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Sample records for regulates sonic hedgehog

  1. Sonic Hedgehog regulates thymic epithelial cell differentiation

    PubMed Central

    Saldaña, José Ignacio; Solanki, Anisha; Lau, Ching-In; Sahni, Hemant; Ross, Susan; Furmanski, Anna L.; Ono, Masahiro; Holländer, Georg; Crompton, Tessa

    2016-01-01

    Sonic Hedgehog (Shh) is expressed in the thymus, where it regulates T cell development. Here we investigated the influence of Shh on thymic epithelial cell (TEC) development. Components of the Hedgehog (Hh) signalling pathway were expressed by TEC, and use of a Gli Binding Site-green fluorescence protein (GFP) transgenic reporter mouse demonstrated active Hh-dependent transcription in TEC in the foetal and adult thymus. Analysis of Shh-deficient foetal thymus organ cultures (FTOC) showed that Shh is required for normal TEC differentiation. Shh-deficient foetal thymus contained fewer TEC than wild type (WT), the proportion of medullary TEC was reduced relative to cortical TEC, and cell surface expression of MHC Class II molecules was increased on both cortical and medullary TEC populations. In contrast, the Gli3-deficient thymus, which shows increased Hh-dependent transcription in thymic stroma, had increased numbers of TEC, but decreased cell surface expression of MHC Class II molecules on both cortical and medullary TEC. Neutralisation of endogenous Hh proteins in WT FTOC led to a reduction in TEC numbers, and in the proportion of mature Aire-expressing medullary TEC, but an increase in cell surface expression of MHC Class II molecules on medullary TEC. Likewise, conditional deletion of Shh from TEC in the adult thymus resulted in alterations in TEC differentiation and consequent changes in T cell development. TEC numbers, and the proportion of mature Aire-expressing medullary TEC were reduced, and cell surface expression of MHC Class II molecules on medullary TEC was increased. Differentiation of mature CD4 and CD8 single positive thymocytes was increased, demonstrating the regulatory role of Shh production by TEC on T cell development. Treatment of human thymus explants with recombinant Shh or neutralising anti-Shh antibody indicated that the Hedgehog pathway is also involved in regulation of differentiation from DP to mature SP T cells in the human thymus. PMID

  2. Regulation of Patched by Sonic Hedgehog in the Developing Neural Tube

    NASA Astrophysics Data System (ADS)

    Marigo, Valeria; Tabin, Clifford J.

    1996-09-01

    Ventral cell fates in the central nervous system are induced by Sonic hedgehog, a homolog of hedgehog, a secreted Drosophila protein. In the central nervous system, Sonic hedgehog has been identified as the signal inducing floor plate, motor neurons, and dopaminergic neurons. Sonic hedgehog is also involved in the induction of ventral cell type in the developing somites. ptc is a key gene in the Drosophila hedgehog signaling pathway where it is involved in transducing the hedgehog signal and is also a transcriptional target of the signal. PTC, a vertebrate homolog of this Drosophila gene, is genetically downstream of Sonic hedgehog (Shh) in the limb bud. We analyze PTC expression during chicken neural and somite development and find it expressed in all regions of these tissues known to be responsive to Sonic hedgehog signal. As in the limb bud, ectopic expression of Sonic hedgehog leads to ectopic induction of PTC in the neural tube and paraxial mesoderm. This conservation of regulation allows us to use PTC as a marker for Sonic hedgehog response. The pattern of PTC expression suggests that Sonic hedgehog may play an inductive role in more dorsal regions of the neural tube than have been previously demonstrated. Examination of the pattern of PTC expression also suggests that PTC may act in a negative feedback loop to attenuate hedgehog signaling.

  3. Regulation of patched by sonic hedgehog in the developing neural tube.

    PubMed Central

    Marigo, V; Tabin, C J

    1996-01-01

    Ventral cell fates in the central nervous system are induced by Sonic hedgehog, a homolog of hedgehog, a secreted Drosophila protein. In the central nervous system, Sonic hedgehog has been identified as the signal inducing floor plate, motor neurons, and dopaminergic neurons. Sonic hedgehog is also involved in the induction of ventral cell type in the developing somites. ptc is a key gene in the Drosophila hedgehog signaling pathway where it is involved in transducing the hedgehog signal and is also a transcriptional target of the signal. PTC, a vertebrate homolog of this Drosophila gene, is genetically downstream of Sonic hedgehog (Shh) in the limb bud. We analyze PTC expression during chicken neural and somite development and find it expressed in all regions of these tissues known to be responsive to Sonic hedgehog signal. As in the limb bud, ectopic expression of Sonic hedgehog leads to ectopic induction of PTC in the neural tube and paraxial mesoderm. This conservation of regulation allows us to use PTC as a marker for Sonic hedgehog response. The pattern of PTC expression suggests that Sonic hedgehog may play an inductive role in more dorsal regions of the neural tube than have been previously demonstrated. Examination of the pattern of PTC expression also suggests that PTC may act in a negative feedback loop to attenuate hedgehog signaling. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8790332

  4. YAP regulates neuronal differentiation through Sonic hedgehog signaling pathway

    SciTech Connect

    Lin, Yi-Ting; Ding, Jing-Ya; Li, Ming-Yang; Yeh, Tien-Shun; Wang, Tsu-Wei; Yu, Jenn-Yah

    2012-09-10

    Tight regulation of cell numbers by controlling cell proliferation and apoptosis is important during development. Recently, the Hippo pathway has been shown to regulate tissue growth and organ size in Drosophila. In mammalian cells, it also affects cell proliferation and differentiation in various tissues, including the nervous system. Interplay of several signaling cascades, such as Notch, Wnt, and Sonic Hedgehog (Shh) pathways, control cell proliferation during neuronal differentiation. However, it remains unclear whether the Hippo pathway coordinates with other signaling cascades in regulating neuronal differentiation. Here, we used P19 cells, a mouse embryonic carcinoma cell line, as a model to study roles of YAP, a core component of the Hippo pathway, in neuronal differentiation. P19 cells can be induced to differentiate into neurons by expressing a neural bHLH transcription factor gene Ascl1. Our results showed that YAP promoted cell proliferation and inhibited neuronal differentiation. Expression of Yap activated Shh but not Wnt or Notch signaling activity during neuronal differentiation. Furthermore, expression of Yap increased the expression of Patched homolog 1 (Ptch1), a downstream target of the Shh signaling. Knockdown of Gli2, a transcription factor of the Shh pathway, promoted neuronal differentiation even when Yap was over-expressed. We further demonstrated that over-expression of Yap inhibited neuronal differentiation in primary mouse cortical progenitors and Gli2 knockdown rescued the differentiation defect in Yap over-expressing cells. In conclusion, our study reveals that Shh signaling acts downstream of YAP in regulating neuronal differentiation. -- Highlights: Black-Right-Pointing-Pointer YAP promotes cell proliferation and inhibits neuronal differentiation in P19 cells. Black-Right-Pointing-Pointer YAP promotes Sonic hedgehog signaling activity during neuronal differentiation. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap

  5. Dendrosomatic Sonic Hedgehog Signaling in Hippocampal Neurons Regulates Axon Elongation

    PubMed Central

    Petralia, Ronald S.; Ott, Carolyn; Wang, Ya-Xian; Lippincott-Schwartz, Jennifer; Mattson, Mark P.

    2015-01-01

    The presence of Sonic Hedgehog (Shh) and its signaling components in the neurons of the hippocampus raises a question about what role the Shh signaling pathway may play in these neurons. We show here that activation of the Shh signaling pathway stimulates axon elongation in rat hippocampal neurons. This Shh-induced effect depends on the pathway transducer Smoothened (Smo) and the transcription factor Gli1. The axon itself does not respond directly to Shh; instead, the Shh signal transduction originates from the somatodendritic region of the neurons and occurs in neurons with and without detectable primary cilia. Upon Shh stimulation, Smo localization to dendrites increases significantly. Shh pathway activation results in increased levels of profilin1 (Pfn1), an actin-binding protein. Mutations in Pfn1's actin-binding sites or reduction of Pfn1 eliminate the Shh-induced axon elongation. These findings indicate that Shh can regulate axon growth, which may be critical for development of hippocampal neurons. SIGNIFICANCE STATEMENT Although numerous signaling mechanisms have been identified that act directly on axons to regulate their outgrowth, it is not known whether signals transduced in dendrites may also affect axon outgrowth. We describe here a transcellular signaling pathway in embryonic hippocampal neurons in which activation of Sonic Hedgehog (Shh) receptors in dendrites stimulates axon growth. The pathway involves the dendritic-membrane-associated Shh signal transducer Smoothened (Smo) and the transcription factor Gli, which induces the expression of the gene encoding the actin-binding protein profilin 1. Our findings suggest scenarios in which stimulation of Shh in dendrites results in accelerated outgrowth of the axon, which therefore reaches its presumptive postsynaptic target cell more quickly. By this mechanism, Shh may play critical roles in the development of hippocampal neuronal circuits. PMID:26658865

  6. Interaction of PACAP with Sonic hedgehog reveals complex regulation of the hedgehog pathway by PKA.

    PubMed

    Niewiadomski, Pawel; Zhujiang, Annie; Youssef, Mary; Waschek, James A

    2013-11-01

    Sonic hedgehog (Shh) signaling is essential for proliferation of cerebellar granule cell progenitors (cGCPs) and its aberrant activation causes a cerebellar cancer medulloblastoma. Pituitary adenylate cyclase activating polypeptide (PACAP) inhibits Shh-driven proliferation of cGCPs and acts as tumor suppressor in murine medulloblastoma. We show that PACAP blocks canonical Shh signaling by a mechanism that involves activation of protein kinase A (PKA) and inhibition of the translocation of the Shh-dependent transcription factor Gli2 into the primary cilium. PKA is shown to play an essential role in inhibiting gene transcription in the absence of Shh, but global PKA activity levels are found to be a poor predictor of the degree of Shh pathway activation. We propose that the core Shh pathway regulates a small compartmentalized pool of PKA in the vicinity of primary cilia. GPCRs that affect global PKA activity levels, such as the PACAP receptor, cooperate with the canonical Shh signal to regulate Gli protein phosphorylation by PKA. This interaction serves to fine-tune the transcriptional and physiological function of the Shh pathway. PMID:23872071

  7. Sonic hedgehog is a regulator of extracellular glutamate levels and epilepsy.

    PubMed

    Feng, Shengjie; Ma, Shaorong; Jia, Caixia; Su, Yujuan; Yang, Shenglian; Zhou, Kechun; Liu, Yani; Cheng, Ju; Lu, Dunguo; Fan, Liu; Wang, Yizheng

    2016-05-01

    Sonic hedgehog (Shh), both as a mitogen and as a morphogen, plays an important role in cell proliferation and differentiation during early development. Here, we show that Shh inhibits glutamate transporter activities in neurons, rapidly enhances extracellular glutamate levels, and affects the development of epilepsy. Shh is quickly released in response to epileptic, but not physiological, stimuli. Inhibition of neuronal glutamate transporters by Shh depends on heterotrimeric G protein subunit Gαi and enhances extracellular glutamate levels. Inhibiting Shh signaling greatly reduces epileptiform activities in both cell cultures and hippocampal slices. Moreover, pharmacological or genetic inhibition of Shh signaling markedly suppresses epileptic phenotypes in kindling or pilocarpine models. Our results suggest that Shh contributes to the development of epilepsy and suppression of its signaling prevents the development of the disease. Thus, Shh can act as a modulator of neuronal activity, rapidly regulating glutamate levels and promoting epilepsy. PMID:27113760

  8. p63 Sustains self-renewal of mammary cancer stem cells through regulation of Sonic Hedgehog signaling.

    PubMed

    Memmi, Elisa Maria; Sanarico, Anna Giulia; Giacobbe, Arianna; Peschiaroli, Angelo; Frezza, Valentina; Cicalese, Angelo; Pisati, Federica; Tosoni, Daniela; Zhou, Huiqing; Tonon, Giovanni; Antonov, Alexey; Melino, Gerry; Pelicci, Pier Giuseppe; Bernassola, Francesca

    2015-03-17

    The predominant p63 isoform, ΔNp63, is a master regulator of normal epithelial stem cell (SC) maintenance. However, in vivo evidence of the regulation of cancer stem cell (CSC) properties by p63 is still limited. Here, we exploit the transgenic MMTV-ErbB2 (v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 2) mouse model of carcinogenesis to dissect the role of p63 in the regulation of mammary CSC self-renewal and breast tumorigenesis. ErbB2 tumor cells enriched for SC-like properties display increased levels of ΔNp63 expression compared with normal mammary progenitors. Down-regulation of p63 in ErbB2 mammospheres markedly restricts self-renewal and expansion of CSCs, and this action is fully independent of p53. Furthermore, transplantation of ErbB2 progenitors expressing shRNAs against p63 into the mammary fat pads of syngeneic mice delays tumor growth in vivo. p63 knockdown in ErbB2 progenitors diminishes the expression of genes encoding components of the Sonic Hedgehog (Hh) signaling pathway, a driver of mammary SC self-renewal. Remarkably, p63 regulates the expression of Sonic Hedgehog (Shh), GLI family zinc finger 2 (Gli2), and Patched1 (Ptch1) genes by directly binding to their gene regulatory regions, and eventually contributes to pathway activation. Collectively, these studies highlight the importance of p63 in maintaining the self-renewal potential of mammary CSCs via a positive modulation of the Hh signaling pathway. PMID:25739959

  9. Sonic hedgehog signaling regulates mode of cell division of early cerebral cortex progenitors and increases astrogliogenesis

    PubMed Central

    Araújo, Geissy L. L.; Araújo, Jessica A. M.; Schroeder, Timm; Tort, Adriano B. L.; Costa, Marcos R.

    2014-01-01

    The morphogen Sonic Hedgehog (SHH) plays a critical role in the development of different tissues. In the central nervous system, SHH is well known to contribute to the patterning of the spinal cord and separation of the brain hemispheres. In addition, it has recently been shown that SHH signaling also contributes to the patterning of the telencephalon and establishment of adult neurogenic niches. In this work, we investigated whether SHH signaling influences the behavior of neural progenitors isolated from the dorsal telencephalon, which generate excitatory neurons and macroglial cells in vitro. We observed that SHH increases proliferation of cortical progenitors and generation of astrocytes, whereas blocking SHH signaling with cyclopamine has opposite effects. In both cases, generation of neurons did not seem to be affected. However, cell survival was broadly affected by blockade of SHH signaling. SHH effects were related to three different cell phenomena: mode of cell division, cell cycle length and cell growth. Together, our data in vitro demonstrate that SHH signaling controls cell behaviors that are important for proliferation of cerebral cortex progenitors, as well as differentiation and survival of neurons and astroglial cells. PMID:24653675

  10. Sonic hedgehog signaling regulates amygdalar neurogenesis and extinction of fear memory.

    PubMed

    Hung, Hui-Chi; Hsiao, Ya-Hsin; Gean, Po-Wu

    2015-10-01

    It is now recognized that neurogenesis occurs throughout life predominantly in the subgranular zone (SGZ) of the hippocampus and the subventricular zone (SVZ) of the lateral ventricle. In the present study, we investigated the relationship between neurogenesis in the amygdala and extinction of fear memory. Mice received 15 tone-footshock pairings. Twenty-four hours after training, the mice were given 15 tone-alone trials (extinction training) once per day for 7 days. Two hours before extinction training, the mice were injected intraperitoneally with 5-bromo-3-deoxyuridine (BrdU). BrdU-positive and NeuN-positive cells were analyzed 52 days after the training. A group of mice that received tone-footshock pairings but no extinction training served as controls (FC+No-Ext). The number of BrdU(+)/NeuN(+) cells was significantly higher in the extinction (FC+Ext) than in the FC+No-Ext mice. Proliferation inhibitor methylazoxymethanol acetate (MAM) or DNA synthesis inhibitor cytosine arabinoside (Ara-C) reduced neurogenesis and retarded extinction. Silencing Sonic hedgehog (Shh) gene with short hairpin interfering RNA (shRNA) by means of a retrovirus expression system to knockdown Shh specifically in the mitotic neurons reduced neurogenesis and retarded extinction. By contrast, over-expression of Shh increased neurogenesis and facilitated extinction. These results suggest that amygdala neurogenesis and Shh signaling are involved in the extinction of fear memory.

  11. Generation of reactive astrocytes from NG2 cells is regulated by sonic hedgehog.

    PubMed

    Honsa, Pavel; Valny, Martin; Kriska, Jan; Matuskova, Hana; Harantova, Lenka; Kirdajova, Denisa; Valihrach, Lukas; Androvic, Peter; Kubista, Mikael; Anderova, Miroslava

    2016-09-01

    NG2 cells, a fourth glial cell type in the adult mammalian central nervous system, produce oligodendrocytes in the healthy nervous tissue, and display wide differentiation potential under pathological conditions, where they could give rise to reactive astrocytes. The factors that control the differentiation of NG2 cells after focal cerebral ischemia (FCI) are largely unknown. Here, we used transgenic Cspg4-cre/Esr1/ROSA26Sortm14(CAG-tdTomato) mice, in which tamoxifen administration triggers the expression of red fluorescent protein (tomato) specifically in NG2 cells and cells derived therefrom. Differentiation potential (in vitro and in vivo) of tomato-positive NG2 cells from control or postischemic brains was determined using the immunohistochemistry, single cell RT-qPCR and patch-clamp method. The ischemic injury was induced by middle cerebral artery occlusion, a model of FCI. Using genetic fate-mapping method, we identified sonic hedgehog (Shh) as an important factor that influences differentiation of NG2 cells into astrocytes in vitro. We also manipulated Shh signaling in the adult mouse brain after FCI. Shh signaling activation significantly increased the number of astrocytes derived from NG2 cells in the glial scar around the ischemic lesion, while Shh signaling inhibition caused the opposite effect. Since Shh signaling modifications did not change the proliferation rate of NG2 cells, we can conclude that Shh has a direct influence on the differentiation of NG2 cells and therefore, on the formation and composition of a glial scar, which consequently affects the degree of the brain damage. GLIA 2016;64:1518-1531. PMID:27340757

  12. Inhibitors of Hedgehog acyltransferase block Sonic Hedgehog signaling.

    PubMed

    Petrova, Elissaveta; Rios-Esteves, Jessica; Ouerfelli, Ouathek; Glickman, J Fraser; Resh, Marilyn D

    2013-04-01

    Inhibition of Sonic hedgehog (Shh) signaling is of great clinical interest. Here we exploit Hedgehog acyltransferase (Hhat)-mediated Shh palmitoylation, a modification critical for Shh signaling, as a new target for Shh pathway inhibition. A target-oriented high-throughput screen was used to identify small-molecule inhibitors of Hhat. In cells, these Hhat inhibitors specifically block Shh palmitoylation and inhibit autocrine and paracrine Shh signaling.

  13. Inhibitors of Hedgehog acyltransferase block Sonic Hedgehog signaling.

    PubMed

    Petrova, Elissaveta; Rios-Esteves, Jessica; Ouerfelli, Ouathek; Glickman, J Fraser; Resh, Marilyn D

    2013-04-01

    Inhibition of Sonic hedgehog (Shh) signaling is of great clinical interest. Here we exploit Hedgehog acyltransferase (Hhat)-mediated Shh palmitoylation, a modification critical for Shh signaling, as a new target for Shh pathway inhibition. A target-oriented high-throughput screen was used to identify small-molecule inhibitors of Hhat. In cells, these Hhat inhibitors specifically block Shh palmitoylation and inhibit autocrine and paracrine Shh signaling. PMID:23416332

  14. LKB1 Regulates Cerebellar Development by Controlling Sonic Hedgehog-mediated Granule Cell Precursor Proliferation and Granule Cell Migration

    PubMed Central

    Men, Yuqin; Zhang, Aizhen; Li, Haixiang; Jin, Yecheng; Sun, Xiaoyang; Li, Huashun; Gao, Jiangang

    2015-01-01

    The Liver Kinase B1 (LKB1) gene plays crucial roles in cell differentiation, proliferation and the establishment of cell polarity. We created LKB1 conditional knockout mice (LKB1Atoh1 CKO) to investigate the function of LKB1 in cerebellar development. The LKB1Atoh1 CKO mice displayed motor dysfunction. In the LKB1Atoh1 CKO cerebellum, the overall structure had a larger volume and morelobules. LKB1 inactivationled to an increased proliferation of granule cell precursors (GCPs), aberrant granule cell migration and overproduction of unipolar brush cells. To investigate the mechanism underlying the abnormal foliation, we examined sonic hedgehog signalling (Shh) by testing its transcriptional mediators, the Gli proteins, which regulate the GCPs proliferation and cerebellar foliation during cerebellar development. The expression levels of Gli genes were significantly increased in the mutant cerebellum. In vitro assays showed that the proliferation of cultured GCPs from mutant cerebellum significantly increased, whereas the proliferation of mutant GCPs significantly decreased in the presence of a Shh inhibitor GDC-0049. Thus, LKB1 deficiency in the LKB1Atoh1 CKO mice enhanced Shh signalling, leading to the excessive GCP proliferation and the formation of extra lobules. We proposed that LKB1 regulates cerebellar development by controlling GCPs proliferation through Shh signalling during cerebellar development. PMID:26549569

  15. Wnt/β-Catenin and Sonic Hedgehog Pathways Interact in the Regulation of the Development of the Dorsal Mesenchymal Protrusion

    PubMed Central

    Briggs, Laura E.; Burns, Tara A.; Lockhart, Marie M.; Phelps, Aimee L.; Van den Hoff, Maurice J.B.; Wessels, Andy

    2016-01-01

    Background The dorsal mesenchymal protrusion (DMP) is a second heart field (SHF) derived tissue involved in cardiac septation. Molecular mechanisms controlling SHF/DMP development include the Bone Morphogenetic Protein and Wnt/β-catenin signaling pathways. Reduced expression of components in these pathways leads to inhibition of proliferation of the SHF/DMP precursor population and failure of the DMP to develop. While the Sonic Hedgehog (Shh) pathway has also been demonstrated to be critically important for SHF/DMP development and atrioventricular septation, its role in the regulation of SHF proliferation is contentious. Results Tissue-specific deletion of the Shh receptor Smoothened from the SHF resulted in compromised DMP formation and atrioventricular septal defects (AVSDs). Immunohistochemical analysis at critical stages of DMP development showed significant proliferation defect as well as reduction in levels of the Wnt/β-catenin pathway-intermediates β-catenin, Lef1, and Axin2. To determine whether the defects seen in the conditional Smoothened knock-out mouse could be attributed to reduced Wnt/β-catenin signaling, LiCl, a pharmacological activator of this Wnt/β-catenin pathway, was administered. This resulted in restoration of proliferation and partial rescue of the AVSD phenotype. Conclusions The data presented suggest that the Wnt/β-catenin pathway interact with the Shh pathway in the regulation of SHF/DMP-precursor proliferation and, hence, the development of the DMP. PMID:26297872

  16. Cell Division Mode Change Mediates the Regulation of Cerebellar Granule Neurogenesis Controlled by the Sonic Hedgehog Signaling

    PubMed Central

    Yang, Rong; Wang, Minglei; Wang, Jia; Huang, Xingxu; Yang, Ru; Gao, Wei-Qiang

    2015-01-01

    Summary Symmetric and asymmetric divisions are important for self-renewal and differentiation of stem cells during neurogenesis. Although cerebellar granule neurogenesis is controlled by sonic hedgehog (SHH) signaling, whether and how this process is mediated by regulation of cell division modes have not been determined. Here, using time-lapse imaging and cell culture from neuronal progenitor-specific and differentiated neuron-specific reporter mouse lines (Math1-GFP and Dcx-DsRed) and Patched+/− mice in which SHH signaling is activated, we find evidence for the existence of symmetric and asymmetric divisions that are closely associated with progenitor proliferation and differentiation. While activation of the SHH pathway enhances symmetric progenitor cell divisions, blockade of the SHH pathway reverses the cell division mode change in Math1-GFP;Dcx-DsRed;Patched+/− mice by promoting asymmetric divisions or terminal neuronal symmetric divisions. Thus, cell division mode change mediates the regulation of cerebellar granule neurogenesis controlled by SHH signaling. PMID:26527387

  17. Foxj1 regulates floor plate cilia architecture and modifies the response of cells to sonic hedgehog signalling

    PubMed Central

    Cruz, Catarina; Ribes, Vanessa; Kutejova, Eva; Cayuso, Jordi; Lawson, Victoria; Norris, Dominic; Stevens, Jonathan; Davey, Megan; Blight, Ken; Bangs, Fiona; Mynett, Anita; Hirst, Elizabeth; Chung, Rachel; Balaskas, Nikolaos; Brody, Steven L.; Marti, Elisa; Briscoe, James

    2010-01-01

    Sonic hedgehog signalling is essential for the embryonic development of many tissues including the central nervous system, where it controls the pattern of cellular differentiation. A genome-wide screen of neural progenitor cells to evaluate the Shh signalling-regulated transcriptome identified the forkhead transcription factor Foxj1. In both chick and mouse Foxj1 is expressed in the ventral midline of the neural tube in cells that make up the floor plate. Consistent with the role of Foxj1 in the formation of long motile cilia, floor plate cells produce cilia that are longer than the primary cilia found elsewhere in the neural tube, and forced expression of Foxj1 in neuroepithelial cells is sufficient to increase cilia length. In addition, the expression of Foxj1 in the neural tube and in an Shh-responsive cell line attenuates intracellular signalling by decreasing the activity of Gli proteins, the transcriptional mediators of Shh signalling. We show that this function of Foxj1 depends on cilia. Nevertheless, floor plate identity and ciliogenesis are unaffected in mouse embryos lacking Foxj1 and we provide evidence that additional transcription factors expressed in the floor plate share overlapping functions with Foxj1. Together, these findings identify a novel mechanism that modifies the cellular response to Shh signalling and reveal morphological and functional features of the amniote floor plate that distinguish these cells from the rest of the neuroepithelium. PMID:21098568

  18. miR-326 is downstream of Sonic hedgehog signaling and regulates the expression of Gli2 and smoothened.

    PubMed

    Jiang, Zhihua; Cushing, Leah; Ai, Xingbin; Lü, Jining

    2014-08-01

    Sonic hedgehog (Shh) is expressed and secreted from the embryonic lung epithelium and acts on the adjacent mesenchymal cells via its receptor Patched (Ptch)/Smoothened (Smo) and transcriptional effectors Gli proteins. Genetic studies showed that the Shh pathway plays critical roles in mouse lung development. However, little is known about microRNAs (miRNAs) downstream of Shh in embryonic lungs. Here we profiled miRNAs in embryonic lung cultures treated with cyclopamine, a specific Smo antagonist or with Smo agonist by next-generation of sequencing. We then performed functional screening to examine whether some of these miRNAs can modulate the induction of Gli-responsive luciferase by Shh treatment. These analyses revealed that expression of miR-326 and its host gene, Arrestin β1, is selectively enriched in embryonic lung mesenchymal cells and is specifically influenced by Shh activity. Furthermore, functional analyses showed that miR-326 acts as a negative modulator for Shh signaling by directly targeting Smo and Gli2. Together, these findings suggest a novel miR-326-negative feedback loop in regulating the activity of Shh signaling.

  19. Polycomb-Mediated Repression and Sonic Hedgehog Signaling Interact to Regulate Merkel Cell Specification during Skin Development.

    PubMed

    Perdigoto, Carolina N; Dauber, Katherine L; Bar, Carmit; Tsai, Pai-Chi; Valdes, Victor J; Cohen, Idan; Santoriello, Francis J; Zhao, Dejian; Zheng, Deyou; Hsu, Ya-Chieh; Ezhkova, Elena

    2016-07-01

    An increasing amount of evidence indicates that developmental programs are tightly regulated by the complex interplay between signaling pathways, as well as transcriptional and epigenetic processes. Here, we have uncovered coordination between transcriptional and morphogen cues to specify Merkel cells, poorly understood skin cells that mediate light touch sensations. In murine dorsal skin, Merkel cells are part of touch domes, which are skin structures consisting of specialized keratinocytes, Merkel cells, and afferent neurons, and are located exclusively around primary hair follicles. We show that the developing primary hair follicle functions as a niche required for Merkel cell specification. We find that intraepidermal Sonic hedgehog (Shh) signaling, initiated by the production of Shh ligand in the developing hair follicles, is required for Merkel cell specification. The importance of Shh for Merkel cell formation is further reinforced by the fact that Shh overexpression in embryonic epidermal progenitors leads to ectopic Merkel cells. Interestingly, Shh signaling is common to primary, secondary, and tertiary hair follicles, raising the possibility that there are restrictive mechanisms that regulate Merkel cell specification exclusively around primary hair follicles. Indeed, we find that loss of Polycomb repressive complex 2 (PRC2) in the epidermis results in the formation of ectopic Merkel cells that are associated with all hair types. We show that PRC2 loss expands the field of epidermal cells competent to differentiate into Merkel cells through the upregulation of key Merkel-differentiation genes, which are known PRC2 targets. Importantly, PRC2-mediated repression of the Merkel cell differentiation program requires inductive Shh signaling to form mature Merkel cells. Our study exemplifies how the interplay between epigenetic and morphogen cues regulates the complex patterning and formation of the mammalian skin structures. PMID:27414999

  20. Polycomb-Mediated Repression and Sonic Hedgehog Signaling Interact to Regulate Merkel Cell Specification during Skin Development

    PubMed Central

    Bar, Carmit; Tsai, Pai-Chi; Valdes, Victor J.; Cohen, Idan; Santoriello, Francis J.; Zhao, Dejian; Hsu, Ya-Chieh; Ezhkova, Elena

    2016-01-01

    An increasing amount of evidence indicates that developmental programs are tightly regulated by the complex interplay between signaling pathways, as well as transcriptional and epigenetic processes. Here, we have uncovered coordination between transcriptional and morphogen cues to specify Merkel cells, poorly understood skin cells that mediate light touch sensations. In murine dorsal skin, Merkel cells are part of touch domes, which are skin structures consisting of specialized keratinocytes, Merkel cells, and afferent neurons, and are located exclusively around primary hair follicles. We show that the developing primary hair follicle functions as a niche required for Merkel cell specification. We find that intraepidermal Sonic hedgehog (Shh) signaling, initiated by the production of Shh ligand in the developing hair follicles, is required for Merkel cell specification. The importance of Shh for Merkel cell formation is further reinforced by the fact that Shh overexpression in embryonic epidermal progenitors leads to ectopic Merkel cells. Interestingly, Shh signaling is common to primary, secondary, and tertiary hair follicles, raising the possibility that there are restrictive mechanisms that regulate Merkel cell specification exclusively around primary hair follicles. Indeed, we find that loss of Polycomb repressive complex 2 (PRC2) in the epidermis results in the formation of ectopic Merkel cells that are associated with all hair types. We show that PRC2 loss expands the field of epidermal cells competent to differentiate into Merkel cells through the upregulation of key Merkel-differentiation genes, which are known PRC2 targets. Importantly, PRC2-mediated repression of the Merkel cell differentiation program requires inductive Shh signaling to form mature Merkel cells. Our study exemplifies how the interplay between epigenetic and morphogen cues regulates the complex patterning and formation of the mammalian skin structures. PMID:27414999

  1. Sonic hedgehog acts as a negative regulator of {beta}-catenin signaling in the adult tongue epithelium.

    PubMed

    Schneider, Fabian T; Schänzer, Anne; Czupalla, Cathrin J; Thom, Sonja; Engels, Knut; Schmidt, Mirko H H; Plate, Karl H; Liebner, Stefan

    2010-07-01

    Wnt/beta-catenin signaling has been implicated in taste papilla development; however, its role in epithelial maintenance and tumor progression in the adult tongue remains elusive. We show Wnt/beta-catenin pathway activation in reporter mice and by nuclear beta-catenin staining in the epithelium and taste papilla of adult mouse and human tongues. beta-Catenin activation in APC(min/+) mice, which carry a mutation in adenomatous poliposis coli (APC), up-regulates Sonic hedgehog (Shh) and Jagged-2 (JAG2) in the tongue epithelium without formation of squamous cell carcinoma (SCC). We demonstrate that Shh suppresses beta-catenin transcriptional activity in a signaling-dependent manner in vitro and in vivo. A similar regulation and function was observed for JAG2, suggesting that both pathways negatively regulate beta-catenin, thereby preventing SCC formation in the tongue. This was supported by reduced nuclear beta-catenin in the tongue epithelium of Patched(+/-) mice, exhibiting dominant active Shh signaling. At the invasive front of human tongue cancer, nuclear beta-catenin and Shh were increased, suggesting their participation in tumor progression. Interestingly, Shh but not JAG2 was able to reduce beta-catenin signaling in SCC cells, arguing for a partial loss of negative feedback on beta-catenin transcription in tongue cancer. We show for the first time that the putative Wnt/beta-catenin targets Shh and JAG2 control beta-catenin signaling in the adult tongue epithelium, a function that is partially lost in lingual SCC. PMID:20508033

  2. The matricellular protein CCN1/Cyr61 is a critical regulator of Sonic Hedgehog in pancreatic carcinogenesis.

    PubMed

    Haque, Inamul; De, Archana; Majumder, Monami; Mehta, Smita; McGregor, Douglas; Banerjee, Sushanta K; Van Veldhuizen, Peter; Banerjee, Snigdha

    2012-11-01

    CCN1 is a matricellular protein and a member of the CCN family of growth factors. CCN1 is associated with the development of various cancers including pancreatic ductal adenocarcinoma (PDAC). Our recent studies found that CCN1 plays a critical role in pancreatic carcinogenesis through the induction of EMT and stemness. CCN1 mRNA and protein were detected in the early precursor lesions, and their expression intensified with disease progression. However, biochemical activity and the molecular targets of CCN1 in pancreatic cancer cells are unknown. Here we show that CCN1 regulates the Sonic Hedgehog (SHh) signaling pathway, which is associated with the PDAC progression and poor prognosis. SHh regulation by CCN1 in pancreatic cancer cells is mediated through the active Notch-1. Notably, active Notch-1is recruited by CCN1 in these cells via the inhibition of proteasomal degradation results in stabilization of the receptor. We find that CCN1-induced activation of SHh signaling might be necessary for CCN1-dependent in vitro pancreatic cancer cell migration and tumorigenicity of the side population of pancreatic cancer cells (cancer stem cells) in a xenograft in nude mice. Moreover, the functional role of CCN1 could be mediated through the interaction with the αvβ3 integrin receptor. These extensive studies propose that targeting CCN1 can provide a new treatment option for patients with pancreatic cancer since blocking CCN1 simultaneously blocks two critical pathways (i.e. SHh and Notch1) associated with the development of the disease as well as drug resistance.

  3. MYB Promotes Desmoplasia in Pancreatic Cancer through Direct Transcriptional Up-regulation and Cooperative Action of Sonic Hedgehog and Adrenomedullin.

    PubMed

    Bhardwaj, Arun; Srivastava, Sanjeev K; Singh, Seema; Tyagi, Nikhil; Arora, Sumit; Carter, James E; Khushman, Moh'd; Singh, Ajay P

    2016-07-29

    Extensive desmoplasia is a prominent pathological characteristic of pancreatic cancer (PC) that not only impacts tumor development, but therapeutic outcome as well. Recently, we demonstrated a novel role of MYB, an oncogenic transcription factor, in PC growth and metastasis. Here we studied its effect on pancreatic tumor histopathology and associated molecular and biological mechanisms. Tumor-xenografts derived from orthotopic-inoculation of MYB-overexpressing PC cells exhibited far-greater desmoplasia in histological analyses compared with those derived from MYB-silenced PC cells. These findings were further confirmed by immunostaining of tumor-xenograft sections with collagen-I, fibronectin (major extracellular-matrix proteins), and α-SMA (well-characterized marker of myofibroblasts or activated pancreatic stellate cells (PSCs)). Likewise, MYB-overexpressing PC cells provided significantly greater growth benefit to PSCs in a co-culture system as compared with the MYB-silenced cells. Interrogation of deep-sequencing data from MYB-overexpressing versus -silenced PC cells identified Sonic-hedgehog (SHH) and Adrenomedullin (ADM) as two differentially-expressed genes among others, which encode for secretory ligands involved in tumor-stromal cross-talk. In-silico analyses predicted putative MYB-binding sites in SHH and ADM promoters, which was later confirmed by chromatin-immunoprecipitation. A cooperative role of SHH and ADM in growth promotion of PSCs was confirmed in co-culture by using their specific-inhibitors and exogenous recombinant-proteins. Importantly, while SHH acted exclusively in a paracrine fashion on PSCs and influenced the growth of PC cells only indirectly, ADM could directly impact the growth of both PC cells and PSCs. In summary, we identified MYB as novel regulator of pancreatic tumor desmoplasia, which is suggestive of its diverse roles in PC pathobiology. PMID:27246849

  4. Modulation of Ciliary Phosphoinositide Content Regulates Trafficking and Sonic Hedgehog Signaling Output.

    PubMed

    Chávez, Marcelo; Ena, Sabrina; Van Sande, Jacqueline; de Kerchove d'Exaerde, Alban; Schurmans, Stéphane; Schiffmann, Serge N

    2015-08-10

    Ciliary transport is required for ciliogenesis, signal transduction, and trafficking of receptors to the primary cilium. Mutations in inositol polyphosphate 5-phosphatase E (INPP5E) have been associated with ciliary dysfunction; however, its role in regulating ciliary phosphoinositides is unknown. Here we report that in neural stem cells, phosphatidylinositol 4-phosphate (PI4P) is found in high levels in cilia whereas phosphatidylinositol (4,5)-bisphosphate (PI(4,5)P2) is not detectable. Upon INPP5E inactivation, PI(4,5)P2 accumulates at the ciliary tip whereas PI4P is depleted. This is accompanied by recruitment of the PI(4,5)P2-interacting protein TULP3 to the ciliary membrane, along with Gpr161. This results in an increased production of cAMP and a repression of the Shh transcription gene Gli1. Our results reveal the link between ciliary regulation of phosphoinositides by INPP5E and Shh regulation via ciliary trafficking of TULP3/Gpr161 and also provide mechanistic insight into ciliary alterations found in Joubert and MORM syndromes resulting from INPP5E mutations.

  5. Sonic hedgehog: restricted expression and limb dysmorphologies

    PubMed Central

    Hill, Robert E; Heaney, Simon JH; Lettice, Laura A

    2003-01-01

    Sonic hedgehog, SHH, is required for patterning the limb. The array of skeletal elements that compose the hands and feet, and the ordered arrangement of these bones to form the pattern of fingers and toes are dependent on SHH. The mechanism of action of SHH in the limb is not fully understood; however, an aspect that appears to be important is the localized, asymmetric expression of Shh. Shh is expressed in the posterior margin of the limb bud in a region defined as the zone of polarizing activity (ZPA). Analysis of mouse mutants which have polydactyly (extra toes) shows that asymmetric expression of Shh is lost due to the appearance of an ectopic domain of expression in the anterior limb margin. One such polydactylous mouse mutant, sasquatch (Ssq), maps to the corresponding chromosomal region of the human condition pre-axial polydactyly (PPD) and thus represents a model for this condition. The mutation responsible for Ssq is located 1 Mb away from the Shh gene; however, the mutation disrupts a long-range cis-acting regulator of Shh expression. By inference, human pre-axial polydactyly results from a similar disruption of Shh expression. Other human congenital abnormalities also map near the pre-axial polydactyly locus, suggesting a major chromosomal region for limb dysmorphologies. The distinct phenotypes range from loss of all bones of the hands and feet to syndactyly of the soft tissue and fusion of the digits. We discuss the role played by Shh expression in mouse mutant phenotypes and the human limb dysmorphologies. PMID:12587915

  6. Sonic hedgehog: restricted expression and limb dysmorphologies.

    PubMed

    Hill, Robert E; Heaney, Simon J H; Lettice, Laura A

    2003-01-01

    Sonic hedgehog, SHH, is required for patterning the limb. The array of skeletal elements that compose the hands and feet, and the ordered arrangement of these bones to form the pattern of fingers and toes are dependent on SHH. The mechanism of action of SHH in the limb is not fully understood; however, an aspect that appears to be important is the localized, asymmetric expression of Shh. Shh is expressed in the posterior margin of the limb bud in a region defined as the zone of polarizing activity (ZPA). Analysis of mouse mutants which have polydactyly (extra toes) shows that asymmetric expression of Shh is lost due to the appearance of an ectopic domain of expression in the anterior limb margin. One such polydactylous mouse mutant, sasquatch (Ssq), maps to the corresponding chromosomal region of the human condition pre-axial polydactyly (PPD) and thus represents a model for this condition. The mutation responsible for Ssq is located 1 Mb away from the Shh gene; however, the mutation disrupts a long-range cis-acting regulator of Shh expression. By inference, human pre-axial polydactyly results from a similar disruption of Shh expression. Other human congenital abnormalities also map near the pre-axial polydactyly locus, suggesting a major chromosomal region for limb dysmorphologies. The distinct phenotypes range from loss of all bones of the hands and feet to syndactyly of the soft tissue and fusion of the digits. We discuss the role played by Shh expression in mouse mutant phenotypes and the human limb dysmorphologies.

  7. Mammalian Gup1, a homolog of Saccharomyces cerevisiae glycerol uptake/transporter 1, acts as a negative regulator for N-terminal palmitoylation of Sonic hedgehog.

    PubMed

    Abe, Yoichiro; Kita, Yoshiko; Niikura, Takako

    2008-01-01

    Mammalian glycerol uptake/transporter 1 (Gup1), a homolog of Saccharomyces cerevisiae Gup1, is predicted to be a member of the membrane-bound O-acyltransferase family and is highly homologous to mammalian hedgehog acyltransferase, known as Skn, the homolog of the Drosophila skinny hedgehog gene product. Although mammalian Gup1 has a sequence conserved among the membrane-bound O-acyltransferase family, the histidine residue in the motif that is indispensable to the acyltransferase activity of the family has been replaced with leucine. In this study, we cloned Gup1 cDNA from adult mouse lung and examined whether Gup1 is involved in the regulation of N-terminal palmitoylation of Sonic hedgehog (Shh). Subcellular localization of mouse Gup1 was indistinguishable from that of mouse Skn detected using the fluorescence of enhanced green fluorescent protein that was fused to each C terminus of these proteins. Gup1 and Skn were co-localized with an endoplasmic reticulum marker, 78 kDa glucose-regulated protein, suggesting that these two molecules interact with overlapped targets, including Shh. In fact, full-length Shh coprecipitated with FLAG-tagged Gup1 by immunoprecipitation using anti-FLAG IgG. Ectopic expression of Gup1 with full-length Shh in cells lacking endogenous Skn showed no hedgehog acyltransferase activity as determined using the monoclonal antibody 5E1, which was found to recognize the palmitoylated N-terminal signaling domain of Shh under denaturing conditions. On the other hand, Gup1 interfered with the palmitoylation of Shh catalyzed by endogenous Skn in COS7 and NSC34. These results suggest that Gup1 is a negative regulator of N-terminal palmitoylation of Shh and may contribute to the variety of biological actions of Shh.

  8. The regulation of tooth morphogenesis is associated with epithelial cell proliferation and the expression of Sonic hedgehog through epithelial-mesenchymal interactions

    SciTech Connect

    Ishida, Kentaro; Murofushi, Mayumi; Nakao, Kazuhisa; Morita, Ritsuko; Ogawa, Miho; Tsuji, Takashi

    2011-02-18

    Research highlights: {yields} Bioengineered teeth regulated the contact area of epithelium and mesenchyme. {yields} The crown width is regulated by the contact area of the epithelium and mesenchyme. {yields} This regulation is associated with cell proliferation and Sonic hedgehog expression. {yields} The cusp number is correlated with the crown width of the bioengineered tooth. {yields} Cell proliferation and Shh expression areas regulate the tooth morphogenesis. -- Abstract: Ectodermal organs, such as the tooth, salivary gland, hair, and mammary gland, develop through reciprocal epithelial-mesenchymal interactions. Tooth morphologies are defined by the crown width and tooth length (macro-morphologies), and by the number and locations of the cusp and roots (micro-morphologies). In our current study, we report that the crown width of a bioengineered molar tooth, which was reconstructed using dissociated epithelial and mesenchymal cells via an organ germ method, can be regulated by the contact area between epithelial and mesenchymal cell layers. We further show that this is associated with cell proliferation and Sonic hedgehog (Shh) expression in the inner enamel epithelium after the germ stage has formed a secondary enamel knot. We also demonstrate that the cusp number is significantly correlated with the crown width of the bioengineered tooth. These findings suggest that the tooth micro-morphology, i.e. the cusp formation, is regulated after the tooth width, or macro-morphology, is determined. These findings also suggest that the spatiotemporal patterning of cell proliferation and the Shh expression areas in the epithelium regulate the crown width and cusp formation of the developing tooth.

  9. Sonic hedgehog, the penis and erectile dysfunction: a review of sonic hedgehog signaling in the penis.

    PubMed

    Podlasek, C A; Meroz, C L; Korolis, H; Tang, Y; McKenna, K E; McVary, K T

    2005-01-01

    The sinusoid anatomy of the penis is complex and requires complicated interaction between smooth muscle and endothelium in order to maintain homeostasis in the adult. The morphogen, Sonic hedgehog (Shh), is a crucial regulator of these processes, along with its down stream targets patched (Ptc), Hox, bone morphogenetic proteins (BMP's), vascular endothelial growth factor (VEGF) and nitric oxide synthase (NOS). Shh is critical for patterning and establishing tissue identity of the penis during embryonic development, is a crucial regulator of penile postnatal differentiation of the sinusoid morphology of the corpora cavernosa, and plays a fundamental role in maintaining sinusoidal structures pertinent to erectile function in the adult rat. Shh and its targets are active in human penes, and decreased in human diabetic penes in parallel with observations in the rat, thus lending clinical significance to the role of abnormal Shh signaling in erectile dysfunction (ED). Application of exogenous Shh protein to rat corpora cavernosa, induces VEGF and NOS proteins, suggesting a potential mechanism through which decreased Shh protein can cause ED. The studies outlined in this review provide in depth analysis of the Shh pathway and signal transduction, its role in penile development, how Shh signaling is altered in a rat model of ED and neuropathy, how abnormal Shh signaling can cause ED, and the clinical significance of the Shh pathway to human ED. These studies will provide valuable insight, at the molecular level, into understanding the mechanisms that under lie ED and lead to new treatment strategies for diabetic impotence. PMID:16378507

  10. Developmental pathways: Sonic hedgehog-Patched-GLI.

    PubMed Central

    Walterhouse, D O; Yoon, J W; Iannaccone, P M

    1999-01-01

    Developmental pathways are networks of genes that act coordinately to establish the body plan. Disruptions of genes in one pathway can have effects in related pathways and may result in serious dysmorphogenesis or cancer. Environmental exposures can be associated with poor pregnancy outcomes, including dysmorphic offspring or children with a variety of diseases. An important goal of environmental science should be reduction of these poor outcomes. This will require an understanding of the genes affected by specific exposures and the consequence of alterations in these genes or their products, which in turn will require an understanding of the pathways critical in development. The ligand Sonic hedgehog, the receptors Patched and Smoothened, and the GLI family of transcription factors represent one such pathway. This pathway illustrates several operating principles important in the consideration of developmental consequences of environmental exposures to toxins. Images Figure 1 Figure 2 PMID:10064544

  11. Expression of Sonic hedgehog and retinal opsin genes in experimentally-induced myopic chick eyes.

    PubMed

    Escaño, M F; Fujii, S; Sekiya, Y; Yamamoto, M; Negi, A

    2000-11-01

    The purpose of this study was to evaluate changes in the expression of different genes in chick retinal tissues after induction of experimental myopia and to evaluate the roles of these genes in the regulation of postnatal eye growth and myopia. Form-deprivation using occlusive goggles and hyperopic defocus by negative spectacle lenses were used to induce myopia in hatched chicks. Expression levels of Sonic hedgehog, its receptor complex, and other retinal cell genes were evaluated by semi-quantitative reverse transcription-polymerase chain reaction. Levels of Sonic hedgehog protein were further evaluated by Western blot analysis. The induction of myopia caused significant increase in expression of Sonic hedgehog mRNA and protein and increased expression of blue and red opsin mRNA. In contrast, the expression of mRNA for Sonic hedgehog receptor complex (Patched-Smoothened), rhodopsin, vimentin, green opsin, violet opsin, and HPC-1 were unaffected by the induction of myopia. The increase in expression of Sonic hedgehog in chick retinas in experimentally-induced myopia suggests involvement in the retina control of postnatal eye growth. Furthermore, Sonic hedgehog may influence the expression of blue and red opsins under myopic conditions.

  12. Probing extracellular Sonic hedgehog in neurons

    PubMed Central

    Eitan, Erez; Petralia, Ronald S.; Wang, Ya-Xian; Indig, Fred E.; Mattson, Mark P.

    2016-01-01

    ABSTRACT The bioactivity of Sonic hedgehog (Shh) depends on specific lipid modifications; a palmitate at its N-terminus and a cholesterol at its C-terminus. This dual-lipid modification makes Shh molecules lipophilic, which prevents them from diffusing freely in extracellular space. Multiple lines of evidence indicate that Shh proteins are carried by various forms of extracellular vesicles (EVs). It also has been shown, for instance, that in some tissues Shh proteins are transported to neighboring cells directly via filopodia. We have previously reported that Shh proteins are expressed in hippocampal neurons. In this study we show that, in the hippocampus and cerebellum of postnatal day (P)2 rats, Shh is mostly found near or on the membrane surface of small neurites or filopodia. We also examined cultured hippocampal neurons where we observed noticeable and widespread Shh-immunolabeled vesicles located outside neurons. Through immunoelectron microscopy and biochemical analysis, we find Shh-containing EVs with a wide range of sizes. Unlike robust Shh activity in EVs isolated from cells overexpressing an N-terminal Shh fragment construct, we did not detect measurable Shh activity in EVs purified from the medium of cultured hippocampal neurons. These results suggest the complexity of the transcellular Shh signaling mechanisms in neurons. PMID:27387534

  13. Probing extracellular Sonic hedgehog in neurons.

    PubMed

    Eitan, Erez; Petralia, Ronald S; Wang, Ya-Xian; Indig, Fred E; Mattson, Mark P; Yao, Pamela J

    2016-01-01

    The bioactivity of Sonic hedgehog (Shh) depends on specific lipid modifications; a palmitate at its N-terminus and a cholesterol at its C-terminus. This dual-lipid modification makes Shh molecules lipophilic, which prevents them from diffusing freely in extracellular space. Multiple lines of evidence indicate that Shh proteins are carried by various forms of extracellular vesicles (EVs). It also has been shown, for instance, that in some tissues Shh proteins are transported to neighboring cells directly via filopodia. We have previously reported that Shh proteins are expressed in hippocampal neurons. In this study we show that, in the hippocampus and cerebellum of postnatal day (P)2 rats, Shh is mostly found near or on the membrane surface of small neurites or filopodia. We also examined cultured hippocampal neurons where we observed noticeable and widespread Shh-immunolabeled vesicles located outside neurons. Through immunoelectron microscopy and biochemical analysis, we find Shh-containing EVs with a wide range of sizes. Unlike robust Shh activity in EVs isolated from cells overexpressing an N-terminal Shh fragment construct, we did not detect measurable Shh activity in EVs purified from the medium of cultured hippocampal neurons. These results suggest the complexity of the transcellular Shh signaling mechanisms in neurons. PMID:27387534

  14. Sonic hedgehog signaling coordinates the proliferation and differentiation of neural stem/progenitor cells by regulating cell cycle kinetics during development of the neocortex.

    PubMed

    Komada, Munekazu

    2012-06-01

    Sonic hedgehog (Shh) acts as a morphogen in normal development of various vertebrate tissues and organs. Shh signaling is essential for patterning and cell-fate specification, particularly in the central nervous system. Shh signaling plays different roles depending on its concentration, area, and timing of exposure. During the development of the neocortex, a low level of Shh is expressed in the neural stem/progenitor cells as well as in mature neurons in the dorsal telencephalon. Shh signaling in neocortex development has been shown to regulate cell cycle kinetics of radial glial cells and intermediate progenitor cells, thereby maintaining the proliferation, survival and differentiation of neurons in the neocortex. During the development of the telencephalon, endogenous Shh signaling is involved in the transition of slow-cycling neural stem cells to fast-cycling neural progenitor cells. It seems that high-level Shh signaling in the ventral telencephalon is essential for ventral specification, while low-level Shh signaling in the dorsal telencephalon plays important roles in the fine-tuning of cell cycle kinetics. The Shh levels and multiple functions of Shh signaling are important for proper corticogenesis in the developing brain. The present paper discusses the roles of Shh signaling in the proliferation and differentiation of neural stem/progenitor cells.

  15. The Role of Sonic Hedgehog Reemergence During Gastric Cancer

    PubMed Central

    Martin, Jason; Donnelly, Jessica M.; Houghton, JeanMarie

    2016-01-01

    Sonic Hedgehog (Shh) signaling has been extensively studied for its role in developmental biology and cancer biology. The association between Shh and cancer development in general is well established but the functional role of Shh in the development and progression of gastric cancer specifically is largely unknown. Bone marrow-derived stem cells, specifically mesenchymal stem cells (MSCs) infiltrate and engraft into the gastric mucosa in response to the chronic inflammatory environment of Helicobacter infection. In this review, MSC infiltration and changes in the cytokine and cellular profiles of later-stage chronic environments will be tied into their interactions with the Shh pathway. We will discuss how these changes shape tumorigenesis and tumor progression in the gastric mucosa. The current review focuses on the Shh signaling pathway and its role in the development of gastric cancer, specifically in response to Helicobacter pylori infection. We follow with an in-depth discussion of the regulation of the Hedgehog pathway during acute and chronic gastric inflammation with a focus on signaling within the MSC compartment. PMID:20437100

  16. MicroRNA-602 and microRNA-608 regulate sonic hedgehog expression via target sites in the coding region in human chondrocytes

    PubMed Central

    Akhtar, Nahid; Makki, Mohammad Shahidul; Haqqi, Tariq M.

    2015-01-01

    Objective Hedgehog(Hh) signaling has recently been associated with cartilage degradation in osteoarthritis(OA). As interleukin-1β(IL-1β) is a critical mediator of OA pathogenesis, here we determined whether IL-1β induces the expression of sonic hedgehog(SHH) and its regulation by microRNAs in human chondrocytes. Methods SHH protein expression in human OA-cartilage and in an animal model of OA was determined by immunohistochemistry and immunofluorescence respectively. Gene and protein expression in IL-1β or SHH-stimulated chondrocytes was determined by TaqMan assays and immunoblotting respectively. Effect of overexpression of miR-602 and miR-608 or their anatgomirs on SHH expression was evaluated by transient transfections of human chondrocytes and HEK-293 cells. Role of signaling pathways was evaluated using small molecule inhibitors. Binding of miRNAs with the putative “seed sequence” in the SHH mRNA was validated with a SHH luciferase reporter assay. Results Expression of SHH, PTCH-1, GLI-1, HHIP, MMP-13, and COL10A1 was high in damaged OAcartilage. Expression of SHH was inversely correlated with the expression of miR-608 in damaged cartilage and in IL-1β-stimulated chondrocytes. Transfection with miR-608 or miR-602 mimics inhibited the reporter activity and mutation of the miRNAs “seed sequences” abolished the repression of reporter activity. Overexpression of miR-602 or miR-608 inhibited the expression of SHH mRNA and protein and this was abrogated by antagomirs. Stimulation with SHH-protein up-regulated the MMP-13 expression and inhibition of Hh signaling blocked MMP-13 expression in OA chondrocytes. Conclusions miR-602 and miR-608 are important regulators of SHH expression in chondrocytes and their suppression by IL-1β may contribute to the enhanced expression of SHH and MMP-13 in OA. PMID:25385442

  17. Sonic hedgehog protein regulates fibroblast growth factor 8 expression in metanephric explant culture from BALB/c mice: Possible mechanisms associated with renal morphogenesis

    PubMed Central

    Chen, Xing; Hou, Xiao-Ming; Fan, You-Fei; Jin, Yu-Ting; Wang, Yu-Lin

    2016-01-01

    The sonic hedgehog (SHH) morphogen regulates cell differentiation and controls a number of genes during renal morphogenesis. To date, the effects of SHH on fibroblast growth factors (Fgfs) in embryonic kidney development remain unclear. In the present study, explants of BALB/c mouse embryonic kidney tissues were used to investigate the role of exogenous SHH on Fgf8 and Fgf10 expression levels ex vivo. Ureteric bud branches and epithelial metanephric derivatives were used to determine the renal morphogenesis with Dolichos biflorus agglutinin or hematoxylineosin staining. mRNA expression levels were determined using reverse transcription-quantitative polymerase chain reaction, while the protein expression levels were examined using immunohistochemistry and western blot analysis. During the initial stages of metanephric development, low levels of SHH, Fgf8, and Fgf10 expression were observed, which were found to increase significantly during more advanced stages of metanephric development. In addition, exogenous SHH protein treatment increased the number of ureteric bud branches and enhanced the formation of nephrons. Exogenous SHH reduced the Fgf8 mRNA and protein expression levels, whereas cyclopamine (an SHH-smoothened receptor inhibitor) interfered with SHH-mediated downregulation of Fgf8 expression. By contrast, exogenous SHH protein was not found to modulate Fgf10 mRNA and protein expression levels. In conclusion, these results indicate that the modulatory effects of SHH on BALB/c mouse metanephric explant cultures may involve the regulation of Fgf8 expression but not Fgf10 expression, which provides evidence for the functional role of Fgf proteins in renal morphogenesis. PMID:27510750

  18. Sonic hedgehog protein regulates fibroblast growth factor 8 expression in metanephric explant culture from BALB/c mice: Possible mechanisms associated with renal morphogenesis.

    PubMed

    Chen, Xing; Hou, Xiao-Ming; Fan, You-Fei; Jin, Yu-Ting; Wang, Yu-Lin

    2016-10-01

    The sonic hedgehog (SHH) morphogen regulates cell differentiation and controls a number of genes during renal morphogenesis. To date, the effects of SHH on fibroblast growth factors (Fgfs) in embryonic kidney development remain unclear. In the present study, explants of BALB/c mouse embryonic kidney tissues were used to investigate the role of exogenous SHH on Fgf8 and Fgf10 expression levels ex vivo. Ureteric bud branches and epithelial metanephric derivatives were used to determine the renal morphogenesis with Dolichos biflorus agglutinin or hematoxylin‑eosin staining. mRNA expression levels were determined using reverse transcription‑quantitative polymerase chain reaction, while the protein expression levels were examined using immunohistochemistry and western blot analysis. During the initial stages of metanephric development, low levels of SHH, Fgf8, and Fgf10 expression were observed, which were found to increase significantly during more advanced stages of metanephric development. In addition, exogenous SHH protein treatment increased the number of ureteric bud branches and enhanced the formation of nephrons. Exogenous SHH reduced the Fgf8 mRNA and protein expression levels, whereas cyclopamine (an SHH‑smoothened receptor inhibitor) interfered with SHH‑mediated downregulation of Fgf8 expression. By contrast, exogenous SHH protein was not found to modulate Fgf10 mRNA and protein expression levels. In conclusion, these results indicate that the modulatory effects of SHH on BALB/c mouse metanephric explant cultures may involve the regulation of Fgf8 expression but not Fgf10 expression, which provides evidence for the functional role of Fgf proteins in renal morphogenesis. PMID:27510750

  19. The Sonic hedgehog gradient in the developing limb.

    PubMed

    Tickle, Cheryll; Barker, Heather

    2013-01-01

    A gradient of Sonic hedgehog (Shh) plays a major role in specifying the antero-posterior pattern of structures that develop in the distal part of the vertebrate limb, in particular, the antero-posterior pattern of the digits. Classical embryological experiments identified the polarizing region (or zone of polarizing activity, ZPA), a signaling region at the posterior margin of the early chick wing bud and, consistent with a model in which production of a diffusible morphogen specifies antero-posterior positional information, polarizing region signaling was shown to be dose dependent and long range. It is now well established that the vertebrate hedgehog gene, Sonic hedgehog (Shh), which encodes a secreted protein, is expressed in the polarizing region of the chick wing and that Shh signaling has the same characteristics as polarizing region signaling. Shh expression at the posterior of the early limb bud and the mechanism of Shh signal transduction are conserved among vertebrates including mammals. However, it is unlikely that a simple Shh gradient is responsible for digit pattern formation in mammalian limbs and there is still little understanding of how positional information specified by Shh signaling is encoded and translated into digit anatomy. Alterations in Shh signaling underlie some congenital limb abnormalities and also changes in timing and extent of Shh signaling appear to be related to the evolution of morphological diversity of vertebrate limbs.

  20. Sonic Hedgehog Signalling Pathway and Ameloblastoma - A Review.

    PubMed

    Mishra, Pallavi; Panda, Abikshyeet; Bandyopadhyay, Alokenath; Kumar, Harish; Mohiddin, Gouse

    2015-11-01

    Ameloblastoma is a benign but aggressive odontogenic neoplasm arising from odontogenic epithelium. Many theories have been proposed to explain the pathogenesis of ameloblatoma. Numerous signalling pathways have been implicated to be associated in the development and progression of this neoplasm. Studies have found association of various signalling molecules of Sonic Hedgehog Pathway, namely SHH, PTCH1, SMO, Gli 1, Gli 2, Gli 3, with ameloblastoma. Knowledge about this pathway will help us to understand the nature and behaviour of this neoplasm. This will open the door towards new treatment modalities. PMID:26674664

  1. Sonic Hedgehog Signalling Pathway and Ameloblastoma – A Review

    PubMed Central

    Mishra, Pallavi; Bandyopadhyay, Alokenath; Kumar, Harish; Mohiddin, Gouse

    2015-01-01

    Ameloblastoma is a benign but aggressive odontogenic neoplasm arising from odontogenic epithelium. Many theories have been proposed to explain the pathogenesis of ameloblatoma. Numerous signalling pathways have been implicated to be associated in the development and progression of this neoplasm. Studies have found association of various signalling molecules of Sonic Hedgehog Pathway, namely SHH, PTCH1, SMO, Gli 1, Gli 2, Gli 3, with ameloblastoma. Knowledge about this pathway will help us to understand the nature and behaviour of this neoplasm. This will open the door towards new treatment modalities. PMID:26674664

  2. 14-3-3 proteins regulate a cell-intrinsic switch from sonic hedgehog-mediated commissural axon attraction to repulsion after midline crossing.

    PubMed

    Yam, Patricia T; Kent, Christopher B; Morin, Steves; Farmer, W Todd; Alchini, Ricardo; Lepelletier, Léa; Colman, David R; Tessier-Lavigne, Marc; Fournier, Alyson E; Charron, Frédéric

    2012-11-21

    Axons must switch responsiveness to guidance cues during development for correct pathfinding. Sonic Hedgehog (Shh) attracts spinal cord commissural axons ventrally toward the floorplate. We show that after crossing the floorplate, commissural axons switch their response to Shh from attraction to repulsion, so that they are repelled anteriorly by a posterior-high/anterior-low Shh gradient along the longitudinal axis. This switch is recapitulated in vitro with dissociated commissural neurons as they age, indicating that the switch is intrinsic and time dependent. 14-3-3 protein inhibition converted Shh-mediated repulsion of aged dissociated neurons to attraction and prevented the correct anterior turn of postcrossing commissural axons in vivo, an effect mediated through PKA. Conversely, overexpression of 14-3-3 proteins was sufficient to drive the switch from Shh-mediated attraction to repulsion both in vitro and in vivo. Therefore, we identify a 14-3-3 protein-dependent mechanism for a cell-intrinsic temporal switch in the polarity of axon turning responses.

  3. Anorectal Malformations Caused by Defects in Sonic Hedgehog Signaling

    PubMed Central

    Mo, Rong; Kim, Jae Hong; Zhang, Jianrong; Chiang, Chin; Hui, Chi-chung; Kim, Peter C. W.

    2001-01-01

    Anorectal malformations are a common clinical problem affecting the development of the distal hindgut in infants. The spectrum of anorectal malformations ranges from the mildly stenotic anus to imperforate anus with a fistula between the urinary and intestinal tracts to the most severe form, persistent cloaca. The etiology, embryology, and pathogenesis of anorectal malformations are poorly understood and controversial. Sonic hedgehog (Shh) is an endoderm-derived signaling molecule that induces mesodermal gene expression in the chick hindgut. However, the role of Shh signaling in mammalian hindgut development is unknown. Here, we show that mutant mice with various defects in the Shh signaling pathway exhibit a spectrum of distal hindgut defects mimicking human anorectal malformations. Shh null-mutant mice display persistent cloaca. Mutant mice lacking Gli2 or Gli3, two zinc finger transcription factors involved in Shh signaling, respectively, exhibit imperforate anus with recto-urethral fistula and anal stenosis. Furthermore, persistent cloaca is also observed in Gli2−/−;Gli3+/−, Gli2+/−;Gli3−/−, and Gli2−/−;Gli3−/− mice demonstrating a gene dose-dependent effect. Therefore, Shh signaling is essential for normal development of the distal hindgut in mice and mutations affecting Shh signaling produce a spectrum of anorectal malformations that may reveal new insights into their human disease equivalents. PMID:11485934

  4. Ontogenetic Expression of Sonic Hedgehog in the Chicken Subpallium

    PubMed Central

    Bardet, Sylvia M.; Ferran, José L. E.; Sanchez-Arrones, Luisa; Puelles, Luis

    2010-01-01

    Sonic hedgehog (SHH) is a secreted signaling factor that is implicated in the molecular patterning of the central nervous system (CNS), somites, and limbs in vertebrates. SHH has a crucial role in the generation of ventral cell types along the entire rostrocaudal axis of the neural tube. It is secreted early in development by the axial mesoderm (prechordal plate and notochord) and the overlying ventral neural tube. Recent studies clarified the impact of SHH signaling mechanisms on dorsoventral patterning of the spinal cord, but the corresponding phenomena in the rostral forebrain are slightly different and more complex. This notably involves separate Shh expression in the preoptic part of the forebrain alar plate, as well as in the hypothalamic floor and basal plates. The present work includes a detailed spatiotemporal description of the singular alar Shh expression pattern in the rostral preoptic forebrain of chick embryos, comparing it with FoxG1, Dlx5, Nkx2.1, and Nkx2.2 mRNA expression at diverse stages of development. As a result of this mapping, we report a subdivision of the preoptic region in dorsal and ventral zones; only the dorsal part shows Shh expression. The positive area impinges as well upon a median septocommissural preoptic domain. Our study strongly suggests tangential migration of Shh-positive cells from the preoptic region into other subpallial domains, particularly into the pallidal mantle and the intermediate septum. PMID:20700498

  5. Sonic Hedgehog Promotes Cementoblastic Differentiation via Activating the BMP Pathways.

    PubMed

    Bae, Won-Jung; Auh, Q-Schick; Lim, Hyun-Chang; Kim, Gyu-Tae; Kim, Hyun-Soo; Kim, Eun-Cheol

    2016-10-01

    Although sonic hedgehog (SHH), an essential molecule in embryogenesis and organogenesis, stimulates proliferation of human periodontal ligament (PDL) stem cells, the effects of recombinant human SHH (rh-SHH) on osteoblastic differentiation are unclear. To reveal the role of SHH in periodontal regeneration, expression of SHH in mouse periodontal tissues and its effects on the osteoblastic/cementoblastic differentiation in human cementoblasts were investigated. SHH is immunolocalized to differentiating cementoblasts, PDL cells, and osteoblasts of the developing mouse periodontium. Addition of rh-SHH increased cell growth, ALP activity, and mineralization nodule formation, and upregulated mRNA expression of osteoblastic and cementoblastic markers. The osteoblastic/cementoblastic differentiation of rh-SHH was abolished by the SHH inhibitor cyclopamine (Cy) and the BMP antagonist noggin. rh-SHH increased the expression of BMP-2 and -4 mRNA, as well as levels of phosphorylated Akt, ERK, p38, and JNK, and of MAPK and NF-κB activation, which were reversed by noggin, Cy, and BMP-2 siRNA. Collectively, this study is the first to demonstrate that SHH can promote cell growth and cell osteoblastic/cementoblastic differentiation via BMP pathway. Thus, SHH plays important roles in the development of periodontal tissue, and might represent a new therapeutic target for periodontitis and periodontal regeneration. PMID:27289556

  6. Sonic hedgehog (shh) expression in developing and regenerating axolotl limbs.

    PubMed

    Torok, M A; Gardiner, D M; Izpisúa-Belmonte, J C; Bryant, S V

    1999-07-01

    Sonic hedgehog (shh) expression is detectable in the posterior mesenchyme of many developing vertebrate limbs. We have isolated an RT-PCR fragment from the axolotl, Ambystoma mexicanum, that has high identity to other vertebrate shh genes. We describe the localization of this transcript during development and regeneration and in response to tissue grafts and retinoic acid (RA) exposure in the axolotl. Even though axolotl digits show a reversed polarity of differentiation (anterior [A] to posterior [P]) when compared to other tetrapods (P to A), shh is nevertheless expressed on the posterior margin of developing and regenerating limb buds. When A cells are grafted adjacent to P cells, an ectopic domain of shh is induced. Exposure to retinoic acid (RA), a molecule known to alter pattern in all three limb axes in urodeles, results in ectopic expression of shh in anterior cells of the regeneration blastema. Prior to this induced expression in response to RA, there is an earlier response by the endogenous domain of shh, which is downregulated within the first few hours of exposure. PMID:10404648

  7. Sonic hedgehog in oral squamous cell carcinoma: An immunohistochemical study

    PubMed Central

    Srinath, Sahana; Iyengar, Asha R; Mysorekar, Vijaya

    2016-01-01

    Background: Recent studies have revealed the involvement of hedgehog (Hh) signaling component in proliferation and invasive behavior of many carcinomas. Aim: This study aims to identify the expression of sonic Hh (SHH) protein of SHH pathway in oral epithelial dysplasia and oral squamous cell carcinoma (OSCC) using SHH (H-160) (Santa Cruz, sc-9042) which could have therapeutic implication in future. Materials and Methods: A total of 250 cases comprising 50 normal oral mucosa, 50 cases of oral epithelial dysplasia, 50 well, 50 moderate and 50 poorly differentiated OSCCs were included in the study. Immunohistochemical evaluation of SHH protein expression was conducted using monoclonal antibody. Interpretation of the expression was done by immunoreactive score of Remmele and Stegner (IRS) scoring method. Statistical Analysis: Chi-Square test was used to analyze the results. Results: The study showed that SHH signaling molecules are highly expressed in OSCC, and their expression was mainly in the cytoplasm of epithelial cells. Conclusion: The SHH signaling component is associated with the pathological parameter in OSCC and oral epithelial dysplasia. PMID:27721600

  8. A critical role for sonic hedgehog signaling in the early expansion of the developing brain.

    PubMed

    Britto, Joanne; Tannahill, David; Keynes, Roger

    2002-02-01

    The mechanisms that coordinate the three-dimensional shape of the vertebrate brain during development are largely unknown. We have found that sonic hedgehog (Shh) is crucial in driving the rapid, extensive expansion of the early vesicles of the developing midbrain and forebrain. Transient displacement of the notochord from the midbrain floor plate resulted in abnormal folding and overall collapse of the vesicles, accompanied by reduced cell proliferation and increased cell death in the midbrain. Simultaneously, expression of Shh decreased locally in the notochord and floor plate, whereas overt patterning and differentiation proceeded normally. Normal midbrain expansion was restored by implantation of Shh-secreting cells in a dose-dependent manner; conversely, expansion was retarded following antagonism of the Shh signaling pathway by cyclopamine. Our results indicate that Shh signaling from the ventral midline is essential for regulating brain morphogenesis during early development.

  9. Notch Activity Modulates the Responsiveness of Neural Progenitors to Sonic Hedgehog Signaling

    PubMed Central

    Kong, Jennifer H.; Yang, Linlin; Dessaud, Eric; Chuang, Katherine; Moore, Destaye M.; Rohatgi, Rajat; Briscoe, James; Novitch, Bennett G.

    2015-01-01

    Summary Throughout the developing nervous system, neural stem and progenitor cells give rise to diverse classes of neurons and glia in a spatially and temporally coordinated manner. In the ventral spinal cord, much of this diversity emerges through the morphogen actions of Sonic hedgehog (Shh). Interpretation of the Shh gradient depends on both the amount of ligand and duration of exposure, but the mechanisms permitting prolonged responses to Shh are not well understood. We demonstrate that Notch signaling plays an essential role in this process, enabling neural progenitors to attain sufficiently high levels of Shh pathway activity needed to direct the ventral-most cell fates. Notch activity regulates subcellular localization of the Shh receptor Patched1, gating the translocation of the key effector Smoothened to primary cilia and its downstream signaling activities. These data reveal an unexpected role for Notch shaping the interpretation of the Shh morphogen gradient and influencing cell fate determination. PMID:25936505

  10. Sonic Hedgehog-activated engineered blood vessels enhance bone tissue formation.

    PubMed

    Rivron, Nicolas C; Raiss, Christian C; Liu, Jun; Nandakumar, Anandkumar; Sticht, Carsten; Gretz, Norbert; Truckenmüller, Roman; Rouwkema, Jeroen; van Blitterswijk, Clemens A

    2012-03-20

    Large bone defects naturally regenerate via a highly vascularized tissue which progressively remodels into cartilage and bone. Current approaches in bone tissue engineering are restricted by delayed vascularization and fail to recapitulate this stepwise differentiation toward bone tissue. Here, we use the morphogen Sonic Hedgehog (Shh) to induce the in vitro organization of an endothelial capillary network in an artificial tissue. We show that endogenous Hedgehog activity regulates angiogenic genes and the formation of vascular lumens. Exogenous Shh further induces the in vitro development of the vasculature (vascular lumen formation, size, distribution). Upon implantation, the in vitro development of the vasculature improves the in vivo perfusion of the artificial tissue and is necessary to contribute to, and enhance, the formation of de novo mature bone tissue. Similar to the regenerating callus, the artificial tissue undergoes intramembranous and endochondral ossification and forms a trabecular-like bone organ including bone-marrow-like cavities. These findings open the door for new strategies to treat large bone defects by closely mimicking natural endochondral bone repair.

  11. Sonic Hedgehog: A Good Gene Gone Bad? Detection and Treatment of Genetic Abnormalities.

    ERIC Educational Resources Information Center

    Yaich, Lauren E.

    2001-01-01

    Presents a case of a baby born with the genetic condition holoprosencephaly in which students explore the "Sonic hedgehog" gene, signal transduction, and the ethics of body and tissue donation. Presents a two-part assignment that features students writing an informed consent document that explains the science behind this congenital abnormality,…

  12. Identification of a family of fatty acid-speciated Sonic Hedgehog proteins, whose members display differential biological properties

    PubMed Central

    Houel, Stephane; Rodgriguez-Blanco, Jezabel; Singh, Samer; Schilling, Neal; J.Capobianco, Anthony; Ahn, Natalie G.; Robbins, David J.

    2015-01-01

    SUMMARY Hedgehog (HH) proteins are proteolytically processed into a biologically active form, which is covalently modified by cholesterol and palmitate. However, most studies of HH biogenesis have characterized protein from cells in which HH is over-expressed. We purified Sonic Hedgehog (SHH) from cells expressing physiologically relevant levels, and showed that it was more potent than SHH isolated from over-expressing cells. Furthermore, the SHH in our preparations were modified with a diverse spectrum of fatty acids on their amino-termini, and this spectrum of fatty acids varied dramatically depending on the growth conditions of the cells. The fatty acid composition of SHH affected its trafficking to lipid rafts, as well as its potency. Our results suggest that HH proteins exist as a family of diverse lipid-speciated proteins, which might be altered in different physiological and pathological contexts to regulate distinct properties of HH proteins. PMID:25732819

  13. Targeting of sonic hedgehog-Gli signaling: A potential therapeutic target for patients with breast cancer

    PubMed Central

    Song, Lingqin; Wang, Weifeng; Liu, Di; Zhao, Yang; He, Jianjun; Wang, Xijing; Dai, Zhijun; Zhang, Huimin; Li, Xiao

    2016-01-01

    Breast cancer is the most common malignant cancer among women. The Hedgehog (Hh) signaling pathway serves a key role in malignant cancer cell growth and migration. However, little is known with regard to the specific function of the Hh signaling pathway in human breast cancer. The current study investigated the specific role of Hh signaling in the human breast cancer cell line MDA-MB-231. Expression of components of Shh-Gli signaling, as well as the Gli-responsive genes B-cell lymphoma 2 (Bcl-2) and cyclin D1, were investigated in MDA-MB-231 cells using western blotting. The effects of Shh-Gli signaling on MDA-MB-231 proliferation were analyzed by MTT assay. The role of E-cadherin in the epithelial-mesenchymal transition process was determined by western blot while matrix metalloproteinase (MMP)-9/MMP-2 secretion was studied by enzyme-linked immunosorbent assay. The results indicated that Shh-Gli signaling was activated in MDA-MB-231 cells, significantly enhancing cell viability. Overexpression of Gli positively regulated the transcription of Bcl-2 and cyclin D1 thereby regulating MDA-MB-231 cell proliferation and survival. Treatment of MDA-MB-231 cells with human sonic hedgehog, n-terminus for 72 h significantly reduced E-cadherin protein levels and enhanced secretion of MMP-9 and MMP-2. These findings suggest that Shh-Gli signaling is significantly activated in human breast cancer cells, and is accompanied by enhanced cell viability, proliferation and migration capacities. PMID:27446389

  14. Sonic hedgehog controls enteric nervous system development by patterning the extracellular matrix.

    PubMed

    Nagy, Nandor; Barad, Csilla; Graham, Hannah K; Hotta, Ryo; Cheng, Lily S; Fejszak, Nora; Goldstein, Allan M

    2016-01-15

    The enteric nervous system (ENS) develops from neural crest cells that migrate along the intestine, differentiate into neurons and glia, and pattern into two plexuses within the gut wall. Inductive interactions between epithelium and mesenchyme regulate gut development, but the influence of these interactions on ENS development is unknown. Epithelial-mesenchymal recombinations were constructed using avian hindgut mesenchyme and non-intestinal epithelium from the bursa of Fabricius. These recombinations led to abnormally large and ectopically positioned ganglia. We hypothesized that sonic hedgehog (Shh), a secreted intestinal epithelial protein not expressed in the bursa, mediates this effect. Inhibition of Shh signaling, by addition of cyclopamine or a function-blocking antibody, resulted in large, ectopic ganglia adjacent to the epithelium. Shh overexpression, achieved in ovo using Shh-encoding retrovirus and in organ culture using recombinant protein, led to intestinal aganglionosis. Shh strongly induced the expression of versican and collagen type IX, whereas cyclopamine reduced expression of these chondroitin sulfate proteoglycans that are known to be inhibitory to neural crest cell migration. Shh also inhibited enteric neural crest-derived cell (ENCC) proliferation, promoted neuronal differentiation, and reduced expression of Gdnf, a key regulator of ENS formation. Ptc1 and Ptc2 were not expressed by ENCCs, and migration of isolated ENCCs was not inhibited by Shh protein. These results suggest that epithelial-derived Shh acts indirectly on the developing ENS by regulating the composition of the intestinal microenvironment. PMID:26674309

  15. Quiescent Sox2+ Cells Drive Hierarchical Growth and Relapse in Sonic Hedgehog Subgroup Medulloblastoma

    PubMed Central

    Vanner, Robert J.; Remke, Marc; Gallo, Marco; Selvadurai, Hayden J.; Coutinho, Fiona; Lee, Lilian; Kushida, Michelle; Head, Renee; Morrissy, Sorana; Zhu, Xueming; Aviv, Tzvi; Voisin, Veronique; Clarke, Ian D.; Li, Yisu; Mungall, Andrew J.; Moore, Richard A.; Ma, Yussanne; Jones, Steven J.M.; Marra, Marco A.; Malkin, David; Northcott, Paul A.; Kool, Marcel; Pfister, Stefan M.; Bader, Gary; Hochedlinger, Konrad; Korshunov, Andrey; Taylor, Michael D.; Dirks, Peter B.

    2015-01-01

    SUMMARY Functional heterogeneity within tumors presents a significant therapeutic challenge. Here we show that quiescent, therapy-resistant Sox2+ cells propagate sonic hedgehog subgroup medulloblastoma by a mechanism that mirrors a neurogenic program. Rare Sox2+ cells produce rapidly cycling doublecortin+ progenitors that, together with their postmitotic progeny expressing NeuN, comprise tumor bulk. Sox2+ cells are enriched following anti-mitotic chemotherapy and Smoothened inhibition, creating a reservoir for tumor regrowth. Lineage traces from Sox2+ cells increase following treatment, suggesting that this population is responsible for relapse. Targeting Sox2+ cells with the antineoplastic mithramycin abrogated tumor growth. Addressing functional heterogeneity and eliminating Sox2+ cells presents a promising therapeutic paradigm for treatment of sonic hedgehog subgroup medulloblastoma. PMID:24954133

  16. Sonic hedgehog functions upstream of disrupted-in-schizophrenia 1 (disc1): implications for mental illness

    PubMed Central

    Boyd, Penelope J.; Cunliffe, Vincent T.; Roy, Sudipto; Wood, Jonathan D.

    2015-01-01

    ABSTRACT DISRUPTED-IN-SCHIZOPHRENIA (DISC1) has been one of the most intensively studied genetic risk factors for mental illness since it was discovered through positional mapping of a translocation breakpoint in a large Scottish family where a balanced chromosomal translocation was found to segregate with schizophrenia and affective disorders. While the evidence for it being central to disease pathogenesis in the original Scottish family is compelling, recent genome-wide association studies have not found evidence for common variants at the DISC1 locus being associated with schizophrenia in the wider population. It may therefore be the case that DISC1 provides an indication of biological pathways that are central to mental health issues and functional studies have shown that it functions in multiple signalling pathways. However, there is little information regarding factors that function upstream of DISC1 to regulate its expression and function. We herein demonstrate that Sonic hedgehog (Shh) signalling promotes expression of disc1 in the zebrafish brain. Expression of disc1 is lost in smoothened mutants that have a complete loss of Shh signal transduction, and elevated in patched mutants which have constitutive activation of Shh signalling. We previously demonstrated that disc1 knockdown has a dramatic effect on the specification of oligodendrocyte precursor cells (OPC) in the hindbrain and Shh signalling is known to be essential for the specification of these cells. We show that disc1 is prominently expressed in olig2-positive midline progenitor cells that are absent in smo mutants, while cyclopamine treatment blocks disc1 expression in these cells and mimics the effect of disc1 knock down on OPC specification. Various features of a number of psychiatric conditions could potentially arise through aberrant Hedgehog signalling. We therefore suggest that altered Shh signalling may be an important neurodevelopmental factor in the pathobiology of mental illness. PMID

  17. Controlled Delivery of Sonic Hedgehog with a Heparin-Based Coacervate.

    PubMed

    Johnson, Noah Ray; Wang, Yadong

    2015-01-01

    Here we describe the preparation of a delivery vehicle for controlled release of Sonic hedgehog (Shh). The vehicle contains a synthetic polycation and heparin which interact by polyvalent charge attraction and rapidly self-assemble into liquid coacervate droplets. The coacervate loads Shh with high efficiency, protects its bioactivity, and provides sustained and localized release at the site of application. Shh coacervate may be injected directly or coated onto a polymeric scaffold for tissue engineering approaches, as described here.

  18. Identification of sonic hedgehog as a candidate gene responsible for the polydactylous mouse mutant Sasquatch.

    PubMed

    Sharpe, J; Lettice, L; Hecksher-Sorensen, J; Fox, M; Hill, R; Krumlauf, R

    1999-01-28

    The mouse mutants of the hemimelia-luxate group (lx, lu, lst, Dh, Xt, and the more recently identified Hx, Xpl and Rim4; [1] [2] [3] [4] [5]) have in common preaxial polydactyly and longbone abnormalities. Associated with the duplication of digits are changes in the regulation of development of the anterior limb bud resulting in ectopic expression of signalling components such as Sonic hedgehog (Shh) and fibroblast growth factor-4 (Fgf4), but little is known about the molecular causes of this misregulation. We generated, by a transgene insertion event, a new member of this group of mutants, Sasquatch (Ssq), which disrupted aspects of both anteroposterior (AP) and dorsoventral (DV) patterning. The mutant displayed preaxial polydactyly in the hindlimbs of heterozygous embryos, and in both hindlimbs and forelimbs of homozygotes. The Shh, Fgf4, Fgf8, Hoxd12 and Hoxd13 genes were all ectopically expressed in the anterior region of affected limb buds. The insertion site was found to lie close to the Shh locus. Furthermore, expression from the transgene reporter has come under the control of a regulatory element that directs a pattern mirroring the endogenous expression pattern of Shh in limbs. In abnormal limbs, both Shh and the reporter were ectopically induced in the anterior region, whereas in normal limbs the reporter and Shh were restricted to the zone of polarising activity (ZPA). These data strongly suggest that Ssq is caused by direct interference with the cis regulation of the Shh gene.

  19. Multiphasic and tissue-specific roles of sonic hedgehog in cloacal septation and external genitalia development

    PubMed Central

    Seifert, Ashley W.; Bouldin, Cortney M.; Choi, Kyung-Suk; Harfe, Brian D.; Cohn, Martin J.

    2009-01-01

    Malformations of the external genitalia are among the most common congenital anomalies in humans. The urogenital and anorectal sinuses develop from the embryonic cloaca, and the penis and clitoris develop from the genital tubercle. Within the genital tubercle, the endodermally derived urethral epithelium functions as an organizer and expresses sonic hedgehog (Shh). Shh knockout mice lack external genitalia and have a persistent cloaca. This identified an early requirement for Shh, but precluded analysis of its later role in the genital tubercle. We conducted temporally controlled deletions of Shh and report that Shh is required continuously through the onset of sexual differentiation. Shh function is divisible into two temporal phases; an anogenital phase, during which Shh regulates outgrowth and patterning of the genital tubercle and septation of the cloaca, and a later external genital phase, during which Shh regulates urethral tube closure. Disruption of Shh function during the anogenital phase causes coordinated anorectal and genitourinary malformations, whereas inactivation during the external genital phase causes hypospadias. Shh directs cloacal septation by promoting cell proliferation in adjacent urorectal septum mesenchyme. Additionally, conditional inactivation of smoothened in the genital ectoderm and cloacal/urethral endoderm shows that the ectoderm is a direct target of Shh and is required for urethral tube closure, highlighting a novel role for genital ectoderm in urethragenesis. Identification of the stages during which disruption of Shh results in either isolated or coordinated malformations of anorectal and external genital organs provides a new tool for investigating the etiology of anogenital malformations in humans. PMID:19906862

  20. Detection of Canonical Hedgehog Signaling in Breast Cancer by 131-Iodine-Labeled Derivatives of the Sonic Hedgehog Protein

    PubMed Central

    Sims-Mourtada, Jennifer; Yang, David; Tworowska, Izabela; Larson, Richard; Smith, Daniel; Tsao, Ning; Opdenaker, Lynn; Mourtada, Firas; Woodward, Wendy

    2012-01-01

    Activation of hedgehog (HH) pathway signaling is observed in many tumors. Due to a feedback loop, the HH receptor Patched (PTCH-1) is overexpressed in tumors with activated HH signaling. Therefore, we sought to radiolabel the PTCH-1 ligand sonic (SHH) for detection of cancer cells with canonical HH activity. Receptor binding of 131I-SHH was increased in cell lines with high HH pathway activation. Our findings also show that PTCH-1 receptor expression is decreased upon treatment with HH signaling inhibitors, and receptor binding of 131I-SHH is significantly decreased following treatment with cyclopamine. In vivo imaging and biodistribution studies revealed significant accumulation of 131I-SHH within tumor tissue as compared to normal organs. Tumor-to-muscle ratios were approximately 8 : 1 at 5 hours, while tumor to blood and tumor to bone were 2 : 1 and 5 : 1, respectively. Significant uptake was also observed in liver and gastrointestinal tissue. These studies show that 131I-SHH is capable of in vivo detection of breast tumors with high HH signaling. We further demonstrate that the hedgehog receptor PTCH-1 is downregulated upon treatment with hedgehog inhibitors. Our data suggests that radiolabeled SHH derivatives may provide a method to determine response to SHH-targeted therapies. PMID:22811598

  1. Mutations of the Sonic Hedgehog Pathway Underlie Hypothalamic Hamartoma with Gelastic Epilepsy.

    PubMed

    Hildebrand, Michael S; Griffin, Nicole G; Damiano, John A; Cops, Elisa J; Burgess, Rosemary; Ozturk, Ezgi; Jones, Nigel C; Leventer, Richard J; Freeman, Jeremy L; Harvey, A Simon; Sadleir, Lynette G; Scheffer, Ingrid E; Major, Heather; Darbro, Benjamin W; Allen, Andrew S; Goldstein, David B; Kerrigan, John F; Berkovic, Samuel F; Heinzen, Erin L

    2016-08-01

    Hypothalamic hamartoma (HH) with gelastic epilepsy is a well-recognized drug-resistant epilepsy syndrome of early life.(1) Surgical resection allows limited access to the small deep-seated lesions that cause the disease. Here, we report the results of a search for somatic mutations in paired hamartoma- and leukocyte-derived DNA samples from 38 individuals which we conducted by using whole-exome sequencing (WES), chromosomal microarray (CMA), and targeted resequencing (TRS) of candidate genes. Somatic mutations were identified in genes involving regulation of the sonic hedgehog (Shh) pathway in 14/38 individuals (37%). Three individuals had somatic mutations in PRKACA, which encodes a cAMP-dependent protein kinase that acts as a repressor protein in the Shh pathway, and four subjects had somatic mutations in GLI3, an Shh pathway gene associated with HH. In seven other individuals, we identified two recurrent and three single brain-tissue-specific, large copy-number or loss-of-heterozygosity (LOH) variants involving multiple Shh genes, as well as other genes without an obvious biological link to the Shh pathway. The Shh pathway genes in these large somatic lesions include the ligand itself (SHH and IHH), the receptor SMO, and several other Shh downstream pathway members, including CREBBP and GLI2. Taken together, our data implicate perturbation of the Shh pathway in at least 37% of individuals with the HH epilepsy syndrome, consistent with the concept of a developmental pathway brain disease. PMID:27453577

  2. Developmental hypothyroidism abolishes bilateral differences in sonic hedgehog gene control in the rat hippocampal dentate gyrus.

    PubMed

    Tanaka, Takeshi; Wang, Liyun; Kimura, Masayuki; Abe, Hajime; Mizukami, Sayaka; Yoshida, Toshinori; Shibutani, Makoto

    2015-03-01

    Both developmental and adult-stage hypothyroidism disrupt rat hippocampal neurogenesis. We previously showed that exposing mouse offspring to manganese permanently disrupts hippocampal neurogenesis and abolishes the asymmetric distribution of cells expressing Mid1, a molecule regulated by sonic hedgehog (Shh) signaling. The present study examined the involvement of Shh signaling on the disruption of hippocampal neurogenesis in rats with hypothyroidism. Pregnant rats were treated with methimazole (MMI) at 0 or 200 ppm in the drinking water from gestation day 10-21 days after delivery (developmental hypothyroidism). Adult male rats were treated with MMI in the same manner from postnatal day (PND) 46 to PND 77 (adult-stage hypothyroidism). Developmental hypothyroidism reduced the number of Mid1(+) cells within the subgranular zone of the dentate gyrus of offspring on PND 21, and consequently abolished the normal asymmetric predominance of Mid1(+) cells on the right side through the adult stage. In control animals, Shh was expressed in a subpopulation of hilar neurons, showing asymmetric distribution with left side predominance on PND 21; however, this asymmetry did not continue through the adult stage. Developmental hypothyroidism increased Shh(+) neurons bilaterally and abolished the asymmetric distribution pattern on PND 21. Adult hypothyroidism also disrupted the asymmetric distribution of Mid1(+) cells but did not affect the distribution of Shh(+) hilar neurons. The results suggest that the hippocampal neurogenesis disruption seen in hypothyroidism involves changes in asymmetric Shh(+) neuron distribution in developmental hypothyroidism and altered Mid1 expression in both developmental and adult-stage hypothyroidism.

  3. Targeting the Sonic Hedgehog Signaling Pathway: Review of Smoothened and GLI Inhibitors

    PubMed Central

    Rimkus, Tadas K.; Carpenter, Richard L.; Qasem, Shadi; Chan, Michael; Lo, Hui-Wen

    2016-01-01

    The sonic hedgehog (Shh) signaling pathway is a major regulator of cell differentiation, cell proliferation, and tissue polarity. Aberrant activation of the Shh pathway has been shown in a variety of human cancers, including, basal cell carcinoma, malignant gliomas, medulloblastoma, leukemias, and cancers of the breast, lung, pancreas, and prostate. Tumorigenesis, tumor progression and therapeutic response have all been shown to be impacted by the Shh signaling pathway. Downstream effectors of the Shh pathway include smoothened (SMO) and glioma-associated oncogene homolog (GLI) family of zinc finger transcription factors. Both are regarded as important targets for cancer therapeutics. While most efforts have been devoted towards pharmacologically targeting SMO, developing GLI-targeted approach has its merit because of the fact that GLI proteins can be activated by both Shh ligand-dependent and -independent mechanisms. To date, two SMO inhibitors (LDE225/Sonidegib and GDC-0449/Vismodegib) have received FDA approval for treating basal cell carcinoma while many clinical trials are being conducted to evaluate the efficacy of this exciting class of targeted therapy in a variety of cancers. In this review, we provide an overview of the biology of the Shh pathway and then detail the current landscape of the Shh-SMO-GLI pathway inhibitors including those in preclinical studies and clinical trials. PMID:26891329

  4. The p53 inhibitor MDM2 facilitates Sonic Hedgehog-mediated tumorigenesis and influences cerebellar foliation.

    PubMed

    Malek, Reem; Matta, Jennifer; Taylor, Natalie; Perry, Mary Ellen; Mendrysa, Susan M

    2011-01-01

    Disruption of cerebellar granular neuronal precursor (GNP) maturation can result in defects in motor coordination and learning, or in medulloblastoma, the most common childhood brain tumor. The Sonic Hedgehog (Shh) pathway is important for GNP proliferation; however, the factors regulating the extent and timing of GNP proliferation, as well as GNP differentiation and migration are poorly understood. The p53 tumor suppressor has been shown to negatively regulate the activity of the Shh effector, Gli1, in neural stem cells; however, the contribution of p53 to the regulation of Shh signaling in GNPs during cerebellar development has not been determined. Here, we exploited a hypomorphic allele of Mdm2 (Mdm2(puro)), which encodes a critical negative regulator of p53, to alter the level of wild-type MDM2 and p53 in vivo. We report that mice with reduced levels of MDM2 and increased levels of p53 have small cerebella with shortened folia, reminiscent of deficient Shh signaling. Indeed, Shh signaling in Mdm2-deficient GNPs is attenuated, concomitant with decreased expression of the Shh transducers, Gli1 and Gli2. We also find that Shh stimulation of GNPs promotes MDM2 accumulation and enhances phosphorylation at serine 166, a modification known to increase MDM2-p53 binding. Significantly, loss of MDM2 in Ptch1(+/-) mice, a model for Shh-mediated human medulloblastoma, impedes cerebellar tumorigenesis. Together, these results place MDM2 at a major nexus between the p53 and Shh signaling pathways in GNPs, with key roles in cerebellar development, GNP survival, cerebellar foliation, and MB tumorigenesis. PMID:21437245

  5. Sonic Hedgehog Signaling in Basal Cell Nevus Syndrome

    PubMed Central

    Athar, Mohammad; Li, Changzhao; Kim, Arianna L.; Spiegelman, Vladimir S; Bickers, David R.

    2014-01-01

    The hedgehog (Hh) signaling pathway is considered to be a major signal transduction pathway during embryonic development but it usually shuts down after birth. Aberrant Shh activation during adulthood leads to neoplastic growth. Basal cell carcinoma (BCC) of the skin is driven by this pathway. Here, we summarize information related to the pathogenesis of this neoplasm, discuss pathways that crosstalk with Shh signaling and the importance of the primary cilium in this neoplastic process. The identification of the basic/translational components of Shh signaling has led to the discovery of potential mechanism-driven druggable targets and subsequent clinical trials have confirmed their remarkable efficacy in treating BCCs particularly in patients with Nevoid Basal Cell Carcinoma Syndrome (NBCCS), an autosomal dominant disorder in which patients inherit a germline mutation in the tumor suppressor gene Patched (Ptch). Patients with NBCCS develop dozens to hundreds of BCCs due to de-repression of the downstream G-protein coupled receptor Smoothened (SMO). Ptch mutations permit transposition of SMO to the primary cilium followed by enhanced expression of transcription factors Glis that drive cell proliferation and tumor growth. Clinical trials with the SMO inhibitor, vismodegib, in patients with NBCCS showing remarkable efficacy finally led to its FDA approval in 2012. PMID:25172843

  6. A specific mutation in the distant sonic hedgehog (SHH) cis-regulator (ZRS) causes Werner mesomelic syndrome (WMS) while complete ZRS duplications underlie Haas type polysyndactyly and preaxial polydactyly (PPD) with or without triphalangeal thumb.

    PubMed

    Wieczorek, Dagmar; Pawlik, Barbara; Li, Yun; Akarsu, Nurten A; Caliebe, Almuth; May, Klaus J W; Schweiger, Bernd; Vargas, Fernando R; Balci, Sevim; Gillessen-Kaesbach, Gabriele; Wollnik, Bernd

    2010-01-01

    Werner mesomelic syndrome (WMS) is an autosomal dominant disorder with unknown molecular etiology characterized by hypo- or aplasia of the tibiae in addition to the preaxial polydactyly (PPD) of the hands and feet and/or five-fingered hand with absence of thumbs. We show that point mutations of a specific nucleotide within the sonic hedgehog (SHH) regulatory region (ZRS) cause WMS. In a previously unpublished WMS family, we identified the causative G>A transition at position 404 of the ZRS, and in six affected family members of a second WMS family we found a 404G>C mutation of the ZRS. The 404G>A ZRS mutation is known as the "Cuban mutation" of PPD type II (PPD2). Interestingly, the index patient of that family had tibial hypoplasia as well. These data provide the first evidence that WMS is caused by a specific ZRS mutation, which leads to strong ectopic SHH expression. In contrast, we show that complete duplications of the ZRS region lead to type Haas polysyndactyly or triphalangeal thumb-polysyndactyly syndrome, but do not affect lower limb development. We suggest the term "ZRS-associated syndromes" and a clinical subclassification for the continuum of limb malformations caused by different molecular alterations of the ZRS.

  7. Development of stratum intermedium and its role as a Sonic hedgehog-signaling structure during odontogenesis.

    PubMed

    Koyama, E; Wu, C; Shimo, T; Iwamoto, M; Ohmori, T; Kurisu, K; Ookura, T; Bashir, M M; Abrams, W R; Tucker, T; Pacifici, M

    2001-10-01

    Stratum intermedium is a transient and subtle epithelial structure closely associated with inner dental epithelium in tooth germs. Little is known about its development and roles. To facilitate analysis, we used bovine tooth germs, predicting that they may contain a more conspicuous stratum intermedium. Indeed, early bell stage bovine tooth germs already displayed an obvious stratum intermedium with a typical multilayered organization and flanking the enamel knot. Strikingly, with further development, the cuspally located stratum intermedium underwent thinning and involution, whereas a multilayered stratum intermedium formed at successive sites along the cusp-to-cervix axis of odontogenesis. In situ hybridization and immunohistochemistry showed that stratum intermedium produces the signaling molecule Sonic hedgehog (Shh). Maximal Shh expression was invariably seen in its thickest multilayered portions. Shh was also produced by inner dental epithelium; expression was not constant but varied with development and cytodifferentiation of ameloblasts along the cusp-to-cervix axis. Interestingly, maximal Shh expression in inner dental epithelium did not coincide with that in stratum intermedium. Both stratum intermedium and inner dental epithelium expressed the Shh receptor Patched2 (Ptch2), an indication of autocrine signaling loops. Shh protein, but not RNA, was present in underlying dental mesenchyme, probably resulting from gradual diffusion from epithelial layers and reflecting paracrine loops of action. To analyze the regulation of Shh expression, epithelial and mesenchymal layers were separated and maintained in organ culture. Shh expression decreased over time, but was maintained in unoperated specimens. Our data show for the first time that stratum intermedium is a highly regulated and Shh-expressing structure. Given its dynamic and apparently interactive properties, stratum intermedium may help orchestrate progression of odontogenesis from cusp to cervix.

  8. Bridging the gap: heparan sulfate and Scube2 assemble Sonic hedgehog release complexes at the surface of producing cells

    PubMed Central

    Jakobs, P.; Schulz, P.; Ortmann, C.; Schürmann, S.; Exner, S.; Rebollido-Rios, R.; Dreier, R.; Seidler, D. G.; Grobe, K.

    2016-01-01

    Decision making in cellular ensembles requires the dynamic release of signaling molecules from the producing cells into the extracellular compartment. One important example of molecules that require regulated release in order to signal over several cell diameters is the Hedgehog (Hh) family, because all Hhs are synthesized as dual-lipidated proteins that firmly tether to the outer membrane leaflet of the cell that produces them. Factors for the release of the vertebrate Hh family member Sonic Hedgehog (Shh) include cell-surface sheddases that remove the lipidated terminal peptides, as well as the soluble glycoprotein Scube2 that cell-nonautonomously enhances this process. This raises the question of how soluble Scube2 is recruited to cell-bound Shh substrates to regulate their turnover. We hypothesized that heparan sulfate (HS) proteoglycans (HSPGs) on the producing cell surface may play this role. In this work, we confirm that HSPGs enrich Scube2 at the surface of Shh-producing cells and that Scube2-regulated proteolytic Shh processing and release depends on specific HS. This finding indicates that HSPGs act as cell-surface assembly and storage platforms for Shh substrates and for protein factors required for their release, making HSPGs critical decision makers for Scube2-dependent Shh signaling from the surface of producing cells. PMID:27199253

  9. Bridging the gap: heparan sulfate and Scube2 assemble Sonic hedgehog release complexes at the surface of producing cells.

    PubMed

    Jakobs, P; Schulz, P; Ortmann, C; Schürmann, S; Exner, S; Rebollido-Rios, R; Dreier, R; Seidler, D G; Grobe, K

    2016-01-01

    Decision making in cellular ensembles requires the dynamic release of signaling molecules from the producing cells into the extracellular compartment. One important example of molecules that require regulated release in order to signal over several cell diameters is the Hedgehog (Hh) family, because all Hhs are synthesized as dual-lipidated proteins that firmly tether to the outer membrane leaflet of the cell that produces them. Factors for the release of the vertebrate Hh family member Sonic Hedgehog (Shh) include cell-surface sheddases that remove the lipidated terminal peptides, as well as the soluble glycoprotein Scube2 that cell-nonautonomously enhances this process. This raises the question of how soluble Scube2 is recruited to cell-bound Shh substrates to regulate their turnover. We hypothesized that heparan sulfate (HS) proteoglycans (HSPGs) on the producing cell surface may play this role. In this work, we confirm that HSPGs enrich Scube2 at the surface of Shh-producing cells and that Scube2-regulated proteolytic Shh processing and release depends on specific HS. This finding indicates that HSPGs act as cell-surface assembly and storage platforms for Shh substrates and for protein factors required for their release, making HSPGs critical decision makers for Scube2-dependent Shh signaling from the surface of producing cells. PMID:27199253

  10. Mesocortical Dopamine Phenotypes in Mice Lacking the Sonic Hedgehog Receptor Cdon.

    PubMed

    Verwey, Michael; Grant, Alanna; Meti, Nicholas; Adye-White, Lauren; Torres-Berrío, Angelica; Rioux, Veronique; Lévesque, Martin; Charron, Frederic; Flores, Cecilia

    2016-01-01

    Motivated behaviors and many psychopathologies typically involve changes in dopamine release from the projections of the ventral tegmental area (VTA) and/or the substantia nigra pars compacta (SNc). The morphogen Sonic Hedgehog (Shh) specifies fates of midbrain dopamine neurons, but VTA-specific effects of Shh signaling are also being uncovered. In this study, we assessed the role of the Shh receptor Cdon in the development of VTA and SNc dopamine neurons. We find that Cdon is expressed in the proliferating progenitor zone of the embryonic ventral midbrain and that the number of proliferating cells in this region is increased in mouse Cdon(-/-) embryos. Consistent with a role of Shh in the regulation of neuronal proliferation in this region, we find that the number of tyrosine hydroxylase (TH)-positive neurons is increased in the VTA of Cdon(-/-) mice at birth and that this effect endures into adulthood. In contrast, the number of TH-positive neurons in the SNc is not altered in Cdon(-/-) mice at either age. Moreover, adult Cdon(-/-) mice have a greater number of medial prefrontal cortex (mPFC) dopamine presynaptic sites, and increased baseline concentrations of dopamine and dopamine metabolites selectively in this region. Finally, consistent with increased dopamine function in the mPFC, we find that adult Cdon(-/-) mice fail to exhibit behavioral plasticity upon repeated amphetamine treatment. Based on these data, we suggest that Cdon plays an important role encoding the diversity of dopamine neurons in the midbrain, influencing both the development of the mesocortical dopamine pathway and behavioral outputs that involve this neural circuitry. PMID:27419218

  11. Mesocortical Dopamine Phenotypes in Mice Lacking the Sonic Hedgehog Receptor Cdon

    PubMed Central

    Grant, Alanna; Meti, Nicholas; Adye-White, Lauren; Rioux, Veronique

    2016-01-01

    Abstract Motivated behaviors and many psychopathologies typically involve changes in dopamine release from the projections of the ventral tegmental area (VTA) and/or the substantia nigra pars compacta (SNc). The morphogen Sonic Hedgehog (Shh) specifies fates of midbrain dopamine neurons, but VTA-specific effects of Shh signaling are also being uncovered. In this study, we assessed the role of the Shh receptor Cdon in the development of VTA and SNc dopamine neurons. We find that Cdon is expressed in the proliferating progenitor zone of the embryonic ventral midbrain and that the number of proliferating cells in this region is increased in mouse Cdon−/− embryos. Consistent with a role of Shh in the regulation of neuronal proliferation in this region, we find that the number of tyrosine hydroxylase (TH)-positive neurons is increased in the VTA of Cdon−/− mice at birth and that this effect endures into adulthood. In contrast, the number of TH-positive neurons in the SNc is not altered in Cdon−/− mice at either age. Moreover, adult Cdon−/− mice have a greater number of medial prefrontal cortex (mPFC) dopamine presynaptic sites, and increased baseline concentrations of dopamine and dopamine metabolites selectively in this region. Finally, consistent with increased dopamine function in the mPFC, we find that adult Cdon−/− mice fail to exhibit behavioral plasticity upon repeated amphetamine treatment. Based on these data, we suggest that Cdon plays an important role encoding the diversity of dopamine neurons in the midbrain, influencing both the development of the mesocortical dopamine pathway and behavioral outputs that involve this neural circuitry. PMID:27419218

  12. The G-protein Alpha Subunit Gsα Is A Tumor Suppressor In Sonic Hedgehog-driven Medulloblastoma

    PubMed Central

    He, Xuelian; Zhang, Liguo; Chen, Ying; Remke, Marc; Shih, David; Lu, Fanghui; Wang, Haibo; Deng, Yaqi; Yu, Yang; Xia, Yong; Wu, Xiaochong; Ramaswamy, Vijay; Hu, Tom; Wang, Fan; Zhou, Wenhao; Burns, Dennis K.; Kim, Se Hoon; Kool, Marcel; Pfister, Stefan M.; Weinstein, Lee S.; Pomeroy, Scott L.; Gilbertson, Richard J.; Rubin, Joshua B.; Hou, Yiping; Wechsler-Reya, Robert; Taylor, Michael D.; Lu, Q. Richard

    2014-01-01

    Medulloblastoma, the most common malignant childhood brain tumor, exhibits distinct molecular subtypes and cellular origins. Genetic alterations driving medulloblastoma initiation and progression remain poorly understood. Herein, we identify GNAS, encoding the G-protein Gsα, as a potent tumor suppressor gene that defines a subset of aggressive Sonic Hedgehog (Shh)-driven human medulloblastomas. Ablation of the single Gnas gene in anatomically-distinct progenitors is sufficient to induce Shh-associated medulloblastomas, which recapitulate their human counterparts. Gsα is highly enriched at the primary cilium of granule neuron precursors and suppresses Shh-signaling by regulating both the cAMP-dependent pathway and ciliary trafficking of Hedgehog pathway components. Elevation of a Gsα effector, cAMP, effectively inhibits tumor cell proliferation and progression in Gnas mutants. Thus, our gain- and loss-of-function studies identify a previously unrecognized tumor suppressor function for Gsα that acts as a molecular link across Shh-group medulloblastomas of disparate cellular and anatomical origins, illuminating G-protein modulation as a potential therapeutic avenue. PMID:25150496

  13. Sonic hedgehog exerts distinct, stage-specific effects on tongue and taste papilla development.

    PubMed

    Liu, Hong-Xiang; Maccallum, Donald K; Edwards, Chris; Gaffield, William; Mistretta, Charlotte M

    2004-12-15

    Taste papillae are ectodermal specializations that serve to house and distribute the taste buds and their renewing cell populations in specific locations on the tongue. We previously showed that Sonic hedgehog (Shh) has a major role in regulating the number and spatial pattern of fungiform taste papillae on embryonic rat tongue, during a specific period of papilla formation from the prepapilla placode. Now we have immunolocalized the Shh protein and the Patched receptor protein (Ptc), and have tested potential roles for Shh in formation of the tongue, emergence of papilla placodes, development of papilla number and size, and maintenance of papillae after morphogenesis is advanced. Cultures of entire embryonic mandible or tongues from gestational days 12 to 18 [gestational or embryonic days (E)12-E18] were used, in which tongues and papillae develop with native spatial, temporal, and molecular characteristics. The Shh signaling pathway was disrupted with addition of cyclopamine, jervine, or the 5E1 blocking antibody. Shh and Ptc proteins are diffuse in prelingual tissue and early tongue swellings, and are progressively restricted to papilla placodes and then to regions of developing papillae. Ptc encircles the dense Shh immunoproduct in papillae at various stages. When the Shh signal is disrupted in cultures of E12 mandible, tongue formation is completely prevented. At later stages of tongue culture initiation, Shh signal disruption alters development of tongue shape (E13) and results in a repatterned fungiform papilla distribution that does not respect normally papilla-free tongue regions (E13-E14). Only a few hours of Shh signal disruption can irreversibly alter number and location of fungiform papillae on anterior tongue and elicit papilla formation on the intermolar eminence. However, once papillae are well formed (E16-E18), Shh apparently does not have a clear role in papilla maintenance, nor does the tongue retain competency to add fungiform papillae in

  14. Controlled delivery of sonic hedgehog morphogen and its potential for cardiac repair.

    PubMed

    Johnson, Noah Ray; Wang, Yadong

    2013-01-01

    The morphogen Sonic hedgehog (Shh) holds great promise for repair or regeneration of tissues suffering ischemic injury, however clinical translation is limited by its short half-life in the body. Here, we describe a coacervate delivery system which incorporates Shh, protects it from degradation, and sustains its release for at least 3 weeks. Shh released from the coacervate stimulates cardiac fibroblasts to upregulate the expression of multiple trophic factors including VEGF, SDF-1α, IGF-1, and Shh itself, for at least 48 hours. Shh coacervate also demonstrates cytoprotective effects for cardiomyocytes in a hydrogen peroxide-induced oxidative stress environment. In each of these studies the bioactivity of the Shh coacervate is enhanced compared to free Shh. These results warrant further investigation of the in vivo efficacy of Shh coacervate for cardiac repair.

  15. Sonic Hedgehog promotes proliferation of Notch-dependent monociliated choroid plexus tumour cells.

    PubMed

    Li, Li; Grausam, Katie B; Wang, Jun; Lun, Melody P; Ohli, Jasmin; Lidov, Hart G W; Calicchio, Monica L; Zeng, Erliang; Salisbury, Jeffrey L; Wechsler-Reya, Robert J; Lehtinen, Maria K; Schüller, Ulrich; Zhao, Haotian

    2016-04-01

    Aberrant Notch signalling has been linked to many cancers including choroid plexus (CP) tumours, a group of rare and predominantly paediatric brain neoplasms. We developed animal models of CP tumours, by inducing sustained expression of Notch1, that recapitulate properties of human CP tumours with aberrant NOTCH signalling. Whole-transcriptome and functional analyses showed that tumour cell proliferation is associated with Sonic Hedgehog (Shh) in the tumour microenvironment. Unlike CP epithelial cells, which have multiple primary cilia, tumour cells possess a solitary primary cilium as a result of Notch-mediated suppression of multiciliate differentiation. A Shh-driven signalling cascade in the primary cilium occurs in tumour cells but not in epithelial cells. Lineage studies show that CP tumours arise from monociliated progenitors in the roof plate characterized by elevated Notch signalling. Abnormal SHH signalling and distinct ciliogenesis are detected in human CP tumours, suggesting the SHH pathway and cilia differentiation as potential therapeutic avenues.

  16. Sonic Hedgehog promotes proliferation of Notch-dependent monociliated choroid plexus tumour cells

    PubMed Central

    Li, Li; Grausam, Katie B.; Wang, Jun; Lun, Melody P.; Ohli, Jasmin; Lidov, Hart G. W.; Calicchio, Monica L.; Zeng, Erliang; Salisbury, Jeffrey L.; Wechsler-Reya, Robert J.; Lehtinen, Maria K.; Schüller, Ulrich; Zhao, Haotian

    2016-01-01

    Aberrant Notch signaling has been linked to many cancers including choroid plexus (CP) tumours, a group of rare and predominantly pediatric brain neoplasms. We developed animal models of CP tumours by inducing sustained expression of Notch1 that recapitulate properties of human CP tumours with aberrant NOTCH signaling. Whole transcriptome and functional analyses showed that tumour cell proliferation is associated with Sonic Hedgehog (Shh) in the tumour microenvironment. Unlike CP epithelial cells, which have multiple primary cilia, tumour cells possess a solitary primary cilium as a result of Notch-mediated suppression of multiciliate diffferentiation. A Shh-driven signaling cascade in the primary cilium occurs in tumour cells but not in epithelial cells. Lineage studies show that CP tumours arise from mono-ciliated progenitors in the roof plate characterized by elevated Notch signaling. Abnormal SHH signaling and distinct ciliogenesis are detected in human CP tumours, suggesting SHH pathway and cilia differentiation as potential therapeutic avenues. PMID:26999738

  17. Sonic hedgehog pathway contributes to gastric cancer cell growth and proliferation.

    PubMed

    Wan, Jianhua; Zhou, Ji; Zhao, Hailong; Wang, Mei; Wei, Zhuanqin; Gao, Hongyan; Wang, Yongzhong; Cui, Hongjuan

    2014-04-01

    The Sonic Hedgehog (Shh) signaling pathway is commonly activated in gastrointestinal cancer. However, our understanding of the Shh pathway in gastric cancer remains limited. Here we examined the effects of cyclopamine, a specific inhibitor of the Shh signaling pathway, on cell growth and proliferation in gastric primary cancer cells GAM-016 and the MKN-45 cell line. The results showed that the Shh signaling molecules SHH, PTCH, SMO, GLI1, and GLI2 were intact and activated in both types of cells. Furthermore, we observed that cyclopamine inhibited gastric cancer cell proliferation through cell cycle arrest and apoptosis. An in vivo study using NOD/SCID mouse xenografts demonstrated that cyclopamine significantly prevented tumor growth and development. Our study indicated that Shh signaling pathway could promote gastric cancer cell proliferation and tumor development, and blocking this pathway may be a potential strategy in gastric cancer treatment.

  18. Sonic Hedgehog promotes proliferation of Notch-dependent monociliated choroid plexus tumour cells.

    PubMed

    Li, Li; Grausam, Katie B; Wang, Jun; Lun, Melody P; Ohli, Jasmin; Lidov, Hart G W; Calicchio, Monica L; Zeng, Erliang; Salisbury, Jeffrey L; Wechsler-Reya, Robert J; Lehtinen, Maria K; Schüller, Ulrich; Zhao, Haotian

    2016-04-01

    Aberrant Notch signalling has been linked to many cancers including choroid plexus (CP) tumours, a group of rare and predominantly paediatric brain neoplasms. We developed animal models of CP tumours, by inducing sustained expression of Notch1, that recapitulate properties of human CP tumours with aberrant NOTCH signalling. Whole-transcriptome and functional analyses showed that tumour cell proliferation is associated with Sonic Hedgehog (Shh) in the tumour microenvironment. Unlike CP epithelial cells, which have multiple primary cilia, tumour cells possess a solitary primary cilium as a result of Notch-mediated suppression of multiciliate differentiation. A Shh-driven signalling cascade in the primary cilium occurs in tumour cells but not in epithelial cells. Lineage studies show that CP tumours arise from monociliated progenitors in the roof plate characterized by elevated Notch signalling. Abnormal SHH signalling and distinct ciliogenesis are detected in human CP tumours, suggesting the SHH pathway and cilia differentiation as potential therapeutic avenues. PMID:26999738

  19. Sonic hedgehog and Wnt: antagonists in morphogenesis but collaborators in axon guidance

    PubMed Central

    Avilés, Evelyn C.; Wilson, Nicole H.; Stoeckli, Esther T.

    2013-01-01

    As indicated by their name, morphogens were first identified for their role in the formation of tissues early in development. Secreted from a source, they spread through the tissue to form gradients by which they affect the differentiation of precursor cells in a concentration-dependent manner. In this context, the antagonistic roles of the morphogens of the Wnt family and Sonic hedgehog (Shh) in the specification of cell types along the dorso-ventral axis of the neural tube have been studied in detail. However, more recently, morphogens have been demonstrated to act well beyond the early stages of nervous system development, as additional roles of morphogen gradients in vertebrate neural circuit formation have been identified. Both Wnt and Shh affect neural circuit formation at several stages by their influence on neurite extension, axon pathfinding and synapse formation. In this review, we will summarize the mechanisms of morphogen function during axon guidance in the vertebrate nervous system. PMID:23772206

  20. Activation of sonic hedgehog signaling enhances cell migration and invasion by induction of matrix metalloproteinase-2 and -9 via the phosphoinositide-3 kinase/AKT signaling pathway in glioblastoma.

    PubMed

    Chang, Liang; Zhao, Dan; Liu, Hui-Bin; Wang, Qiu-Shi; Zhang, Ping; Li, Chen-Long; Du, Wen-Zhong; Wang, Hong-Jun; Liu, Xing; Zhang, Zhi-Ren; Jiang, Chuan-Lu

    2015-11-01

    Aberrant hedgehog signaling contributes to the development of various malignancies, including glioblastoma (GBM). However, the potential mechanism of hedgehog signaling in GBM migration and invasion has remained to be elucidated. The present study showed that enhanced hedgehog signaling by recombinant human sonic hedgehog N‑terminal peptide (rhSHH) promoted the adhesion, invasion and migration of GBM cells, accompanied by increases in mRNA and protein levels of matrix metalloproteinase‑2 (MMP‑2) and MMP‑9. However, inhibition of hedgehog signaling with cyclopamine suppressed the adhesion, invasion and migration of GBM cells, accompanied by decreases in mRNA and protein levels of MMP‑2 and ‑9. Furthermore, it was found that MMP‑2- and MMP‑9-neutralizing antibodies or GAM6001 reversed the inductive effects of rhSHH on cell migration and invasion. In addition, enhanced hedgehog signaling by rhSHH increased AKT phosphorylation, whereas blockade of hedgehog signaling decreased AKT phosphorylations. Further experiments showed that LY294002, an inhibitor of phosphoinositide-3 kinase (PI3K), decreased rhSHH‑induced upregulation of MMP‑2 and ‑9. Finally, the protein expression of glioblastoma-associated oncogene 1 was positively correlated with levels of phosphorylated AKT as well as protein expressions of MMP‑2 and ‑9 in GBM tissue samples. In conclusion, the present study indicated that the hedgehog pathway regulates GBM-cell migration and invasion by increasing MMP-2 and MMP-9 production via the PI3K/AKT pathway. PMID:26299938

  1. Sonic hedgehog shedding results in functional activation of the solubilized protein.

    PubMed

    Ohlig, Stefanie; Farshi, Pershang; Pickhinke, Ute; van den Boom, Johannes; Höing, Susanne; Jakuschev, Stanislav; Hoffmann, Daniel; Dreier, Rita; Schöler, Hans R; Dierker, Tabea; Bordych, Christian; Grobe, Kay

    2011-06-14

    All Hedgehog (Hh) proteins are released from producing cells despite being synthesized as N- and C-terminally lipidated, membrane-tethered molecules. Thus, a cellular mechanism is needed for Hh solubilization. We previously suggested that a disintegrin and metalloprotease (ADAM)-mediated shedding of Sonic hedgehog (ShhNp) from its lipidated N and C termini results in protein solubilization. This finding, however, seemed at odds with the established role of N-terminal palmitoylation for ShhNp signaling activity. We now resolve this paradox by showing that N-palmitoylation of ShhNp N-terminal peptides is required for their proteolytic removal during solubilization. These peptides otherwise block ShhNp zinc coordination sites required for ShhNp binding to its receptor Patched (Ptc), explaining the essential yet indirect role of N-palmitoylation for ShhNp function. We suggest a functional model in which membrane-tethered multimeric ShhNp is at least partially autoinhibited in trans but is processed into fully active, soluble multimers upon palmitoylation-dependent cleavage of inhibitory N-terminal peptides. PMID:21664575

  2. The Role of Sonic Hedgehog Signaling in Osteoclastogenesis and Jaw Bone Destruction.

    PubMed

    Shimo, Tsuyoshi; Matsumoto, Kenichi; Takabatake, Kiyofumi; Aoyama, Eriko; Takebe, Yuichiro; Ibaragi, Soichiro; Okui, Tatsuo; Kurio, Naito; Takada, Hiroyuki; Obata, Kyoichi; Pang, Pai; Iwamoto, Masahiro; Nagatsuka, Hitoshi; Sasaki, Akira

    2016-01-01

    Sonic hedgehog (SHH) and its signaling have been identified in several human cancers, and increased levels of its expression appear to correlate with disease progression and metastasis. However, the role of SHH in bone destruction associated with oral squamous cell carcinomas is still unclear. In this study we analyzed SHH expression and the role played by SHH signaling in gingival carcinoma-induced jawbone destruction. From an analysis of surgically resected lower gingival squamous cell carcinoma mandible samples, we found that SHH was highly expressed in tumor cells that had invaded the bone matrix. On the other hand, the hedgehog receptor Patched and the signaling molecule Gli-2 were highly expressed in the osteoclasts and the progenitor cells. SHH stimulated osteoclast formation and pit formation in the presence of the receptor activator for nuclear factor-κB ligand (RANKL) in CD11b+ mouse bone marrow cells. SHH upregulated phosphorylation of ERK1/2 and p38 MAPK, NFATc1, tartrate-resistant acid phosphatase (TRAP), and Cathepsin K expression in RAW264.7 cells. Our results suggest that tumor-derived SHH stimulated the osteoclast formation and bone resorption in the tumor jawbone microenvironment. PMID:27007126

  3. Sonic hedgehog functions as a mitogen during bell stage of odontogenesis.

    PubMed

    Wu, Changshan; Shimo, Tsuyoshi; Liu, Mufei; Pacifici, Maurizio; Koyama, Eiki

    2003-01-01

    Epithelial-mesenchymal interactions are required for tissue growth and gene expression patterns during odontogenesis. We showed previously that Sonic hedgehog (SHH) is detectable in both dental epithelium and mesenchyme, while Shh transcripts are present in dental epithelium only, suggesting that SHH functions as an autocrine signal in epithelium and a paracrine signal in mesenchyme. This hypothesis was tested here. We found by in situ hybridization that the SHH autocrine receptor Ptch-2 is indeed expressed in dental epithelium whereas the paracrine receptor Ptc is expressed in mesenchyme. Bovine bell stage tooth germs were microsurgically separated into epithelial and mesenchymal portions and the resulting tissue fragments were organ-cultured. In epithelium fragments cultured by themselves, gene expression of Shh and Gli-1 (a putative transcriptional mediator of hedgehog signaling) was significantly decreased in both inner dental epithelium and stratum intermedium layers; this was accompanied by a sharp drop in epithelial cell proliferation. However, in companion control tissue fragments containing both epithelium and mesenchyme, Shh and Gli-1 expression as well as cell proliferation were maintained. Treatment of dental epithelial or mesenchymal cell populations in monolayer cultures with exogenous recombinant SHH stimulated cell proliferation. Together, the data provide clear evidence that Shh is synthesized by dental epithelium, reaches the underlying mesenchyme, and appears to act as an autocrine mitogen for epithelial cells and a paracrine mitogen for mesenchymal cells, thus exerting crucial functions in tooth germ growth, morphogenesis, and tissue-tissue interactions of bell stage of odontogenesis.

  4. The Role of Sonic Hedgehog Signaling in Osteoclastogenesis and Jaw Bone Destruction

    PubMed Central

    Shimo, Tsuyoshi; Matsumoto, Kenichi; Takabatake, Kiyofumi; Aoyama, Eriko; Takebe, Yuichiro; Ibaragi, Soichiro; Okui, Tatsuo; Kurio, Naito; Takada, Hiroyuki; Obata, Kyoichi; Pang, Pai; Iwamoto, Masahiro; Nagatsuka, Hitoshi; Sasaki, Akira

    2016-01-01

    Sonic hedgehog (SHH) and its signaling have been identified in several human cancers, and increased levels of its expression appear to correlate with disease progression and metastasis. However, the role of SHH in bone destruction associated with oral squamous cell carcinomas is still unclear. In this study we analyzed SHH expression and the role played by SHH signaling in gingival carcinoma-induced jawbone destruction. From an analysis of surgically resected lower gingival squamous cell carcinoma mandible samples, we found that SHH was highly expressed in tumor cells that had invaded the bone matrix. On the other hand, the hedgehog receptor Patched and the signaling molecule Gli-2 were highly expressed in the osteoclasts and the progenitor cells. SHH stimulated osteoclast formation and pit formation in the presence of the receptor activator for nuclear factor-κB ligand (RANKL) in CD11b+ mouse bone marrow cells. SHH upregulated phosphorylation of ERK1/2 and p38 MAPK, NFATc1, tartrate-resistant acid phosphatase (TRAP), and Cathepsin K expression in RAW264.7 cells. Our results suggest that tumor-derived SHH stimulated the osteoclast formation and bone resorption in the tumor jawbone microenvironment. PMID:27007126

  5. Holoprosencephaly in RSH/Smith-Lemli-Opitz syndrome: Does abnormal cholesterol metabolism affect the function of sonic hedgehog?

    SciTech Connect

    Kelley, R.I.; Roessler, E.; Muenke, M.

    1996-12-30

    The RAH/Smith-Lemli-Opitz syndrome (RAH/SLOS) is an autosomal recessive malformation syndrome associated with increased levels of 7-dehydrocholesterol (7-DHC) and a defect of cholesterol biosynthesis at the level of 3{beta}-hydroxy-steroid-{Delta}{sup 7}-reductase (7-DHC reductase). Because rats exposed to inhibitors of 7-DHC reductase during development have a high frequency of holoprosencephaly (HPE), we have undertaken a search for biochemical evidence of RSH/SLOS and other possible defects of sterol metabolism among patients with various forms of HPE. We describe 4 patients, one with semilobar HPE and three others with less complete forms of the HPE sequence, in whom we have made a biochemical diagnosis of RAH/SLOS. The clinical and biochemical spectrum of these and other patients with RAH/SLOS suggests a role of abnormal sterol metabolism in the pathogenesis of their malformations. The association of HPE and RAH/SLOS is discussed in light of the recent discoveries that mutations in the embryonic patterning gene, Sonic Hedgehog (SHH), can cause HPE in humans and that the sonic hedgehog protein product undergoes autoproteolysis to form a cholesterol-modified active product. These clinical, biochemical, and molecular studies suggest that HPE and other malformations in SLOS may be caused by incomplete or abnormal modification of the sonic hedgehog protein and, possibly, other patterning proteins of the hedgehog class, a hypothesis testable in somatic cell systems. 37 refs., 1 fig.

  6. The G protein Gαs acts as a tumor suppressor in sonic hedgehog signaling-driven tumorigenesis.

    PubMed

    Rao, Rohit; Salloum, Ralph; Xin, Mei; Lu, Q Richard

    2016-05-18

    G protein-coupled receptors (GPCRs) are critical players in tumor growth and progression. The redundant roles of GPCRs in tumor development confound effective treatment; therefore, targeting a single common signaling component downstream of these receptors may be efficacious. GPCRs transmit signals through heterotrimeric G proteins composed of Gα and Gβγ subunits. Hyperactive Gαs signaling can mediate tumor progression in some tissues; however, recent work in medulloblastoma and basal cell carcinoma revealed that Gαs can also function as a tumor suppressor in neoplasms derived from ectoderm cells including neural and epidermal stem/progenitor cells. In these stem-cell compartments, signaling through Gαs suppresses self-renewal by inhibiting the Sonic Hedgehog (SHH) and Hippo pathways. The loss of GNAS, which encodes Gαs, leads to activation of these pathways, over-proliferation of progenitor cells, and tumor formation. Gαs activates the cAMP-dependent protein kinase A (PKA) signaling pathway and inhibits activation of SHH effectors Smoothened-Gli. In addition, Gαs-cAMP-PKA activation negatively regulates the Hippo pathway by blocking the NF2-LATS1/2-Yap signaling. In this review, we will address the novel function of the signaling network regulated by Gαs in suppression of SHH-driven tumorigenesis and the therapeutic approaches that can be envisioned to harness this pathway to inhibit tumor growth and progression. PMID:27052725

  7. Transposon mutagenesis with coat color genotyping identifies an essential role for Skor2 in sonic hedgehog signaling and cerebellum development

    PubMed Central

    Wang, Baiping; Harrison, Wilbur; Overbeek, Paul A.; Zheng, Hui

    2011-01-01

    Correct development of the cerebellum requires coordinated sonic hedgehog (Shh) signaling from Purkinje to granule cells. How Shh expression is regulated in Purkinje cells is poorly understood. Using a novel tyrosinase minigene-tagged Sleeping Beauty transposon-mediated mutagenesis, which allows for coat color-based genotyping, we created mice in which the Ski/Sno family transcriptional co-repressor 2 (Skor2) gene is deleted. Loss of Skor2 leads to defective Purkinje cell development, a severe reduction of granule cell proliferation and a malformed cerebellum. Skor2 is specifically expressed in Purkinje cells in the brain, where it is required for proper expression of Shh. Skor2 overexpression suppresses BMP signaling in an HDAC-dependent manner and stimulates Shh promoter activity, suggesting that Skor2 represses BMP signaling to activate Shh expression. Our study identifies an essential function for Skor2 as a novel transcriptional regulator in Purkinje cells that acts upstream of Shh during cerebellum development. PMID:21937600

  8. Altered Sonic hedgehog signaling is associated with morphological abnormalities in the penis of the BB/WOR diabetic rat.

    PubMed

    Podlasek, Carol A; Zelner, David J; Harris, Joseph D; Meroz, Cynthia L; Tang, Yi; McKenna, Kevin E; McVary, Kevin T

    2003-09-01

    Erectile dysfunction (ED) is a common and debilitating pathological development that affects up to 75% of diabetic males. Neural stimulation is a crucial aspect of the normal erection process. Nerve injury causes ED and disrupts signaling of the Sonic hedgehog (Shh) cascade in the smooth muscle of the corpora cavernosa. Shh and targets of its signaling establish normal corpora cavernosal morphology during postnatal differentiation of the penis and regulate homeostasis in the adult. Interruption of the Shh cascade in the smooth muscle of the corpora cavernosa results in extensive changes in corpora cavernosal morphology that lead to ED. Our hypothesis is that the neuropathy observed in diabetics causes morphological changes in the corpora cavernosa of the penis that result in ED. Disruption of the Shh cascade may be involved in this process. We tested this hypothesis by examining morphological changes in the penis, altered gene and protein expression, apoptosis, and bromodeoxyuridine incorporation in the BB/WOR rat model of diabetes. Extensive smooth muscle and endothelial degradation was observed in the corpora cavernosa of diabetic penes. This degradation accompanied profound ED, significantly decreased Shh protein in the smooth muscle of the corpora cavernosa, and increased penile Shh RNA expression in the intact penis (nerves, corpora, and urethra). Localization and expression of Shh targets were also disrupted in the corpora cavernosa. Increasing our understanding of the molecular mechanisms that regulate Shh signaling may provide valuable insight into improving treatment options for diabetic impotence. PMID:12748119

  9. Sonic Hedgehog Has a Dual Effect on the Growth of Retinal Ganglion Axons Depending on Its Concentration

    PubMed Central

    Kolpak, Adrianne; Zhang, Jinhua; Bao, Zheng-Zheng

    2006-01-01

    The stereotypical projection of retinal ganglion cell (RGC) axons to the optic disc has served as a good model system for studying axon guidance. By both in vitro and in vivo experiments, we show that a secreted molecule, Sonic hedgehog (Shh), may play a critical role in the process. It is expressed in a dynamic pattern in the ganglion cell layer with a relatively higher expression in the center of the retina. Through gel culture and stripe assays, we show that Shh has a dual effect on RGC axonal growth, acting as a positive factor at low concentrations and a negative factor at high concentrations. Results from time-lapse video microscopic and stripe assay experiments further suggest that the effects of Shh on axons are not likely attributable to indirect transcriptional regulation by Shh. Overexpression of Shh protein or inhibition of Shh function inside the retina resulted in a complete loss of centrally directed projection of RGC axons, suggesting that precise regulation of Shh level inside the retina is critical for the projection of RGC axons to the optic disc. PMID:15800198

  10. Transposon mutagenesis with coat color genotyping identifies an essential role for Skor2 in sonic hedgehog signaling and cerebellum development.

    PubMed

    Wang, Baiping; Harrison, Wilbur; Overbeek, Paul A; Zheng, Hui

    2011-10-01

    Correct development of the cerebellum requires coordinated sonic hedgehog (Shh) signaling from Purkinje to granule cells. How Shh expression is regulated in Purkinje cells is poorly understood. Using a novel tyrosinase minigene-tagged Sleeping Beauty transposon-mediated mutagenesis, which allows for coat color-based genotyping, we created mice in which the Ski/Sno family transcriptional co-repressor 2 (Skor2) gene is deleted. Loss of Skor2 leads to defective Purkinje cell development, a severe reduction of granule cell proliferation and a malformed cerebellum. Skor2 is specifically expressed in Purkinje cells in the brain, where it is required for proper expression of Shh. Skor2 overexpression suppresses BMP signaling in an HDAC-dependent manner and stimulates Shh promoter activity, suggesting that Skor2 represses BMP signaling to activate Shh expression. Our study identifies an essential function for Skor2 as a novel transcriptional regulator in Purkinje cells that acts upstream of Shh during cerebellum development. PMID:21937600

  11. An Emerging Role of Sonic Hedgehog Shedding as a Modulator of Heparan Sulfate Interactions*

    PubMed Central

    Ohlig, Stefanie; Pickhinke, Ute; Sirko, Svetlana; Bandari, Shyam; Hoffmann, Daniel; Dreier, Rita; Farshi, Pershang; Götz, Magdalena; Grobe, Kay

    2012-01-01

    Major developmental morphogens of the Hedgehog (Hh) family act at short range and long range to direct cell fate decisions in vertebrate and invertebrate tissues. To this end, Hhs are released from local sources and act at a distance on target cells that express the Hh receptor Patched. However, morphogen secretion and spreading are not passive processes because all Hhs are synthesized as dually (N- and C-terminal) lipidated proteins that firmly tether to the surface of producing cells. On the cell surface, Hhs associate with each other and with heparan sulfate (HS) proteoglycans. This raises the question of how Hh solubilization and spreading is achieved. We recently discovered that Sonic hedgehog (Shh) is solubilized by proteolytic processing (shedding) of lipidated peptide termini in vitro. Because unprocessed N termini block Patched receptor binding sites in the cluster, we further suggested that their proteolytic removal is required for simultaneous Shh activation. In this work we confirm inactivity of unprocessed protein clusters and demonstrate restored biological Shh function upon distortion or removal of N-terminal amino acids and peptides. We further show that N-terminal Shh processing targets and inactivates the HS binding Cardin-Weintraub (CW) motif, resulting in soluble Shh clusters with their HS binding capacities strongly reduced. This may explain the ability of Shh to diffuse through the HS-containing extracellular matrix, whereas other HS-binding proteins are quickly immobilized. Our in vitro findings are supported by the presence of CW-processed Shh in murine brain samples, providing the first in vivo evidence for Shh shedding and subsequent solubilization of N-terminal-truncated proteins. PMID:23118222

  12. Reiterative pattern of sonic hedgehog expression in the catshark dentition reveals a phylogenetic template for jawed vertebrates.

    PubMed

    Smith, Moya M; Fraser, Gareth J; Chaplin, Natalie; Hobbs, Carl; Graham, Anthony

    2009-04-01

    For a dentition representing the most basal extant gnathostomes, that of the shark can provide us with key insights into the evolution of vertebrate dentitions. To detail the pattern of odontogenesis, we have profiled the expression of sonic hedgehog, a key regulator of tooth induction. We find in the catshark (Scyliorhinus canicula) that intense shh expression first occurs in a bilaterally symmetrical pattern restricted to broad regions in each half of the dentition in the embryo jaw. As in the mouse, there follows a changing temporal pattern of shh spatial restriction corresponding to epithelial bands of left and right dental fields, but also a subfield for symphyseal teeth. Then, intense shh expression is restricted to loci coincident with a temporal series of teeth in iterative jaw positions. The developmental expression of shh reveals previously undetected timing within epithelial stages of tooth formation. Each locus at alternate, even then odd, jaw positions establishes precise sequential timing for successive replacement within each tooth family. Shh appears first in the central cusp, iteratively along the jaw, then reiteratively within each tooth for secondary cusps. This progressive, sequential restriction of shh is shared by toothed gnathostomes and conserved through 500 million years of evolution.

  13. Expression pattern of sonic hedgehog signaling and calcitonin gene-related peptide in the socket healing process after tooth extraction.

    PubMed

    Pang, Pai; Shimo, Tsuyoshi; Takada, Hiroyuki; Matsumoto, Kenichi; Yoshioka, Norie; Ibaragi, Soichiro; Sasaki, Akira

    2015-11-01

    Sonic Hedgehog (SHH), a neural development inducer, plays a significant role in the bone healing process. Calcitonin gene-related peptide (CGRP), a neuropeptide marker of sensory nerves, has been demonstrated to affect bone formation. The roles of SHH signaling and CGRP-positive sensory nerves in the alveolar bone formation process have been unknown. Here we examined the expression patterns of SHH signaling and CGRP in mouse socket by immunohistochemistry and immunofluorescence analysis. We found that the expression level of SHH peaked at day 3 and was then decreased at 5 days after tooth extraction. CGRP, PTCH1 and GLI2 were each expressed in a similar pattern with their highest expression levels at day 5 and day 7 after tooth extraction. CGRP and GLI2 were co-expressed in some inflammatory cells and bone forming cells. In some areas, CGRP-positive neurons expressed GLI2. In conclusion, SHH may affect alveolar bone healing by interacting with CGRP-positive sensory neurons and thus regulate the socket's healing process after tooth extraction. PMID:26427874

  14. Spatial and temporal pattern for the dentition in the Australian lungfish revealed with sonic hedgehog expression profile.

    PubMed

    Smith, Moya M; Okabe, Masataka; Joss, Jean

    2009-02-22

    We report a temporal order of tooth addition in the Australian lungfish where timing of tooth induction is sequential in the same pattern as osteichthyans along the lower jaw. The order of tooth initiation in Neoceratodus starts from the midline tooth, together with left and right ones at jaw position 2, followed by 3 and then 1. This is the pattern order for dentary teeth of several teleosts and what we propose represents a stereotypic initiation pattern shared with all osteichthyans, including the living sister group to all tetrapods, the Australian lungfish. This is contrary to previous opinions that the lungfish dentition is otherwise derived and uniquely different. Sonic hedgehog (shh) expression is intensely focused on tooth positions at different times corresponding with their initiation order. This deployment of shh is required for lungfish tooth induction, as cyclopamine treatment results in complete loss of these teeth when applied before they develop. The temporal sequence of tooth initiation is possibly regulated by shh and is know to be required for dentition pattern in other osteichthyans, including cichlid fish and snakes. This reflects a shared developmental process with jawed vertebrates at the level of the tooth module but differs with the lack of replacement teeth.

  15. Variant allele frequency enrichment analysis in vitro reveals sonic hedgehog pathway to impede sustained temozolomide response in GBM

    PubMed Central

    Biswas, Nidhan K.; Chandra, Vikas; Sarkar-Roy, Neeta; Das, Tapojyoti; Bhattacharya, Rabindra N.; Tripathy, Laxmi N.; Basu, Sunandan K.; Kumar, Shantanu; Das, Subrata; Chatterjee, Ankita; Mukherjee, Ankur; Basu, Pryiadarshi; Maitra, Arindam; Chattopadhyay, Ansuman; Basu, Analabha; Dhara, Surajit

    2015-01-01

    Neoplastic cells of Glioblastoma multiforme (GBM) may or may not show sustained response to temozolomide (TMZ) chemotherapy. We hypothesize that TMZ chemotherapy response in GBM is predetermined in its neoplastic clones via a specific set of mutations that alter relevant pathways. We describe exome-wide enrichment of variant allele frequencies (VAFs) in neurospheres displaying contrasting phenotypes of sustained versus reversible TMZ-responses in vitro. Enrichment of VAFs was found on genes ST5, RP6KA1 and PRKDC in cells showing sustained TMZ-effect whereas on genes FREM2, AASDH and STK36, in cells showing reversible TMZ-effect. Ingenuity pathway analysis (IPA) revealed that these genes alter cell-cycle, G2/M-checkpoint-regulation and NHEJ pathways in sustained TMZ-effect cells whereas the lysine-II&V/phenylalanine degradation and sonic hedgehog (Hh) pathways in reversible TMZ-effect cells. Next, we validated the likely involvement of the Hh-pathway in TMZ-response on additional GBM neurospheres as well as on GBM patients, by extracting RNA-sequencing-based gene expression data from the TCGA-GBM database. Finally, we demonstrated TMZ-sensitization of a TMZ non-responder neurosphere in vitro by treating them with the FDA-approved pharmacological Hh-pathway inhibitor vismodegib. Altogether, our results indicate that the Hh-pathway impedes sustained TMZ-response in GBM and could be a potential therapeutic target to enhance TMZ-response in this malignancy. PMID:25604826

  16. Smoothened regulation in response to Hedgehog stimulation

    PubMed Central

    Jiang, Kai; Jia, Jianhang

    2016-01-01

    The Hedgehog (Hh) signaling pathway play critical roles in embryonic development and adult tissue homeostasis. A critical step in Hh signal transduction is how Hh receptor Patched (Ptc) inhibits the atypical G protein-coupled receptor Smoothened (Smo) in the absence of Hh and how this inhibition is release by Hh stimulation. It is unlikely that Ptc inhibits Smo by direct interaction. Here we discuss how Hh regulates the phosphorylation and ubiquitination of Smo, leading to cell surface and ciliary accumulation of Smo in Drosophila and vertebrate cells, respectively. In addition, we discuss how PI(4)P phospholipid acts in between Ptc and Smo to regulate Smo phosphorylation and activation in response to Hh stimulation. PMID:26973699

  17. Aberrant activation of Sonic hedgehog signaling in chronic cholecystitis and gallbladder carcinoma.

    PubMed

    Xie, Fang; Xu, Xiaoping; Xu, Angao; Liu, Cuiping; Liang, Fenfen; Xue, Minmin; Bai, Lan

    2014-03-01

    Sonic hedgehog (Shh) signaling has been extensively studied and is implicated in various inflammatory diseases and malignant tumors. We summarized the clinicopathological features and performed immunohistochemistry assays to examine expression of Shh signaling proteins in 10 normal mucosa, 32 gallbladder carcinoma (GBC), and 95 chronic cholecystitis (CC) specimens. The CC specimens were classified into three groups according to degree of inflammation. Compared with normal mucosa, CC, and GBC specimens exhibited increased expression of Shh. The immunoreactive score of Shh in the GBC group was higher than that in the mild to moderate CC groups but lower than that in the severe CC group (P < .05). Expression of Patched (Ptch) and Gli1 gradually increased from non-malignant cholecystitis to malignant tumors. Compared with CC specimens, GBC specimens showed higher cytoplasmic and membranous expression for Ptch (P < .05). Gli1 staining showed cytoplasmic expression of Gli1 in both CC (60% for mild, 77% for moderate, and 84% for severe) and GBC specimens (97%). Nuclear expression of Gli1 was detected in 16% of severe CC specimens with moderate to poor atypical hyperplasia, and in 62.5% of GBC specimens. Shh expression strongly correlated with expression of Ptch and Gli1. Furthermore, patients with strongly positive Gli1 staining had significantly lower survival rates than those with weakly positive staining. Our data indicate that the Shh signaling pathway is aberrantly activated in CC and GBC, and altered Shh signaling may be involved in the course of development from CC to gallbladder carcinogenesis.

  18. Sonic Hedgehog Mutations Identified in Holoprosencephaly Patients Can Act in a Dominant Negative Manner

    PubMed Central

    Singh, Samer; Tokhunts, Robert; Baubet, Valerie; Goetz, John A.; Huang, Zhen Jane; Schilling, Neal S.; Black, Kendall E.; MacKenzie, Todd A.; Dahmane, Nadia; Robbins, David J.

    2009-01-01

    Sonic Hedgehog (SHH) plays an important instructional role in vertebrate development, as exemplified by the numerous developmental disorders that occur when the SHH pathway is disrupted. Mutations in the SHH gene are the most common cause of sporadic and inherited Holoprosencephaly (HPE), a developmental disorder that is characterized by defective prosencephalon development. SHH HPE mutations provide a unique opportunity to better understand SHH biogenesis and signaling, and to decipher its role in the development of HPE. Here, we analyzed a panel of SHH HPE missense mutations that encode changes in the amino-terminal active domain of SHH. Our results show that SHH HPE mutations affect SHH biogenesis and signaling at multiple steps, which broadly results in low levels of protein expression, defective processing of SHH into its active form and protein with reduced activity. Additionally, we found that some inactive SHH proteins were able to modulate the activity of wt SHH in a dominant negative manner, both in vitro and in vivo. These findings show for the first time the susceptibility of SHH driven developmental processes to perturbations by low-activity forms of SHH. In conclusion, we demonstrate that SHH mutations found in HPE patients affect distinct steps of SHH biogenesis to attenuate SHH activity to different levels, and suggest that these variable levels of SHH activity might contribute to some of the phenotypic variation found in HPE patients. PMID:19057928

  19. Activation of Sonic hedgehog signaling in ventricular cardiomyocytes exerts cardioprotection against ischemia reperfusion injuries

    PubMed Central

    Paulis, Ludovit; Fauconnier, Jeremy; Cazorla, Olivier; Thireau, Jérome; Soleti, Raffaella; Vidal, Bastien; Ouillé, Aude; Bartholome, Marion; Bideaux, Patrice; Roubille, François; Le Guennec, Jean-Yves; Andriantsitohaina, Ramaroson; Martínez, M. Carmen; Lacampagne, Alain

    2015-01-01

    Sonic hedgehog (SHH) is a conserved protein involved in embryonic tissue patterning and development. SHH signaling has been reported as a cardio-protective pathway via muscle repair–associated angiogenesis. The goal of this study was to investigate the role of SHH signaling pathway in the adult myocardium in physiological situation and after ischemia-reperfusion. We show in a rat model of ischemia-reperfusion that stimulation of SHH pathway, either by a recombinant peptide or shed membranes microparticles harboring SHH ligand, prior to reperfusion reduces both infarct size and subsequent arrhythmias by preventing ventricular repolarization abnormalities. We further demonstrate in healthy animals a reduction of QTc interval mediated by NO/cGMP pathway leading to the shortening of ventricular cardiomyocytes action potential duration due to the activation of an inward rectifying potassium current sharing pharmacological and electrophysiological properties with ATP-dependent potassium current. Besides its effect on both angiogenesis and endothelial dysfunction we demonstrate here a novel cardio-protective effect of SHH acting directly on the cardiomyocytes. This emphasizes the pleotropic effect of SHH pathway as a potential cardiac therapeutic target. PMID:25613906

  20. Hippi is essential for node cilia assembly and Sonic hedgehog signaling

    PubMed Central

    Houde, Caroline; Dickinson, Robin J.; Houtzager, Vicky M.; Cullum, Rebecca; Montpetit, Rachel; Metzler, Martina; Simpson, Elizabeth M.; Roy, Sophie; Hayden, Michael R.; Hoodless, Pamela A.; Nicholson, Donald W.

    2016-01-01

    Hippi functions as an adapter protein that mediates pro-apoptotic signaling from poly-glutamine-expanded huntingtin, an established cause of Huntington disease, to the extrinsic cell death pathway. To explore other functions of Hippi we generated Hippi knock-out mice. This deletion causes randomization of the embryo turning process and heart looping, which are hallmarks of defective left–right (LR) axis patterning. We report that motile monocilia normally present at the surface of the embryonic node, and proposed to initiate the break in LR symmetry, are absent on Hippi−/− embryos. Furthermore, defects in central nervous system development are observed. The Sonic hedgehog (Shh) pathway is downregulated in the neural tube in the absence of Hippi, which results in failure to establish ventral neural cell fate. Together, these findings demonstrate a dual role for Hippi in cilia assembly and Shh signaling during development, in addition to its proposed role in apoptosis signal transduction in the adult brain under pathogenically stressful conditions. PMID:17027958

  1. Sonic hedgehog rescues cranial neural crest from cell death induced by ethanol exposure.

    PubMed

    Ahlgren, Sara C; Thakur, Vijaya; Bronner-Fraser, Marianne

    2002-08-01

    Alcohol is a teratogen that induces a variety of abnormalities including brain and facial defects [Jones, K. & Smith, D. (1973) Lancet 2, 999-1001], with the exact nature of the deficit depending on the time and magnitude of the dose of ethanol to which developing fetuses are exposed. In addition to abnormal facial structures, ethanol-treated embryos exhibit a highly characteristic pattern of cell death. Dying cells are observed in the premigratory and migratory neural crest cells that normally populate most facial structures. The observation that blocking Sonic hedgehog (Shh) signaling results in similar craniofacial abnormalities prompted us to examine whether there was a link between this aspect of fetal alcohol syndrome and loss of Shh. We demonstrate that administration of ethanol to chick embryos results in a dramatic loss of Shh, as well as a loss of transcripts involved in Shh signaling pathways. In contrast, other signaling molecules examined do not demonstrate such dramatic changes. Furthermore, we demonstrate that both the ethanol-induced cranial neural crest cell death and the associated craniofacial growth defect can be rescued by application of Shh. These data suggest that craniofacial anomalies resulting from fetal alcohol exposure are caused at least partially by loss of Shh and subsequent neural crest cell death.

  2. May Sonic Hedgehog proteins be markers for malignancy in uterine smooth muscle tumors?

    PubMed

    Garcia, Natalia; Bozzini, Nilo; Baiocchi, Glauco; da Cunha, Isabela Werneck; Maciel, Gustavo Arantes; Soares Junior, José Maria; Soares, Fernando Augusto; Baracat, Edmund Chada; Carvalho, Katia Candido

    2016-04-01

    Several studies have demonstrated that the Sonic Hedgehog signaling pathway (SHH) plays an important role in tumorigenesis and cellular differentiation. We analyzed the protein expression of SHH pathway components and evaluated whether their profile could be useful for the diagnosis, prognosis, or prediction of the risk of malignancy for uterine smooth muscle tumors (USMTs). A total of 176 samples (20 myometrium, 119 variants of leiomyoma, and 37 leiomyosarcoma) were evaluated for the protein expression of the SHH signaling components, HHIP1 (SHH inhibitor), and BMP4 (SHH target) by immunohistochemistry. Western blot analysis was performed to verify the specificity of the antibodies. We grouped leiomyoma samples into conventional leiomyomas and unusual leiomyomas that comprise atypical, cellular, mitotically active leiomyomas and uterine smooth muscle tumors of uncertain malignant potential. Immunohistochemical analysis showed that SMO, SUFU, GLI1, GLI3, and BMP4 expression gradually increased depending on to the histologic tissue type. The protein expression of SMO, SUFU, and GLI1 was increased in unusual leiomyoma and leiomyosarcoma samples compared to normal myometrium. The inhibitor HHIP1 showed higher expression in myometrium, whereas only negative or basal expression of SMO, SUFU, GLI1, and GLI3 was detected in these samples. Strong expression of SHH was associated with poorer overall survival. Our data suggest that the expression of SHH proteins can be useful for evaluating the potential risk of malignancy for USMTs. Moreover, GLI1 and SMO may serve as future therapeutic targets for women with USMTs. PMID:26997437

  3. Regeneration of the Cavernous Nerve by Sonic Hedgehog Using Aligned Peptide Amphiphile Nanofibers

    PubMed Central

    Angeloni, Nicholas L.; Bond, Christopher W.; Tang, Yi; Harrington, Daniel A.; Zhang, Shuming; Stupp, Samuel I.; McKenna, Kevin E.; Podlasek, Carol A.

    2010-01-01

    Efforts to regenerate the cavernous nerve (CN), which provides innervation to the penis, have been minimally successful, with little translation into improved clinical outcomes. We propose that, Sonic hedgehog (SHH), is critical to maintain CN integrity, and that SHH delivered to the CN by novel peptide amphiphile (PA) nanofibers, will promote CN regeneration, restore physiological function, and prevent penile morphology changes that result in erectile dysfunction (ED). We performed localization studies, inhibition of SHH signaling in the CN, and treatment of crushed CNs with SHH protein via linear PA gels, which are an innovative extended release method of delivery. Morphological, functional and molecular analysis revealed that SHH protein is essential to maintain CN architecture, and that SHH treatment promoted CN regeneration, suppressed penile apoptosis and caused a 58% improvement in erectile function in less than half the time reported in the literature. These studies show that SHH has substantial clinical application to regenerate the CN in prostatectomy and diabetic patients, that this methodology has broad application to regenerate any peripheral nerve that SHH is necessary for maintenance of its structure, and that this nanotechnology method of protein delivery may have wide spread application as an in vivo delivery tool in many organs. PMID:20971506

  4. Sonic hedgehog initiates cochlear hair cell regeneration through downregulation of retinoblastoma protein

    SciTech Connect

    Lu, Na; Chen, Yan; Wang, Zhengmin; Chen, Guoling; Lin, Qin; Chen, Zheng-Yi; Li, Huawei

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer Shh activation in neonatal cochleae enhances sensory cell proliferation. Black-Right-Pointing-Pointer Proliferating supporting cells can transdifferentiate into hair cells. Black-Right-Pointing-Pointer Shh promotes proliferation by transiently modulating pRb activity. Black-Right-Pointing-Pointer Shh inhibits pRb by inhibiting transcription and increasing phosphorylation of pRb. -- Abstract: Cell cycle re-entry by cochlear supporting cells and/or hair cells is considered one of the best approaches for restoring hearing loss as a result of hair cell damage. To identify mechanisms that can be modulated to initiate cell cycle re-entry and hair cell regeneration, we studied the effect of activating the sonic hedgehog (Shh) pathway. We show that Shh signaling in postnatal rat cochleae damaged by neomycin leads to renewed proliferation of supporting cells and hair cells. Further, proliferating supporting cells are likely to transdifferentiate into hair cells. Shh treatment leads to inhibition of retinoblastoma protein (pRb) by increasing phosphorylated pRb and reducing retinoblastoma gene transcription. This results in upregulation of cyclins B1, D2, and D3, and CDK1. These results suggest that Shh signaling induces cell cycle re-entry in cochlear sensory epithelium and the production of new hair cells, in part by attenuating pRb function. This study provides an additional route to modulate pRb function with important implications in mammalian hair cell regeneration.

  5. Sonic hedgehog maintains cellular and neurochemical homeostasis in the adult nigrostriatal circuit.

    PubMed

    Gonzalez-Reyes, Luis E; Verbitsky, Miguel; Blesa, Javier; Jackson-Lewis, Vernice; Paredes, Daniel; Tillack, Karsten; Phani, Sudarshan; Kramer, Edgar R; Przedborski, Serge; Kottmann, Andreas H

    2012-07-26

    Non cell-autonomous processes are thought to play critical roles in the cellular maintenance of the healthy and diseased brain but mechanistic details remain unclear. We report that the interruption of a non cell-autonomous mode of sonic hedgehog (Shh) signaling originating from dopaminergic neurons causes progressive, adult-onset degeneration of dopaminergic, cholinergic, and fast spiking GABAergic neurons of the mesostriatal circuit, imbalance of cholinergic and dopaminergic neurotransmission, and motor deficits reminiscent of Parkinson's disease. Variable Shh signaling results in graded inhibition of muscarinic autoreceptor- and glial cell line-derived neurotrophic factor (GDNF)-expression in the striatum. Reciprocally, graded signals that emanate from striatal cholinergic neurons and engage the canonical GDNF receptor Ret inhibit Shh expression in dopaminergic neurons. Thus, we discovered a mechanism for neuronal subtype specific and reciprocal communication that is essential for neurochemical and structural homeostasis in the nigrostriatal circuit. These results provide integrative insights into non cell-autonomous processes likely at play in neurodegenerative conditions such as Parkinson's disease. PMID:22841315

  6. Expression Profile of Sonic Hedgehog Pathway Members in the Developing Human Fetal Brain

    PubMed Central

    Tichy, Julia; Zinke, Jenny; Bunz, Benedikt; Meyermann, Richard; Harter, Patrick N.; Mittelbronn, Michel

    2015-01-01

    The Sonic Hedgehog (SHH) pathway plays a central role in the developing mammalian CNS. In our study, we aimed to investigate the spatiotemporal SHH pathway expression pattern in human fetal brains. We analyzed 22 normal fetal brains for Shh, Patched, Smoothened, and Gli1-3 expression by immunohistochemistry. In the telencephalon, strongest expression of Shh, Smoothened, and Gli2 was found in the cortical plate (CP) and ventricular zone. Patched was strongly upregulated in the ventricular zone and Gli1 in the CP. In the cerebellum, SHH pathway members were strongly expressed in the external granular layer (EGL). SHH pathway members significantly decreased over time in the ventricular and subventricular zone and in the cerebellar EGL, while increasing levels were found in more superficial telencephalic layers. Our findings show that SHH pathway members are strongly expressed in areas important for proliferation and differentiation and indicate a temporal expression gradient in telencephalic and cerebellar layers probably due to decreased proliferation of progenitor cells and increased differentiation. Our data about the spatiotemporal expression of SHH pathway members in the developing human brain serves as a base for the understanding of both normal and pathological CNS development. PMID:26266257

  7. WIP1 modulates responsiveness to Sonic Hedgehog signaling in neuronal precursor cells and medulloblastoma

    PubMed Central

    Wen, Jing; Lee, Juhyun; Malhotra, Anshu; Nahta, Rita; Arnold, Amanda R.; Buss, Meghan C.; Brown, Briana D.; Maier, Caroline; Kenney, Anna M.; Remke, Marc; Ramaswamy, Vijay; Taylor, Michael D.; Castellino, Robert C.

    2016-01-01

    High-level amplification of the protein phosphatase PPM1D (WIP1) is present in a subset of medulloblastomas (MBs) that have an expression profile consistent with active Sonic Hedgehog (SHH) signaling. We found that WIP1 overexpression increased expression of Shh target genes and cell proliferation in response to Shh stimulation in NIH3T3 and cerebellar granule neuron precursor (cGNP) cells in a p53-independent manner. Thus, we developed a mouse in which WIP1 is expressed in the developing brain under control of the Neurod2 promoter (ND2:WIP1). The external granule layer in early post-natal ND2:WIP1 mice exhibited increased proliferation and expression of Shh downstream targets. MB incidence increased and survival decreased when ND2:WIP1 mice were crossed with a Shh-activated MB mouse model. Conversely, Wip1 knock out significantly suppressed MB formation in two independent mouse models of Shh-activated MB. Furthermore, Wip1 knock-down or treatment with a WIP1 inhibitor suppressed the effects of Shh stimulation and potentiated the growth inhibitory effects of SHH pathway-inhibiting drugs in Shh-activated MB cells in vitro. This suggests an important cross-talk between SHH and WIP1 pathways that accelerates tumorigenesis and supports WIP1 inhibition as a potential treatment strategy for MB. PMID:27086929

  8. GATA4 and GATA6 regulate pancreatic endoderm identity through inhibition of hedgehog signaling.

    PubMed

    Xuan, Shouhong; Sussel, Lori

    2016-03-01

    GATA4 and GATA6 are zinc finger transcription factors that have important functions in several mesodermal and endodermal organs, including heart, liver and pancreas. In humans, heterozygous mutations of either factor are associated with pancreatic agenesis; however, homozygous deletion of both Gata4 and Gata6 is necessary to disrupt pancreas development in mice. In this study, we demonstrate that arrested pancreatic development in Gata4(fl/fl); Gata6(fl/fl); Pdx1:Cre (pDKO) embryos is accompanied by the transition of ventral and dorsal pancreatic fates into intestinal or stomach lineages, respectively. These results indicate that GATA4 and GATA6 play essential roles in maintaining pancreas identity by regulating foregut endodermal fates. Remarkably, pancreatic anlagen derived from pDKO embryos also display a dramatic upregulation of hedgehog pathway components, which are normally absent from the presumptive pancreatic endoderm. Consistent with the erroneous activation of hedgehog signaling, we demonstrate that GATA4 and GATA6 are able to repress transcription through the sonic hedgehog (Shh) endoderm-specific enhancer MACS1 and that GATA-binding sites within this enhancer are necessary for this repressive activity. These studies establish the importance of GATA4/6-mediated inhibition of hedgehog signaling as a major mechanism regulating pancreatic endoderm specification during patterning of the gut tube.

  9. GATA4 and GATA6 regulate pancreatic endoderm identity through inhibition of hedgehog signaling.

    PubMed

    Xuan, Shouhong; Sussel, Lori

    2016-03-01

    GATA4 and GATA6 are zinc finger transcription factors that have important functions in several mesodermal and endodermal organs, including heart, liver and pancreas. In humans, heterozygous mutations of either factor are associated with pancreatic agenesis; however, homozygous deletion of both Gata4 and Gata6 is necessary to disrupt pancreas development in mice. In this study, we demonstrate that arrested pancreatic development in Gata4(fl/fl); Gata6(fl/fl); Pdx1:Cre (pDKO) embryos is accompanied by the transition of ventral and dorsal pancreatic fates into intestinal or stomach lineages, respectively. These results indicate that GATA4 and GATA6 play essential roles in maintaining pancreas identity by regulating foregut endodermal fates. Remarkably, pancreatic anlagen derived from pDKO embryos also display a dramatic upregulation of hedgehog pathway components, which are normally absent from the presumptive pancreatic endoderm. Consistent with the erroneous activation of hedgehog signaling, we demonstrate that GATA4 and GATA6 are able to repress transcription through the sonic hedgehog (Shh) endoderm-specific enhancer MACS1 and that GATA-binding sites within this enhancer are necessary for this repressive activity. These studies establish the importance of GATA4/6-mediated inhibition of hedgehog signaling as a major mechanism regulating pancreatic endoderm specification during patterning of the gut tube. PMID:26932670

  10. Metformin exerts anticancer effects through the inhibition of the Sonic hedgehog signaling pathway in breast cancer

    PubMed Central

    FAN, CONG; WANG, YUNSHAN; LIU, ZIMING; SUN, YING; WANG, XIUWEN; WEI, GUANGWEI; WEI, JUNMIN

    2015-01-01

    Metformin, a widely prescribed antidiabetic drug, has previously been shown to lower the risk of certain types of cancer, including that of breast cancer, and to improve prognosis. Its anticancer effects, which are mediated by the activation of AMP-activated protein kinase (AMPK), have become notable. The Sonic hedgehog (Shh) signaling pathway is involved in changes in mammary ducts and malignant transformation. The aim of the present study was to elucidate the role of the Shh pathway in mediating the anticancer effects of metformin and the correlation between AMPK and the Shh pathway. We investigated the effectiveness of metformin in inhibiting the proliferation, migration, invasion and stemness of breast cancer cells in vitro using RNA extraction and reverse transcription-polymerase chain reaction (RT-PCR), western blot analysis, cell proliferation assay, scratch-wound assay (cell migration assay), cell invasion assay, mammosphere culture and flow cytometry. In in vivo experiments, a tumor xenograft model was used to detect the effects of metformin on cancer cell proliferation. The results revealed that the treatment of breast cancer cells with metformin led to the inhibition of the Shh signaling pathway. Importantly, metformin inhibited recombinant human Shh (rhShh)-induced cell migration, invasion, and stemness, and impaired cell proliferation both in vitro and in vivo. Furthermore, the small interfering RNA (siRNA)-mediated downregulation of AMPK reversed the inhibitory effects of metformin on rhShh-induced Gli-1 expression and stemness. Our findings identified a role of the Shh signaling pathway in the anticancer effects of metformin in breast cancer. Furthermore, we revealed that the metformin-mediated inhibition of the Shh signaling pathway may be dependent on AMPK. PMID:25999130

  11. Metformin exerts anticancer effects through the inhibition of the Sonic hedgehog signaling pathway in breast cancer.

    PubMed

    Fan, Cong; Wang, Yunshan; Liu, Ziming; Sun, Ying; Wang, Xiuwen; Wei, Guangwei; Wei, Junmin

    2015-07-01

    Metformin, a widely prescribed antidiabetic drug, has previously been shown to lower the risk of certain types of cancer, including that of breast cancer, and to improve prognosis. Its anticancer effects, which are mediated by the activation of AMP-activated protein kinase (AMPK), have become notable. The Sonic hedgehog (Shh) signaling pathway is involved in changes in mammary ducts and malignant transformation. The aim of the present study was to elucidate the role of the Shh pathway in mediating the anticancer effects of metformin and the correlation between AMPK and the Shh pathway. We investigated the effectiveness of metformin in inhibiting the proliferation, migration, invasion and stemness of breast cancer cells in vitro using RNA extraction and reverse transcription‑polymerase chain reaction (RT-PCR), western blot analysis, cell proliferation assay, scratch-wound assay (cell migration assay), cell invasion assay, mammosphere culture and flow cytometry. In in vivo experiments, a tumor xenograft model was used to detect the effects of metformin on cancer cell proliferation. The results revealed that the treatment of breast cancer cells with metformin led to the inhibition of the Shh signaling pathway. Importantly, metformin inhibited recombinant human Shh (rhShh)‑induced cell migration, invasion, and stemness, and impaired cell proliferation both in vitro and in vivo. Furthermore, the small interfering RNA (siRNA)‑mediated downregulation of AMPK reversed the inhibitory effects of metformin on rhShh‑induced Gli-1 expression and stemness. Our findings identified a role of the Shh signaling pathway in the anticancer effects of metformin in breast cancer. Furthermore, we revealed that the metformin-mediated inhibition of the Shh signaling pathway may be dependent on AMPK. PMID:25999130

  12. Sonic hedgehog: its expression in a healing cornea and its role in neovascularization

    PubMed Central

    Miyamoto, Takeshi; Saika, Shizuya

    2009-01-01

    Purpose To examine if sonic hedgehog (Shh) is involved in tissue neovascularization by using cell culture and an animal cornea. Methods The effects of exogenous Shh (5.0 nM), vascular endothelial growth factor (VEGF), and/or a Shh signal inhibitor (2.5 or 10.0 μM cyclopamine) on vessel-like tube formation of vascular endothelial cells were examined in vitro. The effects of Shh on the expression of angiogenic cytokines in cultured cell types were examined in cultured cells. The expression of Shh and its receptor, Patched 1 (Ptc), was examined in a vascularized mouse cornea during post-alkali burn healing. The effect of exogenous Shh on corneal neovascularization in vivo was assayed using a rat cornea system. The effect of a topical injection of cyclopamine on cauterization-induced corneal neovascularization was then studied. Results Adding Shh promoted vessel-like tube formation of vascular endothelial cells. This effect was counteracted by addition of cyclopamine. Cyclopamine did not affect VEGF-enhanced tube formation. Shh did not affect the expression levels of angiogenic cytokines in cultured cell types. mRNA and protein expression levels of Shh and Ptc were under the detection limit in an uninjured cornea, but Shh but not Ptc was upregulated in a healing, alkali-burned, vascularized cornea. Exogenous Shh promoted neovascularization (NV) formation in vivo in a rat cornea. Topical cyclopmine blocked Gli signaling (blocked translocation of Gli3) and the length of neovascularization in the peripheral cornea post-cauterization as compared with the control vehicle-treated cornea. Conclusions Shh enhances endothelial tube formation independently through VEGF signaling in vitro. Shh signaling is involved in the development of unfavorable corneal neovascularization in animal corneas. PMID:19471603

  13. Sonic hedgehog cascade is required for penile postnatal morphogenesis, differentiation, and adult homeostasis.

    PubMed

    Podlasek, Carol A; Zelner, David J; Jiang, Hong Bin; Tang, Yi; Houston, John; McKenna, Kevin E; McVary, Kevin T

    2003-02-01

    The penis is unique in that it undergoes morphogenesis and differentiation primarily in the postnatal period. For complex structures such as the penis to be made from undifferentiated precursor cells, proliferation, differentiation, and patterning are required. This process involves coordinated activity of multiple signals. Sonic hedgehog (Shh) forms part of a regulatory cascade that is essential for growth and morphogenesis of many tissues. It is hypothesized that the penis utilizes regulatory mechanisms similar to those of the limb and accessory sex organs to pattern penile postnatal morphogenesis and differentiation and that the Shh cascade is critical to this process. To test this hypothesis, Shh, BMP-4, Ptc, and Hoxa-10 localization and function were examined in Sprague-Dawley rat penes by means of quantitative reverse transcription polymerase chain reaction, in situ hybridization, immunohistochemistry, and Western blotting. These genes were expressed in the penis during postnatal morphogenesis in a spatially and temporally restricted manner in adjacent layers of the corpora cavernosal sinusoids. The function of Shh and BMP-4 is to establish and maintain corpora cavernosal sinusoids. The data suggest that Ptc and Hoxa-10 are also important in penile morphogenesis. The continuing function of Shh and targets of its signaling in maintaining penile homeostasis in the adult is significant because disruption of Shh signaling affects erectile function. This is the first report that demonstrates the significant role that Shh plays in establishing and maintaining penile homeostasis and how this relates to erectile function. These studies provide valuable insight that may be applied to improve treatment options for erectile dysfunction. PMID:12533405

  14. Sonic Hedgehog Signaling Affected by Promoter Hypermethylation Induces Aberrant Gli2 Expression in Spina Bifida.

    PubMed

    Lu, Xiao-Lin; Wang, Li; Chang, Shao-Yan; Shangguan, Shao-Fang; Wang, Zhen; Wu, Li-Hua; Zou, Ji-Zhen; Xiao, Ping; Li, Rui; Bao, Yi-Hua; Qiu, Z-Y; Zhang, Ting

    2016-10-01

    GLI2 is a key mediator of the sonic hedgehog (Shh) signaling pathway and plays an important role in neural tube development during vertebrate embryogenesis; however, the role of gli2 in human folate-related neural tube defects remains unclear. In this study, we compared methylation status and polymorphisms of gli2 between spina bifida patients and a control group to explore the underlying mechanisms related to folate deficiency in spina bifida. No single nucleotide polymorphism was found to be significantly different between the two groups, although gli2 methylation levels were significantly increased in spina bifida samples, accompanied by aberrant GLI2 expression. Moreover, a prominent negative correlation was found between the folate level in brain tissue and the gli2 methylation status (r = -0.41, P = 0.014), and gli2 hypermethylation increased the risk of spina bifida with an odds ratio of 12.45 (95 % confidence interval: 2.71-57.22, P = 0.001). In addition, we established a cell model to illustrate the effect of gli2 expression and the accessibility of chromatin affected by methylation. High gli2 and gli1 mRNA expression was detected in 5-Aza-treated cells, while gli2 hypermethylation resulted in chromatin inaccessibility and a reduced association with nuclear proteins containing transcriptional factors. More meaningful to the pathway, the effect gene of the Shh pathway, gli1, was found to have a reduced level of expression along with a decreased expression of gli2 in our cell model. Aberrant high methylation resulted in the low expression of gli2 in spina bifida, which was affected by the change in chromatin status and the capacity of transcription factor binding. PMID:26446020

  15. Sonic hedgehog is a chemotactic neural crest cell guide that is perturbed by ethanol exposure.

    PubMed

    Tolosa, Ezequiel J; Fernández-Zapico, Martín E; Battiato, Natalia L; Rovasio, Roberto A

    2016-01-01

    Our aim was to understand the involvement of Sonic hedgehog (Shh) morphogen in the oriented distribution of neural crest cells (NCCs) toward the optic vesicle and to look for potential disorders of this guiding mechanism after ethanol exposure. In vitro directional analysis showed the chemotactic response of NCCs up Shh gradients and to notochord co-cultures (Shh source) or to their conditioned medium, a response inhibited by anti-Shh antibody, receptor inhibitor cyclopamine and anti-Smo morpholino (MO). Expression of the Ptch-Smo receptor complex on in vitro NCCs was also shown. In whole embryos, the expression of Shh mRNA and protein was seen in the ocular region, and of Ptch, Smo and Gli/Sufu system on cephalic NCCs. Anti-Smo MO or Ptch-mutated plasmid (Ptch1(Δloop2)) impaired cephalic NCC migration/distribution, with fewer cells invading the optic region and with higher cell density at the homolateral mesencephalic level. Beads embedded with cyclopamine (Smo-blocking) or Shh (ectopic signal) supported the role of Shh as an in vivo guide molecule for cephalic NCCs. Ethanol exposure perturbed in vitro and in vivo NCC migration. Early stage embryos treated with ethanol, in a model reproducing Fetal Alcohol Syndrome, showed later disruptions of craniofacial development associated with abnormal in situ expression of Shh morphogen. The results show the Shh/Ptch/Smo-dependent migration of NCCs toward the optic vesicle, with the support of specific inactivation with genetic and pharmacological tools. They also help to understand mechanisms of accurate distribution of embryonic cells and of their perturbation by a commonly consumed teratogen, and demonstrate, in addition to its other known developmental functions, a new biological activity of cellular guidance for Shh. PMID:26979762

  16. Sonic hedgehog-mediated epithelial-mesenchymal transition in renal tubulointerstitial fibrosis.

    PubMed

    Bai, Yongheng; Lu, Hong; Lin, Chengcheng; Xu, Yaya; Hu, Dannü; Liang, Yong; Hong, Weilong; Chen, Bicheng

    2016-05-01

    The sonic hedgehog (SHH) signaling pathway plays a critical role in embryonic development, tissue regeneration and organogenesis. The activation of SHH signaling produces profibrogenic effects in various tissues, such as the liver and the biliary ducts. However, the role of SHH signaling in renal fibrogenesis remains to be elucidated. For this purpose, in the present study, we evaluated the hypothesis that activated SHH signaling promotes the acquisition of a myofibroblastic phenotype through the epithelial-mesenchymal transition (EMT), resulting in renal interstitial fibrosis (RIF). Kidney samples from rats subjected to unilateral or bilateral ureteral obstruction exhibited the enhanced expression of SHH-pathway proteins, mesenchymal markers and the decreased expression of epithelial markers. Overactive SHH signaling as well as tubular EMT and RIF in the obstructed kidneys were inhibited by recanalization of the ureter. In vitro, SHH signaling was activated during EMT induction and extracellular matrix (ECM) deposition was observed in transforming growth factor-β1 (TGF-β1)-treated renal tubular epithelial cells [RTECs; NRK-52E cell line]. Exogenous SHH activated SHH signaling and resulted in the upregulated expression of mesenchymal genes, the profibrogenic cytokine TGF-β1, and the downregulated expression of epithelial markers. The blockade of SHH signaling with cyclopamine abolished SHH-mediated EMT as well as the acquisition of a myofibroblastic phenotype, and decreased TGF-β1 expression and ECM production. Thus, taken together, these findings demonstrate that the activation of the SHH signaling pathway promotes the induction of EMT and renal tubulointerstitial fibrosis. The pharmacological inhibition of SHH signaling may potentially be of therapeutic value in the management of fibrotic kidney diseases. PMID:27035418

  17. Primary cilia are critical for Sonic hedgehog-mediated dopaminergic neurogenesis in the embryonic midbrain.

    PubMed

    Gazea, Mary; Tasouri, Evangelia; Tolve, Marianna; Bosch, Viktoria; Kabanova, Anna; Gojak, Christian; Kurtulmus, Bahtiyar; Novikov, Orna; Spatz, Joachim; Pereira, Gislene; Hübner, Wolfgang; Brodski, Claude; Tucker, Kerry L; Blaess, Sandra

    2016-01-01

    Midbrain dopaminergic (mDA) neurons modulate various motor and cognitive functions, and their dysfunction or degeneration has been implicated in several psychiatric diseases. Both Sonic Hedgehog (Shh) and Wnt signaling pathways have been shown to be essential for normal development of mDA neurons. Primary cilia are critical for the development of a number of structures in the brain by serving as a hub for essential developmental signaling cascades, but their role in the generation of mDA neurons has not been examined. We analyzed mutant mouse lines deficient in the intraflagellar transport protein IFT88, which is critical for primary cilia function. Conditional inactivation of Ift88 in the midbrain after E9.0 results in progressive loss of primary cilia, a decreased size of the mDA progenitor domain, and a reduction in mDA neurons. We identified Shh signaling as the primary cause of these defects, since conditional inactivation of the Shh signaling pathway after E9.0, through genetic ablation of Gli2 and Gli3 in the midbrain, results in a phenotype basically identical to the one seen in Ift88 conditional mutants. Moreover, the expansion of the mDA progenitor domain observed when Shh signaling is constitutively activated does not occur in absence of Ift88. In contrast, clusters of Shh-responding progenitors are maintained in the ventral midbrain of the hypomorphic Ift88 mouse mutant, cobblestone. Despite the residual Shh signaling, the integrity of the mDA progenitor domain is severely disturbed, and consequently very few mDA neurons are generated in cobblestone mutants. Our results identify for the first time a crucial role of primary cilia in the induction of mDA progenitors, define a narrow time window in which Shh-mediated signaling is dependent upon normal primary cilia function for this purpose, and suggest that later Wnt signaling-dependent events act independently of primary cilia.

  18. Adenohypophysis formation in the zebrafish and its dependence on sonic hedgehog.

    PubMed

    Herzog, Wiebke; Zeng, Xianchun; Lele, Zsolt; Sonntag, Carmen; Ting, Jing-Wen; Chang, Chi-Yao; Hammerschmidt, Matthias

    2003-02-01

    Formation of the adenohypophysis in mammalian embryos occurs via an invagination of the oral ectoderm to form Rathke's pouch, which becomes exposed to opposing dorsoventral gradients of signaling proteins governing specification of the different hormone-producing pituitary cell types. One signal promoting pituitary cell proliferation and differentiation to ventral cell types is Sonic hedgehog (Shh) from the oral ectoderm. To study pituitary formation and patterning in zebrafish, we cloned four cDNAs encoding different pituitary hormones, prolactin (prl), proopiomelancortin (pomc), thyroid stimulating hormone (tsh), and growth hormone (gh), and analyzed their expression patterns relative to that of the pituitary marker lim3. prl and pomc start to be expressed at the lateral edges of the lim3 expression domain, before pituitary cells move into the head. This indicates that patterning of the pituitary anlage and terminal differentiation of pituitary cells starts while cells are still organized in a placodal fashion at the anterior edge of the developing brain. Following the expression pattern of prl and pomc during development, we show that no pituitary-specific invagination equivalent to Rathke's pouch formation takes place. Rather, pituitary cells move inwards together with stomodeal cells during oral cavity formation, with medial cells of the placode ending up posterior and lateral cells ending up anterior, resulting in an anterior-posterior, rather than a dorsoventral, patterning of the adenohypophysis. Carrying out loss- and gain-of-function experiments, we show that Shh from the ventral diencephalon plays a crucial role during induction, patterning, and growth of the zebrafish adenohypophysis. The phenotypes are very similar to those obtained upon pituitary-specific inactivation or overexpression of Shh in mouse embryo, suggesting that the role of Shh during pituitary development has been largely conserved between fish and mice, despite the different modes of

  19. Hedgehog pathway regulators influence cervical cancer cell proliferation, survival and migration

    SciTech Connect

    Samarzija, Ivana; Beard, Peter

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer Unknown cellular mutations complement papillomavirus-induced carcinogenesis. Black-Right-Pointing-Pointer Hedgehog pathway components are expressed by cervical cancer cells. Black-Right-Pointing-Pointer Hedgehog pathway activators and inhibitors regulate cervical cancer cell biology. Black-Right-Pointing-Pointer Cell immortalization by papillomavirus and activation of Hedgehog are independent. -- Abstract: Human papillomavirus (HPV) infection is considered to be a primary hit that causes cervical cancer. However, infection with this agent, although needed, is not sufficient for a cancer to develop. Additional cellular changes are required to complement the action of HPV, but the precise nature of these changes is not clear. Here, we studied the function of the Hedgehog (Hh) signaling pathway in cervical cancer. The Hh pathway can have a role in a number of cancers, including those of liver, lung and digestive tract. We found that components of the Hh pathway are expressed in several cervical cancer cell lines, indicating that there could exists an autocrine Hh signaling loop in these cells. Inhibition of Hh signaling reduces proliferation and survival of the cervical cancer cells and induces their apoptosis as seen by the up-regulation of the pro-apoptotic protein cleaved caspase 3. Our results indicate that Hh signaling is not induced directly by HPV-encoded proteins but rather that Hh-activating mutations are selected in cells initially immortalized by HPV. Sonic Hedgehog (Shh) ligand induces proliferation and promotes migration of the cervical cancer cells studied. Together, these results indicate pro-survival and protective roles of an activated Hh signaling pathway in cervical cancer-derived cells, and suggest that inhibition of this pathway may be a therapeutic option in fighting cervical cancer.

  20. The buccohypophyseal canal is an ancestral vertebrate trait maintained by modulation in sonic hedgehog signaling

    PubMed Central

    2013-01-01

    Background The pituitary gland is formed by the juxtaposition of two tissues: neuroectoderm arising from the basal diencephalon, and oral epithelium, which invaginates towards the central nervous system from the roof of the mouth. The oral invagination that reaches the brain from the mouth is referred to as Rathke’s pouch, with the tip forming the adenohypophysis and the stalk disappearing after the earliest stages of development. In tetrapods, formation of the cranial base establishes a definitive barrier between the pituitary and oral cavity; however, numerous extinct and extant vertebrate species retain an open buccohypophyseal canal in adulthood, a vestige of the stalk of Rathke’s pouch. Little is currently known about the formation and function of this structure. Here we have investigated molecular mechanisms driving the formation of the buccohypophyseal canal and their evolutionary significance. Results We show that Rathke’s pouch is located at a boundary region delineated by endoderm, neural crest-derived oral mesenchyme and the anterior limit of the notochord, using CD1, R26R-Sox17-Cre and R26R-Wnt1-Cre mouse lines. As revealed by synchrotron X-ray microtomography after iodine staining in mouse embryos, the pouch has a lobulated three-dimensional structure that embraces the descending diencephalon during pituitary formation. Polarisfl/fl; Wnt1-Cre, Ofd1-/- and Kif3a-/- primary cilia mouse mutants have abnormal sonic hedgehog (Shh) signaling and all present with malformations of the anterior pituitary gland and midline structures of the anterior cranial base. Changes in the expressions of Shh downstream genes are confirmed in Gas1-/- mice. From an evolutionary perspective, persistence of the buccohypophyseal canal is a basal character for all vertebrates and its maintenance in several groups is related to a specific morphology of the midline that can be related to modulation in Shh signaling. Conclusion These results provide insight into a poorly

  1. Sonic Hedgehog Signaling Protects Human Hepatocellular Carcinoma Cells Against Ionizing Radiation in an Autocrine Manner

    SciTech Connect

    Chen, Yu-Jen; Lin, Chin-Ping; Hsu, Ming-Ling; Shieh, Hui-Ru; Chao, Nicholas K.; Chao, K.S. Clifford

    2011-07-01

    Purpose: Sonic hedgehog (Shh) signaling is critical to embryogenesis and resistance to chemotherapy. We aimed to examine the role of Shh signaling in the response to radiation of human hepatocellular carcinoma (HCC) cells. Methods and Materials: Response to ionizing radiation therapy (RT) was evaluated by clonogenic assay. Quantitative RT-polymerase chain reaction for patched-1 (PTCH-1) expression was performed. Cytosolic accumulation of Shh and nuclear translocation of Gli-1 were assessed by immunofluorescence. Gli-1 knockdown was done by RNA interference (RNAi). Immunoprecipitation was performed to detect Shh ligand in conditioned medium. Immunofluorescent stain for {gamma}-H2AX was used as an index of DNA double strand breaks (DSB). Expression of proteins related to DNA damage repair was assessed by Western blotting. Results: We found that Shh ligand could protect human HCC HA22T and Sk-Hep1 cells against RT. In HA22T cells, Shh ligand activated the Shh signaling with upregulation of Shh, PTCH-1, and Gli-1 expression. The nuclear translocation of Gli-1 further supports the activation of Gli-1. The radioprotection by Shh ligand was partly blocked by Shh antibody neutralization and was abolished by Gli-1 RNAi, suggesting a critical role of Shh signaling in radiation resistance. Furthermore, we noted that soluble factors secreted into conditioned medium, either constitutively or responding to radiation, by HA22T or Sk-Hep1 cells protected subsequent culturing cells against RT. Immunoprecipitation shows the presence of Shh peptide in conditioned medium. Intriguingly, antibody neutralization of Shh ligand or knockdown of Gli-1 reversed the radioprotective effect of conditioned medium. Furthermore, Shh ligand reduced the RT-induced phosphorylation of checkpoint kinase 1 and impaired the repair of DNA DSB. Conclusions: Activation of Shh signaling protects HCC cells against ionizing radiation in an autocrine manner. Impairment of DNA damage repair might involve

  2. Hedgehog signaling regulates E-cadherin expression for the maintenance of the actin cytoskeleton and tight junctions.

    PubMed

    Xiao, Chang; Ogle, Sally A; Schumacher, Michael A; Schilling, Neal; Tokhunts, Robert A; Orr-Asman, Melissa A; Miller, Marian L; Robbins, David J; Hollande, Frederic; Zavros, Yana

    2010-12-01

    In the stomach, strictly regulated cell adherens junctions are crucial in determining epithelial cell differentiation. Sonic Hedgehog (Shh) regulates epithelial cell differentiation in the adult stomach. We sought to identify whether Shh plays a role in regulating adherens junction protein E-cadherin as a mechanism for epithelial cell differentiation. Mouse nontumorigenic gastric epithelial (IMGE-5) cells treated with Hedgehog signaling inhibitor cyclopamine and anti-Shh 5E1 antibody or transduced with short hairpin RNA against Skinny Hedgehog (IMGE-5(Ski)) were cultured. A mouse model expressing a parietal cell-specific deletion of Shh (HKCre/Shh(KO)) was used to identify further changes in adherens and tight junctions. Inhibition of Hedgehog signaling in IMGE-5 cells caused loss of E-cadherin expression accompanied by disruption of F-actin cortical expression and relocalization of zonula occludens-1 (ZO-1). Loss of E-cadherin was also associated with increased proliferation in IMGE-5(Ski) cells and increased expression of the mucous neck cell lineage marker MUC6. Compared with membrane-expressed E-cadherin and ZO-1 protein in controls, dissociation of E-cadherin/β-catenin and ZO-1/occludin protein complexes was observed in HKCre/Shh(KO) mice. In conclusion, we demonstrate that Hedgehog signaling regulates E-cadherin expression that is required for the maintenance of F-actin cortical expression and stability of tight junction protein ZO-1.

  3. Hedgehog regulates cerebellar progenitor cell and medulloblastoma apoptosis.

    PubMed

    Noguchi, Kevin Kiyoshi; Cabrera, Omar Hoseá; Swiney, Brant S; Salinas-Contreras, Patricia; Smith, Julie Kathryn; Farber, Nuri B

    2015-11-01

    The external granule layer (EGL) is a proliferative region that produces over 90% of the neurons in the cerebellum but can also malignantly transform into a cerebellar tumor called the medulloblastoma (the most common malignant brain tumor in children). Current dogma considers Hedgehog stimulation a potent proliferative signal for EGL neural progenitor cells (NPCs) and medulloblastomas. However, the Hedgehog pathway also acts as a survival signal in the neural tube where it regulates dorsoventral patterning by controlling NPC apoptosis. Here we show that Hedgehog stimulation is also a potent survival signal in the EGL and medulloblastomas that produces a massive apoptotic response within hours of signal loss in mice. This toxicity can be produced by numerous Hedgehog antagonists (vismodegib, cyclopamine, and jervine) and is Bax/Bak dependent but p53 independent. Finally, since glucocorticoids can also induce EGL and medulloblastoma apoptosis, we show that Hedgehog's effects on apoptosis can occur independent of glucocorticoid stimulation. This effect may play a major role in cerebellar development by directing where EGL proliferation occurs thereby morphologically sculpting growth. It may also be a previously unknown major therapeutic effect of Hedgehog antagonists during medulloblastoma therapy. Results are discussed in terms of their implications for both cerebellar development and medulloblastoma treatment. PMID:26319366

  4. Involvement of the sonic hedgehog, patched 1 and bmp2 genes in patterning of the zebrafish dermal fin rays.

    PubMed

    Laforest, L; Brown, C W; Poleo, G; Géraudie, J; Tada, M; Ekker, M; Akimenko, M A

    1998-11-01

    The signaling molecule encoded by Sonic hedgehog (shh) participates in the patterning of several embryonic structures including limbs. During early fin development in zebrafish, a subset of cells in the posterior margin of pectoral fin buds express shh. We have shown that regulation of shh in pectoral fin buds is consistent with a role in mediating the activity of a structure analogous to the zone of polarizing activity (ZPA) (Akimenko and Ekker (1995) Dev. Biol. 170, 243-247). During growth of the bony rays of both paired and unpaired fins, and during fin regeneration, there does not seem to be a region equivalent to the ZPA and one would predict that shh would play a different role, if any, during these processes specific to fish fins. We have examined the expression of shh in the developing fins of 4-week old larvae and in regenerating fins of adults. A subset of cells in the basal layer of the epidermis in close proximity to the newly formed dermal bone structures of the fin rays, the lepidotrichia, express shh, and ptc1 which is thought to encode the receptor of the SHH signal. The expression domain of ptc1 is broader than that of shh and adjacent blastemal cells releasing the dermal bone matrix also express ptc1. Further observations indicate that the bmp2 gene, in addition to being expressed in the same cells of the basal layer of the epidermis as shh, is also expressed in a subset of the ptc1-expressing cells of the blastema. Amputations of caudal fins immediately after the first branching point of the lepidotrichia, and global administration of all-trans-retinoic acid, two procedures known to cause fusion of adjacent rays, result in a transient decrease in the expression of shh, ptc1 and bmp2. The effects of retinoic acid on shh expression occur within minutes after the onset of treatment suggesting direct regulation of shh by retinoic acid. These observations suggest a role for shh, ptc1 and bmp2 in patterning of the dermoskeleton of developing and

  5. Peptide Amphiphile Nanofiber Delivery of Sonic Hedgehog Protein to Reduce Smooth Muscle Apoptosis in the Penis after Cavernous Nerve Resection

    PubMed Central

    Bond, Christopher W.; Angeloni, Nicholas L.; Harrington, Daniel A.; Stupp, Samuel I.; McKenna, Kevin E.; Podlasek, Carol A.

    2011-01-01

    Introduction Erectile dysfunction (ED) is a serious medical condition that affects 16–82% of prostate cancer patients treated by radical prostatectomy and current treatments are ineffective in 50–60% of prostatectomy patients. The reduced efficacy of treatments makes novel therapeutic approaches to treat ED essential. The secreted protein Sonic hedgehog (SHH) is a critical regulator of penile smooth muscle and apoptosis that is decreased in cavernous nerve (CN) injury and diabetic ED models. Past studies using Affi-Gel beads have shown SHH protein to be effective in suppressing apoptosis caused by CN injury. Aim We hypothesize that SHH protein delivered via novel peptide amphiphile (PA) nanofibers will be effective in suppressing CN injury-induced apoptosis. Methods Adult Sprague Dawley rats (n = 50) were used to optimize PA injection in vivo. PA with SHH protein (n = 16) or bovine serum albumin (BSA) (control, n = 14) was injected into adult rats that underwent bilateral CN cut. Rats were sacrificed at 2, 4, and 7 days. Alexa Fluor-labeled SHH protein was used to determine the target of SHH signaling (n = 3). Main Outcome Measures Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and semi-quantitative immunohistochemical analysis for SHH protein and cluster differentiation protein three (CD3) were performed. Results SHH-PA caused a 25% and 16% reduction in apoptosis at 4 and 7 days after CN injury and a 9.3% and 19% increase in SHH protein at 4 and 7 days after CN injury. CD3 protein was not observed in SHH-PA-treated penis. In vitro, 73% of SHH protein diffused from PA within 6 days. Labeled SHH was observed in smooth muscle. Conclusions PA technology is effective in delivering SHH protein to the penis and SHH is effective in suppressing CN injury-induced apoptosis. These results suggest substantial translational potential of this methodology and show that only a short duration of SHH treatment is required to impact the apoptotic index

  6. Relationship between Sonic hedgehog protein, brain-derived neurotrophic factor and oxidative stress in autism spectrum disorders.

    PubMed

    Al-Ayadhi, Laila Y

    2012-02-01

    The etiology of autism spectrum disorders (ASD) is not well known but oxidative stress has been suggested to play a pathological role. We report here that the serum levels of Sonic hedgehog (SHH) protein and brain-derived neurotrophic factor (BDNF) might be linked to oxidative stress in ASD. By using the whole blood or polymorphonuclear leukocytes, we demonstrated that autistic children produced a significantly higher level of oxygen free radicals (OFR). In addition, we found significantly higher levels of serum SHH protein in children with mild as well as severe form of autism. We also found that the serum level of BDNF was significantly reduced in autistic children with mild form of the disorder but not with severe form of the disorder. Our findings are the first to report a correlation between SHH, BDNF and OFR in autistic children, suggesting a pathological role of oxidative stress and SHH in autism spectrum disorders. PMID:21984201

  7. Sonic hedgehog-expressing cells in the developing limb measure time by an intrinsic cell cycle clock.

    PubMed

    Chinnaiya, Kavitha; Tickle, Cheryll; Towers, Matthew

    2014-01-01

    How time is measured is an enduring issue in developmental biology. Classical models of somitogenesis and limb development implicated intrinsic cell cycle clocks, but their existence remains controversial. Here we show that an intrinsic cell cycle clock in polarizing region cells of the chick limb bud times the duration of Sonic hedgehog (Shh) expression, which encodes the morphogen specifying digit pattern across the antero-posterior axis (thumb to little finger). Timing by this clock starts when polarizing region cells fall out of range of retinoic acid signalling. We found that timing of Shh transcription by the cell cycle clock can be reset, thus revealing an embryonic form of self-renewal. In contrast, antero-posterior positional values cannot be reset, suggesting that this may be an important constraint on digit regeneration. Our findings provide the first evidence for an intrinsic cell cycle timer controlling duration and patterning activity of a major embryonic signalling centre.

  8. Zfp423 Regulates Sonic Hedgehog Signaling via Primary Cilium Function

    PubMed Central

    Hamilton, Bruce A.

    2016-01-01

    Zfp423 encodes a 30-zinc finger transcription factor that intersects several canonical signaling pathways. Zfp423 mutations result in ciliopathy-related phenotypes, including agenesis of the cerebellar vermis in mice and Joubert syndrome (JBTS19) and nephronophthisis (NPHP14) in humans. Unlike most ciliopathy genes, Zfp423 encodes a nuclear protein and its developmental expression is complex, leading to alternative proposals for cellular mechanisms. Here we show that Zfp423 is expressed by cerebellar granule cell precursors, that loss of Zfp423 in these precursors leads to cell-intrinsic reduction in proliferation, loss of response to Shh, and primary cilia abnormalities that include diminished frequency of both Smoothened and IFT88 localization. Loss of Zfp423 alters expression of several genes encoding key cilium components, including increased expression of Tulp3. Tulp3 is a direct binding target of Zfp423 and reducing the overexpression of Tulp3 in Zfp423-deficient cells suppresses Smoothened translocation defects. These results define Zfp423 deficiency as a bona fide ciliopathy, acting upstream of Shh signaling, and indicate a mechanism intrinsic to granule cell precursors for the resulting cerebellar hypoplasia. PMID:27727273

  9. YB-1 is elevated in medulloblastoma and drives proliferation in Sonic hedgehog - dependent cerebellar granule neuron progenitor cells and medulloblastoma cells

    PubMed Central

    Dey, Abhinav; Robitaille, Mélanie; Remke, Marc; Maier, Caroline; Malhotra, Anshu; Gregorieff, Alex; Wrana, Jeffrey L; Taylor, Michael D; Angers, Stéphane; Kenney, Anna Marie

    2016-01-01

    Post-natal proliferation of cerebellar granule neuron precursors (CGNPs), proposed cells-of-origin for the SHH-associated subgroup of medulloblastoma (MB), is driven by Sonic Hedgehog (Shh) and Insulin-like Growth Factor (IGF) in the developing cerebellum. Shh induces the oncogene Yes-associated protein (YAP), which drives IGF2 expression in CGNPs and mouse Shh-associated medulloblastomas. To determine how IGF2 expression is regulated downstream of YAP, we carried out an unbiased screen for transcriptional regulators bound to IGF2 promoters. We report that Y-box binding protein-1 (YB-1), an onco-protein regulating transcription and translation, binds to IGF2 promoter P3. We observed that YB-1 is up-regulated across human medulloblastoma subclasses as well as in other varieties of pediatric brain tumors. Utilizing the cerebellar progenitor model for the Shh-subgroup of MB in mice, we show for the first time that YB-1 is induced by Shh in CGNPs. Its expression is YAP-dependent and it is required for IGF2 expression in CGNPs. Finally, both gain-of function and loss-of-function experiments reveal that YB-1 activity is required for sustaining CGNP and medulloblastoma cell (MBC) proliferation. Collectively, our findings describe a novel role for YB-1 in driving proliferation in the developing cerebellum and medulloblastoma cells and they identify the SHH:YAP:YB1:IGF2 axis as a powerful target for therapeutic intervention in medulloblastomas. PMID:26725322

  10. Sonic hedgehog stimulates glycolysis and proliferation of breast cancer cells: Modulation of PFKFB3 activation

    SciTech Connect

    Ge, Xin; Lyu, Pengwei; Gu, Yuanting; Li, Lin; Li, Jingruo; Wang, Yan; Zhang, Linfeng; Fu, Chao; Cao, Zhang

    2015-08-28

    Sonic hesgehog (Shh) signaling has been reported to play an essential role in cancer progression. The mechanism of Shh involved in breast cancer carcinogenesis remains unclear. The present study sought to explore whether Shh signaling could regulate the glycolytic metabolism in breast cancers. Overexpression of the smoothed (Smo) and Gli-1 was found in human primary breast cancers. The expressions of Shh and Gli-1 correlated significantly with tumor size and tumor stage. In vitro, human recombinant Shh (rShh) triggered Smo and Gli-1 expression, promoted glucose utilization and lactate production, and accelerated cell proliferation in MCF-7 and MDA-MB-231 cells. Notably, rShh did not alter 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) expression but augmented PFKFB3 phosphorylation on ser{sup 461}, along with elevated fructose-2,6-bisphosphate (F2,6BP) generation by MCF-7 and MDA-MB-231 cells. This effect could be dampened by Smo siRNA but not by Gli-1 siRNA. In addition, our data showed the upregulated expressions of MAPK by rShh and elevatory PFKFB3 phosphorylation by p38/MAPK activated kinase (MK2). In conclusion, our study characterized a novel role of Shh in promoting glycolysis and proliferation of breast cancer cells via PFKFB3 phosphorylation, which was mediated by Smo and p38/MK2. - Highlights: • Overexpression of Smo and Gli-1 was found in human primary breast cancers. • Shh promoted glucose utilization, lactate production, and cell proliferation. • Shh did not alter PFKFB3 expression but augmented PFKFB3 phosphorylation on ser461. • Shh acts on PFKFB3 phosphorylation via Smo and p38 MAPK/MK2.

  11. Reconstruction of the gene regulatory network involved in the sonic hedgehog pathway with a potential role in early development of the mouse brain.

    PubMed

    Liu, Jinhua; Wang, Xuelong; Li, Juan; Wang, Haifang; Wei, Gang; Yan, Jun

    2014-10-01

    The Sonic hedgehog (Shh) signaling pathway is crucial for pattern formation in early central nervous system development. By systematically analyzing high-throughput in situ hybridization data of E11.5 mouse brain, we found that Shh and its receptor Ptch1 define two adjacent mutually exclusive gene expression domains: Shh+Ptch1- and Shh-Ptch1+. These two domains are associated respectively with Foxa2 and Gata3, two transcription factors that play key roles in specifying them. Gata3 ChIP-seq experiments and RNA-seq assays on Gata3-knockdown cells revealed that Gata3 up-regulates the genes that are enriched in the Shh-Ptch1+ domain. Important Gata3 targets include Slit2 and Slit3, which are involved in the process of axon guidance, as well as Slc18a1, Th and Qdpr, which are associated with neurotransmitter synthesis and release. By contrast, Foxa2 both up-regulates the genes expressed in the Shh+Ptch1- domain and down-regulates the genes characteristic of the Shh-Ptch1+ domain. From these and other data, we were able to reconstruct a gene regulatory network governing both domains. Our work provides the first genome-wide characterization of the gene regulatory network involved in the Shh pathway that underlies pattern formation in the early mouse brain. PMID:25299227

  12. Regulation of Hedgehog Signalling Inside and Outside the Cell

    PubMed Central

    Ramsbottom, Simon A.; Pownall, Mary E.

    2016-01-01

    The hedgehog (Hh) signalling pathway is conserved throughout metazoans and plays an important regulatory role in both embryonic development and adult homeostasis. Many levels of regulation exist that control the release, reception, and interpretation of the hedgehog signal. The fatty nature of the Shh ligand means that it tends to associate tightly with the cell membrane, and yet it is known to act as a morphogen that diffuses to elicit pattern formation. Heparan sulfate proteoglycans (HSPGs) play a major role in the regulation of Hh distribution outside the cell. Inside the cell, the primary cilium provides an important hub for processing the Hh signal in vertebrates. This review will summarise the current understanding of how the Hh pathway is regulated from ligand production, release, and diffusion, through to signal reception and intracellular transduction. PMID:27547735

  13. Characterization of the human oncogene SCL/TAL1 interrupting locus (Stil) mediated Sonic hedgehog (Shh) signaling transduction in proliferating mammalian dopaminergic neurons

    SciTech Connect

    Sun, Lei; Carr, Aprell L.; Li, Ping; Lee, Jessica; McGregor, Mary; Li, Lei

    2014-07-11

    Highlights: • Stil is a human oncogene that is conserved in vertebrate species. • Stil functions in the Shh pathway in mammalian cells. • The expression of Stil is required for mammalian dopaminergic cell proliferation. - Abstract: The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in all vertebrate species. In humans, the expression of Stil is involved in cancer cell survival, apoptosis and proliferation. In this research, we investigated the roles of Stil expression in cell proliferation of mammalian dopaminergic (DA) PC12 cells. Stil functions through the Sonic hedgehog (Shh) signal transduction pathway. Co-immunoprecipitation tests revealed that STIL interacts with Shh downstream components, which include SUFU and GLI1. By examining the expression of Stil, Gli1, CyclinD2 (cell-cycle marker) and PCNA (proliferating cell nuclear antigen), we found that up-regulation of Stil expression (transfection with overexpression plasmids) increased Shh signaling transduction and PC12 cell proliferation, whereas down-regulation of Stil expression (by shRNA) inhibited Shh signaling transduction, and thereby decreased PC12 cell proliferation. Transient transfection of PC12 cells with Stil knockdown or overexpression plasmids did not affect PC12 cell neural differentiation, further indicating the specific roles of Stil in cell proliferation. The results from this research suggest that Stil may serve as a bio-marker for neurological diseases involved in DA neurons, such as Parkinson’s disease.

  14. The Teratogenic Effects of Prenatal Ethanol Exposure Are Exacerbated by Sonic Hedgehog or Gli2 Haploinsufficiency in the Mouse

    PubMed Central

    Kietzman, Henry W.; Everson, Joshua L.; Sulik, Kathleen K.; Lipinski, Robert J.

    2014-01-01

    Disruption of the Hedgehog signaling pathway has been implicated as an important molecular mechanism in the pathogenesis of fetal alcohol syndrome. In severe cases, the abnormalities of the face and brain that result from prenatal ethanol exposure fall within the spectrum of holoprosencephaly. Single allele mutations in the Hh pathway genes Sonic Hedgehog (SHH) and GLI2 cause holoprosencephaly with extremely variable phenotypic penetrance in humans. Here, we tested whether mutations in these genes alter the frequency or severity of ethanol-induced dysmorphology in a mouse model. Timed pregnancies were established by mating Shh+/− or Gli2+/− male mice backcrossed to C57BL/6J strain, with wildtype females. On gestational day 7, dams were treated with two ip doses of 2.9 g/kg ethanol (or vehicle alone), administered four hrs apart. Fetuses were then genotyped and imaged, and the severity of facial dysmorphology was assessed. Following ethanol exposure, mean dysmorphology scores were increased by 3.2- and 6.6-fold in Shh+/− and Gli2+/− groups, respectively, relative to their wildtype littermates. Importantly, a cohort of heterozygous fetuses exhibited phenotypes not typically produced in this model but associated with severe holoprosencephaly, including exencephaly, median cleft lip, otocephaly, and proboscis. As expected, a correlation between the severity of facial dysmorphology and medial forebrain deficiency was observed in affected animals. While Shh+/− and Gli2+/− mice have been described as phenotypically normal, these results illustrate a functional haploinsufficiency of both genes in combination with ethanol exposure. By demonstrating an interaction between specific genetic and environmental risk factors, this study provides important insights into the multifactorial etiology and complex pathogenesis of fetal alcohol syndrome and holoprosencephaly. PMID:24586787

  15. The teratogenic effects of prenatal ethanol exposure are exacerbated by Sonic Hedgehog or GLI2 haploinsufficiency in the mouse.

    PubMed

    Kietzman, Henry W; Everson, Joshua L; Sulik, Kathleen K; Lipinski, Robert J

    2014-01-01

    Disruption of the Hedgehog signaling pathway has been implicated as an important molecular mechanism in the pathogenesis of fetal alcohol syndrome. In severe cases, the abnormalities of the face and brain that result from prenatal ethanol exposure fall within the spectrum of holoprosencephaly. Single allele mutations in the Hh pathway genes Sonic Hedgehog (SHH) and GLI2 cause holoprosencephaly with extremely variable phenotypic penetrance in humans. Here, we tested whether mutations in these genes alter the frequency or severity of ethanol-induced dysmorphology in a mouse model. Timed pregnancies were established by mating Shh(+/-) or Gli2(+/-) male mice backcrossed to C57BL/6J strain, with wildtype females. On gestational day 7, dams were treated with two i.p. doses of 2.9 g/kg ethanol (or vehicle alone), administered four hrs apart. Fetuses were then genotyped and imaged, and the severity of facial dysmorphology was assessed. Following ethanol exposure, mean dysmorphology scores were increased by 3.2- and 6.6-fold in Shh(+/-) and Gli2(+/-) groups, respectively, relative to their wildtype littermates. Importantly, a cohort of heterozygous fetuses exhibited phenotypes not typically produced in this model but associated with severe holoprosencephaly, including exencephaly, median cleft lip, otocephaly, and proboscis. As expected, a correlation between the severity of facial dysmorphology and medial forebrain deficiency was observed in affected animals. While Shh(+/-) and Gli2(+/-) mice have been described as phenotypically normal, these results illustrate a functional haploinsufficiency of both genes in combination with ethanol exposure. By demonstrating an interaction between specific genetic and environmental risk factors, this study provides important insights into the multifactorial etiology and complex pathogenesis of fetal alcohol syndrome and holoprosencephaly.

  16. Mycobacteria-responsive sonic hedgehog signaling mediates programmed death-ligand 1- and prostaglandin E2-induced regulatory T cell expansion

    PubMed Central

    Holla, Sahana; Stephen-Victor, Emmanuel; Prakhar, Praveen; Sharma, Meenu; Saha, Chaitrali; Udupa, Vibha; Kaveri, Srinivas V.; Bayry, Jagadeesh; Balaji, Kithiganahalli Narayanaswamy

    2016-01-01

    CD4+CD25+FoxP3+ regulatory T cells (Tregs) are exploited by mycobacteria to subvert the protective host immune responses. The Treg expansion in the periphery requires signaling by professional antigen presenting cells and in particularly dendritic cells (DC). However, precise molecular mechanisms by which mycobacteria instruct Treg expansion via DCs are not established. Here we demonstrate that mycobacteria-responsive sonic hedgehog (SHH) signaling in human DCs leads to programmed death ligand-1 (PD-L1) expression and cyclooxygenase (COX)-2-catalyzed prostaglandin E2 (PGE2) that orchestrate mycobacterial infection-induced expansion of Tregs. While SHH-responsive transcription factor GLI1 directly arbitrated COX-2 transcription, specific microRNAs, miR-324-5p and miR-338-5p, which target PD-L1 were downregulated by SHH signaling. Further, counter-regulatory roles of SHH and NOTCH1 signaling during mycobacterial-infection of human DCs was also evident. Together, our results establish that Mycobacterium directs a fine-balance of host signaling pathways and molecular regulators in human DCs to expand Tregs that favour immune evasion of the pathogen. PMID:27080341

  17. Sonic Hedgehog (SHH) Promotes the Differentiation of Mouse Cochlear Neural Progenitors via the Math1–Brn3.1 Signaling pathway in vitro

    PubMed Central

    Hu, Xiaohua; Huang, Jianmin; Feng, Ling; Fukudome, Shinji; Hamajima, Yuki; Lin, Jizhen

    2009-01-01

    Sonic hedgehog (SHH) is essential for the development of the cochlear duct that harbors the organ of Corti. However, little is known about the molecular signaling pathway through which SHH promotes the development of the organ of Corti, especially cochlear sensory epithelial cells. In this study, we demonstrated that SHH contributes to the differentiation of cochlear neural progenitors (CNPs), which are derived from the postnatal day 1 organ of Corti in mice. Addition of SHH to CNPs increased the formation of epithelial cell islands, simultaneously activated the expression of Math1 that is a transcription factor for the initial differentiation of auditory hair cells. The increased expression of Math1 then regulated the promoter activity of Brn3.1, another transcription factor that controls the further differentiation and survival of auditory hair cells. Taken together, our data suggest that SHH plays an important role in the promotion of auditory hair cell differentiation via the Math1-Brn3.1 signaling pathway. PMID:19908278

  18. Lycium barbarum Polysaccharides Protect against Trimethyltin Chloride-Induced Apoptosis via Sonic Hedgehog and PI3K/Akt Signaling Pathways in Mouse Neuro-2a Cells

    PubMed Central

    Zhao, Wanyun; Pan, Xiaoqi; Li, Tao; Zhang, Changchun; Shi, Nian

    2016-01-01

    Trimethyltin chloride (TMT) is a classic neurotoxicant that can cause severe neurodegenerative diseases. Some signaling pathways involving cell death play pivotal roles in the central nervous system. In this study, the role of Sonic Hedgehog (Shh) and PI3K/Akt pathways in TMT-induced apoptosis and protective effect of Lycium barbarum polysaccharides (LBP) on mouse neuro-2a (N2a) cells were investigated. Results showed that TMT treatment significantly enhanced apoptosis, upregulated proapoptotic Bax, downregulated antiapoptotic Bcl-2 expression, and increased caspase-3 activity in a dose-dependent manner in N2a cells. TMT induced oxidative stress in cells, performing reactive oxygen species (ROS) and malondialdehyde (MDA) excessive generation, and superoxide dismutase (SOD) activity reduction. TMT significantly decreased phosphorylated glycogen synthase kinase-3β (GSK-3β) and inhibited Shh and PI3K/Akt pathways. However, the addition of LBP upregulated GSK-3β phosphorylation, activated Shh and PI3K/Akt pathways, and eventually reduced apoptosis and oxidative stress caused by TMT. The interaction between Shh and PI3K/Akt pathways was clarified by specific PI3K inhibitor LY294002 or Shh inhibitor GDC-0449. Moreover, LY294002 and GDC-0449 pretreatment both induced phosphorylated GSK-3β downregulation and significantly promoted apoptosis induced by TMT. These results suggest that LBP could reduce TMT-induced N2a cells apoptosis by regulating GSK-3β phosphorylation, Shh, and PI3K/Akt signaling pathways. PMID:27143997

  19. A shared role for sonic hedgehog signalling in patterning chondrichthyan gill arch appendages and tetrapod limbs.

    PubMed

    Gillis, J Andrew; Hall, Brian K

    2016-04-15

    Chondrichthyans (sharks, skates, rays and holocephalans) possess paired appendages that project laterally from their gill arches, known as branchial rays. This led Carl Gegenbaur to propose that paired fins (and hence tetrapod limbs) originally evolved via transformation of gill arches. Tetrapod limbs are patterned by asonic hedgehog(Shh)-expressing signalling centre known as the zone of polarising activity, which establishes the anteroposterior axis of the limb bud and maintains proliferative expansion of limb endoskeletal progenitors. Here, we use loss-of-function, label-retention and fate-mapping approaches in the little skate to demonstrate that Shh secretion from a signalling centre in the developing gill arches establishes gill arch anteroposterior polarity and maintains the proliferative expansion of branchial ray endoskeletal progenitor cells. These findings highlight striking parallels in the axial patterning mechanisms employed by chondrichthyan branchial rays and paired fins/limbs, and provide mechanistic insight into the anatomical foundation of Gegenbaur's gill arch hypothesis. PMID:27095494

  20. Expression and secretion of a biologically active mouse sonic hedgehog protein by the methylotrophic yeast Pichia pastoris.

    PubMed

    Sakuma, Y; Kimura, M; Takabatake, T; Takeshima, K; Fujimura, H

    1999-09-01

    We have successfully secreted the amino-terminal functional domain of mouse sonic hedgehog protein (SHH) into culture fluid using a yeast Pichia pastoris expression system. A cDNA fragment encoding the amino-terminal domain of mouse SHH was inserted downstream of the Saccharomyces cerevisiae alpha-mating factor secretion signal. The DNA fragment was introduced into the host genome by the spheroplast transformation method. Transformants were selected based on their resistance to G418: His+ transformants which showed resistance to over 8 mg G418/ml were selected and analyzed for determination of the plasmid copy number. One His+ clone which has eight copies of the expression cassette per genome was cultured in minimal medium deficient for histidine, and further cultured in buffered medium supplemented with methanol which activates the AOX1 promoter. SDS-PAGE analysis indicated efficient expression and secretion of mouse SHH into culture fluid. The yield of secreted SHH was estimated to be 50 micrograms/ml. Purified protein was assayed for biological activity and found to activate the transcription of the Patched genes (Ptc-1 and Ptc-2) encoding receptors for SHH. PMID:10531654

  1. Mutational analysis of the Sonic Hedgehog gene in 220 newborns with oral clefts in a South American (ECLAMC) population.

    PubMed

    Orioli, Iêda M; Vieira, Alexandre R; Castilla, Eduardo E; Ming, Jeffrey E; Muenke, Maximilian

    2002-02-15

    Oral clefts generally have a multifactorial etiology. A number of genes contribute to the formation of the face and palate. Cleft lip and/or palate can occur in pedigrees with autosomal dominant holoprosencephaly due to mutations in Sonic Hedgehog (SHH). In addition, animal models have shown that SHH is involved in face development. We thus examined the human SHH gene in 220 newborn infants with nonsyndromic oral clefts registered by the Estudio Colaborativo Latinoamericano de Malformaciones Congenitas: ECLAMC (Latin American Collaborative Study of Congenital Malformations). We found 15 variant bands in 13 patients with oral clefts, representing five different base changes, all of which were found by sequencing to represent silent polymorphisms. Four occurred in introns. The alteration occurring in an exon, Ser190Ser, may create a consensus sequence for the 3'splice site 6 bp downstream of the original consensus sequence. Thus, we did not identify any clearly disease-causing mutation in SHH in these patients, and conclude that SHH mutations are not a frequent cause of isolated oral clefts in humans.

  2. Lentivirus-mediated delivery of sonic hedgehog into the striatum stimulates neuroregeneration in a rat model of Parkinson disease.

    PubMed

    Zhang, Yi; Dong, Weiren; Guo, Suiqun; Zhao, Shu; He, Suifen; Zhang, Lihua; Tang, Yinjuan; Wang, Haihong

    2014-12-01

    Parkinson disease (PD) is a progressive neurodegenerative disorder in which the nigrostriatal pathway, consisting of dopaminergic neuronal projections from the substantia nigra to the striatum, degenerates. Viral transduction is currently the most promising in vivo strategy for delivery of therapeutic proteins into the brain for treatment of PD. Sonic hedgehog (Shh) is necessary for cell proliferation, differentiation and neuroprotection in the central nervous system. In this study, we investigated the effects of overexpressed N-terminal product of SHH (SHH-N) in a PD model rat. A lentiviral vector containing SHH-N was stereotactically injected into the striatum 24 h after a striatal 6-OHDA lesion. We found that overexpressed SHH-N attenuated behavioral deficits and reduced the loss of dopamine neurons in the substantia nigra and the loss of dopamine fibers in the striatum. In addition, fluoro-ruby-labeled nigrostriatal projections were also repaired. Together, our results demonstrate the feasibility and efficacy of using the strategy of lentivirus-mediated Shh-N delivery to delay nigrostriatal pathway degeneration. This strategy holds the potential for therapeutic application in the treatment of PD.

  3. Effect of methacrylic acid beads on the sonic hedgehog signaling pathway and macrophage polarization in a subcutaneous injection mouse model.

    PubMed

    Lisovsky, Alexandra; Zhang, David K Y; Sefton, Michael V

    2016-08-01

    Poly(methacrylic acid-co-methyl methacrylate) (MAA) beads promote a vascular regenerative response when used in diabetic wound healing. Previous studies reported that MAA beads modulated the expression of sonic hedgehog (Shh) and inflammation related genes in diabetic wounds. The aim of this work was to follow up on these observations in a subcutaneous injection model to study the host response in the absence of the confounding factors of diabetic wound healing. In this model, MAA beads improved vascularization in healthy mice of both sexes compared to control poly(methyl methacrylate) (MM) beads, with a stronger effect seen in males than females. MAA-induced vessels were perfusable, as evidenced from the CLARITY-processed images. In Shh-Cre-eGFP/Ptch1-LacZ non-diabetic transgenic mice, the increased vessel formation was accompanied by a higher density of cells expressing GFP (Shh) and β-Gal (patched 1, Ptch1) suggesting MAA enhanced the activation of the Shh pathway. Ptch1 is the Shh receptor and a target of the pathway. MAA beads also modulated the inflammatory cell infiltrate in CD1 mice: more neutrophils and more macrophages were noted with MAA relative to MM beads at days 1 and 7, respectively. In addition, MAA beads biased macrophages towards a MHCII-CD206+ ("M2") polarization state. This study suggests that the Shh pathway and an altered inflammatory response are two elements of the complex mechanism whereby MAA-based biomaterials effect vascular regeneration. PMID:27264502

  4. Temozolomide resistance in glioblastoma occurs by miRNA-9-targeted PTCH1, independent of sonic hedgehog level.

    PubMed

    Munoz, Jessian L; Rodriguez-Cruz, Vivian; Ramkissoon, Shakti H; Ligon, Keith L; Greco, Steven J; Rameshwar, Pranela

    2015-01-20

    Glioblastoma Multiforme (GBM), the most common and lethal adult primary tumor of the brain, showed a link between Sonic Hedgehog (SHH) pathway in the resistance to temozolomide (TMZ). PTCH1, the SHH receptor, can tonically represses signaling by endocytosis. We asked how the decrease in PTCH1 in GBM cells could lead to TMZ-resistance. TMZ resistant GBM cells have increased PTCH1 mRNA and reduced protein. Knockdown of Dicer, a Type III RNAase, indicated that miRNAs can explain the decreased PTCH1 in TMZ resistant cells. Computational studies, real-time PCR, reporter gene studies, western blots, target protector oligos and ectopic expression identified miR-9 as the target of PTCH1 in resistant GBM cells with concomitant activation of SHH signaling. MiR-9 mediated increases in the drug efflux transporters, MDR1 and ABCG2. MiR-9 was increased in the tissues from GBM patients and in an early passage GBM cell line from a patient with recurrent GBM but not from a naïve patient. Pharmacological inhibition of SHH signaling sensitized the GBM cells to TMZ. Taken together, miR-9 targets PTCH1 in GBM cells by a SHH-independent method in GBM cells for TMZ resistance. The identified pathways could lead to new strategies to target GBM with combinations of drugs. PMID:25595896

  5. Protein kinase A activation inhibits oncogenic Sonic hedgehog signalling and suppresses basal cell carcinoma of the skin.

    PubMed

    Makinodan, Eri; Marneros, Alexander G

    2012-11-01

    Basal cell carcinoma of the skin (BCC) is caused by constitutive activation of the Sonic hedgehog (Shh) pathway, mainly through mutations either in the Shh receptor Patched (PTCH) or in its co-receptor Smoothened (Smo). Inhibitors of this pathway that are currently in clinical trials inhibit Smo. However, mutations in Smo can result in resistance to these inhibitors. To target most BCCs and avoid acquired resistance because of Smo mutations, inhibiting the Shh-pathway downstream of Smo is critical. Attractive downstream targets would be at the level of Gli proteins, the transcriptional activators of this pathway in BCCs. Previously it has been shown that Gli1 and Gli2, when phosphorylated by protein kinase A (PKA), are targeted for proteosomal degradation. Here we show that PKA activation via the cAMP agonist forskolin is sufficient to completely abolish oncogenic Smo activity in vitro. In an inducible BCC mouse model due to a Smo mutation that confers resistance to current Smo inhibitors, topical forskolin treatment significantly reduced Gli1 mRNA levels and resulted in strongly suppressed BCC tumor growth. Our data show that forskolin inhibits the growth of even those BCCs that are resistant to Smo inhibitors and provide a proof-of-principle framework for the development of topically applied human skin-permeable novel pharmacologic inhibitors of oncogenic Shh-signaling through PKA activation. PMID:23163650

  6. Activation of sonic hedgehog signaling attenuates oxidized low-density lipoprotein-stimulated brain microvascular endothelial cells dysfunction in vitro.

    PubMed

    Jiang, Xiu-Long; Chen, Ting; Zhang, Xu

    2015-01-01

    The study was performed to investigate the role of sonic hedgehog (SHH) in the oxidized low-density lipoprotein (oxLDL)-induced blood-brain barrier (BBB) disruption. The primary mouse brain microvascular endothelial cells (MBMECs) were exposed to oxLDL. The results indicated that treatment of MBMECs with oxLDL decreased the cell viability, and oxidative stress was involved in oxLDL-induce MBMECs dysfunction with increasing intracellular ROS and MDA formation as well as decreasing NO release and eNOS mRNA expression. In addition, SHH signaling components, such as SHH, Smo and Gli1, mRNA and protein levels were significantly decreased after incubation with increasing concentrations of oxLDL. Treatment with oxLDL alone or SHH loss-of-function significantly increased the permeability of MBMECs, and overexpression of SHH attenuated oxLDL-induced elevation of permeability in MBMECs. Furthermore, SHH gain-of-function could reverse oxLDL-induced apoptosis through inhibition caspase3 and caspase8 levels in MBMECs. Taken together, these results demonstrated that the suppression of SHH in MBMECs might contribute to the oxLDL-induced disruption of endothelial barrier. However, the overexpression of SHH could reverse oxLDL-induced endothelial cells dysfunction in vitro.

  7. Disruption of sonic hedgehog signaling in Ellis-van Creveld dwarfism confers protection against bipolar affective disorder.

    PubMed

    Ginns, E I; Galdzicka, M; Elston, R C; Song, Y E; Paul, S M; Egeland, J A

    2015-10-01

    Ellis-van Creveld syndrome, an autosomal recessively inherited chondrodysplastic dwarfism, is frequent among Old Order Amish of Pennsylvania. Decades of longitudinal research on bipolar affective disorder (BPAD) revealed cosegregation of high numbers of EvC and Bipolar I (BPI) cases in several large Amish families descending from the same pioneer. Despite the high prevalence of both disorders in these families, no EvC individual has ever been reported with BPI. The proximity of the EVC gene to our previously reported chromosome 4p16 BPAD locus with protective alleles, coupled with detailed clinical observations that EvC and BPI do not occur in the same individuals, led us to hypothesize that the genetic defect causing EvC in the Amish confers protection from BPI. This hypothesis is supported by a significant negative association of these two disorders when contrasted with absence of disease (P=0.029, Fisher's exact test, two-sided, verified by permutation to estimate the null distribution of the test statistic). As homozygous Amish EVC mutations causing EvC dwarfism do so by disrupting sonic hedgehog (Shh) signaling, our data implicate Shh signaling in the underlying pathophysiology of BPAD. Understanding how disrupted Shh signaling protects against BPI could uncover variants in the Shh pathway that cause or increase risk for this and related mood disorders.

  8. Embryonic tongue morphogenesis in an organ culture model of mouse mandibular arches: blocking Sonic hedgehog signaling leads to microglossia.

    PubMed

    Torii, Daisuke; Soeno, Yuuichi; Fujita, Kazuya; Sato, Kaori; Aoba, Takaaki; Taya, Yuji

    2016-01-01

    Mouse tongue development is initiated with the formation of lateral lingual swellings just before fusion between the mediodorsal surfaces of the mandibular arches at around embryonic day 11.0. Here, we investigated the role of Sonic hedgehog (Shh) signaling in embryonic mouse tongue morphogenesis. For this, we used an organ culture model of the mandibular arches from mouse embryos at embryonic day 10.5. When the Shh signaling inhibitor jervine was added to the culture medium for 24-96 h, the formation of lateral lingual swellings and subsequent epithelial invagination into the mesenchyme were impaired markedly, leading to a hypoplastic tongue with an incomplete oral sulcus. Notably, jervine treatment reduced the proliferation of non-myogenic mesenchymal cells at the onset of forming the lateral lingual swellings, whereas it did not affect the proliferation and differentiation of a myogenic cell lineage, which created a cell community at the central circumferential region of the lateral lingual swellings as seen in vivo and in control cultures lacking the inhibitor. Thus, epithelium-derived Shh signaling stimulates the proliferation of non-myogenic mesenchymal cells essential for forming lateral lingual swellings and contributes to epithelial invagination into the mesenchyme during early tongue development.

  9. Distinct Sonic Hedgehog signaling dynamics specify floor plate and ventral neuronal progenitors in the vertebrate neural tube.

    PubMed

    Ribes, Vanessa; Balaskas, Nikolaos; Sasai, Noriaki; Cruz, Catarina; Dessaud, Eric; Cayuso, Jordi; Tozer, Samuel; Yang, Lin Lin; Novitch, Ben; Marti, Elisa; Briscoe, James

    2010-06-01

    The secreted ligand Sonic Hedgehog (Shh) organizes the pattern of cellular differentiation in the ventral neural tube. For the five neuronal subtypes, increasing levels and durations of Shh signaling direct progenitors to progressively more ventral identities. Here we demonstrate that this mode of action is not applicable to the generation of the most ventral cell type, the nonneuronal floor plate (FP). In chick and mouse embryos, FP specification involves a biphasic response to Shh signaling that controls the dynamic expression of key transcription factors. During gastrulation and early somitogenesis, FP induction depends on high levels of Shh signaling. Subsequently, however, prospective FP cells become refractory to Shh signaling, and this is a prerequisite for the elaboration of their identity. This prompts a revision to the model of graded Shh signaling in the neural tube, and provides insight into how the dynamics of morphogen signaling are deployed to extend the patterning capacity of a single ligand. In addition, we provide evidence supporting a common scheme for FP specification by Shh signaling that reconciles mechanisms of FP development in teleosts and amniotes.

  10. CD44, Sonic Hedgehog, and Gli1 Expression Are Prognostic Biomarkers in Gastric Cancer Patients after Radical Resection

    PubMed Central

    Jian-Hui, Chen; Er-Tao, Zhai; Si-Le, Chen; Hui, Wu; Kai-Ming, Wu; Xin-Hua, Zhang; Chuang-Qi, Chen; Shi-Rong, Cai; Yu-Long, He

    2016-01-01

    Aim. CD44 and Sonic Hedgehog (Shh) signaling are important for gastric cancer (GC). However, the clinical impact, survival, and recurrence outcome of CD44, Shh, and Gli1 expressions in GC patients following radical resection have not been elucidated. Patients and Methods. CD44, Shh, and Gli1 protein levels were quantified by immunohistochemistry (IHC). The association between CD44, Shh, and Gli1 expression and clinicopathological features or prognosis of GC patients was determined. The biomarker risk score was calculated by the IHC staining score of CD44, Shh, and Gli1 protein. Results. The IHC positive staining of CD44, Shh, and Gli1 proteins was correlated with larger tumour size, worse gross type and histological type, and advanced TNM stage, which also predicted shorter overall survival (OS) and disease-free survival (DFS) after radical resection. Multivariate analysis indicated the Gli1 protein and Gli1, CD44 proteins were predictive biomarkers for OS and DFS, respectively. If biomarker risk score was taken into analysis, it was the independent prognostic factor for OS and DFS. Conclusions. CD44 and Shh signaling are important biomarkers for tumour aggressiveness, survival, and recurrence in GC. PMID:26839535

  11. Sonic hedgehog, TBX18, and TSHZ3 proteins involved in pyeloureteral motility development are overexpressed in ureteropelvic junction obstruction

    PubMed Central

    Yilmaz, Omer; Nese, Nalan; Dalgic, Mustafa; Kesici, Gonca P.; Genc, Abdulkadir; Taneli, Can; Gunsar, Cuneyt; Sencan, Aydın; Cayirli, Hasan; Isisag, Aydın

    2016-01-01

    Objectives: To compare pathological samples obtained from cases that underwent surgery for ureteropelvic junction (UPJ) obstruction with samples obtained during autopsies of subjects. Methods: Retrospectively, 42 patients who had undergone surgery due to UPJ obstruction (group 1) were included in the study. Histopathological and immunohistochemical features for sonic hedgehog (SHH), TBX18, and TSHZ3 of UPJ were evaluated and findings were compared with 20 autopsy cases (group 2). Results: In group 1, the scores were statistically significantly higher in terms of cytoplasmic SHH, nuclear TBX18, cytoplasmic and nuclear TSHZ3 staining. Statistically, no correlation was found between age and the staining scores belonging to these 3 antibodies in group 1 and group 2. Intense inflammation was found to be related with nuclear staining for TBX18. Conclusion: Gene product expressions of SHH, TBX18 and TSHZ3 are statistically higher in patients with UPJ obstruction, when compared with control group. The explanation may be the reactivation of the processes, which had shown their effects in the embryological period, due to the chronic inflammation and long-term micro-trauma created by the disease. PMID:27381532

  12. Activation of sonic hedgehog signaling attenuates oxidized low-density lipoprotein-stimulated brain microvascular endothelial cells dysfunction in vitro.

    PubMed

    Jiang, Xiu-Long; Chen, Ting; Zhang, Xu

    2015-01-01

    The study was performed to investigate the role of sonic hedgehog (SHH) in the oxidized low-density lipoprotein (oxLDL)-induced blood-brain barrier (BBB) disruption. The primary mouse brain microvascular endothelial cells (MBMECs) were exposed to oxLDL. The results indicated that treatment of MBMECs with oxLDL decreased the cell viability, and oxidative stress was involved in oxLDL-induce MBMECs dysfunction with increasing intracellular ROS and MDA formation as well as decreasing NO release and eNOS mRNA expression. In addition, SHH signaling components, such as SHH, Smo and Gli1, mRNA and protein levels were significantly decreased after incubation with increasing concentrations of oxLDL. Treatment with oxLDL alone or SHH loss-of-function significantly increased the permeability of MBMECs, and overexpression of SHH attenuated oxLDL-induced elevation of permeability in MBMECs. Furthermore, SHH gain-of-function could reverse oxLDL-induced apoptosis through inhibition caspase3 and caspase8 levels in MBMECs. Taken together, these results demonstrated that the suppression of SHH in MBMECs might contribute to the oxLDL-induced disruption of endothelial barrier. However, the overexpression of SHH could reverse oxLDL-induced endothelial cells dysfunction in vitro. PMID:26722472

  13. Hedgehog signaling regulates gene expression in planarian glia

    PubMed Central

    Wang, Irving E; Lapan, Sylvain W; Scimone, M Lucila; Clandinin, Thomas R; Reddien, Peter W

    2016-01-01

    Hedgehog signaling is critical for vertebrate central nervous system (CNS) development, but its role in CNS biology in other organisms is poorly characterized. In the planarian Schmidtea mediterranea, hedgehog (hh) is expressed in medial cephalic ganglia neurons, suggesting a possible role in CNS maintenance or regeneration. We performed RNA sequencing of planarian brain tissue following RNAi of hh and patched (ptc), which encodes the Hh receptor. Two misregulated genes, intermediate filament-1 (if-1) and calamari (cali), were expressed in a previously unidentified non-neural CNS cell type. These cells expressed orthologs of astrocyte-associated genes involved in neurotransmitter uptake and metabolism, and extended processes enveloping regions of high synapse concentration. We propose that these cells are planarian glia. Planarian glia were distributed broadly, but only expressed if-1 and cali in the neuropil near hh+ neurons. Planarian glia and their regulation by Hedgehog signaling present a novel tractable system for dissection of glia biology. DOI: http://dx.doi.org/10.7554/eLife.16996.001 PMID:27612382

  14. Modulation of Sonic hedgehog signaling and WW domain containing oxidoreductase WOX1 expression enhances radiosensitivity of human glioblastoma cells

    PubMed Central

    Chiang, Ming-Fu; Chen, Hsin-Hong; Chi, Chih-Wen; Sze, Chun-I; Hsu, Ming-Ling; Shieh, Hui-Ru; Lin, Chin-Ping; Tsai, Jo-Ting

    2015-01-01

    WW domain containing oxidoreductase, designated WWOX, FOR or WOX1, is a known pro-apoptotic factor when ectopically expressed in various types of cancer cells, including glioblastoma multiforme (GBM). The activation of sonic hedgehog (Shh) signaling, especially paracrine Shh secretion in response to radiation, is associated with impairing the effective irradiation of cancer cells. Here, we examined the role of Shh signaling and WOX1 overexpression in the radiosensitivity of human GBM cells. Our results showed that ionizing irradiation (IR) increased the cytoplasmic Shh and nuclear Gli-1 content in GBM U373MG and U87MG cells. GBM cells with exogenous Shh treatment exhibited similar results. Pretreatment with Shh peptides protected U373MG and U87MG cells against IR in a dose-dependent manner. Cyclopamine, a Hedgehog/Smoothened (SMO) inhibitor, reversed the protective effect of Shh in U87MG cells. Cyclopamine increased Shh plus IR-induced H2AX, a marker of DNA double-strand breaks, in these cells. To verify the role of Shh signaling in the radiosensitivity of GBM cells, we tested the effect of the Gli family zinc finger 1 (Gli-1) inhibitor zerumbone and found that it could sensitize GBM cells to IR. We next examined the role of WOX1 in radiosensitivity. Overexpression of WOX1 enhanced the radiosensitivity of U87MG (possessing wild type p53 or WTp53) but not U373MG (harboring mutant p53 or MTp53) cells. Pretreatment with Shh peptides protected both WOX1-overexpressed U373MG and U87MG cells against IR and increased the cytoplasmic Shh and nuclear Gli-1 content. Zerumbone enhanced the radiosensitivity of WOX1-overexpressed U373MG and U87MG cells. In conclusion, overexpression of WOX1 preferentially sensitized human GBM cells possessing wild type p53 to radiation therapy. Blocking of Shh signaling may enhance radiosensitivity independently of the expression of p53 and WOX1. The crosstalk between Shh signaling and WOX1 expression in human glioblastoma warrants further

  15. Prenatal cadmium exposure dysregulates sonic hedgehog and Wnt/beta-catenin signaling in the thymus resulting in altered thymocyte development

    SciTech Connect

    Hanson, Miranda L.; Brundage, Kathleen M.; Schafer, Rosana; Tou, Janet C.; Barnett, John B.

    2010-01-15

    Cadmium (Cd) is both an environmental pollutant and a component of cigarette smoke. Although evidence demonstrates that adult exposure to Cd causes changes in the immune system, there are limited reports in the literature of immunomodulatory effects of prenatal exposure to Cd. The sonic hedgehog (Shh) and Wnt/beta-catenin pathways are required for thymocyte maturation. Several studies have demonstrated that Cd exposure affects these pathways in different organ systems. This study was designed to investigate the effect of prenatal Cd exposure on thymocyte development, and to determine if these effects were linked to dysregulation of Shh and Wnt/beta-catenin pathways. Pregnant C57Bl/6 mice were exposed to an environmentally relevant dose (10 ppm) of Cd throughout pregnancy and effects on the thymus were assessed on the day of birth. Thymocyte phenotype was determined by flow cytometry. A Gli:luciferase reporter cell line was used to measure Shh signaling. Transcription of target genes and translation of key components of both signaling pathways were assessed using real-time RT-PCR and western blot, respectively. Prenatal Cd exposure increased the number of CD4{sup +} cells and a subpopulation of double-negative cells (DN; CD4{sup -}CD8{sup -}), DN4 (CD44{sup -}CD25{sup -}). Shh and Wnt/beta-catenin signaling were both decreased in the thymus. Target genes of Shh (Patched1 and Gli1) and Wnt/beta-catenin (c-fos, and c-myc) were affected differentially among thymocyte subpopulations. These findings suggest that prenatal exposure to Cd dysregulates two signaling pathways in the thymus, resulting in altered thymocyte development.

  16. Oxidative stress contributes to the impaired sonic hedgehog pathway in type 1 diabetic mice with myocardial infarction

    PubMed Central

    XIAO, QING; YANG, YA; ZHAO, XIAO-YA; HE, LI-SHAN; QIN, YUAN; HE, YAN-HUA; ZHANG, GUI-PING; LUO, JIAN-DONG

    2015-01-01

    Our previous study demonstrated that an impaired sonic hedgehog (Shh) pathway contributed to cardiac dysfunction in type 1 diabetic mice with myocardial infarction (MI). The present study aimed to test the hypothesis that oxidative stress may contribute to the impaired Shh pathway and cardiac dysfunction in type 1 diabetic mice with MI. Streptozotocin-induced type 1 diabetic mice (C57/Bl6, male) and rat neonatal cardiomyocytes were used in the present study. Mice were randomly assigned to undergo ligation of the coronary artery or pseudosurgery. A potent antioxidant Tempol was administered in vivo and in vitro. Cardiac function was assessed by echocardiography, capillary density by immunohistochemisty, percentage of myocardial infarct using Massons trichrome staining, reactive oxygen species detection using dihydroethidium dye or 2,7-dichlorofluorescein diacetate probe and protein expression levels of the Shh pathway by western blot analysis. The antioxidant Tempol was shown to significantly increase myocardial protein expression levels of Shh and patched-1 (Ptc1) at 7–18 weeks and improved cardiac function at 18 weeks in type 1 diabetic mice, as compared with mice receiving no drug treatment. Furthermore, myocardial protein expression levels of Shh and Ptc1 were significantly upregulated on day 7 after MI, and capillary density was enhanced. In addition, the percentage area of myocardial infarct was reduced, and the cardiac dysfunction and survival rate were improved on day 21 in diabetic mice treated with Tempol. In vitro, treatment of rat neonatal cardiomyocytes with a mixture of xanthine oxidase and xanthine decreased protein expression levels of Shh and Ptc1 in a concentration-dependent manner, and Tempol attenuated this effect. These results indicate that oxidative stress may contribute to an impaired Shh pathway in type 1 diabetic mice, leading to diminished myocardial healing and cardiac dysfunction. Antioxidative strategies aimed at restoring the

  17. Duration of culture and sonic hedgehog signaling differentially specify PV versus SST cortical interneuron fates from embryonic stem cells.

    PubMed

    Tyson, Jennifer A; Goldberg, Ethan M; Maroof, Asif M; Xu, Qing; Petros, Timothy J; Anderson, Stewart A

    2015-04-01

    Medial ganglionic eminence (MGE)-derived GABAergic cortical interneurons (cINs) consist of multiple subtypes that are involved in many cortical functions. They also have a remarkable capacity to migrate, survive and integrate into cortical circuitry after transplantation into postnatal cortex. These features have engendered considerable interest in generating distinct subgroups of interneurons from pluripotent stem cells (PSCs) for the study of interneuron fate and function, and for the development of cell-based therapies. Although advances have been made, the capacity to generate highly enriched pools of subgroup fate-committed interneuron progenitors from PSCs has remained elusive. Previous studies have suggested that the two main MGE-derived interneuron subgroups--those expressing somatostatin (SST) and those expressing parvalbumin (PV)--are specified in the MGE from Nkx2.1-expressing progenitors at higher or lower levels of sonic hedgehog (Shh) signaling, respectively. To further explore the role of Shh and other factors in cIN fate determination, we generated a reporter line such that Nkx2.1-expressing progenitors express mCherry and postmitotic Lhx6-expressing MGE-derived interneurons express GFP. Manipulations of Shh exposure and time in culture influenced the subgroup fates of ESC-derived interneurons. Exposure to higher Shh levels, and collecting GFP-expressing precursors at 12 days in culture, resulted in the strongest enrichment for SST interneurons over those expressing PV, whereas the strongest enrichment for PV interneurons was produced by lower Shh and by collecting mCherry-expressing cells after 17 days in culture. These findings confirm that fate determination of cIN subgroups is crucially influenced by Shh signaling, and provide a system for the further study of interneuron fate and function. PMID:25804737

  18. Genetic Variations in the Sonic Hedgehog Pathway Affect Clinical Outcomes in Non-muscle–invasive Bladder Cancer

    PubMed Central

    Chen, Meng; Hildebrandt, Michelle A.T.; Clague, Jessica; Kamat, Ashish M.; Picornell, Antoni; Chang, Joshua; Zhang, Xiaofan; Izzo, Julie; Yang, Hushan; Lin, Jie; Gu, Jian; Chanock, Stephen; Kogevinas, Manolis; Rothman, Nathaniel; Silverman, Debra T.; Garcia-Closas, Montserrat; Grossman, H. Barton; Dinney, Colin P.; Malats, Núria; Wu, Xifeng

    2010-01-01

    Sonic hedgehog (Shh) pathway genetic variations may affect bladder cancer risk and clinical outcomes; therefore, we genotyped 177 SNPs in 11 Shh pathway genes in a study including 803 bladder cancer cases and 803 controls. We assessed SNP associations with cancer risk and clinical outcomes in 419 cases of non-muscle invasive bladder cancer (NMIBC) and 318 cases of muscle invasive and metastatic bladder cancer (MiMBC). Only 3 SNPs (GLI3 rs3823720, rs3735361, rs10951671) reached nominal significance in association with risk (P≤0.05), which became non-significant after adjusting for multiple comparisons. Nine SNPs reached a nominally significant individual association with recurrence of NMIBC in patients who received transurethral resection (TUR) only (P≤0.05), of which 2 (SHH rs1233560 and GLI2 rs11685068) were replicated independently in 356 TUR-only NMIBC patients with P-values of 1.0×10−3 (SHH rs1233560) and 1.3×10−3 (GLI2 rs11685068). Nine SNPs also reached a nominally significant individual association with clinical outcome of NMIBC patients who received Bacillus Calmette-Guérin (BCG; P≤0.05), of which 2, the independent GLI3 variants rs6463089 and rs3801192, remained significant after adjusting for multiple comparisons (P=2×10−4 and 9×10−4, respectively). The wild-type genotype of either of these SNPs was associated with a lower recurrence rate and longer recurrence-free survival (versus the variants). Although 3 SNPs (GLI2 rs735557, GLI2 rs4848632, and SHH rs208684) showed nominal significance in association with overall survival in MiMBC patients (P≤0.05), none remained significant after multiple-comparison adjustments. Germline genetic variations in the Shh pathway predicted clinical outcomes of TUR and BCG for NMIBC patients. PMID:20858759

  19. Inhibition of APP gamma-secretase restores Sonic Hedgehog signaling and neurogenesis in the Ts65Dn mouse model of Down syndrome.

    PubMed

    Giacomini, Andrea; Stagni, Fiorenza; Trazzi, Stefania; Guidi, Sandra; Emili, Marco; Brigham, Elizabeth; Ciani, Elisabetta; Bartesaghi, Renata

    2015-10-01

    Neurogenesis impairment starting from early developmental stages is a key determinant of intellectual disability in Down syndrome (DS). Previous evidence provided a causal relationship between neurogenesis impairment and malfunctioning of the mitogenic Sonic Hedgehog (Shh) pathway. In particular, excessive levels of AICD (amyloid precursor protein intracellular domain), a cleavage product of the trisomic gene APP (amyloid precursor protein) up-regulate transcription of Ptch1 (Patched1), the Shh receptor that keeps the pathway repressed. Since AICD results from APP cleavage by γ-secretase, the goal of the current study was to establish whether treatment with a γ-secretase inhibitor normalizes AICD levels and restores neurogenesis in trisomic neural precursor cells. We found that treatment with a selective γ-secretase inhibitor (ELND006; ELN) restores proliferation in neurospheres derived from the subventricular zone (SVZ) of the Ts65Dn mouse model of DS. This effect was accompanied by reduction of AICD and Ptch1 levels and was prevented by inhibition of the Shh pathway with cyclopamine. Treatment of Ts65Dn mice with ELN in the postnatal period P3-P15 restored neurogenesis in the SVZ and hippocampus, hippocampal granule cell number and synapse development, indicating a positive impact of treatment on brain development. In addition, in the hippocampus of treated Ts65Dn mice there was a reduction in the expression levels of various genes that are transcriptionally regulated by AICD, including APP, its origin substrate. Inhibitors of γ-secretase are currently envisaged as tools for the cure of Alzheimer's disease because they lower βamyloid levels. Current results provide novel evidence that γ-secretase inhibitors may represent a strategy for the rescue of neurogenesis defects in DS. PMID:26254735

  20. Inhibition of APP gamma-secretase restores Sonic Hedgehog signaling and neurogenesis in the Ts65Dn mouse model of Down syndrome.

    PubMed

    Giacomini, Andrea; Stagni, Fiorenza; Trazzi, Stefania; Guidi, Sandra; Emili, Marco; Brigham, Elizabeth; Ciani, Elisabetta; Bartesaghi, Renata

    2015-10-01

    Neurogenesis impairment starting from early developmental stages is a key determinant of intellectual disability in Down syndrome (DS). Previous evidence provided a causal relationship between neurogenesis impairment and malfunctioning of the mitogenic Sonic Hedgehog (Shh) pathway. In particular, excessive levels of AICD (amyloid precursor protein intracellular domain), a cleavage product of the trisomic gene APP (amyloid precursor protein) up-regulate transcription of Ptch1 (Patched1), the Shh receptor that keeps the pathway repressed. Since AICD results from APP cleavage by γ-secretase, the goal of the current study was to establish whether treatment with a γ-secretase inhibitor normalizes AICD levels and restores neurogenesis in trisomic neural precursor cells. We found that treatment with a selective γ-secretase inhibitor (ELND006; ELN) restores proliferation in neurospheres derived from the subventricular zone (SVZ) of the Ts65Dn mouse model of DS. This effect was accompanied by reduction of AICD and Ptch1 levels and was prevented by inhibition of the Shh pathway with cyclopamine. Treatment of Ts65Dn mice with ELN in the postnatal period P3-P15 restored neurogenesis in the SVZ and hippocampus, hippocampal granule cell number and synapse development, indicating a positive impact of treatment on brain development. In addition, in the hippocampus of treated Ts65Dn mice there was a reduction in the expression levels of various genes that are transcriptionally regulated by AICD, including APP, its origin substrate. Inhibitors of γ-secretase are currently envisaged as tools for the cure of Alzheimer's disease because they lower βamyloid levels. Current results provide novel evidence that γ-secretase inhibitors may represent a strategy for the rescue of neurogenesis defects in DS.

  1. Inhibition of APP gamma-secretase restores Sonic Hedgehog signaling and neurogenesis in the Ts65Dn mouse model of Down syndrome

    PubMed Central

    Giacomini, Andrea; Stagni, Fiorenza; Trazzi, Stefania; Guidi, Sandra; Emili, Marco; Brigham, Elizabeth; Ciani, Elisabetta; Bartesaghi, Renata

    2015-01-01

    Neurogenesis impairment starting from early developmental stages is a key determinant of intellectual disability in Down syndrome (DS). Previous evidence provided a causal relationship between neurogenesis impairment and malfunctioning of the mitogenic Sonic Hedgehog (Shh) pathway. In particular, excessive levels of AICD (amyloid precursor protein intracellular domain), a cleavage product of the trisomic gene APP (amyloid precursor protein) up-regulate transcription of Ptch1 (Patched1), the Shh receptor that keeps the pathway repressed. Since AICD results from APP cleavage by γ-secretase, the goal of the current study was to establish whether treatment with a γ-secretase inhibitor normalizes AICD levels and restores neurogenesis in trisomic neural precursor cells. We found that treatment with a selective γ-secretase inhibitor (ELND006; ELN) restores proliferation in neurospheres derived from the subventricular zone (SVZ) of the Ts65Dn mouse model of DS. This effect was accompanied by reduction of AICD and Ptch1 levels and was prevented by inhibition of the Shh pathway with cyclopamine. Treatment of Ts65Dn mice with ELN in the postnatal period P3–P15 restored neurogenesis in the SVZ and hippocampus, hippocampal granule cell number and synapse development, indicating a positive impact of treatment on brain development. In addition, in the hippocampus of treated Ts65Dn mice there was a reduction in the expression levels of various genes that are transcriptionally regulated by AICD, including APP, its origin substrate. Inhibitors of γ-secretase are currently envisaged as tools for the cure of Alzheimer's disease because they lower βamyloid levels. Current results provide novel evidence that γ-secretase inhibitors may represent a strategy for the rescue of neurogenesis defects in DS. PMID:26254735

  2. The influence of metal-ion binding on the structure and surface composition of Sonic Hedgehog: a combined classical and hybrid QM/MM MD study.

    PubMed

    Hitzenberger, Manuel; Hofer, Thomas S

    2016-08-10

    In this work, the influence of the metal ions present in vertebrate Sonic Hedgehog was assessed by a series of molecular mechanics molecular dynamics simulations with differing ionic compositions. The obtained data suggest that Ca(ii) binding has a very distinct influence on the composition of the protein surface surrounding the binding site by shaping several ionic interactions with negatively charged sidechains that otherwise would be pointing towards the solvent, repelling potential ligands. Furthermore, the Ca(ii) ions play an important role in the stability of the loop regions where they are coordinated. In contrast, the removal of the Zn(ii) ion results in no noticeable destabilization of its chemical surrounding, however, it is shown that the destabilizing effect of removed Ca(ii) ions is amplified if Zn(ii) is absent as well. Furthermore, a quantum mechanical/molecular mechanical (QM/MM) molecular dynamics simulation of Sonic Hedgehog with special focus on the Zn(ii) binding site has been conducted. The results indicate that QM/MM in contrast to pure MM accurately reproduces structural features also found by experimental studies and therefore is able to provide credible predictions not only of the dynamical properties of the studied system but also of protein-ligand interactions at the metal ion binding site. PMID:27452578

  3. Set7 mediated Gli3 methylation plays a positive role in the activation of Sonic Hedgehog pathway in mammals

    PubMed Central

    Fu, Lin; Wu, Hailong; Cheng, Steven Y; Gao, Daming; Zhang, Lei; Zhao, Yun

    2016-01-01

    Hedgehog signaling plays very important roles in development and cancers. Vertebrates have three transcriptional factors, Gli1, Gli2 and Gli3. Among them, Gli3 is a very special transcriptional factor which closely resembles Cubitus interruptus (Ci, in Drosophila) structurally and functionally as a ‘double agent’ for Shh target gene expression. Here we show that Gli3 full-length, but not the truncated form, can be methylated at K436 and K595. This methylation is specifically catalyzed by Set7, a lysine methyltransferase (KMT). Methylation at K436 and K595 respectively increases the stability and DNA binding ability of Gli3, resulting in an enhancement of Shh signaling activation. Furthermore, functional experiments indicate that the Gli3 methylation contributes to the tumor growth and metastasis in non-small cell lung cancer in vitro and in vivo. Therefore, we propose that Set7 mediated methylation is a novel PTM of Gli3, which positively regulates the transactivity of Gli3 and the activation of Shh signaling. DOI: http://dx.doi.org/10.7554/eLife.15690.001 PMID:27146893

  4. Autonomous and nonautonomous roles of Hedgehog signaling in regulating limb muscle formation

    PubMed Central

    Hu, Jimmy Kuang-Hsien; McGlinn, Edwina; Harfe, Brian D.; Kardon, Gabrielle; Tabin, Clifford J.

    2012-01-01

    Muscle progenitor cells migrate from the lateral somites into the developing vertebrate limb, where they undergo patterning and differentiation in response to local signals. Sonic hedgehog (Shh) is a secreted molecule made in the posterior limb bud that affects patterning and development of multiple tissues, including skeletal muscles. However, the cell-autonomous and non-cell-autonomous functions of Shh during limb muscle formation have remained unclear. We found that Shh affects the pattern of limb musculature non-cell-autonomously, acting through adjacent nonmuscle mesenchyme. However, Shh plays a cell-autonomous role in maintaining cell survival in the dermomyotome and initiating early activation of the myogenic program in the ventral limb. At later stages, Shh promotes slow muscle differentiation cell-autonomously. In addition, Shh signaling is required cell-autonomously to regulate directional muscle cell migration in the distal limb. We identify neuroepithelial cell transforming gene 1 (Net1) as a downstream target and effector of Shh signaling in that context. PMID:22987639

  5. miR-219 regulates neural progenitors by dampening apical Par protein-dependent Hedgehog signaling.

    PubMed

    Hudish, Laura I; Galati, Domenico F; Ravanelli, Andrew M; Pearson, Chad G; Huang, Peng; Appel, Bruce

    2016-07-01

    The transition of dividing neuroepithelial progenitors to differentiated neurons and glia is essential for the formation of a functional nervous system. Sonic hedgehog (Shh) is a mitogen for spinal cord progenitors, but how cells become insensitive to the proliferative effects of Shh is not well understood. Because Shh reception occurs at primary cilia, which are positioned within the apical membrane of neuroepithelial progenitors, we hypothesized that loss of apical characteristics reduces the Shh signaling response, causing cell cycle exit and differentiation. We tested this hypothesis using genetic and pharmacological manipulation, gene expression analysis and time-lapse imaging of zebrafish embryos. Blocking the function of miR-219, a microRNA that downregulates apical Par polarity proteins and promotes progenitor differentiation, elevated Shh signaling. Inhibition of Shh signaling reversed the effects of miR-219 depletion and forced expression of Shh phenocopied miR-219 deficiency. Time-lapse imaging revealed that knockdown of miR-219 function accelerates the growth of primary cilia, revealing a possible mechanistic link between miR-219-mediated regulation of apical Par proteins and Shh signaling. Thus, miR-219 appears to decrease progenitor cell sensitivity to Shh signaling, thereby driving these cells towards differentiation. PMID:27226318

  6. Hedgehog signaling regulates liver sinusoidal endothelial cell capillarisation

    PubMed Central

    Xie, Guanhua; Choi, Steve S.; Syn, Wing-Kin; Michelotti, Gregory A.; Swiderska-Syn, Marzena; Karaca, Gamze; Chan, Isaac S.; Chen, Yuping; Diehl, Anna Mae

    2013-01-01

    Objective Vascular remodeling during liver damage involves loss of healthy liver sinusoidal endothelial cell (LSEC) phenotype via capillarisation. Hedgehog (Hh) signaling regulates vascular development and increases during liver injury. Therefore, we examined its role in capillarisation. Design Primary LSEC were cultured for 5 days to induce capillarisation. Pharmacologic, antibody-mediated, and genetic approaches were used to manipulate Hh signaling. Effects on mRNA and protein expression of Hh-regulated genes and capillarisation markers were evaluated by qRT-PCR and immunoblot. Changes in LSEC function were assessed by migration and tube forming assay, and gain/loss of fenestrae was examined by electron microscopy. Mice with acute or chronic liver injury were treated with Hh inhibitors; effects on capillarisation were assessed by immunohistochemistry. Results Freshly isolated LSEC expressed Hh ligands, Hh receptors, and Hh ligand antagonist Hhip. Capillarisation was accompanied by repression of Hhip and increased expression of Hh-regulated genes. Treatment with Hh agonist further induced expression of Hh ligands and Hh-regulated genes, and up-regulated capillarisation-associated genes; whereas Hh signaling antagonist or Hh ligand neutralizing antibody each repressed expression of Hh target genes and capillarisation markers. LSEC isolated from SmoloxP/loxP transgenic mice that had been infected with adenovirus expressing Cre-recombinase to delete Smoothened showed over 75% knockdown of Smoothened. During culture, Smoothened-deficient LSEC had inhibited Hh signaling, less induction of capillarisation-associated genes, and retention of fenestrae. In mice with injured livers, inhibiting Hh signaling prevented capillarisation. Conclusions LSEC produce and respond to Hh ligands, and use Hh signaling to regulate complex phenotypic changes that occur during capillarisation. PMID:22362915

  7. Embelin Suppresses Growth of Human Pancreatic Cancer Xenografts, and Pancreatic Cancer Cells Isolated from KrasG12D Mice by Inhibiting Akt and Sonic Hedgehog Pathways

    PubMed Central

    Huang, Minzhao; Tang, Su-Ni; Upadhyay, Ghanshyam; Marsh, Justin L.; Jackman, Christopher P.; Shankar, Sharmila; Srivastava, Rakesh K.

    2014-01-01

    Pancreatic cancer is a deadly disease, and therefore effective treatment and/or prevention strategies are urgently needed. The objectives of this study were to examine the molecular mechanisms by which embelin inhibited human pancreatic cancer cell growth in vitro, and xenografts in Balb C nude mice, and pancreatic cancer cell growth isolated from KrasG12D transgenic mice. XTT assays were performed to measure cell viability. AsPC-1 cells were injected subcutaneously into Balb c nude mice and treated with embelin. Cell proliferation and apoptosis were measured by Ki67 and TUNEL staining, respectively. The expression of Akt, and Sonic Hedgehog (Shh) and their target gene products were measured by the immunohistochemistry, and Western blot analysis. The effects of embelin on pancreatic cancer cells isolated from 10-months old KrasG12D mice were also examined. Embelin inhibited cell viability in pancreatic cancer AsPC-1, PANC-1, MIA PaCa-2 and Hs 766T cell lines, and these inhibitory effects were blocked either by constitutively active Akt or Shh protein. Embelin-treated mice showed significant inhibition in tumor growth which was associated with reduced expression of markers of cell proliferation (Ki67, PCNA and Bcl-2) and cell cycle (cyclin D1, CDK2, and CDK6), and induction of apoptosis (activation of caspase-3 and cleavage of PARP, and increased expression of Bax). In addition, embelin inhibited the expression of markers of angiogenesis (COX-2, VEGF, VEGFR, and IL-8), and metastasis (MMP-2 and MMP-9) in tumor tissues. Antitumor activity of embelin was associated with inhibition of Akt and Shh pathways in xenografts, and pancreatic cancer cells isolated from KrasG12D mice. Furthermore, embelin also inhibited epithelial-to-mesenchymal transition (EMT) by up-regulating E-cadherin and inhibiting the expression of Snail, Slug, and ZEB1. These data suggest that embelin can inhibit pancreatic cancer growth, angiogenesis and metastasis by suppressing Akt and Shh pathways

  8. An Nfic-hedgehog signaling cascade regulates tooth root development.

    PubMed

    Liu, Yang; Feng, Jifan; Li, Jingyuan; Zhao, Hu; Ho, Thach-Vu; Chai, Yang

    2015-10-01

    Coordination between the Hertwig's epithelial root sheath (HERS) and apical papilla (AP) is crucial for proper tooth root development. The hedgehog (Hh) signaling pathway and Nfic are both involved in tooth root development; however, their relationship has yet to be elucidated. Here, we establish a timecourse of mouse molar root development by histological staining of sections, and we demonstrate that Hh signaling is active before and during root development in the AP and HERS using Gli1 reporter mice. The proper pattern of Hh signaling activity in the AP is crucial for the proliferation of dental mesenchymal cells, because either inhibition with Hh inhibitors or constitutive activation of Hh signaling activity in transgenic mice leads to decreased proliferation in the AP and shorter roots. Moreover, Hh activity is elevated in Nfic(-/-) mice, a root defect model, whereas RNA sequencing and in situ hybridization show that the Hh attenuator Hhip is downregulated. ChIP and RNAscope analyses suggest that Nfic binds to the promoter region of Hhip. Treatment of Nfic(-/-) mice with Hh inhibitor partially restores cell proliferation, AP growth and root development. Taken together, our results demonstrate that an Nfic-Hhip-Hh signaling pathway is crucial for apical papilla growth and proper root formation. This discovery provides insight into the molecular mechanisms regulating tooth root development.

  9. An Nfic-hedgehog signaling cascade regulates tooth root development

    PubMed Central

    Liu, Yang; Feng, Jifan; Li, Jingyuan; Zhao, Hu; Ho, Thach-Vu; Chai, Yang

    2015-01-01

    Coordination between the Hertwig's epithelial root sheath (HERS) and apical papilla (AP) is crucial for proper tooth root development. The hedgehog (Hh) signaling pathway and Nfic are both involved in tooth root development; however, their relationship has yet to be elucidated. Here, we establish a timecourse of mouse molar root development by histological staining of sections, and we demonstrate that Hh signaling is active before and during root development in the AP and HERS using Gli1 reporter mice. The proper pattern of Hh signaling activity in the AP is crucial for the proliferation of dental mesenchymal cells, because either inhibition with Hh inhibitors or constitutive activation of Hh signaling activity in transgenic mice leads to decreased proliferation in the AP and shorter roots. Moreover, Hh activity is elevated in Nfic−/− mice, a root defect model, whereas RNA sequencing and in situ hybridization show that the Hh attenuator Hhip is downregulated. ChIP and RNAscope analyses suggest that Nfic binds to the promoter region of Hhip. Treatment of Nfic−/− mice with Hh inhibitor partially restores cell proliferation, AP growth and root development. Taken together, our results demonstrate that an Nfic-Hhip-Hh signaling pathway is crucial for apical papilla growth and proper root formation. This discovery provides insight into the molecular mechanisms regulating tooth root development. PMID:26293299

  10. Phosphoinositides Regulate Ciliary Protein Trafficking to Modulate Hedgehog Signaling

    PubMed Central

    Roberson, Elle C.; Garcia, Galo; Abedin, Monika; Schurmans, Stéphane; Inoue, Takanari; Reiter, Jeremy F.

    2015-01-01

    SUMMARY Primary cilia interpret vertebrate Hedgehog (Hh) signals. Why cilia are essential for signaling is unclear. One possibility is that some forms of signaling require a distinct membrane lipid composition, found at cilia. We found that the ciliary membrane contains a particular phosphoinositide, PI(4)P, whereas a different phosphoinositide, PI(4,5)P2, is restricted to the membrane of the ciliary base. This distribution is created by Inpp5e, a ciliary phosphoinositide 5-phosphatase. Without Inpp5e, ciliary PI(4,5)P2 levels are elevated and Hh signaling is disrupted. Inpp5e limits the ciliary levels of inhibitors of Hh signaling, including Gpr161 and the PI(4,5)P2-binding protein Tulp3. Increasing ciliary PI(4,5)P2 levels or conferring the ability to bind PI(4)P on Tulp3 increases the ciliary localization of Tulp3. Lowering Tulp3 in cells lacking Inpp5e reduces ciliary Gpr161 levels and restores Hh signaling. Therefore, Inpp5e regulates ciliary membrane phosphoinositide composition, and Tulp3 reads out ciliary phosphoinositides to control ciliary protein localization, enabling Hh signaling. PMID:26305592

  11. Hedgehog signaling regulates dental papilla formation and tooth size during zebrafish odontogenesis

    PubMed Central

    Yu, Jeffrey C.; Fox, Zachary D.B.; Crimp, James L.; Littleford, Hana E.; Jowdry, Andrea L.; Jackman, William R.

    2015-01-01

    Background Intercellular communication by the hedgehog cell signaling pathway is necessary for tooth development throughout the vertebrates, but it remains unclear which specific developmental signals control cell behavior at different stages of odontogenesis. To address this issue, we have manipulated hedgehog activity during zebrafish tooth development and visualized the results using confocal microscopy. Results We first established that reporter lines for dlx2b, fli1, NF-κB, and prdm1a are markers for specific subsets of tooth germ tissues. We then blocked hedgehog signaling with cyclopamine and observed a reduction or elimination of the cranial neural crest derived dental papilla, which normally contains the cells that later give rise to dentin-producing odontoblasts. Upon further investigation we observed that the dental papilla begins to form and then regresses in the absence of hedgehog signaling, through a mechanism unrelated to cell proliferation or apoptosis. We also found evidence of an isometric reduction in tooth size that correlates with the time of earliest hedgehog inhibition. Conclusions We hypothesize that these results reveal a previously uncharacterized function of hedgehog signaling during tooth morphogenesis, regulating the number of cells in the dental papilla and thereby controlling tooth size. PMID:25645398

  12. Hedgehog-dependent E3-ligase Midline1 regulates ubiquitin-mediated proteasomal degradation of Pax6 during visual system development.

    PubMed

    Pfirrmann, Thorsten; Jandt, Enrico; Ranft, Swantje; Lokapally, Ashwin; Neuhaus, Herbert; Perron, Muriel; Hollemann, Thomas

    2016-09-01

    Pax6 is a key transcription factor involved in eye, brain, and pancreas development. Although pax6 is expressed in the whole prospective retinal field, subsequently its expression becomes restricted to the optic cup by reciprocal transcriptional repression of pax6 and pax2 However, it remains unclear how Pax6 protein is removed from the eyestalk territory on time. Here, we report that Mid1, a member of the RBCC/TRIM E3 ligase family, which was first identified in patients with the X-chromosome-linked Opitz BBB/G (OS) syndrome, interacts with Pax6. We found that the forming eyestalk is a major domain of mid1 expression, controlled by the morphogen Sonic hedgehog (Shh). Here, Mid1 regulates the ubiquitination and proteasomal degradation of Pax6 protein. Accordantly, when Mid1 levels are knocked down, Pax6 expression is expanded and eyes are enlarged. Our findings indicate that remaining or misaddressed Pax6 protein is cleared from the eyestalk region to properly set the border between the eyestalk territory and the retina via Mid1. Thus, we identified a posttranslational mechanism, regulated by Sonic hedgehog, which is important to suppress Pax6 activity and thus breaks pax6 autoregulation at defined steps during the formation of the visual system. PMID:27555585

  13. Hedgehog Signaling Regulates Telomerase Reverse Transcriptase in Human Cancer Cells

    PubMed Central

    Mazumdar, Tapati; Sandhu, Ranjodh; Qadan, Maha; DeVecchio, Jennifer; Magloire, Victoria; Agyeman, Akwasi; Li, Bibo; Houghton, Janet A.

    2013-01-01

    The Hedgehog (HH) signaling pathway is critical for normal embryonic development, tissue patterning and cell differentiation. Aberrant HH signaling is involved in multiple human cancers. HH signaling involves a multi-protein cascade activating the GLI proteins that transcriptionally regulate HH target genes. We have previously reported that HH signaling is essential for human colon cancer cell survival and inhibition of this signal induces DNA damage and extensive cell death. Here we report that the HH/GLI axis regulates human telomerase reverse transcriptase (hTERT), which determines the replication potential of cancer cells. Suppression of GLI1/GLI2 functions by a C-terminus truncated GLI3 repressor mutant (GLI3R), or by GANT61, a pharmacological inhibitor of GLI1/GLI2, reduced hTERT protein expression in human colon cancer, prostate cancer and Glioblastoma multiforme (GBM) cell lines. Expression of an N-terminus deleted constitutively active mutant of GLI2 (GLI2ΔN) increased hTERT mRNA and protein expression and hTERT promoter driven luciferase activity in human colon cancer cells while GANT61 inhibited hTERT mRNA expression and hTERT promoter driven luciferase activity. Chromatin immunoprecipitation with GLI1 or GLI2 antibodies precipitated fragments of the hTERT promoter in human colon cancer cells, which was reduced upon exposure to GANT61. In contrast, expression of GLI1 or GLI2ΔN in non-malignant 293T cells failed to alter the levels of hTERT mRNA and protein, or hTERT promoter driven luciferase activity. Further, expression of GLI2ΔN increased the telomerase enzyme activity, which was reduced by GANT61 administration in human colon cancer, prostate cancer, and GBM cells. These results identify hTERT as a direct target of the HH signaling pathway, and reveal a previously unknown role of the HH/GLI axis in regulating the replication potential of cancer cells. These findings are of significance in understanding the important regulatory mechanisms that

  14. Activation of Smurf E3 Ligase Promoted by Smoothened Regulates Hedgehog Signaling through Targeting Patched Turnover

    PubMed Central

    Zheng, Xiudeng; Chen, Zhenping; Sun, Liwei; Wang, Hailong; Zhu, Yuanxiang; Zhang, Jing; Yang, Shuyan; Lu, Yi; Sun, Qinmiao; Tao, Yi; Liu, Feng; Zhao, Yun; Chen, Dahua

    2013-01-01

    Hedgehog signaling plays conserved roles in controlling embryonic development; its dysregulation has been implicated in many human diseases including cancers. Hedgehog signaling has an unusual reception system consisting of two transmembrane proteins, Patched receptor and Smoothened signal transducer. Although activation of Smoothened and its downstream signal transduction have been intensively studied, less is known about how Patched receptor is regulated, and particularly how this regulation contributes to appropriate Hedgehog signal transduction. Here we identified a novel role of Smurf E3 ligase in regulating Hedgehog signaling by controlling Patched ubiquitination and turnover. Moreover, we showed that Smurf-mediated Patched ubiquitination depends on Smo activity in wing discs. Mechanistically, we found that Smo interacts with Smurf and promotes it to mediate Patched ubiquitination by targeting the K1261 site in Ptc. The further mathematic modeling analysis reveals that a bidirectional control of activation of Smo involving Smurf and Patched is important for signal-receiving cells to precisely interpret external signals, thereby maintaining Hedgehog signaling reliability. Finally, our data revealed an evolutionarily conserved role of Smurf proteins in controlling Hh signaling by targeting Ptc during development. PMID:24302888

  15. Identification of Small Molecules Which Induce Skeletal Muscle Differentiation in Embryonic Stem Cells via Activation of the Wnt and Inhibition of Smad2/3 and Sonic Hedgehog Pathways.

    PubMed

    Lee, Hyunwoo; Haller, Corinne; Manneville, Carole; Doll, Thierry; Fruh, Isabelle; Keller, Caroline Gubser; Richards, Shola M; Ibig-Rehm, Yvonne; Patoor, Maude; Goette, Marjo; Bouchez, Laure C; Mueller, Matthias

    2016-02-01

    The multilineage differentiation capacity of mouse and human embryonic stem (ES) cells offers a testing platform for small molecules that mediate mammalian lineage determination and cellular specialization. Here we report the identification of two small molecules which drives mouse 129 ES cell differentiation to skeletal muscle with high efficiency without any genetic modification. Mouse embryoid bodies (EBs) were used to screen a library of 1,000 small molecules to identify compounds capable of inducing high levels of Pax3 mRNA. Stimulation of EBs with SMIs (skeletal muscle inducer, SMI1 and SMI2) from the screen resulted in a high percentage of intensively twitching skeletal muscle fibers 3 weeks after induction. Gene expression profiling studies that were carried out for mode of actions analysis showed that SMIs activated genes regulated by the Wnt pathway and inhibited expression of Smad2/3 and Sonic Hedgehog (Shh) target genes. A combination of three small molecules known to modulate these three pathways acted similarly to the SMIs found here, driving ES cells from 129 as well as Balb/c and C57Bl/6 to skeletal muscle. Taken together, these data demonstrate that the SMI drives ES cells to skeletal muscle via concerted activation of the Wnt pathway, and inhibition of Smad2/3 signaling and Shh pathways. This provides important developmental biological information about skeletal muscle differentiation from embryonic stem cells and may lead to the development of new therapeutics for muscle disease.

  16. Identification of Small Molecules Which Induce Skeletal Muscle Differentiation in Embryonic Stem Cells via Activation of the Wnt and Inhibition of Smad2/3 and Sonic Hedgehog Pathways.

    PubMed

    Lee, Hyunwoo; Haller, Corinne; Manneville, Carole; Doll, Thierry; Fruh, Isabelle; Keller, Caroline Gubser; Richards, Shola M; Ibig-Rehm, Yvonne; Patoor, Maude; Goette, Marjo; Bouchez, Laure C; Mueller, Matthias

    2016-02-01

    The multilineage differentiation capacity of mouse and human embryonic stem (ES) cells offers a testing platform for small molecules that mediate mammalian lineage determination and cellular specialization. Here we report the identification of two small molecules which drives mouse 129 ES cell differentiation to skeletal muscle with high efficiency without any genetic modification. Mouse embryoid bodies (EBs) were used to screen a library of 1,000 small molecules to identify compounds capable of inducing high levels of Pax3 mRNA. Stimulation of EBs with SMIs (skeletal muscle inducer, SMI1 and SMI2) from the screen resulted in a high percentage of intensively twitching skeletal muscle fibers 3 weeks after induction. Gene expression profiling studies that were carried out for mode of actions analysis showed that SMIs activated genes regulated by the Wnt pathway and inhibited expression of Smad2/3 and Sonic Hedgehog (Shh) target genes. A combination of three small molecules known to modulate these three pathways acted similarly to the SMIs found here, driving ES cells from 129 as well as Balb/c and C57Bl/6 to skeletal muscle. Taken together, these data demonstrate that the SMI drives ES cells to skeletal muscle via concerted activation of the Wnt pathway, and inhibition of Smad2/3 signaling and Shh pathways. This provides important developmental biological information about skeletal muscle differentiation from embryonic stem cells and may lead to the development of new therapeutics for muscle disease. PMID:26577380

  17. Testing chemotherapeutic agents in the feather follicle identifies a selective blockade of cell proliferation and a key role for sonic hedgehog signaling in chemotherapy-induced tissue damage.

    PubMed

    Xie, Guojiang; Wang, Hangwei; Yan, Zhipeng; Cai, Linyan; Zhou, Guixuan; He, Wanzhong; Paus, Ralf; Yue, Zhicao

    2015-03-01

    Chemotherapeutic agents induce complex tissue responses in vivo and damage normal organ functions. Here we use the feather follicle to investigate details of this damage response. We show that cyclophosphamide treatment, which causes chemotherapy-induced alopecia in mice and man, induces distinct defects in feather formation: feather branching is transiently and reversibly disrupted, thus leaving a morphological record of the impact of chemotherapeutic agents, whereas the rachis (feather axis) remains unperturbed. Similar defects are observed in feathers treated with 5-fluorouracil or taxol but not with doxorubicin or arabinofuranosyl cytidine (Ara-C). Selective blockade of cell proliferation was seen in the feather branching area, along with a downregulation of sonic hedgehog (Shh) transcription, but not in the equally proliferative rachis. Local delivery of the Shh inhibitor, cyclopamine, or Shh silencing both recapitulated this effect. In mouse hair follicles, those chemotherapeutic agents that disrupted feather formation also downregulated Shh gene expression and induced hair loss, whereas doxorubicin or Ara-C did not. Our results reveal a mechanism through which chemotherapeutic agents damage rapidly proliferating epithelial tissue, namely via the cell population-specific, Shh-dependent inhibition of proliferation. This mechanism may be targeted by future strategies to manage chemotherapy-induced tissue damage.

  18. A variant in the sonic hedgehog regulatory sequence (ZRS) is associated with triphalangeal thumb and deregulates expression in the developing limb

    PubMed Central

    Furniss, Dominic; Lettice, Laura A.; Taylor, Indira B.; Critchley, Paul S.; Giele, Henk; Hill, Robert E.; Wilkie, Andrew O.M.

    2008-01-01

    A locus for triphalangeal thumb, variably associated with pre-axial polydactyly, was previously identified in the zone of polarizing activity regulatory sequence (ZRS), a long range limb-specific enhancer of the Sonic Hedgehog (SHH) gene at human chromosome 7q36.3. Here, we demonstrate that a 295T>C variant in the human ZRS, previously thought to represent a neutral polymorphism, acts as a dominant allele with reduced penetrance. We found this variant in three independently ascertained probands from southern England with triphalangeal thumb, demonstrated significant linkage of the phenotype to the variant (LOD = 4.1), and identified a shared microsatellite haplotype around the ZRS, suggesting that the probands share a common ancestor. An individual homozygous for the 295C allele presented with isolated bilateral triphalangeal thumb resembling the heterozygous phenotype, suggesting that the variant is largely dominant to the wild-type allele. As a functional test of the pathogenicity of the 295C allele, we utilized a mutated ZRS construct to demonstrate that it can drive ectopic anterior expression of a reporter gene in the developing mouse forelimb. We conclude that the 295T>C variant is in fact pathogenic and, in southern England, appears to be the most common cause of triphalangeal thumb. Depending on the dispersal of the founding mutation, it may play a wider role in the aetiology of this disorder. PMID:18463159

  19. BRD4 induces cell migration and invasion in HCC cells through MMP-2 and MMP-9 activation mediated by the Sonic hedgehog signaling pathway

    PubMed Central

    WANG, YONG-HUI; SUI, XIAO-MEI; SUI, YA-NA; ZHU, QIN-WEI; YAN, KAI; WANG, LI-SHAN; WANG, FEI; ZHOU, JIA-HUA

    2015-01-01

    Hepatocellular carcinoma (HCC) is a highly aggressive form of carcinoma with poor prognosis, and HCC-associated mortality primarily occurs due to migration and invasion of HCC cells. The manipulation of epigenetic proteins, such as BRD4, has recently emerged as an alternative therapeutic strategy. The present study aimed to investigate the novel mechanism of BRD4 involvement in the migration and invasion of HCC cells. Reverse transcription-quantitative polymerase chain reaction was used to assess BRD4 mRNA expression levels in HCC cell lines. This analysis demonstrated that BRD4 was significantly overexpressed in HCC cell lines compared with a human immortalized normal liver cell line. A short hairpin RNA (shRNA) was then used to suppress BRD4 expression in HCC cells, and resulted in impaired HCC cell proliferation, migration and invasion. When the HepG2 HCC cell line was treated with recombinant human sonic hedgehog (SHH) peptide, the migration and invasion capabilities of HepG2 cells that were inhibited by BRD4 silencing were restored. BRD4 induced cell migration and invasion in HepG2 cells through the activation of matrix metalloproteinase (MMP)-2 and MMP-9, mediated by the SHH signaling pathway. Taken together, the results of the present study demonstrated the importance of BRD4 in HCC cell proliferation and metastasis. Thus, BRD4 is a potential novel target for the development of therapeutic approaches against HCC. PMID:26622824

  20. Network modeling of TGFβ signaling in hepatocellular carcinoma epithelial-to-mesenchymal transition reveals joint Sonic hedgehog and Wnt pathway activation

    PubMed Central

    Steinway, Steven Nathaniel; Zañudo, Jorge Gomez Tejeda; Ding, Wei; Rountree, Carl Bart; Feith, David J.; Loughran, Thomas P.; Albert, Reka

    2014-01-01

    Epithelial-to-mesenchymal transition (EMT) is a developmental process hijacked by cancer cells to leave the primary tumor site, invade surrounding tissue, and establish distant metastases. A hallmark of EMT is the loss of E-cadherin expression, and one major signal for the induction of EMT is transforming growth factor beta (TGFβ, which is dysregulated in up to 40% of hepatocellular carcinoma (HCC). We have constructed an EMT network of 70 nodes and 135 edges by integrating the signaling pathways involved in developmental EMT and known dysregulations in invasive HCC. We then used discrete dynamic modeling to understand the dynamics of the EMT network driven by TGFβ. Our network model recapitulates known dysregulations during the induction of EMT and predicts the activation of the Wnt and Sonic hedgehog (SHH) signaling pathways during this process. We show, across multiple murine (P2E and P2M) and human HCC cell lines (Huh7, PLC/PRF/5, HLE, and HLF), that the TGFβ signaling axis is a conserved driver of mesenchymal phenotype HCC and confirm that Wnt and SHH signaling are induced in these cell lines. Furthermore, we identify by network analysis eight regulatory feedback motifs that stabilize the EMT process and show that these motifs involve cross-talk among multiple major pathways. Our model will be useful in identifying potential therapeutic targets for the suppression of EMT, invasion and metastasis in HCC. PMID:25189528

  1. Activation of Sonic Hedgehog Leads to Survival Enhancement of Astrocytes via the GRP78-Dependent Pathway in Mice Infected with Angiostrongylus cantonensis

    PubMed Central

    Chen, Kuang-Yao; Cheng, Chien-Ju; Wang, Lian-Chen

    2015-01-01

    Angiostrongylus cantonensis infection may cause elevation of ROS and antioxidants in the CSF of infected mice. Astrocytes may protect the surrounding neurons from oxidative stress-induced cell death by secreting Sonic hedgehog (Shh) via the PI3-K/AKT/Bcl-2 pathway. This study was conducted to determine the role of the Shh signaling pathway in A. cantonensis-infected BABL/c mice by coculturing astrocytes with living fifth-stage larvae or soluble antigens. The Shh pathway was activated with corresponding increases in the level of the Shh. Glial fibrillary acidic protein (GFAP) and Shh were increased in astrocyte cocultured with living fifth-stage larvae or soluble antigens. The survival of astrocytes pretreated with Shh was significantly elevated in cocultures with the antigens but reduced by its inhibitor cyclopamine. The expression of GRP78 and Bcl-2 was significantly higher in astrocytes pretreated with recombinant Shh. These findings suggest that the expression of Shh may inhibit cell death by activating Bcl-2 through a GRP78-dependent pathway. PMID:25961032

  2. Hedgehog Signaling Regulates the Survival of Gastric Cancer Cells by Regulating the Expression of Bcl-2

    PubMed Central

    Han, Myoung-Eun; Lee, Young-Suk; Baek, Sun-Yong; Kim, Bong-Seon; Kim, Jae-Bong; Oh, Sae-Ock

    2009-01-01

    Gastric cancer is the second most common cause of cancer deaths worldwide. The underlying molecular mechanisms of its carcinogenesis are relatively poorly characterized. Hedgehog (Hh) signaling, which is critical for development of various organs including the gastrointestinal tract, has been associated with gastric cancer. The present study was undertaken to reveal the underlying mechanism by which Hh signaling controls gastric cancer cell proliferation. Treatment of gastric cancer cells with cyclopamine, a specific inhibitor of Hh signaling pathway, reduced proliferation and induced apoptosis of gastric cancer cells. Cyclopamine treatment induced cytochrome c release from mitochondria and cleavage of caspase 9. Moreover, Bcl-2 expression was significantly reduced by cyclopamine treatment. These results suggest that Hh signaling regulates the survival of gastric cancer cells by regulating the expression of Bcl-2. PMID:19742123

  3. PI3K/AKT/mTOR and sonic hedgehog pathways cooperate together to inhibit human pancreatic cancer stem cell characteristics and tumor growth

    PubMed Central

    Sharma, Narinder; Nanta, Rajesh; Sharma, Jay; Gunewardena, Sumedha; Singh, Karan P.; Shankar, Sharmila; Srivastava, Rakesh K.

    2015-01-01

    Cancer stem cells (CSCs) play major roles in cancer initiation, progression, and metastasis. It is evident from growing reports that PI3K/Akt/mTOR and Sonic Hedgehog (Shh) signaling pathways are aberrantly reactivated in pancreatic CSCs. Here, we examined the efficacy of combining NVP-LDE-225 (PI3K/mTOR inhibitor) and NVP-BEZ-235 (Smoothened inhibitor) on pancreatic CSCs characteristics, microRNA regulatory network, and tumor growth. NVP-LDE-225 co-operated with NVP-BEZ-235 in inhibiting pancreatic CSC's characteristics and tumor growth in mice by acting at the level of Gli. Combination of NVP-LDE-225 and NVP-BEZ-235 inhibited self-renewal capacity of CSCs by suppressing the expression of pluripotency maintaining factors Nanog, Oct-4, Sox-2 and c-Myc, and transcription of Gli. NVP-LDE-225 co-operated with NVP-BEZ-235 to inhibit Lin28/Let7a/Kras axis in pancreatic CSCs. Furthermore, a superior interaction of these drugs was observed on spheroid formation by pancreatic CSCs isolated from Pankras/p53 mice. The combination of these drugs also showed superior effects on the expression of proteins involved in cell proliferation, survival and apoptosis. In addition, NVP-LDE-225 co-operated with NVP-BEZ-235 in inhibiting EMT through modulation of cadherin, vimentin and transcription factors Snail, Slug and Zeb1. In conclusion, these data suggest that the combined inhibition of PI3K/Akt/mTOR and Shh pathways may be beneficial for the treatment of pancreatic cancer. PMID:26451606

  4. The differentiation of amniotic fluid stem cells into sweat glandlike cells is enhanced by the presence of Sonic hedgehog in the conditioned medium.

    PubMed

    Liang, Hansi; Sun, Qing; Zhen, Yunfang; Li, Fang; Xu, YunYun; Liu, Yao; Zhang, Xueguang; Qin, Mingde

    2016-09-01

    After patients suffer severe full-thickness burn injuries, the current treatments cannot lead to the complete self-regeneration of the sweat gland structure and function. Therefore, it is important to identify new methods for acquiring sufficient functional sweat gland cells to restore skin function. In this study, we induced CD117+ human amniotic fluid stem (hAFS) cells to differentiate into sweat glandlike (hAFS-SG) cells based on the use of conditioned medium (CM) from the human sweat gland (hSG) cells. Real-time PCR and immunofluorescent staining were used to confirm the expression of the sweat gland-related genes Ectodysplasin-A (EDA), Ectodysplasin-A receptor (EDAR), keratin 8 (K8) and carcino-embryonic antigen (CEA). Transmission electron microscopy analysis shows that microvilli, the cellular structures that are typical for hSG cells, can also be observed on the membrane of the hAFS-SG cells. Our test for the calcium response to acetylcholine (Ach) proved that hAFS-SG cells have the potential to respond to Ach in a manner similar to normal sweat glands. A three-dimensional culture is an effective approach that stimulates the hAFS-SG cells to form tubular structures and drives hAFS-SG cells to mature into higher stage. We also found that epidermal growth factor enhances the efficiency of differentiation and that Sonic hedgehog is an important factor of the CM that influences sweat gland differentiation. Our study provides the basis for further investigations into novel methods of inducing stem cells to differentiate into sweat glandlike cells. PMID:27120089

  5. PI3K/AKT/mTOR and sonic hedgehog pathways cooperate together to inhibit human pancreatic cancer stem cell characteristics and tumor growth.

    PubMed

    Sharma, Narinder; Nanta, Rajesh; Sharma, Jay; Gunewardena, Sumedha; Singh, Karan P; Shankar, Sharmila; Srivastava, Rakesh K

    2015-10-13

    Cancer stem cells (CSCs) play major roles in cancer initiation, progression, and metastasis. It is evident from growing reports that PI3K/Akt/mTOR and Sonic Hedgehog (Shh) signaling pathways are aberrantly reactivated in pancreatic CSCs. Here, we examined the efficacy of combining NVP-LDE-225 (PI3K/mTOR inhibitor) and NVP-BEZ-235 (Smoothened inhibitor) on pancreatic CSCs characteristics, microRNA regulatory network, and tumor growth. NVP-LDE-225 co-operated with NVP-BEZ-235 in inhibiting pancreatic CSC's characteristics and tumor growth in mice by acting at the level of Gli. Combination of NVP-LDE-225 and NVP-BEZ-235 inhibited self-renewal capacity of CSCs by suppressing the expression of pluripotency maintaining factors Nanog, Oct-4, Sox-2 and c-Myc, and transcription of Gli. NVP-LDE-225 co-operated with NVP-BEZ-235 to inhibit Lin28/Let7a/Kras axis in pancreatic CSCs. Furthermore, a superior interaction of these drugs was observed on spheroid formation by pancreatic CSCs isolated from Pankras/p53 mice. The combination of these drugs also showed superior effects on the expression of proteins involved in cell proliferation, survival and apoptosis. In addition, NVP-LDE-225 co-operated with NVP-BEZ-235 in inhibiting EMT through modulation of cadherin, vimentin and transcription factors Snail, Slug and Zeb1. In conclusion, these data suggest that the combined inhibition of PI3K/Akt/mTOR and Shh pathways may be beneficial for the treatment of pancreatic cancer. PMID:26451606

  6. Comparison of expression patterns of fibroblast growth factor 8, bone morphogenetic protein 4 and sonic hedgehog in jaw development of the house shrew, Suncus murinus.

    PubMed

    Ogi, Hidenao; Tabata, Makoto J; Yamanaka, Atsushi; Yasui, Kinya; Uemura, Masanori

    2002-01-01

    To elucidate the mechanism underlying jaw development in mammals, we used a new laboratory animal, Suncus murinus (house shrew, an insectivore) as the subject for the investigation, because Suncus has all types of teeth (incisor, canine, premolar and molar) in its upper and lower jaws and is thought to be a good model animal having a general mammalian tooth pattern. At the start, by use of degenerate primers we cloned Suncus homologues of fibroblast growth factor 8 (sFgf8), bone morphogenetic protein 4 (sBmp4) and sonic hedgehog (sShh) genes from cDNA library derived from whole Suncus embryos at day 12 (E12). Thereafter, we examined the expression patterns of these genes in the jaw development of Suncus E11-16 embryos (for mouse E9.5-12 embryos). sFgf8 and sBmp4 were expressed in E11 but not in E15 and onward during orofacial development. sShh was expressed from E11 onward, and its expression was increased in the orofacial area. The expression pattern of sFgf8 in the maxillary and mandibular arches of E14 coincided with the area of the presumptive tooth arch. However, sShh and sBmp4 were expressed only in the outer area (= buccal/labial side) of presumptive tooth arch. Thus, these 3 genes showed specific expression pattern in jaw development of Suncus, and their distributions did not overlap each other except in a few regions. These findings suggest that sFgf8, sBmp4 and sShh have a specific function respectively during jaw development in Suncus murinus.

  7. Seminiferous cord formation is regulated by hedgehog signaling in the marsupial.

    PubMed

    Chung, Jin Wei; Pask, Andrew J; Renfree, Marilyn B

    2012-03-01

    The signaling molecule DHH, secreted by Sertoli cells, has essential regulatory functions in testicular differentiation. DHH is required for the differentiation of peritubular myoid cells that line the seminiferous cords and steroidogenic Leydig cells. The testicular cords in Dhh-null male mice lack a basal lamina and develop abnormally. To date, the DHH-signaling pathway has never been examined outside of any eutherian mammals. This study examined the effects of inhibition of DHH signaling in a marsupial mammal, the tammar wallaby, by culturing gonads in vitro in the presence of the hedgehog-signaling inhibitors cyclopamine and forskolin. Disruption of hedgehog signaling in the tammar testes caused highly disorganized cord formation. SOX9 protein remained strongly expressed in Sertoli cells, laminin distribution was highly fragmented, and germ cells were distributed around the cortical regions of treated testes in an ovarianlike morphology. This suggests that hedgehog signaling regulates cord formation in the tammar wallaby testis as it does in eutherian mammals. These data demonstrate that the hedgehog pathway has been highly conserved in mammals for at least 160 million years. PMID:22133695

  8. Hedgehog signaling regulates FOXA2 in esophageal embryogenesis and Barrett’s metaplasia

    PubMed Central

    Wang, David H.; Tiwari, Anjana; Kim, Monica E.; Clemons, Nicholas J.; Regmi, Nanda L.; Hodges, William A.; Berman, David M.; Montgomery, Elizabeth A.; Watkins, D. Neil; Zhang, Xi; Zhang, Qiuyang; Jie, Chunfa; Spechler, Stuart J.; Souza, Rhonda F.

    2014-01-01

    Metaplasia can result when injury reactivates latent developmental signaling pathways that determine cell phenotype. Barrett’s esophagus is a squamous-to-columnar epithelial metaplasia caused by reflux esophagitis. Hedgehog (Hh) signaling is active in columnar-lined, embryonic esophagus and inactive in squamous-lined, adult esophagus. We showed previously that Hh signaling is reactivated in Barrett’s metaplasia and overexpression of Sonic hedgehog (SHH) in mouse esophageal squamous epithelium leads to a columnar phenotype. Here, our objective was to identify Hh target genes involved in Barrett’s pathogenesis. By microarray analysis, we found that the transcription factor Foxa2 is more highly expressed in murine embryonic esophagus compared with postnatal esophagus. Conditional activation of Shh in mouse esophageal epithelium induced FOXA2, while FOXA2 expression was reduced in Shh knockout embryos, establishing Foxa2 as an esophageal Hh target gene. Evaluation of patient samples revealed FOXA2 expression in Barrett’s metaplasia, dysplasia, and adenocarcinoma but not in esophageal squamous epithelium or squamous cell carcinoma. In esophageal squamous cell lines, Hh signaling upregulated FOXA2, which induced expression of MUC2, an intestinal mucin found in Barrett’s esophagus, and the MUC2-processing protein AGR2. Together, these data indicate that Hh signaling induces expression of genes that determine an intestinal phenotype in esophageal squamous epithelial cells and may contribute to the development of Barrett’s metaplasia. PMID:25083987

  9. Centrosomal Protein DZIP1 Regulates Hedgehog Signaling by Promoting Cytoplasmic Retention of Transcription Factor GLI3 and Affecting Ciliogenesis*

    PubMed Central

    Wang, Chengbing; Low, Wee-Chuang; Liu, Aimin; Wang, Baolin

    2013-01-01

    The primary cilium is required for Hedgehog signaling. So far, all known ciliogenic proteins regulate Hedgehog signaling through their role in ciliogenesis. Here we show that the mouse DZIP1 regulates Hedgehog signaling through two mechanisms. First, DZIP1 interacts with GLI3, a transcriptional regulator for Hedgehog signaling, and prevents GLI3 from entering the nucleus. Second, DZIP1 is required for ciliogenesis. We show that DZIP1 colocalizes and interacts with CEP164, a protein localizing at appendages of the mother centrioles, and IFT88, a component of the intraflagellar transport (IFT) machinery. Functionally, both CEP164 and Ninein appendage proteins fail to localize to ciliary appendages in Dzip1 mutant cells; IFT components are not recruited to the basal body of cilia. Importantly, the accumulation of GLI3 in the nucleus is independent of loss of primary cilia in Dzip1 mutant cells. Therefore, DZIP1 is the first known ciliogenic protein that regulates Hedgehog signaling through a dual mechanism and that biochemically links IFT machinery with Hedgehog pathway components. PMID:23955340

  10. Structural basis of SUFU–GLI interaction in human Hedgehog signalling regulation

    PubMed Central

    Cherry, Amy L.; Finta, Csaba; Karlström, Mikael; Jin, Qianren; Schwend, Thomas; Astorga-Wells, Juan; Zubarev, Roman A.; Del Campo, Mark; Criswell, Angela R.; de Sanctis, Daniele; Jovine, Luca; Toftgård, Rune

    2013-01-01

    Hedgehog signalling plays a fundamental role in the control of metazoan development, cell proliferation and differentiation, as highlighted by the fact that its deregulation is associated with the development of many human tumours. SUFU is an essential intracellular negative regulator of mammalian Hedgehog signalling and acts by binding and modulating the activity of GLI transcription factors. Despite its central importance, little is known about SUFU regulation and the nature of SUFU–GLI interaction. Here, the crystal and small-angle X-ray scattering structures of full-length human SUFU and its complex with the key SYGHL motif conserved in all GLIs are reported. It is demonstrated that GLI binding is associated with major conformational changes in SUFU, including an intrinsically disordered loop that is also crucial for pathway activation. These findings reveal the structure of the SUFU–GLI interface and suggest a mechanism for an essential regulatory step in Hedgehog signalling, offering possibilities for the development of novel pathway modulators and therapeutics. PMID:24311597

  11. Structural basis of SUFU–GLI interaction in human Hedgehog signalling regulation

    SciTech Connect

    Cherry, Amy L.; Finta, Csaba; Karlström, Mikael; Jin, Qianren; Schwend, Thomas; Astorga-Wells, Juan; Zubarev, Roman A.; Del Campo, Mark; Criswell, Angela R.; Sanctis, Daniele de; Jovine, Luca Toftgård, Rune

    2013-12-01

    Crystal and small-angle X-ray scattering structures of full-length human SUFU alone and in complex with the conserved SYGHL motif from GLI transcription factors show major conformational changes associated with binding and reveal an intrinsically disordered region crucial for pathway activation. Hedgehog signalling plays a fundamental role in the control of metazoan development, cell proliferation and differentiation, as highlighted by the fact that its deregulation is associated with the development of many human tumours. SUFU is an essential intracellular negative regulator of mammalian Hedgehog signalling and acts by binding and modulating the activity of GLI transcription factors. Despite its central importance, little is known about SUFU regulation and the nature of SUFU–GLI interaction. Here, the crystal and small-angle X-ray scattering structures of full-length human SUFU and its complex with the key SYGHL motif conserved in all GLIs are reported. It is demonstrated that GLI binding is associated with major conformational changes in SUFU, including an intrinsically disordered loop that is also crucial for pathway activation. These findings reveal the structure of the SUFU–GLI interface and suggest a mechanism for an essential regulatory step in Hedgehog signalling, offering possibilities for the development of novel pathway modulators and therapeutics.

  12. MicroRNA Biogenesis and Hedgehog-Patched Signaling Cooperate to Regulate an Important Developmental Transition in Granule Cell Development.

    PubMed

    Constantin, Lena; Constantin, Myrna; Wainwright, Brandon J

    2016-03-01

    The Dicer1, Dcr-1 homolog (Drosophila) gene encodes a type III ribonuclease required for the canonical maturation and functioning of microRNAs (miRNAs). Subsets of miRNAs are known to regulate normal cerebellar granule cell development, in addition to the growth and progression of medulloblastoma, a neoplasm that often originates from granule cell precursors. Multiple independent studies have also demonstrated that deregulation of Sonic Hedgehog (Shh)-Patched (Ptch) signaling, through miRNAs, is causative of granule cell pathologies. In the present study, we investigated the genetic interplay between miRNA biogenesis and Shh-Ptch signaling in granule cells of the cerebellum by way of the Cre/lox recombination system in genetically engineered models of Mus musculus (mouse). We demonstrate that, although the miRNA biogenesis and Shh-Ptch-signaling pathways, respectively, regulate the opposing growth processes of cerebellar hypoplasia and hyperplasia leading to medulloblastoma, their concurrent deregulation was nonadditive and did not bring the growth phenotypes toward an expected equilibrium. Instead, mice developed either hypoplasia or medulloblastoma, but of a greater severity. Furthermore, some genotypes were bistable, whereby subsets of mice developed hypoplasia or medulloblastoma. This implies that miRNAs and Shh-Ptch signaling regulate an important developmental transition in granule cells of the cerebellum. We also conclusively show that the Dicer1 gene encodes a haploinsufficient tumor suppressor gene for Ptch1-induced medulloblastoma, with the monoallielic loss of Dicer1 more severe than biallelic loss. These findings exemplify how genetic interplay between pathways may produce nonadditive effects with a substantial and unpredictable impact on biology. Furthermore, these findings suggest that the functional dosage of Dicer1 may nonadditively influence a wide range of Shh-Ptch-dependent pathologies. PMID:26773048

  13. Sorting Sloppy Sonic

    PubMed Central

    Little, Shawn C.; Gregor, Thomas

    2014-01-01

    In the classic picture of morphogen-mediated patterning, cells acquire the correct spatial arrangement of specified fates by reading a precisely distributed gradient of morphogen. Xiong et al. now provide evidence for an alternate strategy—cells of the zebrafish neural tube actively sort to their correct positions following disordered specification by Sonic hedgehog. PMID:23622235

  14. Hedgehog Signaling Regulates the Ciliary Transport of Odorant Receptors in Drosophila.

    PubMed

    Sanchez, Gonzalo M; Alkhori, Liza; Hatano, Eduardo; Schultz, Sebastian W; Kuzhandaivel, Anujaianthi; Jafari, Shadi; Granseth, Björn; Alenius, Mattias

    2016-01-26

    Hedgehog (Hh) signaling is a key regulatory pathway during development and also has a functional role in mature neurons. Here, we show that Hh signaling regulates the odor response in adult Drosophila olfactory sensory neurons (OSNs). We demonstrate that this is achieved by regulating odorant receptor (OR) transport to and within the primary cilium in OSN neurons. Regulation relies on ciliary localization of the Hh signal transducer Smoothened (Smo). We further demonstrate that the Hh- and Smo-dependent regulation of the kinesin-like protein Cos2 acts in parallel to the intraflagellar transport system (IFT) to localize ORs within the cilium compartment. These findings expand our knowledge of Hh signaling to encompass chemosensory modulation and receptor trafficking.

  15. Epigenetic regulation of human hedgehog interacting protein in glioma cell lines and primary tumor samples

    PubMed Central

    Shahi, Mehdi H.; Zazpe, Idoya; Afzal, Mohammad; Sinha, Subrata; Rebhun, Robert B.; Meléndez, Bárbara; Rey, Juan A.

    2016-01-01

    Glioma constitutes one of the most common groups of brain tumors, and its prognosis is influenced by different genetic and epigenetic modulations. In this study, we demonstrated low or no expression of hedgehog interacting protein (HHIP) in most of the cell lines and primary glioma tumor samples. We further proceeded to promoter methylation study of this gene in the same cell lines and primary tumor samples and found 87 % (7/8) HHIP methylation in glioblastoma cell lines and 75 % (33/44) in primary tumor samples. These methylation pattern correlates with low or unexpressed HHIP in both cell lines and primary tumor samples. Our results suggest the possibility of epigenetic regulation of this gene in glioma, similarly to medulloblastoma, gastric, hepatic, and pancreatic cancers. Also, HHIP might be a diagnostic or prognostic marker in glioma and help to the detection of these tumors in early stages of disease. PMID:25416442

  16. SOX18 Is a Novel Target Gene of Hedgehog Signaling in Cervical Carcinoma Cell Lines

    PubMed Central

    Popovic, Jelena; Schwirtlich, Marija; Rankovic, Branislava; Stevanovic, Milena

    2015-01-01

    Although there is much evidence showing functional relationship between Hedgehog pathway, in particular Sonic hedgehog, and SOX transcription factors during embryonic development, scarce data are available regarding their crosstalk in cancer cells. SOX18 protein plays an important role in promoting tumor angiogenesis and therefore emerged as a promising potential target in antiangiogenic tumor therapy. Recently it became evident that expression of SOX18 gene in tumors is not restricted to endothelium of accompanying blood and lymphatic vessels, but in tumor cells as well.In this paper we have identified human SOX18 gene as a novel target gene of Hedgehog signaling in cervical carcinoma cell lines. We have presented data showing that expression of SOX18 gene is regulated by GLI1 and GLI2 transcription factors, final effectors of Hedgehog signaling, and that modulation of Hedgehog signaling activity in considerably influence SOX18 expression. We consider important that Hedgehog pathway inhibitors reduced SOX18 expression, thus showing, for the first time, possibility for manipulationwith SOX18 gene expression. In addition, we analyzed the role of SOX18 in malignant potential of cervical carcinoma cell line, and showed that its overexpression has no influence on cells proliferation and viability, but substantially promotes migration and invasion of cells in vitro. Pro-migratory effect of SOX18 suggests its role in promoting malignant spreading, possibly in response to Hedgehog activation. PMID:26588701

  17. IFT80 is essential for chondrocyte differentiation by regulating hedgehog and Wnt signaling pathways

    PubMed Central

    Wang, Changdong; Yuan, Xue; Yang, Shuying

    2013-01-01

    Partial mutation of intraflagellar transport 80 (IFT80) in humans causes Jeune asphyxiating thoracic dystrophy (JATD) and short-rib polydactyly (SRP) syndrome type III. These diseases are autosomal recessive chondrodysplasias that share clinical similarities, including shortened long bones and constricted thoracic cage. However, the role and mechanism of IFT80 in the regulation of chondrocyte differentiation and function remain largely unknown. We hypothesize that IFT80 is required for the formation and function of cilia and plays a critical role in chondrogenic differentiation by regulating Hedgehog (Hh) and Wingless (Wnt) signaling pathways. To test this hypothesis, we first analyzed the IFT80 expression pattern and found that IFT80 was predominantly expressed in growth plate chondrocytes and during chondrogenic differentiation. Silencing IFT80 impaired cilia formation and chondrogenic differentiation in mouse bone marrow derived stromal cells (BMSCs), and decreased the expression of chondrocyte marker genes—collagen II and aggrecan. Additionally, silencing IFT80 down-regulated Hh signaling activity whereas up-regulated Wnt signaling activity. The overexpression of Gli2 in IFT80-silenced cells promoted chondrogenesis and recovered the chondrogenic deficiency from IFT80 silencing. Overall, our results demonstrate that IFT80 is essential for chondrocyte differentiation by regulating the Hh and Wnt signaling pathways. PMID:23333501

  18. Regulation of Smoothened Phosphorylation and High-Level Hedgehog Signaling Activity by a Plasma Membrane Associated Kinase.

    PubMed

    Li, Shuangxi; Li, Shuang; Han, Yuhong; Tong, Chao; Wang, Bing; Chen, Yongbin; Jiang, Jin

    2016-06-01

    Hedgehog (Hh) signaling controls embryonic development and adult tissue homeostasis through the G protein coupled receptor (GPCR)-family protein Smoothened (Smo). Upon stimulation, Smo accumulates on the cell surface in Drosophila or primary cilia in vertebrates, which is thought to be essential for its activation and function, but the underlying mechanisms remain poorly understood. Here we show that Hh stimulates the binding of Smo to a plasma membrane-associated kinase Gilgamesh (Gish)/CK1γ and that Gish fine-tunes Hh pathway activity by phosphorylating a Ser/Thr cluster (CL-II) in the juxtamembrane region of Smo carboxyl-terminal intracellular tail (C-tail). We find that CL-II phosphorylation is promoted by protein kinase A (PKA)-mediated phosphorylation of Smo C-tail and depends on cell surface localization of both Gish and Smo. Consistent with CL-II being critical for high-threshold Hh target gene expression, its phosphorylation appears to require higher levels of Hh or longer exposure to the same level of Hh than PKA-site phosphorylation on Smo. Furthermore, we find that vertebrate CK1γ is localized at the primary cilium to promote Smo phosphorylation and Sonic hedgehog (Shh) pathway activation. Our study reveals a conserved mechanism whereby Hh induces a change in Smo subcellular localization to promote its association with and activation by a plasma membrane localized kinase, and provides new insight into how Hh morphogen progressively activates Smo. PMID:27280464

  19. Regulation of Smoothened Phosphorylation and High-Level Hedgehog Signaling Activity by a Plasma Membrane Associated Kinase

    PubMed Central

    Tong, Chao; Wang, Bing; Chen, Yongbin; Jiang, Jin

    2016-01-01

    Hedgehog (Hh) signaling controls embryonic development and adult tissue homeostasis through the G protein coupled receptor (GPCR)-family protein Smoothened (Smo). Upon stimulation, Smo accumulates on the cell surface in Drosophila or primary cilia in vertebrates, which is thought to be essential for its activation and function, but the underlying mechanisms remain poorly understood. Here we show that Hh stimulates the binding of Smo to a plasma membrane-associated kinase Gilgamesh (Gish)/CK1γ and that Gish fine-tunes Hh pathway activity by phosphorylating a Ser/Thr cluster (CL-II) in the juxtamembrane region of Smo carboxyl-terminal intracellular tail (C-tail). We find that CL-II phosphorylation is promoted by protein kinase A (PKA)-mediated phosphorylation of Smo C-tail and depends on cell surface localization of both Gish and Smo. Consistent with CL-II being critical for high-threshold Hh target gene expression, its phosphorylation appears to require higher levels of Hh or longer exposure to the same level of Hh than PKA-site phosphorylation on Smo. Furthermore, we find that vertebrate CK1γ is localized at the primary cilium to promote Smo phosphorylation and Sonic hedgehog (Shh) pathway activation. Our study reveals a conserved mechanism whereby Hh induces a change in Smo subcellular localization to promote its association with and activation by a plasma membrane localized kinase, and provides new insight into how Hh morphogen progressively activates Smo. PMID:27280464

  20. The Hedgehog signalling pathway in bone formation

    PubMed Central

    Yang, Jing; Andre, Philipp; Ye, Ling; Yang, Ying-Zi

    2015-01-01

    The Hedgehog (Hh) signalling pathway plays many important roles in development, homeostasis and tumorigenesis. The critical function of Hh signalling in bone formation has been identified in the past two decades. Here, we review the evolutionarily conserved Hh signalling mechanisms with an emphasis on the functions of the Hh signalling pathway in bone development, homeostasis and diseases. In the early stages of embryonic limb development, Sonic Hedgehog (Shh) acts as a major morphogen in patterning the limb buds. Indian Hedgehog (Ihh) has an essential function in endochondral ossification and induces osteoblast differentiation in the perichondrium. Hh signalling is also involved intramembrane ossification. Interactions between Hh and Wnt signalling regulate cartilage development, endochondral bone formation and synovial joint formation. Hh also plays an important role in bone homeostasis, and reducing Hh signalling protects against age-related bone loss. Disruption of Hh signalling regulation leads to multiple bone diseases, such as progressive osseous heteroplasia. Therefore, understanding the signalling mechanisms and functions of Hh signalling in bone development, homeostasis and diseases will provide important insights into bone disease prevention, diagnoses and therapeutics. PMID:26023726

  1. The T-box transcription factor Midline regulates wing development by repressing wingless and hedgehog in Drosophila

    PubMed Central

    Fu, Chong-Lei; Wang, Xian-Feng; Cheng, Qian; Wang, Dan; Hirose, Susumu; Liu, Qing-Xin

    2016-01-01

    Wingless (Wg) and Hedgehog (Hh) signaling pathways are key players in animal development. However, regulation of the expression of wg and hh are not well understood. Here, we show that Midline (Mid), an evolutionarily conserved transcription factor, expresses in the wing disc of Drosophila and plays a vital role in wing development. Loss or knock down of mid in the wing disc induced hyper-expression of wingless (wg) and yielded cocked and non-flat wings. Over-expression of mid in the wing disc markedly repressed the expression of wg, DE-Cadherin (DE-Cad) and armadillo (arm), and resulted in a small and blistered wing. In addition, a reduction in the dose of mid enhanced phenotypes of a gain-of-function mutant of hedgehog (hh). We also observed repression of hh upon overexpression of mid in the wing disc. Taken together, we propose that Mid regulates wing development by repressing wg and hh in Drosophila. PMID:27301278

  2. The T-box transcription factor Midline regulates wing development by repressing wingless and hedgehog in Drosophila.

    PubMed

    Fu, Chong-Lei; Wang, Xian-Feng; Cheng, Qian; Wang, Dan; Hirose, Susumu; Liu, Qing-Xin

    2016-01-01

    Wingless (Wg) and Hedgehog (Hh) signaling pathways are key players in animal development. However, regulation of the expression of wg and hh are not well understood. Here, we show that Midline (Mid), an evolutionarily conserved transcription factor, expresses in the wing disc of Drosophila and plays a vital role in wing development. Loss or knock down of mid in the wing disc induced hyper-expression of wingless (wg) and yielded cocked and non-flat wings. Over-expression of mid in the wing disc markedly repressed the expression of wg, DE-Cadherin (DE-Cad) and armadillo (arm), and resulted in a small and blistered wing. In addition, a reduction in the dose of mid enhanced phenotypes of a gain-of-function mutant of hedgehog (hh). We also observed repression of hh upon overexpression of mid in the wing disc. Taken together, we propose that Mid regulates wing development by repressing wg and hh in Drosophila. PMID:27301278

  3. The T-box transcription factor Midline regulates wing development by repressing wingless and hedgehog in Drosophila.

    PubMed

    Fu, Chong-Lei; Wang, Xian-Feng; Cheng, Qian; Wang, Dan; Hirose, Susumu; Liu, Qing-Xin

    2016-06-15

    Wingless (Wg) and Hedgehog (Hh) signaling pathways are key players in animal development. However, regulation of the expression of wg and hh are not well understood. Here, we show that Midline (Mid), an evolutionarily conserved transcription factor, expresses in the wing disc of Drosophila and plays a vital role in wing development. Loss or knock down of mid in the wing disc induced hyper-expression of wingless (wg) and yielded cocked and non-flat wings. Over-expression of mid in the wing disc markedly repressed the expression of wg, DE-Cadherin (DE-Cad) and armadillo (arm), and resulted in a small and blistered wing. In addition, a reduction in the dose of mid enhanced phenotypes of a gain-of-function mutant of hedgehog (hh). We also observed repression of hh upon overexpression of mid in the wing disc. Taken together, we propose that Mid regulates wing development by repressing wg and hh in Drosophila.

  4. Analysis of Germline GLI1 Variation Implicates Hedgehog Signalling in the Regulation of Intestinal Inflammatory Pathways

    PubMed Central

    Tremelling, Mark; Noble, Colin L; Nimmo, Elaine R; Tenesa, Albert; Cornelius, Jennine; Torkvist, Leif; Kao, John; Farrington, Susan; Drummond, Hazel E; Ho, Gwo-Tzer; Arnott, Ian D. R; Appelman, Henry D; Diehl, Lauri; Campbell, Harry; Dunlop, Malcolm G; Parkes, Miles; Howie, Sarah E. M; Gumucio, Deborah L; Satsangi, Jack

    2008-01-01

    Background Ulcerative colitis (UC) and Crohn's disease (CD) are polygenic chronic inflammatory bowel diseases (IBD) of high prevalence that are associated with considerable morbidity. The hedgehog (HH) signalling pathway, which includes the transcription factor glioma-associated oncogene homolog 1 (GLI1), plays vital roles in gastrointestinal tract development, homeostasis, and malignancy. We identified a germline variation in GLI1 (within the IBD2 linkage region, 12q13) in patients with IBD. Since this IBD-associated variant encodes a GLI1 protein with reduced function and our expression studies demonstrated down-regulation of the HH response in IBD, we tested whether mice with reduced Gli1 activity demonstrate increased susceptibility to chemically induced colitis. Methods and Findings Using a gene-wide haplotype-tagging approach, germline GLI1 variation was examined in three independent populations of IBD patients and healthy controls from Northern Europe (Scotland, England, and Sweden) totalling over 5,000 individuals. On log-likelihood analysis, GLI1 was associated with IBD, predominantly UC, in Scotland and England (p < 0.0001). A nonsynonymous SNP (rs2228226C→G), in exon 12 of GLI1 (Q1100E) was strongly implicated, with pooled odds ratio of 1.194 (confidence interval = 1.09–1.31, p = 0.0002). GLI1 variants were tested in vitro for transcriptional activity in luciferase assays. Q1100E falls within a conserved motif near the C terminus of GLI1; the variant GLI protein exhibited reduced transactivation function in vitro. In complementary expression studies, we noted the colonic HH response, including GLI1, patched (PTCH), and hedgehog-interacting protein (HHIP), to be down-regulated in patients with UC. Finally, Gli1+/lacZ mice were tested for susceptibility to dextran sodium sulphate (DSS)-induced colitis. Clinical response, histology, and expression of inflammatory cytokines and chemokines were recorded. Gli1+/lacZ mice rapidly developed severe intestinal

  5. Hedgehog Signaling Components Are Expressed in Choroidal Neovascularization in Laser-induced Retinal Lesion.

    PubMed

    Nochioka, Katsunori; Okuda, Hiroaki; Tatsumi, Kouko; Morita, Shoko; Ogata, Nahoko; Wanaka, Akio

    2016-04-28

    Choroidal neovascularization is one of the major pathological changes in age-related macular degeneration, which causes devastating blindness in the elderly population. The molecular mechanism of choroidal neovascularization has been under extensive investigation, but is still an open question. We focused on sonic hedgehog signaling, which is implicated in angiogenesis in various organs. Laser-induced injuries to the mouse retina were made to cause choroidal neovascularization. We examined gene expression of sonic hedgehog, its receptors (patched1, smoothened, cell adhesion molecule down-regulated by oncogenes (Cdon) and biregional Cdon-binding protein (Boc)) and downstream transcription factors (Gli1-3) using real-time RT-PCR. At seven days after injury, mRNAs for Patched1 and Gli1 were upregulated in response to injury, but displayed no upregulation in control retinas. Immunohistochemistry revealed that Patched1 and Gli1 proteins were localized to CD31-positive endothelial cells that cluster between the wounded retina and the pigment epithelium layer. Treatment with the hedgehog signaling inhibitor cyclopamine did not significantly decrease the size of the neovascularization areas, but the hedgehog agonist purmorphamine made the areas significantly larger than those in untreated retina. These results suggest that the hedgehog-signaling cascade may be a therapeutic target for age-related macular degeneration. PMID:27239075

  6. Hedgehog Signaling Components Are Expressed in Choroidal Neovascularization in Laser-induced Retinal Lesion

    PubMed Central

    Nochioka, Katsunori; Okuda, Hiroaki; Tatsumi, Kouko; Morita, Shoko; Ogata, Nahoko; Wanaka, Akio

    2016-01-01

    Choroidal neovascularization is one of the major pathological changes in age-related macular degeneration, which causes devastating blindness in the elderly population. The molecular mechanism of choroidal neovascularization has been under extensive investigation, but is still an open question. We focused on sonic hedgehog signaling, which is implicated in angiogenesis in various organs. Laser-induced injuries to the mouse retina were made to cause choroidal neovascularization. We examined gene expression of sonic hedgehog, its receptors (patched1, smoothened, cell adhesion molecule down-regulated by oncogenes (Cdon) and biregional Cdon-binding protein (Boc)) and downstream transcription factors (Gli1-3) using real-time RT-PCR. At seven days after injury, mRNAs for Patched1 and Gli1 were upregulated in response to injury, but displayed no upregulation in control retinas. Immunohistochemistry revealed that Patched1 and Gli1 proteins were localized to CD31-positive endothelial cells that cluster between the wounded retina and the pigment epithelium layer. Treatment with the hedgehog signaling inhibitor cyclopamine did not significantly decrease the size of the neovascularization areas, but the hedgehog agonist purmorphamine made the areas significantly larger than those in untreated retina. These results suggest that the hedgehog-signaling cascade may be a therapeutic target for age-related macular degeneration. PMID:27239075

  7. Hedgehog signaling pathway regulated the target genes for adipogenesis in silkworm Bombyx mori.

    PubMed

    Liang, Shuang; Chen, Rui-Ting; Zhang, Deng-Pan; Xin, Hu-Hu; Lu, Yan; Wang, Mei-Xian; Miao, Yun-Gen

    2015-10-01

    Hedgehog (Hh) signals regulate invertebrate and vertebrate development, yet the role of the pathway in adipose development remains poorly understood. In this report, we found that Hh pathway components are expressed in the fat body of silkworm larvae. Functional analysis of these components in a BmN cell line model revealed that activation of the Hh gene stimulated transcription of Hh pathway components, but inhibited the expression of the adipose marker gene AP2. Conversely, specific RNA interference-mediated knockdown of Hh resulted in increased AP2 expression. This further showed the regulation of Hh signal on the adipose marker gene. In silkworm larval models, enhanced adipocyte differentiation and an increase in adipocyte cell size were observed in silkworms that had been treated with a specific Hh signaling pathway antagonist, cyclopamine. The fat-body-specific Hh blockade tests were consistent with Hh signaling inhibiting silkworm adipogenesis. Our results indicate that the role of Hh signaling in inhibiting fat formation is conserved in vertebrates and invertebrates.

  8. Secreted HHIP1 interacts with heparan sulfate and regulates Hedgehog ligand localization and function.

    PubMed

    Holtz, Alexander M; Griffiths, Samuel C; Davis, Samantha J; Bishop, Benjamin; Siebold, Christian; Allen, Benjamin L

    2015-06-01

    Vertebrate Hedgehog (HH) signaling is controlled by several ligand-binding antagonists including Patched-1 (PTCH1), PTCH2, and HH-interacting protein 1 (HHIP1), whose collective action is essential for proper HH pathway activity. However, the molecular mechanisms used by these inhibitors remain poorly understood. In this paper, we investigated the mechanisms underlying HHIP1 antagonism of HH signaling. Strikingly, we found evidence that HHIP1 non-cell-autonomously inhibits HH-dependent neural progenitor patterning and proliferation. Furthermore, this non-cell-autonomous antagonism of HH signaling results from the secretion of HHIP1 that is modulated by cell type-specific interactions with heparan sulfate (HS). These interactions are mediated by an HS-binding motif in the cysteine-rich domain of HHIP1 that is required for its localization to the neuroepithelial basement membrane (BM) to effectively antagonize HH pathway function. Our data also suggest that endogenous, secreted HHIP1 localization to HS-containing BMs regulates HH ligand distribution. Overall, the secreted activity of HHIP1 represents a novel mechanism to regulate HH ligand localization and function during embryogenesis. PMID:26056142

  9. SCF (Fbxl17) ubiquitylation of Sufu regulates Hedgehog signaling and medulloblastoma development.

    PubMed

    Raducu, Madalina; Fung, Ella; Serres, Sébastien; Infante, Paola; Barberis, Alessandro; Fischer, Roman; Bristow, Claire; Thézénas, Marie-Laëtitia; Finta, Csaba; Christianson, John C; Buffa, Francesca M; Kessler, Benedikt M; Sibson, Nicola R; Di Marcotullio, Lucia; Toftgård, Rune; D'Angiolella, Vincenzo

    2016-07-01

    Skp1-Cul1-F-box protein (SCF) ubiquitin ligases direct cell survival decisions by controlling protein ubiquitylation and degradation. Sufu (Suppressor of fused) is a central regulator of Hh (Hedgehog) signaling and acts as a tumor suppressor by maintaining the Gli (Glioma-associated oncogene homolog) transcription factors inactive. Although Sufu has a pivotal role in Hh signaling, the players involved in controlling Sufu levels and their role in tumor growth are unknown. Here, we show that Fbxl17 (F-box and leucine-rich repeat protein 17) targets Sufu for proteolysis in the nucleus. The ubiquitylation of Sufu, mediated by Fbxl17, allows the release of Gli1 from Sufu for proper Hh signal transduction. Depletion of Fbxl17 leads to defective Hh signaling associated with an impaired cancer cell proliferation and medulloblastoma tumor growth. Furthermore, we identify a mutation in Sufu, occurring in medulloblastoma of patients with Gorlin syndrome, which increases Sufu turnover through Fbxl17-mediated polyubiquitylation and leads to a sustained Hh signaling activation. In summary, our findings reveal Fbxl17 as a novel regulator of Hh pathway and highlight the perturbation of the Fbxl17-Sufu axis in the pathogenesis of medulloblastoma. PMID:27234298

  10. Androgens regulate Hedgehog signalling and proliferation in androgen-dependent prostate cells.

    PubMed

    Sirab, Nanor; Terry, Stéphane; Giton, Frank; Caradec, Josselin; Chimingqi, Mihelaiti; Moutereau, Stéphane; Vacherot, Francis; de la Taille, Alexandre; Kouyoumdjian, Jean-Claude; Loric, Sylvain

    2012-09-15

    Prostate cancer (PCa) is androgen sensitive in its development and progression to metastatic disease. Hedgehog (Hh) pathway activation is important in the initiation and growth of various carcinomas including PCa. We and others have observed aberrations of Hh pathway during the progression of PCa to the castration-resistant state. The involvement of androgen signalling in Hh pathway activation, however, remains largely elusive. Here we investigate the direct role of androgen signalling on Hh pathway. We examined the effect of Dihydrosterone (DHT), antiandrogen, bicalutamide, and Hh pathway inhibitor, KAAD-cyclopamine in four human prostate cell lines (two cancerous: LNCaP, VCaP, and two normal: PNT2 and PNT2-ARm which harbours a mutant version of androgen receptor (AR) that is commonly found in LNCaP). Cell proliferation as well as Hh pathway members (SHH, IHH, DHH, GLI, PTCH) mRNA expression levels were assessed. We showed that KAAD-cyclopamine decreased cell proliferation of DHT-stimulated LNCaP, VCaP and PNT2-ARm cells. SHH expression was found to be downregulated by DHT in all AR posititve cells. The negative effect of DHT on SHH expression was counteracted when cells were treated by bicalutamide. Importantly, KAAD-cyclopamine treatment seemed to inhibit AR activity. Moreover, bicalutamide as well as KAAD-cyclopamine treatments induced GLI and PTCH expression in VCaP and PNT2-ARm. Our results suggest that Hh pathway activity can be regulated by androgen signalling. Specifically, we show that the DHT-induced inhibition of Hh pathway is AR dependent. The mutual interaction between these two pathways might be important in the regulation of cell proliferation in PCa.

  11. [Primary cilia and hedgehog signaling].

    PubMed

    Fujii, Katsunori

    2015-07-01

    The primary cilium is an immotile organelle protruding from the cell surface in almost all vertebrate cells. Many molecules inside the primary cilia coordinately play a pivotal role, so genetic defects of these components result in diverse congenital malformations of the brain, eye, liver, kidney, and skeleton. Hedgehog signaling is a highly conserved pathway regulating morphogenesis in early development and tumorigenesis postnatally. Recently, advanced molecular biology has revealed that components of hedgehog signaling such as PTCH1, SMO, and GLI specifically translocate within the primary cilium upon the ligand binding of the hedgehog protein, and transduce the biological growth signal from the cell surface to the nucleus. Haploinsufficiency of the components in the primary cilium would inhibit the hedgehog pathway, resulting in developmental anomalies like ventral neural tube defects. Since the hedgehog-dependent pathway is critical for vertebrate development, it is crucial to elucidate the functional roles of hedgehog-related proteins in the primary cilium. PMID:26353446

  12. Lizard tail regeneration: regulation of two distinct cartilage regions by Indian hedgehog.

    PubMed

    Lozito, Thomas P; Tuan, Rocky S

    2015-03-15

    Lizards capable of caudal autotomy exhibit the remarkable ability to "drop" and then regenerate their tails. However, the regenerated lizard tail (RLT) is known as an "imperfect replicate" due to several key anatomical differences compared to the original tail. Most striking of these "imperfections" concerns the skeleton; instead of the vertebrae of the original tail, the skeleton of the RLT takes the form of an unsegmented cartilage tube (CT). Here we have performed the first detailed staging of skeletal development of the RLT CT, identifying two distinct mineralization events. CTs isolated from RLTs of various ages were analyzed by micro-computed tomography to characterize mineralization, and to correlate skeletal development with expression of endochondral ossification markers evaluated by histology and immunohistochemistry. During early tail regeneration, shortly after CT formation, the extreme proximal CT in direct contact with the most terminal vertebra of the original tail develops a growth plate-like region that undergoes endochondral ossification. Proximal CT chondrocytes enlarge, express hypertrophic markers, including Indian hedgehog (Ihh), apoptose, and are replaced by bone. During later stages of tail regeneration, the distal CT mineralizes without endochondral ossification. The sub-perichondrium of the distal CT expresses Ihh, and the perichondrium directly calcifies without cartilage growth plate formation. The calcified CT perichondrium also contains a population of stem/progenitor cells that forms new cartilage in response to TGF-β stimulation. Treatment with the Ihh inhibitor cyclopamine inhibited both proximal CT ossification and distal CT calcification. Thus, while the two mineralization events are spatially, temporally, and mechanistically very different, they both involve Ihh. Taken together, these results suggest that Ihh regulates CT mineralization during two distinct stages of lizard tail regeneration. PMID:25596336

  13. Lizard tail regeneration: regulation of two distinct cartilage regions by Indian hedgehog.

    PubMed

    Lozito, Thomas P; Tuan, Rocky S

    2015-03-15

    Lizards capable of caudal autotomy exhibit the remarkable ability to "drop" and then regenerate their tails. However, the regenerated lizard tail (RLT) is known as an "imperfect replicate" due to several key anatomical differences compared to the original tail. Most striking of these "imperfections" concerns the skeleton; instead of the vertebrae of the original tail, the skeleton of the RLT takes the form of an unsegmented cartilage tube (CT). Here we have performed the first detailed staging of skeletal development of the RLT CT, identifying two distinct mineralization events. CTs isolated from RLTs of various ages were analyzed by micro-computed tomography to characterize mineralization, and to correlate skeletal development with expression of endochondral ossification markers evaluated by histology and immunohistochemistry. During early tail regeneration, shortly after CT formation, the extreme proximal CT in direct contact with the most terminal vertebra of the original tail develops a growth plate-like region that undergoes endochondral ossification. Proximal CT chondrocytes enlarge, express hypertrophic markers, including Indian hedgehog (Ihh), apoptose, and are replaced by bone. During later stages of tail regeneration, the distal CT mineralizes without endochondral ossification. The sub-perichondrium of the distal CT expresses Ihh, and the perichondrium directly calcifies without cartilage growth plate formation. The calcified CT perichondrium also contains a population of stem/progenitor cells that forms new cartilage in response to TGF-β stimulation. Treatment with the Ihh inhibitor cyclopamine inhibited both proximal CT ossification and distal CT calcification. Thus, while the two mineralization events are spatially, temporally, and mechanistically very different, they both involve Ihh. Taken together, these results suggest that Ihh regulates CT mineralization during two distinct stages of lizard tail regeneration.

  14. Expression of Sonic Hedgehog (SHH) in human lung cancer and the impact of YangZheng XiaoJi on SHH-mediated biological function of lung cancer cells and tumor growth.

    PubMed

    Jiang, Wen G; Ye, Lin; Ruge, Fiona; Sun, Ping-Hui; Sanders, Andrew J; Ji, Ki; Lane, Jane; Zhang, Lijian; Satherley, Lucy; Weeks, Hoi P; Zhi, Xiuyi; Gao, Yong; Wei, Cong; Wu, Yiling; Mason, Malcolm D

    2015-03-01

    Sonic Hedgehog (SHH) is a protein that is aberrantly expressed in various human tumors. SHH and its signaling molecules have been indicated as potential therapeutic targets. In the present study, we evaluated the expression of SHH transcript in human non-small cell lung cancer (NSCLC) tissues and investigated the impact of inhibiting SHH together with a traditional Chinese medicine formula, YangZheng XiaoJi (YZXJ), on the function and growth of lung cancer cells. Human NSCLC tissues had significantly higher levels of the SHH transcript compared matched normal lung tissues (n=83). TNM2 tumors and tumors with pleural invasion had higher levels than TNM1 and non-invasive tumors. High SHH levels were associated with a shorter overall survival (OS) of the patients. A SHH inhibitor, cyclopamine, and YZXJ alone or in combination had a marked inhibitory effect on cellular invasion and cellular migration of human lung cancer cells, A549 and SKMES1. YangZheng XiaoJi and its combination with cyclopamine also significantly reduced the growth of lung tumors in vivo together with a reduction of SHH and smoothened (Smo) proteins in the lung tumors. The present study provides evidence that blocking SHH by way of small inhibitor and by YangZheng XiaoJi has a profound influence on lung cancer cells as seen by in vitro invasion and cell migration and in vivo tumor growth. Together with the aberrant expression of SHH in NSCLC tumors in the patients, it is suggested that SHH is a potential target for therapies for NSCLC. PMID:25750281

  15. Folate deprivation enhances invasiveness of human colon cancer cells mediated by activation of sonic hedgehog signaling through promoter hypomethylation and cross action with transcription nuclear factor-kappa B pathway.

    PubMed

    Wang, Tz-Ping; Hsu, Shu-Han; Feng, Hsin-Chun; Huang, Rwei-Fen S

    2012-06-01

    Low folate status is well recognized as one of the metabolic stressors for colorectal cancer carcinogenesis, but its role in colon cancer invasion remains unknown. Activation of the Sonic hedgehog (Shh) signal in interaction with the transcription nuclear factor-kappa B (NF-κB) pathway is crucial for cancer aggressiveness. The aims of this study were to investigate whether and how folate deprivation promotes invasion by colon cancer cells in relation to Shh signaling and NF-κB pathway activation. Cultivation of epithelial colon carcinoma-derived cells (HCT116) in folate-deficient (FD) medium enhanced cellular migration and invasion, in correlation with epithelial-mesenchymal transition (EMT) associated with Snail expression and E-cadherin suppression, increased production of β1 integrin and increased proteolysis by matrix metalloproteinase 2. Blockade of Shh signaling by cyclopamine (CYC) or of NF-κB activation by BAY abolished FD-enhanced EMT and invasion by HCT116 cells. FD cells had 50-80% less intracellular folate, associated with aberrant hypomethylation of the Shh promoter, than control cells, and increased binding of nuclear NF-κB subunit p65 to the Shh promoter region, which coincided with increased Shh expression and protein production of Shh ligand; in addition, the FD-induced Shh signaling targeted Gli1 transcription activator as well as Ptch receptor. The FD-induced Shh induction and activated signaling were blocked by NF-κB inhibitor BAY. Blockade of Shh signaling abrogated FD-promoted NF-κB activation measured by IκBα degradation and by target gene TNFα expression. Taken together, these findings demonstrate that folate deprivation enhanced invasiveness of colon cancer cells mediated by activation of Shh signaling through promoter hypomethylation and cross actions with the NF-κB pathway.

  16. Intestinal cell kinase, a protein associated with endocrine-cerebro-osteodysplasia syndrome, is a key regulator of cilia length and Hedgehog signaling.

    PubMed

    Moon, Heejung; Song, Jieun; Shin, Jeong-Oh; Lee, Hankyu; Kim, Hong-Kyung; Eggenschwiller, Jonathan T; Bok, Jinwoong; Ko, Hyuk Wan

    2014-06-10

    Endocrine-cerebro-osteodysplasia (ECO) syndrome is a recessive genetic disorder associated with multiple congenital defects in endocrine, cerebral, and skeletal systems that is caused by a missense mutation in the mitogen-activated protein kinase-like intestinal cell kinase (ICK) gene. In algae and invertebrates, ICK homologs are involved in flagellar formation and ciliogenesis, respectively. However, it is not clear whether this role of ICK is conserved in mammals and how a lack of functional ICK results in the characteristic phenotypes of human ECO syndrome. Here, we generated Ick knockout mice to elucidate the precise role of ICK in mammalian development and to examine the pathological mechanisms of ECO syndrome. Ick null mouse embryos displayed cleft palate, hydrocephalus, polydactyly, and delayed skeletal development, closely resembling ECO syndrome phenotypes. In cultured cells, down-regulation of Ick or overexpression of kinase-dead or ECO syndrome mutant ICK resulted in an elongation of primary cilia and abnormal Sonic hedgehog (Shh) signaling. Wild-type ICK proteins were generally localized in the proximal region of cilia near the basal bodies, whereas kinase-dead ICK mutant proteins accumulated in the distal part of bulged ciliary tips. Consistent with these observations in cultured cells, Ick knockout mouse embryos displayed elongated cilia and reduced Shh signaling during limb digit patterning. Taken together, these results indicate that ICK plays a crucial role in controlling ciliary length and that ciliary defects caused by a lack of functional ICK leads to abnormal Shh signaling, resulting in congenital disorders such as ECO syndrome.

  17. Zebrafish blowout provides genetic evidence for Patched1 mediated negative regulation of Hedgehog signaling within the proximal optic vesicle of the vertebrate eye

    PubMed Central

    Lee, Jiwoon; Willer, Jason R.; Willer, Gregory B.; Smith, Kierann; Gregg, Ronald G.; Gross, Jeffrey M.

    2008-01-01

    In this study we have characterized the ocular defects in the recessive zebrafish mutant blowout that presents with a variably penetrant coloboma phenotype. blowout mutants develop unilateral or bilateral colobomas and as a result, the retina and retinal pigmented epithelium are not contained within the optic cup. Colobomas result from defects in optic stalk morphogenesis whereby the optic stalk extends into the retina and impedes the lateral edges of the choroid fissure from meeting and fusing. The expression domain of the proximal optic vesicle marker pax2a is expanded in blowout at the expense of the distal optic vesicle marker pax6, suggesting that the initial patterning of the optic vesicle into proximal and distal territories is disrupted in blowout. Later aspects of distal optic cup formation (i.e. retina development) are normal in blowout mutants, however. Positional cloning of blowout identified a nonsense mutation in patched1, a negative regulator of the Hedgehog pathway, as the underlying cause of the blowout phenotype. Expanded domains of expression of the Hedgehog target genes patched1 and patched2 were observed in blowout, consistent with a loss of Patched1 function and upregulation of Hedgehog pathway activity. Moreover, colobomas in blowout could be suppressed by pharmacologically inhibiting the Hedgehog pathway with cyclopamine, and maximal rescue occurred when embryos were exposed to cyclopamine between 5.5 and 13 hours post fertilization. These observations highlight the critical role that Hedgehog pathway activity plays in mediating patterning of the proximal/distal axis of the optic vesicle during the early phases of eye development and they provide genetic confirmation for the integral role that patched1-mediated negative regulation of Hedgehog signaling plays during vertebrate eye development. PMID:18479681

  18. Regulation of hedgehog Ligand Expression by the N-End Rule Ubiquitin-Protein Ligase Hyperplastic Discs and the Drosophila GSK3β Homologue, Shaggy

    PubMed Central

    Moncrieff, Sophie; Moncan, Matthieu; Scialpi, Flavia; Ditzel, Mark

    2015-01-01

    Hedgehog (Hh) morphogen signalling plays an essential role in tissue development and homeostasis. While much is known about the Hh signal transduction pathway, far less is known about the molecules that regulate the expression of the hedgehog (hh) ligand itself. Here we reveal that Shaggy (Sgg), the Drosophila melanogaster orthologue of GSK3β, and the N-end Rule Ubiquitin-protein ligase Hyperplastic Discs (Hyd) act together to co-ordinate Hedgehog signalling through regulating hh ligand expression and Cubitus interruptus (Ci) expression. Increased hh and Ci expression within hyd mutant clones was effectively suppressed by sgg RNAi, placing sgg downstream of hyd. Functionally, sgg RNAi also rescued the adult hyd mutant head phenotype. Consistent with the genetic interactions, we found Hyd to physically interact with Sgg and Ci. Taken together we propose that Hyd and Sgg function to co-ordinate hh ligand and Ci expression, which in turn influences important developmental signalling pathways during imaginal disc development. These findings are important as tight temporal/spatial regulation of hh ligand expression underlies its important roles in animal development and tissue homeostasis. When deregulated, hh ligand family misexpression underlies numerous human diseases (e.g., colorectal, lung, pancreatic and haematological cancers) and developmental defects (e.g., cyclopia and polydactyly). In summary, our Drosophila-based findings highlight an apical role for Hyd and Sgg in initiating Hedgehog signalling, which could also be evolutionarily conserved in mammals. PMID:26334301

  19. BMP and Hedgehog Regulate Distinct AGM Hematopoietic Stem Cells Ex Vivo

    PubMed Central

    Crisan, Mihaela; Solaimani Kartalaei, Parham; Neagu, Alex; Karkanpouna, Sofia; Yamada-Inagawa, Tomoko; Purini, Caterina; Vink, Chris S.; van der Linden, Reinier; van Ijcken, Wilfred; Chuva de Sousa Lopes, Susana M.; Monteiro, Rui; Mummery, Christine; Dzierzak, Elaine

    2016-01-01

    Summary Hematopoietic stem cells (HSC), the self-renewing cells of the adult blood differentiation hierarchy, are generated during embryonic stages. The first HSCs are produced in the aorta-gonad-mesonephros (AGM) region of the embryo through endothelial to a hematopoietic transition. BMP4 and Hedgehog affect their production and expansion, but it is unknown whether they act to affect the same HSCs. In this study using the BRE GFP reporter mouse strain that identifies BMP/Smad-activated cells, we find that the AGM harbors two types of adult-repopulating HSCs upon explant culture: One type is BMP-activated and the other is a non-BMP-activated HSC type that is indirectly controlled by Hedgehog signaling through the VEGF pathway. Transcriptomic analyses demonstrate that the two HSC types express distinct but overlapping genetic programs. These results revealing the bifurcation in HSC types at early embryonic stages in the AGM explant model suggest that their development is dependent upon the signaling molecules in the microenvironment. PMID:26923823

  20. Habenular Neurogenesis in Zebrafish Is Regulated by a Hedgehog, Pax6 Proneural Gene Cascade

    PubMed Central

    Naye, François; Peers, Bernard; Roussigné, Myriam; Blader, Patrick

    2016-01-01

    The habenulae are highly conserved nuclei in the dorsal diencephalon that connect the forebrain to the midbrain and hindbrain. These nuclei have been implicated in a broad variety of behaviours in humans, primates, rodents and zebrafish. Despite this, the molecular mechanisms that control the genesis and differentiation of neural progenitors in the habenulae remain relatively unknown. We have previously shown that, in zebrafish, the timing of habenular neurogenesis is left-right asymmetric and that in the absence of Nodal signalling this asymmetry is lost. Here, we show that habenular neurogenesis requires the homeobox transcription factor Pax6a and the redundant action of two proneural bHLH factors, Neurog1 and Neurod4. We present evidence that Hedgehog signalling is required for the expression of pax6a, which is in turn necessary for the expression of neurog1 and neurod4. Finally, we demonstrate by pharmacological inhibition that Hedgehog signalling is required continuously during habenular neurogenesis and by cell transplantation experiments that pathway activation is required cell autonomously. Our data sheds light on the mechanism underlying habenular development that may provide insights into how Nodal signalling imposes asymmetry on the timing of habenular neurogenesis. PMID:27387288

  1. A review of hedgehog signaling in cranial bone development

    PubMed Central

    Pan, Angel; Chang, Le; Nguyen, Alan; James, Aaron W.

    2013-01-01

    During craniofacial development, the Hedgehog (HH) signaling pathway is essential for mesodermal tissue patterning and differentiation. The HH family consists of three protein ligands: Sonic Hedgehog (SHH), Indian Hedgehog (IHH), and Desert Hedgehog (DHH), of which two are expressed in the craniofacial complex (IHH and SHH). Dysregulations in HH signaling are well documented to result in a wide range of craniofacial abnormalities, including holoprosencephaly (HPE), hypotelorism, and cleft lip/palate. Furthermore, mutations in HH effectors, co-receptors, and ciliary proteins result in skeletal and craniofacial deformities. Cranial suture morphogenesis is a delicate developmental process that requires control of cell commitment, proliferation and differentiation. This review focuses on both what is known and what remains unknown regarding HH signaling in cranial suture morphogenesis and intramembranous ossification. As demonstrated from murine studies, expression of both SHH and IHH is critical to the formation and fusion of the cranial sutures and calvarial ossification. SHH expression has been observed in the cranial suture mesenchyme and its precise function is not fully defined, although some postulate SHH to delay cranial suture fusion. IHH expression is mainly found on the osteogenic fronts of the calvarial bones, and functions to induce cell proliferation and differentiation. Unfortunately, neonatal lethality of IHH deficient mice precludes a detailed examination of their postnatal calvarial phenotype. In summary, a number of basic questions are yet to be answered regarding domains of expression, developmental role, and functional overlap of HH morphogens in the calvaria. Nevertheless, SHH and IHH ligands are integral to cranial suture development and regulation of calvarial ossification. When HH signaling goes awry, the resultant suite of morphologic abnormalities highlights the important roles of HH signaling in cranial development. PMID:23565096

  2. Stromal cells expressing hedgehog-interacting protein regulate the proliferation of myeloid neoplasms

    PubMed Central

    Kobune, M; Iyama, S; Kikuchi, S; Horiguchi, H; Sato, T; Murase, K; Kawano, Y; Takada, K; Ono, K; Kamihara, Y; Hayashi, T; Miyanishi, K; Sato, Y; Takimoto, R; Kato, J

    2012-01-01

    Aberrant reactivation of hedgehog (Hh) signaling has been described in a wide variety of human cancers including cancer stem cells. However, involvement of the Hh-signaling system in the bone marrow (BM) microenvironment during the development of myeloid neoplasms is unknown. In this study, we assessed the expression of Hh-related genes in primary human CD34+ cells, CD34+ blastic cells and BM stromal cells. Both Indian Hh (Ihh) and its signal transducer, smoothened (SMO), were expressed in CD34+ acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS)-derived cells. However, Ihh expression was relatively low in BM stromal cells. Remarkably, expression of the intrinsic Hh-signaling inhibitor, human Hh-interacting protein (HHIP) in AML/MDS-derived stromal cells was markedly lower than in healthy donor-derived stromal cells. Moreover, HHIP expression levels in BM stromal cells highly correlated with their supporting activity for SMO+ leukemic cells. Knockdown of HHIP gene in stromal cells increased their supporting activity although control cells marginally supported SMO+ leukemic cell proliferation. The demethylating agent, 5-aza-2′-deoxycytidine rescued HHIP expression via demethylation of HHIP gene and reduced the leukemic cell-supporting activity of AML/MDS-derived stromal cells. This indicates that suppression of stromal HHIP could be associated with the proliferation of AML/MDS cells. PMID:22961059

  3. Curcumin regulates cell fate and metabolism by inhibiting hedgehog signaling in hepatic stellate cells.

    PubMed

    Lian, Naqi; Jiang, Yuanyuan; Zhang, Feng; Jin, Huanhuan; Lu, Chunfeng; Wu, Xiafei; Lu, Yin; Zheng, Shizhong

    2015-07-01

    Accumulating evidence indicates that Hedgehog (Hh) signaling becomes activated in chronic liver injury and plays a role in the pathogenesis of hepatic fibrosis. Hepatic stellate cells (HSCs) are Hh-responsive cells and activation of the Hh pathway promotes transdifferentiation of HSCs into myofibroblasts. Targeting Hh signaling may be a novel therapeutic strategy for treatment of liver fibrosis. We previously reported that curcumin has potent antifibrotic effects in vivo and in vitro, but the underlying mechanisms are not fully elucidated. This study shows that curcumin downregulated Patched and Smoothened, two key elements in Hh signaling, but restored Hhip expression in rat liver with carbon tetrachloride-induced fibrosis and in cultured HSCs. Curcumin also halted the nuclear translocation, DNA binding, and transcription activity of Gli1. Moreover, the Hh signaling inhibitor cyclopamine, like curcumin, arrested the cell cycle, induced mitochondrial apoptosis, reduced fibrotic gene expression, restored lipid accumulation, and inhibited invasion and migration in HSCs. However, curcumin's effects on cell fate and fibrogenic properties of HSCs were abolished by the Hh pathway agonist SAG. Furthermore, curcumin and cyclopamine decreased intracellular levels of adenosine triphosphate and lactate, and inhibited the expression and/or function of several key molecules controlling glycolysis. However, SAG abrogated the curcumin effects on these parameters of glycolysis. Animal data also showed that curcumin downregulated glycolysis-regulatory proteins in rat fibrotic liver. These aggregated data therefore indicate that curcumin modulated cell fate and metabolism by disrupting the Hh pathway in HSCs, providing novel molecular insights into curcumin reduction of HSC activation.

  4. Regulation of Ci-SCFSlimb binding, Ci proteolysis and Hedgehog pathway activity by Ci phosphorylation

    PubMed Central

    Smelkinson, Margery G.; Zhou, Qianhe; Kalderon, Daniel

    2007-01-01

    SUMMARY Hedgehog (Hh) proteins signal by inhibiting the proteolytic processing of Ci/Gli family transcription factors and by increasing Ci/Gli specific activity. In the absence of Hh, phosphorylation of Ci/Gli triggers binding to SCF ubiquitin ligase complexes and consequent proteolysis. Here we define the principal SCFSlimb binding site in Ci as an extended variant of a canonical Slimb/β-TRCP binding motif that can be created by PKA-priming of five successive CK1 sites. GSK3 enhances binding primarily through a nearby region of Ci, which may contact an SCF component other than Slimb. Studies of Ci variants with altered CK1 and GSK3 sites suggest that the large number of phosphorylation sites that direct SCFSlimb binding confers a Hh response that is both sensitive and graded, and that in the Drosophila wing disc, morphogenetic responses involve changes in both the level and specific activity of Ci. We also show that when Ci proteolysis is compromised, its specific activity is limited principally by Su(fu) and not by Cos2 cytoplasmic tethering or PKA phosphorylation. PMID:17925225

  5. Hedgehog-mediated paracrine interaction between hepatic stellate cells and marrow-derived mesenchymal stem cells

    SciTech Connect

    Lin Nan Tang Zhaofeng; Deng Meihai; Zhong Yuesi; Lin Jizong; Yang Xuhui; Xiang Peng; Xu Ruiyun

    2008-07-18

    During liver injury, bone marrow-derived mesenchymal stem cells (MSCs) can migrate and differentiate into hepatocytes. Hepatic stellate cell (SC) activation is a pivotal event in the development of liver fibrosis. Therefore, we hypothesized that SCs may play an important role in regulating MSC proliferation and differentiation through the paracrine signaling pathway. We demonstrate that MSCs and SCs both express hedgehog (Hh) pathway components, including its ligands, receptors, and target genes. Transwell co-cultures of SCs and MSCs showed that the SCs produced sonic hedgehog (Shh), which enhanced the proliferation and differentiation of MSCs. These findings demonstrate that SCs indirectly modulate the activity of MSCs in vitro via the Hh pathway, and provide a plausible explanation for the mechanisms of transplanted MSCs in the treatment of liver fibrosis.

  6. Protein-bound Polysaccharide-K Inhibits Hedgehog Signaling Through Down-regulation of MAML3 and RBPJ Transcription Under Hypoxia, Suppressing the Malignant Phenotype in Pancreatic Cancer.

    PubMed

    Yamasaki, Akio; Onishi, Hideya; Imaizumi, Akira; Kawamoto, Makoto; Fujimura, Akiko; Oyama, Yasuhiro; Katano, Mitsuo

    2016-08-01

    Hedgehog signaling is activated in pancreatic cancer and could be a therapeutic target. We previously demonstrated that recombination signal binding protein for immunoglobulin-kappa-J region (RBPJ) and mastermind-like 3 (MAML3) contribute to the hypoxia-induced up-regulation of Smoothened (SMO) transcription. We have also shown that protein-bound polysaccharide-K (PSK) could be effective for refractory pancreatic cancer that down-regulates SMO transcription under hypoxia. In this study, we evaluated whether the anticancer mechanism of PSK involves inhibiting RBPJ and MAML3 expression under hypoxia. PSK reduced SMO, MAML3 and RBPJ expression in pancreatic cancer cells under hypoxia. PSK also blocked RBPJ-induced invasiveness under hypoxia by inhibiting matrix metalloproteinase expression. Lastly, we showed that PSK attenuated RBPJ-induced proliferation both in vitro and in vivo. These results suggest that PSK suppresses Hedgehog signaling through down-regulation of MAML3 and RBPJ transcription under hypoxia, inhibiting the induction of a malignant phenotype in pancreatic cancer. Our results may lead to development of new treatments for refractory pancreatic cancer using PSK as a Hedgehog inhibitor. PMID:27466498

  7. A Comparison of Ci/Gli Activity as Regulated by Sufu in Drosophila and Mammalian Hedgehog Response

    PubMed Central

    Zhang, Chi; Guo, Yurong; Beachy, Philip A.

    2015-01-01

    Suppressor of fused (Su(fu)/Sufu), one of the most conserved components of the Hedgehog (Hh) signaling pathway, binds Ci/Gli transcription factors and impedes activation of target gene expression. In Drosophila, the Su(fu) mutation has a minimal phenotype, and we show here that Ci transcriptional activity in large part is regulated independently of Su(fu) by other pathway components. Mutant mice lacking Sufu in contrast show excessive pathway activity and die as embryos with patterning defects. Here we show that in cultured cells Hh stimulation can augment transcriptional activity of a Gli2 variant lacking Sufu interaction and, surprisingly, that regulation of Hh pathway targets is nearly normal in the neural tube of Sufu-/- mutant embryos that also lack Gli1 function. Some degree of Hh-induced transcriptional activation of Ci/Gli thus can occur independently of Sufu in both flies and mammals. We further note that Sufu loss can also reduce Hh induction of high-threshold neural tube fates, such as floor plate, suggesting a possible positive pathway role for Sufu. PMID:26271100

  8. Activities of the Cytoplasmic Domains of Patched-1 Modulate but Are Not Essential for the Regulation of Canonical Hedgehog Signaling.

    PubMed

    Fleet, Andrew; Lee, Jennifer P Y; Tamachi, Aaliya; Javeed, Imaan; Hamel, Paul A

    2016-08-19

    The Hedgehog (Hh) pathway is a highly conserved signaling cascade crucial for cell fate determination during embryogenesis. Response to the Hh ligands is mediated by the receptor Patched-1 (Ptch1), a 12-pass transmembrane glycoprotein. Despite its essential role in Hh signaling and its activity as a tumor suppressor, Ptch1 remains largely uncharacterized. We demonstrate here that Ptch1 binds to itself to form oligomeric structures. Oligomerization is mediated by two distinct, structurally disordered, intracellular domains spanning amino acids 584-734 ("middle loop") and 1162-1432 (C terminus). However, oligomerization is not required for Ptch1-dependent regulation of the canonical Hh pathway operating through Smo. Expression of a mutant protein that deletes both regions represses the Hh pathway and responds to the addition of Hh ligand independent of its inability to bind other factors such as Smurf2. Additionally, deletion of the cytoplasmic middle loop domain generates a Ptch1 mutant that, despite binding to Hh ligand, constitutively suppresses Hh signaling and increases the length of primary cilia. Constitutive activity because of deletion of this region is reversed by further deletion of specific sequences in the cytoplasmic C-terminal domain. These data reveal an interaction between the cytoplasmic domains of Ptch1 and that these domains modulate Ptch1 activity but are not essential for regulation of the Hh pathway. PMID:27325696

  9. Activities of the Cytoplasmic Domains of Patched-1 Modulate but Are Not Essential for the Regulation of Canonical Hedgehog Signaling.

    PubMed

    Fleet, Andrew; Lee, Jennifer P Y; Tamachi, Aaliya; Javeed, Imaan; Hamel, Paul A

    2016-08-19

    The Hedgehog (Hh) pathway is a highly conserved signaling cascade crucial for cell fate determination during embryogenesis. Response to the Hh ligands is mediated by the receptor Patched-1 (Ptch1), a 12-pass transmembrane glycoprotein. Despite its essential role in Hh signaling and its activity as a tumor suppressor, Ptch1 remains largely uncharacterized. We demonstrate here that Ptch1 binds to itself to form oligomeric structures. Oligomerization is mediated by two distinct, structurally disordered, intracellular domains spanning amino acids 584-734 ("middle loop") and 1162-1432 (C terminus). However, oligomerization is not required for Ptch1-dependent regulation of the canonical Hh pathway operating through Smo. Expression of a mutant protein that deletes both regions represses the Hh pathway and responds to the addition of Hh ligand independent of its inability to bind other factors such as Smurf2. Additionally, deletion of the cytoplasmic middle loop domain generates a Ptch1 mutant that, despite binding to Hh ligand, constitutively suppresses Hh signaling and increases the length of primary cilia. Constitutive activity because of deletion of this region is reversed by further deletion of specific sequences in the cytoplasmic C-terminal domain. These data reveal an interaction between the cytoplasmic domains of Ptch1 and that these domains modulate Ptch1 activity but are not essential for regulation of the Hh pathway.

  10. Regulation of Sufu activity by p66β and Mycbp provides new insight into vertebrate Hedgehog signaling

    PubMed Central

    Lin, Chuwen; Yao, Erica; Wang, Kevin; Nozawa, Yoko; Shimizu, Hirohito; Johnson, Jeffrey R.; Chen, Jau-Nian; Krogan, Nevan J.

    2014-01-01

    Control of Gli function by Suppressor of Fused (Sufu), a major negative regulator, is a key step in mammalian Hedgehog (Hh) signaling, but how this is achieved in the nucleus is unknown. We found that Hh signaling results in reduced Sufu protein levels and Sufu dissociation from Gli proteins in the nucleus, highlighting critical functions of Sufu in the nucleus. Through a proteomic approach, we identified several Sufu-interacting proteins, including p66β (a member of the NuRD [nucleosome remodeling and histone deacetylase] repressor complex) and Mycbp (a Myc-binding protein). p66β negatively and Mycbp positively regulate Hh signaling in cell-based assays and zebrafish. They function downstream from the membrane receptors, Patched and Smoothened, and the primary cilium. Sufu, p66β, Mycbp, and Gli are also detected on the promoters of Hh targets in a dynamic manner. Our results support a new model of Hh signaling in the nucleus. Sufu recruits p66β to block Gli-mediated Hh target gene expression. Meanwhile, Mycbp forms a complex with Gli and Sufu without Hh stimulation but remains inactive. Hh pathway activation leads to dissociation of Sufu/p66β from Gli, enabling Mycbp to promote Gli protein activity and Hh target gene expression. These studies provide novel insight into how Sufu controls Hh signaling in the nucleus. PMID:25403183

  11. HEDGEHOG/GLI-E2F1 axis modulates iASPP expression and function and regulates melanoma cell growth.

    PubMed

    Pandolfi, S; Montagnani, V; Lapucci, A; Stecca, B

    2015-12-01

    HEDGEHOG (HH) signaling is a key regulator of tissue development and its aberrant activation is involved in several cancer types, including melanoma. We and others have shown a reciprocal cross talk between HH signaling and p53, whose function is often impaired in melanoma. Here we present evidence that both GLI1 and GLI2, the final effectors of HH signaling, regulate the transcription factor E2F1 in melanoma cells, by binding to a functional non-canonical GLI consensus sequence. Consistently, we find a significant correlation between E2F1 and PATCHED1 (PTCH1), GLI1 and GLI2 expression in human melanomas. Functionally, we find that E2F1 is a crucial mediator of HH signaling and it is required for melanoma cell proliferation and xenograft growth induced by activation of the HH pathway. Interestingly, we present evidence that the HH/GLI-E2F1 axis positively modulates the inhibitor of apoptosis-stimulating protein of p53 (iASPP) at multiple levels. HH activation induces iASPP expression through E2F1, which directly binds to iASPP promoter. HH pathway also contributes to iASPP function, by the induction of Cyclin B1 and by the E2F1-dependent regulation of CDK1, which are both involved in iASPP activation. Our data show that activation of HH signaling enhances proliferation in presence of E2F1 and promotes apoptosis in its absence or upon CDK1 inhibition, suggesting that E2F1/iASPP dictates the outcome of HH signaling in melanoma. Together, these findings identify a novel HH/GLI-E2F1-iASPP axis that regulates melanoma cell growth and survival, providing an additional mechanism through which HH signaling restrains p53 proapoptotic function. PMID:26024388

  12. The zebrafish-secreted matrix protein you/scube2 is implicated in long-range regulation of hedgehog signaling.

    PubMed

    Kawakami, Atsushi; Nojima, Yasuhiro; Toyoda, Atsushi; Takahoko, Mikako; Satoh, Miki; Tanaka, Hideomi; Wada, Hironori; Masai, Ichiro; Terasaki, Harumi; Sakaki, Yoshiyuki; Takeda, Hiroyuki; Okamoto, Hitoshi

    2005-03-01

    The Hedgehog (Hh) signal plays a pivotal role in induction of ventral neuronal and muscle cell types around the midline during vertebrate development [1]. We report that the gene disrupted in zebrafish you mutants, in which Hh signaling is impaired, encodes the secreted matrix protein Scube2. Consistently, epistasis analyses suggested that Scube2 functions upstream of Hh ligands or through a parallel pathway. In addition, overexpression analyses suggested that Scube2 is an essential, but a permissive, mediator of Hh signaling in zebrafish embryos. Surprisingly, the you gene is expressed in the dorsal neural tube, raising the possibility that Scube2 could indirectly act via a long-range regulator of Hh signaling. The dorsal Bmps have a long-range and opposing influence on Hh signaling [2-5]. We show that neural plate patterning is affected in you mutants in a way that is consistent with the aberrant long-range action of a Bmp-dependent signal. We further show that Bmp activity can be attenuated by the coexpression of Scube2. Our data support the idea that Scube2 can modulate the long-range action of Bmp-dependent signaling in the neural tube and somites.

  13. HEDGEHOG-GLI1 signaling regulates human glioma growth, cancer stem cell self-renewal and tumorigenicity

    PubMed Central

    Clement, Virginie; Sanchez, Pilar; de Tribolet, Nicolas; Radovanovic, Ivan; Altaba, Ariel Ruiz i

    2007-01-01

    Summary Cancer stem cells are rare tumor cells characterized by their ability to self-renew and to induce tumorigenesis. They are present in gliomas and may be responsible for the lethality of these incurable brain tumors. In the most aggressive and invasive type, glioblastoma multiforme (GBM), an average of ±1 year spans the period between detection and death (1). The resistence of gliomas to current therapies may be related to the existence of cancer stem cells (2–6). We find that human gliomas display a stemness signature and demonstrate that HEDGEHOG (HH)-GLI signaling regulates the expression of stemness genes in and the self-renewal of CD133+ glioma cancer stem cells. HH-GLI signaling is also required for sustained glioma growth and survival, displaying additive and synergistic effects with temozolomide, the current chemotherapeutic agent of choice, which does not affect glioma stem cell self-renewal. Finally, interference of HH-GLI signaling with cyclopamine or through lentiviral-mediated silencing demonstrates that the tumorigenicity of human gliomas in nude mice requires an active pathway. Our results reveal the essential role of HH-GLI signaling in controlling the behavior of human glioma cancer stem cells and offer new therapeutic possibilities. PMID:17196391

  14. Hedgehog signalling in the mouse requires intraflagellar transport proteins.

    PubMed

    Huangfu, Danwei; Liu, Aimin; Rakeman, Andrew S; Murcia, Noel S; Niswander, Lee; Anderson, Kathryn V

    2003-11-01

    Intraflagellar transport (IFT) proteins were first identified as essential factors for the growth and maintenance of flagella in the single-celled alga Chlamydomonas reinhardtii. In a screen for embryonic patterning mutations induced by ethylnitrosourea, here we identify two mouse mutants, wimple (wim) and flexo (fxo), that lack ventral neural cell types and show other phenotypes characteristic of defects in Sonic hedgehog signalling. Both mutations disrupt IFT proteins: the wim mutation is an allele of the previously uncharacterized mouse homologue of IFT172; and fxo is a new hypomorphic allele of polaris, the mouse homologue of IFT88. Genetic analysis shows that Wim, Polaris and the IFT motor protein Kif3a are required for Hedgehog signalling at a step downstream of Patched1 (the Hedgehog receptor) and upstream of direct targets of Hedgehog signalling. Our data show that IFT machinery has an essential and vertebrate-specific role in Hedgehog signal transduction. PMID:14603322

  15. COP9-Hedgehog axis regulates the function of the germline stem cell progeny differentiation niche in the Drosophila ovary.

    PubMed

    Lu, Tinglin; Wang, Su; Gao, Yuan; Mao, Ying; Yang, Zhihao; Liu, Luping; Song, Xiaoqing; Ni, Jianquan; Xie, Ting

    2015-12-15

    Both stem cell self-renewal and lineage differentiation are controlled extrinsically as well as intrinsically. Germline stem cells (GSCs) in the Drosophila ovary provide an attractive model in which to study both stem cell self-renewal and lineage differentiation at the molecular and cellular level. Recently, we have proposed that escort cells (ECs) form a differentiation niche to control GSC lineage specification extrinsically. However, it remains poorly understood how the maintenance and function of the differentiation niche are regulated at the molecular level. Here, this study reveals a new role of COP9 in the differentiation niche to modulate autocrine Hedgehog (Hh) signaling, thereby promoting GSC lineage differentiation. COP9, which is a highly conserved protein complex composed of eight CSN subunits, catalyzes the removal of Nedd8 protein modification from target proteins. Our genetic results have demonstrated that all the COP9 components and the hh pathway components, including hh itself, are required in ECs to promote GSC progeny differentiation. Interestingly, COP9 is required in ECs to maintain Hh signaling activity, and activating Hh signaling in ECs can partially bypass the requirement for COP9 in GSC progeny differentiation. Finally, both COP9 and Hh signaling in ECs promote GSC progeny differentiation partly by preventing BMP signaling and maintaining cellular processes. Therefore, this study has demonstrated that the COP9-Hh signaling axis operates in the differentiation niche to promote GSC progeny differentiation partly by maintaining EC cellular processes and preventing BMP signaling. This provides new insight into how the function of the differentiation niche is regulated at the molecular level. PMID:26672093

  16. Sonic Watermarking

    NASA Astrophysics Data System (ADS)

    Tachibana, Ryuki

    2004-12-01

    Audio watermarking has been used mainly for digital sound. In this paper, we extend the range of its applications to live performances with a new composition method for real-time audio watermarking. Sonic watermarking mixes the sound of the watermark signal and the host sound in the air to detect illegal music recordings recorded from auditoriums. We propose an audio watermarking algorithm for sonic watermarking that increases the magnitudes of the host signal only in segmented areas pseudorandomly chosen in the time-frequency plane. The result of a MUSHRA subjective listening test assesses the acoustic quality of the method in the range of "excellent quality." The robustness is dependent on the type of music samples. For popular and orchestral music, a watermark can be stably detected from music samples that have been sonic-watermarked and then once compressed in an MPEG[InlineEquation not available: see fulltext.] layer[InlineEquation not available: see fulltext.] file.

  17. Hedgehog Is a Positive Regulator of FGF Signalling during Embryonic Tracheal Cell Migration

    PubMed Central

    Butí, Elisenda; Mesquita, Duarte; Araújo, Sofia J.

    2014-01-01

    Cell migration is a widespread and complex process that is crucial for morphogenesis and for the underlying invasion and metastasis of human cancers. During migration, cells are steered toward target sites by guidance molecules that induce cell direction and movement through complex intracellular mechanisms. The spatio-temporal regulation of the expression of these guidance molecules is of extreme importance for both normal morphogenesis and human disease. One way to achieve this precise regulation is by combinatorial inputs of different transcription factors. Here we used Drosophila melanogaster mutants with migration defects in the ganglionic branches of the tracheal system to further clarify guidance regulation during cell migration. By studying the cellular consequences of overactivated Hh signalling, using ptc mutants, we found that Hh positively regulates Bnl/FGF levels during embryonic stages. Our results show that Hh modulates cell migration non-autonomously in the tissues surrounding the action of its activity. We further demonstrate that the Hh signalling pathway regulates bnl expression via Stripe (Sr), a zinc-finger transcription factor with homology to the Early Growth Response (EGR) family of vertebrate transcription factors. We propose that Hh modulates embryonic cell migration by participating in the spatio-temporal regulation of bnl expression in a permissive mode. By doing so, we provide a molecular link between the activation of Hh signalling and increased chemotactic responses during cell migration. PMID:24651658

  18. Low-level Ga-Al-As laser irradiation enhances osteoblast proliferation through activation of Hedgehog signaling pathway

    NASA Astrophysics Data System (ADS)

    Li, Qiushi; Qu, Zhou; Chen, Yingxin; Liu, Shujie; Zhou, Yanmin

    2014-12-01

    Low-level laser irradiation has been reported to promote bone formation, but the molecular mechanism is still unclear. Hedgehog signaling pathway has been reported to play an important role in promoting bone formation. The aim of the present study was to examine whether low-level Ga-Al-As laser (808 nm) irradiation could have an effect on Hedgehog signaling pathway during osteoblast proliferation in vitro. Mouse osteoblastic cell line MC3T3-E1 was cultured in vitro. The cultures after laser irradiation (3.75J/cm2) were treated with recombinant N-terminals Sonic Hedgehog (N-Shh)or Hedgehog inhibitor cyclopamine (cy). The experiment was divided into 4 group, group 1:laser irradiation, group 2: laser irradiation and N-Shh, group 3: laser irradiation and cy, group 4:control with no laser irradiation. On day 1,2 and 3,cell proliferation was determined by cell counting, Cell Counting Kit-8.On 12 h and 24 h, cell cycle was detected by flow cytometry. Proliferation activity of laser irradiation and N-Shh group was remarkably increased compared with those of laser irradiation group. Proliferation activity of laser irradiation and cy group was remarkably decreased compared with those of laser irradiation group, however proliferation activity of laser irradiation and cy group was remarkably increased compared with those of control group. These results suggest that low-level Ga-Al-As laser irradiation activate Hedgehog signaling pathway during osteoblast proliferation in vitro. Hedgehog signaling pathway is one of the signaling pathways by which low-level Ga-Al-As laser irradiation regulates osteoblast proliferation.

  19. Hedgehog signaling is a potent regulator of liver lipid metabolism and reveals a GLI-code associated with steatosis.

    PubMed

    Matz-Soja, Madlen; Rennert, Christiane; Schönefeld, Kristin; Aleithe, Susanne; Boettger, Jan; Schmidt-Heck, Wolfgang; Weiss, Thomas S; Hovhannisyan, Amalya; Zellmer, Sebastian; Klöting, Nora; Schulz, Angela; Kratzsch, Jürgen; Guthke, Reinhardt; Gebhardt, Rolf

    2016-01-01

    Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease in industrialized countries and is increasing in prevalence. The pathomechanisms, however, are poorly understood. This study assessed the unexpected role of the Hedgehog pathway in adult liver lipid metabolism. Using transgenic mice with conditional hepatocyte-specific deletion of Smoothened in adult mice, we showed that hepatocellular inhibition of Hedgehog signaling leads to steatosis by altering the abundance of the transcription factors GLI1 and GLI3. This steatotic 'Gli-code' caused the modulation of a complex network of lipogenic transcription factors and enzymes, including SREBP1 and PNPLA3, as demonstrated by microarray analysis and siRNA experiments and could be confirmed in other steatotic mouse models as well as in steatotic human livers. Conversely, activation of the Hedgehog pathway reversed the "Gli-code" and mitigated hepatic steatosis. Collectively, our results reveal that dysfunctions in the Hedgehog pathway play an important role in hepatic steatosis and beyond. PMID:27185526

  20. Hedgehog signaling is a potent regulator of liver lipid metabolism and reveals a GLI-code associated with steatosis

    PubMed Central

    Matz-Soja, Madlen; Rennert, Christiane; Schönefeld, Kristin; Aleithe, Susanne; Boettger, Jan; Schmidt-Heck, Wolfgang; Weiss, Thomas S; Hovhannisyan, Amalya; Zellmer, Sebastian; Klöting, Nora; Schulz, Angela; Kratzsch, Jürgen; Guthke, Reinhardt; Gebhardt, Rolf

    2016-01-01

    Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease in industrialized countries and is increasing in prevalence. The pathomechanisms, however, are poorly understood. This study assessed the unexpected role of the Hedgehog pathway in adult liver lipid metabolism. Using transgenic mice with conditional hepatocyte-specific deletion of Smoothened in adult mice, we showed that hepatocellular inhibition of Hedgehog signaling leads to steatosis by altering the abundance of the transcription factors GLI1 and GLI3. This steatotic 'Gli-code' caused the modulation of a complex network of lipogenic transcription factors and enzymes, including SREBP1 and PNPLA3, as demonstrated by microarray analysis and siRNA experiments and could be confirmed in other steatotic mouse models as well as in steatotic human livers. Conversely, activation of the Hedgehog pathway reversed the "Gli-code" and mitigated hepatic steatosis. Collectively, our results reveal that dysfunctions in the Hedgehog pathway play an important role in hepatic steatosis and beyond. DOI: http://dx.doi.org/10.7554/eLife.13308.001 PMID:27185526

  1. Nkx factors specifically regulate expression of Hedgehog receptor isoforms in early embryonic development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    BACKGROUND: NK homeobox family members are tissue-specific transcription factors that regulate developmental genes. Homozygous disruption of Nkx3.2 produces severe developmental defects of the axial skeleton, skull, spleen, and stomach. Murine mutation of Nkx2.5 results in death at E9 with defects i...

  2. Immunohistochemical evaluation of hedgehog signalling in epithelial/mesenchymal interactions in squamous cell carcinoma transformation: a pilot study.

    PubMed

    Gonzalez, Ana Cristina; Ferreira, Maira; Ariel, Tamires; Reis, Sílvia Regina; Andrade, Zilton; Peixoto Medrado, Alena

    2016-03-01

    Precancerous lesions have been studied because of their carcinogenic potential and their association with squamous cell carcinoma (SCC) has been reported. In the tumour microenvironment, the processes of angiogenesis and tissue remodelling are regulated by a family of proteins (Hedgehog) described as being able to modulate epithelial/mesenchymal interactions. The objective of this study was to perform a comparative study of precancerous lesions and SCCs by immunohistochemistry for the presence of Sonic, Gli2, SMO and Patched proteins, members of the Hedgehog pathway. Sixteen cases diagnosed as actinic cheilitis associated with SCC were compared to normal oral mucosa. The sections were subjected to immunohistochemistry and the positively stained cells were counted by morphometric analysis. There was a significant progressive increase in expression of all proteins of the Hedgehog pathway, both in the epithelium and in the connective tissue, when sections of normal mucosa, dysplasia and carcinoma were compared (P < 0.05). Thus, one may suggest that the Hedgehog pathway in tumour transformation influences SCC, and more studies should be conducted to expand the understanding of the role of these proteins in neoplastic transformation. PMID:26947270

  3. Protein kinase A activity and Hedgehog signaling pathway.

    PubMed

    Kotani, Tomoya

    2012-01-01

    Protein kinase A (PKA) is a well-known kinase that plays fundamental roles in a variety of biological processes. In Hedgehog-responsive cells, PKA plays key roles in proliferation and fate specification by modulating the transduction of Hedgehog signaling. In the absence of Hedgehog, a basal level of PKA activity represses the transcription of Hedgehog target genes. The main substrates of PKA in this process are the Ci/Gli family of bipotential transcription factors, which activate and repress Hedgehog target gene expression. PKA phosphorylates Ci/Gli, promoting the production of the repressor forms of Ci/Gli and thus repressing Hedgehog target gene expression. In contrast, the activation of Hedgehog signaling in response to Hedgehog increases the active forms of Ci/Gli, resulting in Hedgehog target gene expression. Because both decreased and increased levels of PKA activity cause abnormal cell proliferation and alter cell fate specification, the basal level of PKA activity in Hedgehog-responsive cells should be precisely regulated. However, the mechanism by which PKA activity is regulated remains obscure and appears to vary between cell types, tissues, and organisms. To date, two mechanisms have been proposed. One is a classical mechanism in which PKA activity is regulated by a small second messenger, cAMP; the other is a novel mechanism in which PKA activity is regulated by a protein, Misty somites. PMID:22391308

  4. EFCAB7 and IQCE regulate Hedgehog signaling by tethering the EVC-EVC2 complex to the base of primary cilia

    PubMed Central

    Pusapati, Ganesh V.; Hughes, Casey E; Dorn, Karolin V.; Zhang, Dapeng; Sugianto, Priscilla; Aravind, L.; Rohatgi, Rajat

    2014-01-01

    The Hedgehog (Hh) pathway depends on primary cilia in vertebrates, but the signaling machinery within cilia remains incompletely defined. We report the identification of a complex between two ciliary proteins, EFCAB7 and IQCE, which positively regulates the Hh pathway. The EFCAB7-IQCE module anchors the EVC-EVC2 complex in a signaling microdomain at the base of cilia. EVC and EVC2 genes are mutated in Ellis van Creveld and Weyers syndromes, characterized by impaired Hh signaling in skeletal, cardiac and orofacial tissues. EFCAB7 binds to a C-terminal disordered region in EVC2 that is deleted in Weyers patients. EFCAB7 depletion mimics the Weyers cellular phenotype— the mis-localization of EVC-EVC2 within cilia and impaired activation of the transcription factor GLI2. Evolutionary analysis suggests that emergence of these complexes might have been important for adaptation of an ancient organelle, the cilium, for an animal-specific signaling network. PMID:24582806

  5. The Hedgehog Inhibitor Cyclopamine Reduces β-Catenin-Tcf Transcriptional Activity, Induces E-Cadherin Expression, and Reduces Invasion in Colorectal Cancer Cells

    PubMed Central

    Qualtrough, David; Rees, Phil; Speight, Beverley; Williams, Ann C.; Paraskeva, Christos

    2015-01-01

    Colorectal cancer is a major global health problem resulting in over 600,000 deaths world-wide every year with the majority of these due to metastatic disease. Wnt signalling, and more specifically β-catenin-related transcription, has been shown to drive both tumorigenesis and the metastatic process in colorectal neoplasia, yet its complex interactions with other key signalling pathways, such as hedgehog, remain to be elucidated. We have previously shown that the Hedgehog (HH) signalling pathway is active in cells from colorectal tumours, and that inhibition of the pathway with cyclopamine induces apoptosis. We now show that cyclopamine treatment reduces β-catenin related transcription in colorectal cancer cell lines, and that this effect can be reversed by addition of Sonic Hedgehog protein. We also show that cyclopamine concomitantly induces expression of the tumour suppressor and prognostic indicator E-cadherin. Consistent with a role for HH in regulating the invasive potential we show that cyclopamine reduces the expression of transcription factors (Slug, Snail and Twist) associated with the epithelial-mesenchymal transition and reduces the invasiveness of colorectal cancer cells in vitro. Taken together, these data show that pharmacological inhibition of the hedgehog pathway has therapeutic potential in the treatment of colorectal cancer. PMID:26393651

  6. Single-molecule imaging of Hedgehog pathway protein Smoothened in primary cilia reveals binding events regulated by Patched1

    PubMed Central

    Milenkovic, Ljiljana; Weiss, Lucien E.; Yoon, Joshua; Roth, Theodore L.; Su, YouRong S.; Sahl, Steffen J.; Scott, Matthew P.; Moerner, W. E.

    2015-01-01

    Accumulation of the signaling protein Smoothened (Smo) in the membrane of primary cilia is an essential step in Hedgehog (Hh) signal transduction, yet the molecular mechanisms of Smo movement and localization are poorly understood. Using ultrasensitive single-molecule tracking with high spatial/temporal precision (30 nm/10 ms), we discovered that binding events disrupt the primarily diffusive movement of Smo in cilia at an array of sites near the base. The affinity of Smo for these binding sites was modulated by the Hh pathway activation state. Activation, by either a ligand or genetic loss of the negatively acting Hh receptor Patched-1 (Ptch), reduced the affinity and frequency of Smo binding at the base. Our findings quantify activation-dependent changes in Smo dynamics in cilia and highlight a previously unknown step in Hh pathway activation. PMID:26100903

  7. Outfoxing the Hedgehog

    ERIC Educational Resources Information Center

    Barbieri, Richard

    2011-01-01

    Jim Collins's "Good to Great" has attained near-scriptural status in organizations, including nonprofits, which Collins says constitute a third of his readers. The pivot point in "Good to Great" is the Hedgehog Concept. The "Hedgehog Concept" (HC), this author claims, is dangerous for schools because it distorts the nature of education. As Collins…

  8. Olfactomedin 4 deficiency promotes prostate neoplastic progression and is associated with upregulation of the hedgehog-signaling pathway

    PubMed Central

    Li, Hongzhen; Liu, Wenli; Chen, Weiping; Zhu, Jianqiong; Deng, Chu-Xia; Rodgers, Griffin P.

    2015-01-01

    Loss of olfactomedin 4 (OLFM4) gene expression is associated with the progression of human prostate cancer, but its role and the molecular mechanisms involved in this process have not been completely understood. In this study, we found that Olfm4-knockout mice developed prostatic intraepithelial neoplasia and prostatic adenocarcinoma. Importantly, we found that the hedgehog-signaling pathway was significantly upregulated in the Olfm4-knockout mouse model. We also found that restoration of OLFM4 in human prostate-cancer cells that lack OLFM4 expression significantly downregulated hedgehog signaling-pathway component expression. Furthermore, we demonstrated that the OLFM4 protein interacts with sonic hedgehog protein, as well as significantly inhibits GLI-reporter activity. Bioinformatic and immunohistochemistry analyses revealed that decreased OLFM4 and increased SHH expression was significantly associated with advanced human prostate cancer. Thus, olfactomedin 4 appears to play a critical role in regulating progression of prostate cancer, and has potential as a new biomarker for prostate cancer. PMID:26581960

  9. Indian hedgehog B function is required for the specification of oligodendrocyte progenitor cells in the zebrafish CNS.

    PubMed

    Chung, Ah-Young; Kim, Suhyun; Kim, Eunmi; Kim, Dohyun; Jeong, Inyoung; Cha, Young Ryun; Bae, Young-ki; Park, Seung Woo; Lee, Jehee; Park, Hae-Chul

    2013-01-23

    A subset of ventral spinal cord precursors, known as pMN precursor cells, initially generate motor neurons and then oligodendrocyte progenitor cells (OPCs), which migrate and differentiate as myelinating oligodendrocytes in the developing neural tube. The switch between motor neuron and oligodendrocyte production by the pMN neural precursors is an important step in building a functional nervous system. However, the precise mechanism that orchestrates the sequential generation of motor neurons and oligodendrocytes within the common population of pMN precursors is still unclear. The current study demonstrates that Indian Hedgehog b (Ihhb), previously known as Echidna Hedgehog, begins to be expressed in the floor plate cells of the ventral spinal cord at the time of OPC specification in zebrafish embryos. Ihhb loss-of-function analysis revealed that Ihhb function is required for OPC specification from pMN precursors by negatively regulating the proliferation of neural precursors. Finally, results showed that Sonic Hedgehog (Shh) could not replace Ihhb function in OPC specification, suggesting that Ihhb and Shh play separate roles in OPC specification. Altogether, data from the present study suggested a novel mechanism, mediated by Ihhb, for the sequential generation of motor neurons and oligodendrocytes from pMN precursors in the ventral spinal cord of zebrafish embryos.

  10. Indian hedgehog B function is required for the specification of oligodendrocyte progenitor cells in the zebrafish CNS.

    PubMed

    Chung, Ah-Young; Kim, Suhyun; Kim, Eunmi; Kim, Dohyun; Jeong, Inyoung; Cha, Young Ryun; Bae, Young-ki; Park, Seung Woo; Lee, Jehee; Park, Hae-Chul

    2013-01-23

    A subset of ventral spinal cord precursors, known as pMN precursor cells, initially generate motor neurons and then oligodendrocyte progenitor cells (OPCs), which migrate and differentiate as myelinating oligodendrocytes in the developing neural tube. The switch between motor neuron and oligodendrocyte production by the pMN neural precursors is an important step in building a functional nervous system. However, the precise mechanism that orchestrates the sequential generation of motor neurons and oligodendrocytes within the common population of pMN precursors is still unclear. The current study demonstrates that Indian Hedgehog b (Ihhb), previously known as Echidna Hedgehog, begins to be expressed in the floor plate cells of the ventral spinal cord at the time of OPC specification in zebrafish embryos. Ihhb loss-of-function analysis revealed that Ihhb function is required for OPC specification from pMN precursors by negatively regulating the proliferation of neural precursors. Finally, results showed that Sonic Hedgehog (Shh) could not replace Ihhb function in OPC specification, suggesting that Ihhb and Shh play separate roles in OPC specification. Altogether, data from the present study suggested a novel mechanism, mediated by Ihhb, for the sequential generation of motor neurons and oligodendrocytes from pMN precursors in the ventral spinal cord of zebrafish embryos. PMID:23345245

  11. Inhibition of CK2α Down-Regulates Hedgehog/Gli Signaling Leading to a Reduction of a Stem-Like Side Population in Human Lung Cancer Cells

    PubMed Central

    Zhang, Shulin; Wang, Yucheng; Mao, Jian-Hua; Hsieh, David; Kim, Il-Jin; Hu, Li-Min; Xu, Zhidong; Long, Hao; Jablons, David M.; You, Liang

    2012-01-01

    Protein kinase CK2 is frequently elevated in a variety of human cancers. The Hedgehog (Hh) signaling pathway has been implicated in stem cell maintenance, and its aberrant activation has been indicated in several types of cancer, including lung cancer. In this study, we show that CK2 is positively involved in Hh/Gli signaling in lung cancer cell lines A549 and H1299. First, we found a correlation between CK2α and Gli1 mRNA levels in 100 primary lung cancer tissues. Down-regulation of Gli1 expression and transcriptional activity were demonstrated after the silencing of CK2α in lung cancer cells. In addition, CK2α siRNA down-regulated the expression of Hh target genes. Furthermore, two small-molecule CK2α inhibitors led to a dose-dependent inhibition of Gli1 expression and transcriptional activity in lung cancer cells. Reversely, forced over-expression of CK2α resulted in an increase both in Gli1 expression and transcriptional activity in A549 cells. Finally, the inhibition of Hh/Gli by CK2α siRNA led to a reduction of a cancer stem cell-like side population that shows higher ABCG2 expression level. Thus, we report that the inhibition of CK2α down-regulates Hh/Gli signaling and subsequently reduces stem-like side population in human lung cancer cells. PMID:22768056

  12. An essential role for Grk2 in Hedgehog signalling downstream of Smoothened.

    PubMed

    Zhao, Zhonghua; Lee, Raymond Teck Ho; Pusapati, Ganesh V; Iyu, Audrey; Rohatgi, Rajat; Ingham, Philip W

    2016-05-01

    The G-protein-coupled receptor kinase 2 (adrbk2/GRK2) has been implicated in vertebrate Hedgehog (Hh) signalling based on the effects of its transient knock-down in mammalian cells and zebrafish embryos. Here, we show that the response to Hh signalling is effectively abolished in the absence of Grk2 activity. Zebrafish embryos lacking all Grk2 activity are refractory to both Sonic hedgehog (Shh) and oncogenic Smoothened (Smo) activity, but remain responsive to inhibition of cAMP-dependent protein kinase (PKA) activity. Mutation of the kinase domain abrogates the rescuing activity of grk2 mRNA, suggesting that Grk2 acts in a kinase-dependent manner to regulate the response to Hh. Previous studies have suggested that Grk2 potentiates Smo activity by phosphorylating its C-terminal tail (CTT). In the zebrafish embryo, however, phosphomimetic Smo does not display constitutive activity, whereas phospho-null mutants retain activity, implying phosphorylation is neither sufficient nor necessary for Smo function. Since Grk2 rescuing activity requires the integrity of domains essential for its interaction with GPCRs, we speculate that Grk2 may regulate Hh pathway activity by downregulation of a GPCR. PMID:27113758

  13. Membrane Topology of Hedgehog Acyltransferase*

    PubMed Central

    Matevossian, Armine; Resh, Marilyn D.

    2015-01-01

    Hedgehog acyltransferase (Hhat) is a multipass transmembrane enzyme that mediates the covalent attachment of the 16-carbon fatty acid palmitate to the N-terminal cysteine of Sonic Hedgehog (Shh). Palmitoylation of Shh by Hhat is critical for short and long range signaling. Knowledge of the topological organization of Hhat transmembrane helices would enhance our understanding of Hhat-mediated Shh palmitoylation. Bioinformatics analysis of transmembrane domains within human Hhat using 10 different algorithms resulted in highly consistent predictions in the C-terminal, but not in the N-terminal, region of Hhat. To empirically determine the topology of Hhat, we designed and exploited Hhat constructs containing either terminal or 12 different internal epitope tags. We used selective permeabilization coupled with immunofluorescence as well as a protease protection assay to demonstrate that Hhat contains 10 transmembrane domains and 2 re-entrant loops. The invariant His and highly conserved Asp residues within the membrane-bound O-acyltransferase (MBOAT) homology domain are segregated on opposite sides of the endoplasmic reticulum membrane. The localization of His-379 on the lumenal membrane surface is consistent with a role for this invariant residue in catalysis. Analysis of the activity and stability of the Hhat constructs revealed that the C-terminal MBOAT domain is especially sensitive to manipulation. Moreover, there was remarkable similarity in the overall topological organization of Hhat and ghrelin O-acyltransferase, another MBOAT family member. Knowledge of the topological organization of Hhat could serve as an important tool for further design of selective Hhat inhibitors. PMID:25488661

  14. The energy sensor AMPK regulates Hedgehog signaling in human cells through a unique Gli1 metabolic checkpoint

    PubMed Central

    Di Magno, Laura; Basile, Alessio; Coni, Sonia; Manni, Simona; Sdruscia, Giulia; D'Amico, Davide; Antonucci, Laura; Infante, Paola; De Smaele, Enrico; Cucchi, Danilo; Ferretti, Elisabetta; Di Marcotullio, Lucia; Screpanti, Isabella; Canettieri, Gianluca

    2016-01-01

    Hedgehog signaling controls proliferation of cerebellar granule cell precursors (GCPs) and its aberrant activation is a leading cause of Medulloblastoma, the most frequent pediatric brain tumor. We show here that the energy sensor AMPK inhibits Hh signaling by phosphorylating a single residue of human Gli1 that is not conserved in other species. Studies with selective agonists and genetic deletion have revealed that AMPK activation inhibits canonical Hh signaling in human, but not in mouse cells. Indeed we show that AMPK phosphorylates Gli1 at the unique residue Ser408, which is conserved only in primates but not in other species. Once phosphorylated, Gli1 is targeted for proteasomal degradation. Notably, we show that selective AMPK activation inhibits Gli1-driven proliferation and that this effect is linked to Ser408 phosphorylation, which represents a key metabolic checkpoint for Hh signaling. Collectively, this data unveil a novel mechanism of inhibition of Gli1 function, which is exclusive for human cells and may be exploited for the treatment of Medulloblastoma or other Gli1 driven tumors. PMID:26843621

  15. Prognostic value of hedgehog signaling pathway in patients with colon cancer.

    PubMed

    Xu, Meihua; Li, Xinhua; Liu, Ting; Leng, Aimin; Zhang, Guiying

    2012-06-01

    Hedgehog signaling pathway plays an important role in normal mammalian gastrointestinal development and is implicated in the oncogenesis of various tumors. However, its correlation with progression and prognosis of colon cancer has not been well documented. This study was designed to investigate expression patterns of related proteins in hedgehog signaling pathway in colon cancer to elucidate its prognostic value in this tumor. Using human colon cancer and their corresponding non-diseased colon from 228 patients' biopsies, the expression of sonic hedgehog, its receptor Patched, and downstream transcription factor Gli1 was investigated by immunohistochemical staining to assess their association with the clinicopathological characteristics of colon cancer. Disease-free survival and overall survival were examined by Kaplan-Meier estimates and the log-rank test. Prognostic factors were determined by multivariate Cox analysis. One hundred and thirty-eight patients (59.6%) had sonic hedgehog-positive tumors and that the disease-free survival (43.5 vs. 73.3%, P < 0.001), and overall survival rates (50.7 vs. 88.9%, P < 0.001) of patients with sonic hedgehog-positive tumors were much lower than those of patients with sonic hedgehog-negative tumors. In addition, 163 patients (71.5%) had Patched-positive tumors, and the disease-free survival (41.7 vs. 76.9%, P < 0.001) and overall survival rates (55.2 vs. 80.0%, P = 0.002) of patients with Patched-positive tumors were also lower than those of patients with Patched-negative tumors. Moreover, positive Gli1 expression had a bad effect on the disease-free survival (41.9 vs. 73.2%, P < 0.001) and overall survival rate of patients with colon cancer (50.0 vs. 89.3%, P < 0.001). In a multivariate analysis, sonic hedgehog, Patched, and Gli1 status were indicators for poor disease-free survival and overall survival. These results have shown that the increasing expression of sonic hedgehog, Patched, and Gli1 are indicators for a poor

  16. Intraflagellar transport, cilia, and mammalian Hedgehog signaling: analysis in mouse embryonic fibroblasts.

    PubMed

    Ocbina, Polloneal Jymmiel R; Anderson, Kathryn V

    2008-08-01

    Genetic studies in the mouse have shown that Intraflagellar Transport (IFT) is essential for mammalian Hedgehog (Hh) signal transduction. In this study, we take advantage of wild type and IFT mutant mouse embryonic fibroblasts (MEFs) to characterize additional aspects of the relationship between IFT and Hh signaling. Exposure to Sonic hedgehog (Shh) ligand or expression of an activated allele of Smo, SmoA1, activates an Hh reporter in wild-type MEFs, but not in MEFs derived from embryos that lack IFT172 or the Dync2h1 subunit of the retrograde IFT motor. Similarly, decreased activity of either Sufu or PKA, two negative regulators of Hh signal transduction, activates the pathway in wild-type, but not IFT mutant, MEFs. In contrast to wild-type MEFs, Smo is constitutively present in the cilia of Dync2h1 mutant MEFs. This finding suggests that IFT-dependent trafficking of Hh pathway components through the cilium is essential for their function.

  17. Hedgehog associated to microparticles inhibits adipocyte differentiation via a non-canonical pathway

    PubMed Central

    Fleury, Audrey; Hoch, Lucile; Martinez, M. Carmen; Faure, Hélène; Taddei, Maurizio; Petricci, Elena; Manetti, Fabrizio; Girard, Nicolas; Mann, André; Jacques, Caroline; Larghero, Jérôme; Ruat, Martial; Andriantsitohaina, Ramaroson; Le Lay, Soazig

    2016-01-01

    Hedgehog (Hh) is a critical regulator of adipogenesis. Extracellular vesicles are natural Hh carriers, as illustrated by activated/apoptotic lymphocytes specifically shedding microparticles (MP) bearing the morphogen (MPHh+). We show that MPHh+ inhibit adipocyte differentiation and orientate mesenchymal stem cells towards a pro-osteogenic program. Despite a Smoothened (Smo)-dependency, MPHh+ anti-adipogenic effects do not activate a canonical Hh signalling pathway in contrast to those elicited either by the Smo agonist SAG or recombinant Sonic Hedgehog. The Smo agonist GSA-10 recapitulates many of the hallmarks of MPHh+ anti-adipogenic effects. The adipogenesis blockade induced by MPHh+ and GSA-10 was abolished by the Smo antagonist LDE225. We further elucidate a Smo/Lkb1/Ampk axis as the non-canonical Hh pathway used by MPHh+ and GSA-10 to inhibit adipocyte differentiation. Our results highlight for the first time the ability of Hh-enriched MP to signal via a non-canonical pathway opening new perspectives to modulate fat development. PMID:27010359

  18. Vertebrate Hedgehog is secreted on two types of extracellular vesicles with different signaling properties

    PubMed Central

    Vyas, Neha; Walvekar, Ankita; Tate, Dhananjay; Lakshmanan, Vairavan; Bansal, Dhiru; Cicero, Alessandra Lo; Raposo, Graca; Palakodeti, Dasaradhi; Dhawan, Jyotsna

    2014-01-01

    Hedgehog (Hh) is a secreted morphogen that elicits differentiation and patterning in developing tissues. Multiple proposed mechanisms to regulate Hh dispersion includes lipoprotein particles and exosomes. Here we report that vertebrate Sonic Hedgehog (Shh) is secreted on two types of extracellular-vesicles/exosomes, from human cell lines and primary chick notochord cells. Although largely overlapping in size as estimated from electron micrographs, the two exosomal fractions exhibited distinct protein and RNA composition. We have probed the functional properties of these vesicles using cell-based assays of Hh-elicited gene expression. Our results suggest that while both Shh-containing exo-vesicular fractions can activate an ectopic Gli-luciferase construct, only exosomes co-expressing Integrins can activate endogenous Shh target genes HNF3β and Olig2 during the differentiation of mouse ES cells to ventral neuronal progenitors. Taken together, our results demonstrate that primary vertebrate cells secrete Shh in distinct vesicular forms, and support a model where packaging of Shh along with other signaling proteins such as Integrins on exosomes modulates target gene activation. The existence of distinct classes of Shh-containing exosomes also suggests a previously unappreciated complexity for fine-tuning of Shh-mediated gradients and pattern formation. PMID:25483805

  19. Zinc Inhibits Hedgehog Autoprocessing

    PubMed Central

    Xie, Jian; Owen, Timothy; Xia, Ke; Singh, Ajay Vikram; Tou, Emiley; Li, Lingyun; Arduini, Brigitte; Li, Hongmin; Wan, Leo Q.; Callahan, Brian; Wang, Chunyu

    2015-01-01

    Zinc is an essential trace element with wide-ranging biological functions, whereas the Hedgehog (Hh) signaling pathway plays crucial roles in both development and disease. Here we show that there is a mechanistic link between zinc and Hh signaling. The upstream activator of Hh signaling, the Hh ligand, originates from Hh autoprocessing, which converts the Hh precursor protein to the Hh ligand. In an in vitro Hh autoprocessing assay we show that zinc inhibits Hh autoprocessing with a Ki of 2 μm. We then demonstrate that zinc inhibits Hh autoprocessing in a cellular environment with experiments in primary rat astrocyte culture. Solution NMR reveals that zinc binds the active site residues of the Hh autoprocessing domain to inhibit autoprocessing, and isothermal titration calorimetry provided the thermodynamics of the binding. In normal physiology, zinc likely acts as a negative regulator of Hh autoprocessing and inhibits the generation of Hh ligand and Hh signaling. In many diseases, zinc deficiency and elevated level of Hh ligand co-exist, including prostate cancer, lung cancer, ovarian cancer, and autism. Our data suggest a causal relationship between zinc deficiency and the overproduction of Hh ligand. PMID:25787080

  20. The hedgehog system in ovarian follicles of cattle selected for twin ovulations and births: evidence of a link between the IGF and hedgehog systems

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hedgehog signaling is involved in regulation of ovarian function in Drosophila but its role in regulating mammalian ovarian folliculogenesis is less clear. Therefore, gene expression of Indian hedgehog (IHH) and its type 1 receptor, patched 1 (PTCH1), were quantified in bovine granulosa (GC) or the...

  1. Up-regulation of 11β-Hydroxysteroid Dehydrogenase Type 2 Expression by Hedgehog Ligand Contributes to the Conversion of Cortisol Into Cortisone.

    PubMed

    Zhu, Haibin; Zou, Chaochun; Fan, Xueying; Xiong, Wenyi; Tang, Lanfang; Wu, Ximei; Tang, Chao

    2016-09-01

    The cortisol-inactivating enzyme 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) that catalyzes the intracellular inactivation of glucocorticoids plays a pivotal role in human pregnant maintenance and normal fetal development. Given the fact that the main components of Hedgehog (HH) signaling pathway are predominantly expressed in syncytial layer of human placental villi where 11β-HSD2 is robustly expressed, in the present study, we have investigated the potential roles and underlying mechanisms of HH signaling in 11β-HSD2 expression. Activation of HH signaling by a variety of approaches robustly induced 11β-HSD2 expression as well as the 11β-HSD2 activity, whereas suppression of HH signaling significantly attenuated 11β-HSD2 expression as well as the 11β-HSD2 activity in both human primary cytotrophoblasts and trophoblast-like BeWo cells. Moreover, among glioma-associated oncogene (GLI) family transcriptional factors in HH signaling, knockdown of GLI2 but not GLI1 and GLI3 significantly attenuated HH-induced 11β-HSD2 expression and activity, and overexpression of GLI2 activator alone was sufficient to induce 11β-HSD2 expression and activity. Finally, GLI2 not only directly bound to the promoter region of gene hsd11b2 to transactivate hsd11b2 but also formed a heterodimer with RNA polymerase II, an enzyme that catalyzes the transcription of DNA to synthesize mRNAs, resulting in up-regulation of hsd11b2 gene transcription. Taken together, the present study has uncovered a hitherto uncharacterized role of HH/GLI2 signaling in 11β-HSD2 regulation, implicating that HH signaling through GLI2 could be required for the human pregnant maintenance and fetal development. PMID:27379371

  2. Use of a Conditional Ubr5 Mutant Allele to Investigate the Role of an N-End Rule Ubiquitin-Protein Ligase in Hedgehog Signalling and Embryonic Limb Development

    PubMed Central

    Kinsella, Elaine; Dora, Natalie; Mellis, David; Lettice, Laura; Deveney, Paul

    2016-01-01

    Hedgehog (Hh) signalling is a potent regulator of cell fate and function. While much is known about the events within a Hh-stimulated cell, far less is known about the regulation of Hh-ligand production. Drosophila Hyperplastic Discs (Hyd), a ubiquitin-protein ligase, represents one of the few non-transcription factors that independently regulates both hh mRNA expression and pathway activity. Using a murine embryonic stem cell system, we revealed that shRNAi of the mammalian homologue of hyd, Ubr5, effectively prevented retinoic-acid-induced Sonic hedgehog (Shh) expression. We next investigated the UBR5:Hh signalling relationship in vivo by generating and validating a mouse bearing a conditional Ubr5 loss-of-function allele. Conditionally deleting Ubr5 in the early embryonic limb-bud mesenchyme resulted in a transient decrease in Indian hedgehog ligand expression and decreased Hh pathway activity, around E13.5. Although Ubr5-deficient limbs and digits were, on average, shorter than control limbs, the effects were not statistically significant. Hence, while loss of UBR5 perturbed Hedgehog signalling in the developing limb, there were no obvious morphological defects. In summary, we report the first conditional Ubr5 mutant mouse and provide evidence for a role for UBR5 in influencing Hh signalling, but are uncertain to whether the effects on Hedgehog signaling were direct (cell autonomous) or indirect (non-cell-autonomous). Elaboration of the cellular/molecular mechanism(s) involved may help our understanding on diseases and developmental disorders associated with aberrant Hh signalling. PMID:27299863

  3. Use of a Conditional Ubr5 Mutant Allele to Investigate the Role of an N-End Rule Ubiquitin-Protein Ligase in Hedgehog Signalling and Embryonic Limb Development.

    PubMed

    Kinsella, Elaine; Dora, Natalie; Mellis, David; Lettice, Laura; Deveney, Paul; Hill, Robert; Ditzel, Mark

    2016-01-01

    Hedgehog (Hh) signalling is a potent regulator of cell fate and function. While much is known about the events within a Hh-stimulated cell, far less is known about the regulation of Hh-ligand production. Drosophila Hyperplastic Discs (Hyd), a ubiquitin-protein ligase, represents one of the few non-transcription factors that independently regulates both hh mRNA expression and pathway activity. Using a murine embryonic stem cell system, we revealed that shRNAi of the mammalian homologue of hyd, Ubr5, effectively prevented retinoic-acid-induced Sonic hedgehog (Shh) expression. We next investigated the UBR5:Hh signalling relationship in vivo by generating and validating a mouse bearing a conditional Ubr5 loss-of-function allele. Conditionally deleting Ubr5 in the early embryonic limb-bud mesenchyme resulted in a transient decrease in Indian hedgehog ligand expression and decreased Hh pathway activity, around E13.5. Although Ubr5-deficient limbs and digits were, on average, shorter than control limbs, the effects were not statistically significant. Hence, while loss of UBR5 perturbed Hedgehog signalling in the developing limb, there were no obvious morphological defects. In summary, we report the first conditional Ubr5 mutant mouse and provide evidence for a role for UBR5 in influencing Hh signalling, but are uncertain to whether the effects on Hedgehog signaling were direct (cell autonomous) or indirect (non-cell-autonomous). Elaboration of the cellular/molecular mechanism(s) involved may help our understanding on diseases and developmental disorders associated with aberrant Hh signalling. PMID:27299863

  4. Katanin p80 Regulates Human Cortical Development by Limiting Centriole and Cilia Number

    PubMed Central

    Hu, Wen F.; Pomp, Oz; Ben-Omran, Tawfeg; Kodani, Andrew; Henke, Katrin; Mochida, Ganeshwaran H.; Yu, Timothy W.; Woodworth, Mollie B.; Bonnard, Carine; Raj, Grace Selva; Tan, Thong Teck; Hamamy, Hanan; Masri, Amira; Shboul, Mohammad; Al Saffar, Muna; Partlow, Jennifer N.; Al-Dosari, Mohammed; Alazami, Anas; Alowain, Mohammed; Alkuraya, Fowzan S.; Reiter, Jeremy F.; Harris, Matthew P.; Reversade, Bruno; Walsh, Christopher A.

    2015-01-01

    SUMMARY Katanin is a microtubule-severing complex whose catalytic activities are well characterized, but whose in vivo functions are incompletely understood. Human mutations in KATNB1, which encodes the noncatalytic regulatory p80 subunit of katanin, cause severe microlissencephaly. Loss of Katnb1 in mice confirms essential roles in neurogenesis and cell survival, while loss of zebrafish katnb1 reveals specific roles for katnin p80 in early and late developmental stages. Surprisingly, Katnb1 null mutant mouse embryos display hallmarks of aberrant Sonic hedgehog signaling, including holoprosencephaly. KATNB1-deficient human cells show defective proliferation and spindle structure, while Katnb1 null fibroblasts also demonstrate a remarkable excess of centrioles, with supernumerary cilia but deficient Hedgehog signaling. Our results reveal unexpected functions for KATNB1 in regulating overall centriole, mother centriole, and cilia number, and as an essential gene for normal Hedgehog signaling during neocortical development. PMID:25521379

  5. Katanin p80 regulates human cortical development by limiting centriole and cilia number.

    PubMed

    Hu, Wen F; Pomp, Oz; Ben-Omran, Tawfeg; Kodani, Andrew; Henke, Katrin; Mochida, Ganeshwaran H; Yu, Timothy W; Woodworth, Mollie B; Bonnard, Carine; Raj, Grace Selva; Tan, Thong Teck; Hamamy, Hanan; Masri, Amira; Shboul, Mohammad; Al Saffar, Muna; Partlow, Jennifer N; Al-Dosari, Mohammed; Alazami, Anas; Alowain, Mohammed; Alkuraya, Fowzan S; Reiter, Jeremy F; Harris, Matthew P; Reversade, Bruno; Walsh, Christopher A

    2014-12-17

    Katanin is a microtubule-severing complex whose catalytic activities are well characterized, but whose in vivo functions are incompletely understood. Human mutations in KATNB1, which encodes the noncatalytic regulatory p80 subunit of katanin, cause severe microlissencephaly. Loss of Katnb1 in mice confirms essential roles in neurogenesis and cell survival, while loss of zebrafish katnb1 reveals specific roles for katnin p80 in early and late developmental stages. Surprisingly, Katnb1 null mutant mouse embryos display hallmarks of aberrant Sonic hedgehog signaling, including holoprosencephaly. KATNB1-deficient human cells show defective proliferation and spindle structure, while Katnb1 null fibroblasts also demonstrate a remarkable excess of centrioles, with supernumerary cilia but deficient Hedgehog signaling. Our results reveal unexpected functions for KATNB1 in regulating overall centriole, mother centriole, and cilia number, and as an essential gene for normal Hedgehog signaling during neocortical development.

  6. Anteroposterior Limb Skeletal Patterning Requires the Bifunctional Action of SWI/SNF Chromatin Remodeling Complex in Hedgehog Pathway.

    PubMed

    Jeon, Shin; Seong, Rho Hyun

    2016-03-01

    Graded Sonic hedgehog (Shh) signaling governs vertebrate limb skeletal patterning along the anteroposterior (AP) axis by regulating the activity of bifunctional Gli transcriptional regulators. The genetic networks involved in this patterning are well defined, however, the epigenetic control of the process by chromatin remodelers remains unknown. Here, we report that the SWI/SNF chromatin remodeling complex is essential for Shh-driven limb AP patterning. Specific inactivation of Srg3/mBaf155, a core subunit of the remodeling complex, in developing limb buds hampered the transcriptional upregulation of Shh/Gli target genes, including the Shh receptor Ptch1 and its downstream effector Gli1 in the posterior limb bud. In addition, Srg3 deficiency induced ectopic activation of the Hedgehog (Hh) pathway in the anterior mesenchyme, resulting in loss of progressive asymmetry. These defects in the Hh pathway accompanied aberrant BMP activity and disruption of chondrogenic differentiation in zeugopod and autopod primordia. Notably, our data revealed that dual control of the Hh pathway by the SWI/SNF complex is essential for spatiotemporal transcriptional regulation of the BMP antagonist Gremlin1, which affects the onset of chondrogenesis. This study uncovers the bifunctional role of the SWI/SNF complex in the Hh pathway to determine the fate of AP skeletal progenitors. PMID:26959361

  7. Anteroposterior Limb Skeletal Patterning Requires the Bifunctional Action of SWI/SNF Chromatin Remodeling Complex in Hedgehog Pathway.

    PubMed

    Jeon, Shin; Seong, Rho Hyun

    2016-03-01

    Graded Sonic hedgehog (Shh) signaling governs vertebrate limb skeletal patterning along the anteroposterior (AP) axis by regulating the activity of bifunctional Gli transcriptional regulators. The genetic networks involved in this patterning are well defined, however, the epigenetic control of the process by chromatin remodelers remains unknown. Here, we report that the SWI/SNF chromatin remodeling complex is essential for Shh-driven limb AP patterning. Specific inactivation of Srg3/mBaf155, a core subunit of the remodeling complex, in developing limb buds hampered the transcriptional upregulation of Shh/Gli target genes, including the Shh receptor Ptch1 and its downstream effector Gli1 in the posterior limb bud. In addition, Srg3 deficiency induced ectopic activation of the Hedgehog (Hh) pathway in the anterior mesenchyme, resulting in loss of progressive asymmetry. These defects in the Hh pathway accompanied aberrant BMP activity and disruption of chondrogenic differentiation in zeugopod and autopod primordia. Notably, our data revealed that dual control of the Hh pathway by the SWI/SNF complex is essential for spatiotemporal transcriptional regulation of the BMP antagonist Gremlin1, which affects the onset of chondrogenesis. This study uncovers the bifunctional role of the SWI/SNF complex in the Hh pathway to determine the fate of AP skeletal progenitors.

  8. Sex and hedgehog: roles of genes in the hedgehog signaling pathway in mammalian sexual differentiation.

    PubMed

    Franco, Heather L; Yao, Humphrey H-C

    2012-01-01

    The chromosome status of the mammalian embryo initiates a multistage process of sexual development in which the bipotential reproductive system establishes itself as either male or female. These events are governed by intricate cell-cell and interorgan communication that is regulated by multiple signaling pathways. The hedgehog signaling pathway was originally identified for its key role in the development of Drosophila, but is now recognized as a critical developmental regulator in many species, including humans. In addition to its developmental roles, the hedgehog signaling pathway also modulates adult organ function, and misregulation of this pathway often leads to diseases, such as cancer. The hedgehog signaling pathway acts through its morphogenetic ligands that signal from ligand-producing cells to target cells over a specified distance. The target cells then respond in a graded manner based on the concentration of the ligands that they are exposed to. Through this unique mechanism of action, the hedgehog signaling pathway elicits cell fate determination, epithelial-mesenchymal interactions, and cellular homeostasis. Here, we review current findings on the roles of hedgehog signaling in the sexually dimorphic development of the reproductive organs with an emphasis on mammals and comparative evidence in other species.

  9. Gli2 protein expression level is a feasible marker of ligand-dependent hedgehog activation in pancreatic neoplasms.

    PubMed

    Sugiyama, Y; Sasajima, J; Mizukami, Y; Koizumi, K; Kawamoto, T; Ono, Y; Karasaki, H; Tanabe, H; Fujiya, M; Kohgo, Y

    2016-06-01

    The hedgehog pathway is known to promote proliferation of pancreatic ductal adenocarcinoma (PDA) and has been shown to restrain tumor progression. To understand how hedgehog causes these effects, we sought to carefully examine protein expression of hedgehog signaling components during different tumor stages. Genetically engineered mice, Pdx1-Cre;LSL-KrasG12D and Pdx1-Cre;LSL-KrasG12D;p53lox/+, were utilized to model distinct phases of tumorigenesis, pancreatic intraepithelial neoplasm (PanIN) and PDA. Human pancreatic specimens of intraductal papillary mucinous neoplasm (IPMN) and PDA were also employed. PanIN and IPMN lesions highly express Sonic Hedgehog, at a level that is slightly higher than that observed in PDA. GLI2 protein is also expressed in both PanIN/IPMN and PDA. Although there was no difference in the nuclear staining, the cytoplasmic GLI2 level in PDA was modest in comparison to that in PanIN/IPMN. Hedgehog interacting protein was strongly expressed in the precursors, whereas the level in PDA was significantly attenuated. There were no differences in expression of Patched1 at early and late stages. Finally, a strong correlation between Sonic Hedgehog and GLI2 staining was found in both human and murine pancreatic tumors. The results indicate that the GLI2 protein level could serve as a feasible marker of ligand-dependent hedgehog activation in pancreatic neoplasms. PMID:27543868

  10. Cellular Cholesterol Directly Activates Smoothened in Hedgehog Signaling.

    PubMed

    Huang, Pengxiang; Nedelcu, Daniel; Watanabe, Miyako; Jao, Cindy; Kim, Youngchang; Liu, Jing; Salic, Adrian

    2016-08-25

    In vertebrates, sterols are necessary for Hedgehog signaling, a pathway critical in embryogenesis and cancer. Sterols activate the membrane protein Smoothened by binding its extracellular, cysteine-rich domain (CRD). Major unanswered questions concern the nature of the endogenous, activating sterol and the mechanism by which it regulates Smoothened. We report crystal structures of CRD complexed with sterols and alone, revealing that sterols induce a dramatic conformational change of the binding site, which is sufficient for Smoothened activation and is unique among CRD-containing receptors. We demonstrate that Hedgehog signaling requires sterol binding to Smoothened and define key residues for sterol recognition and activity. We also show that cholesterol itself binds and activates Smoothened. Furthermore, the effect of oxysterols is abolished in Smoothened mutants that retain activation by cholesterol and Hedgehog. We propose that the endogenous Smoothened activator is cholesterol, not oxysterols, and that vertebrate Hedgehog signaling controls Smoothened by regulating its access to cholesterol. PMID:27545348

  11. Reciprocal interplay between thyroid hormone and microRNA-21 regulates hedgehog pathway–driven skin tumorigenesis

    PubMed Central

    Di Girolamo, Daniela; Ambrosio, Raffaele; De Stefano, Maria A.; Mancino, Giuseppina; Porcelli, Tommaso; Luongo, Cristina; Di Cicco, Emery; Scalia, Giulia; Vecchio, Luigi Del; Colao, Annamaria; Dlugosz, Andrzej A.; Missero, Caterina; Salvatore, Domenico

    2016-01-01

    The thyroid hormone–inactivating (TH-inactivating) enzyme type 3 iodothyronine deiodinase (D3) is an oncofetal protein that is rarely expressed in adult life but has been shown to be reactivated in the context of proliferation and neoplasms. D3 terminates TH action within the tumor microenvironment, thereby enhancing cancer cell proliferation. However, the pathological role of D3 and the contribution of TH metabolism in cancer have yet to be fully explored. Here, we describe a reciprocal regulation between TH action and the cancer-associated microRNA-21 (miR21) in basal cell carcinoma (BCC) skin tumors. We found that, besides being negatively regulated by TH at the transcriptional level, miR21 attenuates the TH signal by increasing D3 levels. The ability of miR21 to positively regulate D3 was mediated by the tumor suppressor gene GRHL3, a hitherto unrecognized D3 transcriptional inhibitor. Finally, in a BCC mouse model, keratinocyte-specific D3 depletion markedly reduced tumor growth. Together, our results establish TH action as a critical hub of multiple oncogenic pathways and provide functional and mechanistic evidence of the involvement of TH metabolism in BCC tumorigenesis. Moreover, our results identify a miR21/GRHL3/D3 axis that reduces TH in the tumor microenvironment and has potential to be targeted as a therapeutic approach to BCC. PMID:27159391

  12. Reciprocal interplay between thyroid hormone and microRNA-21 regulates hedgehog pathway-driven skin tumorigenesis.

    PubMed

    Di Girolamo, Daniela; Ambrosio, Raffaele; De Stefano, Maria A; Mancino, Giuseppina; Porcelli, Tommaso; Luongo, Cristina; Di Cicco, Emery; Scalia, Giulia; Vecchio, Luigi Del; Colao, Annamaria; Dlugosz, Andrzej A; Missero, Caterina; Salvatore, Domenico; Dentice, Monica

    2016-06-01

    The thyroid hormone-inactivating (TH-inactivating) enzyme type 3 iodothyronine deiodinase (D3) is an oncofetal protein that is rarely expressed in adult life but has been shown to be reactivated in the context of proliferation and neoplasms. D3 terminates TH action within the tumor microenvironment, thereby enhancing cancer cell proliferation. However, the pathological role of D3 and the contribution of TH metabolism in cancer have yet to be fully explored. Here, we describe a reciprocal regulation between TH action and the cancer-associated microRNA-21 (miR21) in basal cell carcinoma (BCC) skin tumors. We found that, besides being negatively regulated by TH at the transcriptional level, miR21 attenuates the TH signal by increasing D3 levels. The ability of miR21 to positively regulate D3 was mediated by the tumor suppressor gene GRHL3, a hitherto unrecognized D3 transcriptional inhibitor. Finally, in a BCC mouse model, keratinocyte-specific D3 depletion markedly reduced tumor growth. Together, our results establish TH action as a critical hub of multiple oncogenic pathways and provide functional and mechanistic evidence of the involvement of TH metabolism in BCC tumorigenesis. Moreover, our results identify a miR21/GRHL3/D3 axis that reduces TH in the tumor microenvironment and has potential to be targeted as a therapeutic approach to BCC. PMID:27159391

  13. Dosage-dependent hedgehog signals integrated with Wnt/β-catenin signaling regulate external genitalia formation as an appendicular program

    PubMed Central

    Miyagawa, Shinichi; Moon, Anne; Haraguchi, Ryuma; Inoue, Chie; Harada, Masayo; Nakahara, Chiaki; Suzuki, Kentaro; Matsumaru, Daisuke; Kaneko, Takehito; Matsuo, Isao; Yang, Lei; Taketo, Makoto M.; Iguchi, Taisen; Evans, Sylvia M.; Yamada, Gen

    2009-01-01

    Embryonic appendicular structures, such as the limb buds and the developing external genitalia, are suitable models with which to analyze the reciprocal interactions of growth factors in the regulation of outgrowth. Although several studies have evaluated the individual functions of different growth factors in appendicular growth, the coordinated function and integration of input from multiple signaling cascades is poorly understood. We demonstrate that a novel signaling cascade governs formation of the embryonic external genitalia [genital tubercle (GT)]. We show that the dosage of Shh signal is tightly associated with subsequent levels of Wnt/β-catenin activity and the extent of external genitalia outgrowth. In Shh-null mouse embryos, both expression of Wnt ligands and Wnt/β-catenin signaling activity are downregulated. β-catenin gain-of-function mutation rescues defective GT outgrowth and Fgf8 expression in Shh-null embryos. These data indicate that Wnt/β-catenin signaling in the distal urethral epithelium acts downstream of Shh signaling during GT outgrowth. The current data also suggest that Wnt/β-catenin regulates Fgf8 expression via Lef/Tcf binding sites in a 3′ conserved enhancer. Fgf8 induces phosphorylation of Erk1/2 and cell proliferation in the GT mesenchyme in vitro, yet Fgf4/8 compound-mutant phenotypes indicate dispensable functions of Fgf4/8 and the possibility of redundancy among multiple Fgfs in GT development. Our results provide new insights into the integration of growth factor signaling in the appendicular developmental programs that regulate external genitalia development. PMID:19906864

  14. Hedgehog and Fgf Signaling Pathways Regulate the Development of tphR-Expressing Serotonergic Raphe Neurons in Zebrafish Embryos

    PubMed Central

    Teraoka, H.; Russell, C.; Regan, J.; Chandrasekhar, A.; Concha, M. L.; Yokoyama, R.; Higashi, K.; Take-uchi, M.; Dong, W.; Hiraga, T.; Holder, N.; Wilson, S. W.

    2009-01-01

    Serotonin (5HT) plays major roles in the physiological regulation of many behavioral processes, including sleep, feeding, and mood, but the genetic mechanisms by which serotonergic neurons arise during development are poorly understood. In the present study, we have investigated the development of serotonergic neurons in the zebrafish. Neurons exhibiting 5HT-immunoreactivity (5HT-IR) are detected from 45 h postfertilization (hpf) in the ventral hindbrain raphe, the hypothalamus, pineal organ, and pretectal area. Tryptophan hydroxylases encode rate-limiting enzymes that function in the synthesis of 5HT. As part of this study, we cloned and analyzed a novel zebrafish tph gene named tphR. Unlike two other zebrafish tph genes (tphD1 and tphD2), tphR is expressed in serotonergic raphe neurons, similar to tph genes in mammalian species. tphR is also expressed in the pineal organ where it is likely to be involved in the pathway leading to synthesis of melatonin. To better understand the signaling pathways involved in the induction of the serotonergic phenotype, we analyzed tphR expression and 5HT-IR in embryos in which either Hh or Fgf signals are abrogated. Hindbrain 5HT neurons are severely reduced in mutants lacking activity of either Ace/Fgf8 or the transcription factor Noi/Pax2.1, which regulates expression of ace/fgf8, and probably other genes encoding signaling proteins. Similarly, serotonergic raphe neurons are absent in embryos lacking Hh activity confirming a conserved role for Hh signals in the induction of these cells. Conversely, over-activation of the Hh pathway increases the number of serotonergic neurons. As in mammals, our results are consistent with the transcription factors Nk2.2 and Gata3 acting downstream of Hh activity in the development of serotonergic raphe neurons. Our results show that the pathways involved in induction of hindbrain serotonergic neurons are likely to be conserved in all vertebrates and help establish the zebrafish as a model

  15. Sonic spectrometer and treatment system

    DOEpatents

    Slomka, B.J.

    1997-06-03

    A novel system and method is developed for treating an object with sonic waveforms. A traveling broad-band sonic waveform containing a broad-band of sonic frequencies is radiated at the object. A traveling reflected sonic waveform containing sonic frequencies reflected by the object is received in response to the traveling broad-band sonic waveform. A traveling transmitted sonic waveform containing sonic frequencies transmitted through the object is also received in response to the traveling broad-band sonic waveform. In a resonance mode, the frequency spectra of the broad-band and reflected sonic waveforms is analyzed so as to select one or more sonic frequencies that cause the object to resonate. An electrical resonance treatment sonic waveform containing the sonic frequencies that cause the object to resonate is then radiated at the object so as to treat the object. In an absorption mode, the frequency spectra of the electrical broad-band, reflected, and transmitted sonic waveforms is compared so as to select one or more sonic frequencies that are absorbed by the object. An electrical absorption treatment sonic waveform containing the sonic frequencies that are absorbed by the object is then radiated at the object so as to treat the object. 1 fig.

  16. Sonic spectrometer and treatment system

    DOEpatents

    Slomka, Bogdan J.

    1997-06-03

    A novel system and method for treating an object with sonic waveforms. A traveling broad-band sonic waveform containing a broad-band of sonic frequencies is radiated at the object. A traveling reflected sonic waveform containing sonic frequencies reflected by the object is received in response to the traveling broad-band sonic waveform. A traveling transmitted sonic waveform containing sonic frequencies transmitted through the object is also received in response to the traveling broad-band sonic waveform. In a resonance mode, the frequency spectra of the broad-band and reflected sonic waveforms is analyzed so as to select one or more sonic frequencies that cause the object to resonate. An electrical resonance treatment sonic waveform containing the sonic frequencies that cause the object to resonate is then radiated at the object so as to treat the object. In an absorption mode, the frequency spectra of the electrical broad-band, reflected, and transmitted sonic waveforms is compared so as to select one or more sonic frequencies that are absorbed by the object. An electrical absorption treatment sonic waveform containing the sonic frequencies that are absorbed by the object is then radiated at the object so as to treat the object.

  17. The Insecticide Synergist Piperonyl Butoxide Inhibits Hedgehog Signaling: Assessing Chemical Risks

    PubMed Central

    Chen, Wei

    2012-01-01

    The spread of chemicals, including insecticides, into the environment often raises public health concerns, as exemplified by a recent epidemiologic study associating in utero piperonyl butoxide (PBO) exposure with delayed mental development. The insecticide synergist PBO is listed among the top 10 chemicals detected in indoor dust; a systematic assessment of risks from PBO exposure, as for many toxicants unfortunately, may be underdeveloped when important biological targets that can cause toxicity are unknown. Hedgehog/Smoothened signaling is critical in neurological development. This study was designed to use novel high-throughput in vitro drug screening technology to identify modulators of Hedgehog signaling in environmental chemicals to assist the assessment of their potential risks. A directed library of 1408 environmental toxicants was screened for Hedgehog/Smoothened antagonist activity using a high-content assay that evaluated the interaction between Smoothened and βarrestin2 green fluorescent protein. PBO was identified as a Hedgehog/Smoothened antagonist capable of inhibiting Hedgehog signaling. We found that PBO bound Smoothened and blocked Smoothened overexpression–induced Gli-luciferase reporter activity but had no effect on Gli-1 downstream transcriptional factor–induced Gli activity. PBO inhibited Sonic Hedgehog ligand–induced Gli signaling and mouse cerebellar granular precursor cell proliferation. Moreover, PBO disrupted zebrafish development. Our findings demonstrate the value of high-throughput target-based screening strategies that can successfully evaluate large numbers of environmental toxicants and identify key targets and unknown biological activity that is helpful in properly assessing potential risks. PMID:22552772

  18. Sonic boom acceptability studies

    NASA Astrophysics Data System (ADS)

    Shepherd, Kevin P.; Sullivan, Brenda M.; Leatherwood, Jack D.; McCurdy, David A.

    1992-04-01

    The determination of the magnitude of sonic boom exposure which would be acceptable to the general population requires, as a starting point, a method to assess and compare individual sonic booms. There is no consensus within the scientific and regulatory communities regarding an appropriate sonic boom assessment metric. Loudness, being a fundamental and well-understood attribute of human hearing was chosen as a means of comparing sonic booms of differing shapes and amplitudes. The figure illustrates the basic steps which yield a calculated value of loudness. Based upon the aircraft configuration and its operating conditions, the sonic boom pressure signature which reaches the ground is calculated. This pressure-time history is transformed to the frequency domain and converted into a one-third octave band spectrum. The essence of the loudness method is to account for the frequency response and integration characteristics of the auditory system. The result of the calculation procedure is a numerical description (perceived level, dB) which represents the loudness of the sonic boom waveform.

  19. Sonic boom acceptability studies

    NASA Technical Reports Server (NTRS)

    Shepherd, Kevin P.; Sullivan, Brenda M.; Leatherwood, Jack D.; Mccurdy, David A.

    1992-01-01

    The determination of the magnitude of sonic boom exposure which would be acceptable to the general population requires, as a starting point, a method to assess and compare individual sonic booms. There is no consensus within the scientific and regulatory communities regarding an appropriate sonic boom assessment metric. Loudness, being a fundamental and well-understood attribute of human hearing was chosen as a means of comparing sonic booms of differing shapes and amplitudes. The figure illustrates the basic steps which yield a calculated value of loudness. Based upon the aircraft configuration and its operating conditions, the sonic boom pressure signature which reaches the ground is calculated. This pressure-time history is transformed to the frequency domain and converted into a one-third octave band spectrum. The essence of the loudness method is to account for the frequency response and integration characteristics of the auditory system. The result of the calculation procedure is a numerical description (perceived level, dB) which represents the loudness of the sonic boom waveform.

  20. The Mode of Hedgehog Binding to Ihog Homologues is Not Conserved Across Different Phyla

    SciTech Connect

    McLellan, J.; Zheng, X; Hauk, G; Ghirlando, R; Beachy, P; Leahy, D

    2008-01-01

    Hedgehog (Hh) proteins specify tissue pattern in metazoan embryos by forming gradients that emanate from discrete sites of expression and elicit concentration-dependent cellular differentiation or proliferation responses1, 2. Cellular responses to Hh and the movement of Hh through tissues are both precisely regulated, and abnormal Hh signalling has been implicated in human birth defects and cancer3, 4, 5, 6, 7. Hh signalling is mediated by its amino-terminal domain (HhN), which is dually lipidated and secreted as part of a multivalent lipoprotein particle8, 9, 10. Reception of the HhN signal is modulated by several cell-surface proteins on responding cells, including Patched (Ptc), Smoothened (Smo), Ihog (known as CDO or CDON in mammals) and the vertebrate-specific proteins Hip (also known as Hhip) and Gas1 (ref. 11). Drosophila Ihog and its vertebrate homologues CDO and BOC contain multiple immunoglobulin and fibronectin type III (FNIII) repeats, and the first FNIII repeat of Ihog binds Drosophila HhN in a heparin-dependent manner12, 13. Surprisingly, pull-down experiments suggest that a mammalian Sonic hedgehog N-terminal domain (ShhN) binds a non-orthologous FNIII repeat of CDO12, 14. Here we report biochemical, biophysical and X-ray structural studies of a complex between ShhN and the third FNIII repeat of CDO. We show that the ShhN-CDO interaction is completely unlike the HhN-Ihog interaction and requires calcium, which binds at a previously undetected site on ShhN. This site is conserved in nearly all Hh proteins and is a hotspot for mediating interactions between ShhN and CDO, Ptc, Hip and Gas1. Mutations in vertebrate Hh proteins causing holoprosencephaly and brachydactyly type A1 map to this calcium-binding site and disrupt interactions with these partners.

  1. Hedgehog signal activation coordinates proliferation and differentiation of fetal liver progenitor cells

    SciTech Connect

    Hirose, Yoshikazu; Itoh, Tohru; Miyajima, Atsushi

    2009-09-10

    Hedgehog (Hh) signaling plays crucial roles in development and homeostasis of various organs. In the adult liver, it regulates proliferation and/or viability of several types of cells, particularly under injured conditions, and is also implicated in stem/progenitor cell maintenance. However, the role of this signaling pathway during the normal developmental process of the liver remains elusive. Although Sonic hedgehog (Shh) is expressed in the ventral foregut endoderm from which the liver derives, the expression disappears at the onset of the liver bud formation, and its possible recurrence at the later stages has not been investigated. Here we analyzed the activation and functional relevance of Hh signaling during the mouse fetal liver development. At E11.5, Shh and an activation marker gene for Hh signaling, Gli1, were expressed in Dlk{sup +} hepatoblasts, the fetal liver progenitor cells, and the expression was rapidly decreased thereafter as the development proceeded. In the culture of Dlk{sup +} hepatoblasts isolated from the E11.5 liver, activation of Hh signaling stimulated their proliferation and this effect was cancelled by a chemical Hh signaling inhibitor, cyclopamine. In contrast, hepatocyte differentiation of Dlk{sup +} hepatoblasts in vitro as manifested by the marker gene expression and acquisition of ammonia clearance activity was significantly inhibited by forced activation of Hh signaling. Taken together, these results demonstrate the temporally restricted manner of Hh signal activation and its role in promoting the hepatoblast proliferation, and further suggest that the pathway needs to be shut off for the subsequent hepatic differentiation of hepatoblasts to proceed normally.

  2. Hedgehog Signaling Modulates the Release of Gliotransmitters from Cultured Cerebellar Astrocytes.

    PubMed

    Okuda, Hiroaki; Tatsumi, Kouko; Morita-Takemura, Shoko; Nakahara, Kazuki; Nochioka, Katsunori; Shinjo, Takeaki; Terada, Yuki; Wanaka, Akio

    2016-02-01

    Sonic hedgehog (Shh), a member of the Hedgehog (Hh) family, plays essential roles in the development of the central nervous system. Recent studies suggest that the Hh signaling pathway also functions in mature astrocytes under physiological conditions. We first examined the expression of genes encoding Hh signaling molecules in the adult mouse cerebellum by in situ hybridization histochemistry. mRNA for Patched homolog 1 (Ptch1), a receptor for Hh family members, was expressed in S100β-positive astrocytes and Shh mRNA was expressed in HuC/D-positive neurons, implying that the Hh signaling pathway contributes to neuro-glial interactions. To test this hypothesis, we next examined the effects of recombinant SHH N-terminal protein (rSHH-N) on the functions of cultured cerebellar astrocytes. rSHH-N up-regulated Hh signal target genes such as Ptch1 and Gli-1, a key transcription factor of the Hh signaling pathway. Although activation of Hh signaling by rSHH-N or purmorphamine influenced neither glutamate uptake nor gliotransmitters release, inhibition of the Hh signaling pathway by cyclopamine, neutralizing antibody against SHH or intracellular Ca(2+) chelation decreased glutamate and ATP release from cultured cerebellar astrocytes. On the other hand, cyclopamine, neutralizing antibody against SHH or Ca(2+) chelator hardly affected D-serine secretion. Various kinase inhibitors attenuated glutamate and ATP release, while only U0126 reduced D-serine secretion from the astrocytes. These results suggested that the Hh signaling pathway sustains the release of glutamate and ATP and participates in neuro-glial interactions in the adult mouse brain. We also propose that signaling pathways distinct from the Hh pathway govern D-serine secretion from adult cerebellar astrocytes. PMID:26694649

  3. The Hedgehog Signal Transduction Network

    PubMed Central

    Robbins, David J.; Fei, Dennis Liang; Riobo, Natalia A.

    2013-01-01

    Hedgehog (Hh) proteins regulate the development of a wide range of metazoan embryonic and adult structures, and disruption of Hh signaling pathways results in various human diseases. Here, we provide a comprehensive review of the signaling pathways regulated by Hh, consolidating data from a diverse array of organisms in a variety of scientific disciplines. Similar to the elucidation of many other signaling pathways, our knowledge of Hh signaling developed in a sequential manner centered on its earliest discoveries. Thus, our knowledge of Hh signaling has for the most part focused on elucidating the mechanism by which Hh regulates the Gli family of transcription factors, the so-called “canonical” Hh signaling pathway. However, in the past few years, numerous studies have shown that Hh proteins can also signal through Gli-independent mechanisms collectively referred to as “noncanonical” signaling pathways. Noncanonical Hh signaling is itself subdivided into two distinct signaling modules: (i) those not requiring Smoothened (Smo) and (ii) those downstream of Smo that do not require Gli transcription factors. Thus, Hh signaling is now proposed to occur through a variety of distinct context-dependent signaling modules that have the ability to crosstalk with one another to form an interacting, dynamic Hh signaling network. PMID:23074268

  4. A Hedgehog Survival Pathway in ‘Undead’ Lipotoxic Hepatocytes

    PubMed Central

    Kakisaka, Keisuke; Cazanave, Sophie C.; Werneburg, Nathan W.; Razumilava, Nataliya; Mertens, Joachim C.; Bronk, Steve F.; Gores, Gregory J.

    2012-01-01

    Background & Aims Ballooned hepatocytes in nonalcoholic steatohepatitis (NASH) generate sonic hedgehog (SHH). This observation is consistent with a cellular phenotype in which the cell death program has been initiated but cannot be executed. Our aim was to determine if ballooned hepatocytes have potentially disabled the cell death execution machinery, and if so, can their functional biology be modeled in vitro. Methods Immunohistochemistry was performed on human NASH specimens. In vitro studies were performed using Huh-7 cells with shRNA targeted knockdown of caspase 9 (shC9 cells) or primary hepatocytes from caspase 3−/− mice. Results Ballooned hepatocytes in NASH display diminished expression of the caspase 9. This phenotype was modeled using shC9 cells; these cells were resistant to lipoapoptosis by palmitate (PA) or lysophosphatidylcholine (LPC) despite lipid droplet formation. During lipid loading by either PA or LPC, shC9 cells activate JNK which via AP-1 induces SHH expression. An autocrine hedgehog survival signaling pathway was further delineated in both shC9 and caspase 3−/− cells during lipotoxic stress. Conclusion Ballooned hepatocytes in NASH downregulate caspase 9, a pivotal caspase executing the mitochondrial pathway of apoptosis. Hepatocytes engineered to reduce caspase 9 expression are resistant to lipoapoptosis, in part, due to a hedgehog autocrine survival signaling pathway. PMID:22641094

  5. Ultrasonic/Sonic Jackhammer

    NASA Technical Reports Server (NTRS)

    Bar-Cohen, Yoseph; Sherrit, Stewart; Herz, Jack

    2005-01-01

    An ultrasonic/sonic jackhammer (USJ) is the latest in a series of related devices. Each of these devices cuts into a brittle material by means of hammering and chiseling actions of a tool bit excited with a combination of ultrasonic and sonic vibrations. A small-scale prototype of the USJ has been demonstrated. A fully developed, full-scale version of the USJ would be used for cutting through concrete, rocks, hard asphalt, and other materials to which conventional pneumatic jackhammers are applied, but the USJ would offer several advantages over conventional pneumatic jackhammers.

  6. Sonic levitation apparatus

    NASA Technical Reports Server (NTRS)

    Dunn, S. A.; Pomplum, A. R.; Paquette, E. G.; Ethridge, E. C.; Johnson, J. L. (Inventor)

    1984-01-01

    A sonic levitation apparatus is disclosed which includes a sonic transducer which generates acoustical energy responsive to the level of an electrical amplifier. A duct communicates with an acoustical chamber to deliver an oscillatory motion of air to a plenum section which contains a collimated hole structure having a plurality of parallel orifices. The collimated hole structure converts the motion of the air to a pulsed. Unidirectional stream providing enough force to levitate a material specimen. Particular application to the production of microballoons in low gravity environment is discussed.

  7. Hedgehog signaling in the normal and neoplastic mammary gland.

    PubMed

    Visbal, Adriana P; Lewis, Michael T

    2010-09-01

    The hedgehog signal transduction network is a critical regulator of metazoan development. Inappropriate activation of this network is implicated in several different cancers, including breast. Genetic evidence in mice as well as molecular biological studies in human cells clearly indicate that activated signaling can lead to mammary hyperplasia and, in some cases, tumor formation. However, the exact role(s) activated hedgehog signaling plays in the development or progression of breast cancer also remain unclear. In this review, we have discussed recent data regarding the mechanism(s) by which the hedgehog network may signal in the mammary gland, as well as the data implicating activated signaling as a contributing factor to breast cancer development. Finally, we provide a brief update on the available hedgehog signaling inhibitors with respect to ongoing clinical trials, some of which will include locally advanced or metastatic breast cancers. Given the growing intensity with which the hedgehog signaling network is being studied in the normal and neoplastic mammary gland, a more complete understanding of this network should allow more effective targeting of its activities in breast cancer treatment or prevention.

  8. Deciphering the role of hedgehog signaling in pancreatic cancer

    PubMed Central

    Gu, Dongsheng; Schlotman, Kelly E; Xie, Jingwu

    2016-01-01

    Abstract Pancreatic cancer, mostly pancreatic ductal adenocarcinoma (PDAC), is a leading cause of cancer-related death in the US, with a dismal median survival of 6 months. Thus, there is an urgent unmet need to identify ways to diagnose and to treat this deadly cancer. Although a number of genetic changes have been identified in pancreatic cancer, their mechanisms of action in tumor development, progression and metastasis are not completely understood. Hedgehog signaling, which plays a major role in embryonic development and stem cell regulation, is known to be activated in pancreatic cancer; however, specific inhibitors targeting the smoothened molecule failed to improve the condition of pancreatic cancer patients in clinical trials. Furthermore, results regarding the role of Hh signaling in pancreatic cancer are controversial with some reporting tumor promoting activities whereas others tumor suppressive actions. In this review, we will summarize what we know about hedgehog signaling in pancreatic cancer, and try to explain the contradicting roles of hedgehog signaling as well as the reason(s) behind the failed clinical trials. In addition to the canonical hedgehog signaling, we will also discuss several non-canonical hedgehog signaling mechanisms. PMID:27346466

  9. Piperazic acid derivatives inhibit Gli1 in Hedgehog signaling pathway.

    PubMed

    Khatra, Harleen; Kundu, Jayanta; Khan, Pragya Paramita; Duttagupta, Indranil; Pattanayak, Sankha; Sinha, Surajit

    2016-09-15

    Piperazic acid, a non-proteinogenic amino acid, found in complex secondary metabolites and peptide natural substances, has shown down regulation of Gli1 expression in Hedgehog signaling pathway in cell based assays. Further structure activity relationship study indicated that amide derivatives of piperazic acid are more potent than piperazic acid itself, with little to no toxicity. However, other cellular components involved in the pathway were not affected. To the best of our knowledge, this is the first report on the inhibitory property of piperazic acid in this pathway. Hence, this molecule could serve as a useful tool for studying Hedgehog signaling. PMID:27528433

  10. Homeodomains, Hedgehogs, and Happiness.

    PubMed

    Scott, Matthew P

    2016-01-01

    Developmental biologists have had a spectacular quarter century of discoveries, building on many decades of work earlier, discovering molecular, cellular, and genetic mechanisms that underlie the magical process by which an egg becomes a plant or animal. Among the discoveries were homeodomains, DNA-binding domains that allow transcription factors to recognize their target genes, and the Hedgehog signaling pathway, which is used in many organs and tissues for communication among cells. The experience of unveiling the mechanisms and molecules connected to both of these findings has been remarkable, joyful, difficult, and a time of great teamwork and collaboration within and between laboratory groups. More than ever it is possible to discern the evolutionary processes, and their mechanisms, that led to the diversity of life on earth. A huge amount of work remains to be done to obtain a broad understanding of what happened and how development works.

  11. Homeodomains, Hedgehogs, and Happiness.

    PubMed

    Scott, Matthew P

    2016-01-01

    Developmental biologists have had a spectacular quarter century of discoveries, building on many decades of work earlier, discovering molecular, cellular, and genetic mechanisms that underlie the magical process by which an egg becomes a plant or animal. Among the discoveries were homeodomains, DNA-binding domains that allow transcription factors to recognize their target genes, and the Hedgehog signaling pathway, which is used in many organs and tissues for communication among cells. The experience of unveiling the mechanisms and molecules connected to both of these findings has been remarkable, joyful, difficult, and a time of great teamwork and collaboration within and between laboratory groups. More than ever it is possible to discern the evolutionary processes, and their mechanisms, that led to the diversity of life on earth. A huge amount of work remains to be done to obtain a broad understanding of what happened and how development works. PMID:26969987

  12. The Hedgehog protein family.

    PubMed

    Bürglin, Thomas R

    2008-01-01

    The Hedgehog (Hh) pathway is one of the fundamental signal transduction pathways in animal development and is also involved in stem-cell maintenance and carcinogenesis. The hedgehog (hh) gene was first discovered in Drosophila, and members of the family have since been found in most metazoa. Hh proteins are composed of two domains, an amino-terminal domain HhN, which has the biological signal activity, and a carboxy-terminal autocatalytic domain HhC, which cleaves Hh into two parts in an intramolecular reaction and adds a cholesterol moiety to HhN. HhC has sequence similarity to the self-splicing inteins, and the shared region is termed Hint. New classes of proteins containing the Hint domain have been discovered recently in bacteria and eukaryotes, and the Hog class, of which Hh proteins comprise one family, is widespread throughout eukaryotes. The non-Hh Hog proteins have carboxy-terminal domains (the Hog domain) highly similar to HhC, although they lack the HhN domain, and instead have other amino-terminal domains. Hog proteins are found in many protists, but the Hh family emerged only in early metazoan evolution. HhN is modified by cholesterol at its carboxyl terminus and by palmitate at its amino terminus in both flies and mammals. The modified HhN is released from the cell and travels through the extracellular space. On binding its receptor Patched, it relieves the inhibition that Patched exerts on Smoothened, a G-protein-coupled receptor. The resulting signaling cascade converges on the transcription factor Cubitus interruptus (Ci), or its mammalian counterparts, the Gli proteins, which activate or repress target genes.

  13. A sesquiterpene lactone from Siegesbeckia glabrescens suppresses Hedgehog/Gli-mediated transcription in pancreatic cancer cells

    PubMed Central

    Lee, Hwa Jin; Wu, Qian; Li, Hua; Bae, Gyu-Un; Kim, An Keun; Ryu, Jae-Ha

    2016-01-01

    Pancreatic cancer is aggressive and therefore difficult to treat; however, continued efforts have been made with the aim of developing an effective therapy against the disease. The Hedgehog (Hh) signaling pathway is reportedly involved in the proliferation and survival of pancreatic cancer cells. The transcription factor glioma-associated oncogene (Gli) is a key component of the Hh signaling pathway and the primary effector of pancreatic cancer development. Inhibiting Gli is a proven therapeutic strategy for this disease. The present study examined the regulation of Gli and the expression of its target genes to identify an inhibitor of the Sonic Hh (Shh) pathway. A germacranolide sesquiterpene lactone (GSL) was isolated from Siegesbeckia glabrescens as an inhibitor of Gli-mediated transcription. The results demonstrated that GSL inhibited Shh-induced osteoblast differentiation and Gli homolog 1 (Gli1)-mediated transcriptional activity in mesenchymal C3H10T1/2 stem cells. Furthermore, GSL suppressed Gli-mediated transcriptional activity in human pancreatic cancer PANC-1 and AsPC-1 cells, which resulted in reduced cancer cell proliferation and downregulated expression of the Gli-target genes, Gli1 and cyclin D1. A sesquiterpene lactone from S. glabrescens may therefore serve as a candidate for the treatment of Hh/Gli-dependent pancreatic cancer.

  14. Evolution of the Hedgehog Gene Family

    PubMed Central

    Kumar, S.; Balczarek, K. A.; Lai, Z. C.

    1996-01-01

    Effective intercellular communication is an important feature in the development of multicellular organisms. Secreted hedgehog (hh) protein is essential for both long- and short-range cellular signaling required for body pattern formation in animals. In a molecular evolutionary study, we find that the vertebrate homologs of the Drosophila hh gene arose by two gene duplications: the first gave rise to Desert hh, whereas the second produced the Indian and Sonic hh genes. Both duplications occurred before the emergence of vertebrates and probably before the evolution of chordates. The amino-terminal fragment of the hh precursor, crucial in long- and short-range intercellular communication, evolves two to four times slower than the carboxyl-terminal fragment in both Drosophila hh and its vertebrate homologues, suggesting conservation of mechanism of hh action in animals. A majority of amino acid substitutions in the amino- and carboxyl-terminal fragments are conservative, but the carboxyl-terminal domain has undergone extensive insertion-deletion events while maintaining its autocleavage protease activity. Our results point to similarity of evolutionary constraints among sites of Drosophila and vertebrate hh homologs and suggest some future directions for understanding the role of hh genes in the evolution of developmental complexity in animals. PMID:8849902

  15. Pilot Clinical Trial of Hedgehog Pathway Inhibitor GDC-0449 (Vismodegib) in Combination with Gemcitabine in Patients with Metastatic Pancreatic Adenocarcinoma

    PubMed Central

    Abel, Ethan V.; Griffith, Kent A.; Greenson, Joel K.; Takebe, Naoko; Khan, Gazala N.; Blau, John L.; Craig, Ronald; Balis, Ulysses G.; Zalupski, Mark M.; Simeone, Diane M.

    2014-01-01

    Background The hedgehog (HH) signaling pathway is a key regulator in tumorigenesis of pancreatic adenocarcinoma (PDA) and is up-regulated in PDA cancer stem cells (CSCs). GDC-0449 is an oral small-molecule inhibitor of HH pathway. This study assessed the effect of GDC-0449-mediated HH inhibition in paired biopsies, followed by combined treatment with gemcitabine, in patients with metastatic PDA. Methods Twenty-five patients were enrolled of which 23 underwent core biopsies at baseline and following 3 weeks of GDC-0449. On day 29, 23 patients started weekly gemcitabine while continuing GDC-0449. We evaluated GLI1 and PTCH1 inhibition, change in CSCs, Ki-67, fibrosis, and assessed tumor response, survival and toxicity. Results On pre-treatment biopsy, 75% of patients had elevated sonic hedgehog (SHH) expression. On post-treatment biopsy, GLI1 and PTCH1 decreased in 95.6% and 82.6% of 23 patients, fibrosis decreased in 45.4% of 22 and Ki-67 in 52.9% of 17 evaluable patients. No significant changes were detected in CSCs pre- and post-biopsy. The median progression-free and overall survival for all treated patients was 2.8 and 5.3 months. The response and disease control rate was 21.7% and 65.2%. No significant correlation was noted between CSCs, fibrosis, SHH, Ki-67, GLI1, PTCH1 (baseline values, or relative change on post-treatment biopsy) and survival. Grade >3 adverse events were noted in 56% of patients. Conclusion We show that GDC-0449 for 3 weeks leads to down-modulation of GLI1 and PTCH1, without significant changes in CSCs compared to baseline. GDC-0449 and gemcitabine was not superior to gemcitabine alone in the treatment of metastatic pancreatic cancer. PMID:25278454

  16. Ultrasonic/Sonic Jackhammer

    NASA Technical Reports Server (NTRS)

    Bar-Cohen, Yoseph (Inventor); Sherrit, Stewart (Inventor); Herz, Jack L. (Inventor)

    2014-01-01

    The invention provides a novel jackhammer that utilizes ultrasonic and/or sonic vibrations as source of power. It is easy to operate and does not require extensive training, requiring substantially less physical capabilities from the user and thereby increasing the pool of potential operators. An important safety benefit is that it does not fracture resilient or compliant materials such as cable channels and conduits, tubing, plumbing, cabling and other embedded fixtures that may be encountered along the impact path. While the ultrasonic/sonic jackhammer of the invention is able to cut concrete and asphalt, it generates little back-propagated shocks or vibrations onto the mounting fixture, and can be operated from an automatic platform or robotic system. PNEUMATICS; ULTRASONICS; IMPACTORS; DRILLING; HAMMERS BRITTLE MATERIALS; DRILL BITS; PROTOTYPES; VIBRATION

  17. Hedgehog signaling is synergistically enhanced by nutritional deprivation and ligand stimulation in human fibroblasts of Gorlin syndrome.

    PubMed

    Mizuochi, Hiromi; Fujii, Katsunori; Shiohama, Tadashi; Uchikawa, Hideki; Shimojo, Naoki

    2015-02-13

    Hedgehog signaling is a pivotal developmental pathway that comprises hedgehog, PTCH1, SMO, and GLI proteins. Mutations in PTCH1 are responsible for Gorlin syndrome, which is characterized by developmental defects and tumorigenicity. Although the hedgehog pathway has been investigated extensively in Drosophila and mice, its functional roles have not yet been determined in human cells. In order to elucidate the mechanism by which transduction of the hedgehog signal is regulated in human tissues, we employed human fibroblasts derived from three Gorlin syndrome patients and normal controls. We investigated GLI1 transcription, downstream of hedgehog signaling, to assess native signal transduction, and then treated fibroblasts with a recombinant human hedgehog protein with or without serum deprivation. We also examined the transcriptional levels of hedgehog-related genes under these conditions. The expression of GLI1 mRNA was significantly higher in Gorlin syndrome-derived fibroblasts than in control cells. Hedgehog stimulation and nutritional deprivation synergistically enhanced GLI1 transcription levels, and this was blocked more efficiently by vismodegib, a SMO inhibitor, than by the natural compound, cyclopamine. Messenger RNA profiling revealed the increased expression of Wnt signaling and morphogenetic molecules in these fibroblasts. These results indicated that the hedgehog stimulation and nutritional deprivation synergistically activated the hedgehog signaling pathway in Gorlin syndrome fibroblasts, and this was associated with increments in the transcription levels of hedgehog-related genes such as those involved in Wnt signaling. These fibroblasts may become a significant tool for predicting the efficacies of hedgehog molecular-targeted therapies such as vismodegib. PMID:25576868

  18. Genetic Regulation of Prostate Development

    PubMed Central

    Meeks, Joshua; Schaeffer, Edward M

    2011-01-01

    Prostatic development is a dynamic process in which basic mechanisms of epithelial outgrowth and epithelial-mesenchymal interaction are initiated by androgens and androgen receptor signaling. Even in adulthood, the prostate's function remains tightly regulated by androgens--without them, pathologic diseases including hyperplastic and malignant growth which together plague nearly 50% of aging males does not occur. Unraveling the etiology of these pathologic processes is a complex and important goal. In fact, many insights into these processes have come from an intimate understanding of the complex signaling networks that regulate physiologic prostatic growth in development. This review aims to highlight important key molecules such as Nkx3.1, sonic hedgehog and Sox9 as well as key signaling pathways including the Fibroblast growth factor and Wnt pathways. These molecules and pathways are critical for prostate development with both know and postulated roles in prostatic pathology. PMID:20930191

  19. The role of glypicans in Hedgehog signaling.

    PubMed

    Filmus, Jorge; Capurro, Mariana

    2014-04-01

    Glypicans (GPCs) are a family of proteoglycans that are bound to the cell surface by a glycosylphosphatidylinositol anchor. Six glypicans have been found in the mammalian genome (GPC1 to GPC6). GPCs regulate several signaling pathways, including the pathway triggered by Hedgehogs (Hhs). This regulation, which could be stimulatory or inhibitory, occurs at the signal reception level. In addition, GPCs have been shown to be involved in the formation of Hh gradients in the imaginal wing disks in Drosophila. In this review we will discuss the role of various glypicans in specific developmental events in the embryo that are regulated by Hh signaling. In addition, we will discuss the mechanism by which loss-of-function GPC3 mutations alter Hh signaling in the Simpson-Golabi-Behmel overgrowth syndrome, and the molecular basis of the GPC5-induced stimulation of Hh signaling and tumor progression in rhabdomyosarcomas. PMID:24412155

  20. Hedgehog signaling stimulates the formation of proliferating Müller glia-derived progenitor cells in the chick retina.

    PubMed

    Todd, Levi; Fischer, Andy J

    2015-08-01

    Müller glia can be stimulated to de-differentiate and become proliferating progenitor cells that regenerate neurons in the retina. The signaling pathways that regulate the formation of proliferating Müller glia-derived progenitor cells (MGPCs) are beginning to be revealed. The purpose of this study was to investigate whether Hedgehog (Hh) signaling influences the formation of MGPCs in the chick retina. We find that Hh signaling is increased in damaged retinas where MGPCs are known to form. Sonic Hedgehog (Shh) is normally present in the axons of ganglion cells, but becomes associated with Müller glia and MGPCs following retinal damage. Activation of Hh signaling with recombinant human SHH (rhShh) or smoothened agonist (SAG) increased levels of Ptch1, Gli1, Gli2, Gli3, Hes1 and Hes5, and stimulated the formation of proliferating MGPCs in damaged retinas. In undamaged retinas, SAG or rhShh had no apparent effect upon the Müller glia. However, SAG combined with FGF2 potentiated the formation of MGPCs, whereas SAG combined with IGF1 stimulated the nuclear migration of Müller glia, but not the formation of MGPCs. Conversely, inhibition of Hh signaling with KAAD-cyclopamine, Gli antagonists or antibody to Shh reduced numbers of proliferating MGPCs in damaged and FGF2-treated retinas. Hh signaling potentiates Pax6, Klf4 and cFos expression in Müller glia during the formation of MGPCs. We find that FGF2/MAPK signaling recruits Hh signaling into the signaling network that drives the formation of proliferating MGPCs. Our findings implicate Hh signaling as a key component of the network of signaling pathways that promote the de-differentiation of Müller glia and proliferation of MGPCs. PMID:26116667

  1. Calculating Sonic-Boom Propagation

    NASA Technical Reports Server (NTRS)

    Darden, C. M.; Ting, L.

    1987-01-01

    Nonlinear effects included, enabling more-realistic modeling. Modified Method of Characteristics Sonic Boom Extrapolation Program (MMOC) is computer program for sonic-boom propagation that includes shock coalescence and incorporates effects of asymmetries due to volume and lift. Numerically integrates nonlinear governing equations using data on initial data line approximately one body length from aircraft and yields sonic-boom pressure at ground as function of time or of position at given time. MMOC written in FORTRAN IV.

  2. Sonic Boom Modeling Technical Challenge

    NASA Technical Reports Server (NTRS)

    Sullivan, Brenda M.

    2007-01-01

    This viewgraph presentation reviews the technical challenges in modeling sonic booms. The goal of this program is to develop knowledge, capabilities and technologies to enable overland supersonic flight. The specific objectives of the modeling are: (1) Develop and validate sonic boom propagation model through realistic atmospheres, including effects of turbulence (2) Develop methods enabling prediction of response of and acoustic transmission into structures impacted by sonic booms (3) Develop and validate psychoacoustic model of human response to sonic booms under both indoor and outdoor listening conditions, using simulators.

  3. Sonication standard laboratory module

    DOEpatents

    Beugelsdijk, Tony; Hollen, Robert M.; Erkkila, Tracy H.; Bronisz, Lawrence E.; Roybal, Jeffrey E.; Clark, Michael Leon

    1999-01-01

    A standard laboratory module for automatically producing a solution of cominants from a soil sample. A sonication tip agitates a solution containing the soil sample in a beaker while a stepper motor rotates the sample. An aspirator tube, connected to a vacuum, draws the upper layer of solution from the beaker through a filter and into another beaker. This beaker can thereafter be removed for analysis of the solution. The standard laboratory module encloses an embedded controller providing process control, status feedback information and maintenance procedures for the equipment and operations within the standard laboratory module.

  4. Distinct roles of PTCH2 splice variants in Hedgehog signalling.

    PubMed Central

    Rahnama, Fahimeh; Toftgård, Rune; Zaphiropoulos, Peter G

    2004-01-01

    The human PTCH2 gene is highly similar to PTCH1, a tumour suppressor gene frequently mutated in basal cell carcinoma and several other tumour types. PTCH1 is a transmembrane protein believed to inhibit another transmembrane protein SMO (Smoothened), which mediates HH (Hedgehog) signalling. In this study, we analysed the biological properties of several PTCH2 splice variants. An mRNA form that lacked the last exon was abundantly expressed in all tissues examined, in contrast with the one that included it. Moreover, a transcript lacking exon 9, which is a part of a conserved sterol-sensing domain, was identified in intestine, prostate and cerebellum. In ovary, spleen, testis, cerebellum and skin, an mRNA lacking both exons 9 and 10 could also be observed. The different PTCH2 isoforms localized in the cytoplasm were capable of internalizing the N-terminal fragment of Sonic HH (Shh-N). Additionally, the PTCH2 gene was found to be a target of HH signalling. PTCH2 promoter regulation assays demonstrated that only one of the PTCH2 variants could inhibit the activity of SHH-N, whereas none was capable of inhibiting the activated form of SMO (SMO-M2) and this contrasts with PTCH1. Despite the fact that the PTCH2 isoforms lacked the ability to inhibit SMO-M2 activity, all PTCH2 variants as well as PTCH1, on co-transfection with Smo, were able to change Smo localization from being largely dispersed in the cytoplasm to the juxtanuclear region. Furthermore, the PTCH2 isoforms and PTCH1 co-localized in doubly transfected cells and an interaction between them was confirmed using immunoprecipitation assays. Using Ptch1-/- mouse cells, it was shown that the PTCH2 variants and PTCH1 differentially act to reconstitute not only the SHH but also the Desert HH-dependent transcriptional response. We conclude that in spite of their structural similarities, the PTCH2 isoforms have distinct functional properties when compared with PTCH1. PMID:14613484

  5. Hedgehog Signaling during Appendage Development and Regeneration.

    PubMed

    Singh, Bhairab N; Koyano-Nakagawa, Naoko; Donaldson, Andrew; Weaver, Cyprian V; Garry, Mary G; Garry, Daniel J

    2015-01-01

    Regulatory networks that govern embryonic development have been well defined. While a common hypothesis supports the notion that the embryonic regulatory cascades are reexpressed following injury and tissue regeneration, the mechanistic regulatory pathways that mediate the regenerative response in higher organisms remain undefined. Relative to mammals, lower vertebrates, including zebrafish and newts, have a tremendous regenerative capacity to repair and regenerate a number of organs including: appendages, retina, heart, jaw and nervous system. Elucidation of the pathways that govern regeneration in these lower organisms may provide cues that will enhance the capacity for the regeneration of mammalian organs. Signaling pathways, such as the hedgehog pathway, have been shown to play critical functions during development and during regeneration in lower organisms. These signaling pathways have been shown to modulate multiple processes including cellular origin, positional identity and cellular maturation. The present review will focus on the cellular and molecular regulation of the hedgehog (HH) signaling pathway and its interaction with other signaling factors during appendage development and regeneration. PMID:26110318

  6. Hedgehog Signaling during Appendage Development and Regeneration

    PubMed Central

    Singh, Bhairab N.; Koyano-Nakagawa, Naoko; Donaldson, Andrew; Weaver, Cyprian V.; Garry, Mary G.; Garry, Daniel J.

    2015-01-01

    Regulatory networks that govern embryonic development have been well defined. While a common hypothesis supports the notion that the embryonic regulatory cascades are reexpressed following injury and tissue regeneration, the mechanistic regulatory pathways that mediate the regenerative response in higher organisms remain undefined. Relative to mammals, lower vertebrates, including zebrafish and newts, have a tremendous regenerative capacity to repair and regenerate a number of organs including: appendages, retina, heart, jaw and nervous system. Elucidation of the pathways that govern regeneration in these lower organisms may provide cues that will enhance the capacity for the regeneration of mammalian organs. Signaling pathways, such as the hedgehog pathway, have been shown to play critical functions during development and during regeneration in lower organisms. These signaling pathways have been shown to modulate multiple processes including cellular origin, positional identity and cellular maturation. The present review will focus on the cellular and molecular regulation of the hedgehog (HH) signaling pathway and its interaction with other signaling factors during appendage development and regeneration. PMID:26110318

  7. Protecting the hedgerow: p53 and hedgehog pathway interactions.

    PubMed

    Ho, Louisa; Alman, Benjamin

    2010-02-01

    A common environment for the Hedgehog (Subfamily: erinaceinae) is a row of shrubs and trees often used on farms for enclosing or separating fields, called a hedgerow. Maintenance of a continuous shrub border is important for shielding crops from weather damage, but also provides an ideal protective habitat for the hedgehog. Similar to its mammalian counterpart, the Hedgehog (Hh) signalling pathway requires a controlled environment to regulate proper functioning of the cell. When allowed to run wild, constitutive activation of the Hh pathway results in tumorigenesis in different tissues types, including brain, skin and cartilage. With an additional loss of p53 tumor suppressor activity, an increase in tumor incidence, size and metastasis have been observed. p53 has a number of functions that can suppress tumor formation and growth in most, if not all Hh-related cancers, such as the inhibition of cell cycle progression and cell survival. Furthermore, increasing evidence of an interaction between p53 and Hedgehog signalling pathways suggests a critical role for the tumor suppressor activity of p53 in "protecting the hedgerow".

  8. MicroRNA125b-mediated Hedgehog signaling influences liver regeneration by chorionic plate-derived mesenchymal stem cells.

    PubMed

    Hyun, Jeongeun; Wang, Sihyung; Kim, Jieun; Kim, Gi Jin; Jung, Youngmi

    2015-09-15

    Although chorionic plate-derived mesenchymal stem cells (CP-MSCs) were shown to promote liver regeneration, the mechanisms underlying the effect remain unclear. Hedgehog (Hh) signaling orchestrates tissue reconstruction in damaged liver. MSCs release microRNAs mediating various cellular responses. Hence, we hypothesized that microRNAs from CP-MSCs regulated Hh signaling, which influenced liver regeneration. Livers were obtained from carbon tetrachloride (CCl4)-treated rats transplanted with human CP-MSCs (Tx) or saline (non-Tx). Sonic Hh, one of Hh ligands, increased in CCl4-treated liver, whereas it decreased in CP-MSC-treated liver with CCl4. The expression of Hh-target genes was significantly downregulated in the Tx. Reduced expansion of progenitors and regressed fibrosis were observed in the liver of the Tx rats. CP-MSCs suppressed the expression of Hh and profibrotic genes in co-cultured LX2 (human hepatic stellate cell) with CP-MSCs. MicroRNA-125b targeting smo was retained in exosomes of CP-MSCs. CP-MSCs with microRNA-125b inhibitor failed to attenuate the expression of Hh signaling and profibrotic genes in the activated HSCs. Therefore, these results demonstrated that microRNA-125b from CP-MSCs suppressed the activation of Hh signaling, which promoted the reduced fibrosis, suggesting that microRNA-mediated regulation of Hh signaling contributed to liver regeneration by CP-MSCs.

  9. Arsenic inhibits hedgehog signaling during P19 cell differentiation

    SciTech Connect

    Liu, Jui Tung; Bain, Lisa J.

    2014-12-15

    Arsenic is a toxicant found in ground water around the world, and human exposure mainly comes from drinking water or from crops grown in areas containing arsenic in soils or water. Epidemiological studies have shown that arsenic exposure during development decreased intellectual function, reduced birth weight, and altered locomotor activity, while in vitro studies have shown that arsenite decreased muscle and neuronal cell differentiation. The sonic hedgehog (Shh) signaling pathway plays an important role during the differentiation of both neurons and skeletal muscle. The purpose of this study was to investigate whether arsenic can disrupt Shh signaling in P19 mouse embryonic stem cells, leading to changes muscle and neuronal cell differentiation. P19 embryonic stem cells were exposed to 0, 0.25, or 0.5 μM of sodium arsenite for up to 9 days during cell differentiation. We found that arsenite exposure significantly reduced transcript levels of genes in the Shh pathway in both a time and dose-dependent manner. This included the Shh ligand, which was decreased 2- to 3-fold, the Gli2 transcription factor, which was decreased 2- to 3-fold, and its downstream target gene Ascl1, which was decreased 5-fold. GLI2 protein levels and transcriptional activity were also reduced. However, arsenic did not alter GLI2 primary cilium accumulation or nuclear translocation. Moreover, additional extracellular SHH rescued the inhibitory effects of arsenic on cellular differentiation due to an increase in GLI binding activity. Taken together, we conclude that arsenic exposure affected Shh signaling, ultimately decreasing the expression of the Gli2 transcription factor. These results suggest a mechanism by which arsenic disrupts cell differentiation. - Highlights: • Arsenic exposure decreases sonic hedgehog pathway-related gene expression. • Arsenic decreases GLI2 protein levels and transcriptional activity in P19 cells. • Arsenic exposure does not alter the levels of SHH

  10. Portable Sonic Boom Simulation

    NASA Astrophysics Data System (ADS)

    Salamone, Joe

    2006-05-01

    A method is presented to simulate sonic booms using high fidelity and custom-built audio equipment that output to an acoustically treated listening environment, all of which is contained in a portable vehicle. The audio system has inherent low and high frequency performance limitations and also introduces distortion due to the frequency response of the system. The limitations of the system are compensated for by band-pass filtering a full-fidelity sonic boom signature and applying a system equalization filter. The purpose of the band-pass filter is to remove frequency content above and below the capabilities of the system yet retain the audible and felt characteristics of the full-fidelity waveform. The equalization filter, computed from time-domain Wiener filtering, compensates for the frequency-dependent system response of the audio system at several listening positions. The system performance is evaluated by comparing the PLdB, SEL(A) and SEL(C) of the measured system output to the full-fidelity waveform. Results show good agreement between the loudness levels of the full-fidelity waveform and the corresponding measured system output.

  11. Characterization of two patched receptors for the vertebrate hedgehog protein family

    PubMed Central

    Carpenter, David; Stone, Donna M.; Brush, Jennifer; Ryan, Anne; Armanini, Mark; Frantz, Gretchen; Rosenthal, Arnon; de Sauvage, Frederic J.

    1998-01-01

    The multitransmembrane protein Patched (PTCH) is the receptor for Sonic Hedgehog (Shh), a secreted molecule implicated in the formation of embryonic structures and in tumorigenesis. Current models suggest that binding of Shh to PTCH prevents the normal inhibition of the seven-transmembrane-protein Smoothened (SMO) by PTCH. According to this model, the inhibition of SMO signaling is relieved after mutational inactivation of PTCH in the basal cell nevus syndrome. Recently, PTCH2, a molecule with sequence homology to PTCH, has been identified. To characterize both PTCH molecules with respect to the various Hedgehog proteins, we have isolated the human PTCH2 gene. Biochemical analysis of PTCH and PTCH2 shows that they both bind to all hedgehog family members with similar affinity and that they can form a complex with SMO. However, the expression patterns of PTCH and PTCH2 do not fully overlap. While PTCH is expressed throughout the mouse embryo, PTCH2 is found at high levels in the skin and in spermatocytes. Because Desert Hedgehog (Dhh) is expressed specifically in the testis and is required for germ cell development, it is likely that PTCH2 mediates its activity in vivo. Chromosomal localization of PTCH2 places it on chromosome 1p33–34, a region deleted in some germ cell tumors, raising the possibility that PTCH2 may be a tumor suppressor in Dhh target cells. PMID:9811851

  12. Activation of Hedgehog signaling by the environmental toxicant arsenic may contribute to the etiology of arsenic-induced tumors.

    PubMed

    Fei, Dennis Liang; Li, Hua; Kozul, Courtney D; Black, Kendall E; Singh, Samer; Gosse, Julie A; DiRenzo, James; Martin, Kathleen A; Wang, Baolin; Hamilton, Joshua W; Karagas, Margaret R; Robbins, David J

    2010-03-01

    Exposure to the environmental toxicant arsenic, through both contaminated water and food, contributes to significant health problems worldwide. In particular, arsenic exposure is thought to function as a carcinogen for lung, skin, and bladder cancer via mechanisms that remain largely unknown. More recently, the Hedgehog signaling pathway has also been implicated in the progression and maintenance of these same cancers. Based on these similarities, we tested the hypothesis that arsenic may act in part through activating Hedgehog signaling. Here, we show that arsenic is able to activate Hedgehog signaling in several primary and established tissue culture cells as well as in vivo. Arsenic activates Hedgehog signaling by decreasing the stability of the repressor form of GLI3, one of the transcription factors that ultimately regulate Hedgehog activity. We also show, using tumor samples from a cohort of bladder cancer patients, that high levels of arsenic exposure are associated with high levels of Hedgehog activity. Given the important role Hedgehog signaling plays in the maintenance and progression of a variety of tumors, including bladder cancer, these results suggest that arsenic exposure may in part promote cancer through the activation of Hedgehog signaling. Thus, we provide an important insight into the etiology of arsenic-induced human carcinogenesis, which may be relevant to millions of people exposed to high levels of arsenic worldwide.

  13. A multilayer sonic film

    NASA Astrophysics Data System (ADS)

    Munteanu, L.; Chiroiu, V.; Sireteanu, T.; Dumitriu, D.

    2015-10-01

    A non-periodic multilayer film was analyzed to show that, despite its non-periodicity, the film exhibits full band-gaps and localized modes at its interfaces, as well as in the sonic composites. The film consists of alternating layers of two different materials that follow a triadic Cantor sequence. The Cantor structure shows extremely low thresholds for subharmonic generation of ultrasonic waves, compared with homogeneous and periodic structures. The coupling between the extended-mode (phonon) and the localized-mode (fracton) vibration regimes explains the generation of full band-gaps, for which there are no propagating Lamb waves. The large enhancement of the nonlinear interaction results from a more favorable frequency and spatial matching of coupled modes. A full band-gap that excludes Love waves is also analyzed.

  14. Quiet Sonic Booms: A NASA and Industry Progress Report

    NASA Technical Reports Server (NTRS)

    Larson, David Nils; Martin, Roy; Haering, Edward A.

    2011-01-01

    The purpose of this Oral Presentation is to present a progress report on NASA and Industry efforts related to Quiet Sonic Boom Program activities. This presentation will review changes in aircraft shaping to produce quiet supersonic booms and associated supersonic flight test methods and results. In addition, new flight test profiles have been recently developed that have allowed for the generation of sonic booms of varying intensity. These new flight test profiles have allowed for ground testing of the response of various building structures to sonic booms and the associated public acceptability to various sonic boom intensities. The new flight test profiles and associated ground measurement test methods will be reviewed. Finally, this Oral Presentation will review the International Regulatory requirements that would be involved to change aviation regulation and allow for overland quiet supersonic flight.

  15. Hedgehog and Resident Vascular Stem Cell Fate

    PubMed Central

    Mooney, Ciaran J.; Hakimjavadi, Roya; Fitzpatrick, Emma; Kennedy, Eimear; Walls, Dermot; Morrow, David; Redmond, Eileen M.; Cahill, Paul A.

    2015-01-01

    The Hedgehog pathway is a pivotal morphogenic driver during embryonic development and a key regulator of adult stem cell self-renewal. The discovery of resident multipotent vascular stem cells and adventitial progenitors within the vessel wall has transformed our understanding of the origin of medial and neointimal vascular smooth muscle cells (SMCs) during vessel repair in response to injury, lesion formation, and overall disease progression. This review highlights the importance of components of the Hh and Notch signalling pathways within the medial and adventitial regions of adult vessels, their recapitulation following vascular injury and disease progression, and their putative role in the maintenance and differentiation of resident vascular stem cells to vascular lineages from discrete niches within the vessel wall. PMID:26064136

  16. NASA Ames Sonic Boom Testing

    NASA Technical Reports Server (NTRS)

    Durston, Donald A.; Kmak, Francis J.

    2009-01-01

    Multiple sonic boom wind tunnel models were tested in the NASA Ames Research Center 9-by 7-Foot Supersonic Wind Tunnel to reestablish related test techniques in this facility. The goal of the testing was to acquire higher fidelity sonic boom signatures with instrumentation that is significantly more sensitive than that used during previous wind tunnel entries and to compare old and new data from established models. Another objective was to perform tunnel-to-tunnel comparisons of data from a Gulfstream sonic boom model tested at the NASA Langley Research Center 4-foot by 4-foot Unitary Plan Wind Tunnel.

  17. Sonic crystal acoustic switch device.

    PubMed

    Alagoz, Serkan; Alagoz, Baris Baykant

    2013-06-01

    This study reports a wave-controlled sonic crystal switch device that exhibits a destructive interference-based wave to wave reverse switching effect. By applying control waves, this acoustic device, composed of a two-dimensional square lattice sonic crystal block, reduces acoustic wave transmission from input to output. The finite difference time domain simulation and experimental results confirm the wave-to-wave reverse switching effect at the peak frequencies of the second band. The proposed sonic crystal switch prototype provides a contrast rate of 86% at 11.3 kHz frequency. This wave-to-wave switching effect is useful for controlling wave propagation for smart structure applications.

  18. Evolution of the vertebrate pth2 (tip39) gene family and the regulation of PTH type-2 Receptor (pth2r) and its endogenous ligand pth2 by Hedgehog signaling in zebrafish development

    PubMed Central

    Bhattacharya, Poulomi; Yan, Yi Lin; Postlethwait, John; Rubin, David A.

    2011-01-01

    In mammals, parathyroid hormone (PTH), secreted by parathyroid glands, increases calcium levels in the blood from reservoirs in bone. While mammals have two PTH receptor genes, PTH1R and PTH2R, zebrafish has three, pth1r, pth2r and pth3r. PTH can activate all three zebrafish Pthrs while PTH2 (alias tuberoinfundibular peptide 39, TIP39) preferentially activates zebrafish and mammalian PTH2Rs. We know little about the roles of the PTH2/PTH2R system in the development of any animal. To determine the roles of PTH2 and PTH2R during vertebrate development, we evaluated their expression patterns in developing zebrafish, observed their phylogenetic and conserved synteny relationships with humans, and described the genomic organization of pth2, pth2r, and pth2r splice variants. Expression studies showed that pth2 is expressed in cells adjacent to the ventral part of the posterior tuberculum in the diencephalon, whereas pth2r is robustly expressed throughout the CNS. Otic vesicles express both pth2 and pth2r, but heart expresses only pth2. Analysis of mutants showed that Hedgehog (Hh) signaling regulates the expression of pth2 transcripts more than that of nearby gnrh2-expressing cells. Genomic analysis showed that a lizard, chicken, and zebra finch lack a PTH2 gene, which is associated with an inversion breakpoint. Likewise, chickens lack PTH2R, while humans lack PTH3R, a case of reciprocally missing ohnologs (paralogs derived from a genome duplication). The considerable evolutionary conservation in genomic structure, synteny relationships, and expression of zebrafish pth2 and pth2r provides a foundation for exploring the endocrine roles of this system in developing vertebrate embryos. PMID:21880859

  19. The Sonic Altimeter for Aircraft

    NASA Technical Reports Server (NTRS)

    Draper, C S

    1937-01-01

    Discussed here are results already achieved with sonic altimeters in light of the theoretical possibilities of such instruments. From the information gained in this investigation, a procedure is outlined to determine whether or not a further development program is justified by the value of the sonic altimeter as an aircraft instrument. The information available in the literature is reviewed and condensed into a summary of sonic altimeter developments. Various methods of receiving the echo and timing the interval between the signal and the echo are considered. A theoretical discussion is given of sonic altimeter errors due to uncertainties in timing, variations in sound velocity, aircraft speed, location of the sending and receiving units, and inclinations of the flight path with respect to the ground surface. Plots are included which summarize the results in each case. An analysis is given of the effect of an inclined flight path on the frequency of the echo. A brief study of the acoustical phases of the sonic altimeter problem is carried through. The results of this analysis are used to predict approximately the maximum operating altitudes of a reasonably designed sonic altimeter under very good and very bad conditions. A final comparison is made between the estimated and experimental maximum operating altitudes which shows good agreement where quantitative information is available.

  20. Hedgehog Signaling in Pancreatic Fibrosis and Cancer

    PubMed Central

    Bai, Yongyu; Bai, Yongheng; Dong, Jiaojiao; Li, Qiang; Jin, Yuepeng; Chen, Bicheng; Zhou, Mengtao

    2016-01-01

    Abstract The hedgehog signaling pathway was first discovered in the 1980s. It is a stem cell-related pathway that plays a crucial role in embryonic development, tissue regeneration, and organogenesis. Aberrant activation of hedgehog signaling leads to pathological consequences, including a variety of human tumors such as pancreatic cancer. Multiple lines of evidence indicate that blockade of this pathway with several small-molecule inhibitors can inhibit the development of pancreatic neoplasm. In addition, activated hedgehog signaling has been reported to be involved in fibrogenesis in many tissues, including the pancreas. Therefore, new therapeutic targets based on hedgehog signaling have attracted a great deal of attention to alleviate pancreatic diseases. In this review, we briefly discuss the recent advances in hedgehog signaling in pancreatic fibrogenesis and carcinogenesis and highlight new insights on their potential relationship with respect to the development of novel targeted therapies. PMID:26962810

  1. Hedgehog Signaling in Pancreatic Fibrosis and Cancer.

    PubMed

    Bai, Yongyu; Bai, Yongheng; Dong, Jiaojiao; Li, Qiang; Jin, Yuepeng; Chen, Bicheng; Zhou, Mengtao

    2016-03-01

    The hedgehog signaling pathway was first discovered in the 1980s. It is a stem cell-related pathway that plays a crucial role in embryonic development, tissue regeneration, and organogenesis. Aberrant activation of hedgehog signaling leads to pathological consequences, including a variety of human tumors such as pancreatic cancer. Multiple lines of evidence indicate that blockade of this pathway with several small-molecule inhibitors can inhibit the development of pancreatic neoplasm. In addition, activated hedgehog signaling has been reported to be involved in fibrogenesis in many tissues, including the pancreas. Therefore, new therapeutic targets based on hedgehog signaling have attracted a great deal of attention to alleviate pancreatic diseases. In this review, we briefly discuss the recent advances in hedgehog signaling in pancreatic fibrogenesis and carcinogenesis and highlight new insights on their potential relationship with respect to the development of novel targeted therapies. PMID:26962810

  2. Hedgehog inhibits β-catenin activity in synovial joint development and osteoarthritis

    PubMed Central

    Rockel, Jason S.; Yu, Chunying; Whetstone, Heather; Craft, April M.; Reilly, Katherine; Ma, Henry; Tsushima, Hidetoshi; Puviindran, Vijitha; Al-Jazrawe, Mushriq; Keller, Gordon M.; Alman, Benjamin A.

    2016-01-01

    Both the WNT/β-catenin and hedgehog signaling pathways are important in the regulation of limb development, chondrocyte differentiation, and degeneration of articular cartilage in osteoarthritis (OA). It is not clear how these signaling pathways interact in interzone cell differentiation and synovial joint morphogenesis. Here, we determined that constitutive activation of hedgehog signaling specifically within interzone cells induces joint morphological changes by selectively inhibiting β-catenin–induced Fgf18 expression. Stabilization of β-catenin or treatment with FGF18 rescued hedgehog-induced phenotypes. Hedgehog signaling induced expression of a dominant negative isoform of TCF7L2 (dnTCF7L2) in interzone progeny, which may account for the selective regulation of β-catenin target genes observed. Knockdown of TCF7L2 isoforms in mouse chondrocytes rescued hedgehog signaling–induced Fgf18 downregulation, while overexpression of the human dnTCF7L2 orthologue (dnTCF4) in human chondrocytes promoted the expression of catabolic enzymes associated with OA. Similarly, expression of dnTCF4 in human chondrocytes positively correlated with the aggrecanase ADAMTS4. Consistent with our developmental findings, activation of β-catenin also attenuated hedgehog-induced or surgically induced articular cartilage degeneration in mouse models of OA. Thus, our results demonstrate that hedgehog inhibits selective β-catenin target gene expression to direct interzone progeny fates and articular cartilage development and disease. Moreover, agents that increase β-catenin activity have the potential to therapeutically attenuate articular cartilage degeneration as part of OA. PMID:27018594

  3. Amarogentin regulates self renewal pathways to restrict liver carcinogenesis in experimental mouse model.

    PubMed

    Sur, Subhayan; Pal, Debolina; Banerjee, Kaustav; Mandal, Suvra; Das, Ashes; Roy, Anup; Panda, Chinmay Kumar

    2016-07-01

    Amarogentin, a secoiridoid glycoside isolated from medicinal plant Swertia chirata, was found to restrict CCl4 /N-nitrosodiethyl amine (NDEA) induced mouse liver carcinogenesis by modulating G1/S cell cycle check point and inducing apoptosis. To understand its therapeutic efficacy on stem cell self renewal pathways, prevalence of CD44 positive cancer stem cell (CSC) population, expressions (mRNA/protein) of some key regulatory genes of self renewal Wnt and Hedgehog pathways along with expressions of E-cadherin and EGFR were analyzed during the liver carcinogenesis and in liver cancer cell line HepG2. It was observed that amarogentin could significantly reduce CD44 positive CSCs in both pre and post initiation stages of carcinogenesis than carcinogen control mice. In Wnt pathway, amarogentin could inhibit expressions of β-catenin, phospho β-catenin (Y-654) and activate expressions of antagonists sFRP1/2 and APC in the liver lesions. In Hedgehog pathway, decreased expressions of Gli1, sonic hedgehog ligand, and SMO along with up-regulation of PTCH1 were seen in the liver lesions due to amarogentin treatment. Moreover, amarogentin could up-regulate E-cadherin expression and down-regulate expression of EGFR in the liver lesions. Similarly, amarogentin could inhibit HepG2 cell growth along with expression and prevalence of CD44 positive CSCs. Similar to in vivo analysis, amarogentin could modulate the expressions of the key regulatory genes of the Wnt and hedgehog pathways and EGFR in HepG2 cells. Thus, our data suggests that the restriction of liver carcinogenesis by amarogentin might be due to reduction of CD44 positive CSCs and modulation of the self renewal pathways. © 2015 Wiley Periodicals, Inc. PMID:26154024

  4. Primary cilia integrate hedgehog and Wnt signaling during tooth development.

    PubMed

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

    2014-05-01

    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 Wnt1(Cre+)Kif3a(fl/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.

  5. Functional Interaction between HEXIM and Hedgehog Signaling during Drosophila Wing Development.

    PubMed

    Nguyen, Duy; Fayol, Olivier; Buisine, Nicolas; Lecorre, Pierrette; Uguen, Patricia

    2016-01-01

    Studying the dynamic of gene regulatory networks is essential in order to understand the specific signals and factors that govern cell proliferation and differentiation during development. This also has direct implication in human health and cancer biology. The general transcriptional elongation regulator P-TEFb regulates the transcriptional status of many developmental genes. Its biological activity is controlled by an inhibitory complex composed of HEXIM and the 7SK snRNA. Here, we examine the function of HEXIM during Drosophila development. Our key finding is that HEXIM affects the Hedgehog signaling pathway. HEXIM knockdown flies display strong phenotypes and organ failures. In the wing imaginal disc, HEXIM knockdown initially induces ectopic expression of Hedgehog (Hh) and its transcriptional effector Cubitus interuptus (Ci). In turn, deregulated Hedgehog signaling provokes apoptosis, which is continuously compensated by apoptosis-induced cell proliferation. Thus, the HEXIM knockdown mutant phenotype does not result from the apoptotic ablation of imaginal disc; but rather from the failure of dividing cells to commit to a proper developmental program due to Hedgehog signaling defects. Furthermore, we show that ci is a genetic suppressor of hexim. Thus, HEXIM ensures the integrity of Hedgehog signaling in wing imaginal disc, by a yet unknown mechanism. To our knowledge, this is the first time that the physiological function of HEXIM has been addressed in such details in vivo. PMID:27176767

  6. Functional Interaction between HEXIM and Hedgehog Signaling during Drosophila Wing Development

    PubMed Central

    Nguyen, Duy; Fayol, Olivier; Buisine, Nicolas; Lecorre, Pierrette; Uguen, Patricia

    2016-01-01

    Studying the dynamic of gene regulatory networks is essential in order to understand the specific signals and factors that govern cell proliferation and differentiation during development. This also has direct implication in human health and cancer biology. The general transcriptional elongation regulator P-TEFb regulates the transcriptional status of many developmental genes. Its biological activity is controlled by an inhibitory complex composed of HEXIM and the 7SK snRNA. Here, we examine the function of HEXIM during Drosophila development. Our key finding is that HEXIM affects the Hedgehog signaling pathway. HEXIM knockdown flies display strong phenotypes and organ failures. In the wing imaginal disc, HEXIM knockdown initially induces ectopic expression of Hedgehog (Hh) and its transcriptional effector Cubitus interuptus (Ci). In turn, deregulated Hedgehog signaling provokes apoptosis, which is continuously compensated by apoptosis-induced cell proliferation. Thus, the HEXIM knockdown mutant phenotype does not result from the apoptotic ablation of imaginal disc; but rather from the failure of dividing cells to commit to a proper developmental program due to Hedgehog signaling defects. Furthermore, we show that ci is a genetic suppressor of hexim. Thus, HEXIM ensures the integrity of Hedgehog signaling in wing imaginal disc, by a yet unknown mechanism. To our knowledge, this is the first time that the physiological function of HEXIM has been addressed in such details in vivo. PMID:27176767

  7. Mechanism and evolution of cytosolic Hedgehog signal transduction

    PubMed Central

    Wilson, Christopher W.; Chuang, Pao-Tien

    2010-01-01

    Hedgehog (Hh) signaling is required for embryonic patterning and postnatal physiology in invertebrates and vertebrates. With the revelation that the primary cilium is crucial for mammalian Hh signaling, the prevailing view that Hh signal transduction mechanisms are conserved across species has been challenged. However, more recent progress on elucidating the function of core Hh pathway cytosolic regulators in Drosophila, zebrafish and mice has confirmed that the essential logic of Hh transduction is similar between species. Here, we review Hh signaling events at the membrane and in the cytosol, and focus on parallel and divergent functions of cytosolic Hh regulators in Drosophila and mammals. PMID:20530542

  8. Hedgehog Signal Transduction: Key Players, Oncogenic Drivers, and Cancer Therapy.

    PubMed

    Pak, Ekaterina; Segal, Rosalind A

    2016-08-22

    The Hedgehog (Hh) signaling pathway governs complex developmental processes, including proliferation and patterning within diverse tissues. These activities rely on a tightly regulated transduction system that converts graded Hh input signals into specific levels of pathway activity. Uncontrolled activation of Hh signaling drives tumor initiation and maintenance. However, recent entry of pathway-specific inhibitors into the clinic reveals mixed patient responses and thus prompts further exploration of pathway activation and inhibition. In this review, we share emerging insights into regulated and oncogenic Hh signaling, supplemented with updates on the development and use of Hh pathway-targeted therapies.

  9. Click chemistry armed enzyme-linked immunosorbent assay to measure palmitoylation by hedgehog acyltransferase.

    PubMed

    Lanyon-Hogg, Thomas; Masumoto, Naoko; Bodakh, George; Konitsiotis, Antonio D; Thinon, Emmanuelle; Rodgers, Ursula R; Owens, Raymond J; Magee, Anthony I; Tate, Edward W

    2015-12-01

    Hedgehog signaling is critical for correct embryogenesis and tissue development. However, on maturation, signaling is also found to be aberrantly activated in many cancers. Palmitoylation of the secreted signaling protein sonic hedgehog (Shh) by the enzyme hedgehog acyltransferase (Hhat) is required for functional signaling. To quantify this important posttranslational modification, many in vitro Shh palmitoylation assays employ radiolabeled fatty acids, which have limitations in terms of cost and safety. Here we present a click chemistry armed enzyme-linked immunosorbent assay (click-ELISA) for assessment of Hhat activity through acylation of biotinylated Shh peptide with an alkyne-tagged palmitoyl-CoA (coenzyme A) analogue. Click chemistry functionalization of the alkyne tag with azido-FLAG peptide allows analysis through an ELISA protocol and colorimetric readout. This assay format identified the detergent n-dodecyl β-d-maltopyranoside as an improved solubilizing agent for Hhat activity. Quantification of the potency of RU-SKI small molecule Hhat inhibitors by click-ELISA indicated IC50 values in the low- or sub-micromolar range. A stopped assay format was also employed that allows measurement of Hhat kinetic parameters where saturating substrate concentrations exceed the binding capacity of the streptavidin-coated plate. Therefore, click-ELISA represents a nonradioactive method for assessing protein palmitoylation in vitro that is readily expandable to other classes of protein lipidation.

  10. Click chemistry armed enzyme-linked immunosorbent assay to measure palmitoylation by hedgehog acyltransferase

    PubMed Central

    Lanyon-Hogg, Thomas; Masumoto, Naoko; Bodakh, George; Konitsiotis, Antonio D.; Thinon, Emmanuelle; Rodgers, Ursula R.; Owens, Raymond J.; Magee, Anthony I.; Tate, Edward W.

    2015-01-01

    Hedgehog signaling is critical for correct embryogenesis and tissue development. However, on maturation, signaling is also found to be aberrantly activated in many cancers. Palmitoylation of the secreted signaling protein sonic hedgehog (Shh) by the enzyme hedgehog acyltransferase (Hhat) is required for functional signaling. To quantify this important posttranslational modification, many in vitro Shh palmitoylation assays employ radiolabeled fatty acids, which have limitations in terms of cost and safety. Here we present a click chemistry armed enzyme-linked immunosorbent assay (click–ELISA) for assessment of Hhat activity through acylation of biotinylated Shh peptide with an alkyne-tagged palmitoyl-CoA (coenzyme A) analogue. Click chemistry functionalization of the alkyne tag with azido-FLAG peptide allows analysis through an ELISA protocol and colorimetric readout. This assay format identified the detergent n-dodecyl β-d-maltopyranoside as an improved solubilizing agent for Hhat activity. Quantification of the potency of RU-SKI small molecule Hhat inhibitors by click–ELISA indicated IC50 values in the low- or sub-micromolar range. A stopped assay format was also employed that allows measurement of Hhat kinetic parameters where saturating substrate concentrations exceed the binding capacity of the streptavidin-coated plate. Therefore, click–ELISA represents a nonradioactive method for assessing protein palmitoylation in vitro that is readily expandable to other classes of protein lipidation. PMID:26334609

  11. Seismic detection of sonic booms.

    PubMed

    Cates, Joseph E; Sturtevant, Bradford

    2002-01-01

    The pressure signals from a sonic boom will produce a small, but detectable, ground motion. The extensive seismic network in southern California, consisting of over 200 sites covering over 50000 square kilometers, is used to map primary and secondary sonic boom carpets. Data from the network is used to analyze three supersonic overflights in the western United States. The results are compared to ray-tracing computations using a realistic model of the stratified atmospheric at the time of the measurements. The results show sonic boom ground exposure under the real atmosphere is much larger than previously expected or predicted by ray tracing alone. Finally, seismic observations are used to draw some inferences on the origin of a set of "mystery booms" recorded in 1992-1993 in southern California. PMID:11837967

  12. Anomalous dispersions of `hedgehog' particles

    NASA Astrophysics Data System (ADS)

    Bahng, Joong Hwan; Yeom, Bongjun; Wang, Yichun; Tung, Siu On; Hoff, J. Damon; Kotov, Nicholas

    2015-01-01

    Hydrophobic particles in water and hydrophilic particles in oil aggregate, but can form colloidal dispersions if their surfaces are chemically camouflaged with surfactants, organic tethers, adsorbed polymers or other particles that impart affinity for the solvent and increase interparticle repulsion. A different strategy for modulating the interaction between a solid and a liquid uses surface corrugation, which gives rise to unique wetting behaviour. Here we show that this topographical effect can also be used to disperse particles in a wide range of solvents without recourse to chemicals to camouflage the particles' surfaces: we produce micrometre-sized particles that are coated with stiff, nanoscale spikes and exhibit long-term colloidal stability in both hydrophilic and hydrophobic media. We find that these `hedgehog' particles do not interpenetrate each other with their spikes, which markedly decreases the contact area between the particles and, therefore, the attractive forces between them. The trapping of air in aqueous dispersions, solvent autoionization at highly developed interfaces, and long-range electrostatic repulsion in organic media also contribute to the colloidal stability of our particles. The unusual dispersion behaviour of our hedgehog particles, overturning the notion that like dissolves like, might help to mitigate adverse environmental effects of the use of surfactants and volatile organic solvents, and deepens our understanding of interparticle interactions and nanoscale colloidal chemistry.

  13. Gli1-Mediated Regulation of Sox2 Facilitates Self-Renewal of Stem-Like Cells and Confers Resistance to EGFR Inhibitors in Non-Small Cell Lung Cancer.

    PubMed

    Bora-Singhal, Namrata; Perumal, Deepak; Nguyen, Jonathan; Chellappan, Srikumar

    2015-07-01

    Non-small cell lung cancer (NSCLC) patients have very low survival rates because the current therapeutic strategies are not fully effective. Although EGFR tyrosine kinase inhibitors are effective for NSCLC patients harboring EGFR mutations, patients invariably develop resistance to these agents. Alterations in multiple signaling cascades have been associated with the development of resistance to EGFR inhibitors. Sonic Hedgehog and associated Gli transcription factors play a major role in embryonic development and have recently been found to be reactivated in NSCLC, and elevated Gli1 levels correlate with poor prognosis. The Hedgehog pathway has been implicated in the functions of cancer stem cells, although the underlying molecular mechanisms are not clear. In this context, we demonstrate that Gli1 is a strong regulator of embryonic stem cell transcription factor Sox2. Depletion of Gli1 or inhibition of the Hedgehog signaling significantly abrogated the self-renewal of stem-like side-population cells from NSCLCs as well as vascular mimicry of such cells. Gli1 was found to transcriptionally regulate Sox2 through its promoter region, and Gli1 could be detected on the Sox2 promoter. Inhibition of Hedgehog signaling appeared to work cooperatively with EGFR inhibitors in markedly reducing the viability of NSCLC cells as well as the self-renewal of stem-like cells. Thus, our study demonstrates a cooperative functioning of the EGFR signaling and Hedgehog pathways in governing the stem-like functions of NSCLC cancer stem cells and presents a novel therapeutic strategy to combat NSCLC harboring EGFR mutations.

  14. Real Time Sonic Boom Display

    NASA Technical Reports Server (NTRS)

    Haering, Ed

    2014-01-01

    This presentation will provide general information about sonic boom mitigation technology to the public in order to supply information to potential partners and licensees. The technology is a combination of flight data, atmospheric data and terrain information implemented into a control room real time display for flight planning. This research is currently being performed and as such, any results and conclusions are ongoing.

  15. The Sonic Pathfinder: An Evaluation.

    ERIC Educational Resources Information Center

    Dodds, Allan G.; And Others

    1984-01-01

    An objective evaluation of the Sonic Pathfinder, a new ultrasonic mobility aid, showed that use of the aid changes mobility in many ways. Reduced perception of environmental sounds was not reflected in performance. The majority of users traveled slowly and exhibited less than optimal strategies. (Author/CL)

  16. Sonicated Diagnostic Immunoblot for Bartonellosis

    PubMed Central

    Mallqui, Vania; Speelmon, Emily C.; Verástegui, Manuela; Maguiña-Vargas, Ciro; Pinell-Salles, Paula; Lavarello, Rosa; Delgado, Jose; Kosek, Margaret; Romero, Sofia; Arana, Yanina; Gilman, Robert H.

    2000-01-01

    Two simple Bartonella bacilliformis immunoblot preparation methods were developed. Antigen was prepared by two different methods: sonication of whole organisms or glycine extraction. Both methods were then tested for sensitivity and specificity. Well-defined control sera were utilized in the development of these diagnostic immunoblots, and possible cross-reactions were thoroughly examined. Sera investigated for cross-reaction with these diagnostic antigens were drawn from patients with brucellosis, chlamydiosis, Q fever, and cat scratch disease, all of whom were from regions where bartonellosis is not endemic. While both immunoblots yielded reasonable sensitivity and high specificity, we recommend the use of the sonicated immunoblot, which has a higher sensitivity when used to detect acute disease and produces fewer cross-reactions. The sonicated immunoblot reported here is 94% sensitive to chronic bartonellosis and 70% sensitive to acute bartonellosis. In a healthy group, it is 100% specific. This immunoblot preparation requires a simple sonication protocol for the harvesting of B. bacilliformis antigens and is well suited for use in regions of endemicity. PMID:10618267

  17. Evolutionary Genomics and Adaptive Evolution of the Hedgehog Gene Family (Shh, Ihh and Dhh) in Vertebrates

    PubMed Central

    Pereira, Joana; Johnson, Warren E.; O’Brien, Stephen J.; Jarvis, Erich D.; Zhang, Guojie; Gilbert, M. Thomas P.; Vasconcelos, Vitor; Antunes, Agostinho

    2014-01-01

    The Hedgehog (Hh) gene family codes for a class of secreted proteins composed of two active domains that act as signalling molecules during embryo development, namely for the development of the nervous and skeletal systems and the formation of the testis cord. While only one Hh gene is found typically in invertebrate genomes, most vertebrates species have three (Sonic hedgehog – Shh; Indian hedgehog – Ihh; and Desert hedgehog – Dhh), each with different expression patterns and functions, which likely helped promote the increasing complexity of vertebrates and their successful diversification. In this study, we used comparative genomic and adaptive evolutionary analyses to characterize the evolution of the Hh genes in vertebrates following the two major whole genome duplication (WGD) events. To overcome the lack of Hh-coding sequences on avian publicly available databases, we used an extensive dataset of 45 avian and three non-avian reptilian genomes to show that birds have all three Hh paralogs. We find suggestions that following the WGD events, vertebrate Hh paralogous genes evolved independently within similar linkage groups and under different evolutionary rates, especially within the catalytic domain. The structural regions around the ion-binding site were identified to be under positive selection in the signaling domain. These findings contrast with those observed in invertebrates, where different lineages that experienced gene duplication retained similar selective constraints in the Hh orthologs. Our results provide new insights on the evolutionary history of the Hh gene family, the functional roles of these paralogs in vertebrate species, and on the location of mutational hotspots. PMID:25549322

  18. Evolutionary genomics and adaptive evolution of the Hedgehog gene family (Shh, Ihh and Dhh) in vertebrates.

    PubMed

    Pereira, Joana; Johnson, Warren E; O'Brien, Stephen J; Jarvis, Erich D; Zhang, Guojie; Gilbert, M Thomas P; Vasconcelos, Vitor; Antunes, Agostinho

    2014-01-01

    The Hedgehog (Hh) gene family codes for a class of secreted proteins composed of two active domains that act as signalling molecules during embryo development, namely for the development of the nervous and skeletal systems and the formation of the testis cord. While only one Hh gene is found typically in invertebrate genomes, most vertebrates species have three (Sonic hedgehog--Shh; Indian hedgehog--Ihh; and Desert hedgehog--Dhh), each with different expression patterns and functions, which likely helped promote the increasing complexity of vertebrates and their successful diversification. In this study, we used comparative genomic and adaptive evolutionary analyses to characterize the evolution of the Hh genes in vertebrates following the two major whole genome duplication (WGD) events. To overcome the lack of Hh-coding sequences on avian publicly available databases, we used an extensive dataset of 45 avian and three non-avian reptilian genomes to show that birds have all three Hh paralogs. We find suggestions that following the WGD events, vertebrate Hh paralogous genes evolved independently within similar linkage groups and under different evolutionary rates, especially within the catalytic domain. The structural regions around the ion-binding site were identified to be under positive selection in the signaling domain. These findings contrast with those observed in invertebrates, where different lineages that experienced gene duplication retained similar selective constraints in the Hh orthologs. Our results provide new insights on the evolutionary history of the Hh gene family, the functional roles of these paralogs in vertebrate species, and on the location of mutational hotspots. PMID:25549322

  19. Inflammatory PAF Receptor Signaling Initiates Hedgehog Signaling and Kidney Fibrogenesis During Ethanol Consumption.

    PubMed

    Latchoumycandane, Calivarathan; Hanouneh, Mohamad; Nagy, Laura E; McIntyre, Thomas M

    2015-01-01

    Acute inflammation either resolves or proceeds to fibrotic repair that replaces functional tissue. Pro-fibrotic hedgehog signaling and induction of its Gli transcription factor in pericytes induces fibrosis in kidney, but molecular instructions connecting inflammation to fibrosis are opaque. We show acute kidney inflammation resulting from chronic ingestion of the common xenobiotic ethanol initiates Gli1 transcription and hedgehog synthesis in kidney pericytes, and promotes renal fibrosis. Ethanol ingestion stimulated transcription of TGF-ß, collagens I and IV, and alpha-smooth muscle actin with accumulation of these proteins. This was accompanied by deposition of extracellular fibrils. Ethanol catabolism by CYP2E1 in kidney generates local reactive oxygen species that oxidize cellular phospholipids to phospholipid products that activate the Platelet-activating Factor receptor (PTAFR) for inflammatory phospholipids. Genetically deleting this ptafr locus abolished accumulation of mRNA for TGF-ß, collagen IV, and α-smooth muscle actin. Loss of PTAFR also abolished ethanol-stimulated Sonic (Shh) and Indian hedgehog (Ihh) expression, and abolished transcription and accumulation of Gli1. Shh induced in pericytes and Ihh in tubules escaped to urine of ethanol-fed mice. Neutrophil myeloperoxidase (MPO) is required for ethanol-induced kidney inflammation, and Shh was not present in kidney or urine of mpo-/- mice. Shh also was present in urine of patients with acute kidney injury, but not in normal individuals or those with fibrotic liver cirrhosis We conclude neither endogenous PTAFR signaling nor CYP2E1-generated radicals alone are sufficient to initiate hedgehog signaling, but instead PTAFR-dependent neutrophil infiltration with myeloperoxidase activation is necessary to initiate ethanol-induced fibrosis in kidney. We also show fibrogenic mediators escape to urine, defining a new class of urinary mechanistic biomarkers of fibrogenesis for an organ not commonly

  20. Dataset for phenotypic classification of genetic modifiers of smoothened and Hedgehog.

    PubMed

    Marada, Suresh; Truong, Ashley; Ogden, Stacey K

    2016-06-01

    This data article includes supporting information for the research article entitled "The Small GTPase Rap1 is a Modulator of Hedgehog Signaling" [1]. Drosophila wing phenotypes induced by expression of a dominant negative Smoothened (Smo) mutant were cataloged into five distinct classes. Class distributions observed following expression of dominant negative Smo in control and sensitized backgrounds were quantified to serve as references for strength of phenotypic modification. Shifts in class distribution of Hedgehog (Hh) wing phenotypes resulting from introduction of loss-of-function alleles of select Ras family G protein genes and the Hh pathway regulators Fused and Suppressor of Fused are shown. PMID:27014736

  1. In Vivo Imaging of Hedgehog Pathway Activation with a Nuclear Fluorescent Reporter

    PubMed Central

    Mich, John K.; Payumo, Alexander Y.; Rack, Paul G.; Chen, James K.

    2014-01-01

    The Hedgehog (Hh) pathway is essential for embryonic development and tissue regeneration, and its dysregulation can lead to birth defects and tumorigenesis. Understanding how this signaling mechanism contributes to these processes would benefit from an ability to visualize Hedgehog pathway activity in live organisms, in real time, and with single-cell resolution. We report here the generation of transgenic zebrafish lines that express nuclear-localized mCherry fluorescent protein in a Gli transcription factor-dependent manner. As demonstrated by chemical and genetic perturbations, these lines faithfully report Hedgehog pathway state in individual cells and with high detection sensitivity. They will be valuable tools for studying dynamic Gli-dependent processes in vertebrates and for identifying new chemical and genetic regulators of the Hh pathway. PMID:25068273

  2. Hedgehog inhibitors from Withania somnifera.

    PubMed

    Yoneyama, Tatsuro; Arai, Midori A; Sadhu, Samir K; Ahmed, Firoj; Ishibashi, Masami

    2015-09-01

    The hedgehog (Hh) signaling pathway performs an important role in embryonic development and in cellular proliferation and differentiation. However, aberrant activation of the Hh signaling pathway is associated with tumorigenesis. Hh signal inhibition was evaluated using a cell-based assay system that targets GLI1-mediated transcription. Activity-guided isolation of the Withania somnifera MeOH extract led to the isolation of six compounds: withaferin A (1) and its derivatives (2-6). Compounds 1 and 2 showed strong inhibition of Hh/GLI1-mediated transcriptional activity with IC50 values of 0.5 and 0.6 μM, respectively. Compounds 1, 2, 3, and 6 were cytotoxic toward human pancreatic (PANC-1), prostate (DU145) and breast (MCF7) cancer cells. Furthermore, 1 also inhibited GLI1-DNA complex formation in EMSA.

  3. Midline-derived Shh regulates mesonephric tubule formation through the paraxial mesoderm

    PubMed Central

    Murashima, Aki; Akita, Hiroki; Okazawa, Mika; Kishigami, Satoshi; Nakagata, Naomi; Nishinakamura, Ryuichi; Yamada, Gen

    2014-01-01

    During organogenesis, Sonic hedgehog (Shh) possesses dual functions: Shh emanating from midline structures regulates the positioning of bilateral structures at early stages, whereas organ-specific Shh locally regulates organ morphogenesis at later stages. The mesonephros is a transient embryonic kidney in amniote, whereas it becomes definitive adult kidney in some anamniotes. Thus, elucidating the regulation of mesonephros formation has important implications for our understanding of kidney development and evolution. In Shh knockout (KO) mutant mice, the mesonephros was displaced towards the midline and ectopic mesonephric tubules (MTs) were present in the caudal mesonephros. Mesonephros-specific ablation of Shh in Hoxb7-Cre;Shhflox/− and Sall1CreERT2/+;Shhflox/− mice embryos indicated that Shh expressed in the mesonephros was not required for either the development of the mesonephros or the differentiation of the male reproductive tract. Moreover, stage-specific ablation of Shh in ShhCreERT2/flox mice showed that notochord- and/or floor plate-derived Shh were essential for the regulation of the number and position of MTs. Lineage analysis of hedgehog (Hh)-responsive cells, and analysis of gene expression in Shh KO embryos suggested that Shh regulated nephrogenic gene expression indirectly, possibly through effects on the paraxial mesoderm. These data demonstrate the essential role of midline-derived Shh in local tissue morphogenesis and differentiation. PMID:24370450

  4. High Speed Research Program Sonic Fatigue

    NASA Technical Reports Server (NTRS)

    Rizzi, Stephen A. (Technical Monitor); Beier, Theodor H.; Heaton, Paul

    2005-01-01

    The objective of this sonic fatigue summary is to provide major findings and technical results of studies, initiated in 1994, to assess sonic fatigue behavior of structure that is being considered for the High Speed Civil Transport (HSCT). High Speed Research (HSR) program objectives in the area of sonic fatigue were to predict inlet, exhaust and boundary layer acoustic loads; measure high cycle fatigue data for materials developed during the HSR program; develop advanced sonic fatigue calculation methods to reduce required conservatism in airframe designs; develop damping techniques for sonic fatigue reduction where weight effective; develop wing and fuselage sonic fatigue design requirements; and perform sonic fatigue analyses on HSCT structural concepts to provide guidance to design teams. All goals were partially achieved, but none were completed due to the premature conclusion of the HSR program. A summary of major program findings and recommendations for continued effort are included in the report.

  5. Counterexamples to the Sonic Criterion

    NASA Astrophysics Data System (ADS)

    Elling, Volker

    2009-12-01

    We consider self-similar (pseudo-steady) shock reflection at an oblique wall. There are three parameters: wall corner angle, Mach number, angle of incident shock. Ever since Ernst Mach discovered the irregular reflection named after him, researchers have sought to predict precisely for which parameters the reflection is regular. Three conflicting proposals—the detachment, sonic and von Neumann criteria—have been studied extensively without a clear result. We demonstrate that the sonic criterion is not correct. We consider polytropic potential flow and prove that there is an open nonempty set of parameters that admit a global regular reflection with a reflected shock that is transonic. We also provide a clear physical reason: the flow type (sub- or supersonic) is not decisive; instead the reflected shock type (weak or strong) determines whether structural perturbations decay towards the reflection point.

  6. Review of sonic fatigue technology

    NASA Technical Reports Server (NTRS)

    Clarkson, B. L.

    1994-01-01

    From the early-1960s until the mid-1980s, there was very little theoretical development for sonic fatigue prediction. Design nomographs based on simple theoretical models and results of specially designed tests were developed for most common aircraft structures. The use of advanced composites in the 1980s, however, generated an increased interest in development of more sophisticated theoretical models because of the possibilities for a much wider range of structural designs. The purpose of this report is to review sonic fatigue technology and, in particular, to assess recent developments. It also suggests a plan for a coordinated program of theoretical and experimental work to meet the anticipated needs of future aerospace vehicles.

  7. Georgia Tech sonic boom simulator

    NASA Technical Reports Server (NTRS)

    Ahuja, Krish K.

    1992-01-01

    To examine the building and human response to sonic boom in the range 3 Hz to 30 Hz, Georgia Institute of Technology is building a special acoustic driver system to simulate sonic boom. To support the NASA LaRC program on building and human response, this simulator's capability has been extended to an upper frequency of 4 KHz. A residential test house was made available by Georgia Tech for these tests. At the time of preparation of this document, most of the acoustic drivers and the associated electronics have been built and assembled. The system has, however, not been fully tested. The following pages provide an overview of the progress to date. The acoustic driver systems, and the principle of their operation together with the test house are described. Future plans are also summarized.

  8. Dispersion of Single Walled Carbon Nanotubes Using a Novel Type of Sonication: Focused Sonication.

    PubMed

    Sachin, Bramhe N; Ae, Hwangbo Seon; Chu, Min Cheol

    2016-03-01

    We demonstrate the use of novel type of sonication method, focused sonication, with added advantages over bath and probe type of sonication for the dispersion of single walled carbon nanotubes (SWNT). Di-chloro benzene was used as the solvent for dispersion of SWNT. Results from focused sonication and bath sonication were compared and found that focused sonication results in better dispersion. Also Raman spectroscopy was analysed to ascertain if focused sonication causes any damage to the tubes and it was found that there was no damage to the SWNT. We believe that with the added advantages like in-situ temperature control and large sample volume processing, focused sonication would prove to be the most proficient method of sonication for dispersion of nanoparticles. PMID:27455717

  9. Experimentally determined aeroacoustic performance and control of several sonic inlets

    NASA Technical Reports Server (NTRS)

    Miller, B. A.

    1975-01-01

    Low speed wind tunnel tests were conducted to determine the aeroacoustic performance of several model sonic inlets. The results were analyzed to indicate how inlet aeroacoustic characteristics were affected by inlet design and operating conditions. A system for regulating sonic inlet noise reduction was developed and tested. Results indicate that pressure losses at forward velocity may be substantially less than those at static conditions. This is particularly true for translating centerbody inlets with the centerbody extended in the approach and landing position. Operation to simulated takeoff incidence angles of 50 degrees was demonstrated with good inlet performance. Inlet sound pressure level reduction was regulation was regulated to within approximately + or - 1 dB by controlling inlet surface static pressure measured at the diffuser exit.

  10. Aberrant expression of Sonic hedgehog signaling in Peutz-Jeghers syndrome.

    PubMed

    Xu, Xiaoping; Su, Juan; Li, Ran; Wang, Yadong; Zeng, Di; Wu, Baoping

    2016-04-01

    The SHH signaling pathway is critical for gastrointestinal development and organic patterning, and dysregulation of SHH pathway molecules has been detected in multiple gastrointestinal neoplasms. This study investigated the role of the SHH signaling pathway in PJS. Expression of SHH, PTCH, and GLI1 was examined by real-time PCR and immunohistochemistry in 20 normal tissues and 75 colorectal lesions (25 PJPs, 25 adenomas, and 25 adenocarcinomas). Expression of SHH, PTCH, and GLI1 mRNA was higher in PJPs than in normal tissue (P < .05) and gradually increased along the PJP-adenoma-adenocarcinoma sequence (P < .05). Immunostaining indicated that SHH expression was present in 60% of PJPs, 72% of adenomas, and 84% of carcinomas, whereas 68% of PJPs, 72% of adenomas, and 88% of carcinomas exhibited cytoplasmic expression of PTCH. Moreover, high GLI1 expression was detected in 56% of PJPs, 64% of adenomas, and 80% of carcinomas; and high nuclear expression of GLI1 was observed in 8 adenomas with atypia and 15 carcinomas. Increased SHH, PTCH, and GLI1 protein correlated positively with tumor grade (P = .012, P = .003, and P = .007, respectively), tumor depth (P = .024, P = .007, and P = .01), and lymph node metastasis (P = .05, P = .015, and P = .005). This study identified aberrant expression of SHH pathway molecules in PJS, and the findings may supply a novel mechanism for the development of PJ polyps. PMID:26997450

  11. Roles for Hedgehog signaling in adult organ homeostasis and repair

    PubMed Central

    Petrova, Ralitsa; Joyner, Alexandra L.

    2014-01-01

    The hedgehog (HH) pathway is well known for its mitogenic and morphogenic functions during development, and HH signaling continues in discrete populations of cells within many adult mammalian tissues. Growing evidence indicates that HH regulates diverse quiescent stem cell populations, but the exact roles that HH signaling plays in adult organ homeostasis and regeneration remain poorly understood. Here, we review recently identified functions of HH in modulating the behavior of tissue-specific adult stem and progenitor cells during homeostasis, regeneration and disease. We conclude that HH signaling is a key factor in the regulation of adult tissue homeostasis and repair, acting via multiple different routes to regulate distinct cellular outcomes, including maintenance of plasticity, in a context-dependent manner. PMID:25183867

  12. Hedgehog Cholesterolysis: Specialized Gatekeeper to Oncogenic Signaling

    PubMed Central

    Callahan, Brian P.; Wang, Chunyu

    2015-01-01

    Discussions of therapeutic suppression of hedgehog (Hh) signaling almost exclusively focus on receptor antagonism; however, hedgehog’s biosynthesis represents a unique and potentially targetable aspect of this oncogenic signaling pathway. Here, we review a key biosynthetic step called cholesterolysis from the perspectives of structure/function and small molecule inhibition. Cholesterolysis, also called cholesteroylation, generates cholesterol-modified Hh ligand via autoprocessing of a hedgehog precursor protein. Post-translational modification by cholesterol appears to be restricted to proteins in the hedgehog family. The transformation is essential for Hh biological activity and upstream of signaling events. Despite its decisive role in generating ligand, cholesterolysis remains conspicuously unexplored as a therapeutic target. PMID:26473928

  13. Permeability extraction: A sonic log inversion

    SciTech Connect

    Akbar, N.; Kim, J.J.

    1994-12-31

    In this paper the authors provide the missing important link between permeability and acoustic velocities by generating a permeability-dependent synthetic sonic log in a carbonate reservoir. The computations are based on Akbar`s theory that relates wave velocity to frequency, rock properties (e.g., lithology, permeability, and porosity), and fluid saturation and properties (viscosity, density, and compressibility). An inverted analytical expression of the theory is used to extract permeability from sonic velocity. The synthetic sonic and the computed permeability are compared with the observed sonic log and with plug permeability, respectively. The results demonstrate, as predicted by theory, that permeability can be related directly to acoustic velocities.

  14. Basal cell carcinomas: attack of the hedgehog.

    PubMed

    Epstein, Ervin H

    2008-10-01

    Basal cell carcinomas (BCCs) were essentially a molecular 'black box' until some 12 years ago, when identification of a genetic flaw in a rare subset of patients who have a great propensity to develop BCCs pointed to aberrant Hedgehog signalling as the pivotal defect leading to formation of these tumours. This discovery has facilitated a remarkable increase in our understanding of BCC carcinogenesis and has highlighted the carcinogenic role of this developmental pathway when aberrantly activated in adulthood. Importantly, a phase 1 first-in-human trial of a Hedgehog inhibitor has shown real progress in halting and even reversing the growth of these tumours.

  15. Ultrasonic/Sonic Impacting Penetrators

    NASA Technical Reports Server (NTRS)

    Bao, Xiaoqi; Bar-Cohen, Yoseph; Chang, Zensheu; Sherrit, Stewart; Stark, Randall A.

    2008-01-01

    Ultrasonic/sonic impacting penetrators (USIPs) are recent additions to the series of apparatuses based on ultrasonic/sonic drill corers (USDCs). A USIP enables a rod probe to penetrate packed soil or another substance of similar consistency, without need to apply a large axial force that could result in buckling of the probe or in damage to some buried objects. USIPs were conceived for use in probing and analyzing soil to depths of tens of centimeters in the vicinity of buried barrels containing toxic waste, without causing rupture of the barrels. USIPs could also be used for other purposes, including, for example, searching for pipes, barrels, or other hard objects buried in soil; and detecting land mines. USDCs and other apparatuses based on USDCs have been described in numerous previous NASA Tech Briefs articles. The ones reported previously were designed, variously, for boring into, and/or acquiring samples of, rock or other hard, brittle materials of geological interest. To recapitulate: A USDC can be characterized as a lightweight, low-power, piezoelectrically driven jackhammer in which ultrasonic and sonic vibrations are generated and coupled to a tool bit. As shown in the figure, a basic USDC includes a piezoelectric stack, a backing and a horn connected to the stack, a free mass (free in the sense that it can slide axially a short distance between the horn and the shoulder of tool bit), and a tool bit, i.e., probe for USIP. The piezoelectric stack is driven at the resonance frequency of the stack/horn/backing assembly to create ultrasonic vibrations that are mechanically amplified by the horn. To prevent fracture during operation, the piezoelectric stack is held in compression by a bolt. The bouncing of the free mass between the horn and the tool bit at sonic frequencies generates hammering actions to the bit that are more effective for drilling than is the microhammering action of ultrasonic vibrations in ordinary ultrasonic drills. The hammering actions

  16. Hedgehog pathway gene expression during early development of the molar tooth root in the mouse.

    PubMed

    Khan, Mohammed; Seppala, Maisa; Zoupa, Maria; Cobourne, Martyn T

    2007-01-01

    Sonic hedgehog is a secreted protein important for many aspects of embryonic development. In the developing tooth, Shh expression is restricted to the epithelial compartment and plays an important role during both initiation and subsequent coronal morphogenesis. We have investigated the expression of Shh and constituent members of the signalling pathway during early development of the molar tooth root in the mouse and find the presence of transcripts in Hertwig's epithelial root sheath. These epithelial cells of the root sheath and the surrounding apical mesenchyme of the dental papilla and follicle also expressed the Shh receptor Ptc1, agonist Smo and Gli downstream transcriptional effectors; however, this response occurred over short range. In contrast, the Shh antagonists Hip1 and Gas1 were both expressed at a distance from these responding cells, in more peripheral regions of the developing root. Transcripts of the Skn acyl transferase lacked specific expression in early root structures.

  17. Integration of telencephalic Wnt and hedgehog signaling center activities by Foxg1.

    PubMed

    Danesin, Catherine; Peres, João N; Johansson, Marie; Snowden, Victoria; Cording, Amy; Papalopulu, Nancy; Houart, Corinne

    2009-04-01

    The forebrain is patterned along the dorsoventral (DV) axis by Sonic Hedgehog (Shh). However, previous studies have suggested the presence of an Shh-independent mechanism. Our study identifies Wnt/beta-catenin-activated from the telencephalic roof-as an Shh-independent pathway that is essential for telencephalic pallial (dorsal) specification during neurulation. We demonstrate that the transcription factor Foxg1 coordinates the activity of two signaling centers: Foxg1 is a key downstream effector of the Shh pathway during induction of subpallial (ventral) identity, and it inhibits Wnt/beta-catenin signaling through direct transcriptional repression of Wnt ligands. This inhibition restricts the dorsal Wnt signaling center to the roof plate and consequently limits pallial identities. Concomitantly to these roles, Foxg1 controls the formation of the compartment boundary between telencephalon and basal diencephalon. Altogether, these findings identify a key direct target of Foxg1, and uncover a simple molecular mechanism by which Foxg1 integrates two opposing signaling centers.

  18. A novel function of the human oncogene Stil: Regulation of PC12 cell toxic susceptibility through the Shh pathway

    PubMed Central

    Li, Lei; Carr, Aprell L.; Sun, Lei; Drewing, Audrey; Lee, Jessica; Rao, Zihe

    2015-01-01

    The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in vertebrate species. Here, we report new findings of Stil in the regulation of toxic susceptibility in mammalian dopaminergic (DA)-like PC12 cells. RNAi-mediated knockdown of Stil expression did not affect the survival of proliferating PC12 cells but caused a significant amount of cell death in differentiated neurons after toxic drug treatment. In contrast, overexpression of Stil increased toxic susceptibility only in proliferating cells but produced no effect in mature neurons. Exogenetic inactivation or activation of the Sonic hedgehog (Shh) signaling transduction mimicked the effect of Stil knockdown or overexpression in regulation of PC12 cell toxic susceptibility, suggesting that Stil exerts its role through the Shh pathway. Together, the data provide evidence for novel functions of the human oncogene Stil in neural toxic susceptibility. PMID:26549353

  19. Smoothened transduces Hedgehog signal by forming a complex with Evc/Evc2.

    PubMed

    Yang, Cuiping; Chen, Wenlin; Chen, Yongbin; Jiang, Jin

    2012-11-01

    Hedgehog (Hh) signaling plays pivotal roles in embryonic development and adult tissue homeostasis in species ranging from Drosophila to mammals. The Hh signal is transduced by Smoothened (Smo), a seven-transmembrane protein related to G protein coupled receptors. Despite a conserved mechanism by which Hh activates Smo in Drosophila and mammals, how mammalian Hh signal is transduced from Smo to the Gli transcription factors is poorly understood. Here, we provide evidence that two ciliary proteins, Evc and Evc2, the products of human disease genes responsible for the Ellis-van Creveld syndrome, act downstream of Smo to transduce the Hh signal. We found that loss of Evc/Evc2 does not affect Sonic Hedgehog-induced Smo phosphorylation and ciliary localization but impedes Hh pathway activation mediated by constitutively active forms of Smo. Evc/Evc2 are dispensable for the constitutive Gli activity in Sufu(-/-) cells, suggesting that Evc/Evc2 act upstream of Sufu to promote Gli activation. Furthermore, we demonstrated that Hh stimulates binding of Evc/Evc2 to Smo depending on phosphorylation of the Smo C-terminal intracellular tail and that the binding is abolished in Kif3a(-/-) cilium-deficient cells. We propose that Hh activates Smo by inducing its phosphorylation, which recruits Evc/Evc2 to activate Gli proteins by antagonizing Sufu in the primary cilia.

  20. Chloroquine targets pancreatic cancer stem cells via inhibition of CXCR4 and hedgehog signaling.

    PubMed

    Balic, Anamaria; Sørensen, Morten Dræby; Trabulo, Sara Maria; Sainz, Bruno; Cioffi, Michele; Vieira, Catarina R; Miranda-Lorenzo, Irene; Hidalgo, Manuel; Kleeff, Joerg; Erkan, Mert; Heeschen, Christopher

    2014-07-01

    Pancreatic ductal adenocarcinoma is one of the deadliest carcinomas and is characterized by highly tumorigenic and metastatic cancer stem cells (CSC). CSCs evade available therapies, which preferentially target highly proliferative and more differentiated progenies, leaving behind CSCs as a putative source for disease relapse. Thus, to identify potentially more effective treatment regimens, we screened established and new compounds for their ability to eliminate CSCs in primary pancreatic cancer (stem) cells in vitro and corresponding patient-derived pancreatic cancer tissue xenografts in vivo. Intriguingly, we found that in vitro treatment with the antimalarial agent chloroquine significantly decreased CSCs, translating into diminished in vivo tumorigenicity and invasiveness in a large panel of pancreatic cancers. In vivo treatment in combination with gemcitabine was capable of more effectively eliminating established tumors and improved overall survival. The inhibitory effect of chloroquine was not related to inhibition of autophagy, but was due to inhibition of CXCL12/CXCR4 signaling, resulting in reduced phosphorylation of ERK and STAT3. Furthermore, chloroquine showed potent inhibition of hedgehog signaling by decreasing the production of Smoothened, translating into a significant reduction in sonic hedgehog-induced chemotaxis and downregulation of downstream targets in CSCs and the surrounding stroma. Our study demonstrates that via to date unreported effects, chloroquine is an effective adjuvant therapy to chemotherapy, offering more efficient tumor elimination and improved cure rates. Chloroquine should be further explored in the clinical setting as its success may help to more rapidly improve the poor prognosis of patients with pancreatic cancer. PMID:24785258

  1. Hedgehog signaling pathway is active in GBM with GLI1 mRNA expression showing a single continuous distribution rather than discrete high/low clusters.

    PubMed

    Chandra, Vikas; Das, Tapojyoti; Gulati, Puneet; Biswas, Nidhan K; Rote, Sarang; Chatterjee, Uttara; Ghosh, Samarendra N; Deb, Sumit; Saha, Suniti K; Chowdhury, Anup K; Ghosh, Subhashish; Rudin, Charles M; Mukherjee, Ankur; Basu, Analabha; Dhara, Surajit

    2015-01-01

    Hedgehog (Hh) signaling pathway is a valid therapeutic target in a wide range of malignancies. We focus here on glioblastoma multiforme (GBM), a lethal malignancy of the central nervous system (CNS). By analyzing RNA-sequencing based transcriptomics data on 149 clinical cases of TCGA-GBM database we show here a strong correlation (r = 0.7) between GLI1 and PTCH1 mRNA expression--as a hallmark of the canonical Hh-pathway activity in this malignancy. GLI1 mRNA expression varied in 3 orders of magnitude among the GBM patients of the same cohort showing a single continuous distribution-unlike the discrete high/low-GLI1 mRNA expressing clusters of medulloblastoma (MB). When compared with MB as a reference, the median GLI1 mRNA expression in GBM appeared 14.8 fold lower than that of the "high-Hh" cluster of MB but 5.6 fold higher than that of the "low-Hh" cluster of MB. Next, we demonstrated statistically significant up- and down-regulation of GLI1 mRNA expressions in GBM patient-derived low-passage neurospheres in vitro by sonic hedgehog ligand-enriched conditioned media (shh-CM) and by Hh-inhibitor drug vismodegib respectively. We also showed clinically achievable dose (50 μM) of vismodegib alone to be sufficient to induce apoptosis and cell cycle arrest in these low-passage GBM neurospheres in vitro. Vismodegib showed an effect on the neurospheres, both by down-regulating GLI1 mRNA expression and by inducing apoptosis/cell cycle arrest, irrespective of their relative endogenous levels of GLI1 mRNA expression. We conclude from our study that this single continuous distribution pattern of GLI1 mRNA expression technically puts almost all GBM patients in a single group rather than discrete high- or low-clusters in terms of Hh-pathway activity. That is suggestive of therapies with Hh-pathway inhibitor drugs in this malignancy without a need for further stratification of patients on the basis of relative levels of Hh-pathway activity among them. PMID:25775002

  2. Using homogenization, sonication and thermo-sonication to inactivate fungi

    PubMed Central

    Bevilacqua, Antonio; Sinigaglia, Milena; Corbo, Maria Rosaria

    2016-01-01

    Ultrasound (US), Thermo-sonication (TS) and High Pressure Homogenization (HPH) were studied as tools to inactivate the spores of Penicillium spp. and Mucor spp. inoculated in distilled water. For US, the power ranged from 40% to 100%, pulse from 2 to 10 s, and duration of the treatment from 2 to 10 min. TS was performed combining US (40–80% of power, for 8 min and pulse of 2 s) with a thermal treatment (50, 55 and 60°C at 4, 8 and 12 min). Homogenization was done at 30–150 MPa for 1, 2 and 3 times. Power was the most important factors to determine the antifungal effect of US and TS towards the conidia of Penicillium spp.; on the other hand, in US treatments Mucor spp. was also affected by pulse and time. HPH exerted a significant antifungal effect only if the highest pressures were applied for 2–3 times. PMID:27375964

  3. Twist transition of nematic hyperbolic hedgehogs

    NASA Astrophysics Data System (ADS)

    James, Richard; Fukuda, Jun-ichi

    2014-04-01

    Stability of an idealized hyperbolic hedgehog in a nematic liquid crystal against a twist transition is investigated by extending the methodology of Rüdinger and Stark [Liq. Cryst. 26, 753 (1999), 10.1080/026782999204840], where the hedgehog is confined between two concentric spheres. In the ideal hyperbolic-hedgehog the molecular orientation is assumed to rotate proportionally with respect to the inclination angle, θ (and in the opposite sense). However, when splay, k11, and bend, k33, moduli differ this proportionality is lost and the liquid crystal deforms relative to the ideal with bend and splay. Although slight, these deformations are shown to significantly shift the transition if k11/k33 is small. By increasing the degree of confinement the twist transition can be inhibited, a characteristic both hyperbolic and radial hedgehogs have in common. The twist transition of a hyperbolic defect that accompanies a particle is found to be well predicted by the earlier stability analysis of a thick shell.

  4. Ectoparasites of hedgehogs (Erinaceus concolor) from Turkey.

    PubMed

    Girisgin, Ahmet Onur; Senlik, Bayram; Aydin, Levent; Cirak, Veli Y

    2015-01-01

    Hedgehogs are small, nocturnal, spiny-coated animals that have been growing in popularity as exotic pets. However, these animals are host to a wide variety of viruses, bacteria, fungi and parasites, some of which are of zoonotic character. Thus, because hedgehogs have a potential role to transmit zoonoses including arthropod-borne diseases, we examined them for their ectoparasites. The study was carried out on hedgehogs found dead mainly due to road casualties in the Bursa province of Turkey. The ectoparasites were collected by both insecticide spraying of the body and inspection on a white paper carefully. Totally three species of ticks (Rhipicephalus sanguineus, Hyalomma aegyptium, Haemophysalis parvo) and one flea species (Archeopsylla erinacei) were detected. The prevalence of mixed infestation with both ticks and fleas was 45.5%. Haemaphysalis parva was reported for the first time from hedgehogs (Erinaceus concolor) in Turkey. The occurrence of ectoparasites and their potential role as vectors of certain zoonotic diseases are briefly discussed. PMID:26281445

  5. Status of knowledge of sonic booms

    NASA Technical Reports Server (NTRS)

    Maglieri, D. J.; Carlson, H. W.; Hubbard, H. H.

    1980-01-01

    The effects of primary and secondary sonic boom carpets are reviewed with reference to waveform characteristics and audibility, role of the atmosphere, source characteristics, carpet measurements and predictions, effects of aircraft maneuvers, and sonic boom minimization. It is shown that primary booms which involve only propagation in the lower atmosphere are well predicted by means of current methods and are amendable to minimization.

  6. LRP1 mediates Hedgehog-induced endocytosis of the GPC3–Hedgehog complex

    PubMed Central

    Capurro, Mariana I.; Shi, Wen; Filmus, Jorge

    2013-01-01

    Summary Glypican-3 (GPC3) is a heparan sulfate (HS) proteoglycan that is bound to the cell membrane through a glycosylphosphatidylinositol link. This glypican regulates embryonic growth by inhibiting the hedgehog (Hh) signaling pathway. GPC3 binds Hh and competes with Patched (Ptc), the Hh receptor, for Hh binding. The interaction of Hh with GPC3 triggers the endocytosis and degradation of the GPC3–Hh complex with the consequent reduction of Hh available for binding to Ptc. Currently, the molecular mechanisms by which the GPC3–Hh complex is internalized remains unknown. Here we show that the low-density-lipoprotein receptor-related protein-1 (LRP1) mediates the Hh-induced endocytosis of the GPC3–Hh complex, and that this endocytosis is necessary for the Hh-inhibitory activity of GPC3. Furthermore, we demonstrate that GPC3 binds through its HS chains to LRP1, and that this interaction causes the removal of GPC3 from the lipid rafts domains. PMID:22467855

  7. Effect of sonication on different quality parameters of Pinus massoniana pollen.

    PubMed

    Liu, Xiao-dong; Zhang, Fu-bo; Zhou, Bin; Shan, Hu; Chen, Pu-Yan

    2015-01-01

    A study was initiated with the objective of evaluating the effects of sonication treatment on important quality parameters of extract of Pinus massoniana pollen. Sonication of extract was done (frequency 20kHz and various amplitude levels) for 10, 30, 50min, respectively. As results, total polysaccharide, phenolics and flavonoids significantly increased (P<0.05). And sonicated P.massoniana pollen displays strong immuno-stimulating activity by increasing proliferations of splenic lymphocytes and subsets of CD4+ T cells (CD3+CD4+), CD8 T cells (CD3+CD8+), and increased Ig secretion. Sonicated P. massoniana pollen also showed anti-tumor function by inhibition of tumor cell proliferation, inhibition of ROS production, up-regulation of GSH/GSSG ration, up-regulating the gene expression of P53, Bax and down-regulating the gene expression of Bcl-2. Findings of the present study suggested the sonication treatment of P. massoniana pollen could improve the quality and bioactivity of P. massoniana pollen, indicating that sonication is effective in processing of pollen and could be a potential process in tumor prevention and treatment.

  8. Development of the sonic pump levitation

    NASA Technical Reports Server (NTRS)

    Dunn, S. A.

    1984-01-01

    A prototype levitating/positioning device termed the Sonic Pump Levitator was designed, built and successfully tested in full gravity and in the reduced gravity of the parabolic flight regime of the KC-135. Positioning is achieved by timely and appropriate application of gas momentum from one or more of six sonic pumps. The sonic pumps, which are arranged orthogonally in opposed pairs about the levitation region, are activated by an electro-optical, computer controlled, feedback system. The sonic pump is a transducer which is capable of converting sound energy into a directed flow of gas. It consists of a loudspeaker whose face is sealed by a closure perforated by one or more orifices. The diaphragm of the loudspeaker is the only moving part of the sonic pump, no valves being needed. This very low inertia electromechanical device was developed to provide the short response time necessary to keep pace with the demands of computerized position keeping.

  9. Status of sonic boom methodology and understanding

    NASA Technical Reports Server (NTRS)

    Darden, Christine M.; Powell, Clemans A.; Hayes, Wallace D.; George, Albert R.; Pierce, Allan D.

    1989-01-01

    In January 1988, approximately 60 representatives of industry, academia, government, and the military gathered at NASA-Langley for a 2 day workshop on the state-of-the-art of sonic boom physics, methodology, and understanding. The purpose of the workshop was to assess the sonic boom area, to determine areas where additional sonic boom research is needed, and to establish some strategies and priorities in this sonic boom research. Attendees included many internationally recognized sonic boom experts who had been very active in the Supersonic Transport (SST) and Supersonic Cruise Aircraft Research Programs of the 60's and 70's. Summaries of the assessed state-of-the-art and the research needs in theory, minimization, atmospheric effects during propagation, and human response are given.

  10. Sonic boom interaction with turbulence

    NASA Technical Reports Server (NTRS)

    Rusak, Zvi; Giddings, Thomas E.

    1994-01-01

    A recently developed transonic small-disturbance model is used to analyze the interactions of random disturbances with a weak shock. The model equation has an extended form of the classic small-disturbance equation for unsteady transonic aerodynamics. It shows that diffraction effects, nonlinear steepening effects, focusing and caustic effects and random induced vorticity fluctuations interact simultaneously to determine the development of the shock wave in space and time and the pressure field behind it. A finite-difference algorithm to solve the mixed-type elliptic hyperbolic flows around the shock wave is presented. Numerical calculations of shock wave interactions with various deterministic vorticity and temperature disturbances result in complicate shock wave structures and describe peaked as well as rounded pressure signatures behind the shock front, as were recorded in experiments of sonic booms running through atmospheric turbulence.

  11. The expression of Hedgehog genes (Ihh, Dhh) and Hedgehog target genes (Ptc1, Gli1, Coup-TfII) is affected by estrogenic stimuli in the uterus of immature female rats

    SciTech Connect

    Katayama, Seiichi . E-mail: katayama@ankaken.co.jp; Ashizawa, Koji; Gohma, Hiroshi; Fukuhara, Tadahiro; Narumi, Kazunori; Tsuzuki, Yasuhiro; Tatemoto, Hideki; Nakada, Tadashi; Nagai, Kenji

    2006-12-15

    The objective of this study was to investigate the effects of estrogen receptor (ER) agonists and an ER antagonist on the expression of Hedgehog genes (Indian hedgehog: Ihh; Desert hedgehog: Dhh) and Hedgehog target genes (Patched 1: Ptc1; glioma-associated oncogene homolog 1: Gli1; chicken ovalbumin upstream promoter transcription factor II: Coup-TfII) in the rat uterus. Immature female rats were administered once with 17{alpha}-ethynyl estradiol (EE, an ER agonist), propyl pyrazole triole (PPT, an ER{alpha}-selective agonist), diarylpropionitrile (DPN, an ER{beta}-selective agonist), or ICI 182,780 (an ER antagonist). Expression of mRNA for Ihh, Dhh, and Ptc1 was dose-dependently downregulated by EE in the uterus of immature rats, mediated by ER as confirmed by coadministration of ICI 182,780. The mRNA expression levels of Ptc1, Gli1, and Coup-TfII were simultaneously downregulated during the period in which the mRNA expression levels of Ihh and Dhh were downregulated in the uterus after administration of EE. PPT downregulated the transcription of Ihh, Dhh, Ptc1, Gli1, and Coup-TfII, indicating that expression of these genes was regulated by the ER{alpha}-dependent pathway. DPN also downregulated the transcription of Ihh and Dhh, although the effect was weaker than that of PPT, indicating that the regulation of uterine Ihh and Dhh transcription was also affected by the ER{beta}-dependent pathway. These results suggest that the expression of Hedgehog genes (Ihh, Dhh) and Hedgehog target genes (Ptc1, Gli1, Coup-TfII) is affected by estrogenic stimuli in the uterus of immature female rats.

  12. Hedgehog Receptor Patched Is Expressed in a Tissue and Gestation Specific Manner During Early Human and Murine Development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Hedgehog (Hh) signaling pathway is fundamental for appropriate patterning of nearly every organ system in the developing fetus. The Hh receptor, Patched (Ptc), plays a fundamental role in regulating signal transduction in this pathway. Three main 5 splice forms of the Ptc gene (Ptc1B, Ptc1C, and...

  13. Micro-sonicator for spore lysis

    DOEpatents

    Miles, Robin R.; Belgrader, Phillip; Nasarabadi, Shanavaz L.

    2000-01-01

    A micro-sonicator for spore lysis. Using micromachining technology, the micro-sonicator uses ultrasonic excitation of spores to perform spore and cell lysis. The micro-sonicator comprises a container with a cavity therein for retaining the sample in an ultrasonic transmission medium, the cavity being closed by a silicon membrane to which an electrode and piezoelectric material are attached, with the electrode and piezoelectric material being electrically connected to an AC signal generator which causes the membrane to flex and vibrate at the frequency of the applied voltage.

  14. Real-Time, Interactive Sonic Boom Display

    NASA Technical Reports Server (NTRS)

    Haering, Jr., Edward A. (Inventor); Plotkin, Kenneth J. (Inventor)

    2012-01-01

    The present invention is an improved real-time, interactive sonic boom display for aircraft. By using physical properties obtained via various sensors and databases, the invention determines, in real-time, sonic boom impacts locations and intensities for aircraft traveling at supersonic speeds. The information is provided to a pilot via a display that lists a selectable set of maneuvers available to the pilot to mitigate sonic boom issues. Upon selection of a maneuver, the information as to the result of the maneuver is displayed and the pilot may proceed with making the maneuver, or provide new data to the system in order to calculate a different maneuver.

  15. Sonic fatigue of launch vehicle components

    SciTech Connect

    Wentz, K.R.; Camden, M.P.

    1997-01-01

    Wright Laboratory has long been a leader in the technologies required for aerospace structures. One of these driving technology areas is that of the dynamic environments of acoustics and vibration to which structures are exposed and required to survive. This paper presents an overview of sonic fatigue of launch vehicle components. An experimental program to develop sonic fatigue design criteria for a proposed thermal protection system is reviewed. Wright Laboratory{close_quote}s experimental facilities utilized to subject structures to simulated launch vehicle environments which are necessary to generate the experimental data required to provide sonic fatigue design criteria are described. {copyright} {ital 1997 American Institute of Physics.}

  16. Hedgehog Signaling in the Maintenance of Cancer Stem Cells

    PubMed Central

    Cochrane, Catherine R.; Szczepny, Anette; Watkins, D. Neil; Cain, Jason E.

    2015-01-01

    Cancer stem cells (CSCs) represent a rare population of cells with the capacity to self-renew and give rise to heterogeneous cell lineages within a tumour. Whilst the mechanisms underlying the regulation of CSCs are poorly defined, key developmental signaling pathways required for normal stem and progenitor functions have been strongly implicated. Hedgehog (Hh) signaling is an evolutionarily-conserved pathway essential for self-renewal and cell fate determination. Aberrant Hh signaling is associated with the development and progression of various types of cancer and is implicated in multiple aspects of tumourigenesis, including the maintenance of CSCs. Here, we discuss the mounting evidence suggestive of Hh-driven CSCs in the context of haematological malignancies and solid tumours and the novel strategies that hold the potential to block many aspects of the transformation attributed to the CSC phenotype, including chemotherapeutic resistance, relapse and metastasis. PMID:26270676

  17. Cranking in hedgehog models with vector mesons

    NASA Astrophysics Data System (ADS)

    Broniowski, Wojciech; Cohen, Thomas D.

    1986-09-01

    A cranking calculation is performed in a massive SU(2) × SU(2) × U(1) model with valence quarks and the σ, π, ϱ, A and ω mesons. The nucleon moment of inertia, the N-Δ mass splitting and the proton and neutron charge distributions are obtained. The general framework and the specific ansatz for the cranked fields can be used in any hedgehog model with vector mesons. A possible role of η and δ mesons is also discussed.

  18. Hedgehog Signalling in the Embryonic Mouse Thymus

    PubMed Central

    Saldaña, José Ignacio; Crompton, Tessa

    2016-01-01

    T cells develop in the thymus, which provides an essential environment for T cell fate specification, and for the differentiation of multipotent progenitor cells into major histocompatibility complex (MHC)-restricted, non-autoreactive T cells. Here we review the role of the Hedgehog signalling pathway in T cell development, thymic epithelial cell (TEC) development, and thymocyte–TEC cross-talk in the embryonic mouse thymus during the last week of gestation. PMID:27504268

  19. A sonic transducer to detect fluid leaks

    NASA Technical Reports Server (NTRS)

    Cimerman, I.; Janus, J.

    1972-01-01

    Ultrasonic detector utilizes set of contact transducers and bandpass filters to detect and analyze sonic energy produced by flow or leakage. Detector covers wide frequency range and is operable at cryogenic temperatures and in vacuum.

  20. NASA Engineer Larry Cliatt: Softening Sonic Booms

    NASA Video Gallery

    A sudden sonic boom can startle persons unfamiliar with the phenomenon. As a result, supersonic flight over the United States is currently prohibited, except in several restricted testing areas. La...

  1. Influence of Chair Vibrations on Indoor Sonic Boom Annoyance

    NASA Technical Reports Server (NTRS)

    Rathsam, Jonathan; Klos, Jacob; Loubeau, Alexandra

    2015-01-01

    One goal of NASA’s Commercial Supersonic Technology Project is to identify candidate noise metrics suitable for regulating quiet sonic boom aircraft. A suitable metric must consider the short duration and pronounced low frequency content of sonic booms. For indoor listeners, rattle and creaking sounds and floor and chair vibrations may also be important. The current study examined the effect of such vibrations on the annoyance of test subjects seated indoors. The study involved two chairs exposed to nearly identical acoustic levels: one placed directly on the floor, and the other isolated from floor vibrations by pneumatic elastomeric mounts. All subjects experienced both chairs, sitting in one chair for the first half of the experiment and the other chair for the remaining half. Each half of the experiment consisted of 80 impulsive noises played at the exterior of the sonic boom simulator. When all annoyance ratings were analyzed together there appeared to be no difference in mean annoyance with isolation condition. When the apparent effect of transfer bias was removed, a subtle but measurable effect of vibration on annoyance was identified.

  2. Influence of chair vibrations on indoor sonic boom annoyance

    NASA Astrophysics Data System (ADS)

    Rathsam, Jonathan; Klos, Jacob; Loubeau, Alexandra

    2015-10-01

    One goal of NASA's Commercial Supersonic Technology Project is to identify candidate noise metrics suitable for regulating quiet sonic boom aircraft. A suitable metric must consider the short duration and pronounced low frequency content of sonic booms. For indoor listeners, rattle and creaking sounds and floor and chair vibrations may also be important. The current study examined the effect of such vibrations on the annoyance of test subjects seated indoors. The study involved two chairs exposed to nearly identical acoustic levels: one placed directly on the floor, and the other isolated from floor vibrations by pneumatic elastomeric mounts. All subjects experienced both chairs, sitting in one chair for the first half of the experiment and the other chair for the remaining half. Each half of the experiment consisted of 80 impulsive noises played at the exterior of the sonic boom simulator. When all annoyance ratings were analyzed together there appeared to be no difference in mean annoyance with isolation condition. When the apparent effect of transfer bias was removed, a subtle but measurable effect of vibration on annoyance was identified.

  3. Role of Hedgehog Signaling Pathway in NASH

    PubMed Central

    Verdelho Machado, Mariana; Diehl, Anna Mae

    2016-01-01

    Non-alcoholic fatty liver disease (NAFLD) is the number one cause of chronic liver disease in the Western world. Although only a minority of patients will ultimately develop end-stage liver disease, it is not yet possible to efficiently predict who will progress and, most importantly, effective treatments are still unavailable. Better understanding of the pathophysiology of this disease is necessary to improve the clinical management of NAFLD patients. Epidemiological data indicate that NAFLD prognosis is determined by an individual’s response to lipotoxic injury, rather than either the severity of exposure to lipotoxins, or the intensity of liver injury. The liver responds to injury with a synchronized wound-healing response. When this response is abnormal, it leads to pathological scarring, resulting in progressive fibrosis and cirrhosis, rather than repair. The hedgehog pathway is a crucial player in the wound-healing response. In this review, we summarize the pre-clinical and clinical evidence, which demonstrate the role of hedgehog pathway dysregulation in NAFLD pathogenesis, and the preliminary data that place the hedgehog pathway as a potential target for the treatment of this disease. PMID:27258259

  4. Impacts on Dissipative Sonic Vacuum

    NASA Astrophysics Data System (ADS)

    Xu, Yichao; Nesterenko, Vitali

    We investigate the propagating compression bell shape stress waves generated by the strikers with different masses impacting the sonic vacuum - the discrete dissipative strongly nonlinear metamaterial with zero long wave sound speed. The metamaterial is composed of alternating steel disks and Nitrile O-rings. Being a solid material, it has exceptionally low speed of the investigated stress waves in the range of 50 - 74 m/s, which is a few times smaller than the speed of sound or shock waves in air generated by blast. The shape of propagating stress waves was dramatically changed by the viscous dissipation. It prevented the incoming pulses from splitting into trains of solitary waves, a phenomenon characteristic of the non-dissipative strongly nonlinear discrete systems when the striker mass is larger than the cell mass. Both high-speed camera images and numerical simulations demonstrate the unusual rattling behavior of the top disk between the striker and the rest of the system. The linear momentum and energy from the striker were completely transferred to the metamaterial. This strongly nonlinear dissipative metamaterial can be designed for the optimal attenuation of dynamic loads generated by impact or contact explosion. Author 1 wants to acknowledge the support provided by UCSD.

  5. The ciliary Evc/Evc2 complex interacts with Smo and controls Hedgehog pathway activity in chondrocytes by regulating Sufu/Gli3 dissociation and Gli3 trafficking in primary cilia.

    PubMed

    Caparrós-Martín, Jose A; Valencia, María; Reytor, Edel; Pacheco, María; Fernandez, Margarita; Perez-Aytes, Antonio; Gean, Esther; Lapunzina, Pablo; Peters, Heiko; Goodship, Judith A; Ruiz-Perez, Victor L

    2013-01-01

    Hedgehog (Hh) signaling is involved in patterning and morphogenesis of most organs in the developing mammalian embryo. Despite many advances in understanding core components of the pathway, little is known about how the activity of the Hh pathway is adjusted in organ- and tissue-specific developmental processes. Mutations in EVC or EVC2 disrupt Hh signaling in tooth and bone development. Using mouse models, we show here that Evc and Evc2 are mutually required for localizing to primary cilia and also for maintaining their normal protein levels. Consistent with Evc and Evc2 functioning as a complex, the skeletal phenotypes in either single or double homozygous mutant mice are virtually indistinguishable. Smo translocation to the cilium was normal in Evc2-deficient chondrocytes following Hh activation with the Smo-agonist SAG. However, Gli3 recruitment to cilia tips was reduced and Sufu/Gli3 dissociation was impaired. Interestingly, we found Smo to co-precipitate with Evc/Evc2, indicating that in some cells Hh signaling requires direct interaction of Smo with the Evc/Evc2 complex. Expression of a dominantly acting Evc2 mutation previously identified in Weyer's acrodental dysostosis (Evc2Δ43) caused mislocalization of Evc/Evc2Δ43 within the cilium and also reproduced the Gli3-related molecular defects observed in Evc2(-/-) chondrocytes. Moreover, Evc silencing in Sufu(-/-) cells attenuated the output of the Hh pathway, suggesting that Evc/Evc2 also promote Hh signaling in the absence of Sufu. Together our data reveal that the Hh pathway involves Evc/Evc2-dependent modulations that are necessary for normal endochondral bone formation.

  6. Phase I ResonantSonic CRADA report

    SciTech Connect

    Richterich, L.R.; Amos, L.O.; Fancher, J.D.; McLellan, G.W.; Setzer, W.V.; Tuttle, B.G.; Hockey, R.L.; Ferris, R.H.; Riechers, D.M.; Pitman, S.G.

    1994-03-28

    This test report describes the Phase 1 testing and results of the ResonantSonic drilling method. This effort was conducted as part of a Cooperative Research and Development Agreement (CRADA) between the US Department of Energy, Pacific Northwest Laboratory, Westinghouse Hanford Company and Water Development Corporation. The purpose of this demonstration was to evaluate the Water Development Corporation`s ResonantSonic drilling system, modify components as necessary and determine compatible drilling applications for the ResonantSonic drilling method for use at facilities in the DOE complex and private industry. Initially, the ResonantSonic drill was used to drill several test holes at the Drilling Technology Test Site to assess the feasibility of drilling vertical and angle holes. After this initial phase, a 45 degree angle vapor extraction well was drilled to a depth of 168 feet at the 200 West Carbon Tetrachloride Site. This well was drilled and completed in nine days. Extensive geologic and vapor sampling were conducted while drilling this well. In addition, testing was also conducted at the test site to evaluated drilling with larger diameter casing (8 5/8 inch). Evaluation of the Resonant Sonic drilling method will be continued during the Phase 2 portion of testing to determine if improvements to the ResonantSonic system will make it a more viable method for drilling and sampling.

  7. Pitchfork and Gprasp2 Target Smoothened to the Primary Cilium for Hedgehog Pathway Activation

    PubMed Central

    Jung, Bomi; Padula, Daniela; Burtscher, Ingo; Landerer, Cedric; Lutter, Dominik; Theis, Fabian; Messias, Ana C.; Geerlof, Arie; Sattler, Michael; Kremmer, Elisabeth; Boldt, Karsten; Ueffing, Marius; Lickert, Heiko

    2016-01-01

    The seven-transmembrane receptor Smoothened (Smo) activates all Hedgehog (Hh) signaling by translocation into the primary cilia (PC), but how this is regulated is not well understood. Here we show that Pitchfork (Pifo) and the G protein-coupled receptor associated sorting protein 2 (Gprasp2) are essential components of an Hh induced ciliary targeting complex able to regulate Smo translocation to the PC. Depletion of Pifo or Gprasp2 leads to failure of Smo translocation to the PC and lack of Hh target gene activation. Together, our results identify a novel protein complex that is regulated by Hh signaling and required for Smo ciliary trafficking and Hh pathway activation. PMID:26901434

  8. Pitchfork and Gprasp2 Target Smoothened to the Primary Cilium for Hedgehog Pathway Activation.

    PubMed

    Jung, Bomi; Padula, Daniela; Burtscher, Ingo; Landerer, Cedric; Lutter, Dominik; Theis, Fabian; Messias, Ana C; Geerlof, Arie; Sattler, Michael; Kremmer, Elisabeth; Boldt, Karsten; Ueffing, Marius; Lickert, Heiko

    2016-01-01

    The seven-transmembrane receptor Smoothened (Smo) activates all Hedgehog (Hh) signaling by translocation into the primary cilia (PC), but how this is regulated is not well understood. Here we show that Pitchfork (Pifo) and the G protein-coupled receptor associated sorting protein 2 (Gprasp2) are essential components of an Hh induced ciliary targeting complex able to regulate Smo translocation to the PC. Depletion of Pifo or Gprasp2 leads to failure of Smo translocation to the PC and lack of Hh target gene activation. Together, our results identify a novel protein complex that is regulated by Hh signaling and required for Smo ciliary trafficking and Hh pathway activation. PMID:26901434

  9. Cloning of zebrafish nkx6.2 and a comprehensive analysis of the conserved transcriptional response to Hedgehog/Gli signaling in the zebrafish neural tube.

    PubMed

    Guner, Burcu; Karlstrom, Rolf O

    2007-04-01

    Sonic Hedgehog (Shh) signaling helps pattern the vertebrate neural tube, in part by regulating the dorsal/ventral expression of a number of homeodomain containing transcription factors. These Hh responsive genes have been divided into two classes, with Class II genes being activated by Hh signaling and Class I genes being repressed by Hh signaling. While the transcriptional response to varying Hh levels is well defined in chick and mouse, it is only partially described in zebrafish, despite the fact that zebrafish has emerged as a powerful genetic system for the study of neural patterning. To better characterize the Hh response in the zebrafish neural tube, we cloned the zebrafish Class II Hh target genes nkx2.9 and nkx6.2. We then analyzed the expression of a number of Class I and Class II Hh responsive genes in wild type, Hh mutant, and Hh over-expressing zebrafish embryos. We show that expression of Class I and Class II genes is highly conserved in the vertebrate neural tube. Further, ventral-most Class II gene expression was completely lost in all Hh pathway mutants analyzed, indicating high levels of Hh signaling are blocked in all of these mutants. In contrast, more dorsally expressed genes were variably affected in different Hh pathway mutants, indicating mid-levels of Hh signaling are differentially affected. This comprehensive expression study provides an important tool for the characterization of Hh signaling in zebrafish and provides a sensitive assay for determining the degree to which newly identified zebrafish mutants affect Hh signaling.

  10. Heparan Sulfate Regulates Hair Follicle and Sebaceous Gland Morphogenesis and Homeostasis*

    PubMed Central

    Coulson-Thomas, Vivien Jane; Gesteira, Tarsis Ferreira; Esko, Jeffrey; Kao, Winston

    2014-01-01

    Hair follicle (HF) morphogenesis and cycling are a result of intricate autonomous epithelial-mesenchymal interactions. Once the first HF cycle is complete it repeatedly undergoes cyclic transformations. Heparan sulfate (HS) proteoglycans are found on the cell surface and in the extracellular matrix where they influence a variety of biological processes by interacting with physiologically important proteins, such as growth factors. Inhibition of heparanase (an HS endoglycosidase) in in vitro cultured HFs has been shown to induce a catagen-like process. Therefore, this study aimed to elucidate the precise role of HS in HF morphogenesis and cycling. An inducible tetratransgenic mouse model was generated to excise exostosin glycosyltransferase 1 (Ext1) in keratin 14-positive cells from P21. Interestingly, EXT1StEpiΔ/StEpiΔ mice presented solely anagen HFs. Moreover, waxing the fur to synchronize the HFs revealed accelerated hair regrowth in the EXT1StEpiΔ/StEpiΔ mice and hindered cycling into catagen. The ablation of HS in the interfollicular epidermal cells of mature skin led to the spontaneous formation of new HFs and an increase in Sonic Hedgehog expression resembling wild-type mice at P0, thereby indicating that the HS/Sonic Hedgehog signaling pathway regulates HF formation during embryogenesis and prevents HF formation in mature skin. Finally, the knock-out of HS also led to the morphogenesis and hyperplasia of sebaceous glands and sweat glands in mature mice, leading to exacerbated sebum production and accumulation on the skin surface. Therefore, our findings clearly show that an intricate control of HS levels is required for HF, sebaceous gland, and sweat gland morphogenesis and HF cycling. PMID:25053416

  11. ZnRF3 Induces Apoptosis of Gastric Cancer Cells by Antagonizing Wnt and Hedgehog Signaling.

    PubMed

    Qin, Hongzhen; Cai, Aizhen; Xi, Hongqing; Yuan, Jing; Chen, Lin

    2015-11-01

    A large proportion of malignant cancers of the stomach are gastric adenocarcinoma type. In spite of many studies, the molecular basis for this cancer is still unclear. Deregulated cell proliferative signaling via Wnt/β-catenin and Hedgehog pathways is considered important in the pathogenesis of many cancers including the gastric cancer. Recent studies identified ZnRF3 protein, which is a E3-ubiquitin ligase and which is either deleted or mutated in cancers, to inhibit Wnt signaling. However, the significance of ZnRF3 in the control of gastric cancer and whether it also regulates Hedgehog signaling pathway, is not known. In the present study, we assessed the expression of ZnRF3 in gastric tumors and paracancerous tissues from 58 patients (44 male and 14 female) of different ages and related this to patient survival. We observed a clear relationship between ZnRF3 expression in paracancerous tissue and tumor size. Also, ZnRF3 expression was much higher in tumors from aged patients. Male patients showed higher mortality than the females. Mechanistic studies using normal gastric cells (GES1) and gastric cancer cells (MGC-803) infected with either AdZnRF3 or AdGFP viral vectors, revealed that ZnRF3 overexpression causes significantly more apoptosis and lowered proliferation of cancer cells. ZnRF3 overexpression led to greatly reduced levels of Lgr5, a component of Wnt signaling and also Gli1, a component of Hedgehog signaling. Thus, ZnRF3 negatively influences both the Wnt and Hedgehog proliferative pathways, and probably this way it negatively regulates cancer progression. These results suggest the importance of normal ZnRF3 function in checking the progression of cancer cell growth and indicate that a lack of this protein can lead to poorer clinical outcomes for gastric cancer patients. PMID:27352324

  12. [Hedgehog signaling in the pathogenesis of neuro-oncology diseases].

    PubMed

    Cherepanov, S A; Baklaushev, V P; Gabashvili, A N; Shepeleva, I I; Chekhonin, V P

    2015-01-01

    The review summarizes current knowledge on the Hedgehog signaling pathway, its role in normal embryogenesis and/or initiation and progression of neuro-oncological diseases, especially of high-grade gliomas, the most malignant neuroepithelial tumors. The main proteins forming the Hedgehog signaling pathway include Shh, PTCH1, SMO, HHIP, SUFU and GLI1 isoforms. Effects of other signaling pathways on the family of transcription factors GLI and other proteins are described. The review summarizes modern data about the impact of the Hedgehog signaling pathway on proliferation, migration activity and invasiveness, and also on tumor neoangiogenesis and tumor cell chemoresistance. The role of the Hedgehog signaling pathway in origin of cancer stem cells and epithelial-mesenchymal transition is also analyzed. Some prospects for new anticancer drugs acting on components of the Hedgehog signaling pathway inhibitors are demonstrated. PMID:26215410

  13. ENVIRONMENTAL TECHNOLOGY VERIFICATION REPORT; ULTRASONIC AQUEOUS CLEANING SYSTEMS, SMART SONIC CORPORATION, SMART SONIC

    EPA Science Inventory

    This report is a product of the U.S. EPA's Environmental Technoloy Verification (ETV) Program and is focused on the Smart Sonics Ultrasonic Aqueous Cleaning Systems. The verification is based on three main objectives. (1) The Smart Sonic Aqueous Cleaning Systems, Model 2000 and...

  14. Drosophila genome-wide obesity screen reveals hedgehog as a determinant of brown versus white adipose cell fate.

    PubMed

    Pospisilik, J Andrew; Schramek, Daniel; Schnidar, Harald; Cronin, Shane J F; Nehme, Nadine T; Zhang, Xiaoyun; Knauf, Claude; Cani, Patrice D; Aumayr, Karin; Todoric, Jelena; Bayer, Martina; Haschemi, Arvand; Puviindran, Vijitha; Tar, Krisztina; Orthofer, Michael; Neely, G Gregory; Dietzl, Georg; Manoukian, Armen; Funovics, Martin; Prager, Gerhard; Wagner, Oswald; Ferrandon, Dominique; Aberger, Fritz; Hui, Chi-chung; Esterbauer, Harald; Penninger, Josef M

    2010-01-01

    Over 1 billion people are estimated to be overweight, placing them at risk for diabetes, cardiovascular disease, and cancer. We performed a systems-level genetic dissection of adiposity regulation using genome-wide RNAi screening in adult Drosophila. As a follow-up, the resulting approximately 500 candidate obesity genes were functionally classified using muscle-, oenocyte-, fat-body-, and neuronal-specific knockdown in vivo and revealed hedgehog signaling as the top-scoring fat-body-specific pathway. To extrapolate these findings into mammals, we generated fat-specific hedgehog-activation mutant mice. Intriguingly, these mice displayed near total loss of white, but not brown, fat compartments. Mechanistically, activation of hedgehog signaling irreversibly blocked differentiation of white adipocytes through direct, coordinate modulation of early adipogenic factors. These findings identify a role for hedgehog signaling in white/brown adipocyte determination and link in vivo RNAi-based scanning of the Drosophila genome to regulation of adipocyte cell fate in mammals.

  15. Production Well Performance Enhancement using Sonication Technology

    SciTech Connect

    Adewumi, Michael A; Ityokumbul, M Thaddeus; Watson, Robert W; Eltohami, Eltohami; Farias, Mario; Heckman, Glenn; Houlihan, Brendan; Karoor, Samata Prakash; Miller, Bruce G; Mohammed, Nazia; Olanrewaju, Johnson; Ozdemir, Mine; Rejepov, Dautmamed; Sadegh, Abdallah A; Quammie, Kevin E; Zaghloul, Jose; Hughes, W Jack; Montgomery, Thomas C

    2005-12-31

    The objective of this project was to develop a sonic well performance enhancement technology that focused on near wellbore formation damage. In order to successfully achieve this objective, a three-year project was defined. The entire project was broken into four tasks. The overall objective of all this was to foster a better understanding of the mechanisms involved in sonic energy interactions with fluid flow in porous media and adapt such knowledge for field applications. The fours tasks are: • Laboratory studies • Mathematical modeling • Sonic tool design and development • Field demonstration The project was designed to be completed in three years; however, due to budget cuts, support was only provided for the first year, and hence the full objective of the project could not be accomplished. This report summarizes what was accomplished with the support provided by the US Department of Energy. Experiments performed focused on determining the inception of cavitation, studying thermal dissipation under cavitation conditions, investigating sonic energy interactions with glass beads and oil, and studying the effects of sonication on crude oil properties. Our findings show that the voltage threshold for onset of cavitation is independent of transducer-hydrophone separation distance. In addition, thermal dissipation under cavitation conditions contributed to the mobilization of deposited paraffins and waxes. Our preliminary laboratory experiments suggest that waxes are mobilized when the fluid temperature approaches 40°C. Experiments were conducted that provided insights into the interactions between sonic wave and the fluid contained in the porous media. Most of these studies were carried out in a slim-tube apparatus. A numerical model was developed for simulating the effect of sonication in the nearwellbore region. The numerical model developed was validated using a number of standard testbed problems. However, actual application of the model for scale

  16. Realism Assessment of Sonic Boom Simulators

    NASA Technical Reports Server (NTRS)

    Sullivan, Brenda M.; Davies, Patrica; Hodgdon, Kthleen K.; Salamone, Joseph A., III; Pilon, Anthony

    2008-01-01

    Developments in small supersonic aircraft design are predicted to result in low-intensity sonic booms. Booms generated by current aircraft are similar to those that led to the ban on commercial supersonic fli ght over the US, so are unsuitable for parametric studies of psychoac oustic response to low-intensity booms. Therefore, simulators have be en used to study the impact of predicted low-intensity sonic booms. H owever, simulators have been criticized because, when simulating conv entional-level booms, the sounds were observed to be unrealistic by p eople experienced in listening to sonic booms. Thus, two studies were conducted to measure the perceived realism of three sonic boom simul ators. Experienced listeners rated the realism of conventional sonic boom signatures when played in these simulators. The effects on percei ved realism of factors such as duration of post-boom noise, exclusion of very low frequency components, inclusion of ground reflections, a nd type of simulator were examined. Duration of post-boom noise was f ound to have a strong effect on perceived realism, while type of simu lator had a weak effect. It was determined that post-boom noise had t o be at least 1.5 seconds long for the sound to be rated very realist ic. Loudness level did not affect realism for the range of sounds pla yed in the tests (80-93 dB ASEL).

  17. T-box3 is a ciliary protein and regulates stability of the Gli3 transcription factor to control digit number

    PubMed Central

    Emechebe, Uchenna; Kumar P, Pavan; Rozenberg, Julian M; Moore, Bryn; Firment, Ashley; Mirshahi, Tooraj; Moon, Anne M

    2016-01-01

    Crucial roles for T-box3 in development are evident by severe limb malformations and other birth defects caused by T-box3 mutations in humans. Mechanisms whereby T-box3 regulates limb development are poorly understood. We discovered requirements for T-box at multiple stages of mouse limb development and distinct molecular functions in different tissue compartments. Early loss of T-box3 disrupts limb initiation, causing limb defects that phenocopy Sonic Hedgehog (Shh) mutants. Later ablation of T-box3 in posterior limb mesenchyme causes digit loss. In contrast, loss of anterior T-box3 results in preaxial polydactyly, as seen with dysfunction of primary cilia or Gli3-repressor. Remarkably, T-box3 is present in primary cilia where it colocalizes with Gli3. T-box3 interacts with Kif7 and is required for normal stoichiometry and function of a Kif7/Sufu complex that regulates Gli3 stability and processing. Thus, T-box3 controls digit number upstream of Shh-dependent (posterior mesenchyme) and Shh-independent, cilium-based (anterior mesenchyme) Hedgehog pathway function. DOI: http://dx.doi.org/10.7554/eLife.07897.001 PMID:27046536

  18. T-box3 is a ciliary protein and regulates stability of the Gli3 transcription factor to control digit number.

    PubMed

    Emechebe, Uchenna; Kumar P, Pavan; Rozenberg, Julian M; Moore, Bryn; Firment, Ashley; Mirshahi, Tooraj; Moon, Anne M

    2016-01-01

    Crucial roles for T-box3 in development are evident by severe limb malformations and other birth defects caused by T-box3 mutations in humans. Mechanisms whereby T-box3 regulates limb development are poorly understood. We discovered requirements for T-box at multiple stages of mouse limb development and distinct molecular functions in different tissue compartments. Early loss of T-box3 disrupts limb initiation, causing limb defects that phenocopy Sonic Hedgehog (Shh) mutants. Later ablation of T-box3 in posterior limb mesenchyme causes digit loss. In contrast, loss of anterior T-box3 results in preaxial polydactyly, as seen with dysfunction of primary cilia or Gli3-repressor. Remarkably, T-box3 is present in primary cilia where it colocalizes with Gli3. T-box3 interacts with Kif7 and is required for normal stoichiometry and function of a Kif7/Sufu complex that regulates Gli3 stability and processing. Thus, T-box3 controls digit number upstream of Shh-dependent (posterior mesenchyme) and Shh-independent, cilium-based (anterior mesenchyme) Hedgehog pathway function. PMID:27046536

  19. TEST & EVALUATION REPORT FOR THE HEDGEHOG-II PACKAGING SYSTEMS DOT-7A TYPE A CONTAINER

    SciTech Connect

    KELLY, D.L.

    2003-12-29

    This report documents the US. Department of Transportation Specification 7A (DOT-7A) Type A compliance test and evaluation results for the Hedgehog-II packaging systems. The approved Hedgehog-II packaging configurations provide primary and secondary containment. The approved packaging configurations described within this report are designed to ship Type A quantities of radioactive materials, normal form. Contents may be in solid or liquid form. Liquids transported in the approved 1 L glass bottle assembly shall have a specific gravity of less than or equal to 1.6. Liquids transported in all other approved configurations shall have a specific gravity of less than or equal to 2.0. The solid contents, including packaging, are limited in weight to the gross weight of the as-tested liquids and bottles. The approved Hedgehog-II packaging configurations described in this report may be transported by air, and have been evaluated as meeting the applicable International Air Transport Association/International Civil Aviation Organization (IATA/ICAO) Dangerous Goods Regulations in addition to the DOT requirements.

  20. Disturbance of sleep by sonic booms.

    PubMed

    Griefahn, B; Jansen, G

    1975-05-01

    After a pilot study (2 subjects, 19 nights) we tested two different subjects during 57 nights, administering sonic booms (1 mb, 300 ms; sound level of sonic boom in the bedroom 80-85 dB (A) and recording EEG and peripheral blood volume. After 7 nights without noise, 30 nights with either 2 or 4 sonic booms (alternately) were applied. After 10 more nights without noise, four nights with 8 and 16 bangs followed alternately. The last 6 nights were used as a comparison phase. Results showed that distrubance was obvious during all periods of noise. No adaptation could be observed during any of the experiments. On the contrary, during the night with 4 bangs there was a tendency for compensation, e.g., in the last two thirds of nights with 4 bangs, the total time of deep sleep was comparable with the nights without any noise. PMID:1145178

  1. Hawking radiation in sonic black holes.

    PubMed

    Giovanazzi, S

    2005-02-18

    I present a microscopic description of Hawking radiation in sonic black holes. A one-dimensional Fermi-degenerate liquid squeezed by a smooth barrier forms a transonic flow, a sonic analog of a black hole. The quantum treatment of the noninteracting case establishes a close relationship between sonic Hawking radiation and quantum tunneling through the barrier. Quasiparticle excitations appear at the barrier and are then radiated with a thermal distribution in exact agreement with Hawking's formula. The signature of the radiation can be found in the dynamic structure factor, which can be measured in a scattering experiment. The possibility for experimental verification of this new transport phenomenon for ultracold atoms is discussed. PMID:15783717

  2. Sonic Thermometer for High-Altitude Balloons

    NASA Technical Reports Server (NTRS)

    Bognar, John

    2012-01-01

    The sonic thermometer is a specialized application of well-known sonic anemometer technology. Adaptations have been made to the circuit, including the addition of supporting sensors, which enable its use in the high-altitude environment and in non-air gas mixtures. There is a need to measure gas temperatures inside and outside of superpressure balloons that are flown at high altitudes. These measurements will allow the performance of the balloon to be modeled more accurately, leading to better flight performance. Small thermistors (solid-state temperature sensors) have been used for this general purpose, and for temperature measurements on radiosondes. A disadvantage to thermistors and other physical (as distinct from sonic) temperature sensors is that they are subject to solar heating errors when they are exposed to the Sun, and this leads to issues with their use in a very high-altitude environment

  3. A Flight Research Overview of WSPR, a Pilot Project for Sonic Boom Community Response

    NASA Technical Reports Server (NTRS)

    Cliatt, Larry James; Haering, Ed; Jones, Thomas P.; Waggoner, Erin R.; Flattery, Ashley K.; Wiley, Scott L.

    2014-01-01

    In support of NASAs ongoing effort to bring supersonic commercial travel to the public, NASA Dryden Flight Research Center and NASA Langley Research Center, in cooperation with other industry organizations, conducted a flight research experiment to identify the methods, tools, and best practices for a large-scale quiet (or low) sonic boom community human response test. The name of the effort was Waveforms and Sonic boom Perception and Response. Such tests will go towards building a dataset that governing agencies like the Federal Aviation Administration and International Civil Aviation Organization will use to establish regulations for acceptable sound levels of overland sonic booms. Until WSPR, there had never been an effort that studied the response of people in their own homes and performing daily activities to non-traditional, low sonic booms.WSPR was a NASA collaborative effort with several industry partners, in response to a NASA Aeronautics Research Mission Directorate Research Opportunities in Aeronautics. The primary contractor was Wyle. Other partners included Gulfstream Aerospace Corporation, Pennsylvania State University, Tetra Tech, and Fidell Associates, Inc.A major objective of the effort included exposing a community with the sonic boom magnitudes and occurrences expected in high-air traffic regions with a network of supersonic commercial aircraft in place. Low-level sonic booms designed to simulate those produced by the next generation of commercial supersonic aircraft were generated over a small residential community. The sonic boom footprint was recorded with an autonomous wireless microphone array that spanned the entire community. Human response data was collected using multiple survey methods. The research focused on essential elements of community response testing including subject recruitment, survey methods, instrumentation systems, flight planning and operations, and data analysis methods.This paper focuses on NASAs role in the efforts

  4. A Flight Research Overview of WSPR, a Pilot Project for Sonic Boom Community Response

    NASA Technical Reports Server (NTRS)

    Cliatt, Larry J., II; Haering, Edward A., Jr.; Jones, Thomas P.; Waggoner, Erin R.; Flattery, Ashley K.; Wiley, Scott L.

    2014-01-01

    In support of the ongoing effort by the National Aeronautics and Space Administration (NASA) to bring supersonic commercial travel to the public, the NASA Armstrong Flight Research Center and the NASA Langley Research Center, in cooperation with other industry organizations, conducted a flight research experiment to identify the methods, tools, and best practices for a large-scale quiet (or low) sonic boom community human response test. The name of the effort was Waveforms and Sonic boom Perception and Response (WSPR). Such tests will be applied to building a dataset that governing agencies such as the Federal Aviation Administration and the International Civil Aviation Organization will use to establish regulations for acceptable sound levels of overland sonic booms. The WSPR test was the first such effort that studied responses to non-traditional low sonic booms while the subject persons were in their own homes and performing daily activities.The WSPR test was a NASA collaborative effort with several industry partners, in response to a NASA Aeronautics Research Mission Directorate Research Opportunities in Aeronautics. The primary contractor was Wyle (El Segundo, California). Other partners included Gulfstream Aerospace Corporation (Savannah, Georgia); Pennsylvania State University (University Park, Pennsylvania); Tetra Tech, Inc. (Pasadena, California); and Fidell Associates, Inc. (Woodland Hills, California).A major objective of the effort included exposing a community to the sonic boom magnitudes and occurrences that would be expected to occur in high-air traffic regions having a network of supersonic commercial aircraft in place. Low-level sonic booms designed to simulate those produced by the next generation of commercial supersonic aircraft were generated over a small residential community. The sonic boom footprint was recorded with an autonomous wireless microphone array that spanned the entire community. Human response data were collected using multiple

  5. Hedgehog Related Protein Expression in Breast Cancer: Gli-2 Is Associated with Poor Overall Survival

    PubMed Central

    Im, Soyoung; Yoo, Changyoung; Jung, Ji-Han; Jeon, Ye-Won; Suh, Young Jin; Kang, Chang Suk

    2013-01-01

    Background The hedgehog (Hh) signaling pathway is known to play a critical role in various malignancies, but its clinicopathologic role in breast cancer is yet to be established. Methods Tissue microarray blocks from 334 cases of breast cancer were prepared. The expression of six Hh signaling proteins including sonic hedgehog (Shh), patched (Ptch), smoothened (Smo), and the glioma-associated oncogene (Gli)-1, Gli-2, and Gli-3 were analyzed immunohistochemically. Results The expression of Hh signaling proteins was significantly correlated with some prognostic factors including the correlation of lymph node metastasis with the expression of Shh (p=0.001) and Ptch (p=0.064), the correlation of the stages with Shh and Gli-3 expression (p=0.007 and p=0.024, respectively), the correlation of the nuclear grade with the Smo (p=0.004) and Gli-3 (p=0.000), and the correlation of the histologic grade with the Ptch (p=0.016), Smo (p=0.007), and Gli-3 (p=0.000). The Shh, Ptch, Smo, Gli-1, and Gli-2 expression was significantly different between the phenotypes (p=0.000, p=0.001, p=0.004, p=0.039, and p=0.031, respectively). Gli-2 expression was correlated with a worse overall survival outcome (p=0.012). Conclusions Hh pathway activation is correlated with a more aggressive clinical behavior in breast carcinomas. The comparison of phenotypes suggested that the Hh pathway may be a useful therapeutic target for breast carcinoma. Patients with Gli-2 expression had a significantly lower overall survival rate and, therefore, it showed promise as a prognostic marker. PMID:23667370

  6. Development of the sonic pump levitator

    NASA Technical Reports Server (NTRS)

    Dunn, S. A.

    1985-01-01

    The process and mechanism involved in producing glass microballoons (GMBs) of acceptable quality for laser triggered inertial fusion through use of glass jet levitation and manipulation are considered. The gas jet levitation device, called sonic pumps, provides positioning by timely and appropriate application of gas mementum from one or more of six sonic pumps which are arranged orthogonally in opposed pairs about the levitation region and are activated by an electrooptical, computer controlled, feedback system. The levitation device was fabricated and its associated control systems were assembled into a package and tested in reduced gravity flight regime of the NASA KC-135 aircraft.

  7. Status and capabilities of sonic boom simulators

    NASA Technical Reports Server (NTRS)

    Shepherd, K. P.; Powell, C. A.

    1986-01-01

    The current status and capabilities of sonic boom simulators which might be used in future studies of the effects of sonic boom on people, animals, or structures is summarized. The list of candidate simulators is based on a literature search which was confined to the United States and Canada. Some of the simulators are fully operational, others could be made operational with a modest investment, and still others would require a major investment. For the sake of the completeness, some simulators which were the subject of a previous review, but which no longer exist, are also included herein.

  8. Lhx6 and Lhx8: cell fate regulators and beyond

    PubMed Central

    Zhou, Chen; Yang, Guodong; Chen, Mo; He, Ling; Xiang, Lusai; Ricupero, Christopher; Mao, Jeremy J.; Ling, Junqi

    2015-01-01

    As transcription factors of the lines (LIN)-11/Islet (Isl)-1/mitosis entry checkpoint (MEC)-3 (LIM)-homeobox subfamily, LIM homeobox (Lhx)6 and -8 are remarkably conserved and involved in the morphogenesis of multiple organ systems. Lhx6 and -8 play overlapping and distinctive roles, but in general act as cell fate mediators and in turn are regulated by several transcriptional factors, such as sonic hedgehog, fibroblast growth factors, and wingless-int (Wnt)/β-catenin. In this review, we first summarize Lhx6 and -8 distributions in development and then explore how Lhx6 and -8 act as transcription factors and coregulators of cell lineage specification. Known Lhx6 and -8 functions and targets are outlined in neurogenesis, craniofacial development, and germ cell differentiation. The underlying mechanisms of Lhx6 and -8 in regulating cell fate remain elusive. Whether Lhx6 and -8 affect functions in tissues and organs other than neural, craniofacial, oocytes, and germ cells is largely unexplored. Taken together, Lhx6 and -8 are important regulators of cell lineage specification and may act as one of the pivotal mediators of stem cell fate. Undoubtedly, future investigations of Lhx6 and -8 biology will continue to yield fascinating insights into tissue development and homeostasis, in addition to their putative roles in tissue regeneration and ageing.—Zhou, C., Yang, G., Chen, M., He, L., Xiang, L., Ricupero, C., Mao, J. J., Ling, J. Lhx6 and Lhx8: cell fate regulators and beyond. PMID:26148970

  9. Foxp2 regulates neuronal differentiation and neuronal subtype specification.

    PubMed

    Chiu, Yi-Chi; Li, Ming-Yang; Liu, Yuan-Hsuan; Ding, Jing-Ya; Yu, Jenn-Yah; Wang, Tsu-Wei

    2014-07-01

    Mutations of the transcription factor FOXP2 in humans cause a severe speech and language disorder. Disruption of Foxp2 in songbirds or mice also leads to deficits in song learning or ultrasonic vocalization, respectively. These data suggest that Foxp2 plays important roles in the developing nervous system. However, the mechanism of Foxp2 in regulating neural development remains elusive. In the current study, we found that Foxp2 increased neuronal differentiation without affecting cell proliferation and cell survival in primary neural progenitors from embryonic forebrains. Foxp2 induced the expression of platelet-derived growth factor receptor α, which mediated the neurognic effect of Foxp2. In addition, Foxp2 positively regulated the differentiation of medium spiny neurons derived from the lateral ganglionic eminence and negatively regulated the formation of interneurons derived from dorsal medial ganglionic eminence by interacting with the Sonic hedgehog pathway. Taken together, our results suggest that Foxp2 regulates multiple aspects of neuronal development in the embryonic forebrain.

  10. An ancient yet flexible cis-regulatory architecture allows localized Hedgehog tuning by patched/Ptch1.

    PubMed

    Lorberbaum, David S; Ramos, Andrea I; Peterson, Kevin A; Carpenter, Brandon S; Parker, David S; De, Sandip; Hillers, Lauren E; Blake, Victoria M; Nishi, Yuichi; McFarlane, Matthew R; Chiang, Ason Cy; Kassis, Judith A; Allen, Benjamin L; McMahon, Andrew P; Barolo, Scott

    2016-01-01

    The Hedgehog signaling pathway is part of the ancient developmental-evolutionary animal toolkit. Frequently co-opted to pattern new structures, the pathway is conserved among eumetazoans yet flexible and pleiotropic in its effects. The Hedgehog receptor, Patched, is transcriptionally activated by Hedgehog, providing essential negative feedback in all tissues. Our locus-wide dissections of the cis-regulatory landscapes of fly patched and mouse Ptch1 reveal abundant, diverse enhancers with stage- and tissue-specific expression patterns. The seemingly simple, constitutive Hedgehog response of patched/Ptch1 is driven by a complex regulatory architecture, with batteries of context-specific enhancers engaged in promoter-specific interactions to tune signaling individually in each tissue, without disturbing patterning elsewhere. This structure-one of the oldest cis-regulatory features discovered in animal genomes-explains how patched/Ptch1 can drive dramatic adaptations in animal morphology while maintaining its essential core function. It may also suggest a general model for the evolutionary flexibility of conserved regulators and pathways. PMID:27146892

  11. An ancient yet flexible cis-regulatory architecture allows localized Hedgehog tuning by patched/Ptch1

    PubMed Central

    Lorberbaum, David S; Ramos, Andrea I; Peterson, Kevin A; Carpenter, Brandon S; Parker, David S; De, Sandip; Hillers, Lauren E; Blake, Victoria M; Nishi, Yuichi; McFarlane, Matthew R; Chiang, Ason CY; Kassis, Judith A; Allen, Benjamin L; McMahon, Andrew P; Barolo, Scott

    2016-01-01

    The Hedgehog signaling pathway is part of the ancient developmental-evolutionary animal toolkit. Frequently co-opted to pattern new structures, the pathway is conserved among eumetazoans yet flexible and pleiotropic in its effects. The Hedgehog receptor, Patched, is transcriptionally activated by Hedgehog, providing essential negative feedback in all tissues. Our locus-wide dissections of the cis-regulatory landscapes of fly patched and mouse Ptch1 reveal abundant, diverse enhancers with stage- and tissue-specific expression patterns. The seemingly simple, constitutive Hedgehog response of patched/Ptch1 is driven by a complex regulatory architecture, with batteries of context-specific enhancers engaged in promoter-specific interactions to tune signaling individually in each tissue, without disturbing patterning elsewhere. This structure—one of the oldest cis-regulatory features discovered in animal genomes—explains how patched/Ptch1 can drive dramatic adaptations in animal morphology while maintaining its essential core function. It may also suggest a general model for the evolutionary flexibility of conserved regulators and pathways. DOI: http://dx.doi.org/10.7554/eLife.13550.001 PMID:27146892

  12. Particle-Molecule Collection by Sonic Flow Impingers

    ERIC Educational Resources Information Center

    Jackson, Melbourne L.

    1974-01-01

    The theoretical basis of the sonic-flow impinger is discussed. Details are given for the design, prediction of performance, preliminary evaluation for particle collection, and field use of a sonic-flow impinger train. (DT)

  13. NASA Studies Sonic Booms' Effect on Big Structures

    NASA Video Gallery

    NASA recently conducted flight experiments at Edwards Air Force Base in Southern California to examine the effect of low-amplitude sonic booms on large office buildings. As part of the Sonic Booms ...

  14. Zfx facilitates tumorigenesis caused by activation of the Hedgehog pathway.

    PubMed

    Palmer, Colin J; Galan-Caridad, Jose M; Weisberg, Stuart P; Lei, Liang; Esquilin, Jose M; Croft, Gist F; Wainwright, Brandon; Canoll, Peter; Owens, David M; Reizis, Boris

    2014-10-15

    The Hedgehog (Hh) signaling pathway regulates normal development and cell proliferation in metazoan organisms, but its aberrant activation can promote tumorigenesis. Hh-induced tumors arise from various tissues and they may be indolent or aggressive, as is the case with skin basal cell carcinoma (BCC) or cerebellar medulloblastoma, respectively. Little is known about common cell-intrinsic factors that control the development of such diverse Hh-dependent tumors. Transcription factor Zfx is required for the self-renewal of hematopoietic and embryonic stem cells, as well as for the propagation of acute myeloid and T-lymphoblastic leukemias. We report here that Zfx facilitates the development of experimental BCC and medulloblastoma in mice initiated by deletion of the Hh inhibitory receptor Ptch1. Simultaneous deletion of Zfx along with Ptch1 prevented BCC formation and delayed medulloblastoma development. In contrast, Zfx was dispensable for tumorigenesis in a mouse model of glioblastoma. We used genome-wide expression and chromatin-binding analysis in a human medulloblastoma cell line to characterize direct, evolutionarily conserved targets of Zfx, identifying Dis3L and Ube2j1 as two targets required for the growth of the human medulloblastoma cells. Our results establish Zfx as a common cell-intrinsic regulator of diverse Hh-induced tumors, with implications for the definition of new therapeutic targets in these malignancies. PMID:25164012

  15. Skinny hedgehog, an acyltransferase required for palmitoylation and activity of the hedgehog signal.

    PubMed

    Chamoun, Z; Mann, R K; Nellen, D; von Kessler, D P; Bellotto, M; Beachy, P A; Basler, K

    2001-09-14

    One of the most dominant influences in the patterning of multicellular embryos is exerted by the Hedgehog (Hh) family of secreted signaling proteins. Here, we identify a segment polarity gene in Drosophila melanogaster, skinny hedgehog (ski), and show that its product is required in Hh-expressing cells for production of appropriate signaling activity in embryos and in the imaginal precursors of adult tissues. The ski gene encodes an apparent acyltransferase, and we provide genetic and biochemical evidence that Hh proteins from ski mutant cells retain carboxyl-terminal cholesterol modification but lack amino-terminal palmitate modification. Our results suggest that ski encodes an enzyme that acts within the secretory pathway to catalyze amino-terminal palmitoylation of Hh, and further demonstrate that this lipid modification is required for the embryonic and larval patterning activities of the Hh signal.

  16. Generation of sonic power during welding

    NASA Technical Reports Server (NTRS)

    Mc Campbell, W. M.

    1969-01-01

    Generation of intense sonic and ultrasonic power in the weld zone, close to the puddle, reduces the porosity and refinement of the grain. The ac induction brazing power supply is modified with long cables for deliberate addition of resistance to that circuit. The concept is extensible to the molding of metals and plastics.

  17. Sonic Boom: Six Decades of Research

    NASA Technical Reports Server (NTRS)

    Maglieri, Domenic J.; Bobbitt, Percy J.; Plotkin, Kenneth J.; Shepherd, Kevin P.; Coen, Peter G.; Richwine, David M.

    2014-01-01

    Sonic booms generated by aircraft traveling at supersonic speeds have been the subject of extensive aeronautics research for over 60 years. Hundreds of papers have been published that document the experimental and analytical research conducted during this time period. The purpose of this publication is to assess and summarize this work and establish the state-of-the-art for researchers just entering the field, or for those interested in a particular aspect of the subject. This publication consists of ten chapters that cover the experimental and analytical aspects of sonic boom generation, propagation and prediction with summary remarks provided at the end of each chapter. Aircraft maneuvers, sonic boom minimization, simulation techniques and devices as well as human, structural, and other responses to sonic booms are also discussed. The geometry and boom characteristics of various low-boom concepts, both large civil transports and smaller business-jet concepts, are included. The final chapter presents an assessment of civilian supersonic overland flight and highlights the need for continued research and a low-boom demonstrator vehicle. Summary remarks are provided at the end of each chapter. The studies referenced in this publication have been drawn from over 500 references.

  18. Variability of measured sonic boom signatures

    NASA Technical Reports Server (NTRS)

    Elmer, K. R.; Joshi, M. C.

    1994-01-01

    The topics discussed include the following: atmospheric turbulence; BOOMFILE Database description; BOOMFILE flight conditions; XB-70 Database descriptions; analysis progression; extended database; prediction method; overpressure variability dependence on flight conditions; loudness variability on flight conditions; sonic boom variability in repeat flights; and statistical distributions.

  19. Dhcr7 Regulates Palatal Shelf Fusion through Regulation of Shh and Bmp2 Expression

    PubMed Central

    Xiao, Wen-lin; Zhang, Dai-zun; Xu, Hong; Zhuang, Cui-zhu

    2016-01-01

    The aim of this study was to investigate the effect of the 7-dehydrocholesterol reductase (Dhcr7) gene and identify signaling pathways involved in regulation of embryonic palatogenesis. The expression of Dhcr7 and its protein product were examined during murine normal embryonic palatogenesis via a reverse transcription polymerase chain reaction (RT-PCR) and Western blot (WB). RNA interference (RNAi) technology was used to inhibit Dhcr7 expression in a palatal shelf culture in vitro. The effects of Dhcr7 on palatogenesis and palatal fusion were examined by scanning electron microscopy (SEM). The expression changes of Dhcr7, Sonic Hedgehog (Shh), and bone morphogenetic protein-2 (Bmp2) were measured by RT-PCR and WB after Dhcr7 gene silencing and the addition of exogenous cholesterol. The results showed that the palatal shelf failed to complete normal development and fusion when Dhcr7 expression was inhibited. The inhibitory effect study of RNAi on the development of the palatal shelf supported that cholesterol supplementation did not alter the silencing of Dhcr7. Shh and Bmp2 expressions were reduced after Dhcr7 gene silencing, and administration of exogenous cholesterol did not affect Dhcr7 expression; however Shh and Bmp2 expressions increased. We conclude that Dhcr7 plays a role in growth of the palatal shelf and can regulate palatogenesis through alterations in the levels of Shh and Bmp2. PMID:27066502

  20. Dhcr7 Regulates Palatal Shelf Fusion through Regulation of Shh and Bmp2 Expression.

    PubMed

    Xiao, Wen-lin; Zhang, Dai-zun; Xu, Hong; Zhuang, Cui-zhu

    2016-01-01

    The aim of this study was to investigate the effect of the 7-dehydrocholesterol reductase (Dhcr7) gene and identify signaling pathways involved in regulation of embryonic palatogenesis. The expression of Dhcr7 and its protein product were examined during murine normal embryonic palatogenesis via a reverse transcription polymerase chain reaction (RT-PCR) and Western blot (WB). RNA interference (RNAi) technology was used to inhibit Dhcr7 expression in a palatal shelf culture in vitro. The effects of Dhcr7 on palatogenesis and palatal fusion were examined by scanning electron microscopy (SEM). The expression changes of Dhcr7, Sonic Hedgehog (Shh), and bone morphogenetic protein-2 (Bmp2) were measured by RT-PCR and WB after Dhcr7 gene silencing and the addition of exogenous cholesterol. The results showed that the palatal shelf failed to complete normal development and fusion when Dhcr7 expression was inhibited. The inhibitory effect study of RNAi on the development of the palatal shelf supported that cholesterol supplementation did not alter the silencing of Dhcr7. Shh and Bmp2 expressions were reduced after Dhcr7 gene silencing, and administration of exogenous cholesterol did not affect Dhcr7 expression; however Shh and Bmp2 expressions increased. We conclude that Dhcr7 plays a role in growth of the palatal shelf and can regulate palatogenesis through alterations in the levels of Shh and Bmp2. PMID:27066502

  1. The Sound of Stigmatization: Sonic Habitus, Sonic Styles, and Boundary Work in an Urban Slum.

    PubMed

    Schwarz, Ori

    2015-07-01

    Based on focus groups and interviews with student renters in an Israeli slum, the article explores the contributions of differences in sonic styles and sensibilities to boundary work, social categorization, and evaluation. Alongside visual cues such as broken windows, bad neighborhoods are characterized by sonic cues, such as shouts from windows. Students understand "being ghetto" as being loud in a particular way and use loudness as a central resource in their boundary work. Loudness is read as a performative index of class and ethnicity, and the performance of middle-class studentship entails being appalled by stigmatized sonic practices and participating in their exoticization. However, the sonic is not merely yet another resource of boundary work. Paying sociological attention to senses other than vision reveals complex interactions between structures anchored in the body, structures anchored in language, and actors' identification strategies, which may refine theorizations of the body and the senses in social theory.

  2. The Sound of Stigmatization: Sonic Habitus, Sonic Styles, and Boundary Work in an Urban Slum.

    PubMed

    Schwarz, Ori

    2015-07-01

    Based on focus groups and interviews with student renters in an Israeli slum, the article explores the contributions of differences in sonic styles and sensibilities to boundary work, social categorization, and evaluation. Alongside visual cues such as broken windows, bad neighborhoods are characterized by sonic cues, such as shouts from windows. Students understand "being ghetto" as being loud in a particular way and use loudness as a central resource in their boundary work. Loudness is read as a performative index of class and ethnicity, and the performance of middle-class studentship entails being appalled by stigmatized sonic practices and participating in their exoticization. However, the sonic is not merely yet another resource of boundary work. Paying sociological attention to senses other than vision reveals complex interactions between structures anchored in the body, structures anchored in language, and actors' identification strategies, which may refine theorizations of the body and the senses in social theory. PMID:26430711

  3. Sonic morphology: Aesthetic dimensional auditory spatial awareness

    NASA Astrophysics Data System (ADS)

    Whitehouse, Martha M.

    The sound and ceramic sculpture installation, " Skirting the Edge: Experiences in Sound & Form," is an integration of art and science demonstrating the concept of sonic morphology. "Sonic morphology" is herein defined as aesthetic three-dimensional auditory spatial awareness. The exhibition explicates my empirical phenomenal observations that sound has a three-dimensional form. Composed of ceramic sculptures that allude to different social and physical situations, coupled with sound compositions that enhance and create a three-dimensional auditory and visual aesthetic experience (see accompanying DVD), the exhibition supports the research question, "What is the relationship between sound and form?" Precisely how people aurally experience three-dimensional space involves an integration of spatial properties, auditory perception, individual history, and cultural mores. People also utilize environmental sound events as a guide in social situations and in remembering their personal history, as well as a guide in moving through space. Aesthetically, sound affects the fascination, meaning, and attention one has within a particular space. Sonic morphology brings art forms such as a movie, video, sound composition, and musical performance into the cognitive scope by generating meaning from the link between the visual and auditory senses. This research examined sonic morphology as an extension of musique concrete, sound as object, originating in Pierre Schaeffer's work in the 1940s. Pointing, as John Cage did, to the corporeal three-dimensional experience of "all sound," I composed works that took their total form only through the perceiver-participant's participation in the exhibition. While contemporary artist Alvin Lucier creates artworks that draw attention to making sound visible, "Skirting the Edge" engages the perceiver-participant visually and aurally, leading to recognition of sonic morphology.

  4. Spontaneous neoplasia in four captive greater hedgehog tenrecs (Setifer setosus).

    PubMed

    Khoii, Mina K; Howerth, Elizabeth W; Burns, Roy B; Carmichael, K Paige; Gyimesi, Zoltan S

    2008-09-01

    Little information is available about diseases and pathology of species within the family Tenrecidae, including the greater hedgehog tenrec (Setifer setosus), a Madagascan insectivore. This report summarizes necropsy and histopathologic findings of neoplasia in four captive greater hedgehog tenrecs. Although only four animals are included in this report, neoplasia seems to be a common and significant source of morbidity and mortality in greater hedgehog tenrecs. Types of neoplasia identified include a thyroid follicular-solid carcinoma, two urinary bladder transitional cell carcinomas, uterine endometrial polyps, and multicentric B-cell lymphoma. Due to small sample size, no etiology could be determined, but genetics, viral infection, pesticide treatment, nutrition, or other environmental factors might contribute to the development of neoplasia in this species. This is the first report of neoplasia in greater hedgehog tenrecs.

  5. Impacts of Removing Badgers on Localised Counts of Hedgehogs

    PubMed Central

    Trewby, Iain D.; Young, Richard; McDonald, Robbie A.; Wilson, Gavin J.; Davison, John; Walker, Neil; Robertson, Andrew; Doncaster, C. Patrick; Delahay, Richard J.

    2014-01-01

    Experimental evidence of the interactions among mammalian predators that eat or compete with one another is rare, due to the ethical and logistical challenges of managing wild populations in a controlled and replicated way. Here, we report on the opportunistic use of a replicated and controlled culling experiment (the Randomised Badger Culling Trial) to investigate the relationship between two sympatric predators: European badgers Meles meles and western European hedgehogs Erinaceus europaeus. In areas of preferred habitat (amenity grassland), counts of hedgehogs more than doubled over a 5-year period from the start of badger culling (from 0.9 ha−1 pre-cull to 2.4 ha−1 post-cull), whereas hedgehog counts did not change where there was no badger culling (0.3–0.3 hedgehogs ha−1). This trial provides experimental evidence for mesopredator release as an outcome of management of a top predator. PMID:24736454

  6. Impacts of removing badgers on localised counts of hedgehogs.

    PubMed

    Trewby, Iain D; Young, Richard; McDonald, Robbie A; Wilson, Gavin J; Davison, John; Walker, Neil; Robertson, Andrew; Doncaster, C Patrick; Delahay, Richard J

    2014-01-01

    Experimental evidence of the interactions among mammalian predators that eat or compete with one another is rare, due to the ethical and logistical challenges of managing wild populations in a controlled and replicated way. Here, we report on the opportunistic use of a replicated and controlled culling experiment (the Randomised Badger Culling Trial) to investigate the relationship between two sympatric predators: European badgers Meles meles and western European hedgehogs Erinaceus europaeus. In areas of preferred habitat (amenity grassland), counts of hedgehogs more than doubled over a 5-year period from the start of badger culling (from 0.9 ha-1 pre-cull to 2.4 ha-1 post-cull), whereas hedgehog counts did not change where there was no badger culling (0.3-0.3 hedgehogs ha-1). This trial provides experimental evidence for mesopredator release as an outcome of management of a top predator.

  7. WT1 regulates the development of the posterior taste field.

    PubMed

    Gao, Yankun; Toska, Eneda; Denmon, Dane; Roberts, Stefan G E; Medler, Kathryn F

    2014-06-01

    Despite the importance of taste in determining nutrient intake, our understanding of the processes that control the development of the peripheral taste system is lacking. Several early regulators of taste development have been identified, including sonic hedgehog, bone morphogenetic protein 4 and multiple members of the Wnt/β-catenin signaling pathway. However, the regulation of these factors, including their induction, remains poorly understood. Here, we identify a crucial role for the Wilms' tumor 1 protein (WT1) in circumvallate (CV) papillae development. WT1 is a transcription factor that is important in the normal development of multiple tissues, including both the olfactory and visual systems. In mice, WT1 expression is detectable by E12.5, when the CV taste placode begins to form. In mice lacking WT1, the CV fails to develop normally and markers of early taste development are dysregulated compared with wild type. We demonstrate that expression of the WT1 target genes Lef1, Ptch1 and Bmp4 is significantly reduced in developing tongue tissue derived from Wt1 knockout mice and that, in normal tongue, WT1 is bound to the promoter regions of these genes. Moreover, siRNA knockdown of WT1 in cultured taste cells leads to a reduction in the expression of Lef1 and Ptch1. Our data identify WT1 as a crucial transcription factor in the development of the CV through the regulation of multiple signaling pathways that have established roles in the formation and patterning of taste placodes.

  8. NHR-23 dependent collagen and hedgehog-related genes required for molting

    SciTech Connect

    Kouns, Nathaniel A.; Nakielna, Johana; Behensky, Frantisek; Krause, Michael W.; Kostrouch, Zdenek; Kostrouchova, Marta

    2011-10-07

    Highlights: {yields} NHR-23 is a critical regulator of nematode development and molting. {yields} The manuscript characterizes the loss-of-function phenotype of an nhr-23 mutant. {yields} Whole genome expression analysis identifies new potential targets of NHR-23. {yields} Hedgehog-related genes are identified as NHR-23 dependent genes. {yields} New link between sterol mediated signaling and regulation by NHR-23 is found. -- Abstract: NHR-23, a conserved member of the nuclear receptor family of transcription factors, is required for normal development in Caenorhabditis elegans where it plays a critical role in growth and molting. In a search for NHR-23 dependent genes, we performed whole genome comparative expression microarrays on both control and nhr-23 inhibited synchronized larvae. Genes that decreased in response to nhr-23 RNAi included several collagen genes. Unexpectedly, several hedgehog-related genes were also down-regulated after nhr-23 RNAi. A homozygous nhr-23 deletion allele was used to confirm the RNAi knockdown phenotypes and the changes in gene expression. Our results indicate that NHR-23 is a critical co-regulator of functionally linked genes involved in growth and molting and reveal evolutionary parallels among the ecdysozoa.

  9. Rickettsiae in arthropods collected from the North African Hedgehog (Atelerix algirus) and the desert hedgehog (Paraechinus aethiopicus) in Algeria.

    PubMed

    Khaldi, Mourad; Socolovschi, Cristina; Benyettou, Meryam; Barech, Ghania; Biche, Mohamed; Kernif, Tahar; Raoult, Didier; Parola, Philippe

    2012-03-01

    Hedgehogs have become a popular pet despite their potential role in zoonotic disease transmission. We conducted an entomological study in a mountainous region of northeast Algeria in which we collected 387 fleas (Archeopsylla erinacei) and 342 ticks (Rhipicephalus sanguineus and Haemaphysalis erinacei) from Paraechinus aethiopicus and Atelerix algirus hedgehogs. Of the hedgehogs sampled, 77.7% and 91% were infested with fleas and ticks, respectively. Significantly more ticks and fleas were collected from A. algirus than from P. aethiopicus. Rickettsia felis was detected in 95.5% of fleas and R. massiliae was detected in 6.25% of Rh. sanguineus ticks by molecular tools. A new Rickettsia species of the spotted fever group was detected in 11.25% of Rh. sanguineus and in 77% of H. erinacei ticks. Overall, we show that hedgehogs can act as hosts for ectoparasites infected with several rickettsial agents. These data justify a more detailed investigation of animal reservoirs for Rickettsiae. PMID:22222114

  10. Sonic-boom research: Selected bibliography with annotation

    NASA Astrophysics Data System (ADS)

    Hubbard, H. H.; Maglieri, D. J.; Stephens, D. G.

    1986-09-01

    Citations of selected documents are included which represent the state of the art of technology in each of the following subject areas: prediction, measurement, and minimization of steady-flight sonic booms; prediction and measurement of accelerating-flight sonic booms; sonic-boom propagation; the effects of sonic booms on people, communities, structures, animals, birds, and terrain; and sonic-boom simulator technology. Documents are listed in chronological order in each section of the paper, with key documents and associated annotation listed first. The sources are given along with acquisition numbers, when available, to expedite the acquisition of copies of the documents.

  11. Sonic-boom research: Selected bibliography with annotation

    NASA Technical Reports Server (NTRS)

    Hubbard, H. H.; Maglieri, D. J.; Stephens, D. G.

    1986-01-01

    Citations of selected documents are included which represent the state of the art of technology in each of the following subject areas: prediction, measurement, and minimization of steady-flight sonic booms; prediction and measurement of accelerating-flight sonic booms; sonic-boom propagation; the effects of sonic booms on people, communities, structures, animals, birds, and terrain; and sonic-boom simulator technology. Documents are listed in chronological order in each section of the paper, with key documents and associated annotation listed first. The sources are given along with acquisition numbers, when available, to expedite the acquisition of copies of the documents.

  12. Hedgehog Secretion and Signal Transduction in Vertebrates

    PubMed Central

    Ryan, Kaitlyn E.; Chiang, Chin

    2012-01-01

    Signaling by the Hedgehog (Hh) family of secreted proteins is essential for proper embryonic patterning and development. Dysregulation of Hh signaling is associated with a variety of human diseases ranging from developmental disorders such as holoprosencephaly to certain forms of cancer, including medulloblastoma and basal cell carcinoma. Genetic studies in flies and mice have shaped our understanding of Hh signaling and revealed that nearly all core components of the pathway are highly conserved. Although many aspects of the Drosophila Hh pathway are conserved in vertebrates, mechanistic differences between the two species have begun to emerge. Perhaps the most striking divergence in vertebrate Hh signaling is its dependence on the primary cilium, a vestigial organelle that is largely absent in flies. This minireview will provide an overview of Hh signaling and present recent insights into vertebrate Hh secretion, receptor binding, and signal transduction. PMID:22474285

  13. The Hedgehog pathway: role in cell differentiation, polarity and proliferation.

    PubMed

    Jia, Yanfei; Wang, Yunshan; Xie, Jingwu

    2015-02-01

    Hedgehog (Hh) is first described as a genetic mutation that has "spiked" phenotype in the cuticles of Drosophila in later 1970s. Since then, Hh signaling has been implicated in regulation of differentiation, proliferation, tissue polarity, stem cell population and carcinogenesis. The first link of Hh signaling to cancer was established through discovery of genetic mutations of Hh receptor gene PTCH1 being responsible for Gorlin syndrome in 1996. It was later shown that Hh signaling is associated with many types of cancer, including skin, leukemia, lung, brain and gastrointestinal cancers. Another important milestone for the Hh research field is the FDA approval for the clinical use of Hh inhibitor Erivedge/Vismodegib for treatment of locally advanced and metastatic basal cell carcinomas. However, recent clinical trials of Hh signaling inhibitors in pancreatic, colon and ovarian cancer all failed, indicating a real need for further understanding of Hh signaling in cancer. In this review, we will summarize recent progress in the Hh signaling mechanism and its role in human cancer. PMID:25559776

  14. Hedgehog does not guide migrating Drosophila germ cells

    PubMed Central

    Renault, Andrew D.; Ricardo, Sara; Kunwar, Prabhat S.; Santos, Ana; Starz-Gaiano, Michelle; Stein, Jennifer; Lehmann, Ruth

    2009-01-01

    In many species, the germ cells, precursors of sperm and egg, migrate during embryogenesis. The signals that regulate this migration are thus essential for fertility. In flies, lipid signals have been shown to affect germ cell guidance. In particular, the synthesis of geranylgeranyl pyrophosphate through the 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (Hmgcr) pathway is critical for attracting germ cells to their target tissue. In a genetic analysis of signaling pathways known to affect cell migration of other migratory cells, we failed to find a role for the Hedgehog (Hh) pathway in germ cell migration. However, previous reports had implicated Hh as a germ cell attractant in flies and suggested that Hh signaling is enhanced through the action of the Hmgcr pathway. We therefore repeated several critical experiments and carried out further experiments to test specifically whether Hh is a germ cell attractant in flies. In contrast to previously reported findings and consistent with findings in zebrafish our data do not support the notion that Hh has a direct role in the guidance of migrating germ cells in flies. PMID:19389345

  15. [The Cytoskelrtal Protein Zvxin Interacts with the Hedgehog Receptor Patched].

    PubMed

    Martynova, N U; Ermolina, L V; Eroshkin, F M; Zarayskiy, A G

    2015-01-01

    Earlier, we demonstrated Zyxin influence upon Hedgehog (Hh)-signaling pathway during early patterning of the central neural system (CNS) anlage of the Xenopus laevis embryo. Now we show that Zyxin can physically interact with the transmembrane receptor of Hh, Patched2 (Ptc2). Binding of Hh by this receptor activates signaling pathway, which regulates many events, including numerous types of cell differentiation during the embryonic development. In particular, patterning of the CNS anlage. The ability of Zyxin to interact with Ptc2 have been confirmed by immunoprecipitation experiments, in which we tested mutual binding affinity of Zyxin and Ptc2, as well as mutual affinity of their deletion mutants. As a result, we have established that in Xenopus levis, Zyxin binding to Ptc2 is due to the interaction of Zyxin 2nd LIM-domain (530-590 aa) with the under-membrane region of the cytoplasmic C-terminus of Ptc2 (1159-1412 aa). We have also demonstrated that similar interaction is valid for the homologous regions of the human Zyxin and human Hh receptor, Ptc1. The data obtained allow to hypothesize existence of evolutionary conserved mechanism that modulates Hh-signaling and based on the interaction of Zyxin with Ptc. PMID:27125030

  16. Distinct spatiotemporal roles of hedgehog signalling during chick and mouse cranial base and axial skeleton development

    PubMed Central

    Balczerski, B.; Zakaria, S.; Tucker, A. S.; Borycki, A.G.; Koyama, E.; Pacifici, M.; Francis-West, P.

    2012-01-01

    The cranial base exerts a supportive role for the brain and includes the occipital, sphenoid and ethmoid bones that arise from cartilaginous precursors in the early embryo. As the occipital bone and the posterior part of the sphenoid are mesoderm derivatives that arise in close proximity to the notochord and floor plate, it has been assumed that their development, like the axial skeleton, is dependent on Sonic hedgehog (Shh) and modulation of bone morphogenetic protein (Bmp) signalling. Here we examined the development of the cranial base in chick and mouse embryos to compare the molecular signals that are required for chondrogenic induction in the trunk and head. We found that Shh signalling is required but the molecular network controlling cranial base development is distinct from that in the trunk. In the absence of Shh, the presumptive cranial base did not undergo chondrogenic commitment as determined by the loss of Sox9 expression and there was a decrease in cell survival. In contrast, induction of the otic capsule occurred normally demonstrating that induction of the cranial base is uncoupled from formation of the sensory capsules. Lastly, we found that the early cranial mesoderm is refractory to Shh signalling, likely accounting for why development of the cranial base occurs after the axial skeleton. Our data reveal that cranial and axial skeletal induction is controlled by conserved, yet spatiotemporally distinct mechanisms that co-ordinate development of the cranial base with that of the cranial musculature and the pharyngeal arches. PMID:23009899

  17. Crocetinic acid inhibits hedgehog signaling to inhibit pancreatic cancer stem cells

    PubMed Central

    Rangarajan, Parthasarathy; Subramaniam, Dharmalingam; Paul, Santanu; Kwatra, Deep; Palaniyandi, Kanagaraj; Islam, Shamima; Harihar, Sitaram; Ramalingam, Satish; Gutheil, William; Putty, Sandeep; Pradhan, Rohan; Padhye, Subhash; Welch, Danny R.; Anant, Shrikant; Dhar, Animesh

    2015-01-01

    Pancreatic cancer is the fourth leading cause of cancer deaths in the US and no significant treatment is currently available. Here, we describe the effect of crocetinic acid, which we purified from commercial saffron compound crocetin using high performance liquid chromatography. Crocetinic acid inhibits proliferation of pancreatic cancer cell lines in a dose- and time-dependent manner. In addition, it induced apoptosis. Moreover, the compound significantly inhibited epidermal growth factor receptor and Akt phosphorylation. Furthermore, crocetinic acid decreased the number and size of the pancospheres in a dose-dependent manner, and suppressed the expression of the marker protein DCLK-1 (Doublecortin Calcium/Calmodulin-Dependent Kinase-1) suggesting that crocetinic acid targets cancer stem cells (CSC). To understand the mechanism of CSC inhibition, the signaling pathways affected by purified crocetinic acid were dissected. Sonic hedgehog (Shh) upon binding to its cognate receptor patched, allows smoothened to accumulate and activate Gli transcription factor. Crocetinic acid inhibited the expression of both Shh and smoothened. Finally, these data were confirmed in vivo where the compound at a dose of 0.5 mg/Kg bw suppressed growth of tumor xenografts. Collectively, these data suggest that purified crocetinic acid inhibits pancreatic CSC, thereby inhibiting pancreatic tumorigenesis. PMID:26317547

  18. Hedgehog signaling promotes basal progenitor expansion and the growth and folding of the neocortex.

    PubMed

    Wang, Lei; Hou, Shirui; Han, Young-Goo

    2016-07-01

    The unique mental abilities of humans are rooted in the immensely expanded and folded neocortex, which reflects the expansion of neural progenitors, especially basal progenitors including basal radial glia (bRGs) and intermediate progenitor cells (IPCs). We found that constitutively active Sonic hedgehog (Shh) signaling expanded bRGs and IPCs and induced folding in the otherwise smooth mouse neocortex, whereas the loss of Shh signaling decreased the number of bRGs and IPCs and the size of the neocortex. SHH signaling was strongly active in the human fetal neocortex but Shh signaling was not strongly active in the mouse embryonic neocortex, and blocking SHH signaling in human cerebral organoids decreased the number of bRGs. Mechanistically, Shh signaling increased the initial generation and self-renewal of bRGs and IPC proliferation in mice and the initial generation of bRGs in human cerebral organoids. Thus, robust SHH signaling in the human fetal neocortex may contribute to bRG and IPC expansion and neocortical growth and folding. PMID:27214567

  19. Sonic temperature sensor for food processing

    SciTech Connect

    Akers, D.W.; Porter, A.M.; Tow, D.M.

    1997-09-01

    The lack of adequate temperature measurement is the major barrier to the development of more efficient and better quality food processing methods. The objective of the sonic temperature sensor for food processing project is to develop a prototype sensor to noninvasively measure the interior temperature of particulate foods during processing. This, a joint project with the National Food Processors Association, utilizes the property of materials that when the temperature of a material changes, there is a corresponding change in the speed of sound. The approach taken for the sonic sensor system is to determine the speed of sound inside particulate foods using a tomographic reconstruction process. This work has shown that the speed of sound can be accurately determined to an accuracy of {+-}0.4%, corresponding to a temperature uncertainty of {+-}2{degree}C using tomographic reconstruction methods.

  20. Concorde sonic booms as an atmospheric probe.

    PubMed

    Balachandran, N K; Donn, W L; Rind, D H

    1977-07-01

    Infrasound generated by the sonic boom from the inbound Concorde supersonic transport is recorded at Palisades, New York (Lamont-Doherty Geological Observatory), as a series of impulses from distances varying from 165 to about 1000 kilometers. Refraction effects determined by temperature and wind conditions return the signal to the surface from both stratospheric (40 to 50 kilometers) and thermospheric (100 to 130 kilometers) levels. The frequency of the recorded signal is a function of the level of reflection; the frequency decreases from impulse stretching as the atmosphere becomes more rarified relative to the sound pressure. The horizontal trace velocity of the signal across the array of instruments is equal to the acoustic velocity at the reflection level. The sonic boom can thus be used to provide temperature-wind parameters at reflection levels estimated from the signal frequency. Daily observed signal variations have indicated significant variations in these parameters. PMID:17828888

  1. Sonic Temperature Sensor for Food Processing

    SciTech Connect

    Akers, D. W.; Porter, A. M.; Tow, D. M.

    1997-09-01

    The lack of adequate temperature measurement is the major barrier to the development of more efficient and better quality food processing methods. The objective of the sonic temperature sensor for food processing project is to develop a prototype sensor system to noninvasively measure the interior temperature of particulate foods during processing. The development of the prototype sensor is a collaborative project with the National Food Processors Association. The project is based on the property of materials that involves a change in the temperature of a material having a corresponding change in the speed of sound. The approach for the sonic sensor system is to determine the speed of sound through particulate foods using a tomographic reconstruction process.

  2. Current research in sonic-boom minimization

    NASA Technical Reports Server (NTRS)

    Darden, C. M.; Mack, R. J.

    1976-01-01

    A review is given of several questions as yet unanswered in the area of sonic-boom research. Efforts, both here at Langley and elsewhere, in the area of minimization, human response, design techniques and in developing higher order propagation methods are discussed. In addition, a wind-tunnel test program being conducted to assess the validity of minimization methods based on a forward spike in the F-function is described.

  3. Status of knowledge of sonic booms

    NASA Technical Reports Server (NTRS)

    Maglieri, D. J.; Carlson, H. W.; Hubbard, H. H.

    1979-01-01

    The status of sonic boom technology with emphasis on the recent research results is summarized. Included are definitions of the boom carpets, both primary and secondary, a discussion of existing experience with primary booms including the status of overpressure predictions and boom minimization methodology through airplane design, an indication of the boom waveforms and audibility, and a discussion of focus booms resulting from aircraft maneuvers as well as the effect of abnormal atmospheric conditions on these maneuver booms.

  4. Identification of spontaneous mutations within the long-range limb-specific Sonic Hedgehog enhancer (ZRS) that alter Sonic Hedgehog expression in the chicken limb mutants oligozeugodactly and Silkie Breed

    PubMed Central

    Maas, Sarah A.; Suzuki, Takayuki; Fallon, John F.

    2011-01-01

    The evolutionarily conserved, non-coding ~800 base-pair zone of polarizing activity (ZPA) regulatory sequence (ZRS) controls Shh expression in the posterior limb. We report that the chicken mutant oligozeugodactly (ozd), which lacks limb Shh expression, has a large deletion within the ZRS. Furthermore, the preaxial polydactylous, Silkie Breed chicken, which develops ectopic anterior limb Shh expression, has a single base-pair change within the ZRS. Using an in vivo reporter assay to examine enhancer function in the chick limb, we demonstrate that the wild-type ZRS drives β-galactosidase reporter expression in the ZPA of both wild-type and ozd limbs. The Silkie ZRS drives β-galactosidase in both posterior and anterior Shh domains in wild-type limb buds. These results support the hypothesis that the ZRS integrates positive and negative prepatterned regulatory inputs in the chicken model system and demonstrate the utility of the chicken limb as an efficient genetic system for gene regulatory studies. PMID:21509895

  5. Measured N-wave sonic boom events and sensitivity in sonic boom metrics

    NASA Astrophysics Data System (ADS)

    Palmer, Joshua; Sparrow, Victor W.

    2015-10-01

    Various sonic boom noise metrics have been calculated for a number of sonic boom, N-wave signatures. The newly computed metrics dataset utilized high-quality recordings from the Superboom Caustic Analysis and Measurement Program (SCAMP) experiment conducted by NASA. With this signature dataset comprised of microphone measurements by long linear arrays, one can assess the waveform variability due to atmospheric turbulence influences across the arrays. Preferred boom events from these NASA datasets were then chosen after review of the flight conditions, flight objectives and actual waveforms generated in order to study only the non-focused, N-wave sonic boom signatures. The sonic boom noise metrics calculated for the preferred boom events include Stevens Mark VII Perceived Level (PLdB), un-weighted Sound Exposure Level (SELz) as well as Sound Exposure Level with A, B, C, D, and E weightings applied to the waveforms. The results show, for example, that the A-weighted sound exposure levels and Steven's Mark VII Perceived Levels had standard deviations in the range of 1.4 dB to 6.1 dB for the SCAMP measurements. Such sensitivity results should be helpful in assessing the applicability of sonic boom metrics for use in future en-route certification standards for civilian supersonic aircraft.

  6. A Compilation of Space Shuttle Sonic Boom Measurements - Supplemental STS Sonic Boom Files

    NASA Technical Reports Server (NTRS)

    Maglieri, Domenic J.; Henderson, Herbert R.; Massey, Steven J.; Stansbery, Eugene G.

    2011-01-01

    Supplemental STS Sonic Boom Files for NASA/CR-2011-217080. Data files included on CDROM formatted to ISO 9660 standards. Sonic boom measurements have been obtained on 26 flights of the Space Shuttle system beginning with the launch of STS-1 on April 12, 1981, to the reentry-descent of STS-41 into EAFB on Oct. 10, 1990. A total of 23 boom measurements were acquired within the focus region off the Florida coast during 3 STS launch-ascents and 113 boom measurements were acquired during 23 STS reentry-descent to landing into Florida and California. Sonic boom measurements were made under, and lateral to, the vehicle ground track and cover the Mach-altitude range of about 1.3 to 23 and 54,000 feet to 243,000 feet, respectively. Vehicle operational data, flight profiles and weather data were also gathered during the flights. This STS boom database is contained in 26 documents, some are formal and referenceable but most internal documents. Another 38 documents, also non-referenceable, contain predicted sonic boom footprints for reentry-descent flights on which no measurements were made. The purpose of this report is to provide an overview of the STS sonic boom database and summarize the main findings.

  7. Expression pattern of the Hedgehog signaling pathway in pituitary adenomas.

    PubMed

    Yavropoulou, Maria P; Maladaki, Anna; Topouridou, Konstantina; Kotoula, Vasiliki; Poulios, Chris; Daskalaki, Emily; Foroglou, Nikolaos; Karkavelas, George; Yovos, John G

    2016-01-12

    Several studies have demonstrated the role of Wnt and Notch signaling in the pathogenesis of pituitary adenomas, but data are scarce regarding the role of Hedgehog signaling. In this study we investigated the differential expression of gene targets of the Hedgehog signaling pathway. Formalin-fixed, paraffin-embedded specimens from adult patients who underwent transphenoidal resection and normal human pituitary tissues that were obtained from autopsies were used. Clinical information and data from pre-operative MRI scan (extracellular tumor extension, tumor size, displacement of the optic chiasm) were retrieved from the Hospital's database. We used a customized RT(2) Profiler PCR Array, to investigate the expression of genes related to Notch and Hedgehog signaling pathways (PTCH1, PTCH2, GLI1, GLI3, NOTCH3, JAG1, HES1, and HIP). A total of 52 pituitary adenomas (32 non-functioning adenomas, 15 somatotropinomas and 5 prolactinomas) were used in the final analysis. In non-functioning pituitary adenomas there was a significant decrease (approximately 75%) in expression of all Hedgehog related genes that were tested, while Notch3 and Jagged-1 expression was found significantly increased, compared with normal pituitary tissue controls. In contrast, somatotropinomas demonstrated a significant increase in expression of all Hedgehog related genes and a decrease in the expression of Notch3 and Jagged-1. There was no significant difference in the expression of Hedgehog and Notch related genes between prolactinomas and healthy pituitary tissues. Hedgehog signalling appears to be activated in somatotropinomas but not in non-functioning pituitary adenomas in contrast to the expression pattern of Notch signalling pathway. PMID:26620835

  8. High-Quality Seismic Observations of Sonic Booms

    NASA Astrophysics Data System (ADS)

    Wurman, G.; Haering, E. A.; Price, M.

    2011-12-01

    The SonicBREWS project (Sonic Boom Resistant Earthquake Warning Systems) is a collaborative effort between Seismic Warning Systems, Inc. and NASA Dryden Flight Research Center. This project aims to evaluate the effects of sonic booms on Earthquake Warning Systems in order to prevent such systems from experiencing false alarms due to sonic booms. The airspace above the Antelope Valley, California includes the High Altitude Supersonic Corridor and the Black Mountain Supersonic Corridor. These corridors are among the few places in the US where supersonic flight is permitted, and sonic booms are commonplace in the Antelope Valley. One result of this project is a rich dataset of high-quality accelerometer records of sonic booms which can shed light on the interaction between these atmospheric phenomena and the solid earth. Nearly 100 sonic booms were recorded with low-noise triaxial MEMS accelerometers recording 1000 samples per second. The sonic booms had peak overpressures ranging up to approximately 10 psf and were recorded in three flight series in 2010 and 2011. Each boom was recorded with up to four accelerometers in various array configurations up to 100 meter baseline lengths, both in the built environment and the free field. All sonic booms were also recorded by nearby microphones. We present the results of the project in terms of the potential for sonic-boom-induced false alarms in Earthquake Warning Systems, and highlight some of the interesting features of the dataset.

  9. High-Quality Seismic Observations of Sonic Booms

    NASA Technical Reports Server (NTRS)

    Wurman, Gilead; Haering, Edward A., Jr.; Price, Michael J.

    2011-01-01

    The SonicBREWS project (Sonic Boom Resistant Earthquake Warning Systems) is a collaborative effort between Seismic Warning Systems, Inc. and NASA Dryden Flight Research Center. This project aims to evaluate the effects of sonic booms on Earthquake Warning Systems in order to prevent such systems from experiencing false alarms due to sonic booms. The airspace above the Antelope Valley, California includes the High Altitude Supersonic Corridor and the Black Mountain Supersonic Corridor. These corridors are among the few places in the US where supersonic flight is permitted, and sonic booms are commonplace in the Antelope Valley. One result of this project is a rich dataset of high-quality accelerometer records of sonic booms which can shed light on the interaction between these atmospheric phenomena and the solid earth. Nearly 100 sonic booms were recorded with low-noise triaxial MEMS accelerometers recording 1000 samples per second. The sonic booms had peak overpressures ranging up to approximately 10 psf and were recorded in three flight series in 2010 and 2011. Each boom was recorded with up to four accelerometers in various array configurations up to 100 meter baseline lengths, both in the built environment and the free field. All sonic booms were also recorded by nearby microphones. We present the results of the project in terms of the potential for sonic-boom-induced false alarms in Earthquake Warning Systems, and highlight some of the interesting features of the dataset.

  10. Sonic Booms on Big Structures (SonicBOBS) Phase I Database; NASA Dryden Sensors

    NASA Technical Reports Server (NTRS)

    Haering, Edward A., Jr.; Arnac, Sarah Renee

    2010-01-01

    This DVD contains 13 channels of microphone and up to 22 channels of pressure transducer data collected in September, 2009 around several buildings located at Edwards Air Force Base. These data were recorded by NASA Dryden. Not included are data taken by NASA Langley and Gulfstream. Each day's data is in a separate folder and each pass is in a file beginning with "SonicBOBS_" (for microphone data) or "SonicBOBSBB_" (for BADS and BASS data) followed by the month, day, year as two digits each, followed by the hour, minute, sec after midnight GMT. The filename time given is for the END time of the raw recording file. In the case of the microphone data, this time may be several minutes after the sonic boom, and is according to the PC's uncalibrated clock. The Matlab data files have the actual time as provided by a GPS-based IRIG-B signal recorded concurrently with the data. Microphone data is given for 5 seconds prior to 20 seconds after the sonic boom. BADS and BASS data is given for the full recording, 6 seconds for the BADS and 10 seconds for the BASS. As an example of the naming convention, file "SonicBOBS_091209154618.mat" is from September 12, 2009 at 15:46:18 GMT. Note that data taken on September 12, 2009 prior to 01:00:00 GMT was of the Space Shuttle Discovery (a sonic boom of opportunity), which was on September 11, 2009 in local Pacific Daylight Time.

  11. Targeting the Sonic Hedgehog-Gli1 Pathway as a Potential New Therapeutic Strategy for Myelodysplastic Syndromes

    PubMed Central

    Zou, Jixue; Zhou, Zhigang; Wan, Liping; Tong, Yin; Qin, Youwen; Wang, Chun; Zhou, Kun

    2015-01-01

    The complex mechanistic array underlying the pathogenesis of myelodysplastic syndrome (MDS) is still unclear. Although dysregulations of different signaling pathways involved in MDS have been described, the identification of specific biomarkers and therapy targets remains an important task in order to establish novel therapeutic approaches. Here, we demonstrated that the Shh signaling pathway is active in MDS and correlated it with disease progression. Additionally, the knockdown of Gli1 significantly inhibited cell proliferation in vitro and in vivo. Gli1 silencing also induced apoptosis and G0/G1 phase arrest. Furthermore, Gli1 silencing enhanced the demethylating effect of 5-aza-2'-deoxycytidine on the p15 gene promoter and subsequently promoted its expression by inhibiting DNA methyltransferase 1(DNMT1). Our findings show that the Shh signaling pathway plays a role in the pathogenesis and disease progression of MDS, and proceeds by modulating DNA methylation. This pathway may prove to be a potential therapeutic target for enhancing the therapeutic effects of 5-azacytidine on malignant transformation of MDS. PMID:26317501

  12. Induction of MITF expression in human cholangiocarcinoma cells and hepatocellular carcinoma cells by cyclopamine, an inhibitor of the Hedgehog signaling.

    PubMed

    Samatiwat, Papavee; Takeda, Kazuhisa; Satarug, Soisungwan; Ohba, Koji; Kukongviriyapan, Veerapol; Shibahara, Shigeki

    2016-01-29

    Microphthalmia-associated transcription factor (MITF) is a key regulator of differentiation of melanocytes and retinal pigment epithelial cells, but it also has functions in non-pigment cells. MITF consists of multiple isoforms, including widely expressed MITF-A and MITF-H. In the present study, we explored the potential role played by the Hedgehog signaling on MITF expression in two common types of primary liver cancer, using human cholangiocarcinoma cell lines, the KKU-100 and HuCCT1, along with the HepG2 human hepatocellular carcinoma cell line. Importantly, cholangiocarcinoma is characterized by the activated Hedgehog signaling. Here we show that MITF-A mRNA is predominantly expressed in all three human liver cancer cell lines examined. Moreover, cyclopamine, an inhibitor of the Hedgehog signalling, increased the expression levels of MITF proteins in HuCCT1 and HepG2 cells, but not in KKU-100 cells, suggesting that MITF expression may be down-regulated in some liver cancer cases. PMID:26773496

  13. Lithium chloride modulates chondrocyte primary cilia and inhibits Hedgehog signaling.

    PubMed

    Thompson, Clare L; Wiles, Anna; Poole, C Anthony; Knight, Martin M

    2016-02-01

    Lithium chloride (LiCl) exhibits significant therapeutic potential as a treatment for osteoarthritis. Hedgehog signaling is activated in osteoarthritis, where it promotes chondrocyte hypertrophy and cartilage matrix catabolism. Hedgehog signaling requires the primary cilium such that maintenance of this compartment is essential for pathway activity. Here we report that LiCl (50 mM) inhibits Hedgehog signaling in bovine articular chondrocytes such that the induction of GLI1 and PTCH1 expression is reduced ​ by 71 and 55%, respectively. Pathway inhibition is associated with a 97% increase in primary cilia length from 2.09 ± 0.7 μm in untreated cells to 4.06 ± 0.9 μm in LiCl-treated cells. We show that cilia elongation disrupts trafficking within the axoneme with a 38% reduction in Arl13b ciliary localization at the distal region of the cilium, consistent with the role of Arl13b in modulating Hedgehog signaling. In addition, we demonstrate similar increases in cilia length in human chondrocytes in vitro and after administration of dietary lithium to Wistar rats in vivo. Our data provide new insights into the effects of LiCl on chondrocyte primary cilia and Hedgehog signaling and shows for the first time that pharmaceutical targeting of the primary cilium may have therapeutic benefits in the treatment of osteoarthritis. PMID:26499268

  14. Effect of sonication on eliminating of phorate in apple juice.

    PubMed

    Zhang, Yuanyuan; Zhang, Zhe; Chen, Fang; Zhang, Hui; Hu, Xiaosong

    2012-01-01

    The degradation of phorate in apple juice by sonication was investigated in the present study. Results showed that sonication was effective in eliminating phorate in apple juice, and the ultrasonic power and sonication time significantly influenced the degradation of phorate (p<0.05). The degradation of phorate followed the first-order kinetics model well. Phorate-oxon and phorate sulfoxide were identified as the degradation products of phorate by gas chromatography-mass spectrometry (GC-MS). Moreover, the toxicity of apple juice samples spiked with phorate was significantly reduced by sonication (p<0.05). The quality indexes of apple juice including pH, titratable acidity (TA), electrical conductivity (EC), total soluble solids (TSS), and the contents of sucrose, glucose and fructose were not affected by sonication, and no visible difference in color was observed between the sonicated samples and the control.

  15. Tachykinin acts upstream of autocrine Hedgehog signaling during nociceptive sensitization in Drosophila

    PubMed Central

    Im, Seol Hee; Takle, Kendra; Jo, Juyeon; Babcock, Daniel T; Ma, Zhiguo; Xiang, Yang; Galko, Michael J

    2015-01-01

    Pain signaling in vertebrates is modulated by neuropeptides like Substance P (SP). To determine whether such modulation is conserved and potentially uncover novel interactions between nociceptive signaling pathways we examined SP/Tachykinin signaling in a Drosophila model of tissue damage-induced nociceptive hypersensitivity. Tissue-specific knockdowns and genetic mutant analyses revealed that both Tachykinin and Tachykinin-like receptor (DTKR99D) are required for damage-induced thermal nociceptive sensitization. Electrophysiological recording showed that DTKR99D is required in nociceptive sensory neurons for temperature-dependent increases in firing frequency upon tissue damage. DTKR overexpression caused both behavioral and electrophysiological thermal nociceptive hypersensitivity. Hedgehog, another key regulator of nociceptive sensitization, was produced by nociceptive sensory neurons following tissue damage. Surprisingly, genetic epistasis analysis revealed that DTKR function was upstream of Hedgehog-dependent sensitization in nociceptive sensory neurons. Our results highlight a conserved role for Tachykinin signaling in regulating nociception and the power of Drosophila for genetic dissection of nociception. DOI: http://dx.doi.org/10.7554/eLife.10735.001 PMID:26575288

  16. Hedgehog pathway maintains cell survival under stress conditions, and drives drug resistance in lung adenocarcinoma

    PubMed Central

    Lin, Erh-Hsuan; Kao, Yu-Rung; Lin, Chih-An; Kuo, Ting-Yu; Yang, Sheng-Ping; Hsu, Chiung-Fang; Chou, Teh-Ying; Ho, Chao-Chi; Wu, Cheng-Wen

    2016-01-01

    Hedgehog (HH) pathway plays an important role in embryonic development, but is largely inactive in adult except for tissue repair. Aberrant activation of HH pathway has been found in a variety of cancer types. In non-small cell lung cancer, however, the role and importance of HH pathway remain controversial. In the current study, we found that HH pathway was maintained in low activity in lung adenocarcinoma (LAC) cells under normal culture condition, but was highly induced in response to stress conditions. Activation of HH pathway promoted cell survival, growth, and invasion partially through HGF and MET signaling. Hedgehog-Interacting Protein (HHIP), a cell-surface negative regulator of HH pathway, was epigenetically silenced in LAC. Overexpression of HHIP blocked the activation of HH and HGF/MET pathways, and made cells significantly more susceptible to stress conditions. In LAC cells with acquired resistance to Epidermal Growth Factor Receptor Tyrosin Kinase Inhibitor (EGFR-TKI), we found that a part of tumor cells were much more sensitive to HH or HGF/MET inhibitors, suggesting an oncogenic addiction shift from EGFR to HH and HGF/MET pathways. In conclusion, this study showed that HH pathway is a survival signaling that drives LAC cell growth under stress conditions, and HHIP is a key regulator to block the induction of HH pathway. Targeting the HH pathway through inhibitors or HHIP thus holds promise to address EGFR-TKI resistance in LAC in clinic. PMID:27015549

  17. Tachykinin acts upstream of autocrine Hedgehog signaling during nociceptive sensitization in Drosophila.

    PubMed

    Im, Seol Hee; Takle, Kendra; Jo, Juyeon; Babcock, Daniel T; Ma, Zhiguo; Xiang, Yang; Galko, Michael J

    2015-11-17

    Pain signaling in vertebrates is modulated by neuropeptides like Substance P (SP). To determine whether such modulation is conserved and potentially uncover novel interactions between nociceptive signaling pathways we examined SP/Tachykinin signaling in a Drosophila model of tissue damage-induced nociceptive hypersensitivity. Tissue-specific knockdowns and genetic mutant analyses revealed that both Tachykinin and Tachykinin-like receptor (DTKR99D) are required for damage-induced thermal nociceptive sensitization. Electrophysiological recording showed that DTKR99D is required in nociceptive sensory neurons for temperature-dependent increases in firing frequency upon tissue damage. DTKR overexpression caused both behavioral and electrophysiological thermal nociceptive hypersensitivity. Hedgehog, another key regulator of nociceptive sensitization, was produced by nociceptive sensory neurons following tissue damage. Surprisingly, genetic epistasis analysis revealed that DTKR function was upstream of Hedgehog-dependent sensitization in nociceptive sensory neurons. Our results highlight a conserved role for Tachykinin signaling in regulating nociception and the power of Drosophila for genetic dissection of nociception.

  18. Hedgehog pathway maintains cell survival under stress conditions, and drives drug resistance in lung adenocarcinoma.

    PubMed

    Lin, Erh-Hsuan; Kao, Yu-Rung; Lin, Chih-An; Kuo, Ting-Yu; Yang, Sheng-Ping; Hsu, Chiung-Fang; Chou, Teh-Ying; Ho, Chao-Chi; Wu, Cheng-Wen

    2016-04-26

    Hedgehog (HH) pathway plays an important role in embryonic development, but is largely inactive in adult except for tissue repair. Aberrant activation of HH pathway has been found in a variety of cancer types. In non-small cell lung cancer, however, the role and importance of HH pathway remain controversial. In the current study, we found that HH pathway was maintained in low activity in lung adenocarcinoma (LAC) cells under normal culture condition, but was highly induced in response to stress conditions. Activation of HH pathway promoted cell survival, growth, and invasion partially through HGF and MET signaling. Hedgehog-Interacting Protein (HHIP), a cell-surface negative regulator of HH pathway, was epigenetically silenced in LAC. Overexpression of HHIP blocked the activation of HH and HGF/MET pathways, and made cells significantly more susceptible to stress conditions. In LAC cells with acquired resistance to Epidermal Growth Factor Receptor Tyrosin Kinase Inhibitor (EGFR-TKI), we found that a part of tumor cells were much more sensitive to HH or HGF/MET inhibitors, suggesting an oncogenic addiction shift from EGFR to HH and HGF/MET pathways. In conclusion, this study showed that HH pathway is a survival signaling that drives LAC cell growth under stress conditions, and HHIP is a key regulator to block the induction of HH pathway. Targeting the HH pathway through inhibitors or HHIP thus holds promise to address EGFR-TKI resistance in LAC in clinic. PMID:27015549

  19. The Effect of Sonic Booms on Earthquake Warning Systems

    NASA Technical Reports Server (NTRS)

    Wurman, Gilead; Haering, Edward A, Jr.; Price, Michael J.

    2011-01-01

    Several aerospace companies are designing quiet supersonic business jets for service over the United States. These aircraft have the potential to increase the occurrence of mild sonic booms across the country. This leads to interest among earthquake warning (EQW) developers and the general seismological community in characterizing the effect of sonic booms on seismic sensors in the field, their potential impact on EQW systems, and means of discriminating their signatures from those of earthquakes. The SonicBREWS project (Sonic Boom Resistant Earthquake Warning Systems) is a collaborative effort between Seismic Warning Systems, Inc. (SWS) and NASA Dryden Flight Research Center. This project aims to evaluate the effects of sonic booms on EQW sensors. The study consists of exposing high-sample-rate (1000 sps) triaxial accelerometers to sonic booms with overpressures ranging from 10 to 600 Pa in the free field and the built environment. The accelerometers record the coupling of the sonic boom to the ground and surrounding structures, while microphones record the acoustic wave above ground near the sensor. Sonic booms are broadband signals with more high-frequency content than earthquakes. Even a 1000 sps accelerometer will produce a significantly aliased record. Thus the observed peak ground velocity is strongly dependent on the sampling rate, and increases as the sampling rate is reduced. At 1000 sps we observe ground velocities that exceed those of P-waves from ML 3 earthquakes at local distances, suggesting that sonic booms are not negligible for EQW applications. We present the results of several experiments conducted under SonicBREWS showing the effects of typical-case low amplitude sonic booms and worst-case high amplitude booms. We show the effects of various sensor placements and sensor array geometries. Finally, we suggest possible avenues for discriminating sonic booms from earthquakes for the purposes of EQW.

  20. Research at NASA on Human Response to Sonic Booms

    NASA Technical Reports Server (NTRS)

    Sullivan, Brenda M.

    2008-01-01

    NASA used its sonic boom simulator to study human response to shaped sonic booms and concluded that a loudness metric, such as Perceived Level, predicts human reaction to outdoor booms more accurately than overpressure. To investigate the importance of indoor phenomena (rattle, reverberation) under controlled laboratory conditions, NASA is building an "indoor sonic boom simulator." The intention is to develop a psychoacoustic model that describes human response as a function of boom shape (spectrum), boom intensity, reverberation, and varying rattle characteristics.

  1. Simulator Study of Indoor Annoyance Caused by Shaped Sonic Boom Stimuli With and Without Rattle Augmentation

    NASA Technical Reports Server (NTRS)

    Rathsam, Jonathan; Loubeau, Alexandra; Klos, Jacob

    2013-01-01

    The National Aeronautics and Space Administration's High Speed Project is developing a predictive capability for annoyance caused by shaped sonic booms transmitted indoors. The predictive capability is intended for use by aircraft designers as well as by aircraft noise regulators who are considering lifting the current prohibition on overland civil supersonic flight. The goal of the current study is to use an indoor simulator to validate two models developed using headphone tests for annoyance caused by sonic booms with and without rattle augmentation. The predictors in the proposed models include Moore and Glasberg's Stationary Loudness Level, the time derivative of Moore and Glasberg's time-varying short-term Loudness Level, and the difference between two weighted sound exposure levels, CSEL-ASEL. The indoor simulator provides a more realistic listening environment than headphones due to lowfrequency sound reproduction down to 6 Hz, which also causes perceptible tactile vibration. The results of this study show that a model consisting of {PL + (CSEL-ASEL)} is a reliable predictor of annoyance caused by shaped sonic booms alone, rattle sounds alone, and shaped sonic booms and rattle sounds together.

  2. Amygdalar connections in the lesser hedgehog tenrec.

    PubMed

    Künzle, Heinz

    2012-01-01

    The present study analyses the overall extrinsic connectivity of the non-olfactory amygdala (Ay) in the lesser hedgehog tenrec. The data were obtained from tracer injections into the lateral and intermediate portions of the Ay as well as several non-amygdalar brain regions. Both the solitary and the parabrachial nucleus receive descending projections from the central nucleus of the Ay, but only the parabrachial nucleus appears to project to the Ay. There is one prominent region in the ventromedial hypothalamus connected reciprocally with the medial and central Ay. Amygdalar afferents clearly arise from the dorsomedial thalamus, the subparafascicular nuclei and the medial geniculate complex (GM). Similar to other subprimate species, the latter projections originate in the dorsal and most caudal geniculate portions and terminate in the dorsolateral Ay. Unusual is the presence of amygdalo-projecting cells in the marginal geniculate zone and their virtual absence in the medial GM. As in other species, amygdalo-striatal projections mainly originate in the basolateral Ay and terminate predominantly in the ventral striatum. Given the poor differentiation of the tenrec's neocortex, there is a remarkable similarity with regard to the amygdalo-cortical connectivity between tenrec and rat, particularly as to prefrontal, limbic and somatosensorimotor areas as well as the rhinal cortex throughout its length. The tenrec's isocortex dorsomedial to the caudal rhinal cortex, on the other hand, may not be connected with the Ay. An absence of such connections is expected for primary auditory and visual fields, but it is unusual for their secondary fields.

  3. The dawn of hedgehog inhibitors: Vismodegib.

    PubMed

    Sandhiya, Selvarajan; Melvin, George; Kumar, Srinivasamurthy Suresh; Dkhar, Steven Aibor

    2013-01-01

    Cancer, one of the leading causes of death worldwide is estimated to increase to approximately 13.1 million by 2030. This has amplified the research in oncology towards the exploration of novel targets. Recently there has been lots of interest regarding the hedgehog (Hh) pathway, which plays a significant role in the development of organs and tissues during embryonic and postnatal periods. In a normal person, the Hh signaling pathway is under inhibition and gets activated upon the binding of Hh ligand to a transmembrane receptor called Patched (PTCH1) thus allowing the transmembrane protein, smoothened (SMO) to transfer signals through various proteins. One of the newer drugs namely vismodegib involves the inhibition of Hh pathway and has shown promising results in the treatment of advanced basal-cell carcinoma as well as medulloblastoma. It has been granted approval by US Food and Drug Administration's (US FDA) priority review program on January 30, 2012 for the treatment of advanced basal-cell carcinoma. The drug is also being evaluated in malignancies like medulloblastoma, pancreatic cancer, multiple myeloma, chondrosarcoma and prostate cancer. Moreover various Hh inhibitors namely LDE 225, saridegib, BMS 833923, LEQ 506, PF- 04449913 and TAK-441 are also undergoing phase I and II trials for different neoplasms. Hence this review will describe briefly the Hh pathway and the novel drug vismodegib.

  4. dachshund Potentiates Hedgehog Signaling during Drosophila Retinogenesis

    PubMed Central

    Aerts, Stein; Casares, Fernando; Janody, Florence

    2016-01-01

    Proper organ patterning depends on a tight coordination between cell proliferation and differentiation. The patterning of Drosophila retina occurs both very fast and with high precision. This process is driven by the dynamic changes in signaling activity of the conserved Hedgehog (Hh) pathway, which coordinates cell fate determination, cell cycle and tissue morphogenesis. Here we show that during Drosophila retinogenesis, the retinal determination gene dachshund (dac) is not only a target of the Hh signaling pathway, but is also a modulator of its activity. Using developmental genetics techniques, we demonstrate that dac enhances Hh signaling by promoting the accumulation of the Gli transcription factor Cubitus interruptus (Ci) parallel to or downstream of fused. In the absence of dac, all Hh-mediated events associated to the morphogenetic furrow are delayed. One of the consequences is that, posterior to the furrow, dac- cells cannot activate a Roadkill-Cullin3 negative feedback loop that attenuates Hh signaling and which is necessary for retinal cells to continue normal differentiation. Therefore, dac is part of an essential positive feedback loop in the Hh pathway, guaranteeing the speed and the accuracy of Drosophila retinogenesis. PMID:27442438

  5. Production of systemically circulating Hedgehog by the intestine couples nutrition to growth and development.

    PubMed

    Rodenfels, Jonathan; Lavrynenko, Oksana; Ayciriex, Sophie; Sampaio, Julio L; Carvalho, Maria; Shevchenko, Andrej; Eaton, Suzanne

    2014-12-01

    In Drosophila larvae, growth and developmental timing are regulated by nutrition in a tightly coordinated fashion. The networks that couple these processes are far from understood. Here, we show that the intestine responds to nutrient availability by regulating production of a circulating lipoprotein-associated form of the signaling protein Hedgehog (Hh). Levels of circulating Hh tune the rates of growth and developmental timing in a coordinated fashion. Circulating Hh signals to the fat body to control larval growth. It regulates developmental timing by controlling ecdysteroid production in the prothoracic gland. Circulating Hh is especially important during starvation, when it is also required for mobilization of fat body triacylglycerol (TAG) stores. Thus, we demonstrate that Hh, previously known only for its local morphogenetic functions, also acts as a lipoprotein-associated endocrine hormone, coordinating the response of multiple tissues to nutrient availability.

  6. Multidisciplinary design optimization for sonic boom mitigation

    NASA Astrophysics Data System (ADS)

    Ozcer, Isik A.

    Automated, parallelized, time-efficient surface definition and grid generation and flow simulation methods are developed for sharp and accurate sonic boom signal computation in three dimensions in the near and mid-field of an aircraft using Euler/Full-Potential unstructured/structured computational fluid dynamics. The full-potential mid-field sonic boom prediction code is an accurate and efficient solver featuring automated grid generation, grid adaptation and shock fitting, and parallel processing. This program quickly marches the solution using a single nonlinear equation for large distances that cannot be covered with Euler solvers due to large memory and long computational time requirements. The solver takes into account variations in temperature and pressure with altitude. The far-field signal prediction is handled using the classical linear Thomas Waveform Parameter Method where the switching altitude from the nonlinear to linear prediction is determined by convergence of the ground signal pressure impulse value. This altitude is determined as r/L ≈ 10 from the source for a simple lifting wing, and r/L ≈ 40 for a real complex aircraft. Unstructured grid adaptation and shock fitting methodology developed for the near-field analysis employs an Hessian based anisotropic grid adaptation based on error equidistribution. A special field scalar is formulated to be used in the computation of the Hessian based error metric which enhances significantly the adaptation scheme for shocks. The entire cross-flow of a complex aircraft is resolved with high fidelity using only 500,000 grid nodes after only about 10 solution/adaptation cycles. Shock fitting is accomplished using Roe's Flux-Difference Splitting scheme which is an approximate Riemann type solver and by proper alignment of the cell faces with respect to shock surfaces. Simple to complex real aircraft geometries are handled with no user-interference required making the simulation methods suitable tools for

  7. Control of inertial acoustic cavitation in pulsed sonication using a real-time feedback loop system.

    PubMed

    Desjouy, Cyril; Poizat, Adrien; Gilles, Bruno; Inserra, Claude; Bera, Jean-Christophe

    2013-08-01

    Owing to the complex behavior of ultrasound-induced bubble clouds (nucleation, linear and nonlinear oscillations, collapse), acoustic cavitation remains a hardly controllable phenomenon, leading to poorly reproducible ultrasound-based therapies. A better control of the various aspects of cavitation phenomena for in vivo applications is a key requirement to improve emerging ultrasound therapies. Previous publications have reported on systems performing regulation of acoustic cavitation in continuous sonication when applied in vitro, but the main challenge today is to achieve real-time control of cavitation activity in pulsed sonication when used in vivo. The present work aims at developing a system to control acoustic cavitation in a pulsed wave condition using a real-time feedback loop. The experimental setup consists of a water bath in which is submerged a focused transducer (pulsed waves, frequency 550 kHz) used for sonication and a hydrophone used to listen to inertial cavitation. The designed regulation process allows the cavitation activity to be controlled through a 300 μs feedback loop. Without regulation, cavitation exhibits numerous bursts of intense activity and large variations of inertial cavitation level over time. In a regulated regime, the control of inertial cavitation activity within a pulse leads to consistent cavitation levels over time with an enhancement of the reproducibility.

  8. Novel Hedgehog pathway targets against basal cell carcinoma

    SciTech Connect

    Tang, Jean Y. So, P.-L.; Epstein, Ervin H.

    2007-11-01

    The Hedgehog signaling pathway plays a key role in directing growth and patterning during embryonic development and is required in vertebrates for the normal development of many structures, including the neural tube, axial skeleton, skin, and hair. Aberrant activation of the Hedgehog (Hh) pathway in adult tissue is associated with the development of basal cell carcinoma (BCC), medulloblastoma, and a subset of pancreatic, gastrointestinal, and other cancers. This review will provide an overview of what is known about the mechanisms by which activation of Hedgehog signaling leads to the development of BCCs and will review two recent papers suggesting that agents that modulate sterol levels might influence the Hh pathway. Thus, sterols may be a new therapeutic target for the treatment of BCCs, and readily available agents such as statins (HMG-CoA reductase inhibitors) or vitamin D might be helpful in reducing BCC incidence.

  9. High-Speed Research: Sonic Boom, volume 2

    NASA Technical Reports Server (NTRS)

    Darden, Christine M. (Compiler)

    1992-01-01

    A High-Speed Sonic Boom Workshop was held at NASA Langley Research Center on February 25-27, 1992. The purpose of the workshop was to make presentations on current research activities and accomplishments and to assess progress in the area of sonic boom since the program was initiated in FY-90. Twenty-nine papers were presented during the 2-1/2 day workshop. Attendees included representatives from academia, industry, and government who are actively involved in sonic-boom research. Volume 2 contains papers related to low sonic-boom design and analysis using both linear theory and higher order computational fluid dynamics (CFD) methods.

  10. High-Speed Research: Sonic Boom, volume 1

    NASA Technical Reports Server (NTRS)

    Darden, Christine M. (Compiler)

    1992-01-01

    A High-Speed Sonic Boom Workshop was held at LaRC of Feb. 25-27, 1992. The purpose was to make presentations on current research activities and accomplishments and to assess progress in the area of sonic boom since the program was initiated in FY-90. Twenty-nine papers were presented during the 2-1/2 day workshop. Attendees included representatives from academia, industry, and government who are actively involved in sonic-boom research. Volume 1 contains papers related to atmospheric effects on the sonic-boom signature during propagation and on acceptability studies.

  11. Sonic anemometers in aeolian sediment transport research

    NASA Astrophysics Data System (ADS)

    van Boxel, J. H.; Sterk, G.; Arens, S. M.

    2004-04-01

    Fast-response wind and turbulence instruments, including sonic anemometers, are used more and more in aeolian sediment transport research. These instruments give information on mean wind, but also on fluctuations and turbulent statistics, such as the uw covariance, which is a direct measure of Reynolds' stress (RS) and friction velocity. This paper discusses the interpretation of sonic anemometer data, the transformations needed to get proper results and turbulence spectra, and how they are influenced by instrument size, sampling frequency, and measurement height. Turbulence spectra characterize how much the different frequencies in the turbulent signals contribute to the variance of wind speed, or to the covariance of horizontal and vertical wind speed. They are important in determining the measurement strategy when working with fast-response instruments, such as sonic anemometers, and are useful for interpreting the measurement results. Choices on the type of sonic anemometer, observation height, sampling period, sampling frequency, and filtering can be made on the basis of expected high and low-frequency losses in turbulent signals, which are affected by those variables, as well as wind speed and atmospheric stability. Friction velocity and RS, important variables in aeolian sediment transport research, are very sensitive to tilt or slope errors. During a field experiment, the slope sensitivity of the RS was established as 9% per degree of slope, which is 1.5 times the value reported in literature on the basis of theoretical considerations. An important reason for the difference probably is the large influence of streamline curvature on turbulence statistics and thereby on the slope sensitivity of the RS. An error of 9% per degree of slope in the RS will translate into an error of approximately 4% per degree of slope in the calculated friction velocity. Space-time correlation of the horizontal wind speed is much larger than that of the vertical wind speed and

  12. Hedgehog Pathway Inhibition Radiosensitizes Non-Small Cell Lung Cancers

    SciTech Connect

    Zeng, Jing; Aziz, Khaled; Chettiar, Sivarajan T.; Aftab, Blake T.; Armour, Michael; Gajula, Rajendra; Gandhi, Nishant; Salih, Tarek; Herman, Joseph M.; Wong, John; Rudin, Charles M.; Tran, Phuoc T.; Hales, Russell K.

    2013-05-01

    Purpose: Despite improvements in chemoradiation, local control remains a major clinical problem in locally advanced non-small cell lung cancer. The Hedgehog pathway has been implicated in tumor recurrence by promoting survival of tumorigenic precursors and through effects on tumor-associated stroma. Whether Hedgehog inhibition can affect radiation efficacy in vivo has not been reported. Methods and Materials: We evaluated the effects of a targeted Hedgehog inhibitor (HhAntag) and radiation on clonogenic survival of human non-small cell lung cancer lines in vitro. Using an A549 cell line xenograft model, we examined tumor growth, proliferation, apoptosis, and gene expression changes after concomitant HhAntag and radiation. In a transgenic mouse model of Kras{sup G12D}-induced and Twist1-induced lung adenocarcinoma, we assessed tumor response to radiation and HhAntag by serial micro-computed tomography (CT) scanning. Results: In 4 human lung cancer lines in vitro, HhAntag showed little or no effect on radiosensitivity. By contrast, in both the human tumor xenograft and murine inducible transgenic models, HhAntag enhanced radiation efficacy and delayed tumor growth. By use of the human xenograft model to differentiate tumor and stromal effects, mouse stromal cells, but not human tumor cells, showed significant and consistent downregulation of Hedgehog pathway gene expression. This was associated with increased tumor cell apoptosis. Conclusions: Targeted Hedgehog pathway inhibition can increase in vivo radiation efficacy in lung cancer preclinical models. This effect is associated with pathway suppression in tumor-associated stroma. These data support clinical testing of Hedgehog inhibitors as a component of multimodality therapy for locally advanced non-small cell lung cancer.

  13. Ultrasonic/Sonic Rotary-Hammer Drills

    NASA Technical Reports Server (NTRS)

    Badescu, Mircea; Sherrit, Stewart; Bar-Cohen, Yoseph; Bao, Xiaoqi; Kassab, Steve

    2010-01-01

    Ultrasonic/sonic rotary-hammer drill (USRoHD) is a recent addition to the collection of apparatuses based on ultrasonic/sonic drill corer (USDC). As described below, the USRoHD has several features, not present in a basic USDC, that increase efficiency and provide some redundancy against partial failure. USDCs and related apparatuses were conceived for boring into, and/or acquiring samples of, rock or other hard, brittle materials of geological interest. They have been described in numerous previous NASA Tech Briefs articles. To recapitulate: A USDC can be characterized as a lightweight, lowpower, piezoelectrically driven jackhammer in which ultrasonic and sonic vibrations are generated and coupled to a tool bit. A basic USDC includes a piezoelectric stack, an ultrasonic transducer horn connected to the stack, a free mass ( free in the sense that it can bounce axially a short distance between hard stops on the horn and the bit), and a tool bit. The piezoelectric stack creates ultrasonic vibrations that are mechanically amplified by the horn. The bouncing of the free mass between the hard stops generates the sonic vibrations. The combination of ultrasonic and sonic vibrations gives rise to a hammering action (and a resulting chiseling action at the tip of the tool bit) that is more effective for drilling than is the microhammering action of ultrasonic vibrations alone. The hammering and chiseling actions are so effective that unlike in conventional twist drilling, little applied axial force is needed to make the apparatus advance into the material of interest. There are numerous potential applications for USDCs and related apparatuses in geological exploration on Earth and on remote planets. In early USDC experiments, it was observed that accumulation of cuttings in a drilled hole causes the rate of penetration of the USDC to decrease steeply with depth, and that the rate of penetration can be increased by removing the cuttings. The USRoHD concept provides for

  14. Crossed thalamocortical connections in the Madagascan hedgehog tenrec: dissimilarities to erinaceous hedgehog, similarities to mammals with more differentiated brains.

    PubMed

    Künzle, H

    1995-04-14

    The adult erinaceous hedgehog, unlike other mammals, has recently been shown to have prominent crossed projections from the thalamus to the motor cortex. There are suggestions relating this unique pattern of connectivity to the overall degree of brain differentiation and/or the poorly developed corpus callosum. The present tracing study demonstrates that the Madagascan lesser hedgehog tenrec, with its tiny corpus callosum and one of the lowest neocorticalization indices among insectivores, has extensive crossed cortico-thalamic projections, but essentially the same sparse thalamic projections to the contralateral cortex as have placental mammals with more differentiated brains. The implications of the findings and the relevance of extracallosal pathways are discussed.

  15. A different kind of hedgehog pathway: tinea manus due to Trichophyton erinacei transmitted by an African pygmy hedgehog (Atelerix albiventris).

    PubMed

    Weishaupt, Julia; Kolb-Mäurer, Annette; Lempert, Sigrid; Nenoff, Pietro; Uhrlaß, Silke; Hamm, Henning; Goebeler, Matthias

    2014-02-01

    The unusual case of a 29-year-old woman with tinea manus caused by infection due to Trichophyton erinacei is described. The patient presented with marked erosive inflammation of the entire fifth finger of her right hand. Mycological and genomic diagnostics resulted in identification of T. erinacei as the responsible pathogen, which had been transmitted by a domestic African pygmy hedgehog, Atelerix albiventris. Upon prolonged treatment with topical and systemic antifungal agents skin lesions slowly resolved. This case illustrates that the increasingly popular keeping of extraordinary pets such as hedgehogs may bear the risk of infections with uncommon dermatophytes.

  16. The Hedgehog gene family of the cnidarian, Nematostella vectensis, and implications for understanding metazoan Hedgehog pathway evolution

    PubMed Central

    Matus, David Q.; Magie, Craig; Pang, Kevin; Martindale, Mark Q; Thomsen, Gerald H.

    2008-01-01

    Hedgehog signaling is an important component of cell-cell communication during bilaterian development, and abnormal Hedgehog signaling contributes to disease and birth defects. Hedgehog genes are composed of a ligand (“hedge”) domain and an autocatalytic intein (“hog”) domain. Hedgehog (hh) ligands bind to a conserved set of receptors and activate downstream signal transduction pathways terminating with Gli/Ci transcription factors. We have identified five intein-containing genes in the anthozoan cnidarian Nematostella vectensis, two of which (NvHh1 and NvHh2) contain definitive hedgehog ligand domains, suggesting that to date, cnidarians are the earliest branching metazoan phylum to possess definitive Hh orthologs. Expression analysis of NvHh1 and NvHh2, the receptor NvPatched and a downstream transcription factor NvGli (a Gli3/Ci ortholog) indicate that these genes may have conserved roles in planar and trans-epithelial signaling during gut and germline development, while the three remaining intein-containing genes (NvHint1,2,3) are expressed in a cell-type specific manner in putative neural precursors. Metazoan intein-containing genes that lack a ligand domain have previously only been identified within nematodes. However, phylogenetic analyses suggest that these nematode inteins may be derived from an ancestral nematode true hedgehog gene, and that the non-bilaterian intein-containing genes identified here may represent an ancestral state prior to the domain swapping events that resulted in the formation of true hedgehog genes in the cnidarian-bilaterian ancestor. Genomic surveys of N. vectensis suggest that most of the components of both protostome and deuterostome Hh signaling pathways are present in anthozoans and that some appear to have been lost in ecdysozoan lineages. Cnidarians possess many bilaterian cell-cell signaling pathways (Wnt, TGFß, FGF and Hh) that appear to act in concert to pattern tissues along the oral-aboral axis of the polyp

  17. Hedgehog signaling is required for cranial neural crest morphogenesis and chondrogenesis at the midline in the zebrafish skull.

    PubMed

    Wada, Naoyuki; Javidan, Yashar; Nelson, Sarah; Carney, Thomas J; Kelsh, Robert N; Schilling, Thomas F

    2005-09-01

    Neural crest cells that form the vertebrate head skeleton migrate and interact with surrounding tissues to shape the skull, and defects in these processes underlie many human craniofacial syndromes. Signals at the midline play a crucial role in the development of the anterior neurocranium, which forms the ventral braincase and palate, and here we explore the role of Hedgehog (Hh) signaling in this process. Using sox10:egfp transgenics to follow neural crest cell movements in the living embryo, and vital dye labeling to generate a fate map, we show that distinct populations of neural crest form the two main cartilage elements of the larval anterior neurocranium: the paired trabeculae and the midline ethmoid. By analyzing zebrafish mutants that disrupt sonic hedgehog (shh) expression, we demonstrate that shh is required to specify the movements of progenitors of these elements at the midline, and to induce them to form cartilage. Treatments with cyclopamine, to block Hh signaling at different stages, suggest that although requirements in morphogenesis occur during neural crest migration beneath the brain, requirements in chondrogenesis occur later, as cells form separate trabecular and ethmoid condensations. Cell transplantations indicate that these also reflect different sources of Shh, one from the ventral neural tube that controls trabecular morphogenesis and one from the oral ectoderm that promotes chondrogenesis. Our results suggest a novel role for Shh in the movements of neural crest cells at the midline, as well as in their differentiation into cartilage, and help to explain why both skeletal fusions and palatal clefting are associated with the loss of Hh signaling in holoprosencephalic humans.

  18. 14 CFR 91.817 - Civil aircraft sonic boom.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 14 Aeronautics and Space 2 2012-01-01 2012-01-01 false Civil aircraft sonic boom. 91.817 Section....817 Civil aircraft sonic boom. (a) No person may operate a civil aircraft in the United States at a... may operate a civil aircraft for which the maximum operating limit speed MM0 exceeds a Mach number...

  19. 14 CFR 91.817 - Civil aircraft sonic boom.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 14 Aeronautics and Space 2 2013-01-01 2013-01-01 false Civil aircraft sonic boom. 91.817 Section....817 Civil aircraft sonic boom. (a) No person may operate a civil aircraft in the United States at a... may operate a civil aircraft for which the maximum operating limit speed MM0 exceeds a Mach number...

  20. 14 CFR 91.817 - Civil aircraft sonic boom.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 14 Aeronautics and Space 2 2014-01-01 2014-01-01 false Civil aircraft sonic boom. 91.817 Section....817 Civil aircraft sonic boom. (a) No person may operate a civil aircraft in the United States at a... may operate a civil aircraft for which the maximum operating limit speed MM0 exceeds a Mach number...

  1. 14 CFR 91.817 - Civil aircraft sonic boom.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 2 2010-01-01 2010-01-01 false Civil aircraft sonic boom. 91.817 Section....817 Civil aircraft sonic boom. (a) No person may operate a civil aircraft in the United States at a... may operate a civil aircraft for which the maximum operating limit speed MM0 exceeds a Mach number...

  2. 14 CFR 91.817 - Civil aircraft sonic boom.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 14 Aeronautics and Space 2 2011-01-01 2011-01-01 false Civil aircraft sonic boom. 91.817 Section....817 Civil aircraft sonic boom. (a) No person may operate a civil aircraft in the United States at a... may operate a civil aircraft for which the maximum operating limit speed MM0 exceeds a Mach number...

  3. Cell context-specific expression of primary cilia in the human testis and ciliary coordination of Hedgehog signalling in mouse Leydig cells.

    PubMed

    Nygaard, Marie Berg; Almstrup, Kristian; Lindbæk, Louise; Christensen, Søren Tvorup; Svingen, Terje

    2015-01-01

    Primary cilia are sensory organelles that coordinate numerous cellular signalling pathways during development and adulthood. Defects in ciliary assembly or function lead to a series of developmental disorders and diseases commonly referred to as ciliopathies. Still, little is known about the formation and function of primary cilia in the mammalian testis. Here, we characterized primary cilia in adult human testis and report a constitutive expression of cilia in peritubular myoid cells and a dynamic expression of cilia in differentiating Leydig cells. Primary cilia are generally absent from cells of mature seminiferous epithelium, but present in Sertoli cell-only tubules in Klinefelter syndrome testis. Peritubular cells in atrophic testis produce overly long cilia. Furthermore cultures of growth-arrested immature mouse Leydig cells express primary cilia that are enriched in components of Hedgehog signalling, including Smoothened, Patched-1, and GLI2, which are involved in regulating Leydig cell differentiation. Stimulation of Hedgehog signalling increases the localization of Smoothened to the cilium, which is followed by transactivation of the Hedgehog target genes, Gli1 and Ptch1. Our findings provide new information on the spatiotemporal formation of primary cilia in the testis and show that primary cilia in immature Leydig cells mediate Hedgehog signalling. PMID:25992706

  4. A local difference in Hedgehog signal transduction increases mechanical cell bond tension and biases cell intercalations along the Drosophila anteroposterior compartment boundary.

    PubMed

    Rudolf, Katrin; Umetsu, Daiki; Aliee, Maryam; Sui, Liyuan; Jülicher, Frank; Dahmann, Christian

    2015-11-15

    Tissue organization requires the interplay between biochemical signaling and cellular force generation. The formation of straight boundaries separating cells with different fates into compartments is important for growth and patterning during tissue development. In the developing Drosophila wing disc, maintenance of the straight anteroposterior (AP) compartment boundary involves a local increase in mechanical tension at cell bonds along the boundary. The biochemical signals that regulate mechanical tension along the AP boundary, however, remain unknown. Here, we show that a local difference in Hedgehog signal transduction activity between anterior and posterior cells is necessary and sufficient to increase mechanical tension along the AP boundary. This difference in Hedgehog signal transduction is also required to bias cell rearrangements during cell intercalations to keep the characteristic straight shape of the AP boundary. Moreover, severing cell bonds along the AP boundary does not reduce tension at neighboring bonds, implying that active mechanical tension is upregulated, cell bond by cell bond. Finally, differences in the expression of the homeodomain-containing protein Engrailed also contribute to the straight shape of the AP boundary, independently of Hedgehog signal transduction and without modulating cell bond tension. Our data reveal a novel link between local differences in Hedgehog signal transduction and a local increase in active mechanical tension of cell bonds that biases junctional rearrangements. The large-scale shape of the AP boundary thus emerges from biochemical signals inducing patterns of active tension on cell bonds.

  5. Cell context-specific expression of primary cilia in the human testis and ciliary coordination of Hedgehog signalling in mouse Leydig cells.

    PubMed

    Nygaard, Marie Berg; Almstrup, Kristian; Lindbæk, Louise; Christensen, Søren Tvorup; Svingen, Terje

    2015-01-01

    Primary cilia are sensory organelles that coordinate numerous cellular signalling pathways during development and adulthood. Defects in ciliary assembly or function lead to a series of developmental disorders and diseases commonly referred to as ciliopathies. Still, little is known about the formation and function of primary cilia in the mammalian testis. Here, we characterized primary cilia in adult human testis and report a constitutive expression of cilia in peritubular myoid cells and a dynamic expression of cilia in differentiating Leydig cells. Primary cilia are generally absent from cells of mature seminiferous epithelium, but present in Sertoli cell-only tubules in Klinefelter syndrome testis. Peritubular cells in atrophic testis produce overly long cilia. Furthermore cultures of growth-arrested immature mouse Leydig cells express primary cilia that are enriched in components of Hedgehog signalling, including Smoothened, Patched-1, and GLI2, which are involved in regulating Leydig cell differentiation. Stimulation of Hedgehog signalling increases the localization of Smoothened to the cilium, which is followed by transactivation of the Hedgehog target genes, Gli1 and Ptch1. Our findings provide new information on the spatiotemporal formation of primary cilia in the testis and show that primary cilia in immature Leydig cells mediate Hedgehog signalling.

  6. Netrin 1 regulates blood-brain barrier function and neuroinflammation.

    PubMed

    Podjaski, Cornelia; Alvarez, Jorge I; Bourbonniere, Lyne; Larouche, Sandra; Terouz, Simone; Bin, Jenea M; Lécuyer, Marc-André; Saint-Laurent, Olivia; Larochelle, Catherine; Darlington, Peter J; Arbour, Nathalie; Antel, Jack P; Kennedy, Timothy E; Prat, Alexandre

    2015-06-01

    Blood-brain barrier function is driven by the influence of astrocyte-secreted factors. During neuroinflammatory responses the blood-brain barrier is compromised resulting in central nervous system damage and exacerbated pathology. Here, we identified endothelial netrin 1 induction as a vascular response to astrocyte-derived sonic hedgehog that promotes autocrine barrier properties during homeostasis and increases with inflammation. Netrin 1 supports blood-brain barrier integrity by upregulating endothelial junctional protein expression, while netrin 1 knockout mice display disorganized tight junction protein expression and barrier breakdown. Upon inflammatory conditions, blood-brain barrier endothelial cells significantly upregulated netrin 1 levels in vitro and in situ, which prevented junctional breach and endothelial cell activation. Finally, netrin 1 treatment during experimental autoimmune encephalomyelitis significantly reduced blood-brain barrier disruption and decreased clinical and pathological indices of disease severity. Our results demonstrate that netrin 1 is an important regulator of blood-brain barrier maintenance that protects the central nervous system against inflammatory conditions such as multiple sclerosis and experimental autoimmune encephalomyelitis.

  7. Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) Pathway Is Induced by Mechanical Load and Reduces the Activity of Hedgehog Signaling in Chondrogenic Micromass Cell Cultures.

    PubMed

    Juhász, Tamás; Szentléleky, Eszter; Somogyi, Csilla Szűcs; Takács, Roland; Dobrosi, Nóra; Engler, Máté; Tamás, Andrea; Reglődi, Dóra; Zákány, Róza

    2015-01-01

    Pituitary adenylate cyclase activating polypeptide (PACAP) is a neurohormone exerting protective function during various stress conditions either in mature or developing tissues. Previously we proved the presence of PACAP signaling elements in chicken limb bud-derived chondrogenic cells in micromass cell cultures. Since no data can be found if PACAP signaling is playing any role during mechanical stress in any tissues, we aimed to investigate its contribution in mechanotransduction during chondrogenesis. Expressions of the mRNAs of PACAP and its major receptor, PAC1 increased, while that of other receptors, VPAC1, VPAC2 decreased upon mechanical stimulus. Mechanical load enhanced the expression of collagen type X, a marker of hypertrophic differentiation of chondrocytes and PACAP addition attenuated this elevation. Moreover, exogenous PACAP also prevented the mechanical load evoked activation of hedgehog signaling: protein levels of Sonic and Indian Hedgehogs and Gli1 transcription factor were lowered while expressions of Gli2 and Gli3 were elevated by PACAP application during mechanical load. Our results suggest that mechanical load activates PACAP signaling and exogenous PACAP acts against the hypertrophy inducing effect of mechanical load. PMID:26230691