Anti-Inflammatory Activity of Glabralactone, a Coumarin Compound from Angelica sinensis, via Suppression of TRIF-Dependent IRF-3 Signaling and NF-κB Pathways.

The dried root of Angelica sinensis (A. sinensis) has been widely used in Chinese traditional medicine for various diseases such as inflammation, osteoarthritis, infections, mild anemia, fatigue, and high blood pressure. Searching for the secondary metabolites of A. sinensis has been mainly conducted. However, the bioactivity of coumarins in the plant remains unexplored. Therefore, this study was designed to evaluate the anti-inflammatory activity of glabralactone, a coumarin compound from A. sinensis, using in vitro and in vivo models, and to elucidate the underlying molecular mechanisms of action. Glabralactone effectively inhibited nitric oxide production in lipopolysaccharide- (LPS-) stimulated RAW264.7 macrophage cells. The downregulation of LPS-induced mRNA and protein expression of iNOS, TNF-α, IL-1ß, and miR-155 was found by glabralactone. The activation of NF-κB and TRIF-dependent IRF-3 pathway was also effectively suppressed by glabralactone in LPS-stimulated macrophages. Glabralactone (5 and 10 mg/kg) exhibited an in vivo anti-inflammatory activity with the reduction of paw edema volume in carrageenan-induced rat model, and the expressions of iNOS and IL-1ß proteins were suppressed by glabralactone in the paw soft tissues of the animal model. Taken together, glabralactone exhibited an anti-inflammatory activity in in vitro and in vivo models. These findings reveal that glabralactone might be one of the potential components for the anti-inflammatory activity of A. sinensis and may be prioritized in the development of a chemotherapeutic agent for the treatment of inflammatory diseases.

Amentoflavone Promotes Cellular Uptake and Degradation of Amyloid-Beta in Neuronal Cells.

Deposition of fibrillar forms of amyloid ß-protein (Aß) is commonly found in patients with Alzheimer's disease (AD) associated with cognitive decline. Impaired clearance of Aß species is thought to be a major cause of late-onset sporadic AD. Aß secreted into the extracellular milieu can be cleared from the brain through multiple pathways, including cellular uptake in neuronal and non-neuronal cells. Recent studies have showed that the naturally-occurring polyphenol amentoflavone (AMF) exerts anti-amyloidogenic effects. However, its effects on metabolism and cellular clearance of Aß remain to be tested. In the present study, we demonstrated that AMF significantly increased the cellular uptake of both Aß1-40 and Aß1-42, but not inverted Aß42-1 in mouse neuronal N2a cells. Though AMF promoted internalization of cytotoxic Aß1-42, it significantly reduced cell death in our assay condition. Our data further revealed that the internalized Aß is translocated to lysosomes and undergoes enzymatic degradation. The saturable kinetic of Aß uptake and our pharmacologic experiments showed the involvement of receptor-mediated endocytosis, in part, through the class A scavenger receptors as a possible mechanism of action of AMF. Taken together, our findings indicate that AMF can lower the levels of extracellular Aß by increasing their cellular uptake and clearance, suggesting the therapeutic potential of AMF for the treatment of AD.

Antitumor Activity of Americanin A Isolated from the Seeds of Phytolacca americana by Regulating the ATM/ATR Signaling Pathway and the Skp2-p27 Axis in Human Colon Cancer Cells.

The antiproliferative and antitumor activities of americanin A (1), a neolignan isolated from the seeds of Phytolacca americana, were investigated in human colon cancer cells. Compound 1 inhibited the proliferation of HCT116 human colon cancer cells both in vitro and in vivo. The induction of G2/M cell-cycle arrest by 1 was concomitant with regulation of the ataxia telangiectasia-mutated/ATM and Rad3-related (ATM/ATR) signaling pathway. Treatment with 1 activated ATM and ATR, initiating the subsequent signal transduction cascades that include checkpoint kinase 1 (Chk1), checkpoint kinase 2 (Chk2), and tumor suppressor p53. Another line of evidence underlined the significance of 1 in regulation of the S phase kinase-associated protein 2 (Skp2)-p27 axis. Compound 1 targeted selectively Skp2 for degradation and thereby stabilized p27. Therefore, compound 1 suppressed the activity of cyclin B1 and its partner cell division cycle 2 (cdc2) to prevent entry into mitosis. Furthermore, prolonged treatment with 1 induced apoptosis by producing excessive reactive oxygen species. The intraperitoneal administration of 1 inhibited the growth of HCT116 tumor xenografts in nude mice without any overt toxicity. Modulation of the ATM/ATR signaling pathway and the Skp2-p27 axis might be plausible mechanisms of action for the antiproliferative and antitumor activities of 1 in human colon cancer cells.

Correction: Kang, K.A.; et al., Myricetin Protects Cells against Oxidative Stress-Induced Apoptosis via Regulation of PI3K/Akt and MAPK Signaling Pathways. Int. J. Mol. Sci. 2010, 11, 4348-4360.

The authors want to change Figure 1 of the paper published in IJMS [1]. In Figure 1, 5-position of OH was at 6-position. Therefore, Figure 1 is revised as follows. The authors would like to apologize for any inconvenience caused to the readers by this change.[...].

Antitumor activity of jujuboside B and the underlying mechanism via induction of apoptosis and autophagy.

Jujuboside B (1) is one of the saponins isolated from the seeds of Zizyphus jujuba var. spinosa, which are used as a well-known traditional medicine for the treatment of insomnia and anxiety in East Asian countries. This is the first study to investigate the antitumor mechanism of 1 in vivo and in vitro. The results showed that 1 induced apoptosis and autophagy in AGS and HCT 116 human cancer cells and also effectively suppressed tumor growth in a nude mouse xenograft model bearing HCT 116 cells. The apoptosis-inducing effect of 1 was characterized by annexin V/propidium iodide staining, sub-G1 phase increase, and caspase-3 activation. Mechanistic studies showed that 1-induced apoptosis is associated with the extrinsic pathway through an increase in FasL and caspase-8 activation. Moreover, 1 activated p38/c-Jun N-terminal kinase (JNK), and the extrinsic pathway-mediated apoptosis was attenuated by both SB202190 (a p38 inhibitor) and SP600125 (a JNK inhibitor). The autophagy-inducing effect was indicated by the formation of cytoplasmic vacuoles and microtubule-associated protein 1 light chain-3 II (LC3-II) conversion. The autophagy inhibitor bafilomycin A1 (BaF) decreased 1-induced cell viability and increased pp38, pJNK, FasL, caspase-8 activation, and caspase-3 activation. Taken together, these results demonstrate that 1 induced protective autophagy to retard extrinsic pathway-mediated apoptosis.

Suppression of inflammatory responses by handelin, a guaianolide dimer from Chrysanthemum boreale, via downregulation of NF-κB signaling and pro-inflammatory cytokine production.

The anti-inflammatory activity of handelin (1), a guaianolide dimer from Chrysanthemum boreale flowers, was evaluated in vivo, and the effects on mediators nitric oxide (NO), prostaglandin E2 (PGE2), tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß) and the nuclear factor-κB (NF-κB) and ERK/JNK signaling pathways were investigated in vitro. Compound 1 inhibited lipopolysaccharide (LPS)-induced production of NO and PGE2 in cultured mouse macrophage RAW 264.7 cells. The suppression of NO and PGE2 production by 1 was correlated with the downregulation of mRNA and protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Compound 1 also suppressed the induction of pro-inflammatory cytokines TNF-α and IL-1ß in LPS-stimulated RAW 264.7 cells. To further clarify the transcriptional regulatory pathway in the expression of iNOS and COX-2 by 1, the role of NF-κB was determined in RAW 264.7 cells. Compound 1 inhibits the binding activity of NF-κB into the nuclear proteins. The transcriptional activity of NF-κB stimulated with LPS was also suppressed by 1, which coincided with the inhibition of IκB degradation. Compound 1 also suppressed the activation of mitogen-activated protein kinases, including ERK and JNK signaling. In addition, the LPS-stimulated upregulation of miRNA-155 expression was suppressed by 1. The oral administration of 1 inhibited acute inflammation in carrageenan-induced paw and 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced ear edema models. The serum level of IL-1ß was also inhibited by 1 in a carrageenan-induced paw edema model. These findings suggest that the suppression of NF-κB activation and pro-inflammatory cytokine production may be a plausible mechanism of action for the anti-inflammatory activity of handelin.

Anti-proliferative activity of hydnocarpin, a natural lignan, is associated with the suppression of Wnt/ß-catenin signaling pathway in colon cancer cells.

Based on the Wnt inhibitors as potential targets in the development of anticancer agents, natural compounds were evaluated for ß-catenin-mediated transcriptional activity. A natural lignan hydnocarpin isolated from Lonicera japonica was considered a potential inhibitor for Wnt/ß-catenin signalings. The anti-proliferative activity of hydnocarpin was also found to be associated with the suppression of Wnt/ß-catenin-mediated signaling pathway in human colon cancer cells. These data suggest that hydnocarpin might be a novel Wnt inhibitor and has a potential of signaling regulator in ß-catenin-mediated signaling pathways.

Zizyphus jujuba and its active component jujuboside B inhibit platelet aggregation.

The seeds of Zizyphus jujuba (SZJ), a famous oriental traditional medicine, have been reported to exhibit diverse activities in biological systems including the cardiovascular system. However, little information is available on its antiplatelet activity. This study was undertaken to investigate the antiplatelet effects of the ethanolic extract of SZJ (ESZJ) and of its principal components jujuboside A and B. In the in vitro platelet aggregation study, ESZJ exhibited significant and concentration-dependent inhibitory effects on collagen-, thrombin-, and AA-induced platelet aggregation. In addition, ESZJ-treated mice showed significantly the prolongation of bleeding times and the protection against thromboembolic attack. A comparison of the effects of jujuboside A and B on platelet aggregation revealed that only jujuboside B had potent inhibitory effects on collagen-, thrombin-, AA-, and ADP-induced aggregation. Jujuboside B also exhibited superior protection on thromboembolic model. Furthermore, jujuboside B had a significant inhibitory effect on collagen-induced thromboxane A2 production in rat platelets. This study describes the antiplatelet effects of ESZJ and of its active component jujuboside B, and its findings suggest that these agents be considered as components of preventive and therapeutic herbal drugs targeting cardiovascular diseases associated with platelet hyperaggregation.

Glycosides from the aerial parts of Patrinia villosa.

An investigation of the Korean medicinal plant Patrinia villosa (THUNB.) JUSS. (Valerianaceae) led to the isolation of two new flavonoid glycosides, patrivilosides 1 (1) and 2 (2), a new iridoid glycoside, patrinovalerosidate (3), and two new saponins, patrinovilosides A (4) and B (5), along with six known compounds including three flavonoid glycosides and three iridoid glycosides. The structures of the new compounds were elucidated based on analysis of their one dimensional (1D)- and 2D-NMR spectra along with their mass spectrometric data and the results of acid hydrolysis.

Separation of two cytotoxic saponins from the roots of Adenophora triphylla var. japonica by high-speed counter-current chromatography.

INTRODUCTION: The roots of Adenophorae species have been reported to exhibit anti-obese, anti-oxidant, anti-cancer, and anti-bacterial activities. However, there has been no single report regarding the preparative isolation and biological activities of the chemical components from Adenophora triphylla. OBJECTIVE: To develop an efficient method for the determination of the active fraction from the methanol extract from the roots of Adenophora triphylla and for the preparative isolation and purification of target compounds having cytotoxicity on carcinoma cells from the active fraction by high-speed counter-current chromatography (HSCCC). METHODS: The Plant (5 kg, dry weight) was extracted with methanol. Three hundred grams of the dried methanol extract (885 g) were fractionated by open-column chromatography with a stepwise gradient of water-methanol. Preparative isolation of bioactive components was performed by HSCCC with a two-phase solvent system composed of ethyl acetate-n-butanol-0.2% trifluoroacetic acid in water (5:5:10, v/v). The cytotoxicity of column fractions and isolated compounds was evaluated by 2-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide assay. RESULTS: The 70% MeOH column fraction showed inhibitory effects against three human carcinoma cells A549, AGS and HepG2. Two saponins were separated from 400 mg of the active fraction by HSCCC. After further purification with solid phase extraction column, 25 mg of peak fraction 1 and 20 mg of peak fraction 2 were obtained. Their structures were identified by ¹H-NMR, ¹³C-NMR, Fourier transform infrared, fast atom bombardment-MS and electrospray ionisation-MS/MS. They exhibited strong cytotoxic effects against three cancer cells. CONCLUSION: Two cytotoxic saponins were isolated for the first time from the roots of Adenophora triphylla by HSCCC.

Baicalein (5,6,7-trihydroxyflavone) reduces oxidative stress-induced DNA damage by upregulating the DNA repair system.

Oxidative stress caused by reactive oxygen species (ROS) induces DNA base modifications and DNA strand breaks. In this study, the protective effect of baicalein against H(2)O(2)-induced DNA damage was investigated in V79-4 Chinese hamster fibroblast cells. H(2)O(2) treatment increased the levels of intracellular ROS and DNA double-strand breaks (DSBs) and decreased the level of Ku70 protein and the phosphorylation (activation) of DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which are involved in the repair of DSBs by nonhomologous end joining. Baicalein effectively scavenged intracellular ROS induced by H(2)O(2), reduced DSBs, and rescued Ku70 protein level and phosphorylation of DNA-PKcs. In cellular response to DNA base damage, 8-oxoguanine DNA glycosylase 1 (OGG1) plays a vital role in the removal of 8-oxoguanine (8-OxoG), which is formed mainly by oxidative stress. Baicalein significantly decreased the levels of 8-OxoG induced by H(2)O(2), and this correlated with increases in OGG1 promoter activity and OGG1 mRNA and protein expression. The phosphorylated form of Akt kinase, which is a regulator of OGG1, was sharply decreased by H(2)O(2), but was prevented by baicalein. A specific Akt inhibitor abolished the cytoprotective effects of baicalein, suggesting that OGG1 induction by baicalein involves the Akt pathway. In conclusion, baicalein exerted protective effects against DNA damage induced by oxidative stress by activating DNA repair systems and scavenging ROS.

Anti-inflammatory mechanism of 15,16-epoxy-3α-hydroxylabda-8,13(16),14-trien-7-one via inhibition of LPS-induced multicellular signaling pathways.

Phytochemical investigation of Leonurus japonicus has led to the isolation of a labdane diterpene derivative, 15,16-epoxy-3α-hydroxylabda-8,13(16),14-trien-7-one (1), which was tested for its in vitro anti-inflammatory effects. The results demonstrated that 1 exhibits an inhibitory effect on LPS-stimulated RAW 264.7 macrophages. The anti-inflammatory action shown by 1 suppressed LPS-induced NF-κB activation, resulting in the down-regulation of iNOS and COX-2 protein expression, attributable to the inhibitory action of LPS-induced NO and PGE(2) production. Compound 1 inhibited LPS-induced phosphorylation and the degradation of inhibitory kappa B (IκBα) and decreased the nuclear translocation of p50 and p65. In addition, 1 exhibited an inhibitory effect on LPS-induced NF-κB-DNA and AP-1-DNA binding activity, using an electrophoretic mobility shift assay with NF-κB- and AP-1-specific (32)P-labeled probes. The LPS-induced mitogen-activated protein kinases (p-JNK, p-p38, and p-ERK) and p-Akt were inhibited after 30 and 10 min of LPS stimulation, respectively. In addition, TNF-α production was suppressed by 1.

Sesquiterpenes and other constituents from Dendranthema zawadskii var. latilobum.

Six new germacranolides, zawadskinolides A-F (1-6), and a new eudesmane glucoside, chrysantiloboside (7) were isolated from the aerial parts of Dendranthema zawadskii var. latilobum, along with thirteen known constituents. Their structures were elucidated by means of spectroscopic evidence. Bioassay showed that flavonoids such as apigenin (9), (-)-eriodictyol (10) and nepetin (12), as well as the sesquiterpene lactone, zawadskinolide F (6), inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophage cells with IC50 values of 66.15, 132.55, 35.44, and 91.32 µM, respectively.

A new polyoxygenated triterpene and two new aeginetic acid quinovosides from the roots of Rehmannia glutinosa.

A new minor polyoxygenated triterpene named glutinolic acid (1) and two new aeginetic acid quinovosides (2, 3) were isolated from the roots of Rehmannia glutinosa LIBOSCH. (Scrophulariaceae) cultivated in Gunwi-gun, Korea. The structures of these compounds were established as 3α,19α,20ß,24,30-pentahydroxyurs-12-en-28-oic acid (1, glutinolic acid), aeginetic acid 5-O-ß-D-quinovoside (2) and aeginetoyl ajugol 5″-O-ß-D-quinovoside (3) on the basis of chemical and spectroscopic evidence.

Hepatoprotective effect of pinoresinol on carbon tetrachloride-induced hepatic damage in mice.

Forsythiae Fructus is known to have diuretic, anti-bacterial, and anti-inflammatory activities. This study examined the hepatoprotective effects of pinoresinol, a lignan isolated from Forsythiae Fructus, against carbon tetrachloride (CCl(4))-induced liver injury. Mice were treated intraperitoneally with vehicle or pinoresinol (25, 50, 100, and 200 mg/kg) 30 min before and 2 h after CCl4 (20 microl/kg) injection. In the vehicle-treated CCl(4 )group, serum aminotransferase activities were significantly increased 24 h after CCl4 injection, and these increases were attenuated by pinoresinol at all doses. Hepatic glutathione contents were significantly decreased and lipid peroxidation was increased after CCl4 treatment. These changes were attenuated by 50 and 100 mg/kg of pinoresinol. The levels of protein and mRNA expression of inflammatory mediators, including tumor necrosis factor-alpha, inducible nitric oxide synthase, and cyclooxygenase-2, were significantly increased after CCl4 injection; and these increases were attenuated by pinoresinol. Nuclear translocation of nuclear factor-kappaB (NF-kappaB) and phosphorylation of c-Jun, one of the components of activating protein 1 (AP-1), were inhibited by pinoresinol. Our results suggest that pinoresinol ameliorates CCl4)-induced acute liver injury, and this protection is likely due to anti-oxidative activity and down-regulation of inflammatory mediators through inhibition of NF-kappaB and AP-1.

Growth inhibition and cell cycle arrest in the G0/G1 by schizandrin, a dibenzocyclooctadiene lignan isolated from Schisandra chinensis, on T47D human breast cancer cells.

Schizandrin is one of the main dibenzocyclooctadiene lignans present in the fruit of Schisandra chinensis (Schisandraceae). Biological activities including hepatoprotective, antiviral and neuroprotective effects of schizandrin and other dibenzocyclooctadiene lignans have been reported previously. However, the antiproliferative effect of schizandrin against human cancer cells has been poorly determined to date. This study examined the antiproliferative effect of schizandrin in human breast cancer cells. Schizandrin exhibited growth inhibitory activities in cultured human breast cancer cells, and the effect was the more profound in estrogen receptor (ER)-positive T47D cells than in ER-negative MDA-MB-231 cells. When treated with the compound in T47D cells, schizandrin induced the accumulation of a cell population in the G0/G1 phase, which was further demonstrated by the induction of CDK inhibitors p21 and p27 and the inhibition of the expression of cell cycle checkpoint proteins including cyclin D1, cyclin A, CDK2 and CDK4. These results suggest that schizandrin inhibits cell proliferation through the induction of cell cycle arrest with modulating cell cycle-related proteins in human breast cancer cells.

Myricetin protects cells against oxidative stress-induced apoptosis via regulation of PI3K/Akt and MAPK signaling pathways.

Recently, we demonstrated that myricetin exhibits cytoprotective effects against H(2)O(2)-induced cell damage via its antioxidant properties. In the present study, myricetin was found to inhibit H(2)O(2)-induced apoptosis in Chinese hamster lung fibroblast (V79-4) cells, as shown by decreased apoptotic bodies, nuclear fragmentation, sub-G(1) cell population, and disruption of mitochondrial membrane potential (Δψ(m)), which are increased in H(2)O(2)-treated cells. Western blot data showed that in H(2)O(2)-treated cells, myricetin increased the level of Bcl-2, which is an anti-apoptotic factor, and decreased the levels of Bax, active caspase-9 and -3, which are pro-apoptotic factors. And myricetin inhibited release of cytochrome c from mitochondria to cytosol in H(2)O(2)-treated cells. Myricetin-induced survival correlated with Akt activity, and the rescue of cells by myricetin treatment against H(2)O(2)-induced apoptosis was inhibited by the specific PI3K (phosphoinositol-3-kinase) inhibitor. Myricetin-mediated survival also inhibited the activation of p38 mitogen activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK), which are members of MAPK. Our studies suggest that myricetin prevents oxidative stress-induced apoptosis via regulation of PI3K/Akt and MAPK signaling pathways.

Platelet anti-aggregatory and blood anti-coagulant effects of compounds isolated from Paeonia lactiflora and Paeonia suffruticosa.

The roots of two Paeoniaceae family members have long been used as traditional medicines in Korea, China, and Japan. Dry roots of Paeonia lactiflora and dry root bark of P. suffruticosa are used under the traditional names of Paeoniae Radix and Moutan Cortex, respectively. Both Paeoniae Radix and Moutan Cortex have been used as remedies for cardiovascular diseases, for improving blood circulation, or for other uses. It was postulated that both plants may contain common active constituents that contribute to inhibiting blood coagulation and/or platelet aggregation. Eighteen compounds, which have been reported to be present in both plant medicines, were evaluated for their effects on platelet aggregation and blood coagulation. Paeonol (5), paeoniflorin (9), benzoylpaeoniflorin (11), and benzoyloxypaeoniflorin (12) were found to be the major common active constituents and they would collectively contribute to improving blood circulation through their inhibitory effects on both platelet aggregation and blood coagulation. In addition, methylgallate (4), (+)-catechin (7), paeoniflorigenone (8), galloylpaeoniflorin (13), and daucosterol (16) may also take part in improving blood circulation by inhibiting ether platelet aggregation and/or blood coagulation.

Polyphenolic Biflavonoids Inhibit Amyloid-Beta Fibrillation and Disaggregate Preformed Amyloid-Beta Fibrils.

Alzheimer's disease (AD) is a devastating neurodegenerative disease and a major cause of dementia in elderly individuals worldwide. Increased deposition of insoluble amyloid ß (Aß) fibrils in the brain is thought be a key neuropathological hallmark of AD. Many recent studies show that natural products such as polyphenolic flavonoids inhibit the formation of insoluble Aß fibrils and/or destabilize ß-sheet-rich Aß fibrils to form non-cytotoxic aggregates. In the present study, we explored the structure-activity relationship of naturally-occurring biflavonoids on Aß amyloidogenesis utilizing an in vitro thioflavin T assay with Aß1-42 peptide which is prone to aggregate more rapidly to fibrils than Aß1-40 peptide. Among the biflavonoids we tested, we found amentoflavone revealed the most potent effects on inhibiting Aß1-42 fibrillization (IC50: 0.26 µM), as well as on disassembling preformed Aß1-42 fibrils (EC50: 0.59 µM). Our structure-activity relationship study suggests that the hydroxyl groups of biflavonoid compounds play an essential role in their molecular interaction with the dynamic process of Aß1-42 fibrillization. Our atomic force microscopic imaging analysis demonstrates that amentoflavone directly disrupts the fibrillar structure of preformed Aß1-42 fibrils, resulting in conversion of those fibrils to amorphous Aß1-42 aggregates. These results indicate that amentoflavone affords the most potent anti-amyloidogenic effects on both inhibition of Aß1-42 fibrillization and disaggregation of preformed mature Aß1-42 fibrils.