PW06 suppresses cancer cell metastasis in human pancreatic carcinoma MIA PaCa-2 cells via the inhibitions of p-Akt/mTOR/NF-κB and MMP2/MMP9 signaling pathways in vitro.

PW06 [(E)-3-(9-ethyl-9H-carbazol-3-yl)-1-(2,5-dimethoxyphenyl) prop-2-en-1-one], a kind of the carbazole derivative containing chalcone moiety, induced cell apoptosis in human pancreatic carcinoma in vitro. There is no investigation to show that PW06 inhibits cancer cell metastasis in human pancreatic carcinoma in vitro. Herein, PW06 (0.1-0.8 µM) significantly exists in the antimetastatic activities of human pancreatic carcinoma MIA PaCa-2 cells in vitro. Wound healing assay shows PW06 at 0.2 µM suppressed cell mobility by 7.45 and 16.55% at 6 and 24 hours of treatments. PW06 at 0.1 and 0.2 µM reduced cell mobility by 14.72 and 21.8% for 48 hours of treatment. Transwell chamber assay indicated PW06 (0.1-0.2 µM) suppressed the cell migration (decreased 26.67-35.42%) and invasion (decreased 48.51-68.66%). Atomic force microscopy assay shows PW06 (0.2 µM) significantly changed the shape of cell morphology. The gelatin zymography assay indicates PW06 decreased MMP2's and MMP9's activities at 48 hours of treatment. Western blotting assay further confirms PW06 reduced levels of MMP2 and MMP9 and increased protein expressions of EGFR, SOS1, and Ras. PW06 also increased the p-JNK, p-ERK, and p-p38. PW06 increased the expression of PI3K, PTEN, Akt, GSK3α/ß, and E-cadherin. Nevertheless, results also show PW06 decreased p-Akt, mTOR, NF-κB, p-GSK3ß, ß-catenin, Snail, N-cadherin, and vimentin in MIA PaCa-2 cells. The confocal laser microscopy examination shows PW06 increased E-cadherin but decreased vimentin in MIA PaCa-2 cells. Together, our findings strongly suggest that PW06 inhibited the p-Akt/mTOR/NF-κB/MMPs pathways, increased E-cadherin, and decreased N-cadherin/vimentin, suppressing the migration and invasion in MIA PaCa-2 cells in vitro.

PW06 Triggered Fas-FADD to Induce Apoptotic Cell Death In Human Pancreatic Carcinoma MIA PaCa-2 Cells through the Activation of the Caspase-Mediated Pathway.

Pancreatic cancer has higher incidence and mortality rates worldwide. PW06 [(E)-3-(9-ethyl-9H-carbazol-3-yl)-1-(2,5-dimethoxyphenyl) prop-2-en-1-one] is a carbazole derivative containing chalcone moiety which was designed for inhibiting tumorigenesis in human pancreatic cancer. This study is aimed at investigating PW06-induced anticancer effects in human pancreatic cancer MIA PaCa-2 cells in vitro. The results showed PW06 potent antiproliferative/cytotoxic activities and induced cell morphological changes in a human pancreatic cancer cell line (MIA PaCa-2), and these effects are concentration-dependent (IC50 is 0.43 µM). Annexin V and DAPI staining assays indicated that PW06 induced apoptotic cell death and DNA condensation. Western blotting indicated that PW06 increased the proapoptotic proteins such as Bak and Bad but decreased the antiapoptotic protein such as Bcl-2 and Bcl-xL. Moreover, PW06 increased the active form of caspase-8, caspase-9, and caspase-3, PARP, releasing cytochrome c, AIF, and Endo G from mitochondria in MIA PaCa-2 cells. Confocal laser microscopy assay also confirmed that PW06 increased Bak and decreased Bcl-xL. Also, the cells were pretreated with inhibitors of caspase-3, caspase-8, and caspase-9 and then were treated with PW06, resulting in increased viable cell number compared to PW06 treated only. Furthermore, PW06 showed a potent binding ability with hydrophobic interactions in the core site of the Fas-Fas death domains (FADD). In conclusion, PW06 can potent binding ability to the Fas-FADD which led to antiproliferative, cytotoxic activities, and apoptosis induction accompanied by the caspase-dependent and mitochondria-dependent pathways in human pancreatic cancer MIA PaCa-2 cells.

Osthole Antagonizes Microglial Activation in an NRF2-Dependent Manner.

Microglia are neuroglia in the brain with an innate immune function and participate in the progress of neurodegenerative diseases. Osthole (OST) is a coumarin derivative extracted from Cnidium monnieri and bears a microglia-antagonizing ability. However, the underlying mechanism of the antagonism is not clear. The lipopolysaccharides-induced microglial BV2 cell line and amyloid-overexpressing fruit fly were used as models to study OST treatment. We found that OST treatment is sufficient to evoke NRF2 cascade under an LPS-induced inflammatory environment, and silencing NRF2 is sufficient to abolish the process. Moreover, we found that OST is sufficient to antagonize microglial activation in both LPS-induced BV2 cells and Aß-overexpressing fruit flies, and silencing NRF2 abolishes OST's antagonism. Furthermore, OST treatment rescued survival, climbing, and the learning ability of Aß-overexpressing fruit flies and relieved oxidative stress. In conclusion, we proved that OST antagonizes microglial activation induced by either LPS or Aß and that NRF2 is necessary for OST's antagonism.

Ergosta-7,9(11),22-trien-3ß-ol Attenuates Inflammatory Responses via Inhibiting MAPK/AP-1 Induced IL-6/JAK/STAT Pathways and Activating Nrf2/HO-1 Signaling in LPS-Stimulated Macrophage-like Cells.

Chronic inflammation induces autoimmune disorders and chronic diseases. Several natural products activate nuclear factor erythroid 2-related factor 2 (Nrf2) signaling, attenuating inflammatory responses. Ergosta-7,9(11),22-trien-3ß-ol (EK100) isolated from Cordyceps militaris showed anti-inflammatory and antioxidative activity, but those mechanisms are still unclear. This study is the first to investigate EK100 on antioxidant Nrf2 relative genes expression in LPS-stimulated macrophage-like cell lines. The results showed that EK100 reduced IL-6 (interleukin-6) and tumor necrosis factor-α production. EK100 also attenuated a mitogen-activated protein kinase/activator protein-1 (MAPK/AP-1) pathway and interleukin-6/Janus kinase/signal transducer and activator of transcription (IL-6/JAK/STAT) pathway in LPS-stimulated cells. Toll-like receptor 4 (TLR4) inhibitor CLI-095 and MAPK inhibitors can synergize the anti-inflammatory response of EK100 in LPS-stimulated cells. Moreover, EK100 activated Nrf2/HO-1 (heme oxygenase-1) signaling in LPS-stimulated murine macrophage-like RAW 264.7 cells, murine microglial BV2 cells, and human monocytic leukemia THP-1 cells. However, Nrf2 small interfering RNA (Nrf2 siRNA) reversed EK100-induced antioxidative proteins expressions. In conclusion, EK100 showed anti-inflammatory responses via activating the antioxidative Nrf2/HO-1 signaling and inhibiting TLR4 related MAPK/AP-1 induced IL-6/JAK/STAT pathways in the LPS-stimulated cells in vitro. The results suggest EK100 acts as a novel antioxidant with multiple therapeutic targets that can potentially be developed to treat chronic inflammation-related diseases.

Ergosta-7, 9 (11), 22-trien-3ß-ol Interferes with LPS Docking to LBP, CD14, and TLR4/MD-2 Co-Receptors to Attenuate the NF-κB Inflammatory Pathway In Vitro and Drosophila.

Ergosta-7, 9 (11), 22-trien-3ß-ol (EK100) was isolated from Cordyceps militaris, which has been used as a traditional anti-inflammatory medicine. EK100 has been reported to attenuate inflammatory diseases, but its anti-inflammatory mechanism is still unclear. We were the first to investigate the effect of EK100 on the Toll-like receptor 4 (TLR4)/nuclear factor of the κ light chain enhancer of B cells (NF-κB) signaling in the lipopolysaccharide (LPS)-stimulated RAW264.7 cells and the green fluorescent protein (GFP)-labeled NF-κB reporter gene of Drosophila. EK100 suppressed the release of the cytokine and attenuated the mRNA and protein expression of pro-inflammatory mediators. EK100 inhibited the inhibitor kappa B (IκB)/NF-κB signaling pathway. EK100 also inhibited phosphatidylinositol-3-kinase (PI3K)/Protein kinase B (Akt) signal transduction. Moreover, EK100 interfered with LPS docking to the LPS-binding protein (LBP), transferred to the cluster of differentiation 14 (CD14), and bonded to TLR4/myeloid differentiation-2 (MD-2) co-receptors. Compared with the TLR4 antagonist, resatorvid (CLI-095), and dexamethasone (Dexa), EK100 suppressed the TLR4/AKT signaling pathway. In addition, we also confirmed that EK100 attenuated the GFP-labeled NF-κB reporter gene expression in Drosophila. In summary, EK100 might alter LPS docking to LBP, CD14, and TLR4/MD-2 co-receptors, and then it suppresses the TLR4/NF-κB inflammatory pathway in LPS-stimulated RAW264.7 cells and Drosophila.

Imperatorin Interferes with LPS Binding to the TLR4 Co-Receptor and Activates the Nrf2 Antioxidative Pathway in RAW264.7 Murine Macrophage Cells.

Imperatorin (IMP) could downregulate several inflammatory transcription factor signaling pathways. Some studies have pointed out that IMP could interfere with toll-like receptor 4 (TLR4) signaling. This study evaluates how IMP interferes with the TLR4 co-receptors signaling through the protein-ligand docking model, Western blotting, immunofluorescence (IF), and atomic force microscopy (AFM) assays in lipopolysaccharide (LPS) stimulated macrophage-like RAW264.7 cells in vitro. The results of the protein-ligand docking demonstrate that IMP interferes with LPS binding to the LPS-binding protein (LBP), the cluster of differentiation 14 (CD14), and the toll-like receptor 4/myeloid differentiation factor 2 (TLR4/MD-2) co-receptors in LPS-stimulated RAW264.7 cells. Compared with TLR4 antagonist CLI-095 or dexamethasone, IMP could suppress the protein expressions of LBP, CD14, and TLR4/MD-2 in LPS-stimulated cells. Furthermore, the three-dimensional (3D) image assay of the AFM showed IMP could prevent the LPS-induced morphological change in RAW264.7 cells. Additionally, IMP could activate the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, and it increased the antioxidative protein expression of heme oxygenase-1 (HO-1), superoxidase dismutase (SOD), and catalase (CAT). Our results are the first to reveal that the anti-inflammatory effect of IMP interferes with LPS binding to TLR4 co-receptor signaling and activates the antioxidative Nrf2 signaling pathway.

Estimated left ventricular pressure-myocardial strain loop as an index of cardiac work predicts all-cause mortality in patients receiving regular hemodialysis.

BACKGROUND: A non-invasive method for left ventricular pressure-strain analysis has recently been introduced to provide information on cardiac work and detect subtler changes in cardiac function. This study aims to verify and construct a novel index that could accurately and independently predict the prognosis of patients with end-stage kidney disease (ESRD) receiving regular hemodialysis. METHODS: Patients with end-stage kidney disease (ESRD) receiving maintenance hemodialysis (4-h sessions, 3 times weekly for 3 months or more) and who underwent echocardiography between 2009 and 2014 in China Medical University Hospital, Taichung, Taiwan, were enrolled. Conventional (left ventricular ejection fraction, LVEF) and strain echocardiography parameters (global longitudinal strain, GLS; cardiac work index, CWI) in 102 eligible patients were analyzed and compared. CWI was calculated from estimated LV pressure-myocardial strain loop area. RESULTS: Results show that, while no significant differences were found between LVEF (0.57 ±â€¯0.12 vs. 0.59 ±â€¯0.09, P = 0.27) and GLS (-16.12 ±â€¯6.57% vs. -18.44 ±â€¯5.54%, P = 0.07), deceased patients had significantly lower CWI (1339 ±â€¯683.05 mmHg% vs. 1883.38 ±â€¯640.99 mmHg%, P = 0.0002) than surviving patients. The predictive values defined by area under the curve (AUC) of LVEF, GLS and CWI were 0.499, 0.619 and 0.724, respectively. CONCLUSION: In conclusion, CWI is an accurately independent predictor of all-cause mortality in ESRD patients receiving regular hemodialysis and may superior to the current predictors such as LVEF and GLS.

Anti-lymphangiogenic diterpenes from the bark of Calocedrus macrolepis var. formosana.

Eight new diterpenes, 6α,7ß-dihydroxyferruginol (1), 6α,7α-dihydroxyferruginol (2), 6α-hydroxyhinokiol (3), 4α-hydroxy-7-oxo-18-norabieta-8,11,13-trien-4α-ol (4a), 15,16-dehydrosugiol (5), 7-methoxy-6,7-secoabieta-8,11,13-triene-6,12-diol (6), 7α-acetoxyabieta-8,12-diene-11,14-dione (7), 7α-butyloxyethyloxyabieta-8,12-diene-11,14-dione (8), along with four known compounds, 6,7-dehydroferruginol (9), 12-hydroxy-6,7-secoabieta-8,11,13-triene-6,7-dial (10), 7α-11-dihydroxy-12-methoxy-8,11,13-abietatriene (11), and 11,14-dihydroxy-8,11,13-abietatrien-7-one (12) were successfully isolated from the bark of Calocedrus macrolepis var. formosana. The structures of all isolates were elucidated by physical data (appearance, UV, IR, optical rotation) and spectroscopic data (1D, 2D NMR, and HREIMS). Compounds 9, 10, 11, and 12 showed promising growth-inhibitory effect on human lymphatic endothelial cells (LECs). Among these compounds, compound 10 exerted the most potent anti-lymphangiogenesis property by suppressing cell growth and tube formation of LECs. In conclusion, the results revealed the anti-lymphangiogenic potentials of Formosan C. macrolepis var. formosana.

Pipoxolan suppresses the inflammatory factors of NF-κB, AP-1, and STATs, but activates the antioxidative factor Nrf2 in LPS-stimulated RAW 264.7 murine macrophage cells.

Although pipoxolan (PIPO) is a smooth muscle relaxant, its anti-inflammatory capability has not been studied. Therefore, we investigated the anti-inflammatory molecular mechanisms of PIPO in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. In this study, we used the MTT assay to evaluate the cytotoxicity, applied the enzyme-linked immunosorbent assay to determine the inflammatory cytokines, and performed Western blotting to assess protein expression. The results showed that PIPO significantly inhibited cytokine production, including nitric oxide, prostaglandin E2 , tumor necrosis factor-α, and interleukin-6. PIPO also suppressed the pro-inflammatory mediator expression with inducible nitric oxide synthase and cyclooxygenase-2. Moreover, PIPO prohibited the multiple inflammatory transcription factor pathways, including inhibitor kappa B/nuclear factor of the κ light chain enhancer of B cells (NF-κB), mitogen-activated protein kinase/activator protein-1 (AP-1), Janus kinase/signal transducer and activator of transcription (STAT), and toll-like receptor 4 (TLR4)/serine/threonine kinase (AKT). Besides, PIPO effectively activated the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 antioxidative pathway. Collectively, PIPO may attenuate the inflammatory effects via influencing the LPS/TLR4 receptor binding; suppress the expression of anti-inflammatory transcription factors NF-κB, AP-1, and STAT; and activating the antioxidative transcription factor Nrf2 in LPS-stimulated mouse RAW 264.7 cells.

Physalin A attenuates inflammation through down-regulating c-Jun NH2 kinase phosphorylation/Activator Protein 1 activation and up-regulating the antioxidant activity.

Physalin A (PA), a withanolide, was isolated from Physalis angulata L. In this study, it is shown that PA can inhibit the production of inflammatory cytokines such as PGE2, NO, IL-1ß, IL-6, and TNF-α in LPS-induced RAW 264.7 cells. Furthermore, the results indicated that PA suppressed the IκB/NF-κB and JNK/ AP-1 inflammatory signaling pathways and inhibited the levels of pro-inflammatory factors iNOS and COX-2 in LPS-stimulated RAW 264.7 cells. In the carrageenan-induced mouse hind paw edema study, PA was shown to inhibit the production of inflammatory mediators such as NO, MDA, and TNF-α production. Conversely, the antioxidant factor levels of SOD, CAT, and GPx were all increased by the treated PA. According to the data, we are suggesting that the anti-inflammatory effects of PA may be through the suppressions of the JNK/AP-1 and IκB/NF-κB signaling pathways and up-regulation of the anti-oxidative activity.

Aloe-Emodin Enhances Tamoxifen Cytotoxicity by Suppressing Ras/ERK and PI3K/mTOR in Breast Cancer Cells.

Aloe-emodin (AE) is derived from Aloe vera and rhubarb (Rheum palmatum) and exhibits anticancer activities via multiple regulatory mechanisms in various cancers. AE can also enhance the anticancer efficacy of cisplatin, doxorubicin, docetaxel, and 5-fluorouracil; however, its effects remain poorly characterized. MCF-7, MDA-MB-231, MDA-MB-468, BT-474, and HCC-1954 breast cancer cell lines were treated with the indicated conditions of AE, and cell viability assays were performed. The expression levels of signaling proteins were determined by western blot analysis, intracellular reactive oxygen species (ROS), cell cycle distributions, and rates of apoptosis as estimated by flow cytometry. In comparison with other cells, MCF-7 cells were more sensitive to AE treatment; AE enhanced the cytotoxicity of 9[Formula: see text][Formula: see text]g/ml tamoxifen by reducing EGFR, ER[Formula: see text], Ras, ERK, c-Myc, and mTOR protein expression and blocking PI3K and mTOR activation. Finally, although co-treatment of AE with tamoxifen increased intracellular ROS, there were no effects on cell cycle progression. Besides facilitating tamoxifen-induced cell death, AE also enhanced the antiproliferative activity of tamoxifen by blocking Ras/ERK and PI3K/mTOR pathways in breast cancer cells, thus demonstrating the chemosensitizing potential of AE.

Roles of Peroxisome Proliferator-Activated Receptor α in Bitter Melon Seed Oil-Corrected Lipid Disorders and Conversion of α-Eleostearic Acid into Rumenic Acid in C57BL/6J Mice.

We previously reported that bitter melon seed oil (BMSO) was an effective anti-steatosis and antiobesity agent. Since the major fatty acid α-eleostearic acid (α-ESA) in BMSO is a peroxisome proliferator-activated receptor α (PPARα) activator, the objective was to investigate the role of PPARα in BMSO-modulated lipid disorders and α-ESA metabolism. C57BL/6J wild (WD) and PPARα knockout (KO) mice were fed a high-fat diet containing BMSO (15% soybean oil + 15% BMSO, HB) or not (30% soybean oil, HS) for 5 weeks. The HB diet significantly reduced hepatic triglyceride concentrations and increased acyl-CoA oxidase activity in WD, but not in KO mice. However, regardless of genotype, body fat percentage was lowered along with upregulated protein levels of uncoupling protein 1 (UCP1) and tyrosine hydroxylase, as well as signaling pathway of cAMP-dependent protein kinase and AMP-activated protein kinase in the white adipose tissue of HB-treated groups compared to HS cohorts. In WD-HB and KO-HB groups, white adipose tissue had autophagy, apoptosis, inflammation, and browning characteristics. Without PPARα, in vivo reduction of α-ESA into rumenic acid was slightly but significantly lowered, along with remarkable reduction of hepatic retinol saturase (RetSat) expression. We concluded that BMSO-mediated anti-steatosis depended on PPARα, whereas the anti-adiposity effect was PPARα-independent. In addition, PPARα-dependent enzymes may participate in α-ESA conversion, but only have a minor role.

Lobeline improves acute lung injury via nuclear factor-κB-signaling pathway and oxidative stress.

Acute lung injury (ALI) is a severe, life-threatening medical condition whose pathogenesis is linked to neutrophil infiltration of the lung. Activation and recruitment of neutrophils to the lung is mostly attributed to the production of chemokines NO, IL-6, for instance. This study aims to investigate lobeline ability in reducing NO production, and nitric oxide synthase (iNOs) expression. Lobeline was tested by inhibiting phosphorylation of mitogen-activated protein kinases (MAPKs), NF-κB and IκBα in LPS-stimulated RAW 264.7 cells. When RAW 264.7 macrophages were given lobeline with LPS, a significant concentration-dependent inhibition of NO production was detected. In vivo tests, mice were either treated with normal saline, 10mg/kg dexmethasone or 5, 10, 20mg/kg lobeline intraperitoneally, and after an hour, the administration of 5mg/kg of LPS was given intratracheally. External performance, cytokines, MAPK pathways and antioxidative enzymes (AOEs) were also carried out to evaluate the effects of these drugs. This is the first investigation in which lobeline was found to effectively inhibit acute lung edema, which may provide a potential target for treating ALI. Lobeline may utilize MAPKs pathways as well as AOEs activity to attenuate LPS-induced nonspecific pulmonary inflammation.

Suppression of the TNF-alpha level is mediated by Gan-Lu-Yin (traditional Chinese medicine) in human oral cancer cells through the NF-kappa B, AKT, and ERK-dependent pathways.

Oral cancer is one of the major causes of deaths in the male population of Taiwan. Gan-Lu-Yin (GLY) is used for an adjuvant treatment of Traditional Chinese Medicine in clinical patients. In this study, we investigated the molecular mechanisms in oral cancer cell lines after exposure to GLY. The cytometric bead-based array (CBA) method was used for the examining and analyzing of tumor necrosis factor-alpha (TNF-α) secretion level. TNF-α mRNA expression was determined by real-time PCR analysis. Nuclear factor kappa B (NF-κB) activity and other relative proteins were determined by NF-κB promoter assay, Western blotting, electrophoretic mobility shift assay (EMSA), and immuno-staining analyses. GLY decreased the secretion of TNF-α from the oral cancer CAL 27 cells. Furthermore, 2000 µg/mL of GLY significantly suppressed TNF-α mRNA expression of CAL 27 cells in a time-dependent manner. GLY reduced the levels of proteins, including nuclear NF-κB (p65 and p50), p-IKK (ser176), p-IκB, p-AKT, p-ERK, and nuclear Egr-1 in a time and dose-dependent manner. GLY also suppressed the NF-κB activity and translocation in CAL 27 cells. We suggest that GLY might promote the cure of oral cancer through decreasing the level of TNF-α cytokine, and these actions were mediated partially through the NF-κB, AKT, and ERK-dependent pathways in vitro. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1196-1205, 2016.

FARE-CAFE: a database of functional and regulatory elements of cancer-associated fusion events.

Chromosomal translocation (CT) is of enormous clinical interest because this disorder is associated with various major solid tumors and leukemia. A tumor-specific fusion gene event may occur when a translocation joins two separate genes. Currently, various CT databases provide information about fusion genes and their genomic elements. However, no database of the roles of fusion genes, in terms of essential functional and regulatory elements in oncogenesis, is available. FARE-CAFE is a unique combination of CTs, fusion proteins, protein domains, domain-domain interactions, protein-protein interactions, transcription factors and microRNAs, with subsequent experimental information, which cannot be found in any other CT database. Genomic DNA information including, for example, manually collected exact locations of the first and second break points, sequences and karyotypes of fusion genes are included. FARE-CAFE will substantially facilitate the cancer biologist's mission of elucidating the pathogenesis of various types of cancer. This database will ultimately help to develop 'novel' therapeutic approaches. Database URL: http://ppi.bioinfo.asia.edu.tw/FARE-CAFE.

Transcription factor and microRNA-regulated network motifs for cancer and signal transduction networks.

BACKGROUND: Molecular networks are the basis of biological processes. Such networks can be decomposed into smaller modules, also known as network motifs. These motifs show interesting dynamical behaviors, in which co-operativity effects between the motif components play a critical role in human diseases. We have developed a motif-searching algorithm, which is able to identify common motif types from the cancer networks and signal transduction networks (STNs). Some of the network motifs are interconnected which can be merged together and form more complex structures, the so-called coupled motif structures (CMS). These structures exhibit mixed dynamical behavior, which may lead biological organisms to perform specific functions. RESULTS: In this study, we integrate transcription factors (TFs), microRNAs (miRNAs), miRNA targets and network motifs information to build the cancer-related TF-miRNA-motif networks (TMMN). This allows us to examine the role of network motifs in cancer formation at different levels of regulation, i.e. transcription initiation (TF → miRNA), gene-gene interaction (CMS), and post-transcriptional regulation (miRNA → target genes). Among the cancer networks and STNs we considered, it is found that there is a substantial amount of crosstalking through motif interconnections, in particular, the crosstalk between prostate cancer network and PI3K-Akt STN.To validate the role of network motifs in cancer formation, several examples are presented which demonstrated the effectiveness of the present approach. A web-based platform has been set up which can be accessed at: http://ppi.bioinfo.asia.edu.tw/pathway/. It is very likely that our results can supply very specific CMS missing information for certain cancer types, it is an indispensable tool for cancer biology research.

Apigenin-7-glycoside prevents LPS-induced acute lung injury via downregulation of oxidative enzyme expression and protein activation through inhibition of MAPK phosphorylation.

Apigenin-7-glycoside (AP7Glu) with multiple biological activities is a flavonoid that is currently prescribed to treat inflammatory diseases such as upper respiratory infections. Recently, several studies have shown that its anti-inflammatory activities have been strongly linked to the inhibition of secretion of pro-inflammatory proteins, such as inducible nitric oxide synthase (iNOs) and cyclooxygenase-2 (COX-2) induced through phosphorylation nuclear factor-κB (NF-κB) and mitogen-activated protein kinases (MAPK) pathways. Additionally, inflammation, which can decrease the activities of antioxidative enzymes (AOEs) is also observed in these studies. At the same time, flavonoids are reported to promote the activities of heme oxygenase-1 (HO-1) decreased by LPS. The purpose of this study was to assess these theories in a series of experiments on the suppressive effects of AP7Glu based on LPS-induced nitric oxide production in RAW264.7 macrophages in vitro and acute lung injury in mice in vivo. After six hours of lipopolysaccharide (LPS) stimulation, pulmonary pathological, myeloperoxidase (MPO) activity, total polymorphonuclear leukocytes (PMN) cells, cytokines in bronchoalveolar lavage fluid (BALF) and AOEs, are all affected and changed. Meanwhile, our data revealed that AP7Glu not only did significantly inhibit the LPS-enhanced inflammatory activity in lung, but also exhibited anti-inflammatory effect through the MAPK and inhibitor NF-κB (IκB) pathways.

Proteomic analysis of urethral protein expression in an estrogen receptor α-deficient murine model of stress urinary incontinence.

PURPOSE: The roles of estrogen receptor α (ERα) in stress urinary incontinence (SUI) remain elusive. This study was conducted to understand the molecular mechanism of ERα against SUI. METHODS: Wild-type (ERα(+/+)) and ACTB-cre ERα knockout (ERα(-/-)) female mice were generated. Urethral function and protein expression were measured. Leak point pressures (LPP) and maximum urethral closure pressure (MUCP) were assessed in mice under urethane anesthesia. After the measurements, the urethras were removed for proteomic analysis using the two-dimensional differential gel electrophoresis and liquid chromatography-mass spectrometry technology. Interaction between these ERα pathway proteins was further analyzed by using MetaCore. Lastly, Western blot and immunochemistry (IHC) were used to confirm the candidate protein expression levels and locations, respectively. RESULTS: Compared with the ERα(+/+) group, the LPP and MUCP values of the ERα(-/-) group were significantly decreased. Additionally, we identified 11 differentially expressed proteins in the urethra of ERα(-/-) female mice; five proteins were down-regulated and six were up-regulated. The majority of the ERα knockout-modified proteins were involved in muscle development, contraction, and regulation, as well as immune response (amphoterin signaling and phagocytosis), proteolysis, and cell adhesion (platelet aggregation and integrin-mediated cell-matrix adhesion). IHC and Western blot confirmed the down-regulation of tropomyosin and up-regulation of myosin in urethra. CONCLUSIONS: This is the first study to estimate protein expression changes in urethras from ERα(-/-) female mice. These changes could be related to the molecular mechanism of ERα in SUI.

Latex of Euphorbia antiquorum-induced S-phase arrest via active ATM kinase and MAPK pathways in human cervical cancer HeLa cells.

Latex of Euphorbia antiquorum (EA) has demonstrated great chemotherapeutic potential for cancer. However, the mechanisms of anti-proliferation of EA on cancer cell remain to be further investigated. The purpose of this study was to explore the influence of EA in human cervical cancer cells. Here, the cell cycle distribution by flow cytometry was examined and the protein expression by the western blotting methods was analyzed. From the cytometric results it was shown that EA-induced S-phase arrest in a concentration manner both in human cervical cancer HeLa and CaSki cells. According the western blot results it was illustrated that EA could downregulate early cyclin E1-Cdk2; and cyclin A-Cdc2 provides a significant additional quantity of S-phase promotion, that in turn promoted the expression of p21(waf1/cip1) and p27(kip1) which were the inhibitors in the complex of cyclin A and Cdc2 that led to cell cycle arrest. Moreover, EA promoted the activation of ataxia telangiectasia mutated (ATM) and check-point kinase-2 (Chk2); however, it negatively regulated the expression of Topoisomerases I and II, Cdc25A, and Cdc25C signaling. Caffeine, an ATM/ATR inhibitor significantly reversed EA downregulation in the levels of Cdc25A. Furthermore, JNK inhibitor SP600125 and p38 MAPK inhibitor SB203580 both could reverse the EA upregulation of the protein of Chk2 level, significantly. This study, therefore, revealed that EA could downregulate topoisomerase, and activate ATM kinase, which then induce parallel Chk 1/2 and MAPK signaling pathways to promote the degradation of Cdc25A to induced S-phase arrest in human cervical cancer HeLa cells.

Urethral dysfunction in female mice with estrogen receptor ß deficiency.

Estrogen has various regulatory functions in the growth, development, and differentiation of the female urogenital system. This study investigated the roles of ERß in stress urinary incontinence (SUI). Wild-type (ERß(+/+)) and knockout (ERß(-/-)) female mice were generated (aged 6-8 weeks, n = 6) and urethral function and protein expression were measured. Leak point pressures (LPP) and maximum urethral closure pressure (MUCP) were assessed in mice under urethane anesthesia. After the measurements, the urethras were removed for proteomic analysis using label-free quantitative proteomics by nano-liquid chromatography-mass spectrometry (LC-MS/MS) analysis. The interaction between these proteins was further analysed using MetaCore. Lastly, Western blot was used to confirm the candidate proteins. Compared with the ERß(+/+) group, the LPP and MUCP values of the ERß(-/-) group were significantly decreased. Additionally, we identified 85 differentially expressed proteins in the urethra of ERß(-/-) female mice; 57 proteins were up-regulated and 28 were down-regulated. The majority of the ERß knockout-modified proteins were involved in cell-matrix adhesion, metabolism, immune response, signal transduction, nuclear receptor translational regelation, and muscle contraction and development. Western blot confirmed the up-regulation of myosin and collagen in urethra. By contrast, elastin was down-regulated in the ERß(-/-) mice. This study is the first study to estimate protein expression changes in urethras from ERß(-/-) female mice. These changes could be related to the molecular mechanism of ERß in SUI.