Cell suspension culture extract of Eriobotrya japonica attenuates growth and induces apoptosis in prostate cancer cells via targeting SREBP-1/FASN-driven metabolism and AR.

BACKGROUND: Castration-resistant prostate cancer (CRPC) is one of the main causes of male cancer mortality. There is currently no effective treatment to cure this deadly prostate cancer (PCa) progression. However, recent research showed that activation of lipogenesis leads to CRPC progression. It provides a rationale to target the highly lipogenic activity as a novel and promising therapy against lethal CRPC. PURPOSES: The present study aims to evaluate the anticancer efficacy and the molecular mechanism of cell suspension culture extract from Eriobotrya japonica (EJCE) in PCa, including CRPC. METHODS: Cell growth, migration and invasion analyses were performed by MTT method, a wound healing assay and the transwell method, respectively. Apoptosis was assessed by a flow cytometry-based Annexin V-FITC/PI assay, caspase enzymatic activity and Western blot analyses. Lipogenesis was determined by a Fatty Acid Quantification Kit and an Oil Red O staining. The in vivo experiment was conducted by a xenograft mouse model. RESULTS: PCa cell growth, migration and invasion were significantly affected by EJCE. EJCE decreased expression of sterol regulatory element-binding protein-1 (SREBP-1) and fatty acid synthase (FASN) in PCa cells, two main factors for lipogenesis. By inhibiting SREBP-1/FASN, EJCE reduced the intracellular fatty acid levels and lipid droplet accumulation in PCa. Moreover, EJCE down-regulated the androgen receptor (AR) and prostate-specific antigen (PSA) in PCa cells. Significantly, EJCE exhibited the potential anticancer activity by suppressing the growth and leading to apoptosis of CRPC tumors in a xenograft mouse model. CONCLUSION: These results reveal a novel therapeutic molecular mechanism of EJCE in PCa. Blockade of SREBP-1/FASN-driven metabolism and AR by EJCE could be employed as a potent opportunity to cure malignant PCa.

Emerging Therapeutic Activity of Davallia formosana on Prostate Cancer Cells through Coordinated Blockade of Lipogenesis and Androgen Receptor Expression.

BACKGROUND: Prostate cancer (PCa) is the most prevalent malignancy diagnosed in men in Western countries. There is currently no effective therapy for advanced PCa aggressiveness, including castration-resistant progression. The aim of this study is to evaluate the potential efficacy and determine the molecular basis of Davallia formosana (DF) in PCa. Methods: LNCaP (androgen-sensitive) and C4-2 (androgen-insensitive/castration-resistant) PCa cells were utilized in this study. An MTT-based method, a wound healing assay, and the transwell method were performed to evaluate cell proliferation, migration, and invasion. Intracellular fatty acid levels and lipid droplet accumulation were analyzed to determine lipogenesis. Moreover, apoptotic assays and in vivo experiments were conducted. RESULTS: DF ethanol extract (DFE) suppressed proliferation, migration, and invasion in PCa cells. DFE attenuated lipogenesis through inhibition of the expression of sterol regulatory element-binding protein-1 (SREBP-1) and fatty acid synthase (FASN). Moreover, DFE decreased androgen receptor (AR) and prostate-specific antigen (PSA) expression in PCa cells. We further showed the potent therapeutic activity of DFE by repressing the growth and leading to apoptosis of subcutaneous C4-2 tumors in a xenograft mouse model. CONCLUSIONS: These data provide a new molecular basis of DFE in PCa cells, and co-targeting SREBP-1/FASN/lipogenesis and the AR axis by DFE could be employed as a novel and promising strategy for the treatment of PCa.

Anti-arthritic and anti-inflammatory effects of (-)-Epicatechin-3-O-ß-d-allopyranoside, a constituent of Davallia formosana.

BACKGROUND: (-)-Epicatechin-3-O-ß-d-allopyranoside (ECAP) is isolated from the popular Chinese herbal medicine Davallia formosana, which has been used to treat bone diseases including bone fracture, arthritis, and osteoporosis. PURPOSE: To investigate the antiarthritic and the anti-inflammatory effect of ECAP on a mouse model of collagen-induced arthritis (CIA) and in vitro. METHODS: Male DBA/1 J mice were immunized by administering an intradermal injection of 100 µg of type II collagen in Freund's complete adjuvant. The control groups (vehicle) and ECAP were administered orally at doses of 1 ml/kg (H2O), 50 and 100 mg/ml/kg once a day from Day 22 to Day 42 after primary immunization. Paw swelling, arthritis severity score, and histological changes were examined. Enzyme-linked immunosorbent assay was used to measure the levels of cytokines, including tumor necrosis factor alpha (TNF-α), interleukin (IL)-10, IL-17, IL-4, and interferon-γ (IFN-γ), in splenocytes. Furthermore, the anti-inflammatory activities of ECAP were investigated in vitro by measuring nitric oxide (NO) levels in lipopolysaccharide (LPS)-activated RAW264.7 macrophages. RESULTS: In the CIA model, the oral administration of ECAP ameliorated paw edema and reduced the arthritis severity score and disease incidence. Histopathological examination demonstrated that ECAP treatment effectively protected the bone and cartilage of knee joints from erosion, lesion formation, and deformation compared with the vehicle treatment. ECAP also reduced IL-1ß and MMP-9 expression in inflamed joints. Compared with the vehicle-treated mice with CIA, the reduced severity of the disease in ECAP-treated mice was associated with decreased levels of TNF-α and IL-17 and increased levels of IL-10 and IL-4 in the supernatants of splenocyte cultures. Flow cytometry analysis demonstrated that ECAP increased the population of CD4+CD25+ regulatory T cells, thereby inhibiting the B cell population. Anticollagen IgG1 and IgG2a levels decreased in the serum of ECAP-treated mice. ECAP suppressed LPS-induced NO production in RAW264.7 macrophages. CONCLUSION: The administration of ECAP effectively suppressed inflammation and inflammatory pain and adjuvant-induced arthritis, indicating its therapeutic potential in the treatment of rheumatoid arthritis.

Baicalin, Baicalein, and Lactobacillus Rhamnosus JB3 Alleviated Helicobacter pylori Infections in Vitro and in Vivo.

Helicobacter pylori infection is associated with chronic gastritis, peptic ulcers, and gastric cancer. The flavonoid compounds baicalin and baicalein found in many medicinal plants exhibit an anti-inflammatory effect. The administration of Lactobacillus strains reducing the risk of H. pylori infection is well accepted. In this study, the therapeutic effects against H. pylori infection of baicalin, baicalein, and L. rhamnosus JB3 (LR-JB3), isolated from a dairy product, were investigated. Compared to baicalin, baicalein exhibited stronger anti-H. pylori activity and cytotoxicity on human gastric cancer epithelial AGS cells. Baicalin and baicalein both suppressed the vacA gene expression of H. pylori and interfered with the adhesion and invasion ability of H. pylori to AGS cells, as well as decreased H. pylori-induced interleukin (IL)-8 expression. In the mice infection model, high dosages of baicalin and baicalein inhibited H. pylori growth in the mice stomachs. Serum IL-1ß levels and H. pylori-specific serum IgM and IgA levels in mice treated with baicalin and baicalein were decreased. Moreover, a synergistic therapeutic effect of baicalein and LR-JB3 on eradicating H. pylori infections was observed. Thus, administrating baicalin, baicalein, or LR-JB3 for an H. pylori infection could offer similar therapeutic effects to administering antibiotics while not disturbing the balance of gut microbiota. This study revealed the effects of baicalin, baicalein, and LR-JB3 on attenuating the virulence of H. pylori. The synergistic effect with baicalein and LR-JB3 provides the experimental rationale for testing the reliability, safety, and efficacy of this approach in higher animals and perhaps ultimately in humans to eradicate H. pylori infections. PRACTICAL APPLICATION: Baicalin and baicalein exert health promotion and avoidance of H. pylori infections by interfering with H. pylori growth and virulence. Lactobacillus rhamnosus JB3 was used to reduce the gastric inflammation caused by H. pylori infection.

Protective Effects of Tormentic Acid, a Major Component of Suspension Cultures of Eriobotrya japonica Cells, on Acetaminophen-Induced Hepatotoxicity in Mice.

An acetaminophen (APAP) overdose can cause hepatotoxicity and lead to fatal liver damage. The hepatoprotective effects of tormentic acid (TA) on acetaminophen (APAP)-induced liver damage were investigated in mice. TA was intraperitoneally (i.p.) administered for six days prior to APAP administration. Pretreatment with TA prevented the elevation of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), total bilirubin (T-Bil), total cholesterol (TC), triacylglycerol (TG), and liver lipid peroxide levels in APAP-treated mice and markedly reduced APAP-induced histological alterations in liver tissues. Additionally, TA attenuated the APAP-induced production of nitric oxide (NO), reactive oxygen species (ROS), tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1ß), and IL-6. Furthermore, the Western blot analysis showed that TA blocked the protein expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2), as well as the inhibition of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPKs) activation in APAP-injured liver tissues. TA also retained the superoxidase dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) in the liver. These results suggest that the hepatoprotective effects of TA may be related to its anti-inflammatory effect by decreasing thiobarbituric acid reactive substances (TBARS), iNOS, COX-2, TNF-α, IL-1ß, and IL-6, and inhibiting NF-κB and MAPK activation. Antioxidative properties were also observed, as shown by heme oxygenase-1 (HO-1) induction in the liver, and decreases in lipid peroxides and ROS. Therefore, TA may be a potential therapeutic candidate for the prevention of APAP-induced liver injury by inhibiting oxidative stress and inflammation.

(-)-Epicatechin 3-O-ß-D-allopyranoside prevent ovariectomy-induced bone loss in mice by suppressing RANKL-induced NF-κB and NFATc-1 signaling pathways.

BACKGROUND: Davallia formosana Hayata is a herb that has been used in Chinese medicine to treat bone diseases, including arthritis, bone fractures and osteoporosis. The rhizome of D. formosana H. has been found to be rich in (-)-Epicatechin 3-O-ß-D-allopyranoside (ECAP), which is considered to be the active component of the plant in terms of its antiosteoporotic effect. This study investigated the molecular mechanism of the antiosteoporotic property of ECAP isolated from the roots of D. formosana H. using both in vitro and in vivo models. METHODS: We studied the effects of ECAP on the signaling pathways of the receptor activator of nuclear factor-κB ligand (RANKL)-stimulated osteoclastogenesis and ovariectomy-induced osteoporosis. In the in vitro study, the inhibitory action of ECAP on RANKL-induced osteoclastogenesis and the expression of osteoclast-related marker genes were investigated, and in the in vivo study, the effects of ECAP on bone were evaluated using ovariectomized (OVX) mice orally-administered ECAP for 4 weeks. RESULTS: We demonstrated that ECAP dose-dependently inhibited RANKL- and nuclear factor of activated T-cells, and cytoplasmic 1 (NFATc-1)-induced osteoclastogenesis by RAW 264.7 cells, and reduced the extent of bone resorption. Furthermore, µCT images and TRAP staining showed that oral administration of ECAP to OVX mice prevented bone loss. ECAP administration also exerted recovery effects on serum C-terminal telopeptide of type I collagen and osteocalcin levels in OVX mice. In addition, we also found that MMP-9 expression was decreased in vivo and in vitro. CONCLUSIONS: Overall, our findings suggested that ECAP suppresses RANKL-induced osteoclastogenesis through NF-κB and NFATc-1 signaling pathways, and has the potential for use in osteoporosis treatment.

(-)-Epicatechin-3-O-ß-D-allopyranoside from Davallia formosana prevents diabetes and dyslipidemia in streptozotocin-induced diabetic mice.

The objective of this study was to evaluate the effects and molecular mechanism of (-)-epicatechin-3-O-ß-D-allopyranoside from Davallia formosana (BB) (also known as Gu-Sui-Bu) on type 1 diabetes mellitus and dyslipidemia in streptozotocin (STZ)-induced diabetic mice. This plant was demonstrated to display antioxidant activities and possess polyphenol contents. Diabetic mice were randomly divided into six groups and were given daily oral gavage doses of either BB (at three dosage levels), metformin (Metf) (at 0.3 g/kg body weight), fenofibrate (Feno) (at 0.25 g/kg body weight) or vehicle (distilled water) and a group of control (CON) mice were gavaged with vehicle over a period of 4 weeks. Treatment with BB led to reduced levels of blood glucose, HbA1C, triglycerides and leptin and to increased levels of insulin and adiponectin compared with the vehicle-treated STZ group. The diabetic islets showed retraction from their classic round-shaped as compared with the control islets. The BB-treated groups (at middle and high dosages) showed improvement in islets size and number of Langerhans islet cells. The membrane levels of skeletal muscular glucose transporter 4 (GLUT4) were significantly higher in BB-treated mice. This resulted in a net glucose lowering effect among BB-treated mice. Moreover, BB enhanced the expression of skeletal muscle phospho-AMPK in treated mice. BB-treated mice increased expression of fatty acid oxidation enzymes, including peroxisome proliferator-activated receptor α (PPARα) and mRNA levels of carnitine palmitoyl transferase Ia (CPT1a). These mice also expressed lower levels of lipogenic genes such as fatty acid synthase (FAS), as well as lower mRNA levels of sterol regulatory element binding protein 1c (SREBP1c) and liver adipocyte fatty acid binding protein 2 (aP2). This resulted in a reduction in plasma triglyceride levels. BB-treated mice also expressed lower levels of PPARγ and FAS protein. This led to reduced adipogenesis, fatty acid synthesis and lipid accumulation within adipose tissue, and consequently, to lower triglyceride levels in liver, blood, and adipose tissue. Moreover, BB treatment not only displayed the activation Akt in liver tissue and skeletal muscle, but also in C2C12 myotube to cause an increase in phosphorylation of Akt in the absence of insulin. These results demonstrated that BB act as an activator of AMPK and /or regulation of insulin pathway (Akt), and the antioxidant activity within the pancreas. Therefore, BB treatment ameliorated the diabetic and dyslipidemic state in STZ-induced diabetic mice.

Serralongamines B-D, three new Lycopodium alkaloids from Lycopodium serratum var. longipetiolatum, and their inhibitory effects on foam cell formation in macrophages.

Three new Lycopodium alkaloids, serralongamines B-D (1-3), have been isolated from the club moss Lycopodium serratum var. longipetiolatum, and the structures were elucidated on the basis of spectroscopic data and chemical transformation. 1 and 3 significantly exhibited the inhibitory activity against foam cell formation in human macrophages, one of characteristic features of early atherosclerotic lesions.

Pervanadate induces Mammalian Ste20 Kinase 3 (MST3) tyrosine phosphorylation but not activation.

The yeast Ste20 (sterile) protein kinase, which is a serine/threonine kinase, responds to the stimulation of the G proteincoupled receptor (GPCR) pheromone receptor. Ste20 protein kinase serves as the critical component that links signaling from the GPCR/G proteins to the mitogen-activated protein kinase (MAPK) cascade in yeast. The yeast Ste20p functions as a MAP kinase kinase kinase kinase (MAP4K) in the pheromone response. Ste20-like kinases are structurally conserved from yeast to mammals. The mechanism by which MAP4K links GPCR to the MAPK pathway is less clearly defined in vertebrates. In addition to MAP4K, the tyrosine kinase cascade bridges G proteins and the MAPK pathway in vertebrate cells. Mammalian Ste20 Kinase 3 (MST3) has been categorized into the Ste20 family and has been reported to function in the regulation of cell polarity and migration. However, whether MST3 tyrosine phosphorylation regulates diverse signaling pathways is unknown. In this study, the tyrosine phosphatase inhibitor pervanadate was found to induce MST3 tyrosine phosphorylation in intact cells, and the activity of tyrosine-phosphorylated MST3 was measured. This tyrosine-directed phosphorylation was independent of MST3 activity. Parameters including protein conformation, Triton concentration and ionic concentration influenced the sensitivity of MST3 activity. Taken together, our data suggests that the serine/threonine kinase MST3 undergoes tyrosinedirected phosphorylation. The tyrosine-phosphorylated MST3 may create a docking site for the structurally conserved SH2/SH3 (Src Homology 2 and 3) domains within the Src oncoprotein. The unusual tyrosinephosphorylated MST3 may recruit MST3 to various signaling components.

Kinsenoside inhibits the inflammatory mediator release in a type-II collagen induced arthritis mouse model by regulating the T cells responses.

BACKGROUND: Anoectochilus formosanus has been used as a Chinese folk medicine and is known as the "King of medicine" in Chinese society due to its versatile pharmacological effects such as anti-hypertension, anti-diabetes, anti-heart disease, anti-lung and liver diseases, anti-nephritis and anti-Rheumatoid arthritis. Kinsenoside is an essential and active compound of A. formosanus (Orchidaceae). However, the anti-arthritic activity of kinsenoside has still not been demonstrated. In the present study, we confirmed that the kinsenoside treatment rheumatoid arthritis induced by collagen-induced arthritis in mice. METHODS: Male DBA/1 J mice were immunized by intradermal injection of 100 µg of type II collagen in CFA. Kinsenoside was administered orally at a dose of 100 and 300 mg/kg once a day after 2nd booster injection. Paw swelling, arthritic score and histological change were measured. ELISA was used to measure cytokines including tumor necrosis factor alpha (TNF-α), interleukin-10 (IL-10), interleukin-17 (IL-17) and interferon-γ (IFN-γ) in the splenocyte according to the manufacturer's instructions. RESULTS: Compared with model group, kinsenoside significantly inhibited paw edema and decreased the arthritis score and disease incidence. Histopathological examination demonstrated that kinsenoside effectively protected bone and cartilage of knee joint from erosion, lesion and deformation versus those from the CIA group. Kinsenoside also decreased IL-1ß, TNF-α, and MMP-9 expression, and increased the expression of IL-10 in inflamed joints. The administration of kinsenoside significantly suppressed levels of TNF-α, IFN-γ, and IL-17, but increased concentrations of IL-10 in the supernatants of each of the splenocytes in CIA mice compared with that in the H2O-treated mice with CIA. Using flow cytometric analysis, we demonstrated that kinsenoside increases the population of CD4(+)CD25(+) regulatory T cells, thereby inhibiting the Th1 cell and B cell populations. Anticollagen IgG1 and IgG2a levels decreased in the serum of kinsenoside-treated mice. CONCLUSIONS: These results suggest that the administration of kinsenoside effectively suppressed inflammatory mediators' production and bone erosion in mice with collagen-induced arthritis showing the potential as an anti-arthritis agent.

(-)-Epicatechin-3-O-ß-D-allopyranoside from Davallia formosana, Prevents Diabetes and Hyperlipidemia by Regulation of Glucose Transporter 4 and AMP-Activated Protein Kinase Phosphorylation in High-Fat-Fed Mice.

The purpose of this experiment was to determine the antidiabetic and lipid-lowering effects of (-)-epicatechin-3-O-ß-D-allopyranoside (BB) from the roots and stems of Davallia formosana in mice. Animal treatment was induced by high-fat diet (HFD) or low-fat diet (control diet, CD). After eight weeks of HFD or CD exposure, the HFD mice were treating with BB or rosiglitazone (Rosi) or fenofibrate (Feno) or water through gavage for another four weeks. However, at 12 weeks, the HFD-fed group had enhanced blood levels of glucose, triglyceride (TG), and insulin. BB treatment significantly decreased blood glucose, TG, and insulin levels. Moreover, visceral fat weights were enhanced in HFD-fed mice, accompanied by increased blood leptin concentrations and decreased adiponectin levels, which were reversed by treatment with BB. Muscular membrane protein levels of glucose transporter 4 (GLUT4) were reduced in HFD-fed mice and significantly enhanced upon administration of BB, Rosi, and Feno. Moreover, BB treatment markedly increased hepatic and skeletal muscular expression levels of phosphorylation of AMP-activated (adenosine monophosphate) protein kinase (phospho-AMPK). BB also decreased hepatic mRNA levels of phosphenolpyruvate carboxykinase (PEPCK), which are associated with a decrease in hepatic glucose production. BB-exerted hypotriglyceridemic activity may be partly associated with increased mRNA levels of peroxisome proliferator activated receptor α (PPARα), and with reduced hepatic glycerol-3-phosphate acyltransferase (GPAT) mRNA levels in the liver, which decreased triacylglycerol synthesis. Nevertheless, we demonstrated BB was a useful approach for the management of type 2 diabetes and dyslipidemia in this animal model.

A New Bithiophene from the Root of Echinops grijsii.

A new bithiophene, 5-(4-hydroxy-3-methoxy-1-butyny)-2,2'-bithiophene (1), and sixteen known thiophenes: 2-(3,4-dihydroxybut-1-ynyl)-5-(penta-1,3-diynyl)thiophene (2), α-terthienyl (3), 5-(3,4-dihydroxybut-1-ynyl)-2,2'-bithiophene (4), 5-acetyl-2,2'-bithiophene (5), 5-formyl-2,2'-bithiophene (6), methyl 2,2'-bithiophene-5-carboxylate (7), 5-(but-3-en-1-ynyl)-2,2'-bithiophene (8), 5-(4-isovaleroyloxybut-1-ynyl)-2,2'-bithiophene (9), cardopatine (10), isocardopatine (11), 5-(3-hydroxy-4-isovaleroyloxybut-1-ynyl)-2,2'-bithiophene (12), 5-(3-hydroxymethyl-3-isovaleroyloxyprop-1-ynyl)-2,2'-bithiophene (13), 5-(4-hydroxy-1-butynyl)-2,2'-bithiophene (14), 5-(4-acetoxy-1-butynl)-2,2'-bithiophene (15), 2,2'-bithiophene-5-carboxylic acid (16) and 2-(4-hydroxybut-1-ynyl)-5-(penta-1,3-diynyl)thiophene (17) were isolated from the roots of Echinops grjisii Hance. Among them, compounds 6, 7 and 16 were isolated from a natural source for the first time. Compounds 2, 4 and 14 exhibited significant anti-inflammatory activity against nitrite of LPS-stimulated production in the RAW 264.7 cell line.

8-Alkylcoumarins from the fruits of Cnidium monnieri protect against hydrogen peroxide induced oxidative stress damage.

Three new 8-alkylcoumarins, 7-O-methylphellodenol-B (1), 7-methoxy-8-(3-methyl- 2,3-epoxy-1-oxobutyl)chromen-2-one (2), and 3'-O-methylvaginol (3), together with seven known compounds (4-10) were isolated from the fruits of Cnidium monnieri. Their structures were determined by detailed analysis of spectroscopic data and comparison with the data of known analogues. All the isolates were evaluated the cytoprotective activity by MTS cell proliferation assay and the results showed that all the three new 8-alkylcoumarins exhibited cytoprotective effect on Neuro-2a neuroblastoma cells injured by hydrogen peroxide.

The prebiotic effect of Anoectochilus formosanus and its consequences on bone health.

The present study evaluated the prebiotic effect of a standardised aqueous extract of Anoectochilus formosanus (SAEAF) and its effects on osteoporosis in ovariectomised (OVX) rats. The OVX rats were randomly divided into five groups and orally treated with water, SAEAF (200 and 400 mg/kg daily) and inulin (400 mg/kg daily) for 12 weeks. The sham group was orally treated with water. The SAEAF treatment enhanced the number of faecal bifidobacteria in OVX rats. The results of a Ca-balance experiment showed that SAEAF increased apparent Ca absorption and retention. The OVX rats were killed after SAEAF treatment lasting 12 weeks. The SAEAF decreased the caecal pH values and increased the caecal wall weight, caecal mucosa calbindin-D9k mRNA expression, free-Ca concentration and levels of SCFA in the caecum. The mineral content, density and biomechanical strength of bones were lower in OVX rats than the sham group, but these bone losses were prevented by SAEAF administration. Microtomography scanning showed that the SAEAF-treated rats had higher trabecular bone volume than the OVX rats. These results suggest that SAEAF prevented bone loss associated with ovarian hormone deficiency in the rats.

Antitumor effects of the novel quinazolinone MJ-33: inhibition of metastasis through the MAPK, AKT, NF-κB and AP-1 signaling pathways in DU145 human prostate cancer cells.

Quinazolinone compounds have been shown to have antitumor activity in many human cancer cell lines. In the present study, we investigated the anti-metastatic activity of MJ-33 (2-(3-ethoxyphenyl)-6-pyrrolidinylquinazolinone), a novel quinazolinone derivate, and the signaling pathway of MJ-33 in human prostate cells. MJ-33 exhibited a growth inhibitory effect on DU145, LNCaP and PC-3 cells by MTT assay. DU145 cells showed greater sensitivity to the growth inhibition of MJ-33 than that of LNCaP and PC-3 cells. MJ-33 also had an inhibitory effect on the invasion, migration and adhesion of DU145 cells using Boyden chamber transwell assays, wound-healing and adhesion assay. In addition, MJ-33 inhibited cell metastasis through the reduction of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) and urokinase-type plasminogen activator (u-PA) enzyme activities and protein levels by gelatin zymography assay and western blot analysis, respectively. MJ-33 reduced the protein levels of p-JNK, p-p38, p-ERK, p-AKT and nuclear NF-κB (p65), c-fos and c-Jun protein levels by western blotting. Using electrophoretic mobility-shift assay (EMSA), we demonstrated that MJ-33 blocked the activation of transcription factor AP-1 (activator protein-1) and NF-κB, which led to the inhibition of MMP-2 and MMP-9 expression. Collectively, our data showed that MJ-33 decreased protein levels of MAPKs (mitogen-activated protein kinases), AKT, AP-1 and NF-κB, resulting in the inhibition of matrix metalloproteinases. Downregulation of MMP-2 and MMP-9 reduces the invasion, migration and adhesion activities of DU145 cells. MJ-33 may be a promising agent against prostate cancer metastasis.

Antiosteoporotic activity of Davallia formosana.

ETHNOPHARMACOLOGICAL RELEVANCE: [corrected] In Taiwanese folk medicine, Davallia formosana is used to treat bone diseases, including osteoporosis. AIM: This study evaluated the anti-osteoporotic effect of ethanolic extract derived from Davallia formosana (DFE). MATERIALS AND METHODS: In this in vitro study, we investigated the inhibitory action of DFE on RANKL-stimulated osteoclastogenesis. The in vivo effects of DFE on bone metabolism were evaluated using ovariectomized (OVX) rats orally administered DFE (200, 500 mg/kg), alendronate (2.5 mg/kg, three times a week) or its vehicle for 12 weeks. RESULTS: This in vitro study demonstrated that DFE inhibited osteoclast differentiation, and also isolated the active component, (-)-epicatechin 3-O-ß-D-allopyranoside (ECAP). DFE did not affect the body or vaginal weight in OVX rats. The bone mineral density and bone calcium content in OVX rats were lower in the control group showing that DFE was able to prevent significant bone loss. In addition, the three point bending test and the microcomputer tomography scanning showed that DFE treatment enhanced bone strength and inhibited the deterioration of trabecular microarchitecture. In the biochemical assay, DFE decreased urinary deoxypyridinoline and calcium concentrations, but did not inhibit serum alkaline phosphatase activities, indicating that it ameliorated bone loss via inhibition of bone reabsorption. CONCLUSIONS: These results suggest that DFE may represent a useful remedy for the treatment of bone reabsorption diseases such as osteoporosis. In addition, ECAP could be used as a marker compound to control the quality of DFE.

Synthesis and antiproliferative evaluation of 3,5-disubstituted 1,2,4-triazoles containing flurophenyl and trifluoromethanephenyl moieties.

An efficient 1,3-dipolar cycloaddition method was performed for the synthesis of a series of monofluoro- and trifluoromethane-3,5-disubstituted 1,2,4-triazoles. This efficient cycloaddition method was to react hydrazonoyl hydrochlorides with a series of aldehydes in the presence of NEt(3) as catalytic basic agent to provide the corresponding product in 28-94%. Their growth inhibitory results against cancer cells indicated that some of the fluorine- and trifluoromethane-containing compounds could effectively inhibit the growth of NCI-H226 and T-cell leukemia (Jurkat) cells. Among the compounds, trifluoromethane-containing 1,2,4-triazoles possessed the five-membered ring groups on the C-5 position of the triazolic ring, including cyclopentyl, 3-furyl, 3-thienyl, and 2-pyrrolyl, possessed the significant inhibitory activity for NCI-H226 cancer cells.

Specific microtubule-depolymerizing agents augment efficacy of dendritic cell-based cancer vaccines.

BACKGROUND: Damage-associated molecular patterns (DAMPs) are associated with immunogenic cell death and have the ability to enhance maturation and antigen presentation of dendritic cells (DCs). Specific microtubule-depolymerizing agents (MDAs) such as colchicine have been shown to confer anti-cancer activity and also trigger activation of DCs. METHODS: In this study, we evaluated the ability of three MDAs (colchicine and two 2-phenyl-4-quinolone analogues) to induce immunogenic cell death in test tumor cells, activate DCs, and augment T-cell proliferation activity. These MDAs were further evaluated for use as an adjuvant in a tumor cell lysate-pulsed DC vaccine. RESULTS: The three test phytochemicals considerably increased the expression of DAMPs including HSP70, HSP90 and HMGB1, but had no effect on expression of calreticulin (CRT). DC vaccines pulsed with MDA-treated tumor cell lysates had a significant effect on tumor growth, showed cytotoxic T-lymphocyte activity against tumors, and increased the survival rate of test mice. In vivo antibody depletion experiments suggested that CD8+ and NK cells, but not CD4+ cells, were the main effector cells responsible for the observed anti-tumor activity. In addition, culture of DCs with GM-CSF and IL-4 during the pulsing and stimulation period significantly increased the production of IL-12 and decreased production of IL-10. MDAs also induced phenotypic maturation of DCs and augmented CD4+ and CD8+ T-cell proliferation when co-cultured with DCs. CONCLUSIONS: Specific MDAs including the clinical drug, colchicine, can induce immunogenic cell death in tumor cells, and DCs pulsed with MDA-treated tumor cell lysates (TCLs) can generate potent anti-tumor immunity in mice. This approach may warrant future clinical evaluation as a cancer vaccine.

Anti-inflammatory activities of tormentic acid from suspension cells of Eriobotrya Japonicaex vivo and in vivo.

Anti-inflammatory effects of tormentic acid (TA) were investigated ex vivo and in vivo. TA decreased the paw edema at the 4th and 5thhour after λ-carrageenin (Carr) administration, and increased the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the liver tissue. TA also significantly attenuated the thiobarbituric acid reactive substances (TBARS) level in the edematous paw at the 5thhour after Carr injection. TA decreased the nitric oxide (NO) levels on the serum level and diminished the serum tumour necrosis factor (TNF-α) at the 5thhour after Carr injection. Western blotting revealed that the TA decreased Carr-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expressions. As per results, the anti-inflammatory mechanisms of TA might be correlated to the decrease in the level of TBARS, iNOS, and COX-2 in the edema paw via increasing the activities of CAT, SOD, and GPx in the liver.

A standardized aqueous extract of Anoectochilus formosanus ameliorated thioacetamide-induced liver fibrosis in mice: the role of Kupffer cells.

Anoectochilus formosanus is used in traditional folk medicine as an hepatoprotective agent. The purpose of this study was to investigate the effects of a standardized aqueous extract of A. formosanus (SAEAF) on thioacetamide (TAA)-induced liver fibrosis. An in vitro study showed that the inhibitive effect of kinsenoside, a major component of SAEAF, on tumor necrosis factor alpha (TNF-alpha) secretion from Kupffer cells might be derived at least partly from downregulation of LPS-receptor Toll-like receptor 4 (TLR4) signaling. Hepatic fibrosis was produced by TAA (200 mg/kg, i.p.) 3 times per week for 12 weeks. Mice in the three TAA groups were treated daily with distilled water and SAEAF (1.0, 0.2 g/kg) via gastrogavage throughout the experimental period. The mice that received the SAEAF treatment had significantly reduced plasma alanine aminotransferase activity, relative liver weights, and hepatic hydroxyproline contents. A histological examination also confirmed that SAEAF reduced the degree of fibrosis caused by TAA treatment. RT-PCR analysis showed that SAEAF treatment reduced mRNA expression of collagen (alpha1)(I), lipopolysaccharide-binding protein, CD14, TLR4, and TNF receptor 1. An immunohistochemical examination also indicated that SAEAF reduced the number of CD68-positive cells (macrophages). In conclusion, oral administration of SAEAF significantly reduced TAA-induced hepatic fibrosis in mice, probably through inhibition of hepatic Kupffer cell activation.