Enhancement of the apoptotic effects of Arctii Fructus extracts on cancer cells by the enzymatic bioconversion of lignans.

The fruit of Arctium lappa L. (Arctii Fructus) is one of the most popularly used medicinal plant components in Asia. To enhance the functionality of Arctii Fructus extract, a bioconversion method was developed to produce arctigenin from arctiin. Treatment with ß-glucosidase increased the arctigenin content by >5 fold in Arctii Fructus extracts. The bioconversion products enhanced the apoptosis of cancer cells. The cell viabilities of gefitinib-resistant lung cancer HCC827 (HCC827GR) cells and colon cancer cells (DLD1) were decreased by 40% and 35%, respectively. The bioconversion products also decreased anchorage-independent growth of cancer cells. In addition, the increase of apoptosis in cancer cells by bioconversion was confirmed by the flow cytometry analysis. These results indicated that arctigenin exerts anticancer effects on lung and colon cancer cells and that Arctii Fructus can potentially function as a chemopreventive agent. In addition, bioconverted Arctii Fructus extract displayed higher anticancer activity than the same levels of purified arctigenin, indicating the advantage of consuming Arctii Fructus itself as a food or medicinal material.

Arctium lappa root extract containing L-arginine prevents TNF-α-induced early atherosclerosis in vitro and in vivo.

Atherosclerosis is a chronic inflammatory disease affecting the aorta and is a major cause of cardiovascular disease. Arctium lappa root is a plant widely used in traditional Chinese medicine (TCM), and Arctium lappa root extract (ALE) has been reported to exhibit anti-inflammatory capacity and to ameliorate endothelial dysfunction. Thus, we hypothesized that ALE would inhibit the early atherosclerotic stage. In this study, we evaluated the inhibitory effect of ALE on early arteriosclerosis and its mechanisms of action. ALE suppressed TNF-α-induced monocyte adhesion to the vascular endothelium by suppressing NF-κB signaling in HUVECs. In an acute mouse model of atherosclerosis, ALE suppressed TNF-α-induced monocyte infiltration of the vascular endothelium and the expression of genes encoding inflammatory cytokines including IL-1ß, IL-6, TNF-α, and MCP-1 in the mouse aorta. Moreover, inulin-type fructan and amino acids, especially L-aspartate and L-arginine (60.27 and 42.17 mg/g, respectively) were detected by NMR, MALDI-TOF MS, and HPLC analysis as the main components of ALE. Notably, L-arginine suppressed TNF-α-induced monocyte adhesion to HUVECs. Therefore, these results suggest that ALE may be a functional food for the suppression or prevention of early stages of atherosclerosis.

Syringic acid prevents skin carcinogenesis via regulation of NoX and EGFR signaling.

Validation of nutraceutical and pharmaceutical targets is essential for the prediction of physiological and side effects. Epidemiologic evidence and molecular studies suggest that non-melanoma skin cancer is directly associated with excessive exposure to ultraviolet (UV) radiation. The aim of the present study was to evaluate the inhibitory effects of syringic acid on UVB-induced signaling and skin carcinogenesis, and determine the molecular targets. Treatment of human epidermal keratinocytes (HaCaT) cells with syringic acid resulted in the suppression of UVB-induced cyclooxygenase-2, matrix metalloproteinase-1, and prostaglandin E2 expression as well as activator protein-1 activity. Moreover, syringic acid inhibited the UVB-induced phosphorylation of mitogen-activated protein kinases and Akt signaling pathways as well as epidermal growth factor receptor (EGFR). Syringic acid treatment further inhibited intracellular reactive oxygen species and protein-tyrosine phosphatase-κ activity, a regulator of EGFR activation. Syringic acid and the antioxidant N-acetyl-l-cysteine inhibited UVB-induced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity. In vivo, pretreatment of mouse skin with syringic acid significantly suppressed UVB-induced skin tumor incidence in a dose-dependent manner. Overall, these results indicate that syringic acid exerts potent chemopreventive activity in skin carcinogenesis mainly by inhibition of the Nox/PTP-κ/EGFR axis. Syringic acid might serve as an effective chemopreventive and therapeutic agent against UVB-mediated skin cancer.

Ultrasonicated Lespedeza cuneata extract prevents TNF-α-induced early atherosclerosis in vitro and in vivo.

This study evaluated the use of ultrasonication to extract Lespedeza cuneata as a potential nutraceutical for preventing vascular inflammation. Ultrasonicated L. cuneata extract (ULCE) was prepared using 20% ethanol and 2 h of ultrasonication at room temperature, and its effects were investigated using relevant in vitro and in vivo models. ULCE suppressed tumor necrosis factor-alpha (TNF-α)-induced adhesion capacity, vascular cell adhesion protein 1 (VCAM-1) expression, and nuclear factor kappa-B (NF-κB) activity in human umbilical vein endothelial cells (HUVECs). ULCE also suppressed TNF-α-induced NF-κB signaling pathways and p65 translocation from the cytosol to the nucleus, as well as the mRNA expression of IL-1ß, IL-6, and TNF-α in HUVECs. Oral administration of ULCE suppressed TNF-α-induced monocyte infiltration into the intima and VCAM-1 expression, as well as the IL-1ß, IL-6, TNF-α, and monocyte chemoattractant protein-1 (MCP-1) mRNA expression in the main artery in mice. Among the compounds identified in the hydrolyzed ULCE, quercetin exhibited the strongest inhibitory effect against TNF-α-induced cell adhesion capacity. These results demonstrate that ULCE contains potent preventive factors against early atherosclerosis, which act by suppressing the NF-κB and VCAM-1 signaling axis.

1,8-cineole prevents UVB-induced skin carcinogenesis by targeting the aryl hydrocarbon receptor.

1,8-cineole is a natural monoterpene cyclic ether present in Eucalyptus, and has been reported to exhibit anti-inflammatory and antioxidant effects. However, the preventive effect of 1,8-cineole on skin carcinogenesis and the molecular mechanism of action responsible remains unknown. In the present study, we investigated the effect of 1,8-cineole on UVB-induced skin carcinogenesis. 1,8-cineole inhibited UVB-induced cyclooxygenase-2 (COX-2) protein and mRNA expression and prostaglandin E2 (PGE2) generation in HaCaT cells. 1,8-cineole also inhibited phosphorylation of extracellular signal-regulated kinase (ERK) 1/2, and phosphorylation of its upstream kinases, c-Src and epidermal growth factor receptor (EGFR). Quantitative real-time RT-PCR (qRT-PCR) and drug affinity responsive target stability (DARTS) assay results showed that 1,8-cineole suppressed UVB-induced expression of a target gene of the aryl hydrocarbon receptor (AhR), cyp1a1, and directly binds to AhR. Knockdown of AhR suppressed COX-2 expression as well as phosphorylation of ERK1/2 in HaCaT cells. Furthermore, topical treatment of 1,8-cineole on mouse skin delayed tumor incidence and reduced tumor numbers, while inhibiting COX-2 expression in vivo. Taken together, these results suggest that 1,8-cineole is a potent chemopreventive agent that inhibits UVB-induced COX-2 expression by targeting AhR to suppress UVB-induced skin carcinogenesis.

Cyanidin-3-glucoside isolated from mulberry fruit protects pancreatic ß-cells against oxidative stress-induced apoptosis.

The extract obtained from berries contains high amounts of anthocyanins, and this extract is used as a phytotherapeutic agent for different types of diseases. In this study, we examined the cytoprotective effects of cyanidin-3-glucoside (C3G) isolated from mulberry fruit against pancreatic ß-cell apoptosis caused by hydrogen peroxide (H2O2)-induced oxidative stress. The MIN6 pancreatic ß-cells were used to investigate the cytoprotective effects of C3G on the oxidative stress-induced apoptosis of cells. Cell viability was examined by MTT assay and lipid peroxidation was assayed by thiobarbituric acid (TBA) reaction. Immunofluorescence staining, flow cytometry and western blot analysis were also used to determine apoptosis and the expression of proteins associated with apoptosis. Our results revealed that H2O2 increased the rate of apoptosis by stimulating various pro-apoptotic processes, such as the generation of intracellular reactive oxygen species (ROS), lipid peroxidation, DNA fragmentation and caspase-3 activation. However, C3G reduced the H2O2-induced cell death in the MIN6N pancreatic ß-cells. In addition, we confirmed that H2O2 activated mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK) and p38 MAPK. C3G inhibited the phosphorylation of ERK and p38 without inducing the phosphorylation of JNK. Furthermore, C3G regulated the intrinsic apoptotic pathway-associated proteins, such as proteins belonging to the Bcl-2 family, cytochrome c and caspase-3. Taken together, our results suggest that C3G isolated from mulberry fruit has potential for use as a phytotherapeutic agent for the prevention of diabetes by preventing oxidative stress-induced ß-cell apoptosis.

Cyanidin-3-glucoside isolated from mulberry fruits protects pancreatic ß-cells against glucotoxicity-induced apoptosis.

The present study investigated the cytoprotective effects of cyanidin­3­glucoside (C3G), isolated from mulberry fruits, on the glucotoxicity­induced apoptosis of pancreatic ß­cells to evaluate the antidiabetic effects of this compound. MIN6N pancreatic ß­cells were used to investigate the cytoprotective effects of C3G. In addition, the effects of C3G on the glucotoxicity­induced apoptosis of pancreatic ß­cells was evaluated using MTT assay, immunofluorescent staining, flow cytometric and western blot analyses. The pancreatic ß­cells cultured under high glucose conditions exhibited distinct apoptotic features. C3G decreased the generation of intracellular reactive oxygen species, DNA fragmentation and the rate of apoptosis. C3G also prevented pancreatic ß­cell apoptosis induced by high glucose conditions by interfering with the intrinsic apoptotic pathways. In addition, C3G treatment resulted in increased insulin secretion compared with treatment with high glucose only. In conclusion, the results of the present study suggested that C3G obtained from mulberry fruits may be a potential phytotherapeutic agent for the prevention of diabetes.

Neuroprotective effect of prenylated arylbenzofuran and flavonoids from morus alba fruits on glutamate-induced oxidative injury in HT22 hippocampal cells.

A prenylated arylbenzofuran and six flavonoids were isolated from the fruits of Morus alba L. through silica gel, octadecyl silica gel, and Diaion HP-20 column chromatography. Based on the nuclear magnetic resonance, mass spectrometry, and infrared spectroscopic data, the chemical structures of the compounds were determined to be artoindonesianin O (1), isobavachalcone (2), morachalcone A (3), quercetin (4), astragalin (5), isoquercetin (6), and rutin (7). The isolated compounds were evaluated for protection of HT22-immortalized hippocampal cells against glutamate-induced oxidative stress. Compounds 1 and 3 exhibited protective effects with EC(50) values of 19.7±1.2 and 35.5±2.1 µM, respectively. The major compounds 1-3 and 7 were quantified using liquid chromatography/mass spectrometry analysis and were determined to be 1.88±2.1, 1.90±1.8, 0.78±1.5, and 37.29±2.2 mg/kg, respectively, in the ethanol extract of M. alba L. fruits.

Mulberry fruit extract protects pancreatic ß-cells against hydrogen peroxide-induced apoptosis via antioxidative activity.

Among the many environmental stresses, excessive production of reactive oxygen species (ROS) and the ensuring oxidative stress are known to cause significant cellular damage. This has clinical implications in the onset of type 1 diabetes, which is triggered by the destruction of pancreatic ß-cells and is associated with oxidative stress. In this study, we investigated the protective and antioxidative effects of mulberry extract (ME) in insulin-producing pancreatic ß-cells. We found that ME protects pancreatic ß-cells against hydrogen peroxide (H2O2)-induced oxidative stress and the associated apoptotic cell death. ME treatment significantly reduced the levels of H2O2-induced 2-diphenyl-1-picrylhydrazyl (DPPH) radicals, and lipid peroxidation and intracellular ROS accumulation. In addition, ME inhibited DNA condensation and/or fragmentation induced by H2O2. These results suggest that ME protects pancreatic ß-cells against hydrogen peroxide-induced oxidative stress.

Effects of mulberry ethanol extracts on hydrogen peroxide-induced oxidative stress in pancreatic ß-cells.

Reactive oxygen species (ROS) are key mediators of mammalian cellular damage and are associated with diseases such as aging, arteriosclerosis, inflammation, rheumatoid arthritis and diabetes. Type 1 diabetes develops upon the destruction of pancreatic ß-cells, which is partly due to ROS activity. In this study, we investigated the cytoprotective and anti-oxidative effects of fractionated mulberry extracts in mouse insulin-producing pancreatic ß-cells (MIN6N cells). Treatment with hydrogen peroxide (H2O2) induced significant cell death and increased intracellular ROS levels, lipid peroxidation and DNA fragmentation in the MIN6N cells. Fractionated mulberry extracts significantly reduced the H2O2-dependent production of intracellular ROS, 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals and lipid peroxidation. In addition, mulberry extracts inhibited DNA fragmentation induced by H2O2. Thus, the antioxidant properties of mulberry extracts in pancreatic ß-cells may be exploited for the prevention or treatment of type 1 diabetes.

Optimization of ultrasonic extraction of phenolic antioxidants from green tea using response surface methodology.

Response surface methodology (RSM) has been used to optimize the extraction conditions of antioxidants with relatively low caffeine content from green tea by using ultrasonic extraction. The predicted optimal conditions for the highest antioxidant activity and minimum caffeine level were found at 19.7% ethanol, 26.4 min extraction time, and 24.0 ° C extraction temperature. In the predicted optimal conditions, the experimental values were very close to the predicted values. Moreover, the ratio of (EGCg + ECg)/EGC was identified a major factor contributing to the antioxidant activity of green tea extracts. In this study, ultrasonic extraction showed that the ethanol concentration and extraction time used for antioxidant extraction could be remarkably reduced without a decrease in antioxidant activity compared to the conventional extraction conditions.

Anti-obesity effect of Artemisia capillaris extracts in high-fat diet-induced obese rats.

This study evaluated the anti-obesity effects of Artemisia capillaris extracts in high-fat diet (HFD)-induced obese rats. After six weeks feeding with HFD, Wistar male rats (12-weeks-old) were divided into three groups: HFD-control group and HFD mixed with 0.4% and 0.8% Artemisia capillaris extracts treated groups. After seven weeks of treatments, the body weight gain of the 0.4% and 0.8% A. capillaris extracts treated groups were significantly less than that of the HFD-control group by 11.8% and 15.4%, respectively. Also, A. capillaris extracts treated groups showed significantly lower serum TG, TC and LDL-c levels in a dose-related manner, while causing the reverse effect in serum HDL-c, and exhibited a hepatoprotective effects in vivo, indicated by reduced hepatic lipid contents, and serum ALT and AST levels. These results show that A. capillaris extracts may prevent body weight increases and improve dyslipidemia in HFD-induced obese rats by enhancing their lipid metabolism.

Effects of the Cynanchum wilfordii Ethanol Extract on the Serum Lipid Profile in Hypercholesterolemic Rats.

The purpose of this study was to investigate the effects of the ethanol extract of Cynanchum wilfordii (ECW) on the blood lipid profile of hypercholesterolemic rats. Thirty 7-week-old male Sprague-Dawley rats were allowed free access to either a normal diet (AIN-93 diet), or 1% high-cholesterol diet with or without 0.5% or 1% ECW for 5 weeks. After sacrifice, the rat serum lipid profile was analyzed. The diets containing ECW decreased body weight gains compared to the normal diet. Serum HDL-cholesterol levels of ECW-fed groups were significantly increased in the hypercholesterolemic groups and normal groups (P<0.05). When 1% ECW was fed to the normal group, total cholesterol level was increased. Moreover, treatment of ECW in hypercholesterolemic groups yielded a dose-dependent and highly significant decrease in the atherogenic index as compared to the control. These results suggest that intake of Cynanchum wilfordii may help reduce the risks of hypercholesterolemia by increasing blood HDL-cholesterol and lowering the atherogenic index.

Process development for mycelial growth and polysaccharide production in Tricholoma matsutake liquid culture.

In this study, the effects of agitation and aeration on mycelial growth and exo-polysaccharide production were examined in batch cultures of Tricholoma matsutake. Agitation was varied from 100 to 300 rpm and aeration was varied from 0.5 to 1.5 vvm. Mycelial growth was 21.87 g/l at 150 rpm, and exo-polysaccharide production was 8.79 g/l at 1.5 vvm. When we analyzed the polysaccharide extractions from the cultured mycelium and the culture broth of T. matsutake, 1.4 g of crude polysaccharide was found per 100 g of dried weight in the cultured mycelium, and 1.47 g/l of polysaccharides was found in the culture broth. In addition, the amounts of beta-Glucan in the soluble polysaccharide fractions of the cultured mycelium and culture broth were 75.4% and 83.6%, respectively. The cultured mycelium and the culture broth contained a higher amount of beta-Glucan than that of the fruiting body.

Flavonoids from the buds of Rosa damascena inhibit the activity of 3-hydroxy-3-methylglutaryl-coenzyme a reductase and angiotensin I-converting enzyme.

Rosa damascena has been manufactured as various food products, including tea, in Korea. A new flavonoid glycoside, kaempferol-3-O-beta-D-glucopyranosyl(1-->4)-beta-D-xylopyranoside, named roxyloside A was isolated from the buds of this plant, along with four known compounds, isoquercitrin, afzelin, cyanidin-3-O-beta-glucoside, and quercetin gentiobioside. The chemical structures of these compounds were determined by spectroscopic analyses, including FAB-MS, UV, IR, (1)H and (13)C NMR, DEPT, and 2D NMR (COSY, HSQC, and HMBC). All the isolated compounds except cyanidin-3-O-beta-glucoside exhibited high levels of inhibitory activity against 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase with IC(50) values ranging from 47.1 to 80.6 microM. Cyanidin-3-O-beta-glucoside significantly suppressed angiotensin I-converting enzyme (ACE) activity, with an IC(50) value of 138.8 microM, while the other four compounds were ineffective. These results indicate that R. damascena and its flavonoids may be effective to improve the cardiovascular system.

Study on immunostimulating activity of macrophage treated with purified polysaccharides from liquid culture and fruiting body of Lentinus edodes.

Lentinus edodes is a well-known edible and medicinal mushroom used in Oriental cultures. Recently, L. edodes has attracted a lot of attention owing to its antifungal activity, antibacterial activity, antiviral activity, hepatoprotective effect, antitumor activities, and immunomodulatory and cytotoxic effects. In this study, the water-soluble crude polysaccharides, CPF and CPB, which were obtained from the fruiting body and culture cell-free broth of L. edodes by hot-water extraction and ethanol precipitation, were fractionated by DEAE cellulose and Sepharose CL-6B column chromatography, resulting in six polysaccharide fractions, CPFN-G-I, CPFN-G-II, CPFN-G-III, CPFA-G, CPBN-G, and CPBA-G. Among these fractions, CPFN-GI, CPBN-G, and CPBA-G were shown to stimulate the functional activation of macrophages including NO production cytokine expression and phagocytosis.

Structural characteristics of immunostimulating polysaccharides from lentinus edodes.

There is a significant amount of experimental evidence suggesting that polysaccharides from mushrooms enhance the host immune system by activating various mechanisms in immune cells, including macrophages. In this study, polysaccharides from Lentinus edodes were found to stimulate the functional activation of macrophages to secrete inflammatory mediators and cytokines and increase the phagocytotic uptake. The chemical properties of the stimulatory polysaccharides, CPFN-G-I, CPBN-G, and CPBA-G, were determined based on their monosaccharide composition, which mainly consisted of glucose and mannose. According to FT-IR and GC/MS, the structure of CPFN-G-I, purified from the fruiting body of L. edodes, was found to consist of a beta-1,6-branched-beta-1,4-glucan, whereas CPBN-G and CPBA-G, purified from the liquid.

Functional activation of macrophages, monocytes and splenic lymphocytes by polysaccharide fraction from Tricholoma matsutake.

Mushroom-derived polysaccharides (beta-glucans) are considered as a valuable biopharmaceutical principle without displaying side effects. Although Tricholoma matsutake is well-known mushroom in Korea, Japan and China, the immunoregulatory roles of T. matsutake-derived polysaccharides were not fully elucidated yet. In this study, we continued to evaluate the immunomodulatory effect of T. matsutake-derived polysaccharide fraction (TmC-2) using functional activation models of macrophages, monocytes and splenic lymphocytes. TmC-2 treatment strongly increased the production of NO and TNF-alpha. Phagocytic uptake and ROS generation was also up-regulated by TmC-2. Interestingly, TmC-2 stimulated CD29-mediated cell-cell or cell-finbronectin adhesions in monocytes, while CD43-mediated cell adhesion was down-regulated. Interestingly, the enhancement of proliferation and IFN-gamma production was striking observed in TmC-2-treated splenic lymphocytes. The activation seemed to be mediated by up-regulating intracellular signaling cascades such as PI3K/Akt and MAPK (ERK and p38) and by the involvement of surface receptors (dectin-1 and TLR-2). Therefore, our results suggest that this TmC-2 from T. matsutake can be developed as a promising immunostimulatory principle, applicable to people with lowered immunomodulatory potentials.

Antifatigue effect of Rubus coreanus Miquel extract in mice.

The antifatigue properties of six Korean medicinal herb extracts were studied by evaluating forced swimming capacity and biochemical parameters in ICR mice. The treatment groups were orally administered 30% ethanolic extracts (500 mg/kg/day) of Rubus coreanus Miquel, Cyperus rotundus Linn., Acanthopanax sessiliflorus, Saururus chinensis Baili, Epimedium koreanumNakai, or Houttuynia cordata Thunb. for 4 weeks. Swimming time to exhaustion was found to be longer for the group fed R. coreanus than for the control group (P < .05). No significant differences were found in the plasma levels of either glucose or lactate between the control group and the group fed R. coreanus, which swam longer than the control. The plasma ammonia levels were significantly lower in the groups fed R. coreanus and A. sessiliflorus, when compared to the control group (P < .05). No significant differences were found in gastrocnemius muscle or liver glycogen content between the control group and any treatment group. These results suggest that R. coreanus extract, and none of the other herbs, has antifatigue effects in mice, as demonstrated by the increased forced swimming capacity and decreased plasma ammonia accumulation.

Cardiovascular protective properties of kiwifruit extracts in vitro.

It is currently accepted that the consumption of fruit-derived antioxidants such as vitamin C, carotenoids, and flavonoids provides a preventive effect against cardiovascular disease. The purpose of the present study was to investigate potential cardiovascular protective properties of aqueous and 70% ethanol extracts from kiwifruit by analyzing the antioxidative, antihypertensive, hypocholesterolemic, and fibrinolytic activities in vitro. Aqueous and 70% ethanol extracts at 50 mg/ml showed DPPH-radical scavenging activities of 72.31% and 70.75%, respectively. Total antioxidant activity in linoleic acid emulsion was 85-88% at 10 mg/ml and 96-98% at 50 mg/ml of kiwifruit extract. Inhibitory activities against angiogensin I-converting enzyme of kiwifruit extracts were 21-26% at 10 mg/ml and 46-49% at 50 mg/ml, and inhibitory activities on HMG-CoA reductase were 13-14% at 10 mg/ml and 19-30% at 50 mg/ml. Fibrinolytic activity of kiwifruit was also observed at a high concentration of 100 mg/ml in both aqueous and 70% EtOH extracts. Based on our results, kiwifruit have potential cardiovascular protective properties in vitro.