Extracts of Musa basjoo induce growth inhibition and changes in the protein expression of cell cycle control molecules in human colorectal cancer cell lines.

Musa basjoo (MB) is a species of the banana plant belonging to the genus Musa that has been used as a folk medicine. However, evidence-based biological activities and the molecular mechanism of action of MB are unknown. Thus, the aim of the present study was to examine whether the crude dried leaf extracts of MB inhibit the growth of colorectal (HT29 and HCT116) and other types (HepG2, MCF-7 and PC-3) of human cancer cell lines. Crude extracts of MB inhibited the growth of cells with IC50 values of 136 µg/ml (acetone extract, HT29), 51 µg/ml (acetone extract, HCT116), 45 µg/ml (acetone extract, HepG2), 40 µg/ml (acetone extract, MCF-7), 29 µg/ml (acetone extract, PC-3), 175 µg/ml (methanol extract, HT29), 137 µg/ml (methanol extract, HCT116), 102 µg/ml (methanol extract, HepG2), 85 µg/ml (methanol extract, MCF-7), and 85 µg/ml (methanol extract, PC-3) in colony formation assays, and 126 µg/ml (acetone extract, HT29), 68 µg/ml (acetone extract, HCT116), 260 µg/ml (methanol extract, HT29), and 216 µg/ml (methanol extract, HCT116) in MTT assays. Thin layer chromatography analysis revealed the potential existence of aromatic compounds in the acetone extract of MB. Flow cytometric analysis indicated that the percentage of cells in G1 increased, and this was associated with a concomitant decrease of cells in the S and/or G2-M phases of the cell cycle. When colorectal cancer cells were treated with acetone extract of MB, there was a marked decrease in the levels of expression of the cyclin D1, cyclin E, cdk2 and cdk4 proteins and a marked increase in the levels of the expression of the p21CIP1, p27KIP1, and p53 proteins, but those of apoptosis-associated protein PARP did not change. There was a tendency for acetone extract of MB to inhibit xenograft tumor growth in mice. Collectively, the crude extracts of MB contain active components that exert growth inhibition of human cancer cells. This is the first systematic study of the anticancer activity of MB and may broaden insights into the possible clinical approach of specific herbal medicines.

Ikarisoside A inhibits acetylcholine-induced catecholamine secretion and synthesis by suppressing nicotinic acetylcholine receptor-ion channels in cultured bovine adrenal medullary cells.

Ikarisoside A is a natural flavonol glycoside derived from plants of the genus Epimedium, which have been used in Traditional Chinese Medicine as tonics, antirheumatics, and aphrodisiacs. Here, we report the effects of ikarisoside A and three other flavonol glycosides on catecholamine secretion and synthesis in cultured bovine adrenal medullary cells. We found that ikarisoside A (1-100 µM), but not icariin, epimedin C, or epimedoside A, concentration-dependently inhibited the secretion of catecholamines induced by acetylcholine, a physiological secretagogue and agonist of nicotinic acetylcholine receptors. Ikarisoside A had little effect on catecholamine secretion induced by veratridine and 56 mM K(+). Ikarisoside A (1-100 µM) also inhibited (22)Na(+) influx and (45)Ca(2+) influx induced by acetylcholine in a concentration-dependent manner similar to that of catecholamine secretion. In Xenopus oocytes expressing α3ß4 nicotinic acetylcholine receptors, ikarisoside A (0.1-100 µM) directly inhibited the current evoked by acetylcholine. It also suppressed (14)C-catecholamine synthesis and tyrosine hydroxylase activity induced by acetylcholine at 1-100 µM and 10-100 µM, respectively. The present findings suggest that ikarisoside A inhibits acetylcholine-induced catecholamine secretion and synthesis by suppression of nicotinic acetylcholine receptor-ion channels in bovine adrenal medullary cells.

Anti-Androgenic Activity of Icarisid II from Epimedium Herb in Prostate Cancer LNCaP Cells.

Anti-androgens are regarded as potential therapeutic agents for the treatment of prostate cancer. We determined that an epimedium herb (EH) extract exhibited anti-androgenic activity in a luciferase assay using androgen receptor-positive prostate cancer LNCaP cells. Nine EH-derived flavonoids were examined. The results identified icarisid II as a very potent anti-androgenic EH-derived flavonoid. A quantitative RT-PCR analysis confirmed that the flavonol suppressed the expression of the androgen-responsive KLK3 gene.

Anti-androgenic activity of hydroxyxanthones in prostate cancer LNCaP cells.

Anti-androgens are used to treat prostate cancer. Here, we report that hydroxyxanthones from a plant extract act as anti-androgens in androgen receptor (AR)-positive prostate cancer LNCaP cells. Anti-androgenic activity of the ethanol extract from Garcinia subelliptica was observed in a luciferase assay using LNCaP/MMTV cells with a stably integrated mouse mammary tumor virus (MMTV) promoter. HPLC-based activity profiling followed by a chemical library-based assay strategy enabled the rapid identification of several active principles bearing a xanthone core substituted with hydroxyl and isoprenyl groups. Among the active compounds, 2-(1,1-dimethyl-allyl)-1,4,5,6-tetrahydroxyxanthone (subelliptenone F) was identified as a potent inhibitor of AR transcriptional activity. The structure-activity relationship of some substituents on the xanthone core was also determined using the chemical library-based bioassay. A quantitative RT-PCR analysis revealed that treatment with the compound resulted in a significant reduction in AR-induced gene (KLK3) expression. Hydroxyxanthone may be a possible candidate for the development of a new anti-androgenic molecule.

Isolation of six isoprenylated biflavonoids from the leaves of Garcinia subelliptica.

The acetone-soluble parts of Garcinia subelliptica leaves were analyzed and six new biflavonoids were isolated, i.e., garciniaflavones A-F (1-6), as well as the five known biflavonoids amentoflavone (7), podocarpusflavone A (8), (+)-morelloflavone (9), (+)-morelloflavone-7"-O-ß-glucopyranoside (10), and (+)-4'''-O-methylmorelloflavone (11) and the three triterpenoids oleanan-3-one, ß-amyrin, and cycloartenol. The structures of the isolates were established based on spectroscopic analyses, including a detailed NMR spectroscopic investigation. The new biflavonoids are rare mono-isoprenylated derivatives that have a flavone-(3'-8")-flavone core (1-4: amentoflavone type) and a flavanone-(3-8")-flavone core (5, 6: morelloflavone type). The absolute configurations of the morelloflavone-type biflavonoids (5, 6) were confirmed by circular dichroism to be 2R,3S. The biflavonoids with an isoprenyloxy group (1) and a 2-hydroxy-3-methyl-3-butenyl group (2), and the morelloflavone-type biflavonoids with a C(5) unit are the first examples in nature. We found that 7, one of the major biflavonoids, strongly inhibited hypoxia-inducible factor-1 in human embryonic kidney 293 cells under hypoxic conditions.

Anti-degranulating activity in rat basophil leukemia RBL-2H3 cells of flavanone glycosides and their aglycones in citrus fruits.

The anti-degranulating activity of flavonoids present in Citrus fruits was comprehensively evaluated. Among these, hesperetin and naringenin, respectively aglycones of hesperidin and narirutin, showed significant activity. The targets of hesperetin and naringenin were found: hesperetin inhibited phosphorylation of Syk and Akt, while naringenin suppressed the expression of Lyn and inhibited the phosphorylation of Akt. These results suggest that hesperetin and naringenin inhibit degranulation by suppression of pathway signals and reduce the symptoms of allergy by inhibiting phosphorylation of Akt, which leads to the suppression of cytokines. In addition, hesperetin showed inhibitory activity against the degranulation induced by calcium ionophores, indicating that hesperetin exerts its inhibitory activity by stabilizing the membrane structure.

New glycosides of acetophenone derivatives and phenylpropanoids from Juniperus occidentalis.

New glycosides of seven acetophenone derivatives (1-7) and two phenylpropanoids (8, 9), named juniperosides III-XI, have been isolated from the MeOH extract of the leaves and stems of Juniperus occidentalis Hook. (Cupressaceae), together with eleven other known compounds. The structures of these compounds have been successfully elucidated using a variety of spectroscopic techniques.

Sesquiterpenes from the aerial part of Chloranthus japonicus and their cytotoxicities.

Four new sesquiterpenes, chlorajapolides F-I (1-4), along with nine known terpenoids (5-13) were isolated from the aerial part of Chloranthus japonicus. Their structures were elucidated on the basis of spectroscopic analysis, and a lindenane sesquiterpene, named 9-hydroxy-heterogorgiolide, previously isolated from the C. japonicus, was revised as its 8-epimer (1a). Moreover, methanol extract (ME), EtOAc fraction (EF), water fraction (WF), and all isolates (1a, 1-13) were evaluated for their cytotoxicities using two human cancer cell lines.

Quantification of polyphenols and pharmacological analysis of water and ethanol-based extracts of cultivated agarwood leaves.

Mangiferin (3) and genkwanin 5-O-ß-primeveroside (5) are the two major bioactive polyphenols with laxative property present in the extracts of agarwood (Aquilaria sinensis) leaves (AL). Here we developed an HPLC method to determine these bioactive components and four other major polyphenols in AL extracts and evaluated the pharmacological equivalence of organic and water extracts. Using mobile phase gradient conditions combined with UV detection at 330 nm, all six compounds were separated and we determined the relative extraction ratios of the six compounds present in A. sinensis extracts that were prepared under different conditions and compared the contents of the two laxative polyphenols present in the 60% ethanol extracts of A. sinensis and A. crassna. The polyphenols present in water extracts of 13 commercially cultivated A. crassna plants have also been analyzed. The laxative properties of 60% ethanol and four water extracts of A. crassna were evaluated by the frequency and weight of stools in loperamide-induced constipation model mice. The pharmacological equivalence of 60% ethanol extract and hot water (95°C) extract was identified in mice.

Annatto prevents retinal degeneration induced by endoplasmic reticulum stress in vitro and in vivo.

SCOPE: Annatto (Bixa orellana) seeds have been used as a colorant in butter and in a variety of other foods. In this study, we investigated the amelioration of retinal damage by an acetone extract of annatto (A-ext.), bixin (a main component of annatto), and four bixin derivatives (Bx-1, Bx-2, Bx-3, and Bx-4) that we have synthesized. METHODS AND RESULTS: We used cultured retinal ganglion cells (RGC-5) to examine in vitro effects of A-ext. on stress pathways, focusing on intracellular oxidation induced by reactive oxygen species, expression of endoplasmic reticulum (ER) stress-related proteins, caspase-3 activation, and cell membrane damage. In vivo retinal damage in mice following intravitreous injection of tunicamycin was evaluated by counting the cell numbers in the ganglion cell layer (GCL) and measuring the thickness of outer nuclear layer (ONL). A-ext., bixin, and Bx-1 treatment inhibited both tunicamycin- and H2O2-induced cell death. Bixin derivatives also inhibited tunicamycin-induced cell death. Treatment with A-ext., bixin, and Bx-1 reduced tunicamycin-induced caspase-3 activity and inhibited the inversion of phosphatidylserine, an early apoptotic event without antioxidant effect or reduction of ER stress itself. A-ext., bixin, and Bx-1 significantly inhibited the tunicamycin-induced loss of cells from the GCL, and these materials also suppressed the tunicamycin-induced thinning of ONL. CONCLUSION: A-ext., its main component bixin, and bixin derivatives may therefore be useful for preventive and therapeutic treatment of retinal-related diseases.

Alterations in cell cycle and induction of apoptotic cell death in breast cancer cells treated with α-mangostin extracted from mangosteen pericarp.

The development of molecularly targeted drugs has greatly advanced cancer therapy, despite these drugs being associated with some serious problems. Recently, increasing attention has been paid to the anticancer effects of natural products. α-Mangostin, a xanthone isolated from the pericarp of mangosteen fruit, has been shown to induce apoptosis in various cancer cell lines and to exhibit antitumor activity in a mouse mammary cancer model. In this study, we investigated the influence of α-mangostin on apoptosis and cell cycle in the human breast cancer cell line MDA-MB231 (carrying a p53 mutation, and HER2, ER, and PgR negative) in order to elucidate its anticancer mechanisms. In α-mangostin-treated cells, induction of mitochondria-mediated apoptosis was observed. On cell-cycle analysis, G1-phase arrest, increased p21(cip1) expression and decreases in cyclins, cdc(s), CDKs and PCNA were observed. In conclusion, α-mangostin may be useful as a therapeutic agent for breast cancer carrying a p53 mutation and having HER2- and hormone receptor-negative subtypes.

Anti-inflammatory sesquiterpenes and sesquiterpene dimers from Chloranthus fortunei.

A novel lindenane sesquiterpene with an unprecedented 18-membered triester ring, named chlorafortulide (1), along with one known lindenane sesquiterpene (2) and nine known lindenane sesquiterpene dimers (3-11), was isolated from the whole plant of Chloranthus fortunei. Their structures and relative configurations were elucidated on the basis of spectroscopic analysis. All the isolates were evaluated for their inhibitory effects on lipopolysaccharide-induced nitric oxide production in RAW264.7 cells. Henriol D (4), shizukaols E (8), G (9), M (10), and O (11) showed significant anti-inflammatory activities with IC(50) values of 1.90, 3.68, 1.95, 7.01, and 1.95 µM, respectively.

Terpenoids from Chloranthus serratus and their anti-inflammatory activities.

Seven new terpenoids, including two sesquiterpene dimers (1, 2), two norditerpenoids (3, 4), and three sesquiterpenes (5-7), along with six known sesquiterpene dimers and four known sesquiterpenes were isolated from the whole plant of Chloranthus serratus. Their structures and relative configurations were elucidated on the basis of spectroscopic data analysis. The absolute configuration of 1 was determined by the CD exciton chirality method. These isolates were evaluated for their inhibitory effects on lipopolysaccharide-induced nitric oxide production in RAW264.7 cells. Compound 2 and two known compounds, shizukaols B and D, showed significant anti-inflammatory activities, with IC(50) values of 0.22, 0.15, and 7.22 µM, respectively.

Occurrence of stilbene oligomers in Cyperus rhizomes.

Investigation of the chemical constituents of Rhizoma Cyperi (Cyperus rotundus Linneus)resulted in the isolation of novel enantiomeric andmeso-stilbene trimers [i.e., (+)- and (−)-(E)-cyperusphenol A (1, 2 respectively) and (E)-mesocyperusphenol A (3)], a trimer bearing a novelhexacyclic ring system [cyperusphenol B (5)], aswell as knownstilbenoids (cyperusphenols C (4)and D (6), scirpusins A (7) and B (8), and piceid (9)) and luteolin. HPLC was used for the opticalresolution of 1 and 2 as well as for the identification of cooccurrence of enantiomers of 7. Thestructures of the isolates were established by spectroscopic analyses, including a detailed NMRspectroscopic investigation. The isolates were evaluated in terms of their antiproliferative activityemploying the Jurkat cell line (human T-cell leukemia cells), while the IC50 potencies of aracemate of 1 and 2, 3, 5, and 6 were estimated as 27.4, 40.5, 26.4, and 26.3 µM, respectively. Thesuppression of cell growth by 6 was due to the induction of apoptosis,whichwas characterized bynuclear changes and PARP-1 cleavage determined bywestern blotting.We also evaluated the freeradical scavenging activity of the isolates.

α-Mangostin extracted from the pericarp of the mangosteen (Garcinia mangostana Linn) reduces tumor growth and lymph node metastasis in an immunocompetent xenograft model of metastatic mammary cancer carrying a p53 mutation.

BACKGROUND: The mangosteen fruit has a long history of medicinal use in Chinese and Ayurvedic medicine. Recently, the compound α-mangostin, which is isolated from the pericarp of the fruit, was shown to induce cell death in various types of cancer cells in in vitro studies. This led us to investigate the antitumor growth and antimetastatic activities of α-mangostin in an immunocompetent xenograft model of mouse metastatic mammary cancer having a p53 mutation that induces a metastatic spectrum similar to that seen in human breast cancers. METHODS: Mammary tumors, induced by inoculation of BALB/c mice syngeneic with metastatic BJMC3879luc2 cells, were subsequently treated with α-mangostin at 0, 10 and 20 mg/kg/day using mini-osmotic pumps and histopathologically examined. To investigate the mechanisms of antitumor ability by α-mangostin, in vitro studies were also conducted. RESULTS: Not only were in vivo survival rates significantly higher in the 20 mg/kg/day α-mangostin group versus controls, but both tumor volume and the multiplicity of lymph node metastases were significantly suppressed. Apoptotic levels were significantly increased in the mammary tumors of mice receiving 20 mg/kg/day and were associated with increased expression of active caspase-3 and -9. Other significant effects noted at this dose level were decreased microvessel density and lower numbers of dilated lymphatic vessels containing intraluminal tumor cells in mammary carcinoma tissues. In vitro, α-mangostin induced mitochondria-mediated apoptosis and G1-phase arrest and S-phase suppression in the cell cycle. Since activation by Akt phosphorylation plays a central role in a variety of oncogenic processes, including cell proliferation, anti-apoptotic cell death, angiogenesis and metastasis, we also investigated alterations in Akt phosphorylation induced by α-mangostin treatment both in vitro and in vivo. Quantitative analysis and immunohistochemistry showed that α-mangostin significantly decreased the levels of phospho-Akt-threonine 308 (Thr308), but not serine 473 (Ser473), in both mammary carcinoma cell cultures and mammary carcinoma tissues in vivo. CONCLUSIONS: Since lymph node involvement is the most important prognostic factor in breast cancer patients, the antimetastatic activity of α-mangostin as detected in mammary cancers carrying a p53 mutation in the present study may have specific clinical applications. In addition, α-mangostin may have chemopreventive benefits and/or prove useful as an adjuvant therapy, or as a complementary alternative medicine in the treatment of breast cancer.

Occurrence of C-glucoside of resveratrol oligomers in Hopea parviflora.

Investigation of the highly polar chemical constituents in the stem of Hopea parviflora (Dipterocarpaceae) resulted in the isolation of four new resveratrol derivatives, hopeasides A and B (1, 2) (resveratrol pentamers), C (3) (resveratrol trimer), and D (4) (resveratrol dimer) together with nine known resveratrol oligomers (5-13). The new structures have a common partial structure of the 1-hydroxy-1-(3,5-dihydroxy-2-C-glucopyranosylphenyl)-2-(4-hydroxyphenyl)ethane-2-yl group after oxidative condensation of (E)-resveratrol-10-C-ß-glucopyranoside (14). The structures were determined by spectroscopic analysis including 2D-NMR and computer-aided molecular modeling. The biogenetic relationship of the isolates and NMR characteristics caused by steric hindrance are also discussed in this paper.

Resveratrol oligomers from Vatica albiramis.

Five new stilbenoids, vatalbinosides A-E (1-5), and 13 known compounds (6-18) were isolated from the stem of Vatica albiramis. The effects of these new compounds on interleukin-1ß-induced production of matrix metalloproteinase-1 (MMP-1) in human dermal fibroblasts were examined. Three resveratrol tetramers, (-)-hopeaphenol (6), vaticanol C (13), and stenophyllol C (14), were identified as strong inhibitors of MMP-1 production.

Agarwood induced laxative effects via acetylcholine receptors on loperamide-induced constipation in mice.

Agarwood (Aquilaria sinensis, Aquilaria crasna) is well known as an incense in the oriental region such as Thailand, Taiwan, and Cambodia, and is used as a digestive in traditional medicine. We investigated the laxative effects and mechanism of agarwood leaves extracted with ethanol (EEA-1, Aquilaria sinensis; EEA-2, Aquilaria crasna). EEA-1, EEA-2, the main constituents of EEAs (mangiferin, and genkwanin-5-O-primeveroside), and senna increased the frequency and weight of stools in loperamide-induced constipation model mice. EEA-1 and EEA-2 did not induce diarrhea as a side effect, but senna induced severe diarrhea. EEA-1 and senna increased gastro-intestinal (GI) transit in the model mice. EEA-1, but not senna, also increased the intestinal tension of isolated jejunum and ileum in guinea pigs, and the tension increase was blocked by atropine, a muscarinic receptor antagonist, but not by other inhibitors (granicetron, pyrilamine, or bradykinin-antagonist peptide). Furthermore, the increase in frequency and weight of stools induced by EEA-1 were blocked by pre-administration of atropine in the model mice. These findings indicate that EEAs exerted a laxative effect via acetylcholine receptors in the mouse constipation model.

Allergy-preventive effects of the flowers of Impatiens textori.

The allergy-preventive activity of a 35% EtOH extract (IT) of flowers of Impatiens textori MIQ. was demonstrated in a continuing search for allergy-preventive substances from natural sources. The evaluation of its activity used an in vivo assay method for monitoring the blood flow decrease in the tail vein microcirculation of mice subjected to sensitization with hen-egg white lysozyme. Among the principal compounds in IT, apigenin (1), luteolin (3), and luteolin 7-glucoside (4) showed significant allergy-preventive effects.