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Sample records for blocks wnt processing

  1. Epitope mapping by a Wnt-blocking antibody: evidence of the Wnt binding domain in heparan sulfate.

    PubMed

    Gao, Wei; Xu, Yongmei; Liu, Jian; Ho, Mitchell

    2016-01-01

    Heparan sulfate (HS) is a polysaccharide known to modulate many important biological processes, including Wnt signaling. However, the biochemical interaction between HS and Wnt molecules is not well characterized largely due to the lack of suitable methods. To determine the Wnt binding domain in HS, we used a Wnt signaling-inhibitory antibody (HS20) and a panel of synthetic HS oligosaccharides with distinct lengths and sulfation modifications. We found that the binding of HS20 to heparan sulfate required sulfation at both the C2 position (2-O-sulfation) and C6 position (6-O-sulfation). The oligosaccharides with the greatest competitive effect for HS20 binding were between six and eight saccharide residues in length. Additionally, a four residue-long oligosaccharide could also be recognized by HS20 if an additional 3-O-sulfation modification was present. Furthermore, similar oligosaccharides with 2-O, 6-O and 3-O-sulfations showed inhibition for Wnt activation. These results have revealed that HS20 and Wnt recognize a HS structure containing IdoA2S and GlcNS6S, and that the 3-O-sulfation in GlcNS6S3S significantly enhances the binding of both HS20 and Wnt. This study provides the evidence for identifying the Wnt binding domain in HS and suggests a therapeutic approach to target the interaction of Wnt and HS in cancer and other diseases. PMID:27185050

  2. Epitope mapping by a Wnt-blocking antibody: evidence of the Wnt binding domain in heparan sulfate

    PubMed Central

    Gao, Wei; Xu, Yongmei; Liu, Jian; Ho, Mitchell

    2016-01-01

    Heparan sulfate (HS) is a polysaccharide known to modulate many important biological processes, including Wnt signaling. However, the biochemical interaction between HS and Wnt molecules is not well characterized largely due to the lack of suitable methods. To determine the Wnt binding domain in HS, we used a Wnt signaling-inhibitory antibody (HS20) and a panel of synthetic HS oligosaccharides with distinct lengths and sulfation modifications. We found that the binding of HS20 to heparan sulfate required sulfation at both the C2 position (2-O-sulfation) and C6 position (6-O-sulfation). The oligosaccharides with the greatest competitive effect for HS20 binding were between six and eight saccharide residues in length. Additionally, a four residue-long oligosaccharide could also be recognized by HS20 if an additional 3-O-sulfation modification was present. Furthermore, similar oligosaccharides with 2-O, 6-O and 3-O-sulfations showed inhibition for Wnt activation. These results have revealed that HS20 and Wnt recognize a HS structure containing IdoA2S and GlcNS6S, and that the 3-O-sulfation in GlcNS6S3S significantly enhances the binding of both HS20 and Wnt. This study provides the evidence for identifying the Wnt binding domain in HS and suggests a therapeutic approach to target the interaction of Wnt and HS in cancer and other diseases. PMID:27185050

  3. RING finger protein PLR-1 blocks Wnt signaling by altering trafficking of Wnt Receptors

    NASA Astrophysics Data System (ADS)

    Robinson, Ryan E.

    Secreted Wnt proteins control a wide range of essential developmental processes, including axon guidance and establishment of anteroposterior neuronal polarity. We identified a transmembrane RING finger protein, PLR-1, that governs the response to Wnts by reducing the cell surface levels of Wnt receptors Frizzled, CAM-1 and LIN-18 in Caenorhabditis elegans. Frizzled, CAM-1 and LIN-18 are normally enriched at the plasma membrane where they are capable of detecting and responding to extracellular Wnts. However, when PLR-1 is expressed Frizzled, CAM-1 and LIN-18 are no longer detected at the cell surface and instead colocalize with PLR-1 in endosomes and Golgi. PLR-1 is related to a broad family of transmembrane proteins that contain a lumenal protease associated domain and a cytosolic RING finger. The RING finger is a hallmark of one type of E3 ubiquitin ligase and monoubiquitination is commonly used to regulate protein trafficking. Protease associated domains are largely thought to mediate interactions between proteins. To identify the domains responsible for PLR-1 regulation of Frizzled from the cell surface we utilized a series of fluorescently tagged fusion proteins and protein truncations containing various domains from PLR-1 and Frizzled. Our data suggests that PLR-1 and Frizzled interact and form a complex via their respective extracellular/lumenal domains, and that ubiqiuitination of Frizzled by PLR-1 targets the Frizzled/PLR-1 complex to the endosome.

  4. Axitinib blocks Wnt/β-catenin signaling and directs asymmetric cell division in cancer.

    PubMed

    Qu, Yi; Gharbi, Naouel; Yuan, Xing; Olsen, Jan Roger; Blicher, Pernille; Dalhus, Bjørn; Brokstad, Karl A; Lin, Biaoyang; Øyan, Anne Margrete; Zhang, Weidong; Kalland, Karl-Henning; Ke, Xisong

    2016-08-16

    Oncogenic mutations of the Wnt (wingless)/β-catenin pathway are frequently observed in major cancer types. Thus far, however, no therapeutic agent targeting Wnt/β-catenin signaling is available for clinical use. Here we demonstrate that axitinib, a clinically approved drug, strikingly blocks Wnt/β-catenin signaling in cancer cells, zebrafish, and Apc(min/+) mice. Notably, axitinib dramatically induces Wnt asymmetry and nonrandom DNA segregation in cancer cells by promoting nuclear β-catenin degradation independent of the GSK3β (glycogen synthase kinase3β)/APC (adenomatous polyposis coli) complex. Using a DARTS (drug affinity-responsive target stability) assay coupled to 2D-DIGE (2D difference in gel electrophoresis) and mass spectrometry, we have identified the E3 ubiquitin ligase SHPRH (SNF2, histone-linker, PHD and RING finger domain-containing helicase) as the direct target of axitinib in blocking Wnt/β-catenin signaling. Treatment with axitinib stabilizes SHPRH and thereby increases the ubiquitination and degradation of β-catenin. Our findings suggest a previously unreported mechanism of nuclear β-catenin regulation and indicate that axitinib, a clinically approved drug, would provide therapeutic benefits for cancer patients with aberrant nuclear β-catenin activation. PMID:27482107

  5. Mesd Is a Universal Inhibitor of Wnt Co-receptor LRP5/6 and Blocks Wnt/β-catenin Signaling in Cancer Cells†

    PubMed Central

    Lu, Wenyan; Liu, Chia-Chen; Thottassery, Jaideep V.; Bu, Guojun; Li, Yonghe

    2010-01-01

    Mesd is a specialized chaperone for the low-density lipoprotein receptor-related protein-5 (LRP5) and LRP6. In our previous studies, we found that Mesd binds to mature LRP6 on the cell surface and blocks the binding of Wnt antagonist Dickkopf-1(Dkk1) to LRP6. Herein, we demonstrated that Mesd also binds to LRP5 with a high affinity, and is a universal inhibitor of LRP5/6 ligands. Mesd not only blocks Wnt antagonists Dkk1 and Sclerostin binding to LRP5/6, but also inhibits Wnt3A and Rspondin1-induced Wnt/β-catenin signaling in LRP5/6 expressing cells. We also found that Mesd, Dkk1 and Sclerostin compete with one another for binding to LRP5 and LRP6 at the cell surface. More importantly, we demonstrated that Mesd is able to suppress LRP6 phosphorylation and Wnt/β-catenin signaling in prostate cancer PC-3 cells, and inhibits PC-3 cell proliferation. Our results indicate that recombinant Mesd protein is a useful tool for studying Wnt/β-catenin signaling on the cell surface, and has a potential therapeutic role in Wnt-dependent cancers. PMID:20446724

  6. Inhibition of tankyrases induces Axin stabilization and blocks Wnt signalling in breast cancer cells.

    PubMed

    Bao, Renyue; Christova, Tania; Song, Siyuan; Angers, Stephane; Yan, Xiaojun; Attisano, Liliana

    2012-01-01

    Constitutive Wnt signalling is characterized by excessive levels of β-catenin protein and is a frequent occurrence in cancer. APC and Axin are key components of the β-catenin destruction complex that acts to promote β-catenin degradation. The levels of Axin are in turn controlled by tankyrases, members of the PARP-family of poly-ADP-ribosylation enzymes. In colorectal cancer cells, which typically harbor APC mutations, inhibition of tankyrase activity promotes Axin stabilization and attenuates Wnt signalling. Here, we examined the effect of inhibiting tankyrases in breast cancer cells with normal APC. We show that application of the small molecule tankyrase inhibitor, XAV939 or siRNA-mediated abrogation of tankyrase expression increases Axin1 and Axin2 protein levels and attenuates Wnt-induced transcriptional responses in several breast cancer lines. In MDA-MB-231 cells, inhibiton of tankyrase activity also attenuate Wnt3a induced cell migration. Moreover, in both MDA-MB-231 and colorectal cancer cells, XAV939 inhibits cell growth under conditions of serum-deprivation. However, the presence of serum prevents this growth inhibitory effect, although inhibition of Wnt-induced transcriptional and migratory responses was maintained. These results indicate that stabilization of Axin by inhibition of tankyrases alone, may not be an effective means to block tumor cell growth and that combinatorial therapeutic approaches should be considered. PMID:23144924

  7. Stearoyl CoA desaturase is required to produce active, lipid-modified Wnt proteins.

    PubMed

    Rios-Esteves, Jessica; Resh, Marilyn D

    2013-09-26

    Wnt proteins contain palmitoleic acid, an unusual lipid modification. Production of an active Wnt signal requires the acyltransferase Porcupine and depends on the attachment of palmitoleic acid to Wnt. The source of this monounsaturated fatty acid has not been identified, and it is not known how Porcupine recognizes its substrate and whether desaturation occurs before or after fatty acid transfer to Wnt. Here, we show that stearoyl desaturase (SCD) generates a monounsaturated fatty acid substrate that is then transferred by Porcupine to Wnt. Treatment of cells with SCD inhibitors blocked incorporation of palmitate analogs into Wnt3a and Wnt5a and reduced Wnt secretion as well as autocrine and paracrine Wnt signaling. The SCD inhibitor effects were rescued by exogenous addition of monounsaturated fatty acids. We propose that SCD is a key molecular player responsible for Wnt biogenesis and processing and that SCD inhibition provides an alternative mechanism for blocking Wnt pathway activation. PMID:24055053

  8. Stearoyl CoA desaturase is required to produce active, lipid-modified Wnt proteins

    PubMed Central

    Rios-Esteves, Jessica; Resh, Marilyn D.

    2013-01-01

    Summary Wnt proteins contain an unusual lipid modification, palmitoleic acid. Production of an active Wnt signal requires the acyltransferase Porcupine and depends on attachment of palmitoleic acid to Wnt. The source of this monounsaturated fatty acid has not been identified, and it is not known how Porcupine recognizes its substrate and whether desaturation occurs before or after fatty acid transfer to Wnt. Here we show that stearoyl desaturase (SCD) generates a monounsaturated fatty acid substrate which is then transferred by Porcupine to Wnt. Treatment of cells with SCD inhibitors blocked incorporation of palmitate analogs into Wnt3a and Wnt5a, and reduced Wnt secretion as well as autocrine and paracrine Wnt signaling. The SCD inhibitor effects were rescued by exogenous addition of monounsaturated fatty acids. We propose that SCD is a key molecular player responsible for Wnt biogenesis and processing and that SCD inhibition provides an alternative mechanism for blocking Wnt pathway activation. PMID:24055053

  9. PTH1-34 blocks radiation-induced osteoblast apoptosis by enhancing DNA repair through canonical Wnt pathway.

    PubMed

    Chandra, Abhishek; Lin, Tiao; Zhu, Ji; Tong, Wei; Huo, Yanying; Jia, Haoruo; Zhang, Yejia; Liu, X Sherry; Cengel, Keith; Xia, Bing; Qin, Ling

    2015-01-01

    Focal radiotherapy for cancer patients has detrimental effects on bones within the radiation field and the primary clinical signs of bone damage include the loss of functional osteoblasts. We reported previously that daily injection of parathyroid hormone (PTH, 1-34) alleviates radiation-induced osteopenia in a preclinical radiotherapy model by improving osteoblast survival. To elucidate the molecular mechanisms, we irradiated osteoblastic UMR 106-01 cells and calvarial organ culture and demonstrated an anti-apoptosis effect of PTH1-34 on these cultures. Inhibitor assay indicated that PTH exerts its radioprotective action mainly through protein kinase A/β-catenin pathway. γ-H2AX foci staining and comet assay revealed that PTH efficiently promotes the repair of DNA double strand breaks (DSBs) in irradiated osteoblasts via activating the β-catenin pathway. Interestingly, Wnt3a alone also blocked cell death and accelerated DNA repair in primary osteoprogenitors, osteoblastic and osteocytic cells after radiation through the canonical signaling. Further investigations revealed that both Wnt3a and PTH increase the amount of Ku70, a core protein for initiating the assembly of DSB repair machinery, in osteoblasts after radiation. Moreover, down-regulation of Ku70 by siRNA abrogated the prosurvival effect of PTH and Wnt3a on irradiated osteoblasts. In summary, our results identify a novel role of PTH and canonical Wnt signaling in regulating DSB repair machinery and apoptosis in osteoblasts and shed light on using PTH1-34 or Wnt agonist as possible therapy for radiation-induced osteoporosis. PMID:25336648

  10. Wnt5a-mediated non-canonical Wnt signalling regulates human endothelial cell proliferation and migration

    SciTech Connect

    Cheng Chingwen Yeh Juching; Fan Taiping; Smith, Stephen K.; Charnock-Jones, D. Stephen

    2008-01-11

    Cell to cell interaction is one of the key processes effecting angiogenesis and endothelial cell function. Wnt signalling is mediated through cell-cell interaction and is involved in many developmental processes and cellular functions. In this study, we investigated the possible function of Wnt5a and the non-canonical Wnt pathway in human endothelial cells. We found that Wnt5a-mediated non-canonical Wnt signalling regulated endothelial cell proliferation. Blocking this pathway using antibody, siRNA or a down-stream inhibitor led to suppression of endothelial cell proliferation, migration, and monolayer wound closure. We also found that the mRNA level of Wnt5a is up-regulated when endothelial cells are treated with a cocktail of inflammatory cytokines. Our findings suggest non-canonical Wnt signalling plays a role in regulating endothelial cell growth and possibly in angiogenesis.

  11. Norcantharidin blocks Wnt/β-catenin signaling via promoter demethylation of WIF-1 in glioma.

    PubMed

    Xie, Dajiang; Xie, Jixi; Wan, Yingfeng; Ma, Li; Qi, Xuchen; Wang, Kun; Yang, Shuxu

    2016-04-01

    Glioma is one of the most common primary intracranial tumors, and the prognosis is poor even though much treatment management is employed. Wnt/β-catenin signaling has been reported to be associated with glioma. Norcantharidin (NCTD) is the demethylated analog of cantharidin isolated from blister beetles, and it is reported to possess anticancer activity but less nephrotoxicity than cantharidin. Accordingly, we aimed to investigate NCTD as an anti-neoplastic drug that inhibits the Wnt/β‑catenin pathway via promoter demethylation of Wnt inhibitory factor-1 (WIF-1) in glioma growth in vitro. In the present study, we report that NCTD inhibited cell proliferation, induced apoptosis and cell cycle arrest, and suppressed cell migration and invasion in vitro. Moreover, we observed that the expression levels of WIF-1 mRNA and protein in the NCTD-treated cells were increased significantly compared with these levels in the negative control (NC) cells. Promoter demethylation was observed in the NCTD‑treated cells. In contrast, aberrant methylation was observed in the NC cells. Additionally, more investigation revealed that NCTD suppressed activity of Wnt/β-catenin signaling and transcription of β-catenin/TCF-4. Furthermore, the expression of apoptosis-related proteins Bcl-2 and cleaved caspase-3 indicated significant cell apoptosis. We provide initial evidence that NCTD reactivates WIF-1 from a methylation state, and downregulates canonical Wnt/β-catenin signaling. Our findings revealed that NCTD is effective for glioma in vitro and may be a new therapeutic option in vivo. PMID:26781164

  12. Wnt pathway activation by ADP-ribosylation

    PubMed Central

    Yang, Eungi; Tacchelly-Benites, Ofelia; Wang, Zhenghan; Randall, Michael P.; Tian, Ai; Benchabane, Hassina; Freemantle, Sarah; Pikielny, Claudio; Tolwinski, Nicholas S.; Lee, Ethan; Ahmed, Yashi

    2016-01-01

    Wnt/β-catenin signalling directs fundamental processes during metazoan development and can be aberrantly activated in cancer. Wnt stimulation induces the recruitment of the scaffold protein Axin from an inhibitory destruction complex to a stimulatory signalosome. Here we analyse the early effects of Wnt on Axin and find that the ADP-ribose polymerase Tankyrase (Tnks)—known to target Axin for proteolysis—regulates Axin's rapid transition following Wnt stimulation. We demonstrate that the pool of ADP-ribosylated Axin, which is degraded under basal conditions, increases immediately following Wnt stimulation in both Drosophila and human cells. ADP-ribosylation of Axin enhances its interaction with the Wnt co-receptor LRP6, an essential step in signalosome assembly. We suggest that in addition to controlling Axin levels, Tnks-dependent ADP-ribosylation promotes the reprogramming of Axin following Wnt stimulation; and propose that Tnks inhibition blocks Wnt signalling not only by increasing destruction complex activity, but also by impeding signalosome assembly. PMID:27138857

  13. Wnt pathway activation by ADP-ribosylation.

    PubMed

    Yang, Eungi; Tacchelly-Benites, Ofelia; Wang, Zhenghan; Randall, Michael P; Tian, Ai; Benchabane, Hassina; Freemantle, Sarah; Pikielny, Claudio; Tolwinski, Nicholas S; Lee, Ethan; Ahmed, Yashi

    2016-01-01

    Wnt/β-catenin signalling directs fundamental processes during metazoan development and can be aberrantly activated in cancer. Wnt stimulation induces the recruitment of the scaffold protein Axin from an inhibitory destruction complex to a stimulatory signalosome. Here we analyse the early effects of Wnt on Axin and find that the ADP-ribose polymerase Tankyrase (Tnks)--known to target Axin for proteolysis-regulates Axin's rapid transition following Wnt stimulation. We demonstrate that the pool of ADP-ribosylated Axin, which is degraded under basal conditions, increases immediately following Wnt stimulation in both Drosophila and human cells. ADP-ribosylation of Axin enhances its interaction with the Wnt co-receptor LRP6, an essential step in signalosome assembly. We suggest that in addition to controlling Axin levels, Tnks-dependent ADP-ribosylation promotes the reprogramming of Axin following Wnt stimulation; and propose that Tnks inhibition blocks Wnt signalling not only by increasing destruction complex activity, but also by impeding signalosome assembly. PMID:27138857

  14. Blocking the Wnt/β-Catenin Pathway by Lentivirus-Mediated Short Hairpin RNA Targeting β-Catenin Gene Suppresses Silica-Induced Lung Fibrosis in Mice.

    PubMed

    Wang, Xin; Dai, Wujing; Wang, Yanrang; Gu, Qing; Yang, Deyi; Zhang, Ming

    2015-09-01

    Silicosis is a form of occupational lung disease caused by inhalation of crystalline silica dust. While the pathogenesis of silicosis is not clearly understood, the Wnt/β-catenin signaling pathway is thought to play a major role in lung fibrosis. To explore the role of Wnt/β-catenin pathway in silicosis, we blocked Wnt/β-catenin pathway both in silica-treated MLE-12 cells (a mouse pulmonary epithelial cell line) and in a mouse silicosis model by using a lentiviral vector expressing a short hairpin RNA silencing β-catenin (Lv-shβ-catenin). In vitro, Lv-shβ-catenin significantly decreased the expression of β-catenin, MMP2 and MMP9, and secretion of TGF-β1. In vivo, intratracheal treatment with Lv-shβ-catenin significantly reduced expression of β-catenin in the lung and levels of TGF-β1 in bronchoalveolar lavage fluid, and notably attenuated pulmonary fibrosis as evidenced by hydroxyproline content and collagen I\\III synthesis in silica-administered mice. These results indicate that blockade of the Wnt/β-catenin pathway can prevent the development of silica-induced lung fibrosis. Thus Wnt/β-catenin pathway may be a target in prevention and treatment of silicosis. PMID:26340635

  15. Blocking the Wnt/β-Catenin Pathway by Lentivirus-Mediated Short Hairpin RNA Targeting β-Catenin Gene Suppresses Silica-Induced Lung Fibrosis in Mice

    PubMed Central

    Wang, Xin; Dai, Wujing; Wang, Yanrang; Gu, Qing; Yang, Deyi; Zhang, Ming

    2015-01-01

    Silicosis is a form of occupational lung disease caused by inhalation of crystalline silica dust. While the pathogenesis of silicosis is not clearly understood, the Wnt/β-catenin signaling pathway is thought to play a major role in lung fibrosis. To explore the role of Wnt/β-catenin pathway in silicosis, we blocked Wnt/β-catenin pathway both in silica-treated MLE-12 cells (a mouse pulmonary epithelial cell line) and in a mouse silicosis model by using a lentiviral vector expressing a short hairpin RNA silencing β-catenin (Lv-shβ-catenin). In vitro, Lv-shβ-catenin significantly decreased the expression of β-catenin, MMP2 and MMP9, and secretion of TGF-β1. In vivo, intratracheal treatment with Lv-shβ-catenin significantly reduced expression of β-catenin in the lung and levels of TGF-β1 in bronchoalveolar lavage fluid, and notably attenuated pulmonary fibrosis as evidenced by hydroxyproline content and collagen I\\III synthesis in silica-administered mice. These results indicate that blockade of the Wnt/β-catenin pathway can prevent the development of silica-induced lung fibrosis. Thus Wnt/β-catenin pathway may be a target in prevention and treatment of silicosis. PMID:26340635

  16. Non-canonical Wnt signaling through Wnt5a/b and a novel Wnt11 gene, Wnt11b, regulates cell migration during avian gastrulation

    PubMed Central

    Hardy, Katharine M.; Garriock, Robert J.; Yatskievych, Tatiana A.; D’Agostino, Susan L.; Antin, Parker B.; Krieg, Paul A.

    2008-01-01

    Knowledge of the molecular mechanisms regulating cell ingression, epithelial-mesenchymal transition and migration movements during amniote gastrulation is steadily improving. In the frog and fish embryo, Wnt5 and Wnt11 ligands are expressed around the blastopore and play an important role in regulating cell movements associated with gastrulation. In the chicken embryo, although Wnt5a and Wnt5b are expressed in the primitive streak, the known Wnt11 gene is expressed in paraxial and intermediate mesoderm and in differentiated myocardial cells. Here, we identify a previously uncharacterized chicken Wnt11 gene, Wnt11b, that is orthologous to the frog Wnt11 and zebrafish Wnt11 (silberblick) genes. Chicken Wnt11b is expressed in the primitive streak in a pattern similar to chicken Wnt5a and Wnt5b. When non-canonical Wnt signaling is blocked using a Dishevelled dominant negative protein, gastrulation movements are inhibited and cells accumulate in the primitive streak. Furthermore, disruption of non-canonical Wnt signaling by overexpression of full length or dominant negative Wnt11b or Wnt5a constructions abrogates normal cell migration through the primitive streak. We conclude that non-canonical Wnt signaling, mediated in part by Wnt11b, is important for regulation of gastrulation cell movements in the avian embryo. PMID:18602094

  17. Updating the Wnt pathways

    PubMed Central

    Yu, Jia; Virshup, David M.

    2014-01-01

    In the three decades since the discovery of the Wnt1 proto-oncogene in virus-induced mouse mammary tumours, our understanding of the signalling pathways that are regulated by the Wnt proteins has progressively expanded. Wnts are involved in an complex signalling network that governs multiple biological processes and cross-talk with multiple additional signalling cascades, including the Notch, FGF (fibroblast growth factor), SHH (Sonic hedgehog), EGF (epidermal growth factor) and Hippo pathways. The Wnt signalling pathway also illustrates the link between abnormal regulation of the developmental processes and disease manifestation. Here we provide an overview of Wnt-regulated signalling cascades and highlight recent advances. We focus on new findings regarding the dedicated Wnt production and secretion pathway with potential therapeutic targets that might be beneficial for patients with Wnt-related diseases. PMID:25208913

  18. Secreted and transmembrane wnt inhibitors and activators.

    PubMed

    Cruciat, Cristina-Maria; Niehrs, Christof

    2013-03-01

    Signaling by the Wnt family of secreted glycoproteins plays important roles in embryonic development and adult homeostasis. Wnt signaling is modulated by a number of evolutionarily conserved inhibitors and activators. Wnt inhibitors belong to small protein families, including sFRP, Dkk, WIF, Wise/SOST, Cerberus, IGFBP, Shisa, Waif1, APCDD1, and Tiki1. Their common feature is to antagonize Wnt signaling by preventing ligand-receptor interactions or Wnt receptor maturation. Conversely, the Wnt activators, R-spondin and Norrin, promote Wnt signaling by binding to Wnt receptors or releasing a Wnt-inhibitory step. With few exceptions, these antagonists and agonists are not pure Wnt modulators, but also affect additional signaling pathways, such as TGF-β and FGF signaling. Here we discuss their interactions with Wnt ligands and Wnt receptors, their role in developmental processes, as well as their implication in disease. PMID:23085770

  19. Wnt6, Wnt10a and Wnt10b inhibit adipogenesis and stimulate osteoblastogenesis through a β-catenin-dependent mechanism.

    PubMed

    Cawthorn, William P; Bree, Adam J; Yao, Yao; Du, Baowen; Hemati, Nahid; Martinez-Santibañez, Gabriel; MacDougald, Ormond A

    2012-02-01

    Wnt10b is an established regulator of mesenchymal stem cell (MSC) fate that inhibits adipogenesis and stimulates osteoblastogenesis, thereby impacting bone mass in vivo. However, downstream mechanisms through which Wnt10b exerts these effects are poorly understood. Moreover, whether other endogenous Wnt ligands also modulate MSC fate remains to be fully addressed. In this study, we identify Wnt6 and Wnt10a as additional Wnt family members that, like Wnt10b, are downregulated during development of white adipocytes in vivo and in vitro, suggesting that Wnt6 and/or Wnt10a may also inhibit adipogenesis. To assess the relative activities of Wnt6, Wnt10a and Wnt10b to regulate mesenchymal cell fate, we used gain- and loss-of function approaches in bipotential ST2 cells and in 3T3-L1 preadipocytes. Enforced expression of Wnt10a stabilizes β-catenin, suppresses adipogenesis and stimulates osteoblastogenesis to a similar extent as Wnt10b, whereas stable expression of Wnt6 has a weaker effect on these processes than Wnt10a or Wnt10b. In contrast, knockdown of endogenous Wnt6 is associated with greater preadipocyte differentiation and impaired osteoblastogenesis than knockdown of Wnt10a or Wnt10b, suggesting that, among these Wnt ligands, Wnt6 is the most potent endogenous regulator of MSC fate. Finally, we show that knockdown of β-catenin completely prevents the inhibition of adipogenesis and stimulation of osteoblast differentiation by Wnt6, Wnt10a or Wnt10b. Potential mechanisms whereby Wnts regulate fate of MSCs downstream of β-catenin are also investigated. In conclusion, this study identifies Wnt10a and Wnt6 as additional regulators of MSC fate and demonstrates that mechanisms downstream of β-catenin are required for Wnt6, Wnt10a and Wnt10b to influence differentiation of mesenchymal precursors. PMID:21872687

  20. Processing of IN-718 Lattice Block Castings

    NASA Technical Reports Server (NTRS)

    Hebsur, Mohan G.

    2002-01-01

    Recently a low cost casting method known as lattice block casting has been developed by JAM Corporation, Wilmington, Massachusetts for engineering materials such as aluminum and stainless steels that has shown to provide very high stiffness and strength with only a fraction of density of the alloy. NASA Glenn Research Center has initiated research to investigate lattice block castings of high temperature Ni-base superalloys such as the model system Inconel-718 (IN-718) for lightweight nozzle applications. Although difficulties were encountered throughout the manufacturing process , a successful investment casting procedure was eventually developed. Wax formulation and pattern assembly, shell mold processing, and counter gravity casting techniques were developed. Ten IN-718 lattice block castings (each measuring 15-cm wide by 30-cm long by 1.2-cm thick) have been successfully produced by Hitchiner Gas Turbine Division, Milford, New Hampshire, using their patented counter gravity casting techniques. Details of the processing and resulting microstructures are discussed in this paper. Post casting processing and evaluation of system specific mechanical properties of these specimens are in progress.

  1. Wnt4 is required for ostia development in the Drosophila heart.

    PubMed

    Chen, Zhimin; Zhu, Jun-Yi; Fu, Yulong; Richman, Adam; Han, Zhe

    2016-05-15

    The Drosophila ostia are valve-like structures in the heart with functional similarity to vertebrate cardiac valves. The Wnt/β-catenin signaling pathway is critical for valve development in zebrafish and mouse, but the key ligand(s) for valve induction remains unclear. We observed high levels of Wnt4 gene expression in Drosophila ostia progenitor cells, immediately prior to morphological differentiation of these cells associated with ostia formation. This differentiation was blocked in Wnt4 mutants and in flies expressing canonical Wnt signaling pathway inhibitors but not inhibitors of the planar cell polarity pathway. High levels of Wnt4 dependent activation of a canonical Wnt signaling reporter was observed specifically in ostia progenitor cells. In vertebrate valve formation Wnt signaling is active in cells undergoing early endothelial-mesenchymal transition (EMT) and the Wnt9 homolog of Drosophila Wnt4 is expressed in valve progenitors. In demonstrating an essential role for Wnt4 in ostia development we have identified similarities between molecular and cellular events associated with early EMT during vertebrate valve development and the differentiation and partial delamination of ostia progenitor cells in the process of ostia formation. PMID:26994311

  2. Ehrlichia chaffeensis Exploits Canonical and Noncanonical Host Wnt Signaling Pathways To Stimulate Phagocytosis and Promote Intracellular Survival

    PubMed Central

    Luo, Tian; Dunphy, Paige S.; Lina, Taslima T.

    2015-01-01

    Ehrlichia chaffeensis invades and survives in phagocytes by modulating host cell processes and evading innate defenses, but the mechanisms are not fully defined. Recently we have determined that E. chaffeensis tandem repeat proteins (TRPs) are type 1 secreted effectors involved in functionally diverse interactions with host targets, including components of the evolutionarily conserved Wnt signaling pathways. In this study, we demonstrated that induction of host canonical and noncanonical Wnt pathways by E. chaffeensis TRP effectors stimulates phagocytosis and promotes intracellular survival. After E. chaffeensis infection, canonical and noncanonical Wnt signalings were significantly stimulated during early stages of infection (1 to 3 h) which coincided with dephosphorylation and nuclear translocation of β-catenin, a major canonical Wnt signal transducer, and NFATC1, a noncanonical Wnt transcription factor. In total, the expression of ∼44% of Wnt signaling target genes was altered during infection. Knockdown of TRP120-interacting Wnt pathway components/regulators and other critical components, such as Wnt5a ligand, Frizzled 5 receptor, β-catenin, nuclear factor of activated T cells (NFAT), and major signaling molecules, resulted in significant reductions in the ehrlichial load. Moreover, small-molecule inhibitors specific for components of canonical and noncanonical (Ca2+ and planar cell polarity [PCP]) Wnt pathways, including IWP-2, which blocks Wnt secretion, significantly decreased ehrlichial infection. TRPs directly activated Wnt signaling, as TRP-coated microspheres triggered phagocytosis which was blocked by Wnt pathway inhibitors, demonstrating a key role of TRP activation of Wnt pathways to induce ehrlichial phagocytosis. These novel findings reveal that E. chaffeensis exploits canonical and noncanonical Wnt pathways through TRP effectors to facilitate host cell entry and promote intracellular survival. PMID:26712203

  3. Ehrlichia chaffeensis Exploits Canonical and Noncanonical Host Wnt Signaling Pathways To Stimulate Phagocytosis and Promote Intracellular Survival.

    PubMed

    Luo, Tian; Dunphy, Paige S; Lina, Taslima T; McBride, Jere W

    2016-03-01

    Ehrlichia chaffeensis invades and survives in phagocytes by modulating host cell processes and evading innate defenses, but the mechanisms are not fully defined. Recently we have determined that E. chaffeensis tandem repeat proteins (TRPs) are type 1 secreted effectors involved in functionally diverse interactions with host targets, including components of the evolutionarily conserved Wnt signaling pathways. In this study, we demonstrated that induction of host canonical and noncanonical Wnt pathways by E. chaffeensis TRP effectors stimulates phagocytosis and promotes intracellular survival. After E. chaffeensis infection, canonical and noncanonical Wnt signalings were significantly stimulated during early stages of infection (1 to 3 h) which coincided with dephosphorylation and nuclear translocation of β-catenin, a major canonical Wnt signal transducer, and NFATC1, a noncanonical Wnt transcription factor. In total, the expression of ∼44% of Wnt signaling target genes was altered during infection. Knockdown of TRP120-interacting Wnt pathway components/regulators and other critical components, such as Wnt5a ligand, Frizzled 5 receptor, β-catenin, nuclear factor of activated T cells (NFAT), and major signaling molecules, resulted in significant reductions in the ehrlichial load. Moreover, small-molecule inhibitors specific for components of canonical and noncanonical (Ca(2+) and planar cell polarity [PCP]) Wnt pathways, including IWP-2, which blocks Wnt secretion, significantly decreased ehrlichial infection. TRPs directly activated Wnt signaling, as TRP-coated microspheres triggered phagocytosis which was blocked by Wnt pathway inhibitors, demonstrating a key role of TRP activation of Wnt pathways to induce ehrlichial phagocytosis. These novel findings reveal that E. chaffeensis exploits canonical and noncanonical Wnt pathways through TRP effectors to facilitate host cell entry and promote intracellular survival. PMID:26712203

  4. Wnt Signaling in Bone

    PubMed Central

    Kubota, Takuo; Michigami, Toshimi; Ozono, Keiichi

    2010-01-01

    Wnt signaling is involved not only in embryonic development but also in maintenance of homeostasis in postnatal tissues. Multiple lines of evidence have increased understanding of the roles of Wnt signaling in bone since mutations in the LRP5 gene were identified in human bone diseases. Canonical Wnt signaling promotes mesenchymal progenitor cells to differentiate into osteoblasts. The canonical Wnt/β-catenin pathway possibly through Lrp6, a co-receptor for Wnts as well as Lrp5, in osteoblasts regulates bone resorption by increasing the OPG/RANKL ratio. However, endogenous inhibitors of Wnt signaling including sclerostin block bone formation. Regulation of sclerostin appears to be one of the mechanisms of PTH anabolic actions on bone. Since sclerostin is almost exclusively expressed in osteocytes, inhibition of sclerostin is the most promising design. Surprisingly, Lrp5 controls bone formation by inhibiting serotonin synthesis in the duodenum, but not by directly promoting bone formation. Pharmacological intervention may be considered in many components of the canonical Wnt signaling pathway, although adverse effects and tumorigenicity to other tissues are important. More studies will be needed to fully understand how the Wnt signaling pathway actually influences bone metabolism and to assure the safety of new interventions. PMID:23926379

  5. Downregulation of the CXCR4/CXCL12 axis blocks the activation of the Wnt/β-catenin pathway in human colon cancer cells.

    PubMed

    Song, Zhi-Yu; Gao, Zu-Hua; Chu, Jia-Hui; Han, Xiu-Zhen; Qu, Xian-Jun

    2015-04-01

    Chemokine CXCL12 is an extracellular chemokine, which binds to its cell surface receptor CXCR4. High expressions of CXCR4 and CXCL12 are associated with biological malignant potential in colon cancers. We aimed to investigate the roles of the CXCR4/CXCL12 axis in activation of the Wnt/β-catenin pathway in the development of colon cancers. Using colon cancer cell line, we performed the RNA interference assay to downregulate the expression of CXCR4. Cells were exposed to CXCL12 and their growth and metastatic activity were examined. Matrix metalloproteinases (MMPs) activity were analyzed by the gelatin zymography assay. Cell migration ability was estimated by assays of scratch wound and transwell chamber. The expression of CXCR4 and molecules relevant to the Wnt/β-catenin pathway were analyzed by the western blotting and real-time PCR assays. Human colon cancer HT-29 cells identified high expression of CXCR4. HT-29 cells highly responded to CXCL12 stimulation, showing the increase of cell proliferation, invasion and migration through the Matrigeal. The secretion and activity of MMP-2 and MMP-9 were also stimulated in HT-29 cells exposure to CXCL12. However, the CXCR4 knockdown HT-29 cells did not response to CXCL12 stimulation. We suggested that the activation of the CXCR4/CXCL12 axis be blocked in the CXCR4 knockdown cells. This study indicated that one key to the role of the CXCR4/CXCL12 axis is activation of the Wnt/β-catenin pathway. Downregulation of the CXCR4/CXCL12 axis thus reduces cancer growth and metastasis. Targeted therapy utilizing the CXCR4/CXCL12 axis could be an effective strategy for treatment of colon cancers. PMID:25960214

  6. Wnt signaling in osteosarcoma.

    PubMed

    Lin, Carol H; Ji, Tao; Chen, Cheng-Fong; Hoang, Bang H

    2014-01-01

    Osteosarcoma (OS) is the most common primary bone malignancy diagnosed in children and adolescents with a high propensity for local invasion and distant metastasis. Despite current multidisciplinary treatments, there has not been a drastic change in overall prognosis within the last two decades. With current treatments, 60-70 % of patients with localized disease survive. Given a propensity of Wnt signaling to control multiple cellular processes, including proliferation, cell fate determination, and differentiation, it is a critical pathway in OS disease progression. At the same time, this pathway is extremely complex with vast arrays of cross-talk. Even though decades of research have linked the role of Wnt to tumorigenesis, there are still outstanding areas that remain poorly understood and even controversial. The canonical Wnt pathway functions to regulate the levels of the transcriptional co-activator β-catenin, which ultimately controls key developmental gene expressions. Given the central role of this mediator, inhibition of Wnt/β-catenin signaling has been investigated as a potential strategy for cancer control. In OS, several secreted protein families modulate the Wnt/β-catenin signaling, including secreted Frizzled-related proteins (sFRPs), Wnt inhibitory protein (WIF), Dickkopf proteins (DKK-1,2,3), sclerostin, and small molecules. This chapter focuses on our current understanding of Wnt/β-catenin signaling in OS, based on recent in vitro and in vivo data. Wnt activates noncanonical signaling pathways as well that are independent of β-catenin which will be discussed. In addition, stem cells and their association with Wnt/β-catenin are important factors to consider. Ultimately, the multiple canonical and noncanonical Wnt/β-catenin agonists and antagonists need to be further explored for potential targeted therapies. PMID:24924167

  7. R26-WntVis reporter mice showing graded response to Wnt signal levels.

    PubMed

    Takemoto, Tatsuya; Abe, Takaya; Kiyonari, Hiroshi; Nakao, Kazuki; Furuta, Yasuhide; Suzuki, Hitomi; Takada, Shinji; Fujimori, Toshihiko; Kondoh, Hisato

    2016-06-01

    The canonical Wnt signaling pathway plays a major role in the regulation of embryogenesis and organogenesis, where signal strength-dependent cellular responses are of particular importance. To assess Wnt signal levels in individual cells, and to circumvent the integration site-dependent bias shown in previous Wnt reporter lines, we constructed a new Wnt signal reporter mouse line R26-WntVis. Heptameric TCF/LEF1 binding sequences were combined with a viral minimal promoter to confer a graded response to the reporter depending on Wnt signal strengths. The histone H2B-EGFP fusion protein was chosen as the fluorescent reporter to facilitate single-cell resolution analyses. This WntVis reporter gene was then inserted into the ROSA26 locus in an orientation opposite to that of the endogenous gene. The R26-WntVis allele was introduced into Wnt3a(-/-) and Wnt3a(vt/-) mutant mouse embryos and compared with wild-type embryos to assess its performance. The R26-WntVis reporter was activated in known Wnt-dependent tissues and responded in a graded fashion to signal intensity. This analysis also indicated that the major Wnt activity early in embryogenesis switched from Wnt3 to Wnt3a around E7.5. The R26-WntVis mouse line will be widely useful for the study of Wnt signal-dependent processes. PMID:27030109

  8. Wnt5a attenuates Wnt3a-induced alkaline phosphatase expression in dental follicle cells

    SciTech Connect

    Sakisaka, Yukihiko; Tsuchiya, Masahiro; Nakamura, Takashi; Tamura, Masato; Shimauchi, Hidetoshi; Nemoto, Eiji

    2015-08-01

    Wnt signaling regulates multiple cellular events such as cell proliferation, differentiation, and apoptosis through β-catenin-dependent canonical and β-catenin-independent noncanonical pathways. Canonical Wnt/β-catenin signaling can promote the differentiation of dental follicle cells, putative progenitor cells for cementoblasts, osteoblasts, and periodontal ligament cells, toward a cementoblast/osteoblast phenotype during root formation, but little is known about the biological significance of noncanonical Wnt signaling in this process. We identified the expression of Wnt5a, a representative noncanonical Wnt ligand, in tooth root lining cells (i.e. precementoblasts/cementoblasts) and dental follicle cells during mouse tooth root development, as assessed by immunohistochemistry. Silencing expression of the Wnt5a gene in a dental follicle cell line resulted in enhancement of the Wnt3a (a representative canonical Wnt ligand)-mediated increase in alkaline phosphatase (ALP) expression. Conversely, treatment with recombinant Wnt5a inhibited the increase in ALP expression, suggesting that Wnt5a signaling functions as a negative regulator of canonical Wnt-mediated ALP expression of dental follicle cells. Wnt5a did not affect the nuclear translocation of β-catenin as well as β-catenin-mediated transcriptional activation of T-cell factor (Tcf) triggered by Wnt3a, suggesting that Wnt5a inhibits the downstream part of the β-catenin-Tcf pathway. These findings suggest the existence of a feedback mechanism between canonical and noncanonical Wnt signaling during the differentiation of dental follicle cells. - Highlights: • Dental follicle cells express Wnt5a during tooth root development. • Silencing of Wnt5a enhances Wnt3a-mediated ALP expression of dental follicle cells. • Conversely, treatment with rWnt5a inhibited the increase in ALP expression. • Wnt5a functions as a negative regulator of Wnt3a-mediated ALP expression.

  9. Blocking Wnt signaling by SFRP-like molecules inhibits in vivo cell proliferation and tumor growth in cells carrying active β-catenin

    PubMed Central

    Lavergne, Elise; Hendaoui, Ismaïl; Coulouarn, Cédric; Ribault, Catherine; Leseur, Julie; Eliat, Pierre-Antoine; Mebarki, Sihem; Corlu, Anne; Clément, Bruno; Musso, Orlando

    2011-01-01

    Constitutive activation of Wnt/β-catenin signaling in cancer results from mutations in pathway components, which frequently coexist with autocrine Wnt signaling or epigenetic silencing of extracellular Wnt antagonists. Among the extracellular Wnt inhibitors, the secreted frizzled-related proteins (SFRPs) are decoy receptors that contain soluble Wnt-binding frizzled domains. In addition to SFRPs, other endogenous molecules harboring frizzled motifs bind to and inhibit Wnt signaling. One of such molecules is V3Nter, a soluble SFRP-like frizzled polypeptide that binds to Wnt3a and inhibits Wnt signaling and expression of the β-catenin target genes cyclin D1 and c-myc. V3Nter is derived from the cell surface extracellular matrix component collagen XVIII. Here, we used HCT116 human colon cancer cells carrying the ΔS45 activating mutation in one of the alleles of β-catenin to show that V3Nter and SFRP-1 decrease baseline and Wnt3a-induced β-catenin stabilization. Consequently, V3Nter reduces the growth of human colorectal cancer xenografts by specifically controlling cell proliferation and cell cycle progression, without affecting angiogenesis or apoptosis, as shown by decreased [3H] thymidine (in vitro) or BrdU (in vivo) incorporation, clonogenesis assays, cell cycle analysis and magnetic resonance imaging in living mice. Additionally, V3Nter switches off the β-catenin target gene expression signature in vivo. Moreover, experiments with β-catenin allele-targeted cells showed that the ΔS45 β-catenin allele hampers, but does not abrogate inhibition of Wnt signaling by SFRP-1 or by the SFRP-like frizzled domain. Finally, neither SFRP-1 nor V3Nter affect β-catenin signaling in SW480 cells carrying non functional APC. Thus, SFRP-1 and the SFRP-like molecule V3Nter can inhibit tumor growth of β-catenin-activated tumor cells in vivo. PMID:20856206

  10. Blocking Wnt signaling by SFRP-like molecules inhibits in vivo cell proliferation and tumor growth in cells carrying active β-catenin.

    PubMed

    Lavergne, E; Hendaoui, I; Coulouarn, C; Ribault, C; Leseur, J; Eliat, P-A; Mebarki, S; Corlu, A; Clément, B; Musso, O

    2011-01-27

    Constitutive activation of Wnt/β-catenin signaling in cancer results from mutations in pathway components, which frequently coexist with autocrine Wnt signaling or epigenetic silencing of extracellular Wnt antagonists. Among the extracellular Wnt inhibitors, the secreted frizzled-related proteins (SFRPs) are decoy receptors that contain soluble Wnt-binding frizzled domains. In addition to SFRPs, other endogenous molecules harboring frizzled motifs bind to and inhibit Wnt signaling. One of such molecules is V3Nter, a soluble SFRP-like frizzled polypeptide that binds to Wnt3a and inhibits Wnt signaling and expression of the β-catenin target genes cyclin D1 and c-myc. V3Nter is derived from the cell surface extracellular matrix component collagen XVIII. Here, we used HCT116 human colon cancer cells carrying the ΔS45 activating mutation in one of the alleles of β-catenin to show that V3Nter and SFRP-1 decrease baseline and Wnt3a-induced β-catenin stabilization. Consequently, V3Nter reduces the growth of human colorectal cancer xenografts by specifically controlling cell proliferation and cell cycle progression, without affecting angiogenesis or apoptosis, as shown by decreased [(3)H]-thymidine (in vitro) or BrdU (in vivo) incorporation, clonogenesis assays, cell cycle analysis and magnetic resonance imaging in living mice. Additionally, V3Nter switches off the β-catenin target gene expression signature in vivo. Moreover, experiments with β-catenin allele-targeted cells showed that the ΔS45 β-catenin allele hampers, but does not abrogate, inhibition of Wnt signaling by SFRP-1 or by the SFRP-like frizzled domain. Finally, neither SFRP-1 nor V3Nter affect β-catenin signaling in SW480 cells carrying nonfunctional Adenomatous polyposis coli. Thus, SFRP-1 and the SFRP-like molecule V3Nter can inhibit tumor growth of β-catenin-activated tumor cells in vivo. PMID:20856206

  11. The antidepressant roles of Wnt2 and Wnt3 in stress-induced depression-like behaviors.

    PubMed

    Zhou, W-J; Xu, N; Kong, L; Sun, S-C; Xu, X-F; Jia, M-Z; Wang, Y; Chen, Z-Y

    2016-01-01

    Wnts-related signaling pathways have been reported to play roles in the pathogenesis of stress-induced depression-like behaviors. However, there is relatively few direct evidence to indicate the effect of Wnt ligands on this process. Here, we investigated the role of Wnts in mediating chronic restraint stress (CRS)-induced depression-like behaviors. We found that CRS induced a significant decrease in the expression of Wnt2 and Wnt3 in the ventral hippocampus (VH) but not in the dorsal hippocampus. Knocking down Wnt2 or Wnt3 in the VH led to impaired Wnt/β-catenin signaling, neurogenesis deficits and depression-like behaviors. In contrast, overexpression of Wnt2 or Wnt3 reversed CRS-induced depression-like behaviors. Moreover, Wnt2 and Wnt3 activated cAMP response element-binding protein (CREB) and there was CREB-dependent positive feedback between Wnt2 and Wnt3. Finally, fluoxetine treatment increased Wnt2 and Wnt3 levels in the VH and knocking down Wnt2 or Wnt3 abolished the antidepressant effect of fluoxetine. Taken together, our study indicates essential roles for Wnt2 and Wnt3 in CRS-induced depression-like behaviors and antidepressant. PMID:27622936

  12. Specific functions of the Wnt signaling system in gene regulatory networks throughout the early sea urchin embryo

    PubMed Central

    Cui, Miao; Siriwon, Natnaree; Li, Enhu; Davidson, Eric H.; Peter, Isabelle S.

    2014-01-01

    Wnt signaling affects cell-fate specification processes throughout embryonic development. Here we take advantage of the well-studied gene regulatory networks (GRNs) that control pregastrular sea urchin embryogenesis to reveal the gene regulatory functions of the entire Wnt-signaling system. Five wnt genes, three frizzled genes, two secreted frizzled-related protein 1 genes, and two Dickkopf genes are expressed in dynamic spatial patterns in the pregastrular embryo of Strongylocentrotus purpuratus. We present a comprehensive analysis of these genes in each embryonic domain. Total functions of the Wnt-signaling system in regulatory gene expression throughout the embryo were studied by use of the Porcupine inhibitor C59, which interferes with zygotic Wnt ligand secretion. Morpholino-mediated knockdown of each expressed Wnt ligand demonstrated that individual Wnt ligands are functionally distinct, despite their partially overlapping spatial expression. They target specific embryonic domains and affect particular regulatory genes. The sum of the effects of blocking expression of individual wnt genes is shown to equal C59 effects. Remarkably, zygotic Wnt-signaling inputs are required for only three general aspects of embryonic specification: the broad activation of endodermal GRNs, the regional specification of the immediately adjacent stripe of ectoderm, and the restriction of the apical neurogenic domain. All Wnt signaling in this pregastrular embryo is short range (and/or autocrine). Furthermore, we show that the transcriptional drivers of wnt genes execute important specification functions in the embryonic domains targeted by the ligands, thus connecting the expression and function of wnt genes by encoded cross-regulatory interactions within the specific regional GRNs. PMID:25385617

  13. Second hit in cervical carcinogenesis process: involvement of wnt/beta catenin pathway

    PubMed Central

    Perez-Plasencia, Carlos; Duenas-Gonzalez, Alfonso; Alatorre-Tavera, Brenda

    2008-01-01

    The Human papillomavirus plays an important role in the initiation and progression of cervical cancer. However, it is a necessary but not sufficient cause to develop invasive carcinoma; hence, other factors are required in the pathogenesis of this malignancy. In this review we explore the hypothesis of the deregulation of wnt/β-catenin signaling pathway as a "second hit" required to develop cervical cancer. PMID:18606007

  14. Liposomal packaging generates Wnt protein with in vivo biological activity.

    PubMed

    Morrell, Nathan T; Leucht, Philipp; Zhao, Ludan; Kim, Jae-Beom; ten Berge, Derk; Ponnusamy, Karthik; Carre, A Lyonel; Dudek, Henryk; Zachlederova, Marie; McElhaney, Michael; Brunton, Shirley; Gunzner, Janet; Callow, Marinella; Polakis, Paul; Costa, Mike; Zhang, Xiaoyan M; Helms, Jill A; Nusse, Roel

    2008-01-01

    Wnt signals exercise strong cell-biological and regenerative effects of considerable therapeutic value. There are, however, no specific Wnt agonists and no method for in vivo delivery of purified Wnt proteins. Wnts contain lipid adducts that are required for activity and we exploited this lipophilicity by packaging purified Wnt3a protein into lipid vesicles. Rather than being encapsulated, Wnts are tethered to the liposomal surface, where they enhance and sustain Wnt signaling in vitro. Molecules that effectively antagonize soluble Wnt3a protein but are ineffective against the Wnt3a signal presented by a cell in a paracrine or autocrine manner are also unable to block liposomal Wnt3a activity, suggesting that liposomal packaging mimics the biological state of active Wnts. When delivered subcutaneously, Wnt3a liposomes induce hair follicle neogenesis, demonstrating their robust biological activity in a regenerative context. PMID:18698373

  15. Processing by data and program blocks

    NASA Technical Reports Server (NTRS)

    Schaffner, M. R.

    1978-01-01

    A processing system is presented that implements simultaneously the efficiency of the special-purpose processor and the total applicability of the general-purpose computer - characteristics commonly thought of as being mutually exclusive. This is achieved through specializing the machine by programming the hardware structure, rather than by adding software systems to it. Data are organized in circulating pages which form a multiplicity of local dynamic memories for each process. Programs are made up of modules, each describing a transient special-purpose machine. A characteristic of this approach is that the processes are data-driven, rather than program-driven. The programming language presents significant flexibility and efficiency in modeling certain classes of problems, and it may be of interest as an implementation model in a broader context. Applications to real-time processing of radar signals are reported. The relevance of characteristics of this system to problems in multiprogramming and multiprocessing systems is discussed.

  16. Signal transduction by the Wnt family of ligands.

    PubMed Central

    Dale, T C

    1998-01-01

    The Wnt genes encode a large family of secreted polypeptides that mediate cell-cell communication in diverse developmental processes. The loss or inappropriate activation of Wnt expression has been shown to alter cell fate, morphogenesis and mitogenesis. Recent progress has identified Wnt receptors and components of an intracellular signalling pathway that mediate Wnt-dependent transcription. This review will highlight this 'core' Wnt signal-transduction pathway, but also aims to reveal the potential diversity of Wnt signalling targets. Particular attention will be paid to the overlap between developmental biology and oncogenesis, since recent progress shows Wnt signalling forms a paradigm for an interdisciplinary approach. PMID:9425102

  17. Light-emitting block copolymers composition, process and use

    DOEpatents

    Ferraris, John P.; Gutierrez, Jose J.

    2006-11-14

    Generally, and in one form, the present invention is a composition of light-emitting block copolymer. In another form, the present invention is a process producing a light-emitting block copolymers that intends polymerizing a first di(halo-methyl) aromatic monomer compound in the presence of an anionic initiator and a base to form a polymer and contacting a second di(halo-methyl) aromatic monomer compound with the polymer to form a homopolymer or block copolymer wherein the block copolymer is a diblock, triblock, or star polymer. In yet another form, the present invention is an electroluminescent device comprising a light-emitting block copolymer, wherein the electroluminescent device is to be used in the manufacturing of optical and electrical devices.

  18. Structural Insights into the Inhibition of Wnt Signaling by Cancer Antigen 5T4/Wnt-Activated Inhibitory Factor 1

    PubMed Central

    Zhao, Yuguang; Malinauskas, Tomas; Harlos, Karl; Jones, E. Yvonne

    2014-01-01

    Summary The tumor antigen 5T4/WAIF1 (Wnt-activated inhibitory factor 1; also known as Trophoblast glycoprotein TPBG) is a cell surface protein targeted in multiple cancer immunotherapy clinical trials. Recently, it has been shown that 5T4/WAIF1 inhibits Wnt/β-catenin signaling, a signaling system central to many developmental and pathological processes. Wnt/β-catenin signaling is controlled by multiple inhibitors and activators. Here, we report crystal structures for the extracellular domain of 5T4/WAIF1 at 1.8 Å resolution. They reveal a highly glycosylated, rigid core, comprising eight leucine-rich repeats (LRRs), which serves as a platform to present evolutionarily conserved surface residues in the N-terminal LRR1. Structural and cell-based analyses, coupled with previously reported in vivo data, suggest that Tyr325 plus the LRR1 surface centered on a second exposed aromatic residue, Phe97, are essential for inhibition of Wnt/β-catenin signaling. These results provide a structural basis for the development of 5T4/WAIF1-targeted therapies that preserve or block 5T4/WAIF1-mediated inhibition of Wnt/β-catenin signaling. PMID:24582434

  19. A Wnt5a signaling pathway in the pathogenesis of HIV-1 gp120-induced pain

    PubMed Central

    Yuan, Su-Bo; Ji, Guangchen; Li, Bei; Andersson, Tommy; Neugebauer, Volker; Tang, Shao-Jun

    2015-01-01

    Pathological pain is one of the most common neurological complications in HIV-1/AIDS patients. However, the pathogenic process is unclear. Our recent studies show that Wnt5a is up-regulated in the spinal cord dorsal horn of the HIV patients who develop pain and that HIV-1 gp120, a potential causal factor of the HIV-associated pain, rapidly up-regulates Wnt5a in the mouse SDH. Using a mouse model, we show here that a specific Wnt5a antagonist, Box-5, attenuated gp120-induced mechanical allodynia. Conversely, a Wnt5a agonist, Foxy5, facilitated the allodynia. To elucidate the molecular mechanism by which Wnt5a regulates gp120-induced allodynia, we tested the role of the JNK/TNF-α pathway. We observed that the JNK-specific inhibitor SP600125 blocked either gp120- or Foxy5-induced allodynia. Similarly, the TNF-α-specific antagonist Enbrel also reversed either gp120- or Foxy5-induced allodynia. These data suggest that JNK and TNF-α mediate the biological effects of Wnt5a in regulating gp120-induced allodynia. To investigate the cellular mechanism, we performed extracellular single-unit recording from SDH neurons in anesthetized mice. Both Box5 and SP600125 negated gp120-induced potentiation of SDH neuron spiking evoked by mechanical stimulation of the hindpaw. Furthermore, while Foxy5 potentiated spike frequency of SDH neurons, either SP600125 or Enbrel blocked the potentiation. The data indicate that Wnt5a potentiates the activity of SDH neurons via the JNK-TNF-α pathway. Collectively, our findings suggest that Wnt5a regulates the pathogenesis of gp120-induced pain, likely by sensitizing pain-processing SDH neurons via JNK/TNF-α signaling. PMID:25840108

  20. Wnt5a Signaling in Cancer.

    PubMed

    Asem, Marwa S; Buechler, Steven; Wates, Rebecca Burkhalter; Miller, Daniel L; Stack, M Sharon

    2016-01-01

    Wnt5a is involved in activating several non-canonical WNT signaling pathways, through binding to different members of the Frizzled- and Ror-family receptors. Wnt5a signaling is critical for regulating normal developmental processes, including proliferation, differentiation, migration, adhesion and polarity. However, the aberrant activation or inhibition of Wnt5a signaling is emerging as an important event in cancer progression, exerting both oncogenic and tumor suppressive effects. Recent studies show the involvement of Wnt5a in regulating cancer cell invasion, metastasis, metabolism and inflammation. In this article, we review findings regarding the molecular mechanisms and roles of Wnt5a signaling in various cancer types, and highlight Wnt5a in ovarian cancer. PMID:27571105

  1. Wnt and the Wnt signaling pathway in bone development and disease

    PubMed Central

    Wang, Yiping; Li, Yi-Ping; Paulson, Christie; Shao, Jian-Zhong; Zhang, Xiaoling; Wu, Mengrui; Chen, Wei

    2014-01-01

    Wnt signaling affects both bone modeling, which occurs during development, and bone remodeling, which is a lifelong process involving tissue renewal. Wnt signals are especially known to affect the differentiation of osteoblasts. In this review, we summarize recent advances in understanding the mechanisms of Wnt signaling, which is divided into two major branches: the canonical pathway and the noncanonical pathway. The canonical pathway is also called the Wnt/β-catenin pathway. There are two major noncanonical pathways: the Wnt-planar cell polarity pathway (Wnt-PCP pathway) and the Wnt-calcium pathway (Wnt-Ca2+ pathway). This review also discusses how Wnt ligands, receptors, intracellular effectors, transcription factors, and antagonists affect both the bone modeling and bone remodeling processes. We also review the role of Wnt ligands, receptors, intracellular effectors, transcription factors, and antagonists in bone as demonstrated in mouse models. Disrupted Wnt signaling is linked to several bone diseases, including osteoporosis, van Buchem disease, and sclerosteosis. Studying the mechanism of Wnt signaling and its interactions with other signaling pathways in bone will provide potential therapeutic targets to treat these bone diseases. PMID:24389191

  2. NOTUM is a potential pharmacodynamic biomarker of Wnt pathway inhibition

    PubMed Central

    Madan, Babita; Ke, Zhiyuan; Lei, Zheng Deng; Oliver, Frois Ashley; Oshima, Masanobu; Lee, May Ann; Rozen, Steve; Virshup, David M.

    2016-01-01

    Activation of Wnt signaling due to Wnt overexpression or mutations of Wnt pathway components is associated with various cancers. Blocking Wnt secretion by inhibiting PORCN enzymatic activity has shown efficacy in a subset of cancers with elevated Wnt signaling. Predicting response to upstream Wnt inhibitors and monitoring response to therapeutics is challenging due to the paucity of well-defined biomarkers. In this study we identify Notum as a potential biomarker for Wnt driven cancers and show that coordinate regulation of NOTUM and AXIN2 expression may be a useful predictor of response to PORCN inhibitors. Most importantly, as NOTUM is a secreted protein and its levels in blood correlate with tumor growth, it has potential as a pharmacodynamic biomarker for PORCN and other Wnt pathway inhibitors. PMID:26848981

  3. Wnt Signalling Promotes Actin Dynamics during Axon Remodelling through the Actin-Binding Protein Eps8

    PubMed Central

    Salinas, Patricia C.

    2015-01-01

    Upon arrival at their synaptic targets, axons slow down their growth and extensively remodel before the assembly of presynaptic boutons. Wnt proteins are target-derived secreted factors that promote axonal remodelling and synaptic assembly. In the developing spinal cord, Wnts secreted by motor neurons promote axonal remodelling of NT-3 responsive dorsal root ganglia neurons. Axon remodelling induced by Wnts is characterised by growth cone pausing and enlargement, processes that depend on the re-organisation of microtubules. However, the contribution of the actin cytoskeleton has remained unexplored. Here, we demonstrate that Wnt3a regulates the actin cytoskeleton by rapidly inducing F-actin accumulation in growth cones from rodent DRG neurons through the scaffold protein Dishevelled-1 (Dvl1) and the serine-threonine kinase Gsk3β. Importantly, these changes in actin cytoskeleton occurs before enlargement of the growth cones is evident. Time-lapse imaging shows that Wnt3a increases lamellar protrusion and filopodia velocity. In addition, pharmacological inhibition of actin assembly demonstrates that Wnt3a increases actin dynamics. Through a yeast-two hybrid screen, we identified the actin-binding protein Eps8 as a direct interactor of Dvl1, a scaffold protein crucial for the Wnt signalling pathway. Gain of function of Eps8 mimics Wnt-mediated axon remodelling, whereas Eps8 silencing blocks the axon remodelling activity of Wnt3a. Importantly, blockade of the Dvl1-Eps8 interaction completely abolishes Wnt3a-mediated axonal remodelling. These findings demonstrate a novel role for Wnt-Dvl1 signalling through Eps8 in the regulation of axonal remodeling. PMID:26252776

  4. Decoupling Processes in Block-and-ash Flows

    NASA Astrophysics Data System (ADS)

    Hanenkamp, E.; Davies, T.

    2009-05-01

    Lava dome collapse and collapse of lava flow fronts generate short-lived, highly mobile block-and-ash flows, usually comprising three components, a high-density, ground-hugging basal avalanche, a low-density ash cloud surge and a more dilute ash cloud on top. Generally the basal avalanche is valley-confined whereas the overlying surge has the ability to decouple from the dense basal part, overtop topographic barriers and affect greater areas than the basal avalanche, hence posing a larger hazard to the population. These processes have been observed at several volcanoes, e.g. Unzen (Japan), Merapi (Indonesia) and Soufriere Hills (Montserrat), resulting often in the loss of lives. Decoupling is often accentuated by topographic obstructions causing blocking or deflecting of the basal avalanche. Currently laboratory flume experiments are being undertaken to examine the factors initiating and influencing the decoupling processes of block-and-ash flows. A better understanding of the flow dynamics of a moving block-and-ash flow and of the interaction with the underlying substrate and topographic irregularities will provide better hazard zone delineation maps for the future. The experimental results will be used to investigate the emplacement of the block-and-ash flows at Mount Tarawera, New Zealand, where the deposits show indicators of possible decoupling processes during the AD 1305 Kaharoa eruptive episode.

  5. Wnt trafficking: new insights into Wnt maturation, secretion and spreading.

    PubMed

    Port, Fillip; Basler, Konrad

    2010-10-01

    Proteins of the Wnt family are secreted signaling molecules that regulate multiple processes in animal development and control tissue homeostasis in the adult. Wnts spread over considerable distances to regulate gene expression in cells located at distant sites. Paradoxically, Wnts are poorly mobile because of their posttranslational modification with lipids. Recent evidence suggests that several pathways exist that are capable of transforming hydrophobic, insoluble Wnts into long-range signaling molecules. Furthermore, the discovery of Wntless as a protein specifically required for the secretion of Wnt suggests that Wnt trafficking through the secretory pathway is already under special scrutiny. Here, we review recent data on the molecular machinery that controls Wnt secretion and discuss how Wnts can be mobilized for long-range signaling. PMID:20477987

  6. Role of Wnt signaling in fracture healing

    PubMed Central

    Xu, Huiyun; Duan, Jing; Ning, Dandan; Li, Jingbao; Liu, Ruofei; Yang, Ruixin; Jiang, Jean X.; Shang, Peng

    2014-01-01

    The Wnt signaling pathway is well known to play major roles in skeletal development and homeostasis. In certain aspects, fracture repair mimics the process of bone embryonic development. Thus, the importance of Wnt signaling in fracture healing has become more apparent in recent years. Here, we summarize recent research progress in the area, which may be conducive to the development of Wnt-based therapeutic strategies for bone repair. [BMB Reports 2014; 47(12): 666-672] PMID:25301020

  7. Inhibition of tumorigenesis driven by different Wnt proteins requires blockade of distinct ligand-binding regions by LRP6 antibodies

    PubMed Central

    Ettenberg, Seth A.; Charlat, Olga; Daley, Michael P.; Liu, Shanming; Vincent, Karen J.; Stuart, Darrin D.; Schuller, Alwin G.; Yuan, Jing; Ospina, Beatriz; Green, John; Yu, Qunyan; Walsh, Renee; Schmitz, Rita; Heine, Holger; Bilic, Sanela; Ostrom, Lance; Mosher, Rebecca; Hartlepp, K. Felix; Zhu, Zhenping; Fawell, Stephen; Yao, Yung-Mae; Stover, David; Finan, Peter M.; Porter, Jeffery A.; Sellers, William R.; Klagge, Ingo M.; Cong, Feng

    2010-01-01

    Disregulated Wnt/β-catenin signaling has been linked to various human diseases, including cancers. Inhibitors of oncogenic Wnt signaling are likely to have a therapeutic effect in cancers. LRP5 and LRP6 are closely related membrane coreceptors for Wnt proteins. Using a phage-display library, we identified anti-LRP6 antibodies that either inhibit or enhance Wnt signaling. Two classes of LRP6 antagonistic antibodies were discovered: one class specifically inhibits Wnt proteins represented by Wnt1, whereas the second class specifically inhibits Wnt proteins represented by Wnt3a. Epitope-mapping experiments indicated that Wnt1 class-specific antibodies bind to the first propeller and Wnt3a class-specific antibodies bind to the third propeller of LRP6, suggesting that Wnt1- and Wnt3a-class proteins interact with distinct LRP6 propeller domains. This conclusion is further supported by the structural functional analysis of LRP5/6 and the finding that the Wnt antagonist Sclerostin interacts with the first propeller of LRP5/6 and preferentially inhibits the Wnt1-class proteins. We also show that Wnt1 or Wnt3a class-specific anti-LRP6 antibodies specifically block growth of MMTV-Wnt1 or MMTV-Wnt3 xenografts in vivo. Therapeutic application of these antibodies could be limited without knowing the type of Wnt proteins expressed in cancers. This is further complicated by our finding that bivalent LRP6 antibodies sensitize cells to the nonblocked class of Wnt proteins. The generation of a biparatopic LRP6 antibody blocks both Wnt1- and Wnt3a-mediated signaling without showing agonistic activity. Our studies provide insights into Wnt-induced LRP5/6 activation and show the potential utility of LRP6 antibodies in Wnt-driven cancer. PMID:20713706

  8. Glucagon Like Peptide-1 Promotes Adipocyte Differentiation via the Wnt4 Mediated Sequestering of Beta-Catenin

    PubMed Central

    Liu, Rui; Li, Na; Lin, Yi; Wang, Mei; Peng, Yongde; Lewi, Keidren; Wang, Qinghua

    2016-01-01

    Glucagon-like peptide-1 (GLP-1) plays a role in the regulation of adipogenesis; however, the precise underlying molecular mechanism has not been fully defined. Wnt was recently identified as an important regulator of adipogenesis. This study aimed to investigate the involvement of the Wnt signaling pathway in the effects of GLP-1 on adipocyte differentiation. 3T3-L1 cells were induced to differentiate. The changes in the expression levels of adipogenic transcription factors and Wnts and the phosphorylation level and subcellular localization of β-catenin were quantified after GLP-1 treatment. GLP-1 stimulated adipocyte differentiation and lipid accumulation, which were accompanied by the expression of adipocyte marker genes. The expression of Wnt4 was upregulated in the process of adipocyte differentiation, which was further enhanced by treatment with GLP-1. β-catenin, an important mediator of the Wnt pathway, was immediately dephosphorylated and translocated from cytoplasm to nucleus when differentiation was induced. In the presence of GLP-1, however, β-catenin was redirected to the cell plasma membrane leading to its decreased accumulation in the nucleus. Knockdown of Wnt4 blocked the effect of GLP-1 on the cellular localization of β-catenin and expression level of adipogenic transcription factors. Our findings showed that GLP-1 promoted adipogenesis through the modulation of the Wnt4/β-catenin signaling pathway, suggesting that the GLP-1-Wntβ-catenin system might be a new target for the treatment of metabolic disease. PMID:27504979

  9. Wingless homolog Wnt11 suppresses bacterial invasion and inflammation in intestinal epithelial cells.

    PubMed

    Liu, Xingyin; Wu, Shaoping; Xia, Yinglin; Li, Xi Emma; Xia, Yuxuan; Zhou, Zhongren David; Sun, Jun

    2011-12-01

    Wnt11 plays an essential role in gastrointestinal epithelial proliferation, and previous investigations have focused on development and immune responses. However, the roles of how enteric bacteria regulate Wnt11 and how Wnt11 modulates the host response to pathogenic bacteria remain unexplored. This study investigated the effects of Salmonella infection on Wnt activation in intestinal epithelial cells. We found that Wnt11 mRNA and protein expression were elevated after Salmonella colonization. Wnt11 protein secretion in epithelial cells was also elevated after bacterial infection. Furthermore, we demonstrated that pathogenic Salmonella regulated Wnt11 expression and localization in vivo. We found a decrease in Salmonella invasion in cells with Wnt11 overexpression compared with cells with normal Wnt11 level. IL-8 mRNA in Wnt11-transfected cells was low; however, it was enhanced in cells with a low level of Wnt11 expression. Functionally, Wnt11 overexpression inhibited Salmonella-induced apoptosis. AvrA is a known bacterial effector protein that stabilizes β-catenin, the downstream regulator of Wnt signaling, and inhibits bacterially induced intestinal inflammation. We observed that Wnt11 expression, secretion, and transcriptional activity were regulated by Salmonella AvrA. Overall, Wnt11 is involved in the protection of the host intestinal cells by blocking the invasion of pathogenic bacteria, suppressing inflammation, and inhibiting apoptosis. Wnt11 is a novel and important contributor to intestinal homeostasis and host defense. PMID:21903761

  10. Wnt signaling pathway in non-small cell lung cancer.

    PubMed

    Stewart, David J

    2014-01-01

    Wnt/β-catenin alterations are prominent in human malignancies. In non-small cell lung cancer (NSCLC), β-catenin and APC mutations are uncommon, but Wnt signaling is important in NSCLC cell lines, and Wnt inhibition reduces proliferation. Overexpression of Wnt-1, -2, -3, and -5a and of Wnt-pathway components Frizzled-8, Dishevelled, Porcupine, and TCF-4 is common in resected NSCLC and is associated with poor prognosis. Conversely, noncanonical Wnt-7a suppresses NSCLC development and is often downregulated. Although β-catenin is often expressed in NSCLCs, it was paradoxically associated with improved prognosis in some series, possibly because of E-cadherin interactions. Downregulation of Wnt inhibitors (eg, by hypermethylation) is common in NSCLC tumor cell lines and resected samples; may be associated with high stage, dedifferentiation, and poor prognosis; and has been reported for AXIN, sFRPs 1-5, WIF-1, Dkk-1, Dkk-3, HDPR1, RUNX3, APC, CDX2, DACT2, TMEM88, Chibby, NKD1, EMX2, ING4, and miR-487b. AXIN is also destabilized by tankyrases, and GSK3β may be inactivated through phosphorylation by EGFR. Preclinically, restoration of Wnt inhibitor function is associated with reduced Wnt signaling, decreased cell proliferation, and increased apoptosis. Wnt signaling may also augment resistance to cisplatin, docetaxel, and radiotherapy, and Wnt inhibitors may restore sensitivity. Overall, available data indicate that Wnt signaling substantially impacts NSCLC tumorigenesis, prognosis, and resistance to therapy, with loss of Wnt signaling inhibitors by promoter hypermethylation or other mechanisms appearing to be particularly important. Wnt pathway antagonists warrant exploration clinically in NSCLC. Agents blocking selected specific β-catenin interactions and approaches to increase expression of downregulated Wnt inhibitors may be of particular interest. PMID:24309006

  11. Increased Expression of Colonic Wnt9A through Sp1-mediated Transcriptional Effects involving Arylsulfatase B, Chondroitin 4-Sulfate, and Galectin-3

    PubMed Central

    Bhattacharyya, Sumit; Feferman, Leo; Tobacman, Joanne K.

    2014-01-01

    In cultured human colonic epithelial cells and mouse colonic tissue, exposure to the common food additive carrageenan leads to inflammation, activation of Wnt signaling, increased Wnt9A expression, and decline in the activity of the enzyme arylsulfatase B (ARSB; N-acetylgalactosamine-4-sulfatase). In this study, the novel transcriptional mechanism by which carrageenan and decline in ARSB increase Wnt9A expression in NCM460 and HT-29 human colonic epithelial cells and in mouse colon is presented. Increased expression of Wnt9A has been associated with multiple malignancies, including colon carcinoma, and with ectodermal and mesoendodermal morphogenesis. When ARSB activity was reduced by siRNA or by exposure to carrageenan (1 μg/ml for 24 h), degradation of chondroitin 4-sulfate (C4S) was inhibited, leading to accumulation of more highly sulfated C4S, which binds less galectin-3, a β-galactoside-binding protein. Nuclear galectin-3 increased and mediated increased binding of Sp1 to the Sp1 consensus sequence in the Wnt9A promoter, shown by oligonucleotide-binding assay and by chromatin immunoprecipitation assay. When galectin-3 was silenced, the increases in Sp1 binding to the Wnt9A promoter and in Wnt9A expression, which followed carrageenan or ARSB silencing, were inhibited. Mithramycin A, a specific inhibitor of Sp1 oligonucleotide binding, and Sp1 siRNA blocked the carrageenan- and ARSB siRNA-induced increases in Wnt9A expression. These studies reveal how carrageenan exposure can lead to transcriptional events in colonic epithelial cells through decline in arylsulfatase B activity, with subsequent impact on C4S, galectin-3, Sp1, and Wnt9A and can exert significant effects on Wnt-initiated signaling and related vital cell processes. PMID:24778176

  12. Wnt your brain be inflamed? Yes, it Wnt!

    PubMed Central

    Marchetti, Bianca; Pluchino, Stefano

    2013-01-01

    The roles of Wnts in neural development, synaptogenesis, and cancer are generally well characterized. Nonetheless, evidence exists that interactions between the immune and nervous systems control major brain regenerative processes ranging from physiological or pathological (reparative) regeneration to neurogenesis and synaptic plasticity. Recent studies describe deregulated Wnt-Fzd signaling in degenerative and inflammatory central nervous system (CNS) disorders, and the expression of Wnt signaling components in the immune system, and in immune-like cells of the mammalian CNS. This would suggest a likely involvement of Wnts in inflammation-driven brain damage and inflammation-directed brain repair. Here, we review how Wnts modulate neuroimmune interactions and offer a perspective on the most challenging therapeutic opportunities for those CNS diseases where injury-reactive Wnt-flavored inflammation precedes secondary neurodegeneration. PMID:23312954

  13. Wnt pathway regulation of intestinal stem cells.

    PubMed

    Mah, Amanda T; Yan, Kelley S; Kuo, Calvin J

    2016-09-01

    Wnt signalling is involved in multiple aspects of embryonic development and adult tissue homeostasis, notably via controlling cellular proliferation and differentiation. Wnt signalling is subject to stringent positive and negative regulation to promote proper development and homeostasis yet avoid aberrant growth. Such multi-layer regulation includes post-translational modification and processing of Wnt proteins themselves, R-spondin (Rspo) amplification of Wnt signalling, diverse receptor families, and intracellular and extracellular antagonists and destruction and transcription complexes. In the gastrointestinal tract, Wnt signalling is crucial for development and renewal of the intestinal epithelium. Intestinal stem cells (ISCs) undergo symmetric division and neutral drift dynamics to renew the intestinal epithelium. Sources of Wnts and Wnt amplifers such as R-spondins are beginning to be elucidated as well as their functional contribution to intestinal homeostasis. In this review we focus on regulation of ISCs and intestinal homeostasis by the Wnt/Rspo pathway, the potential cellular sources of Wnt signalling regulators and highlight potential future areas of study. PMID:27581568

  14. Canonical Wnt signaling in the oligodendroglial lineage--puzzles remain.

    PubMed

    Guo, Fuzheng; Lang, Jordan; Sohn, Jiho; Hammond, Elizabeth; Chang, Marcello; Pleasure, David

    2015-10-01

    The straightforward concept that accentuated Wnt signaling via the Wnt-receptor-β-catenin-TCF/LEF cascade (also termed canonical Wnt signaling or Wnt/β-catenin signaling) delays or blocks oligodendrocyte differentiation is very appealing. According to this concept, canonical Wnt signaling is responsible for remyelination failure in multiple sclerosis and for persistent hypomyelination in periventricular leukomalacia. This has given rise to the hope that pharmacologically inhibiting this signaling will be of therapeutic potential in these disabling neurological disorders. But current studies suggest that Wnt/β-catenin signaling plays distinct roles in oligodendrogenesis, oligodendrocyte differentiation, and myelination in a context-dependent manner (central nervous system regions, developmental stages), and that Wnt/β-catenin signaling interplays with, and is subjected to regulation by, other central nervous system factors and signaling pathways. On this basis, we propose the more nuanced concept that endogenous Wnt/β-catenin activity is delicately and temporally regulated to ensure the seamless development of oligodendroglial lineage cells in different contexts. In this review, we discuss the role Wnt/β-catenin signaling in oligodendrocyte development, focusing on the interpretation of disparate results, and highlighting areas where important questions remain to be answered about oligodendroglial lineage Wnt/β-catenin signaling. PMID:25782433

  15. Roles of Wnt/{beta}-catenin signaling in epithelial differentiation of mesenchymal stem cells

    SciTech Connect

    Wang, Yajing; Sun, Zhaorui; Qiu, Xuefeng; Li, Yan; Qin, Jizheng; Han, Xiaodong

    2009-12-25

    Bone marrow-derived mesenchymal stem cells (MSCs) have been demonstrated to be able to differentiate into epithelial lineage, but the precise mechanisms controlling this process are unclear. Our aim is to explore the roles of Wnt/{beta}-catenin in the epithelial differentiation of MSCs. Using indirect co-culture of rat MSCs with rat airway epithelial cells (RTE), MSCs expressed several airway epithelial markers (cytokeratin 18, tight junction protein occudin, cystic fibrosis transmembrance regulator). The protein levels of some important members in Wnt/{beta}-catenin signaling were determined, suggested down-regulation of Wnt/{beta}-catenin with epithelial differentiation of MSCs. Furthermore, Wnt3{alpha} can inhibit the epithelial differentiation of MSCs. A loss of {beta}-catenin induced by Dickkopf-1 can enhance MSCs differentiation into epithelial cells. Lithium chloride transiently activated {beta}-catenin expression and subsequently decreased {beta}-catenin level and at last inhibited MSCs to differentiate into airway epithelium. Taken together, our study indicated that RTE cells can trigger epithelial differentiation of MSCs. Blocking Wnt/{beta}-catenin signaling may promote MSCs to differentiate towards airway epithelial cells.

  16. Molecular cloning, characterization and expression analysis of Wnt4, Wnt5, Wnt6, Wnt7, Wnt10 and Wnt16 from Litopenaeus vannamei.

    PubMed

    Zhang, Shuang; Li, Chao-Zheng; Yang, Qi-Hui; Dong, Xiao-Hui; Chi, Shu-Yan; Liu, Hong-Yu; Shi, Li-Li; Tan, Bei-Ping

    2016-07-01

    The Wnt (Wg-type MMTV integration site) signaling represents as the negative regulator of virus-induced innate immune responses. Wnt genes act as ligands to activate the Wnt signaling. To know more about the information of Wnt genes in invertebrates, Litopenaeus vannamei Wnt genes (LvWnts) were identified and characterized. In this study, Six Wnt genes (LvWnt4, LvWnt5, LvWnt6, LvWnt7, LvWnt10 and LvWnt16) were obtained in L. vannamei. The complete cDNAs open reading frames (ORF) of LvWnt4, LvWnt5, LvWnt6, LvWnt7, LvWnt10 and LvWnt16 were 1077 bp, 1107 bp, 1350 bp, 1047 bp, 1509 bp and 1158 bp (GenBank accession no. KU169896, KU169897, KU169898, KU169899, KU169900 and KU169901), encoding 358, 368, 449, 348, 502 and 385 amino acid (aa) residues respectively. All the six members of LvWnts contain a Wnt1 domain, which is considered as an important feature of Wnt gene family. ClustalW analysis with amino acid sequences revealed that the proportion of identity with other species was more than 48% for all the LvWnts except LvWnt10 (36-41%). The phylogenetic relationship analysis illustrated that different subtype of Wnts formed their own separate branches and were placed in branch of invertebrates respectively with strong bootstrap support. The constitutive expressions of LvWnts were confirmed by RT-PCR in all the examined five developmental stages and eleven tissues of L. vannamei with different express patterns. LvWnt4, LvWnt5 and LvWnt10 were expressed highest in nerve while LvWnt6, LvWnt7 and LvWnt16 were expressed highest in intestine, stomach and gill, respectively. In addition, all the LvWnts were regulated by white spot syndrome virus (WSSV) challenges at different levels in hepatopancreas, gill and hemocytes, suggesting that Wnt genes may play a role in the defense against pathogenic virus infection in innate immune of L. vannamei. PMID:27153750

  17. Wnt/β-Catenin and noncanonical Wnt signaling interact in tissue evagination in the simple eumetazoan Hydra

    PubMed Central

    Philipp, Isabelle; Aufschnaiter, Roland; Özbek, Suat; Pontasch, Stefanie; Jenewein, Marcell; Watanabe, Hiroshi; Rentzsch, Fabian; Holstein, Thomas W.; Hobmayer, Bert

    2009-01-01

    In and evaginations of 2D cell sheets are major shape generating processes in animal development. They result from directed movement and intercalation of polarized cells associated with cell shape changes. Work on several bilaterian model organisms has emphasized the role of noncanonical Wnt signaling in cell polarization and movement. However, the molecular processes responsible for generating tissue and body shape in ancestral, prebilaterian animals are unknown. We show that noncanonical Wnt signaling acts in mass tissue movements during bud and tentacle evagination and regeneration in the cnidarian polyp Hydra. The wnt5, wnt8, frizzled2 (fz2), and dishevelled-expressing cell clusters define the positions, where bud and tentacle evaginations are initiated; wnt8, fz2, and dishevelled remain up-regulated in those epithelial cells, undergoing cell shape changes during the entire evagination process. Downstream of wnt and dsh expression, JNK activity is required for the evagination process. Multiple ectopic wnt5, wnt8, fz2, and dishevelled-expressing centers and the subsequent evagination of ectopic tentacles are induced throughout the body column by activation of Wnt/β-Catenin signaling. Our results indicate that integration of axial patterning and tissue morphogenesis by the coordinated action of canonical and noncanonical Wnt pathways was crucial for the evolution of eumetazoan body plans. PMID:19237582

  18. The Wnt pathway: emerging anticancer strategies.

    PubMed

    Gupta, Aman; Verma, Anukriti; Mishra, Ashutosh K; Wadhwa, Gulshan; Sharma, Sanjeev K; Jain, Chakresh K

    2013-05-01

    The canonical Wnt cascade has emerged as a critical regulator of cancer cells. Activation of the Wnt signaling pathway has also been associated with stem cell, thus raising the possibility of its role in embryogenesis and in the proliferation of malignant cancer cells. Wnt pathway has been reported to be involved in normal physiological processes in adult animals and integrally associated with cancer cell growth and maintenance, thus has been harnessed to devise strategies for anticancer therapy. The presence or absence of some members in this pathway, such as β-catenin, Axin or APC, has been found to involve in different types of tumors in human beings. Dysregulation of the canonical Wnt/β-catenin signaling pathway, mostly by inactivating mutations of the APC tumor suppressor, or oncogenic mutations of β-catenin, has been implicated in colorectal tumorigenesis. Further, elevated levels of β-catenin protein, a hallmark of activated canonical Wnt pathway, have been significantly observed in common forms of human malignancies, indicating that activation of the Wnt pathway may play an important role in tumor development and hence could be a crucial consideration for drug development. The paper discusses the potential therapeutic and diagnostic strategies directing on Wnt pathways on the basis of recent patents and their analysis. PMID:23432158

  19. Studies with WNT Genes and Nonsyndromic Cleft Lip and Palate

    PubMed Central

    Menezes, Renato; Letra, Ariadne; Kim, Ana H.; Kuchler, Erika C.; Day, Alicia; Tannure, Patricia N.; da Motta, Luise Gomes; Paiva, Katiucia Batista; Granjeiro, Jose M.; Vieira, Alexandre R.

    2010-01-01

    Background Clefts of the lip and/or palate (cleft lip/palate) are notable for their complex etiology. The WNT pathway regulates multiple developmental processes including craniofacial development and may play a role in cleft lip/palate and other defects of craniofacial development such as tooth agenesis. Variations in WNT genes have been recently associated with cleft lip/palate in humans. In addition, two WNT genes, Wnt3 and Wnt9B, are located in the clf1 cleft locus in mice. Methods We investigated 13 SNPs located in WNT3A, WNT5A, WNT8A, WNT11, WNT3 and WNT9B genes, for association with cleft lip/palate subphenotypes in 500 cleft cases and 500 unrelated controls. Genotyping of selected polymorphisms was carried out using Taqman assays. PLINK 1.06 software was used to test for differences in allele frequencies of each polymorphism between affected and unaffected individuals. Haplotype analysis was also performed. Results Individuals carrying variant alleles in WNT3 presented an increased risk for cleft lip/palate (P=0.0003; OR=1.61 95% C.I: 1.29 -2.02) in the population studied. Conclusion Our results continue to support a role for WNT genes in the pathogenesis of cleft lip/palate. Although much remains to be learned about the function of individual WNT genes during craniofacial development, additional studies should focus in the identification of potentially functional variants in these genes as contributors to human clefting. PMID:20890934

  20. Variation in WNT genes is associated with non-syndromic cleft lip with or without cleft palate

    PubMed Central

    Chiquet, Brett T.; Blanton, Susan H.; Burt, Amber; Ma, Deqiong; Stal, Samuel; Mulliken, John B.; Hecht, Jacqueline T.

    2008-01-01

    Non-syndromic cleft lip with or without cleft palate (NSCLP) is a common birth defect. Genetic and environmental factors have been causally implicated and studies have begun to delineate genetic contributions. The Wnt genes are involved in regulating mid-face development and upper lip fusion and are therefore strong candidates for an etiological role in NSCLP. Furthermore, the clf1 region in A/WyN clefting susceptible mice contains the Wnt3 and Wnt9B genes. To assess the role of the Wnt family of genes in NSCLP, we interrogated seven Wnt genes (Wnt3, Wnt3A, Wnt5A, Wnt7A, Wnt8A, Wnt9B and Wnt11) in our well-defined NSCLP dataset. Thirty-eight single nucleotide polymorphisms were genotyped in 132 multiplex NSCLP families and 354 simplex parent–child trios. In the entire dataset, single-nucleotide polymorphisms (SNPs) in three genes, Wnt3A (P = 0.006), Wnt 5A (P = 0.002) and Wnt11 (P = 0.0001) were significantly associated with NSCLP after correction for multiple testing. When stratified by ethnicity, the strongest associations were found for SNPs in Wnt3A (P = 0.0007), Wnt11 (P = 0.0012) and Wnt8A (P = 0.0013). Multiple haplotypes in Wnt genes were associated with NSCLP, and gene–gene interactions were observed between Wnt3A and both Wnt3 and Wnt5A (P = 0.004 and P = 0.039, respectively). This data suggests that alteration in Wnt gene function may perturb formation and/or fusion of the facial processes and predispose to NSCLP. PMID:18413325

  1. Variation in WNT genes is associated with non-syndromic cleft lip with or without cleft palate.

    PubMed

    Chiquet, Brett T; Blanton, Susan H; Burt, Amber; Ma, Deqiong; Stal, Samuel; Mulliken, John B; Hecht, Jacqueline T

    2008-07-15

    Non-syndromic cleft lip with or without cleft palate (NSCLP) is a common birth defect. Genetic and environmental factors have been causally implicated and studies have begun to delineate genetic contributions. The Wnt genes are involved in regulating mid-face development and upper lip fusion and are therefore strong candidates for an etiological role in NSCLP. Furthermore, the clf1 region in A/WyN clefting susceptible mice contains the Wnt3 and Wnt9B genes. To assess the role of the Wnt family of genes in NSCLP, we interrogated seven Wnt genes (Wnt3, Wnt3A, Wnt5A, Wnt7A, Wnt8A, Wnt9B and Wnt11) in our well-defined NSCLP dataset. Thirty-eight single nucleotide polymorphisms were genotyped in 132 multiplex NSCLP families and 354 simplex parent-child trios. In the entire dataset, single-nucleotide polymorphisms (SNPs) in three genes, Wnt3A (P = 0.006), Wnt 5A (P = 0.002) and Wnt11 (P = 0.0001) were significantly associated with NSCLP after correction for multiple testing. When stratified by ethnicity, the strongest associations were found for SNPs in Wnt3A (P = 0.0007), Wnt11 (P = 0.0012) and Wnt8A (P = 0.0013). Multiple haplotypes in Wnt genes were associated with NSCLP, and gene-gene interactions were observed between Wnt3A and both Wnt3 and Wnt5A (P = 0.004 and P = 0.039, respectively). This data suggests that alteration in Wnt gene function may perturb formation and/or fusion of the facial processes and predispose to NSCLP. PMID:18413325

  2. Distinct Patterns of Wnt3a and Wnt5a Signaling Pathway in the Lung from Rats with Endotoxic Shock.

    PubMed

    Hii, Hiong-Ping; Liao, Mei-Hui; Chen, Shiu-Jen; Wu, Chin-Chen; Shih, Chih-Chin

    2015-01-01

    Septic shock is a syndrome with severe hypotension and multiple organ dysfunction caused by an imbalance between pro-inflammatory and anti-inflammatory response. The most common risk factor of acute lung injury is severe sepsis. Patients with sepsis-related acute respiratory distress syndrome have higher mortality. Recent studies reveal regulatory roles of Wnt3a and Wnt5a signaling in inflammatory processes. Wnt3a signaling has been implicated in anti-inflammatory effects, whereas Wnt5a signaling has been postulated to have pro-inflammatory properties. However, the balance between Wnt3a and Wnt5a signaling pathway in the lung of rats with endotoxic shock has not been determined. Thus, we investigated the major components of Wnt3a and Wnt5a signaling pathway in the lung of endotoxemic rats. Male Wistar rats were intravenously infused with saline or lipopolysaccharide (LPS, 10 mg/kg). The changes of hemodynamics, biochemical variables, and arterial blood gas were examined during the experimental period. At 6 h after saline or LPS, animals were sacrificed, and lungs were obtained for analyzing superoxide production, water accumulation, histologic assessment, and protein expressions of Wnt3a and Wnt5a signaling pathway. Animals that received LPS showed circulatory failure, multiple organ dysfunction, metabolic acidosis, hyperventilation, lung edema, and high mortality. The lung from rats with endotoxic shock exhibited significant decreases in the levels of Wnt3a, Fzd1, Dsh1, phosphorylated GSK-3β at Ser9, and β-catenin. In contrast, the expressions of Wnt5a, Fzd5, and CaMKII were up-regulated in the lung of endotoxemic rats. These findings indicate the major components of Wnt3a and Wnt5a signaling in the lung are disturbed under endotoxic insult. PMID:26218875

  3. Wnt signaling and the control of human stem cell fate.

    PubMed

    Van Camp, J K; Beckers, S; Zegers, D; Van Hul, W

    2014-04-01

    Wnt signaling determines major developmental processes in the embryonic state and regulates maintenance, self-renewal and differentiation of adult mammalian tissue stem cells. Both β-catenin dependent and independent Wnt pathways exist, and both affect stem cell fate in developing and adult tissues. In this review, we debate the response to Wnt signal activation in embryonic stem cells and human, adult stem cells of mesenchymal, hematopoetic, intestinal, gastric, epidermal, mammary and neural lineages, and discuss the need for Wnt signaling in these cell types. Due to the vital actions of Wnt signaling in developmental and maintenance processes, deregulation of the pathway can culminate into a broad spectrum of developmental and genetic diseases, including cancer. The way in which Wnt signals can feed tumors and maintain cancer stem stells is discussed as well. Manipulation of Wnt signals both in vivo and in vitro thus carries potential for therapeutic approaches such as tissue engineering for regenerative medicine and anti-cancer treatment. Although many questions remain regarding the complete Wnt signal cell-type specific response and interplay of Wnt signaling with pathways such as BMP, Hedgehog and Notch, we hereby provide an overview of current knowledge on Wnt signaling and its control over human stem cell fate. PMID:24323281

  4. Wnt/β-Catenin Signaling Determines the Vasculogenic Fate of Postnatal Mesenchymal Stem Cells.

    PubMed

    Zhang, Zhaocheng; Nör, Felipe; Oh, Min; Cucco, Carolina; Shi, Songtao; Nör, Jacques E

    2016-06-01

    Vasculogenesis is the process of de novo blood vessel formation observed primarily during embryonic development. Emerging evidence suggest that postnatal mesenchymal stem cells are capable of recapitulating vasculogenesis when these cells are engaged in tissue regeneration. However, the mechanisms underlining the vasculogenic differentiation of mesenchymal stem cells remain unclear. Here, we used stem cells from human permanent teeth (dental pulp stem cells [DPSC]) or deciduous teeth (stem cells from human exfoliated deciduous teeth [SHED]) as models of postnatal primary human mesenchymal stem cells to understand mechanisms regulating their vasculogenic fate. GFP-tagged mesenchymal stem cells seeded in human tooth slice/scaffolds and transplanted into immunodeficient mice differentiate into human blood vessels that anastomize with the mouse vasculature. In vitro, vascular endothelial growth factor (VEGF) induced the vasculogenic differentiation of DPSC and SHED via potent activation of Wnt/β-catenin signaling. Further, activation of Wnt signaling is sufficient to induce the vasculogenic differentiation of postnatal mesenchymal stem cells, while Wnt inhibition blocked this process. Notably, β-catenin-silenced DPSC no longer differentiate into endothelial cells in vitro, and showed impaired vasculogenesis in vivo. Collectively, these data demonstrate that VEGF signaling through the canonical Wnt/β-catenin pathway defines the vasculogenic fate of postnatal mesenchymal stem cells. Stem Cells 2016;34:1576-1587. PMID:26866635

  5. The Wnt/β-catenin signaling pathway regulates the development of airway remodeling in patients with asthma

    PubMed Central

    Kwak, Hyun Jung; Park, Dong Won; Seo, Ji-Young; Moon, Ji-Yong; Kim, Tae Hyung; Sohn, Jang Won; Shin, Dong Ho; Yoon, Ho Joo; Park, Sung Soo; Kim, Sang-Heon

    2015-01-01

    Airway remodeling is a key characteristic of chronic asthma, particularly in patients with a fixed airflow limitation. The mechanisms underlying airway remodeling are poorly understood, and no therapeutic option is available. The Wnt/β-catenin signaling pathway is involved in various physiological and pathological processes, including fibrosis and smooth muscle hypertrophy. In this study, we investigated the roles of Wnt/β-catenin signaling in airway remodeling in patients with asthma. Wnt7a mRNA expression was prominent in induced sputum from patients with asthma compared with that from healthy controls. Next, we induced a chronic asthma mouse model with airway remodeling features, including subepithelial fibrosis and airway smooth muscle hyperplasia. Higher expression of Wnt family proteins and β-catenin was detected in the lung tissue of mice with chronic asthma compared to control mice. Blocking β-catenin expression with a specific siRNA attenuated airway inflammation and airway remodeling. Decreased subepithelial fibrosis and collagen accumulation in the β-catenin siRNA-treated mice was accompanied by reduced expression of transforming growth factor-β. We further showed that suppressing β-catenin in the chronic asthma model inhibited smooth muscle hyperplasia by downregulating the tenascin C/platelet-derived growth factor receptor pathway. Taken together, these findings demonstrate that the Wnt/β-catenin signaling pathway is highly expressed and regulates the development of airway remodeling in chronic asthma. PMID:26655831

  6. Integration of Shh and Wnt Signaling Pathways Regulating Hematopoiesis.

    PubMed

    Zhou, Zhigang; Wan, Liping; Wang, Chun; Zhou, Kun

    2015-12-01

    To investigate the spatial and temporal programmed expression of Shh and Wnt members during key stages of definitive hematopoiesis and the possible mechanism of Shh and Wnt signaling pathways regulating the proliferation of hematopoietic progenitor cells (HPCs). Spatial and temporal programmed gene expression of Shh and Wnt signaling during hematopoiesis corresponded with c-kit(+)lin(-) HPCs proliferation. C-kit(+)Lin(-) populations derived from aorta-gonad-mesonephros (AGM) of Balb/c mice at E10.5 with increased expression of Shh and Wnt3a demonstrated a greater potential for proliferation. Additionally, supplementation with soluble Shh N-terminal peptide promoted the proliferation of c-kit(+)Lin(-) populations by activating the Wnt signaling pathway, an effect which was inhibited by blocking Shh signaling. A specific inhibitor of wnt signaling was capable of inhibiting Shh-induced proliferation in a similar manner to shh inhibitor. Our results provide valuable information on Shh and Wnt signaling involved in hematopoiesis and highlight the importance of interaction of Shh and Wnt signaling in regulating HPCs proliferation. PMID:26378473

  7. [A role of WNT in kidney development and function].

    PubMed

    Mesar, Ines; Kes, Petar; Jukić, Nikolina Basić

    2012-10-01

    WNT 4 is a secreted glycoprotein that is critical for nephrogenesis during mesenchymal to epithelial transformation. Lately there are some experimental modles witch confirm a role of Wnt 4 during regeneration process in acute renal failure. On the other hand there are some evidence that Wnt 4 plays important role in renal fibrosis during experimental renal injury in rats that provide tubuloinerstitial fibrosis. When will Wnt 4 have a protective role or when will induce fibrosis still is not known and therefore futher studies will be necessary to gain a more precise understanding. PMID:23513417

  8. Roles of Wnt proteins in neural development and maintenance

    PubMed Central

    Patapoutian, Ardem; Reichardt, Louis F

    2013-01-01

    Many constituents of Wnt signaling pathways are expressed in the developing and mature nervous systems. Recent work has shown that Wnt signaling controls initial formation of the neural plate and many subsequent patterning decisions in the embryonic nervous system, including formation of the neural crest. Wnt signaling continues to be important at later stages of development. Wnts have been shown to regulate the anatomy of the neuronal cytoskeleton and the differentiation of synapses in the cerebellum. Wnt signaling has been demonstrated to regulate apoptosis and may participate in degenerative processes leading to cell death in the aging brain. PMID:10851180

  9. Inhibiting DNA methylation switches adipogenesis to osteoblastogenesis by activating Wnt10a

    PubMed Central

    Chen, Yii-Shyuan; Wu, Rui; Yang, Xiaosong; Kou, Shuping; MacDougald, Ormond A.; Yu, Liqing; Shi, Hang; Xue, Bingzhong

    2016-01-01

    Both adipocytes and osteoblasts share the mesodermal lineage that derives from mesenchymal stem cells. Most studies investigating the mechanisms underlying the regulation of adipogenic or osteoblastogenic development focus on transcriptional pathways; little is known about the epigenetic mechanisms in this process. We thus determined the role of 5-aza-2′-deoxycytidine (5-Aza-dC), an inhibitor of DNA methylation, in the lineage determination between adipogenesis and osteoblastogenesis. Inhibiting DNA methylation in 3T3-L1 preadipocytes by 5-Aza-dC significantly inhibited adipogenesis whereas promoted osteoblastogenesis. This dual effect of 5-Aza-dC was associated with up-regulation of Wnt10a, a key factor determining the fate of the mesenchymal lineage towards osteoblasts. Consistently, IWP-2, an inhibitor of Wnt proteins, was found to prevent the anti-adipogenic effect of 5-Aza-dC in 3T3-L1 preadipocytes and block the osteoblastogenic effect of 5-Aza-dC in ST2 mesenchymal stem cell line. Finally, the Wnt10a 5′-region is enriched with CpG sites, whose methylation levels were markedly reduced by 5-Aza-dC. Thus we conclude that inhibiting DNA methylation by 5-Aza-dC mutual-exclusively regulates the lineage determination of adipogenesis and osteoblastogenesis by demethylating Wnt10a gene and upregulating its expression. Our study defines DNA methylation as a novel mechanism underlying adipocyte and bone cell development. PMID:27136753

  10. Adipocytes WNT5a mediated dedifferentiation: a possible target in pancreatic cancer microenvironment

    PubMed Central

    Zoico, Elena; Darra, Elena; Rizzatti, Vanni; Budui, Simona; Franceschetti, Guido; Mazzali, Gloria; Rossi, Andrea P; Fantin, Francesco; Menegazzi, Marta; Cinti, Saverio; Zamboni, Mauro

    2016-01-01

    A significant epidemiological association between obesity and pancreatic ductal adenocarcinoma (PDAC) has previously been described, as well as a correlation between the degree of pancreatic steatosis, PDAC risk and prognosis. The underlying mechanisms are still not completely known. After co-culture of 3T3-L1 adipocytes and MiaPaCa2 with an in vitro transwell system we observed the appearance of fibroblast-like cells, along with a decrease in number and size of remaining adipocytes. RT-PCR analyses of 3T3-L1 adipocytes in co-culture showed a decrease in gene expression of typical markers of mature adipocytes, in parallel with an increased expression of fibroblast-specific and reprogramming genes. We found an increased WNT5a gene and protein expression early in MiaPaCa2 cells in co-culture. Additionally, EMSA of c-Jun and AP1 in 3T3-L1 demonstrated an increased activation in adipocytes after co-culture. Treatment with WNT5a neutralizing antibody completely reverted the activation of c-Jun and AP1 observed in co-cultured adipocytes. Increasing doses of recombinant SFRP-5, a competitive inhibitor for WNT5a receptor, added to the co-culture medium, were able to block the dedifferentiation of adipocytes in co-culture. These data support a WNT5a-mediated dedifferentiation process with adipocytes reprogramming toward fibroblast-like cells that might profoundly influence cancer microenvironment. PMID:26958939

  11. Adipocytes WNT5a mediated dedifferentiation: a possible target in pancreatic cancer microenvironment.

    PubMed

    Zoico, Elena; Darra, Elena; Rizzatti, Vanni; Budui, Simona; Franceschetti, Guido; Mazzali, Gloria; Rossi, Andrea P; Fantin, Francesco; Menegazzi, Marta; Cinti, Saverio; Zamboni, Mauro

    2016-04-12

    A significant epidemiological association between obesity and pancreatic ductal adenocarcinoma (PDAC) has previously been described, as well as a correlation between the degree of pancreatic steatosis, PDAC risk and prognosis. The underlying mechanisms are still not completely known.After co-culture of 3T3-L1 adipocytes and MiaPaCa2 with an in vitro transwell system we observed the appearance of fibroblast-like cells, along with a decrease in number and size of remaining adipocytes. RT-PCR analyses of 3T3-L1 adipocytes in co-culture showed a decrease in gene expression of typical markers of mature adipocytes, in parallel with an increased expression of fibroblast-specific and reprogramming genes. We found an increased WNT5a gene and protein expression early in MiaPaCa2 cells in co-culture. Additionally, EMSA of c-Jun and AP1 in 3T3-L1 demonstrated an increased activation in adipocytes after co-culture. Treatment with WNT5a neutralizing antibody completely reverted the activation of c-Jun and AP1 observed in co-cultured adipocytes.Increasing doses of recombinant SFRP-5, a competitive inhibitor for WNT5a receptor, added to the co-culture medium, were able to block the dedifferentiation of adipocytes in co-culture.These data support a WNT5a-mediated dedifferentiation process with adipocytes reprogramming toward fibroblast-like cells that might profoundly influence cancer microenvironment. PMID:26958939

  12. Can Wnt5a and Wnt non-canonical pathways really mediate adipocyte de-differentiation in a tumour microenvironment?

    PubMed

    Chirumbolo, Salvatore; Bjørklund, Geir

    2016-09-01

    Wnt5a has been recently reported as a possible triggering factor of adipocyte de-differentiation into an adipocyte-derived fibroblast in the tumour microenvironment of pancreas cancer. The Wnt/β-catenin pathway was described in processes involving de-differentiation and epithelial-mesenchymal transition but some Wnt family-belonging molecules exert an adipogenic role on adipocyte, while other ones, such as Wnt10b or Wnt3a, an anti-adipogenic role. Although this ability depends on the different tumoural microenvironments, it is intriguing to ascertain if some Wnt molecules, participating in the non-canonical pathway, may be targeted as fundamental factors able to trigger the desmoplastic reaction of peritumoural white adipose tissue. PMID:27391920

  13. Canonical Wnt Signaling Activity in Early Stages of Chick Lung Development

    PubMed Central

    daMota, Paulo; Correia-Pinto, Jorge

    2014-01-01

    Wnt signaling pathway is an essential player during vertebrate embryonic development which has been associated with several developmental processes such as gastrulation, body axis formation and morphogenesis of numerous organs, namely the lung. Wnt proteins act through specific transmembrane receptors, which activate intracellular pathways that regulate cellular processes such as cell proliferation, differentiation and death. Morphogenesis of the fetal lung depends on epithelial-mesenchymal interactions that are governed by several growth and transcription factors that regulate cell proliferation, fate, migration and differentiation. This process is controlled by different signaling pathways such as FGF, Shh and Wnt among others. Wnt signaling is recognized as a key molecular player in mammalian pulmonary development but little is known about its function in avian lung development. The present work characterizes, for the first time, the expression pattern of several Wnt signaling members, such as wnt-1, wnt-2b, wnt-3a, wnt-5a, wnt-7b, wnt-8b, wnt-9a, lrp5, lrp6, sfrp1, dkk1, β-catenin and axin2 at early stages of chick lung development. In general, their expression is similar to their mammalian counterparts. By assessing protein expression levels of active/total β-catenin and phospho-LRP6/LRP6 it is revealed that canonical Wnt signaling is active in this embryonic tissue. In vitro inhibition studies were performed in order to evaluate the function of Wnt signaling pathway in lung branching. Lung explants treated with canonical Wnt signaling inhibitors (FH535 and PK115-584) presented an impairment of secondary branch formation after 48 h of culture along with a decrease in axin2 expression levels. Branching analysis confirmed this inhibition. Wnt-FGF crosstalk assessment revealed that this interaction is preserved in the chick lung. This study demonstrates that Wnt signaling is crucial for precise chick lung branching and further supports the avian lung as a good

  14. TRIB3 mediates the expression of Wnt5a and activation of nuclear factor-κB in Porphyromonas endodontalis lipopolysaccharide-treated osteoblasts.

    PubMed

    Yu, Y; Qiu, L; Guo, J; Yang, D; Qu, L; Yu, J; Zhan, F; Xue, M; Zhong, M

    2015-08-01

    Porphyromonas endodontalis lipopolysaccharide (LPS) is considered to be correlated with the progression of bone resorption in periodontal and periapical diseases. Wnt5a has recently been implicated in inflammatory processes, but its role is unclear as a P. endodontalis LPS-induced mediator in osteoblasts. Tribbles homolog 3 (TRIB3) encodes a pseudokinase and has been linked to inflammation in certain situations. Here, we found that P. endodontalis LPS induced Wnt5a expression in a dose- and time-dependent manner and it also upregulated translocation, phosphorylation and transcriptional activity of nuclear factor-κB (NF-κB) in MC3T3-E1 cells. Bay 11-7082 blocked the translocation of NF-κB and Wnt5a expression induced by P. endodontalis LPS. Chromatin immunoprecipitation assay further established that induction of Wnt5a by P. endodontalis LPS was mediated through the NF-κB p65 subunit. Additionally, P. endodontalis LPS increased expression of TRIB3 in osteoblasts after 10 h simulated time. Overexpression of TRIB3 enhanced NF-κB phosphorylation and Wnt5a induction, whereas knockdown of TRIB3 inhibited NF-κB phosphorylation and Wnt5a expression in P. endodontalis LPS-stimulated osteoblasts. These results suggest that P. endodontalis LPS has the ability to promote the expression of Wnt5a in mouse osteoblasts, and this induction is mainly mediated by NF-κB pathway. TRIB3 seems to modulate the sustained expression of Wnt5a in osteoblasts stimulated by P. endodontalis LPS, as well as regulating NF-κB phosphorylation. PMID:25601649

  15. Block copolymers useful for enhanced oil recovery processes

    SciTech Connect

    Shu, P.

    1989-10-03

    This patent describes a method for permeability control of a subterranean formation and for enhancing oil recovery from the subterranean formation. The method comprising injecting into the subterranean formation an aqueous solution comprised of a block copolymer having the structural form of ABA triblock or (AB){sub n} multiblock wherein A is a crosslinkable polymer, B is a non-crosslinkable polymer or a polymer having low reactivity to crosslinking agents, n is a number of at least 2 and a crosslinking agent. Wherein A is selected from the group consisting of polyacrylic acid, partially hydrolyzed polyacrylamide, highly hydrolyzed polyacrylamide, partially hydrolyzed polyacrylic ester, highly hydrolyzed polyacrylic ester, polyvinyl alcohol and mixtures thereof and B is selected from the group consisting of polyacrylamide, partially hydrolyzed polyacrylamide, polyalkylene ethers, polyvinyl alcohol and polyvinylpyridine.

  16. Down-Regulation of Canonical and Up-Regulation of Non-Canonical Wnt Signalling in the Carcinogenic Process of Squamous Cell Lung Carcinoma

    PubMed Central

    Weich, Alexander; Kiss, Edit; Barko, Szilvia; Kovacs, Tamas; Avdicevic, Monika; D’Souza, Vijay K.; Rapp, Judit; Kvell, Krisztian; Jakab, Laszlo; Nyitrai, Miklos; Molnar, Tamas F.; Thickett, David R.; Laszlo, Terezia; Pongracz, Judit E.

    2013-01-01

    The majority of lung cancers (LC) belong to the non-small cell lung carcinoma (NSCLC) type. The two main NSCLC sub-types, namely adenocarcinoma (AC) and squamous cell carcinoma (SCC), respond differently to therapy. Whereas the link between cigarette smoke and lung cancer risk is well established, the relevance of non-canonical Wnt pathway up-regulation detected in SCC remains poorly understood. The present study was undertaken to investigate further the molecular events in canonical and non-canonical Wnt signalling during SCC development. A total of 20 SCC and AC samples with matched non-cancerous controls were obtained after surgery. TaqMan array analysis confirmed up-regulation of non-canonical Wnt5a and Wnt11 and identified down-regulation of canonical Wnt signalling in SCC samples. The molecular changes were tested in primary small airway epithelial cells (SAEC) and various lung cancer cell lines (e.g. A549, H157, etc). Our studies identified Wnt11 and Wnt5a as regulators of cadherin expression and potentiated relocation of β-catenin to the nucleus as an important step in decreased cellular adhesion. The presented data identifies additional details in the regulation of SCC that can aid identification of therapeutic drug targets in the future. PMID:23505429

  17. Sclerostin inhibition of Wnt-3a-induced C3H10T1/2 cell differentiation is indirect and mediated by bone morphogenetic proteins.

    PubMed

    Winkler, David G; Sutherland, May S Kung; Ojala, Ethan; Turcott, Eileen; Geoghegan, James C; Shpektor, Diana; Skonier, John E; Yu, Changpu; Latham, John A

    2005-01-28

    High bone mass diseases are caused both by activating mutations in the Wnt pathway and by loss of SOST, a bone morphogenetic protein (BMP) antagonist, leading to the activation of BMP signaling. Given the phenotypic similarity between mutations that activate these signaling pathways, it seems likely that BMPs and Wnts operate in parallel or represent components of the same pathway, modulating osteoblast differentiation. In this study, we show that in C3H10T1/2 cells, Wnt-3A and BMP-6 proteins were inducers of osteoblast differentiation, as measured by alkaline phosphatase (ALP) induction. Surprisingly, sclerostin, noggin, and human BMP receptor 1A (BMPR1A)-FC fusion proteins blocked Wnt-3A-induced ALP as well as BMP-6-induced ALP activity. Dkk-1, a Wnt inhibitor, blocked Wnt-induced ALP activity but not BMP-induced ALP activity. Early Wnt-3A signaling as measured by beta-catenin accumulation was not affected by the BMP antagonists but was blocked by Dkk-1. Wnt-3A induced the appearance of BMP-4 mRNA 12 h prior to that of ALP in C3H10T1/2 cells. We propose that sclerostin and other BMP antagonists do not block Wnt signaling directly. Sclerostin blocks Wnt-induced ALP activity by blocking the activity of BMP proteins produced by Wnt treatment. The expression of BMP proteins in this autocrine loop is essential for Wnt-3A-induced osteoblast differentiation. PMID:15545262

  18. A rare human syndrome provides genetic evidence that WNT signaling is required for reprogramming of fibroblasts to induced pluripotent stem cells

    PubMed Central

    Ross, Jason; Busch, Julia; Mintz, Ellen; Ng, Damian; Stanley, Alexandra; Brafman, David; Sutton, V. Reid; Van den Veyver, Ignatia; Willert, Karl

    2015-01-01

    SUMMARY WNT signaling promotes the reprogramming of somatic cells to an induced pluripotent state. We provide genetic evidence that WNT signaling is a requisite step during the induction of pluripotency. Fibroblasts from individuals with Focal Dermal Hypoplasia (FDH), a rare genetic syndrome caused by mutations in the essential WNT processing enzyme PORCN, fail to reprogram using standard methods. This blockade in reprogramming is overcome by ectopic WNT signaling and by PORCN overexpression, thus demonstrating that WNT signaling is essential for reprogramming. The rescue of reprogramming is critically dependent on the level of WNT signaling: steady baseline activation of the WNT pathway yields karyotypically normal iPS cells, whereas daily stimulation with Wnt3a produces FDH-iPS cells with severely abnormal karyotypes. Therefore, although WNT signaling is required for cellular reprogramming, inappropriate activation of WNT signaling induces chromosomal instability, highlighting the precarious nature of ectopic WNT activation, and its tight relationship with oncogenic transformation. PMID:25464842

  19. Parallel Block Structured Adaptive Mesh Refinement on Graphics Processing Units

    SciTech Connect

    Beckingsale, D. A.; Gaudin, W. P.; Hornung, R. D.; Gunney, B. T.; Gamblin, T.; Herdman, J. A.; Jarvis, S. A.

    2014-11-17

    Block-structured adaptive mesh refinement is a technique that can be used when solving partial differential equations to reduce the number of zones necessary to achieve the required accuracy in areas of interest. These areas (shock fronts, material interfaces, etc.) are recursively covered with finer mesh patches that are grouped into a hierarchy of refinement levels. Despite the potential for large savings in computational requirements and memory usage without a corresponding reduction in accuracy, AMR adds overhead in managing the mesh hierarchy, adding complex communication and data movement requirements to a simulation. In this paper, we describe the design and implementation of a native GPU-based AMR library, including: the classes used to manage data on a mesh patch, the routines used for transferring data between GPUs on different nodes, and the data-parallel operators developed to coarsen and refine mesh data. We validate the performance and accuracy of our implementation using three test problems and two architectures: an eight-node cluster, and over four thousand nodes of Oak Ridge National Laboratory’s Titan supercomputer. Our GPU-based AMR hydrodynamics code performs up to 4.87× faster than the CPU-based implementation, and has been scaled to over four thousand GPUs using a combination of MPI and CUDA.

  20. Metastases and Colon Cancer Tumor Growth Display Divergent Responses to Modulation of Canonical WNT Signaling

    PubMed Central

    Seth, Chandan; Ruiz i Altaba, Ariel

    2016-01-01

    Human colon cancers commonly harbor loss of function mutations in APC, a repressor of the canonical WNT pathway, thus leading to hyperactive WNT-TCF signaling. Re-establishment of Apc function in mice, engineered to conditionally repress Apc through RNAi, resolve the intestinal tumors formed due to hyperactivated Wnt-Tcf signaling. These and other results have prompted the search for specific WNT pathway antagonists as therapeutics for clinically problematic human colon cancers and associated metastases, which remain largely incurable. This widely accepted view seems at odds with a number of findings using patient-derived material: Canonical TCF targets are repressed, instead of being hyperactivated, in advanced colon cancers, and repression of TCF function does not generally result in tumor regression in xenografts. The results of a number of genetic mouse studies have also suggested that canonical WNT-TCF signaling drives metastases, but direct in vivo tests are lacking, and, surprisingly, TCF repression can enhance directly seeded metastatic growth. Here we have addressed the abilities of enhanced and blocked WNT-TCF signaling to alter tumor growth and distant metastases using xenografts of advanced human colon cancers in mice. We find that endogenous WNT-TCF signaling is mostly anti-metastatic since downregulation of TCF function with dnTCF generally enhances metastatic spread. Consistently, elevating the level of WNT signaling, by increasing the levels of WNT ligands, is not generally pro-metastatic. Our present and previous data reveal a heterogeneous response to modulating WNT-TCF signaling in human cancer cells. Nevertheless, the findings that a fraction of colon cancers tested require WNT-TCF signaling for tumor growth but all respond to repressed signaling by increasing metastases beg for a reevaluation of the goal of blocking WNT-TCF signaling to universally treat colon cancers. Our data suggest that WNT-TCF blockade may be effective in inhibiting tumor

  1. WNT signaling controls expression of pro-apoptotic BOK and BAX in intestinal cancer

    SciTech Connect

    Zeilstra, Jurrit; Joosten, Sander P.J.; Wensveen, Felix M.; Dessing, Mark C.; Schuetze, Denise M.; Eldering, Eric; Spaargaren, Marcel; Pals, Steven T.

    2011-03-04

    Research highlights: {yields} Intestinal adenomas initiated by aberrant activation of the WNT pathway displayed an increased sensitivity to apoptosis. {yields} Expression profiling of apoptosis-related genes in Apc{sup Min/+} mice revealed the differential expression of pro-apoptotic Bok and Bax. {yields} APC-mutant adenomatous crypts in FAP patients showed strongly increased BAX immunoreactivity. {yields} Blocking of {beta}-catenin/TCF-4-mediated signaling in colon cancer cells reduced the expression of BOK and BAX. -- Abstract: In a majority of cases, colorectal cancer is initiated by aberrant activation of the WNT signaling pathway. Mutation of the genes encoding the WNT signaling components adenomatous polyposis coli or {beta}-catenin causes constitutively active {beta}-catenin/TCF-mediated transcription, driving the transformation of intestinal crypts to cancer precursor lesions, called dysplastic aberrant crypt foci. Deregulated apoptosis is a hallmark of adenomatous colon tissue. However, the contribution of WNT signaling to this process is not fully understood. We addressed this role by analyzing the rate of epithelial apoptosis in aberrant crypts and adenomas of the Apc{sup Min/+} mouse model. In comparison with normal crypts and adenomas, aberrant crypts displayed a dramatically increased rate of apoptotic cell death. Expression profiling of apoptosis-related genes along the crypt-villus axis and in Apc mutant adenomas revealed increased expression of two pro-apoptotic Bcl-2 family members in intestinal adenomas, Bok and Bax. Analysis of the colon of familial adenomatous polyposis (FAP) patients along the crypt-to-surface axis, and of dysplastic crypts, corroborated this expression pattern. Disruption of {beta}-catenin/TCF-4-mediated signaling in the colorectal cancer cell line Ls174T significantly decreased BOK and BAX expression, confirming WNT-dependent regulation in intestinal epithelial cells. Our results suggest a feedback mechanism by which

  2. Synthesis, characterization, and evaluation of a novel inhibitor of WNT/β-catenin signaling pathway

    PubMed Central

    2013-01-01

    Background Wnt/β-catenin signaling is a highly conserved pathway in organism evolution and is important in many biological processes. Overactivation of Wnt/β-catenin signaling is closely related to tumor development and progression. To identify potent small molecules that can fight aberrant Wnt/β-catenin-mediated cancer, we synthesized a novel pyrazoline derivative (N-(4-hydroxybenzyl)-1,3,4-triphenyl-4,5-dihydro-1H-pyrazole-5-carboxamide, BHX) to block Wnt signaling, and determined the absolute configuration of its precursor (ethyl 1,3,4-triphenyl-4,5-dihydro-1H-pyrazole-5-carboxylate). We then evaluated the inhibitory effect of BHX in vitro and in vivo. Results Cell proliferation was assessed in three human cancer cell lines (A549, HT29, and MGC803) in the presence and absence of BHX using MTS assays. BHX effectively inhibited A549, HT29, and MGC803 cell proliferation with IC50 of 5.43 ± 1.99, 6.95 ± 0.24, and 7.62 ± 1.31 μM, respectively. BHX significantly induced apoptosis and G1 phase arrest in A549 and MGC803 cells. The β-catenin protein level was markedly reduced in A549 and MGC803 cells under BHX treatment. The inhibitory effect of BHX in vivo was investigated using a mouse xenograft model. A549 xenograft growth was suppressed by 50.96% in nude mice treated continuously with 100 mg/kg BHX for 21 d. Weight remained almost unchanged, which indicates the low toxicity of the compound. Conclusions Our data suggest that BHX is a new drug candidate for cancer treatment because of its potent effect on the Wnt/β-catenin pathway and low toxicity. PMID:24098916

  3. Low-Temperature Processable Block Copolymers That Preserve the Function of Blended Proteins.

    PubMed

    Iwasaki, Yasuhiko; Takemoto, Kyohei; Tanaka, Shinya; Taniguchi, Ikuo

    2016-07-11

    Low-temperature processable polymers have attracted increasing interest as ecological materials because of their reduced energy consumption during processing and suitability for making composites with heat-sensitive biomolecules at ambient temperature. In the current study, low-temperature processable biodegradable block copolymers were synthesized by ring-opening polymerization of l-lactide (LLA) using polyphosphoester as a macroinitiator. The polymer films could be processed under a hydraulic pressure of 35 MPa. The block copolymer films swelled in water because the polyphosphoester block was partially hydrated. Interestingly, the swelling ratio of the films changed with temperature. The pressure-induced order-to-disorder transition of the block copolymers was characterized by small-angle X-ray scattering; a crystallinity reduction in the block copolymers was observed after application of pressure. The crystallinity of the block copolymers was recovered after removing the applied pressure. The Young's modulus of the block copolymer films increased as the LLA unit content increased. Moreover, the modulus did not change after multiple processing cycles and the recyclability of the block copolymers was also confirmed. Finally, polymer films with embedded proteinase K as a model protein were prepared. The activity of catalase loaded into the polymer films was evaluated after processing at different temperatures. The activity of catalase was preserved when the polymer films were processed at room temperature but was significantly reduced after high-temperature processing. The suitability of low-temperature processable biodegradable polymers for making biofunctional composites without reducing protein activity was clarified. These materials will be useful for biomedical and therapeutic applications. PMID:27280847

  4. Wnt4 Participates in the Formation of Vertebrate Neuromuscular Junction

    PubMed Central

    Strochlic, Laure; Falk, Julien; Goillot, Evelyne; Sigoillot, Séverine; Bourgeois, Francine; Delers, Perrine; Rouvière, Jérôme; Swain, Amanda; Castellani, Valérie; Schaeffer, Laurent; Legay, Claire

    2012-01-01

    Neuromuscular junction (NMJ) formation requires the highly coordinated communication of several reciprocal signaling processes between motoneurons and their muscle targets. Identification of the early, spatially restricted cues in target recognition at the NMJ is still poorly documented, especially in mammals. Wnt signaling is one of the key pathways regulating synaptic connectivity. Here, we report that Wnt4 contributes to the formation of vertebrate NMJ in vivo. Results from a microarray screen and quantitative RT-PCR demonstrate that Wnt4 expression is regulated during muscle cell differentiation in vitro and muscle development in vivo, being highly expressed when the first synaptic contacts are formed and subsequently downregulated. Analysis of the mouse Wnt4−/− NMJ phenotype reveals profound innervation defects including motor axons overgrowing and bypassing AChR aggregates with 30% of AChR clusters being unapposed by nerve terminals. In addition, loss of Wnt4 function results in a 35% decrease of the number of prepatterned AChR clusters while Wnt4 overexpression in cultured myotubes increases the number of AChR clusters demonstrating that Wnt4 directly affects postsynaptic differentiation. In contrast, muscle structure and the localization of several synaptic proteins including acetylcholinesterase, MuSK and rapsyn are not perturbed in the Wnt4 mutant. Finally, we identify MuSK as a Wnt4 receptor. Wnt4 not only interacts with MuSK ectodomain but also mediates MuSK activation. Taken together our data reveal a new role for Wnt4 in mammalian NMJ formation that could be mediated by MuSK, a key receptor in synaptogenesis. PMID:22253844

  5. AP1- and NF-kappaB-binding sites conserved among mammalian WNT10B orthologs elucidate the TNFalpha-WNT10B signaling loop implicated in carcinogenesis and adipogenesis.

    PubMed

    Katoh, Masuko; Katoh, Masaru

    2007-04-01

    WNT signals are context-dependently transduced to canonical and non-canonical signaling cascades. We cloned and characterized wild-type human WNT10B, while another group cloned aberrant human WNT10B with Gly60Asp amino-acid substitution. Proto-oncogene WNT10B is expressed in gastric cancer, pancreatic cancer, breast cancer, esophageal cancer, and cervical cancer. Because WNT10B blocks adipocyte differentiation, coding SNP of WNT10B gene is associated with familial obesity. In 2001, we reported WNT10B upregulation by TNFalpha. Here, comparative integromics analyses on WNT10B orthologs were performed to elucidate the transcriptional mechanism of WNT10B. Chimpanzee WNT10B and cow Wnt10b genes were identified within NW_001223159.1 and AC150975.2 genome sequences, respectively, by using bioinformatics (Techint) and human intelligence (Humint). Chimpanzee WNT10B and cow Wnt10b showed 98.7% and 95.1% total-amino-acid identity with human WNT10B, respectively. N-terminal signal peptide, 24 Cys residues, two Asn-linked glycosylation sites, and Gly60 of human WNT10B were conserved among mammalian WNT10B orthologs. Transcription start site of human WNT10B gene was 106-bp upstream of NM_003394.2 RefSeq 5'-end. Number of GC di-nucleotide repeats just down-stream of WNT10B transcription start site varied among primates and human population. Comparative genomics analyses revealed that double AP1-binding sites in the 5'-flanking promoter region and NF-kappaB-binding site in intron 3 were conserved among human, chimpanzee, cow, mouse, and rat WNT10B orthologs. Because TNFalpha signaling through TNFR1 and TRADD/RIP/TRAF2 complex activates JUN kinase (JNK) and IkappaB kinase (IKK) signaling cascades, conserved AP1- and NF-kappaB-binding sites explain the mechanism of TNFalpha-induced WNT10B upregulation. TNFalpha-WNT10B signaling loop is the negative feedback mechanism of adipogenesis to prevent obesity and metabolic syndrome. On the other hand, TNFalpha-WNT10B signaling loop is

  6. Extracellular modulators of Wnt signalling.

    PubMed

    Malinauskas, Tomas; Jones, E Yvonne

    2014-12-01

    Wnt morphogens are secreted signalling proteins that play leading roles in embryogenesis and tissue homeostasis throughout life. Wnt signalling is controlled by multiple mechanisms, including posttranslational modification of Wnts, antagonist binding (to Wnts or their receptors), and regulation of the availability of Wnt receptors. Recent crystallographic, structure-guided biophysical and cell-based studies have advanced our understanding of how Wnt signalling is regulated at the cell surface. Structures include Wnt in complex with the cysteine-rich domain (CRD) of Frizzled, extracellular fragments of Wnt co-receptor LRP6, LRP6-binding antagonists Dickkopf and Sclerostin, antagonists 5T4/WAIF1 and Wnt inhibitory factor 1 (WIF-1), as well as Frizzled-ubiquitin ligases ZNRF3/RNF43 (in isolation and in complexes with Wnt signalling promoters R-spondins and LGR5). We review recent discoveries and remaining questions. PMID:25460271

  7. Stimulation of Wnt/beta-Catenin Signaling Pathway with Wnt Agonist Reduces Organ Injury after Hemorrhagic Shock

    PubMed Central

    Kuncewitch, Michael; Yang, Weng-Lang; Jacob, Asha; Khader, Adam; Giangola, Matthew; Nicastro, Jeffrey; Coppa, Gene F.; Wang, Ping

    2014-01-01

    Background Hemorrhagic shock is a leading cause of morbidity and mortality in surgery and trauma patients. Despite a large number of preclinical trials conducted to develop therapeutic strategies against hemorrhagic shock, there is still an unmet need exist for effective therapy for hemorrhage victims. Wnt/β-catenin signaling controls developmental processes and cellular regeneration owing to its central role in cell survival and proliferation. We therefore hypothesized that the activation of Wnt signaling reduces systemic injury caused by hemorrhagic shock. Methods Adult male Sprague-Dawley rats underwent hemorrhagic shock by controlled bleeding of the femoral artery to maintain a mean arterial pressure (MAP) of 30 mmHg for 90 min, followed by resuscitation with crystalloid equal to two times the shed blood volume. After resuscitation, animals were infused with Wnt agonist (5 mg/kg) or Vehicle (20% DMSO in saline). Blood and tissue samples were collected 6 h after resuscitation for analysis. Results Hemorrhagic shock increased serum levels of AST, lactate, and LDH. Treatment with Wnt agonist significantly reduced these levels by 40%, 36%, and 77%, respectively. Wnt agonist also decreased BUN and creatinine by 34% and 56%, respectively. Treatment reduced lung myeloperoxidase activity and IL-6 mRNA by 55% and 68% respectively and, significantly improved lung histology. Wnt agonist treatment increased Bcl-2 protein to Sham values and decreased cleaved caspase-3 by 46% indicating attenuation of hemorrhage-induced apoptosis in the lungs. Hemorrhage resulted in significant reductions of β-catenin protein levels in the lungs as well as down-regulation of a Wnt target gene, Cyclin-D1, while Wnt agonist treatment preserved these levels. Conclusions The administration of Wnt agonist attenuated hemorrhage-induced organ injury, inflammation and apoptosis. This was correlated with preservation of the Wnt signaling pathway. Thus, Wnt/β-catenin activation could be protective

  8. The role of Ppt/Wnt5 in regulating cell shape and movement during zebrafish gastrulation.

    PubMed

    Kilian, Beate; Mansukoski, Hannu; Barbosa, Filipa Carreira; Ulrich, Florian; Tada, Masazumi; Heisenberg, Carl Philipp

    2003-04-01

    Wnt genes play important roles in regulating patterning and morphogenesis during vertebrate gastrulation. In zebrafish, slb/wnt11 is required for convergence and extension movements, but not cell fate specification during gastrulation. To determine if other Wnt genes functionally interact with slb/wnt11, we analysed the role of ppt/wnt5 during zebrafish gastrulation. ppt/wnt5 is maternally provided and zygotically expressed at all stages during gastrulation. The analysis of ppt mutant embryos reveals that Ppt/Wnt5 regulates cell elongation and convergent extension movements in posterior regions of the gastrula, while its function in more anterior regions is largely redundant to that of Slb/Wnt11. Frizzled-2 functions downstream of ppt/wnt5, indicating that it might act as a receptor for Ppt/Wnt5 in this process. The characterisation of the role of Ppt/Wnt5 provides insight into the functional diversity of Wnt genes in regulating vertebrate gastrulation movements. PMID:12676324

  9. Downregulation of human Wnt3 in gastric cancer suppresses cell proliferation and induces apoptosis

    PubMed Central

    Wang, Hai-Sheng; Nie, Xiaobo; Wu, Rui-Bing; Yuan, Hong-Wei; Ma, Yue-Hong; Liu, Xiu-Lan; Zhang, Jian-Yu; Deng, Xiu-Ling; Na, Qin; Jin, Hai-Yan; Bian, Yan-Chao; Gao, Yu-Min; Wang, Yan-Dong; Chen, Wei-Dong

    2016-01-01

    Aberrant activation of Wnt/β-catenin signaling pathways is closely involved in the occurrence and progression of several types of human malignancies. However, as a fundamental component in this cascade, Wnt3 has not been well understood for the expression level and pathogenic mechanism in gastric carcinogenesis. Here, this research was undertaken to elucidate the important role of Wnt3 in gastric cancer. Wnt3 expression in gastric carcinomas and their respective normal tissues was examined by immunoblotting and immunohistochemistry. In all cases, Wnt3 expression was significantly elevated in gastric carcinomas compared with normal tissues. Knocking down Wnt3 in MGC-803 gastric cancer cells by small interfering RNAs transfection led to an obvious decrease in both transcript and protein levels. Silence of Wnt3 expression in gastric cancer cells inhibited the expression of β-catenin and cyclin D1 genes in Wnt/β-catenin pathway, significantly blocked cellular proliferation, delayed cell cycle, suppressed cell invasion and metastasis, accompanied by a higher apoptosis rate. Together, we conclude that upregulation of Wnt3 plays a crucial role in gastric tumorigenesis by inducing proliferation, migration, and invasion and inhibiting apoptosis of cancer cells, and Wnt3 might be a potential target for the treatment of gastric cancer. PMID:27390525

  10. Casein kinase iepsilon in the wnt pathway: regulation of beta-catenin function.

    PubMed

    Sakanaka, C; Leong, P; Xu, L; Harrison, S D; Williams, L T

    1999-10-26

    Wnt and its intracellular effector beta-catenin regulate developmental and oncogenic processes. Using expression cloning to identify novel components of the Wnt pathway, we isolated casein kinase Iepsilon (CKIepsilon). CKIepsilon mimicked Wnt in inducing a secondary axis in Xenopus, stabilizing beta-catenin, and stimulating gene transcription in cells. Inhibition of endogenous CKIepsilon by kinase-defective CKIepsilon or CKIepsilon antisense-oligonucleotides attenuated Wnt signaling. CKIepsilon was in a complex with axin and other downstream components of the Wnt pathway, including Dishevelled. CKIepsilon appears to be a positive regulator of the pathway and a link between upstream signals and the complexes that regulate beta-catenin. PMID:10535959

  11. Noncanonical Wnt signaling promotes osteoclast differentiation and is facilitated by the human immunodeficiency virus protease inhibitor ritonavir

    SciTech Connect

    Santiago, Francisco; Oguma, Junya; Brown, Anthony M.C.; Laurence, Jeffrey

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer First demonstration of direct role for noncanonical Wnt in osteoclast differentiation. Black-Right-Pointing-Pointer Demonstration of Ryk as a Wnt5a/b receptor in inhibition of canonical Wnt signaling. Black-Right-Pointing-Pointer Modulation of noncanonical Wnt signaling by a clinically important drug, ritonavir. Black-Right-Pointing-Pointer Establishes a mechanism for an important clinical problem: HIV-associated bone loss. -- Abstract: Wnt proteins that signal via the canonical Wnt/{beta}-catenin pathway directly regulate osteoblast differentiation. In contrast, most studies of Wnt-related effects on osteoclasts involve indirect changes. While investigating bone mineral density loss in the setting of human immunodeficiency virus (HIV) infection and its treatment with the protease inhibitor ritonavir (RTV), we observed that RTV decreased nuclear localization of {beta}-catenin, critical to canonical Wnt signaling, in primary human and murine osteoclast precursors. This occurred in parallel with upregulation of Wnt5a and Wnt5b transcripts. These Wnts typically stimulate noncanonical Wnt signaling, and this can antagonize the canonical Wnt pathway in many cell types, dependent upon Wnt receptor usage. We now document RTV-mediated upregulation of Wnt5a/b protein in osteoclast precursors. Recombinant Wnt5b and retrovirus-mediated expression of Wnt5a enhanced osteoclast differentiation from human and murine monocytic precursors, processes facilitated by RTV. In contrast, canonical Wnt signaling mediated by Wnt3a suppressed osteoclastogenesis. Both RTV and Wnt5b inhibited canonical, {beta}-catenin/T cell factor-based Wnt reporter activation in osteoclast precursors. RTV- and Wnt5-induced osteoclast differentiation were dependent upon the receptor-like tyrosine kinase Ryk, suggesting that Ryk may act as a Wnt5a/b receptor in this context. This is the first demonstration of a direct role for Wnt signaling pathways and Ryk in

  12. Osteoclast TGF-β Receptor Signaling Induces Wnt1 Secretion and Couples Bone Resorption to Bone Formation

    PubMed Central

    Weivoda, Megan M; Ruan, Ming; Pederson, Larry; Hachfeld, Christine; Davey, Rachel A; Zajac, Jeffrey D; Westendorf, Jennifer J; Khosla, Sundeep; Oursler, Merry Jo

    2016-01-01

    Osteoblast-mediated bone formation is coupled to osteoclast-mediated bone resorption. These processes become uncoupled with age, leading to increased risk for debilitating fractures. Therefore, understanding how osteoblasts are recruited to sites of resorption is vital to treating age-related bone loss. Osteoclasts release and activate TGF-β from the bone matrix. Here we show that osteoclastspecific inhibition of TGF-β receptor signaling in mice results in osteopenia due to reduced osteoblast numbers with no significant impact on osteoclast numbers or activity. TGF-β induced osteoclast expression of Wnt1, a protein crucial to normal bone formation, and this response was blocked by impaired TGF-β receptor signaling. Osteoclasts in aged murine bones had lower TGF-β signaling and Wnt1 expression in vivo. Ex vivo stimulation of osteoclasts derived from young or old mouse bone marrow macrophages showed no difference in TGF-β–induced Wnt1 expression. However, young osteoclasts expressed reduced Wnt1 when cultured on aged mouse bone chips compared to young mouse bone chips, consistent with decreased skeletal TGF-β availability with age. Therefore, osteoclast responses to TGF-β are essential for coupling bone resorption to bone formation, and modulating this pathway may provide opportunities to treat age-related bone loss. PMID:26108893

  13. LIG4 mediates Wnt signalling-induced radioresistance.

    PubMed

    Jun, Sohee; Jung, Youn-Sang; Suh, Han Na; Wang, Wenqi; Kim, Moon Jong; Oh, Young Sun; Lien, Esther M; Shen, Xi; Matsumoto, Yoshihisa; McCrea, Pierre D; Li, Lei; Chen, Junjie; Park, Jae-Il

    2016-01-01

    Despite the implication of Wnt signalling in radioresistance, the underlying mechanisms are unknown. Here we find that high Wnt signalling is associated with radioresistance in colorectal cancer (CRC) cells and intestinal stem cells (ISCs). We find that LIG4, a DNA ligase in DNA double-strand break repair, is a direct target of β-catenin. Wnt signalling enhances non-homologous end-joining repair in CRC, which is mediated by LIG4 transactivated by β-catenin. During radiation-induced intestinal regeneration, LIG4 mainly expressed in the crypts is conditionally upregulated in ISCs, accompanied by Wnt/β-catenin signalling activation. Importantly, among the DNA repair genes, LIG4 is highly upregulated in human CRC cells, in correlation with β-catenin hyperactivation. Furthermore, blocking LIG4 sensitizes CRC cells to radiation. Our results reveal the molecular mechanism of Wnt signalling-induced radioresistance in CRC and ISCs, and further unveils the unexpected convergence between Wnt signalling and DNA repair pathways in tumorigenesis and tissue regeneration. PMID:27009971

  14. LIG4 mediates Wnt signalling-induced radioresistance

    PubMed Central

    Jun, Sohee; Jung, Youn-Sang; Suh, Han Na; Wang, Wenqi; Kim, Moon Jong; Oh, Young Sun; Lien, Esther M.; Shen, Xi; Matsumoto, Yoshihisa; McCrea, Pierre D.; Li, Lei; Chen, Junjie; Park, Jae-Il

    2016-01-01

    Despite the implication of Wnt signalling in radioresistance, the underlying mechanisms are unknown. Here we find that high Wnt signalling is associated with radioresistance in colorectal cancer (CRC) cells and intestinal stem cells (ISCs). We find that LIG4, a DNA ligase in DNA double-strand break repair, is a direct target of β-catenin. Wnt signalling enhances non-homologous end-joining repair in CRC, which is mediated by LIG4 transactivated by β-catenin. During radiation-induced intestinal regeneration, LIG4 mainly expressed in the crypts is conditionally upregulated in ISCs, accompanied by Wnt/β-catenin signalling activation. Importantly, among the DNA repair genes, LIG4 is highly upregulated in human CRC cells, in correlation with β-catenin hyperactivation. Furthermore, blocking LIG4 sensitizes CRC cells to radiation. Our results reveal the molecular mechanism of Wnt signalling-induced radioresistance in CRC and ISCs, and further unveils the unexpected convergence between Wnt signalling and DNA repair pathways in tumorigenesis and tissue regeneration. PMID:27009971

  15. Wnt signaling induces epithelial differentiation during cutaneous wound healing

    PubMed Central

    Houschyar, Khosrow S; Momeni, Arash; Pyles, Malcolm N; Maan, Zeshaan N; Whittam, Alexander J; Siemers, Frank

    2015-01-01

    ABSTRACT Cutaneous wound repair in adult mammals typically does not regenerate original dermal architecture. Skin that has undergone repair following injury is not identical to intact uninjured skin. This disparity may be caused by differences in the mechanisms that regulate postnatal cutaneous wound repair compared to embryonic skin development and thus we seek a deeper understanding of the role that Wnt signaling plays in the mechanisms of skin repair in both fetal and adult wounds. The influence of secreted Wnt signaling proteins in tissue homeostasis has galvanized efforts to identify small molecules that target Wnt-mediated cellular responses. Wnt signaling is activated by wounding and participates in every subsequent stage of the healing process from the control of inflammation and programmed cell death, to the mobilization of stem cell reservoirs within the wound site. Endogenous Wnt signaling augmentation represents an attractive option to aid in the restoration of cutaneous wounds, as the complex mechanisms of the Wnt pathway have been increasingly investigated over the years. In this review, we summarize recent data elucidating the roles that Wnt signaling plays in cutaneous wound healing process. PMID:26309090

  16. Structural dynamics and inhibitor searching for Wnt-4 protein using comparative computational studies

    PubMed Central

    Hammad, Mirza A; Azam, Syed Sikander

    2015-01-01

    Wnt-4 (wingless mouse mammary tumor virus integration site-4) protein is involved in many crucial embryonic pathways regulating essential processes. Aberrant Wnt-4 activity causes various anomalies leading to gastric, colon, or breast cancer. Wnt-4 is a conserved protein in structure and sequence. All Wnt proteins contain an unusual fold comprising of a thumb (or N-terminal domain) and index finger (or C-terminal domain) bifurcated by a palm domain. The aim of this study was to identify the best inhibitors of Wnt-4 that not only interact with Wnt-4 protein but also with the covalently bound acyl group to inhibit aberrant Wnt-4 activity. A systematic computational approach was used to analyze inhibition of Wnt-4. Palmitoleic acid was docked into Wnt-4 protein, followed by ligand-based virtual screening of nearly 209,847 compounds; conformer generation of 271 compounds resulted from extensive virtual screening and comparative docking of 10,531 conformers of 271 unique compounds through GOLD (Genetic Optimization for Ligand Docking), AutoDock-Vina, and FRED (Fast Rigid Exhaustive Docking) was subsequently performed. Linux scripts was used to handle the libraries of compounds. The best compounds were selected on the basis of having maximum interactions to protein with bound palmitoleic acid. These represented lead inhibitors in further experiments. Palmitoleic acid is important for efficient Wnt activity, but aberrant Wnt-4 expression can be inhibited by designing inhibitors interacting with both protein and palmitoleic acid. PMID:25995617

  17. WNT5A Knock-Out Mouse As A New Model of Anorectal Malformation

    PubMed Central

    Tai, Cindy C.; Sala, Frederic G.; Ford, Henri R.; Wang, Kasper S.; Minoo, Parviz; Grikscheit, Tracy C.; Bellusci, Saverio

    2009-01-01

    Background Anorectal malformations (ARM) represent a variety of congenital disorders that involve abnormal termination of the anorectum. Mutations in Shh signaling and Fgf10 produce a variety of ARM phenotypes. Wnt signaling has been shown to be crucial during gastrointestinal development. We therefore hypothesized that Wnt5a may play a role in anorectal development. Methods Wild type (WT), Wnt5a+/-, and Wnt5a-/- embryos were harvested from timed pregnant mice from E15.5 to E18.5 and analyzed for anorectal phenotype. Tissues were processed for whole-mount in situ hybridization and histology. Results Wnt5a is expressed in the embryonic WT colon and rectum. Wnt5a-/- mutants exhibit multiple deformities including anorectal malformation. A fistula between the urinary and intestinal tracts can be identified as early as E15.5. By E18.5, the majority of the Wnt5a-/- mutants display a blind-ending pouch of the distal gut. Conclusions The expression pattern of Wnt5a and the ARM phenotype seen in Wnt5a-/- mutants demonstrate the critical role of Wnt5a during anorectal development. This study establishes a new model of ARM involving the Wnt5a pathway. PMID:19577771

  18. Cilia, Wnt signaling, and the cytoskeleton.

    PubMed

    May-Simera, Helen L; Kelley, Matthew W

    2012-01-01

    Primary cilia have recently been highlighted as key regulators in development and disease. This review focuses on current work demonstrating the broad role of cilia-related proteins in developmental signaling systems. Of particular consideration is the importance of the basal body region, located at the base of the cilium, in its role as a focal point for many signaling pathways and as a microtubule organizing center. As the cilium is effectively a microtubular extension of the cytoskeleton, investigating connections between the cilium and the cytoskeleton provides greater insight into signaling and cell function. Of the many signaling pathways associated with primary cilia, the most extensively studied in association with the cytoskeleton and cytoskeletal rearrangements are both canonical and non-canonical Wnt pathways. One of the key concepts currently emerging is a possible additional role for the traditionally 'cilia-related' proteins in other aspects of cellular processes. In many cases, disruption of such processes manifests at the level of the cilium. While the involvement of cilia and cilia-related proteins in signaling pathways is currently being unraveled, there is a growing body of evidence to support the notion that ciliary proteins are required not only for regulation of Wnt signaling, but also as downstream effectors of Wnt signaling. This review summarizes recent advances in our understanding of the involvement of cilia and basal body proteins in Wnt signaling pathways. PMID:23351924

  19. Involvement of Wnt, Eda and Shh at defined stages of sweat gland development.

    PubMed

    Cui, Chang-Yi; Yin, Mingzhu; Sima, Jian; Childress, Victoria; Michel, Marc; Piao, Yulan; Schlessinger, David

    2014-10-01

    To maintain body temperature, sweat glands develop from embryonic ectoderm by a poorly defined mechanism. We demonstrate a temporal cascade of regulation during mouse sweat gland formation. Sweat gland induction failed completely when canonical Wnt signaling was blocked in skin epithelium, and was accompanied by sharp downregulation of downstream Wnt, Eda and Shh pathway genes. The Wnt antagonist Dkk4 appeared to inhibit this induction: Dkk4 was sharply downregulated in β-catenin-ablated mice, indicating that it is induced by Wnt/β-catenin; however, its overexpression repressed Wnt target genes and significantly reduced gland numbers. Eda signaling succeeded Wnt. Wnt signaling was still active and nascent sweat gland pre-germs were still seen in Eda-null mice, but the pre-germs failed to develop further and the downstream Shh pathway was not activated. When Wnt and Eda were intact but Shh was ablated, germ induction and subsequent duct formation occurred normally, but the final stage of secretory coil formation failed. Thus, sweat gland development shows a relay of regulatory steps initiated by Wnt/β-catenin - itself modulated by Dkk4 - with subsequent participation of Eda and Shh pathways. PMID:25249463

  20. Proinflammatory Cytokines Regulate Cementogenic Differentiation of Periodontal Ligament Cells by Wnt/Ca(2+) Signaling Pathway.

    PubMed

    Han, Pingping; Lloyd, Tain; Chen, Zetao; Xiao, Yin

    2016-05-01

    Periodontal inflammation can inhibit cell differentiation of periodontal ligament cells (PDLCs), resulting in decreased bone/cementum regeneration ability. The Wnt signaling pathway, including canonical Wnt/β-catenin signaling and noncanonical Wnt/Ca(2+) signaling, plays essential roles in cell proliferation and differentiation during tooth development. However, little is still known whether noncanonical Wnt/Ca(2+) signaling cascade could regulate cementogenic/osteogenic differentiation capability of PDLCs within an inflammatory environment. Therefore, in this study, human PDLCs (hPDLCs) and their cementogenic differentiation potential were investigated in the presence of cytokines. The data demonstrated that both cytokines interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) inhibited cell proliferation, relative alkaline phosphatase activity, bone/cementum-related gene/protein expression, and canonical Wnt pathway-related gene/protein expression in hPDLCs. Interestingly, both cytokines upregulated the noncanonical Wnt/Ca(2+) signaling-related gene and protein expression in hPDLCs. When the Wnt/Ca(2+) pathway was blocked by Ca(2+)/calmodulin-dependent protein kinase II inhibitor KN93, even in the presence of IL-6 and TNF-α, cementogenesis could be stimulated in hPDLCs. Our data indicate that the Wnt/Ca(2+) pathway plays an inhibitory role on PDLC cementogenic differentiation in inflammatory microenvironments. Therefore, targeting the Wnt/Ca(2+) pathway may provide a novel therapeutic approach to improve periodontal regeneration for periodontal diseases. PMID:27074616

  1. Perturbation of zebrafish swimbladder development by enhancing Wnt signaling in Wif1 morphants.

    PubMed

    Yin, Ao; Korzh, Vladimir; Gong, Zhiyuan

    2012-02-01

    Wnt signaling plays critical roles in development of both tetrapod lung and fish swimbladder, which are the two evolutionary homologous organs. Our previous data reveal that down-regulation of Wnt signaling leads to defective swimbladder development. However, the effects of up-regulation of Wnt signaling on swimbladder development remain unclear. By knockdown of the Wnt protein inhibitory gene wif1, we demonstrated that up-regulation of Wnt signaling also resulted in perturbed development of the swimbladder. Specifically, the growth of epithelium and mesenchyme was greatly inhibited, the smooth muscle differentiation was abolished, and the organization of mesothelium was disturbed. Furthermore, our data reveal that it is the reduced cell proliferation, but not enhanced apoptosis, that contributes to the disturbance of swimbladder development in wif1 morphants. Blocking Wnt signaling by the Wnt antagonist IWR-1 did not affect wif1 expression in the swimbladder, but complete suppression of Hedgehog signaling in smo-/- mutants abolished wif expression, consistent with our earlier report of a negative feedback regulation of Wnt signaling in the swimbladder by the Hedgehog signaling. Our works established the importance of proper level of Wnt signaling for normal development of swimbladder in zebrafish. PMID:22008465

  2. Involvement of Wnt, Eda and Shh at defined stages of sweat gland development

    PubMed Central

    Cui, Chang-Yi; Yin, Mingzhu; Sima, Jian; Childress, Victoria; Michel, Marc; Piao, Yulan; Schlessinger, David

    2014-01-01

    To maintain body temperature, sweat glands develop from embryonic ectoderm by a poorly defined mechanism. We demonstrate a temporal cascade of regulation during mouse sweat gland formation. Sweat gland induction failed completely when canonical Wnt signaling was blocked in skin epithelium, and was accompanied by sharp downregulation of downstream Wnt, Eda and Shh pathway genes. The Wnt antagonist Dkk4 appeared to inhibit this induction: Dkk4 was sharply downregulated in β-catenin-ablated mice, indicating that it is induced by Wnt/β-catenin; however, its overexpression repressed Wnt target genes and significantly reduced gland numbers. Eda signaling succeeded Wnt. Wnt signaling was still active and nascent sweat gland pre-germs were still seen in Eda-null mice, but the pre-germs failed to develop further and the downstream Shh pathway was not activated. When Wnt and Eda were intact but Shh was ablated, germ induction and subsequent duct formation occurred normally, but the final stage of secretory coil formation failed. Thus, sweat gland development shows a relay of regulatory steps initiated by Wnt/β-catenin – itself modulated by Dkk4 – with subsequent participation of Eda and Shh pathways. PMID:25249463

  3. Analysis of the Wnt gene repertoire in an onychophoran provides new insights into the evolution of segmentation

    PubMed Central

    2014-01-01

    Background The Onychophora are a probable sister group to Arthropoda, one of the most intensively studied animal phyla from a developmental perspective. Pioneering work on the fruit fly Drosophila melanogaster and subsequent investigation of other arthropods has revealed important roles for Wnt genes during many developmental processes in these animals. Results We screened the embryonic transcriptome of the onychophoran Euperipatoides kanangrensis and found that at least 11 Wnt genes are expressed during embryogenesis. These genes represent 11 of the 13 known subfamilies of Wnt genes. Conclusions Many onychophoran Wnt genes are expressed in segment polarity gene-like patterns, suggesting a general role for these ligands during segment regionalization, as has been described in arthropods. During early stages of development, Wnt2, Wnt4, and Wnt5 are expressed in broad multiple segment-wide domains that are reminiscent of arthropod gap and Hox gene expression patterns, which suggests an early instructive role for Wnt genes during E. kanangrensis segmentation. PMID:24708787

  4. Wnt Signaling and Its Contribution to Craniofacial Tissue Homeostasis.

    PubMed

    Yin, X; Li, J; Salmon, B; Huang, L; Lim, W H; Liu, B; Hunter, D J; Ransom, R C; Singh, G; Gillette, M; Zou, S; Helms, J A

    2015-11-01

    A new field of dental medicine seeks to exploit nature's solution for repairing damaged tissues, through the process of regeneration. Most adult mammalian tissues have limited regenerative capacities, but in lower vertebrates, the molecular machinery for regeneration is an elemental part of their genetic makeup. Accumulating data suggest that the molecular pathways responsible for the regenerative capacity of teleosts, amphibians, and reptiles have fallen into disuse in mammals but that they can be "jumpstarted" by the selective activation of key molecules. The Wnt family of secreted proteins constitutes one such critical pathway: Wnt proteins rank among the most potent and ubiquitous stem cell self-renewing factors, with tremendous potential for promoting human tissue regeneration. Wnt reporter and lineage-tracing strains of mice have been employed to create molecular maps of Wnt responsiveness in the craniofacial tissues, and these patterns of Wnt signaling colocalize with stem/progenitor populations in the rodent incisor apex, the dental pulp, the alveolar bone, the periodontal ligament, the cementum, and oral mucosa. The importance of Wnt signaling in both the maintenance and healing of these craniofacial tissues is summarized, and the therapeutic potential of Wnt-based strategies to accelerate healing through activation of endogenous stem cells is highlighted. PMID:26285808

  5. Isolation and characterization of WNT8B, a novel human Wnt gene that maps to 10q24

    SciTech Connect

    Lako, Majlinda; Strachan, T.; Curtis, A.R.J.; Lindsay, S.

    1996-07-15

    Wnt genes encode intercellular signalling molecules that play important roles in key processes of embryonic development such as mesoderm induction, specification of the embryonic axis, and patterning of the central nervous system, spinal cord, and limb. Multiple such genes are known to exist in each of several species that have been investigated, and they have been classified into various groups and subgroups on the basis of high sequence homology and common expression patterns. The vertebrate Wnt8 subfamily includes genes from Xenopus, zebrafish, and chicken, but, to data, no mammalian homologues have been described. We now report cloning and characterization of a novel human member of this family that we have termed WNT8B on the basis of high sequence homology and common expression patterns. The vertebrate Wnt8 subfamily includes genes from Xenopus, zebrafish, and chicken, but, to date no mammalian homologues have been described. We now report cloning and characterization of a novel human member of this family that we have termed WNT8B on the basis of the very high sequence similarity of the inferred protein to those encoded by the Xenopus and zebrafish Wnt8b genes. PCR typing of a human monochromosomal hybrid cell panel mapped the gene to chromosome 10, and FISH mapping provided a subchromosomal location at 10q24. Northern blotting and RT-PCR assays indicated that the WNT8B gene is expressed in several human tissues during fetal and adult stages. 14 refs., 2 figs.

  6. Confusion and the Older Adult. Module A-8. Block A. Basic Knowledge of the Aging Process.

    ERIC Educational Resources Information Center

    Harvey, Dexter; Cap, Orest

    This instructional module on confusion and the older adult is one in a block of 10 modules designed to provide the human services worker who works with older adults with basic information regarding the aging process. An introduction provides an overview of the module content. A listing of general objectives follows. Three sections present…

  7. Military Curricula for Vocational & Technical Education. Metals Processing Specialist, Block VII, Classroom Course 13-8.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. National Center for Research in Vocational Education.

    These curriculum materials are the fourth section of a four-part, secondary-postsecondary-level course in metals processing. The course is one of a number of military-developed curriculum packages selected for adaptation to vocational instruction and curriculum development in a civilian setting. Block VII deals with heat treating, hardness…

  8. Nutrition and the Older Adult. Module A-9. Block A. Basic Knowledge of the Aging Process.

    ERIC Educational Resources Information Center

    Harvey, Dexter; Cap, Orest

    This instructional module on nutrition and the older adult is one in a block of 10 modules designed to provide the human services worker who works with older adults with basic information regarding the aging process. An introduction provides an overview of the module content. A listing of general objectives follows. Five sections present…

  9. Physiological Aspects of Aging. Module A-5. Block A. Basic Knowledge of the Aging Process.

    ERIC Educational Resources Information Center

    Harvey, Dexter; Cap, Orest

    This instructional module on physiological aspects of aging is one in a block of 10 modules designed to provide the human services worker who works with older adults with basic information regarding the aging process. An introduction provides an overview of the module content. A listing of general objectives follows. Nine sections present…

  10. Psychological Aspects of Aging. Module A-7. Block A. Basic Knowledge of the Aging Process.

    ERIC Educational Resources Information Center

    Harvey, Dexter; Cap, Orest

    This instructional module on psychological aspects of aging is one in a block of 10 modules designed to provide the human services worker who works with older adults with basic information regarding the aging process. An introduction provides an overview of the module content. A listing of general objectives follows. Six sections present…

  11. Aberrant Wnt Signaling in Leukemia.

    PubMed

    Staal, Frank J T; Famili, Farbod; Garcia Perez, Laura; Pike-Overzet, Karin

    2016-01-01

    The Wnt signaling pathway is essential in the development and homeostasis of blood and immune cells, but its exact role is still controversial and is the subject of intense research. The malignant counterpart of normal hematopoietic cells, leukemic (stem) cells, have hijacked the Wnt pathway for their self-renewal and proliferation. Here we review the multiple ways dysregulated Wnt signaling can contribute to leukemogenesis, both cell autonomously as well as by changes in the microenvironment. PMID:27571104

  12. Newly Constructed Network Models of Different WNT Signaling Cascades Applied to Breast Cancer Expression Data

    PubMed Central

    Kramer, Frank; Pukrop, Tobias; Beißbarth, Tim; Bleckmann, Annalen

    2015-01-01

    Introduction WNT signaling is a complex process comprising multiple pathways: the canonical β-catenin-dependent pathway and several alternative non-canonical pathways that act in a β-catenin-independent manner. Representing these intricate signaling mechanisms through bioinformatic approaches is challenging. Nevertheless, a simplified but reliable bioinformatic WNT pathway model is needed, which can be further utilized to decipher specific WNT activation states within e.g. high-throughput data. Results In order to build such a model, we collected, parsed, and curated available WNT signaling knowledge from different pathway databases. The data were assembled to construct computationally suitable models of different WNT signaling cascades in the form of directed signaling graphs. This resulted in four networks representing canonical WNT signaling, non-canonical WNT signaling, the inhibition of canonical WNT signaling and the regulation of WNT signaling pathways, respectively. Furthermore, these networks were integrated with microarray and RNA sequencing data to gain deeper insight into the underlying biology of gene expression differences between MCF-7 and MDA-MB-231 breast cancer cell lines, representing weakly and highly invasive breast carcinomas, respectively. Differential genes up-regulated in the MDA-MB-231 compared to the MCF-7 cell line were found to display enrichment in the gene set originating from the non-canonical network. Moreover, we identified and validated differentially regulated modules representing canonical and non-canonical WNT pathway components specific for the aggressive basal-like breast cancer subtype. Conclusions In conclusion, we demonstrated that these newly constructed WNT networks reliably reflect distinct WNT signaling processes. Using transcriptomic data, we shaped these networks into comprehensive modules of the genes implicated in the aggressive basal-like breast cancer subtype and demonstrated that non-canonical WNT signaling is

  13. Processes of subduction and exhumation of continental blocks in collisional orogeny

    NASA Astrophysics Data System (ADS)

    Tirel, C.; Brun, J.; Burov, E. B.; Wortel, M. J.; Lebedev, S.

    2009-12-01

    Understanding the mechanism of accretion, subduction and exhumation of rocks during orogeny is a fundamental issue for plate tectonics. Numerous models have been proposed in order to explain the tectonic events that accompany continental subduction. Here, in order to examine the different processes in a simple, self-consistent manner, the fully coupled thermo-mechanical numerical code PARAFLAM is used to perform a parametric study on the subduction mode and, particularly, on the continental subduction processes. A wide range of parameters including slab-pull magnitude, temperature and viscosity distribution, initial geometry of the subduction zone and rheology, is explored to understand the dynamics of accretion, the different processes of exhumation, the deformation of the slab and the deformation of the overriding plate. The first results of this study show the evolution of one or two small continental blocks (terranes) reaching the subduction-zone continental margin. A set of experiments demonstrates that the progressive incorporation of a continental block into the over-riding plate is governed by the dynamics of subduction. Initially, the continental block is partly or entirely subducted, which increases its buoyancy and induces its detachment from the subducting slab. By the time the emplacement of the block into the continental margin is completed, exhumation is likely to have occurred, depending mainly on the slab pull magnitude and the rheology of the continental block itself. As an example, some experiments show the continuous subduction of a continental block followed by an UHP-HP exhumation and a metamorphic-core-complex type of exhumation. These cases can be compared with the pattern of deformation observed in the Aegean-Sea domain or in the Tyrrhenian one.

  14. Evolutionary Dynamics of the wnt Gene Family: A Lophotrochozoan Perspective

    PubMed Central

    Cho, Sung-Jin; Vallès, Yvonne; Giani, Vincent C.; Seaver, Elaine C.; Weisblat, David A.

    2010-01-01

    The wnt gene family encodes a set of secreted glycoproteins involved in key developmental processes, including cell fate specification and regulation of posterior growth (Cadigan KM, Nusse R. 1997. Wnt signaling: a common theme in animal development. Genes Dev. 11:3286–3305.; Martin BL, Kimelman D. 2009. Wnt signaling and the evolution of embryonic posterior development. Curr Biol. 19:R215–R219.). As for many other gene families, evidence for expansion and/or contraction of the wnt family is available from deuterostomes (e.g., echinoderms and vertebrates [Nusse R, Varmus HE. 1992. Wnt genes. Cell. 69:1073–1087.; Schubert M, Holland LZ, Holland ND, Jacobs DK. 2000. A phylogenetic tree of the Wnt genes based on all available full-length sequences, including five from the cephalochordate amphioxus. Mol Biol Evol. 17:1896–1903.; Croce JC, Wu SY, Byrum C, Xu R, Duloquin L, Wikramanayake AH, Gache C, McClay DR. 2006. A genome-wide survey of the evolutionarily conserved Wnt pathways in the sea urchin Strongylocentrotus purpuratus. Dev Biol. 300:121–131.]) and ecdysozoans (e.g., arthropods and nematodes [Eisenmann DM. 2005. Wnt signaling. WormBook. 1–17.; Bolognesi R, Farzana L, Fischer TD, Brown SJ. 2008. Multiple Wnt genes are required for segmentation in the short-germ embryo of Tribolium castaneum. Curr Biol. 18:1624–1629.]), but little is known from the third major bilaterian group, the lophotrochozoans (e.g., mollusks and annelids [Prud'homme B, Lartillot N, Balavoine G, Adoutte A, Vervoort M. 2002. Phylogenetic analysis of the Wnt gene family. Insights from lophotrochozoan members. Curr Biol. 12:1395.]). To obtain a more comprehensive scenario of the evolutionary dynamics of this gene family, we exhaustively mined wnt gene sequences from the whole genome assemblies of a mollusk (Lottia gigantea) and two annelids (Capitella teleta and Helobdella robusta) and examined them by phylogenetic, genetic linkage, intron–exon structure, and embryonic

  15. Wnt4 coordinates directional cell migration and extension of the Müllerian duct essential for ontogenesis of the female reproductive tract

    PubMed Central

    Prunskaite-Hyyryläinen, Renata; Skovorodkin, Ilya; Xu, Qi; Miinalainen, Ilkka; Shan, Jingdong; Vainio, Seppo J.

    2016-01-01

    The Müllerian duct (MD) is the anlage of the oviduct, uterus and upper part of the vagina, the main parts of the female reproductive tract. Several wingless-type mouse mammary tumor virus (MMTV) integration site family member (Wnt) genes, including Wnt4, Wnt5a and Wnt7a, are involved in the development of MD and its derivatives, with Wnt4 particularly critical, since the MD fails to develop in its absence. We use, here, Wnt4EGFPCre-based fate mapping to demonstrate that the MD tip cells and the subsequent MD cells are derived from Wnt4+ lineage cells. Moreover, Wnt4 is required for the initiation of MD-forming cell migration. Application of anti-Wnt4 function-blocking antibodies after the initiation of MD elongation indicated that Wnt4 is necessary for the elongation as well, and consistent with this, cell culture wound-healing assays with NIH3T3 cells overexpressing Wnt4 promoted cell migration by comparison with controls. In contrast to the Wnt4 null embryos, some Wnt4monomeric cherry/monomeric cherry (Wnt4mCh/mCh) hypomorphic mice survived to adulthood and formed MD in ∼45% of cases. Nevertheless, the MD of the Wnt4mCh/mCh females had altered cell polarization and basement membrane deposition relative to the controls. Examination of the reproductive tract of the Wnt4mCh/mCh females indicated a poorly coiled oviduct, absence of the endometrial glands and an undifferentiated myometrium, and these mice were prone to develop a hydro-uterus. In conclusion, the results suggest that the Wnt4 gene encodes signals that are important for various aspects of female reproductive tract development. PMID:26721931

  16. Wnt4 coordinates directional cell migration and extension of the Müllerian duct essential for ontogenesis of the female reproductive tract.

    PubMed

    Prunskaite-Hyyryläinen, Renata; Skovorodkin, Ilya; Xu, Qi; Miinalainen, Ilkka; Shan, Jingdong; Vainio, Seppo J

    2016-03-15

    The Müllerian duct (MD) is the anlage of the oviduct, uterus and upper part of the vagina, the main parts of the female reproductive tract. Several wingless-type mouse mammary tumor virus (MMTV) integration site family member (Wnt) genes, including Wnt4, Wnt5a and Wnt7a, are involved in the development of MD and its derivatives, with Wnt4 particularly critical, since the MD fails to develop in its absence. We use, here, Wnt4(EGFPCre)-based fate mapping to demonstrate that the MD tip cells and the subsequent MD cells are derived from Wnt4+ lineage cells. Moreover, Wnt4 is required for the initiation of MD-forming cell migration. Application of anti-Wnt4 function-blocking antibodies after the initiation of MD elongation indicated that Wnt4 is necessary for the elongation as well, and consistent with this, cell culture wound-healing assays with NIH3T3 cells overexpressing Wnt4 promoted cell migration by comparison with controls. In contrast to the Wnt4 null embryos, some Wnt4(monomeric cherry/monomeric cherry) (Wnt4(mCh/mCh)) hypomorphic mice survived to adulthood and formed MD in ∼45% of cases. Nevertheless, the MD of the Wnt4(mCh/mCh) females had altered cell polarization and basement membrane deposition relative to the controls. Examination of the reproductive tract of the Wnt4(mCh/mCh) females indicated a poorly coiled oviduct, absence of the endometrial glands and an undifferentiated myometrium, and these mice were prone to develop a hydro-uterus. In conclusion, the results suggest that the Wnt4 gene encodes signals that are important for various aspects of female reproductive tract development. PMID:26721931

  17. Automated Processing of LASCO Coronal Images: Spurious Point-Source-Filtering and Missing-Blocks Correction

    NASA Astrophysics Data System (ADS)

    Pagot, E.; Lamy, P.; Llebaria, A.; Boclet, B.

    2014-04-01

    We report on automated procedures for correcting the images of the LASCO coronagraph for i) spurious quasi-point-sources such as the impacts of cosmic rays, stars, and planets, and ii) the absence of signal due to transmission errors or dropouts, which results in blocks of missing information in the images. Correcting for these undesirable artifacts is mandatory for all quantitative works on the solar corona that require data inversion and/or long series of images, for instance. The nonlinear filtering of spike noise or point-like objects is based on mathematical morphology and implements the procedure opening by morphological reconstruction. However, a simple opening filter is applied whenever the fractional area of corrupted pixels exceeds 50 % of the original image. We describe different strategies for reconstructing the missing information blocks. In general, it is possible to implement the method of averaged neighbors using the two images obtained immediately before and after the corrupted image. For the other cases, and in particular when missing blocks overlapped in three images, we developed an original procedure of weighted interpolation along radial profiles from the center of the Sun that intercept the missing block(s). This procedure is also adequate for the saturated images of bright planets (such as Venus) that bleed along the neighboring pixels. Missing blocks in polarized images may generally be reconstructed using the associated unpolarized image of the same format. But in the case of overlapping missing blocks, we implemented our procedure of weighted interpolation. All tests performed on numerous LASCO-C2 images at various periods of solar activity ( i.e. varying complexity of the structure of the corona) demonstrate the excellent performance of these new procedures, with results vastly superior to the methods implemented so far in the pipeline-processing of the LASCO images.

  18. Bisphenol-A Mediated Inhibition of Hippocampal Neurogenesis Attenuated by Curcumin via Canonical Wnt Pathway.

    PubMed

    Tiwari, Shashi Kant; Agarwal, Swati; Tripathi, Anurag; Chaturvedi, Rajnish Kumar

    2016-07-01

    Bisphenol A (BPA) is an environmental xenoestrogenic endocrine disruptor, utilized for production of consumer products, and exerts adverse effects on the developing nervous system. Recently, we found that BPA impairs the finely tuned dynamic processes of neurogenesis (generation of new neurons) in the hippocampus of the developing rat brain. Curcumin is a natural polyphenolic compound, which provides neuroprotection against various environmental neurotoxicants and in the cellular and animal models of neurodegenerative disorders. Here, we have assessed the neuroprotective efficacy of curcumin against BPA-mediated reduced neurogenesis and the underlying cellular and molecular mechanism(s). Both in vitro and in vivo studies showed that curcumin protects against BPA-induced hippocampal neurotoxicity. Curcumin protects against BPA-mediated reduced neural stem cells (NSC) proliferation and neuronal differentiation and enhanced neurodegeneration. Curcumin also enhances the expression/levels of neurogenic and the Wnt pathway genes/proteins, which were reduced due to BPA exposure in the hippocampus. Curcumin-mediated neuroprotection against BPA-induced neurotoxicity involved activation of the Wnt/β-catenin signaling pathway, which was confirmed by the use of Wnt specific activators (LiCl and GSK-3β siRNA) and inhibitor (Dkk-1). BPA-mediated increased β-catenin phosphorylation, decreased GSK-3β levels, and β-catenin nuclear translocation were significantly reversed by curcumin, leading to enhanced neurogenesis. Curcumin-induced protective effects on neurogenesis were blocked by Dkk-1 in NSC culture treated with BPA. Curcumin-mediated enhanced neurogenesis was correlated well with improved learning and memory in BPA-treated rats. Overall, our results conclude that curcumin provides neuroprotection against BPA-mediated impaired neurogenesis via activation of the Wnt/β-catenin signaling pathway. PMID:25963729

  19. Thin Isoporous Block Copolymer Membranes: It Is All about the Process.

    PubMed

    Hahn, Janina; Clodt, Juliana I; Abetz, Clarissa; Filiz, Volkan; Abetz, Volker

    2015-09-30

    The combination of the self-assembly of amphiphilic block copolymers and the nonsolvent induced phase inversion process offers an efficient way to isoporous integral-asymmetric membranes. In this context we report fast, easily upscalable and material reducing ways to thin self-assembled membranes. Therefore, we succeeded to implement a spray or dip coating step into the membrane formation process of different diblock copolymers like polystyrene-block-poly(4-vinylpyridine), poly(α-methylstyrene)-bock-poly(4-vinylpyridine), and polystyrene-block-poly(iso-propylglycidyl methacrylate). The formation of hexagonal pore structures was possible using a highly diluted one solvent system allowing the reduction of diblock copolymer consumption and therefore the production costs are minimized compared to conventional blade casting approaches. The broad applicability of the process was proven by using different flat and hollow fiber support materials. Furthermore, the membranes made by this new method showed a more than 6-fold increase in water flux compared to conventional polystyrene-block-poly(4-vinylpyridine) membranes with similar pore sizes prepared by blade casting. The membranes could be proven to be stable at transmembrane pressures of 2 bar and showed a pH responsive flux behavior over several cycles. PMID:26349610

  20. Block Copolymer Modified Epoxy Amine System for Reactive Rotational Molding: Structures, Properties and Processability

    NASA Astrophysics Data System (ADS)

    Lecocq, Eva; Nony, Fabien; Tcharkhtchi, Abbas; Gérard, Jean-François

    2011-05-01

    Poly(styrene-butadiene-methylmethacrylate) (SBM) and poly(methylmethacrylate-butyle-acrylate-methylmethacrylate) (MAM) triblock copolymers have been dissolved in liquid DGEBA epoxy resin which is subsequently polymerized by meta-xylene diamine (MXDA) or Jeffamine EDR-148. A chemorheology study of these formulations by plate-plate rheology and by thermal analysis has allowed to conclude that the addition of these copolymer blocks improve the reactive rotational moulding processability without affecting the processing time. Indeed, it prevents the pooling of the formulation at the bottom of the mould and a too rapid build up of resin viscosity of these thermosetting systems. The morphology of the cured blends examined by scanning electron microscopy (SEM) shows an increase of fracture surface area and thereby a potential increase of the toughness with the modification of epoxy system. Dynamic mechanical spectroscopy (DMA) and opalescence of final material show that the block PMMA, initially miscible, is likely to induce phase separation from the epoxy-amine matrix. Thereby, the poor compatibilisation between the toughener and the matrix has a detrimental effect on the tensile mechanical properties. The compatibilisation has to be increased to improve in synergy the processability and the final properties of these block copolymer modified formulations. First attempts could be by adapting the length and ratio of each block.

  1. A wound-induced Wnt expression program controls planarian regeneration polarity.

    PubMed

    Petersen, Christian P; Reddien, Peter W

    2009-10-01

    Regeneration requires specification of the identity of new tissues to be made. Whether this process relies only on intrinsic regulative properties of regenerating tissues or whether wound signaling provides input into tissue repatterning is not known. The head-versus-tail regeneration polarity decision in planarians, which requires Wnt signaling, provides a paradigm to study the process of tissue identity specification during regeneration. The Smed-wntP-1 gene is required for regeneration polarity and is expressed at the posterior pole of intact animals. Surprisingly, wntP-1 was expressed at both anterior- and posterior-facing wounds rapidly after wounding. wntP-1 expression was induced by all types of wounds examined, regardless of whether wounding prompted tail regeneration. Regeneration polarity was found to require new expression of wntP-1. Inhibition of the wntP-2 gene enhanced the polarity phenotype due to wntP-1 inhibition, with new expression of wntP-2 in regeneration occurring subsequent to expression of wntP-1 and localized only to posterior-facing wounds. New expression of wntP-2 required wound-induced wntP-1. Finally, wntP-1 and wntP-2 expression changes occurred even in the absence of neoblast stem cells, which are required for regeneration, suggesting that the role of these genes in polarity is independent of and instructive for tail formation. These data indicate that wound-induced input is involved in resetting the normal polarized features of the body axis during regeneration. PMID:19805089

  2. Role of the Wnt-Frizzled system in cardiac pathophysiology: a rapidly developing, poorly understood area with enormous potential

    PubMed Central

    Dawson, Kristin; Aflaki, Mona; Nattel, Stanley

    2013-01-01

    The Wnt-Frizzled (Fzd) G-protein-coupled receptor system, involving 19 distinct Wnt ligands and 10 Fzd receptors, plays key roles in the development and functioning of many organ systems. There is increasing evidence that Wnt-Fzd signalling is important in regulating cardiac function. Wnt-Fzd signalling primarily involves a canonical pathway, with dishevelled-1-dependent nuclear translocation of β-catenin that derepresses Wnt-sensitive gene transcription, but can also include non-canonical pathways via phospholipase-C/Ca2+ mobilization and dishevelled-protein activation of small GTPases. Wnt-Fzd effects vary with specific ligand/receptor interactions and associated downstream pathways. This paper reviews the biochemistry and physiology of the Wnt-Fzd complex, and presents current knowledge of Wnt signalling in cardiac remodelling processes such as hypertrophy and fibrosis, as well as disease states such as myocardial infarction (MI), heart failure and arrhythmias. Wnt signalling is activated during hypertrophy; inhibiting Wnt signalling by activating glycogen synthase kinase attenuates the hypertrophic response. Wnt signalling has complex and time-dependent actions post-MI, so that either beneficial or harmful effects might result from Wnt-directed interventions. Stem cell biology, a promising area for therapeutic intervention, is highly regulated by Wnt signalling. The Wnt system regulates fibroblast function, and is prominently altered in arrhythmogenic ventricular cardiomyopathy, a familial disease involving excess deposition of fibroadipose tissue. Wnt signalling controls connexin43 expression, thereby contributing to the regulation of cardiac electrical stability and arrhythmia generation. Although much has been learned about Wnt-Fzd signalling in hypertrophy and infarction, its role is poorly understood for a broad range of other heart disorders. Much more needs to be learned for its contributions to be fully appreciated, and to permit more effective

  3. Pharmacophore Model for Wnt/Porcupine Inhibitors and Its Use in Drug Design.

    PubMed

    Poulsen, Anders; Ho, Soo Yei; Wang, Weiling; Alam, Jenefer; Jeyaraj, Duraiswamy A; Ang, Shi Hua; Tan, Eldwin Sum Wai; Lin, Grace Ruiting; Cheong, Vivien Wei Wen; Ke, Zhiyuan; Lee, May Ann; Keller, Thomas H

    2015-07-27

    Porcupine is a component of the Wnt pathway which regulates cell proliferation, migration, stem cell self-renewal, and differentiation. The Wnt pathway has been shown to be dysregulated in a variety of cancers. Porcupine is a membrane bound O-acyltransferase that palmitoylates Wnt. Inhibiting porcupine blocks the secretion of Wnt and effectively inhibits the Wnt pathway. Using high throughput screening, we have identified a number of novel porcupine inhibitors with diverse scaffolds. The pharmacophore requirements for our porcupine inhibitors were elucidated, and a pharmacophore model is proposed. Our compounds as well as all currently published porcupine inhibitors may be fitted to this model in low energy conformations with good superimposition of the pharmacophore elements. The model also explains the stereochemical requirements of our chiral porcupine inhibitors. The pharmacophore model was successfully used for designing 3 new series of porcupine inhibitors having a tricyclic xantine, a phtalimide, or a piperidine-maleimide scaffold. PMID:26024410

  4. Wnt-Dependent Control of Cell Polarity in Cultured Cells.

    PubMed

    Runkle, Kristin B; Witze, Eric S

    2016-01-01

    The secreted ligand Wnt5a regulates cell polarity and polarized cell movement during development by signaling through the poorly defined noncanonical Wnt pathway. Cell polarity regulates most aspects of cell behavior including the organization of apical/basolateral membrane domains of epithelial cells, polarized cell divisions along a directional plane, and front rear polarity during cell migration. These characteristics of cell polarity allow coordinated cell movements required for tissue formation and organogenesis during embryonic development. Genetic model organisms have been used to identify multiple signaling pathways including Wnt5a that are required to establish cell polarity and regulate polarized cell behavior. However, the downstream signaling events that regulate these complex cellular processes are still poorly understood. The methods below describe assays to study Wnt5a-induced cell polarity in cultured cells, which may facilitate our understanding of these complex signaling pathways. PMID:27590152

  5. How targets select activation or repression in response to Wnt.

    PubMed

    Murgan, Sabrina; Bertrand, Vincent

    2015-01-01

    In metazoans, the Wnt signaling pathway plays a key role in the regulation of binary decisions during development. During this process different sets of target genes are activated in cells where the Wnt pathway is active (classic target genes) versus cells where the pathway is inactive (opposite target genes). While the mechanism of transcriptional activation is well understood for classic target genes, how opposite target genes are activated in the absence of Wnt remains poorly characterized. Here we discuss how the key transcriptional mediator of the Wnt pathway, the TCF family member POP-1, regulates opposite target genes during C. elegans development. We examine recent findings suggesting that the direction of the transcriptional output (activation or repression) can be determined by the way TCF is recruited and physically interacts with its target gene. PMID:27123368

  6. GSK-3 and Wnt Signaling in Neurogenesis and Bipolar Disorder

    PubMed Central

    Valvezan, Alexander J.; Klein, Peter S.

    2012-01-01

    The canonical Wnt signaling pathway is critical for development of the mammalian central nervous system and regulates diverse processes throughout adulthood, including adult neurogenesis. Glycogen synthase kinase-3 (GSK-3) antagonizes the canonical Wnt pathway and therefore also plays a central role in neural development and adult neurogenesis. Lithium, the first line of therapy for bipolar disorder, inhibits GSK-3, activates Wnt signaling and stimulates adult neurogenesis, which may be important for its therapeutic effects. GSK-3 also regulates other critical signaling pathways which may contribute to the therapeutic effects of lithium, including growth factor/neurotrophin signaling downstream of Akt. Here we will review the roles of GSK-3 in CNS development and adult neurogenesis, with a focus on the canonical Wnt pathway. We will also discuss the validation of GSK-3 as the relevant target of lithium and the mechanisms downstream of GSK-3 that influence mammalian behavior. PMID:22319467

  7. Silibinin inhibits Wnt/β-catenin signaling by suppressing Wnt co-receptor LRP6 expression in human prostate and breast cancer cells.

    PubMed

    Lu, Wenyan; Lin, Cuihong; King, Taj D; Chen, Honghong; Reynolds, Robert C; Li, Yonghe

    2012-12-01

    Silibinin is a natural compound isolated from milk thistle seed extracts, and has traditionally been used as a hepatoprotectant. A number of studies have also established the cancer therapeutic and chemopreventive role of silibinin in both in vitro and in vivo models. The low density lipoprotein receptor-related protein-6 (LRP6) is an essential Wnt co-receptor for the Wnt/β-catenin pathway and represents a promising target for cancer prevention and therapy. In the present study, we found that silibinin was able to repress endogenous LRP6 expression and block Wnt3A-induced LRP6 phosphorylation and Wnt/β-catenin signaling activation in HEK293 cells. Importantly, silibinin was also able to suppress endogenous LRP6 expression and phosphorylation and block Wnt/β-catenin signaling in prostate cancer PC-3 and DU-145 cells and breast cancer MDA-MB-231 and T-47D cells. Mechanistically, silibinin inhibited LRP6 promoter activity and decreased LRP6 mRNA levels in prostate and breast cancer cells. Finally, we demonstrated that silibinin displayed anticancer activity with IC(50) values comparable to those shown to suppress LRP6 expression and Wnt/β-catenin signaling activities in prostate and breast cancer cells. Our data indicate that silibinin is a novel small molecule Wnt/β-catenin signaling inhibitor by suppressing Wnt co-receptor LRP6 expression at the transcription level, and that the anti-cancer activity of silibinin is associated with its inhibitory effect on Wnt/LRP6 signaling. PMID:22820499

  8. Hear the Wnt Ror: How Melanoma Cells Adjust to Changes in Wnt

    PubMed Central

    O’Connell, Michael P.; Weeraratna, Ashani T.

    2009-01-01

    The interplay between canonical and non-canonical Wnt pathways in development and tumorigenesis is tightly regulated. In this review we will describe the yin and the yang of canonical and non-canonical Wnt signaling pathways during melanocyte development, and melanoma genesis. Canonical Wnt signaling, represented by Wnts such as Wnt1 and Wnt3A, signals via β-catenin to promote melanocyte differentiation and tumor development. Non-canonical Wnt signaling, specifically Wnt5A, regulates canonical pathways, and signals to induce melanoma metastasis. This review will focus on the role of Wnt5A during melanoma progression, and its relationship to canonical Wnt signaling. PMID:19708915

  9. CD44 regulates Wnt signaling at the level of LRP6

    PubMed Central

    Orian-Rousseau, Véronique; Schmitt, Mark

    2015-01-01

    CD44 was recently identified as a positive feedback regulator of Wnt/β-catenin signaling. This regulation occurs at the level of low-density lipoprotein receptor-related protein 6 phosphorylation and membrane targeting. These findings broaden our understanding of the Wnt pathway activation process and open new perspectives for anti-CD44 therapies in diseases associated with Wnt induction, including colorectal cancer. PMID:27308483

  10. Wnt5a promotes migration of human osteosarcoma cells by triggering a phosphatidylinositol-3 kinase/Akt signals

    PubMed Central

    2014-01-01

    Wnt5a is classified as a non-transforming Wnt family member and plays complicated roles in oncogenesis and cancer metastasis. However, Wnt5a signaling in osteosarcoma progression remains poorly defined. In this study, we found that Wnt5a stimulated the migration of human osteosarcoma cells (MG-63), with the maximal effect at 100 ng/ml, via enhancing phosphorylation of phosphatidylinositol-3 kinase (PI3K)/Akt. PI3K and Akt showed visible signs of basal phosphorylation and elevated phosphorylation at 15 min after stimulation with Wnt5a. Pharmaceutical inhibition of PI3K with LY294002 significantly blocked the Wnt5a-induced activation of Akt (p-Ser473) and decreased Wnt5a-induced cell migration. Akt siRNA remarkably inhibited Wnt5a-induced cell migration. Additionally, Wnt5a does not alter the total expression and phosphorylation of β-catenin in MG-63 cells. Taken together, we demonstrated for the first time that Wnt5a promoted osteosarcoma cell migration via the PI3K/Akt signaling pathway. These findings could provide a rationale for designing new therapy targeting osteosarcoma metastasis. PMID:24524196

  11. Wnt signaling during tooth replacement in zebrafish (Danio rerio): pitfalls and perspectives.

    PubMed

    Huysseune, Ann; Soenens, Mieke; Elderweirdt, Fien

    2014-01-01

    The canonical (β-catenin dependent) Wnt signaling pathway has emerged as a likely candidate for regulating tooth replacement in continuously renewing dentitions. So far, the involvement of canonical Wnt signaling has been experimentally demonstrated predominantly in amniotes. These studies tend to show stimulation of tooth formation by activation of the Wnt pathway, and inhibition of tooth formation when blocking the pathway. Here, we report a strong and dynamic expression of the soluble Wnt inhibitor dickkopf1 (dkk1) in developing zebrafish (Danio rerio) tooth germs, suggesting an active repression of Wnt signaling during morphogenesis and cytodifferentiation of a tooth, and derepression of Wnt signaling during start of replacement tooth formation. To further analyse the role of Wnt signaling, we used different gain-of-function approaches. These yielded disjunct results, yet none of them indicating enhanced tooth replacement. Thus, masterblind (mbl) mutants, defective in axin1, mimic overexpression of Wnt, but display a normally patterned dentition in which teeth are replaced at the appropriate times and positions. Activating the pathway with LiCl had variable outcomes, either resulting in the absence, or the delayed formation, of first-generation teeth, or yielding a regular dentition with normal replacement, but no supernumerary teeth or accelerated tooth replacement. The failure so far to influence tooth replacement in the zebrafish by perturbing Wnt signaling is discussed in the light of (i) potential technical pitfalls related to dose- or time-dependency, (ii) the complexity of the canonical Wnt pathway, and (iii) species-specific differences in the nature and activity of pathway components. Finally, we emphasize the importance of in-depth knowledge of the wild-type pattern for reliable interpretations. It is hoped that our analysis can be inspiring to critically assess and elucidate the role of Wnt signaling in tooth development in polyphyodonts. PMID

  12. Wnt signaling during tooth replacement in zebrafish (Danio rerio): pitfalls and perspectives

    PubMed Central

    Huysseune, Ann; Soenens, Mieke; Elderweirdt, Fien

    2014-01-01

    The canonical (β-catenin dependent) Wnt signaling pathway has emerged as a likely candidate for regulating tooth replacement in continuously renewing dentitions. So far, the involvement of canonical Wnt signaling has been experimentally demonstrated predominantly in amniotes. These studies tend to show stimulation of tooth formation by activation of the Wnt pathway, and inhibition of tooth formation when blocking the pathway. Here, we report a strong and dynamic expression of the soluble Wnt inhibitor dickkopf1 (dkk1) in developing zebrafish (Danio rerio) tooth germs, suggesting an active repression of Wnt signaling during morphogenesis and cytodifferentiation of a tooth, and derepression of Wnt signaling during start of replacement tooth formation. To further analyse the role of Wnt signaling, we used different gain-of-function approaches. These yielded disjunct results, yet none of them indicating enhanced tooth replacement. Thus, masterblind (mbl) mutants, defective in axin1, mimic overexpression of Wnt, but display a normally patterned dentition in which teeth are replaced at the appropriate times and positions. Activating the pathway with LiCl had variable outcomes, either resulting in the absence, or the delayed formation, of first-generation teeth, or yielding a regular dentition with normal replacement, but no supernumerary teeth or accelerated tooth replacement. The failure so far to influence tooth replacement in the zebrafish by perturbing Wnt signaling is discussed in the light of (i) potential technical pitfalls related to dose- or time-dependency, (ii) the complexity of the canonical Wnt pathway, and (iii) species-specific differences in the nature and activity of pathway components. Finally, we emphasize the importance of in-depth knowledge of the wild-type pattern for reliable interpretations. It is hoped that our analysis can be inspiring to critically assess and elucidate the role of Wnt signaling in tooth development in polyphyodonts. PMID

  13. The regulation of stem cell aging by Wnt signaling.

    PubMed

    Fujimaki, Shin; Wakabayashi, Tamami; Takemasa, Tohru; Asashima, Makoto; Kuwabara, Tomoko

    2015-12-01

    Aging is an inevitable physiological process that leads to the dysfunction of various tissues, and these changes may contribute to certain diseases, and ultimately death. Recent research has discovered biological pathways that promote aging. This review focuses on Wnt signaling, Wnt is a highly conserved secreted signaling molecule that plays an essential role in the development and function of various tissues, and is a notable factor that regulates aging. Although Wnt signaling influences aging in various tissues, its effects are particularly prominent in neuronal tissue and skeletal muscle. In neuronal tissue, neurogenesis is attenuated by the downregulation of Wnt signaling with aging. Skeletal muscle can also become weaker with aging, in a process known as sarcopenia. A notable cause of sarcopenia is the myogenic-to-fibrogenic trans-differentiation of satellite cells by excessive upregulation of Wnt signaling with aging, resulting in the impaired regenerative capacity of aged skeletal muscle. However, exercise is very useful for preventing the age-related alterations in neuronal tissue and skeletal muscle. Upregulation of Wnt signaling is implicated in the positive effects of exercise, resulting in the activation of neurogenesis in adult neuronal tissue and myogenesis in mature skeletal muscle. Although more investigations are required to thoroughly understand age-related changes and their biological mechanisms in a variety of tissues, this review proposes exercise as a useful therapy for the elderly, to prevent the negative effects of aging and maintain their quality of life. PMID:26322973

  14. Comminution and sizing processes of concrete block waste as recycled aggregates.

    PubMed

    Gomes, P C C; Ulsen, C; Pereira, F A; Quattrone, M; Angulo, S C

    2015-11-01

    Due to the environmental impact of construction and demolition waste (CDW), recycling is mandatory. It is also important that recycled concrete aggregates (RCA) are used in concrete to meet market demands. In the literature, the influence of RCAs on concrete has been investigated, but very limited studies have been conducted on how the origin of concrete waste and comminution processes influence RCA characteristics. This paper aims to investigate the influence of three different comminution and sizing processes (simple screening, crushing and grinding) on the composition, shape and porosity characteristics of RCA obtained from concrete block waste. Crushing and grinding implies a reduction of RCA porosity. However, due to the presence of coarse quartz rounded river pebbles in the original concrete block mixtures, the shape characteristics deteriorated. A large amount of powder (<0.15 mm) without detectable anhydrous cement was also generated. PMID:26168872

  15. Wnt5 is required for notochord cell intercalation in the ascidian Halocynthia roretzi

    PubMed Central

    Niwano, Tomoko; Takatori, Naohito; Kumano, Gaku; Nishida, Hiroki

    2009-01-01

    Background information. In the embryos of various animals, the body elongates after gastrulation by morphogenetic movements involving convergent extension. The Wnt/PCP (planar cell polarity) pathway plays roles in this process, particularly mediolateral polarization and intercalation of the embryonic cells. In ascidians, several factors in this pathway, including Wnt5, have been identified and found to be involved in the intercalation process of notochord cells. Results. In the present study, the role of the Wnt5 genes, Hr-Wnt5α (Halocynthia roretzi Wnt5α) and Hr-Wnt5β, in convergent extension was investigated in the ascidian H. roretzi by injecting antisense oligonucleotides and mRNAs into single precursor blastomeres of various tissues, including notochord, at the 64-cell stage. Hr-Wnt5α is expressed in developing notochord and was essential for notochord morphogenesis. Precise quantitative control of its expression level was crucial for proper cell intercalation. Overexpression of Wnt5 proteins in notochord and other tissues that surround the notochord indicated that Wnt5α plays a role within the notochord, and is unlikely to be the source of polarizing cues arising outside the notochord. Detailed mosaic analysis of the behaviour of individual notochord cells overexpressing Wnt5α indicated that a Wnt5α-manipulated cell does not affect the behaviour of neighbouring notochord cells, suggesting that Wnt5α works in a cell-autonomous manner. This is further supported by comparison of the results of Wnt5α and Dsh (Dishevelled) knockdown experiments. In addition, our results suggest that the Wnt/PCP pathway is also involved in mediolateral intercalation of cells of the ventral row of the nerve cord (floor plate) and the endodermal strand. Conclusion. The present study highlights the role of the Wnt5α signal in notochord convergent extension movements in ascidian embryos. Our results raise the novel possibility that Wnt5α functions in a cell-autonomous manner

  16. Efficient block processing of long duration biotelemetric brain data for health care monitoring.

    PubMed

    Soumya, I; Rahman, M Zia Ur; Reddy, D V Rama Koti; Lay-Ekuakille, A

    2015-03-01

    In real time clinical environment, the brain signals which doctor need to analyze are usually very long. Such a scenario can be made simple by partitioning the input signal into several blocks and applying signal conditioning. This paper presents various block based adaptive filter structures for obtaining high resolution electroencephalogram (EEG) signals, which estimate the deterministic components of the EEG signal by removing noise. To process these long duration signals, we propose Time domain Block Least Mean Square (TDBLMS) algorithm for brain signal enhancement. In order to improve filtering capability, we introduce normalization in the weight update recursion of TDBLMS, which results TD-B-normalized-least mean square (LMS). To increase accuracy and resolution in the proposed noise cancelers, we implement the time domain cancelers in frequency domain which results frequency domain TDBLMS and FD-B-Normalized-LMS. Finally, we have applied these algorithms on real EEG signals obtained from human using Emotive Epoc EEG recorder and compared their performance with the conventional LMS algorithm. The results show that the performance of the block based algorithms is superior to the LMS counter-parts in terms of signal to noise ratio, convergence rate, excess mean square error, misadjustment, and coherence. PMID:25832268

  17. Efficient block processing of long duration biotelemetric brain data for health care monitoring

    SciTech Connect

    Soumya, I.; Zia Ur Rahman, M.; Rama Koti Reddy, D. V.; Lay-Ekuakille, A.

    2015-03-15

    In real time clinical environment, the brain signals which doctor need to analyze are usually very long. Such a scenario can be made simple by partitioning the input signal into several blocks and applying signal conditioning. This paper presents various block based adaptive filter structures for obtaining high resolution electroencephalogram (EEG) signals, which estimate the deterministic components of the EEG signal by removing noise. To process these long duration signals, we propose Time domain Block Least Mean Square (TDBLMS) algorithm for brain signal enhancement. In order to improve filtering capability, we introduce normalization in the weight update recursion of TDBLMS, which results TD-B-normalized-least mean square (LMS). To increase accuracy and resolution in the proposed noise cancelers, we implement the time domain cancelers in frequency domain which results frequency domain TDBLMS and FD-B-Normalized-LMS. Finally, we have applied these algorithms on real EEG signals obtained from human using Emotive Epoc EEG recorder and compared their performance with the conventional LMS algorithm. The results show that the performance of the block based algorithms is superior to the LMS counter-parts in terms of signal to noise ratio, convergence rate, excess mean square error, misadjustment, and coherence.

  18. Activation of intracellular calcium by multiple Wnt ligands and translocation of β-catenin into the nucleus: a convergent model of Wnt/Ca2+ and Wnt/β-catenin pathways.

    PubMed

    Thrasivoulou, Christopher; Millar, Michael; Ahmed, Aamir

    2013-12-13

    Ca(2+) and β-catenin, a 92-kDa negatively charged transcription factor, transduce Wnt signaling via the non-canonical, Wnt/Ca(2+) and canonical, Wnt/β-catenin pathways independently. The nuclear envelope is a barrier to large protein entry, and this process is regulated by intracellular calcium [Ca(2+)]i and trans-nuclear potential. How β-catenin traverses the nuclear envelope is not well known. We hypothesized that Wnt/Ca(2+) and Wnt/β-catenin pathways act in a coordinated manner and that [Ca(2+)]i release facilitates β-catenin entry into the nucleus in mammalian cells. In a live assay using calcium dyes in PC3 prostate cancer cells, six Wnt peptides (3A, 4, 5A, 7A, 9B, and 10B) mobilized [Ca(2+)]i but Wnt11 did not. Based upon dwell time (range = 15-30 s) of the calcium waveform, these Wnts could be classified into three classes: short, 3A and 5A; long, 7A and 10B; and very long, 4 and 9B. Wnt-activated [Ca(2+)]i release was followed by an increase in intranuclear calcium and the depolarization of both the cell and nuclear membranes, determined by using FM4-64. In cells treated with Wnts 5A, 9B, and 10B, paradigm substrates for each Wnt class, increased [Ca(2+)]i was followed by β-catenin translocation into the nucleus in PC3, MCF7, and 253J, prostate, breast, and bladder cancer cell lines; both the increase in Wnt 5A, 9B, and 10B induced [Ca(2+)]i release and β-catenin translocation are suppressed by thapsigargin in PC3 cell line. We propose a convergent model of Wnt signaling network where Ca(2+) and β-catenin pathways may act in a coordinated, interdependent, rather than independent, manner. PMID:24158438

  19. Secreted Frizzled-related Protein 2 (sFRP2) Redirects Non-canonical Wnt Signaling from Fz7 to Ror2 during Vertebrate Gastrulation.

    PubMed

    Brinkmann, Eva-Maria; Mattes, Benjamin; Kumar, Rahul; Hagemann, Anja I H; Gradl, Dietmar; Scholpp, Steffen; Steinbeisser, Herbert; Kaufmann, Lilian T; Özbek, Suat

    2016-06-24

    Convergent extension movements during vertebrate gastrulation require a balanced activity of non-canonical Wnt signaling pathways, but the factors regulating this interplay on the molecular level are poorly characterized. Here we show that sFRP2, a member of the secreted frizzled-related protein (sFRP) family, is required for morphogenesis and papc expression during Xenopus gastrulation. We further provide evidence that sFRP2 redirects non-canonical Wnt signaling from Frizzled 7 (Fz7) to the receptor tyrosine kinase-like orphan receptor 2 (Ror2). During this process, sFRP2 promotes Ror2 signal transduction by stabilizing Wnt5a-Ror2 complexes at the membrane, whereas it inhibits Fz7 signaling, probably by blocking Fz7 receptor endocytosis. The cysteine-rich domain of sFRP2 is sufficient for Ror2 activation, and related sFRPs can substitute for this function. Notably, direct interaction of the two receptors via their cysteine-rich domains also promotes Ror2-mediated papc expression but inhibits Fz7 signaling. We propose that sFRPs can act as a molecular switch, channeling the signal input for different non-canonical Wnt pathways during vertebrate gastrulation. PMID:27129770

  20. Wnt signaling in adult intestinal stem cells and cancer.

    PubMed

    Krausova, Michaela; Korinek, Vladimir

    2014-03-01

    Signaling initiated by secreted glycoproteins of the Wnt family regulates many aspects of embryonic development and it is involved in homeostasis of adult tissues. In the gastrointestinal (GI) tract the Wnt pathway maintains the self-renewal capacity of epithelial stem cells. The stem cell attributes are conferred by mutual interactions of the stem cell with its local microenvironment, the stem cell niche. The niche ensures that the threshold of Wnt signaling in the stem cell is kept in physiological range. In addition, the Wnt pathway involves various feedback loops that balance the opposing processes of cell proliferation and differentiation. Today, we have compelling evidence that mutations causing aberrant activation of the Wnt pathway promote expansion of undifferentiated progenitors and lead to cancer. The review summarizes recent advances in characterization of adult epithelial stem cells in the gut. We mainly focus on discoveries related to molecular mechanisms regulating the output of the Wnt pathway. Moreover, we present novel experimental approaches utilized to investigate the epithelial cell signaling circuitry in vivo and in vitro. Pivotal aspects of tissue homeostasis are often deduced from studies of tumor cells; therefore, we also discuss some latest results gleaned from the deep genome sequencing studies of human carcinomas of the colon and rectum. PMID:24308963

  1. Monitoring Wnt/β-Catenin Signaling in Skin.

    PubMed

    Ku, Amy T; Miao, Qi; Nguyen, Hoang

    2016-01-01

    Wnt signaling through β-catenin plays a crucial role in skin development and homeostasis. Disruption or hyperactivation of this pathway results in skin defects and diseases (Lim and Nusse, Cold Spring Harb Perspect Biol 5(2), 2013). Monitoring Wnt signaling in skin under normal and abnormal conditions is therefore critical to understand the role of this pathway in development and homeostasis.In this chapter, we provide methods to detect Wnt/β-catenin (canonical) signaling in the skin. We present a comprehensive list of Wnt reporter mice and detail the processing of skin tissue to detect reporter genes. From this list, we focus on the three most recent lines that, according to reports, are the most sensitive in skin. Additionally, we describe a protocol to detect nuclear β-catenin, a hallmark of active Wnt signaling, although this technique should be used with caution due to its limited sensitivity. The techniques outlined below will be useful for detecting active Wnt signaling in skin. PMID:27590159

  2. WNT signaling and cartilage: of mice and men.

    PubMed

    Ma, Bin; Landman, Ellie B M; Miclea, Razvan L; Wit, Jan M; Robanus-Maandag, Els C; Post, Janine N; Karperien, Marcel

    2013-05-01

    In adult articular cartilage, the extracellular matrix is maintained by a balance between the degradation and the synthesis of matrix components. Chondrocytes that sparsely reside in the matrix and rarely proliferate are the key cellular mediators for cartilage homeostasis. There are indications for the involvement of the WNT signaling pathway in maintaining articular cartilage. Various WNTs are involved in the subsequent stages of chondrocyte differentiation during development, and deregulation of WNT signaling was observed in cartilage degeneration. Even though gene expression and protein synthesis can be activated upon injury, articular cartilage has a limited ability of self-repair and efforts to regenerate articular cartilage have so far not been successful. Because WNT signaling was found to be involved in the development and maintenance of cartilage as well as in the degeneration of cartilage, interfering with this pathway might contribute to improving cartilage regeneration. However, most of the studies on elucidating the role of WNT signaling in these processes were conducted using in vitro or in vivo animal models. Discrepancies have been found in the role of WNT signaling between chondrocytes of mouse and human origin, and extrapolation of results from mouse models to the human situation remains a challenge. Elucidation of detailed WNT signaling functions will provide knowledge to improve cartilage regeneration. PMID:23212543

  3. Improved cost-effectiveness of the block co-polymer anneal process for DSA

    NASA Astrophysics Data System (ADS)

    Pathangi, Hari; Stokhof, Maarten; Knaepen, Werner; Vaid, Varun; Mallik, Arindam; Chan, Boon Teik; Vandenbroeck, Nadia; Maes, Jan Willem; Gronheid, Roel

    2016-04-01

    This manuscript first presents a cost model to compare the cost of ownership of DSA and SAQP for a typical front end of line (FEoL) line patterning exercise. Then, we proceed to a feasibility study of using a vertical furnace to batch anneal the block co-polymer for DSA applications. We show that the defect performance of such a batch anneal process is comparable to the process of record anneal methods. This helps in increasing the cost benefit for DSA compared to the conventional multiple patterning approaches.

  4. Wnt5a can both activate and repress Wnt/β-catenin signaling during mouse embryonic development

    PubMed Central

    van Amerongen, Renée; Fuerer, Christophe; Mizutani, Makiko; Nusse, Roel

    2012-01-01

    Embryonic development is controlled by a small set of signal transduction pathways, with vastly different phenotypic outcomes depending on the time and place of their recruitment. How the same molecular machinery can elicit such specific and distinct responses, remains one of the outstanding questions in developmental biology. Part of the answer may lie in the high inherent genetic complexity of these signaling cascades, as observed for the Wnt-pathway. The mammalian genome encodes multiple Wnt proteins and receptors, each of which show dynamic and tightly controlled expression patterns in the embryo. Yet how these components interact in the context of the whole organism remains unknown. Here we report the generation of a novel, inducible transgenic mouse model that allows spatiotemporal control over the expression of Wnt5a, a protein implicated in many developmental processes and multiple Wnt-signaling responses. We show that ectopic Wnt5a expression from E10.5 onwards results in a variety of developmental defects, including loss of hair follicles and reduced bone formation in the skull. Moreover, we find that Wnt5a can have dual signaling activities during mouse embryonic development. Specifically, Wnt5a is capable of both inducing and repressing β-catenin/TCF signaling in vivo, depending on the time and site of expression and the receptors expressed by receiving cells. These experiments show for the first time that a single mammalian Wnt protein can have multiple signaling activities in vivo, thereby furthering our understanding of how signaling specificity is achieved in a complex developmental context. PMID:22771246

  5. Quality Prediction for a Fed-Batch Fermentation Process Using Multi-Block PLS

    NASA Astrophysics Data System (ADS)

    Hong, Jeong Jin; Zhang, Jie

    Quality prediction is usually required for product quality monitoring and setting up control strategy can reduce operating cost and improve production efficiency. Partial least square (PLS) regression is a popular statistical method for predictive modelling. The amount of data measured and stored in a typical industrial process is dramatically increased due to the fast development of computer and measuring system. It is hard to analyse all measured data using one matrix for its complexity. Multi-Block PLS model allows the data to be separated into sub-blocks and the sub-blocks can be analysed independently. Data from the fed-batch fermentation process is used to build models. Data is divided by different modes and different phases and model parameters are used to select variables that can be used as good predictors. The new set of data after variable selections is used to build a new model again. In most cases, new models show improved prediction performances compared with results from the conventional method.

  6. Impact of p53 loss on reversal and recurrence of conditional Wnt-induced tumorigenesis

    PubMed Central

    Gunther, Edward J.; Moody, Susan E.; Belka, George K.; Hahn, Kristina T.; Innocent, Nathalie; Dugan, Katherine D.; Cardiff, Robert D.; Chodosh, Lewis A.

    2003-01-01

    Aberrant activation of Wnt signaling is oncogenic and has been implicated in a variety of human cancers. We have developed a doxycycline-inducible Wnt1 transgenic mouse model to determine the dependence of established mammary adenocarcinomas on continued Wnt signaling. Using this model we show that targeted down-regulation of the Wnt pathway results in the rapid disappearance of essentially all Wnt-initiated invasive primary tumors as well as pulmonary metastases. Tumor regression does not require p53 and occurs even in highly aneuploid tumors. However, despite the dependence of primary mammary tumors and metastases on continued Wnt signaling and the dispensability of p53 for tumor regression, we find that a substantial fraction of tumors progress to a Wnt-independent state and that p53 suppresses this process. Specifically, loss of one p53 allele dramatically facilitates the progression of mammary tumors to a Wnt1-independent state both by impairing the regression of primary tumors following doxycycline withdrawal and by promoting the recurrence of fully regressed tumors in the absence of doxycycline. Thus, although p53 itself is dispensable for tumor regression, it nevertheless plays a critical role in the suppression of tumor recurrence. Our findings demonstrate that although even advanced stages of epithelial malignancy remain dependent upon continued Wnt signaling for maintenance and growth, loss of p53 facilitates tumor escape and the acquisition of oncogene independence. PMID:12600942

  7. Wnt5a inhibits K(+) currents in hippocampal synapses through nitric oxide production.

    PubMed

    Parodi, Jorge; Montecinos-Oliva, Carla; Varas, Rodrigo; Alfaro, Iván E; Serrano, Felipe G; Varas-Godoy, Manuel; Muñoz, Francisco J; Cerpa, Waldo; Godoy, Juan A; Inestrosa, Nibaldo C

    2015-09-01

    Hippocampal synapses play a key role in memory and learning processes by inducing long-term potentiation and depression. Wnt signaling is essential in the development and maintenance of synapses via several mechanisms. We have previously found that Wnt5a induces the production of nitric oxide (NO), which modulates NMDA receptor expression in the postsynaptic regions of hippocampal neurons. Here, we report that Wnt5a selectively inhibits a voltage-gated K(+) current (Kv current) and increases synaptic activity in hippocampal slices. Further supporting a specific role for Wnt5a, the soluble Frizzled receptor protein (sFRP-2; a functional Wnt antagonist) fully inhibits the effects of Wnt5a. We additionally show that these responses to Wnt5a are mediated by activation of a ROR2 receptor and increased NO production because they are suppressed by the shRNA-mediated knockdown of ROR2 and by 7-nitroindazole, a specific inhibitor of neuronal NOS. Together, our results show that Wnt5a increases NO production by acting on ROR2 receptors, which in turn inhibit Kv currents. These results reveal a novel mechanism by which Wnt5a may regulate the excitability of hippocampal neurons. PMID:26311509

  8. Lhx2 and Lhx9 determine neuronal differentiation and compartition in the caudal forebrain by regulating Wnt signaling.

    PubMed

    Peukert, Daniela; Weber, Sabrina; Lumsden, Andrew; Scholpp, Steffen

    2011-12-01

    Initial axial patterning of the neural tube into forebrain, midbrain, and hindbrain primordia occurs during gastrulation. After this patterning phase, further diversification within the brain is thought to proceed largely independently in the different primordia. However, mechanisms that maintain the demarcation of brain subdivisions at later stages are poorly understood. In the alar plate of the caudal forebrain there are two principal units, the thalamus and the pretectum, each of which is a developmental compartment. Here we show that proper neuronal differentiation of the thalamus requires Lhx2 and Lhx9 function. In Lhx2/Lhx9-deficient zebrafish embryos the differentiation process is blocked and the dorsally adjacent Wnt positive epithalamus expands into the thalamus. This leads to an upregulation of Wnt signaling in the caudal forebrain. Lack of Lhx2/Lhx9 function as well as increased Wnt signaling alter the expression of the thalamus specific cell adhesion factor pcdh10b and lead subsequently to a striking anterior-posterior disorganization of the caudal forebrain. We therefore suggest that after initial neural tube patterning, neurogenesis within a brain compartment influences the integrity of the neuronal progenitor pool and border formation of a neuromeric compartment. PMID:22180728

  9. Lhx2 and Lhx9 Determine Neuronal Differentiation and Compartition in the Caudal Forebrain by Regulating Wnt Signaling

    PubMed Central

    Peukert, Daniela; Weber, Sabrina; Lumsden, Andrew; Scholpp, Steffen

    2011-01-01

    Initial axial patterning of the neural tube into forebrain, midbrain, and hindbrain primordia occurs during gastrulation. After this patterning phase, further diversification within the brain is thought to proceed largely independently in the different primordia. However, mechanisms that maintain the demarcation of brain subdivisions at later stages are poorly understood. In the alar plate of the caudal forebrain there are two principal units, the thalamus and the pretectum, each of which is a developmental compartment. Here we show that proper neuronal differentiation of the thalamus requires Lhx2 and Lhx9 function. In Lhx2/Lhx9-deficient zebrafish embryos the differentiation process is blocked and the dorsally adjacent Wnt positive epithalamus expands into the thalamus. This leads to an upregulation of Wnt signaling in the caudal forebrain. Lack of Lhx2/Lhx9 function as well as increased Wnt signaling alter the expression of the thalamus specific cell adhesion factor pcdh10b and lead subsequently to a striking anterior-posterior disorganization of the caudal forebrain. We therefore suggest that after initial neural tube patterning, neurogenesis within a brain compartment influences the integrity of the neuronal progenitor pool and border formation of a neuromeric compartment. PMID:22180728

  10. Wnt Signaling in Renal Cell Carcinoma.

    PubMed

    Xu, Qi; Krause, Mirja; Samoylenko, Anatoly; Vainio, Seppo

    2016-01-01

    Renal cell carcinoma (RCC) accounts for 90% of all kidney cancers. Due to poor diagnosis, high resistance to the systemic therapies and the fact that most RCC cases occur sporadically, current research switched its focus on studying the molecular mechanisms underlying RCC. The aim is the discovery of new effective and less toxic anti-cancer drugs and novel diagnostic markers. Besides the PI3K/Akt/mTOR, HGF/Met and VHL/hypoxia cellular signaling pathways, the involvement of the Wnt/β-catenin pathway in RCC is commonly studied. Wnt signaling and its targeted genes are known to actively participate in different biological processes during embryonic development and renal cancer. Recently, studies have shown that targeting this pathway by alternating/inhibiting its intracellular signal transduction can reduce cancer cells viability and inhibit their growth. The targets and drugs identified show promising potential to serve as novel RCC therapeutics and prognostic markers. This review aims to summarize the current status quo regarding recent research on RCC focusing on the involvement of the Wnt/β-catenin pathway and how its understanding could facilitate the identification of potential therapeutic targets, new drugs and diagnostic biomarkers. PMID:27322325

  11. Wnt Signaling in Renal Cell Carcinoma

    PubMed Central

    Xu, Qi; Krause, Mirja; Samoylenko, Anatoly; Vainio, Seppo

    2016-01-01

    Renal cell carcinoma (RCC) accounts for 90% of all kidney cancers. Due to poor diagnosis, high resistance to the systemic therapies and the fact that most RCC cases occur sporadically, current research switched its focus on studying the molecular mechanisms underlying RCC. The aim is the discovery of new effective and less toxic anti-cancer drugs and novel diagnostic markers. Besides the PI3K/Akt/mTOR, HGF/Met and VHL/hypoxia cellular signaling pathways, the involvement of the Wnt/β-catenin pathway in RCC is commonly studied. Wnt signaling and its targeted genes are known to actively participate in different biological processes during embryonic development and renal cancer. Recently, studies have shown that targeting this pathway by alternating/inhibiting its intracellular signal transduction can reduce cancer cells viability and inhibit their growth. The targets and drugs identified show promising potential to serve as novel RCC therapeutics and prognostic markers. This review aims to summarize the current status quo regarding recent research on RCC focusing on the involvement of the Wnt/β-catenin pathway and how its understanding could facilitate the identification of potential therapeutic targets, new drugs and diagnostic biomarkers. PMID:27322325

  12. Analysis of backward error recovery for concurrent processes with recovery blocks

    NASA Technical Reports Server (NTRS)

    Shin, K. G.; Lee, Y. H.

    1982-01-01

    Three different methods of implementing recovery blocks (RB's). These are the asynchronous, synchronous, and the pseudo recovery point implementations. Pseudo recovery points so that unbounded rollback may be avoided while maintaining process autonomy are proposed. Probabilistic models for analyzing these three methods under standard assumptions in computer performance analysis, i.e., exponential distributions for related random variables were developed. The interval between two successive recovery lines for asynchronous RB's mean loss in computation power for the synchronized method, and additional overhead and rollback distance in case PRP's are used were estimated.

  13. Instantaneous Formation of Block Copolymer Patterns via Solvo-Thermal Casting Process

    NASA Astrophysics Data System (ADS)

    Jung, Hyun Jung; Woo, Sanghoon; Huh, June; Bang, Joona

    2015-03-01

    A self-assembly of block copolymers (BCPs) exhibits one of the most promising alternative methods for the next-generation lithography. Many semiconductor companies have explored the possibility of implementing this process in actual chip process, whereas the critical challenges such as feature size control, defect density, and long processing time need to be overcome. Regarding the BCP process, the formation of BCP patterns usually requires long processing time via thermal or solvent annealing. Herein we developed a simple processing method to promote a microphase separation of BCPs using solvo-thermal spin casting process. Spin casting has a very similar mechanism to solvent vapor annealing but its short process time prevents BCP chains from reaching equilibrium morphology. To maximize the chain mobility, we employed a high boiling point solvent and also applied the heat during spin casting. As a result, a well ordered BCP patterns were obtained within less than 5 min via solvo-thermal casting process without further additional annealing step.

  14. MOVIE: a hardware building block for software-only real-time video processing

    NASA Astrophysics Data System (ADS)

    Barzic, Ronan; Bouville, Christian; Charot, Francois; Le Fol, Gwendal; Lemonnier, Pascal; Wagner, Charles

    1996-03-01

    The goal of the MOVIE VLSI chip is to facilitate the development of software-only solutions for real time video processing applications. This chip can be seen as a building block for SIMD arrays of processing elements and its architecture has been designed so as to facilitate high level language programming. The basic architecture building block associates a sub-array of computational processors with a I/O processor. A module can be seen as a small linear, systolic-like array of processing elements, connected at each end to the I/O processor. The module can communicate with its two nearest neighbors via two communication ports. The chip architecture also includes three 16-bit video ports. One important aspect in the programming environment is the C-stolic programming language. C-stolic is a C-like language augmented with parallel constructs which allow to differentiate between the array controller variables (scalar variables) and the local variables in the array structure (systolic variables). A statement operating on systolic variables implies a simultaneous execution on all the cells of the structure. Implementation examples of MOVIE-based architectures dealing with video compression algorithms are given.

  15. Process Controlled Multiscale Morphologies in Metal-containing Block Copolymer Thin Films

    SciTech Connect

    Ramanathan, Nathan Muruganathan; Kilbey, II, S Michael; Darling, Seth B.

    2014-01-01

    Poly(styrene-block-ferrocenyldimethylsilane) (PS-b-PFS) is a metal-containing block copolymer that exhibits certain advantages as a mask for lithographic applications. These advantages include compatibility with a wide range of substrates, ease of control over domain morphologies and robust stability to etch plasma, which aid in the development of high-aspect-ratio patterns. An asymmetric cylinder-forming PS-b-PFS copolymer is subjected to different processing to manipulate the morphology of the phase-separated domains. Control of film structure and domain morphology is achieved by adjusting the film thickness, mode of annealing, and/or annealing time. Changing the process from thermal or solvent annealing to hybrid annealing (thermal and then solvent annealing in sequence) leads to the formation of mesoscale spherulitic and dendritic morphologies. In this communication, we show that reversing the order of the hybrid annealing (solvent annealing first and then thermal annealing) of relatively thick films (>100 nm) on homogeneously thick substrates develops disordered lamellar structure. Furthermore, the same processing applied on a substrate with a thin, mechanically flexible window in the center leads to the formation of sub-micron scale concentric ring patterns. Enhanced material mobility in the thick film during hybrid annealing along with dynamic rippling effects that may arise from the vibration of the thin window during spin casting are likely causes for these morphologies.

  16. Participation of WNT and β-Catenin in Physiological and Pathological Endometrial Changes: Association with Angiogenesis

    PubMed Central

    Kiewisz, Jolanta; Wasniewski, Tomasz; Kmiec, Zbigniew

    2015-01-01

    WNT proteins are involved in embryonic development, sex determination, stem cell recruitment, angiogenesis, and cancer. They take part in morphological changes in the endometrium during development, regulate processes of endometrial proliferation and differentiation. This review presents current knowledge about implication of WNT proteins and β-catenin in physiological endometrial functions as well as their involvement in uterine carcinogenesis. Influence of WNT proteins on the formation of blood vessel, taking place both under healthy and pathological conditions, is also considered. Participation of WNT proteins, β-catenin, and inhibitors and inducers of WNT signaling in the process of endometrial angiogenesis is largely unknown. Thus, confirmation of their local and systemic participation in the process of endometrial angiogenesis may in the long term help to establish new diagnostic and therapeutic approaches in conditions associated with the pathology of the female reproductive system. PMID:26366420

  17. MiR-146a-5p suppresses activation and proliferation of hepatic stellate cells in nonalcoholic fibrosing steatohepatitis through directly targeting Wnt1 and Wnt5a

    PubMed Central

    Du, Jinghua; Niu, Xuemin; Wang, Yang; Kong, Lingbo; Wang, Rongqi; Zhang, Yuguo; Zhao, Suxian; Nan, Yuemin

    2015-01-01

    Nonalcoholic fibrosing steatohepatitis is a uniform process throughout nonalcoholic fatty liver disease (NAFLD). MicroRNAs (miRNAs) have been suggested to modulate cellular processes in liver diseases. However, the functional role of miRNAs in nonalcoholic fibrosing steatohepatitis is largely unclear. In this study, we systematically analyzed the hepatic miRNAs by microarray analysis in nonalcoholic fibrosing steatohepatitis in C57BL/6J mice induced by methionine-choline deficient (MCD) diet. We identified 19 up-regulated and 18 down-regulated miRNAs in liver with fibrosis. Among these dysregulated miRNAs, miR-146a-5p was the most significant down-regulated miRNA. Luciferase activity assay confirmed that Wnt1 and Wnt5a were both the target genes of miR-146a-5p. Hepatic miR-146a-5p was down-regulated in fibrosing steatohepatitis, but its target genes Wnt1 and Wnt5a and their consequent effectors α-SMA and Col-1 were significantly up-regulated. In addition, miR-146a-5p was downregulated, whilst Wnt1 and Wnt5a were up-regulated in the activated primary hepatic stellate cells (HSCs) compared to the quiescent primary HSCs. Overexpression of miR-146a-5p in HSCs inhibited HSC activation and proliferation, which concomitant with the decreased expressions of Wnt1, Wnt5a, α-SMA and Col-1. In conclusion, miR-146a-5p suppresses activation and proliferation of HSCs in the progress of nonalcoholic fibrosing steatohepatitis through targeting Wnt1 and Wnt5a and consequent effectors α-SMA and Col-1. PMID:26537990

  18. Wnt-5a Ligand Modulates Mitochondrial Fission-Fusion in Rat Hippocampal Neurons*

    PubMed Central

    Godoy, Juan A.; Arrázola, Macarena S.; Ordenes, Daniela; Silva-Alvarez, Carmen; Braidy, Nady; Inestrosa, Nibaldo C.

    2014-01-01

    The Wnt signaling pathway plays an important role in developmental processes, including embryonic patterning, cell specification, and cell polarity. Wnt components participate in the development of the central nervous system, and growing evidence indicates that this pathway also regulates the function of the adult nervous system. In this study, we report that Wnt-5a, a noncanonical Wnt ligand, is a potent activator of mitochondrial dynamics and induces acute fission and fusion events in the mitochondria of rat hippocampal neurons. The effect of Wnt-5a was inhibited in the presence of sFRP, a Wnt scavenger. Similarly, the canonical Wnt-3a ligand had no effect on mitochondrial fission-fusion events, suggesting that this effect is specific for Wnt-5a alone. We also show that the Wnt-5a effects on mitochondrial dynamics occur with an increase in both intracellular and mitochondrial calcium (Ca2+), which was correlated with an increased phosphorylation of Drp1(Ser-616) and a decrease of Ser-637 phosphorylation, both indicators of mitochondrial dynamics. Electron microscope analysis of hippocampal tissues in the CA1 region showed an increase in the number of mitochondria present in the postsynaptic region, and this finding correlated with a change in mitochondrial morphology. We conclude that Wnt-5a/Ca2+ signaling regulates the mitochondrial fission-fusion process in hippocampal neurons, a feature that might help to further understand the role of Wnt-related pathologies, including neurodegenerative diseases associated with mitochondrial dysfunction, and represents a potentially important link between impaired metabolic function and degenerative disorders. PMID:25336659

  19. Silica Nanoparticles Target a Wnt Signal Transducer for Degradation and Impair Embryonic Development in Zebrafish.

    PubMed

    Yi, Hongyang; Wang, Zhuyao; Li, Xiaojiao; Yin, Min; Wang, Lihua; Aldalbahi, Ali; El-Sayed, Nahed Nasser; Wang, Hui; Chen, Nan; Fan, Chunhai; Song, Haiyun

    2016-01-01

    Many types of biocompatible nanomaterials have proven of low cytotoxicity and hold great promise for various applications in nanomedicine. Whereas they generally do not cause apparent organ toxicity or tissue damage in adult animals, it is yet to determine their biological consequences in more general contexts. In this study, we investigate how silica nanoparticles (NPs) affect cellular activities and functions under several physiological or pathological conditions. Although silica NPs are generally regarded as "inert" nanocarriers and widely employed in biomedical studies, we find that they actively affect Wnt signaling in various types of cell lines, diminishing its anti-adipogenic effect in preadipocytes and pro-invasive effect in breast cancer cells, and more significantly, impair Wnt-regulated embryonic development in Zebrafish. We further demonstrate that intracellular silica NPs block Wnt signal transduction in a way resembling signaling molecules. Specifically, silica NPs target the Dvl protein, a key component of Wnt signaling cascade, for lysosomal degradation. As Wnt signaling play significant roles in embryonic development and adipogenesis, the observed physiological effects beyond toxicity imply potential risk of obesity, or developmental defects in somitogenesis and osteogenesis upon exposure to silica NPs. In addition, given the clinical implications of Wnt signaling in tumorigenesis and cancer metastasis, our work also establishes for the first time a molecular link between nanomaterials and the Wnt signaling pathway, which opens new door for novel applications of unmodified silica NPs in targeted therapy for cancers and other critical illness. PMID:27570552

  20. Silica Nanoparticles Target a Wnt Signal Transducer for Degradation and Impair Embryonic Development in Zebrafish

    PubMed Central

    Yi, Hongyang; Wang, Zhuyao; Li, Xiaojiao; Yin, Min; Wang, Lihua; Aldalbahi, Ali; El-Sayed, Nahed Nasser; Wang, Hui; Chen, Nan; Fan, Chunhai; Song, Haiyun

    2016-01-01

    Many types of biocompatible nanomaterials have proven of low cytotoxicity and hold great promise for various applications in nanomedicine. Whereas they generally do not cause apparent organ toxicity or tissue damage in adult animals, it is yet to determine their biological consequences in more general contexts. In this study, we investigate how silica nanoparticles (NPs) affect cellular activities and functions under several physiological or pathological conditions. Although silica NPs are generally regarded as “inert” nanocarriers and widely employed in biomedical studies, we find that they actively affect Wnt signaling in various types of cell lines, diminishing its anti-adipogenic effect in preadipocytes and pro-invasive effect in breast cancer cells, and more significantly, impair Wnt-regulated embryonic development in Zebrafish. We further demonstrate that intracellular silica NPs block Wnt signal transduction in a way resembling signaling molecules. Specifically, silica NPs target the Dvl protein, a key component of Wnt signaling cascade, for lysosomal degradation. As Wnt signaling play significant roles in embryonic development and adipogenesis, the observed physiological effects beyond toxicity imply potential risk of obesity, or developmental defects in somitogenesis and osteogenesis upon exposure to silica NPs. In addition, given the clinical implications of Wnt signaling in tumorigenesis and cancer metastasis, our work also establishes for the first time a molecular link between nanomaterials and the Wnt signaling pathway, which opens new door for novel applications of unmodified silica NPs in targeted therapy for cancers and other critical illness. PMID:27570552

  1. WNT3 Inhibits Cerebellar Granule Neuron Progenitor Proliferation and Medulloblastoma Formation via MAPK Activation

    PubMed Central

    Ayrault, Olivier; Kim, Jee Hae; Zhu, Xiaodong; Murphy, David A.; Van Aelst, Linda; Roussel, Martine F.; Hatten, Mary E.

    2013-01-01

    During normal cerebellar development, the remarkable expansion of granule cell progenitors (GCPs) generates a population of granule neurons that outnumbers the total neuronal population of the cerebral cortex, and provides a model for identifying signaling pathways that may be defective in medulloblastoma. While many studies focus on identifying pathways that promote growth of GCPs, a critical unanswered question concerns the identification of signaling pathways that block mitogenic stimulation and induce early steps in differentiation. Here we identify WNT3 as a novel suppressor of GCP proliferation during cerebellar development and an inhibitor of medulloblastoma growth in mice. WNT3, produced in early postnatal cerebellum, inhibits GCP proliferation by down-regulating pro-proliferative target genes of the mitogen Sonic Hedgehog (SHH) and the bHLH transcription factor Atoh1. WNT3 suppresses GCP growth through a non-canonical Wnt signaling pathway, activating prototypic mitogen-activated protein kinases (MAPKs), the Ras-dependent extracellular-signal-regulated kinases 1/2 (ERK1/2) and ERK5, instead of the classical β-catenin pathway. Inhibition of MAPK activity using a MAPK kinase (MEK) inhibitor reversed the inhibitory effect of WNT3 on GCP proliferation. Importantly, WNT3 inhibits proliferation of medulloblastoma tumor growth in mouse models by a similar mechanism. Thus, the present study suggests a novel role for WNT3 as a regulator of neurogenesis and repressor of neural tumors. PMID:24303070

  2. Neutral wetting brush layers for block copolymer thin films using homopolymer blends processed at high temperatures

    NASA Astrophysics Data System (ADS)

    Ceresoli, M.; Palermo, M.; Ferrarese Lupi, F.; Seguini, G.; Perego, M.; Zuccheri, G.; Phadatare, S. D.; Antonioli, D.; Gianotti, V.; Sparnacci, K.; Laus, M.

    2015-10-01

    Binary homopolymer blends of two hydroxyl-terminated polystyrene (PS-OH) and polymethylmethacrylate (PMMA-OH) homopolymers (Mn ˜ 16000 g mol-1) were grafted on SiO2 substrates by high-temperature (T > 150 °C), short-time (t < 600 s) thermal treatments. The resulting brush layer was tested to screen preferential interactions of the SiO2 substrate with the different symmetric and asymmetric PS-b-PMMA block copolymers deposited on top of the grafted molecules. By properly adjusting the blend composition and the processing parameters, an efficient surface neutralization path was identified, enabling the formation, in the block copolymer film, of homogeneous textures of lamellae or cylinders perpendicularly oriented with respect to the substrate. A critical interplay between the phase segregation of the homopolymer blends and their grafting process on the SiO2 was observed. In fact, the polar SiO2 is preferential for the PMMA-rich phase that forms a homogeneous layer on the substrate, while the PS-rich phase is located at the polymer-air interface. During the thermal treatment, phase segregation and grafting proceed simultaneously. Complete wetting of the PS rich phase on the PMMA rich phase leads to the formation of a PS/PMMA bilayer. In this case, the progressive diffusion of PS chains toward the polymer-SiO2 interface during the thermal treatment allows tuning of the brush layer composition.

  3. Biochemical Methods to Analyze Wnt Protein Secretion.

    PubMed

    Glaeser, Kathrin; Boutros, Michael; Gross, Julia Christina

    2016-01-01

    Wnt proteins act as potent morphogens in various aspects of embryonic development and adult tissue homeostasis. However, in addition to its physiological importance, aberrant Wnt signaling has been linked to the onset and progression of different types of cancer. On the cellular level, the secretion of Wnt proteins involves trafficking of lipid-modified Wnts from the endoplasmic reticulum (ER) to Golgi and further compartments via the Wnt cargo receptor evenness interrupted. Others and we have recently shown that Wnt proteins are secreted on extracellular vesicles (EVs) such as microvesicles and exosomes. Although more details about specific regulation of Wnt secretion steps are emerging, it remains largely unknown how Wnt proteins are channeled into different release pathways such as lipoprotein particles, EVs and cytonemes. Here, we describe protocols to purify and quantify Wnts from the supernatant of cells by either assessing total Wnt proteins in the supernatant or monitoring Wnt proteins on EVs. Purified Wnts from the supernatant as well as total cellular protein content can be investigated by immunoblotting. Additionally, the relative activity of canonical Wnts in the supernatant can be assessed by a dual-luciferase Wnt reporter assay. Quantifying the amount of secreted Wnt proteins and their activity in the supernatant of cells allows the investigation of intracellular trafficking events that regulate Wnt secretion and the role of extracellular modulators of Wnt spreading. PMID:27590148

  4. Wnt Signaling Inhibits Osteoclast Differentiation by Activating Canonical and Noncanonical cAMP/PKA Pathways

    PubMed Central

    Weivoda, Megan M; Ruan, Ming; Hachfeld, Christine M; Pederson, Larry; Howe, Alan; Davey, Rachel A; Zajac, Jeffrey D; Kobayashi, Yasuhiro; Williams, Bart O; Westendorf, Jennifer J; Khosla, Sundeep; Oursler, Merry Jo

    2016-01-01

    Although there has been extensive characterization of the Wnt signaling pathway in the osteoblast lineage, the effects of Wnt proteins on the osteoclast lineage are less well studied. We found that osteoclast lineage cells express canonical Wnt receptors. Wnt3a reduced osteoclast formation when applied to early bone-marrow macrophage (BMM) osteoclast differentiation cultures, whereas late addition did not suppress osteoclast formation. Early Wnt3a treatment inactivated the crucial transcription factor NFATc1 in osteoclast progenitors. Wnt3a led to the accumulation of nuclear β-catenin, confirming activation of canonical Wnt signaling. Reducing low-density lipoprotein receptor-related proteins (Lrp) 5 and Lrp6 protein expression prevented Wnt3a-induced inactivation of NFATc1; however, deletion of β-catenin did not block Wnt3a inactivation of NFATc1, suggesting that this effect was mediated by a noncanonical pathway. Wnt3a rapidly activated the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) pathway and pharmacological stimulation of cAMP/PKA signaling suppressed osteoclast differentiation; Wnt3a-induced NFATc1 phosphorylation was blocked by inhibiting interactions between PKA and A-kinase anchoring proteins (AKAPs). These data indicate that Wnt3a directly suppresses osteoclast differentiation through both canonical (β-catenin) and noncanonical (cAMP/PKA) pathways in osteoclast precursors. In vivo reduction of Lrp5 and Lrp6 expressions in the early osteoclast lineage via Rank promoter Cre recombination reduced trabecular bone mass, whereas disruption of Lrp5/6 expression in late osteoclast precursors via cathepsin K (Ctsk) promoter Cre recombination did not alter the skeletal phenotype. Surprisingly, reduction of Lrp5/6 in the early osteoclast lineage decreased osteoclast numbers, as well as osteoblast numbers. Published studies have previously noted that β-catenin signaling is required for osteoclast progenitor proliferation. Our in vivo data

  5. Reporting methods for processing and analysis of data from serial block face scanning electron microscopy.

    PubMed

    Borrett, S; Hughes, L

    2016-07-01

    Serial block face scanning electron microscopy is rapidly becoming a popular tool for collecting large three-dimensional data sets of cells and tissues, filling the resolution and volume gap between fluorescence microscopy and high-resolution electron microscopy. The automated collection of data within the instrument occupies the smallest proportion of the time required to prepare and analyse biological samples. It is the processing of data once it has been collected that proves the greatest challenge. In this review we discuss different methods that are used to process data. We suggest potential workflows that can be used to facilitate the transfer of raw image stacks into quantifiable data as well as propose a set of criteria for reporting methods for data analysis to enable replication of work. PMID:26800017

  6. Skeletal metastasis: treatments, mouse models, and the Wnt signaling

    PubMed Central

    Valkenburg, Kenneth C.; Steensma, Matthew R.; Williams, Bart O.; Zhong, Zhendong

    2013-01-01

    Skeletal metastases result in significant morbidity and mortality. This is particularly true of cancers with a strong predilection for the bone, such as breast, prostate, and lung cancers. There is currently no reliable cure for skeletal metastasis, and palliative therapy options are limited. The Wnt signaling pathway has been found to play an integral role in the process of skeletal metastasis and may be an important clinical target. Several experimental models of skeletal metastasis have been used to find new biomarkers and test new treatments. In this review, we discuss pathologic process of bone metastasis, the roles of the Wnt signaling, and the available experimental models and treatments. PMID:23327798

  7. Wnt-/-β-catenin pathway signaling in human hepatocellular carcinoma

    PubMed Central

    Waisberg, Jaques; Saba, Gabriela Tognini

    2015-01-01

    The molecular basis of the carcinogenesis of hepatocellular carcinoma (HCC) has not been adequately clarified, which negatively impacts the development of targeted therapy protocols for this overwhelming neoplasia. The aberrant activation of signaling in the HCC is primarily due to the deregulated expression of the components of the Wnt-/-β-catenin. This leads to the activation of β-catenin/T-cell factor-dependent target genes that control cell proliferation, cell cycle, apoptosis, and cell motility. The deregulation of the Wnt pathway is an early event in hepatocarcinogenesis. An aggressive phenotype was associated with HCC, since this pathway is implicated in the proliferation, migration, and invasiveness of cancer cells, regarding the cell’s own survival. The disruption of the signaling cascade Wnt-/-β-catenin has shown anticancer properties in HCC’s clinical evaluations of therapeutic molecules targeted for blocking the Wnt signaling pathway for the treatment of HCC, and it represents a promising perspective. The key to bringing this strategy in to clinical practice is to identify new molecules that would be effective only in tumor cells with aberrant signaling β-catenin. PMID:26609340

  8. Wnt-/-β-catenin pathway signaling in human hepatocellular carcinoma.

    PubMed

    Waisberg, Jaques; Saba, Gabriela Tognini

    2015-11-18

    The molecular basis of the carcinogenesis of hepatocellular carcinoma (HCC) has not been adequately clarified, which negatively impacts the development of targeted therapy protocols for this overwhelming neoplasia. The aberrant activation of signaling in the HCC is primarily due to the deregulated expression of the components of the Wnt-/-β-catenin. This leads to the activation of β-catenin/T-cell factor-dependent target genes that control cell proliferation, cell cycle, apoptosis, and cell motility. The deregulation of the Wnt pathway is an early event in hepatocarcinogenesis. An aggressive phenotype was associated with HCC, since this pathway is implicated in the proliferation, migration, and invasiveness of cancer cells, regarding the cell's own survival. The disruption of the signaling cascade Wnt-/-β-catenin has shown anticancer properties in HCC's clinical evaluations of therapeutic molecules targeted for blocking the Wnt signaling pathway for the treatment of HCC, and it represents a promising perspective. The key to bringing this strategy in to clinical practice is to identify new molecules that would be effective only in tumor cells with aberrant signaling β-catenin. PMID:26609340

  9. Sequence Analysis and Molecular Characterization of Wnt4 Gene in Metacestodes of Taenia solium

    PubMed Central

    Hou, Junling; Luo, Xuenong; Wang, Shuai; Yin, Cai; Zhang, Shaohua; Zhu, Xueliang; Dou, Yongxi

    2014-01-01

    Wnt proteins are a family of secreted glycoproteins that are evolutionarily conserved and considered to be involved in extensive developmental processes in metazoan organisms. The characterization of wnt genes may improve understanding the parasite's development. In the present study, a wnt4 gene encoding 491amino acids was amplified from cDNA of metacestodes of Taenia solium using reverse transcription PCR (RT-PCR). Bioinformatics tools were used for sequence analysis. The conserved domain of the wnt gene family was predicted. The expression profile of Wnt4 was investigated using real-time PCR. Wnt4 expression was found to be dramatically increased in scolex evaginated cysticerci when compared to invaginated cysticerci. In situ hybridization showed that wnt4 gene was distributed in the posterior end of the worm along the primary body axis in evaginated cysticerci. These findings indicated that wnt4 may take part in the process of cysticerci evagination and play a role in scolex/bladder development of cysticerci of T. solium. PMID:24850959

  10. Novel surface treatment materials for aligning block-co-polymer in directed self-assembly processes

    NASA Astrophysics Data System (ADS)

    Someya, Yasunobu; Wakayama, Hiroyuki; Endo, Takafumi; Sakamoto, Rikimaru

    2014-03-01

    Directed Self-Assembly (DSA) process is one of the attractive processes for creating the very fine pitch pattern. Especially, the contact hole shrink processes with block-co-polymer (BCP) or polymer blend materials were attractive processes for creating very small size hole patterns with better CD uniformity compare to general photo-lithography patterning. In general contact hole shrink process, the pattern of Spin-on Carbon Hardmask (SOC) or the photo Resist pattern created by Negative-Tone Development (NTD) process were selected for guide patterns. Since the alignment property of BCP was affected by the surface of these guide materials, it is important to control the surface condition of guide in order to obtain good shrunk contact hole patterns. In this study, we will report the surface treatment materials to control the surface condition of guide patterns such as SOC or NTD resist to achieve the better contact hole shrink performance. These materials were attached to guide pattern surface and controlled the surface energy.

  11. Autoplan: A self-processing network model for an extended blocks world planning environment

    NASA Technical Reports Server (NTRS)

    Dautrechy, C. Lynne; Reggia, James A.; Mcfadden, Frank

    1990-01-01

    Self-processing network models (neural/connectionist models, marker passing/message passing networks, etc.) are currently undergoing intense investigation for a variety of information processing applications. These models are potentially very powerful in that they support a large amount of explicit parallel processing, and they cleanly integrate high level and low level information processing. However they are currently limited by a lack of understanding of how to apply them effectively in many application areas. The formulation of self-processing network methods for dynamic, reactive planning is studied. The long-term goal is to formulate robust, computationally effective information processing methods for the distributed control of semiautonomous exploration systems, e.g., the Mars Rover. The current research effort is focusing on hierarchical plan generation, execution and revision through local operations in an extended blocks world environment. This scenario involves many challenging features that would be encountered in a real planning and control environment: multiple simultaneous goals, parallel as well as sequential action execution, action sequencing determined not only by goals and their interactions but also by limited resources (e.g., three tasks, two acting agents), need to interpret unanticipated events and react appropriately through replanning, etc.

  12. Opposing activities of Notch and Wnt signaling regulate intestinal stem cells and gut homeostasis.

    PubMed

    Tian, Hua; Biehs, Brian; Chiu, Cecilia; Siebel, Christian W; Wu, Yan; Costa, Mike; de Sauvage, Frederic J; Klein, Ophir D

    2015-04-01

    Proper organ homeostasis requires tight control of adult stem cells and differentiation through the integration of multiple inputs. In the mouse small intestine, Notch and Wnt signaling are required both for stem cell maintenance and for a proper balance of differentiation between secretory and absorptive cell lineages. In the absence of Notch signaling, stem cells preferentially generate secretory cells at the expense of absorptive cells. Here, we use function-blocking antibodies against Notch receptors to demonstrate that Notch blockade perturbs intestinal stem cell function by causing a derepression of the Wnt signaling pathway, leading to misexpression of prosecretory genes. Importantly, attenuation of the Wnt pathway rescued the phenotype associated with Notch blockade. These studies bring to light a negative regulatory mechanism that maintains stem cell activity and balanced differentiation, and we propose that the interaction between Wnt and Notch signaling described here represents a common theme in adult stem cell biology. PMID:25818302

  13. Cell cycle control of Wnt/β-catenin signalling by conductin/axin2 through CDC20

    PubMed Central

    Hadjihannas, Michel V; Bernkopf, Dominic B; Brückner, Martina; Behrens, Jürgen

    2012-01-01

    Wnt/β-catenin signalling regulates cell proliferation by modulating the cell cycle and is negatively regulated by conductin/axin2/axil. We show that conductin levels peak at G2/M followed by a rapid decline during return to G1. In line with this, Wnt/β-catenin target genes are low at G2/M and high at G1/S, and β-catenin phosphorylation oscillates during the cell cycle in a conductin-dependent manner. Conductin is degraded by the anaphase-promoting complex/cyclosome cofactor CDC20. Knockdown of CDC20 blocks Wnt signalling through conductin. CDC20-resistant conductin inhibits Wnt signalling and attenuates colony formation of colorectal cancer cells. We propose that CDC20-mediated degradation of conductin regulates Wnt/β-catenin signalling for maximal activity during G1/S. PMID:22322943

  14. Cloning and characterization of wnt4a gene and evidence for positive selection in half-smooth tongue sole (Cynoglossus semilaevis)

    NASA Astrophysics Data System (ADS)

    Hu, Qiaomu; Zhu, Ying; Liu, Yang; Wang, Na; Chen, Songlin

    2014-11-01

    Wnt4 gene plays a role in developmental processes in mammals. However, little is known regarding its function in teleosts. We cloned and characterized the full-length half-smooth tongue sole (Cynoglossus semilaevis) wnt4a gene (CS-wnt4a). CS-wnt4a cDNA was 1746 bp in length encoding 353aa. CS-wnt4a expression level was highest in the testis, and gradually increased in the developing gonads until 1 year of age. In situ hybridization revealed that CS-wnt4a expression level was highest in stage II oocytes and sperm in the adult ovary and testis, respectively. CS-wnt4a expression level was significantly up-regulated in the gonads after exposure to high temperature. The level of methylation of the CS-wnt4a first exon was negatively correlated with the expression of CS-wnt4a. The branch-site model suggested that vertebrate wnt4a differed significantly from that of wnt4b, and that the selective pressures differed between ancestral aquatic and terrestrial organisms. Two positively selected sites were found in the ancestral lineages of teleost fish, but none in the ancestral lineages of mammals. One positively selected site was located on the α-helices of the 3D structure, the other on the random coil. Our results are of value for further study of the function of wnt4 and the mechanism of selection.

  15. Cloning and characterization of wnt4a gene and evidence for positive selection in half-smooth tongue sole (Cynoglossus semilaevis)

    PubMed Central

    Hu, Qiaomu; Zhu, Ying; Liu, Yang; Wang, Na; Chen, Songlin

    2014-01-01

    Wnt4 gene plays a role in developmental processes in mammals. However, little is known regarding its function in teleosts. We cloned and characterized the full-length half-smooth tongue sole (Cynoglossus semilaevis) wnt4a gene (CS-wnt4a). CS-wnt4a cDNA was 1746 bp in length encoding 353aa. CS-wnt4a expression level was highest in the testis, and gradually increased in the developing gonads until 1 year of age. In situ hybridization revealed that CS-wnt4a expression level was highest in stage II oocytes and sperm in the adult ovary and testis, respectively. CS-wnt4a expression level was significantly up-regulated in the gonads after exposure to high temperature. The level of methylation of the CS-wnt4a first exon was negatively correlated with the expression of CS-wnt4a. The branch-site model suggested that vertebrate wnt4a differed significantly from that of wnt4b, and that the selective pressures differed between ancestral aquatic and terrestrial organisms. Two positively selected sites were found in the ancestral lineages of teleost fish, but none in the ancestral lineages of mammals. One positively selected site was located on the α-helices of the 3D structure, the other on the random coil. Our results are of value for further study of the function of wnt4 and the mechanism of selection. PMID:25418599

  16. WNT-3A Regulates an Axin1/NRF2 Complex That Regulates Antioxidant Metabolism in Hepatocytes

    PubMed Central

    Rada, Patricia; Rojo, Ana I.; Offergeld, Anika; Feng, Gui Jie; Velasco-Martín, Juan P.; González-Sancho, José Manuel; Valverde, Ángela M.; Dale, Trevor; Regadera, Javier

    2015-01-01

    Abstract Aims: Nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a master regulator of oxidant and xenobiotic metabolism, but it is unknown how it is regulated to provide basal expression of this defense system. Here, we studied the putative connection between NRF2 and the canonical WNT pathway, which modulates hepatocyte metabolism. Results: WNT-3A increased the levels of NRF2 and its transcriptional signature in mouse hepatocytes and HEK293T cells. The use of short interfering RNAs in hepatocytes and mouse embryonic fibroblasts which are deficient in the redox sensor Kelch-like ECH-associated protein 1 (KEAP1) indicated that WNT-3A activates NRF2 in a β-Catenin- and KEAP1-independent manner. WNT-3A stabilized NRF2 by preventing its GSK-3-dependent phosphorylation and subsequent SCF/β-TrCP-dependent ubiquitination and proteasomal degradation. Axin1 and NRF2 were physically associated in a protein complex that was regulated by WNT-3A, involving the central region of Axin1 and the Neh4/Neh5 domains of NRF2. Axin1 knockdown increased NRF2 protein levels, while Axin1 stabilization with Tankyrase inhibitors blocked WNT/NRF2 signaling. The relevance of this novel pathway was assessed in mice with a conditional deletion of Axin1 in the liver, which showed upregulation of the NRF2 signature in hepatocytes and disruption of liver zonation of antioxidant metabolism. Innovation: NRF2 takes part in a protein complex with Axin1 that is regulated by the canonical WNT pathway. This new WNT-NRF2 axis controls the antioxidant metabolism of hepatocytes. Conclusion: These results uncover the participation of NRF2 in a WNT-regulated signalosome that participates in basal maintenance of hepatic antioxidant metabolism. Antioxid. Redox Signal. 22, 555–571. PMID:25336178

  17. Expression of Gpr177, a Wnt trafficking regulator, in mouse embryogenesis

    PubMed Central

    Yu, Hsiao-Man Ivy; Jin, Ying; Fu, Jiang; Hsu, Wei

    2010-01-01

    Wls/Evi/Srt encoding a multipass transmembrane protein has been identified as a regulator for proper sorting and secretion of Wnt in flies. We have previously demonstrated that Gpr177 is the mouse orthologue required for axis determination. Gpr177 is a transcriptional target of Wnt which is activated to assist its subcellular distribution in a feedback regulatory loop. We therefore proposed that reciprocal regulation of Wnt and Gpr177 is essential for the Wnt-dependent developmental and pathogenic processes. Here we examine the expression pattern of Gpr177 in mouse development. Gpr177 is expressed in a variety of tissues and cell types during organogenesis. Furthermore, Gpr177 is a glycoprotein primarily accumulating in the Golgi apparatus in signal-producing cells. The glycosylation of Gpr177 is necessary for proper transportation in the secretory pathway. Our findings suggest that the Gpr177-mediated regulation of Wnt is crucial for organogenesis in health and disease. PMID:20549736

  18. Expression of Gpr177, a Wnt trafficking regulator, in mouse embryogenesis.

    PubMed

    Yu, Hsiao-Man Ivy; Jin, Ying; Fu, Jiang; Hsu, Wei

    2010-07-01

    Wls/Evi/Srt encoding a multipass transmembrane protein has been identified as a regulator for proper sorting and secretion of Wnt in flies. We have previously demonstrated that Gpr177 is the mouse ortholog required for axis determination. Gpr177 is a transcriptional target of Wnt that is activated to assist its subcellular distribution in a feedback regulatory loop. We, therefore, proposed that reciprocal regulation of Wnt and Gpr177 is essential for the Wnt-dependent developmental and pathogenic processes. Here, we examine the expression pattern of Gpr177 in mouse development. Gpr177 is expressed in a variety of tissues and cell types during organogenesis. Furthermore, Gpr177 is a glycoprotein primarily accumulating in the Golgi apparatus in signal-producing cells. The glycosylation of Gpr177 is necessary for proper transportation in the secretory pathway. Our findings suggest that the Gpr177-mediated regulation of Wnt is crucial for organogenesis in health and disease. PMID:20549736

  19. KLF4 transcriptionally activates non-canonical WNT5A to control epithelial stratification

    PubMed Central

    Tetreault, Marie-Pier; Weinblatt, Daniel; Shaverdashvili, Khvaramze; Yang, Yizeng; Katz, Jonathan P.

    2016-01-01

    Epithelial differentiation and stratification are essential for normal homeostasis, and disruption of these processes leads to both injury and cancer. The zinc-finger transciption factor KLF4 is a key driver of epithelial differentiation, yet the mechanisms and targets by which KLF4 controls differentiation are not well understood. Here, we define WNT5A, a non-canonical Wnt ligand implicated in epithelial differentiation, repair, and cancer, as a direct transcriptional target that is activated by KLF4 in squamous epithelial cells. Further, we demonstrate functionally that WNT5A mediates KLF4 control of epithelial differentiation and stratification, as treatment of keratinocytes with WNT5A rescues defective epithelial stratification resulting from KLF4 loss. Finally, we show that the small GTPase CDC42 is regulated by KLF4 in a WNT5A dependent manner. As such, we delineate a novel pathway for epithelial differentiation and stratification and define potential therapeutic targets for epithelial diseases. PMID:27184424

  20. Expression of secreted Wnt pathway components reveals unexpected complexity of the planarian amputation response.

    PubMed

    Gurley, Kyle A; Elliott, Sarah A; Simakov, Oleg; Schmidt, Heiko A; Holstein, Thomas W; Sánchez Alvarado, Alejandro

    2010-11-01

    Regeneration is widespread throughout the animal kingdom, but our molecular understanding of this process in adult animals remains poorly understood. Wnt/β-catenin signaling plays crucial roles throughout animal life from early development to adulthood. In intact and regenerating planarians, the regulation of Wnt/β-catenin signaling functions to maintain and specify anterior/posterior (A/P) identity. Here, we explore the expression kinetics and RNAi phenotypes for secreted members of the Wnt signaling pathway in the planarian Schmidtea mediterranea. Smed-wnt and sFRP expression during regeneration is surprisingly dynamic and reveals fundamental aspects of planarian biology that have been previously unappreciated. We show that after amputation, a wounding response precedes rapid re-organization of the A/P axis. Furthermore, cells throughout the body plan can mount this response and reassess their new A/P location in the complete absence of stem cells. While initial stages of the amputation response are stem cell independent, tissue remodeling and the integration of a new A/P address with anatomy are stem cell dependent. We also show that WNT5 functions in a reciprocal manner with SLIT to pattern the planarian mediolateral axis, while WNT11-2 patterns the posterior midline. Moreover, we perform an extensive phylogenetic analysis on the Smed-wnt genes using a method that combines and integrates both sequence and structural alignments, enabling us to place all nine genes into Wnt subfamilies for the first time. PMID:20707997

  1. Boehmenan, a lignan from Hibiscus ficulneus, showed Wnt signal inhibitory activity.

    PubMed

    Shono, Takumi; Ishikawa, Naoki; Toume, Kazufumi; Arai, Midori A; Ahmed, Firoj; Sadhu, Samir K; Ishibashi, Masami

    2015-07-15

    The Wnt signal pathway modulates numerous biological processes, and its aberrant activation is related to various diseases. Therefore, inhibition of the Wnt signal may provide an effective (or efficient) strategy for these diseases. Cell-based luciferase assay targeting the Wnt signal (TOP assay) revealed that Hibiscus ficulneus extract inhibited the Wnt signal. The activity-guided isolation of the MeOH extract of H. ficulneus stems yielded four known (1-4) lignans along with myriceric acid (5). Compounds 1-4 potently inhibited the Wnt signal with TOPflash IC50 values of 1.0, 4.5, 6.3, and 1.9 μM, respectively. Compound 1 exhibited cytotoxicity against both Wnt-dependent (HCT116) and Wnt-independent (RKO) cells. Western blot analysis showed that 1 decreased the expression of full, cytosolic and nuclear β-catenin along with c-myc in STF/293 cells. Our results suggested that 1 may have inhibited the Wnt signal by decreasing β-catenin levels. PMID:26026364

  2. Wnt5a Suppresses β-catenin Signaling during Hair Follicle Regeneration

    PubMed Central

    Xing, Yizhan; Ma, Xiaogen; Guo, Haiying; Deng, Fang; Yang, Jin; Li, Yuhong

    2016-01-01

    Hair follicles display periodic growth. Wnt signaling is a critical regulator for hair follicle regeneration. Previously, we reported that Wnt5a inhibits the telogen-to-anagen transition of hair follicles, but the mechanism by which this process occurs has not yet been reported. Here, we determined the expression patterns of Wnt signaling pathway molecules by quantitative reverse transcription polymerase chain reaction, western blot, and immunohistochemistry and found that β-catenin signaling was suppressed by Wnt5a. We then compared the phenotypes and expression patterns following β-catenin knockdown and Wnt5a overexpression during hair follicle regeneration induced by hair depilation and observed similar patterns. In addition, we performed a rescue experiment in the JB6 cell line and found that the inhibitory effect of Wnt5a on cell proliferation could be rescued by the addition of Wnt3a. Our data reveal that Wnt5a suppresses the activation of β-catenin signaling during hair follicle regeneration. PMID:27499692

  3. Inhibition of Wnt/β-catenin pathway promotes regenerative repair of cutaneous and cartilage injury.

    PubMed

    Bastakoty, Dikshya; Saraswati, Sarika; Cates, Justin; Lee, Ethan; Nanney, Lillian B; Young, Pampee P

    2015-12-01

    Wound healing in mammals is a fibrotic process. The mechanisms driving fibrotic (as opposed to regenerative) repair are poorly understood. Herein we report that therapeutic Wnt inhibition with topical application of small-molecule Wnt inhibitors can reduce fibrosis and promote regenerative cutaneous wound repair. In the naturally stented model of ear punch injury, we found that Wnt/β-catenin pathway is activated most notably in the dermis of the wound bed early (d 2) after injury and subsides to baseline levels by d10. Topical application of either of 2 mechanistically distinct small-molecule Wnt pathway inhibitors (a tankyrase inhibitor, XAV-939, and the U.S. Food and Drug Administration-approved casein kinase activator, pyrvinium) in C57Bl/6J mice resulted in significantly increased rates of wound closure (72.3 ± 14.7% with XAV-939; and 52.1 ± 20.9% with pyrvinium) compared with contralateral controls (38.1 ± 23.0 and 40.4.± 16.7%, respectively). Histologically, Wnt inhibition reduced fibrosis as measured by α-smooth muscle actin positive myofibroblasts and collagen type I α1 synthesis. Wnt inhibition also restored skin architecture including adnexal structures in ear wounds and dermal-epidermal junction with rete pegs in excisional wounds. Additionally, in ear punch injury Wnt inhibitor treatment enabled regeneration of auricular cartilage. Our study shows that pharmacologic Wnt inhibition holds therapeutic utility for regenerative repair of cutaneous wounds. PMID:26268926

  4. Making sense of Wnt signaling—linking hair cell regeneration to development

    PubMed Central

    Jansson, Lina; Kim, Grace S.; Cheng, Alan G.

    2015-01-01

    Wnt signaling is a highly conserved pathway crucial for development and homeostasis of multicellular organisms. Secreted Wnt ligands bind Frizzled receptors to regulate diverse processes such as axis patterning, cell division, and cell fate specification. They also serve to govern self-renewal of somatic stem cells in several adult tissues. The complexity of the pathway can be attributed to the myriad of Wnt and Frizzled combinations as well as its diverse context-dependent functions. In the developing mouse inner ear, Wnt signaling plays diverse roles, including specification of the otic placode and patterning of the otic vesicle. At later stages, its activity governs sensory hair cell specification, cell cycle regulation, and hair cell orientation. In regenerating sensory organs from non-mammalian species, Wnt signaling can also regulate the extent of proliferative hair cell regeneration. This review describes the current knowledge of the roles of Wnt signaling and Wnt-responsive cells in hair cell development and regeneration. We also discuss possible future directions and the potential application and limitation of Wnt signaling in augmenting hair cell regeneration. PMID:25814927

  5. Expression of secreted Wnt pathway components reveals unexpected complexity of the planarian amputation response

    PubMed Central

    Gurley, Kyle A.; Elliott, Sarah A.; Simakov, Oleg; Schmidt, Heiko A.; Holstein, Thomas W.; Sánchez Alvarado, Alejandro

    2010-01-01

    Regeneration is widespread throughout the animal kingdom, but our molecular understanding of this process in adult animals remains poorly understood. Wnt/β-catenin signaling plays crucial roles throughout animal life from early development to adulthood. In intact and regenerating planarians, the regulation of Wnt/β-catenin signaling functions to maintain and specify anterior/posterior (A/P) identity. Here, we explore the expression kinetics and RNAi phenotypes for secreted members of the Wnt signaling pathway in the planarian Schmidtea mediterranea. Smed-wnt and sFRP expression during regeneration is surprisingly dynamic and reveals fundamental aspects of planarian biology that have been previously unappreciated. We show that after amputation, a wounding response precedes rapid reorganization of the A/P axis. Furthermore, cells throughout the body plan can mount this response and reassess their new A/P location in the complete absence of stem cells. While initial stages of the amputation response are stem cell independent, tissue remodeling and the integration of new A/P address with anatomy are stem cell dependent. We also show that WNT5 functions in a reciprocal manner with SLIT to pattern the planarian mediolateral axis, while WNT11-2 patterns the posterior midline. Moreover, we perform an extensive phylogenetic analysis on the Smed-wnt genes using a method that combines and integrates both sequence and structural alignments, enabling us to place all nine genes into Wnt subfamilies for the first time. PMID:20707997

  6. Process-directed self-assembly of block copolymers: a computer simulation study

    NASA Astrophysics Data System (ADS)

    Müller, Marcus; Sun, De-Wen

    2015-05-01

    The free-energy landscape of self-assembling block copolymer systems is characterized by a multitude of metastable minima and concomitant protracted relaxation times of the morphology. Tailoring rapid changes (quench) of thermodynamic conditions, one can reproducibly trap the ensuing kinetics of self-assembly in a specific metastable state. To this end, it is necessary to (1) control the generation of well-defined, highly unstable states and (2) design the unstable state such that the ensuing spontaneous kinetics of structure formation reaches the desired metastable morphology. This process-directed self-assembly provides an alternative to fine-tuning molecular architecture by synthesis or blending, for instance, in order to fabricate complex network structures. Comparing our simulation results to recently developed free-energy techniques, we highlight the importance of non-equilibrium molecular conformations in the starting state and motivate the significance of the local conservation of density.

  7. Selective inhibition of farnesyl-protein transferase blocks ras processing in vivo.

    PubMed

    Gibbs, J B; Pompliano, D L; Mosser, S D; Rands, E; Lingham, R B; Singh, S B; Scolnick, E M; Kohl, N E; Oliff, A

    1993-04-15

    The ras oncogene product, Ras, is synthesized in vivo as a precursor protein that requires post-translational processing to become biologically active and to be capable of transforming mammalian cells. Farnesylation appears to be a critical modification of Ras, and thus inhibitors of the farnesyl-protein transferase (FPTase) that catalyzes this reaction may block ras-dependent tumorigenesis. Three structural classes of FPTase inhibitors were identified: (alpha-hydroxyfarnesyl)phosphonic acid, chaetomellic acids, and zaragozic acids. By comparison, these compounds were weaker inhibitors of geranylgeranyl-protein transferases. Each of these inhibitors was competitive with respect to farnesyl diphosphate in the FPTase reaction. All compounds were assayed for inhibition of Ras processing in Ha-ras-transformed NIH3T3 fibroblasts. Ras processing was inhibited by 1 microM (alpha-hydroxyfarnesyl)phosphonic acid. Neither chaetomellic acid nor zaragozic acid were active in this assay. These results are the first demonstration that a small organic chemical selected for inhibition of FPTase can inhibit Ras processing in vivo. PMID:8463291

  8. Wnt signalling tunes neurotransmitter release by directly targeting Synaptotagmin-1

    PubMed Central

    Ciani, Lorenza; Marzo, Aude; Boyle, Kieran; Stamatakou, Eleanna; Lopes, Douglas M.; Anane, Derek; McLeod, Faye; Rosso, Silvana B.; Gibb, Alasdair; Salinas, Patricia C.

    2015-01-01

    The functional assembly of the synaptic release machinery is well understood; however, how signalling factors modulate this process remains unknown. Recent studies suggest that Wnts play a role in presynaptic function. To examine the mechanisms involved, we investigated the interaction of release machinery proteins with Dishevelled-1 (Dvl1), a scaffold protein that determines the cellular locale of Wnt action. Here we show that Dvl1 directly interacts with Synaptotagmin-1 (Syt-1) and indirectly with the SNARE proteins SNAP25 and Syntaxin (Stx-1). Importantly, the interaction of Dvl1 with Syt-1, which is regulated by Wnts, modulates neurotransmitter release. Moreover, presynaptic terminals from Wnt signalling-deficient mice exhibit reduced release probability and are unable to sustain high-frequency release. Consistently, the readily releasable pool size and formation of SNARE complexes are reduced. Our studies demonstrate that Wnt signalling tunes neurotransmitter release and identify Syt-1 as a target for modulation by secreted signalling proteins. PMID:26400647

  9. Wnt inhibition enhances browning of mouse primary white adipocytes

    PubMed Central

    Lo, Kinyui Alice; Ng, Pei Yi; Kabiri, Zahra; Virshup, David; Sun, Lei

    2016-01-01

    ABSTRACT The global epidemic in obesity and metabolic syndrome requires novel approaches to tackle. White adipose tissue, traditionally seen as a passive energy-storage organ, can be induced to take on certain characteristics of brown fat in a process called browning. The “browned” white adipose tissue, or beige fat, is a potential anti-obesity target. Various signaling pathways can enhance browning. Wnt is a key regulator of adipocyte biology, but its role in browning has not been explored. In this study, we found that in primary mouse adipocytes derived from the inguinal depot, Wnt inhibition by both chemical and genetic methods significantly enhanced browning. The effect of Wnt inhibition on browning most likely targets the beige precursor cells in selected adipose depots. PMID:27386162

  10. Apc bridges Wnt/{beta}-catenin and BMP signaling during osteoblast differentiation of KS483 cells

    SciTech Connect

    Miclea, Razvan L.; Horst, Geertje van der; Robanus-Maandag, Els C.; Loewik, Clemens W.G.M.; Oostdijk, Wilma; Wit, Jan M.; Karperien, Marcel

    2011-06-10

    The canonical Wnt signaling pathway influences the differentiation of mesenchymal cell lineages in a quantitative and qualitative fashion depending on the dose of {beta}-catenin signaling. Adenomatous polyposis coli (Apc) is the critical intracellular regulator of {beta}-catenin turnover. To better understand the molecular mechanisms underlying the role of Apc in regulating the differentiation capacity of skeletal progenitor cells, we have knocked down Apc in the murine mesenchymal stem cell-like KS483 cells by stable expression of Apc-specific small interfering RNA. In routine culture, KSFrt-Apc{sub si} cells displayed a mesenchymal-like spindle shape morphology, exhibited markedly decreased proliferation and increased apoptosis. Apc knockdown resulted in upregulation of the Wnt/{beta}-catenin and the BMP/Smad signaling pathways, but osteogenic differentiation was completely inhibited. This effect could be rescued by adding high concentrations of BMP-7 to the differentiation medium. Furthermore, KSFrt-Apc{sub si} cells showed no potential to differentiate into chondrocytes or adipocytes. These results demonstrate that Apc is essential for the proliferation, survival and differentiation of KS483 cells. Apc knockdown blocks the osteogenic differentiation of skeletal progenitor cells, a process that can be overruled by high BMP signaling.

  11. Can we safely target the WNT pathway?

    PubMed Central

    Kahn, Michael

    2015-01-01

    WNT–β-catenin signalling is involved in a multitude of developmental processes and the maintenance of adult tissue homeostasis by regulating cell proliferation, differentiation, migration, genetic stability and apoptosis, as well as by maintaining adult stem cells in a pluripotent state. Not surprisingly, aberrant regulation of this pathway is therefore associated with a variety of diseases, including cancer, fibrosis and neurodegeneration. Despite this knowledge, therapeutic agents specifically targeting the WNT pathway have only recently entered clinical trials and none has yet been approved. This Review examines the problems and potential solutions to this vexing situation and attempts to bring them into perspective. PMID:24981364

  12. Hipk2 and PP1c cooperate to maintain Dvl protein levels required for Wnt signal transduction.

    PubMed

    Shimizu, Nobuyuki; Ishitani, Shizuka; Sato, Atsushi; Shibuya, Hiroshi; Ishitani, Tohru

    2014-09-11

    The phosphoprotein Dishevelled (Dvl) is a common essential component of Wnt/β-catenin and Wnt/planar cell polarity (PCP) signaling pathways. However, the regulation and significance of Dvl phosphorylation are not fully understood. Here, we show that homeodomain-interacting protein kinase 2 (Hipk2) facilitates protein phosphatase 1 catalytic subunit (PP1c)-mediated dephosphorylation of Dvl via its C-terminal domain and that this dephosphorylation blocks ubiquitination and consequent degradation mediated by the E3 ubiquitin ligase Itch, which targets the phosphorylated form of Dvl proteins. Inhibition of Hipk2 or PP1c function reduces Dvl protein levels and suppresses Wnt/β-catenin and Wnt/PCP pathway-dependent events in mammalian cells and zebrafish embryos, suggesting that Hipk2 and PP1c are essential for maintaining Dvl protein levels that are sufficient to activate Wnt signaling. We also show that Wnt-3a, a Wnt/β-catenin ligand, induces dissociation of the Dvl-Hipk2-PP1c complex and Dvl degradation under high-cell-density conditions. This regulation may be a negative feedback mechanism that fine-tunes Wnt/β-catenin signaling. PMID:25159144

  13. Blockade of Wnt/β-Catenin Pathway Aggravated Silica-Induced Lung Inflammation through Tregs Regulation on Th Immune Responses

    PubMed Central

    Dai, Wujing; Liu, Fangwei; Li, Chao; Lu, Yiping; Lu, Xiaowei; Du, Sitong; Chen, Ying; Weng, Dong; Chen, Jie

    2016-01-01

    CD4+ T cells play an important role in regulating silica-induced inflammation and fibrosis. Recent studies showed that Wnt/β-catenin pathway could modulate the function and the differentiation of CD4+ T cells. Therefore, Wnt/β-catenin pathway may participate in the development and progress of silicosis. To investigate the role of Wnt/β-catenin pathway, we used lentivirus expressing β-catenin shRNA to block the Wnt/β-catenin pathway by intratracheal instillation to the mice model of silicosis. Treatment of lentivirus could significantly aggravate the silica-induced lung inflammation and attenuated the fibrosis at the late stage. By analyzing CD4+ T cells, we found that blockade of Wnt/β-catenin pathway suppressed regulatory T cells (Tregs). Reciprocally, enhanced Th17 response was responsible for the further accumulation of neutrophils and production of proinflammatory cytokines. In addition, blockade of Wnt/β-catenin pathway delayed the Th1/Th2 polarization by inhibiting Tregs and Th2 response. These results indicated that Wnt/β-catenin pathway could regulate Tregs to modulate Th immune response, which finally altered the pathological character of silicosis. Our study suggested that Wnt/β-catenin pathway might be a potential target to treat the silica-induced inflammation and fibrosis. PMID:27069316

  14. Blockade of Wnt/β-Catenin Pathway Aggravated Silica-Induced Lung Inflammation through Tregs Regulation on Th Immune Responses.

    PubMed

    Dai, Wujing; Liu, Fangwei; Li, Chao; Lu, Yiping; Lu, Xiaowei; Du, Sitong; Chen, Ying; Weng, Dong; Chen, Jie

    2016-01-01

    CD4(+) T cells play an important role in regulating silica-induced inflammation and fibrosis. Recent studies showed that Wnt/β-catenin pathway could modulate the function and the differentiation of CD4(+) T cells. Therefore, Wnt/β-catenin pathway may participate in the development and progress of silicosis. To investigate the role of Wnt/β-catenin pathway, we used lentivirus expressing β-catenin shRNA to block the Wnt/β-catenin pathway by intratracheal instillation to the mice model of silicosis. Treatment of lentivirus could significantly aggravate the silica-induced lung inflammation and attenuated the fibrosis at the late stage. By analyzing CD4(+) T cells, we found that blockade of Wnt/β-catenin pathway suppressed regulatory T cells (Tregs). Reciprocally, enhanced Th17 response was responsible for the further accumulation of neutrophils and production of proinflammatory cytokines. In addition, blockade of Wnt/β-catenin pathway delayed the Th1/Th2 polarization by inhibiting Tregs and Th2 response. These results indicated that Wnt/β-catenin pathway could regulate Tregs to modulate Th immune response, which finally altered the pathological character of silicosis. Our study suggested that Wnt/β-catenin pathway might be a potential target to treat the silica-induced inflammation and fibrosis. PMID:27069316

  15. Norcantharidin inhibits Wnt signal pathway via promoter demethylation of WIF-1 in human non-small cell lung cancer.

    PubMed

    Xie, Junran; Zhang, Yaping; Hu, Xuming; Lv, Ran; Xiao, Dongju; Jiang, Li; Bao, Qi

    2015-05-01

    Wingless-type (Wnt) family of secreted glycoproteins is a group of signal molecules implicated in oncogenesis. Abnormal activation of Wnt signal pathway is associated with a variety of human cancers, including non-small cell lung cancer (NSCLC). Wnt antagonists, such as the secreted frizzled-related protein (SFRP) family, Wnt inhibitory factor-1 (WIF-1) and cerberus, inhibit Wnt signal pathway by directly binding to Wnt molecules. Norcantharidin (NCTD) is known to possess anticancer activity but less nephrotoxicity than cantharidin. In this study, we found that NCTD inhibited cell proliferation, induced apoptosis, arrested cell cycle and suppressed cell invasion/migration in vitro. Additionally, Wnt signal pathway transcription was also suppressed. NCTD treatment blocked cytoplasmic translocation of beta-catenin into the nucleus. Alterations of apoptosis-related proteins, such as Bax, cleaved caspase-3 (pro-apoptotic) and Bcl-2 (anti-apoptotic), had been detected. Furthermore, the expression levels of WIF-1 and SFRP1 were significantly increased in NCTD-treated groups compared with negative control (NC) groups. Abnormal methylation was observed in NC groups, while NCTD treatment promoted WIF-1 demethylation. The present study revealed that NCTD activated WIF-1 via promoter demethylation, inhibiting the canonical Wnt signal pathway in NSCLC, which may present a new therapeutic target in vivo. PMID:25814287

  16. Wnt addiction of genetically defined cancers reversed by PORCN inhibition

    PubMed Central

    Madan, B; Ke, Z; Harmston, N; Ho, S Y; Frois, A O; Alam, J; Jeyaraj, D A; Pendharkar, V; Ghosh, K; Virshup, I H; Manoharan, V; Ong, E H Q; Sangthongpitag, K; Hill, J; Petretto, E; Keller, T H; Lee, M A; Matter, A; Virshup, D M

    2016-01-01

    Enhanced sensitivity to Wnts is an emerging hallmark of a subset of cancers, defined in part by mutations regulating the abundance of their receptors. Whether these mutations identify a clinical opportunity is an important question. Inhibition of Wnt secretion by blocking an essential post-translational modification, palmitoleation, provides a useful therapeutic intervention. We developed a novel potent, orally available PORCN inhibitor, ETC-1922159 (henceforth called ETC-159) that blocks the secretion and activity of all Wnts. ETC-159 is remarkably effective in treating RSPO-translocation bearing colorectal cancer (CRC) patient-derived xenografts. This is the first example of effective targeted therapy for this subset of CRC. Consistent with a central role of Wnt signaling in regulation of gene expression, inhibition of PORCN in RSPO3-translocated cancers causes a marked remodeling of the transcriptome, with loss of cell cycle, stem cell and proliferation genes, and an increase in differentiation markers. Inhibition of Wnt signaling by PORCN inhibition holds promise as differentiation therapy in genetically defined human cancers. PMID:26257057

  17. Strabismus-mediated primary archenteron invagination is uncoupled from Wnt/β-catenin-dependent endoderm cell fate specification in Nematostella vectensis (Anthozoa, Cnidaria): Implications for the evolution of gastrulation

    PubMed Central

    2011-01-01

    Background Gastrulation is a uniquely metazoan character, and its genesis was arguably the key step that enabled the remarkable diversification within this clade. The process of gastrulation involves two tightly coupled events during embryogenesis of most metazoans. Morphogenesis produces a distinct internal epithelial layer in the embryo, and this epithelium becomes segregated as an endoderm/endomesodermal germ layer through the activation of a specific gene regulatory program. The developmental mechanisms that induced archenteron formation and led to the segregation of germ layers during metazoan evolution are unknown. But an increased understanding of development in early diverging taxa at the base of the metazoan tree may provide insights into the origins of these developmental mechanisms. Results In the anthozoan cnidarian Nematostella vectensis, initial archenteron formation begins with bottle cell-induced buckling of the blastula epithelium at the animal pole. Here, we show that bottle cell formation and initial gut invagination in Nematostella requires NvStrabismus (NvStbm), a maternally-expressed core component of the Wnt/Planar Cell Polarity (PCP) pathway. The NvStbm protein is localized to the animal pole of the zygote, remains asymmetrically expressed through the cleavage stages, and becomes restricted to the apical side of invaginating bottle cells at the blastopore. Antisense morpholino-mediated NvStbm-knockdown blocks bottle cell formation and initial archenteron invagination, but it has no effect on Wnt/ß-catenin signaling-mediated endoderm cell fate specification. Conversely, selectively blocking Wnt/ß-catenin signaling inhibits endoderm cell fate specification but does not affect bottle cell formation and initial archenteron invagination. Conclusions Our results demonstrate that Wnt/PCP-mediated initial archenteron invagination can be uncoupled from Wnt/ß-catenin-mediated endoderm cell fate specification in Nematostella, and provides evidence

  18. Wnt5a signaling is a substantial constituent in bone morphogenetic protein-2-mediated osteoblastogenesis

    SciTech Connect

    Nemoto, Eiji; Ebe, Yukari; Kanaya, Sousuke; Tsuchiya, Masahiro; Nakamura, Takashi; Tamura, Masato; Shimauchi, Hidetoshi

    2012-06-15

    Highlights: Black-Right-Pointing-Pointer Wnt5a is identified in osteoblasts in tibial growth plate and bone marrow. Black-Right-Pointing-Pointer Osteoblastic differentiation is associated with increased expression of Wnt5a/Ror2. Black-Right-Pointing-Pointer Wnt5a/Ror2 signaling is important for BMP-2-mediated osteoblastic differentiation. Black-Right-Pointing-Pointer Wnt5a/Ror2 operates independently of BMP-Smad pathway. -- Abstract: Wnts are secreted glycoproteins that mediate developmental and post-developmental physiology by regulating cellular processes including proliferation, differentiation, and apoptosis through {beta}-catenin-dependent canonical and {beta}-catenin-independent noncanonical pathway. It has been reported that Wnt5a activates noncanonical Wnt signaling through receptor tyrosine kinase-like orphan receptor 2 (Ror2). Although it appears that Wnt5a/Ror2 signaling supports normal bone physiology, the biological significance of noncanonical Wnts in osteogenesis is essentially unknown. In this study, we identified expression of Wnt5a in osteoblasts in the ossification zone of the tibial growth plate as well as bone marrow of the rat tibia as assessed by immunohistochemistry. In addition, we show that osteoblastic differentiation mediated by BMP-2 is associated with increased expression of Wnt5a and Ror2 using cultured pre-osteoblasts, MC3T3-E1 cells. Silencing gene expression of Wnt5a and Ror2 in MC3T3-E1 cells results in suppression of BMP-2-mediated osteoblastic differentiation, suggesting that Wnt5a and Ror2 signaling are of substantial importance for BMP-2-mediated osteoblastic differentiation. BMP-2 stimulation induced phosphorylation of Smad1/5/8 in a similar fashion in both siWnt5a-treated cells and control cells, suggesting that Wnt5a was dispensable for the phosphorylation of Smads by BMP-2. Taken together, our results suggest that Wnt5a/Ror2 signaling appears to be involved in BMP-2-mediated osteoblast differentiation in a Smad independent

  19. Wnt Signaling in Cancer Stem Cell Biology

    PubMed Central

    de Sousa e Melo, Felipe; Vermeulen, Louis

    2016-01-01

    Aberrant regulation of Wnt signaling is a common theme seen across many tumor types. Decades of research have unraveled the epigenetic and genetic alterations that result in elevated Wnt pathway activity. More recently, it has become apparent that Wnt signaling levels identify stem-like tumor cells that are responsible for fueling tumor growth. As therapeutic targeting of these tumor stem cells is an intense area of investigation, a concise understanding on how Wnt activity relates to cancer stem cell traits is needed. This review attempts at summarizing the intricacies between Wnt signaling and cancer stem cell biology with a special emphasis on colorectal cancer. PMID:27355964

  20. Visualizing Wnt Palmitoylation in Single Cells.

    PubMed

    Gao, Xinxin; Hannoush, Rami N

    2016-01-01

    Wnt palmitoylation regulates its secretion and signaling activity in cells. Methods to monitor cellular Wnt palmitoylation are instrumental in investigating Wnt activity, secretion, and its interaction with cellular membrane compartments. This protocol describes a method we have recently developed to detect cellular Wnt palmitoylation. The method, combining click chemistry, bio-orthogonal fatty acid probes, and proximity ligation assay (PLA), provides high sensitivity and subcellular resolution for detection of Wnt palmitoylation. It is also compatible with multiple imaging platforms, and is applicable to detecting palmitoylated forms of other fatty acylated proteins. PMID:27590146

  1. Wnt isoform-specific interactions with coreceptor specify inhibition or potentiation of signaling by LRP6 antibodies.

    PubMed

    Gong, Yan; Bourhis, Eric; Chiu, Cecilia; Stawicki, Scott; DeAlmeida, Venita I; Liu, Bob Y; Phamluong, Khanhky; Cao, Tim C; Carano, Richard A D; Ernst, James A; Solloway, Mark; Rubinfeld, Bonnee; Hannoush, Rami N; Wu, Yan; Polakis, Paul; Costa, Mike

    2010-01-01

    β-Catenin-dependent Wnt signaling is initiated as Wnt binds to both the receptor FZD and coreceptor LRP5/6, which then assembles a multimeric complex at the cytoplasmic membrane face to recruit and inactivate the kinase GSK3. The large number and sequence diversity of Wnt isoforms suggest the possibility of domain-specific ligand-coreceptor interactions, and distinct binding sites on LRP6 for Wnt3a and Wnt9b have recently been identified in vitro. Whether mechanistically different interactions between Wnts and coreceptors might mediate signaling remains to be determined. It is also not clear whether coreceptor homodimerization induced extracellularly can activate Wnt signaling, as is the case for receptor tyrosine kinases. We generated monoclonal antibodies against LRP6 with the unexpected ability to inhibit signaling by some Wnt isoforms and potentiate signaling by other isoforms. In cell culture, two antibodies characterized further show reciprocal activities on most Wnts, with one antibody antagonizing and the other potentiating. We demonstrate that these antibodies bind to different regions of LRP6 protein, and inhibition of signaling results from blocking Wnt binding. Antibody-mediated dimerization of LRP6 can potentiate signaling only when a Wnt isoform is also able to bind the complex, presumably recruiting FZD. Endogenous autocrine Wnt signaling in different tumor cell lines can be either antagonized or enhanced by the LRP6 antibodies, indicating expression of different Wnt isoforms. As anticipated from the roles of Wnt signaling in cancer and bone development, antibody activities can also be observed in mice for inhibition of tumor growth and in organ culture for enhancement of bone mineral density. Collectively, our results indicate that separate binding sites for different subsets of Wnt isoforms determine the inhibition or potentiation of signaling conferred by LRP6 antibodies. This complexity of coreceptor-ligand interactions may allow for

  2. Two mutations in human BICC1 resulting in Wnt pathway hyperactivity associated with cystic renal dysplasia.

    PubMed

    Kraus, Marine R-C; Clauin, Séverine; Pfister, Yvan; Di Maïo, Massimo; Ulinski, Tim; Constam, Daniel; Bellanné-Chantelot, Christine; Grapin-Botton, Anne

    2012-01-01

    Bicaudal C homologue 1 (Bicc1) knockout in mice causes polycystic kidney disease and pancreas development defects, including a reduction in insulin-producing β-cells and ensuing diabetes. We therefore screened 137 patients with renal abnormalities or association of early-onset diabetes and renal disease for genetic alterations in BICC1. We identified two heterozygous mutations, one nonsense in the first K Homology (KH) domain and one missense in the sterile alpha motif (SAM) domain. In mice, Bicc1 blocks canonical Wnt signaling, mostly via its SAM domain. We show that the human BICC1, similar to its mouse counterpart, blocks canonical Wnt signaling. The nonsense mutation identified results in a complete loss of Wnt inhibitory activity. The point mutation in the SAM domain has a similar effect to a complete SAM domain deletion, resulting in a 22% loss of activity. PMID:21922595

  3. Construction and Experimental Validation of a Petri Net Model of Wnt/β-Catenin Signaling

    PubMed Central

    Heijmans, Nika; Verkaar, Folkert; Smit, Martine J.; Heringa, Jaap

    2016-01-01

    The Wnt/β-catenin signaling pathway is important for multiple developmental processes and tissue maintenance in adults. Consequently, deregulated signaling is involved in a range of human diseases including cancer and developmental defects. A better understanding of the intricate regulatory mechanism and effect of physiological (active) and pathophysiological (hyperactive) WNT signaling is important for predicting treatment response and developing novel therapies. The constitutively expressed CTNNB1 (commonly and hereafter referred to as β-catenin) is degraded by a destruction complex, composed of amongst others AXIN1 and GSK3. The destruction complex is inhibited during active WNT signaling, leading to β-catenin stabilization and induction of β-catenin/TCF target genes. In this study we investigated the mechanism and effect of β-catenin stabilization during active and hyperactive WNT signaling in a combined in silico and in vitro approach. We constructed a Petri net model of Wnt/β-catenin signaling including main players from the plasma membrane (WNT ligands and receptors), cytoplasmic effectors and the downstream negative feedback target gene AXIN2. We validated that our model can be used to simulate both active (WNT stimulation) and hyperactive (GSK3 inhibition) signaling by comparing our simulation and experimental data. We used this experimentally validated model to get further insights into the effect of the negative feedback regulator AXIN2 upon WNT stimulation and observed an attenuated β-catenin stabilization. We furthermore simulated the effect of APC inactivating mutations, yielding a stabilization of β-catenin levels comparable to the Wnt-pathway activities observed in colorectal and breast cancer. Our model can be used for further investigation and viable predictions of the role of Wnt/β-catenin signaling in oncogenesis and development. PMID:27218469

  4. Construction and Experimental Validation of a Petri Net Model of Wnt/β-Catenin Signaling.

    PubMed

    Jacobsen, Annika; Heijmans, Nika; Verkaar, Folkert; Smit, Martine J; Heringa, Jaap; van Amerongen, Renée; Feenstra, K Anton

    2016-01-01

    The Wnt/β-catenin signaling pathway is important for multiple developmental processes and tissue maintenance in adults. Consequently, deregulated signaling is involved in a range of human diseases including cancer and developmental defects. A better understanding of the intricate regulatory mechanism and effect of physiological (active) and pathophysiological (hyperactive) WNT signaling is important for predicting treatment response and developing novel therapies. The constitutively expressed CTNNB1 (commonly and hereafter referred to as β-catenin) is degraded by a destruction complex, composed of amongst others AXIN1 and GSK3. The destruction complex is inhibited during active WNT signaling, leading to β-catenin stabilization and induction of β-catenin/TCF target genes. In this study we investigated the mechanism and effect of β-catenin stabilization during active and hyperactive WNT signaling in a combined in silico and in vitro approach. We constructed a Petri net model of Wnt/β-catenin signaling including main players from the plasma membrane (WNT ligands and receptors), cytoplasmic effectors and the downstream negative feedback target gene AXIN2. We validated that our model can be used to simulate both active (WNT stimulation) and hyperactive (GSK3 inhibition) signaling by comparing our simulation and experimental data. We used this experimentally validated model to get further insights into the effect of the negative feedback regulator AXIN2 upon WNT stimulation and observed an attenuated β-catenin stabilization. We furthermore simulated the effect of APC inactivating mutations, yielding a stabilization of β-catenin levels comparable to the Wnt-pathway activities observed in colorectal and breast cancer. Our model can be used for further investigation and viable predictions of the role of Wnt/β-catenin signaling in oncogenesis and development. PMID:27218469

  5. TGF-β-induced profibrotic signaling is regulated in part by the WNT receptor Frizzled-8.

    PubMed

    Spanjer, Anita I R; Baarsma, Hoeke A; Oostenbrink, Lisette M; Jansen, Sepp R; Kuipers, Christine C; Lindner, Michael; Postma, Dirkje S; Meurs, Herman; Heijink, Irene H; Gosens, Reinoud; Königshoff, Melanie

    2016-05-01

    TGF-β is important in lung injury and remodeling processes. TGF-β and Wingless/integrase-1 (WNT) signaling are interconnected; however, the WNT ligand-receptor complexes involved are unknown. Thus, we aimed to identify Frizzled (FZD) receptors that mediate TGF-β-induced profibrotic signaling. MRC-5 and primary human lung fibroblasts were stimulated with TGF-β1, WNT-5A, or WNT-5B in the presence and absence of specific pathway inhibitors. Specific small interfering RNA was used to knock down FZD8. In vivo studies using bleomycin-induced lung fibrosis were performed in wild-type and FZD8-deficient mice. TGF-β1 induced FZD8 specifically via Smad3-dependent signaling in MRC-5 and primary human lung fibroblasts. It is noteworthy that FZD8 knockdown reduced TGF-β1-induced collagen Iα1, fibronectin, versican, α-smooth muscle (sm)-actin, and connective tissue growth factor. Moreover, bleomycin-induced lung fibrosis was attenuated in FZD8-deficient mice in vivo Although inhibition of canonical WNT signaling did not affect TGF-β1-induced gene expression in vitro, noncanonical WNT-5B mimicked TGF-β1-induced fibroblast activation. FZD8 knockdown reduced both WNT-5B-induced gene expression of fibronectin and α-sm-actin, as well as WNT-5B-induced changes in cellular impedance. Collectively, our findings demonstrate a role for FZD8 in TGF-β-induced profibrotic signaling and imply that WNT-5B may be the ligand for FZD8 in these responses.-Spanjer, A. I. R., Baarsma, H. A., Oostenbrink, L. M., Jansen, S. R., Kuipers, C. C., Lindner, M., Postma, D. S., Meurs, H., Heijink, I. H., Gosens, R., Königshoff, M. TGF-β-induced profibrotic signaling is regulated in part by the WNT receptor Frizzled-8. PMID:26849959

  6. WNT signaling in glioblastoma and therapeutic opportunities.

    PubMed

    Lee, Yeri; Lee, Jin-Ku; Ahn, Sun Hee; Lee, Jeongwu; Nam, Do-Hyun

    2016-02-01

    WNTs and their downstream effectors regulate proliferation, death, and migration and cell fate decision. Deregulation of WNT signaling is associated with various cancers including GBM, which is the most malignant primary brain cancer. In this review, we will summarize the experimental evidence supporting oncogenic roles of WNT signaling in GBM and discuss current progress in the targeting of WNT signaling as an anti-cancer approach. In particular, we will focus on (1) genetic and epigenetic alterations that lead to aberrant WNT pathway activation in GBM, (2) WNT-mediated control of GBM stem cell maintenance and invasion, and (3) cross-talk between WNT and other signaling pathways in GBM. We will then review the discovery of agents that can inhibit WNT signaling in preclinical models and the current status of human clinical trials. PMID:26641068

  7. A progress report for the large block test of the coupled thermal-mechanical-hydrological-chemical processes

    SciTech Connect

    Lin, W.; Wilder, D.G.; Blink, J.

    1994-10-01

    This is a progress report on the Large Block Test (LBT) project. The purpose of the LBT is to study some of the coupled thermal-mechanical-hydrological-chemical (TMHC) processes in the near field of a nuclear waste repository under controlled boundary conditions. To do so, a large block of Topopah Spring tuff will be heated from within for about 4 to 6 months, then cooled down for about the same duration. Instruments to measure temperature, moisture content, stress, displacement, and chemical changes will be installed in three directions in the block. Meanwhile, laboratory tests will be conducted on small blocks to investigate individual thermal-mechanical, thermal-hydrological, and thermal-chemical processes. The fractures in the large block will be characterized from five exposed surfaces. The minerals on fracture surfaces will be studied before and after the test. The results from the LBT will be useful for testing and building confidence in models that will be used to predict TMHC processes in a repository. The boundary conditions to be controlled on the block include zero moisture flux and zero heat flux on the sides, constant temperature on the top, and constant stress on the outside surfaces of the block. To control these boundary conditions, a load-retaining frame is required. A 3 x 3 x 4.5 m block of Topopah Spring tuff has been isolated on the outcrop at Fran Ridge, Nevada Test Site. Pre-test model calculations indicate that a permeability of at least 10{sup -15} m{sup 2} is required so that a dryout zone can be created within a practical time frame when the block is heated from within. Neutron logging was conducted in some of the vertical holes to estimate the initial moisture content of the block. It was found that about 60 to 80% of the pore volume of the block is saturated with water. Cores from the vertical holes have been used to map the fractures and to determine the properties of the rock. A current schedule is included in the report.

  8. Wnt5a and Wnt11 regulate mammalian anterior-posterior axis elongation

    PubMed Central

    Andre, Philipp; Song, Hai; Kim, Wantae; Kispert, Andreas; Yang, Yingzi

    2015-01-01

    Mesoderm formation and subsequent anterior-posterior (A-P) axis elongation are fundamental aspects of gastrulation, which is initiated by formation of the primitive streak (PS). Convergent extension (CE) movements and epithelial-mesenchymal transition (EMT) are important for A-P axis elongation in vertebrate embryos. The evolutionarily conserved planar cell polarity (PCP) pathway regulates CE, and Wnts regulate many aspects of gastrulation including CE and EMT. However, the Wnt ligands that regulate A-P axis elongation in mammalian development remain unknown. Wnt11 and Wnt5a regulate axis elongation in lower vertebrates, but only Wnt5a, not Wnt11, regulates mammalian PCP signaling and A-P axis elongation in development. Here, by generating Wnt5a; Wnt11 compound mutants, we show that Wnt11 and Wnt5a play redundant roles during mouse A-P axis elongation. Both genes regulate trunk notochord extension through PCP-controlled CE of notochord cells, establishing a role for Wnt11 in mammalian PCP. We show that Wnt5a and Wnt11 are required for proper patterning of the neural tube and somites by regulating notochord formation, and provide evidence that both genes are required for the generation and migration of axial and paraxial mesodermal precursor cells by regulating EMT. Axial and paraxial mesodermal precursors ectopically accumulate in the PS at late gastrula stages in Wnt5a−/−; Wnt11−/− embryos and these cells ectopically express epithelial cell adhesion molecules. Our data suggest that Wnt5a and Wnt11 regulate EMT by inducing p38 (Mapk14) phosphorylation. Our findings provide new insights into the role of Wnt5a and Wnt11 in mouse early development and also in cancer metastasis, during which EMT plays a crucial role. PMID:25813538

  9. JNK signalling is necessary for a Wnt- and stem cell-dependent regeneration programme

    PubMed Central

    Tejada-Romero, Belen; Carter, Jean-Michel; Mihaylova, Yuliana; Neumann, Bjoern; Aboobaker, A. Aziz

    2015-01-01

    Regeneration involves the integration of new and old tissues in the context of an adult life history. It is clear that the core conserved signalling pathways that orchestrate development also play central roles in regeneration, and further study of conserved signalling pathways is required. Here we have studied the role of the conserved JNK signalling cascade during planarian regeneration. Abrogation of JNK signalling by RNAi or pharmacological inhibition blocks posterior regeneration and animals fail to express posterior markers. While the early injury-induced expression of polarity markers is unaffected, the later stem cell-dependent phase of posterior Wnt expression is not established. This defect can be rescued by overactivation of the Hh or Wnt signalling pathway to promote posterior Wnt activity. Together, our data suggest that JNK signalling is required to establish stem cell-dependent Wnt expression after posterior injury. Given that Jun is known to be required in vertebrates for the expression of Wnt and Wnt target genes, we propose that this interaction may be conserved and is an instructive part of planarian posterior regeneration. PMID:26062938

  10. Wnt-mediated reciprocal regulation between cartilage and bone development during endochondral ossification.

    PubMed

    Lu, Cheng; Wan, Yong; Cao, Jingjing; Zhu, Xuming; Yu, Jian; Zhou, Rujiang; Yao, Yiyun; Zhang, Lingling; Zhao, Haixia; Li, Hanjun; Zhao, Jianzhi; He, Lin; Ma, Gang; Yang, Xiao; Yao, Zhengju; Guo, Xizhi

    2013-04-01

    The role of Wnt signaling is extensively studied in skeletal development and postnatal bone remodeling, mostly based on the genetic approaches of β-catenin manipulation. However, given their independent function, a requirement for β-catenin is not the same as that for Wnt. Here, we investigated the effect of Wnt proteins in both tissues through generating cartilage- or bone-specific Wls null mice, respectively. Depletion of Wls by Col2-Cre, which would block Wnt secretion in the chondrocytes and perichondrium, delayed chondrocyte hypertrophy in the growth plate and impaired perichondrial osteogenesis. Loss of Wls in chondrocytes also disturbed the proliferating chondrocyte morphology and division orientation, which was similar to the defect observed in Wnt5a null mice. On the other hand, inactivation of Wls in osteoblasts by Col1-Cre resulted in a shorter hypertrophic zone and an increase of TRAP positive cell number in the chondro-osseous junction of growth plate, coupled with a decrease in bone mass. Taken together, our studies reveal that Wnt proteins not only modulate differentiation and cellular communication within populations of chondrocytes, but also mediate the cross regulation between the chondrocytes and osteoblasts in growth plate. PMID:23274346

  11. JNK signalling is necessary for a Wnt- and stem cell-dependent regeneration programme.

    PubMed

    Tejada-Romero, Belen; Carter, Jean-Michel; Mihaylova, Yuliana; Neumann, Bjoern; Aboobaker, A Aziz

    2015-07-15

    Regeneration involves the integration of new and old tissues in the context of an adult life history. It is clear that the core conserved signalling pathways that orchestrate development also play central roles in regeneration, and further study of conserved signalling pathways is required. Here we have studied the role of the conserved JNK signalling cascade during planarian regeneration. Abrogation of JNK signalling by RNAi or pharmacological inhibition blocks posterior regeneration and animals fail to express posterior markers. While the early injury-induced expression of polarity markers is unaffected, the later stem cell-dependent phase of posterior Wnt expression is not established. This defect can be rescued by overactivation of the Hh or Wnt signalling pathway to promote posterior Wnt activity. Together, our data suggest that JNK signalling is required to establish stem cell-dependent Wnt expression after posterior injury. Given that Jun is known to be required in vertebrates for the expression of Wnt and Wnt target genes, we propose that this interaction may be conserved and is an instructive part of planarian posterior regeneration. PMID:26062938

  12. The Wnt/beta-catenin pathway regulates Gli-mediated Myf5 expression during somitogenesis.

    PubMed

    Borello, Ugo; Berarducci, Barbara; Murphy, Paula; Bajard, Lola; Buffa, Viviana; Piccolo, Stefano; Buckingham, Margaret; Cossu, Giulio

    2006-09-01

    Canonical Wnt/beta-catenin signaling regulates the activation of the myogenic determination gene Myf5 at the onset of myogenesis, but the underlying molecular mechanism is unknown. Here, we report that the Wnt signal is transduced in muscle progenitor cells by at least two Frizzled (Fz) receptors (Fz1 and/or Fz6), through the canonical beta-catenin pathway, in the epaxial domain of newly formed somites. We show that Myf5 activation is dramatically reduced by blocking the Wnt/beta-catenin pathway in somite progenitor cells, whereas expression of activated beta-catenin is sufficient to activate Myf5 in somites but not in the presomitic mesoderm. In addition, we identified Tcf/Lef sequences immediately 5' to the Myf5 early epaxial enhancer. These sites determine the correct spatiotemporal expression of Myf5 in the epaxial domain of the somite, mediating the synergistic action of the Wnt/beta-catenin and the Shh/Gli pathways. Taken together, these results demonstrate that Myf5 is a direct target of Wnt/beta-catenin, and that its full activation requires a cooperative interaction between the canonical Wnt and the Shh/Gli pathways in muscle progenitor cells. PMID:16936075

  13. Processing-structure-mechanical Property Relationships of Semicrystalline Polyolefin-based Block Copolymers

    SciTech Connect

    Deplace, F.; Wang, Z; Lynd, N; Hotta, A; Rose, J; Hustad, P; Tian, J; Ohtaki, H; Coates, J; et. al.

    2010-01-01

    The incremental plastic deformation of the crystals of block copolymers made of semicrystalline polypropylene endblocks and amorphous ethylene-r-propylene midblocks occurring during step cycle tensile tests has dramatic effects on the stress-strain curves. This can be understood from the evolution of the morphology and of the microstructure of the crystalline blocks revealed by X-ray scattering experiments.

  14. Butyrate and bioactive proteolytic form of Wnt-5a regulate colonic epithelial proliferation and spatial development.

    PubMed

    Uchiyama, Kazuhiko; Sakiyama, Toshio; Hasebe, Takumu; Musch, Mark W; Miyoshi, Hiroyuki; Nakagawa, Yasushi; He, Tong-Chuan; Lichtenstein, Lev; Naito, Yuji; Itoh, Yoshito; Yoshikawa, Toshikazu; Jabri, Bana; Stappenbeck, Thaddeus; Chang, Eugene B

    2016-01-01

    Proliferation and spatial development of colonic epithelial cells are highly regulated along the crypt vertical axis, which, when perturbed, can result in aberrant growth and carcinogenesis. In this study, two key factors were identified that have important and counterbalancing roles regulating these processes: pericrypt myofibroblast-derived Wnt-5a and the microbial metabolite butyrate. Cultured YAMC cell proliferation and heat shock protein induction were analzyed after butryate, conditioned medium with Wnt5a activity, and FrzB containing conditioned medium. In vivo studies to modulate Hsp25 employed intra-colonic wall Hsp25 encoding lentivirus. To silence Wnt-5a in vivo, intra-colonic wall Wnt-5a silencing RNA was used. Wnt-5a, secreted by stromal myofibroblasts of the lower crypt, promotes proliferation through canonical β-catenin activation. Essential to this are two key requirements: (1) proteolytic conversion of the highly insoluble ~40 kD Wnt-5a protein to a soluble 36 mer amino acid peptide that activates epithelial β-catenin and cellular proliferation, and (2) the simultaneous inhibition of butyrate-induced Hsp25 by Wnt-5a which is necessary to arrest the proliferative process in the upper colonic crypt. The interplay and spatial gradients of these factors insures that crypt epithelial cell proliferation and development proceed in an orderly fashion, but with sufficient plasticity to adapt to physiological perturbations including inflammation. PMID:27561676

  15. Butyrate and bioactive proteolytic form of Wnt-5a regulate colonic epithelial proliferation and spatial development

    PubMed Central

    Uchiyama, Kazuhiko; Sakiyama, Toshio; Hasebe, Takumu; Musch, Mark W.; Miyoshi, Hiroyuki; Nakagawa, Yasushi; He, Tong-Chuan; Lichtenstein, Lev; Naito, Yuji; Itoh, Yoshito; Yoshikawa, Toshikazu; Jabri, Bana; Stappenbeck, Thaddeus; Chang, Eugene B.

    2016-01-01

    Proliferation and spatial development of colonic epithelial cells are highly regulated along the crypt vertical axis, which, when perturbed, can result in aberrant growth and carcinogenesis. In this study, two key factors were identified that have important and counterbalancing roles regulating these processes: pericrypt myofibroblast-derived Wnt-5a and the microbial metabolite butyrate. Cultured YAMC cell proliferation and heat shock protein induction were analzyed after butryate, conditioned medium with Wnt5a activity, and FrzB containing conditioned medium. In vivo studies to modulate Hsp25 employed intra-colonic wall Hsp25 encoding lentivirus. To silence Wnt-5a in vivo, intra-colonic wall Wnt-5a silencing RNA was used. Wnt-5a, secreted by stromal myofibroblasts of the lower crypt, promotes proliferation through canonical β-catenin activation. Essential to this are two key requirements: (1) proteolytic conversion of the highly insoluble ~40 kD Wnt-5a protein to a soluble 36 mer amino acid peptide that activates epithelial β-catenin and cellular proliferation, and (2) the simultaneous inhibition of butyrate-induced Hsp25 by Wnt-5a which is necessary to arrest the proliferative process in the upper colonic crypt. The interplay and spatial gradients of these factors insures that crypt epithelial cell proliferation and development proceed in an orderly fashion, but with sufficient plasticity to adapt to physiological perturbations including inflammation. PMID:27561676

  16. A multiscale modeling study of loss processes in block-copolymer-based solar cell nanodevices

    NASA Astrophysics Data System (ADS)

    Donets, Sergii; Pershin, Anton; Christlmaier, Martin J. A.; Baeurle, Stephan A.

    2013-03-01

    Flexible photovoltaic devices possess promising perspectives in opto-electronic technologies, where high mobility and/or large-scale applicability are important. However, their usefulness in such applications is currently still limited due to the low level of optimization of their performance and durability. For the improvement of these properties, a better understanding and control of small-scale annihilation phenomena involved in the photovoltaic process, such as exciton loss and charge carrier loss, is necessary, which typically implicates multiple length- and time-scales. Here, we study the causes for their occurrence on the example of nanostructured diblock- and triblock-copolymer systems by making use of a novel solar-cell simulation algorithm and explore new routes to optimize their photovoltaic properties. A particular focus is set on the investigation of exciton and charge carrier loss phenomena and their dependence on the inter-monomeric interaction strength, chain architecture, and external mechanical loading. Our simulation results reveal that in the regime from low up to intermediate χ-parameters an increasing number of continuous percolation paths is created. In this parameter range, the internal quantum efficiency (IQE) increases up to a maximum, characterized by a minimum in the number of charge losses due to charge recombination. In the regime of high χ-parameters both block-copolymer systems form nanostructures with a large number of bottlenecks and dead ends. These lead to a large number of charge losses due to charge recombination, charge trapping, and a deteriorated exciton dissociation, resulting in a significant drop in the IQE. Moreover, we find that the photovoltaic performance of the triblock-copolymer material decreases with increasing mechanical loading, caused by a growing number of charge losses due to charge recombination and charge accumulation. Finally, we demonstrate that the process of charge trapping in defects can be reversed

  17. Wnt signaling-mediated redox regulation maintains the germ line stem cell differentiation niche

    PubMed Central

    Wang, Su; Gao, Yuan; Song, Xiaoqing; Ma, Xing; Zhu, Xiujuan; Mao, Ying; Yang, Zhihao; Ni, Jianquan; Li, Hua; Malanowski, Kathryn E; Anoja, Perera; Park, Jungeun; Haug, Jeff; Xie, Ting

    2015-01-01

    Adult stem cells continuously undergo self-renewal and generate differentiated cells. In the Drosophila ovary, two separate niches control germ line stem cell (GSC) self-renewal and differentiation processes. Compared to the self-renewing niche, relatively little is known about the maintenance and function of the differentiation niche. In this study, we show that the cellular redox state regulated by Wnt signaling is critical for the maintenance and function of the differentiation niche to promote GSC progeny differentiation. Defective Wnt signaling causes the loss of the differentiation niche and the upregulated BMP signaling in differentiated GSC progeny, thereby disrupting germ cell differentiation. Mechanistically, Wnt signaling controls the expression of multiple glutathione-S-transferase family genes and the cellular redox state. Finally, Wnt2 and Wnt4 function redundantly to maintain active Wnt signaling in the differentiation niche. Therefore, this study has revealed a novel strategy for Wnt signaling in regulating the cellular redox state and maintaining the differentiation niche. DOI: http://dx.doi.org/10.7554/eLife.08174.001 PMID:26452202

  18. Distinctive microRNA signature associated of neoplasms with the Wnt/β-catenin signaling pathway.

    PubMed

    Sun, Xu; He, Yong; Huang, Cheng; Ma, Tao-Tao; Li, Jun

    2013-12-01

    As the crucial biological regulators, microRNAs that act by suppressing their target genes are involved in a variety of pathophysiological processes. It is generally accepted that microRNAs are often dysregulated in many types of neoplasm and other human diseases. In neoplasm, microRNAs may function as oncogenes or tumor suppressors. As constitutive activation of the Wnt signaling pathway is a common feature of neoplasm and contributes to its development, progression and metastasis in various cancers, numerous studies have revealed that microRNA-mediated gene regulation are interconnected with the Wnt/β-catenin signaling pathway, forming a Wnt/β-catenin-microRNA regulatory network, which is critical to successful targeting of the Wnt/β-catenin pathway for oncotherapy. In this review, we aim to accumulate recent advances on microRNAs that work in tandem with Wnt/β-catenin signaling in tumorigenesis, with particular focus on how microRNAs affect Wnt/β-catenin activity as well as how microRNAs are regulated through the Wnt/β-catenin pathway. PMID:24041653

  19. Military Curricula for Vocational & Technical Education. Metals Processing Specialist, Blocks V and VI, Classroom Course 13-7.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. National Center for Research in Vocational Education.

    These curriculum materials are the third section of a four-part, secondary-postsecondary-level course in metals processing. The course is one of a number of military-developed curriculum packages selected for adaptation to vocational instruction and curriculum development in a civilian setting. Block V, Inert Gas Shielded Welding of High…

  20. Military Curricula for Vocational & Technical Education. Metals Processing Specialist, Blocks III and IV, Classroom Course 13-6.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. National Center for Research in Vocational Education.

    These curriculum materials are the second section of a four-part, secondary-postsecondary-level course in metals processing. The course is one of a number of military-developed curriculum packages selected for adaptation to vocational instruction and curriculum development in a civilian setting. Block III, Introduction to Metallic Arc Welding,…

  1. Military Curricula for Vocational & Technical Education. Metals Processing Specialist, Blocks I and II, Classroom Course 13-5.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. National Center for Research in Vocational Education.

    These curriculum materials are the first section of a four-part, secondary-postsecondary-level course in metals processing. The course is one of a number of military-developed curriculum packages selected for adaptation to vocational instruction and curriculum development in a civilian setting. Block I, Introduction to Oxyacetylene Welding,…

  2. Noncanonical Wnt5a enhances Wnt/β-catenin signaling during osteoblastogenesis.

    PubMed

    Okamoto, Masanori; Udagawa, Nobuyuki; Uehara, Shunsuke; Maeda, Kazuhiro; Yamashita, Teruhito; Nakamichi, Yuko; Kato, Hiroyuki; Saito, Naoto; Minami, Yasuhiro; Takahashi, Naoyuki; Kobayashi, Yasuhiro

    2014-01-01

    Wnt regulates bone formation through β-catenin-dependent canonical and -independent noncanonical signaling pathways. However, the cooperation that exists between the two signaling pathways during osteoblastogenesis remains to be elucidated. Here, we showed that the lack of Wnt5a in osteoblast-lineage cells impaired Wnt/β-catenin signaling due to the reduced expression of Lrp5 and Lrp6. Pretreatment of ST2 cells, a stromal cell line, with Wnt5a enhanced canonical Wnt ligand-induced Tcf/Lef transcription activity. Short hairpin RNA-mediated knockdown of Wnt5a, but not treatment with Dkk1, an antagonist of Wnt/β-catenin signaling, reduced the expression of Lrp5 and Lrp6 in osteoblast-lineage cells under osteogenic culture conditions. Osteoblast-lineage cells from Wnt5a-deficient mice exhibited reduced Wnt/β-catenin signaling, which impaired osteoblast differentiation and enhanced adipocyte differentiation. Adenovirus-mediated gene transfer of Lrp5 into Wnt5a-deficient osteoblast-lineage cells rescued their phenotypic features. Therefore, Wnt5a-induced noncanonical signaling cooperates with Wnt/β-catenin signaling to achieve proper bone formation. PMID:24670389

  3. Primary Cilia Integrate Hedgehog and Wnt Signaling during Tooth Development

    PubMed Central

    Liu, B.; Chen, S.; Cheng, D.; Jing, W.; Helms, J.A.

    2014-01-01

    Many ciliopathies have clinical features that include tooth malformations but how these defects come about is not clear. Here we show that genetic deletion of the motor protein Kif3a in dental mesenchyme results in an arrest in odontogenesis. Incisors are completely missing, and molars are enlarged in Wnt1Cre+Kif3afl/fl embryos. Although amelogenesis and dentinogenesis initiate in the molar tooth bud, both processes terminate prematurely. We demonstrate that loss of Kif3a in dental mesenchyme results in loss of Hedgehog signaling and gain of Wnt signaling in this same tissue. The defective dental mesenchyme then aberrantly signals to the dental epithelia, which prompts an up-regulation in the Hedgehog and Wnt responses in the epithelia and leads to multiple attempts at invagination and an expanded enamel organ. Thus, the primary cilium integrates Hedgehog and Wnt signaling between dental epithelia and mesenchyme, and this cilia-dependent integration is required for proper tooth development. PMID:24659776

  4. Immune Modulation of the T Cell Response in Asthma through Wnt10b.

    PubMed

    Trischler, Jordis; Shiomi, Takayuki; Turner, Damian L; Sklepkiewicz, Piotr L; Goldklang, Monica P; Tanaka, Kenji F; Xu, Ming; Farber, Donna L; D'Armiento, Jeanine M

    2016-04-01

    Asthma is a chronic inflammatory disease, which is characterized by activation of CD4(+) T helper 2 cells orchestrating an allergic airway response. Whereas the role of Wnt family members in regulating T cell maintenance and maturation is established, their contribution to T cell activation in allergic asthma is not known. We hypothesized that Wnt10b plays a role in the modulation of the allergic airway response and affects T cell activation and polarization. Using an in vivo house dust mite asthma model, Wnt10b-deficient (Wnt10b(-/-)) mice were allergen-sensitized and inflammation, as well as T cell activation, was studied in vivo and in vitro. Wnt10b(-/-) mice exhibited an augmented inflammatory phenotype with an increase in eosinophils in the bronchoalveolar lavage and IL-4 and IL-13 in the lungs when compared with wild-type mice. In vitro studies confirmed an increased T helper type 2 polarization and increased T cell activation of Wnt10b(-/-) cells. Accordingly, the percentage of naive T cells was elevated by the addition of recombinant Wnt10b protein. Finally, Wnt10b(-/-) mice exhibited an increase in the percentage of effector T cells in the lungs after house dust mite sensitization, which indicated a heightened activation state, measured by an increased percentage of CD69(hi)CD11a(hi) cells. These findings suggest that Wnt10b plays an important role in regulating asthmatic airway inflammation through modification of the T cell response and is a prospective target in the disease process. PMID:26436894

  5. Epigenetic mediated transcriptional activation of WNT5A participates in arsenical-associated malignant transformation

    SciTech Connect

    Jensen, Taylor J.; Wozniak, Ryan J.; Eblin, Kylee E.; Wnek, Sean M.; Gandolfi, A. Jay; Futscher, Bernard W.

    2009-02-15

    Arsenic is a human carcinogen with exposure associated with cancer of the lung, skin, and bladder. Many potential mechanisms have been implicated as playing a role in the process of arsenical-induced malignancy including the perturbation of signaling pathways and aberrant epigenetic regulation. We initiated studies to examine the role of a member of the non-canonical WNT signaling pathway, WNT5A, in UROtsa cells and arsenite [URO-ASSC] and monomethylarsonous acid [URO-MSC] malignantly transformed variants. We present data herein that suggest that WNT5A is transcriptionally activated during arsenical-induced malignant transformation. This WNT5A transcriptional activation is correlated with the enrichment of permissive histone modifications and the reduction of repressive modifications in the WNT5A promoter region. The epigenetic activation of WNT5A expression and acetylation of its promoter remain after the removal of the arsenical, consistent with the maintenance of an anchorage independent growth phenotype in these cells. Additionally, treatment with epigenetic modifying drugs supports a functional role for these epigenetic marks in controlling gene expression. Reduction of WNT5A using lentiviral shRNA greatly attenuated the ability of these cells to grow in an anchorage independent fashion. Extension of our model into human bladder cancer cell lines indicates that each of the cell lines examined also express WNT5A. Taken together, these data suggest that the epigenetic remodeling of the WNT5A promoter is correlated with its transcriptional activation and this upregulation likely participates in arsenical-induced malignant transformation.

  6. Wnt5a and Ror2 expression associate with the disease progress of primary thyroid lymphoma.

    PubMed

    Wang, Lei; Yang, Dong; Wang, Ying-Hou; Li, Xi; Gao, Hong-Ming; Lv, Jun-Yuan; Wang, Lei; Xin, Shi-Jie

    2016-05-01

    Primary thyroid lymphoma (PTL) is a rare malignant thyroid tumor; its pathogenesis is closely related to chronic lymphocytic thyroiditis. The different pathological subtypes and stages of PTL have distinct clinical characteristics and prognosis, but the specific reasons are not clear. Wnt5a is a representative protein of non-canonical Wnt signaling. It plays an important role in many different types of tumors. This study is to explore the changes of Wnt5a and its receptor Ror2 in PTL development process and the clinical significance of their represent. We collected 22 PTL patient tumor specimens and clinical data. We observed the expression of Wnt5a and Ror2 in PTL tumor tissues by immunohistochemistry. Wnt5a was expressed positively in 12 (54.5 %) cases, and Ror2 was expressed positively in 18 (81.8 %) cases. The expression of Wnt5a had a significant difference in different pathological subtypes of PTL (P < 0.05). Wnt5a and Ror2 expression were associated with local invasion and clinical stage, respectively (P < 0.05), and had no significant correlation with age, gender, and tumor size. Although, no significant difference in overall survival was found between positive and negative groups of Wnt5a (P = 0.416) or Ror2 (P = 0.256), respectively. We still consider that Wnt5a and Ror2 play a complex and subtle role in the pathogenesis and progression of PTL and may become potential biomarkers and therapeutic targets of PTL. PMID:26614433

  7. Wnt5a and Ror2 expression associate with the disease progress of primary thyroid lymphoma.

    PubMed

    Wang, Lei; Yang, Dong; Wang, Ying-Hou; Li, Xi; Gao, Hong-Ming; Lv, Jun-Yuan; Wang, Lei; Xin, Shi-Jie

    2016-05-01

    Primary thyroid lymphoma (PTL) is a rare malignant thyroid tumor; its pathogenesis is closely related to chronic lymphocytic thyroiditis. The different pathological subtypes and stages of PTL have distinct clinical characteristics and prognosis, but the specific reasons are not clear. Wnt5a is a representative protein of non-canonical Wnt signaling. It plays an important role in many different types of tumors. This study is to explore the changes of Wnt5a and its receptor Ror2 in PTL development process and the clinical significance of their represent. We collected 22 PTL patient tumor specimens and clinical data. We observed the expression of Wnt5a and Ror2 in PTL tumor tissues by immunohistochemistry. Wnt5a was expressed positively in 12 (54.5 %) cases, and Ror2 was expressed positively in 18 (81.8 %) cases. The expression of Wnt5a had a significant difference in different pathological subtypes of PTL (P < 0.05). Wnt5a and Ror2 expression were associated with local invasion and clinical stage, respectively (P < 0.05), and had no significant correlation with age, gender, and tumor size. Although, no significant difference in overall survival was found between positive and negative groups of Wnt5a (P = 0.416) or Ror2 (P = 0.256), respectively. We still consider that Wnt5a and Ror2 play a complex and subtle role in the pathogenesis and progression of PTL and may become potential biomarkers and therapeutic targets of PTL. PMID:26608372

  8. Functional analysis of centipede development supports roles for Wnt genes in posterior development and segment generation.

    PubMed

    Hayden, Luke; Schlosser, Gerhard; Arthur, Wallace

    2015-01-01

    The genes of the Wnt family play important and highly conserved roles in posterior growth and development in a wide range of animal taxa. Wnt genes also operate in arthropod segmentation, and there has been much recent debate regarding the relationship between arthropod and vertebrate segmentation mechanisms. Due to its phylogenetic position, body form, and possession of many (11) Wnt genes, the centipede Strigamia maritima is a useful system with which to examine these issues. This study takes a functional approach based on treatment with lithium chloride, which causes ubiquitous activation of canonical Wnt signalling. This is the first functional developmental study performed in any of the 15,000 species of the arthropod subphylum Myriapoda. The expression of all 11 Wnt genes in Strigamia was analyzed in relation to posterior development. Three of these genes, Wnt11, Wnt5, and WntA, were strongly expressed in the posterior region and, thus, may play important roles in posterior developmental processes. In support of this hypothesis, LiCl treatment of S. maritima embryos was observed to produce posterior developmental defects and perturbations in AbdB and Delta expression. The effects of LiCl differ depending on the developmental stage treated, with more severe effects elicited by treatment during germband formation than by treatment at later stages. These results support a role for Wnt signalling in conferring posterior identity in Strigamia. In addition, data from this study are consistent with the hypothesis of segmentation based on a "clock and wavefront" mechanism operating in this species. PMID:25627713

  9. LRP5 negatively regulates differentiation of monocytes through abrogation of Wnt signalling

    PubMed Central

    Borrell-Pagès, Maria; Romero, July Carolina; Badimon, Lina

    2014-01-01

    Molecular changes involved in cell differentiation are only partially known. Circulating inflammatory cells need to differentiate to perform specialized functions in target tissues. Here, we hypothesized that low-density lipoprotein receptor–related protein 5 (LRP5) is involved, through its participation in the canonical Wnt/β-catenin signalling, in the differentiation process of monocytic cells. To this aim, we characterized differentiation mechanisms of HL60 cells and primary human monocytes. We show that silencing the LRP5 gene increased differentiation of HL60 cells and human monocytes, suggesting that LRP5 signalling abrogates differentiation. We demonstrate that the mechanisms behind this blockade include sequestration of β-catenin at the cellular membrane, inhibition of the Wnt signalling and increase of apoptosis. We further demonstrate the involvement of LRP5 and the Wnt/β-catenin signalling in the process because cellular differentiation can be rescued by the addition of downstream Wnt target genes to the monocytic cells. PMID:24266894

  10. List blocking and longer retention intervals reveal an influence of gist processing for lexically ambiguous critical lures

    PubMed Central

    McNabb, Jaimie; Hutchison, Keith A.

    2016-01-01

    In two experiments, we examined veridical and false memory for lists of associates from two meanings (e.g., stumble, trip, harvest, pumpkin, etc.) that converged upon a single, lexically ambiguous critical lure (e.g., fall), in order to compare the activation-monitoring and fuzzy-trace false memory accounts. In Experiment 1, we presented study lists that were blocked or alternated by meaning (within subjects), followed by a free recall test completed immediately or after a 2.5-min delay. Correct recall was greater for blocked than for alternated lists. Critical-lure false recall was greater for blocked lists on an immediate test, whereas both list types produced equivalent false recall on a delayed test. In Experiment 2, lists blocked and alternated by meaning were presented via a between-subjects design, in order to eliminate possible list-type carryover effects. Correct recall replicated the result from Experiment 1; however, blocking lists increased false recall on delayed, but not on immediate, tests. Across the experiments, clustering correct recall by meaning increased across the delay selectively for the alternated lists. Our results suggest that thematic (i.e., gist) processes are influential for false recall, especially following a delay, a pattern consistent with fuzzy-trace theory. PMID:26105976

  11. Rescuing failed oral implants via Wnt activation

    PubMed Central

    Yin, Xing; Li, Jingtao; Chen, Tao; Mouraret, Sylvain; Dhamdhere, Girija; Brunski, John B.; Zou, Shujuan; Helms, Jill A.

    2016-01-01

    Aim Implant osseointegration is not always guaranteed and once fibrous encapsulation occurs clinicians have few options other than implant removal. Our goal was to test whether a WNT protein therapeutic could rescue such failed implants. Material and Methods Titanium implants were placed in over-sized murine oral osteotomies. A lack of primary stability was verified by mechanical testing. Interfacial strains were estimated by finite element modelling and histology coupled with histomorphometry confirmed the lack of peri-implant bone. After fibrous encapsulation was established peri-implant injections of a liposomal formulation of WNT3A protein (L-WNT3A) or liposomal PBS (L-PBS) were then initiated. Quantitative assays were employed to analyse the effects of L-WNT3A treatment. Results Implants in gap-type interfaces exhibited high interfacial strains and no primary stability. After verification of implant failure, L-WNT3A or L-PBS injections were initiated. L-WNT3A induced a rapid, significant increase in Wnt responsiveness in the peri-implant environment, cell proliferation and osteogenic protein expression. The amount of peri-implant bone and bone in contact with the implant were significantly higher in L-WNT3A cases. Conclusions These data demonstrate L-WNT3A can induce peri-implant bone formation even in cases where fibrous encapsulation predominates. PMID:26718012

  12. Wnt pathway in atypical teratoid rhabdoid tumors

    PubMed Central

    Chakravadhanula, Madhavi; Hampton, Chris N.; Chodavadia, Parth; Ozols, Victor; Zhou, Li; Catchpoole, Daniel; Xu, Jingying; Erdreich-Epstein, Anat; Bhardwaj, Ratan D.

    2015-01-01

    Background Atypical teratoid rhabdoid tumor (ATRT) is an aggressive pediatric brain tumor with limited therapeutic options. The hypothesis for this study was that the Wnt pathway triggered by the Wnt5B ligand plays an important role in ATRT biology. To address this hypothesis, the role of WNT5B and other Wnt pathway genes was analyzed in ATRT tissues and ATRT primary cell lines. Methods Transcriptome-sequencing analyses were performed using nanoString platforms, immunohistochemistry, Western blotting, quantitative reverse transcriptase PCR, immunoprecipitation, short interference RNA studies, cell viability studies, and drug dose response (DDR) assays. Results Our transcriptome-sequencing results of Wnt pathway genes from ATRT tissues and cell lines indicated that the WNT5B gene is significantly upregulated in ATRT samples compared with nontumor brain samples. These results also indicated a differential expression of both canonical and noncanonical Wnt genes. Imunoprecipitation studies indicated that Wnt5B binds to Frizzled1 and Ryk receptors. Inhibition of WNT5B by short interference RNA decreased the expression of FRIZZLED1 and RYK. Cell viability studies a indicated significant decrease in cell viability by inhibiting Frizzled1 receptor. DDR assays showed promising results with some inhibitors. Conclusions These promising therapeutic options will be studied further before starting a translational clinical trial. The success of these options will improve care for these patients. PMID:25246426

  13. Wnt signaling in skin organogenesis.

    PubMed

    Widelitz, Randall B

    2008-04-01

    While serving as the interface between an organism and its environment, the skin also can elaborate a wide range of skin appendages to service specific purposes in a region-specific fashion. As in other organs, Wnt signaling plays a key role in regulating the proliferation, differentiation and motility of skin cells during their morphogenesis. Here I will review some of the recent work that has been done on skin organogenesis. I will cover dermis formation, the development of skin appendages, cycling of appendages in the adult, stem cell regulation, patterning, orientation, regional specificity and modulation by sex hormone nuclear receptors. I will also cover their roles in wound healing, hair regeneration and skin related diseases. It appears that Wnt signaling plays essential but distinct roles in different hierarchical levels of morphogenesis and organogenesis. Many of these areas have not yet been fully explored but are certainly promising areas of future research. PMID:19279724

  14. Wnt signaling in skin organogenesis

    PubMed Central

    2008-01-01

    While serving as the interface between an organism and its environment, the skin also can elaborate a wide range of skin appendages to service specific purposes in a region-specific fashion. As in other organs, Wnt signaling plays a key role in regulating the proliferation, differentiation and motility of skin cells during their morphogenesis. Here I will review some of the recent work that has been done on skin organogenesis. I will cover dermis formation, the development of skin appendages, cycling of appendages in the adult, stem cell regulation, patterning, orientation, regional specificity and modulation by sex hormone nuclear receptors. I will also cover their roles in wound healing, hair regeneration and skin related diseases. It appears that Wnt signaling plays essential but distinct roles in different hierarchical levels of morphogenesis and organogenesis. Many of these areas have not yet been fully explored but are certainly promising areas of future research. PMID:19279724

  15. Rapid thermal processing of self-assembling block copolymer thin films on flat surfaces and topographically defined patterns

    NASA Astrophysics Data System (ADS)

    Perego, Michele; Ferrarese Lupi, Federico; Giammaria, Tommaso J.; Seguini, Gabriele; Gianotti, Valentina; Antonioli, Diego; Sparnacci, Katia; Laus, Michele; Enrico, Emanuele; de Leo, Natascia; Boarino, Luca; Ober, Christopher K.

    2014-03-01

    Self-assembling block copolymers generate nanostructured patterns, which are potentially useful for a wide range of applications. However, their technological implementation is prevented by the very long time required to drive the process. In this contribution, we demonstrate the capability to control the morphology of the self-assembling process of cylinder forming PS-b-PMMA diblock copolymer (DBC) thin films deposited on un-patterned and topographically patterned surfaces by means of a Rapid Thermal Processing (RTP) machine. Highly ordered patterns were obtained on flat surfaces for perpendicular-oriented cylindrical PS-b-PMMA block copolymers in less than 60 s. The BCs morphology evolution within topographically defined structures was systematically investigated as well. Irrespective of the surface neutralization, an irreversible orientational flipping of the BCP microdomains inside the trenches was observed. This effect was attributed to de-swelling of the polymeric film as a consequence of a progressive desorption of the solvent retained inside the film.

  16. Probing Wnt Receptor Turnover: A Critical Regulatory Point of Wnt Pathway.

    PubMed

    Jiang, Xiaomo; Cong, Feng

    2016-01-01

    Wnt pathways are critical for embryonic development and adult tissue homeostasis in all multicellular animals. Many regulatory mechanisms exist to control proper signaling output. Recent studies suggest that cell surface Wnt receptor level is controlled by ubiquitination, and serve as a critical regulatory point of Wnt pathway activity as it determines the responsiveness of cells to Wnt signal. Here, we describe flow cytometry, cell surface protein biotinylation, and immunofluorescence pulse-chase methods to probe the surface expression, ubiquitination, and internalization of the Wnt receptors FZD and LRP6. PMID:27590150

  17. Bite-Block Perturbation in People Who Stutter: Immediate Compensatory and Delayed Adaptive Processes

    ERIC Educational Resources Information Center

    Namasivayam, Aravind Kumar; van Lieshout, Pascal; De Nil, Luc

    2008-01-01

    This exploratory study investigated sensory-motor mechanisms in five people who stutter (PWS) and five people who do not (PNS). Lip kinematic and coordination data were recorded as they produced bi-syllabic nonwords at two rates (normal and fast) in three conditions (jaw-free, immediately after insertion of a bite-block, and after a 10-min…

  18. Wnt-11 signaling leads to down-regulation of the Wnt/{beta}-catenin, JNK/AP-1 and NF-{kappa}B pathways and promotes viability in the CHO-K1 cells

    SciTech Connect

    Railo, Antti; Nagy, Irina I.; Kilpelaeinen, Pekka Vainio, Seppo

    2008-08-01

    The Wnt family of glycoprotein growth factors controls a number of central cellular processes such as proliferation, differentiation and ageing. All the Wnt proteins analyzed so far either activate or inhibit the canonical {beta}-catenin signaling pathway that regulates transcription of the target genes. In addition, some of them activate noncanonical signaling pathways that involve components such as the JNK, heterotrimeric G proteins, protein kinase C, and calmodulin-dependent protein kinase II, although the precise signaling mechanisms are only just beginning to be revealed. We demonstrate here that Wnt-11 signaling is sufficient to inhibit not only the canonical {beta}-catenin mediated Wnt signaling but also JNK/AP-1 and NF-{kappa}B signaling in the CHO cells, thus serving as a noncanonical Wnt ligand in this system. Inhibition of the JNK/AP-1 pathway is mediated in part by the MAPK kinase MKK4 and Akt. Moreover, protein kinase C is involved in the regulation of JNK/AP-1 by Wnt-11, but not of the NF-{kappa}B pathway. Consistent with the central role of Akt, JNK and NF-{kappa}B in cell survival and stress responses, Wnt-11 signaling promotes cell viability. Hence Wnt-11 is involved in coordination of key signaling pathways.

  19. Renal Hypodysplasia Associates with a Wnt4 Variant that Causes Aberrant Canonical Wnt Signaling

    PubMed Central

    Vivante, Asaf; Mark-Danieli, Michal; Davidovits, Miriam; Harari-Steinberg, Orit; Omer, Dorit; Gnatek, Yehudit; Cleper, Roxana; Landau, Daniel; Kovalski, Yael; Weissman, Irit; Eisenstein, Israel; Soudack, Michalle; Wolf, Haike Reznik; Issler, Naomi; Lotan, Danny; Anikster, Yair

    2013-01-01

    Abnormal differentiation of the renal stem/progenitor pool into kidney tissue can lead to renal hypodysplasia (RHD), but the underlying causes of RHD are not well understood. In this multicenter study, we identified 20 Israeli pedigrees with isolated familial, nonsyndromic RHD and screened for mutations in candidate genes involved in kidney development, including PAX2, HNF1B, EYA1, SIX1, SIX2, SALL1, GDNF, WNT4, and WT1. In addition to previously reported RHD-causing genes, we found that two affected brothers were heterozygous for a missense variant in the WNT4 gene. Functional analysis of this variant revealed both antagonistic and agonistic canonical WNT stimuli, dependent on cell type. In HEK293 cells, WNT4 inhibited WNT3A induced canonical activation, and the WNT4 variant significantly enhanced this inhibition of the canonical WNT pathway. In contrast, in primary cultures of human fetal kidney cells, which maintain WNT activation and more closely represent WNT signaling in renal progenitors during nephrogenesis, this mutation caused significant loss of function, resulting in diminished canonical WNT/β-catenin signaling. In conclusion, heterozygous WNT4 variants are likely to play a causative role in renal hypodysplasia. PMID:23520208

  20. Wnt Lipidation and Modifiers in Intestinal Carcinogenesis and Cancer

    PubMed Central

    Kaemmerer, Elke; Gassler, Nikolaus

    2016-01-01

    The wingless (Wnt) signaling is suggested as a fundamental hierarchical pathway in regulation of proliferation and differentiation of cells. The Wnt ligands are small proteins of about 40 kDa essentially for regulation and initiation of the Wnt activity. They are secreted proteins requiring acylation for activity in the Wnt signaling cascade and for functional interactivity with transmembrane proteins. Dual lipidation is important for posttranslational activation of the overwhelming number of Wnt proteins and is probably involved in their spatial distribution. The intestinal mucosa, where Wnt signaling is essential in configuration and maintenance, is an established model to study Wnt proteins and their role in carcinogenesis and cancer. The intestinal crypt-villus/crypt-plateau axis, a cellular system with self-renewal, proliferation, and differentiation, is tightly coordinated by a Wnt gradient. In the review, some attention is given to Wnt3, Wnt3A, and Wnt2B as important members of the Wnt family to address the role of lipidation and modifiers of Wnt proteins in intestinal carcinogenesis. Wnt3 is an important player in establishing the Wnt gradient in intestinal crypts and is mainly produced by Paneth cells. Wnt2B is characterized as a mitochondrial protein and shuttles between mitochondria and the nucleus. Porcupine and ACSL5, a long-chain fatty acid activating enzyme, are introduced as modifiers of Wnts and as interesting strategy to targeting Wnt-driven carcinogenesis. PMID:27438855

  1. Wnt Lipidation and Modifiers in Intestinal Carcinogenesis and Cancer.

    PubMed

    Kaemmerer, Elke; Gassler, Nikolaus

    2016-01-01

    The wingless (Wnt) signaling is suggested as a fundamental hierarchical pathway in regulation of proliferation and differentiation of cells. The Wnt ligands are small proteins of about 40 kDa essentially for regulation and initiation of the Wnt activity. They are secreted proteins requiring acylation for activity in the Wnt signaling cascade and for functional interactivity with transmembrane proteins. Dual lipidation is important for posttranslational activation of the overwhelming number of Wnt proteins and is probably involved in their spatial distribution. The intestinal mucosa, where Wnt signaling is essential in configuration and maintenance, is an established model to study Wnt proteins and their role in carcinogenesis and cancer. The intestinal crypt-villus/crypt-plateau axis, a cellular system with self-renewal, proliferation, and differentiation, is tightly coordinated by a Wnt gradient. In the review, some attention is given to Wnt3, Wnt3A, and Wnt2B as important members of the Wnt family to address the role of lipidation and modifiers of Wnt proteins in intestinal carcinogenesis. Wnt3 is an important player in establishing the Wnt gradient in intestinal crypts and is mainly produced by Paneth cells. Wnt2B is characterized as a mitochondrial protein and shuttles between mitochondria and the nucleus. Porcupine and ACSL5, a long-chain fatty acid activating enzyme, are introduced as modifiers of Wnts and as interesting strategy to targeting Wnt-driven carcinogenesis. PMID:27438855

  2. A WntD-Dependent Integral Feedback Loop Attenuates Variability in Drosophila Toll Signaling.

    PubMed

    Rahimi, Neta; Averbukh, Inna; Haskel-Ittah, Michal; Degani, Neta; Schejter, Eyal D; Barkai, Naama; Shilo, Ben-Zion

    2016-02-22

    Patterning by morphogen gradients relies on the capacity to generate reproducible distribution profiles. Morphogen spread depends on kinetic parameters, including diffusion and degradation rates, which vary between embryos, raising the question of how variability is controlled. We examined this in the context of Toll-dependent dorsoventral (DV) patterning of the Drosophila embryo. We find that low embryo-to-embryo variability in DV patterning relies on wntD, a Toll-target gene expressed initially at the posterior pole. WntD protein is secreted and disperses in the extracellular milieu, associates with its receptor Frizzled4, and inhibits the Toll pathway by blocking the Toll extracellular domain. Mathematical modeling predicts that WntD accumulates until the Toll gradient narrows to its desired spread, and we support this feedback experimentally. This circuit exemplifies a broadly applicable induction-contraction mechanism, which reduces patterning variability through a restricted morphogen-dependent expression of a secreted diffusible inhibitor. PMID:26906736

  3. Cardamonin suppresses melanogenesis by inhibition of Wnt/beta-catenin signaling.

    PubMed

    Cho, Munju; Ryu, Minjung; Jeong, Yongsu; Chung, Young-Hwa; Kim, Dong-Eun; Cho, Ho-Song; Kang, Sangjin; Han, Jong-Sub; Chang, Min-Youl; Lee, Cheon-Koo; Jin, Muhyun; Kim, Ho-Jeoung; Oh, Sangtaek

    2009-12-18

    Wnt/beta-catenin signaling plays important roles in many developmental processes, including neural crest-derived melanocyte development. Here we show that cardamonin, a calchone from Aplinia katsumadai Hayata, inhibited pigmentation in melanocytes through suppression of Wnt/beta-catenin signaling pathway. Cardamonin significantly suppressed the expression of microphthalmia-associated transcription factor (MITF) and tyrosinase, which are melanocyte differentiation-associated markers, in human normal melanocytes, thereby decreasing intracellular melanin production. In addition, cardamonin promoted the degradation of intracellular beta-catenin that was accumulated by Wnt3a-conditioned medium (Wnt3a CM) or bromoindirubin-3'-oxime (BIO), a glycogen synthase kinase-3beta (GSK-3beta) inhibitor, in HEK293 reporter cells and human normal melanocytes. Our findings indicate that cardamonin may be a potential whitening agent for use in cosmetics and in the medical treatment of hyperpigmentation disorders. PMID:19800318

  4. Mechanism of Wnt signaling induced down regulation of mrhl long non-coding RNA in mouse spermatogonial cells

    PubMed Central

    Akhade, Vijay Suresh; Dighe, Shrinivas Nivrutti; Kataruka, Shubhangini; Rao, Manchanahalli R. Satyanarayana

    2016-01-01

    Long non coding RNAs (lncRNAs) have emerged as important regulators of various biological processes. LncRNAs also behave as response elements or targets of signaling pathway(s) mediating cellular function. Wnt signaling is important in regulating mammalian spermatogenesis. Mrhl RNA negatively regulates canonical Wnt pathway and gets down regulated upon Wnt signaling activation in mouse spermatogonial cells. Also, mrhl RNA regulates expression of genes pertaining to Wnt pathway and spermatogenesis by binding to chromatin. In the present study, we delineate the detailed molecular mechanism of Wnt signaling induced mrhl RNA down regulation in mouse spermatogonial cells. Mrhl RNA has an independent transcription unit and our various experiments like Chromatin Immunoprecipitation (in cell line as well as mouse testis) and shRNA mediated down regulation convincingly show that β-catenin and TCF4, which are the key effector proteins of the Wnt signaling pathway are required for down regulation of mrhl RNA. We have identified Ctbp1 as the co-repressor and its occupancy on mrhl RNA promoter depends on both β-catenin and TCF4. Upon Wnt signaling activation, Ctbp1 mediated histone repression marks increase at the mrhl RNA promoter. We also demonstrate that Wnt signaling induced mrhl RNA down regulation results in an up regulation of various meiotic differentiation marker genes. PMID:26446991

  5. Mechanism of Wnt signaling induced down regulation of mrhl long non-coding RNA in mouse spermatogonial cells.

    PubMed

    Akhade, Vijay Suresh; Dighe, Shrinivas Nivrutti; Kataruka, Shubhangini; Rao, Manchanahalli R Satyanarayana

    2016-01-01

    Long non coding RNAs (lncRNAs) have emerged as important regulators of various biological processes. LncRNAs also behave as response elements or targets of signaling pathway(s) mediating cellular function. Wnt signaling is important in regulating mammalian spermatogenesis. Mrhl RNA negatively regulates canonical Wnt pathway and gets down regulated upon Wnt signaling activation in mouse spermatogonial cells. Also, mrhl RNA regulates expression of genes pertaining to Wnt pathway and spermatogenesis by binding to chromatin. In the present study, we delineate the detailed molecular mechanism of Wnt signaling induced mrhl RNA down regulation in mouse spermatogonial cells. Mrhl RNA has an independent transcription unit and our various experiments like Chromatin Immunoprecipitation (in cell line as well as mouse testis) and shRNA mediated down regulation convincingly show that β-catenin and TCF4, which are the key effector proteins of the Wnt signaling pathway are required for down regulation of mrhl RNA. We have identified Ctbp1 as the co-repressor and its occupancy on mrhl RNA promoter depends on both β-catenin and TCF4. Upon Wnt signaling activation, Ctbp1 mediated histone repression marks increase at the mrhl RNA promoter. We also demonstrate that Wnt signaling induced mrhl RNA down regulation results in an up regulation of various meiotic differentiation marker genes. PMID:26446991

  6. A Possible Zebrafish Model of Polycystic Kidney Disease: Knockdown of wnt5a Causes Cysts in Zebrafish Kidneys

    PubMed Central

    Huang, Liwei; Xiao, An; Wecker, Andrea; McBride, Daniel A.; Choi, Soo Young; Zhou, Weibin; Lipschutz, Joshua H.

    2015-01-01

    Polycystic kidney disease (PKD) is one of the most common causes of end-stage kidney disease, a devastating disease for which there is no cure. The molecular mechanisms leading to cyst formation in PKD remain somewhat unclear, but many genes are thought to be involved. Wnt5a is a non-canonical glycoprotein that regulates a wide range of developmental processes. Wnt5a works through the planar cell polarity (PCP) pathway that regulates oriented cell division during renal tubular cell elongation. Defects of the PCP pathway have been found to cause kidney cyst formation. Our paper describes a method for developing a zebrafish cystic kidney disease model by knockdown of the wnt5a gene with wnt5a antisense morpholino (MO) oligonucleotides. Tg(wt1b:GFP) transgenic zebrafish were used to visualize kidney structure and kidney cysts following wnt5a knockdown. Two distinct antisense MOs (AUG - and splice-site) were used and both resulted in curly tail down phenotype and cyst formation after wnt5a knockdown. Injection of mouse Wnt5a mRNA, resistant to the MOs due to a difference in primary base pair structure, rescued the abnormal phenotype, demonstrating that the phenotype was not due to “off-target” effects of the morpholino. This work supports the validity of using a zebrafish model to study wnt5a function in the kidney. PMID:25489842

  7. Orchestrating Wnt signalling for metabolic liver zonation.

    PubMed

    Birchmeier, Walter

    2016-04-27

    Wnt/β-catenin signalling is an important regulator of liver development, zonation and regeneration. The cell surface complex RSPO-LGR4/5-ZNF3/RNF43 is now shown to direct Wnt/β-catenin signalling in orchestrating the division of the liver into functionally distinct metabolic zones, providing insights into the mechanisms that influence organ development and regeneration. PMID:27117330

  8. GATA2 regulates Wnt signaling to promote primitive red blood cell fate.

    PubMed

    Mimoto, Mizuho S; Kwon, Sunjong; Green, Yangsook Song; Goldman, Devorah; Christian, Jan L

    2015-11-01

    Primitive erythropoiesis is regulated in a non cell-autonomous fashion across evolution from frogs to mammals. In Xenopus laevis, signals from the overlying ectoderm are required to induce the mesoderm to adopt an erythroid fate. Previous studies in our lab identified the transcription factor GATA2 as a key regulator of this ectodermal signal. To identify GATA2 target genes in the ectoderm required for red blood cell formation in the mesoderm, we used microarray analysis to compare gene expression in ectoderm from GATA2 depleted and wild type embryos. Our analysis identified components of the non-canonical and canonical Wnt pathways as being reciprocally up- and down-regulated downstream of GATA2 in both mesoderm and ectoderm. We show that up-regulation of canonical Wnt signaling during gastrulation blocks commitment to a hematopoietic fate while down-regulation of non-canonical Wnt signaling impairs erythroid differentiation. Our results are consistent with a model in which GATA2 contributes to inhibition of canonical Wnt signaling, thereby permitting progenitors to exit the cell cycle and commit to a hematopoietic fate. Subsequently, activation of non-canonical Wnt signaling plays a later role in enabling these progenitors to differentiate as mature red blood cells. PMID:26365900

  9. Notch1 counteracts WNT/β-catenin signaling through chromatin modification in colorectal cancer

    PubMed Central

    Kim, Hyun-A; Koo, Bon-Kyoung; Cho, Ji-Hoon; Kim, Yoon-Young; Seong, Jinwoo; Chang, Hee Jin; Oh, Young Min; Stange, Daniel E.; Park, Jae-Gahb; Hwang, Daehee; Kong, Young-Yun

    2012-01-01

    Crosstalk between the Notch and wingless-type MMTV integration site (WNT) signaling pathways has been investigated for many developmental processes. However, this negative correlation between Notch and WNT/β-catenin signaling activity has been studied primarily in normal developmental and physiological processes in which negative feedback loops for both signaling pathways are intact. We found that Notch1 signaling retained the capability of suppressing the expression of WNT target genes in colorectal cancers even when β-catenin destruction by the adenomatous polyposis coli (APC) complex was disabled. Activation of Notch1 converted high-grade adenoma into low-grade adenoma in an Apcmin mouse colon cancer model and suppressed the expression of WNT target genes in human colorectal cancer cells through epigenetic modification recruiting histone methyltransferase SET domain bifurcated 1 (SETDB1). Extensive microarray analysis of human colorectal cancers also showed a negative correlation between the Notch1 target gene, Notch-regulated ankyrin repeat protein 1 (NRARP), and WNT target genes. Notch is known to be a strong promoter of tumor initiation, but here we uncovered an unexpected suppressive role of Notch1 on WNT/β-catenin target genes involved in colorectal cancer. PMID:22863622

  10. Wnt activation promotes neuronal differentiation of Glioblastoma

    PubMed Central

    Rampazzo, E; Persano, L; Pistollato, F; Moro, E; Frasson, C; Porazzi, P; Della Puppa, A; Bresolin, S; Battilana, G; Indraccolo, S; Te Kronnie, G; Argenton, F; Tiso, N; Basso, G

    2013-01-01

    One of the biggest challenges in tumour research is the possibility to reprogram cancer cells towards less aggressive phenotypes. In this study, we reprogrammed primary Glioblastoma multiforme (GBM)-derived cells towards a more differentiated and less oncogenic phenotype by activating the Wnt pathway in a hypoxic microenvironment. Hypoxia usually correlates with malignant behaviours in cancer cells, but it has been recently involved, together with Wnt signalling, in the differentiation of embryonic and neural stem cells. Here, we demonstrate that treatment with Wnt ligands, or overexpression of β-catenin, mediate neuronal differentiation and halt proliferation in primary GBM cells. An hypoxic environment cooperates with Wnt-induced differentiation, in line with our finding that hypoxia inducible factor-1α (HIF-1α) is instrumental and required to sustain the expression of β-catenin transcriptional partners TCF-1 and LEF-1. In addition, we also found that Wnt-induced GBM cell differentiation inhibits Notch signalling, and thus gain of Wnt and loss of Notch cooperate in the activation of a pro-neuronal differentiation program. Intriguingly, the GBM sub-population enriched of cancer stem cells (CD133+ fraction) is the primary target of the pro-differentiating effects mediated by the crosstalk between HIF-1α, Wnt, and Notch signalling. By using zebrafish transgenics and mutants as model systems to visualize and manipulate in vivo the Wnt pathway, we confirm that Wnt pathway activation is able to promote neuronal differentiation and inhibit Notch signalling of primary human GBM cells also in this in vivo set-up. In conclusion, these findings shed light on an unsuspected crosstalk between hypoxia, Wnt and Notch signalling in GBM, and suggest the potential to manipulate these microenvironmental signals to blunt GBM malignancy. PMID:23429286

  11. Wnt signalling pathway parameters for mammalian cells.

    PubMed

    Tan, Chin Wee; Gardiner, Bruce S; Hirokawa, Yumiko; Layton, Meredith J; Smith, David W; Burgess, Antony W

    2012-01-01

    Wnt/β-catenin signalling regulates cell fate, survival, proliferation and differentiation at many stages of mammalian development and pathology. Mutations of two key proteins in the pathway, APC and β-catenin, have been implicated in a range of cancers, including colorectal cancer. Activation of Wnt signalling has been associated with the stabilization and nuclear accumulation of β-catenin and consequential up-regulation of β-catenin/TCF gene transcription. In 2003, Lee et al. constructed a computational model of Wnt signalling supported by experimental data from analysis of time-dependent concentration of Wnt signalling proteins in Xenopus egg extracts. Subsequent studies have used the Xenopus quantitative data to infer Wnt pathway dynamics in other systems. As a basis for understanding Wnt signalling in mammalian cells, a confocal live cell imaging measurement technique is developed to measure the cell and nuclear volumes of MDCK, HEK293T cells and 3 human colorectal cancer cell lines and the concentrations of Wnt signalling proteins β-catenin, Axin, APC, GSK3β and E-cadherin. These parameters provide the basis for formulating Wnt signalling models for kidney/intestinal epithelial mammalian cells. There are significant differences in concentrations of key proteins between Xenopus extracts and mammalian whole cell lysates. Higher concentrations of Axin and lower concentrations of APC are present in mammalian cells. Axin concentrations are greater than APC in kidney epithelial cells, whereas in intestinal epithelial cells the APC concentration is higher than Axin. Computational simulations based on Lee's model, with this new data, suggest a need for a recalibration of the model.A quantitative understanding of Wnt signalling in mammalian cells, in particular human colorectal cancers requires a detailed understanding of the concentrations of key protein complexes over time. Simulations of Wnt signalling in mammalian cells can be initiated with the parameters

  12. Wnt3a suppresses Wnt/β-catenin signaling and cancer cell proliferation following serum deprivation.

    PubMed

    He, Qingqing; Yan, Hongwei; Wo, Da; Liu, Junjun; Liu, Peng; Zhang, Jiankang; Li, Limei; Zhou, Bin; Ge, Jin; Li, Huashun; Liu, Shangfeng; Zhu, Weidong

    2016-02-01

    Canonical Wnt/β-catenin signaling is often aberrantly activated in tumor cells and required for tumor growth. The internalization of Wnt co-receptor low-density lipoprotein receptor-related protein 6 (LRP6) induced by Wnt ligands is commonly thought to be critical for Wnt/β-catenin signaling activation. However, in contrast to theses previous studies, we here show that persistent excessive stimulation with a canonical Wnt ligand Wnt3a could induce a long-term decreased expression level of membrane LRP6, which prevented nuclear β-catenin accumulation and tumor cell;proliferation. Importantly, Wnt3a was robustly upregulated following serum deprivation. The upregulated Wnt3a under serum deprivation was responsible for LRP6 internalization, decreased accumulation of nuclear β-catenin, and further inhibition of tumor cell proliferation. It has well been known that insufficient blood supply during tumor development occurs frequently, causing a worsening environment for tumor growth. Therefore, these results reveal a novel inhibitory role of Wnt3a on canonical Wnt/β-catenin signaling and cancer cell proliferation when there is an insufficient blood supply during tumor development, which might be a potential mechanism for tumor evasion within a worsening environment. PMID:26643293

  13. Autocrine stimulation of osteoblast activity by Wnt5a in response to TNF-α in human mesenchymal stem cells

    SciTech Connect

    Briolay, A.; Lencel, P.; Caverzasio, J.; Buchet, R.; Magne, D.

    2013-01-18

    Highlights: ► Ankylosing spondylitis (AS) leads to bone fusions and ankylosis. ► TNF-α stimulates osteoblasts through growth factors in AS. ► We compare the involvement of canonical vs non-canonical Wnt signaling. ► Canonical Wnt signaling is not involved in TNF-α effects in differentiating hMSCs. ► TNF-α stimulates osteoblasts through Wnt5a autocrine secretion in hMSCs. -- Abstract: Although anti-tumor necrosis factor (TNF)-α treatments efficiently block inflammation in ankylosing spondylitis (AS), they are inefficient to prevent excessive bone formation. In AS, ossification seems more prone to develop in sites where inflammation has resolved following anti-TNF therapy, suggesting that TNF-α indirectly stimulates ossification. In this context, our objectives were to determine and compare the involvement of Wnt proteins, which are potent growth factors of bone formation, in the effects of TNF-α on osteoblast function. In human mesenchymal stem cells (MSCs), TNF-α significantly increased the levels of Wnt10b and Wnt5a. Associated with this effect, TNF-α stimulated tissue-non specific alkaline phosphatase (TNAP) and mineralization. This effect was mimicked by activation of the canonical β-catenin pathway with either anti-Dkk1 antibodies, lithium chloride (LiCl) or SB216763. TNF-α reduced, and activation of β-catenin had little effect on expression of osteocalcin, a late marker of osteoblast differentiation. Surprisingly, TNF-α failed to stabilize β-catenin and Dkk1 did not inhibit TNF-α effects. In fact, Dkk1 expression was also enhanced in response to TNF-α, perhaps explaining why canonical signaling by Wnt10b was not activated by TNF-α. However, we found that Wnt5a also stimulated TNAP in MSCs cultured in osteogenic conditions, and increased the levels of inflammatory markers such as COX-2. Interestingly, treatment with anti-Wnt5a antibodies reduced endogenous TNAP expression and activity. Collectively, these data suggest that increased

  14. NFAT5 regulates the canonical Wnt pathway and is required for cardiomyogenic differentiation

    SciTech Connect

    Adachi, Atsuo; Takahashi, Tomosaburo; Ogata, Takehiro; Imoto-Tsubakimoto, Hiroko; Nakanishi, Naohiko; Ueyama, Tomomi; Matsubara, Hiroaki

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer NFAT5 protein expression is downregulated during cardiomyogenesis. Black-Right-Pointing-Pointer Inhibition of NFAT5 function suppresses canonical Wnt signaling. Black-Right-Pointing-Pointer Inhibition of NFAT5 function attenuates mesodermal induction. Black-Right-Pointing-Pointer NFAT5 function is required for cardiomyogenesis. -- Abstract: While nuclear factor of activated T cells 5 (NFAT5), a transcription factor implicated in osmotic stress response, is suggested to be involved in other processes such as migration and proliferation, its role in cardiomyogenesis is largely unknown. Here, we examined the role of NFAT5 in cardiac differentiation of P19CL6 cells, and observed that it was abundantly expressed in undifferentiated P19CL6 cells, and its protein expression was significantly downregulated by enhanced proteasomal degradation during DMSO-induced cardiomyogenesis. Expression of a dominant negative mutant of NFAT5 markedly attenuated cardiomyogenesis, which was associated with the inhibition of mesodermal differentiation. TOPflash reporter assay revealed that the transcriptional activity of canonical Wnt signaling was activated prior to mesodermal differentiation, and this activation was markedly attenuated by NFAT5 inhibition. Pharmacological activation of canonical Wnt signaling by [2 Prime Z, 3 Prime E]-6-bromoindirubin-3 Prime -oxime (BIO) restored Brachyury expression in NFAT5DN-expressing cells. Inhibition of NFAT5 markedly attenuated Wnt3 and Wnt3a induction. Expression of Dkk1 and Cerberus1, which are secreted Wnt antagonists, was also inhibited by NFAT5 inhibition. Thus, endogenous NFAT5 regulates the coordinated expression of Wnt ligands and antagonists, which are essential for cardiomyogenesis through the canonical Wnt pathway. These results demonstrated a novel role of NFAT5 in cardiac differentiation of stem cells.

  15. Regulation of TCF3 by Wnt-dependent phosphorylation during vertebrate axis specification.

    PubMed

    Hikasa, Hiroki; Ezan, Jerome; Itoh, Keiji; Li, Xiaotong; Klymkowsky, Michael W; Sokol, Sergei Y

    2010-10-19

    A commonly accepted model of Wnt/β-catenin signaling involves target gene activation by a complex of β-catenin with a T-cell factor (TCF) family member. TCF3 is a transcriptional repressor that has been implicated in Wnt signaling and plays key roles in embryonic axis specification and stem cell differentiation. Here we demonstrate that Wnt proteins stimulate TCF3 phosphorylation in gastrulating Xenopus embryos and mammalian cells. This phosphorylation event involves β-catenin-mediated recruitment of homeodomain-interacting protein kinase 2 (HIPK2) to TCF3 and culminates in the dissociation of TCF3 from a target gene promoter. Mutated TCF3 proteins resistant to Wnt-dependent phosphorylation function as constitutive inhibitors of Wnt-mediated activation of Vent2 and Cdx4 during anteroposterior axis specification. These findings reveal an alternative in vivo mechanism of Wnt signaling that involves TCF3 phosphorylation and subsequent derepression of target genes and link this molecular event to a specific developmental process. PMID:20951344

  16. Transposon Dysregulation Modulates dWnt4 Signaling to Control Germline Stem Cell Differentiation in Drosophila

    PubMed Central

    Upadhyay, Maitreyi; Martino Cortez, Yesenia; Wong-Deyrup, SiuWah; Tavares, Leticia; Schowalter, Sean; Flora, Pooja; Hill, Corinne; Nasrallah, Mohamad Ali; Chittur, Sridar; Rangan, Prashanth

    2016-01-01

    Germline stem cell (GSC) self-renewal and differentiation are required for the sustained production of gametes. GSC differentiation in Drosophila oogenesis requires expression of the histone methyltransferase dSETDB1 by the somatic niche, however its function in this process is unknown. Here, we show that dSETDB1 is required for the expression of a Wnt ligand, Drosophila Wingless type mouse mammary virus integration site number 4 (dWnt4) in the somatic niche. dWnt4 signaling acts on the somatic niche cells to facilitate their encapsulation of the GSC daughter, which serves as a differentiation cue. dSETDB1 is known to repress transposable elements (TEs) to maintain genome integrity. Unexpectedly, we found that independent upregulation of TEs also downregulated dWnt4, leading to GSC differentiation defects. This suggests that dWnt4 expression is sensitive to the presence of TEs. Together our results reveal a chromatin-transposon-Wnt signaling axis that regulates stem cell fate. PMID:27019121

  17. Promising Druggable Target in Head and Neck Squamous Cell Carcinoma: Wnt Signaling

    PubMed Central

    Aminuddin, Amnani; Ng, Pei Yuen

    2016-01-01

    Canonical Wnt signaling pathway, also known as Wnt/β-catenin signaling pathway, is a crucial mechanism for cellular maintenance and development. It regulates cell cycle progression, apoptosis, proliferation, migration, and differentiation. Dysregulation of this pathway correlates with oncogenesis in various tissues including breast, colon, pancreatic as well as head and neck cancers. Furthermore, the canonical Wnt signaling pathway has also been described as one of the critical signaling pathways for regulation of normal stem cells as well as cancer cells with stem cell-like features, termed cancer stem cells (CSC). In this review, we will briefly describe the basic mechanisms of Wnt signaling pathway and its crucial roles in the normal regulation of cellular processes as well as in the development of cancer. Next, we will highlight the roles of canonical Wnt signaling pathway in the regulation of CSC properties namely self-renewal, differentiation, metastasis, and drug resistance abilities, particularly in head and neck squamous cell carcinoma. Finally, we will examine the findings of several recent studies which explore druggable targets in the canonical Wnt signaling pathway which could be valuable to improve the treatment outcome for head and neck cancer. PMID:27570510

  18. An HMG1-like protein facilitates Wnt signaling in Caenorhabditis elegans

    PubMed Central

    Jiang, Lily I.; Sternberg, Paul W.

    1999-01-01

    We show that during Caenorhabditis elegans male spicule development, the specification of a glial versus neuronal cell fate in a canonical neurogenic sublineage is dependent on Wnt signaling. Inactivation of a Wnt signaling pathway mediated by the Wnt receptor LIN-17 transforms the SPD sheath cell into its sister, the SPD neuron. We discovered a new mutant, son-1, that displays this same cell fate transformation. The son-1 mutation enhances the phenotypes of reduction-of-function lin-17 mutants in several developmental processes, including vulva development, somatic gonad development, and male tail patterning. son-1 encodes an HMG1/2-like DNA-binding protein and is localized in all cell nuclei through development as revealed by a GFP reporter construct. Disruption of son-1 function by RNA-mediated interference results in the same spicule defect as caused by overexpression of POP-1, a TCF/LEF class HMG protein known to act downstream of the Wnt signaling pathway. Our results provide in vivo evidence for the functional involvement of an HMG1/2-like protein, SON-1, in Wnt signaling. The sequence nonspecific HMG protein SON-1 and the sequence specific HMG protein POP-1 might both act in the Wnt responding cells to regulate gene transcription in opposite directions. PMID:10197987

  19. Aberrant levels of Wnt/β-catenin pathway components in a rat model of endometriosis.

    PubMed

    de Mattos, Rômulo Medina; Pereira, Paula Rodrigues; Barros, Eliane Gouvêa de Oliveira; da Silva, Julianna Henriques; Palmero, Celia Yelimar; da Costa, Nathália Meireles; Pinto, Luis Felipe Ribeiro; Gimba, Etel Rodrigues Pereira; Hecht, Fabio; Ferreira, Luciana Bueno; Machado, Daniel Escorsim; de Oliveira, Felipe Leite; Nasciutti, Luiz Eurico

    2016-08-01

    Endometriosis is a benign gynecological disease affecting approximately 10-15% of women of reproductive age and 25-50% of all infertile women. It is characterized by the presence of glands and/or endometrial stroma outside the uterine cavity. Angiogenesis is a crucial process for the development and maintenance of endometriotic lesions. The Wnt/β-catenin pathway is a major promoter of angiogenesis in both physiological and pathological conditions. In the present study, we evaluated the expression of molecules related to the Wnt/β-catenin pathway in a rat model of peritoneal endometriosis. mRNA analyses showed significantly increased expression of Wnt4 and Wnt7b and decreased expression of Gsk3beta and E-cadherin in endometriotic lesions. However, there were no differences in β-catenin and Fzd2 mRNA expression. In addition, we observed a significant increase of nuclear β-catenin in endometriotic lesions, a hallmark of Wnt/ β-catenin pathway activation. Stromal β-catenin staining was found in 45.4% of endometrial tissues and 77.8% of endometriotic lesions. β-catenin nuclear localization was found in 18.2% of the endometrial tissues and 33.3% of endometriotic lesions. Finally, the expression of galectin-3, a regulator of this pathway, was increased in endometriosis. In summary, this pattern of Wnt/β-catenin components expression suggests an increased activity of this pathway in endometriosis. PMID:26853489

  20. Promising Druggable Target in Head and Neck Squamous Cell Carcinoma: Wnt Signaling.

    PubMed

    Aminuddin, Amnani; Ng, Pei Yuen

    2016-01-01

    Canonical Wnt signaling pathway, also known as Wnt/β-catenin signaling pathway, is a crucial mechanism for cellular maintenance and development. It regulates cell cycle progression, apoptosis, proliferation, migration, and differentiation. Dysregulation of this pathway correlates with oncogenesis in various tissues including breast, colon, pancreatic as well as head and neck cancers. Furthermore, the canonical Wnt signaling pathway has also been described as one of the critical signaling pathways for regulation of normal stem cells as well as cancer cells with stem cell-like features, termed cancer stem cells (CSC). In this review, we will briefly describe the basic mechanisms of Wnt signaling pathway and its crucial roles in the normal regulation of cellular processes as well as in the development of cancer. Next, we will highlight the roles of canonical Wnt signaling pathway in the regulation of CSC properties namely self-renewal, differentiation, metastasis, and drug resistance abilities, particularly in head and neck squamous cell carcinoma. Finally, we will examine the findings of several recent studies which explore druggable targets in the canonical Wnt signaling pathway which could be valuable to improve the treatment outcome for head and neck cancer. PMID:27570510

  1. Seafloor character and sedimentary processes in eastern Long Island Sound and western Block Island Sound

    NASA Astrophysics Data System (ADS)

    Poppe, L. J.; Digiacomo-Cohen, M. L.; Smith, S. M.; Stewart, H. F.; Forfinski, N. A.

    2006-06-01

    Multibeam bathymetric data and seismic-reflection profiles collected in eastern Long Island Sound and western Block Island Sound reveal previously unrecognized glacial features and modern bedforms. Glacial features include an ice-sculptured bedrock surface, a newly identified recessional moraine, exposed glaciolacustrine sediments, and remnants of stagnant-ice-contact deposits. Modern bedforms include fields of transverse sand waves, barchanoid waves, giant scour depressions, and pockmarks. Bedform asymmetry and scour around obstructions indicate that net sediment transport is westward across the northern part of the study area near Fishers Island, and eastward across the southern part near Great Gull Island.

  2. Interplay Between Residual Stresses, Microstructure, Process Variables and Engine Block Casting Integrity

    NASA Astrophysics Data System (ADS)

    Lombardi, Anthony; D'Elia, Francesco; Ravindran, Comondore; Sediako, Dimitry; Murty, B. S.; MacKay, Robert

    2012-12-01

    The replacement of nodular cast iron with 319 type aluminum (Al) alloys in gasoline engine blocks is an example of the shift towards the use of lighter alloys in the automotive industry. However, excessive residual stress along the cylinder bore may lead to bore distortion, significantly reducing engine operating efficiency. In the current study, microstructure, mechanical properties and residual stress were characterized along the cylinder bridge of engine blocks following thermal sand reclamation (TSR), T7 heat treatment, and service testing of the casting. Neutron diffraction was effectively used to quantify the residual stress along both the Al cylinder bridge and the adjacent gray cast iron cylinder liners in the hoop, radial, and axial orientations with respect to the cylinder axis. The results suggest that an increase in cooling rate along the cylinder caused a significant refinement in microstructure at the bottom of the cylinder. In turn, this suggested an increase in alloy strength at the bottom of the cylinder relative to the top. This increased strength at the bottom of the cylinder likely reduced the susceptibility of the cylinder to rapid relief of residual stress at elevated temperature. In contrast, the coarse microstructure at the top of the cylinder likely triggered stress relief at an elevated temperature.

  3. Reversible Morphology Control in Block Copolymer Films via Solvent Vapor Processing: An In Situ GISAXS study

    PubMed Central

    Paik, Marvin Y.; Bosworth, Joan K.; Smilges, Detlef-M.; Schwartz, Evan L.; Andre, Xavier; Ober, Christopher K.

    2010-01-01

    The real time changes occurring within films of cylinder-forming poly(α-methylstyrene-block-4-hydroxystyrene) (PαMS-b-PHOST) were monitored as they were swollen in tetrahydrofuran (THF) and acetone solvent vapors. In situ information was obtained by combining grazing incidence small angle X-ray scattering (GISAXS) with film thickness monitoring of the solvent vapor swollen films. We show that for self assembly to occur, the polymer thin film must surpass a swollen thickness ratio of 212% of its original thickness when swollen in THF vapors and a ratio of 268% for acetone vapor annealing. As the polymer becomes plasticized by solvent vapor uptake, the polymer chains must become sufficiently mobile to self assemble, or reorganize, at room temperature. Using vapors of a solvent selective to one of the blocks, in our case PHOST-selective acetone, an order-order transition occured driven by the shift in volume fraction. The BCC spherical phase assumed in the highly swollen state can be quenched by rapid drying. Upon treatment with vapor of a non-selective solvent, THF, the film maintained the cylindrical morphology suggested by its dry-state volume fraction. In situ studies indicate that self-assembly occurs spontaneously upon attaining the threshold swelling ratios. PMID:21116459

  4. Heart Block

    MedlinePlus

    ... Block Explore Heart Block What Is... Electrical System & EKG Results Types Causes Who Is at Risk Signs & ... heart block. Doctors use a test called an EKG (electrocardiogram) to help diagnose heart block. This test ...

  5. The WIF domain of the human and Drosophila Wif-1 secreted factors confers specificity for Wnt or Hedgehog.

    PubMed

    Sánchez-Hernández, David; Sierra, Javier; Ortigão-Farias, João Ramalho; Guerrero, Isabel

    2012-10-01

    The Hedgehog (Hh) and Wnt signaling pathways are crucial for development as well as for adult stem cell maintenance in all organisms from Drosophila to humans. Aberrant activation of these pathways has been implicated in many types of human cancer. During evolution, organisms have developed numerous ways to fine-tune Wnt and Hh signaling. One way is through extracellular modulators that directly interact with Wnt or Hh, such as the Wnt inhibitory factor (Wif-1) family of secreted factors. Interestingly, Wif-1 family members have divergent functions in the Wnt and Hh pathways in different organisms. Whereas vertebrate Wif-1 blocks Wnt signaling, Drosophila Wif-1 [Shifted (Shf)] regulates only Hh distribution and spreading through the extracellular matrix. Here, we investigate which parts of the Shf and human Wif-1 (WIF1) proteins are responsible for functional divergence. We analyze the behavior of domain-swap (the Drosophila and human WIF domain and EGF repeats) chimeric constructs during wing development. We demonstrate that the WIF domain confers the specificity for Hh or Wg morphogen. The EGF repeats are important for the interaction of Wif-1 proteins with the extracellular matrix; Drosophila EGF repeats preferentially interact with the glypican Dally-like (Dlp) when the WIF domain belongs to human WIF1 and with Dally when the WIF domain comes from Shf. These results are important both from the evolutionary perspective and for understanding the mechanisms of morphogen distribution in a morphogenetic field. PMID:22951645

  6. Crosstalk between Wnt/β-Catenin and Estrogen Receptor Signaling Synergistically Promotes Osteogenic Differentiation of Mesenchymal Progenitor Cells

    PubMed Central

    Gao, Yanhong; Huang, Enyi; Zhang, Hongmei; Wang, Jinhua; Wu, Ningning; Chen, Xian; Wang, Ning; Wen, Sheng; Nan, Guoxin; Deng, Fang; Liao, Zhan; Wu, Di; Zhang, Bosi; Zhang, Junhui; Haydon, Rex C.; Luu, Hue H.; Shi, Lewis L.; He, Tong-Chuan

    2013-01-01

    Osteogenic differentiation from mesenchymal progenitor cells (MPCs) are initiated and regulated by a cascade of signaling events. Either Wnt/β-catenin or estrogen signaling pathway has been shown to play an important role in regulating skeletal development and maintaining adult tissue homeostasis. Here, we investigate the potential crosstalk and synergy of these two signaling pathways in regulating osteogenic differentiation of MPCs. We find that the activation of estrogen receptor (ER) signaling by estradiol (E2) or exogenously expressed ERα in MPCs synergistically enhances Wnt3A-induced early and late osteogenic markers, as well as matrix mineralization. The E2 or ERα-mediated synergy can be effectively blocked by ERα antagonist tamoxifen. E2 stimulation can enhance endochondral ossification of Wnt3A-transduced mouse fetal limb explants. Furthermore, exogenously expressed ERα significantly enhances the maturity and mineralization of Wnt3A-induced subcutaneous and intramuscular ectopic bone formation. Mechanistically, we demonstrate that E2 does not exert any detectable effect on β-catenin/Tcf reporter activity. However, ERα expression is up-regulated within the first 48h in AdWnt3A-transduced MPCs, whereas ERβ expression is significantly inhibited within 24h. Moreover, the key enzyme for the biosynthesis of estrogens aromatase is modulated by Wnt3A in a biphasic manner, up-regulated at 24h but reduced after 48h. Our results demonstrate that, while ER signaling acts synergistically with Wnt3A in promoting osteogenic differentiation, Wnt3A may crosstalk with ER signaling by up-regulating ERα expression and down-regulating ERβ expression in MPCs. Thus, the signaling crosstalk and synergy between these two pathways should be further explored as a potential therapeutic approach to combating bone and skeletal disorders, such as fracture healing and osteoporosis. PMID:24340027

  7. How the Wnt signaling pathway protects from neurodegeneration: the mitochondrial scenario.

    PubMed

    Arrázola, Macarena S; Silva-Alvarez, Carmen; Inestrosa, Nibaldo C

    2015-01-01

    Alzheimer's disease (AD) is the most common neurodegenerative disorder and is characterized by progressive memory loss and cognitive decline. One of the hallmarks of AD is the overproduction of amyloid-beta aggregates that range from the toxic soluble oligomer (Aβo) form to extracellular accumulations in the brain. Growing evidence indicates that mitochondrial dysfunction is a common feature of neurodegenerative diseases and is observed at an early stage in the pathogenesis of AD. Reports indicate that mitochondrial structure and function are affected by Aβo and can trigger neuronal cell death. Mitochondria are highly dynamic organelles, and the balance between their fusion and fission processes is essential for neuronal function. Interestingly, in AD, the process known as "mitochondrial dynamics" is also impaired by Aβo. On the other hand, the activation of the Wnt signaling pathway has an essential role in synaptic maintenance and neuronal functions, and its deregulation has also been implicated in AD. We have demonstrated that canonical Wnt signaling, through the Wnt3a ligand, prevents the permeabilization of mitochondrial membranes through the inhibition of the mitochondrial permeability transition pore (mPTP), induced by Aβo. In addition, we showed that non-canonical Wnt signaling, through the Wnt5a ligand, protects mitochondria from fission-fusion alterations in AD. These results suggest new approaches by which different Wnt signaling pathways protect neurons in AD, and support the idea that mitochondria have become potential therapeutic targets for the treatment of neurodegenerative disorders. Here we discuss the neuroprotective role of the canonical and non-canonical Wnt signaling pathways in AD and their differential modulation of mitochondrial processes, associated with mitochondrial dysfunction and neurodegeneration. PMID:25999816

  8. How the Wnt signaling pathway protects from neurodegeneration: the mitochondrial scenario

    PubMed Central

    Arrázola, Macarena S.; Silva-Alvarez, Carmen; Inestrosa, Nibaldo C.

    2015-01-01

    Alzheimer’s disease (AD) is the most common neurodegenerative disorder and is characterized by progressive memory loss and cognitive decline. One of the hallmarks of AD is the overproduction of amyloid-beta aggregates that range from the toxic soluble oligomer (Aβo) form to extracellular accumulations in the brain. Growing evidence indicates that mitochondrial dysfunction is a common feature of neurodegenerative diseases and is observed at an early stage in the pathogenesis of AD. Reports indicate that mitochondrial structure and function are affected by Aβo and can trigger neuronal cell death. Mitochondria are highly dynamic organelles, and the balance between their fusion and fission processes is essential for neuronal function. Interestingly, in AD, the process known as “mitochondrial dynamics” is also impaired by Aβo. On the other hand, the activation of the Wnt signaling pathway has an essential role in synaptic maintenance and neuronal functions, and its deregulation has also been implicated in AD. We have demonstrated that canonical Wnt signaling, through the Wnt3a ligand, prevents the permeabilization of mitochondrial membranes through the inhibition of the mitochondrial permeability transition pore (mPTP), induced by Aβo. In addition, we showed that non-canonical Wnt signaling, through the Wnt5a ligand, protects mitochondria from fission-fusion alterations in AD. These results suggest new approaches by which different Wnt signaling pathways protect neurons in AD, and support the idea that mitochondria have become potential therapeutic targets for the treatment of neurodegenerative disorders. Here we discuss the neuroprotective role of the canonical and non-canonical Wnt signaling pathways in AD and their differential modulation of mitochondrial processes, associated with mitochondrial dysfunction and neurodegeneration. PMID:25999816

  9. Connective tissue growth factor (CTGF) is regulated by Wnt and bone morphogenetic proteins signaling in osteoblast differentiation of mesenchymal stem cells.

    PubMed

    Luo, Qing; Kang, Quan; Si, Weike; Jiang, Wei; Park, Jong Kyung; Peng, Ying; Li, Xinmin; Luu, Hue H; Luo, Jeffrey; Montag, Anthony G; Haydon, Rex C; He, Tong-Chuan

    2004-12-31

    Osteoblast lineage-specific differentiation of mesenchymal stem cells is a well regulated but poorly understood process. Both bone morphogenetic proteins (BMPs) and Wnt signaling are implicated in regulating osteoblast differentiation and bone formation. Here we analyzed the expression profiles of mesenchymal stem cells stimulated with Wnt3A and osteogenic BMPs, and we identified connective tissue growth factor (CTGF) as a potential target of Wnt and BMP signaling. We confirmed the microarray results, and we demonstrated that CTGF was up-regulated at the early stage of BMP-9 and Wnt3A stimulations and that Wnt3A-regulated CTGF expression was beta-catenin-dependent. RNA interference-mediated knockdown of CTGF expression significantly diminished BMP-9-induced, but not Wnt3A-induced, osteogenic differentiation, suggesting that Wnt3A may also regulate osteoblast differentiation in a CTGF-independent fashion. However, constitutive expression of CTGF was shown to inhibit both BMP-9- and Wnt3A-induced osteogenic differentiation. Exogenous expression of CTGF was shown to promote cell migration and recruitment of mesenchymal stem cells. Our findings demonstrate that CTGF is up-regulated by Wnt3A and BMP-9 at the early stage of osteogenic differentiation, which may regulate the proliferation and recruitment of osteoprogenitor cells; however, CTGF is down-regulated as the differentiation potential of committed pre-osteoblasts increases, strongly suggesting that tight regulation of CTGF expression may be essential for normal osteoblast differentiation of mesenchymal stem cells. PMID:15496414

  10. Protective effects of miR-29a on diabetic glomerular dysfunction by modulation of DKK1/Wnt/β-catenin signaling

    PubMed Central

    Hsu, Yung-Chien; Chang, Pey-Jium; Ho, Cheng; Huang, Yu-Ting; Shih, Ya-Hsueh; Wang, Ching-Jen; Lin, Chun-Liang

    2016-01-01

    Dysregulation of specific microRNAs or Wnt/β-catenin signaling pathway is critically implicated in the pathogenesis of various renal diseases. However, the relationship between microRNAs and Wnt/β-catenin signaling in diabetes-induced glomerular sclerosis remains unknown. Here, we found that decreased miR-29a expression and attenuated Wnt/β-catenin signaling were concomitantly detected in glomeruli of streptozotocin-induced diabetic mice. Gain of miR-29a function in diabetic mice substantially increased the expression of β-catenin and blocked the expressions of profibrotic gene markers, including DKK1 (a Wnt antagonist), TGF-β1 and fibronectin, in glomerular mesangium. Moreover, in the normal mice treated with miR-29a inhibitor, renal fibrosis was induced with an attenuated Wnt/β-catenin signaling activity. Consistently, the constructed miR-29a transgenic mice that supported sustained Wnt/β-catenin signaling had the ability to block the expressions of profibrotic genes after induction of diabetes. We also demonstrated that miR-29a acts as a positive regulator of Wnt/β-catenin signaling in cultured mesangial cells and functions to protect cell apoptosis and fibrosis. Importantly, we showed that activation of Wnt/β-catenin signaling in cultured mesangial cells by transfecting the β-catenin (Δ45) mutant or by a GSK-3β inhibitor reversely upregulated miR29a. Our findings suggest that the reciprocal relationship between miR-29a and DKK1/Wnt/β-catenin signaling may play an important part in protecting renal fibrogenesis. PMID:27460630

  11. Protective effects of miR-29a on diabetic glomerular dysfunction by modulation of DKK1/Wnt/β-catenin signaling.

    PubMed

    Hsu, Yung-Chien; Chang, Pey-Jium; Ho, Cheng; Huang, Yu-Ting; Shih, Ya-Hsueh; Wang, Ching-Jen; Lin, Chun-Liang

    2016-01-01

    Dysregulation of specific microRNAs or Wnt/β-catenin signaling pathway is critically implicated in the pathogenesis of various renal diseases. However, the relationship between microRNAs and Wnt/β-catenin signaling in diabetes-induced glomerular sclerosis remains unknown. Here, we found that decreased miR-29a expression and attenuated Wnt/β-catenin signaling were concomitantly detected in glomeruli of streptozotocin-induced diabetic mice. Gain of miR-29a function in diabetic mice substantially increased the expression of β-catenin and blocked the expressions of profibrotic gene markers, including DKK1 (a Wnt antagonist), TGF-β1 and fibronectin, in glomerular mesangium. Moreover, in the normal mice treated with miR-29a inhibitor, renal fibrosis was induced with an attenuated Wnt/β-catenin signaling activity. Consistently, the constructed miR-29a transgenic mice that supported sustained Wnt/β-catenin signaling had the ability to block the expressions of profibrotic genes after induction of diabetes. We also demonstrated that miR-29a acts as a positive regulator of Wnt/β-catenin signaling in cultured mesangial cells and functions to protect cell apoptosis and fibrosis. Importantly, we showed that activation of Wnt/β-catenin signaling in cultured mesangial cells by transfecting the β-catenin (Δ45) mutant or by a GSK-3β inhibitor reversely upregulated miR29a. Our findings suggest that the reciprocal relationship between miR-29a and DKK1/Wnt/β-catenin signaling may play an important part in protecting renal fibrogenesis. PMID:27460630

  12. Wnt3a mitigates acute lung injury by reducing P2X7 receptor-mediated alveolar epithelial type I cell death

    PubMed Central

    Guo, Y; Mishra, A; Weng, T; Chintagari, N R; Wang, Y; Zhao, C; Huang, C; Liu, L

    2014-01-01

    Acute lung injury (ALI) is characterized by pulmonary endothelial and epithelial cell damage, and loss of the alveolar–capillary barrier. We have previously shown that P2X7 receptor (P2X7R), a cell death receptor, is specifically expressed in alveolar epithelial type I cells (AEC I). In this study, we hypothesized that P2X7R-mediated purinergic signaling and its interaction with Wnt/β-catenin signaling contributes to AEC I death. We examined the effect of P2X7R agonist 2′-3′-O-(4-benzoylbenzoyl)-ATP (BzATP) and Wnt agonist Wnt3a on AEC I death in vitro and in vivo. We also assessed the therapeutic potential of Wnt3a in a clinically relevant ALI model of intratracheal lipopolysaccharide (LPS) exposure in ventilated mice. We found that the activation of P2X7R by BzATP caused the death of AEC I by suppressing Wnt/β-catenin signaling through stimulating glycogen synthase kinase-3β (GSK-3β) and proteasome. On the other hand, the activation of Wnt/β-catenin signaling by Wnt3a, GSK-3β inhibitor, or proteasome inhibitor blocked the P2X7R-mediated cell death. More importantly, Wnt3a attenuated the AEC I damage caused by intratracheal instillation of BzATP in rats or LPS in ventilated mice. Our results suggest that Wnt3a overrides the effect of P2X7R on the Wnt/β-catenin signaling to prevent the AEC I death and restrict the severity of ALI. PMID:24922070

  13. Wnt5a induces ROR1/ROR2 heterooligomerization to enhance leukemia chemotaxis and proliferation

    PubMed Central

    Yu, Jian; Chen, Liguang; Cui, Bing; Widhopf, George F.; Shen, Zhouxin; Wu, Rongrong; Zhang, Ling; Zhang, Suping; Briggs, Steven P.; Kipps, Thomas J.

    2015-01-01

    Evolutionarily conserved receptor tyrosine kinase–like orphan receptor-1 and -2 (ROR1/2) are considered distinct receptors for Wnt5a and are implicated in noncanonical Wnt signaling in organogenesis and cancer metastasis. We found that Wnt5a enhanced proliferation and migration of chronic lymphocytic leukemia (CLL) cells and that these effects were blocked by the humanized anti-ROR1 mAb cirmtuzumab (UC-961). Treatment of CLL cells with Wnt5a induced ROR1 to oligomerize with ROR2 and recruit guanine exchange factors (GEFs), which activated Rac1 and RhoA; siRNA-mediated silencing of either ROR1 or ROR2 or treatment with UC-961 inhibited these effects. Using the ROR1-deficient CLL cell line MEC1, we demonstrated that ectopic ROR1 expression induced ROR1/ROR2 heterooligomers, which recruited GEFs, and enhanced proliferation, cytokine-directed migration, and engraftment potential of MEC1 cells in immune-deficient mice. Notably, treatment with UC-961 inhibited engraftment of ROR1+ leukemia cells in immune-competent ROR1-transgenic mice. Molecular analysis revealed that the extracellular Kringle domain is required for ROR1/ROR2 heterooligomerization and the cysteine-rich domain or intracellular proline-rich domain is required for Wnt5a-induced recruitment of GEFs to ROR1/ROR2. This study identifies an interaction between ROR1 and ROR2 that is required for Wnt5a signaling that promotes leukemia chemotaxis and proliferation. PMID:26690702

  14. Wnt5a Regulates Ventral Midbrain Morphogenesis and the Development of A9–A10 Dopaminergic Cells In Vivo

    PubMed Central

    Andersson, Emma R.; Prakash, Nilima; Bryja, Vitezslav; Bryjova, Lenka; Yamaguchi, Terry P.; Hall, Anita C.

    2008-01-01

    Wnt5a is a morphogen that activates the Wnt/planar cell polarity (PCP) pathway and serves multiple functions during development. PCP signaling controls the orientation of cells within an epithelial plane as well as convergent extension (CE) movements. Wnt5a was previously reported to promote differentiation of A9–10 dopaminergic (DA) precursors in vitro. However, the signaling mechanism in DA cells and the function of Wnt5a during midbrain development in vivo remains unclear. We hereby report that Wnt5a activated the GTPase Rac1 in DA cells and that Rac1 inhibitors blocked the Wnt5a-induced DA neuron differentiation of ventral midbrain (VM) precursor cultures, linking Wnt5a-induced differentiation with a known effector of Wnt/PCP signaling. In vivo, Wnt5a was expressed throughout the VM at embryonic day (E)9.5, and was restricted to the VM floor and basal plate by E11.5–E13.5. Analysis of Wnt5a−/− mice revealed a transient increase in progenitor proliferation at E11.5, and a precociously induced NR4A2+ (Nurr1) precursor pool at E12.5. The excess NR4A2+ precursors remained undifferentiated until E14.5, when a transient 25% increase in DA neurons was detected. Wnt5a−/− mice also displayed a defect in (mid)brain morphogenesis, including an impairment in midbrain elongation and a rounded ventricular cavity. Interestingly, these alterations affected mostly cells in the DA lineage. The ventral Sonic hedgehog-expressing domain was broadened and flattened, a typical CE phenotype, and the domains occupied by Ngn2+ DA progenitors, NR4A2+ DA precursors and TH+ DA neurons were rostrocaudally reduced and laterally expanded. In summary, we hereby describe a Wnt5a regulation of Wnt/PCP signaling in the DA lineage and provide evidence for multiple functions of Wnt5a in the VM in vivo, including the regulation of VM morphogenesis, DA progenitor cell division, and differentiation of NR4A2+ DA precursors. PMID:18953410

  15. Wnt proteins regulate acetylcholine receptor clustering in muscle cells

    PubMed Central

    2012-01-01

    Background The neuromuscular junction (NMJ) is a cholinergic synapse that rapidly conveys signals from motoneurons to muscle cells and exhibits a high degree of subcellular specialization characteristic of chemical synapses. NMJ formation requires agrin and its coreceptors LRP4 and MuSK. Increasing evidence indicates that Wnt signaling regulates NMJ formation in Drosophila, C. elegans and zebrafish. Results In the study we systematically studied the effect of all 19 different Wnts in mammals on acetylcholine receptor (AChR) cluster formation. We identified five Wnts (Wnt9a, Wnt9b, Wnt10b, Wnt11, and Wnt16) that are able to stimulate AChR clustering, of which Wnt9a and Wnt11 are expressed abundantly in developing muscles. Using Wnt9a and Wnt11 as example, we demonstrated that Wnt induction of AChR clusters was dose-dependent and non-additive to that of agrin, suggesting that Wnts may act via similar pathways to induce AChR clusters. We provide evidence that Wnt9a and Wnt11 bind directly to the extracellular domain of MuSK, to induce MuSK dimerization and subsequent tyrosine phosphorylation of the kinase. In addition, Wnt-induced AChR clustering requires LRP4. Conclusions These results identify Wnts as new players in AChR cluster formation, which act in a manner that requires both MuSK and LRP4, revealing a novel function of LRP4. PMID:22309736

  16. RHOA inactivation enhances Wnt signaling and promotes colorectal cancer

    PubMed Central

    Rodrigues, Paulo; Macaya, Irati; Bazzocco, Sarah; Mazzolini, Rocco; Andretta, Elena; Dopeso, Higinio; Mateo-Lozano, Silvia; Bilić, Josipa; Cartón-García, Fernando; Nieto, Rocio; Suárez-López, Lucia; Afonso, Elsa; Landolfi, Stefania; Hernandez-Losa, Javier; Kobayashi, Kazuto; Cajal, Santiago Ramón y; Tabernero, Josep; Tebbutt, Niall C.; Mariadason, John M.; Schwartz, Simo; Arango, Diego

    2014-01-01

    Activation of the small GTPase RHOA has strong oncogenic effects in many tumor types, although its role in colorectal cancer remains unclear. Here we show that RHOA inactivation contributes to colorectal cancer progression/metastasis, largely through the activation of Wnt/β-catenin signaling. RhoA inactivation in the murine intestine accelerates the tumorigenic process and in human colon cancer cells leads to the redistribution of β-catenin from the membrane to the nucleus and enhanced Wnt/β-catenin signaling, resulting in increased proliferation, invasion and de-differentiation. In mice, RHOA inactivation contributes to colon cancer metastasis and reduced RHOA levels were observed at metastatic sites compared to primary human colon tumors. Therefore, we have identified a new mechanism of activation of Wnt/β-catenin signaling and characterized the role of RHOA as a novel tumor suppressor in colorectal cancer. These results constitute a shift from the current paradigm and demonstrate that RHO GTPases can suppress tumor progression and metastasis. PMID:25413277

  17. TGIF function in oncogenic Wnt signaling.

    PubMed

    Razzaque, Mohammed S; Atfi, Azeddine

    2016-04-01

    Transforming growth-interacting factor (TGIF) has been implicated in the pathogenesis of many types of human cancer, but the underlying mechanisms remained mostly enigmatic. Our recent study has revealed that TGIF functions as a mediator of oncogenic Wnt/β-catenin signaling. We found that TGIF can interact with and sequesters Axin1 and Axin2 into the nucleus, thereby culminating in disassembly of the β-catenin-destruction complex and attendant accumulation of β-catenin in the nucleus, where it activates expression of Wnt target genes, including TGIF itself. We have provided proof-of-concept evidences that high levels of TGIF expression correlate with poor prognosis in patients with triple negative breast cancer (TNBC), and that TGIF empowers Wnt-driven mammary tumorigenesis in vivo. Here, we will briefly summarize how TGIF influences Wnt signaling to promote tumorigenesis. PMID:26522669

  18. Measured Effects of Wnt3a on Proliferation of HEK293T Cells Depend on the Applied Assay

    PubMed Central

    Reischmann, Patricia; Fiebeck, Johanna; von der Weiden, Nadine; Müller, Oliver

    2015-01-01

    The Wnt signaling pathway has been associated with many essential cell processes. This study aims to examine the effects of Wnt signaling on proliferation of cultured HEK293T cells. Cells were incubated with Wnt3a, and the activation of the Wnt pathway was followed by analysis of the level of the β-catenin protein and of the expression levels of the target genes MYC and CCND1. The level of β-catenin protein increased up to fourfold. While the mRNA levels of c-Myc and cyclin D1 increased slightly, the protein levels increased up to a factor of 1.5. Remarkably, MTT and BrdU assays showed different results when measuring the proliferation rate of Wnt3a stimulated HEK293T cells. In the BrdU assays an increase of the proliferation rate could be detected, which correlated to the applied Wnt3a concentration. Oppositely, this correlation could not be shown in the MTT assays. The MTT results, which are based on the mitochondrial activity, were confirmed by analysis of the succinate dehydrogenase complex by immunofluorescence and by western blotting. Taken together, our study shows that Wnt3a activates proliferation of HEK293 cells. These effects can be detected by measuring DNA synthesis rather than by measuring changes of mitochondrial activity. PMID:26798342

  19. Role of the Wnt/β-catenin pathway in gastric cancer: An in-depth literature review

    PubMed Central

    Chiurillo, Miguel Angel

    2015-01-01

    Gastric cancer remains one of the most common cancers worldwide and one of the leading cause for cancer-related deaths. Gastric adenocarcinoma is a multifactorial disease that is genetically, cytologically and architecturally more heterogeneous than other gastrointestinal carcinomas. The aberrant activation of the Wnt/β-catenin signaling pathway is involved in the development and progression of a significant proportion of gastric cancer cases. This review focuses on the participation of the Wnt/β-catenin pathway in gastric cancer by offering an analysis of the relevant literature published in this field. Indeed, it is discussed the role of key factors in Wnt/β-catenin signaling and their downstream effectors regulating processes involved in tumor initiation, tumor growth, metastasis and resistance to therapy. Available data indicate that constitutive Wnt signalling resulting from Helicobacter pylori infection and inactivation of Wnt inhibitors (mainly by inactivating mutations and promoter hypermethylation) play an important role in gastric cancer. Moreover, a number of recent studies confirmed CTNNB1 and APC as driver genes in gastric cancer. The identification of specific membrane, intracellular, and extracellular components of the Wnt pathway has revealed potential targets for gastric cancer therapy. High-throughput “omics” approaches will help in the search for Wnt pathway antagonist in the near future. PMID:25992323

  20. The PTK7 and ROR2 Protein Receptors Interact in the Vertebrate WNT/Planar Cell Polarity (PCP) Pathway*

    PubMed Central

    Martinez, Sébastien; Scerbo, Pierluigi; Giordano, Marilyn; Daulat, Avais M.; Lhoumeau, Anne-Catherine; Thomé, Virginie; Kodjabachian, Laurent; Borg, Jean-Paul

    2015-01-01

    The non-canonical WNT/planar cell polarity (WNT/PCP) pathway plays important roles in morphogenetic processes in vertebrates. Among WNT/PCP components, protein tyrosine kinase 7 (PTK7) is a tyrosine kinase receptor with poorly defined functions lacking catalytic activity. Here we show that PTK7 associates with receptor tyrosine kinase-like orphan receptor 2 (ROR2) to form a heterodimeric complex in mammalian cells. We demonstrate that PTK7 and ROR2 physically and functionally interact with the non-canonical WNT5A ligand, leading to JNK activation and cell movements. In the Xenopus embryo, Ptk7 functionally interacts with Ror2 to regulate protocadherin papc expression and morphogenesis. Furthermore, we show that Ptk7 is required for papc activation induced by Wnt5a. Interestingly, we find that Wnt5a stimulates the release of the tagged Ptk7 intracellular domain, which can translocate into the nucleus and activate papc expression. This study reveals novel molecular mechanisms of action of PTK7 in non-canonical WNT/PCP signaling that may promote cell and tissue movements. PMID:26499793

  1. A new WNT on the bone: WNT16, cortical bone thickness, porosity and fractures

    PubMed Central

    Gori, Francesca; Lerner, Ulf; Ohlsson, Claes; Baron, Roland

    2015-01-01

    The last decade has provided abundant data implicating the WNT pathway in bone development and in the regulation of skeletal homeostasis. Rare human mutations together with gain- and loss-of-function approaches in mice have clearly demonstrated that disrupted regulation of this pathway leads to altered bone mass. In addition to these rare human and mice mutations, large population-based genome-wide association studies (GWASs) have identified single-nucleotide polymorphisms in ∼60 loci strongly associated with variations in bone mineral density (BMD) at different skeletal sites. Among the loci/genes identified by BMD GWAS, components of the WNT signaling pathway are numerous and have been shown to contribute to skeletal development and homeostasis. Within the components of WNT signaling, the gene coding for WNT16, one of the 19 WNT ligands of the human genome, has been found strongly associated with specific bone traits such as cortical bone thickness, cortical porosity and fracture risk. Recently, the first functional characterization of Wnt16 has confirmed the critical role of Wnt16 in the regulation of cortical bone mass and bone strength in mice. These reports have extended our understanding of Wnt16 function in bone homeostasis and have not only confirmed the unique association of Wnt16 with cortical bone and fracture susceptibility, as suggested by GWAS in human populations, but have also provided novel insights into the biology of this WNT ligand and the mechanism(s) by which it regulates cortical but not trabecular bone homeostasis. Most interestingly, Wnt16 appears to be a strong anti-resorptive soluble factor acting on both osteoblasts and osteoclast precursors. PMID:25987984

  2. Wnt Modulators in the Biotech Pipeline

    PubMed Central

    Rey, Jean-Philippe; Ellies, Debra L.

    2011-01-01

    The purpose of this review is to provide a better understanding for the LRP co-receptor mediated Wnt pathway signaling. Using proteomics we have also subdivided the LRP receptor family into six subfamilies, encompassing the twelve family members. This review includes a discussion of proteins containing a cystine-knot protein motif (i.e. Sclerostin, Dan, Sostdc1, Vwf, Norrin, Pdgf, Mucin) and discusses how this motif plays a role in mediating Wnt signaling through interactions with LRP. PMID:20014100

  3. Paracrine activation of WNT/β-catenin pathway in uterine leiomyoma stem cells promotes tumor growth

    PubMed Central

    Ono, Masanori; Yin, Ping; Navarro, Antonia; Moravek, Molly B.; Coon, John S.; Druschitz, Stacy A.; Serna, Vanida Ann; Qiang, Wenan; Brooks, David C.; Malpani, Saurabh S.; Ma, Jiajia; Ercan, Cihangir Mutlu; Mittal, Navdha; Monsivais, Diana; Dyson, Matthew T.; Yemelyanov, Alex; Maruyama, Tetsuo; Chakravarti, Debabrata; Kim, J. Julie; Kurita, Takeshi; Gottardi, Cara J.; Bulun, Serdar E.

    2013-01-01

    Uterine leiomyomas are extremely common estrogen and progesterone-dependent tumors of the myometrium and cause irregular uterine bleeding, severe anemia, and recurrent pregnancy loss in 15–30% of reproductive-age women. Each leiomyoma is thought to arise from a single mutated myometrial smooth muscle stem cell. Leiomyoma side-population (LMSP) cells comprising 1% of all tumor cells and displaying tumor-initiating stem cell characteristics are essential for estrogen- and progesterone-dependent in vivo growth of tumors, although they have remarkably lower estrogen/progesterone receptor levels than mature myometrial or leiomyoma cells. However, how estrogen/progesterone regulates the growth of LMSP cells via mature neighboring cells is unknown. Here, we demonstrate a critical paracrine role of the wingless-type (WNT)/β-catenin pathway in estrogen/progesterone-dependent tumorigenesis, involving LMSP and differentiated myometrial or leiomyoma cells. Estrogen/progesterone treatment of mature myometrial cells induced expression of WNT11 and WNT16, which remained constitutively elevated in leiomyoma tissues. In LMSP cells cocultured with mature myometrial cells, estrogen-progesterone selectively induced nuclear translocation of β-catenin and induced transcriptional activity of its heterodimeric partner T-cell factor and their target gene AXIN2, leading to the proliferation of LMSP cells. This effect could be blocked by a WNT antagonist. Ectopic expression of inhibitor of β-catenin and T-cell factor 4 in LMSP cells, but not in mature leiomyoma cells, blocked the estrogen/progesterone-dependent growth of human tumors in vivo. We uncovered a paracrine role of the WNT/β-catenin pathway that enables mature myometrial or leiomyoma cells to send mitogenic signals to neighboring tissue stem cells in response to estrogen and progesterone, leading to the growth of uterine leiomyomas. PMID:24082114

  4. Effect of Wnt-1 inducible signaling pathway protein-2 (WISP-2/CCN5), a downstream protein of Wnt signaling, on adipocyte differentiation

    SciTech Connect

    Inadera, Hidekuni Shimomura, Akiko; Tachibana, Shinjiro

    2009-02-20

    Wnt signaling negatively regulates adipocyte differentiation, and ectopic expression of Wnt-1 in 3T3-L1 cells induces several downstream molecules of Wnt signaling, including Wnt-1 inducible signaling pathway protein (WISP)-2. In this study, we examined the role of WISP-2 in the process of adipocyte differentiation using an in vitro cell culture system. In the differentiation of 3T3-L1 cells, WISP-2 expression was observed in growing cells and declined thereafter. In the mitotic clonal expansion phase of adipocyte differentiation, WISP-2 expression was transiently down-regulated concurrently with up-regulation of CCAAT/enhancer-binding protein {delta} expression. Treatment of 3T3-L1 cells in the differentiation medium with lithium, an activator of Wnt signaling, inhibited the differentiation process with concomitant induction of WISP-2. Treatment of differentiated cells with lithium induced de-differentiation as evidenced by profound reduction of peroxisome proliferator-activator receptor {gamma} expression and concomitant induction of WISP-2. However, de-differentiation of differentiated cells induced by tumor necrosis factor-{alpha} did not induce WISP-2 expression. To directly examine the effect of WISP-2 on adipocyte differentiation, 3T3-L1 cells were infected with a retrovirus carrying WISP-2. Although forced expression of WISP-2 inhibited preadipocyte proliferation, it had no effect on adipocyte differentiation. Thus, although WISP-2 is a downstream protein of Wnt signaling, the role of WISP-2 on adipocyte differentiation may be marginal, at least in this in vitro culture model.

  5. METABOLISM Wnt Signaling Regulates Hepatic Metabolism

    PubMed Central

    Liu, Hongjun; Fergusson, Maria M.; Wu, J. Julie; Rovira, Ilsa I.; Liu, Jie; Gavrilova, Oksana; Lu, Teng; Bao, Jianjun; Han, Donghe; Sack, Michael N.; Finkel, Toren

    2011-01-01

    The contribution of the Wnt pathway has been extensively characterized in embryogenesis, differentiation, and stem cell biology but not in mammalian metabolism. Here, using in vivo gain- and loss-of-function models, we demonstrate an important role for Wnt signaling in hepatic metabolism. In particular, β-Catenin, the downstream mediator of canonical Wnt signaling, altered serum glucose concentrations and regulated hepatic glucose production. β-catenin also modulated hepatic insulin signaling. Furthermore, β-catenin interacted with the transcription factor FoxO1 in livers from mice under starved conditions. The interaction of FoxO1 with β-catenin regulated the transcriptional activation of the genes encoding glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK), the two rate-limiting enzymes in hepatic gluconeogenesis. Moreover, starvation induced the hepatic expression of mRNAs encoding different Wnt isoforms. In addition, nutrient deprivation appeared to favor the association of β-catenin with FoxO family members, rather than with members of the T cell factor of transcriptional activators. Notably, in a model of diet-induced obesity, hepatic deletion of β-catenin improved overall metabolic homeostasis. These observations implicate Wnt signaling in the modulation of hepatic metabolism and raise the possibility that Wnt signaling may play a similar role in the metabolic regulation of other tissues. PMID:21285411

  6. Processing Cyclic Peptide-polymer Conjugates in Block Copolymer Thin Films for Sub-nm Porous Membranes

    NASA Astrophysics Data System (ADS)

    Zhang, Chen; Xu, Ting

    2014-03-01

    Porous thin films containing subnanometer channels oriented normal to the surface exhibit unique transport and separation properties and can serve as selective membranes for separation. Inspired by natural protein channels, we have developed an approach using cyclic peptide nanotubes (CPNs) embedded in polymeric matrix to mimic the transport of natural channels. The co-assembly of polymer-covered CPNs in a block copolymer (BCP) thin film requires the synchronization of two self-assembly processes, namely the microphase separation of BCP and the nanotube growth of CP-polymer conjugates. We systematically investigated the co-assembly of isolated CP-poly(ethylene glycol) (CP-PEG) conjugates and polystyrene-b-poly (methyl methacrylate) (PS-b-PMMA) in thin films as a function of CP-PEG loading (fCP-PEG) and solvent-polymer interactions. We find that there is a strong dependence of the co-assembly process on fCP-PEG due to thermodynamic limit of incorporating one CPN in one PMMA microdomain, as well as the kinetic pathway in which favorable PEG-solvent interaction helps to disperse CPNs and thus lowers the activation energy barrier of the system. This study presents critical insights in guided assemblies of functional building blocks within nanoscopic frameworks. DOE-EFRC-Gas Separation, Army Research Office.

  7. Wnt/β-catenin signaling: components, mechanisms, and diseases

    PubMed Central

    MacDonald, Bryan T.; Tamai, Keiko

    2010-01-01

    Signaling by the Wnt family of secreted glycolipoproteins via the transcription co-activator β-catenin controls embryonic development and adult homeostasis. Here we review recent progresses in this so-called canonical Wnt signaling pathway. We discuss Wnt ligands, agonists and antagonists and their interactions with Wnt receptors. We also dissect critical events that regulate β-catenin stability from Wnt receptors to the cytoplasmic β-catenin destruction complex, and nuclear machinery that mediates β-catenin-dependent transcription. Finally we highlight some key aspects of Wnt/β-catenin signaling in human diseases including congenital malformations, cancer and osteoporosis and potential therapeutic implications. PMID:19619488

  8. Mapping of Wnt, Frizzled and Wnt inhibitor gene expression domains in the avian otic primordium

    PubMed Central

    Sienknecht, Ulrike J.; Fekete, Donna M.

    2010-01-01

    Wnt signaling activates at least three different pathways involved in development and disease. Interactions of secreted ligands and inhibitors with cell-surface receptors result in the activation or regulation of particular downstream intracellular cascades. During the developmental stages of otic vesicle closure and beginning morphogenesis, the forming inner ear transcribes a plethora of Wnt-related genes. We report expression of 23 genes out of 25 tested in situ hybridization probes on tissue serial sections. Sensory primordia and Frizzled gene expression share domains, with Fzd1 being a continuous marker. Prospective nonsensory domains express Wnts, whose transcripts mainly flank prosensory regions. Finally, Wnt inhibitor domains are superimposed over both prosensory and nonsensory otic regions. Three Wnt antagonists, Dkk1, SFRP2, and Frzb are prominent. Their gene expression patterns partly overlap and change over time, which adds to the diversity of molecular micro-environments. Strikingly, prosensory domains express Wnts transiently. This includes (1) the prosensory otic region of high proliferation, neuroblast delamination, and programmed cell death at stage 20/21 (Wnt3, -5b, -7b, -8b, -9a, -11), and (2) sensory primordia at stage 25 (Wnt7a, Wnt9a). In summary, robust Wnt-related gene expression shows both spatial and temporal tuning during inner ear development as the otic vesicle initiates morphogenesis and prosensory cell fate determination. PMID:19842206

  9. Bioinformatic Evaluation of Transcriptional Regulation of WNT Pathway Genes with reference to Diabetic Nephropathy

    PubMed Central

    McKay, Gareth J.; Kavanagh, David H.; Crean, John K.; Maxwell, Alexander P.

    2016-01-01

    Objective. WNT/β-catenin pathway members have been implicated in interstitial fibrosis and glomerular sclerosis disease processes characteristic of diabetic nephropathy (DN), processes partly controlled by transcription factors (TFs) that bind to gene promoter regions attenuating regulation. We sought to identify predicted cis-acting transcription factor binding sites (TFBSs) overrepresented within WNT pathway members. Methods. We assessed 62 TFBS motif frequencies from the JASPAR databases in 65 WNT pathway genes. P values were estimated on the hypergeometric distribution for each TF. Gene expression profiles of enriched motifs were examined in DN-related datasets to assess clinical significance. Results. Transcription factor AP-2 alpha (TFAP2A), myeloid zinc finger 1 (MZF1), and specificity protein 1 (SP1) were significantly enriched within WNT pathway genes (P values < 6.83 × 10−29, 1.34 × 10−11, and 3.01 × 10−6, resp.). MZF1 expression was significantly increased in DN in a whole kidney dataset (fold change = 1.16; 16% increase; P = 0.03). TFAP2A expression was decreased in an independent dataset (fold change = −1.02; P = 0.03). No differential expression of SP1 was detected. Conclusions. Three TFBS profiles are significantly enriched within WNT pathway genes highlighting the potential of in silico analyses for identification of pathway regulators. Modification of TF binding may possibly limit DN progression, offering potential therapeutic benefit. PMID:26697505

  10. Compute-unified device architecture implementation of a block-matching algorithm for multiple graphical processing unit cards.

    PubMed

    Massanes, Francesc; Cadennes, Marie; Brankov, Jovan G

    2011-07-01

    In this paper we describe and evaluate a fast implementation of a classical block matching motion estimation algorithm for multiple Graphical Processing Units (GPUs) using the Compute Unified Device Architecture (CUDA) computing engine. The implemented block matching algorithm (BMA) uses summed absolute difference (SAD) error criterion and full grid search (FS) for finding optimal block displacement. In this evaluation we compared the execution time of a GPU and CPU implementation for images of various sizes, using integer and non-integer search grids.The results show that use of a GPU card can shorten computation time by a factor of 200 times for integer and 1000 times for a non-integer search grid. The additional speedup for non-integer search grid comes from the fact that GPU has built-in hardware for image interpolation. Further, when using multiple GPU cards, the presented evaluation shows the importance of the data splitting method across multiple cards, but an almost linear speedup with a number of cards is achievable.In addition we compared execution time of the proposed FS GPU implementation with two existing, highly optimized non-full grid search CPU based motion estimations methods, namely implementation of the Pyramidal Lucas Kanade Optical flow algorithm in OpenCV and Simplified Unsymmetrical multi-Hexagon search in H.264/AVC standard. In these comparisons, FS GPU implementation still showed modest improvement even though the computational complexity of FS GPU implementation is substantially higher than non-FS CPU implementation.We also demonstrated that for an image sequence of 720×480 pixels in resolution, commonly used in video surveillance, the proposed GPU implementation is sufficiently fast for real-time motion estimation at 30 frames-per-second using two NVIDIA C1060 Tesla GPU cards. PMID:22347787

  11. Compute-unified device architecture implementation of a block-matching algorithm for multiple graphical processing unit cards

    NASA Astrophysics Data System (ADS)

    Massanes, Francesc; Cadennes, Marie; Brankov, Jovan G.

    2011-07-01

    We describe and evaluate a fast implementation of a classical block-matching motion estimation algorithm for multiple graphical processing units (GPUs) using the compute unified device architecture computing engine. The implemented block-matching algorithm uses summed absolute difference error criterion and full grid search (FS) for finding optimal block displacement. In this evaluation, we compared the execution time of a GPU and CPU implementation for images of various sizes, using integer and noninteger search grids. The results show that use of a GPU card can shorten computation time by a factor of 200 times for integer and 1000 times for a noninteger search grid. The additional speedup for a noninteger search grid comes from the fact that GPU has built-in hardware for image interpolation. Further, when using multiple GPU cards, the presented evaluation shows the importance of the data splitting method across multiple cards, but an almost linear speedup with a number of cards is achievable. In addition, we compared the execution time of the proposed FS GPU implementation with two existing, highly optimized nonfull grid search CPU-based motion estimations methods, namely implementation of the Pyramidal Lucas Kanade Optical flow algorithm in OpenCV and simplified unsymmetrical multi-hexagon search in H.264/AVC standard. In these comparisons, FS GPU implementation still showed modest improvement even though the computational complexity of FS GPU implementation is substantially higher than non-FS CPU implementation. We also demonstrated that for an image sequence of 720 × 480 pixels in resolution commonly used in video surveillance, the proposed GPU implementation is sufficiently fast for real-time motion estimation at 30 frames-per-second using two NVIDIA C1060 Tesla GPU cards.

  12. Wnt some lose some: transcriptional governance of stem cells by Wnt/β-catenin signaling

    PubMed Central

    Lien, Wen-Hui; Fuchs, Elaine

    2014-01-01

    In mammals, Wnt/β-catenin signaling features prominently in stem cells and cancers, but how and for what purposes have been matters of much debate. In this review, we summarize our current knowledge of Wnt/β-catenin signaling and its downstream transcriptional regulators in normal and malignant stem cells. We centered this review largely on three types of stem cells—embryonic stem cells, hair follicle stem cells, and intestinal epithelial stem cells—in which the roles of Wnt/β-catenin have been extensively studied. Using these models, we unravel how many controversial issues surrounding Wnt signaling have been resolved by dissecting the diversity of its downstream circuitry and effectors, often leading to opposite outcomes of Wnt/β-catenin-mediated regulation and differences rooted in stage- and context-dependent effects. PMID:25030692

  13. Take the Wnt out of the inflammatory sails: modulatory effects of Wnt in airway diseases.

    PubMed

    Reuter, Sebastian; Beckert, Hendrik; Taube, Christian

    2016-02-01

    Bronchial asthma and chronic obstructive pulmonary disease (COPD) are chronic diseases that are associated with inflammation and structural changes in the airways and lungs. Recent findings have implicated Wnt pathways in critically regulating inflammatory responses, especially in asthma. Furthermore, canonical and noncanonical Wnt pathways are involved in structural changes such as airway remodeling, goblet cell metaplasia, and airway smooth muscle (ASM) proliferation. In COPD, Wnt pathways are not only associated with structural changes in the airways but also involved in the development of emphysema. The present review summarizes the role and function of the canonical and noncanonical Wnt pathway with regard to airway inflammation and structural changes in asthma and COPD. Further identification of the role and function of different Wnt molecules and pathways could help to develop novel therapeutic options for these diseases. PMID:26595171

  14. Ethosuximide Induces Hippocampal Neurogenesis and Reverses Cognitive Deficits in an Amyloid-β Toxin-induced Alzheimer Rat Model via the Phosphatidylinositol 3-Kinase (PI3K)/Akt/Wnt/β-Catenin Pathway.

    PubMed

    Tiwari, Shashi Kant; Seth, Brashket; Agarwal, Swati; Yadav, Anuradha; Karmakar, Madhumita; Gupta, Shailendra Kumar; Choubey, Vinay; Sharma, Abhay; Chaturvedi, Rajnish Kumar

    2015-11-20

    Neurogenesis involves generation of new neurons through finely tuned multistep processes, such as neural stem cell (NSC) proliferation, migration, differentiation, and integration into existing neuronal circuitry in the dentate gyrus of the hippocampus and subventricular zone. Adult hippocampal neurogenesis is involved in cognitive functions and altered in various neurodegenerative disorders, including Alzheimer disease (AD). Ethosuximide (ETH), an anticonvulsant drug is used for the treatment of epileptic seizures. However, the effects of ETH on adult hippocampal neurogenesis and the underlying cellular and molecular mechanism(s) are yet unexplored. Herein, we studied the effects of ETH on rat multipotent NSC proliferation and neuronal differentiation and adult hippocampal neurogenesis in an amyloid β (Aβ) toxin-induced rat model of AD-like phenotypes. ETH potently induced NSC proliferation and neuronal differentiation in the hippocampus-derived NSC in vitro. ETH enhanced NSC proliferation and neuronal differentiation and reduced Aβ toxin-mediated toxicity and neurodegeneration, leading to behavioral recovery in the rat AD model. ETH inhibited Aβ-mediated suppression of neurogenic and Akt/Wnt/β-catenin pathway gene expression in the hippocampus. ETH activated the PI3K·Akt and Wnt·β-catenin transduction pathways that are known to be involved in the regulation of neurogenesis. Inhibition of the PI3K·Akt and Wnt·β-catenin pathways effectively blocked the mitogenic and neurogenic effects of ETH. In silico molecular target prediction docking studies suggest that ETH interacts with Akt, Dkk-1, and GSK-3β. Our findings suggest that ETH stimulates NSC proliferation and differentiation in vitro and adult hippocampal neurogenesis via the PI3K·Akt and Wnt·β-catenin signaling. PMID:26420483

  15. Emerging roles of microRNAs in the Wnt signaling network.

    PubMed

    Schepeler, Troels

    2013-01-01

    The Wnt signaling network is known to regulate many cellular processes and is of crucial importance during development and in pathological conditions, including cancer. Small noncoding RNAs from the microRNA family (miRNAs) are important elements in the post-transcriptional control of gene expression. In this work, I review the cross talk between miRNAs and the canonical Wnt signaling pathway in various biological processes with particular emphasis on carcinogenesis. Because alterations of miRNA activity and aberrant Wnt signaling are each intimately linked to tumor biology, deciphering the complex interplay between these two regulatory modules is essential to advance our understanding of the integrated functions of miRNAs in signal transduction cascades and develop rational treatment regimens against cancer. PMID:23614621

  16. wnt3a but not wnt11 supports self-renewal of embryonic stem cells

    SciTech Connect

    Singla, Dinender K. . E-mail: Dinender.Kumar@uvm.edu; Schneider, David J.; LeWinter, Martin M.; Sobel, Burton E.

    2006-06-30

    wnt proteins (wnts) promote both differentiation of midbrain dopaminergic cells and self-renewal of haematopoietic stem cells. Mouse embryonic stem (ES) cells can be maintained and self-renew on mouse feeder cell layers or in media containing leukemia inhibitory factor (LIF). However, the effects of wnts on ES cells self-renewal and differentiation are not clearly understood. In the present study, we found that conditioned medium prepared from L cells expressing wnt3a can replace feeder cell layers and medium containing LIF in maintaining ES cells in the proliferation without differentiation (self-renewal) state. By contrast, conditioned medium from NIH3T3 cells expressing wnt11 did not. Alkaline phosphatase staining and compact colony formation were used as criteria of cells being in the undifferentiated state. ES cells maintained in medium conditioned by Wnt3a expressing cells underwent freezing and thawing while maintaining properties seen with LIF maintained ES cells. Purified wnt3a did not maintain self-renewal of ES cells for prolonged intervals. Thus, other factors in the medium conditioned by wnt3a expressing cells may have contributed to maintenance of ES cells in a self-renewal state. Pluripotency of ES cells was determined with the use of embryoid bodies in vitro. PD98059, a MEK specific inhibitor, promoted the growth of undifferentiated ES cells maintained in conditioned medium from wnt3a expressing cells. By contrast, the P38 MAPK inhibitor SB230580 did not, suggesting a role for the MEK pathway in self-renewal and differentiation of ES cells maintained in the wnt3a cell conditioned medium. Thus, our results show that conditioned medium from wnt3a but not wnt11 expressing cells can maintain ES cells in self-renewal and in a pluripotent state.

  17. Coating Bores of Light Metal Engine Blocks with a Nanocomposite Material using the Plasma Transferred Wire Arc Thermal Spray Process

    NASA Astrophysics Data System (ADS)

    Bobzin, K.; Ernst, F.; Zwick, J.; Schlaefer, T.; Cook, D.; Nassenstein, K.; Schwenk, A.; Schreiber, F.; Wenz, T.; Flores, G.; Hahn, M.

    2008-09-01

    Engine blocks of modern passenger car engines are generally made of light metal alloys, mostly hypoeutectic AlSi-alloys. Due to their low hardness, these alloys do not meet the tribological requirements of the system cylinder running surface—piston rings—lubricating oil. In order to provide a suitable cylinder running surface, nowadays cylinder liners made of gray cast iron are pressed in or cast into the engine block. A newer approach is to apply thermal spray coatings onto the cylinder bore walls. Due to the geometric conditions, the coatings are applied with specifically designed internal diameter thermal spray systems. With these processes a broad variety of feedstock can be applied, whereas mostly low-alloyed carbon steel feedstock is being used for this application. In the context of this work, an iron-based wire feedstock has been developed, which leads to a nanocrystalline coating. The application of this material was carried out with the Plasma Transferred Wire Arc system. AlMgSi0.5 liners were used as substrates. The coating microstructure and the properties of the coatings were analyzed.

  18. Inorganic solution-processed hole-injecting and electron-blocking layers in polymer light-emitting diodes

    NASA Astrophysics Data System (ADS)

    Reynolds, K. J.; Barker, J. A.; Greenham, N. C.; Friend, R. H.; Frey, G. L.

    2002-12-01

    The use of the solution-processed layered transition metal dichalcogenide (LTMDC) MoS2 as a hole-injecting electrode in polymer light-emitting diodes (LEDs) is reported. MoS2 functions as a very high work function metal and, in combination with an electron-blocking layer in the form of MoO3, provides good LED performance. In this study we investigated model LED devices with a single semiconductor layer, namely, the electron transporting polymer poly-[2,7-(9,9'-di-n-octylfluorene)-3,6-benzothiadiazole]. LED operation was successfully modeled using experimentally determined work functions, carrier mobilities, and barrier properties. Good agreement between experiment and model allows us to demonstrate that the MoS2 and the MoO3 layers act as a high work function hole-injection layer (MoS2) and an electron extraction barrier layer (MoO3), respectively. They improve device performance by allowing the buildup of electron density at the oxide/emissive layer interface which generates a local field, enhancing hole injection and recombination. Furthermore, the model shows the importance of controlling the thickness of the blocking layer to optimize device performance. The wide variety of polymeric emitters available and the range of electronic properties displayed by the LTMDC family and their corresponding oxides, provides the potential to tailor device performance through the selection of suitable organic and inorganic components.

  19. Modulating effects of acyl-CoA synthetase 5-derived mitochondrial Wnt2B palmitoylation on intestinal Wnt activity

    PubMed Central

    Klaus, Christina; Schneider, Ursula; Hedberg, Christian; Schütz, Anke K; Bernhagen, Jürgen; Waldmann, Herbert; Gassler, Nikolaus; Kaemmerer, Elke

    2014-01-01

    AIM: To investigate the role of acyl-CoA synthetase 5 (ACSL5) activity in Wnt signaling in intestinal surface epithelia. METHODS: Several cell lines were used to investigate the ACSL5-dependent expression and synthesis of Wnt2B, a mitochondrially expressed protein of the Wnt signaling family. Wnt activity was functionally assessed with a luciferase reporter assay. ACSL5-related biochemical Wnt2B modifications were investigated with a modified acyl-exchange assay. The findings from the cell culture models were verified using an Apcmin/+ mouse model as well as normal and neoplastic diseased human intestinal tissues. RESULTS: In the presence of ACSL5, Wnt2B was unable to translocate into the nucleus and was enriched in mitochondria, which was paralleled by a significant decrease in Wnt activity. ACSL5-dependent S-palmitoylation of Wnt2B was identified as a molecular reason for mitochondrial Wnt2B accumulation. In cell culture systems, a strong relation of ACSL5 expression, Wnt2B palmitoylation, and degree of malignancy were found. Using normal mucosa, the association of ACSL5 and Wnt2B was seen, but in intestinal neoplasias the mechanism was only rudimentarily observed. CONCLUSION: ACSL5 mediates antiproliferative activities via Wnt2B palmitoylation with diminished Wnt activity. The molecular pathway is probably relevant for intestinal homeostasis, overwhelmed by other pathways in carcinogenesis. PMID:25356045

  20. Hippocampal Wnt Signaling: Memory Regulation and Hormone Interactions.

    PubMed

    Fortress, Ashley M; Frick, Karyn M

    2016-06-01

    Wnt signaling has emerged in recent years as a major player in both nervous system development and adult synaptic plasticity. Of particular relevance to researchers studying learning and memory, Wnt signaling is critical for normal functioning of the hippocampus, a brain region that is essential for many types of memory formation and whose dysfunction is implicated in numerous neurodegenerative and psychiatric conditions. Impaired hippocampal Wnt signaling is implicated in several of these conditions, however, little is known about how Wnt signaling mediates hippocampal memory formation. This review will provide a general overview of Wnt signaling and discuss evidence demonstrating a key role for Wnt signaling in hippocampal memory formation in both normal and disease states. The regulation of Wnt signaling by ovarian sex steroid hormones will also be highlighted, given that the neuroprotection afforded by Wnt-hormone interactions may have significant implications for cognitive function in aging, neurodegenerative disease, and ischemic injury. PMID:25717070

  1. Wear Protection of AJ62 Mg Engine Blocks using Plasma Electrolytic Oxidation Process

    NASA Astrophysics Data System (ADS)

    Zhang, Peng

    2011-12-01

    In order to reduce the fuel consumption and pollution, automotive companies are developing magnesium-intensive components. However, due to the low wear resistance of the magnesium (Mg) alloys, Mg cylinder bores are vulnerable to the sliding wear attack. In this thesis, two approaches were used to protect the cylinder bores, made of a new developed Mg engine alloy AJ62 (MgA16Mn0.34Sr2). The first one was to use a Plasma Electrolytic Oxidation (PEO) process to produce oxide coatings on the Mg bores. The wear properties of the PEO coatings were evaluated by sliding wear tests under the boundary lubrication condition at the room and elevated temperatures. It was found that due to the substrate softening and the vaporization loss of the lubricant, the tribological properties of the PEO coatings were deteriorated at the elevated temperature. In order to optimize the PEO process, a statistical method (Response surface method) was used to analyze the effects of the 4 main PEO process parameters with 2 levels for each and their interactions on the tribological properties of the PEO coatings at the room and elevated temperatures, individually. A cylinder liner made of an economical metal-matrix composite (MMC) was another approach to improve the wear resistance of the Mg cylinder bore. In this thesis, an A1383/SiO2 MMC was designed to replace the expensive Alusil alloy used in the BMW Mg/Al composite engine to build the cylinder liner. To further increase the wear resistance of the MMC, PEO process was also used to form an oxide coating on the MMC. The effects of the SiO 2 content and coating thickness on the tribological properties of the MMC were studied. To evaluate the wear properties of the optimal PEO coated Mg coupons and the MMC with the oxide coatings, Alusil and cast iron, currently used on the cylinder bores of the commercial aluminum engines, were used as reference materials. The optimal PEO coated Mg coupons and the oxidized MMC showed their advantages over the

  2. Air-stable, nanostructured electronic and plasmonic materials from solution-processable, silver nanocrystal building blocks.

    PubMed

    Fafarman, Aaron T; Hong, Sung-Hoon; Oh, Soong Ju; Caglayan, Humeyra; Ye, Xingchen; Diroll, Benjamin T; Engheta, Nader; Murray, Christopher B; Kagan, Cherie R

    2014-03-25

    Herein we describe a room-temperature, chemical process to transform silver nanocrystal solids, deposited from colloidal solutions, into highly conductive, corrosion-resistant, optical and electronic materials with nanometer-scale architectures. After assembling the nanocrystal solids, we treated them with a set of simple, compact, organic and inorganic reagents: ammonium thiocyanate, ammonium chloride, potassium hydrogen sulfide, and ethanedithiol. We find that each reagent induces unique changes in the structure and composition of the resulting solid, giving rise to films that vary from insulating to, in the case of thiocyanate, conducting with a remarkably low resistivity of 8.8×10(-6) Ω·cm, only 6 times that of bulk silver. We show that thiocyanate mediates the spontaneous sintering of nanocrystals into structures with a roughness of less than 1/10th of the wavelength of visible light. We demonstrate that these solution-processed, low-resistivity, optically smooth films can be patterned, using imprint lithography, into conductive electrodes and plasmonic mesostructures with programmable resonances. We observe that thiocyanate-treated solids exhibit significantly retarded atmospheric corrosion, a feature that dramatically increases the feasibility of employing silver for electrical and plasmonic applications. PMID:24484271

  3. Curcumin-loaded nanoparticles potently induce adult neurogenesis and reverse cognitive deficits in Alzheimer's disease model via canonical Wnt/β-catenin pathway.

    PubMed

    Tiwari, Shashi Kant; Agarwal, Swati; Seth, Brashket; Yadav, Anuradha; Nair, Saumya; Bhatnagar, Priyanka; Karmakar, Madhumita; Kumari, Manisha; Chauhan, Lalit Kumar Singh; Patel, Devendra Kumar; Srivastava, Vikas; Singh, Dhirendra; Gupta, Shailendra Kumar; Tripathi, Anurag; Chaturvedi, Rajnish Kumar; Gupta, Kailash Chand

    2014-01-28

    Neurogenesis, a process of generation of new neurons, is reported to be reduced in several neurodegenerative disorders including Alzheimer's disease (AD). Induction of neurogenesis by targeting endogenous neural stem cells (NSC) could be a promising therapeutic approach to such diseases by influencing the brain self-regenerative capacity. Curcumin, a neuroprotective agent, has poor brain bioavailability. Herein, we report that curcumin-encapsulated PLGA nanoparticles (Cur-PLGA-NPs) potently induce NSC proliferation and neuronal differentiation in vitro and in the hippocampus and subventricular zone of adult rats, as compared to uncoated bulk curcumin. Cur-PLGA-NPs induce neurogenesis by internalization into the hippocampal NSC. Cur-PLGA-NPs significantly increase expression of genes involved in cell proliferation (reelin, nestin, and Pax6) and neuronal differentiation (neurogenin, neuroD1, neuregulin, neuroligin, and Stat3). Curcumin nanoparticles increase neuronal differentiation by activating the Wnt/β-catenin pathway, involved in regulation of neurogenesis. These nanoparticles caused enhanced nuclear translocation of β-catenin, decreased GSK-3β levels, and increased promoter activity of the TCF/LEF and cyclin-D1. Pharmacological and siRNA-mediated genetic inhibition of the Wnt pathway blocked neurogenesis-stimulating effects of curcumin. These nanoparticles reverse learning and memory impairments in an amyloid beta induced rat model of AD-like phenotypes, by inducing neurogenesis. In silico molecular docking studies suggest that curcumin interacts with Wif-1, Dkk, and GSK-3β. These results suggest that curcumin nanoparticles induce adult neurogenesis through activation of the canonical Wnt/β-catenin pathway and may offer a therapeutic approach to treating neurodegenerative diseases such as AD, by enhancing a brain self-repair mechanism. PMID:24467380

  4. Wnt Signaling Translocates Lys48-Linked Polyubiquitinated Proteins to the Lysosomal Pathway.

    PubMed

    Kim, Hyunjoon; Vick, Philipp; Hedtke, Joshua; Ploper, Diego; De Robertis, Edward M

    2015-05-26

    Cellular proteins are degraded in either proteasomes or lysosomes depending on the types of ubiquitin chains that covalently modify them. It is not known whether the choice between these two pathways is physiologically regulated. The Lys48-polyubiquitin chain is the major signal directing proteins for degradation in proteasomes. Here, we report the unexpected finding that canonical Wnt signaling translocates some K48-linked polyubiquitinated proteins to the endolysosomal pathway. Proteasomal target proteins, such as b-catenin, Smad1, and Smad4, were targeted into endolysosomes in a process dependent on GSK3 activity. Relocalization was also dependent on Axin1 and the multivesicular body (MVB) proteins HRS/Vps27 and Vps4. The Wnt-induced accumulation of K48-linked polyubiquitinated proteins in endolysosomal organelles was accompanied by a transient decrease in cellular levels of free mono-ubiquitin, which may contribute to Wnt-regulated stabilization of proteins (Wnt/ STOP). We conclude that Wnt redirects Lys48-polyubiquitinated proteins that are normally degraded in proteasomes to endolysosomes. PMID:26004177

  5. Initiation of synapse formation by Wnt-induced MuSK endocytosis

    PubMed Central

    Gordon, Laura R.; Gribble, Katherine D.; Syrett, Camille M.; Granato, Michael

    2012-01-01

    In zebrafish, the MuSK receptor initiates neuromuscular synapse formation by restricting presynaptic growth cones and postsynaptic acetylcholine receptors (AChRs) to the center of skeletal muscle cells. Increasing evidence suggests a role for Wnts in this process, yet how muscle cells respond to Wnt signals is unclear. Here, we show that in vivo, wnt11r and wnt4a initiate MuSK translocation from muscle membranes to recycling endosomes and that this transition is crucial for AChR accumulation at future synaptic sites. Moreover, we demonstrate that components of the planar cell polarity pathway colocalize to recycling endosomes and that this localization is MuSK dependent. Knockdown of several core components disrupts MuSK translocation to endosomes, AChR localization and axonal guidance. We propose that Wnt-induced trafficking of the MuSK receptor to endosomes initiates a signaling cascade to align pre- with postsynaptic elements. Collectively, these findings suggest a general mechanism by which Wnt signals shape synaptic connectivity through localized receptor endocytosis. PMID:22318632

  6. Plakophilin-1, a Novel Wnt Signaling Regulator, Is Critical for Tooth Development and Ameloblast Differentiation.

    PubMed

    Miyazaki, Kanako; Yoshizaki, Keigo; Arai, Chieko; Yamada, Aya; Saito, Kan; Ishikawa, Masaki; Xue, Han; Funada, Keita; Haruyama, Naoto; Yamada, Yoshihiko; Fukumoto, Satoshi; Takahashi, Ichiro

    2016-01-01

    Tooth morphogenesis is initiated by reciprocal interactions between the ectoderm and neural crest-derived mesenchyme, and the Wnt signaling pathway is involved in this process. We found that Plakophilin (PKP)1, which is associated with diseases such as ectodermal dysplasia/skin fragility syndrome, was highly expressed in teeth and skin, and was upregulated during tooth development. We hypothesized that PKP1 regulates Wnt signaling via its armadillo repeat domain in a manner similar to β-catenin. To determine its role in tooth development, we performed Pkp1 knockdown experiments using ex vivo organ cultures and cell cultures. Loss of Pkp1 reduced the size of tooth germs and inhibited dental epithelial cell proliferation, which was stimulated by Wnt3a. Furthermore, transfected PKP1-emerald green fluorescent protein was translocated from the plasma membrane to the nucleus upon stimulation with Wnt3a and LiCl, which required the PKP1 N terminus (amino acids 161 to 270). Localization of PKP1, which is known as an adhesion-related desmosome component, shifted to the plasma membrane during ameloblast differentiation. In addition, Pkp1 knockdown disrupted the localization of Zona occludens 1 in tight junctions and inhibited ameloblast differentiation; the two proteins were shown to directly interact by immunoprecipitation. These results implicate the participation of PKP1 in early tooth morphogenesis as an effector of canonical Wnt signaling that controls ameloblast differentiation via regulation of the cell adhesion complex. PMID:27015268

  7. Wnt7a stimulates myogenic stem cell motility and engraftment resulting in improved muscle strength.

    PubMed

    Bentzinger, C Florian; von Maltzahn, Julia; Dumont, Nicolas A; Stark, Danny A; Wang, Yu Xin; Nhan, Kevin; Frenette, Jérôme; Cornelison, D D W; Rudnicki, Michael A

    2014-04-14

    Wnt7a/Fzd7 signaling stimulates skeletal muscle growth and repair by inducing the symmetric expansion of satellite stem cells through the planar cell polarity pathway and by activating the Akt/mTOR growth pathway in muscle fibers. Here we describe a third level of activity where Wnt7a/Fzd7 increases the polarity and directional migration of mouse satellite cells and human myogenic progenitors through activation of Dvl2 and the small GTPase Rac1. Importantly, these effects can be exploited to potentiate the outcome of myogenic cell transplantation into dystrophic muscles. We observed that a short Wnt7a treatment markedly stimulated tissue dispersal and engraftment, leading to significantly improved muscle function. Moreover, myofibers at distal sites that fused with Wnt7a-treated cells were hypertrophic, suggesting that the transplanted cells deliver activated Wnt7a/Fzd7 signaling complexes to recipient myofibers. Taken together, we describe a viable and effective ex vivo cell modulation process that profoundly enhances the efficacy of stem cell therapy for skeletal muscle. PMID:24711502

  8. Wnt7a stimulates myogenic stem cell motility and engraftment resulting in improved muscle strength

    PubMed Central

    Bentzinger, C. Florian; von Maltzahn, Julia; Dumont, Nicolas A.; Stark, Danny A.; Wang, Yu Xin; Nhan, Kevin; Frenette, Jérôme; Cornelison, DDW

    2014-01-01

    Wnt7a/Fzd7 signaling stimulates skeletal muscle growth and repair by inducing the symmetric expansion of satellite stem cells through the planar cell polarity pathway and by activating the Akt/mTOR growth pathway in muscle fibers. Here we describe a third level of activity where Wnt7a/Fzd7 increases the polarity and directional migration of mouse satellite cells and human myogenic progenitors through activation of Dvl2 and the small GTPase Rac1. Importantly, these effects can be exploited to potentiate the outcome of myogenic cell transplantation into dystrophic muscles. We observed that a short Wnt7a treatment markedly stimulated tissue dispersal and engraftment, leading to significantly improved muscle function. Moreover, myofibers at distal sites that fused with Wnt7a-treated cells were hypertrophic, suggesting that the transplanted cells deliver activated Wnt7a/Fzd7 signaling complexes to recipient myofibers. Taken together, we describe a viable and effective ex vivo cell modulation process that profoundly enhances the efficacy of stem cell therapy for skeletal muscle. PMID:24711502

  9. Noncanonical Wnt signaling pathways in C. elegans converge on POP-1/TCF and control cell polarity.

    PubMed

    Herman, Michael A; Wu, Mingfu

    2004-05-01

    In the nematode Caenorhabditis elegans, a canonical Wnt signaling pathway controls a cell migration whereas noncanonical Wnt pathways control the polarities of individual cells. Despite the differences in the identities and interactions among canonical and noncanonical Wnt pathway components, as well as the processes they regulate, almost all C. elegans Wnt pathways involve the sole Tcf homolog, POP-1. Intriguingly, POP-1 is asymmetrically distributed between the daughters of an asymmetric cell division, with the anterior sister cell usually having a higher level of nuclear POP-1 than its posterior sister. At some divisions, asymmetric distribution of POP-1 is controlled by noncanonical Wnt signaling, but at others the asymmetry is generated independently. Recent experiments suggest that despite this elaborate anterior-posterior POP-1 asymmetry, the quantity of POP-1 protein may have less to do with the subsequent determination of fate than does the quality of the POP-1 protein in the cell. In this review, we will embark on a quest to understand Quality (1), at least from the standpoint of the effect POP/Tcf quality has on the control of cell polarity in C. elegans. PMID:14977564

  10. Plakophilin-1, a Novel Wnt Signaling Regulator, Is Critical for Tooth Development and Ameloblast Differentiation

    PubMed Central

    Arai, Chieko; Yamada, Aya; Saito, Kan; Ishikawa, Masaki; Xue, Han; Funada, Keita; Haruyama, Naoto; Yamada, Yoshihiko; Fukumoto, Satoshi; Takahashi, Ichiro

    2016-01-01

    Tooth morphogenesis is initiated by reciprocal interactions between the ectoderm and neural crest-derived mesenchyme, and the Wnt signaling pathway is involved in this process. We found that Plakophilin (PKP)1, which is associated with diseases such as ectodermal dysplasia/skin fragility syndrome, was highly expressed in teeth and skin, and was upregulated during tooth development. We hypothesized that PKP1 regulates Wnt signaling via its armadillo repeat domain in a manner similar to β-catenin. To determine its role in tooth development, we performed Pkp1 knockdown experiments using ex vivo organ cultures and cell cultures. Loss of Pkp1 reduced the size of tooth germs and inhibited dental epithelial cell proliferation, which was stimulated by Wnt3a. Furthermore, transfected PKP1-emerald green fluorescent protein was translocated from the plasma membrane to the nucleus upon stimulation with Wnt3a and LiCl, which required the PKP1 N terminus (amino acids 161 to 270). Localization of PKP1, which is known as an adhesion-related desmosome component, shifted to the plasma membrane during ameloblast differentiation. In addition, Pkp1 knockdown disrupted the localization of Zona occludens 1 in tight junctions and inhibited ameloblast differentiation; the two proteins were shown to directly interact by immunoprecipitation. These results implicate the participation of PKP1 in early tooth morphogenesis as an effector of canonical Wnt signaling that controls ameloblast differentiation via regulation of the cell adhesion complex. PMID:27015268

  11. Specification of hepatopancreas progenitors in zebrafish by hnf1ba and wnt2bb

    PubMed Central

    Lancman, Joseph J.; Zvenigorodsky, Natasha; Gates, Keith P.; Zhang, Danhua; Solomon, Keely; Humphrey, Rohan K.; Kuo, Taiyi; Setiawan, Linda; Verkade, Heather; Chi, Young-In; Jhala, Ulupi S.; Wright, Christopher V. E.; Stainier, Didier Y. R.; Dong, P. Duc Si

    2013-01-01

    Although the liver and ventral pancreas are thought to arise from a common multipotent progenitor pool, it is unclear whether these progenitors of the hepatopancreas system are specified by a common genetic mechanism. Efforts to determine the role of Hnf1b and Wnt signaling in this crucial process have been confounded by a combination of factors, including a narrow time frame for hepatopancreas specification, functional redundancy among Wnt ligands, and pleiotropic defects caused by either severe loss of Wnt signaling or Hnf1b function. Using a novel hypomorphic hnf1ba zebrafish mutant that exhibits pancreas hypoplasia, as observed in HNF1B monogenic diabetes, we show that hnf1ba plays essential roles in regulating β-cell number and pancreas specification, distinct from its function in regulating pancreas size and liver specification, respectively. By combining Hnf1ba partial loss of function with conditional loss of Wnt signaling, we uncover a crucial developmental window when these pathways synergize to specify the entire ventrally derived hepatopancreas progenitor population. Furthermore, our in vivo genetic studies demonstrate that hnf1ba generates a permissive domain for Wnt signaling activity in the foregut endoderm. Collectively, our findings provide a new model for HNF1B function, yield insight into pancreas and β-cell development, and suggest a new mechanism for hepatopancreatic specification. PMID:23720049

  12. A Genome-Wide RNA Interference Screen Identifies a Role for Wnt/β-Catenin Signaling during Rift Valley Fever Virus Infection

    PubMed Central

    Harmon, Brooke; Bird, Sara W.; Schudel, Benjamin R.; Hatch, Anson V.; Rasley, Amy

    2016-01-01

    ABSTRACT Rift Valley fever virus (RVFV) is an arbovirus within the Bunyaviridae family capable of causing serious morbidity and mortality in humans and livestock. To identify host factors involved in bunyavirus replication, we employed genome-wide RNA interference (RNAi) screening and identified 381 genes whose knockdown reduced infection. The Wnt pathway was the most represented pathway when gene hits were functionally clustered. With further investigation, we found that RVFV infection activated Wnt signaling, was enhanced when Wnt signaling was preactivated, was reduced with knockdown of β-catenin, and was blocked using Wnt signaling inhibitors. Similar results were found using distantly related bunyaviruses La Crosse virus and California encephalitis virus, suggesting a conserved role for Wnt signaling in bunyaviral infection. We propose a model where bunyaviruses activate Wnt-responsive genes to regulate optimal cell cycle conditions needed to promote efficient viral replication. The findings in this study should aid in the design of efficacious host-directed antiviral therapeutics. IMPORTANCE RVFV is a mosquito-borne bunyavirus that is endemic to Africa but has demonstrated a capacity for emergence in new territories (e.g., the Arabian Peninsula). As a zoonotic pathogen that primarily affects livestock, RVFV can also cause lethal hemorrhagic fever and encephalitis in humans. Currently, there are no treatments or fully licensed vaccines for this virus. Using high-throughput RNAi screening, we identified canonical Wnt signaling as an important host pathway regulating RVFV infection. The beneficial role of Wnt signaling was observed for RVFV, along with other disparate bunyaviruses, indicating a conserved bunyaviral replication mechanism involving Wnt signaling. These studies supplement our knowledge of the fundamental mechanisms of bunyavirus infection and provide new avenues for countermeasure development against pathogenic bunyaviruses. PMID:27226375

  13. Delivery of the Porcupine Inhibitor WNT974 in Mice

    PubMed Central

    Zhang, Li-shu; Lum, Lawrence

    2016-01-01

    We describe here a technique for delivering the porcupine inhibitor WNT974 (formerly LGK974) in mice. The protocol entails once-a-day oral delivery of WNT974 for up to 3 months at a concentration sufficient to achieve systemic Wnt pathway inhibition with limited toxicity as measured by weight change. This route of delivery enables extended durations of Wnt signaling inhibition in a mammalian model organism. PMID:27590157

  14. Delivery of the Porcupine Inhibitor WNT974 in Mice.

    PubMed

    Zhang, Li-Shu; Lum, Lawrence

    2016-01-01

    We describe here a technique for delivering the porcupine inhibitor WNT974 (formerly LGK974) in mice. The protocol entails once-a-day oral delivery of WNT974 for up to 3 months at a concentration sufficient to achieve systemic Wnt pathway inhibition with limited toxicity as measured by weight change. This route of delivery enables extended durations of Wnt signaling inhibition in a mammalian model organism. PMID:27590157

  15. Fibroblast growth factor and canonical WNT/β-catenin signaling cooperate in suppression of chondrocyte differentiation in experimental models of FGFR signaling in cartilage.

    PubMed

    Buchtova, Marcela; Oralova, Veronika; Aklian, Anie; Masek, Jan; Vesela, Iva; Ouyang, Zhufeng; Obadalova, Tereza; Konecna, Zaneta; Spoustova, Tereza; Pospisilova, Tereza; Matula, Petr; Varecha, Miroslav; Balek, Lukas; Gudernova, Iva; Jelinkova, Iva; Duran, Ivan; Cervenkova, Iveta; Murakami, Shunichi; Kozubik, Alois; Dvorak, Petr; Bryja, Vitezslav; Krejci, Pavel

    2015-05-01

    Aberrant fibroblast growth factor (FGF) signaling disturbs chondrocyte differentiation in skeletal dysplasia, but the mechanisms underlying this process remain unclear. Recently, FGF was found to activate canonical WNT/β-catenin pathway in chondrocytes via Erk MAP kinase-mediated phosphorylation of WNT co-receptor Lrp6. Here, we explore the cellular consequences of such a signaling interaction. WNT enhanced the FGF-mediated suppression of chondrocyte differentiation in mouse limb bud micromass and limb organ cultures, leading to inhibition of cartilage nodule formation in micromass cultures, and suppression of growth in cultured limbs. Simultaneous activation of the FGF and WNT/β-catenin pathways resulted in loss of chondrocyte extracellular matrix, expression of genes typical for mineralized tissues and alteration of cellular shape. WNT enhanced the FGF-mediated downregulation of chondrocyte proteoglycan and collagen extracellular matrix via inhibition of matrix synthesis and induction of proteinases involved in matrix degradation. Expression of genes regulating RhoA GTPase pathway was induced by FGF in cooperation with WNT, and inhibition of the RhoA signaling rescued the FGF/WNT-mediated changes in chondrocyte cellular shape. Our results suggest that aberrant FGF signaling cooperates with WNT/β-catenin in suppression of chondrocyte differentiation. PMID:25558817

  16. Hairy Math: Add Wnt-3a to Multiply Bulge Cells

    PubMed Central

    Hossain, M. Zulfiquer; Garza, Luis A.

    2015-01-01

    Canonical Wnt signals are important for activation of epithelial skin stem cells, but the role of individual Wnt ligands remains uncertain. Ouji et al. demonstrate a key role for Wnt-3a in partial maintenance and long-term expansion of epithelial skin stem cells in vitro. They also report a method for expanding these cells in vitro without feeder cells. PMID:25964269

  17. Citrobacter Infection and Wnt signaling

    PubMed Central

    Umar, Shahid

    2012-01-01

    Gut flora generally contributes to a healthy environment while both commensal and pathogenic bacteria that influence the innate and adaptive immune responses, can cause acute and/or chronic mucosal inflammation. Citrobacter rodentium (C. rodentium) is a member of the family of enteropathogens that provide an excellent in vivo model to investigate the host-pathogen interactions in real-time. It is the etiologic agent for transmissible murine colonic hyperplasia (TMCH) while inflammation following C. rodentium infection is dependent upon the genetic background. Ongoing and completed studies in this model have so far established that Wnt/β-catenin, Notch and PI3K pathways regulate colonic crypt hyperplasia while epithelial-stromal cross-talk, mediated by MEK/ERK/NF-κB signaling, regulates inflammation and/or colitis in susceptible strains. The C. rodentium-induced hyperplastic state also increases the susceptibility to either mutagenic insult or in mice heterozygous for Apc gene. The ability to modulate the host response to C. rodentium infection therefore provides an opportunity to delineate the mechanisms that determine mucosal hyperplasia, intestinal inflammation, and/or neoplasia as disease outcomes. PMID:24358033

  18. Pentagalloylglucose Blocks the Nuclear Transport and the Process of Nucleocapsid Egress to Inhibit HSV-1 Infection.

    PubMed

    Jin, Fujun; Ma, Kaiqi; Chen, Maoyun; Zou, Muping; Wu, Yanting; Li, Feng; Wang, Yifei

    2016-03-23

    Herpes simplex virus type 1 (HSV-1), a widespread virus, causes a variety of human viral diseases worldwide. The serious threat of drug-resistance highlights the extreme urgency to develop novel antiviral drugs with different mechanisms of action. Pentagalloylglucose (PGG) is a natural polyphenolic compound with significant anti-HSV activity; however, the mechanisms underlying its antiviral activity need to be defined by further studies. In this study, we found that PGG treatment delays the nuclear transport process of HSV-1 particles by inhibiting the upregulation of dynein (a cellular major motor protein) induced by HSV-1 infection. Furthermore, PGG treatment affects the nucleocapsid egress of HSV-1 by inhibiting the expression and disrupting the cellular localization of pEGFP-UL31 and pEGFP-UL34, which are indispensable for HSV-1 nucleocapsid egress from the nucleus. However, the over-expression of pEGFP-UL31 and pEGFP-UL34 could decrease the antiviral effect of PGG. In this study, for the first time, the antiviral activity of PGG against acyclovir-resistant virus was demonstrated in vitro, and the possible mechanisms of its anti-HSV activities were identified based on the inhibition of nuclear transport and nucleocapsid egress in HSV-1. It was further confirmed that PGG could be a promising candidate for HSV therapy, especially for drug-resistant strains. PMID:26166506

  19. Processing Effects on Block-Copolymer Based Pressure-Sensitive Adhesives

    NASA Astrophysics Data System (ADS)

    O'Connor, A. E.; Macosko, C. W.

    2000-03-01

    The goal of this work is to investigate how the variables in the hot-melt coating process affect the microstructure and properties of pressure-sensitive adhesives based on a styrene isoprene styrene triblock copolymer. This polymer is a thermoplastic elastomer, able to be coated at high temperatures and physically crosslinked at lower temperatures. Adhesive tape samples have been made through hot-melt and solvent coating methods. Hot-melt coatings are prepared at speeds up to 110 feet/minute. Materials with the same thermal history have been coated using both methods and then tested for comparison of properties. PSA properties are strongly dependent on the time scale of application and debonding, as revealed by shear rheology data, and three types of performance tests (tack, peel, and shear holding power) are used to capture the various responses. Solvent-coated tape has superior shear strength, while hot-melt-coated tape performs better in peel tests. It is expected that the varying flow and deformation histories of the samples will lead to distinct chain orientations, while the rate of cooling of hot-melt-coated samples may influence the degree of phase separation achieved. These factors will cause the adhesive coatings to have different microstructures and therefore different properties.

  20. Simulation of plasma based semiconductor processing using block structured locally refined grids

    SciTech Connect

    Wake, D.D.

    1998-01-01

    We have described a new numerical method for plasma simulation. Calculations have been presented which show that the method is accurate and suggest the regimes in which the method provides savings in CPU time and memory requirements. A steady state simulation of a four centimeter domain was modeled with sheath scale (150 microns) resolution using only 40 grid points. Simulations of semiconductor processing equipment have been performed which imply the usefulness of the method for engineering applications. It is the author`s opinion that these accomplishments represent a significant contribution to plasma simulation and the efficient numerical solution of certain systems of non-linear partial differential equations. More work needs to be done, however, for the algorithm to be of practical use in an engineering environment. Despite our success at avoiding the dielectric relaxation timestep restrictions the algorithm is still conditionally stable and requires timesteps which are relatively small. This represents a prohibitive runtime for steady state solutions on high resolution grids. Current research suggests that these limitations may be overcome and the use of much larger timesteps will be possible.

  1. A Second WNT for Old Drugs: Drug Repositioning against WNT-Dependent Cancers

    PubMed Central

    Ahmed, Kamal; Shaw, Holly V.; Koval, Alexey; Katanaev, Vladimir L.

    2016-01-01

    Aberrant WNT signaling underlies cancerous transformation and growth in many tissues, such as the colon, breast, liver, and others. Downregulation of the WNT pathway is a desired mode of development of targeted therapies against these cancers. Despite the urgent need, no WNT signaling-directed drugs currently exist, and only very few candidates have reached early phase clinical trials. Among different strategies to develop WNT-targeting anti-cancer therapies, repositioning of existing drugs previously approved for other diseases is a promising approach. Nonsteroidal anti-inflammatory drugs like aspirin, the anti-leprotic clofazimine, and the anti-trypanosomal suramin are among examples of drugs having recently revealed WNT-targeting activities. In total, 16 human-use drug compounds have been found to be working through the WNT pathway and show promise for their prospective repositioning against various cancers. Advances, hurdles, and prospects of developing these molecules as potential drugs against WNT-dependent cancers, as well as approaches for discovering new ones for repositioning, are the foci of the current review. PMID:27429001

  2. Gene regulatory networks mediating canonical Wnt signal directed control of pluripotency and differentiation in embryo stem cells

    PubMed Central

    Zhang, Xiaoxiao; Peterson, Kevin A.; Liu, X. Shirley; McMahon, Andrew P.; Ohba, Shinsuke

    2013-01-01

    Canonical Wnt signaling supports the pluripotency of embryonic stem cells (ESCs) but also promotes differentiation of early mammalian cell lineages. To explain these paradoxical observations, we explored the gene regulatory networks at play. Canonical Wnt signaling is intertwined with the pluripotency network comprising Nanog, Oct4, and Sox2 in mouse ESCs. In defined media supporting the derivation and propagation of ESCs, Tcf3 and β-catenin interact with Oct4; Tcf3 binds to Sox motif within Oct-Sox composite motifs that are also bound by Oct4-Sox2 complexes. Further, canonical Wnt signaling up-regulates the activity of the Pou5f1 distal enhancer via the Sox motif in ESCs. When viewed in the context of published studies on Tcf3 and β-catenin mutants, our findings suggest Tcf3 counters pluripotency by competition with Sox2 at these sites, and Tcf3 inhibition is blocked by β-catenin entry into this complex. Wnt pathway stimulation also triggers β-catenin association at regulatory elements with classic Lef/Tcf motifs associated with differentiation programs. The failure to activate these targets in the presence of a MEK/ERK inhibitor essential for ESC culture suggests MEK/ERK signaling and canonical Wnt signaling combine to promote ESC differentiation. PMID:23505158

  3. Interfering with Wnt signalling alters the periodicity of the segmentation clock

    PubMed Central

    Gibb, Sarah; Zagorska, Anna; Melton, Kristin; Tenin, Gennady; Vacca, Irene; Trainor, Paul; Maroto, Miguel; Dale, J. Kim

    2009-01-01

    Somites are embryonic precursors of the ribs, vertebrae and certain dermis tissue. Somite formation is a periodic process regulated by a molecular clock which drives cyclic expression of a number of clock genes in the presomitic mesoderm. To date the mechanism regulating the period of clock gene oscillations is unknown. Here we show that chick homologues of the Wnt pathway genes that oscillate in mouse do not cycle across the chick presomitic mesoderm. Strikingly we find that modifying Wnt signalling changes the period of Notch driven oscillations in both mouse and chick but these oscillations continue. We propose that the Wnt pathway is a conserved mechanism that is involved in regulating the period of cyclic gene oscillations in the presomitic mesoderm. PMID:19272372

  4. Activation of the canonical Wnt/{beta}-catenin pathway enhances monocyte adhesion to endothelial cells

    SciTech Connect

    Lee, Dong Kun . E-mail: leedk@memorialhealthsource.com; Nathan Grantham, R.; Trachte, Aaron L.; Mannion, John D.; Wilson, Colleen L.

    2006-08-18

    Monocyte adhesion to vascular endothelium has been reported to be one of the early processes in the development of atherosclerosis. In an attempt to develop strategies to prevent or delay atherosclerosis progression, we analyzed effects of the Wnt/{beta}-catenin signaling pathway on monocyte adhesion to various human endothelial cells. Adhesion of fluorescein-labeled monocytes to various human endothelial cells was analyzed under a fluorescent microscope. Unlike sodium chloride, lithium chloride enhanced monocyte adhesion to endothelial cells in a dose-dependent manner. We further demonstrated that inhibitors for glycogen synthase kinase (GSK)-3{beta} or proteosome enhanced monocyte-endothelial cell adhesion. Results of semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) indicated that activation of Wnt/{beta}-catenin pathway did not change expression levels of mRNA for adhesion molecules. In conclusion, the canonical Wnt/{beta}-catenin pathway enhanced monocyte-endothelial cell adhesion without changing expression levels of adhesion molecules.

  5. Parkin protects dopaminergic neurons from excessive Wnt/{beta}-catenin signaling

    SciTech Connect

    Rawal, Nina; Corti, Olga; Sacchetti, Paola; Ardilla-Osorio, Hector; Sehat, Bita; Brice, Alexis; Arenas, Ernest

    2009-10-23

    Parkinson's disease (PD) is caused by degeneration of the dopaminergic (DA) neurons of the substantia nigra but the molecular mechanisms underlying the degenerative process remain elusive. Several reports suggest that cell cycle deregulation in post-mitotic neurons could lead to neuronal cell death. We now show that Parkin, an E3 ubiquitin ligase linked to familial PD, regulates {beta}-catenin protein levels in vivo. Stabilization of {beta}-catenin in differentiated primary ventral midbrain neurons results in increased levels of cyclin E and proliferation, followed by increased levels of cleaved PARP and loss of DA neurons. Wnt3a signaling also causes death of post-mitotic DA neurons in parkin null animals, suggesting that both increased stabilization and decreased degradation of {beta}-catenin results in DA cell death. These findings demonstrate a novel regulation of Wnt signaling by Parkin and suggest that Parkin protects DA neurons against excessive Wnt signaling and {beta}-catenin-induced cell death.

  6. Wnt Signaling in Cell Motility and Invasion: Drawing Parallels between Development and Cancer.

    PubMed

    Sedgwick, Alanna E; D'Souza-Schorey, Crislyn

    2016-01-01

    The importance of canonical and non-canonical Wnt signal transduction cascades in embryonic development and tissue homeostasis is well recognized. The aberrant activation of these pathways in the adult leads to abnormal cellular behaviors, and tumor progression is frequently a consequence. Here we discuss recent findings and analogies between Wnt signaling in developmental processes and tumor progression, with a particular focus on cell motility and matrix invasion and highlight the roles of the ARF (ADP-Ribosylation Factor) and Rho-family small GTP-binding proteins. Wnt-regulated signal transduction from cell surface receptors, signaling endosomes and/or extracellular vesicles has the potential to profoundly influence cell movement, matrix degradation and paracrine signaling in both development and disease. PMID:27589803

  7. Induction of WNT11 by hypoxia and hypoxia-inducible factor-1α regulates cell proliferation, migration and invasion

    PubMed Central

    Mori, Hiroyuki; Yao, Yao; Learman, Brian S.; Kurozumi, Kazuhiko; Ishida, Joji; Ramakrishnan, Sadeesh K.; Overmyer, Katherine A.; Xue, Xiang; Cawthorn, William P.; Reid, Michael A.; Taylor, Matthew; Ning, Xiaomin; Shah, Yatrik M.; MacDougald, Ormond A.

    2016-01-01

    Changes in cellular oxygen tension play important roles in physiological processes including development and pathological processes such as tumor promotion. The cellular adaptations to sustained hypoxia are mediated by hypoxia-inducible factors (HIFs) to regulate downstream target gene expression. With hypoxia, the stabilized HIF-α and aryl hydrocarbon receptor nuclear translocator (ARNT, also known as HIF-β) heterodimer bind to hypoxia response elements (HREs) and regulate expression of target genes. Here, we report that WNT11 is induced by hypoxia in many cell types, and that transcription of WNT11 is regulated primarily by HIF-1α. We observed induced WNT11 expression in the hypoxic area of allograft tumors. In addition, in mice bearing orthotopic malignant gliomas, inhibition with bevacizumab of vascular endothelial growth factor, which is an important stimulus for angiogenesis, increased nuclear HIF-1α and HIF-2α, and expression of WNT11. Gain- and loss-of-function approaches revealed that WNT11 stimulates proliferation, migration and invasion of cancer-derived cells, and increases activity of matrix metalloproteinase (MMP)-2 and 9. Since tumor hypoxia has been proposed to increase tumor aggressiveness, these data suggest WNT11 as a possible target for cancer therapies, especially for tumors treated with antiangiogenic therapy. PMID:26861754

  8. Daam2-PIP5K is a regulatory pathway for Wnt signaling and therapeutic target for remyelination in the CNS.

    PubMed

    Lee, Hyun Kyoung; Chaboub, Lesley S; Zhu, Wenyi; Zollinger, Daniel; Rasband, Matthew N; Fancy, Stephen P J; Deneen, Benjamin

    2015-03-18

    Wnt signaling plays an essential role in developmental and regenerative myelination of the CNS; however, contributions of proximal regulators of the Wnt receptor complex to these processes remain undefined. To identify components of the Wnt pathway that regulate these processes, we applied a multifaceted discovery platform and found that Daam2-PIP5K comprise a novel pathway regulating Wnt signaling and myelination. Using dorsal patterning of the chick spinal cord we found that Daam2 promotes Wnt signaling and receptor complex formation through PIP5K-PIP2. Analysis of Daam2 function in oligodendrocytes (OLs) revealed that it suppresses OL differentiation during development, after white matter injury (WMI), and is expressed in human white matter lesions. These findings suggest a pharmacological strategy to inhibit Daam2-PIP5K function, application of which stimulates remyelination after WMI. Put together, our studies integrate information from multiple systems to identify a novel regulatory pathway for Wnt signaling and potential therapeutic target for WMI. PMID:25754822

  9. Daam2-PIP5K Is a Regulatory Pathway for Wnt Signaling and Therapeutic Target for Remyelination in the CNS

    PubMed Central

    Lee, Hyun Kyoung; Chaboub, Lesley S.; Zhu, Wenyi; Zollinger, Daniel; Rasband, Matthew N.; Fancy, Stephen P.J.; Deneen, Benjamin

    2015-01-01

    SUMMARY Wnt signaling plays an essential role in developmental and regenerative myelination of the CNS; however, contributions of proximal regulators of the Wnt receptor complex to these processes remain undefined. To identify components of the Wnt pathway that regulate these processes, we applied a multifaceted discovery platform and found that Daam2-PIP5K comprise a novel pathway regulating Wnt signaling and myelination. Using dorsal patterning of the chick spinal cord we found that Daam2 promotes Wnt signaling and receptor complex formation through PIP5K-PIP2. Analysis of Daam2 function in oligodendrocytes (OLs) revealed that it suppresses OL differentiation during development, after white matter injury (WMI), and is expressed in human white matter lesions. These findings suggest a pharmacological strategy to inhibit Daam2-PIP5K function, application of which stimulates remyelination after WMI. Put together, our studies integrate information from multiple systems to identify a novel regulatory pathway for Wnt signaling and potential therapeutic target for WMI. PMID:25754822

  10. Block co-polymer approach for CD uniformity and placement error improvement in DSA hole grapho-epitaxy process

    NASA Astrophysics Data System (ADS)

    Matsumiya, Tasuku; Kurosawa, Tsuyoshi; Yahagi, Masahito; Yamano, Hitoshi; Miyagi, Ken; Maehashi, Takaya; Suzuki, Issei; Kawaue, Akiya; Komuro, Yoshitaka; Hirayama, Taku; Ohmori, Katsumi

    2015-03-01

    Directed Self-Assembly (DSA) of Block Co-Polymer (BCP) with conventional lithography is being thought as one of the potential patterning solution for future generation devices manufacturing. Many studies have been reported to fabricate the aligned patterns both on grapho and chemoepitaxy for semiconductor application1, 2. The hole shrink and multiplication by graphoepitaxy are one of the DSA implementation candidates in terms of relatively realistic process and versatility of chip design. The critical challenges on hole shrink and multiplication by using conventional Poly (styrene-b-methyl methacrylate) (PS-b-PMMA) BCP have been reported such as CD uniformity, placement error3 and defectivity. It is needed to overcome these challenging issues by improving not only whole process but materials. From the material aspect, the surface treatment material for guide structure, and process friendly BCP material are key development items on graphoepitaxy. In this paper, it will be shown in BCP approach about conventional PS-b-PMMA with additives and new casting solvent as PS-b-PMMA extension for CD uniformity and placement error improvement and then it'll be discussed on what is the key factor and solution from BCP material approach.

  11. Wnt and the Cancer Niche: Paracrine Interactions with Gastrointestinal Cancer Cells Undergoing Asymmetric Cell Division

    PubMed Central

    Xin, Hong-Wu; Ambe, Chenwi M.; Ray, Satyajit; Kim, Bo-Kyu; Koizumi, Tomotake; Wiegand, Gordon W.; Hari, Danielle; Mullinax, John E.; Jaiswal, Kshama R.; Garfield, Susan H.; Stojadinovic, Alexander; Rudloff, Udo; Thorgeirsson, Snorri S.; Avital, Itzhak

    2013-01-01

    Objective: Stem-like cancer cells contribute to cancer initiation and maintenance. Stem cells can self-renew by asymmetric cell division (ACD). ACD with non-random chromosomal cosegregation (ACD-NRCC) is one possible self-renewal mechanism. There is a paucity of evidence supporting ACD-NRCC in human cancer. Our aim was to investigate ACD-NRCC and its potential interactions with the cancer niche (microenvironment) in gastrointestinal cancers. Design: We used DNA double and single labeling approaches with FACS to isolate live cells undergoing ACD-NRCC. Results: Gastrointestinal cancers contain rare subpopulations of cells capable of ACD-NRCC. ACD-NRCC was detected preferentially in subpopulations of cells previously suggested to be stem-like/tumor-initiating cancer cells. ACD-NRCC was independent of cell-to-cell contact, and was regulated by the cancer niche in a heat-sensitive paracrine fashion. Wnt pathway genes and proteins are differentially expressed in cells undergoing ACD-NRCC vs. symmetric cell division. Blocking the Wnt pathway with IWP2 (WNT antagonist) or siRNA-TCF4 resulted in suppression of ACD-NRCC. However, using a Wnt-agonist did not increase the relative proportion of cells undergoing ACD-NRCC. Conclusion: Gastrointestinal cancers contain subpopulations of cells capable of ACD-NRCC. Here we show for the first time that ACD-NRCC can be regulated by the Wnt pathway, and by the cancer niche in a paracrine fashion. However, whether ACD-NRCC is exclusively associated with stem-like cancer cells remains to be determined. Further study of these findings might generate novel insights into stem cell and cancer biology. Targeting the mechanism of ACD-NRCC might engender novel approaches for cancer therapy. PMID:23901343

  12. Wnt Signaling in Neuromuscular Junction Development

    PubMed Central

    Koles, Kate

    2012-01-01

    Wnt proteins are best known for their profound roles in cell patterning, because they are required for the embryonic development of all animal species studied to date. Besides regulating cell fate, Wnt proteins are gaining increasing recognition for their roles in nervous system development and function. New studies indicate that multiple positive and negative Wnt signaling pathways take place simultaneously during the formation of vertebrate and invertebrate neuromuscular junctions. Although some Wnts are essential for the formation of NMJs, others appear to play a more modulatory role as part of multiple signaling pathways. Here we review the most recent findings regarding the function of Wnts at the NMJ from both vertebrate and invertebrate model systems. PMID:22510459

  13. Wnt signaling and tbx16 form a bistable switch to commit bipotential progenitors to mesoderm

    PubMed Central

    Bouldin, Cortney M.; Manning, Alyssa J.; Peng, Yu-Hsuan; Farr, Gist H.; Hung, King L.; Dong, Alice; Kimelman, David

    2015-01-01

    Anterior to posterior growth of the vertebrate body is fueled by a posteriorly located population of bipotential neuro-mesodermal progenitor cells. These progenitors have a limited rate of proliferation and their maintenance is crucial for completion of the anterior-posterior axis. How they leave the progenitor state and commit to differentiation is largely unknown, in part because widespread modulation of factors essential for this process causes organism-wide effects. Using a novel assay, we show that zebrafish Tbx16 (Spadetail) is capable of advancing mesodermal differentiation cell-autonomously. Tbx16 locks cells into the mesodermal state by not only activating downstream mesodermal genes, but also by repressing bipotential progenitor genes, in part through a direct repression of sox2. We demonstrate that tbx16 is activated as cells move from an intermediate Wnt environment to a high Wnt environment, and show that Wnt signaling activates the tbx16 promoter. Importantly, high-level Wnt signaling is able to accelerate mesodermal differentiation cell-autonomously, just as we observe with Tbx16. Finally, because our assay for mesodermal commitment is quantitative we are able to show that the acceleration of mesodermal differentiation is surprisingly incomplete, implicating a potential separation of cell movement and differentiation during this process. Together, our data suggest a model in which high levels of Wnt signaling induce a transition to mesoderm by directly activating tbx16, which in turn acts to irreversibly flip a bistable switch, leading to maintenance of the mesodermal fate and repression of the bipotential progenitor state, even as cells leave the initial high-Wnt environment. PMID:26062939

  14. Alteration of canonical and non-canonical WNT-signaling by crystalline silica in human lung epithelial cells.

    PubMed

    Perkins, Timothy N; Dentener, Mieke A; Stassen, Frank R; Rohde, Gernot G; Mossman, Brooke T; Wouters, Emiel F M; Reynaert, Niki L

    2016-06-15

    Growth and development of the mature lung is a complex process orchestrated by a number of intricate developmental signaling pathways. Wingless-type MMTV-integration site (WNT) signaling plays critical roles in controlling branching morphogenesis cell differentiation, and formation of the conducting and respiratory airways. In addition, WNT pathways are often re-activated in mature lungs during repair and regeneration. WNT- signaling has been elucidated as a crucial contributor to the development of idiopathic pulmonary fibrosis as well as other hyper-proliferative lung diseases. Silicosis, a detrimental occupational lung disease caused by excessive inhalation of crystalline silica dust, is hallmarked by repeated cycles of damaging inflammation, epithelial hyperplasia, and formation of dense, hyalinized nodules of whorled collagen. However, mechanisms of epithelial cell hyperplasia and matrix deposition are not well understood, as most research efforts have focused on the pronounced inflammatory response. Microarray data from our previous studies has revealed a number of WNT-signaling and WNT-target genes altered by crystalline silica in human lung epithelial cells. In the present study, we utilize pathway analysis to designate connections between genes altered by silica in WNT-signaling networks. Furthermore, we confirm microarray findings by QRT-PCR and demonstrate both activation of canonical (β-catenin) and down-regulation of non-canonical (WNT5A) signaling in immortalized (BEAS-2B) and primary (PBEC) human bronchial epithelial cells. These findings suggest that WNT-signaling and cross-talk with other pathways (e.g. Notch), may contribute to proliferative, fibrogenic and inflammatory responses to silica in lung epithelial cells. PMID:27095093

  15. MicroRNA-27a-3p Inhibits Melanogenesis in Mouse Skin Melanocytes by Targeting Wnt3a.

    PubMed

    Zhao, Yuanyuan; Wang, Pengchao; Meng, Jinzhu; Ji, Yuankai; Xu, Dongmei; Chen, Tianzhi; Fan, Ruiwen; Yu, Xiuju; Yao, Jianbo; Dong, Changsheng

    2015-01-01

    MicroRNAs (miRNAs) play an essential role in the regulation of almost all the biological processes, including melanogenesis. MiR-27a-3p is nearly six times higher in white alpaca skin compared to brown skin, which indicates that miR-27a-3p may be a candidate regulator for melanogenesis. Wnt3a plays an important role in promoting melanoblasts to differentiate into melanocytes and melanogenesis. To confirm the function of miR-27a-3p to melanogenesis in mammals, miR-27a-3p mimic, inhibitor and their negative control were transfected into mouse melanocytes. As a result, miR-27a-3p inhibits melanogenesis by repressing Wnt3a at post-transcriptional level. A significant decrease in Wnt3a luciferase activity was observed in 293T cells co-transfected with the matched luciferase reporter vector and pre-miR-27a. Furthermore, the presence of exogenous miR-27a-3p significantly decreased Wnt3a protein expression rather than mRNA and reduced β-catenin mRNA levels in melanocytes. The over-expression of miR-27a-3p significantly increased the melanin content of melanocytes. However, miR-27a-3p inhibitor performs an opposite effect on melanogenesis. Wnt3a is one target of miR-27a-3p. MiR-27a-3p could inhibit Wnt3a protein amount by post-transcriptional regulation and melanogenesis in mouse melanocytes. Previous studies reported that Wnt3a promoted melanogenensis in mouse melanocytes. Thus, miR-27-3p inhibits melanogenesis by repressing Wnt3a protein expression. PMID:26006230

  16. MicroRNA-27a-3p Inhibits Melanogenesis in Mouse Skin Melanocytes by Targeting Wnt3a

    PubMed Central

    Zhao, Yuanyuan; Wang, Pengchao; Meng, Jinzhu; Ji, Yuankai; Xu, Dongmei; Chen, Tianzhi; Fan, Ruiwen; Yu, Xiuju; Yao, Jianbo; Dong, Changsheng

    2015-01-01

    MicroRNAs (miRNAs) play an essential role in the regulation of almost all the biological processes, including melanogenesis. MiR-27a-3p is nearly six times higher in white alpaca skin compared to brown skin, which indicates that miR-27a-3p may be a candidate regulator for melanogenesis. Wnt3a plays an important role in promoting melanoblasts to differentiate into melanocytes and melanogenesis. To confirm the function of miR-27a-3p to melanogenesis in mammals, miR-27a-3p mimic, inhibitor and their negative control were transfected into mouse melanocytes. As a result, miR-27a-3p inhibits melanogenesis by repressing Wnt3a at post-transcriptional level. A significant decrease in Wnt3a luciferase activity was observed in 293T cells co-transfected with the matched luciferase reporter vector and pre-miR-27a. Furthermore, the presence of exogenous miR-27a-3p significantly decreased Wnt3a protein expression rather than mRNA and reduced β-catenin mRNA levels in melanocytes. The over-expression of miR-27a-3p significantly increased the melanin content of melanocytes. However, miR-27a-3p inhibitor performs an opposite effect on melanogenesis. Wnt3a is one target of miR-27a-3p. MiR-27a-3p could inhibit Wnt3a protein amount by post-transcriptional regulation and melanogenesis in mouse melanocytes. Previous studies reported that Wnt3a promoted melanogenensis in mouse melanocytes. Thus, miR-27-3p inhibits melanogenesis by repressing Wnt3a protein expression. PMID:26006230

  17. Energy Efficient Heat Treatment for Linerless Hypereutectic Al-Si Engine Blocks Made Using Vacuum HPDC Process

    NASA Astrophysics Data System (ADS)

    Kasprzak, W.; Sokolowski, J. H.; Yamagata, H.; Aniolek, M.; Kurita, H.

    2011-02-01

    Heat treatment standards developed by the aluminum industry over the last several decades are often not well optimized when applied to components cast by high cooling rate processes such as High Pressure Die Casting (HPDC), Low Pressure Permanent Mold (LPPM), Squeeze Casting, etc. The inherently finer as-cast structures should not require long solution times for the effective dissolution of intermetallic phases and the adequate thermal modification of structural constituents. Hence, long and expensive T6 and T7 treatments should not be required. Heat treatment studies involving as-cast laboratory samples with SDAS = 13.6 μm (equivalent to a thick-section HPDC casting) were conducted. Traditional and modified solution and aging treatments were compared. These studies suggest that a reduction of up to 92% in thermal processing time is possible while maintaining and/or improving the cast component's metallurgical characteristics including hardness (≥75 HRB), dissolution of secondary phases, and spheroidization of the eutectic Si as well as overall homogeneity. Vacuum HPDC of an actual hypereutectic Al-20%Si motorcycle engine block confirmed the potential for significantly reduced heat treatment times, energy consumption, and overall costs.

  18. The Importance of the Canonical Wnt Signaling Pathway in the Porcine Endometrial Stromal Stem/Progenitor Cells: Implications for Regeneration

    PubMed Central

    Bukowska, Joanna; Ziecik, Adam Janusz; Laguna, Joanna; Gawronska-Kozak, Barbara

    2015-01-01

    The regenerative ability of the endometrium is strongly associated with the presence of adult stem/progenitor cells. Purposes of the present study were (1) to establish the presence of stem/progenitor cells in porcine endometrial stroma using a clonogenic assay and (2) to investigate whether the canonical Wnt pathway affects the potential of stem/progenitor cells to undergo self-renewal or differentiation. The utility of endometrial stromal clones as a model for stem/progenitor studies was evaluated based on these cells' increased expression of mesenchymal stem cell (MSC) marker genes, including CD29, CD73, CD90, and CD105, compared with primary cultured cells. Small molecules were introduced to activate (BIO) or inhibit (XAV939) the canonical Wnt pathway during stromal clone formation. Cloning efficiency assays revealed that activation of the Wnt/β-catenin pathway promoted formation of more differentiated small clones. Moreover, activation of the Wnt/β-catenin pathway decreased, whereas inhibition of the pathway increased MSC marker expression. Additionally, we confirmed the importance of canonical Wnt pathway stimulation in endometrial stromal cells through observing the appropriate changes in β-catenin cellular localization. These data indicate that modulation of the canonical Wnt pathway effects the process of regeneration in the porcine endometrium during the course of the estrous cycle. PMID:26414529

  19. The evolving roles of canonical WNT signaling in stem cells and tumorigenesis: Implications in targeted cancer therapies

    PubMed Central

    Yang, Ke; Wang, Xin; Zhang, Hongmei; Wang, Zhongliang; Nan, Guoxin; Li, Yasha; Zhang, Fugui; Mohammed, Maryam K.; Haydon, Rex C.; Luu, Hue H.; Bi, Yang; He, Tong-Chuan

    2015-01-01

    The canonical WNT/β-catenin signaling pathway governs a myriad of biological processes underlying development and maintenance of adult tissue homeostasis, including regulation of stem cell self-renewal, cell proliferation, differentiation, and apoptosis. WNTs are secreted lipid-modified glycoproteins that act as short-range ligands to activate receptor-mediated signaling pathways. The hallmark of the canonical pathway is the activation of β-catenin mediated transcriptional activity. Canonical WNTs control the β-catenin dynamics as the cytoplasmic level of β-catenin is tightly regulated via phosphorylation by the ‘destruction complex’, consisting of glycogen synthase kinase 3β (GSK3β), casein kinase 1α (CK1α), the scaffold protein AXIN, and the tumor suppressor adenomatous polyposis coli (APC). Aberrant regulation of this signaling cascade is associated with varieties of human diseases, especially cancers. Over the past decade, significant progress has been made in understanding the mechanisms of canonical WNT signaling. In this review, we focus on the current understanding of WNT signaling at the extracellular, cytoplasmic membrane, and intracellular/nuclear levels, including the emerging knowledge of crosstalk with other pathways. Recent progresses in developing novel WNT pathway-targeted therapies will also be reviewed. Thus, this review is intended to serve as a refresher of the current understanding about the physiologic and pathogenic roles of WNT/β-catenin signaling pathway, and to outline potential therapeutic opportunities by targeting the canonical WNT pathway. PMID:26618721

  20. The evolving roles of canonical WNT signaling in stem cells and tumorigenesis: implications in targeted cancer therapies.

    PubMed

    Yang, Ke; Wang, Xin; Zhang, Hongmei; Wang, Zhongliang; Nan, Guoxin; Li, Yasha; Zhang, Fugui; Mohammed, Maryam K; Haydon, Rex C; Luu, Hue H; Bi, Yang; He, Tong-Chuan

    2016-02-01

    The canonical WNT/β-catenin signaling pathway governs a myriad of biological processes underlying the development and maintenance of adult tissue homeostasis, including regulation of stem cell self-renewal, cell proliferation, differentiation, and apoptosis. WNTs are secreted lipid-modified glycoproteins that act as short-range ligands to activate receptor-mediated signaling pathways. The hallmark of the canonical pathway is the activation of β-catenin-mediated transcriptional activity. Canonical WNTs control the β-catenin dynamics as the cytoplasmic level of β-catenin is tightly regulated via phosphorylation by the 'destruction complex', consisting of glycogen synthase kinase 3β (GSK3β), casein kinase 1α (CK1α), the scaffold protein AXIN, and the tumor suppressor adenomatous polyposis coli (APC). Aberrant regulation of this signaling cascade is associated with varieties of human diseases, especially cancers. Over the past decade, significant progress has been made in understanding the mechanisms of canonical WNT signaling. In this review, we focus on the current understanding of WNT signaling at the extracellular, cytoplasmic membrane, and intracellular/nuclear levels, including the emerging knowledge of cross-talk with other pathways. Recent progresses in developing novel WNT pathway-targeted therapies will also be reviewed. Thus, this review is intended to serve as a refresher of the current understanding about the physiologic and pathogenic roles of WNT/β-catenin signaling pathway, and to outline potential therapeutic opportunities by targeting the canonical WNT pathway. PMID:26618721

  1. Targeting Wnt signaling at the neuroimmune interface for dopaminergic neuroprotection/repair in Parkinson’s disease

    PubMed Central

    L’Episcopo, Francesca; Tirolo, Cataldo; Caniglia, Salvo; Testa, Nuccio; Morale, Maria Concetta; Serapide, Maria Francesca; Pluchino, Stefano; Marchetti, Bianca

    2014-01-01

    During the past three decades, the Wingless-type MMTV integration site (Wnt) signaling cascade has emerged as an essential system regulating multiple processes in developing and adult brain. Accumulating evidence points to a dysregulation of Wnt signaling in major neurodegenerative pathologies including Parkinson’s disease (PD), a common neurodegenerative disorder characterized by the progressive loss of midbrain dopaminergic (mDA) neurons and deregulated activation of astrocytes and microglia. This review highlights the emerging link between Wnt signaling and key inflammatory pathways during mDA neuron damage/repair in PD progression. In particular, we summarize recent evidence documenting that aging and neurotoxicant exposure strongly antagonize Wnt/β-catenin signaling in mDA neurons and subventricular zone (SVZ) neuroprogenitors via astrocyte–microglial interactions. Dysregulation of the crosstalk between Wnt/β-catenin signaling and anti-oxidant/anti-inflammatory pathways delineate novel mechanisms driving the decline of SVZ plasticity with age and the limited nigrostriatal dopaminergic self-repair in PD. These findings hold a promise in developing therapies that target Wnt/β-catenin signaling to enhance endogenous restoration and neuronal outcome in age-dependent diseases, such as PD. PMID:24431301

  2. Structure and functional properties of Norrin mimic Wnt for signalling with Frizzled4, Lrp5/6, and proteoglycan

    PubMed Central

    Chang, Tao-Hsin; Hsieh, Fu-Lien; Zebisch, Matthias; Harlos, Karl; Elegheert, Jonathan; Jones, E Yvonne

    2015-01-01

    Wnt signalling regulates multiple processes including angiogenesis, inflammation, and tumorigenesis. Norrin (Norrie Disease Protein) is a cystine-knot like growth factor. Although unrelated to Wnt, Norrin activates the Wnt/β-catenin pathway. Signal complex formation involves Frizzled4 (Fz4), low-density lipoprotein receptor related protein 5/6 (Lrp5/6), Tetraspanin-12 and glycosaminoglycans (GAGs). Here, we report crystallographic and small-angle X-ray scattering analyses of Norrin in complex with Fz4 cysteine-rich domain (Fz4CRD), of this complex bound with GAG analogues, and of unliganded Norrin and Fz4CRD. Our structural, biophysical and cellular data, map Fz4 and putative Lrp5/6 binding sites to distinct patches on Norrin, and reveal a GAG binding site spanning Norrin and Fz4CRD. These results explain numerous disease-associated mutations. Comparison with the Xenopus Wnt8–mouse Fz8CRD complex reveals Norrin mimics Wnt for Frizzled recognition. The production and characterization of wild-type and mutant Norrins reported here open new avenues for the development of therapeutics to combat abnormal Norrin/Wnt signalling. DOI: http://dx.doi.org/10.7554/eLife.06554.001 PMID:26158506

  3. Isolation and characterization of recombinant murine Wnt3a

    PubMed Central

    Witkowski, Andrzej; Krishnamoorthy, Aparna; Su, Betty; Beckstead, Jennifer A.; Ryan, Robert O.

    2014-01-01

    Wnt proteins are a family of morphogens that possess potent biological activity. Structure – function studies have been impeded by poor yield of biologically active recombinant Wnt as well as a propensity of isolated Wnt to self-associate in the absence of detergent. Using stably transfected Drosophila S2 cells, studies have been conducted to improve recovery of recombinant murine Wnt3a, establish conditions for a detergent-free Wnt preparation and examine the effects of limited proteolysis. S2 cell culture conditioned media was subjected to a 3-step protocol including dye-ligand chromatography, immobilized metal affinity chromatography and gel filtration chromatography. Through selective pooling of column fractions, homogeneous and purified Wnt3a preparations were obtained. Limited proteolysis of Wnt3a with thrombin resulted in site-specific cleavage within the N-terminal saposin-like motif. To generate detergent-free protein, Wnt3a was immobilized on Cu2+-charged, iminodiacetic acid-derivatized Sepharose beads, detergent-free buffer was applied and Wnt3a eluted from the beads with buffer containing imidazole plus 30 mM methyl-β-cyclodextrin (MβCD). Wnt3a recovered in MβCD-containing buffer was soluble and biologically active. Insofar as MβCD is a member of a family of non-toxic, low molecular weight compounds capable of binding and solubilizing small hydrophobic ligands, Wnt-cyclodextrin complexes may facilitate structure-activity studies in the absence of adverse detergent effects. PMID:25448592

  4. Isolation and characterization of recombinant murine Wnt3a.

    PubMed

    Witkowski, Andrzej; Krishnamoorthy, Aparna; Su, Betty; Beckstead, Jennifer A; Ryan, Robert O

    2015-02-01

    Wnt proteins are a family of morphogens that possess potent biological activity. Structure-function studies have been impeded by poor yield of biologically active recombinant Wnt as well as a propensity of isolated Wnt to self-associate in the absence of detergent. Using stably transfected Drosophila S2 cells, studies have been conducted to improve recovery of recombinant murine Wnt3a, establish conditions for a detergent-free Wnt preparation and examine the effects of limited proteolysis. S2 cell culture conditioned media was subjected to a 3-step protocol including dye-ligand chromatography, immobilized metal affinity chromatography and gel filtration chromatography. Through selective pooling of column fractions, homogeneous and purified Wnt3a preparations were obtained. Limited proteolysis of Wnt3a with thrombin resulted in site-specific cleavage within the N-terminal saposin-like motif. To generate detergent-free protein, Wnt3a was immobilized on Cu(2+)-charged, iminodiacetic acid-derivatized Sepharose beads, detergent-free buffer was applied and Wnt3a eluted from the beads with buffer containing imidazole plus 30mM methyl-ß-cyclodextrin (MßCD). Wnt3a recovered in MßCD-containing buffer was soluble and biologically active. Insofar as MßCD is a member of a family of non-toxic, low molecular weight compounds capable of binding and solubilizing small hydrophobic ligands, Wnt-cyclodextrin complexes may facilitate structure-activity studies in the absence of adverse detergent effects. PMID:25448592

  5. USP6 oncogene promotes Wnt signaling by deubiquitylating Frizzleds.

    PubMed

    Madan, Babita; Walker, Matthew P; Young, Robert; Quick, Laura; Orgel, Kelly A; Ryan, Meagan; Gupta, Priti; Henrich, Ian C; Ferrer, Marc; Marine, Shane; Roberts, Brian S; Arthur, William T; Berndt, Jason D; Oliveira, Andre M; Moon, Randall T; Virshup, David M; Chou, Margaret M; Major, Michael B

    2016-05-24

    The Wnt signaling pathways play pivotal roles in carcinogenesis. Modulation of the cell-surface abundance of Wnt receptors is emerging as an important mechanism for regulating sensitivity to Wnt ligands. Endocytosis and degradation of the Wnt receptors Frizzled (Fzd) and lipoprotein-related protein 6 (LRP6) are regulated by the E3 ubiquitin ligases zinc and ring finger 3 (ZNRF3) and ring finger protein 43 (RNF43), which are disrupted in cancer. In a genome-wide small interfering RNA screen, we identified the deubiquitylase ubiquitin-specific protease 6 (USP6) as a potent activator of Wnt signaling. USP6 enhances Wnt signaling by deubiquitylating Fzds, thereby increasing their cell-surface abundance. Chromosomal translocations in nodular fasciitis result in USP6 overexpression, leading to transcriptional activation of the Wnt/β-catenin pathway. Inhibition of Wnt signaling using Dickkopf-1 (DKK1) or a Porcupine (PORCN) inhibitor significantly decreased the growth of USP6-driven xenograft tumors, indicating that Wnt signaling is a key target of USP6 during tumorigenesis. Our study defines an additional route to ectopic Wnt pathway activation in human disease, and identifies a potential approach to modulate Wnt signaling for therapeutic benefit. PMID:27162353

  6. TCF3, a novel positive regulator of osteogenesis, plays a crucial role in miR-17 modulating the diverse effect of canonical Wnt signaling in different microenvironments

    PubMed Central

    Liu, W; Liu, Y; Guo, T; Hu, C; Luo, H; Zhang, L; Shi, S; Cai, T; Ding, Y; Jin, Y

    2013-01-01

    Wnt signaling pathways are a highly conserved pathway, which plays an important role from the embryonic development to bone formation. The effect of Wnt pathway on osteogenesis relies on their cellular environment and the expression of target genes. However, the molecular mechanism of that remains unclear. On the basis of the preliminary results, we observed the contrary effect of canonical Wnt signaling on osteogenic differentiation of periodontal ligament stem cells (PDLSCs) in the different culture environment. Furthermore, we found that the expression level of miR-17 was also varied with the change in the culture environment. Therefore, we hypothesized that miR-17 and canonical Wnt signaling may have potential interactions, particularly the inner regulation relationship in different microenvironments. In this paper, we observed that canonical Wnt signaling promoted osteogenesis of PDLSCs in the fully culture medium, while inhibited it in the osteogenic differentiation medium. Interestingly, alteration in the expression level of endogenous miR-17 could partially reverse the different effect of canonical Wnt signaling. Furthermore, the role of miR-17 was because of its target gene TCF3 (transcription factor 3), a key transcription factor of canonical Wnt pathway. Overexpression of TCF3 attenuated the effect of miR-17 on modulating canonical Wnt signaling. Finally, we elucidated that TCF3 enhanced osteogenesis both in vitro and in vivo. In brief, the different level of miR-17 was the main cause of the different effect of canonical Wnt signaling, and TCF3 was the crucial node of miR-17–canonial Wnt signaling regulation loop. This understanding of microRNAs regulating signaling pathways in different microenvironments may pave the way for fine-tuning the process of osteogenesis in bone-related disorders. PMID:23492770

  7. Jarid2 Coordinates Nanog Expression and PCP/Wnt Signaling Required for Efficient ESC Differentiation and Early Embryo Development

    PubMed Central

    Landeira, David; Bagci, Hakan; Malinowski, Andrzej R.; Brown, Karen E.; Soza-Ried, Jorge; Feytout, Amelie; Webster, Zoe; Ndjetehe, Elodie; Cantone, Irene; Asenjo, Helena G.; Brockdorff, Neil; Carroll, Thomas; Merkenschlager, Matthias; Fisher, Amanda G.

    2015-01-01

    Summary Jarid2 is part of the Polycomb Repressor complex 2 (PRC2) responsible for genome-wide H3K27me3 deposition. Unlike other PRC2-deficient embryonic stem cells (ESCs), however, Jarid2-deficient ESCs show a severe differentiation block, altered colony morphology, and distinctive patterns of deregulated gene expression. Here, we show that Jarid2−/− ESCs express constitutively high levels of Nanog but reduced PCP signaling components Wnt9a, Prickle1, and Fzd2 and lowered β-catenin activity. Depletion of Wnt9a/Prickle1/Fzd2 from wild-type ESCs or overexpression of Nanog largely phenocopies these cellular defects. Co-culture of Jarid2−/− with wild-type ESCs restores variable Nanog expression and β-catenin activity and can partially rescue the differentiation block of mutant cells. In addition, we show that ESCs lacking Jarid2 or Wnt9a/Prickle1/Fzd2 or overexpressing Nanog induce multiple ICM formation when injected into normal E3.5 blastocysts. These data describe a previously unrecognized role for Jarid2 in regulating a core pluripotency and Wnt/PCP signaling circuit that is important for ESC differentiation and for pre-implantation development. PMID:26190104

  8. Jarid2 Coordinates Nanog Expression and PCP/Wnt Signaling Required for Efficient ESC Differentiation and Early Embryo Development.

    PubMed

    Landeira, David; Bagci, Hakan; Malinowski, Andrzej R; Brown, Karen E; Soza-Ried, Jorge; Feytout, Amelie; Webster, Zoe; Ndjetehe, Elodie; Cantone, Irene; Asenjo, Helena G; Brockdorff, Neil; Carroll, Thomas; Merkenschlager, Matthias; Fisher, Amanda G

    2015-07-28

    Jarid2 is part of the Polycomb Repressor complex 2 (PRC2) responsible for genome-wide H3K27me3 deposition. Unlike other PRC2-deficient embryonic stem cells (ESCs), however, Jarid2-deficient ESCs show a severe differentiation block, altered colony morphology, and distinctive patterns of deregulated gene expression. Here, we show that Jarid2(-/-) ESCs express constitutively high levels of Nanog but reduced PCP signaling components Wnt9a, Prickle1, and Fzd2 and lowered β-catenin activity. Depletion of Wnt9a/Prickle1/Fzd2 from wild-type ESCs or overexpression of Nanog largely phenocopies these cellular defects. Co-culture of Jarid2(-/-) with wild-type ESCs restores variable Nanog expression and β-catenin activity and can partially rescue the differentiation block of mutant cells. In addition, we show that ESCs lacking Jarid2 or Wnt9a/Prickle1/Fzd2 or overexpressing Nanog induce multiple ICM formation when injected into normal E3.5 blastocysts. These data describe a previously unrecognized role for Jarid2 in regulating a core pluripotency and Wnt/PCP signaling circuit that is important for ESC differentiation and for pre-implantation development. PMID:26190104

  9. Molecular modeling of directed self-assembly of block copolymers: Fundamental studies of processing conditions and evolutionary pattern design

    NASA Astrophysics Data System (ADS)

    Khaira, Gurdaman Singh

    Rapid progress in the semi-conductor industry has pushed for smaller feature sizes on integrated electronic circuits. Current photo-lithographic techniques for nanofabrication have reached their technical limit and are problematic when printing features small enough to meet future industrial requirements. "Bottom-up'' techniques, such as the directed self-assembly (DSA) of block copolymers (BCP), are the primary contenders to compliment current "top-down'' photo-lithography ones. For industrial requirements, the defect density from DSA needs to be less than 1 defect per 10 cm by 10 cm. Knowledge of both material synthesis and the thermodynamics of the self-assembly process are required before optimal operating conditions can be found to produce results adequate for industry. The work present in this thesis is divided into three chapters, each discussing various aspects of DSA as studied via a molecular model that contains the essential physics of BCP self-assembly. Though there are various types of guiding fields that can be used to direct BCPs over large wafer areas with minimum defects, this study focuses only on chemically patterned substrates. The first chapter addresses optimal pattern design by describing a framework where molecular simulations of various complexities are coupled with an advanced optimization technique to find a pattern that directs a target morphology. It demonstrates the first ever study where BCP self-assembly on a patterned substrate is optimized using a three-dimensional description of the block-copolymers. For problems pertaining to DSA, the methodology is shown to converge much faster than the traditional random search approach. The second chapter discusses the metrology of BCP thin films using TEM tomography and X-ray scattering techniques, such as CDSAXS and GISAXS. X-ray scattering has the advantage of being able to quickly probe the average structure of BCP morphologies over large wafer areas; however, deducing the BCP morphology

  10. IL-4 Causes Hyperpermeability of Vascular Endothelial Cells through Wnt5A Signaling

    PubMed Central

    Skaria, Tom; Burgener, Julia; Bachli, Esther; Schoedon, Gabriele

    2016-01-01

    Microvascular leakage due to endothelial barrier dysfunction is a prominent feature of T helper 2 (Th2) cytokine mediated allergic inflammation. Interleukin-4 (IL-4) is a potent Th2 cytokine, known to impair the barrier function of endothelial cells. However, the effectors mediating IL-4 induced cytoskeleton remodeling and consequent endothelial barrier dysfunction remain poorly defined. Here we have used whole genome transcriptome profiling and gene ontology analyses to identify the genes and processes regulated by IL-4 signaling in human coronary artery endothelial cells (HCAEC). The study revealed Wnt5A as an effector that can mediate actin cytoskeleton remodeling in IL-4 activated HCAEC through the regulation of LIM kinase (LIMK) and Cofilin (CFL). Following IL-4 treatment, LIMK and CFL were phosphorylated, thereby indicating the possibility of actin stress fiber formation. Imaging of actin showed the formation of stress fibers in IL-4 treated live HCAEC. Stress fiber formation was notably decreased in the presence of Wnt inhibitory factor 1 (WIF1). Non-invasive impedance measurements demonstrated that IL-4 increased the permeability and impaired the barrier function of HCAEC monolayers. Silencing Wnt5A significantly reduced permeability and improved the barrier function of HCAEC monolayers upon IL-4 treatment. Our study identifies Wnt5A as a novel marker of IL-4 activated vascular endothelium and demonstrates a critical role for Wnt5A in mediating IL-4 induced endothelial barrier dysfunction. Wnt5A could be a potential therapeutic target for reducing microvascular leakage and edema formation in Th2 driven inflammatory diseases. PMID:27214384

  11. Haplotype association analysis of genes within the WNT signalling pathways in diabetic nephropathy

    PubMed Central

    2013-01-01

    Background Renal interstitial fibrosis and glomerular sclerosis are hallmarks of diabetic nephropathy (DN) and several studies have implicated members of the WNT pathways in these pathological processes. This study comprehensively examined common genetic variation within the WNT pathway for association with DN. Methods Genes within the WNT pathways were selected on the basis of nominal significance and consistent direction of effect in the GENIE meta-analysis dataset. Common SNPs and common haplotypes were examined within the selected WNT pathway genes in a white population with type 1 diabetes, discordant for DN (cases: n = 718; controls: n = 749). SNPs were genotyped using Sequenom or Taqman assays. Association analyses were performed using PLINK, to compare allele and haplotype frequencies in cases and controls. Correction for multiple testing was performed by either permutation testing or using false discovery rate. Results A logistic regression model including collection centre, duration of diabetes, and average HbA1c as covariates highlighted three SNPs in GSK3B (rs17810235, rs17471, rs334543), two in DAAM1 (rs1253192, rs1252906) and one in NFAT5 (rs17297207) as being significantly (P < 0.05) associated with DN, however these SNPs did not remain significant after correction for multiple testing. Logistic regression of haplotypes, with ESRD as the outcome, and pairwise interaction analyses did not yield any significant results after correction for multiple testing. Conclusions These results indicate that both common SNPs and common haplotypes of WNT pathway genes are not strongly associated with DN. However, this does not completely exclude these or the WNT pathways from association with DN, as unidentified rare genetic or copy number variants could still contribute towards the genetic architecture of DN. PMID:23777469

  12. Activation of the wnt/β-Catenin Signaling Pathway in Polymyositis, Dermatomyositis and Duchenne Muscular Dystrophy

    PubMed Central

    Liu, Fuchen; Liang, Zonglai; Xu, Jingwen; Li, Wei; Zhao, Dandan; Zhao, Yuying

    2016-01-01

    Background and Purpose The wnt/β-catenin signaling pathway plays a critical role in embryonic development and adult-tissue homeostasis. Recent investigations implicate the importance of wnt/β-catenin signaling in normal wound healing and its sustained activation being associated with fibrogenesis. We investigated the immunolocalization and activation of wnt/β-catenin in polymyositis (PM), dermatomyositis (DM), and Duchenne muscular dystrophy (DMD). Methods Immunofluorescence staining and Western blot analysis of β-catenin were performed in muscle specimens from 6 PM, 8 DM, and 6 DMD subjects. The β-catenin/Tcf4 DNA-binding activity in muscle was studied using an electrophoretic mobility shift assay (EMSA), and serum wnt/β-catenin/Tcf transcriptional activity was measured using a luciferase reporter gene assay. Results Immunoreactivity for β-catenin was found in the cytoplasm and nuclei of muscle fibers in PM, DM, and DMD. The protein level of β-catenin was elevated, and EMSA analysis confirmed the activation of wnt/β-catenin signaling. The transcriptional activities of β-catenin/Tcf in the circulation were increased in patients with PM, DM, and DMD, especially in those with interstitial lung disease, and these transcriptional activities decreased when PM or DM patients exhibited obvious clinical improvements. Conclusions Our findings indicate that wnt/β-catenin signaling is activated in PM, DM, and DMD. Its activation in muscle tissue and the circulation may play a role in modulating muscle regeneration and be at least partly involved in the process of muscle and pulmonary fibrosis. PMID:27165423

  13. SFRP2 augments WNT16B signaling to promote therapeutic resistance in the damaged tumor microenvironment.

    PubMed

    Sun, Y; Zhu, D; Chen, F; Qian, M; Wei, H; Chen, W; Xu, J

    2016-08-18

    Most tumors initially respond to cytotoxic treatments, but acquired resistance often follows. The tumor microenvironment (TME) is a major barrier to clinical success by compromising therapeutic efficacy, and pathological relevance of multiple soluble factors released by a therapeutically remodeled TME remains largely unexplored. Here we show that the secreted frizzled-related protein 2 (SFRP2), a Wnt pathway modulator, is produced by human primary fibroblasts after genotoxic treatments. SFRP2 induction is remarkable in tumor stroma, with transcription mainly modulated by the nuclear factor-κB (NF-κB) complex, a property shared by several effectors of the DNA damage secretory program. Instead of directly altering canonical Wnt signaling, SFRP2 augments β-catenin activities initiated by WNT16B, another soluble factor from DNA-damaged stroma. WNT16B recognizes cancer cell surface receptors including frizzled (FZD) 3/4/6, a process enhanced by SFRP2, coordinated by the co-receptor LRP6 but subject to abrogation by DKK1. Importantly, we found WNT16B plays a central role in promoting advanced malignancies particularly acquired resistance by counteracting cell death, an effect that can be minimized by a neutralizing antibody co-administered with classical chemotherapy. Furthermore, DNA damage-triggered expression of WNT16B is systemic, imaged by significant induction among diverse solid organs and circulation in peripheral blood, thereby holding promise as not only a TME-derived anticancer target but also a novel biomarker for clinical evaluation of treatment efficacy. Overall, our study substantiates the biological complexity and pathological implication of a therapy-activated TME, and provides the proof of principle of co-targeting tumor and the TME to prevent acquired resistance, with the aim of improving intervention outcome in an era of precision medicine. PMID:26751775

  14. SFRP2 augments WNT16B signaling to promote therapeutic resistance in the damaged tumor microenvironment

    PubMed Central

    Sun, Y; Zhu, D; Chen, F; Qian, M; Wei, H; Chen, W; Xu, J

    2016-01-01

    Most tumors initially respond to cytotoxic treatments, but acquired resistance often follows. The tumor microenvironment (TME) is a major barrier to clinical success by compromising therapeutic efficacy, and pathological relevance of multiple soluble factors released by a therapeutically remodeled TME remains largely unexplored. Here we show that the secreted frizzled-related protein 2 (SFRP2), a Wnt pathway modulator, is produced by human primary fibroblasts after genotoxic treatments. SFRP2 induction is remarkable in tumor stroma, with transcription mainly modulated by the nuclear factor-κB (NF-κB) complex, a property shared by several effectors of the DNA damage secretory program. Instead of directly altering canonical Wnt signaling, SFRP2 augments β-catenin activities initiated by WNT16B, another soluble factor from DNA-damaged stroma. WNT16B recognizes cancer cell surface receptors including frizzled (FZD) 3/4/6, a process enhanced by SFRP2, coordinated by the co-receptor LRP6 but subject to abrogation by DKK1. Importantly, we found WNT16B plays a central role in promoting advanced malignancies particularly acquired resistance by counteracting cell death, an effect that can be minimized by a neutralizing antibody co-administered with classical chemotherapy. Furthermore, DNA damage-triggered expression of WNT16B is systemic, imaged by significant induction among diverse solid organs and circulation in peripheral blood, thereby holding promise as not only a TME-derived anticancer target but also a novel biomarker for clinical evaluation of treatment efficacy. Overall, our study substantiates the biological complexity and pathological implication of a therapy-activated TME, and provides the proof of principle of co-targeting tumor and the TME to prevent acquired resistance, with the aim of improving intervention outcome in an era of precision medicine. PMID:26751775

  15. Capsaicin inhibits the Wnt/β-catenin signaling pathway by down-regulating PP2A.

    PubMed

    Park, Dong-Seok; Yoon, Gang-Ho; Lee, Hyun-Shik; Choi, Sun-Cheol

    2016-09-01

    Xenopus embryo serves as an ideal model for teratogenesis assays to examine the effects of any substances on the cellular processes critical for early development and adult tissue homeostasis. In our chemical library screening with frog embryo, capsaicin was found to repress the Wnt/β-catenin signaling. Depending on the stages at which embryos became exposed to capsaicin, it could disrupt formation of dorsal or posterior body axis of embryo, which is associated with inhibition of maternal or zygotic Wnt signal in early development. In agreement with these phenotypes, capsaicin suppressed the expression of Wnt target genes such as Siamois and Chordin in the organizer region of embryo and in Wnt signals-stimulated tissue explants. In addition, the cellular level of β-catenin, a key component of Wnt pathway, was down-regulated in capsaicin-treated embryonic cells. Unlike wild-type β-catenin, its non-phosphorylatable mutant in which serine and threonine residues phosphorylated by GSK3 are substituted with alanine was not destabilized by capsaicin, indicative of the effect of this chemical on the phosphorylation status of β-catenin. In support of this, capsaicin up-regulated the level of GSK3- or CK1-phosphorylated β-catenin, concomitantly lowering that of its de-phosphorylated version. Notably, capsaicin augmented the phosphorylation of a phosphatase, PP2A at tyrosine 307, suggesting its repression of the enzymatic activity of the phosphatase. Furthermore, capsaicin still enhanced β-catenin phosphorylation in cells treated with a GSK3 inhibitor, LiCl but not in those treated with a phosphatase inhibitor, okadaic acid. Together, these results indicate that capsaicin inhibits the patterning of the dorso-ventral and anterior-posterior body axes of embryo by repressing PP2A and thereby down-regulating the Wnt/β-catenin signaling. PMID:27318088

  16. Characterization of a Wnt-binding site of the WIF-domain of Wnt inhibitory factor-1.

    PubMed

    Bányai, László; Kerekes, Krisztina; Patthy, László

    2012-09-21

    A Wnt-binding site of the WIF-domain of Wnt inhibitory factor-1 was localized by structure-guided arginine-scanning mutagenesis in combination with surface plasmon resonance assays. Our observation that substitution of some residues of WIF resulted in an increased affinity for Wnt5a, but decreased affinity for Wnt3a, suggests that these residues may define the specificity spectrum of WIF for Wnts. These results hold promise for a more specific targeting of Wnt family members with WIF variants in various forms of cancer. PMID:22986341

  17. Modular mechanism of Wnt signaling inhibition by Wnt inhibitory factor 1

    PubMed Central

    Malinauskas, Tomas; Aricescu, A. Radu; Lu, Weixian; Siebold, Christian; Jones, E. Yvonne

    2012-01-01

    Wnt morphogens control embryonic development and homeostasis in adult tissues. In vertebrates the N-terminal WIF domain (WIF-1WD) of Wnt inhibitory factor 1 (WIF-1) binds Wnt ligands. Our crystal structure of WIF-1WD reveals a previously unidentified binding site for phospholipid; two acyl chains extend deep into the domain, and the head group is exposed to the surface. Biophysical and cellular assays indicate that there is a WIF-1WD Wnt-binding surface proximal to the lipid head group but also implicate the five epidermal growth factor (EGF)-like domains (EGFs I–V) in Wnt binding. The six-domain WIF-1 crystal structure shows that EGFs I–V are wrapped back, interfacing with WIF-1WD at EGF III. EGFs II–V contain a heparan sulfate proteoglycan (HSPG)-binding site, consistent with conserved positively charged residues on EGF IV. This combination of HSPG- and Wnt-binding properties suggests a modular model for the localization of WIF-1 and for signal inhibition within morphogen gradients. PMID:21743455

  18. Population Blocks.

    ERIC Educational Resources Information Center

    Smith, Martin H.

    1992-01-01

    Describes an educational game called "Population Blocks" that is designed to illustrate the concept of exponential growth of the human population and some potential effects of overpopulation. The game material consists of wooden blocks; 18 blocks are painted green (representing land), 7 are painted blue (representing water); and the remaining…

  19. Releasing Activity Disengages Cohesin’s Smc3/Scc1 Interface in a Process Blocked by Acetylation

    PubMed Central

    Beckouët, Frederic; Srinivasan, Madhusudhan; Roig, Maurici Brunet; Chan, Kok-Lung; Scheinost, Johanna C.; Batty, Paul; Hu, Bin; Petela, Naomi; Gligoris, Thomas; Smith, Alexandra C.; Strmecki, Lana; Rowland, Benjamin D.; Nasmyth, Kim

    2016-01-01

    Summary Sister chromatid cohesion conferred by entrapment of sister DNAs within a tripartite ring formed between cohesin’s Scc1, Smc1, and Smc3 subunits is created during S and destroyed at anaphase through Scc1 cleavage by separase. Cohesin’s association with chromosomes is controlled by opposing activities: loading by Scc2/4 complex and release by a separase-independent releasing activity as well as by cleavage. Coentrapment of sister DNAs at replication is accompanied by acetylation of Smc3 by Eco1, which blocks releasing activity and ensures that sisters remain connected. Because fusion of Smc3 to Scc1 prevents release and bypasses the requirement for Eco1, we suggested that release is mediated by disengagement of the Smc3/Scc1 interface. We show that mutations capable of bypassing Eco1 in Smc1, Smc3, Scc1, Wapl, Pds5, and Scc3 subunits reduce dissociation of N-terminal cleavage fragments of Scc1 (NScc1) from Smc3. This process involves interaction between Smc ATPase heads and is inhibited by Smc3 acetylation. PMID:26895425

  20. Effects of thermal fluctuations and block copolymers compositions on defects in directed self-assembly hole shrink process

    NASA Astrophysics Data System (ADS)

    Fukawatase, Ken; Yoshimoto, Kenji; Ohshima, Masahiro

    2015-06-01

    We investigated the two critical defects on the directed self-assembly hole shrink process, i.e., polystyrene (PS) residue and placement error, by performing dynamic simulations with the Ohta-Kawasaki model. In the simulations, the thermal noise was added to generate stochastic variations in shape and location of the poly(methyl methacrylate) (PMMA) cylindrical domains. For the PS residue issue, we found that the volume fraction of the PMMA minor block, fPMMA, was an effective parameter, and that the PS residue could be minimized by increasing fPMMA from a conventional value of 0.30 to 0.40. On the other hand, the placement error of the PMMA cylindrical domain was affected little by the change in shape and size of the guide hole and by the connectivity to the guide bottom wall. It is speculated that the interfacial stiffness between the PMMA and PS domains would be essential to control the placement error of the PMMA cylindrical domains.

  1. Lattice Boltzmann study of hydrodynamic effects in lamellar ordering process of two-dimensional quenched block copolymers

    NASA Astrophysics Data System (ADS)

    Song, Kai-Xu; Jia, Yu-Xi; Sun, Zhao-Yan; An, Li-Jia

    2008-10-01

    By incorporating self-consistent field theory with lattice Boltzmann method, a model for polymer melts is proposed. Compared with models based on Ginzburg-Landau free energy, our model does not employ phenomenological free energies to describe systems and can consider the chain topological details of polymers. We use this model to study the effects of hydrodynamic interactions on the dynamics of microphase separation for block copolymers. In the early stage of phase separation, an exponential growth predicted by Cahn-Hilliard treatment is found. Simulation results also show that the effect of hydrodynamic interactions can be neglected in the early stage. For the late stage of phase separation, it is easy to see the effects of hydrodynamic interactions on the ordering process of lamellae phase. From the analysis of structure factor curves, we find that the growth of domains is faster if hydrodynamic interactions are introduced. Furthermore, the scaling of the pattern dynamics is investigated for the late stage at zero thermal noise. By studying the behavior of scaling exponents of the structure factor and the nematic order-parameter correlation function Cnn, we can see that the effects of hydrodynamic interactions lead to bigger growth exponent for both functions.

  2. Valves - current operating experience of slurry valves (block and letdown) in coal liquefaction processes. Third quarter report

    SciTech Connect

    1996-07-01

    This paper summarizes the recent letdown and block valve experience in the liquefaction pilot plants. Also included is a brief description of the research and development activities on valves which are conducted in supporting laboratories. The purpose of the summary is to concentrate on critical component problems common to all liquefaction plants, to avoid duplication of efforts, and to help provide timely solutions to the valve problems. The main source of information used in this paper is the Minutes of the Critical Component and Materials Meeting which is sponsored by the Office of Coal Processing, Fossil Energy, Department of Energy. Other sources of information such as the technical progress reports are also included based on availability and relevance to topics covered in this paper. It is intended that this report will be followed by updates as pertinent information concerning valves becomes available. In the subsequent sections of this paper a brief outline of past valve studies is given as background material followed by a summary of the most recent valve operating experience at the liquefaction plants.

  3. Role of Wnt Signaling in Central Nervous System Injury.

    PubMed

    Lambert, Catherine; Cisternas, Pedro; Inestrosa, Nibaldo C

    2016-05-01

    The central nervous system (CNS) is highly sensitive to external mechanical damage, presenting a limited capacity for regeneration explained in part by its inability to restore either damaged neurons or the synaptic network. The CNS may suffer different types of external injuries affecting its function and/or structure, including stroke, spinal cord injury, and traumatic brain injury. These pathologies critically affect the quality of life of a large number of patients worldwide and are often fatal because available therapeutics are ineffective and produce limited results. Common effects of the mentioned pathologies involves the triggering of several cellular and metabolic responses against injury, including infiltration of blood cells, inflammation, glial activation, and neuronal death. Although some of the underlying molecular mechanisms of those responses have been elucidated, the mechanisms driving these processes are poorly understood in the context of CNS injury. In the last few years, it has been suggested that the activation of the Wnt signaling pathway could be important in the regenerative response after CNS injury, activating diverse protective mechanisms including the stimulation of neurogenesis, blood brain structure consolidation and the recovery of cognitive brain functions. Because Wnt signaling is involved in several physiological processes, the putative positive role of its activation after injury could be the basis for novel therapeutic approaches to CNS injury. PMID:25976365

  4. Wnt3 and Wnt3a are required for induction of the mid-diencephalic organizer in the caudal forebrain

    PubMed Central

    2012-01-01

    Background A fundamental requirement for development of diverse brain regions is the function of local organizers at morphological boundaries. These organizers are restricted groups of cells that secrete signaling molecules, which in turn regulate the fate of the adjacent neural tissue. The thalamus is located in the caudal diencephalon and is the central relay station between the sense organs and higher brain areas. The mid-diencephalic organizer (MDO) orchestrates the development of the thalamus by releasing secreted signaling molecules such as Shh. Results Here we show that canonical Wnt signaling in the caudal forebrain is required for the formation of the Shh-secreting MD organizer in zebrafish. Wnt signaling induces the MDO in a narrow time window of 4 hours - between 10 and 14 hours post fertilization. Loss of Wnt3 and Wnt3a prevents induction of the MDO, a phenotype also observed upon blockage of canonical Wnt signaling per se. Pharmaceutical activation of the canonical Wnt pathways in Wnt3/Wnt3a compound morphant embryos is able to restore the lack of the MDO. After blockage of Wnt signaling or knock-down of Wnt3/Wnt3a we find an increase of apoptotic cells specifically within the organizer primordium. Consistently, blockage of apoptosis restores the thalamus organizer MDO in Wnt deficient embryos. Conclusion We have identified canonical Wnt signaling as a novel pathway, that is required for proper formation of the MDO and consequently for the development of the major relay station of the brain - the thalamus. We propose that Wnt ligands are necessary to maintain the primordial tissue of the organizer during somitogenesis by suppressing Tp53-mediated apoptosis. PMID:22475147

  5. Wnt signaling in heart valve development and osteogenic gene induction

    PubMed Central

    Alfieri, Christina M.; Cheek, Jonathan; Chakraborty, Santanu; Yutzey, Katherine E.

    2009-01-01

    Wnt signaling mediated by beta-catenin has been implicated in early endocardial cushion development, but its roles in later stages of heart valve maturation and homeostasis have not been identified. Multiple Wnt ligands and pathway genes are differentially expressed during heart valve development. At E12.5, Wnt2 is expressed in cushion mesenchyme, whereas Wnt4 and Wnt9b are predominant in overlying endothelial cells. At E17.5, both Wnt3a and Wnt7b are expressed in the remodeling atrioventricular (AV) and semilunar valves. In addition, the TOPGAL Wnt reporter transgene is active throughout the developing AV and semilunar valves at E16.5, with more localized expression in the stratified valve leaflets after birth. In chicken embryo aortic valves, genes characteristic of osteogenic cell lineages including periostin, osteonectin, and Id2 are expressed specifically in the collagen-rich fibrosa layer at E14. Treatment of E14 aortic valve interstitial cells (VIC) in culture with osteogenic media results in increased expression of multiple genes associated with bone formation. Treatment of VIC with Wnt3a leads to nuclear localization of beta-catenin and induction of periostin and matrix gla-protein, but does not induce genes associated with later stages of osteogenesis. Together, these studies provide evidence for Wnt signaling as a regulator of endocardial cushion maturation as well as valve leaflet stratification, homeostasis and pathogenesis. PMID:19961844

  6. Wnt Signaling Interacts with Bmp and Edn1 to Regulate Dorsal-Ventral Patterning and Growth of the Craniofacial Skeleton

    PubMed Central

    Alexander, Courtney; Piloto, Sarah; Le Pabic, Pierre; Schilling, Thomas F.

    2014-01-01

    Craniofacial development requires signals from epithelia to pattern skeletogenic neural crest (NC) cells, such as the subdivision of each pharyngeal arch into distinct dorsal (D) and ventral (V) elements. Wnt signaling has been implicated in many aspects of NC and craniofacial development, but its roles in D-V arch patterning remain unclear. To address this we blocked Wnt signaling in zebrafish embryos in a temporally-controlled manner, using transgenics to overexpress a dominant negative Tcf3, (dntcf3), (Tg(hsp70I:tcf3-GFP), or the canonical Wnt inhibitor dickkopf1 (dkk1), (Tg(hsp70i:dkk1-GFP) after NC migration. In dntcf3 transgenics, NC cells in the ventral arches of heat-shocked embryos show reduced proliferation, expression of ventral patterning genes (hand2, dlx3b, dlx5a, msxe), and ventral cartilage differentiation (e.g. lower jaws). These D-V patterning defects resemble the phenotypes of zebrafish embryos lacking Bmp or Edn1 signaling, and overexpression of dntcf3 dramatically reduces expression of a subset of Bmp receptors in the arches. Addition of ectopic BMP (or EDN1) protein partially rescues ventral development and expression of dlx3b, dlx5a, and msxe in Wnt signaling-deficient embryos, but surprisingly does not rescue hand2 expression. Thus Wnt signaling provides ventralizing patterning cues to arch NC cells, in part through regulation of Bmp and Edn1 signaling, but independently regulates hand2. Similarly, heat-shocked dkk1+ embryos exhibit ventral arch reductions, but also have mandibular clefts at the ventral midline not seen in dntcf3+ embryos. Dkk1 is expressed in pharyngeal endoderm, and cell transplantation experiments reveal that dntcf3 must be overexpressed in pharyngeal endoderm to disrupt D-V arch patterning, suggesting that distinct endodermal roles for Wnts and Wnt antagonists pattern the developing skeleton. PMID:25058015

  7. Wnt-3a is critical for caudal embryonic development

    SciTech Connect

    Camper, S.A.; Greco, T.L.; Newhouse, M.M.

    1994-09-01

    Skeletal and neural tube defects represent an important class of birth defects. The majority of mouse mutants with neural tube defects also have malformations of the tail. Vestigial tail (vt) is an autosomal recessive mouse mutation characterized by reduction or absence of the tail, vertebral abnormalities, and reduced fertility. The phenotype has been described as the result of failure of cell migration through the primitive streak, causing abnormalities in the development of the neural tube and a reduction in the ventral ectodermal ridge. Wnt3a is an excellent candidate gene for vt because Wnt3a is expressed in the primitive streak and in the embryonic mesoderm, and it is thought to be involved in cell-to-cell communication and formation of the dorsal-ventral axis in the CNS. A lack of Wnt3a might be expected to result in overdorsalization of the neural tube and reduction of the ventral ectodermal ridge characteristic of vt/vt embryos. In a high resolution backcross segregating vt, we observed no recombination between vt and Wnt3a in 363 individuals analyzed. In vt/vt mice, Southern blot analysis revealed no abnormalities in the Wnt3a gene, and the Wnt3a cDNA sequence does not encode any amino acid changes. Whole mount in situ hybridization analysis demonstrated that Wnt3a expression is severely reduced in the developing tailbud of day 9.5 vt/vt embryos, suggestive of a lesion in the regulation on Wnt3a expression. An alleleism test, carried out by mating vt/vt males with Wnt3a +/Wnt3a- females, demonstrated that vt and Wnt3a are noncomplementing alleles. All of the compound heterozygotes exhibited severe tail defects, including occasional examples of hind limb parlaysis and spina bifida. The vertebral defects are intermediate between those of vt and Wnt3a homozygotes, suggesting that the concentration of Wnt3a correlates with the severity of the defect.

  8. Slb/Wnt11 controls hypoblast cell migration and morphogenesis at the onset of zebrafish gastrulation

    PubMed Central

    Ulrich, Florian; Concha, Miguel L.; Heid, Paul J.; Voss, Ed; Witzel, Sabine; Roehl, Henry; Tada, Masazumi; Wilson, Stephen W.; Adams, Richard J.; Soll, David R.; Heisenberg, Carl-Philipp

    2005-01-01

    Summary During vertebrate gastrulation, highly coordinated cellular rearrangements lead to the formation of the three germ layers, ectoderm, mesoderm and endoderm. In zebrafish, silberblick (slb)/wnt11 regulates normal gastrulation movements by activating a signalling pathway similar to the Frizzled-signalling pathway, which establishes epithelial planar cell polarity (PCP) in Drosophila. However, the cellular mechanisms by which slb/wnt11 functions during zebrafish gastrulation are still unclear. Using high-resolution two-photon confocal imaging followed by computer-assisted reconstruction and motion analysis, we have analysed the movement and morphology of individual cells in three dimensions during the course of gastrulation. We show that in slb-mutant embryos, hypoblast cells within the forming germ ring have slower, less directed migratory movements at the onset of gastrulation. These aberrant cell movements are accompanied by defects in the orientation of cellular processes along the individual movement directions of these cells. We conclude that slb/wnt11-mediated orientation of cellular processes plays a role in facilitating and stabilising movements of hypoblast cells in the germ ring, thereby pointing at a novel function of the slb/wnt11 signalling pathway for the regulation of migratory cell movements at early stages of gastrulation. PMID:13129848

  9. Use of an Activated Beta-Catenin to Identify Wnt Pathway Target Genes in Caenorhabditis elegans, Including a Subset of Collagen Genes Expressed in Late Larval Development

    PubMed Central

    Jackson, Belinda M.; Abete-Luzi, Patricia; Krause, Michael W.; Eisenmann, David M.

    2014-01-01

    The Wnt signaling pathway plays a fundamental role during metazoan development, where it regulates diverse processes, including cell fate specification, cell migration, and stem cell renewal. Activation of the beta-catenin−dependent/canonical Wnt pathway up-regulates expression of Wnt target genes to mediate a cellular response. In the nematode Caenorhabditis elegans, a canonical Wnt signaling pathway regulates several processes during larval development; however, few target genes of this pathway have been identified. To address this deficit, we used a novel approach of conditionally activated Wnt signaling during a defined stage of larval life by overexpressing an activated beta-catenin protein, then used microarray analysis to identify genes showing altered expression compared with control animals. We identified 166 differentially expressed genes, of which 104 were up-regulated. A subset of the up-regulated genes was shown to have altered expression in mutants with decreased or increased Wnt signaling; we consider these genes to be bona fide C. elegans Wnt pathway targets. Among these was a group of six genes, including the cuticular collagen genes, bli-1col-38, col-49, and col-71. These genes show a peak of expression in the mid L4 stage during normal development, suggesting a role in adult cuticle formation. Consistent with this finding, reduction of function for several of the genes causes phenotypes suggestive of defects in cuticle function or integrity. Therefore, this work has identified a large number of putative Wnt pathway target genes during larval life, including a small subset of Wnt-regulated collagen genes that may function in synthesis of the adult cuticle. PMID:24569038

  10. A novel Wnt5a-Frizzled4 signaling pathway mediates activity-independent dendrite morphogenesis via the distal PDZ motif of Frizzled 4.

    PubMed

    Bian, Wen-Jie; Miao, Wan-Ying; He, Shun-Ji; Wan, Zong-Fang; Luo, Zhen-Ge; Yu, Xiang

    2015-08-01

    The morphology of the dendritic tree is critical to neuronal function and neural circuit wiring. Several Wnt family members have been demonstrated to play important roles in dendrite development. However, the Wnt receptors responsible for mediating this process remain largely elusive. Using primary hippocampal neuronal cultures as a model system, we report that Frizzled4 (Fzd4), a member of the Fzd family of Wnt receptors, specifically signals downstream of Wnt5a to promote dendrite branching and growth. Interestingly, the less conserved distal PDZ binding motif of Fzd4, and not its conserved proximal Dvl-interacting PDZ motif, is required for mediating this effect. We further showed that Dvl signaled parallel to and independent of Fzd4 in promoting dendrite growth. Unlike most previously described pathways, Wnt5a/Fzd4 signaling promoted dendrite development in an activity-independent and autocrine fashion. Together, these results provide the first identification of a Wnt receptor for regulating dendrite development in the mammalian system, and demonstrate a novel function of the distal PDZ motif of Fzd4 in dendrite morphogenesis, thereby expanding our knowledge of the complex roles of Wnt signaling in neural development. PMID:25424568

  11. Tip cell-specific requirement for an atypical Gpr124- and Reck-dependent Wnt/β-catenin pathway during brain angiogenesis

    PubMed Central

    Vanhollebeke, Benoit; Stone, Oliver A; Bostaille, Naguissa; Cho, Chris; Zhou, Yulian; Maquet, Emilie; Gauquier, Anne; Cabochette, Pauline; Fukuhara, Shigetomo; Mochizuki, Naoki; Nathans, Jeremy; Stainier, Didier YR

    2015-01-01

    Despite the critical role of endothelial Wnt/β-catenin signaling during central nervous system (CNS) vascularization, how endothelial cells sense and respond to specific Wnt ligands and what aspects of the multistep process of intra-cerebral blood vessel morphogenesis are controlled by these angiogenic signals remain poorly understood. We addressed these questions at single-cell resolution in zebrafish embryos. We identify the GPI-anchored MMP inhibitor Reck and the adhesion GPCR Gpr124 as integral components of a Wnt7a/Wnt7b-specific signaling complex required for brain angiogenesis and dorsal root ganglia neurogenesis. We further show that this atypical Wnt/β-catenin signaling pathway selectively controls endothelial tip cell function and hence, that mosaic restoration of single wild-type tip cells in Wnt/β-catenin-deficient perineural vessels is sufficient to initiate the formation of CNS vessels. Our results identify molecular determinants of ligand specificity of Wnt/β-catenin signaling and provide evidence for organ-specific control of vascular invasion through tight modulation of tip cell function. DOI: http://dx.doi.org/10.7554/eLife.06489.001 PMID:26051822

  12. Wnt Signaling in Skin Development, Homeostasis, and Disease

    PubMed Central

    Lim, Xinhong; Nusse, Roel

    2013-01-01

    The skin and its appendages constitute the largest organ of the body. Its stratified epithelia offer protection from environmental stresses such as dehydration, irradiation, mechanical trauma, and pathogenic infection, whereas its appendages, like hair and sebaceous glands, help regulate body temperature as well as influence animal interaction and social behavior through camouflage and sexual signaling. To respond to and function effectively in a dynamic external environment, the skin and its appendages possess a remarkable ability to regenerate in a carefully controlled fashion. When this finely tuned homeostatic process is disrupted, skin diseases such as cancers may result. At present, the molecular signals that orchestrate cell proliferation, differentiation, and patterning in the skin remain incompletely understood. It is increasingly apparent that many morphogenetic pathways with key roles in development are also important in regulating skin biology. Of these, Wnt signaling has emerged as the dominant pathway controlling the patterning of skin and influencing the decisions of embryonic and adult stem cells to adopt the various cell lineages of the skin and its appendages, as well as subsequently controlling the function of differentiated skin cells. Here we will review established concepts and present recent advances in our understanding of the diverse roles that Wnt signaling plays in skin development, homeostasis, and disease. PMID:23209129

  13. High Relief Block Printing.

    ERIC Educational Resources Information Center

    Foster, Michael

    1989-01-01

    Explains a method of block printing using styrofoam shapes to make high relief. Describes the creation of the block design as well as the actual printing process. Uses a range of paper types for printing so children can see the results of using different media. (LS)

  14. Gonad differentiation in zebrafish is regulated by the canonical Wnt signaling pathway.

    PubMed

    Sreenivasan, Rajini; Jiang, Junhui; Wang, Xingang; Bártfai, Richárd; Kwan, Hsiao Yuen; Christoffels, Alan; Orbán, László

    2014-02-01

    Zebrafish males undergo a "juvenile ovary-to-testis" gonadal transformation process. Several genes, including nuclear receptor subfamily 5, group A (nr5a) and anti-Müllerian hormone (amh), and pathways such as Tp53-mediated germ-cell apoptosis have been implicated in zebrafish testis formation. However, our knowledge of the regulation of this complex process is incomplete, and much remains to be investigated about the molecular pathways and network of genes that control it. Using a microarray-based analysis of transforming zebrafish male gonads, we demonstrated that their transcriptomes undergo transition from an ovary-like pattern to an ovotestis to a testis-like profile. Microarray results also validated the previous histological and immunohistochemical observation that there is high variation in the duration and extent of commitment to the juvenile ovary phase among individuals. Interestingly, global gene expression profiling of diverging zebrafish juvenile ovaries and transforming ovotestes revealed that some members of the canonical Wnt/beta-catenin signaling pathway were differentially expressed between these two phases. To investigate whether Wnt/beta-catenin signaling plays a role in zebrafish gonad differentiation, we used the Tg (hsp70l:dkk1b-GFP)w32 line to inhibit Wnt/beta-catenin signaling during gonad differentiation. Activation of dkk1b-GFP expression by heat shock resulted in an increased proportion of males and corresponding decrease in gonadal aromatase gene (cyp19a1a) expression. The Wnt target gene, lymphocyte enhancer binding factor 1 (lef1), was also down-regulated in the process. Together, these results provide the first functional evidence that, similarly to mammals, Wnt/beta-catenin signaling is a "pro-female" pathway that regulates gonad differentiation in zebrafish. PMID:24174574

  15. Expression of WNT genes in cervical cancer-derived cells: Implication of WNT7A in cell proliferation and migration

    SciTech Connect

    Ramos-Solano, Moisés; Meza-Canales, Ivan D.; Torres-Reyes, Luis A.; Alvarez-Zavala, Monserrat; and others

    2015-07-01

    According to the multifactorial model of cervical cancer (CC) causation, it is now recognized that other modifications, in addition to Human papillomavirus (HPV) infection, are necessary for the development of this neoplasia. Among these, it has been proposed that a dysregulation of the WNT pathway might favor malignant progression of HPV-immortalized keratinocytes. The aim of this study was to identify components of the WNT pathway differentially expressed in CC vs. non-tumorigenic, but immortalized human keratinocytes. Interestingly, WNT7A expression was found strongly downregulated in cell lines and biopsies derived from CC. Restoration of WNT7A in CC-derived cell lines using a lentiviral gene delivery system or after adding a recombinant human protein decreases cell proliferation. Likewise, WNT7A silencing in non-tumorigenic cells markedly accelerates proliferation. Decreased WNT7A expression was due to hypermethylation at particular CpG sites. To our knowledge, this is the first study reporting reduced WNT7A levels in CC-derived cells and that ectopic WNT7A restoration negatively affects cell proliferation and migration. - Highlights: • WNT7A is expressed in normal keratinocytes or cervical cells without lesion. • WNT7A is significantly reduced in cervical cancer-derived cells. • Restoration of WNT7A expression in HeLa decreases proliferation and cell migration. • Silencing of WNT7A in HaCaT induces an increased proliferation and migration rate. • Decreased WNT7A expression in this model is due to hypermethylation.

  16. Heterochromatin Blocks Constituting the Entire Short Arms of Acrocentric Chromosomes of Azara's Owl Monkey: Formation Processes Inferred From Chromosomal Locations

    PubMed Central

    Prakhongcheep, Ornjira; Chaiprasertsri, Nampech; Terada, Shoko; Hirai, Yuriko; Srikulnath, Kornsorn; Hirai, Hirohisa; Koga, Akihiko

    2013-01-01

    Centromeres and telomeres of higher eukaryotes generally contain repetitive sequences, which often form pericentric or subtelomeric heterochromatin blocks. C-banding analysis of chromosomes of Azara's owl monkey, a primate species, showed that the short arms of acrocentric chromosomes consist mostly or solely of constitutive heterochromatin. The purpose of the present study was to determine which category, pericentric, or subtelomeric is most appropriate for this heterochromatin, and to infer its formation processes. We cloned and sequenced its DNA component, finding it to be a tandem repeat sequence comprising 187-bp repeat units, which we named OwlRep. Subsequent hybridization analyses revealed that OwlRep resides in the pericentric regions of a small number of metacentric chromosomes, in addition to the short arms of acrocentric chromosomes. Further, in the pericentric regions of the acrocentric chromosomes, OwlRep was observed on the short-arm side only. This distribution pattern of OwlRep among chromosomes can be simply and sufficiently explained by assuming (i) OwlRep was transferred from chromosome to chromosome by the interaction of pericentric heterochromatin, and (ii) it was amplified there as subtelomeric heterochromatin. OwlRep carries several direct and inverted repeats within its repeat units. This complex structure may lead to a higher frequency of chromosome scission and may thus be a factor in the unique distribution pattern among chromosomes. Neither OwlRep nor similar sequences were found in the genomes of the other New World monkey species we examined, suggesting that OwlRep underwent rapid amplification after the divergence of the owl monkey lineage from lineages of the other species. PMID:23761219

  17. Hepatic Progenitor Cells Contribute to the Progression of 2-Acetylaminofluorene/Carbon Tetrachloride-Induced Cirrhosis via the Non-Canonical Wnt Pathway.

    PubMed

    Chen, Jiamei; Zhang, Xiao; Xu, Ying; Li, Xuewei; Ren, Shuang; Zhou, Yaning; Duan, Yuyou; Zern, Mark; Zhang, Hua; Chen, Gaofeng; Liu, Chenghai; Mu, Yongping; Liu, Ping

    2015-01-01

    Hepatic progenitor cells (HPCs) appear to play an important role in chronic liver injury. In this study, cirrhosis was induced in F-344 rats (n = 32) via subcutaneous injection of 50% carbon tetrachloride (CCl4) twice a week for 8 weeks. Then, 30% CCl4 was administered in conjunction with intragastric 2-acetylaminofluorine (2-AAF) for 4 weeks to induce activation of HPCs. WB-F344 cells were used to provide direct evidence for differentiation of HPCs to myofibroblasts. The results showed that after administration of 2-AAF, the hydroxyproline content and the expressions of α-SMA, Col I, Col IV, TGF-β1, CD68, TNF-α, CK19 and OV6 were significantly increased. OV6 and α-SMA were largely co-expressed in fibrous septum and the expressions of Wnt5b, frizzled2, frizzled3 and frizzled6 were markedly increased, while β-catenin expression was not statistically different among the different groups. Consistent with the above results, WB-F344 cells, treated with TGF-β1 in vitro, differentiated into myofibroblasts and α-SMA, Col I, Col IV, Wnt5b and frizzled2 expressions were significantly increased, while β-catenin expression was decreased. After blocking the non-canonical Wnt pathway via WIF-1, the Wnt5b level was down regulated, and α-SMA and F-actin expressions were significantly decreased in the WIF-1-treated cells. In conclusion, these results indicate that HPCs appear to differentiate into myofibroblasts and exhibit a profibrotic effect in progressive cirrhosis via activation of the non-canonical Wnt pathway. Blocking the non-canonical Wnt pathway can inhibit the differentiation of HPCs into myofibroblasts, suggesting that blocking this pathway and changing the fate of differentiated HPCs may be a potential treatment for cirrhosis. PMID:26087010

  18. Inhibition of Wnt Signaling by Silymarin in Human Colorectal Cancer Cells.

    PubMed

    Eo, Hyun Ji; Park, Gwang Hun; Jeong, Jin Boo

    2016-07-01

    Silymarin from milk thistle (Silybum marianum) has been reported to show an anti-cancer activity. In previous study, we reported that silymarin induces cyclin D1 proteasomal degradation through NF-κB-mediated threonine-286 phosphorylation. However, mechanism for the inhibition of Wnt signaling by silymarin still remains unanswered. Thus, we investigated whether silymarin affects Wnt signaling in human colorectal cancer cells to elucidate the additional anti-cancer mechanism of silymarin. Transient transfection with a TOP and FOP FLASH luciferase construct indicated that silymarin suppressed the transcriptional activity of β-catenin/TCF. Silymarin treatment resulted in a decrease of intracellular β-catenin protein but not mRNA. The inhibition of proteasome by MG132 and GSK3β inhibition by SB216763 blocked silymarin-mediated downregulation of β-catenin. In addition, silymarin increased phosphorylation of β-catenin and a point mutation of S33Y attenuated silymarin-mediated β-catenin downregulation. In addition, silymarin decreased TCF4 and increased Axin expression in both protein and mRNA level. From these results, we suggest that silymarin-mediated downregulation of β-catenin and TCF4 may result in the inhibition of Wnt signaling in human colorectal cancer cells. PMID:27068260

  19. Inhibition of Wnt Signaling by Silymarin in Human Colorectal Cancer Cells

    PubMed Central

    Eo, Hyun Ji; Park, Gwang Hun; Jeong, Jin Boo

    2016-01-01

    Silymarin from milk thistle (Silybum marianum) has been reported to show an anti-cancer activity. In previous study, we reported that silymarin induces cyclin D1 proteasomal degradation through NF-κB-mediated threonine-286 phosphorylation. However, mechanism for the inhibition of Wnt signaling by silymarin still remains unanswered. Thus, we investigated whether silymarin affects Wnt signaling in human colorectal cancer cells to elucidate the additional anti-cancer mechanism of silymarin. Transient transfection with a TOP and FOP FLASH luciferase construct indicated that silymarin suppressed the transcriptional activity of β-catenin/TCF. Silymarin treatment resulted in a decrease of intracellular β-catenin protein but not mRNA. The inhibition of proteasome by MG132 and GSK3β inhibition by SB216763 blocked silymarin-mediated downregulation of β-catenin. In addition, silymarin increased phosphorylation of β-catenin and a point mutation of S33Y attenuated silymarin-mediated β-catenin downregulation. In addition, silymarin decreased TCF4 and increased Axin expression in both protein and mRNA level. From these results, we suggest that silymarin-mediated downregulation of β-catenin and TCF4 may result in the inhibition of Wnt signaling in human colorectal cancer cells. PMID:27068260

  20. Wnt/β-Catenin Signaling Mediated-UCH-L1 Expression in Podocytes of Diabetic Nephropathy.

    PubMed

    Zhang, Hongxia; Luo, Weili; Sun, Yonghong; Qiao, Yanchun; Zhang, Liying; Zhao, Zhilian; Lv, Shijun

    2016-01-01

    Increasing studies identified podocyte injury as a key early risk factor resulting in diabetic nephropathy (DN). The ubiquitin carboxy-terminal hydrolase 1 (UCH-L1) participates in podocyte differentiation and injury, which is elevated in the podocytes of a variety of nephritis. Whether UCH-L1 expression is positively related to podocyte injury of DN remains unclear. In this study, elevated expression of UCH-L1 and its intrinsic mechanism in high glucose (HG)-stimulated murine podocytes were investigated using western blot and real-time quantitative PCR. Kidney biopsies of DN patients and health individuals were stained by immunofluorescence (IF) method. The morphological and functional changes of podocytes were tested by F-actin staining and cell migration assay. Results demonstrated that HG induced upregulation of UCH-L1 and activation of the Wnt/β-catenin signaling pathway in podocytes. However, blocking of the Wnt pathway by dickkopf related protein 1 (DKK1) eliminated the above changes. Furthermore, IF staining confirmed that, compared with healthy individuals, the expression of UCH-L1 and β-catenin were obviously increased in kidney biopsy of DN patients. Overexpression of UCH-L1 remodeled its actin cytoskeleton, increased its cell migration and impacted its important proteins. All the findings manifested that Wnt/β-catenin/UCH-L1 may be a new potential therapy method in the treatment of DN in future. PMID:27571062

  1. Activated Wnt signaling induces myofibroblast differentiation of mesenchymal stem cells, contributing to pulmonary fibrosis.

    PubMed

    Sun, Zhaorui; Wang, Cong; Shi, Chaowen; Sun, Fangfang; Xu, Xiaomeng; Qian, Weiping; Nie, Shinan; Han, Xiaodong

    2014-05-01

    Acute lung injury may lead to fibrogenesis. However, no treatment is currently available. This study was conducted to determine the effects of bone marrow-derived mesenchymal stem cells (MSCs) in a model of HCl-induced acute lung injury in Sprague-Dawley (SD) rats. Stromal cell-derived factor (SDF)-1 and its receptor CXC chemokine receptor (CXCR)4 have been shown to participate in mobilizing MSCs. Adenovirus carrying the CXCR4 gene was used to transfect MSCs in order to increase the engraftment numbers of MSCs at injured sites. Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis. The results showed that engraftment of MSCs almost differentiated into myofibroblasts, but rarely differentiated into lung epithelial cells. Additionally, it was demonstrated that activated canonical Wnt/β-catenin signaling in injured lung tissue regulated the myofibroblast differentiation of MSCs in vivo. The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1. Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury. PMID:24573542

  2. A possible role for the canonical Wnt pathway in endocrine cell development in chicks

    SciTech Connect

    Pedersen, Anna Hauntoft; Heller, R. Scott . E-mail: shll@hagedorn.dk

    2005-08-05

    Wnt signalling is involved in many developmental processes such as proliferation, differentiation, cell fate decisions, and morphogenesis. However, little is known about Wnt signalling during pancreas development. Multiple Wnt ligands and Frizzled receptors are expressed in the embryonic mouse pancreas, the surrounding mesenchyme, and have also been detected in the chicken endoderm during development. The aim of this study was to investigate the role of canonical Wnt signalling on endocrine cell development by use of the in ovo electroporation of the chicken endoderm. Overexpression with a constitutive active form of {beta}-catenin in combination with Ngn3 resulted in reduced numbers of glucagon cells. dnLEF-1 or naked-1 did not alter endocrine cell differentiation when co-expressed with Ngn3, but dnLEF-1 appeared to have some potential for inhibiting delamination of Ngn3 cells. In addition, neuronal {beta}-III-tubulin, which had previously been considered a specific marker for neuronal cells, was observed in the pancreas and was upregulated in the electroporated Ngn3 cells and thus may be a new endocrine marker in the chicken.

  3. Modulation of Wnt Signaling Enhances Inner Ear Organoid Development in 3D Culture.

    PubMed

    DeJonge, Rachel E; Liu, Xiao-Ping; Deig, Christopher R; Heller, Stefan; Koehler, Karl R; Hashino, Eri

    2016-01-01

    Stem cell-derived inner ear sensory epithelia are a promising source of tissues for treating patients with hearing loss and dizziness. We recently demonstrated how to generate inner ear sensory epithelia, designated as inner ear organoids, from mouse embryonic stem cells (ESCs) in a self-organizing 3D culture. Here we improve the efficiency of this culture system by elucidating how Wnt signaling activity can drive the induction of otic tissue. We found that a carefully timed treatment with the potent Wnt agonist CHIR99021 promotes induction of otic vesicles-a process that was previously self-organized by unknown mechanisms. The resulting otic-like vesicles have a larger lumen size and contain a greater number of Pax8/Pax2-positive otic progenitor cells than organoids derived without the Wnt agonist. Additionally, these otic-like vesicles give rise to large inner ear organoids with hair cells whose morphological, biochemical and functional properties are indistinguishable from those of vestibular hair cells in the postnatal mouse inner ear. We conclude that Wnt signaling plays a similar role during inner ear organoid formation as it does during inner ear development in the embryo. PMID:27607106

  4. The miR-20-Rest-Wnt signaling axis regulates neural progenitor cell differentiation

    PubMed Central

    Cui, Yi; Han, Jin; Xiao, Zhifeng; Chen, Tong; Wang, Bin; Chen, Bing; Liu, Sumei; Han, Sufang; Fang, Yongxiang; Wei, Jianshu; Wang, Xiujie; Ma, Xu; Dai, Jianwu

    2016-01-01

    Increasing evidence suggests that three dimensional (3-D) cell cultures are an improvement over traditional two dimensional (2-D) cell cultures. Current researches have extensively focused on the study of utilizing biomaterial-based 3-D culture systems to study and direct stem-cell fate both in vitro and in vivo. Here in our study, we screened the differential expression patterns of miRNAs between 2-D cultured and 3-D cultured NPCs using microarray analysis. Among these differentially expressed miRNAs, miR-20 was found to increase during differentiation of NPCs. Specifically, the facilitative effect on neural differentiation of miR-20 is mediated, at least in part by directly target the Rest gene, which is essential for preventing neural differentiation and maintaining NPCs self-renewal. Furthermore, the expression of miR-20 was decreased when the WNT pathway was inhibited by knock down of β-catenin or by exogenous Dkk protein, whereas it increased when the WNT pathway was activated by exogenous Wnt3a protein. Overall, miR-20, Rest and Wnt signaling are suggested to be involved in a regulatory circuit that can modulate the neural differention of NPCs. This novel regulatory circuit provides additional insight into how microRNAs interact with signaling molecules during neural differentiation of NPCs, allowing for fine-tuning of intricate cellular processes. PMID:26996236

  5. Molecular Genetics of Intracranial Meningiomas with Emphasis on Canonical Wnt Signalling

    PubMed Central

    Pećina-Šlaus, Nives; Kafka, Anja; Lechpammer, Mirna

    2016-01-01

    Research over the last decade recognized the importance of novel molecular pathways in pathogenesis of intracranial meningiomas. In this review, we focus on human brain tumours meningiomas and the involvement of Wnt signalling pathway genes and proteins in this common brain tumour, describing their known functional effects. Meningiomas originate from the meningeal layers of the brain and the spinal cord. Most meningiomas have benign clinical behaviour and are classified as grade I by World Health Organization (WHO). However, up to 20% histologically classified as atypical (grade II) or anaplastic (grade III) are associated with higher recurrent rate and have overall less favourable clinical outcome. Recently, there is emerging evidence that multiple signalling pathways including Wnt pathway contribute to the formation and growth of meningiomas. In the review we present the synopsis on meningioma histopathology and genetics and discuss our research regarding Wnt in meningioma. Epithelial-to-mesenchymal transition, a process in which Wnt signalling plays an important role, is shortly discussed. PMID:27429002

  6. BMP-FGF Signaling Axis Mediates Wnt-Induced Epidermal Stratification in Developing Mammalian Skin

    PubMed Central

    Zhu, Xiao-Jing; Liu, YuDong; Dai, Zhong-Min; Zhang, Xiaoyun; Yang, XueQin; Li, Yan; Qiu, Mengsheng; Fu, Jiang; Hsu, Wei; Chen, YiPing; Zhang, Zunyi

    2014-01-01

    Epidermal stratification of the mammalian skin requires proliferative basal progenitors to generate intermediate cells that separate from the basal layer and are replaced by post-mitotic cells. Although Wnt signaling has been implicated in this developmental process, the mechanism underlying Wnt-mediated regulation of basal progenitors remains elusive. Here we show that Wnt secreted from proliferative basal cells is not required for their differentiation. However, epidermal production of Wnts is essential for the formation of the spinous layer through modulation of a BMP-FGF signaling cascade in the dermis. The spinous layer defects caused by disruption of Wnt secretion can be restored by transgenically expressed Bmp4. Non-cell autonomous BMP4 promotes activation of FGF7 and FGF10 signaling, leading to an increase in proliferative basal cell population. Our findings identify an essential BMP-FGF signaling axis in the dermis that responds to the epidermal Wnts and feedbacks to regulate basal progenitors during epidermal stratification. PMID:25329657

  7. A Fully Human Inhibitory Monoclonal Antibody to the Wnt Receptor RYK

    PubMed Central

    Parish, Clare L.; Takano, Elena A.; Fox, Stephen; Layton, Daniel; Nice, Edouard; Stacker, Steven A.

    2013-01-01

    RYK is an unusual member of the receptor tyrosine kinase (RTK) family that is classified as a putative pseudokinase. RYK regulates fundamental biological processes including cell differentiation, migration and target selection, axon outgrowth and pathfinding by transducing signals across the plasma membrane in response to the high affinity binding of Wnt family ligands to its extracellular Wnt inhibitory factor (WIF) domain. Here we report the generation and initial characterization of a fully human inhibitory monoclonal antibody to the human RYK WIF domain. From a naïve human single chain fragment variable (scFv) phage display library, we identified anti-RYK WIF domain–specific scFvs then screened for those that could compete with Wnt3a for binding. Production of a fully human IgG1κ from an inhibitory scFv yielded a monoclonal antibody that inhibits Wnt5a-responsive RYK function in a neurite outgrowth assay. This antibody will have immediate applications for modulating RYK function in a range of settings including development and adult homeostasis, with significant potential for therapeutic use in human pathologies. PMID:24058687

  8. Molecular Genetics of Intracranial Meningiomas with Emphasis on Canonical Wnt Signalling.

    PubMed

    Pećina-Šlaus, Nives; Kafka, Anja; Lechpammer, Mirna

    2016-01-01

    Research over the last decade recognized the importance of novel molecular pathways in pathogenesis of intracranial meningiomas. In this review, we focus on human brain tumours meningiomas and the involvement of Wnt signalling pathway genes and proteins in this common brain tumour, describing their known functional effects. Meningiomas originate from the meningeal layers of the brain and the spinal cord. Most meningiomas have benign clinical behaviour and are classified as grade I by World Health Organization (WHO). However, up to 20% histologically classified as atypical (grade II) or anaplastic (grade III) are associated with higher recurrent rate and have overall less favourable clinical outcome. Recently, there is emerging evidence that multiple signalling pathways including Wnt pathway contribute to the formation and growth of meningiomas. In the review we present the synopsis on meningioma histopathology and genetics and discuss our research regarding Wnt in meningioma. Epithelial-to-mesenchymal transition, a process in which Wnt signalling plays an important role, is shortly discussed. PMID:27429002

  9. A fully human inhibitory monoclonal antibody to the Wnt receptor RYK.

    PubMed

    Halford, Michael M; Macheda, Maria L; Parish, Clare L; Takano, Elena A; Fox, Stephen; Layton, Daniel; Nice, Edouard; Stacker, Steven A

    2013-01-01

    RYK is an unusual member of the receptor tyrosine kinase (RTK) family that is classified as a putative pseudokinase. RYK regulates fundamental biological processes including cell differentiation, migration and target selection, axon outgrowth and pathfinding by transducing signals across the plasma membrane in response to the high affinity binding of Wnt family ligands to its extracellular Wnt inhibitory factor (WIF) domain. Here we report the generation and initial characterization of a fully human inhibitory monoclonal antibody to the human RYK WIF domain. From a naïve human single chain fragment variable (scFv) phage display library, we identified anti-RYK WIF domain-specific scFvs then screened for those that could compete with Wnt3a for binding. Production of a fully human IgG1κ from an inhibitory scFv yielded a monoclonal antibody that inhibits Wnt5a-responsive RYK function in a neurite outgrowth assay. This antibody will have immediate applications for modulating RYK function in a range of settings including development and adult homeostasis, with significant potential for therapeutic use in human pathologies. PMID:24058687

  10. The Use of Chick Embryos to Study Wnt Activity Gradients.

    PubMed

    Galli, Lisa M; Barnes, Tiffany; Burrus, Laura W

    2016-01-01

    The chick spinal cord provides a valuable model for assessing Wnt signaling activity. Loss or gain of function constructs that are transfected by electroporation can be directed to a single side of the spinal cord, thus leaving the contralateral side as an internal control. Here, we describe a method for measuring Wnt signaling via the use of BAT-Gal, a β-catenin dependent Wnt reporter. PMID:27590153

  11. Preclinical validation and imaging of Wnt-induced repair in human 3D lung tissue cultures.

    PubMed

    Uhl, Franziska E; Vierkotten, Sarah; Wagner, Darcy E; Burgstaller, Gerald; Costa, Rita; Koch, Ina; Lindner, Michael; Meiners, Silke; Eickelberg, Oliver; Königshoff, Melanie

    2015-10-01

    Chronic obstructive pulmonary disease (COPD) is characterised by a progressive loss of lung tissue. Inducing repair processes within the adult diseased lung is of major interest and Wnt/β-catenin signalling represents a promising target for lung repair. However, the translation of novel therapeutic targets from model systems into clinical use remains a major challenge.We generated murine and patient-derived three-dimensional (3D) ex vivo lung tissue cultures (LTCs), which closely mimic the 3D lung microenvironment in vivo. Using two well-known glycogen synthase kinase-3β inhibitors, lithium chloride (LiCl) and CHIR 99021 (CT), we determined Wnt/β-catenin-driven lung repair processes in high spatiotemporal resolution using quantitative PCR, Western blotting, ELISA, (immuno)histological assessment, and four-dimensional confocal live tissue imaging.Viable 3D-LTCs exhibited preserved lung structure and function for up to 5 days. We demonstrate successful Wnt/β-catenin signal activation in murine and patient-derived 3D-LTCs from COPD patients. Wnt/β-catenin signalling led to increased alveolar epithelial cell marker expression, decreased matrix metalloproteinase-12 expression, as well as altered macrophage activity and elastin remodelling. Importantly, induction of surfactant protein C significantly correlated with disease stage (per cent predicted forced expiratory volume in 1 s) in patient-derived 3D-LTCs.Patient-derived 3D-LTCs represent a valuable tool to analyse potential targets and drugs for lung repair. Enhanced Wnt/β-catenin signalling attenuated pathological features of patient-derived COPD 3D-LTCs. PMID:25929950

  12. Wnt-regulated dynamics of positional information in zebrafish somitogenesis

    PubMed Central

    Bajard, Lola; Morelli, Luis G.; Ares, Saúl; Pécréaux, Jacques; Jülicher, Frank; Oates, Andrew C.

    2014-01-01

    How signaling gradients supply positional information in a field of moving cells is an unsolved question in patterning and morphogenesis. Here, we ask how a Wnt signaling gradient regulates the dynamics of a wavefront of cellular change in a flow of cells during somitogenesis. Using time-controlled perturbations of Wnt signaling in the zebrafish embryo, we changed segment length without altering the rate of somite formation or embryonic elongation. This result implies specific Wnt regulation of the wavefront velocity. The observed Wnt signaling gradient dynamics and timing of downstream events support a model for wavefront regulation in which cell flow plays a dominant role in transporting positional information. PMID:24595291

  13. Ionic Blocks

    ERIC Educational Resources Information Center

    Sevcik, Richard S.; Gamble, Rex; Martinez, Elizabet; Schultz, Linda D.; Alexander, Susan V.

    2008-01-01

    "Ionic Blocks" is a teaching tool designed to help middle school students visualize the concepts of ions, ionic compounds, and stoichiometry. It can also assist high school students in reviewing their subject mastery. Three dimensional blocks are used to represent cations and anions, with color indicating charge (positive or negative) and size…

  14. Structures of Wnt-Antagonist ZNRF3 and Its Complex with R-Spondin 1 and Implications for Signaling

    PubMed Central

    Peng, Weng Chuan; de Lau, Wim; Madoori, Pramod K.; Forneris, Federico; Granneman, Joke C. M.; Clevers, Hans; Gros, Piet

    2013-01-01

    Zinc RING finger 3 (ZNRF3) and its homolog RING finger 43 (RNF43) antagonize Wnt signaling in adult stem cells by ubiquitinating Frizzled receptors (FZD), which leads to endocytosis of the Wnt receptor. Conversely, binding of ZNRF3/RNF43 to LGR4-6 – R-spondin blocks Frizzled ubiquitination and enhances Wnt signaling. Here, we present crystal structures of the ZNRF3 ectodomain and its complex with R-spondin 1 (RSPO1). ZNRF3 binds RSPO1 and LGR5-RSPO1 with micromolar affinity via RSPO1 furin-like 1 (Fu1) domain. Anonychia-related mutations in RSPO4 support the importance of the observed interface. The ZNRF3-RSPO1 structure resembles that of LGR5-RSPO1-RNF43, though Fu2 of RSPO1 is variably oriented. The ZNRF3-binding site overlaps with trans-interactions observed in 2:2 LGR5-RSPO1 complexes, thus binding of ZNRF3/RNF43 would disrupt such an arrangement. Sequence conservation suggests a single ligand-binding site on ZNRF3, consistent with the proposed competing binding role of ZNRF3/RNF43 in Wnt signaling. PMID:24349440

  15. Chronic skin inflammation leads to bone loss by IL-17-mediated inhibition of Wnt signaling in osteoblasts.

    PubMed

    Uluçkan, Özge; Jimenez, Maria; Karbach, Susanne; Jeschke, Anke; Graña, Osvaldo; Keller, Johannes; Busse, Björn; Croxford, Andrew L; Finzel, Stephanie; Koenders, Marije; van den Berg, Wim; Schinke, Thorsten; Amling, Michael; Waisman, Ari; Schett, Georg; Wagner, Erwin F

    2016-03-16

    Inflammation has important roles in tissue regeneration, autoimmunity, and cancer. Different inflammatory stimuli can lead to bone loss by mechanisms that are not well understood. We show that skin inflammation induces bone loss in mice and humans. In psoriasis, one of the prototypic IL-17A-mediated inflammatory human skin diseases, low bone formation and bone loss correlated with increased serum IL-17A levels. Similarly, in two mouse models with chronic IL-17A-mediated skin inflammation,K14-IL17A(ind)andJunB(Δep), strong inhibition of bone formation was observed, different from classical inflammatory bone loss where osteoclast activation leads to bone degradation. We show that under inflammatory conditions, skin-resident cells such as keratinocytes, γδ T cells, and innate lymphoid cells were able to express IL-17A, which acted systemically to inhibit osteoblast and osteocyte function by a mechanism involving Wnt signaling. IL-17A led to decreased Wnt signaling in vitro, and importantly, pharmacological blockade of IL-17A rescued Wnt target gene expression and bone formation in vivo. These data provide a mechanism where IL-17A affects bone formation by regulating Wnt signaling in osteoblasts and osteocytes. This study suggests that using IL-17A blocking agents in psoriasis could be beneficial against bone loss in these patients. PMID:27089206

  16. Wnt-dependent osteogenic commitment of bone marrow stromal cells using a novel GSK3β inhibitor.

    PubMed

    Cook, David A; Fellgett, Simon W; Pownall, Mary E; O'Shea, Patrick J; Genever, Paul G

    2014-03-01

    Bone marrow stromal cells (BMSCs, also known as bone marrow-derived mesenchymal stem cells) can differentiate into multiple lineages including osteogenic and adipogenic cells. Wnt signalling has been implicated in controlling BMSC fate, but the mechanisms are unclear and apparently conflicting data exist. Here we show that a novel glycogen synthase kinase 3β inhibitor, AR28, is a potent activator of canonical Wnt signalling using in vitro β-catenin translocation studies and TCF-reporter assays. In vivo, AR28 induced characteristic axis duplication and secondary regions of chordin expression in Xenopus laevis embryos. Using human BMSCs grown in adipogenic medium, we confirmed that AR28-mediated Wnt signalling caused a significant (p<0.05) dose-dependent reduction of adipogenic markers. In osteogenic media, including dexamethasone, AR28 caused significant (p<0.05) decreases in alkaline phosphatase (ALP) activity compared to vehicle controls, indicative of a reduced osteogenic response. However, when excluding dexamethasone from the osteogenic media, increases in both ALP and mineralisation were identified following AR28 treatment, which was blocked by mitomycin C. Pre-treatment of BMSCs with AR28 for 7 days before osteogenic induction also increased ALP activity and mineralisation. Furthermore, BMP2-induced osteogenic differentiation was strongly enhanced by AR28 addition within 3 days, but without concomitant changes in cell number, therefore revealing BMP-dependent and independent mechanisms for Wnt-induced osteogenesis. PMID:24382458

  17. Canonical Wnt activity regulates trunk neural crest delamination linking BMP/noggin signaling with G1/S transition.

    PubMed

    Burstyn-Cohen, Tal; Stanleigh, Jonathan; Sela-Donenfeld, Dalit; Kalcheim, Chaya

    2004-11-01

    Delamination of premigratory neural crest cells depends on a balance between BMP/noggin and on successful G1/S transition. Here, we report that BMP regulates G1/S transition and consequent crest delamination through canonical Wnt signaling. Noggin overexpression inhibits G1/S transition and blocking G1/S abrogates BMP-induced delamination; moreover, transcription of Wnt1 is stimulated by BMP and by the developing somites, which concomitantly inhibit noggin production. Interfering with beta-catenin and LEF/TCF inhibits G1/S transition, neural crest delamination and transcription of various BMP-dependent genes, which include Cad6B, Pax3 and Msx1, but not that of Slug, Sox9 or FoxD3. Hence, we propose that developing somites inhibit noggin transcription in the dorsal tube, resulting in activation of BMP and consequent Wnt1 production. Canonical Wnt signaling in turn stimulates G1/S transition and generation of neural crest cell motility independently of its proposed role in earlier neural crest specification. PMID:15456730

  18. Systematic mapping of WNT-FZD protein interactions reveals functional selectivity by distinct WNT-FZD pairs.

    PubMed

    Dijksterhuis, Jacomijn P; Baljinnyam, Bolormaa; Stanger, Karen; Sercan, Hakki O; Ji, Yun; Andres, Osler; Rubin, Jeffrey S; Hannoush, Rami N; Schulte, Gunnar

    2015-03-13

    The seven-transmembrane-spanning receptors of the FZD1-10 class are bound and activated by the WNT family of lipoglycoproteins, thereby inducing a complex network of signaling pathways. However, the specificity of the interaction between mammalian WNT and FZD proteins and the subsequent signaling cascade downstream of the different WNT-FZD pairs have not been systematically addressed to date. In this study, we determined the binding affinities of various WNTs for different members of the FZD family by using bio-layer interferometry and characterized their functional selectivity in a cell system. Using purified WNTs, we show that different FZD cysteine-rich domains prefer to bind to distinct WNTs with fast on-rates and slow off-rates. In a 32D cell-based system engineered to overexpress FZD2, FZD4, or FZD5, we found that WNT-3A (but not WNT-4, -5A, or -9B) activated the WNT-β-catenin pathway through FZD2/4/5 as measured by phosphorylation of LRP6 and β-catenin stabilization. Surprisingly, different WNT-FZD pairs showed differential effects on phosphorylation of DVL2 and DVL3, revealing a previously unappreciated DVL isoform selectivity by different WNT-FZD pairs in 32D cells. In summary, we present extensive mapping of WNT-FZD cysteine-rich domain interactions complemented by analysis of WNT-FZD pair functionality in a unique cell system expressing individual FZD isoforms. Differential WNT-FZD binding and selective functional readouts suggest that endogenous WNT ligands evolved with an intrinsic natural bias toward different downstream signaling pathways, a phenomenon that could be of great importance in the design of FZD-targeting drugs. PMID:25605717

  19. Systematic Mapping of WNT-FZD Protein Interactions Reveals Functional Selectivity by Distinct WNT-FZD Pairs*

    PubMed Central

    Dijksterhuis, Jacomijn P.; Baljinnyam, Bolormaa; Stanger, Karen; Sercan, Hakki O.; Ji, Yun; Andres, Osler; Rubin, Jeffrey S.; Hannoush, Rami N.; Schulte, Gunnar

    2015-01-01

    The seven-transmembrane-spanning receptors of the FZD1–10 class are bound and activated by the WNT family of lipoglycoproteins, thereby inducing a complex network of signaling pathways. However, the specificity of the interaction between mammalian WNT and FZD proteins and the subsequent signaling cascade downstream of the different WNT-FZD pairs have not been systematically addressed to date. In this study, we determined the binding affinities of various WNTs for different members of the FZD family by using bio-layer interferometry and characterized their functional selectivity in a cell system. Using purified WNTs, we show that different FZD cysteine-rich domains prefer to bind to distinct WNTs with fast on-rates and slow off-rates. In a 32D cell-based system engineered to overexpress FZD2, FZD4, or FZD5, we found that WNT-3A (but not WNT-4, -5A, or -9B) activated the WNT-β-catenin pathway through FZD2/4/5 as measured by phosphorylation of LRP6 and β-catenin stabilization. Surprisingly, different WNT-FZD pairs showed differential effects on phosphorylation of DVL2 and DVL3, revealing a previously unappreciated DVL isoform selectivity by different WNT-FZD pairs in 32D cells. In summary, we present extensive mapping of WNT-FZD cysteine-rich domain interactions complemented by analysis of WNT-FZD pair functionality in a unique cell system expressing individual FZD isoforms. Differential WNT-FZD binding and selective functional readouts suggest that endogenous WNT ligands evolved with an intrinsic natural bias toward different downstream signaling pathways, a phenomenon that could be of great importance in the design of FZD-targeting drugs. PMID:25605717

  20. Caught in a Wnt storm: Complexities of Wnt signaling in hematopoiesis.

    PubMed

    Staal, Frank J T; Chhatta, Amiet; Mikkers, Harald

    2016-06-01

    The Wnt signaling pathway is an evolutionary conserved pathway that is involved in the development of almost every organ system in the body and provides self-renewal signals for most, if not all, adult stem cell systems. In recent years, this pathway has been studied by various research groups working on hematopoietic stem cells, resulting in contradicting conclusions. Here, we discuss and interpret the results of these studies and propose that Wnt dosage, the source of hematopoietic stem cells, and interactions with other pathways explain these disparate results. PMID:27016274

  1. Rabconnectin-3a Regulates Vesicle Endocytosis and Canonical Wnt Signaling in Zebrafish Neural Crest Migration

    PubMed Central

    Tuttle, Adam M.; Hoffman, Trevor L.; Schilling, Thomas F.

    2014-01-01

    Cell migration requires dynamic regulation of cell–cell signaling and cell adhesion. Both of these processes involve endocytosis, lysosomal degradation, and recycling of ligand–receptor complexes and cell adhesion molecules from the plasma membrane. Neural crest (NC) cells in vertebrates are highly migratory cells, which undergo an epithelial–mesenchymal transition (EMT) to leave the neural epithelium and migrate throughout the body to give rise to many different derivatives. Here we show that the v-ATPase interacting protein, Rabconnectin-3a (Rbc3a), controls intracellular trafficking events and Wnt signaling during NC migration. In zebrafish embryos deficient in Rbc3a, or its associated v-ATPase subunit Atp6v0a1, many NC cells fail to migrate and misregulate expression of cadherins. Surprisingly, endosomes in Rbc3a- and Atp6v0a1-deficient NC cells remain immature but still acidify. Rbc3a loss-of-function initially downregulates several canonical Wnt targets involved in EMT, but later Frizzled-7 accumulates at NC cell membranes, and nuclear B-catenin levels increase. Presumably due to this later Wnt signaling increase, Rbc3a-deficient NC cells that fail to migrate become pigment progenitors. We propose that Rbc3a and Atp6v0a1 promote endosomal maturation to coordinate Wnt signaling and intracellular trafficking of Wnt receptors and cadherins required for NC migration and cell fate determination. Our results suggest that different v-ATPases and associated proteins may play cell-type-specific functions in intracellular trafficking in many contexts. PMID:24802872

  2. WNT/PCP signaling pathway and human cancer (review).

    PubMed

    Katoh, Masaru

    2005-12-01

    WNT/planar cell polarity (PCP) signaling pathway controls tissue polarity and cell movement through the activation of RHOA, c-Jun N-terminal kinase (JNK), and nemo-like kinase (NLK) signaling cascades. PCP is induced in Drosophila by the asymmetrical localization of Frizzled-Dishevelled-Diego-Starry night (Flamingo) complex and Van Gogh (Strabismus)-Prickle complex. Here, WNT/PCP signaling pathway implicated in human carcinogenesis is reviewed. Human WNT5A, WNT5B, and WNT11 are representative non-canonical WNTs transducing PCP signals through FZD3 or FZD6 receptors, and ROR1, ROR2 or PTK7 co-receptors. Human VANGL1, VANGL2 (Van Gogh homologs), CELSR1, CELSR2, CELSR3 (Starry night homologs), DVL1, DVL2, DVL3 (Dishevelled homologs), PRICKLE1, PRICKLE2 (Prickle homologs), and ANKRD6 (Diego homolog) are core PCP signaling molecules. MAGI3 assembles FZD, VANGL, PTEN, and adhesion molecules. Dishevelled-dependent WNT/PCP signals are transduced to the RHOA signaling cascade through Formin homology proteins DAAM1 and DAAM2, and to the JNK signaling cascade through MAPKKKs and MAPKK4/7. Dishevelled-independent WNT/ PCP signals are transduced to the NLK signaling cascade through MAP3K7 (TAK1). ANKRD6, NKD1 and NKD2 induce class switch from the WNT/GSK3beta signaling pathway to the WNT/PCP signaling pathway. WNT5A is up-regulated in various types of human cancer, such as gastric cancer, lung cancer, and melanoma. FZD3/FZD6 receptor and ROR2 co-receptor transduce WNT5A signal in gastric cancer. Aberrant activation of WNT/PCP signaling pathway in human cancer leads to more malignant phenotypes, such as abnormal tissue polarity, invasion, and metastasis. cDNA-PCR, microarray or ELISA reflecting aberrant activation of WNT/PCP signaling pathway could be developed as novel cancer prognostics. Single nucleotide polymorphism (SNP) and copy number polymorphism (CNP) of WNT/PCP signaling molecules mentioned above are suitable for use in screening of cancer predisposition, especially

  3. Functional Wnt Signaling Is Increased in Idiopathic Pulmonary Fibrosis

    PubMed Central

    Königshoff, Melanie; Balsara, Nisha; Pfaff, Eva-Maria; Kramer, Monika; Chrobak, Izabella; Seeger, Werner; Eickelberg, Oliver

    2008-01-01

    Background Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease, characterized by distorted lung architecture and loss of respiratory function. Alveolar epithelial cell injury and hyperplasia, enhanced extracellular matrix deposition, and (myo)fibroblast activation are features of IPF. Wnt/β-catenin signaling has been shown to determine epithelial cell fate during development. As aberrant reactivation of developmental signaling pathways has been suggested to contribute to IPF pathogenesis, we hypothesized that Wnt/β-catenin signaling is activated in epithelial cells in IPF. Thus, we quantified and localized the expression and activity of the Wnt/β-catenin pathway in IPF. Methodology/Principal Findings The expression of Wnt1, 3a, 7b, and 10b, the Wnt receptors Fzd1-4, Lrp5-6, as well as the intracellular signal transducers Gsk-3β, β-catenin, Tcf1, 3, 4, and Lef1 was analyzed in IPF and transplant donor lungs by quantitative real-time (q)RT-PCR. Wnt1, 7b and 10b, Fzd2 and 3, β-catenin, and Lef1 expression was significantly increased in IPF. Immunohistochemical analysis localized Wnt1, Wnt3a, β-catenin, and Gsk-3β expression largely to alveolar and bronchial epithelium. This was confirmed by qRT-PCR of primary alveolar epithelial type II (ATII) cells, demonstrating a significant increase of Wnt signaling in ATII cells derived from IPF patients. In addition, Western blot analysis of phospho-Gsk-3β, phospho-Lrp6, and β-catenin, and qRT-PCR of the Wnt target genes cyclin D1, Mmp 7, or Fibronectin 1 demonstrated increased functional Wnt/β-catenin signaling in IPF compared with controls. Functional in vitro studies further revealed that Wnt ligands induced lung epithelial cell proliferation and (myo)fibroblast activation and collagen synthesis. Conclusions/Significance Our study demonstrates that the Wnt/β-catenin pathway is expressed and operative in adult lung epithelium. Increased Wnt/β-catenin signaling may be involved in epithelial cell injury and

  4. Wnt/β-catenin signaling in bone marrow niche.

    PubMed

    Ahmadzadeh, Ahmad; Norozi, Fatemeh; Shahrabi, Saeid; Shahjahani, Mohammad; Saki, Najmaldin

    2016-02-01

    The bone marrow (BM) niche is a specific physiological environment for hematopoietic and non-hematopoietic stem cells (HSCs). Several signaling pathways (including Wnt/β-catenin) regulate various aspects of stem cell growth, function and death in the BM niche. In addition, the canonical Wnt pathway is crucial for directing self-renewal and differentiation as important mechanisms in many types of stem cells. We review the role of the Wnt/β-catenin pathway in the BM niche and its importance in stem cells. Relevant literature was identified by a PubMed search (1997-2014) of English-language literature by using the following keywords: BM niche, Wnt/β-catenin signaling, osteoblast, osteoclast and bone disease. The Wnt/β-catenin pathway regulates the stability of the β-catenin proto-oncogene. The stabilized β-catenin then translocates to the nucleus, forming a β-catenin-TCF/LEF complex regulating the transcription of specific target genes. Stem cells require β-catenin to mediate their response to Wnt signaling for maintenance and transition from the pluripotent state during embryogenesis. In adult stem cells, Wnt signaling functions at various hierarchical levels to contribute to the specification of the diverse tissues. Aberrant Wnt/β-catenin signaling and its downstream transcriptional regulators are observed in several malignant stem cells and human cancers. Because Wnt signaling can maintain stem cells and cancer cells, the ability to modulate the Wnt pathway either positively or negatively may be of therapeutic relevance. The controlled activation of Wnt signaling might allow us to enhance stem and progenitor cell activity when regeneration is needed. PMID:26475718

  5. Wnt pathway in Dupuytren disease: connecting profibrotic signals.

    PubMed

    van Beuge, Marike M; Ten Dam, Evert-Jan P M; Werker, Paul M N; Bank, Ruud A

    2015-12-01

    A role of Wnt signaling in Dupuytren disease, a fibroproliferative disease of the hand and fingers, has not been fully elucidated. We examined a large set of Wnt pathway components and signaling targets and found significant dysregulation of 41 Wnt-related genes in tissue from the Dupuytren nodules compared with patient-matched control tissue. A large proportion of genes coding for Wnt proteins themselves was downregulated. However, both canonical Wnt targets and components of the noncanonical signaling pathway were upregulated. Immunohistochemical analysis revealed that protein expression of Wnt1-inducible secreted protein 1 (WISP1), a known Wnt target, was increased in nodules compared with control tissue, but knockdown of WISP1 using small interfering RNA (siRNA) in the Dupuytren myofibroblasts did not confirm a functional role. The protein expression of noncanonical pathway components Wnt5A and VANGL2 as well as noncanonical coreceptors Ror2 and Ryk was increased in nodules. On the contrary, the strongest downregulated genes in this study were 4 antagonists of Wnt signaling (DKK1, FRZB, SFRP1, and WIF1). Downregulation of these genes in the Dupuytren tissue was mimicked in vitro by treating normal fibroblasts with transforming growth factor β1 (TGF-β1), suggesting cross talk between different profibrotic pathways. Furthermore, siRNA-mediated knockdown of these antagonists in normal fibroblasts led to increased nuclear translocation of Wnt target β-catenin in response to TGF-β1 treatment. In conclusion, we have shown extensive dysregulation of Wnt signaling in affected tissue from Dupuytren disease patients. Components of both the canonical and the noncanonical pathways are upregulated, whereas endogenous antagonists are downregulated, possibly via interaction with other profibrotic pathways. PMID:26470681

  6. Positioning of the midbrain-hindbrain boundary organizer through global posteriorization of the neuroectoderm mediated by Wnt8 signaling.

    PubMed

    Rhinn, Muriel; Lun, Klaus; Luz, Marta; Werner, Michaela; Brand, Michael

    2005-03-01

    The organizing center located at the midbrain-hindbrain boundary (MHB) patterns the midbrain and hindbrain primordia of the neural plate. Studies in several vertebrates showed that the interface between cells expressing Otx and Gbx transcription factors marks the location in the neural plate where the organizer forms, but it is unclear how this location is set up. Using mutant analyses and shield ablation experiments in zebrafish, we find that axial mesendoderm, as a candidate tissue, has only a minor role in positioning the MHB. Instead, the blastoderm margin of the gastrula embryo acts as a source of signal(s) involved in this process. We demonstrate that positioning of the MHB organizer is tightly linked to overall neuroectodermal posteriorization, and specifically depends on Wnt8 signaling emanating from lateral mesendodermal precursors. Wnt8 is required for the initial subdivision of the neuroectoderm, including onset of posterior gbx1 expression and establishment of the posterior border of otx2 expression. Cell transplantation experiments further show that Wnt8 signaling acts directly and non-cell-autonomously. Consistent with these findings, a GFP-Wnt8 fusion protein travels from donor cells through early neural plate tissue. Our findings argue that graded Wnt8 activity mediates overall neuroectodermal posteriorization and thus determines the location of the MHB organizer. PMID:15703279

  7. HuR represses Wnt/β-catenin-mediated transcriptional activity by promoting cytoplasmic localization of β-catenin

    SciTech Connect

    Kim, Inae; Hur, Jung; Jeong, Sunjoo

    2015-01-30

    Highlights: • Wnt signaling as well as β-catenin overexpression enhance HuR cytoplasmic export. • HuR overexpression promotes cytoplasmic localization of β-catenin from the perinuclear fraction. • Wnt/β-catenin-mediated transcriptional activity is repressesed by HuR. - Abstract: β-Catenin is the key transcriptional activator of canonical Wnt signaling in the nucleus; thus, nuclear accumulation of β-catenin is a critical step for expressing target genes. β-Catenin accumulates in the nucleus of cancer cells where it activates oncogenic target genes. Hu antigen R (HuR) is a RNA binding protein that regulates multiple post-transcriptional processes including RNA stability. Thus, cytoplasmic HuR protein may be involved in tumorigenesis by stabilizing oncogenic transcripts, but the molecular mechanism remains unclear. Here, we observed that Wnt/β-catenin signaling induced export of the HuR protein, whereas HuR overexpression promoted accumulation of the β-catenin protein in the cytoplasm. Thus, Wnt/β-catenin-mediated transcriptional activity in the nucleus was reduced by overexpressing HuR. These results suggest novel and uncharacterized cytoplasmic β-catenin functions related to HuR-mediated RNA metabolism in cancer cells.

  8. PLD1 regulates Xenopus convergent extension movements by mediating Frizzled7 endocytosis for Wnt/PCP signal activation.

    PubMed

    Lee, Hyeyoon; Lee, Seung Joon; Kim, Gun-Hwa; Yeo, Inchul; Han, Jin-Kwan

    2016-03-01

    Phospholipase D (PLD) is involved in the regulation of receptor-associated signaling, cell movement, cell adhesion and endocytosis. However, its physiological role in vertebrate development remains poorly understood. In this study, we show that PLD1 is required for the convergent extension (CE) movements during Xenopus gastrulation by activating Wnt/PCP signaling. Xenopus PLD1 protein is specifically enriched in the dorsal region of Xenopus gastrula embryo and loss or gain-of-function of PLD1 induce defects in gastrulation and CE movements. These defective phenotypes are due to impaired regulation of Wnt/PCP signaling pathway. Biochemical and imaging analysis using Xenopus tissues reveal that PLD1 is required for Fz7 receptor endocytosis upon Wnt11 stimulation. Moreover, we show that Fz7 endocytosis depends on dynamin and regulation of GAP activity of dynamin by PLD1 via its PX domain is crucial for this process. Taken together, our results suggest that PLD1 acts as a new positive mediator of Wnt/PCP signaling by promoting Wnt11-induced Fz7 endocytosis for precise regulation of Xenopus CE movements. PMID:26806705

  9. Wnt7a interaction with Fzd5 and detection of signaling activation using a split eGFP

    SciTech Connect

    Carmon, Kendra S.; Loose, David S.

    2008-04-04

    Wnts are secreted glycoproteins that regulate important cellular processes including proliferation, differentiation, and cell fate. In the {beta}-catenin/canonical pathway, Wnt interacts with Fzd receptors to inhibit degradation of {beta}-catenin and promote its translocation into the nucleus where it regulates transcription of a number of genes. Dysregulation of this pathway has been attributed to a host of diseases including cancer. As a result, components of the {beta}-catenin/canonical pathway have been gaining recognition as promising targets for the discovery of novel therapeutic agents. Here, we show, using an ELISA-based protein-protein binding assay that purified Wnt7a binds to the extracellular cysteine-rich domain of Fzd5 in the nanomolar range. We have developed a novel split eGFP complementation assay to visually detect Wnt7a-Fzd5 interactions and subsequent pathway activation in cells. These biological tools could help lead to a better understanding of Wnt-Fzd interactions and the identification of new modulators of Wnt signaling.

  10. Hypothalamic radial glia function as self-renewing neural progenitors in the absence of Wnt/β-catenin signaling.

    PubMed

    Duncan, Robert N; Xie, Yuanyuan; McPherson, Adam D; Taibi, Andrew V; Bonkowsky, Joshua L; Douglass, Adam D; Dorsky, Richard I

    2016-01-01

    The vertebrate hypothalamus contains persistent radial glia that have been proposed to function as neural progenitors. In zebrafish, a high level of postembryonic hypothalamic neurogenesis has been observed, but the role of radial glia in generating these new neurons is unclear. We have used inducible Cre-mediated lineage labeling to show that a population of hypothalamic radial glia undergoes self-renewal and generates multiple neuronal subtypes at larval stages. Whereas Wnt/β-catenin signaling has been demonstrated to promote the expansion of other stem and progenitor cell populations, we find that Wnt/β-catenin pathway activity inhibits this process in hypothalamic radial glia and is not required for their self-renewal. By contrast, Wnt/β-catenin signaling is required for the differentiation of a specific subset of radial glial neuronal progeny residing along the ventricular surface. We also show that partial genetic ablation of hypothalamic radial glia or their progeny causes a net increase in their proliferation, which is also independent of Wnt/β-catenin signaling. Hypothalamic radial glia in the zebrafish larva thus exhibit several key characteristics of a neural stem cell population, and our data support the idea that Wnt pathway function may not be homogeneous in all stem or progenitor cells. PMID:26603385

  11. Loss of Wnt5a disrupts second heart field cell deployment and may contribute to OFT malformations in DiGeorge syndrome

    PubMed Central

    Sinha, Tanvi; Li, Ding; Théveniau-Ruissy, Magali; Hutson, Mary R.; Kelly, Robert G.; Wang, Jianbo

    2015-01-01

    Outflow tract (OFT) malformation accounts for ∼30% of human congenital heart defects and manifests frequently in TBX1 haplo-insufficiency associated DiGeorge (22q11.2 deletion) syndrome. OFT myocardium originates from second heart field (SHF) progenitors in the pharyngeal and splanchnic mesoderm (SpM), but how these progenitors are deployed to the OFT is unclear. We find that SHF progenitors in the SpM gradually gain epithelial character and are deployed to the OFT as a cohesive sheet. Wnt5a, a non-canonical Wnt, is expressed specifically in the caudal SpM and may regulate oriented cell intercalation to incorporate SHF progenitors into an epithelial-like sheet, thereby generating the pushing force to deploy SHF cells rostrally into the OFT. Using enhancer trap and Cre transgenes, our lineage tracing experiments show that in Wnt5a null mice, SHF progenitors are trapped in the SpM and fail to be deployed to the OFT efficiently, resulting in a reduction in the inferior OFT myocardial wall and its derivative, subpulmonary myocardium. Concomitantly, the superior OFT and subaortic myocardium are expanded. Finally, in chick embryos, blocking the Wnt5a function in the caudal SpM perturbs polarized elongation of SHF progenitors, and compromises their deployment to the OFT. Collectively, our results highlight a critical role for Wnt5a in deploying SHF progenitors from the SpM to the OFT. Given that Wnt5a is a putative transcriptional target of Tbx1, and the similar reduction of subpulmonary myocardium in Tbx1 mutant mice, our results suggest that perturbing Wnt5a-mediated SHF deployment may be an important pathogenic mechanism contributing to OFT malformations in DiGeorge syndrome. PMID:25410658

  12. Self-assembly of high-resolutions PS-b-PMMA block-copolymers: processes capabilities and integration on 300mm track

    NASA Astrophysics Data System (ADS)

    Chevalier, X.; Nicolet, C.; Tiron, R.; Gharbi, A.; Chamiot-Maitral, G.; Jullian, K.; Pimenta-Barros, P.; Argoud, M.; Peyre, J.-L.; Van Spaandonk, R.; Fleury, G.; Hadziioannou, G.; Navarro, C.

    2014-03-01

    Careful control and reproducibility of BCP's synthesis are mandatory parameters to push-down PS-b-PMMA block-copolymer systems toward its lowest dimensions for microelectronic applications. The self-assembly process optimization of different high-resolution cylindrical PS-b-PMMA block-copolymers (i.e. L0 period below 25 nm) is studied to highlight processes-variations as regard to more classical PS-b-PMMA systems while the characterizations of bulk materials provide deeper insights on the parameters addressing the self-assembly of such materials. The integration of a high-resolution BCP on 300 mm track is then studied to check the capabilities of such materials in terms of lithographic applications. CD uniformity measurements in contact hole shrink approach, as well as the transfer of the BCP mask into typical industrial under-layer stacks leading to aggressive features, show that these materials exhibit promising potentials for advanced lithographic nodes.

  13. Blocked randomization with randomly selected block sizes.

    PubMed

    Efird, Jimmy

    2011-01-01

    When planning a randomized clinical trial, careful consideration must be given to how participants are selected for various arms of a study. Selection and accidental bias may occur when participants are not assigned to study groups with equal probability. A simple random allocation scheme is a process by which each participant has equal likelihood of being assigned to treatment versus referent groups. However, by chance an unequal number of individuals may be assigned to each arm of the study and thus decrease the power to detect statistically significant differences between groups. Block randomization is a commonly used technique in clinical trial design to reduce bias and achieve balance in the allocation of participants to treatment arms, especially when the sample size is small. This method increases the probability that each arm will contain an equal number of individuals by sequencing participant assignments by block. Yet still, the allocation process may be predictable, for example, when the investigator is not blind and the block size is fixed. This paper provides an overview of blocked randomization and illustrates how to avoid selection bias by using random block sizes. PMID:21318011

  14. Wnt and FGF mediated epithelial mesenchymal crosstalk during lung development

    PubMed Central

    Volckaert, Thomas; De Langhe, Stijn P.

    2014-01-01

    The adaptation to terrestrial life required the development of an organ capable of efficient air-blood gas exchange. To meet the metabolic load of cellular respiration, the mammalian respiratory system has evolved from a relatively simple structure, similar to the two-tube amphibian lung, to a highly complex tree-like system of branched epithelial airways connected to a vast network of gas exchanging units called alveoli. The development of such an elaborate organ in a relatively short time window is therefore an extraordinary feat and involves an intimate crosstalk between mesodermal and endodermal cell lineages. This review describes the molecular processes governing lung development with an emphasis on the current knowledge on the role of Wnt and Fgf signaling in lung epithelial differentiation. PMID:25470458

  15. The Generation of Organoids for Studying Wnt Signaling.

    PubMed

    Drost, Jarno; Artegiani, Benedetta; Clevers, Hans

    2016-01-01

    We established an in vitro culture model in which intestinal epithelial stem cells can grow into three-dimensional, ever-expanding epithelial organoids that retain their original organ identity and genetic stability. Moreover, organoids can easily be genetically modified using different genome modification strategies, including viral delivery of transgenes and CRISPR/Cas9 technology. These combined characteristics make them a useful in vitro model system to study many biological processes including the contribution of cellular signaling pathways to tissue homeostasis and disease. Here we describe our current laboratory protocols to establish human intestinal organoids and how to genetically modify both mouse and human intestinal organoids to study cellular signaling pathways, specifically Wnt signaling. Moreover, we provide a detailed protocol for lentiviral transduction and CRISPR/Cas9-mediated genome modification of organoid cultures. PMID:27590160

  16. Wnt signaling: the β-cat(enin)'s meow.

    PubMed

    Bauer, Matthieu; Willert, Karl

    2012-01-15

    In a study in the December 15, 2011, issue of Genes & Development, Valenta and colleagues (pp. 2631-2643) constructed a series of β-catenin mutants that allowed them to separate β-catenin's activity as a mediator of Wnt signaling from its activity as cell adhesion component. In doing so, they uncovered some surprising properties of Wnt signaling. PMID:22279043

  17. Role of TAZ as mediator of Wnt signaling.

    PubMed

    Azzolin, Luca; Zanconato, Francesca; Bresolin, Silvia; Forcato, Mattia; Basso, Giuseppe; Bicciato, Silvio; Cordenonsi, Michelangelo; Piccolo, Stefano

    2012-12-21

    Wnt growth factors are fundamental regulators of cell fate, but how the Wnt signal is translated into biological responses is incompletely understood. Here, we report that TAZ, a biologically potent transcriptional coactivator, serves as a downstream element of the Wnt/β-catenin cascade. This function of TAZ is independent from its well-established role as mediator of Hippo signaling. In the absence of Wnt activity, the components of the β-catenin destruction complex--APC, Axin, and GSK3--are also required to keep TAZ at low levels. TAZ degradation depends on phosphorylated β-catenin that bridges TAZ to its ubiquitin ligase β-TrCP. Upon Wnt signaling, escape of β-catenin from the destruction complex impairs TAZ degradation and leads to concomitant accumulation of β-catenin and TAZ. At the genome-wide level, a substantial portion of Wnt transcriptional responses is mediated by TAZ. TAZ activation is a general feature of Wnt signaling and is functionally relevant to mediate Wnt biological effects. PMID:23245942

  18. Sperm Motility Requires Wnt/GSK3 Stabilization of Proteins.

    PubMed

    De Robertis, Edward M; Ploper, Diego

    2015-11-23

    Inhibition of GSK3 by Wnt signaling stabilizes many cellular proteins, but proof that this effect is independent of β-catenin-mediated transcription is lacking. Koch, Acebron, and colleagues (2015) now demonstrate that transcriptionally silent mammalian sperm require Wnt signaling via exosomes to prevent protein degradation during their lengthy travels through the epididymis. PMID:26609954

  19. The sclerostin-neutralizing antibody AbD09097 recognizes an epitope adjacent to sclerostin's binding site for the Wnt co-receptor LRP6

    PubMed Central

    Boschert, V.; Frisch, C.; Back, J. W.; van Pee, K.; Weidauer, S. E.; Muth, E.-M.; Schmieder, P.; Beerbaum, M.; Knappik, A.; Timmerman, P.

    2016-01-01

    The glycoprotein sclerostin has been identified as a negative regulator of bone growth. It exerts its function by interacting with the Wnt co-receptor LRP5/6, blocks the binding of Wnt factors and thereby inhibits Wnt signalling. Neutralizing anti-sclerostin antibodies are able to restore Wnt activity and enhance bone growth thereby presenting a new osteoanabolic therapy approach for diseases such as osteoporosis. We have generated various Fab antibodies against human and murine sclerostin using a phage display set-up. Biochemical analyses have identified one Fab developed against murine sclerostin, AbD09097 that efficiently neutralizes sclerostin's Wnt inhibitory activity. In vitro interaction analysis using sclerostin variants revealed that this neutralizing Fab binds to sclerostin's flexible second loop, which has been shown to harbour the LRP5/6 binding motif. Affinity maturation was then applied to AbD09097, providing a set of improved neutralizing Fab antibodies which particularly bind human sclerostin with enhanced affinity. Determining the crystal structure of AbD09097 provides first insights into how this antibody might recognize and neutralize sclerostin. Together with the structure–function relationship derived from affinity maturation these new data will foster the rational design of new and highly efficient anti-sclerostin antibodies for the therapy of bone loss diseases such as osteoporosis. PMID:27558933

  20. The sclerostin-neutralizing antibody AbD09097 recognizes an epitope adjacent to sclerostin's binding site for the Wnt co-receptor LRP6.

    PubMed

    Boschert, V; Frisch, C; Back, J W; van Pee, K; Weidauer, S E; Muth, E-M; Schmieder, P; Beerbaum, M; Knappik, A; Timmerman, P; Mueller, T D

    2016-08-01

    The glycoprotein sclerostin has been identified as a negative regulator of bone growth. It exerts its function by interacting with the Wnt co-receptor LRP5/6, blocks the binding of Wnt factors and thereby inhibits Wnt signalling. Neutralizing anti-sclerostin antibodies are able to restore Wnt activity and enhance bone growth thereby presenting a new osteoanabolic therapy approach for diseases such as osteoporosis. We have generated various Fab antibodies against human and murine sclerostin using a phage display set-up. Biochemical analyses have identified one Fab developed against murine sclerostin, AbD09097 that efficiently neutralizes sclerostin's Wnt inhibitory activity. In vitro interaction analysis using sclerostin variants revealed that this neutralizing Fab binds to sclerostin's flexible second loop, which has been shown to harbour the LRP5/6 binding motif. Affinity maturation was then applied to AbD09097, providing a set of improved neutralizing Fab antibodies which particularly bind human sclerostin with enhanced affinity. Determining the crystal structure of AbD09097 provides first insights into how this antibody might recognize and neutralize sclerostin. Together with the structure-function relationship derived from affinity maturation these new data will foster the rational design of new and highly efficient anti-sclerostin antibodies for the therapy of bone loss diseases such as osteoporosis. PMID:27558933

  1. Property Blocks: Games and Activities.

    ERIC Educational Resources Information Center

    Humphreys, Alan, Ed.; Dailey, Jean, Ed.

    This pamphlet describes the property blocks produced by MINNEMAST, and discusses their use in the development of thinking processes. Classification systems, including block diagrams and tree diagrams, are discussed. Sixteen classroom activities and eleven games which use the blocks are described. Suggestions to the teacher for further reading are…

  2. Frat is dispensable for canonical Wnt signaling in mammals

    PubMed Central

    van Amerongen, Renée; Nawijn, Martijn; Franca-Koh, Jonathan; Zevenhoven, John; van der Gulden, Hanneke; Jonkers, Jos; Berns, Anton

    2005-01-01

    Wnt-signal transduction through β-catenin is thought to require the inhibition of GSK3 by Frat/GBP. To investigate the role of Frat in mammalian development, we have generated mice with targeted mutations in all three murine Frat homologs. We show that Frat is normally expressed at sites of active Wnt signaling. Surprisingly, Frat-deficient mice do not display gross abnormalities. Moreover, canonical Wnt signaling in primary cells is unaffected by the loss of Frat. These studies show that Frat is not an essential component of the canonical Wnt pathway in higher organisms, despite the strict requirement of Frat/GBP for maternal Wnt signaling in Xenopus. PMID:15681612

  3. Keeping Wnt Signalosome in Check by Vesicular Traffic

    PubMed Central

    FENG, QIANG; GAO, NAN

    2015-01-01

    Wg/Wnts are paracrine and autocrine ligands that activate distinct signaling pathways while being internalized through surface receptors. Converging and contrasting views are shaping our understanding of whether, where, and how endocytosis may modulate Wnt signaling. We gather considerable amount of evidences to elaborate the point that signal-receiving cells utilize distinct, flexible, and sophisticated vesicular trafficking mechanisms to keep Wnt signaling activity in check. Same molecules in a highly context-dependent fashion serve as regulatory hub for various signaling purposes: amplification, maintenance, inhibition, and termination. Updates are provided for the regulatory mechanisms related to the three critical cell surface complexes, Wnt-Fzd-LRP6, Dkk1-Kremen-LRP6, and R-spondin-LGR5-RNF43, which potently influence Wnt signaling. We pay particular attentions to how cells achieve sustained and delicate control of Wnt signaling strength by employing comprehensive aspects of vesicular trafficking. PMID:25336320

  4. Notum deacylates Wnt proteins to suppress signalling activity.

    PubMed

    Kakugawa, Satoshi; Langton, Paul F; Zebisch, Matthias; Howell, Steven A; Chang, Tao-Hsin; Liu, Yan; Feizi, Ten; Bineva, Ganka; O'Reilly, Nicola; Snijders, Ambrosius P; Jones, E Yvonne; Vincent, Jean-Paul

    2015-03-12

    Signalling by Wnt proteins is finely balanced to ensure normal development and tissue homeostasis while avoiding diseases such as cancer. This is achieved in part by Notum, a highly conserved secreted feedback antagonist. Notum has been thought to act as a phospholipase, shedding glypicans and associated Wnt proteins from the cell surface. However, this view fails to explain specificity, as glypicans bind many extracellular ligands. Here we provide genetic evidence in Drosophila that Notum requires glypicans to suppress Wnt signalling, but does not cleave their glycophosphatidylinositol anchor. Structural analyses reveal glycosaminoglycan binding sites on Notum, which probably help Notum to co-localize with Wnt proteins. They also identify, at the active site of human and Drosophila Notum, a large hydrophobic pocket that accommodates palmitoleate. Kinetic and mass spectrometric analyses of human proteins show that Notum is a carboxylesterase that removes an essential palmitoleate moiety from Wnt proteins and thus constitutes the first known extracellular protein deacylase. PMID:25731175

  5. Methods to Manipulate and Monitor Wnt Signaling in Human Pluripotent Stem Cells.

    PubMed

    Huggins, Ian J; Brafman, David; Willert, Karl

    2016-01-01

    Human pluripotent stem cells (hPSCs) may revolutionize medical practice by providing: (a) a renewable source of cells for tissue replacement therapies, (b) a powerful system to model human diseases in a dish, and (c) a platform for examining efficacy and safety of novel drugs. Furthermore, these cells offer a unique opportunity to study early human development in vitro, in particular, the process by which a seemingly uniform cell population interacts to give rise to the three main embryonic lineages: ectoderm, endoderm. and mesoderm. This process of lineage allocation is regulated by a number of inductive signals that are mediated by growth factors, including FGF, TGFβ, and Wnt. In this book chapter, we introduce a set of tools, methods, and protocols to specifically manipulate the Wnt signaling pathway with the intention of altering the cell fate outcome of hPSCs. PMID:27590161

  6. Prednisolone induces the Wnt signalling pathway in 3T3-L1 adipocytes

    PubMed Central

    Fleuren, Wilco W. M.; Linssen, Margot M. L.; Toonen, Erik J. M.; van der Zon, Gerard C. M.; Guigas, Bruno; de Vlieg, Jacob; Dokter, Wim H. A.; Ouwens, D. Margriet

    2013-01-01

    Synthetic glucocorticoids are potent anti-inflammatory drugs but show dose-dependent metabolic side effects such as the development of insulin resistance and obesity. The precise mechanisms involved in these glucocorticoid-induced side effects, and especially the participation of adipose tissue in this are not completely understood. We used a combination of transcriptomics, antibody arrays and bioinformatics approaches to characterize prednisolone-induced alterations in gene expression and adipokine secretion, which could underlie metabolic dysfunction in 3T3-L1 adipocytes. Several pathways, including cytokine signalling, Akt signalling, and Wnt signalling were found to be regulated at multiple levels, showing that these processes are targeted by prednisolone. These results suggest that mechanisms by which prednisolone induce insulin resistance include dysregulation of wnt signalling and immune response processes. These pathways may provide interesting targets for the development of improved glucocorticoids. PMID:23506355

  7. Inhibition of Melanogenesis by the Pyridinyl Imidazole Class of Compounds: Possible Involvement of the Wnt/β-Catenin Signaling Pathway

    PubMed Central

    Bellei, Barbara; Pitisci, Angela; Izzo, Enzo; Picardo, Mauro

    2012-01-01

    While investigating the role of p38 MAPK in regulating melanogenesis, we found that pyridinyl imidazole inhibitors class compounds as well as the analog compound SB202474, which does not inhibit p38 MAPK, suppressed both α-MSH-induced melanogenesis and spontaneous melanin synthesis. In this study, we demonstrated that the inhibitory activity of the pyridinyl imidazoles correlates with inhibition of the canonical Wnt/β-catenin pathway activity. Imidazole-treated cells showed a reduction in the level of Tcf/Lef target genes involved in the β-catenin signaling network, including ubiquitous genes such as Axin2, Lef1, and Wisp1 as well as cell lineage-restricted genes such as microphthalmia-associated transcription factor and dopachrome tautomerase. Although over-expression of the Wnt signaling pathway effector β-catenin slightly restored the melanogenic program, the lack of complete reversion suggested that the imidazoles interfered with β-catenin-dependent transcriptional activity rather than with β-catenin expression. Accordingly, we did not observe any significant change in β-catenin protein expression. The independence of p38 MAPK activity from the repression of Wnt/β-catenin signaling pathway was confirmed by small interfering RNA knockdown of p38 MAPK expression, which by contrast, stimulated β-catenin-driven gene expression. Our data demonstrate that the small molecule pyridinyl imidazoles possess two distinct and opposite mechanisms that modulate β-catenin dependent transcription: a p38 inhibition-dependent effect that stimulates the Wnt pathway by increasing β-catenin protein expression and an off-target mechanism that inhibits the pathway by repressing β-catenin protein functionality. The p38-independent effect seems to be dominant and, at least in B16-F0 cells, results in a strong block of the Wnt/β-catenin signaling pathway. PMID:22427932

  8. Wnt3a regulates proliferation and migration of HUVEC via canonical and non-canonical Wnt signaling pathways

    SciTech Connect

    Samarzija, Ivana; Sini, Patrizia; Schlange, Thomas; MacDonald, Gwen; Hynes, Nancy E.

    2009-08-28

    Untangling the signaling pathways involved in endothelial cell biology is of central interest for the development of antiangiogenesis based therapies. Here we report that Wnt3a induces the proliferation and migration of HUVECs, but does not affect their survival. Wnt3a-induced proliferation was VEGFR signaling independent, but reduced upon CamKII inhibition. In a search for the downstream mediators of Wnt3a's effects on HUVEC biology, we found that Wnt3a treatment leads to phosphorylation of DVL3 and stabilization of {beta}-catenin. Moreover, under the same conditions we observed an upregulation in c-MYC, TIE-2 and GJA1 mRNA transcripts. Although treatment of HUVECs with Wnt5a induced DVL3 phosphorylation, we did not observe any of the other effects seen upon Wnt3a stimulation. Taken together, our data indicate that Wnt3a induces canonical and non-canonical Wnt signaling in HUVECs, and stimulates their proliferation and migration.

  9. IGF-1R inhibition in mammary epithelia promotes canonical Wnt signaling and Wnt1-driven tumors

    PubMed Central

    Rota, Lauren M.; Albanito, Lidia; Shin, Marcus E.; Goyeneche, Corey L.; Shushanov, Sain; Gallagher, Emily J.; LeRoith, Derek; Lazzarino, Deborah A.; Wood, Teresa L.

    2014-01-01

    Triple-negative breast cancers (TNBC) are an aggressive disease subtype which unlike other subtypes lack an effective targeted therapy. Inhibitors of the insullin-like growth factor receptor (IGF-1R) have been considered for use in treating TNBC. Here we provide genetic evidence that IGF-1R inhibition promotes development of Wnt1-mediated murine mammary tumors that offer a model of TNBC. We found that in a double transgenic mouse model carrying activated Wnt-1 and mutant IGF-1R, a reduction in IGF-1R signaling reduced tumor latency and promoted more aggressive phenotypes. These tumors displayed a squamal cell phenotype with increased expression of keratins 5/6 and β-catenin. Notably, cell lineage analyses revealed an increase in basal (CD29hi/CD24+) and luminal (CD24+/CD61+/CD29lo) progenitor cell populations, along with increased Nanog expression and decreased Elf5 expression. In these doubly transgenic mice, lung metastases developed with characteristics of the primary tumors, unlike MMTV-Wnt1 mice. Mechanistic investigations showed that pharmacological inhibition of the IGF-1R in vitro was sufficient to increase the tumorsphere-forming efficiency of MMTV-Wnt1 tumor cells. Tumors from doubly transgenic mice also exhibited an increase in the expression ratio of the IGF-II-sensitive, A isoform of the insulin receptor vs the IR-B isoform, which in vitro resulted in enhanced expression of β-catenin. Overall, our results revealed that in Wnt-driven tumors an attenuation of IGF-1R signaling accelerates tumorigenesis and promotes more aggressive phenotypes, with potential implications for understanding TNBC pathobiology and treatment. PMID:25092896

  10. p38 MAPK regulates the Wnt inhibitor Dickkopf-1 in osteotropic prostate cancer cells

    PubMed Central

    Browne, A J; Göbel, A; Thiele, S; Hofbauer, L C; Rauner, M; Rachner, T D

    2016-01-01

    The Wnt inhibitor Dickkopf-1 (DKK-1) has been associated with the occurrence of bone metastases in osteotropic prostate cancer by inhibiting osteoblastogenesis. P38 mitogen-activated protein kinase (MAPK) activity is also dysregulated in advanced prostate cancer. However, the impact of p38 MAPK signaling on DKK-1 remains unknown. Inhibition of p38 MAPK signaling in osteolytic PC3 cells by small molecule inhibitors (doramapimod, LY2228820 and SB202190) suppressed DKK-1 expression, whereas activation of p38 MAPK by anisomycin increased DKK-1. Further dissection by targeting individual p38 MAPK isoforms with siRNA revealed a stronger role for MAPK11 than MAPK14 and MAPK12 in the regulation of DKK-1. Moreover, prostate cancer cells with a predominantly osteolytic phenotype produced sufficient amounts of DKK-1 to inhibit Wnt3a-induced osteoblastic differentiation in C2C12 cells. This inhibition was blocked directly by neutralizing DKK-1 using a specific antibody and also indirectly by blocking p38 MAPK. Furthermore, tissue expression in human prostate cancer revealed a correlation between p38 MAPK and DKK-1 expression with higher expression in tumor compared with normal tissues. These results reveal that p38 MAPK regulates DKK-1 in prostate cancer and may present a potential target in osteolytic prostate cancers. PMID:26913608

  11. Sporadic Nucleation and Growth in the Microphase Separation Process of an I2S Miktoarm Star Block Copolymer and its Blends with Homopolymer

    NASA Astrophysics Data System (ADS)

    Yang, Lizhang; Pochan, Darrin J.; Gido, Samuel P.; Pispas, Stergios; Hong, Kunlun; Mays, Jimmy W.

    2000-03-01

    A selective solvent and annealing study was done to investigate the morphology behavior of an I2S miktoarm star block copolymer and its blends with homopolyisoprene. Casting from cyclohexane, a selective solvent for polyisoprene, the neat star shaped I2S block copolymer only partially microphase separated, and formed a unique layered morphology inside a homogeneous media. During annealing, the layered phase and the homogeneous phase both transformed into a randomly oriented worm morphology. The path of this transformation is different depending on whether the starting state is layered or homogenous. The I2S/homopolyisoprene blend formed a mostly homogeneous phase after casting from cyclohexane. Annealing produced slow microphase separation which was observed at various stages by TEM. Based on these observations, a sporadic nucleation process of microphase separation is proposed.

  12. Endpoint-based parallel data processing with non-blocking collective instructions in a parallel active messaging interface of a parallel computer

    DOEpatents

    Archer, Charles J; Blocksome, Michael A; Cernohous, Bob R; Ratterman, Joseph D; Smith, Brian E

    2014-11-18

    Methods, apparatuses, and computer program products for endpoint-based parallel data processing with non-blocking collective instructions in a parallel active messaging interface (`PAMI`) of a parallel computer are provided. Embodiments include establishing by a parallel application a data communications geometry, the geometry specifying a set of endpoints that are used in collective operations of the PAMI, including associating with the geometry a list of collective algorithms valid for use with the endpoints of the geometry. Embodiments also include registering in each endpoint in the geometry a dispatch callback function for a collective operation and executing without blocking, through a single one of the endpoints in the geometry, an instruction for the collective operation.

  13. Endpoint-based parallel data processing with non-blocking collective instructions in a parallel active messaging interface of a parallel computer

    DOEpatents

    Archer, Charles J; Blocksome, Michael A; Cernohous, Bob R; Ratterman, Joseph D; Smith, Brian E

    2014-11-11

    Endpoint-based parallel data processing with non-blocking collective instructions in a PAMI of a parallel computer is disclosed. The PAMI is composed of data communications endpoints, each including a specification of data communications parameters for a thread of execution on a compute node, including specifications of a client, a context, and a task. The compute nodes are coupled for data communications through the PAMI. The parallel application establishes a data communications geometry specifying a set of endpoints that are used in collective operations of the PAMI by associating with the geometry a list of collective algorithms valid for use with the endpoints of the geometry; registering in each endpoint in the geometry a dispatch callback function for a collective operation; and executing without blocking, through a single one of the endpoints in the geometry, an instruction for the collective operation.

  14. Role of Sulf1A in Wnt1- and Wnt6-induced growth regulation and myoblast hyper-elongation.

    PubMed

    Hitchins, L; Fletcher, F; Allen, S; Dhoot, G K

    2013-01-01

    Sulf1A expression, which is a characteristic of embryonic muscle, is undetectable in mature muscle fibres and quiescent satellite cells, but is re-activated in vivo upon injury and in vitro following activation of satellite cells. Sulf1A is known to enhance canonical Wnt signalling, and its association with Wnt1-induced satellite cell proliferation in vitro in the present study further confirmed this. However, exogenous Wnt6 decreased satellite cell proliferation but promoted the adoption of a hyper-elongated cell morphology in myoblasts on isolated single fibres in culture. Such Wnt6-induced cellular hyper-elongation and inhibition of proliferation was found to be dependent upon Sulf1A, as treatment with Sulf1A neutralising antibodies abolished both these effects. This indicates that Sulf1A can regulate Wnt6 signalling and cellular differentiation in skeletal muscle. PMID:23772371

  15. Identification of Wnt Pathway Target Genes Regulating the Division and Differentiation of Larval Seam Cells and Vulval Precursor Cells in Caenorhabditis elegans

    PubMed Central

    Gorrepati, Lakshmi; Krause, Michael W.; Chen, Weiping; Brodigan, Thomas M.; Correa-Mendez, Margarita; Eisenmann, David M.

    2015-01-01

    The evolutionarily conserved Wnt/β-catenin signaling pathway plays a fundamental role during metazoan development, regulating numerous processes including cell fate specification, cell migration, and stem cell renewal. Wnt ligand binding leads to stabilization of the transcriptional effector β-catenin and upregulation of target gene expression to mediate a cellular response. During larval development of the nematode Caenorhabditis elegans, Wnt/β-catenin pathways act in fate specification of two hypodermal cell types, the ventral vulval precursor cells (VPCs) and the lateral seam cells. Because little is known about targets of the Wnt signaling pathways acting during larval VPC and seam cell differentiation, we sought to identify genes regulated by Wnt signaling in these two hypodermal cell types. We conditionally activated Wnt signaling in larval animals and performed cell type–specific "mRNA tagging" to enrich for VPC and seam cell–specific mRNAs, and then used microarray analysis to examine gene expression compared to control animals. Two hundred thirty-nine genes activated in response to Wnt signaling were identified, and we characterized 50 genes further. The majority of these genes are expressed in seam and/or vulval lineages during normal development, and reduction of function for nine genes caused defects in the proper division, fate specification, fate execution, or differentiation of seam cells and vulval cells. Therefore, the combination of these techniques was successful at identifying potential cell type–specific Wnt pathway target genes from a small number of cells and at increasing our knowledge of the specification and behavior of these C. elegans larval hypodermal cells. PMID:26048561

  16. The effect of different component ratios in block polymers and processing conditions on electrodeposition efficiency onto titanium

    NASA Astrophysics Data System (ADS)

    Fukuhara, Yusuke; Kyuzo, Megumi; Tsutsumi, Yusuke; Nagai, Akiko; Chen, Peng; Hanawa, Takao

    2015-11-01

    2-Methacryloyloxyethyl phosphorylcholine (MPC) polymers for electrodeposition to titanium surfaces were synthesized. The polymers were block-type copolymers composed of a poly(MPC) segment and a poly(2-aminoethylmethacrylate (AEMA)) segment, which could electronically adsorb to a titanium oxide film on the titanium surface. The polymer was synthesized as expected by nuclear magnetic resonance and gel permeation chromatography. In a 0.26 mmol L-1 PMbA solution adjusted to pH 11, -3.0 V (vs. an Ag/AgCl electrode) was applied to a titanium substrate for 300 s. We evaluated the effects of the molecular structure of poly(MPC-block-AEMA) (PMbA) with a different polymerization degree of MPC unit, whereas the polymerization degree of the AEMA units was fixed. The 15-min electrodeposition of PMbA100 was the most efficient condition in this study. On the other hand, the results of the water contact angle and the amount of adsorbed protein did not change, even when altering the MPC unit number and electrodeposition time. This indicates that the immobilization by electrodeposition of PMbA is important for the inhibition of protein adsorption, while the polymerization degree of the MPC unit and the electrodeposition time do not influence them. This study will enhance the understanding of effective polymer structures for electrodeposition and electrodeposition conditions.

  17. R-spondin 2 facilitates differentiation of proliferating chondrocytes into hypertrophic chondrocytes by enhancing Wnt/β-catenin signaling in endochondral ossification.

    PubMed

    Takegami, Yasuhiko; Ohkawara, Bisei; Ito, Mikako; Masuda, Akio; Nakashima, Hiroaki; Ishiguro, Naoki; Ohno, Kinji

    2016-04-22

    Endochondral ossification is a crucial process for longitudinal growth of bones. Differentiating chondrocytes in growth cartilage form four sequential zones of proliferation, alignment into column, hypertrophy, and substitution of chondrocytes with osteoblasts. Wnt/β-catenin signaling is essential for differentiation of proliferating chondrocytes into hypertrophic chondrocytes in growth cartilage. R-spondin 2 (Rspo2), a member of R-spondin family, is an agonist for Wnt signaling, but its role in chondrocyte differentiation remains unknown. Here we report that growth cartilage of Rspo2-knockout mice shows a decreased amount of β-catenin and increased amounts collagen type II (CII) and Sox9 in the abnormally extended proliferating zone. In contrast, expression of collagen type X (CX) in the hypertrophic zone remains unchanged. Differentiating chondrogenic ATDC5 cells, mimicking proliferating chondrocytes, upregulate Rspo2 and its putative receptor, Lgr5, in parallel. Addition of recombinant human Rspo2 to differentiating ATDC5 cells decreases expressions of Col2a1, Sox9, and Acan, as well as production of proteoglycans. In contrast, lentivirus-mediated knockdown of Rspo2 has the opposite effect. The effect of Rspo2 on chondrogenic differentiation is mediated by Wnt/β-catenin signaling, and not by Wnt/PCP or Wnt/Ca(2+) signaling. We propose that Rspo2 activates Wnt/β-catenin signaling to reduce Col2a1 and Sox9 and to facilitate differentiation of proliferating chondrocytes into hypertrophic chondrocytes in growth cartilage. PMID:27012200

  18. The low-density lipoprotein receptor-related protein 10 is a negative regulator of the canonical Wnt/{beta}-catenin signaling pathway

    SciTech Connect

    Jeong, Young-Hee; Sekiya, Manami; Hirata, Michiko; Ye, Mingjuan; Yamagishi, Azumi; Lee, Sang-Mi; Kang, Man-Jong; Hosoda, Akemi; Fukumura, Tomoe; Kim, Dong-Ho; Saeki, Shigeru

    2010-02-19

    Wnt signaling pathways play fundamental roles in the differentiation, proliferation and functions of many cells as well as developmental, growth, and homeostatic processes in animals. Low-density lipoprotein receptor (LDLR)-related protein (LRP) 5 and LRP6 serve as coreceptors of Wnt proteins together with Frizzled receptors, triggering activation of canonical Wnt/{beta}-catenin signaling. Here, we found that LRP10, a new member of the LDLR gene family, inhibits the canonical Wnt/{beta}-catenin signaling pathway. The {beta}-catenin/T cell factor (TCF) transcriptional activity in HEK293 cells was activated by transfection with Wnt3a or LRP6, which was then inhibited by co-transfection with LRP10. Deletion of the extracellular domain of LRP10 negated its inhibitory effect. The inhibitory effect of LRP10 was consistently conserved in HEK293 cells even when GSK3{beta} phosphorylation was inhibited by incubation with lithium chloride and co-transfection with constitutively active S33Y-mutated {beta}-catenin. Nuclear {beta}-catenin accumulation was unaffected by LRP10. The present studies suggest that LRP10 may interfere with the formation of the {beta}-catenin/TCF complex and/or its binding to target DNA in the nucleus, and that the extracellular domain of LRP10 is critical for inhibition of the canonical Wnt/{beta}-catenin signaling pathway.

  19. The PTK7-Related Transmembrane Proteins Off-track and Off-track 2 Are Co-receptors for Drosophila Wnt2 Required for Male Fertility

    PubMed Central

    Honemann-Capito, Mona; Brechtel-Curth, Katja; Hedderich, Marie; Wodarz, Andreas

    2014-01-01

    Wnt proteins regulate many developmental processes and are required for tissue homeostasis in adult animals. The cellular responses to Wnts are manifold and are determined by the respective Wnt ligand and its specific receptor complex in the plasma membrane. Wnt receptor complexes contain a member of the Frizzled family of serpentine receptors and a co-receptor, which commonly is a single-pass transmembrane protein. Vertebrate protein tyrosine kinase 7 (PTK7) was identified as a Wnt co-receptor required for control of planar cell polarity (PCP) in frogs and mice. We found that flies homozygous for a complete knock-out of the Drosophila PTK7 homolog off track (otk) are viable and fertile and do not show PCP phenotypes. We discovered an otk paralog (otk2, CG8964), which is co-expressed with otk throughout embryonic and larval development. Otk and Otk2 bind to each other and form complexes with Frizzled, Frizzled2 and Wnt2, pointing to a function as Wnt co-receptors. Flies lacking both otk and otk2 are viable but male sterile due to defective morphogenesis of the ejaculatory duct. Overexpression of Otk causes female sterility due to malformation of the oviduct, indicating that Otk and Otk2 are specifically involved in the sexually dimorphic development of the genital tract. PMID:25010066

  20. The bone-sparing effects of estrogen and WNT16 are independent of each other

    PubMed Central

    Movérare-Skrtic, Sofia; Wu, Jianyao; Henning, Petra; Gustafsson, Karin L.; Sjögren, Klara; Windahl, Sara H.; Koskela, Antti; Tuukkanen, Juha; Börjesson, Anna E.; Lagerquist, Marie K.; Lerner, Ulf H.; Zhang, Fu-Ping; Gustafsson, Jan-Åke; Poutanen, Matti; Ohlsson, Claes

    2015-01-01

    Wingless-type MMTV integration site family (WNT)16 is a key regulator of bone mass with high expression in cortical bone, and Wnt16−/− mice have reduced cortical bone mass. As Wnt16 expression is enhanced by estradiol treatment, we hypothesized that the bone-sparing effect of estrogen in females is WNT16-dependent. This hypothesis was tested in mechanistic studies using two genetically modified mouse models with either constantly high osteoblastic Wnt16 expression or no Wnt16 expression. We developed a mouse model with osteoblast-specific Wnt16 overexpression (Obl-Wnt16). These mice had several-fold elevated Wnt16 expression in both trabecular and cortical bone compared with wild type (WT) mice. Obl-Wnt16 mice displayed increased total body bone mineral density (BMD), surprisingly caused mainly by a substantial increase in trabecular bone mass, resulting in improved bone strength of vertebrae L3. Ovariectomy (ovx) reduced the total body BMD and the trabecular bone mass to the same degree in Obl-Wnt16 mice and WT mice, suggesting that the bone-sparing effect of estrogen is WNT16-independent. However, these bone parameters were similar in ovx Obl-Wnt16 mice and sham operated WT mice. The role of WNT16 for the bone-sparing effect of estrogen was also evaluated in Wnt16−/− mice. Treatment with estradiol increased the trabecular and cortical bone mass to a similar extent in both Wnt16−/− and WT mice. In conclusion, the bone-sparing effects of estrogen and WNT16 are independent of each other. Furthermore, loss of endogenous WNT16 results specifically in cortical bone loss, whereas overexpression of WNT16 surprisingly increases mainly trabecular bone mass. WNT16-targeted therapies might be useful for treatment of postmenopausal trabecular bone loss. PMID:26627248

  1. The bone-sparing effects of estrogen and WNT16 are independent of each other.

    PubMed

    Movérare-Skrtic, Sofia; Wu, Jianyao; Henning, Petra; Gustafsson, Karin L; Sjögren, Klara; Windahl, Sara H; Koskela, Antti; Tuukkanen, Juha; Börjesson, Anna E; Lagerquist, Marie K; Lerner, Ulf H; Zhang, Fu-Ping; Gustafsson, Jan-Åke; Poutanen, Matti; Ohlsson, Claes

    2015-12-01

    Wingless-type MMTV integration site family (WNT)16 is a key regulator of bone mass with high expression in cortical bone, and Wnt16(-/-) mice have reduced cortical bone mass. As Wnt16 expression is enhanced by estradiol treatment, we hypothesized that the bone-sparing effect of estrogen in females is WNT16-dependent. This hypothesis was tested in mechanistic studies using two genetically modified mouse models with either constantly high osteoblastic Wnt16 expression or no Wnt16 expression. We developed a mouse model with osteoblast-specific Wnt16 overexpression (Obl-Wnt16). These mice had several-fold elevated Wnt16 expression in both trabecular and cortical bone compared with wild type (WT) mice. Obl-Wnt16 mice displayed increased total body bone mineral density (BMD), surprisingly caused mainly by a substantial increase in trabecular bone mass, resulting in improved bone strength of vertebrae L3. Ovariectomy (ovx) reduced the total body BMD and the trabecular bone mass to the same degree in Obl-Wnt16 mice and WT mice, suggesting that the bone-sparing effect of estrogen is WNT16-independent. However, these bone parameters were similar in ovx Obl-Wnt16 mice and sham operated WT mice. The role of WNT16 for the bone-sparing effect of estrogen was also evaluated in Wnt16(-/-) mice. Treatment with estradiol increased the trabecular and cortical bone mass to a similar extent in both Wnt16(-/-) and WT mice. In conclusion, the bone-sparing effects of estrogen and WNT16 are independent of each other. Furthermore, loss of endogenous WNT16 results specifically in cortical bone loss, whereas overexpression of WNT16 surprisingly increases mainly trabecular bone mass. WNT16-targeted therapies might be useful for treatment of postmenopausal trabecular bone loss. PMID:26627248

  2. lncRNA DANCR suppresses odontoblast-like differentiation of human dental pulp cells by inhibiting wnt/β-catenin pathway.

    PubMed

    Chen, Lingling; Song, Zhi; Huang, Shuheng; Wang, Runfu; Qin, Wei; Guo, Jia; Lin, Zhengmei

    2016-05-01

    Long noncoding RNAs (lncRNAs) have recently emerged as an important class of regulatory molecules in diverse biological processes, although lncRNA involvement in the odontoblast-like differentiation of human dental pulp cells (hDPCs) is poorly understood. We investigate the expression of lncRNAs in this differentiation and explore their underlying role and the involved mechanism. Integrated comparative lncRNA microarray profiling was used to examine lncRNA expression during this differentiation. The differential expression of lncRNAs was validated by quantitative real-time reverse transcription plus the polymerase chain reaction. Differential lncRNA overexpression was performed with an adenoviral vector and the role and mechanism was then investigated in odontoblast-like differentiation. We identified 139 differentially expressed lncRNAs during this differentiation. Among them, five lncRNAs differentially expressed in microarray analysis were validated. Notably, lncRNA DANCR expression was significantly downregulated during hDPC differentiation to odontoblast-like cells in a time-dependent manner. Moreover, lncRNA DANCR overexpression blocked mineralized nodule formation and the expression of DSPP and DMP-1 in hDPCs after 14 days of odontogenic induction. Importantly, the upregulation of DANCR significantly decreased the expression levels of p-GSK-3β and β-catenin expression indicating that lncRNA DANCR can inhibit the activation of the Wnt/β-catenin signal pathway during the odontoblast-like differentiation of hDPCs. Thus, the modulation of Wnt/β-catenin signaling by lncRNA DANCR represents a potential therapeutic option for reparative dentin formation and regenerative endodontics. PMID:26646542

  3. Extracellular matrix stiffness dictates Wnt expression through integrin pathway

    PubMed Central

    Du, Jing; Zu, Yan; Li, Jing; Du, Shuyuan; Xu, Yipu; Zhang, Lang; Jiang, Li; Wang, Zhao; Chien, Shu; Yang, Chun

    2016-01-01

    It is well established that extracellular matrix (ECM) stiffness plays a significant role in regulating the phenotypes and behaviors of many cell types. However, the mechanism underlying the sensing of mechanical cues and subsequent elasticity-triggered pathways remains largely unknown. We observed that stiff ECM significantly enhanced the expression level of several members of the Wnt/β-catenin pathway in both bone marrow mesenchymal stem cells and primary chondrocytes. The activation of β-catenin by stiff ECM is not dependent on Wnt signals but is elevated by the activation of integrin/ focal adhesion kinase (FAK) pathway. The accumulated β-catenin then bound to the wnt1 promoter region to up-regulate the gene transcription, thus constituting a positive feedback of the Wnt/β-catenin pathway. With the amplifying effect of positive feedback, this integrin-activated β-catenin/Wnt pathway plays significant roles in mediating the enhancement of Wnt signal on stiff ECM and contributes to the regulation of mesenchymal stem cell differentiation and primary chondrocyte phenotype maintenance. The present integrin-regulated Wnt1 expression and signaling contributes to the understanding of the molecular mechanisms underlying the regulation of cell behaviors by ECM elasticity. PMID:26854061

  4. SOX9 drives WNT pathway activation in prostate cancer

    PubMed Central

    Ma, Fen; Ye, Huihui; He, Housheng Hansen; Gerrin, Sean J.; Chen, Sen; Tanenbaum, Benjamin A.; Sowalsky, Adam G.; He, Lingfeng; Wang, Hongyun; Balk, Steven P.; Yuan, Xin

    2016-01-01

    The transcription factor SOX9 is critical for prostate development, and dysregulation of SOX9 is implicated in prostate cancer (PCa). However, the SOX9-dependent genes and pathways involved in both normal and neoplastic prostate epithelium are largely unknown. Here, we performed SOX9 ChIP sequencing analysis and transcriptome profiling of PCa cells and determined that SOX9 positively regulates multiple WNT pathway genes, including those encoding WNT receptors (frizzled [FZD] and lipoprotein receptor-related protein [LRP] family members) and the downstream β-catenin effector TCF4. Analyses of PCa xenografts and clinical samples both revealed an association between the expression of SOX9 and WNT pathway components in PCa. Finally, treatment of SOX9-expressing PCa cells with a WNT synthesis inhibitor (LGK974) reduced WNT pathway signaling in vitro and tumor growth in murine xenograft models. Together, our data indicate that SOX9 expression drives PCa by reactivating the WNT/β−catenin signaling that mediates ductal morphogenesis in fetal prostate and define a subgroup of patients who would benefit from WNT-targeted therapy. PMID:27043282

  5. Canonical Wnt Signaling Regulates Atrioventricular Junction Programming and Electrophysiological Properties

    PubMed Central

    Gillers, Benjamin S; Chiplunkar, Aditi; Aly, Haytham; Valenta, Tomas; Basler, Konrad; Christoffels, Vincent M.; Efimov, Igor R; Boukens, Bastiaan J; Rentschler, Stacey

    2014-01-01

    Rationale Proper patterning of the atrioventricular canal (AVC) is essential for delay of electrical impulses between atria and ventricles, and defects in AVC maturation can result in congenital heart disease. Objective To determine the role of canonical Wnt signaling in the myocardium during AVC development. Methods and Results We utilized a novel allele of β-catenin that preserves β-catenin’s cell adhesive functions but disrupts canonical Wnt signaling, allowing us to probe the effects of Wnt loss of function independently. We show that loss of canonical Wnt signaling in the myocardium results in tricuspid atresia with hypoplastic right ventricle associated with loss of AVC myocardium. In contrast, ectopic activation of Wnt signaling was sufficient to induce formation of ectopic AV junction-like tissue as assessed by morphology, gene expression, and electrophysiologic criteria. Aberrant AVC development can lead to ventricular preexcitation, a characteristic feature of Wolff-Parkinson-White syndrome. We demonstrate that postnatal activation of Notch signaling downregulates canonical Wnt targets within the AV junction. Stabilization of β-catenin protein levels can rescue Notch-mediated ventricular preexcitation and dysregulated ion channel gene expression. Conclusions Our data demonstrate that myocardial canonical Wnt signaling is an important regulator of AVC maturation and electrical programming upstream of Tbx3. Our data further suggests that ventricular preexcitation may require both morphologic patterning defects, as well as myocardial lineage reprogramming, to allow robust conduction across accessory pathway tissue. PMID:25599332

  6. Extracellular matrix stiffness dictates Wnt expression through integrin pathway.

    PubMed

    Du, Jing; Zu, Yan; Li, Jing; Du, Shuyuan; Xu, Yipu; Zhang, Lang; Jiang, Li; Wang, Zhao; Chien, Shu; Yang, Chun

    2016-01-01

    It is well established that extracellular matrix (ECM) stiffness plays a significant role in regulating the phenotypes and behaviors of many cell types. However, the mechanism underlying the sensing of mechanical cues and subsequent elasticity-triggered pathways remains largely unknown. We observed that stiff ECM significantly enhanced the expression level of several members of the Wnt/β-catenin pathway in both bone marrow mesenchymal stem cells and primary chondrocytes. The activation of β-catenin by stiff ECM is not dependent on Wnt signals but is elevated by the activation of integrin/ focal adhesion kinase (FAK) pathway. The accumulated β-catenin then bound to the wnt1 promoter region to up-regulate the gene transcription, thus constituting a positive feedback of the Wnt/β-catenin pathway. With the amplifying effect of positive feedback, this integrin-activated β-catenin/Wnt pathway plays significant roles in mediating the enhancement of Wnt signal on stiff ECM and contributes to the regulation of mesenchymal stem cell differentiation and primary chondrocyte phenotype maintenance. The present integrin-regulated Wnt1 expression and signaling contributes to the understanding of the molecular mechanisms underlying the regulation of cell behaviors by ECM elasticity. PMID:26854061

  7. Targets of Wnt/ß-Catenin Transcription in Penile Carcinoma

    PubMed Central

    Henrique, Rui; Millar, Michael; Hamblin, Ruth; Davda, Reena; Aare, Kristina; Masters, John R.; Thomson, Calum; Muneer, Asif; Patel, Hitendra R. H.; Ahmed, Aamir

    2015-01-01

    Penile squamous cell carcinoma (PeCa) is a rare malignancy and little is known regarding the molecular mechanisms involved in carcinogenesis of PeCa. The Wnt signaling pathway, with the transcription activator ß-catenin as a major transducer, is a key cellular pathway during development and in disease, particularly cancer. We have used PeCa tissue arrays and multi-fluorophore labelled, quantitative, immunohistochemistry to interrogate the expression of WNT4, a Wnt ligand, and three targets of Wnt-ß-catenin transcription activation, namely, MMP7, cyclinD1 (CD1) and c-MYC in 141 penile tissue cores from 101 unique samples. The expression of all Wnt signaling proteins tested was increased by 1.6 to 3 fold in PeCa samples compared to control tissue (normal or cancer adjacent) samples (p<0.01). Expression of all proteins, except CD1, showed a significant decrease in grade II compared to grade I tumors. High magnification, deconvolved confocal images were used to measure differences in co-localization between the four proteins. Significant (p<0.04-0.0001) differences were observed for various permutations of the combinations of proteins and state of the tissue (control, tumor grades I and II). Wnt signaling may play an important role in PeCa and proteins of the Wnt signaling network could be useful targets for diagnosis and prognostic stratification of disease. PMID:25901368

  8. RhoA Controls Wnt Upregulation on Microstructured Titanium Surfaces

    PubMed Central

    Mazzotta, Silvia; Piergianni, Maddalena; Piemontese, Marilina; Passeri, Giovanni

    2014-01-01

    Rough topography enhances the activation of Wnt canonical signaling in vitro, and this mediates its effects on cell differentiation. However, the molecular mechanisms underlying topography-dependent control of Wnt signaling are still poorly understood. As the small GTPase RhoA controls cytoskeletal reorganization and actomyosin-induced tensional forces, we hypothesized that RhoA could affect the activation of Wnt signaling in cells on micropatterned titanium surfaces. G-LISA assay revealed that RhoA activation was higher in C2C12 cells on rough (SLA) surfaces under basal conditions than on smooth (Polished) titanium. Transfection with dominant negative RhoA decreased Wnt activation by normalized TCF-Luc activity on SLA, whilst transfection with constitutively active RhoA increased TCF-Luc activation on Polished titanium. One mM Myosin II inhibitor Blebbistatin increased RhoA activation but decreased Wnt activation on SLA surfaces, indicating that tension-generating structures are required for canonical Wnt modulation on titanium surfaces. Actin inhibitor Cytochalasin markedly enhanced RhoA and TCF-Luc activation on both surfaces and increased the expression of differentiation markers in murine osteoblastic MC3T3 cells. Taken together, these data show that RhoA is upregulated in cells on rough surfaces and it affects the activation of Wnt canonical signaling through Myosin II modulation. PMID:24949442

  9. Wnt Acts as a Prosurvival Signal to Enhance Dentin Regeneration.

    PubMed

    Hunter, Daniel J; Bardet, Claire; Mouraret, Sylvain; Liu, Bo; Singh, Gurpreet; Sadoine, Jérémy; Dhamdhere, Girija; Smith, Andrew; Tran, Xuan Vinh; Joy, Adrienne; Rooker, Scott; Suzuki, Shigeki; Vuorinen, Annukka; Miettinen, Susanna; Chaussain, Catherine; Helms, Jill A

    2015-07-01

    Wnt proteins are lipid-modified, short-range signals that control stem cell self-renewal and tissue regeneration. We identified a population of Wnt responsive cells in the pulp cavity, characterized their function, and then created a pulp injury. The repair response was evaluated over time using molecular, cellular, and quantitative assays. We tested how healing was impacted by wound environments in which Wnt signaling was amplified. We found that a Wnt-amplified environment was associated with superior pulp healing. Although cell death was still rampant, the number of cells undergoing apoptosis was significantly reduced. This resulted in significantly better survival of injured pulp cells, and resulted in the formation of more tertiary dentin. We engineered a liposome-reconstituted form of WNT3A then tested whether this biomimetic compound could activate cells in the injured tooth pulp and stimulate dentin regeneration. Pulp cells responded to the elevated Wnt stimulus by differentiating into secretory odontoblasts. Thus, transiently amplifying the body's natural Wnt response resulted in improved pulp vitality. These data have direct clinical implications for treating dental caries, the most prevalent disease affecting mankind. PMID:25556760

  10. The Tale of the Three Brothers – Shh, Wnt, and Fgf during Development of the Thalamus

    PubMed Central

    Hagemann, Anja I. H.; Scholpp, Steffen

    2012-01-01

    The thalamic complex is an essential part of the brain that requires a combination of specialized activities to attain its final complexity. In the following review we will describe the induction process of the mid-diencephalic organizer (MDO) where three different signaling pathways merge: Wnt, Shh, and Fgf. Here, we dissect the function of each signaling pathway in the thalamus in chronological order of their appearance. First we describe the Wnt mediated induction of the MDO and compartition of the caudal forebrain, then the Shh mediated determination of proneural gene expression before discussing recent progress in characterizing Fgf function during thalamus development. Then, we focus on transcription factors, which are regulated by these pathways and which play a pivotal role in neurogenesis in the thalamus. The three signaling pathways act together in a strictly regulated chronology to orchestrate the development of the entire thalamus. PMID:22654733

  11. Continuous tooth generation in mouse is induced by activated epithelial Wnt/β-catenin signaling

    PubMed Central

    Järvinen, Elina; Salazar-Ciudad, Isaac; Birchmeier, Walter; Taketo, Makoto M.; Jernvall, Jukka; Thesleff, Irma

    2006-01-01

    The single replacement from milk teeth to permanent teeth makes mammalian teeth different from teeth of most nonmammalian vertebrates and other epithelial organs such as hair and feathers, whose continuous replacement has been linked to Wnt signaling. Here we show that mouse tooth buds expressing stabilized β-catenin in epithelium give rise to dozens of teeth. The molar crowns, however, are typically simplified unicusped cones. We demonstrate that the supernumerary teeth develop by a renewal process where new signaling centers, the enamel knots, bud off from the existing dental epithelium. The basic aspects of the unlocked tooth renewal can be reproduced with a computer model on tooth development by increasing the intrinsic level of activator production, supporting the role of β-catenin pathway as an upstream activator of enamel knot formation. These results may implicate Wnt signaling in tooth renewal, a capacity that was all but lost when mammals evolved progressively more complicated tooth shapes. PMID:17121988

  12. Continuous tooth generation in mouse is induced by activated epithelial Wnt/beta-catenin signaling.

    PubMed

    Järvinen, Elina; Salazar-Ciudad, Isaac; Birchmeier, Walter; Taketo, Makoto M; Jernvall, Jukka; Thesleff, Irma

    2006-12-01

    The single replacement from milk teeth to permanent teeth makes mammalian teeth different from teeth of most nonmammalian vertebrates and other epithelial organs such as hair and feathers, whose continuous replacement has been linked to Wnt signaling. Here we show that mouse tooth buds expressing stabilized beta-catenin in epithelium give rise to dozens of teeth. The molar crowns, however, are typically simplified unicusped cones. We demonstrate that the supernumerary teeth develop by a renewal process where new signaling centers, the enamel knots, bud off from the existing dental epithelium. The basic aspects of the unlocked tooth renewal can be reproduced with a computer model on tooth development by increasing the intrinsic level of activator production, supporting the role of beta-catenin pathway as an upstream activator of enamel knot formation. These results may implicate Wnt signaling in tooth renewal, a capacity that was all but lost when mammals evolved progressively more complicated tooth shapes. PMID:17121988

  13. SOXC proteins amplify canonical WNT signaling to secure nonchondrocytic fates in skeletogenesis.

    PubMed

    Bhattaram, Pallavi; Penzo-Méndez, Alfredo; Kato, Kenji; Bandyopadhyay, Kaustav; Gadi, Abhilash; Taketo, Makoto M; Lefebvre, Véronique

    2014-12-01

    Canonical WNT signaling stabilizes β-catenin to determine cell fate in many processes from development onwards. One of its main roles in skeletogenesis is to antagonize the chondrogenic transcription factor SOX9. We here identify the SOXC proteins as potent amplifiers of this pathway. The SOXC genes, i.e., Sox4, Sox11, and Sox12, are coexpressed in skeletogenic mesenchyme, including presumptive joints and perichondrium, but not in cartilage. Their inactivation in mouse embryo limb bud caused massive cartilage fusions, as joint and perichondrium cells underwent chondrogenesis. SOXC proteins govern these cells cell autonomously. They replace SOX9 in the adenomatous polyposis coli-Axin destruction complex and therein inhibit phosphorylation of β-catenin by GSK3. This inhibition, a crucial, limiting step in canonical WNT signaling, thus becomes a constitutive event. The resulting SOXC/canonical WNT-mediated synergistic stabilization of β-catenin contributes to efficient repression of Sox9 in presumptive joint and perichondrium cells and thereby ensures proper delineation and articulation of skeletal primordia. This synergy may determine cell fate in many processes besides skeletogenesis. PMID:25452386

  14. Maintaining embryonic stem cell pluripotency with Wnt signaling.

    PubMed

    Sokol, Sergei Y

    2011-10-01

    Wnt signaling pathways control lineage specification in vertebrate embryos and regulate pluripotency in embryonic stem (ES) cells, but how the balance between progenitor self-renewal and differentiation is achieved during axis specification and tissue patterning remains highly controversial. The context- and stage-specific effects of the different Wnt pathways produce complex and sometimes opposite outcomes that help to generate embryonic cell diversity. Although the results of recent studies of the Wnt/β-catenin pathway in ES cells appear to be surprising and controversial, they converge on the same conserved mechanism that leads to the inactivation of TCF3-mediated repression. PMID:21903672

  15. Effects of temperature, packaging and electron beam irradiation processing conditions on the property behaviour of Poly (ether-block-amide) blends.

    PubMed

    Murray, Kieran A; Kennedy, James E; McEvoy, Brian; Vrain, Olivier; Ryan, Damien; Cowman, Richard; Higginbotham, Clement L

    2014-06-01

    The radiation stability of Poly (ether-block-amide) (PEBA) blended with a multifunctional phenolic antioxidant and a hindered amide light stabiliser was examined under various temperatures, packaging and electron beam processing conditions. FTIR revealed that there were slight alterations to the PEBA before irradiation; however, these became more pronounced following irradiation. The effect of varying the temperature, packaging and processing conditions on the resultant PEBA properties was apparent. For example, rheology demonstrated that the structural properties could be enhanced by manipulating the aforementioned criteria. Mechanical testing exhibited less radiation resistance when the PEBA samples were vacuum packed and exposed to irradiation. MFI and AFM confirmed that the melting strength and surface topography could be reduced/increased depending on the conditions employed. From this study it was concluded that virgin PEBA submerged in dry ice with non-vacuum packaging during the irradiation process, provided excellent radiation resistance (20.9% improvement) in contrast to the traditional method. PMID:24863239

  16. Structural geometry and kinematic processes at the intracontinental Daloushan mountain chain: Implications for tectonic transfer in the Yangtze Block interior

    NASA Astrophysics Data System (ADS)

    Deng, Bin; Li, Zhi-Wu; Liu, Shu-Gen; Wang, Guo-Zhi; Li, Shuang-Jian; Qin, Zuo-Pen; Li, Jing-Xi; Jansa, Luba

    2016-02-01

    The Daloushan mountain chain, located in the centre of the upper Yangtze continental block, is considered to represent the locus of the tectonic shortening resulting from the eastward growth of the Tibetan Plateau and NW-thrusting of the Xuefeng Orogen. Structural data and apatite fission-track ages have been used to decipher the geometry and the kinematic evolution of the Daloushan. The latter is subdivided into two domains: the eastern domain, governed by west- to NW-verging thrusting and deformation with dextral transpression, and a western domain, governed by south-verging thrusting and deformation. Both domains experienced four episodes of deformation, synchronous with the four stages of post-Cretaceous denudation, marked by rapid cooling propagating eastward from 20 to 5 Ma, at a rate of ∼0.1 mm/year. In particular, the last two episodes of denudation are closely related to the growth of the Tibetan Plateau. This indicates an intra-continental transfer of tectonic forcing from the Palaeo-Pacific to the Tethys-Himalayan Tectonic Domain across the Daloushan.

  17. Mapping the dynamic expression of Wnt11 and the lineage contribution of Wnt11-expressing cells during early mouse development.

    PubMed

    Sinha, Tanvi; Lin, Lizhu; Li, Ding; Davis, Jennifer; Evans, Sylvia; Wynshaw-Boris, Anthony; Wang, Jianbo

    2015-02-15

    Planar cell polarity (PCP) signaling is an evolutionarily conserved mechanism that coordinates polarized cell behavior to regulate tissue morphogenesis during vertebrate gastrulation, neurulation and organogenesis. In Xenopus and zebrafish, PCP signaling is activated by non-canonical Wnts such as Wnt11, and detailed understanding of Wnt11 expression has provided important clues on when, where and how PCP may be activated to regulate tissue morphogenesis. To explore the role of Wnt11 in mammalian development, we established a Wnt11 expression and lineage map with high spatial and temporal resolution by creating and analyzing a tamoxifen-inducible Wnt11-CreER BAC (bacterial artificial chromosome) transgenic mouse line. Our short- and long-term lineage tracing experiments indicated that Wnt11-CreER could faithfully recapitulate endogenous Wnt11 expression, and revealed for the first time that cells transiently expressing Wnt11 at early gastrulation were fated to become specifically the progenitors of the entire endoderm. During mid-gastrulation, Wnt11-CreER expressing cells also contribute extensively to the endothelium in both embryonic and extraembryonic compartments, and the endocardium in all chambers of the developing heart. In contrast, Wnt11-CreER expression in the myocardium starts from late-gastrulation, and occurs in three transient, sequential waves: first in the precursors of the left ventricular (LV) myocardium from E7.0 to 8.0; subsequently in the right ventricular (RV) myocardium from E8.0 to 9.0; and finally in the superior wall of the outflow tract (OFT) myocardium from E8.5 to 10.5. These results provide formal genetic proof that the majority of the endocardium and myocardium diverge by mid-gastrulation in the mouse, and suggest a tight spatial and temporal control of Wnt11 expression in the myocardial lineage to coordinate with myocardial differentiation in the first and second heart field progenitors to form the LV, RV and OFT. The insights gained

  18. Transient Expression of WNT2 Promotes Somatic Cell Reprogramming by Inducing β-Catenin Nuclear Accumulation.

    PubMed

    Kimura, Mizuki; Nakajima-Koyama, May; Lee, Joonseong; Nishida, Eisuke

    2016-06-14

    Treatment with several Wnt/β-catenin signaling pathway regulators can change the cellular reprogramming efficiency; however, the dynamics and role of endogenous Wnt/β-catenin signaling in reprogramming remain largely unanswered. Here we identify the upregulation of WNT2 and subsequent β-catenin nuclear accumulation as key events in reprogramming. Transient nuclear accumulation of β-catenin occurs early in MEF reprogramming. Wnt2 is strongly expressed in the early stage of reprogramming. Wnt2 knockdown suppresses the nuclear accumulation of β-catenin and reduces the reprogramming efficiency. WNT2 overexpression promotes β-catenin nuclear accumulation and enhances the reprogramming efficiency. WNT2 contributes to the promotion of cell proliferation. Experiments with several drugs that control the Wnt pathway also indicate the importance of β-catenin nuclear accumulation in reprogramming. Our findings reveal the role of WNT2/β-catenin signaling in reprogramming. PMID:27211212

  19. WNT2 Promotes Cervical Carcinoma Metastasis and Induction of Epithelial-Mesenchymal Transition

    PubMed Central

    Xu, Jing; Zhang, Lan; Wang, Jianhua; Huang, Long; Huang, Shuting; Yuan, Linjing; Jia, Weihua; Yu, Xingjuan; Luo, Rongzhen; Zheng, Min

    2016-01-01

    Background Previously, we found an 11-gene signature could predict pelvic lymph node metastasis (PLNM), and WNT2 is one of the key genes in the signature. This study explored the expression and underlying mechanism of WNT2 in PLNM of cervical cancer. Methods WNT2 expression level in cervical cancer was detected using western blotting, quantitative PCR, and immunohistochemistry. Two WNT2-specific small interfering RNAs (siRNAs) were used to explore the effects of WNT2 on invasive and metastatic ability of cancer cells, and to reveal the possible mechanism of WNT2 affecting epithelial—mesenchymal transition (EMT). The correlation between WNT2 expression and PLNM was further investigated in clinical cervical specimens. Results Both WNT2 mRNA and protein expression was upregulated in cervical cancer. High WNT2 expression was significantly associated with tumor size, lymphovascular space involvement, positive parametrium, and most importantly, PLNM. PLNM and WNT2 expression were independent prognostic factors for overall survival and disease-free survival. WNT2 knockdown inhibited SiHa cell motility and invasion and reversed EMT by inhibiting the WNT2/β-catenin pathway. WNT2 overexpression in cervical cancer was associated with β-catenin activation and induction of EMT, which further contributed to metastasis in cervical cancer. Conclusion WNT2 might be a novel predictor of PLNM and a promising prognostic indicator in cervical cancer. PMID:27513465

  20. Epithelial Wnt/βcatenin signalling is essential for epididymal coiling.

    PubMed

    Kumar, Manish; Syed, Shafiq M; Taketo, Makoto M; Tanwar, Pradeep S

    2016-04-15

    Organ shape and size are important determinants of their physiological functions. Epithelial tubes are anlagen of many complex organs. How these tubes acquire their complex shape and size is a fundamental question in biology. In male mice, the Wolffian duct (WD; postnatally known as epididymis) undergoes an astonishing transformation, where a straight tube only a few millimetres long elongates to over 1000 times its original length and fits into a very small space, due to extensive coiling of epithelium, to perform the highly specialized function of sperm maturation. Defective coiling disrupts sperm maturation and leads to male infertility. Recent work has shown that epithelial cell proliferation is a major driver of WD coiling. Still, very little is known about the molecular signals involved in this process. Testicular androgens are known regulators of WD development. However, epithelial androgen receptor signalling is dispensable for WD coiling. In this study, we have shown that Wnt signalling is highly active in the entire WD epithelium during its coiling, and is limited to only a few segments of the epididymis in later life. Pharmacological and genetic suppression of Wnt signalling inhibited WD coiling by decreasing cell proliferation and promoting apoptosis. Comparative gene expression analysis identified Fibroblast growth factor 7 (Fgf7) as a prime Wnt target gene involved in WD coiling and in vitro treatment with Fgf7 protein increased coiling of WDs. In summary, our work has established that epithelial canonical Wnt signalling is a critical regulator of WD coiling and its precise regulation is essential for WD/epididymal differentiation. PMID:26934381

  1. Identification and expression characterization of three Wnt signaling genes in pearl oyster (Pinctada fucata).

    PubMed

    Gao, Jing; Liu, Jun; Yang, Yi; Liang, Jian; Xie, Jun; Li, Shiguo; Zheng, Guilang; Xie, Liping; Zhang, Rongqing

    2016-01-01

    The Wnt signaling pathway plays an important role in animal development and in the biomineralization process. At present, although the biomineralization mechanism in pearl oyster (Pinctada fucata) has been extensively studied, there is little research on the Wnt signaling pathway in pearl oyster. To understand the potential role of the Wnt signaling pathway in pearl oyster, we cloned and sequenced three genes from the Wnt signaling pathway in pearl oyster that encode the following proteins: β-catenin, Dishevelled (Dvl) and T-cell factor (TCF). Genomic structure analysis revealed that Pf-β-catenin genomic DNA contained 15 exons, Pf-Dvl genomic DNA contained 16 exons, and Pf-TCF genomic DNA contained 7 exons. Their deduced amino acid sequences all showed the highest identity with homologs in Crassostrea gigas. Yeast two-hybrid analysis verified that Pf-β-catenin interacted with Pf-TCF. These three genes were ubiquitously expressed in seven pearl oyster tissues analyzed with the highest expression in the gill and a certain amount of expression in the mantle, a tissue related to shell formation. After shell notching, the dynamic changes in expression of these three genes showed that they reached a maximum at 4days, indicating their response to shell regeneration. All three genes were constitutively expressed during five developmental stages of the pearl oyster, with high levels at the early embryonic development stage. Taken together, these results suggested that Pf-β-catenin, Pf-Dvl and Pf-TCF might participate in shell formation and early embryonic and larval development in the pearl oyster. PMID:26969109

  2. Wnt and FGF signals interact to coordinate growth with cell fate specification during limb development

    PubMed Central

    ten Berge, Derk; Brugmann, Samantha A.; Helms, Jill A.; Nusse, Roel

    2009-01-01

    A fundamental question in developmental biology is how does an undifferentiated field of cells acquire spatial pattern and undergo coordinated differentiation? The development of the vertebrate limb is an important paradigm for understanding these processes. The skeletal and connective tissues of the developing limb all derive from a population of multipotent progenitor cells located in its distal tip. During limb outgrowth, these progenitors segregate into a chondrogenic lineage, located in the center of the limb bud, and soft connective tissue lineages located in its periphery. We report that the interplay of two families of signaling proteins, fibroblast growth factors (FGFs) and Wnts, coordinate the growth of the multipotent progenitor cells with their simultaneous segregation into these lineages. FGF and Wnt signals act together to synergistically promote proliferation while maintaining the cells in an undifferentiated, multipotent state, but act separately to determine cell lineage specification. Withdrawal of both signals results in cell cycle withdrawal and chondrogenic differentiation. Continued exposure to Wnt, however, maintains proliferation and re-specifies the cells towards the soft connective tissue lineages. We have identified target genes that are synergistically regulated by Wnts and FGFs, and show how these factors actively suppress differentiation and promote growth. Finally, we show how the spatial restriction of Wnt and FGF signals to the limb ectoderm, and to a specialized region of it, the apical ectodermal ridge, controls the distribution of cell behaviors within the growing limb, and guides the proper spatial organization of the differentiating tissues. PMID:18776145

  3. Orchestrating A/P and D/V guidance - A Wnt/Netrin tale

    PubMed Central

    Levy-Strumpf, Naomi

    2016-01-01

    ABSTRACT While ample information was gathered in identifying guidance cues and their downstream mediators, very little is known about how the information from multiple extracellular cues is intracellularly to generate normal patterning. Netrin and Wnt signaling pathways play key roles in normal development as well as in malignancies. In C. elegans, as in vertebrates, dorso-ventral (D/V) graded distributions of UNC-6/Netrin and antero-posterior (A/P) graded distributions of Wnts provide instructive polarity information to guide cells and axons along their respective gradients. In this commentary, I will discuss recent findings demonstrating that these 2 signaling pathways also function redundantly to regulate polarity orthogonal to the axis of their gradation. Thus, Wnt signaling components contribute to D/V polarity, while Netrin signaling components contribute to A/P polarity and their joint action collaboratively governs migratory transitions from one axis to the other. These findings pave the way to unraveling broader roles of Wnt and Netrin signaling pathways, roles that are masked due to their redundant nature, and provide a conceptually novel view of how antero-posterior and dorso-ventral guidance mechanisms are orchestrated to establish polarity in multiple biological processes. PMID:27073738

  4. Wnt5a is essential for intestinal elongation in mice

    PubMed Central

    Cervantes, Sara; Yamaguchi, Terry P.; Hebrok, Matthias

    2009-01-01

    Summary Morphogenesis of the mammalian small intestine entails extensive elongation and folding of the primitive gut into a tightly coiled digestive tube. Surprisingly, little is known about the cellular and molecular mechanisms that mediate the morphological aspects of small intestine formation. Here, we demonstrate that Wnt5a, a member of the Wnt family of secreted proteins, is essential for the development and elongation of the small intestine from the midgut region. We found that the small intestine in mice lacking Wnt5a was dramatically shortened and duplicated, forming a bifurcated lumen instead of a single tube. In addition, cell proliferation was reduced and re-intercalation of post-mitotic cells into the elongating gut tube epithelium was disrupted. Thus, our study demonstrates that Wnt5a functions as a critical regulator of midgut formation and morphogenesis in mammals. PMID:19100728

  5. Dysregulation of Wnt/β-catenin Signaling in Gastrointestinal Cancers

    PubMed Central

    White, Bryan D.; Chien, Andy J.; Dawson, David W.

    2012-01-01

    Aberrant Wnt/β-catenin signaling is widely implicated in numerous malignancies, including cancers of the gastrointestinal (GI) tract. Dysregulation of signaling is traditionally attributed to mutations in Axin, APC (adenomatous polyposis coli), and β-catenin that lead to constitutive hyperactivation of the pathway. However, Wnt/β-catenin signaling is also modulated through various other mechanisms in cancer, including crosstalk with other altered signaling pathways. A more complex view of Wnt/β-catenin signaling and its role in GI cancers is now emerging as divergent phenotypic outcomes are found to be dictated by temporospatial context and relative levels of pathway activation. This review summarizes the dysregulation of Wnt/β-catenin signaling in colorectal carcinoma, hepatocellular carcinoma, and pancreatic ductal adenocarcinoma, with particular emphasis on the latter two. We conclude by addressing some of the major challenges faced in attempting to target the pathway in the clinic. PMID:22155636

  6. Wnt Signaling in Neurogenesis during Aging and Physical Activity

    PubMed Central

    Chen, Michael; Do, Huong

    2012-01-01

    Over the past decade, much progress has been made regarding our understanding of neurogenesis in both you