and the responses of reactive T cells against self-antigens are hallmarks. Glycyrrhizae Radix (GR) has been used for detoxification and reducing inflammation. However, very few reports have described the effects of GR on MS. PURPOSE: The immunomodulatory effects of GR extract on autoimmune responses were evaluated through in vitro, ex vivo, and in vivo assays using primary mouse splenocytes (SPLC), mouse microglia BV2 cell line, and a mouse model of experimental autoimmune encephalomyelitis (EAE). STUDY DESIGN: Ethanol extract of GR was used in vitro with primary SPLC in the condition of anti-CD3/CD28 stimulation and interferon (IFN)-γ-producing CD4+ (TH1)/CD8+ (TC1) polarization as well as IFN-γ-stimulated BV2 cells. For EAE induction, female C57BL/6 mice were immunized with 200 µg of myelin oligodendrocyte glycoprotein (MOG)35-55 without pertussis toxin. EAE SPLC (ex vivo) and EAE mice (in vivo) were treated with GR extract to evaluate the changes in antigen-specific responses. SPLC media containing antigen-specific responses were used to stimulate BV2 cells. RESULTS: GR extract effectively modulated the responses of reactive splenic T cells through the reduction in IFN-γ+ T cell populations, the expressions of cell adhesion molecules (CAMs), and secretions of cytokines containing IFN-γ and a chemokine IFN-γ-induced protein 10 (IP-10) in vitro. In addition, GR extract significantly decreased nitric oxide production and secretion of tumor necrosis factor (TNF)-α and IP-10 in IFN-γ-stimulated BV2 cells. The antigen-specific TH1 and TC1 populations were decreased following administration of 100 mg/kg of GR extract, whereas CD8+IL-17A+ (TC17) population was increased on day 36 after EAE induction. Moreover, IFN-γ, which showed the highest secretion among examined cytokines, and IP-10 decreased on day 36. SPLC media derived from 100 mg/kg GR extract-administered EAE mice revealed the ameliorative effects on BV2 cell stimulation. CONCLUSION: This is the first report on the immunomodulatory effects of GR extract on antigen-specific SPLC responses in EAE. These results could be helpful for the discovery of drug candidates for MS by focusing on IFN-γ-related autoimmune responses.

Simultaneous Determination of Five Cytochrome P450 Probe Substrates and Their Metabolites and Organic Anion Transporting Polypeptide Probe Substrate in Human Plasma Using Liquid Chromatography-Tandem Mass Spectrometry.

A rapid and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of organic anion transporting polypeptide 1B1 (OATP1B1) and cytochrome P450 (P450) probe substrates and their phase I metabolites in human plasma was developed. The OATP1B1 (pitavastatin) and five P450 probe substrates, caffeine (CYP1A2), losartan (CYP2C9), omeprazole (CYP2C19), dextromethorphan (CYP2D6), and midazolam (CYP3A) and their metabolites were extracted from human plasma (50 µL) using methanol. Analytes were separated on a C18 column followed by selected reaction monitoring detection using MS/MS. All analytes were separated simultaneously within a 9 min run time. The developed method was fully validated over the expected clinical concentration range for all analytes tested. The intra- and inter-day precisions for all analytes were lower than 11.3% and 8.82%, respectively, and accuracy was 88.5⁻117.3% and 96.1⁻109.2%, respectively. The lower limit of quantitation was 0.05 ng/mL for dextromethorphan, dextrorphan, midazolam, and 1'-hydroxymidazolam; 0.5 ng/mL for losartan, EXP-3174, omeprazole, 5'-hydroxyomeprazole, and pitavastatin; and 5 ng/mL for caffeine and paraxanthine. The method was successfully used in a pharmacokinetic study in healthy subjects after oral doses of five P450 and OATP1B1 probes. This analytical method provides a simple, sensitive, and accurate tool for the determination of OATP1B1 and five major P450 activities in vivo drug interaction studies.

Esculetin from Fraxinus rhynchophylla attenuates atopic skin inflammation by inhibiting the expression of inflammatory cytokines.

Atopic dermatitis (AD) is a common chronic inflammatory skin disorder afflicting from infancy to adults with itching, scratching, and lichenification. We aimed to investigate the effects of esculetin from Fraxinus rhynchophylla on atopic skin inflammation. For induction of atopic skin inflammation, we exposed the ears of female BALB/c mice to house dust mite (Dermatophagoides farinae extract, DFE) and 2,4-dinitrochlorobenzene (DNCB) for 4 weeks. Oral administration of esculetin reduced the symptoms of DFE/DNCB-induced atopic skin inflammation, which were evaluated based on ear swelling and number of scratch bouts. The immunoglobulin (Ig) E, IgG2a, and histamine levels in serum were decreased and inflammatory cell infiltration in skin tissue was reduced by the esculetin. It suppressed production of Th1, Th2 and Th17-related cytokines such as tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-4, IL-13, IL-31 and IL-17 in the ear tissue. Furthermore, we investigated the effects of esculetin on activated keratinocytes, which are representative cells used for studying the pathogenesis of acute and chronic atopic skin inflammation. As results, esculetin suppressed gene expression of Th1, Th2 and Th17 cytokines and the activation of nuclear factor-κB and signal transducer and activator of transcription 1 in TNF-α/IFN-γ-stimulated keratinocytes. Taken together, these results imply that esculetin attenuated atopic skin inflammation, suggesting that esculetin could be a potential therapeutic candidate for the treatment of AD.

Protective Effect of Isoliquiritigenin against Ethanol-Induced Hepatic Steatosis by Regulating the SIRT1-AMPK Pathway.

Ethanol-induced fat accumulation, the earliest and most common response of the liver to ethanol exposure, may be involved in the pathogenesis of liver diseases. Isoliquiritigenin (ISL), an important constituent of Glycyrrhizae Radix, is a chalcone derivative that exhibits antioxidant, anti-inflammatory, and phytoestrogenic activities. However, the effect of ISL treatment on lipid accumulation in hepatocytes and alcoholic hepatitis remains unclear. Therefore, we evaluated the effect and underlying mechanism of ISL on ethanol-induced hepatic steatosis by treating AML-12 cells with 200 mM ethanol and/or ISL (0~50 µM) for 72 hr. Lipid accumulation was assayed by oil red O staining, and the expression of sirtuin1 (SIRT1), sterol regulatory element-binding protein-1c (SREBP-1c), AMP-activated protein kinase (AMPK), and peroxisome proliferator-activated receptor alpha (PPARα) was studied by western blotting. Our results indicated that ISL treatment upregulated SIRT1 expression and downregulated SREBP-1c expression in ethanol-treated cells. Similarly, oil red O staining revealed a decrease in ethanol-induced fat accumulation upon co-treatment of ethanol-treated cells with 10, 20, and 50 µM of ISL. These findings suggest that ISL can reduce ethanol induced-hepatic lipogenesis by activating the SIRT1-AMPK pathway and thus improve lipid metabolism in alcoholic fatty livers.

Neuroprotective and Anti-Neuroinflammatory Activities of Anthraquinones Isolated from Photorhabdus temperata Culture Broth.

Photorhabdus temperata (PT), a gram-negative bacterium, lives symbiotically within entomopathogenic nematodes. The insecticidal compounds derived from Photorhabdus are used as biopesticides in agriculture. However, the physiological properties are not well characterized. In the course of our screening for neuroprotective and anti-neuroinflammatory substances from natural products, the culture broth of PT showed considerable activities. By activity-guided purification, five anthraquinones, namely, 3-methoxychrysazine (1), 1,3-dimethoxy-8-hydroxy-9,10-anthraquinone (2), 1,3,8-trihydroxy-9,10-anthraquinone (3), 3,8-dihydroxy-1-methoxy-9,10-anthraquinone (4), and 1,3,4-trimethoxy-8-hydroxy-9,10-anthraquinone (5), were isolated from the ethyl acetate fraction of the PT culture broth. Among the isolated compounds, 75 µM 3 significantly protected mouse hippocampal neuronal cells (HT22) against 5 mM glutamate-induced cell death via the inhibition of reactive oxygen species production, Ca²âº influx, and lipid peroxidation. Additionally, 3 and 4 effectively suppressed the interferon-γ-induced neuroinflammation of mouse-derived microglial cells (BV2) at 10 ng/ml, via the reduction of nitric oxide, interleukin-6, and tumor necrosis factor-α. Anthraquinones 3 and 4 derived from the PT culture broth are a potential starting point to discover neuroprotective and anti-neuroinflammatory drug leads. The novel compound 5 is reported for the first time in this study.

Oleuropein isolated from Fraxinus rhynchophylla inhibits glutamate-induced neuronal cell death by attenuating mitochondrial dysfunction.

Glutamate-induced neurotoxicity is related to excessive oxidative stress accumulation and results in the increase of neuronal cell death. In addition, glutamate has been reported to lead to neurodegenerative diseases, including Parkinson's and Alzheimer's diseases.It is well known that Fraxinus rhynchophylla contains a significant level of oleuropein (Ole), which exerts various pharmacological effects. However, the mechanism of neuroprotective effects of Ole is still poorly defined. In this study, we aimed to investigate whether Ole prevents glutamate-induced toxicity in HT-22 hippocampal neuronal cells. The exposure of the glutamate treatment caused neuronal cell death through an alteration of Bax/Bcl-2 expression and translocation of mitochondrial apoptosis-inducing factor (AIF) to the cytoplasm of HT-22 cells. In addition, glutamate induced an increase in dephosphorylation of dynamin-related protein 1 (Drp1), mitochondrial fragmentation, and mitochondrial dysfunction. The pretreatment of Ole decreased Bax expression, increased Bcl-2 expression, and inhibited the translocation of mitochondrial AIF to the cytoplasm. Furthermore, Ole amended a glutamate-induced mitochondrial dynamic imbalance and reduced the number of cells with fragmented mitochondria, regulating the phosphorylation of Drp1 at amino acid residue serine 637. In conclusion, our results show that Ole has a preventive effect against glutamate-induced toxicity in HT-22 hippocampal neuronal cells. Therefore, these data imply that Ole may be an efficient approach for the treatment of neurodegenerative diseases.

Identification of acetylshikonin as the novel CYP2J2 inhibitor with anti-cancer activity in HepG2 cells.

BACKGROUND: Acetylshikonin is one of the biologically active compounds derived from the root of Lithospermum erythrorhizon, a medicinal plant with anti-cancer and anti-inflammation activity. Although there have been a few previous reports demonstrating that acetylshikonin exerts anti-cancer activity in vitro and in vivo, it is still not clear what is the exact molecular target protein of acetylshikonin in cancer cells. PURPOSE: The purpose of this study is to evaluate the inhibitory effect of acetylshikonin against CYP2J2 enzyme which is predominantly expressed in human tumor tissues and carcinoma cell lines. STUDY DESIGN: The inhibitory effect of acetylshikonin on the activities of CYP2J2-mediated metabolism were investigated using human liver microsomes (HLMs), and its cytotoxicity against human hepatoma HepG2 cells was also evaluated. METHOD: Astemizole, a representative CYP2J2 probe substrate, was incubated in HLMs in the presence or absence of acetylshikonin. After incubation, the samples were analyzed by liquid chromatography and triple quadrupole mass spectrometry. The anti-cancer activity of acetylshikonin was evaluated on human hepatocellular carcinoma HepG2 cells. WST-1, cell counting, and colony formation assays were further adopted for the estimation of the growth rate of HepG2 cells treated with acetylshikonin. RESULTS: Acetylshikonin inhibited CYP2J2-mediated astemizole O-demethylation activity (Ki = 2.1µM) in a noncompetitive manner. The noncompetitive inhibitory effect of acetylshikonin on CYP2J2 enzyme was also demonstrated using this 3D structure, which showed different binding location of astemizole and acetylshikonin in CYP2J2 model. It showed cytotoxic effects against human hepatoma HepG2 cells (IC50 = 2µM). In addition, acetylshikonin treatment inhibited growth of human hepatocellular carcinoma HepG2 cells leading to apoptosis accompanied with p53, bax, and caspase3 activation as well as bcl2 down-regulation. CONCLUSION: Taken together, our present study elucidates acetylshikonin displays the inhibitory effects against CYP2J2 in HLMs and anti-cancer activity in human hepatocellular carcinoma HepG2 cells.

ß-Secretase (BACE1) inhibitory and neuroprotective effects of p-terphenyls from Polyozellus multiplex.

Alzheimer's disease (AD), a major neurodegenerative disorder, is associated with the enzymatic reaction of ß-secretase (BACE1) on the amyloid precursor protein (APP) for the generation of neurotoxic amyloid-ß (Aß). Therefore, Aß accumulation and oxidative stress-induced neuronal cell death are the pathogenic hallmarks of AD. In this study, we tried to identify BACE1 inhibitors and neuroprotectants from natural products, in particular, from the Korean mushroom Polyozellus multiplex. Four p-terphenyls were identified from the ethanolic extract of P. multiplex; polyozellin (1), thelephoric acid (2), polyozellic acid (3), and kynapcin-12 (4). Compounds 1-4 effectively inhibited BACE1 activity with a half-maximal inhibitory concentration (IC50) of 3.08, 3.50, 4.78, and 15.79 µM, respectively. Compounds 1-3 reduced the production of neurotoxic Aß1-42 production in APPswe-N2a cells in a concentration-dependent manner. When HT22 cells were stressed with 5 mM glutamate, compounds 2 and 3 significantly recovered cell viability. It was correlated with their inhibitory properties against glutamate-mediated Ca(2+) influx, intracellular reactive oxygen species (ROS) generation, lipid peroxidation, reduction in Bcl-2 and Bid levels, and enhanced phosphorylation of mitogen-activated protein kinase (MAPK). Thus, P. multiplex and the isolated p-terphenyls might be useful in the development of lead compounds for the prevention of neurodegenerative diseases, especially AD.

Ameliorative effect of a rarely occurring C-methylrotenoid on HMGB1-induced septic responses in vitro and in vivo.

The ubiquitous nuclear protein, high mobility group box-1 (HMGB1) functions as a late mediator of sepsis. A new rarely occurring C-methylrotenoid, named boeravinone X (comp 1), was isolated and identified from Abronia nana suspension cultures during our continuous works on the discovery of anti-septic natural products. Here, we investigated the antiseptic effects and underlying mechanisms of comp 1 against HMGB1-mediated septic responses. According to the results, comp 1 effectively inhibited lipopolysaccharide (LPS)-induced release of HMGB1, and suppressed HMGB1-mediated septic responses, such as hyperpermeability, adhesion and migration of leukocytes, and expression of cell adhesion molecules. And, comp 1 suppressed the production of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6), and the activation of nuclear factor-κB (NF-κB) and extracellular signal-regulated kinases 1 and 2 (ERK1/2) by HMGB1. Collectively, these results indicate that comp 1 could be potential therapeutic agents for the treatment of various severe vascular inflammatory diseases via inhibition of the HMGB1 signaling pathway.

Enhanced anti-oxidative effect of fermented Korean mistletoe is originated from an increase in the contents of caffeic acid and lyoniresinol.

Viscum album var. coloratum (Korean mistletoe; KM) is an herbal medicine that is used worldwide for the treatment of various immunological disorders and cancers. KM extract showed enhanced anti-oxidative effects in 2,2-diphenyl-1-picrylhydrazyl, Trolox equivalent antioxidant capacity, and 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate acetyl ester assays after being fermented with a crude enzyme extract from a soybean paste fungus, Aspergillus kawachii. High-performance liquid chromatography analysis showed four increased peaks in enzyme treated KM. The increased peaks were isolated and identified as caffeic acid (1), hesperetin (2), syringaldehyde (3), and lyoniresinol (4). Among the four compounds, only 1 and 4 showed strong anti-oxidative activity. Therefore, the fermentation increased the contents of 1 and 4, which consequently increased the anti-oxidative activity of KM.

Polyozellin, a key constituent of the edible mushroom Polyozellus multiplex, attenuates glutamate-induced mouse hippocampal neuronal HT22 cell death.

Polyozellus multiplex (PM), a Korean edible mushroom, has biological activities such as chemoprevention of stomach cancer, inhibition of lipid peroxidation, and reduction of prolyl endopeptidase activity. However, there are little reports on the protective effects of PM or its constituents against glutamate-induced mouse hippocampal neuronal cell (HT22) death. In this study, polyozellin (PZ), a key constituent of PM, was applied to glutamate-treated HT22 cells to evaluate its neuroprotective mechanisms. PZ (25 µM) dramatically increased the HT22 cell viability when the cell death was induced by 5 mM glutamate for 12 h, which was mediated by inhibition of Ca(2+) influx, intracellular reactive oxygen species (ROS) production, and lipid peroxidation. PZ also regulated expression of Bid, Bcl-2, and apoptosis-inducing factor (AIF), as well as phosphorylation of mitogen-activated protein kinases (MAPKs). These data suggest that PM and its constituent PZ might be useful for prevention and treatment of neurodegenerative disorders.

Inhibitory effects of polyozellin from Polyozellus multiplex on HMGB1-mediated septic responses.

AIM AND OBJECTIVE: The ubiquitous nuclear protein, high-mobility group box 1 (HMGB1), is released by activated macrophages and human umbilical vein endothelial cells (HUVECs) and functions as a late mediator of experimental sepsis. Polyozellin, which has been reported to have a variety of biological activities including antioxidant and anticancer activity, is the major active compound found in edible mushroom (Polyozellus multiplex). In this study, we investigated the antiseptic effects and underlying mechanisms of polyozellin against HMGB1-mediated septic responses in HUVECs and mice. METHODS: The anti-inflammatory activities of polyozellin were determined by measuring permeability, human neutrophil adhesion and migration, and activation of proinflammatory proteins in HMGB1-activated HUVECs and mice. RESULTS: According to the results, polyozellin effectively inhibited lipopolysaccharide (LPS)-induced release of HMGB1, and suppressed HMGB1-mediated septic responses, such as hyperpermeability, adhesion and migration of leukocytes, and expression of cell adhesion molecules. In addition, polyozellin suppressed the production of tumor necrosis factor-α and interleukin (IL)-6, and the activation of nuclear factor-κB and extracellular signal-regulated kinases 1/2 by HMGB1. CONCLUSION: Collectively, these results indicate that P. multiplex containing polyozellin could be commercialized as functional food for preventing and treatment of various severe vascular inflammatory diseases via inhibition of the HMGB1 signaling pathway.

Protective effect of dieckol against chemical hypoxia-induced cytotoxicity in primary cultured mouse hepatocytes.

Hepatic ischemic injury is a major complication arising from liver surgery, transplantation, or other ischemic diseases, and both reactive oxygen species (ROS) and pro-inflammatory mediators play the role of key mediators in hepatic ischemic injury. In this study, we examined the effect of dieckol in chemical hypoxia-induced injury in mouse hepatocytes. Cell viability was significantly decreased after treatment with cobalt chloride (CoCl2), a well-known hypoxia mimetic agent in a time- and dose-dependent manner. Pretreatment with dieckol before exposure to CoCl2 significantly attenuated the CoCl2-induced decrease of cell viability. Additionally, pretreatment with dieckol potentiated the CoCl2-induced decrease of Bcl-2 expression and attenuated the CoCl2-induced increase in the expression of Bax and caspase-3. Treatment with CoCl2 resulted in an increased intracellular ROS generation, which is inhibited by dieckol or N-acetyl cysteine (NAC, a ROS scavenger), and p38 MAPK phosphorylation, which is also blocked by dieckol or NAC. In addition, dieckol and SB203580 (p38 MAPK inhibitor) increased the CoCl2-induced decrease of Bcl-2 expression and decreased the CoCl2-induced increase of Bax and caspase-3 expressions. CoCl2-induced decrease of cell viability was attenuated by pretreatment with dieckol, NAC, and SB203580. Furthermore, dieckol attenuated CoCl2-induced COX-2 expression. Similar to the effect of dieckol, NAC also blocked CoCl2-induced COX-2 expression. Additionally, CoCl2-induced decrease of cell viability was attenuated not only by dieckol and NAC but also by NS-398 (a selective COX-2 inhibitor). In conclusion, dieckol protects primary cultured mouse hepatocytes against CoCl2-induced cell injury through inhibition of ROS-activated p38 MAPK and COX-2 pathway.

Potential of decursin to inhibit the human cytochrome P450 2J2 isoform.

CYP2J2 enzyme is highly expressed in human tumors and carcinoma cell lines, and epoxyeicosatrienoic acids, CYP2J2-mediated metabolites, have been implicated in the pathologic development of human cancers. To identify a CYP2J2 inhibitor, 50 natural products obtained from plants were screened using astemizole as a CYP2J2 probe substrate in human liver microsomes. Of these, decursin noncompetitively inhibited CYP2J2-mediated astemizole O-demethylation and terfenadine hydroxylation activities with Ki values of 8.34 and 15.8µM, respectively. It also showed cytotoxic effects against human hepatoma HepG2 cells in a dose-dependent manner while it did not show cytotoxicity against mouse hepatocytes. The present data suggest that decursin is a potential candidate for further evaluation for its CYP2J2 targeting anti-cancer activities. Studies are currently underway to test decursin as a potential therapeutic agent for cancer.