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Sample records for receptor mediates migration

  1. Extracellular acidosis impairs P2Y receptor-mediated Ca(2+) signalling and migration of microglia.

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    Langfelder, Antonia; Okonji, Emeka; Deca, Diana; Wei, Wei-Chun; Glitsch, Maike D

    2015-04-01

    Microglia are the resident macrophage and immune cell of the brain and are critically involved in combating disease and assaults on the brain. Virtually all brain pathologies are accompanied by acidosis of the interstitial fluid, meaning that microglia are exposed to an acidic environment. However, little is known about how extracellular acidosis impacts on microglial function. The activity of microglia is tightly controlled by 'on' and 'off' signals, the presence or absence of which results in generation of distinct phenotypes in microglia. Activation of G protein coupled purinergic (P2Y) receptors triggers a number of distinct behaviours in microglia, including activation, migration, and phagocytosis. Using pharmacological tools and fluorescence imaging of the murine cerebellar microglia cell line C8B4, we show that extracellular acidosis interferes with P2Y receptor-mediated Ca(2+) signalling in these cells. Distinct P2Y receptors give rise to signature intracellular Ca(2+) signals, and Ca(2+) release from stores and Ca(2+) influx are differentially affected by acidotic conditions: Ca(2+) release is virtually unaffected, whereas Ca(2+) influx, mediated at least in part by store-operated Ca(2+) channels, is profoundly inhibited. Furthermore, P2Y1 and P2Y6-mediated stimulation of migration is inhibited under conditions of extracellular acidosis, whereas basal migration independent of P2Y receptor activation is not. Taken together, our results demonstrate that an acidic microenvironment impacts on P2Y receptor-mediated Ca(2+) signalling, thereby influencing microglial responses and responsiveness to extracellular signals. This may result in altered behaviour of microglia under pathological conditions compared with microglial responses in healthy tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. The Innate Immune Receptor CD14 Mediates Lymphocyte Migration in EAE

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    Ramona Halmer

    2015-08-01

    Full Text Available Background: Multiple sclerosis is the most common autoimmune disease of the central nervous system in young adults and histopathologically characterized by inflammation, demyelination and gliosis. It is considered as a CD4+ T cell-mediated disease, but also a disease-promoting role of the innate immune system has been proposed, based e.g. on the observation that innate immune receptors modulate disease severity of experimental autoimmune encephalomyelitis. Recent studies of our group provided first evidence for a key role of the innate immune LPS receptor (CD14 in pathophysiology of experimental autoimmune encephalomyelitis. CD14-deficient experimental autoimmune encephalomyelitis mice showed increased clinical symptoms and enhanced infiltration of monocytes and neutrophils in brain and spinal cord. Methods: In the current study, we further investigated the causes of the disease aggravation by CD14-deficiency and examined T cell activation, also focusing on the costimulatory molecules CTLA-4 and CD28, and T cell migration capacity over the blood brain barrier by FACS analysis, in vitro adhesion and transmigration assays. Results: In the results, we observed a significantly increased migration of CD14-deficient lymphocytes across an endothelial monolayer. In contrast, we did not see any differences in expression levels of TCR/CTLA-4 or TCR/CD28 and lymphocyte adhesion to endothelial cells from CD14-deficient compared to wildtype mice. Conclusion: The results demonstrate an important role of CD14 in migration of lymphocytes, and strengthen the importance of innate immune receptors in adaptive immune disorders, such as multiple sclerosis.

  3. Neuroblast Migration and P2Y1 Receptor Mediated Calcium Signalling Depend on 9-O-Acetyl GD3 Ganglioside

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    Marcelo F Santiago

    2012-08-01

    Full Text Available Previous studies indicated that a ganglioside 9acGD3 (9-O-acetyl GD3 antibody [the J-Ab (Jones antibody] reduces GCP (granule cell progenitor migration in vitro and in vivo. We here investigated, using cerebellar explants of postnatal day (P 6 mice, the mechanism by which 9acGD3 reduces GCP migration. We found that immunoblockade of the ganglioside with the J-Ab or the lack of GD3 synthase reduced GCP in vitro migration and the frequency of Ca2+ oscillations. Immunocytochemistry and pharmacological assays indicated that GCPs expressed P2Y1Rs (P2Y1 receptors and that deletion or blockade of these receptors decreased the migration rate of GCPs and the frequency of Ca2+ oscillations. The reduction in P2Y1-mediated calcium signals seen in Jones-treated and GD3 synthase-null GCPs were paralleled by P2Y1R internalization. We conclude that 9acGD3 controls GCP migration by influencing P2Y1R cellular distribution and function.

  4. Modulation of the leptin receptor mediates tumor growth and migration of pancreatic cancer cells.

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    Alisha M Mendonsa

    Full Text Available Obesity has been implicated as a significant risk factor for development of pancreatic cancer. In the setting of obesity, a systemic chronic inflammatory response is characterized by alterations in the production and secretion of a wide variety of growth factors. Leptin is a hormone whose level increases drastically in the serum of obese patients. High fat diet induced obesity in mice leads to an overall increased body weight, pancreatic weight, serum leptin, and pancreatic tissue leptin levels. Here we report the contribution of obesity and leptin to pancreatic cancer growth utilizing an in vivo orthotopic murine pancreatic cancer model, which resulted in increased tumor proliferation with concomitant increased tumor burden in the diet induced obese mice compared to lean mice. Human and murine pancreatic cancer cell lines were found to express the short as well as the long form of the leptin receptor and functionally responded to leptin induced activation through an increased phosphorylation of AKT473. In vitro, leptin stimulation increased cellular migration which was blocked by addition of a PI3K inhibitor. In vivo, depletion of the leptin receptor through shRNA knockdown partially abrogated increased orthotopic tumor growth in obese mice. These findings suggest that leptin contributes to pancreatic tumor growth through activation of the PI3K/AKT pathway, which promotes pancreatic tumor cell migration.

  5. Modulation of the leptin receptor mediates tumor growth and migration of pancreatic cancer cells.

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    Mendonsa, Alisha M; Chalfant, Madeleine C; Gorden, Lee D; VanSaun, Michael N

    2015-01-01

    Obesity has been implicated as a significant risk factor for development of pancreatic cancer. In the setting of obesity, a systemic chronic inflammatory response is characterized by alterations in the production and secretion of a wide variety of growth factors. Leptin is a hormone whose level increases drastically in the serum of obese patients. High fat diet induced obesity in mice leads to an overall increased body weight, pancreatic weight, serum leptin, and pancreatic tissue leptin levels. Here we report the contribution of obesity and leptin to pancreatic cancer growth utilizing an in vivo orthotopic murine pancreatic cancer model, which resulted in increased tumor proliferation with concomitant increased tumor burden in the diet induced obese mice compared to lean mice. Human and murine pancreatic cancer cell lines were found to express the short as well as the long form of the leptin receptor and functionally responded to leptin induced activation through an increased phosphorylation of AKT473. In vitro, leptin stimulation increased cellular migration which was blocked by addition of a PI3K inhibitor. In vivo, depletion of the leptin receptor through shRNA knockdown partially abrogated increased orthotopic tumor growth in obese mice. These findings suggest that leptin contributes to pancreatic tumor growth through activation of the PI3K/AKT pathway, which promotes pancreatic tumor cell migration.

  6. Zinc Ions Mediate Gastrin Expression, Proliferation, and Migration Downstream of the Cholecystokinin-2 Receptor.

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    Chang, Mike; Xiao, Lin; Shulkes, Arthur; Baldwin, Graham S; Patel, Oneel

    2016-12-01

    Gastrin, acting via the cholecystokinin-2 receptor (CCK2R), activates its own promoter in a positive-feed-forward loop that may result in hypergastrinemia. Activity of the gastrin promoter is also stimulated by exogenous Zn2+ ions. Here, the role of intracellular zinc and calcium signaling in the gastrin positive-feed-forward loop was investigated. Gastrin promoter activity was measured in the human gastric carcinoma cell line AGS-CCK2R and in Jurkat cells transfected with various gastrin promoter-luciferase constructs after treatment with gastrin in the presence and absence of zinc- and calcium-chelating agents. The free intracellular zinc ion concentrations were measured in the same cells with the fluorescent indicator FluoZin-3. Cell proliferation and migration/invasion were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide cell proliferation assay and in Boyden chamber assays, respectively. The zinc chelator N,N,N,N-tetrakis-(2-pyridylmethyl)-ethylenediamine (TPEN) abolished gastrin-stimulated gastrin promoter activity, and the inhibition was completely reversed by exogenous Zn2+ ions. In contrast, the calcium chelator 1,2-Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) (BAPTA-AM) potentiated gastrin-stimulated gastrin promoter activity. Treatment with gastrin increased the intracellular concentration of free Zn2+ ions, and the increase was blocked by TPEN, but not by BAPTA-AM. TPEN also inhibited the stimulation of cell proliferation and migration/invasion by gastrin, but BAPTA-AM had no effect. These results, which are the first report of the existence of Zn2+ signaling downstream of CCK2R activation, suggest that zinc chelation therapies may be effective in counteracting gastrin-dependent tumor growth.

  7. Netrin-1 induces the migration of Schwann cells via p38 MAPK and PI3K-Akt signaling pathway mediated by the UNC5B receptor

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    Lv, Jianwei [General Hospital of Tianjin Medical University, No. 154, Anshan Road, Heping District, Tianjin 300052 (China); Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China); Sun, Xiaolei; Ma, Jianxiong [Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China); Ma, Xinlong, E-mail: gengxiao502@163.com [General Hospital of Tianjin Medical University, No. 154, Anshan Road, Heping District, Tianjin 300052 (China); Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China); Zhang, Yang; Li, Fengbo; Li, Yanjun; Zhao, Zhihu [Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China)

    2015-08-14

    Schwann cells (SCs) play an essentially supportive role in the regeneration of injured peripheral nerve system (PNS). As Netrin-1 is crucial for the normal development of nervous system (NS) and can direct the process of damaged PNS regeneration, our study was designed to determine the role of Netrin-1 in RSC96 Schwann cells (an immortalized rat Schwann cell line) proliferation and migration. Our studies demonstrated that Netrin-1 had no effect on RSC96 cells proliferation, while significantly promoted RSC96 cells migration. The Netrin-1-induced RSC96 cells migration was significantly attenuated by inhibition of p38 and PI3K through pretreatment with SB203580 and LY294002 respectively, but not inhibition of MEK1/2 and JNK by U0126-EtOH and SP600125 individually. Treatment with Netrin-1 enhanced the phosphorylation of p38 and Akt. QRT-PCR indicated that Netrin-1 and only its receptors Unc5a, Unc5b and Neogenin were expressed in RSC96 cells, among which Unc5b expressed the most. And UNC5B protein was significantly increased after stimulated by Netrin-1. In conclusion, we show here that Netrin-1-enhanced SCs migration is mediated by activating p38 MAPK and PI3K-Akt signal cascades via receptor UNC5B, which suggests that Netrin-1 could serve as a new therapeutic strategy and has potential application value for PNS regeneration. - Highlights: • Netrin-1 attracts RSC96 Schwann cells migration in a dose dependent manner. • Netrin-1 induced Schwann cells migration is p38 and PI3K-Akt signaling dependent. • UNC5B may be dominant receptor mediating Netrin-1′ effect on RSC96 cells motility. • Netrin-1 may promote peripheral nerve repair by enhancing Schwann cells motility.

  8. Promotion of formyl peptide receptor 1-mediated neutrophil chemotactic migration by antimicrobial peptides isolated from the centipede Scolopendra subspinipes mutilans.

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    Park, Yoo Jung; Lee, Sung Kyun; Jung, Young Su; Lee, Mingyu; Lee, Ha Young; Kim, Sang Doo; Park, Joon Seong; Koo, JaeHyung; Hwang, Jae Sam; Bae, Yoe-Sik

    2016-09-01

    We investigated the effects of two antimicrobial peptides (AMPs) isolated from Scolopendra subspinipes mutilans on neutrophil activity. Stimulation of mouse neutrophils with the two AMPs elicited chemotactic migration of the cells in a pertussis toxin-sensitive manner. The two AMPs also stimulated activation of ERK and Akt, which contribute to chemotactic migration of neutrophils. We found that AMP-stimulated neutrophil chemotaxis was blocked by a formyl peptide receptor (FPR) 1 antagonist (cyclosporin H); moreover the two AMPs stimulated the chemotactic migration of FPR1-expressing RBL-2H3 cells but not of vector-expressing RBL-2H3 cells. We also found that the two AMPs stimulate neutrophil migration in vivo, and that this effect is blocked in FPR1-deficient mice. Taken together, our results suggest that the two AMPs stimulate neutrophils, leading to chemotactic migration through FPR1, and the two AMPs will be useful for the study of FPR1 signaling and neutrophil activation. [BMB Reports 2016; 49(9): 520-525].

  9. Measurement of chondrocyte chemotaxis using a Boyden chamber: a model of receptor-mediated cell migration combined with cell sedimentation.

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    Chen, Chih-Yuan; Hsiau, Kuan-Chih; Chung, C A

    2013-09-01

    The Boyden chamber assay measures the coefficients of cell motility by fitting the experiments with theoretical calculations. Under the circumstance of rapid receptor kinetics, the distribution of chemical-receptor complexes on the cell surface can be treated as being quasi-steady and chemotaxis is directly related to the biochemical concentration, leading to the celebrated Keller-Segel model, which has been shown to be an approximation to the full receptor-mediated form. No matter approximate or full, these approaches have ignored cell sedimentation in the upper chamber, assuming that all the cells have already resided on the filter top at the beginning of the test. However, the time required for all the cells to settle through the suspension can be close to the entire incubation time of just several hours. In order to amend such a deficiency, the present work combines the receptor-based model with cell sedimentation for modeling the chemotaxis assay using the Boyden chamber. Simulations were performed to fit the experimental data in the literature, which tested the chondrocyte chemotactic motility in response to collagen. Results show that once cell sedimentation is involved, the assumption of quasi-steady receptor distribution may be invalid for the Boyden assay. This is because the formation of the chemical-receptor complexes is profoundly retarded by the process of cell sedimentation. To estimate the parameters of cell motility and receptor kinetics, cell sedimentation should be incorporated in modeling the chemotaxis assay using the Boyden chamber.

  10. The cell surface glycoprotein CUB domain-containing protein 1 (CDCP1) contributes to epidermal growth factor receptor-mediated cell migration.

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    Dong, Ying; He, Yaowu; de Boer, Leonore; Stack, M Sharon; Lumley, John W; Clements, Judith A; Hooper, John D

    2012-03-23

    Epidermal growth factor (EGF) activation of the EGF receptor (EGFR) is an important mediator of cell migration, and aberrant signaling via this system promotes a number of malignancies including ovarian cancer. We have identified the cell surface glycoprotein CDCP1 as a key regulator of EGF/EGFR-induced cell migration. We show that signaling via EGF/EGFR induces migration of ovarian cancer Caov3 and OVCA420 cells with concomitant up-regulation of CDCP1 mRNA and protein. Consistent with a role in cell migration CDCP1 relocates from cell-cell junctions to punctate structures on filopodia after activation of EGFR. Significantly, disruption of CDCP1 either by silencing or the use of a function blocking antibody efficiently reduces EGF/EGFR-induced cell migration of Caov3 and OVCA420 cells. We also show that up-regulation of CDCP1 is inhibited by pharmacological agents blocking ERK but not Src signaling, indicating that the RAS/RAF/MEK/ERK pathway is required downstream of EGF/EGFR to induce increased expression of CDCP1. Our immunohistochemical analysis of benign, primary, and metastatic serous epithelial ovarian tumors demonstrates that CDCP1 is expressed during progression of this cancer. These data highlight a novel role for CDCP1 in EGF/EGFR-induced cell migration and indicate that targeting of CDCP1 may be a rational approach to inhibit progression of cancers driven by EGFR signaling including those resistant to anti-EGFR drugs because of activating mutations in the RAS/RAF/MEK/ERK pathway.

  11. Roles of taurine-mediated tonic GABAA receptor activation in the radial migration of neurons in the fetal mouse cerebral cortex

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    Tomonori eFurukawa

    2014-03-01

    Full Text Available γ-Aminobutyric acid (GABA depolarizes embryonic cerebrocortical neurons and continuous activation of the GABAA receptor (GABAAR contributes to their tonic depolarization. Although multiple reports have demonstrated a role of GABAAR activation in neocortical development, including in migration, most of these studies have used pharmacological blockers. Herein, we performed in utero electroporation in GABA synthesis-lacking homozygous GAD67-GFP knock-in mice (GAD67GFP/GFP to label neurons born in the ventricular zone. Three days after electroporation, there were no differences in the distribution of labeled cells between the genotypes. The dose-response properties of cells labeled to detect GABA were equivalent among genotypes. However, continuous blockade of GABAAR with the GABAAR antagonist SR95531 accelerated radial migration. This effect of GABAAR blockade in GAD67GFP/GFP mice suggested a role for alternative endogenous GABAAR agonists. Thus, we tested the role of taurine, which is derived from maternal blood but is abundant in the fetal brain. The taurine-evoked currents in labeled cells were mediated by GABAAR. Taurine uptake was blocked by a taurine transporter inhibitor, 2-(guanidinoethanesulfonic acid (GES, and taurine release was blocked by a volume-sensitive anion channel blocker, 4-(2-butyl-6,7-dichlor-2-cyclopentylindan-1-on-5-yl oxobutyric acid (DCPIB, as examined through high-performance liquid chromatography (HPLC. GES increased the extracellular taurine concentration and induced an inward shift of the holding current, which was reversed by SR95531. In a taurine-deficient mouse model, the GABAAR-mediated tonic currents were greatly reduced, and radial migration was accelerated. As the tonic currents were equivalent among the genotypes of GAD67-GFP knock-in mice, taurine, rather than GABA, might play a major role as an endogenous agonist of embryonic tonic GABAAR conductance, regulating the radial migration of neurons in the

  12. Role of laminin receptor in tumor cell migration

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    Wewer, U M; Taraboletti, G; Sobel, M E

    1987-01-01

    ). These antisera were used to study the potential role of laminin receptor in laminin-mediated attachment and haptotactic migration of human A2058 melanoma cells. The anti-laminin receptor antisera reacted with the surface of suspended, nonpermeabilized melanoma and carcinoma cells. The anti-laminin receptor...... antisera blocked the surface interaction of A2058 cells with endogenous laminin, resulting in the inhibition of laminin-mediated cell attachment. The A2058 melanoma cells migrated toward a gradient of solid phase laminin or fibronectin (haptotaxis). Anti-laminin antiserum abolished haptotaxis on laminin...... but not on fibronectin. Synthetic peptide GRGDS corresponding to the fibronectin cell-binding domain inhibited haptotaxis on fibronectin but not on laminin. Both types of anti-laminin receptor antisera inhibited haptotaxis on laminin but not on fibronectin. Using immunohistochemistry, invading human carcinoma cells...

  13. The HLA-DR mediated signalling increases the migration and invasion of melanoma cells, the expression and lipid raft recruitment of adhesion receptors, PD-L1 and signal transduction proteins.

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    Costantini, Francesca; Barbieri, Giovanna

    2017-08-01

    The constitutive expression of Major Histocompatibility Complex (MHC) class II molecules is restricted to professional Antigen-Presenting Cells (APCs), nevertheless almost 50% of melanomas express constitutively the MHC class II molecules. Therefore, in two MHC class II constitutive expressing melanoma cell lines we studied the signalling mediated by the HLA-DR molecules in the aim to understand the consequence of class II mediated signalling on metastatic dissemination of melanoma. In particular, we reported that the HLA-DR mediated signalling play a new role in melanoma progression, increasing the migration and invasion of melanoma cells. Furthermore, we showed that the HLA-DR mediated signalling increases the expression and the lipid raft localisation of class II molecules, PD-L1 receptor, Integrin and CAM adhesion receptors, FAK, AKT and STAT3 signalling proteins. We also showed that the HLA-DR mediated signalling increases the activation of FAK, AKT, ERK, PKC and STAT3 signalling proteins and the expression of ILK, PAX, BRAF, ERK and PKC. Indeed, the results showed suggest that the HLA-DR mediated signalling provides a platform useful to frustrate an effective anti-tumour response and to increase melanoma migration and metastatic dissemination of this cancer. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Role of formic receptors in soluble urokinase receptor-induced human vascular smooth muscle migration.

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    Duru, Enrico A; Fu, Yuyang; Davies, Mark G

    2015-05-15

    Vascular smooth muscle cell (VSMC) migration in response to urokinase is dependent on binding of the urokinase molecule to the urokinase plasminogen receptor (uPAR) and cleavage of the receptor. The aim of this study was to examine the role of the soluble uPAR (suPAR) in VSMC migration. Human VSMCs were cultured in vitro. Linear wound and Boyden microchemotaxis assays of migration were performed in the presence of suPAR. Inhibitors to G-protein signaling and kinase activation were used to study these pathways. Assays were performed for mitogen-activated protein kinase and epidermal growth factor receptor activation. suPAR induced concentration-dependent migration of VSMC, which was G protein-dependent and was blocked by Gαi and Gβγ inhibitors. Removal of the full uPAR molecule by incubation of the cells with a phospholipase did not interfere with this response. suPAR induced ERK1/2, p38(MAPK), and c-Jun N-terminal kinase [JNK] activation in a Gαi/Gβγ-dependent manner, and interruption of these signaling pathways prevented suPAR-mediated migration. suPAR activity was independent of plasmin activity. suPAR did not activate epidermal growth factor receptor. Interruption of the low affinity N-formyl-Met-Leu-Phe receptor (FPRL1) but not high affinity N-formyl-Met-Leu-Phe receptor (FPR) prevented cell migration and activation in response to suPAR. suPAR increased matrix metalloproteinase-2 expression and activity, and this was dependent on the low affinity N-formyl-Met-Leu-Phe receptor (FPRL1) and ERK1/2. suPAR induces human smooth muscle cell activation and migration independent of the full uPAR through activation of the G protein-coupled receptor FPRL1, which is not linked to the plasminogen activation cascade. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Pancreatic secretory trypsin inhibitor causes autocrine-mediated migration and invasion in bladder cancer and phosphorylates the EGF receptor, Akt2 and Akt3, and ERK1 and ERK2.

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    Marchbank, Tania; Mahmood, Asif; Playford, Raymond J

    2013-08-01

    Pancreatic secretory trypsin inhibitor (PSTI) is expressed in most bladder carcinomas, where its pathophysiological relevance is unclear. Using recombinant normal sequence PSTI/tumor-associated trypsin inhibitor (TATI), a variant associated with familial pancreatitis (N34S), an active site-inactivated variant (R18/V19), and immunoneutralization and RNA interference-mediated knockdown techniques, we investigated the actions of PSTI/TATI on cell migration (wounding monolayers), collagen invasion (gel invasion assays), and proliferation (Alamar blue) on 253J, RT4, and HT1376 human bladder carcinoma cell lines. All three forms of PSTI/TATI stimulated migration twofold, and normal sequence PSTI/TATI showed synergistic promigratory effects when added with EGF. Addition of structurally unrelated soybean trypsin inhibitor had no promigratory activity. Similar results were seen using collagen invasion assays, although the active site mutated variant had no proinvasive activity, probably due to reduced Akt2 activation. PSTI/TATI did not stimulate proliferation despite acting, at least partially, through the EGF receptor, as effects of PSTI/TATI were truncated by the addition of an EGF receptor blocking antibody or the tyrosine kinase inhibitor tyrphostin. Cell lines produced endogenous PSTI/TATI, and PSTI/TATI RNA interference knockdown or the addition of PSTI/TATI, EGF receptor, or tyrphostin blocking agents reduced migration and invasion below baseline. PSTI/TATI induced phosphorylation of the EGF receptor, ERK1 and ERK2, Akt2 and Akt3, JNK1, MKK3, and ribosomal protein S6 kinase 1. This profile was more limited than that induced by EGF and did not include Akt1, probably explaining the lack of proproliferative activity. Our findings of autocrine stimulation and synergistic responses between EGF and PSTI/TATI at concentrations found in urine and tissue suggest that PSTI/TATI has pathophysiological relevance.

  16. PDGF mediates TGFβ-induced migration during development of the spinous process.

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    Wang, Ying; Serra, Rosa

    2012-05-01

    Mechanisms mediating closure of the dorsal vertebrae are not clear. Previously, we showed that deletion of TGFβ type II receptor (Tgfbr2) in sclerotome in mice results in failure in the formation of the spinous process, mimicking spina bifida occulta, a common malformation in humans. In this study, we aimed to determine whether missing dorsal structures in Tgfbr2 mutant mice were due to defects in mesenchymal migration and to clarify mechanism of TGFβ-mediated migration. First, we showed that gross alterations in dorsal vertebrae were apparent by E16.5days in Tgfbr2 mutants. In addition, histological staining showed that the mesenchyme adjacent to the developing cartilage was thin compared to controls likely due to reduced proliferation and migration of these cells. Next, we used a chemotaxis migration assay to show that TGFβ promotes migration in mixed cultures of embryonic sclerotome and associated mesenchyme. TGFβ stimulated expression of PDGF ligands and receptors in the cultures and intact PDGF signaling was required for TGFβ-mediated migration. Since PDGF ligands are expressed in the sclerotome-derived cartilage where Tgfbr2 is deleted and the receptors are predominantly expressed in the adjacent mesenchyme, we propose that TGFβ acts on the sclerotome to regulate expression of PDGF ligands, which then act on the associated mesenchyme in a paracrine fashion to mediate proliferation, migration and subsequent differentiation of the adjacent sclerotome. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Integrin-mediated cell migration is blocked by inhibitors of human neuraminidase.

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    Jia, Feng; Howlader, Md Amran; Cairo, Christopher W

    2016-09-01

    Integrins are critical receptors in cell migration and adhesion. A number of mechanisms are known to regulate the function of integrins, including phosphorylation, conformational change, and cytoskeletal anchoring. We investigated whether native neuraminidase (Neu, or sialidase) enzymes which modify glycolipids could play a role in regulating integrin-mediated cell migration. Using a scratch assay, we found that exogenously added Neu3 and Neu4 activity altered rates of cell migration. We observed that Neu4 increased the rate of migration in two cell lines (HeLa, A549); while Neu3 only increased migration in HeLa cells. A bacterial neuraminidase was able to increase the rate of migration in HeLa, but not in A549 cells. Treatment of cells with complex gangliosides (GM1, GD1a, GD1b, and GT1b) resulted in decreased cell migration rates, while LacCer was able to increase rates of migration in both lines. Importantly, our results show that treatment of cells with inhibitors of native Neu enzymes had a dramatic effect on the rates of cell migration. The most potent compound tested targeted the human Neu4 isoenzyme, and was able to substantially reduce the rate of cell migration. We found that the lateral mobility of integrins was reduced by treatment of cells with Neu3, suggesting that Neu3 enzyme activity resulted in changes to integrin-co-receptor or integrin-cytoskeleton interactions. Finally, our results support the hypothesis that inhibitors of human Neu can be used to investigate mechanisms of cell migration and for the development of anti-adhesive therapies. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Melanocortin 1 Receptor Regulates Melanoma Cell Migration by Controlling Syndecan-2 Expression*

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    Chung, Heesung; Lee, Jung-hyun; Jeong, Dayun; Han, Inn-Oc; Oh, Eok-Soo

    2012-01-01

    The melanocortin 1 receptor (MC1R), a key regulator of melanogenesis, is known to control inflammation, acting in concert with the MC1R ligand α-melanocyte-stimulating hormone. Although cell migration is a key event in inflammation, few studies have addressed the function of MC1R in this context. Using highly motile melanoma cells, we found that the expression level of MC1R was associated with the extent of migration of mouse melanoma cells, suggesting that MC1R plays a functional role in controlling this migration. Overexpression of MC1R enhanced melanoma cell migration, whereas the opposite was true when MC1R levels were knocked down using small inhibitory RNAs. Interestingly, MC1R expression enhanced the synthesis of syndecan-2, a cell surface heparan sulfate proteoglycan known to be involved in melanoma cell migration. Knockdown of syndecan-2 expression decreased MC1R-mediated cell migration. Further, MC1R inhibited the activation of p38 MAPK, subsequently enhancing expression of sydnecan-2, in parallel with an increase in the extent of cell migration. Consistently, activation of p38 by H2O2 inhibited syndecan-2 expression and cell migration, whereas inhibition of p38 activation enhanced syndecan-2 expression and cell migration. Finally, we found that α-melanocyte-stimulating hormone inhibited MC1R-mediated cell migration via activation of p38 and inhibition of syndecan-2 expression. Together, the data strongly suggest that MC1R regulates melanoma cell migration via inhibition of syndecan-2 expression. PMID:22493442

  19. Glycine Receptor α2 Subunit Activation Promotes Cortical Interneuron Migration

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    Ariel Avila

    2013-08-01

    Full Text Available Glycine receptors (GlyRs are detected in the developing CNS before synaptogenesis, but their function remains elusive. This study demonstrates that functional GlyRs are expressed by embryonic cortical interneurons in vivo. Furthermore, genetic disruption of these receptors leads to interneuron migration defects. We discovered that extrasynaptic activation of GlyRs containing the α2 subunit in cortical interneurons by endogenous glycine activates voltage-gated calcium channels and promotes calcium influx, which further modulates actomyosin contractility to fine-tune nuclear translocation during migration. Taken together, our data highlight the molecular events triggered by GlyR α2 activation that control cortical tangential migration during embryogenesis.

  20. Androgen-induced cell migration: role of androgen receptor/filamin A association.

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    Gabriella Castoria

    Full Text Available BACKGROUND: Androgen receptor (AR controls male morphogenesis, gametogenesis and prostate growth as well as development of prostate cancer. These findings support a role for AR in cell migration and invasiveness. However, the molecular mechanism involved in AR-mediated cell migration still remains elusive. METHODOLOGY/PRINCIPAL FINDINGS: Mouse embryo NIH3T3 fibroblasts and highly metastatic human fibrosarcoma HT1080 cells harbor low levels of transcriptionally incompetent AR. We now report that, through extra nuclear action, AR triggers migration of both cell types upon stimulation with physiological concentrations of the androgen R1881. We analyzed the initial events leading to androgen-induced cell migration and observed that challenging NIH3T3 cells with 10 nM R1881 rapidly induces interaction of AR with filamin A (FlnA at cytoskeleton. AR/FlnA complex recruits integrin beta 1, thus activating its dependent cascade. Silencing of AR, FlnA and integrin beta 1 shows that this ternary complex controls focal adhesion kinase (FAK, paxillin and Rac, thereby driving cell migration. FAK-null fibroblasts migrate poorly and Rac inhibition by EHT impairs motility of androgen-treated NIH3T3 cells. Interestingly, FAK and Rac activation by androgens are independent of each other. Findings in human fibrosarcoma HT1080 cells strengthen the role of Rac in androgen signaling. The Rac inhibitor significantly impairs androgen-induced migration in these cells. A mutant AR, deleted of the sequence interacting with FlnA, fails to mediate FAK activation and paxillin tyrosine phosphorylation in androgen-stimulated cells, further reinforcing the role of AR/FlnA interaction in androgen-mediated motility. CONCLUSIONS/SIGNIFICANCE: The present report, for the first time, indicates that the extra nuclear AR/FlnA/integrin beta 1 complex is the key by which androgen activates signaling leading to cell migration. Assembly of this ternary complex may control organ development

  1. Role of ErbB receptors in cancer cell migration and invasion

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    Aline eAppert-Collin

    2015-11-01

    Full Text Available Growth factors mediate their diverse biologic responses (regulation of cellular proliferation, differentiation, migration and survival by binding to and activating cell-surface receptors with intrinsic protein kinase activity named Receptor Tyrosine Kinases (RTKs. About 60 RTKs have been identified and can be classified into more than 16 different receptor families. Their activity is normally tightly controlled and regulated. Overexpression of RTK proteins or functional alterations caused by mutations in the corresponding genes or abnormal stimulation by autocrine growth factor loops contribute to constitutive RTK signaling, resulting in alterations in the physiological activities of cells. The ErbB receptor family of RTKs comprises four distinct receptors: the EGFR (also known as ErbB1/HER1, ErbB2 (neu, HER2, ErbB3 (HER3 and ErbB4 (HER4. ErbB family members are often overexpressed, amplified, or mutated in many forms of cancer, making them important therapeutic targets. EGFR has been found to be amplified in gliomas and non-small-cell lung carcinoma while ErbB2 amplifications are seen in breast, ovarian, bladder, non-small-cell lung carcinoma, as well as several other tumor types. Several data have shown that ErbB receptor family and its downstream pathway regulate epithelial-mesenchymal transition, migration, and tumor invasion by modulating extracellular matrix components. Recent findings indicate that extracellular matrix components such as matrikines bind specifically to EGF receptor and promote cell invasion. In this review, we will present an in-depth overview of the structure, mechanisms, cell signaling, and functions of ErbB family receptors in cell adhesion and migration. Furthermore, we will describe in a last part the new strategies developed in anti-cancer therapy to inhibit ErbB family receptor activation.

  2. Serotonin receptor 3A controls interneuron migration into the neocortex.

    Science.gov (United States)

    Murthy, Sahana; Niquille, Mathieu; Hurni, Nicolas; Limoni, Greta; Frazer, Sarah; Chameau, Pascal; van Hooft, Johannes A; Vitalis, Tania; Dayer, Alexandre

    2014-11-20

    Neuronal excitability has been shown to control the migration and cortical integration of reelin-expressing cortical interneurons (INs) arising from the caudal ganglionic eminence (CGE), supporting the possibility that neurotransmitters could regulate this process. Here we show that the ionotropic serotonin receptor 3A (5-HT(3A)R) is specifically expressed in CGE-derived migrating interneurons and upregulated while they invade the developing cortex. Functional investigations using calcium imaging, electrophysiological recordings and migration assays indicate that CGE-derived INs increase their response to 5-HT(3A)R activation during the late phase of cortical plate invasion. Using genetic loss-of-function approaches and in vivo grafts, we further demonstrate that the 5-HT(3A)R is cell autonomously required for the migration and proper positioning of reelin-expressing CGE-derived INs in the neocortex. Our findings reveal a requirement for a serotonin receptor in controlling the migration and laminar positioning of a specific subtype of cortical IN.

  3. Particle aggregation during receptor-mediated endocytosis

    Science.gov (United States)

    Mao, Sheng; Kosmrlj, Andrej

    Receptor-mediated endocytosis of particles is driven by large binding energy between ligands on particles and receptors on a membrane, which compensates for the membrane bending energy and for the cost due to the mixing entropy of receptors. While the receptor-mediated endocytosis of individual particle is well understood, much less is known about the joint entry of multiple particles. Here, we demonstrate that the endocytosis of multiple particles leads to a kinetically driven entropic attraction, which may cause the aggregation of particles observed in experiments. During the endocytosis particles absorb nearby receptors and thus produce regions, which are depleted of receptors. When such depleted regions start overlapping, the corresponding particles experience osmotic-like attractive entropic force. If the attractive force between particles is large enough to overcome the repulsive interaction due to membrane bending, then particles tend to aggregate provided that they are sufficiently close, such that they are not completely engulfed before they come in contact. We discuss the necessary conditions for the aggregation of cylindrical particles during receptor-mediated endocytosis and comment on the generalization to spherical particles.

  4. Involvement of LPA Receptor 3 in LPA-induced BGC- 803 Cell Migration

    Directory of Open Access Journals (Sweden)

    Erdene Oyungerel

    2013-12-01

    Full Text Available Lysophosphatidic acid ˄ LPA ˅ is a bioactive phospholipid mediator, which elicits a variety of biological functions mainly through G-protein coupled receptors. Although LPA is shown to stimulate proliferation and motility via LPA receptors, LPAR1 and LPAR3 in several cancer cell lines, but the role of LPA receptors in gastric cancer cells is still being unknown. However, several researches reported that LPAR2 play an important role in the carcinogenesis of gastric cancer, but there is no report to show the LPAR3 involvement in the carcinogenesis. For this reason, we examined LPA receptors (LPAR1, LPAR2 and LPAR3 in BGC-803 cells along with real time PCR method. Real-time PCR analyses were used to evaluate the expression of LPA receptors in BGC-803 cells. Among these receptors, LPAR3 was shown to be highly expressed in BGC-803 cells, a human gastric cancer cell line. Transient transfection with LPAR3 siRNA was observed to reduce LPAR3 mRNA in BGC-803 cells and eliminate the LPA-induced cell migration. The results suggest that the LPAR3 regulates LPA-induced BGC-803 cell migration.

  5. Adaptor protein2 (AP2) orchestrates CXCR2-mediated cell migration.

    Science.gov (United States)

    Raman, Dayanidhi; Sai, Jiqing; Hawkins, Oriana; Richmond, Ann

    2014-04-01

    The chemokine receptor CXCR2 is vital for inflammation, wound healing, angiogenesis, cancer progression and metastasis. Adaptor protein 2 (AP2), a clathrin binding heterotetrameric protein comprised of α, β2, μ2 and σ2 subunits, facilitates clathrin-mediated endocytosis. Mutation of the LLKIL motif in the CXCR2 carboxyl-terminal domain (CTD) results in loss of AP2 binding to the receptor and loss of ligand-mediated receptor internalization and chemotaxis. AP2 knockdown also results in diminished ligand-mediated CXCR2 internalization, polarization and chemotaxis. Using knockdown/rescue approaches with AP2-μ2 mutants, the binding domains were characterized in reference to CXCR2 internalization and chemotaxis. When in an open conformation, μ2 Patch 1 and Patch 2 domains bind tightly to membrane PIP2 phospholipids. When AP2-μ2, is replaced with μ2 mutated in Patch 1 and/or Patch 2 domains, ligand-mediated receptor binding and internalization are not lost. However, chemotaxis requires AP2-μ2 Patch 1, but not Patch 2. AP2-σ2 has been demonstrated to bind dileucine motifs to facilitate internalization. Expression of AP2-σ2 V88D and V98S dominant negative mutants resulted in loss of CXCR2 mediated chemotaxis. Thus, AP2 binding to both membrane phosphatidylinositol phospholipids and dileucine motifs is crucial for directional migration or chemotaxis. Moreover, AP2-mediated receptor internalization can be dissociated from AP2-mediated chemotaxis. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. Integrin-mediated cell surface recruitment of autotaxin promotes persistent directional cell migration

    Science.gov (United States)

    Wu, Tao; Kooi, Craig Vander; Shah, Pritom; Charnigo, Richard; Huang, Cai; Smyth, Susan S.; Morris, Andrew J.

    2014-01-01

    Autotaxin (ATX) is a secreted lysophospholipase D (lysoPLD) that binds to integrin adhesion receptors. We dissected the roles of integrin binding and lysoPLD activity in stimulation of human breast cancer and mouse aortic vascular smooth muscle cell migration by ATX. We compared effects of wild-type human ATX, catalytically inactive ATX, an integrin binding-defective ATX variant with wild-type lysoPLD activity, the isolated ATX integrin binding N-terminal domain, and a potent ATX selective lysoPLD inhibitor on cell migration using transwell and single-cell tracking assays. Stimulation of transwell migration was reduced (18 or 27% of control, respectively) but not ablated by inactivation of integrin binding or inhibition of lysoPLD activity. The N-terminal domain increased transwell migration (30% of control). ATX lysoPLD activity and integrin binding were necessary for a 3.8-fold increase in the fraction of migrating breast cancer cell step velocities >0.7 μm/min. ATX increased the persistent directionality of single-cell migration 2-fold. This effect was lysoPLD activity independent and recapitulated by the integrin binding N-terminal domain. Integrin binding enables uptake and intracellular sequestration of ATX, which redistributes to the front of migrating cells. ATX binding to integrins and lysoPLD activity therefore cooperate to promote rapid persistent directional cell migration.—Wu, T., Kooi, C. V., Shah, P., Charnigo, R., Huang, C., Smyth, S. S., Morris, A. J. Integrin-mediated cell surface recruitment of autotaxin promotes persistent directional cell migration. PMID:24277575

  7. Quantification of receptor-mediated endocytosis.

    Science.gov (United States)

    Wettey, Frank R; Jackson, Antony P

    2006-01-01

    This method allows measuring of the receptor-mediated internalization of 125I-labeled conalbumin, the chicken egg white isoform of transferrin. Kinetic data, i.e. the rate constant k(i) for the initial internalization process, can be extracted from the data by linear curve fitting using an In/Sur plot (intracellular label/cell surface label over time).

  8. Receptor-mediated signalling at plasmodesmata

    Directory of Open Access Journals (Sweden)

    Christine eFaulkner

    2013-12-01

    Full Text Available Plasmodesmata (PD generate continuity between plant cells via the cytoplasm, endoplasmic reticulum (ER and plasma membrane (PM, allowing movement of different classes of molecules between cells. Proteomic data indicates that the PD PM hosts many receptors and receptor kinases, as well as lipid raft and tetraspanin enriched microdomain associated proteins, suggesting the hypothesis that the PD PM is specialised with respect to both composition and function. PD-located receptor proteins and receptor kinases are responsible for perception of microbe associated molecular patterns at PD and initiate signalling that mediates changes to PD flux. In addition, developmentally relevant receptor kinases have different interactions dependent upon whether located at the PD PM or the cellular PM. The implications of these findings are that receptor-mediated signalling in PD membranes differs from that in the cellular PM and, in light the identification of PD-located proteins associated with membrane microdomains and the role of membrane microdomains in analogous signalling processes in animals, suggests that the PD PM contains specialised signalling platforms.

  9. Receptor protein tyrosine phosphatase alpha is essential for hippocampal neuronal migration and long-term potentiation

    DEFF Research Database (Denmark)

    Petrone, Angiola; Battaglia, Fortunato; Wang, Cheng

    2003-01-01

    Despite clear indications of their importance in lower organisms, the contributions of protein tyrosine phosphatases (PTPs) to development or function of the mammalian nervous system have been poorly explored. In vitro studies have indicated that receptor protein tyrosine phosphatase alpha (RPTPa....... However, these synapses are unable to undergo long-term potentiation. Mice lacking RPTPalpha also underperform in the radial-arm water-maze test. These studies identify RPTPalpha as a key mediator of neuronal migration and synaptic plasticity....... neuronal migration. The migratory abnormality likely results from a radial glial dysfunction rather than from a neuron-autonomous defect. In spite of this aberrant development, basic synaptic transmission from the Schaffer collateral pathway to CA1 pyramidal neurons remains intact in Ptpra(-/-) mice...

  10. c-Cbl regulates αPix-mediated cell migration and invasion

    Energy Technology Data Exchange (ETDEWEB)

    Seong, Min Woo; Park, Ji Ho; Yoo, Hee Min; Yang, Seung Wook; Oh, Kyu Hee; Ka, Seung Hyeun; Park, Dong Eun [School of Biological Sciences and Institute for Protein Metabolism, Seoul National University, Seoul 151-742 (Korea, Republic of); Lee, Soon-Tae [Department of Neurology, Seoul National University Hospital, Seoul 110-744 (Korea, Republic of); Chung, Chin Ha, E-mail: chchung@snu.ac.kr [School of Biological Sciences and Institute for Protein Metabolism, Seoul National University, Seoul 151-742 (Korea, Republic of)

    2014-12-12

    Highlights: • c-Cbl ubiquitinates αPix for proteasome-mediated degradation. • C6 and A172 glioma cells lack c-Cbl, which leads to stabilization of αPix. • The accumulated αPix promotes migration and invasion of the cancer cells. • The lack of c-Cbl in the cells appears responsible for their malignant behavior. - Abstract: c-Cbl, a RING-type ubiquitin E3 ligase, down-regulates receptor tyrosine kinases, including EGF receptor, and inhibits cell proliferation. Moreover, c-Cbl mutations are frequently found in patients with myeloid neoplasm. Therefore, c-Cbl is known as a tumor suppressor. αPix is expressed only in highly proliferative and mobile cells, including immune cells, and up-regulated in certain invasive tumors, such as glioblastoma multiforme. Here, we showed that c-Cbl serves as an ubiquitin E3 ligase for proteasome-mediated degradation of αPix, but not βPix. Remarkably, the rat C6 and human A172 glioma cells were unable to express c-Cbl, which leads to a dramatic accumulation of αPix. Depletion of αPix by shRNA markedly reduced the ability of the glioma cells to migrate and invade, whereas complementation of shRNA-insensitive αPix promoted it. These results indicate that c-Cbl negatively regulates αPix-mediated cell migration and invasion and the lack of c-Cbl in the C6 and A172 glioma cells is responsible for their malignant behavior.

  11. ADAMTS1-mediated targeting of TSP-1 by PPARδ suppresses migration and invasion of breast cancer cells.

    Science.gov (United States)

    Ham, Sun Ah; Yoo, Taesik; Lee, Won Jin; Hwang, Jung Seok; Hur, Jinwoo; Paek, Kyung Shin; Lim, Dae-Seog; Han, Sung Gu; Lee, Chi-Ho; Seo, Han Geuk

    2017-11-07

    Migration and invasion of cancer cells into surrounding tissue is a key stage of cancer metastasis. Here, we show that peroxisome proliferator-activated receptor (PPAR) δ regulates migration and invasion of human breast cancer cells via thrombospondin-1 (TSP-1) and its degrading protease, a disintegrin and metalloprotease domains with thrombospondin motifs 1 (ADAMTS1). Activation of PPARδ by GW501516, a specific ligand for PPARδ, led to marked inhibition in the cell migration and TSP-1 expression of breast cancer. These effects were suppressed by small interfering RNA-mediated knock-down of ADAMTS1, indicating that ADAMTS1 is involved in PPARδ-mediated inhibition of migration and TSP-1 expression in breast cancer cells. In addition, ligand-activated PPARδ upregulated expression of ADAMTS1 at the transcriptional level via binding of PPARδ to a direct repeat-1 site within the ADAMTS1 gene promoter. Furthermore, ligand-activated PPARδ suppressed invasion of breast cancer cells in an ADAMTS1-dependent manner. Taken together, these results demonstrate that PPARδ suppresses migration and invasion of breast cancer cells by downregulating TSP-1 in a process mediated by upregulation of ADAMTS1.

  12. Toll-like receptor 9 mediated responses in cardiac fibroblasts.

    Directory of Open Access Journals (Sweden)

    Ingrid Kristine Ohm

    Full Text Available Altered cardiac Toll-like receptor 9 (TLR9 signaling is important in several experimental cardiovascular disorders. These studies have predominantly focused on cardiac myocytes or the heart as a whole. Cardiac fibroblasts have recently been attributed increasing significance in mediating inflammatory signaling. However, putative TLR9-signaling through cardiac fibroblasts remains non-investigated. Thus, our aim was to explore TLR9-signaling in cardiac fibroblasts and investigate the consequence of such receptor activity on classical cardiac fibroblast cellular functions. Cultivated murine cardiac fibroblasts were stimulated with different TLR9 agonists (CpG A, B and C and assayed for the secretion of inflammatory cytokines (tumor necrosis factor α [TNFα], CXCL2 and interferon α/β. Expression of functional cardiac fibroblast TLR9 was proven as stimulation with CpG B and -C caused significant CXCL2 and TNFα-release. These responses were TLR9-specific as complete inhibition of receptor-stimulated responses was achieved by co-treatment with a TLR9-antagonist (ODN 2088 or chloroquine diphosphate. TLR9-stimulated responses were also found more potent in cardiac fibroblasts when compared with classical innate immune cells. Stimulation of cardiac fibroblasts TLR9 was also found to attenuate migration and proliferation, but did not influence myofibroblast differentiation in vitro. Finally, results from in vivo TLR9-stimulation with subsequent fractionation of specific cardiac cell-types (cardiac myocytes, CD45+ cells, CD31+ cells and cardiac fibroblast-enriched cell-fractions corroborated our in vitro data and provided evidence of differentiated cell-specific cardiac responses. Thus, we conclude that cardiac fibroblast may constitute a significant TLR9 responder cell within the myocardium and, further, that such receptor activity may impact important cardiac fibroblast cellular functions.

  13. Discoidin Domain Receptor 1 Mediates Myosin-Dependent Collagen Contraction

    Directory of Open Access Journals (Sweden)

    Nuno M. Coelho

    2017-02-01

    Full Text Available Discoidin domain receptor 1 (DDR1 is a tyrosine kinase collagen adhesion receptor that mediates cell migration through association with non-muscle myosin IIA (NMIIA. Because DDR1 is implicated in cancer fibrosis, we hypothesized that DDR1 interacts with NMIIA to enable collagen compaction by traction forces. Mechanical splinting of rat dermal wounds increased DDR1 expression and collagen alignment. In periodontal ligament of DDR1 knockout mice, collagen mechanical reorganization was reduced >30%. Similarly, cultured cells with DDR1 knockdown or expressing kinase-deficient DDR1d showed 50% reduction of aligned collagen. Tractional remodeling of collagen was dependent on DDR1 clustering, activation, and interaction of the DDR1 C-terminal kinase domain with NMIIA filaments. Collagen remodeling by traction forces, DDR1 tyrosine phosphorylation, and myosin light chain phosphorylation were increased on stiff versus soft substrates. Thus, DDR1 clustering, activation, and interaction with NMIIA filaments enhance the collagen tractional remodeling that is important for collagen compaction in fibrosis.

  14. Single amino acid substitutions in the chemotactic sequence of urokinase receptor modulate cell migration and invasion.

    Directory of Open Access Journals (Sweden)

    Katia Bifulco

    Full Text Available The receptor for urokinase-type plasminogen activator (uPAR plays an important role in controlling cell migration. uPAR binds urokinase and vitronectin extracellular ligands, and signals in complex with transmembrane receptors such as Formyl-peptide Receptors (FPRs and integrins. Previous work from this laboratory has shown that synthetic peptides, corresponding to the uPAR(88-92 chemotactic sequence, when carrying the S90P or S90E substitutions, up- or down-regulate cell migration, respectively. To gain mechanistic insights into these opposite cell responses, the functional consequences of S90P and S90E mutations in full-length uPAR were evaluated. First, (HEK-293 embryonic kidney cells expressing uPAR(S90P exhibit enhanced FPR activation, increased random and directional cell migration, long-lasting Akt phosphorylation, and increased adhesion to vitronectin, as well as uPAR/vitronectin receptor association. In contrast, the S90E substitution prevents agonist-triggered FPR activation and internalization, decreases binding and adhesion to vitronectin, and inhibits uPAR/vitronectin receptor association. Also, 293/uPAR(S90P cells appear quite elongated and their cytoskeleton well organized, whereas 293/uPAR(S90E cells assume a large flattened morphology, with random orientation of actin filaments. Interestingly, when HT1080 cells co-express wild type uPAR with uPAR S90E, the latter behaves as a dominant-negative, impairing uPAR-mediated signaling and reducing cell wound repair as well as lung metastasis in nude mice. In contrast, signaling, wound repair and in vivo lung metastasis of HT1080 cells bearing wild type uPAR are enhanced when they co-express uPAR(S90P. In conclusion, our findings indicate that Ser(90 is a critical residue for uPAR signaling and that the S90P and S90E exert opposite effects on uPAR activities. These findings may be accommodated in a molecular model, in which uPAR(S90E and uPAR(S90P are forced into inactive and active

  15. Transcriptional profiling of human monocytes identifies the inhibitory receptor CD300a as regulator of transendothelial migration.

    Directory of Open Access Journals (Sweden)

    Sharang Ghavampour

    Full Text Available Local inflammatory responses are characterized by the recruitment of circulating leukocytes from the blood to sites of inflammation, a process requiring the directed migration of leukocytes across the vessel wall and hence a penetration of the endothelial lining. To identify underlying signalling events and novel factors involved in these processes we screened for genes differentially expressed in human monocytes following their adhesion to and passage through an endothelial monolayer. Functional annotation clustering of the genes identified revealed an overrepresentation of those associated with inflammation/immune response, in particular early monocyte to macrophage differentiation. Among the gene products so far not implicated in monocyte transendothelial migration was the inhibitory immune receptor CD300a. CD300a mRNA and protein levels were upregulated following transmigration and engagement of the receptor by anti-CD300a antibodies markedly reduced monocyte transendothelial migration. In contrast, siRNA mediated downregulation of CD300a in human monocytes increased their rate of migration. CD300a colocalized and cosedimented with actin filaments and, when activated, caused F-actin cytoskeleton alterations. Thus, monocyte transendothelial migration is accompanied by an elevation of CD300a which serves an inhibitory function possibly required for termination of the actual transmigration.

  16. Neutrophil transepithelial migration: role of toll-like receptors in mucosal inflammation

    Directory of Open Access Journals (Sweden)

    Titus A Reaves

    2005-03-01

    Full Text Available The symptomatic phases of many inflammatory diseases are characterized by migration of large numbers of neutrophils (PMN across a polarized epithelium and accumulation within a lumen. For example, acute PMN influx is common in diseases of the gastrointestinal system (ulcerative colitis, Crohn's disease, bacterial enterocolitis, gastritis, hepatobiliary system (cholangitis, acute cholecystitis, respiratory tract (bronchial pneumonia, bronchitis, cystic fibrosis, bronchiectasis, and urinary tract (pyelonephritis, cystitis. Despite these observations, the molecular basis of leukocyte interactions with epithelial cells is incompletely understood. In vitro models of PMN transepithelial migration typically use N-formylated bacterial peptides such as fMLP in isolation to drive human PMNs across epithelial monolayers. However, other microbial products such as lipopolysaccharide (LPS are major constituents of the intestinal lumen and have potent effects on the immune system. In the absence of LPS, we have shown that transepithelial migration requires sequential adhesive interactions between the PMN beta2 integrin CD11b/CD18 and JAM protein family members. Other epithelial ligands appear to be abundantly represented as fucosylated proteoglycans. Further studies indicate that the rate of PMN migration across mucosal surfaces can be regulated by the ubiquitously expressed transmembrane protein CD47 and microbial-derived factors, although many of the details remain unclear. Current data suggests that Toll-like receptors (TLR, which recognize specific pathogen-associated molecular patterns (PAMPs, are differentially expressed on both leukocytes and mucosal epithelial cells while serving to modulate leukocyte-epithelial interactions. Exposure of epithelial TLRs to microbial ligands has been shown to result in transcriptional upregulation of inflammatory mediators whereas ligation of leukocyte TLRs modulate specific antimicrobial responses. A better

  17. Tre1, a G protein-coupled receptor, directs transepithelial migration of Drosophila germ cells.

    Directory of Open Access Journals (Sweden)

    Prabhat S Kunwar

    2003-12-01

    Full Text Available In most organisms, germ cells are formed distant from the somatic part of the gonad and thus have to migrate along and through a variety of tissues to reach the gonad. Transepithelial migration through the posterior midgut (PMG is the first active step during Drosophila germ cell migration. Here we report the identification of a novel G protein-coupled receptor (GPCR, Tre1, that is essential for this migration step. Maternal tre1 RNA is localized to germ cells, and tre1 is required cell autonomously in germ cells. In tre1 mutant embryos, most germ cells do not exit the PMG. The few germ cells that do leave the midgut early migrate normally to the gonad, suggesting that this gene is specifically required for transepithelial migration and that mutant germ cells are still able to recognize other guidance cues. Additionally, inhibiting small Rho GTPases in germ cells affects transepithelial migration, suggesting that Tre1 signals through Rho1. We propose that Tre1 acts in a manner similar to chemokine receptors required during transepithelial migration of leukocytes, implying an evolutionarily conserved mechanism of transepithelial migration. Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells. Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

  18. Coagulation Factor Xa inhibits cancer cell migration via LIMK1-mediated cofilin inactivation

    NARCIS (Netherlands)

    Borensztajn, Keren; Peppelenbosch, Maikel P.; Spek, C. Arnold

    2010-01-01

    Previously, we showed that activated coagulation factor X (FXa) inhibits migration of breast, lung and colon cancer cells. We showed that the effect of FXa on migration was protease-activated receptor (PAR)-1-dependent, but the subsequent cellular signaling routes remained elusive. In the current

  19. Coagulation Factor Xa inhibits cancer cell migration via LIMK1-mediated cofilin inactivation

    NARCIS (Netherlands)

    Borensztajn, Keren; Peppelenbosch, Maikel P.; Spek, C. Arnold

    Previously, we showed that activated coagulation factor X (FXa) inhibits migration of breast, lung and colon cancer cells. We showed that the effect of FXa on migration was protease-activated receptor (PAR)-1-dependent, but the subsequent cellular signaling routes remained elusive. In the current

  20. Cooperation of tyrosine kinase receptor TrkB and epidermal growth factor receptor signaling enhances migration and dispersal of lung tumor cells.

    Directory of Open Access Journals (Sweden)

    Rudolf Götz

    Full Text Available TrkB mediates the effects of brain-derived neurotrophic factor (BDNF in neuronal and nonnneuronal cells. Based on recent reports that TrkB can also be transactivated through epidermal growth-factor receptor (EGFR signaling and thus regulates migration of early neurons, we investigated the role of TrkB in migration of lung tumor cells. Early metastasis remains a major challenge in the clinical management of non-small cell lung cancer (NSCLC. TrkB receptor signaling is associated with metastasis and poor patient prognosis in NSCLC. Expression of this receptor in A549 cells and in another adenocarcinoma cell line, NCI-H441, promoted enhanced migratory capacity in wound healing assays in the presence of the TrkB ligand BDNF. Furthermore, TrkB expression in A549 cells potentiated the stimulatory effect of EGF in wound healing and in Boyden chamber migration experiments. Consistent with a potential loss of cell polarity upon TrkB expression, cell dispersal and de-clustering was induced in A549 cells independently of exogeneous BDNF. Morphological transformation involved extensive cytoskeletal changes, reduced E-cadherin expression and suppression of E-cadherin expression on the cell surface in TrkB expressing tumor cells. This function depended on MEK and Akt kinase activity but was independent of Src. These data indicate that TrkB expression in lung adenoma cells is an early step in tumor cell dissemination, and thus could represent a target for therapy development.

  1. High density lipoprotein stimulated migration of macrophages depends on the scavenger receptor class B, type I, PDZK1 and Akt1 and is blocked by sphingosine 1 phosphate receptor antagonists.

    Directory of Open Access Journals (Sweden)

    Aishah Al-Jarallah

    Full Text Available HDL carries biologically active lipids such as sphingosine-1-phosphate (S1P and stimulates a variety of cell signaling pathways in diverse cell types, which may contribute to its ability to protect against atherosclerosis. HDL and sphingosine-1-phosphate receptor agonists, FTY720 and SEW2871 triggered macrophage migration. HDL-, but not FTY720-stimulated migration was inhibited by an antibody against the HDL receptor, SR-BI, and an inhibitor of SR-BI mediated lipid transfer. HDL and FTY720-stimulated migration was also inhibited in macrophages lacking either SR-BI or PDZK1, an adaptor protein that binds to SR-BI's C-terminal cytoplasmic tail. Migration in response to HDL and S1P receptor agonists was inhibited by treatment of macrophages with sphingosine-1-phosphate receptor type 1 (S1PR1 antagonists and by pertussis toxin. S1PR1 activates signaling pathways including PI3K-Akt, PKC, p38 MAPK, ERK1/2 and Rho kinases. Using selective inhibitors or macrophages from gene targeted mice, we demonstrated the involvement of each of these pathways in HDL-dependent macrophage migration. These data suggest that HDL stimulates the migration of macrophages in a manner that requires the activities of the HDL receptor SR-BI as well as S1PR1 activity.

  2. TGF-α/HA complex promotes tympanic membrane keratinocyte migration and proliferation via ErbB1 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Mei Teh, Bing, E-mail: bing.teh@earscience.org.au [Ear Sciences Centre, School of Surgery, The University of Western Australia, Nedlands, WA (Australia); Ear Science Institute Australia, Subiaco, WA (Australia); Department of Otolaryngology, Head, Neck and Skull Base Surgery, Sir Charles Gairdner Hospital, Nedlands, WA (Australia); Redmond, Sharon L. [Ear Sciences Centre, School of Surgery, The University of Western Australia, Nedlands, WA (Australia); Ear Science Institute Australia, Subiaco, WA (Australia); Shen, Yi [Ear Sciences Centre, School of Surgery, The University of Western Australia, Nedlands, WA (Australia); Ear Science Institute Australia, Subiaco, WA (Australia); Department of Otolaryngology, Head and Neck, Ningbo Lihuili Hospital (Ningbo Medical Centre), Ningbo, Zhejiang (China); Atlas, Marcus D. [Ear Sciences Centre, School of Surgery, The University of Western Australia, Nedlands, WA (Australia); Ear Science Institute Australia, Subiaco, WA (Australia); Department of Otolaryngology, Head, Neck and Skull Base Surgery, Sir Charles Gairdner Hospital, Nedlands, WA (Australia); Marano, Robert J.; Dilley, Rodney J. [Ear Sciences Centre, School of Surgery, The University of Western Australia, Nedlands, WA (Australia); Ear Science Institute Australia, Subiaco, WA (Australia)

    2013-04-01

    Tympanic membrane perforations are common and represent a management challenge to clinicians. Current treatments for chronic perforations involve a graft surgery and require general anaesthesia, including associated costs and morbidities. Bioactive molecules (e.g. growth factors, cytokines) play an important role in promoting TM wound healing following perforation and the use of growth factors as a topical treatment for tympanic membrane perforations has been suggested as an alternative to surgery. However, the choice of bioactive molecules best suited to promote wound healing has yet to be identified. We investigated the effects of hyaluronic acid, vitronectin, TGF-α, IL-24 and their combinations on migration, proliferation and adhesion of cultured human tympanic membrane-derived keratinocytes (hTM), in addition to their possible mechanisms of action. We found that TGF-α, TGF-α/HA and TGF-α/IL-24 promoted wound healing by significantly increasing both migration and proliferation. TGF-α and/or HA treated cells showed comparable cell–cell adhesion whilst maintaining an epithelial cell phenotype. With the use of receptor binding inhibitors for ErbB1 (AG1478) and CD44 (BRIC235), we revealed that the activation of ErbB1 is required for TGF-α/HA-mediated migration and proliferation. These results suggest factors that may be incorporated into a tissue-engineered membrane or directly as topical treatment for tympanic membrane perforations and hence reduce the need for a surgery. - Highlights: ► TGF-α, TGF-α/HA and TGF-α/IL-24 improved hTM keratinocyte migration and proliferation. ► TGF-α and/or HA maintained epithelial cell phenotype. ► TGF-α/HA-mediated migration and proliferation requires activation of ErbB1 receptor.

  3. RhoC Mediates Epidermal Growth Factor-Stimulated Migration and Invasion in Head and Neck Squamous Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Zohra Tumur

    2015-01-01

    Full Text Available Epidermal growth factor receptor (EGFR is overexpressed in head and neck squamous cell carcinoma (HNSCC where it has been shown to promote tumor cell invasion upon phosphorylation. One mechanism by which EGFR promotes tumor progression is by activating signal cascades that lead to loss of E-cadherin, a transmembrane glycoprotein of the cell-cell adherence junctions; however mediators of these signaling cascades are not fully understood. One such mediator, RhoC, is activated upon a number of external stimuli, such as epidermal growth factor (EGF, but its role as a mediator of EGF-stimulated migration and invasion has not been elucidated in HNSCC. In the present study, we investigate the role of RhoC as a mediator of EGF-stimulated migration and invasion in HNSCC. We show that upon EGF stimulation, EGFR and RhoC were strongly activated in HNSCC. This resulted in activation of the phosphatidylinositol 3-Kinase Akt pathway (PI3K-Akt, phosphorylation of GSK-3β at the Ser9 residue, and subsequent down regulation of E-cadherin cell surface expression resulting in increased tumor cell invasion. Knockdown of RhoC restored E-cadherin expression and inhibited EGF-stimulated migration and invasion. This is the first report in HNSCC demonstrating the role RhoC plays in mediating EGF-stimulated migration and invasion by down-regulating the PI3K-Akt pathway and E-cadherin expression. RhoC may serve as a treatment target for HNSCC.

  4. Scavenger receptor class B type 1 regulates neuroblastoma cell proliferation, migration and invasion.

    Science.gov (United States)

    Panchoo, Marlyn; Lacko, Andras

    2018-01-01

    Neuroblastoma (NB) is an extra cranial pediatric embryonal tumor most prevalent in children less than 1 year of age. NB accounts for 7% of all pediatric cancers but accounts for 15% of all childhood cancer deaths. Scavenger receptor class B type 1 (SR-B1), a mediator of cellular cholesterol uptake, is overexpressed in and have been linked to the aggressiveness of many cancers. Nevertheless, no studies have so far investigated the relationship between SR-B1 and NB. Elucidation of receptors that promote NB may pave the way for discovery of new therapeutic targets. Here we show that inhibition of SR-B1 reduced cell survival, migration and invasion, and cholesterol content in NB cell lines. Additionally analysis of SR-B1 levels in NB patient biopsies using the R2: Genomics Analysis and Visualization Platform showed that high SR-B1 expression correlated with decreased overall and event-free survival. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Receptor-mediated transport of oligodeoxynucleotides into hepatic cells.

    Science.gov (United States)

    Reinis, M; Damková, M; Korec, E

    1993-04-01

    Receptor-mediated endocytosis was employed for a highly efficient transport of oligodeoxynucleotides into hepatoma cell line PLC/PRF/5. The oligodeoxynucleotides were bound to the asialofetuin-poly-L-lysine conjugate and this complex was internalized by the cells via asialoglycoprotein receptor, an endocytic receptor unique for hepatocytes. Binding of the oligodeoxynucleotides to the complex dramatically increased their cellular uptake more than 20-fold. Chloroquine, a lysosomatropic agent, further increased the transport of the complex but not of the free oligodeoxynucleotides.

  6. Loss of Cannabinoid CB1 Receptors Induces Cortical Migration Malformations and Increases Seizure Susceptibility.

    Science.gov (United States)

    Díaz-Alonso, Javier; de Salas-Quiroga, Adán; Paraíso-Luna, Juan; García-Rincón, Daniel; Garcez, Patricia P; Parsons, Maddy; Andradas, Clara; Sánchez, Cristina; Guillemot, François; Guzmán, Manuel; Galve-Roperh, Ismael

    2017-11-01

    Neuronal migration is a fundamental process of brain development, and its disruption underlies devastating neurodevelopmental disorders. The transcriptional programs governing this process are relatively well characterized. However, how environmental cues instruct neuronal migration remains poorly understood. Here, we demonstrate that the cannabinoid CB1 receptor is strictly required for appropriate pyramidal neuron migration in the developing cortex. Acute silencing of the CB1 receptor alters neuronal morphology and impairs radial migration. Consequently, CB1 siRNA-electroporated mice display cortical malformations mimicking subcortical band heterotopias and increased seizure susceptibility in adulthood. Importantly, rescuing the CB1 deficiency-induced radial migration arrest by knockdown of the GTPase protein RhoA restored the hyperexcitable neuronal network and seizure susceptibility. Our findings show that CB1 receptor/RhoA signaling regulates pyramidal neuron migration, and that deficient CB1 receptor signaling may contribute to cortical development malformations leading to refractory epilepsy independently of its canonical neuromodulatory role in the adult brain. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  7. RTN3 Regulates the Expression Level of Chemokine Receptor CXCR4 and is Required for Migration of Primordial Germ Cells

    Directory of Open Access Journals (Sweden)

    Haitao Li

    2016-04-01

    Full Text Available CXCR4 is a crucial chemokine receptor that plays key roles in primordial germ cell (PGC homing. To further characterize the CXCR4-mediated migration of PGCs, we screened CXCR4-interacting proteins using yeast two-hybrid screening. We identified reticulon3 (RTN3, a member of the reticulon family, and considered an apoptotic signal transducer, as able to interact directly with CXCR4. Furthermore, we discovered that the mRNA and protein expression levels of CXCR4 could be regulated by RTN3. We also found that RTN3 altered CXCR4 translocation and localization. Moreover, increasing the signaling of either CXCR4b or RTN3 produced similar PGC mislocalization phenotypes in zebrafish. These results suggested that RTN3 modulates PGC migration through interaction with, and regulation of, CXCR4.

  8. An evolutionarily conserved arginine is essential for Tre1 G protein-coupled receptor function during germ cell migration in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Angela R Kamps

    Full Text Available BACKGROUND: G protein-coupled receptors (GPCRs play central roles in mediating cellular responses to environmental signals leading to changes in cell physiology and behaviors, including cell migration. Numerous clinical pathologies including metastasis, an invasive form of cell migration, have been linked to abnormal GPCR signaling. While the structures of some GPCRs have been defined, the in vivo roles of conserved amino acid residues and their relationships to receptor function are not fully understood. Trapped in endoderm 1 (Tre1 is an orphan receptor of the rhodopsin class that is necessary for primordial germ cell migration in Drosophila melanogaster embryos. In this study, we employ molecular genetic approaches to identify residues in Tre1 that are critical to its functions in germ cell migration. METHODOLOGY/PRINCIPAL FINDINGS: First, we show that the previously reported scattershot mutation is an allele of tre1. The scattershot allele results in an in-frame deletion of 8 amino acids at the junction of the third transmembrane domain and the second intracellular loop of Tre1 that dramatically impairs the function of this GPCR in germ cell migration. To further refine the molecular basis for this phenotype, we assayed the effects of single amino acid substitutions in transgenic animals and determined that the arginine within the evolutionarily conserved E/N/DRY motif is critical for receptor function in mediating germ cell migration within an intact developing embryo. CONCLUSIONS/SIGNIFICANCE: These structure-function studies of GPCR signaling in native contexts will inform future studies into the basic biology of this large and clinically important family of receptors.

  9. Src tyrosine kinase mediates platelet-derived growth factor BB-induced and redox-dependent migration in metanephric mesenchymal cells

    Science.gov (United States)

    Gorin, Yves

    2013-01-01

    The adult kidney is derived from the interaction between the metanephric blastema and the ureteric bud. Platelet-derived growth factor (PDGF) receptor β is essential for the development of the mature glomerular tuft, as mice deficient for this receptor lack mesangial cells. This study investigated the role of Src tyrosine kinase in PDGF-mediated reactive oxygen species (ROS) generation and migration of metanephric mesenchymal cells (MMCs). Cultured embryonic MMCs from wild-type and PDGF receptor-deficient embryos were established. Migration was determined via wound-healing assay. Unlike PDGF AA, PDGF BB-induced greater migration in MMCs with respect to control. This was abrogated by neutralizing an antibody to PDGF BB. Phosphatidylinositol 3-kinase (PI3K) inhibitors suppressed PDGF BB-induced migration. Conversely, mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) inhibitors had no effect. Src inhibitors inhibited PDGF-induced cell migration, PI3K activity, and Akt phosphorylation. Adenoviral dominant negative Src (AD DN Src) abrogated PDGF BB-induced Akt phosphorylation. Hydrogen peroxide stimulated cell migration. PDGF BB-induced wound closure was inhibited by the antioxidants N-acetyl-l-cysteine, tiron, and the flavoprotein inhibitor diphenyleneiodonium. These cells express the NADPH oxidase homolog Nox4. Inhibiting Nox4 with antisense oligonucleotides or small interfering RNA (siRNA) suppressed PDGF-induced wound closure. Inhibition of Src with siRNA reduced PDGF BB-induced ROS generation as assessed by 2′,7′-dichlorodihydrofluorescein diacetate fluorescence. Furthermore, PDGF BB-stimulated ROS generation and migration were similarly suppressed by Ad DN Src. In MMCs, PDGF BB-induced migration is mediated by PI3K and Src in a redox-dependent manner involving Nox4. Src may be upstream to PI3K and Nox4. PMID:24197068

  10. C-X-C chemokine receptor type 4 (CXCR4) is a key receptor for chicken primordial germ cell migration.

    Science.gov (United States)

    Lee, Jeong Hyo; Park, Jeong-Woong; Kim, Si Won; Park, Joonghoon; Park, Tae Sub

    2017-12-15

    In mammals, germ cells originate outside of the developing gonads and follow a unique migration pattern through the embryonic tissue toward the genital ridges. Many studies have attempted to identify critical receptors and factors involved in germ cell migration. However, relatively few reports exist on germ cell receptors and chemokines that are involved in germ cell migration in avian species. In the present study, we investigated the specific migratory function of C-X-C chemokine receptor type 4 (CXCR4) in chicken primordial germ cells (PGCs). We induced loss-of-function via a frameshift mutation in the CXCR4 gene in chicken PGCs using clustered regularly interspaced short palindromic repeat-CRISPR-associated protein 9 (CRISPR/Cas9) genome editing. The migratory capacity of CXCR4 knockout PGCs was significantly reduced in vivo after transplantation into recipient embryos. However, CXCR4-expressing somatic cell lines, such as chicken DT40 and DF1, failed to migrate into the developing gonads, suggesting that another key factor(s) is necessary for targeting and settlement of PGCs into the genital ridges. In conclusion, we show that CXCR4 plays a critical role in the migration of chicken germ cells.

  11. Serotonin receptor 3A controls interneuron migration into the neocortex

    NARCIS (Netherlands)

    Murthy, S.; Niquille, M.; Hurni, N.; Limoni, G.; Frazer, S.; Chameau, P.; van Hooft, J.A.; Vitalis, T.; Dayer, A.

    2014-01-01

    Neuronal excitability has been shown to control the migration and cortical integration of reelin-expressing cortical interneurons (INs) arising from the caudal ganglionic eminence (CGE), supporting the possibility that neurotransmitters could regulate this process. Here we show that the ionotropic

  12. Molecular Mechanisms of Dopamine Receptor Mediated Neuroprotection

    National Research Council Canada - National Science Library

    Sealfon, Stuart

    2000-01-01

    ... of the cellular changes characteristic of this process. Evidence from our laboratory and others suggest that activation of dopamine receptors can oppose the induction of apoptosis in dopamine neurons...

  13. High glucose-mediated oxidative stress impairs cell migration.

    Directory of Open Access Journals (Sweden)

    Marcelo L Lamers

    Full Text Available Deficient wound healing in diabetic patients is very frequent, but the cellular and molecular causes are poorly defined. In this study, we evaluate the hypothesis that high glucose concentrations inhibit cell migration. Using CHO.K1 cells, NIH-3T3 fibroblasts, mouse embryonic fibroblasts and primary skin fibroblasts from control and diabetic rats cultured in 5 mM D-glucose (low glucose, LG, 25 mM D-glucose (high glucose, HG or 25 mM L-glucose medium (osmotic control--OC, we analyzed the migration speed, protrusion stability, cell polarity, adhesion maturation and the activity of the small Rho GTPase Rac1. We also analyzed the effects of reactive oxygen species by incubating cells with the antioxidant N-Acetyl-Cysteine (NAC. We observed that HG conditions inhibited cell migration when compared to LG or OC. This inhibition resulted from impaired cell polarity, protrusion destabilization and inhibition of adhesion maturation. Conversely, Rac1 activity, which promotes protrusion and blocks adhesion maturation, was increased in HG conditions, thus providing a mechanistic basis for the HG phenotype. Most of the HG effects were partially or completely rescued by treatment with NAC. These findings demonstrate that HG impairs cell migration due to an increase in oxidative stress that causes polarity loss, deficient adhesion and protrusion. These alterations arise, in large part, from increased Rac1 activity and may contribute to the poor wound healing observed in diabetic patients.

  14. The urokinase receptor takes control of cell migration by recruiting integrins and FPR1 on the cell surface.

    Directory of Open Access Journals (Sweden)

    Anna Gorrasi

    Full Text Available The receptor (uPAR of the urokinase-type plasminogen activator (uPA is crucial in cell migration since it concentrates uPA proteolytic activity at the cell surface, binds vitronectin and associates to integrins. uPAR cross-talk with receptors for the formylated peptide fMLF (fMLF-Rs has been reported; however, cell-surface uPAR association to fMLF-Rs on the cell membrane has never been explored in detail. We now show that uPAR co-localizes at the cell-surface and co-immunoprecipitates with the high-affinity fMLF-R, FPR1, in uPAR-transfected HEK-293 (uPAR-293 cells. uPAR/β1 integrin and FPR1/β1 integrin co-localization was also observed. Serum or the WKYMVm peptide (W Pep, a FPR1 ligand, strongly increased all observed co-localizations in uPAR-293 cells, including FPR1/β1 integrin co-localization. By contrast, a low FPR1/β1 integrin co-localization was observed in uPAR-negative vector-transfected HEK-293 (V-293 cells, that was not increased by serum or W Pep stimulations. The role of uPAR interactions in cell migration was then explored. Both uPAR-293 and V-293 control cells efficiently migrated toward serum or purified EGF. However, cell treatments impairing uPAR interactions with fMLF-Rs or integrins, or inhibiting specific cell-signaling mediators abrogated uPAR-293 cell migration, without exerting any effect on V-293 control cells. Accordingly, uPAR depletion by a uPAR-targeting siRNA or uPAR blocking with an anti-uPAR polyclonal antibody in cells constitutively expressing high uPAR levels totally impaired their migration toward serum. Altogether, these results suggest that both uPAR-positive and uPAR-negative cells are able to migrate toward serum; however, uPAR expression renders cell migration totally and irreversibly uPAR-dependent, since it is completely inhibited by uPAR blocking. We propose that uPAR takes control of cell migration by recruiting fMLF-Rs and β1 integrins, thus promoting their co-localization at the cell-surface and

  15. Myosin II governs collective cell migration behaviour downstream of guidance receptor signalling.

    Science.gov (United States)

    Combedazou, Anne; Choesmel-Cadamuro, Valérie; Gay, Guillaume; Liu, Jiaying; Dupré, Loïc; Ramel, Damien; Wang, Xiaobo

    2017-01-01

    Border cell migration during Drosophila oogenesis is a potent model to study collective cell migration, a process involved in development and metastasis. Border cell clusters adopt two main types of behaviour during migration: linear and rotational. However, the molecular mechanism controlling the switch from one to the other is unknown. Here, we demonstrate that non-muscle Myosin II (NMII, also known as Spaghetti squash) activity controls the linear-to-rotational switch. Furthermore, we show that the regulation of NMII takes place downstream of guidance receptor signalling and is critical to ensure efficient collective migration. This study thus provides new insight into the molecular mechanism coordinating the different cell behaviours in a migrating cluster. © 2017. Published by The Company of Biologists Ltd.

  16. NMDA receptors mediate synaptic competition in culture.

    Directory of Open Access Journals (Sweden)

    Kevin She

    Full Text Available Activity through NMDA type glutamate receptors sculpts connectivity in the developing nervous system. This topic is typically studied in the visual system in vivo, where activity of inputs can be differentially regulated, but in which individual synapses are difficult to visualize and mechanisms governing synaptic competition can be difficult to ascertain. Here, we develop a model of NMDA-receptor dependent synaptic competition in dissociated cultured hippocampal neurons.GluN1 -/- (KO mouse hippocampal neurons lacking the essential NMDA receptor subunit were cultured alone or cultured in defined ratios with wild type (WT neurons. The absence of functional NMDA receptors did not alter neuron survival. Synapse development was assessed by immunofluorescence for postsynaptic PSD-95 family scaffold and apposed presynaptic vesicular glutamate transporter VGlut1. Synapse density was specifically enhanced onto minority wild type neurons co-cultured with a majority of GluN1 -/- neighbour neurons, both relative to the GluN1 -/- neighbours and relative to sister pure wild type cultures. This form of synaptic competition was dependent on NMDA receptor activity and not conferred by the mere physical presence of GluN1. In contrast to these results in 10% WT and 90% KO co-cultures, synapse density did not differ by genotype in 50% WT and 50% KO co-cultures or in 90% WT and 10% KO co-cultures.The enhanced synaptic density onto NMDA receptor-competent neurons in minority coculture with GluN1 -/- neurons represents a cell culture paradigm for studying synaptic competition. Mechanisms involved may include a retrograde 'reward' signal generated by WT neurons, although in this paradigm there was no 'punishment' signal against GluN1 -/- neurons. Cell culture assays involving such defined circuits may help uncover the rules and mechanisms of activity-dependent synaptic competition in the developing nervous system.

  17. Cell-mediated immune response in rotavirus-infected calves: leucocyte migration inhibition assay.

    Science.gov (United States)

    Chauhan, R S; Singh, N P

    1992-07-01

    The cell-mediated immune (CMI) response was determined in rotavirus-infected calves by leucocyte migration inhibition assay with blood, spleen, mesenteric lymph node and intestinal lymphocytes. The inhibition of migration was more prominent in intestinal and mesenteric lymph node lymphocytes than in spleen and blood. In rotavirus-infected calves, the assay indicated the presence of CMI response which was more prominent at the local site of infection.

  18. Discovery of the migrasome, an organelle mediating release of cytoplasmic contents during cell migration.

    Science.gov (United States)

    Ma, Liang; Li, Ying; Peng, Junya; Wu, Danni; Zhao, Xiaoxin; Cui, Yitong; Chen, Lilian; Yan, Xiaojun; Du, Yanan; Yu, Li

    2015-01-01

    Cells communicate with each other through secreting and releasing proteins and vesicles. Many cells can migrate. In this study, we report the discovery of migracytosis, a cell migration-dependent mechanism for releasing cellular contents, and migrasomes, the vesicular structures that mediate migracytosis. As migrating cells move, they leave long tubular strands, called retraction fibers, behind them. Large vesicles, which contain numerous smaller vesicles, grow on the tips and intersections of retraction fibers. These fibers, which connect the vesicles with the main cell body, eventually break, and the vesicles are released into the extracellular space or directly taken up by surrounding cells. Since the formation of these vesicles is migration-dependent, we named them "migrasomes". We also found that cytosolic contents can be transported into migrasomes and released from the cell through migrasomes. We named this migration-dependent release mechanism "migracytosis".

  19. Relations between immune and mediator receptors of mouse lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Ado, A.D.; Alekseeva, T.A.; Kravchenko, S.A.

    1985-10-01

    This paper examines the action of the specific muscarinic antogonist tritium-quinuclidinyl benzilate (tritium-QNB) on immune rosette formation in mice. It is shown that since the specific muscarini antagonist tritium-QNB inhibits immune rosette formation, this process must be regarded as interconnected with muscarinic receptors of lymphocytes. Interaction of immune (antigen-binding) and mediator receptors, however, is an important factor maintaining immune homeostasis at a certain level.

  20. FASN, ErbB2-mediated glycolysis is required for breast cancer cell migration.

    Science.gov (United States)

    Zhou, Lan; Jiang, Sufang; Fu, Qiang; Smith, Kelly; Tu, Kailing; Li, Hua; Zhao, Yuhua

    2016-05-01

    Both fatty acid synthase (FASN) and ErbB2 have been shown to promote breast cancer cell migration. However, the underlying molecular mechanism remains poorly understood and there is no reported evidence that directly links glycolysis to breast cancer cell migration. In this study, we investigated the role of FASN, ErbB2-mediated glycolysis in breast cancer cell migration. First, we compared lactate dehydrogenase A (LDHA) protein levels, glycolysis and cell migration between FASN, ErbB2-overexpressing SK-BR-3 cells and FASN, ErbB2-low-expressing MCF7 cells. Then, SK-BR-3 cells were treated with cerulenin (Cer), an inhibitor of FASN, and ErbB2, LDHA protein levels, glycolysis, and cell migration were detected. Next, we transiently transfected ErbB2 plasmid into MCF7 cells and detected FASN, LDHA protein levels, glycolysis and cell migration. Heregulin-β1 (HRG-β1) is an activator of ErbB2 and 2-deoxyglucose (2-DG) and oxamate (OX) are inhibitors of glycolysis. MCF7 cells were treated with HRG-β1 alone, HRG-β1 plus 2-DG, OX or cerulenin and glycolysis, and cell migration were measured. We found that FASN, ErbB2-high-expressing SK-BR-3 cells displayed higher levels of glycolysis and migration than FASN, ErbB2-low-expressing MCF7 cells. Inhibition of FASN by cerulenin impaired glycolysis and migration in SK-BR-3 cells. Transient overexpression of ErbB2 in MCF7 cells promotes glycolysis and migration. Moreover, 2-deoxyglucose (2-DG), oxamate (OX), or cerulenin partially reverses heregulin-β1 (HRG-β1)-induced glycolysis and migration in MCF7 cells. In conclusion, this study demonstrates that FASN, ErbB2-mediated glycolysis is required for breast cancer cell migration. These novel findings indicate that targeting FASN, ErbB2-mediated glycolysis may be a new approach to reverse breast cancer cell migration.

  1. Neutrophil Activation of Endothelial Cell-Expressed TRPM2 Mediates Transendothelial Neutrophil Migration and Vascular Injury.

    Science.gov (United States)

    Mittal, Manish; Nepal, Saroj; Tsukasaki, Yoshikazu; Hecquet, Claudie M; Soni, Dheeraj; Rehman, Jalees; Tiruppathi, Chinnaswamy; Malik, Asrar B

    2017-10-13

    TRPM2 (transient receptor potential melastatin-2) expressed in endothelial cells (ECs) is a cation channel mediating Ca2+ entry in response to intracellular generation of adenosine diphosphoribose-the TRPM2 ligand. Because polymorphonuclear neutrophils (PMN) interaction with ECs generates reactive oxygen species, we addressed the possible role of TRPM2 expressed in ECs in the mechanism of transendothelial migration of PMNs. We observed defective PMN transmigration in response to lipopolysaccharide challenge in adult mice in which the EC expressed TRPM2 is conditionally deleted (Trpm2iΔEC ). PMN interaction with ECs induced the entry of Ca2+ in ECs via the EC-expressed TRPM2. Prevention of generation of adenosine diphosphoribose in ECs significantly reduced Ca2+ entry in response to PMN activation of TRPM2 in ECs. PMNs isolated from gp91phox-/- mice significantly reduced Ca2+ entry in ECs via TRPM2 as compared with wild-type PMNs and failed to induce PMN transmigration. Overexpression of the adenosine diphosphoribose insensitive TRPM2 mutant channel (C1008→A) in ECs suppressed the Ca2+ entry response. Further, the forced expression of TRPM2 mutant channel (C1008→A) or silencing of poly ADP-ribose polymerase in ECs of mice prevented PMN transmigration. Thus, endotoxin-induced transmigration of PMNs was secondary to TRPM2-activated Ca2+ signaling and VE-cadherin phosphorylation resulting in the disassembly of adherens junctions and opening of the paracellular pathways. These results suggest blocking TRPM2 activation in ECs is a potentially important means of therapeutically modifying PMN-mediated vascular inflammation. © 2017 American Heart Association, Inc.

  2. Probing Biased Signaling in Chemokine Receptors

    DEFF Research Database (Denmark)

    Amarandi, Roxana Maria; Hjortø, Gertrud Malene; Rosenkilde, Mette Marie

    2016-01-01

    The chemokine system mediates leukocyte migration during homeostatic and inflammatory processes. Traditionally, it is described as redundant and promiscuous, with a single chemokine ligand binding to different receptors and a single receptor having several ligands. Signaling of chemokine receptors...

  3. A Peptide Antagonist of the ErbB1 Receptor Inhibits Receptor Activation, Tumor Cell Growth and Migration In Vitro and Xenograft Tumor Growth In Vivo

    Science.gov (United States)

    Xu, Ruodan; Povlsen, Gro Klitgaard; Soroka, Vladislav; Bock, Elisabeth; Berezin, Vladimir

    2010-01-01

    The epidermal growth factor family of receptor tyrosine kinases (ErbBs) plays essential roles in tumorigenesis and cancer disease progression, and therefore has become an attractive target for structure-based drug design. ErbB receptors are activated by ligand-induced homo- and heterodimerization. Structural studies have revealed that ErbB receptor dimers are stabilized by receptor–receptor interactions, primarily mediated by a region in the second extracellular domain, termed the “dimerization arm”. The present study is the first biological characterization of a peptide, termed Inherbin3, which constitutes part of the dimerization arm of ErbB3. Inherbin3 binds to the extracellular domains of all four ErbB receptors, with the lowest peptide binding affinity for ErbB4. Inherbin3 functions as an antagonist of epidermal growth factor (EGF)-ErbB1 signaling. We show that Inherbin3 inhibits EGF-induced ErbB1 phosphorylation, cell growth, and migration in two human tumor cell lines, A549 and HN5, expressing moderate and high ErbB1 levels, respectively. Furthermore, we show that Inherbin3 inhibits tumor growth in vivo and induces apoptosis in a tumor xenograft model employing the human non-small cell lung cancer cell line A549. The Inherbin3 peptide may be a useful tool for investigating the mechanisms of ErbB receptor homo- and heterodimerization. Moreover, the here described biological effects of Inherbin3 suggest that peptide-based targeting of ErbB receptor dimerization is a promising anti-cancer therapeutic strategy. PMID:20364069

  4. A Peptide Antagonist of the ErbB1 Receptor Inhibits Receptor Activation, Tumor Cell Growth and Migration In Vitro and Xenograft Tumor Growth In Vivo

    Directory of Open Access Journals (Sweden)

    Ruodan Xu

    2010-01-01

    Full Text Available The epidermal growth factor family of receptor tyrosine kinases (ErbBs plays essential roles in tumorigenesis and cancer disease progression, and therefore has become an attractive target for structure-based drug design. ErbB receptors are activated by ligand-induced homo- and heterodimerization. Structural studies have revealed that ErbB receptor dimers are stabilized by receptor–receptor interactions, primarily mediated by a region in the second extracellular domain, termed the “dimerization arm”. The present study is the first biological characterization of a peptide, termed Inherbin3, which constitutes part of the dimerization arm of ErbB3. Inherbin3 binds to the extracellular domains of all four ErbB receptors, with the lowest peptide binding affinity for ErbB4. Inherbin3 functions as an antagonist of epidermal growth factor (EGF-ErbB1 signaling. We show that Inherbin3 inhibits EGF-induced ErbB1 phosphorylation, cell growth, and migration in two human tumor cell lines, A549 and HN5, expressing moderate and high ErbB1 levels, respectively. Furthermore, we show that Inherbin3 inhibits tumor growth in vivo and induces apoptosis in a tumor xenograft model employing the human non-small cell lung cancer cell line A549. The Inherbin3 peptide may be a useful tool for investigating the mechanisms of ErbB receptor homo- and heterodimerization. Moreover, the here described biological effects of Inherbin3 suggest that peptide-based targeting of ErbB receptor dimerization is a promising anti-cancer therapeutic strategy.

  5. Migration of mesenchymal stem cells towards glioblastoma cells depends on hepatocyte-growth factor and is enhanced by aminolaevulinic acid-mediated photodynamic treatment.

    Science.gov (United States)

    Vogel, Sebastian; Peters, Corinna; Etminan, Nima; Börger, Verena; Schimanski, Adrian; Sabel, Michael C; Sorg, Rüdiger V

    2013-02-15

    Hepatocyte-growth factor (HGF) is expressed by glioblastomas and contributes to their growth, migration and invasion. HGF also mediates migration of mesenchymal stem cells (MSC) to sites of apoptotic cell death. Moreover, MSC show tropism for glioblastomas, which is exploited in gene therapy to deliver the therapeutics to the tumor cells. Here, we have studied whether HGF contributes to the recruitment of MSC by glioblastoma cells and whether aminolaevulinic acid-mediated photodynamic therapy (ALA/PDT), a novel therapeutic approach that induces apoptosis in glioblastoma cells, affects HGF release and this migratory response. MSC expressed the HGF receptor MET and migrated towards U87 and U251 glioblastoma spheroids. Migration increased significantly when spheroids were subjected to ALA/PDT, which was associated with induction of apoptosis and up-regulation of HGF. Neutralizing HGF resulted in significant inhibition of MSC migration towards untreated as well as ALA/PDT-treated spheroids. Thus, glioblastoma cells express HGF, which contributes to the attraction of MSC. ALA/PDT induces apoptosis and augments HGF release causing enhanced MSC migration towards the tumor cells. ALA/PDT may therefore be exploited to improve targeting of MSC delivered gene therapy, but it may also constitute a risk in terms of beneficial effects for the tumor. Copyright © 2013 Elsevier Inc. All rights reserved.

  6. Syk-mediated translocation of PI3Kdelta to the leading edge controls lamellipodium formation and migration of leukocytes.

    Directory of Open Access Journals (Sweden)

    Jürgen Schymeinsky

    2007-11-01

    Full Text Available The non-receptor tyrosine kinase Syk is mainly expressed in the hematopoietic system and plays an essential role in beta(2 integrin-mediated leukocyte activation. To elucidate the signaling pathway downstream of Syk during beta2 integrin (CD11/CD18-mediated migration and extravasation of polymorphonuclear neutrophils (PMN, we generated neutrophil-like differentiated HL-60 (dHL-60 cells expressing a fluorescently tagged Syk mutant lacking the tyrosine residue at the position 323 (Syk-Tyr323 that is known to be required for the binding of the regulatory subunit p85 of the phosphatidylinositol 3-kinase (PI3K class I(A. Syk-Tyr323 was found to be critical for the enrichment of the catalytic subunit p110delta of PI3K class I(A as well as for the generation of PI3K products at the leading edge of the majority of polarized cells. In accordance, the translocation of PI3K p110delta to the leading edge was diminished in Syk deficient murine PMN. Moreover, the expression of EGFP-Syk Y323F interfered with proper cell polarization and it impaired efficient migration of dHL-60 cells. In agreement with a major role of beta2 integrins in the recruitment of phagocytic cells to sites of lesion, mice with a Syk-deficient hematopoietic system demonstrated impaired PMN infiltration into the wounded tissue that was associated with prolonged cutaneous wound healing. These data imply a novel role of Syk via PI3K p110delta signaling for beta2 integrin-mediated migration which is a prerequisite for efficient PMN recruitment in vivo.

  7. [Effect of androgen receptor on IgG expression, proliferation and migration of prostate cancer cells in vitro].

    Science.gov (United States)

    Deng, Yu-Lin; Guo, Kai; Zeng, Ying-Ke; Wu, Kai-Hui; Tang, Chen; Zheng, Shao-Bo

    2017-03-20

    To investigate the effect of androgen receptor (AR) on IgG protein expression and the proliferation and migration of prostate cancer cells. Western blotting was used to detect the expression of AR protein and IgG in androgen-dependent prostate cancer LNCap cells and castration-resistant prostate cancer PC-3 cells. In AR-overexpressing cells (PC-3-AR cells) established by transfecting PC-3 with AR gene (pCDNA3.1) and LNCap cells with small interfering RNA-mediated AR silencing (LNCap-siAR cells) were analyzed for expressions of AR protein and IgG with Western blotting; the expression of IgG mRNA was detected by Q-PCR, and the cell proliferation and migration were assessed with MTT assay and wound healing assay, respectively. Compared with PC-3 cells, LNCap cells expressed a higher level of AR protein and a lower level of IgG (PIgG (PIgG (PIgG and is associated with the proliferation and migration of prostate cancer cells in vitro.

  8. Src homology 2-domain containing leukocyte-specific phosphoprotein of 76 kDa is mandatory for TCR-mediated inside-out signaling, but dispensable for CXCR4-mediated LFA-1 activation, adhesion, and migration of T cells.

    Science.gov (United States)

    Horn, Jessica; Wang, Xiaoqian; Reichardt, Peter; Stradal, Theresia E; Warnecke, Nicole; Simeoni, Luca; Gunzer, Matthias; Yablonski, Deborah; Schraven, Burkhart; Kliche, Stefanie

    2009-11-01

    Engagement of the TCR or of chemokine receptors such as CXCR4 induces adhesion and migration of T cells via so-called inside-out signaling pathways. The molecular processes underlying inside-out signaling events are as yet not completely understood. In this study, we show that TCR- and CXCR4-mediated activation of integrins critically depends on the membrane recruitment of the adhesion- and degranulation-promoting adapter protein (ADAP)/Src kinase-associated phosphoprotein of 55 kDa (SKAP55)/Rap1-interacting adapter protein (RIAM)/Rap1 module. We further demonstrate that the Src homology 2 domain containing leukocyte-specific phosphoprotein of 76 kDa (SLP76) is crucial for TCR-mediated inside-out signaling and T cell/APC interaction. Besides facilitating membrane recruitment of ADAP, SKAP55, and RIAM, SLP76 regulates TCR-mediated inside-out signaling by controlling the activation of Rap1 as well as Rac-mediated actin polymerization. Surprisingly, however, SLP76 is not mandatory for CXCR4-mediated inside-out signaling. Indeed, both CXCR4-induced T cell adhesion and migration are not affected by loss of SLP76. Moreover, after CXCR4 stimulation, the ADAP/SKAP55/RIAM/Rap1 module is recruited to the plasma membrane independently of SLP76. Collectively, our data indicate a differential requirement for SLP76 in TCR- vs CXCR4-mediated inside-out signaling pathways regulating T cell adhesion and migration.

  9. Fcγ receptor-mediated inflammation inhibits axon regeneration.

    Directory of Open Access Journals (Sweden)

    Gang Zhang

    Full Text Available Anti-glycan/ganglioside antibodies are the most common immune effectors found in patients with Guillain-Barré Syndrome, which is a peripheral autoimmune neuropathy. We previously reported that disease-relevant anti-glycan autoantibodies inhibited axon regeneration, which echo the clinical association of these antibodies and poor recovery in Guillain-Barré Syndrome. However, the specific molecular and cellular elements involved in this antibody-mediated inhibition of axon regeneration are not previously defined. This study examined the role of Fcγ receptors and macrophages in the antibody-mediated inhibition of axon regeneration. A well characterized antibody passive transfer sciatic nerve crush and transplant models were used to study the anti-ganglioside antibody-mediated inhibition of axon regeneration in wild type and various mutant and transgenic mice with altered expression of specific Fcγ receptors and macrophage/microglia populations. Outcome measures included behavior, electrophysiology, morphometry, immunocytochemistry, quantitative real-time PCR, and western blotting. We demonstrate that the presence of autoantibodies, directed against neuronal/axonal cell surface gangliosides, in the injured mammalian peripheral nerves switch the proregenerative inflammatory environment to growth inhibitory milieu by engaging specific activating Fcγ receptors on recruited monocyte-derived macrophages to cause severe inhibition of axon regeneration. Our data demonstrate that the antibody orchestrated Fcγ receptor-mediated switch in inflammation is one mechanism underlying inhibition of axon regeneration. These findings have clinical implications for nerve repair and recovery in antibody-mediated immune neuropathies. Our results add to the complexity of axon regeneration in injured peripheral and central nervous systems as adverse effects of B cells and autoantibodies on neural injury and repair are increasingly recognized.

  10. TLR4 induces CREB-mediated IL-6 production via upregulation of F-spondin to promote vascular smooth muscle cell migration

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    Lee, Guan-Lin [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Wu, Jing-Yiing [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Yeh, Chang-Ching [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Kuo, Cheng-Chin, E-mail: kuocc@nhri.org.tw [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan (China)

    2016-05-13

    Toll-like receptor 4 (TLR4) is important in promoting inflammation and vascular smooth muscle cell (VSMC) migration, both of which contribute to atherosclerosis development and progression. But the mechanism underlying the regulation of TLR4 in VSMC migration remains unclear. Stimulation of VSMCs with LPS increased the cellular level of F-spondin which is associated with the regulation of proinflammatory cytokine production. The LPS-induced F-spondin expression depended on TLR4-mediated PI3K/Akt pathway. Suppression of F-spondin level by siRNA inhibited not only F-spondin expression but also LPS-induced phosphorylation of cAMP response element binding protein (CREB) and IL-6 expression, VSMC migration and proliferation as well as MMP9 expression. Moreover, suppression of CREB level by siRNA inhibited TLR4-induced IL-6 production and VSMC migration. Inhibition of F-spondin siRNA on LPS-induced migration was restored by addition of exogenous recombinant mouse IL-6. We conclude that upon ligand binding, TLR4 activates PI3K/Akt signaling to induce F-spondin expression, subsequently control CREB-mediated IL-6 production to promote VSMC migration. These findings provide vital insights into the essential role of F-spondin in VSMC function and will be valuable for developing new therapeutic strategies against atherosclerosis. -- Highlights: •LPS-induced F-spondin expression of VSMCs is via a TLR4/PI3K/Akt signaling. •F-spondin is pivotal for LPS-induced CREB-mediated IL-6 production. •F-spondin is required for LPS-induced VSMC migration and proliferation.

  11. XB130 translocation to microfilamentous structures mediates NNK-induced migration of human bronchial epithelial cells.

    Science.gov (United States)

    Wu, Qifei; Nadesalingam, Jeya; Moodley, Serisha; Bai, Xiaohui; Liu, Mingyao

    2015-07-20

    Cigarette smoking contributes to the pathogenesis of chronic obstructive pulmonary disease and lung cancer. Nicotine-derived nitrosamine ketone (NNK) is the most potent carcinogen among cigarette smoking components, and is known to enhance migration of cancer cells. However, the effect of NNK on normal human bronchial epithelial cells is not well studied. XB130 is a member of actin filament associated protein family and is involved in cell morphology changes, cytoskeletal rearrangement and outgrowth formation, as well as cell migration. We hypothesized that XB130 mediates NNK-induced migration of normal human bronchial epithelial cells. Our results showed that, after NNK stimulation, XB130 was translocated to the cell periphery and enriched in cell motility-associated structures, such as lamellipodia, in normal human bronchial epithelial BEAS2B cells. Moreover, overexpression of XB130 significantly enhanced NNK-induced migration, which requires both the N- and C-termini of XB130. Overexpression of XB130 enhanced NNK-induced protein tyrosine phosphorylation and promoted matrix metalloproteinase-14 translocation to cell motility-associated cellular structures after NNK stimulation. XB130-mediated NNK-induced cell migration may contribute to airway epithelial repair; however, it may also be involved in cigarette smoking-related chronic obstructive pulmonary disease and lung cancer.

  12. Bisphenol A triggers proliferation and migration of laryngeal squamous cell carcinoma via GPER mediated upregulation of IL-6.

    Science.gov (United States)

    Li, Shisheng; Wang, Bin; Tang, Qinglai; Liu, Jiajia; Yang, Xinming

    2017-06-01

    Bisphenol A (BPA) can be accumulated into the human body via food intake and inhalation. Numerous studies indicated that BPA can trigger the tumorigenesis and progression of cancer cells. Laryngeal cancer cells can be exposed to BPA directly via food digestion, while there were very limited data concerning the effect of BPA on the development of laryngeal squamous cell carcinoma (LSCC). Our present study revealed that nanomolar BPA can trigger the proliferation of LSCC cells. Bisphenol A also increased the in vitro migration and invasion of LSCC cells and upregulated the expression of matrix metallopeptidase 2. Among various chemokines tested, the expression of IL-6 was significantly increased in LSCC cells treated with BPA for 24 hours. Neutralization antibody of IL-6 or si-IL-6 can attenuate BPA-induced proliferation and migration of LSCC cells. Targeted inhibition of G protein-coupled estrogen receptor, while not estrogen receptor (ERα), abolished BPA-induced IL-6 expression, proliferation, and migration of LSCC cells. The increased IL-6 can further activate its downstream signal molecule STAT3, which was evidenced by the results of increased phosphorylation and nuclear translocation of STAT3, while si-IL-6 and si-GPER can both reverse BPA-induced activation of STAT3. Collectively, our present study revealed that BPA can trigger the progression of LSCC via GPER-mediated upregulation of IL-6. Therefore, more attention should be paid for the BPA exposure on the development of laryngeal cancer. Copyright © 2017 John Wiley & Sons, Ltd.

  13. Hypoxia Regulates mTORC1-Mediated Keratinocyte Motility and Migration via the AMPK Pathway.

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    Tiantian Yan

    Full Text Available Keratinocyte migration, the initial event and rate-limiting step in wound healing, plays a vital role in restoration of the intact skin barrier, also known as re-epithelialization. After acute tissue injury, hypoxic microenvironment gradually develops and acts as an early stimulus to initiate the healing process. Although we have previously found that hypoxia induces keratinocyte migration, the underlying mechanism remains unknown. Here, we first observed that hypoxia increased mTORC1 activity. Recombinant lentivirus vector and Rapamycin were used for silencing mTORC1 in HaCaT cells and primary mouse keratinocytes (MKs. Using cell migration assay and a Zeiss chamber equipped with imaging system, we also demonstrated that mTORC1 downregulation reversed hypoxia-induced keratinocyte motility and lateral migration. Importantly, hypoxia-activated mTORC1 was accompanied by the AMPK downregulation, and we found that the AMPK pathway activators Metformin (Met and 5-Aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR decreased the mTORC1 activity, cell motility and lateral migration. Thus, our results suggest that hypoxia regulates mTORC1-mediated keratinocyte motility and migration via the AMPK pathway.

  14. Raf Kinase Inhibitory Protein protects cells against locostatin-mediated inhibition of migration.

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    Anne N Shemon

    2009-06-01

    Full Text Available Raf Kinase Inhibitory Protein (RKIP, also PEBP1, a member of the Phosphatidylethanolamine Binding Protein family, negatively regulates growth factor signaling by the Raf/MAP kinase pathway. Since an organic compound, locostatin, was reported to bind RKIP and inhibit cell migration by a Raf-dependent mechanism, we addressed the role of RKIP in locostatin function.We analyzed locostatin interaction with RKIP and examined the biological consequences of locostatin binding on RKIP function. NMR studies show that a locostatin precursor binds to the conserved phosphatidylethanolamine binding pocket of RKIP. However, drug binding to the pocket does not prevent RKIP association with its inhibitory target, Raf-1, nor affect RKIP phosphorylation by Protein Kinase C at a regulatory site. Similarly, exposure of wild type, RKIP-depleted HeLa cells or RKIP-deficient (RKIP(-/- mouse embryonic fibroblasts (MEFs to locostatin has no effect on MAP kinase activation. Locostatin treatment of wild type MEFs causes inhibition of cell migration following wounding. RKIP deficiency impairs migration further, indicating that RKIP protects cells against locostatin-mediated inhibition of migration. Locostatin treatment of depleted or RKIP(-/- MEFs reveals cytoskeletal disruption and microtubule abnormalities in the spindle.These results suggest that locostatin's effects on cytoskeletal structure and migration are caused through mechanisms independent of its binding to RKIP and Raf/MAP kinase signaling. The protective effect of RKIP against drug inhibition of migration suggests a new role for RKIP in potentially sequestering toxic compounds that may have deleterious effects on cells.

  15. TRPV2 mediates adrenomedullin stimulation of prostate and urothelial cancer cell adhesion, migration and invasion.

    Directory of Open Access Journals (Sweden)

    Agathe Oulidi

    Full Text Available Adrenomedullin (AM is a 52-amino acid peptide initially isolated from human pheochromocytoma. AM is expressed in a variety of malignant tissues and cancer cell lines and was shown to be a mitogenic factor capable of stimulating growth of several cancer cell types. In addition, AM is a survival factor for certain cancer cells. Some data suggest that AM might be involved in the progression cancer metastasis via angiogenesis and cell migration and invasion control. The Transient Receptor Potential channel TRPV2 is known to promote in prostate cancer cell migration and invasive phenotype and is correlated with the stage and grade of bladder cancer. In this work we show that AM induces prostate and urothelial cancer cell migration and invasion through TRPV2 translocation to plasma membrane and the subsequent increase in resting calcium level.

  16. Low CCR7-mediated migration of human monocyte derived dendritic cells in response to human respiratory syncytial virus and human metapneumovirus.

    Directory of Open Access Journals (Sweden)

    Cyril Le Nouën

    2011-06-01

    Full Text Available Human respiratory syncytial virus (HRSV and, to a lesser extent, human metapneumovirus (HMPV and human parainfluenza virus type 3 (HPIV3, can re-infect symptomatically throughout life without significant antigenic change, suggestive of incomplete or short-lived immunity. In contrast, re-infection by influenza A virus (IAV largely depends on antigenic change, suggestive of more complete immunity. Antigen presentation by dendritic cells (DC is critical in initiating the adaptive immune response. Antigen uptake by DC induces maturational changes that include decreased expression of the chemokine receptors CCR1, CCR2, and CCR5 that maintain DC residence in peripheral tissues, and increased expression of CCR7 that mediates the migration of antigen-bearing DC to lymphatic tissue. We stimulated human monocyte-derived DC (MDDC with virus and found that, in contrast to HPIV3 and IAV, HMPV and HRSV did not efficiently decrease CCR1, 2, and 5 expression, and did not efficiently increase CCR7 expression. Consistent with the differences in CCR7 mRNA and protein expression, MDDC stimulated with HRSV or HMPV migrated less efficiently to the CCR7 ligand CCL19 than did IAV-stimulated MDDC. Using GFP-expressing recombinant virus, we showed that the subpopulation of MDDC that was robustly infected with HRSV was particularly inefficient in chemokine receptor modulation. HMPV- or HRSV-stimulated MDDC responded to secondary stimulation with bacterial lipopolysaccharide or with a cocktail of proinflammatory cytokines by increasing CCR7 and decreasing CCR1, 2 and 5 expression, and by more efficient migration to CCL19, suggesting that HMPV and HRSV suboptimally stimulate rather than irreversibly inhibit MDDC migration. This also suggests that the low concentration of proinflammatory cytokines released from HRSV- and HMPV-stimulated MDDC is partly responsible for the low CCR7-mediated migration. We propose that inefficient migration of HRSV- and HMPV-stimulated DC to

  17. Receptor-Mediated Transport of Insulin across Endothelial Cells

    Science.gov (United States)

    King, George L.; Johnson, Sandra M.

    1985-03-01

    Hormones such as insulin are transported from the interior to the exterior of blood vessels. Whether endothelial cells, which line the inner walls of blood vessels have a role in this transport of hormones is not clear, but it is known that endothelial cells can internalize and release insulin rapidly with little degradation. The transport of iodine-125-labeled insulin was measured directly through the use of dual chambers separated by a horizontal monolayer of cultured bovine aortic endothelial cells. In this setting, endothelial cells took up and released the labeled insulin, thereby transporting it across the cells. The transport of insulin across the endothelial cells was temperature sensitive and was inhibited by unlabeled insulin and by antibody to insulin receptor in proportion to the ability of these substances to inhibit insulin binding to its receptor. More than 80 percent of the transported insulin was intact. These data suggest that insulin is rapidly transported across endothelial cells by a receptor-mediated process.

  18. Purinergic Receptors: Key Mediators of HIV-1 infection and inflammation

    Directory of Open Access Journals (Sweden)

    Talia H Swartz

    2015-11-01

    Full Text Available Human immunodeficiency virus (HIV-1 causes a chronic infection that afflicts more than 38 million individuals worldwide. While the infection can be suppressed with potent anti-retroviral therapies, individuals infected with HIV have elevated levels of inflammation as indicated by increased T cell activation, soluble biomarkers, and associated morbidity and mortality. A single mechanism linking HIV pathogenesis to this inflammation has yet to be identified. Purinergic receptors are known to mediate inflammation and have been shown to be required for HIV-1 infection at the level of HIV-1 membrane fusion. Here we review the literature on the role of purinergic receptors in HIV-1 infection and associated inflammation and describe a role for these receptors as potential therapeutic targets.

  19. Receptor-mediated endocytosis of insect lipoprotein : insight into LDL receptor functioning

    NARCIS (Netherlands)

    Hoof, Dennis van

    2004-01-01

    The extracellular transport of water-insoluble lipids through the aqueous circulatory system of animals is mediated by lipoproteins. The lipoprotein of insects, lipophorin (Lp), is homologous to that of mammalian low-density lipoprotein (LDL). Moreover, an endocytic receptor for Lp has been

  20. Notch activation is dispensable for D, L-sulforaphane-mediated inhibition of human prostate cancer cell migration.

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    Eun-Ryeong Hahm

    Full Text Available D, L-Sulforaphane (SFN, a synthetic racemic analog of broccoli constituent L-sulforaphane, is a highly promising cancer chemopreventive agent with in vivo efficacy against chemically-induced as well as oncogene-driven cancer in preclinical rodent models. Cancer chemopreventive effect of SFN is characterized by G(2/M phase cell cycle arrest, apoptosis induction, and inhibition of cell migration and invasion. Moreover, SFN inhibits multiple oncogenic signaling pathways often hyperactive in human cancers, including nuclear factor-κB, Akt, signal transducer and activator of transcription 3, and androgen receptor. The present study was designed to determine the role of Notch signaling, which is constitutively active in many human cancers, in anticancer effects of SFN using prostate cancer cells as a model. Exposure of human prostate cancer cells (PC-3, LNCaP, and/or LNCaP-C4-2B to SFN as well as its naturally-occurring thio-, sulfinyl-, and sulfonyl-analogs resulted in cleavage (activation of Notch1, Notch2, and Notch4, which was accompanied by a decrease in levels of full-length Notch forms especially at the 16- and 24-hour time points. The SFN-mediated cleavage of Notch isoforms was associated with its transcriptional activation as evidenced by RBP-Jk-, HES-1A/B- and HEY-1 luciferase reporter assays. Migration of PC-3 and LNCaP cells was decreased significantly by RNA interference of Notch1 and Notch2, but not Notch4. Furthermore, SFN-mediated inhibition of PC-3 and LNCaP cell migration was only marginally affected by knockdown of Notch1 and Notch2. Strikingly, SFN administration to Transgenic Adenocarcinoma of Mouse Prostate transgenic mice failed to increase levels of cleaved Notch1, cleaved Notch2, and HES-1 proteins in vivo in prostatic intraepithelial neoplasia, well-differentiated carcinoma or poorly-differentiated prostate cancer lesions. These results indicate that Notch activation is largely dispensable for SFN-mediated inhibition of cell

  1. RNase L Suppresses Androgen Receptor Signaling, Cell Migration and Matrix Metalloproteinase Activity in Prostate Cancer Cells

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    Shubham Dayal

    2017-03-01

    Full Text Available The interferon antiviral pathways and prostate cancer genetics converge on a regulated endoribonuclease, RNase L. Positional cloning and linkage studies mapped Hereditary Prostate Cancer 1 (HPC1 to RNASEL. To date, there is no correlation of viral infections with prostate cancer, suggesting that RNase L may play additional roles in tumor suppression. Here, we demonstrate a role of RNase L as a suppressor of androgen receptor (AR signaling, cell migration and matrix metalloproteinase activity. Using RNase L mutants, we show that its nucleolytic activity is dispensable for both AR signaling and migration. The most prevalent HPC1-associated mutations in RNase L, R462Q and E265X, enhance AR signaling and cell migration. RNase L negatively regulates cell migration and attachment on various extracellular matrices. We demonstrate that RNase L knockdown cells promote increased cell surface expression of integrin β1 which activates Focal Adhesion Kinase-Sarcoma (FAK-Src pathway and Ras-related C3 botulinum toxin substrate 1-guanosine triphosphatase (Rac1-GTPase activity to increase cell migration. Activity of matrix metalloproteinase (MMP-2 and -9 is significantly increased in cells where RNase L levels are ablated. We show that mutations in RNase L found in HPC patients may promote prostate cancer by increasing expression of AR-responsive genes and cell motility and identify novel roles of RNase L as a prostate cancer susceptibility gene.

  2. RNase L Suppresses Androgen Receptor Signaling, Cell Migration and Matrix Metalloproteinase Activity in Prostate Cancer Cells

    Science.gov (United States)

    Dayal, Shubham; Zhou, Jun; Manivannan, Praveen; Siddiqui, Mohammad Adnan; Ahmad, Omaima Farid; Clark, Matthew; Awadia, Sahezeel; Garcia-Mata, Rafael; Shemshedini, Lirim; Malathi, Krishnamurthy

    2017-01-01

    The interferon antiviral pathways and prostate cancer genetics converge on a regulated endoribonuclease, RNase L. Positional cloning and linkage studies mapped Hereditary Prostate Cancer 1 (HPC1) to RNASEL. To date, there is no correlation of viral infections with prostate cancer, suggesting that RNase L may play additional roles in tumor suppression. Here, we demonstrate a role of RNase L as a suppressor of androgen receptor (AR) signaling, cell migration and matrix metalloproteinase activity. Using RNase L mutants, we show that its nucleolytic activity is dispensable for both AR signaling and migration. The most prevalent HPC1-associated mutations in RNase L, R462Q and E265X, enhance AR signaling and cell migration. RNase L negatively regulates cell migration and attachment on various extracellular matrices. We demonstrate that RNase L knockdown cells promote increased cell surface expression of integrin β1 which activates Focal Adhesion Kinase-Sarcoma (FAK-Src) pathway and Ras-related C3 botulinum toxin substrate 1-guanosine triphosphatase (Rac1-GTPase) activity to increase cell migration. Activity of matrix metalloproteinase (MMP)-2 and -9 is significantly increased in cells where RNase L levels are ablated. We show that mutations in RNase L found in HPC patients may promote prostate cancer by increasing expression of AR-responsive genes and cell motility and identify novel roles of RNase L as a prostate cancer susceptibility gene. PMID:28257035

  3. MMP13 mediates cell cycle progression in melanocytes and melanoma cells: in vitro studies of migration and proliferation.

    NARCIS (Netherlands)

    Meierjohann, S.; Hufnagel, A.; Wende, E.; Kleinschmidt, M.A.; Wolf, K. van der; Friedl, P.H.A.; Gaubatz, S.; Schartl, M.

    2010-01-01

    BACKGROUND: Melanoma cells are usually characterized by a strong proliferative potential and efficient invasive migration. Among the multiple molecular changes that are recorded during progression of this disease, aberrant activation of receptor tyrosine kinases (RTK) is often observed. Activation

  4. Regulatory T Cell Migration Is Dependent on Glucokinase-Mediated Glycolysis.

    Science.gov (United States)

    Kishore, Madhav; Cheung, Kenneth C P; Fu, Hongmei; Bonacina, Fabrizia; Wang, Guosu; Coe, David; Ward, Eleanor J; Colamatteo, Alessandra; Jangani, Maryam; Baragetti, Andrea; Matarese, Giuseppe; Smith, David M; Haas, Robert; Mauro, Claudio; Wraith, David C; Okkenhaug, Klaus; Catapano, Alberico L; De Rosa, Veronica; Norata, Giuseppe D; Marelli-Berg, Federica M

    2017-11-21

    Migration of activated regulatory T (Treg) cells to inflamed tissue is crucial for their immune-modulatory function. While metabolic reprogramming during Treg cell differentiation has been extensively studied, the bioenergetics of Treg cell trafficking remains undefined. We have investigated the metabolic demands of migrating Treg cells in vitro and in vivo. We show that glycolysis was instrumental for their migration and was initiated by pro-migratory stimuli via a PI3K-mTORC2-mediated pathway culminating in induction of the enzyme glucokinase (GCK). Subsequently, GCK promoted cytoskeletal rearrangements by associating with actin. Treg cells lacking this pathway were functionally suppressive but failed to migrate to skin allografts and inhibit rejection. Similarly, human carriers of a loss-of-function GCK regulatory protein gene-leading to increased GCK activity-had reduced numbers of circulating Treg cells. These cells displayed enhanced migratory activity but similar suppressive function, while conventional T cells were unaffected. Thus, GCK-dependent glycolysis regulates Treg cell migration. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  5. Role of the receptor Mas in macrophage-mediated inflammation in vivo.

    Science.gov (United States)

    Hammer, Anna; Yang, Guang; Friedrich, Juliane; Kovacs, Agnes; Lee, De-Hyung; Grave, Katharina; Jörg, Stefanie; Alenina, Natalia; Grosch, Janina; Winkler, Jürgen; Gold, Ralf; Bader, Michael; Manzel, Arndt; Rump, Lars C; Müller, Dominik N; Linker, Ralf A; Stegbauer, Johannes

    2016-12-06

    Recently, an alternative renin-angiotensin system pathway has been described, which involves binding of angiotensin-(1-7) to its receptor Mas. The Mas axis may counterbalance angiotensin-II-mediated proinflammatory effects, likely by affecting macrophage function. Here we investigate the role of Mas in murine models of autoimmune neuroinflammation and atherosclerosis, which both involve macrophage-driven pathomechanisms. Mas signaling affected macrophage polarization, migration, and macrophage-mediated T-cell activation. Mas deficiency exacerbated the course of experimental autoimmune encephalomyelitis and increased macrophage infiltration as well as proinflammatory gene expression in the spleen and spinal cord. Furthermore, Mas deficiency promoted atherosclerosis by affecting macrophage infiltration and migration and led to increased oxidative stress as well as impaired endothelial function in ApoE-deficient mice. In summary, we identified the Mas axis as an important factor in macrophage function during inflammation of the central nervous and vascular system in vivo. Modulating the Mas axis may constitute an interesting therapeutic target in multiple sclerosis and/or atherosclerosis.

  6. Estrogen Receptor α Is Crucial in Zearalenone-Induced Invasion and Migration of Prostate Cancer Cells

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    Karolina Kowalska

    2018-02-01

    Full Text Available Zearalenone (ZEA, a mycotoxin produced in the genus Fusarium, binds to estrogen receptors (ER and is therefore regarded as an endocrine disruptor. ZEA has also been found to modulate the proliferation and apoptosis of prostate cancer cells in a dose-dependent manner. This study evaluates whether the effect of a low dose of ZEA (0.1 and 0.001 nM on the invasion and migration of prostate cancer cell line PC3 is associated with ERs expression. The invasion and migration was evaluated by modified Boyden chamber assay, scratch assay, gelatin zymography, Real Time qPCR (RTqPCR and Western blot. The involvement of ERs was evaluated with the selective ER antagonists: estrogen receptor α (ERα antagonist 1,3-bis (4-hydroxyphenyl-4-methyl-5-[4-(2-piperidinylethoxy phenol]-1H-pyrazole dihydrochloride (MPP and estrogen receptor β (ERβ antagonist 4-[2–phenyl-5,7–bis (trifluoromethyl pyrazolo [1,5-a]-pyrimidin-3-yl] phenol (PHTPP. ZEA was found to modulate cell motility dependent on estrogen receptors, particularly ERα. Increased cell migration and invasion were associated with increased MMP-2 and MMP-9 activity as well as the up-regulation of the EMT-associated genes vimentin (VIM, zinc finger E-box-binding homeobox 1/2 (ZEB1/2 and transforming growth factor β 1 (TGFβ1. In conclusion, ZEA might modulate the invasiveness of prostate cancer cells dependently on ERα expression.

  7. N-methyl D-aspartate receptor synaptonuclear signaling and neuronal migration factor (Nsmf) plays a novel role in myoblast proliferation.

    Science.gov (United States)

    Moon, Hyo Youl

    2015-01-01

    Myogenesis, the formation and regeneration of muscular tissue, is a fundamental factor in embryonic development. N-methyl D-aspartate (NMDA) receptor synaptonuclear signaling and neuronal migration factor (Nsmf) mediates NMDA receptor endocytosis in GnRH neuronal cells. NMDA receptor is involved in myoblast differentiation by regulating Ca2 (+) dependent fusion of myocytes. In this study, we investigated the role of Nsmf in myoblast proliferation and differentiation. Quantitative-PCR, immunoblotting, and immunohistochemistry results showed that the Nsmf expression levels increased during both the differentiation and proliferation of myocytes. Knockdown of Nsmf in myocytes by siRNA did not affect the myocyte differentiation marker myogenin. However, flow cytometry showed that the proliferation rate of the Nsmf-knockdown cells was reduced compared to the control cells. Therefore, our results indicate that Nsmf is a novel myogenic factor that can enhance myoblast proliferation. Furthermore, Nsmf may be an important therapeutic target in diseases associated with aging, muscular dystrophy, or cachexia.

  8. ADAM10 controls collagen signaling and cell migration on collagen by shedding the ectodomain of discoidin domain receptor 1 (DDR1)

    Science.gov (United States)

    Shitomi, Yasuyuki; Thøgersen, Ida B.; Ito, Noriko; Leitinger, Birgit; Enghild, Jan J.; Itoh, Yoshifumi

    2015-01-01

    Discoidin domain receptor 1 (DDR1) is a receptor tyrosine kinase that binds and transmits signals from various collagens in epithelial cells. However, how DDR1–dependent signaling is regulated has not been understood. Here we report that collagen binding induces ADAM10-dependent ectodomain shedding of DDR1. DDR1 shedding is not a result of an activation of its signaling pathway, since DDR1 mutants defective in signaling were shed in an efficient manner. DDR1 and ADAM10 were found to be in a complex on the cell surface, but shedding did not occur unless collagen bound to DDR1. Using a shedding-resistant DDR1 mutant, we found that ADAM10-dependent DDR1 shedding regulates the half-life of collagen-induced phosphorylation of the receptor. Our data also revealed that ADAM10 plays an important role in regulating DDR1-mediated cell adhesion to achieve efficient cell migration on collagen matrices. PMID:25540428

  9. The inflammatory mediator leukotriene D{sub 4} induces subcellular β-catenin translocation and migration of colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Salim, Tavga [Division of Cell and Experimental Pathology, Department of Laboratory Medicine, Lund University, Clinical Research Center, Skåne University Hospital, Malmö (Sweden); Sand-Dejmek, Janna [Division of Cell and Experimental Pathology, Department of Laboratory Medicine, Lund University, Clinical Research Center, Skåne University Hospital, Malmö (Sweden); Section of Surgery, Department of Clinical Sciences, Lund University, Skåne University Hospital, Malmö (Sweden); Bayer HealthCare, Pharmaceuticals Medical Affairs, Solna (Sweden); Sjölander, Anita, E-mail: anita.sjolander@med.lu.se [Division of Cell and Experimental Pathology, Department of Laboratory Medicine, Lund University, Clinical Research Center, Skåne University Hospital, Malmö (Sweden)

    2014-02-15

    The abnormal activation of the Wnt/β-catenin pathway frequently occurs in colorectal cancer. The nuclear translocation of β-catenin activates the transcription of target genes that promote cell proliferation, survival, and invasion. The pro-inflammatory mediator leukotriene D{sub 4} (LTD{sub 4}) exerts its effects through the CysLT{sub 1} receptor. We previously reported an upregulation of CysLT{sub 1}R in patients with colon cancer, suggesting the importance of leukotrienes in colon cancer. The aim of this study was to investigate the impact of LTD{sub 4} on Wnt/β-catenin signaling and its effects on proliferation and migration of colon cancer cells. LTD{sub 4} stimulation led to an increase in β-catenin expression, β-catenin nuclear translocation and the subsequent transcription of MYC and CCND1. Furthermore, LTD{sub 4} significantly reduced the expression of E-cadherin and β-catenin at the plasma membrane and increased the migration and proliferation of HCT116 colon cancer cells. The effects of LTD{sub 4} can be blocked by the inhibition of CysLT{sub 1}R. Furthermore, LTD{sub 4} induced the inhibition of glycogen synthase kinase 3 (GSK)-3β activity, indicating a crosstalk between the G-protein-coupled receptor CysLT{sub 1} and the Wnt/β-catenin pathway. In conclusion, LTD{sub 4}, which can be secreted from macrophages and leukocytes in the tumor microenvironment, induces β-catenin translocation and the activation of β-catenin target genes, resulting in the increased proliferation and migration of colon cancer cells. - Highlights: • Leukotriene D{sub 4} (LTD{sub 4}) lowers membrane β-catenin but increases nuclear β-catenin levels in colon cancer cells. • In agreement, LTD{sub 4} triggers inactivation of GSK-3β, activation of TCF/LEF and increased expression of Cyclin D1 and c-Myc. • LTD{sub 4} also caused a significant reduction in the expression of E-cadherin and an increased migration of colon cancer cells.

  10. NMDA receptors mediate calcium accumulation in myelin during chemical ischaemia.

    Science.gov (United States)

    Micu, I; Jiang, Q; Coderre, E; Ridsdale, A; Zhang, L; Woulfe, J; Yin, X; Trapp, B D; McRory, J E; Rehak, R; Zamponi, G W; Wang, W; Stys, P K

    2006-02-23

    Central nervous system myelin is a specialized structure produced by oligodendrocytes that ensheaths axons, allowing rapid and efficient saltatory conduction of action potentials. Many disorders promote damage to and eventual loss of the myelin sheath, which often results in significant neurological morbidity. However, little is known about the fundamental mechanisms that initiate myelin damage, with the assumption being that its fate follows that of the parent oligodendrocyte. Here we show that NMDA (N-methyl-d-aspartate) glutamate receptors mediate Ca2+ accumulation in central myelin in response to chemical ischaemia in vitro. Using two-photon microscopy, we imaged fluorescence of the Ca2+ indicator X-rhod-1 loaded into oligodendrocytes and the cytoplasmic compartment of the myelin sheath in adult rat optic nerves. The AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid)/kainate receptor antagonist NBQX completely blocked the ischaemic Ca2+ increase in oligodendroglial cell bodies, but only modestly reduced the Ca2+ increase in myelin. In contrast, the Ca2+ increase in myelin was abolished by broad-spectrum NMDA receptor antagonists (MK-801, 7-chlorokynurenic acid, d-AP5), but not by more selective blockers of NR2A and NR2B subunit-containing receptors (NVP-AAM077 and ifenprodil). In vitro ischaemia causes ultrastructural damage to both axon cylinders and myelin. NMDA receptor antagonism greatly reduced the damage to myelin. NR1, NR2 and NR3 subunits were detected in myelin by immunohistochemistry and immunoprecipitation, indicating that all necessary subunits are present for the formation of functional NMDA receptors. Our data show that the mature myelin sheath can respond independently to injurious stimuli. Given that axons are known to release glutamate, our finding that the Ca2+ increase was mediated in large part by activation of myelinic NMDA receptors suggests a new mechanism of axo-myelinic signalling. Such a mechanism may represent a

  11. Moesin and merlin regulate urokinase receptor-dependent endothelial cell migration, adhesion and angiogenesis.

    Science.gov (United States)

    Degryse, Bernard; Britto, Mishan; Shan, Chun Xu; Wallace, Robert G; Rochfort, Keith D; Cummins, Philip M; Meade, Gerardene; Murphy, Ronan P

    2017-07-01

    The glycosyl-phosphatidyl-inositol (GPI)-anchored urokinase receptor (uPAR) has no intracellular domain, but nevertheless initiates signalling through proximal interactions with other membrane receptors including integrins. The relationships between uPAR and ezrin/radixin/moesin (ERM) proteins, moesin and merlin have never been explored. Moesin and merlin are versatile membrane-actin links and regulators of receptors signalling, respectively. We show that uPAR controls moesin and merlin, which propagate uPAR-initiated signals and modulate integrin functions, thereby regulating uPAR activity. uPAR rapidly de-phosphorylates moesin and phosphorylates merlin inactivating both proteins, and enhancing cell migration and angiogenesis. Moesin behaves as a molecular switch turning either on or off uPAR signalling through cycles of de-activation/activation, or sustained activation, respectively. Furthermore, moesin is at the crossroads of uPAR-initiated outside-in and inside-out signalling promoting integrin-dependent cell adhesion suggesting that uPAR also activates integrins distally through moesin. Knocking down merlin expression enhanced cell migration and adhesion through different regulation of fibronectin- and vitronectin-binding integrins. Copyright © 2017. Published by Elsevier Ltd.

  12. DMPD: Modulation of Toll-interleukin 1 receptor mediated signaling. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15662540 Modulation of Toll-interleukin 1 receptor mediated signaling. Li X, Qin J.... J Mol Med. 2005 Apr;83(4):258-66. Epub 2005 Jan 21. (.png) (.svg) (.html) (.csml) Show Modulation of Toll-interleukin... 1 receptor mediated signaling. PubmedID 15662540 Title Modulation of Toll-interleukin 1 receptor

  13. The Cannabinoid Delta-9-tetrahydrocannabinol Mediates Inhibition of Macrophage Chemotaxis to RANTES/CCL5 through the CB2 Receptor

    Science.gov (United States)

    Raborn, Erinn S.; Marciano-Cabral, Francine; Buckley, Nancy E.; Martin, Billy R.; Cabral, Guy A.

    2009-01-01

    The chemotactic response of murine peritoneal macrophages to RANTES/CCL5 was inhibited significantly following pretreatment with delta-9-tetrahydrocannabinol (THC), the major psychoactive component in marijuana. Significant inhibition of this chemokine directed migratory response was obtained also when the full cannabinoid agonist CP55940 was used. The CB2 receptor-selective ligand O-2137 exerted a robust inhibition of chemotaxis while the CB1 receptor-selective ligand ACEA had a minimal effect. The THC-mediated inhibition was reversed by the CB2 receptor-specific antagonist SR144528 but not by the CB1 receptor-specific antagonist SR141716A. In addition, THC treatment had a minimal effect on the chemotactic response of peritoneal macrophages from CB2 knockout mice. Collectively, these results suggest that cannabinoids act through the CB2 receptor to trans-deactivate migratory responsiveness to RANTES/CCL5. Furthermore, the results suggest that the CB2 receptor may be a constituent element of a network of G protein-coupled receptor signal transductional systems, inclusive of chemokine receptors, that act coordinately to modulate macrophage migration. PMID:18247131

  14. Bombesin receptor-mediated imaging and cytotoxicity: review and current status

    Science.gov (United States)

    Sancho, Veronica; Di Florio, Alessia; Moody, Terry W.; Jensen, Robert T.

    2010-01-01

    The three mammalian bombesin (Bn) receptors (gastrin-releasing peptide [GRP] receptor, neuromedin B [NMB] receptor, BRS-3) are one of the classes of G protein-coupled receptors that are most frequently over-express/ectopically expressed by common, important malignancies. Because of the clinical success of somatostatin receptor-mediated imaging and cytotoxicity with neuroendocrine tumors, there is now increasing interest in pursuing a similar approach with Bn receptors. In the last few years then have been more than 200 studies in this area. In the present paper, the in vitro and in vivo results, as well as results of human studies from many of these studies are reviewed and the current state of Bn receptor-mediated imaging or cytotoxicity is discussed. Both Bn receptor-mediated imaging studies as well as Bn receptor-mediated tumoral cytotoxic studies using radioactive and non-radioactive Bn-based ligands are covered. PMID:21034419

  15. Receptor-Mediated Endocytosis and Brain Delivery of Therapeutic Biologics

    Directory of Open Access Journals (Sweden)

    Guangqing Xiao

    2013-01-01

    Full Text Available Transport of macromolecules across the blood-brain-barrier (BBB requires both specific and nonspecific interactions between macromolecules and proteins/receptors expressed on the luminal and/or the abluminal surfaces of the brain capillary endothelial cells. Endocytosis and transcytosis play important roles in the distribution of macromolecules. Due to the tight junction of BBB, brain delivery of traditional therapeutic proteins with large molecular weight is generally not possible. There are multiple pathways through which macromolecules can be taken up into cells through both specific and nonspecific interactions with proteins/receptors on the cell surface. This review is focused on the current knowledge of receptor-mediated endocytosis/transcytosis and brain delivery using the Angiopep-2-conjugated system and the molecular Trojan horses. In addition, the role of neonatal Fc receptor (FcRn in regulating the efflux of Immunoglobulin G (IgG from brain to blood, and approaches to improve the pharmacokinetics of therapeutic biologics by generating Fc fusion proteins, and increasing the pH dependent binding affinity between Fc and FcRn, are discussed.

  16. Cyclization of the urokinase receptor-derived ser-arg-ser-arg-tyr Peptide generates a potent inhibitor of trans-endothelial migration of monocytes.

    Directory of Open Access Journals (Sweden)

    Ali Munaim Yousif

    Full Text Available The receptor for the urokinase-type plasminogen activator (uPAR is a widely recognized master regulator of cell migration and uPAR88-92 is the minimal sequence required to induce cell motility. We and others have previously documented that the uPAR88-92 sequence, even in the form of synthetic linear peptide (SRSRY, interacts with the formyl peptide receptor type 1 (FPR1, henceforth inducing cell migration of several cell lines, including monocytes. FPR1 is mainly expressed by mammalian phagocytic leukocytes and plays a crucial role in chemotaxis. In this study, we present evidence that the cyclization of the SRSRY sequence generates a new potent and stable inhibitor of monocyte trafficking. In rat basophilic leukaemia RBL-2H3/ETFR cells expressing high levels of constitutively activated FPR1, the cyclic SRSRY peptide ([SRSRY] blocks FPR1 mediated cell migration by interfering with both internalization and ligand-uptake of FPR1. Similarly to RBL-2H3/ETFR cells, [SRSRY] competes with fMLF for binding to FPR1 and prevents agonist-induced FPR1 internalization in human monocyte THP-1 cells. Unlike scramble [RSSYR], [SRSRY] inhibits fMLF-directed migration of monocytes in a dose-dependent manner, with IC50 value of 0.01 nM. PMA-differentiated THP-1 cell exposure to fMLF gradient causes a marked cytoskeletal re-organization with the formation of F-actin rich pseudopodia that are prevented by the addition of [SRSRY]. Furthermore, [SRSRY] prevents migration of human primary monocytes and trans-endothelial migration of monocytes. Our findings indicate that [SRSRY] is a new FPR1 inhibitor which may suggest the development of new drugs for treating pathological conditions sustained by increased motility of monocytes, such as chronic inflammatory diseases.

  17. miR-503 inhibits platelet-derived growth factor-induced human aortic vascular smooth muscle cell proliferation and migration through targeting the insulin receptor.

    Science.gov (United States)

    Bi, Rui; Ding, Fangbao; He, Yi; Jiang, Lianyong; Jiang, Zhaolei; Mei, Ju; Liu, Hao

    2016-12-01

    Abnormal proliferation and migration of vascular smooth muscle cells (VSMC) is a common feature of disease progression in atherosclerosis. Here, we investigated the potential role of miR-503 in platelet-derived growth factor (PDGF)-induced proliferation and migration of human aortic smooth muscle cells and the underlying mechanisms of action. miR-503 expression was significantly downregulated in a dose- and time-dependent manner following PDGF treatment. Introduction of miR-503 mimics into cultured SMCs significantly attenuated cell proliferation and migration induced by PDGF. Bioinformatics analyses revealed that the insulin receptor (INSR) is a target candidate of miR-503. miR-503 suppressed luciferase activity driven by a vector containing the 3'-untranslated region of INSR in a sequence-specific manner. Downregulation of INSR appeared critical for miR-503-mediated inhibitory effects on PDGF-induced cell proliferation and migration in human aortic SMCs. Based on the collective data, we suggest a novel role of miR-503 as a regulator of VSMC proliferation and migration through modulating INSR. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  18. Regulation and binding of pregnane X receptor by nuclear receptor corepressor silencing mediator of retinoid and thyroid hormone receptors (SMRT).

    Science.gov (United States)

    Johnson, David R; Li, Chia-Wei; Chen, Liuh-Yow; Ghosh, Jagadish C; Chen, J Don

    2006-01-01

    The pregnane X receptor (PXR) is an orphan nuclear receptor predominantly expressed in liver and intestine. PXR coordinates hepatic responses to prevent liver injury induced by environmental toxins. PXR activates cytochrome P450 3A4 gene expression upon binding to rifampicin (Rif) and clotrimazole (CTZ) by recruiting transcriptional coactivators. It remains unclear whether and how PXR regulates gene expression in the absence of ligand. In this study, we analyzed interactions between PXR and the silencing mediator of retinoid and thyroid hormone receptors (SMRT) and determined the role of SMRT in regulating PXR activity. We show that SMRT interacts with PXR in glutathione S-transferase pull-down, yeast two-hybrid, and mammalian two-hybrid assays. The interaction is mediated through the ligand-binding domain of PXR and the SMRTs' nuclear receptor-interacting domain 2. The PXR-SMRT interaction is sensitive to species-specific ligands, and Rif causes an exchange of the corepressor SMRT with the p160 coactivator known as receptor-associated coactivator 3 (RAC3). Deletion of the PXR's activation function 2 helix enhances SMRT binding and abolishes ligand-dependent dissociation of SMRT. Coexpression of PXR with SMRT results in colocalization at discrete nuclear foci. Finally, transient transfection assays show that overexpression of SMRT inhibits PXR's transactivation of the Cyp3A4 promoter, whereas silencing of SMRT enhances the reporter expression. Taken together, our results suggest that the corepressor SMRT may bind to and regulate the transcriptional activity of PXR.

  19. Monocyte scintigraphy in rheumatoid arthritis: the dynamics of monocyte migration in immune-mediated inflammatory disease.

    Directory of Open Access Journals (Sweden)

    Rogier M Thurlings

    Full Text Available BACKGROUND: Macrophages are principal drivers of synovial inflammation in rheumatoid arthritis (RA, a prototype immune-mediated inflammatory disease. Conceivably, synovial macrophages are continuously replaced by circulating monocytes in RA. Animal studies from the 1960s suggested that macrophage replacement by monocytes is a slow process in chronic inflammatory lesions. Translation of these data into the human condition has been hampered by the lack of available techniques to analyze monocyte migration in man. METHODS/PRINCIPAL FINDINGS: We developed a technique that enabled us to analyze the migration of labelled autologous monocytes in RA patients using single photon emission computer tomography (SPECT. We isolated CD14+ monocytes by CliniMACS in 8 patients and labeled these with technetium-99m (99mTc-HMPAO. Monocytes were re-infused into the same patient. Using SPECT we calculated that a very small but specific fraction of 3.4 x 10(-3 (0.95-5.1 x 10(-3 % of re-infused monocytes migrated to the inflamed joints, being detectable within one hour after re-infusion. CONCLUSIONS/SIGNIFICANCE: The results indicate monocytes migrate continuously into the inflamed synovial tissue of RA patients, but at a slow macrophage-replacement rate. This suggests that the rapid decrease in synovial macrophages that occurs after antirheumatic treatment might rather be explained by an alteration in macrophage retention than in monocyte influx and that RA might be particularly sensitive to treatments targeting inflammatory cell retention.

  20. Adenosine inhibits neutrophil vascular endothelial growth factor release and transendothelial migration via A2B receptor activation.

    LENUS (Irish Health Repository)

    Wakai, A

    2012-02-03

    The effects of adenosine on neutrophil (polymorphonuclear neutrophils; PMN)-directed changes in vascular permeability are poorly characterized. This study investigated whether adenosine modulates activated PMN vascular endothelial growth factor (vascular permeability factor; VEGF) release and transendothelial migration. PMN activated with tumour necrosis factor-alpha (TNF-alpha, 10 ng\\/mL) were incubated with adenosine and its receptor-specific analogues. Culture supernatants were assayed for VEGF. PMN transendothelial migration across human umbilical vein endothelial cell (HUVEC) monolayers was assessed in vitro. Adhesion molecule receptor expression was assessed flow cytometrically. Adenosine and some of its receptor-specific analogues dose-dependently inhibited activated PMN VEGF release. The rank order of potency was consistent with the affinity profile of human A2B receptors. The inhibitory effect of adenosine was reversed by 3,7-dimethyl-1-propargylxanthine, an A2 receptor antagonist. Adenosine (100 microM) or the A2B receptor agonist 5\\'-N-ethylcarboxamidoadenosine (NECA, 100 microM) significantly reduced PMN transendothelial migration. However, expression of activated PMN beta2 integrins and HUVEC ICAM-1 were not significantly altered by adenosine or NECA. Adenosine attenuates human PMN VEGF release and transendothelial migration via the A2B receptor. This provides a novel target for the modulation of PMN-directed vascular hyperpermeability in conditions such as the capillary leak syndrome.

  1. Toll-like receptor 9 mediates invasion and predicts prognosis in squamous cell carcinoma of the mobile tongue.

    Science.gov (United States)

    Kauppila, Joonas H; Korvala, Johanna; Siirilä, Kristiina; Manni, Marika; Mäkinen, Laura K; Hagström, Jaana; Atula, Timo; Haglund, Caj; Selander, Katri S; Saarnio, Juha; Karttunen, Tuomo J; Lehenkari, Petri P; Salo, Tuula

    2015-09-01

    Toll-like receptor 9 (TLR9) is a cellular receptor, which recognizes bacterial and host-derived DNA. Stimulation of TLR9 induces cellular invasion via matrix metalloproteinase 13 (MMP-13). The aim of this study was to evaluate the role of TLR9 in invasion of oral tongue squamous cell carcinoma (OTSCC). The effects of TLR9 ligands on oral squamous cell carcinoma cell lines were studied with invasion and migration assays, as well as in a myoma organotypic model. The TLR9 ligand, CpG-ODN, increased invasion and migration in OTSCC lines. These effects were reduced by TLR9 siRNA or inhibition with TLR9 antibodies. Immunohistochemical analysis of tissues from 195 patients with OTSCC revealed that TLR9 was expressed in 181/195 carcinomas. The expression of TLR9 was higher in the malignant cells than in the normal epithelium. High TLR9 expression was associated with high MMP-13 expression and poor differentiation. High TLR9 expression was also identified as an independent predictor of poor prognosis (HR 1.810, 95% CI [1.053-3.112]). Toll-like receptor 9 mediates OTSCC invasion and migration in vitro and is an independent prognostic factor of OTSCC. Inhibition of TLR9 may be a novel therapeutic opportunity in oral cancer. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  2. DJ-1 protein regulates CD3+ T cell migration via overexpression of CXCR4 receptor.

    Science.gov (United States)

    Jung, Seung Hyo; Won, Kyung Jong; Lee, Kang Pa; Lee, Dong Hyun; Yu, Suyeol; Lee, Dong-Youb; Seo, Eun-Hye; Kang, Hyun; Park, Eun-Seok; Kim, Hyun-Joong; Lee, Seung Hyun; Kim, Bokyung

    2014-08-01

    DJ-1-a multifunctional protein responding to oxidative stress-is a possible regulator of the inflammatory response that plays an important role in atherosclerosis. Stromal cell-derived factor (SDF)-1 and its receptor, chemokine receptor type 4 (CXCR4), have been implicated in the recruitment of inflammatory cells during atherosclerosis. Here we investigated the hypothesis that DJ-1 protein might participate in CD3+ T cell functions in response to SDF-1 and contribute to the pathogenesis of atherosclerosis. SDF-1 stimulated migration in mouse CD3+ T cells in a dose-dependent manner. SDF-1 also elevated the phosphorylation level of extracellular-regulated kinase (ERK) 1/2 in CD3+ T cells. These SDF-1-induced responses were greater in CD3+ T cells from DJ-1 gene knockout (DJ-1(-/-)) mice than in those from wild type (DJ-1(+/+)) mice and were abolished by treatment with WZ811 and PD98059, inhibitors of CXCR4 and ERK1/2, respectively. Flow cytometry revealed that expression of the CXCR4 receptor was greater in CD3+ T cells from DJ-1(-/-) mice than in those from the controls. Moreover, expression of the CD3 protein was observed in the neointimal plaque from carotid artery-ligated mice and was stronger in DJ-1(-/-) mice compared with controls. The CD3+ T cell subsets, Th1 and Th17, showed increased production of interferon-γ and interleukin-17 in DJ-1(-/-) compared with DJ-1(+/+) mice. DJ-1 protein is involved in the SDF-1-induced CD3+ T cell migration via overexpression of the CXCR4 receptor, and that DJ-1 acts as an inhibitory regulator in vascular remodeling such as neointima formation. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  3. Selective activation of KCa3.1 and CRAC channels by P2Y2 receptors promotes Ca(2+ signaling, store refilling and migration of rat microglial cells.

    Directory of Open Access Journals (Sweden)

    Roger Ferreira

    Full Text Available Microglial activation involves Ca(2+ signaling, and numerous receptors can evoke elevation of intracellular Ca(2+. ATP released from damaged brain cells can activate ionotropic and metabotropic purinergic receptors, and act as a chemoattractant for microglia. Metabotropic P2Y receptors evoke a Ca(2+ rise through release from intracellular Ca(2+ stores and store-operated Ca(2+ entry, and some have been implicated in microglial migration. This Ca(2+ rise is expected to activate small-conductance Ca(2+-dependent K(+ (SK channels, if present. We previously found that SK3 (KCa2.3 and KCa3.1 (SK4/IK1 are expressed in rat microglia and contribute to LPS-mediated activation and neurotoxicity. However, neither current has been studied by elevating Ca(2+ during whole-cell recordings. We hypothesized that, rather than responding only to Ca(2+, each channel type might be coupled to different receptor-mediated pathways. Here, our objective was to determine whether the channels are differentially activated by P2Y receptors, and, if so, whether they play differing roles. We used primary rat microglia and a rat microglial cell line (MLS-9 in which riluzole robustly activates both SK3 and KCa3.1 currents. Using electrophysiological, Ca(2+ imaging and pharmacological approaches, we show selective functional coupling of KCa3.1 to UTP-mediated P2Y2 receptor activation. KCa3.1 current is activated by Ca(2+ entry through Ca(2+-release-activated Ca(2+ (CRAC/Orai1 channels, and both CRAC/Orai1 and KCa3.1 channels facilitate refilling of Ca(2+ stores. The Ca(2+ dependence of KCa3.1 channel activation was skewed to abnormally high concentrations, and we present evidence for a close physical association of the two channel types. Finally, migration of primary rat microglia was stimulated by UTP and inhibited by blocking either KCa3.1 or CRAC/Orai1 channels. This is the first report of selective coupling of one type of SK channel to purinergic stimulation of microglia

  4. Ang-(1-7) promotes the migration and invasion of human renal cell carcinoma cells via Mas-mediated AKT signaling pathway.

    Science.gov (United States)

    Zheng, Shuai; Yang, Ying; Song, Ran; Yang, Xiaomei; Liu, Hua; Ma, Qian; Yang, Longyan; Meng, Ran; Tao, Tao; Wang, Songlin; He, Junqi

    2015-05-01

    Ang-(1-7) is an active peptide component of renin-angiotensin system and endogenous ligand for Mas receptor. In the current study, we showed that Ang-(1-7) enhanced migratory and invasive abilities of renal cell carcinoma cells 786-O and Caki-1 by wound-healing, transwell migration and transwell invasion assays. Mas antagonist A779 pretreatment or shRNA-mediated Mas knockdown abolished the stimulatory effect of Ang-(1-7). Furthermore, Ang-(1-7)-stimulated AKT activation was inhibited by either A779 pretreatment or Mas knockdown. Blockage of AKT signaling by AKT inhibitor VIII inhibited Ang-(1-7)-induced migration and invasion in 786-O cells. Taken together, our results provided the first evidence for the pro-metastatic role of Ang-(1-7) in RCC, which may help to better understand the molecular mechanism underlying the progression of this tumor. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Ligand Receptor-Mediated Regulation of Growth in Plants.

    Science.gov (United States)

    Haruta, Miyoshi; Sussman, Michael R

    2017-01-01

    Growth and development of multicellular organisms are coordinately regulated by various signaling pathways involving the communication of inter- and intracellular components. To form the appropriate body patterns, cellular growth and development are modulated by either stimulating or inhibiting these pathways. Hormones and second messengers help to mediate the initiation and/or interaction of the various signaling pathways in all complex multicellular eukaryotes. In plants, hormones include small organic molecules, as well as larger peptides and small proteins, which, as in animals, act as ligands and interact with receptor proteins to trigger rapid biochemical changes and induce the intracellular transcriptional and long-term physiological responses. During the past two decades, the availability of genetic and genomic resources in the model plant species, Arabidopsis thaliana, has greatly helped in the discovery of plant hormone receptors and the components of signal transduction pathways and mechanisms used by these immobile but highly complex organisms. Recently, it has been shown that two of the most important plant hormones, auxin and abscisic acid (ABA), act through signaling pathways that have not yet been recognized in animals. For example, auxins stimulate cell elongation by bringing negatively acting transcriptional repressor proteins to the proteasome to be degraded, thus unleashing the gene expression program required for increasing cell size. The "dormancy" inducing hormone, ABA, binds to soluble receptor proteins and inhibits a specific class of protein phosphatases (PP2C), which activates phosphorylation signaling leading to transcriptional changes needed for the desiccation of the seeds prior to entering dormancy. While these two hormone receptors have no known animal counterparts, there are also many similarities between animal and plant signaling pathways. For example, in plants, the largest single gene family in the genome is the protein kinase

  6. USP33 mediates Slit-Robo signaling in inhibiting colorectal cancer cell migration.

    Science.gov (United States)

    Huang, Zhaohui; Wen, Pushuai; Kong, Ruirui; Cheng, Haipeng; Zhang, Binbin; Quan, Cao; Bian, Zehua; Chen, Mengmeng; Zhang, Zhenfeng; Chen, Xiaoping; Du, Xiang; Liu, Jianghong; Zhu, Li; Fushimi, Kazuo; Hua, Dong; Wu, Jane Y

    2015-04-15

    Originally discovered in neuronal guidance, the Slit-Robo pathway is emerging as an important player in human cancers. However, its involvement and mechanism in colorectal cancer (CRC) remains to be elucidated. Here, we report that Slit2 expression is reduced in CRC tissues compared with adjacent noncancerous tissues. Extensive promoter hypermethylation of the Slit2 gene has been observed in CRC cells, which provides a mechanistic explanation for the Slit2 downregulation in CRC. Functional studies showed that Slit2 inhibits CRC cell migration in a Robo-dependent manner. Robo-interacting ubiquitin-specific protease 33 (USP33) is required for the inhibitory function of Slit2 on CRC cell migration by deubiquitinating and stabilizing Robo1. USP33 expression is downregulated in CRC samples, and reduced USP33 mRNA levels are correlated with increased tumor grade, lymph node metastasis and poor patient survival. Taken together, our data reveal USP33 as a previously unknown tumor-suppressing gene for CRC by mediating the inhibitory function of Slit-Robo signaling on CRC cell migration. Our work suggests the potential value of USP33 as an independent prognostic marker of CRC. © 2014 UICC.

  7. NFAT regulates calcium-sensing receptor-mediated TNF production

    Energy Technology Data Exchange (ETDEWEB)

    abdullah, huda ismail; Pedraza, Paulina L.; Hao, Shoujin; Rodland, Karin D.; McGiff, John C.; Ferreri, Nicholas R.

    2006-05-01

    Because nuclear factor of activated T cells (NFAT) has been implicated in TNF production as well as osmoregulation and salt and water homeostasis, we addressed whether calcium-sensing receptor (CaR)-mediated TNF production in medullary thick ascending limb (mTAL) cells was NFAT dependent. TNF production in response to addition of extracellular Ca2+ (1.2 mM) was abolished in mTAL cells transiently transfected with a dominant-negative CaR construct (R796W) or pretreated with the phosphatidylinositol phospholipase C (PI-PLC) inhibitor U-73122. Cyclosporine A (CsA), an inhibitor of the serine/threonine phosphatase calcineurin, and a peptide ligand, VIVIT, that selectively inhibits calcineurin-NFAT signaling, also prevented CaR-mediated TNF production. Increases in calcineurin activity in cells challenged with Ca2+ were inhibited after pretreatment with U-73122 and CsA, suggesting that CaR activation increases calcineurin activity in a PI-PLC-dependent manner. Moreover, U-73122, CsA, and VIVIT inhibited CaR-dependent activity of an NFAT construct that drives expression of firefly luciferase in transiently transfected mTAL cells. Collectively, these data verify the role of calcineurin and NFAT in CaR-mediated TNF production by mTAL cells. Activation of the CaR also increased the binding of NFAT to a consensus oligonucleotide, an effect that was blocked by U-73122 and CsA, suggesting that a calcineurin- and NFAT-dependent pathway increases TNF production in mTAL cells. This mechanism likely regulates TNF gene transcription as U-73122, CsA, and VIVIT blocked CaR-dependent activity of a TNF promoter construct. Elucidating CaR-mediated signaling pathways that regulate TNF production in the mTAL will be crucial to understanding mechanisms that regulate extracellular fluid volume and salt balance.

  8. DMPD: Translational mini-review series on Toll-like receptors: networks regulated byToll-like receptors mediate innate and adaptive immunity. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available e receptors: networks regulated byToll-like receptors mediate innate and adaptive...ed byToll-like receptors mediate innate and adaptive immunity. Authors Parker LC, Prince LR, Sabroe I. Publi...d byToll-like receptors mediate innate and adaptive immunity. Parker LC, Prince LR, Sabroe I. Clin Exp Immun...17223959 Translational mini-review series on Toll-like receptors: networks regulate

  9. Efficient mucosal vaccination mediated by the neonatal Fc receptor.

    Science.gov (United States)

    Ye, Lilin; Zeng, Rongyu; Bai, Yu; Roopenian, Derry C; Zhu, Xiaoping

    2011-02-01

    Almost all infectious diseases are initiated at mucosal surfaces, yet intramuscular or subcutaneous vaccination usually provides only minimal protection at sites of infection owing to suboptimal activation of the mucosal immune system. The neonatal Fc receptor (FcRn) mediates the transport of IgG across polarized epithelial cells lining mucosal surfaces. We mimicked this process by fusing a model antigen, herpes simplex virus type-2 (HSV-2) glycoprotein gD, to an IgG Fc fragment. Intranasal immunization, together with the adjuvant CpG, completely protected wild-type, but not FcRn knockout, mice after intravaginal challenge with virulent HSV-2 186. This immunization strategy induced efficient mucosal and systemic antibody, B- and T-cell immune responses, with stable protection for at least 6 months after vaccination in most of the immunized animals. The FcRn-IgG transcellular transport pathway may provide a general delivery route for subunit vaccines against many mucosal pathogens.

  10. Sphingosine-1-phosphate/S1P Receptors Signaling Modulates Cell Migration in Human Bone Marrow-Derived Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Yaxian Kong

    2014-01-01

    Full Text Available The recruitment of bone marrow-derived mesenchymal stem cells (BMSCs to damaged tissues and sites of inflammation is an essential step for clinical therapy. However, the signals regulating the motility of these cells are still not fully understood. Sphingosine-1-phosphate (S1P, a bioactive sphingolipid metabolite, is known to have a variety of biological effects on various cells. Here, we investigated the roles of S1P and S1P receptors (S1PRs in migration of human BMSCs. We found that S1P exerted a powerful migratory action on human BMSCs. Moreover, by employing RNA interference technology and pharmacological tools, we demonstrated that S1PR1 and S1PR3 are responsible for S1P-induced migration of human BMSCs. In contrast, S1PR2 mediates the inhibition of migration. Additionally, we explored the downstream signaling pathway of the S1P/S1PRs axis and found that activation of S1PR1 or S1PR3 increased migration of human BMSCs through a Gi/extracellular regulated protein kinases 1/2- (ERK1/2- dependent pathway, whereas activation of S1PR2 decreased migration through the Rho/Rho-associated protein kinase (ROCK pathway. In conclusion, we reveal that the S1P/S1PRs signaling axis regulates the migration of human BMSCs via a dual-directional mechanism. Thus, selective modulation of S1PR’s activity on human BMSCs may provide an effective approach to immunotherapy or tissue regeneration.

  11. Hepatoprotective actions of melatonin: Possible mediation by melatonin receptors

    Science.gov (United States)

    Mathes, Alexander M

    2010-01-01

    Melatonin, the hormone of darkness and messenger of the photoperiod, is also well known to exhibit strong direct and indirect antioxidant properties. Melatonin has previously been demonstrated to be a powerful organ protective substance in numerous models of injury; these beneficial effects have been attributed to the hormone’s intense radical scavenging capacity. The present report reviews the hepatoprotective potential of the pineal hormone in various models of oxidative stress in vivo, and summarizes the extensive literature showing that melatonin may be a suitable experimental substance to reduce liver damage after sepsis, hemorrhagic shock, ischemia/reperfusion, and in numerous models of toxic liver injury. Melatonin’s influence on hepatic antioxidant enzymes and other potentially relevant pathways, such as nitric oxide signaling, hepatic cytokine and heat shock protein expression, are evaluated. Based on recent literature demonstrating the functional relevance of melatonin receptor activation for hepatic organ protection, this article finally suggests that melatonin receptors could mediate the hepatoprotective actions of melatonin therapy. PMID:21182223

  12. Intracellular formyl peptide receptor regulates naïve CD4 T cell migration.

    Science.gov (United States)

    Lee, Ha Young; Jeong, Yu Sun; Lee, Mingyu; Kweon, Hee-Seok; Huh, Yang Hoon; Park, Joon Seong; Hwang, Ji Eun; Kim, Kyuseok; Bae, Yoe-Sik

    2018-02-08

    We found that formyl peptide receptor (FPR) 1 and FPR3 were expressed intracellularly and/or the nucleus of naïve CD4 T cell. Activation of naïve CD4 T cells with synthetic intracellular agonists dTAT-WKYMVm and CTP-WKYMVm for FPR members stimulated CD4 T cell migration via pertussis toxin-sensitive manner. Knockdown of FPR1, but not knockdown of FPR3, blocked dTAT-WKYMVm-induced naïve CD4 T cell migration. Stimulation of naïve CD4 T cells with dTAT-WKYMVm elicited the activation of ERK, p38 MAPK, and Akt. Activation of CD4 T cells with anti-CD3 and anti-CD28 antibodies caused surface expression of FPR1 and FPR3, but not FPR2. CD4 T cells isolated from sepsis patients expressed the three members of FPR family on their cell surface. Taken together, our results suggest that intracellular FPR in naïve CD4 T cells and surface FPRs in activated CD4 T cells might regulate immune responses by regulating CD4 T cell activity. Copyright © 2018 Elsevier Inc. All rights reserved.

  13. Secretory phospholipase A2-mediated neuronal cell death involves glutamate ionotropic receptors

    DEFF Research Database (Denmark)

    Kolko, Miriam; de Turco, Elena B; Diemer, Nils Henrik

    2002-01-01

    To define the significance of glutamate ionotropic receptors in sPLA -mediated neuronal cell death we used the NMDA receptor antagonist MK-801 and the AMPA receptor antagonist PNQX. In primary neuronal cell cultures both MK-801 and PNQX inhibited sPLA - and glutamate-induced neuronal death. [ H]A...

  14. Toll-Like Receptor Pathway as Mediator of Bisphosphonate Effects in Breast Cancer

    Science.gov (United States)

    2005-07-01

    receptors TLR4, TLR5 and TLR9 on the gastric epithelium in Helicobacter pylori infection. Clin Exp Immunol 136, 521-526. 10. Leifer, C.A., Kennedy...AD Award Number: W81XWH-04-1-0600 TITLE: Toll -Like Receptor Pathway’as Mediator of Bisphosphonate Effects in Breast Cancer PRINCIPAL INVESTIGATOR...2004 - 14 Jun 2005 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Toll -Like Receptor Pathway as Mediator of Bisphosphonate Effects in Breast Cancer 5b

  15. The Wnt Co-Receptor Lrp5 Is Required for Cranial Neural Crest Cell Migration in Zebrafish.

    Directory of Open Access Journals (Sweden)

    Bernd Willems

    Full Text Available During vertebrate neurulation, cranial neural crest cells (CNCCs undergo epithelial to mesenchymal transition (EMT, delaminate from the neural plate border, and migrate as separate streams into different cranial regions. There, they differentiate into distinct parts of the craniofacial skeleton. Canonical Wnt signaling has been shown to be essential for this process at different levels but the involved receptors remained unclear. Here we show that the frizzled co-receptor low-density-lipoprotein (LDL receptor-related protein 5 (Lrp5 plays a crucial role in CNCC migration and morphogenesis of the cranial skeleton. Early during induction and migration of CNCCs, lrp5 is expressed ubiquitously but later gets restricted to CNCC derivatives in the ventral head region besides different regions in the CNS. A knock-down of lrp5 does not interfere with induction of CNCCs but leads to reduced proliferation of premigratory CNCCs. In addition, cell migration is disrupted as CNCCs are found in clusters at ectopic positions in the dorsomedial neuroepithelium after lrp5 knock-down and transient CRISPR/Cas9 gene editing. These migratory defects consequently result in malformations of the craniofacial skeleton. To date, Lrp5 has mainly been associated with bone homeostasis in mammals. Here we show that in zebrafish, lrp5 also controls cell migration during early morphogenetic processes and contributes to shaping the craniofacial skeleton.

  16. The tyrosine phosphatase STEP mediates AMPA receptor endocytosis after metabotropic glutamate receptor stimulation.

    Science.gov (United States)

    Zhang, Yang; Venkitaramani, Deepa V; Gladding, Clare M; Zhang, Yongfang; Kurup, Pradeep; Molnar, Elek; Collingridge, Graham L; Lombroso, Paul J

    2008-10-15

    Although it is well established that AMPA receptor (AMPAR) trafficking is a central event in several forms of synaptic plasticity, the mechanisms that regulate the surface expression of AMPARs are poorly understood. Previous work has shown that striatal-enriched protein tyrosine phosphatase (STEP) mediates NMDAR endocytosis. This protein tyrosine phosphatase is enriched in the synapses of the striatum, hippocampus, cerebral cortex, and other brain regions. In the present investigation, we have explored whether STEP also regulates AMPAR internalization. We found that (RS)-3,5-dihydroxyphenylglycine (DHPG) stimulation triggered a dose-dependent increase in STEP translation in hippocampal slices and synaptoneurosomes, a process that requires stimulation of mGluR5 (metabotropic glutamate receptor 5) and activation of mitogen-activated protein kinases and phosphoinositide-3 kinase pathways. DHPG-induced AMPAR internalization and tyrosine dephosphorylation of GluR2 (glutamate receptor 2) was blocked by a substrate-trapping TAT-STEP [C/S] protein in hippocampal slices and cultures. Moreover, DHPG-triggered AMPAR internalization was abolished in STEP knock-out mice and restored after replacement of wild-type STEP. These results suggest a role for STEP in the regulation of AMPAR trafficking.

  17. MicroRNA-217 suppresses homocysteine-induced proliferation and migration of vascular smooth muscle cells via N-methyl-D-aspartic acid receptor inhibition.

    Science.gov (United States)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling

    2016-10-01

    Hyperhomocysteine has become a critical risk for atherosclerosis and can stimulate proliferation and migration of vascular smooth muscle cells (VSMCs). N-methyl-D-aspartic acid receptor (NMDAR) is a receptor of homocysteine and mediates the effects of homocysteine on VSMCs. Bioinformatics analysis has shown NMDAR is a potential target of microRNA-217 (miR-217), which exerts multiple functions in cancer tumorigenesis and carotid plaque progression. In this study, we sought to investigate the role of miR-217 in VSMCs phenotype transition under homocysteine exposure and elucidate its effect on atherosclerotic plaque formation. After treating with several doses of homocysteine (0-8 × 10(-4)  mol/L) for 24 hours, the expression of miR-217 in HA-VSMCs and rat aortic VSMCs was not altered. Intriguingly, the expression of NMDAR mRNA and protein was reduced by homocysteine in a dose-dependent manner. Transfection of miR-217 mimic significantly inhibited the proliferation and migration of VSMCs with homocysteine treatment, while transfection of miR-217 inhibitor promoted VSMCs migration. Moreover, miR-217 mimic down-regulated while miR-217 inhibitor up-regulated NMDAR protein expression but not NMDAR mRNA expression. Through luciferase reporter assay, we showed that miR-217 could directly bind to the 3'-UTR of NMDAR. MiR-217 mimic transfection also released the inhibition of cAMP-response element-binding protein (CREB)-PGC-1α signalling induced by homocysteine. Additionally, restoration of PGC-1α expression via AdPGC-1α infection markedly suppressed VSMCs proliferation through the degradation of NADPH oxidase (NOX1) and reduction of reactive oxygen species (ROS). Collectively, our study identified the role of miR-217 in regulating VSMCs proliferation and migration, which might serve as a target for atherosclerosis therapy. © 2016 John Wiley & Sons Australia, Ltd.

  18. Drosophila tensin plays an essential role in cell migration and planar polarity formation during oogenesis by mediating integrin-dependent extracellular signals to actin organization.

    Science.gov (United States)

    Cha, In Jun; Lee, Jang Ho; Cho, Kyoung Sang; Lee, Sung Bae

    2017-03-11

    Oogenesis in Drosophila involves very dynamic cellular changes such as cell migration and polarity formation inside an ovary during short period. Previous studies identified a number of membrane-bound receptors directly receiving certain types of extracellular inputs as well as intracellular signalings to be involved in the regulation of these dynamic cellular changes. However, yet our understanding on exactly how these receptor-mediated extracellular inputs lead to dynamic cellular changes remains largely unclear. Here, we identified Drosophila tensin encoded by blistery (by) as a novel regulator of cell migration and planar polarity formation and characterized the genetic interaction between tensin and integrin during oogenesis. Eggs from by mutant showed decreased hatching rate and morphological abnormality, a round-shape, compared to the wild-type eggs. Further analyses revealed that obvious cellular defects such as defective border cell migration and planar polarity formation might be primarily associated with the decreased hatching rate and the round-shape phenotype of by mutant eggs, respectively. Moreover, by mutation also induced marked defects in F-actin organization closely associated with both cell migration and planar polarity formation during oogenesis of Drosophila. Notably, all these defective phenotypes observed in by mutant eggs became much severer by reduced level of integrin, indicative of a close functional association between integrin and tensin during oogenesis. Collectively, our findings suggest that tensin acts as a crucial regulator of dynamic cellular changes during oogenesis by bridging integrin-dependent extracellular signals to intracellular cytoskeletal organization. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. The E3 ubiquitin ligase NEDD4 mediates cell migration signaling of EGFR in lung cancer cells.

    Science.gov (United States)

    Shao, Genbao; Wang, Ranran; Sun, Aiqin; Wei, Jing; Peng, Ke; Dai, Qian; Yang, Wannian; Lin, Qiong

    2018-02-19

    EGFR-dependent cell migration plays an important role in lung cancer progression. Our previous study observed that the HECT E3 ubiquitin ligase NEDD4 is significantly correlated with tumor metastasis and required for migration and invasion signaling of EGFR in gastric cancer cells. However, how NEDD4 promotes the EGFR-dependent lung cancer cell migration is unknown. This study is to elucidate the mechanism by which NEDD4 mediates the EGFR lung cancer migration signaling. Lentiviral vector-loaded NEDD4 shRNA was used to deplete endogenous NEDD4 in lung cancer cell lines. Effects of the NEDD4 knockdown on the EGFR-dependent or independent lung cancer cell migration were determined using the wound-healing and transwell assays. Association of NEDD4 with activated EGFR was assayed by co-immunoprecipitation. Co-expression of NEDD4 with EGFR or PTEN was determined by immunohistochemical (IHC) staining in 63 lung adenocarcinoma tissue samples. Effects of NEDD4 ectopic expression or knockdown on PTEN ubiquitination and down-regulation, AKT activation and lysosomal secretion were examined using the GST-Uba pulldown assay, immunoblotting, immunofluorescent staining and a human cathepsin B ELISA assay respectively. The specific cathepsin B inhibitor CA-074Me was used for assessing the role of cathepsin B in lung cancer cell migration. Knockdown of NEDD4 significantly reduced EGF-stimulated cell migration in non-small cell lung carcinoma (NSCLC) cells. Co-immunoprecipitation assay found that NEDD4 is associated with EGFR complex upon EGF stimulation, and IHC staining indicates that NEDD4 is co-expressed with EGFR in lung adenocarcinoma tumor tissues, suggesting that NEDD4 might mediate lung cancer cell migration by interaction with the EGFR signaling complex. Interestingly, NEDD4 promotes the EGF-induced cathepsin B secretion, possibly through lysosomal exocytosis, as overexpression of the ligase-dead mutant of NEDD4 impedes lysosomal secretion, and knockdown of NEDD4

  20. Stimulation of neutrophil FC-receptor-mediated ingestion

    Energy Technology Data Exchange (ETDEWEB)

    Gresham, H.; Blackburn, W.; Clement, L.; Volanakis, J.

    1986-03-05

    Neutrophils (PMN) treated with amphotericin B (AmB), chemotactic peptide (FMLP), phorbol ester (PMA), a low molecular weight (< 10,000 M/sub r/), heat-labile cytokine (CK), or media were incubated with sheep erythrocytes coated with IgG (EIgG) and ingestion assessed as a phagocytic index (PI=number of EIgG ingested/100 PMN). PMN incubated with media bound but did not ingest EIgG (PI=35) as efficiently as PMN stimulated with the following reagents: 4..mu..M AmB (PI=150), 10/sup -9/M FMLP (PI=135), 5ng/ml PMA (PI=310), and CK (PI=350). PMN treated with .25% butanol bound but did not ingest EIgG even after stimulation by the above agents. Monoclonal anti-PMN Fc receptor (3G8) treatment of non-stimulated and stimulated PMN inhibited both attachment and ingestion of EIgG. 1C2, a monoclonal antibody which precipitates a 54,000 M/sub r/ protein from PMN, had no effect on EIgG attachment or ingestion by non-stimulated PMN, yet inhibited EIgG ingestion but not attachment by PMN stimulated with CK (PI=40), AmB (PI=35), and FMLP (PI=60), but not PMA (PI=280). These data indicate that phagocytosis mediated by 3G8-positive Fc receptors may be inhibited by agents which decrease membrane fluidity (e.g. butanol) and may be enhanced by agents which require the molecule recognized by 1C2 (AmB, FMLP, and CK) and an agent which does not (PMA).

  1. Crop domestication, global human-mediated migration, and the unresolved role of geography in pest control

    Directory of Open Access Journals (Sweden)

    Yolanda H. Chen

    2016-05-01

    Full Text Available Abstract Ecological pest management seeks to improve pest control through the manipulation of ecological processes that promote natural enemies and suppress pests. These approaches can involve cultural practices such as reduced tillage, increased use of non-crop plants that provide food and shelter for natural enemies, and intercropping to enhance the abundance and diversity of natural enemies. A major assumption of ecological pest management is that these activities can be equally effective for all insect herbivores. Here, I propose that these strategies may only be effective for a subset of pests and geographic regions because most insect pests have complex evolutionary histories that make them difficult to manage. I discuss how crop domestication and human-mediated migration are major evolutionary events that shape the geography of interactions between plants, herbivores, and natural enemies. Insect herbivores can evolve to be pests through three major modes: 1 herbivores associated with the crop wild ancestor may shift onto the domesticated crop, 2 herbivores may host-shift from native host plants onto an introduced crop, or 3 human-mediated migration can introduce insect pests into new cropping regions. The resulting geographic structure can influence the success of pest management by altering ecological factors such as: species distributions, patterns of biodiversity, community structure, and natural enemy attack rates. I discuss how the different modes of insect pest evolution structure a set of relevant questions and approaches for ecological pest management. By acknowledging how agricultural history and geography shape the ecology and evolution of insect pests, we may collectively develop a better capacity to identify where and how ecological pest management approaches can be most broadly effective.

  2. A collapsin response mediator protein 2 isoform controls myosin II-mediated cell migration and matrix assembly by trapping ROCK II

    DEFF Research Database (Denmark)

    Yoneda, Atsuko; Morgan-Fisher, Marie; Wait, Robin

    2012-01-01

    nonneuronal cells, regulates myosin II-mediated cellular functions, including cell migration. While the CRMP-2 short form (CRMP-2S) is recognized as a substrate of the Rho-GTP downstream kinase ROCK in neuronal cells, a CRMP-2 complex containing 2L not only bound the catalytic domain of ROCK II through two...... binding domains but also trapped and inhibited the kinase. CRMP-2L protein levels profoundly affected haptotactic migration and the actin-myosin cytoskeleton of carcinoma cells as well as nontransformed epithelial cell migration in a ROCK activity-dependent manner. Moreover, the ectopic expression of CRMP......-2L but not -2S inhibited fibronectin matrix assembly in fibroblasts. Underlying these responses, CRMP-2L regulated the kinase activity of ROCK II but not ROCK I, independent of GTP-RhoA levels. This study provides a new insight into CRMP-2 as a controller of myosin II-mediated cellular functions...

  3. cAMP-Induced Desensitization of Surface cAMP Receptors in Dictyostelium : Different Second Messengers Mediate Receptor Phosphorylation, Loss of Ligand Binding, Degradation of Receptor, and Reduction of Receptor mRNA Levels

    NARCIS (Netherlands)

    Haastert, Peter J.M. van; Wang, Mei; Bominaar, Anthony A.; Devreotes, Peter N.; Schaap, Pauline

    Surface cAMP receptors on Dictyostelium cells are linked to several second messenger systems and mediate multiple physiological responses, including chemotaxis and differentiation. Activation of the receptor also triggers events which desensitize signal transduction. These events include the

  4. CCR7 Mediates TGF-β1-Induced Human Malignant Glioma Invasion, Migration, and Epithelial-Mesenchymal Transition by Activating MMP2/9 Through the Nuclear Factor KappaB Signaling Pathway.

    Science.gov (United States)

    Zheng, Yanyan; Miu, Yiting; Yang, Xiaokai; Yang, Xiaoguo; Zhu, Meijia

    2017-10-01

    Chemokine receptor 7 (CCR7) has emerged as an inducer of invasion, migration, and epithelial-mesenchymal transition (EMT) in cancer. In this research, human malignant glioma cells were stimulated with transforming growth factor beta 1 (TGF-β1) and siCCR7. The data show that CCR7 mediates TGF-β1-induced EMT, migration, and invasion in U251 and U87 cells and that these effects of TGF-β1 were reversed by treatment with siCCR7 or a CCR7 neutralizing antibody. Importantly, the TGF-β1-mediated increase in nuclear factor kappaB (NF-κB) activity in human glioma cells was reduced by treatment with siCCR7 or a CCR7 neutralizing antibody. Furthermore, CCR7 was shown to mediate TGF-β1-induced glioma cancer cell migration by activating matrix metalloproteinase 2 (MMP2)/9. Our results indicate that CCR7 mediates TGF-β1-induced MMP2/9 expression through NF-κB signaling, thus facilitating glioma cell migration, invasion, and EMT, all of which progressively increase with glioblastoma progression. These findings indicate that CCR7 is a potential therapeutic target for malignant glioma.

  5. Visfatin Mediates SCLC Cells Migration across Brain Endothelial Cells through Upregulation of CCL2

    Directory of Open Access Journals (Sweden)

    Tingting Liu

    2015-05-01

    Full Text Available Small-cell lung cancer (SCLC is characterized as an aggressive tumor with brain metastasis. Although preventing SCLC metastasis to the brain is immensely important for survival, the molecular mechanisms of SCLC cells penetrating the blood–brain barrier (BBB are largely unknown. Recently, visfatin has been considered as a novel pro-inflammatory adipocytokine involved in various cancers. Herein, we present evidence that elevated levels of visfatin in the serum of SCLC patients were associated with brain metastasis, and visfain was increased in NCI-H446 cells, a SCLC cell line, during interacting with human brain microvascular endothelial cells (HBMEC. Using in vitro BBB model, we found that visfatin could promote NCI-H446 cells migration across HBMEC monolayer, while the effect was inhibited by knockdown of visfatin. Furthermore, our findings indicated that CC chemokine ligand 2 (CCL2 was involved in visfatin-mediated NCI-H446 cells transendothelial migtation. Results also showed that the upregulation of CCL2 in the co-culture system was reversed by blockade of visfatin. In particular, visfatin-induced CCL2 was attenuated by specific inhibitor of PI3K/Akt signaling in NCI-H446 cells. Taken together, we demonstrated that visfatin was a prospective target for SCLC metastasis to brain, and understanding the molecular mediators would lead to effective strategies for inhibition of SCLC brain metastasis.

  6. WNT5A-ROR2 is induced by inflammatory mediators and is involved in the migration of human ovarian cancer cell line SKOV-3.

    Science.gov (United States)

    Arabzadeh, Somayeh; Hossein, Ghamartaj; Salehi-Dulabi, Zahra; Zarnani, Amir Hassan

    2016-01-01

    Wnt5A, which is a member of the non-transforming Wnt protein family, is implicated in inflammatory processes. It is also highly expressed by ovarian cancer cells. ROR2, which is a member of the Ror-family of receptor tyrosine kinases, acts as a receptor or co-receptor for Wnt5A. The Wnt5A-ROR2 signaling pathway plays essential roles in the migration and invasion of several types of tumor cell and influences their cell polarity. We investigated the modulation of Wnt5A-ROR2 by inflammatory mediators and its involvement in the migration of the human ovarian cancer cell line SKOV-3. SKOV-3 cells were treated with LPS (lipopolysaccharide), LTA (lipoteichoic acid) and recombinant human IL-6 alone or in combination with STAT3 inhibitor (S1155S31-201) or NF-kB inhibitor (BAY11-7082) for 4, 8, 12, 24 and 48 h. The Wnt5A and ROR2 expression levels were determined at the gene and protein levels. Cells were transfected with specific siRNA against Wnt5A in the absence or presence of human anti-ROR2 antibody and cell migration was assessed using transwells. There was a strong downregulation of Wnt5A expression in the presence of STAT3 or NF-kB inhibitors. Cell stimulation with LTA or IL-6 for 8 h led to significantly increased levels of Wnt5A (5- and 3-fold higher, respectively). LPS, LTA or IL-6 treatment significantly increased ROR2 expression (2-fold after 48 h). LPS- or LTA-induced Wnt5A or ROR2 expression was abrogated in the presence of STAT3 inhibitor (p migration was decreased by 80 % in siRNA Wnt5A-transfected cells in the presence of anti-human ROR2 antibody (p migration. The results described here provide new insight into the role of the Wnt5A-ROR2 complex in ovarian cancer progression in relation to inflammation.

  7. Trypanosoma cruzi: antigen-receptor mediated endocytosis of antibody

    Directory of Open Access Journals (Sweden)

    Judith Abelha

    1981-06-01

    Full Text Available Trypanomastigote forms of Trypanosoma cruzi were derived from tissue culture and incubated with immune and non-immune human sera. All immune sera showed high titers of specific humoral antibodies of the IgM or the IgG type. Agglutination and swelling of parasites were observed after incubation at 37ºC, but many trypomastigotes remained free-swimming in the sera for two to three days. The quantitiy of immune serum capable of lysing a maximum of 10 x 10 [raised to the power of 6] sensitized red cells was not capable of lysing 4 x 10 [raised to the power of 3] tripomastigotes. Typically, the parasites underwent cyclical changes with the formation of clumps of amastigotes and the appearance of epimastigote forms. Multiplication of the parasites was observed in immune sera. Further, the infectivity of the parasites to susceptible mice was not lost. All sera used produced similar general effects on the growth of the parasite. The antibody bound to T. cruzi appeard to enter cells by antigen-receptor mediated endocytosis. The ferritin-conjugated antibody was internalized and delivered to phagolysosomes where they might be completely degraded to amino-acids. This seemed to be a coupled process by which the immunoglobulin is first bound to specific parasite surface receptor and then rapidly endocytosed by the cell.Formas tripomastigotas de Trypanosoma cruzi derivadas de cultura de tecido foram encubadas com soros humanos imunes e não-imunes.Todos os soros humanos usados tinham títulos elevados de anticorpos das classes IgM ou IgG. Aglutinação e entumescimento dos parasitos eram observados apos encubação a 37ºC mas muitos tripomastigotas permaneceram circulando livremente nos soros por dois a três dias. A quantidade de soro imune capaz de lisar um máximo de 10 x 10 [elevado a 6] hemácias sensibilizadas não foi capaz de lisar 4 x 10 [elevado a 3] tripomastigotas. Tipicamente, os parasitos apresentavam alterações cíclicas com formação de

  8. JIP3 regulates neuronal radial migration by mediating TrkB axonal anterograde transport in the developing cerebral cortex.

    Science.gov (United States)

    Ma, Huixian; Yu, Hui; Li, Ting; Zhao, Yan; Hou, Ming; Chen, Zheyu; Wang, Yue; Sun, Tao

    2017-04-15

    Radial migration is essential for the precise lamination and the coordinated function of the cerebral cortex. However, the molecular mechanisms for neuronal radial migration are not clear. Here, we report that c-Jun NH2-terminal kinase (JNK)-interacting protein-3 (JIP3) is highly expressed in the brain of embryonic mice and essential for radial migration. Knocking down JIP3 by in utero electroporation specifically perturbs the radial migration of cortical neurons but has no effect on neurogenesis and neuronal differentiation. Furthermore, we illustrate that JIP3 knockdown delays but does not block the migration of cortical neurons by investigating the distribution of neurons with JIP3 knocked down in the embryo and postnatal mouse. Finally, we find that JIP3 regulates cortical neuronal migration by mediating TrkB axonal anterograde transport during brain development. These findings deepen our understanding of the regulation of neuronal development by JIP3 and provide us a novel view on the regulating mechanisms of neuronal radial migration. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Chlamydia pneumoniae Infection Promotes Vascular Smooth Muscle Cell Migration through a Toll-Like Receptor 2-Related Signaling Pathway

    Science.gov (United States)

    Wang, Beibei; Zhang, Tengteng; Wang, Haiwei; Zhang, Junxia; Wei, Junyan; Shen, Bingling; Liu, Xin; Xu, Zhelong; Zhang, Lijun

    2013-01-01

    The migration of vascular smooth muscle cells (VSMCs) from the media to the intima is proposed to be a key event in the development of atherosclerosis. Recently, we reported that Chlamydia pneumoniae infection is involved in VSMC migration. However, the exact mechanisms for C. pneumoniae infection-induced VSMC migration are not yet well elucidated. In this study, we examined the role of the Toll-like receptor 2 (TLR2) activation-related signaling pathway in VSMC migration induced by C. pneumoniae infection. An Affymetrix-based gene expression array was conducted to identify the changes of gene expression in rat primary VSMCs (rVSMCs) infected with C. pneumoniae. Both the microarray analysis and quantitative real-time reverse transcription (RT)-PCR revealed that TLR2 mRNA expression was strongly upregulated 12 h after C. pneumoniae infection. RT-PCR and Western blot analysis further showed that the expression levels of TLR2 mRNA and protein significantly increased at the different time points after infection. Immunocytochemical analysis suggested a TLR2 recruitment to the vicinity of C. pneumoniae inclusions. Cell migration assays showed that the TLR2-neutralizing antibody could significantly inhibit C. pneumoniae infection-induced rVSMC migration. In addition, C. pneumoniae infection stimulated Akt phosphorylation at Ser 473, which was obviously suppressed by the PI3K inhibitor LY294002, thereby inhibiting rVSMC migration caused by C. pneumoniae infection. Furthermore, both the infection-induced Akt phosphorylation and rVSMC migration were suppressed by the TLR2-neutralizing antibody. Taken together, these data suggest that C. pneumoniae infection can promote VSMC migration possibly through the TLR2-related signaling pathway. PMID:24082081

  10. The Caenorhabditis elegans matrix non-peptidase MNP-1 is required for neuronal cell migration and interacts with the Ror receptor tyrosine kinase CAM-1.

    Science.gov (United States)

    Craft, Teresa R; Forrester, Wayne C

    2017-04-01

    Directed cell migration is critical for metazoan development. During Caenorhabditis elegans development many neuronal, muscle and other cell types migrate. Multiple classes of proteins have been implicated in cell migration including secreted guidance cues, receptors for guidance cues and intracellular proteins that respond to cues to polarize cells and produce the forces that move them. In addition, cell surface and secreted proteases have been identified that may clear the migratory route and process guidance cues. We report here that mnp-1 is required for neuronal cell and growth cone migrations. MNP-1 is expressed by migrating cells and functions cell autonomously for cell migrations. We also find a genetic interaction between mnp-1 and cam-1, which encodes a Ror receptor tyrosine kinase required for some of the same cell migrations. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Resveratrol Prevents High Fluence Red Light-Emitting Diode Reactive Oxygen Species-Mediated Photoinhibition of Human Skin Fibroblast Migration.

    Directory of Open Access Journals (Sweden)

    Andrew Mamalis

    Full Text Available Skin fibrosis is a significant medical problem that leads to a functional, aesthetic, and psychosocial impact on quality-of-life. Light-emitting diode-generated 633-nm red light (LED-RL is part of the visible light spectrum that is not known to cause DNA damage and is considered a safe, non-invasive, inexpensive, and portable potential alternative to ultraviolet phototherapy that may change the treatment paradigm of fibrotic skin disease.The goal of our study was to investigate the how reactive oxygen species (ROS free radicals generated by high fluence LED-RL inhibit the migration of skin fibroblasts, the main cell type involved in skin fibrosis. Fibroblast migration speed is increased in skin fibrosis, and we studied cellular migration speed of cultured human skin fibroblasts as a surrogate measure of high fluence LED-RL effect on fibroblast function. To ascertain the inhibitory role of LED-RL generated ROS on migration speed, we hypothesized that resveratrol, a potent antioxidant, could prevent the photoinhibitory effects of high fluence LED-RL on fibroblast migration speed.High fluence LED-RL generated ROS were measured by flow cytometry analysis using dihydrorhodamine (DHR. For purposes of comparison, we assessed the effects of ROS generated by hydrogen peroxide (H2O2 on fibroblast migration speed and the ability of resveratrol, a well known antioxidant, to prevent LED-RL and H2O2 generated ROS-associated changes in fibroblast migration speed. To determine whether resveratrol could prevent the high fluence LED-RL ROS-mediated photoinhibition of human skin fibroblast migration, treated cells were incubated with resveratrol at concentrations of 0.0001% and 0.001% for 24 hours, irradiated with high fluences LED-RL of 480, 640, and 800 J/cm2.High fluence LED-RL increases intracellular fibroblast ROS and decreases fibroblast migration speed. LED-RL at 480, 640 and 800 J/cm2 increased ROS levels to 132.8%, 151.0%, and 158.4% relative to matched

  12. Resveratrol Prevents High Fluence Red Light-Emitting Diode Reactive Oxygen Species-Mediated Photoinhibition of Human Skin Fibroblast Migration.

    Science.gov (United States)

    Mamalis, Andrew; Koo, Eugene; Isseroff, R Rivkah; Murphy, William; Jagdeo, Jared

    2015-01-01

    Skin fibrosis is a significant medical problem that leads to a functional, aesthetic, and psychosocial impact on quality-of-life. Light-emitting diode-generated 633-nm red light (LED-RL) is part of the visible light spectrum that is not known to cause DNA damage and is considered a safe, non-invasive, inexpensive, and portable potential alternative to ultraviolet phototherapy that may change the treatment paradigm of fibrotic skin disease. The goal of our study was to investigate the how reactive oxygen species (ROS) free radicals generated by high fluence LED-RL inhibit the migration of skin fibroblasts, the main cell type involved in skin fibrosis. Fibroblast migration speed is increased in skin fibrosis, and we studied cellular migration speed of cultured human skin fibroblasts as a surrogate measure of high fluence LED-RL effect on fibroblast function. To ascertain the inhibitory role of LED-RL generated ROS on migration speed, we hypothesized that resveratrol, a potent antioxidant, could prevent the photoinhibitory effects of high fluence LED-RL on fibroblast migration speed. High fluence LED-RL generated ROS were measured by flow cytometry analysis using dihydrorhodamine (DHR). For purposes of comparison, we assessed the effects of ROS generated by hydrogen peroxide (H2O2) on fibroblast migration speed and the ability of resveratrol, a well known antioxidant, to prevent LED-RL and H2O2 generated ROS-associated changes in fibroblast migration speed. To determine whether resveratrol could prevent the high fluence LED-RL ROS-mediated photoinhibition of human skin fibroblast migration, treated cells were incubated with resveratrol at concentrations of 0.0001% and 0.001% for 24 hours, irradiated with high fluences LED-RL of 480, 640, and 800 J/cm2. High fluence LED-RL increases intracellular fibroblast ROS and decreases fibroblast migration speed. LED-RL at 480, 640 and 800 J/cm2 increased ROS levels to 132.8%, 151.0%, and 158.4% relative to matched controls

  13. Migration of myeloid cells during inflammation is differentially regulated by the cell surface receptors Slamf1 and Slamf8.

    Directory of Open Access Journals (Sweden)

    Guoxing Wang

    Full Text Available Previous studies have demonstrated that the cell surface receptor Slamf1 (CD150 is requisite for optimal NADPH-oxidase (Nox2 dependent reactive oxygen species (ROS production by phagocytes in response to Gram- bacteria. By contrast, Slamf8 (CD353 is a negative regulator of ROS in response to Gram+ and Gram- bacteria. Employing in vivo migration after skin sensitization, induction of peritonitis, and repopulation of the small intestine demonstrates that in vivo migration of Slamf1-/- dendritic cells and macrophages is reduced, as compared to wt mice. By contrast, in vivo migration of Slamf8-/- dendritic cells, macrophages and neutrophils is accelerated. These opposing effects of Slamf1 and Slamf8 are cell-intrinsic as judged by in vitro migration in transwell chambers in response to CCL19, CCL21 or CSF-1. Importantly, inhibiting ROS production of Slamf8-/- macrophages by diphenyleneiodonium chloride blocks this in vitro migration. We conclude that Slamf1 and Slamf8 govern ROS-dependent innate immune responses of myeloid cells, thus modulating migration of these cells during inflammation in an opposing manner.

  14. BRK targets Dok1 for ubiquitin-mediated proteasomal degradation to promote cell proliferation and migration.

    Directory of Open Access Journals (Sweden)

    Sayem Miah

    Full Text Available Breast tumor kinase (BRK, also known as protein tyrosine kinase 6 (PTK6, is a non-receptor tyrosine kinase overexpressed in more that 60% of human breast carcinomas. The overexpression of BRK has been shown to sensitize mammary epithelial cells to mitogenic signaling and to promote cell proliferation and tumor formation. The molecular mechanisms of BRK have been unveiled by the identification and characterization of BRK target proteins. Downstream of tyrosine kinases 1 or Dok1 is a scaffolding protein and a substrate of several tyrosine kinases. Herein we show that BRK interacts with and phosphorylates Dok1 specifically on Y362. We demonstrate that this phosphorylation by BRK significantly downregulates Dok1 in a ubiquitin-proteasome-mediated mechanism. Together, these results suggest a novel mechanism of action of BRK in the promotion of tumor formation, which involves the targeting of tumor suppressor Dok1 for degradation through the ubiquitin proteasomal pathway.

  15. The pH-sensing receptor OGR1 improves barrier function of epithelial cells and inhibits migration in an acidic environment.

    Science.gov (United States)

    de Vallière, Cheryl; Vidal, Solange; Clay, Ieuan; Jurisic, Giorgia; Tcymbarevich, Irina; Lang, Silvia; Ludwig, Marie-Gabrielle; Okoniewski, Michal; Eloranta, Jyrki J; Kullak-Ublick, Gerd A; Wagner, Carsten A; Rogler, Gerhard; Seuwen, Klaus

    2015-09-15

    The pH-sensing receptor ovarian cancer G protein-coupled receptor 1 (OGR1; GPR68) is expressed in the gut. Inflammatory bowel disease is typically associated with a decrease in local pH, which may lead to altered epithelial barrier function and subsequent gastrointestinal repair involving epithelial cell adhesion and migration. As the mechanisms underlying the response to pH changes are not well understood, we have investigated OGR1-mediated, pH-dependent signaling pathways in intestinal epithelial cells. Caco-2 cells stably overexpressing OGR1 were created and validated as tools to study OGR1 signaling. Barrier function, migration, and proliferation were measured using electric cell-substrate impedance-sensing technology. Localization of the tight junction proteins zonula occludens protein 1 and occludin and the rearrangement of cytoskeletal actin were examined by confocal microscopy. Paracellular permeability and protein and gene expression analysis using DNA microarrays were performed on filter-grown Caco-2 monolayers. We report that an acidic pH shift from pH 7.8 to 6.6 improved barrier function and stimulated reorganization of filamentous actin with prominent basal stress fiber formation. Cell migration and proliferation during in vitro wound healing were inhibited. Gene expression analysis revealed significant upregulation of genes related to cytoskeleton remodeling, cell adhesion, and growth factor signaling. We conclude that acidic extracellular pH can have a signaling function and impact the physiology of intestinal epithelial cells. The deconstruction of OGR1-dependent signaling may aid our understanding of mucosal inflammation mechanisms. Copyright © 2015 the American Physiological Society.

  16. Odorant Receptor 51E2 Agonist β-ionone Regulates RPE Cell Migration and Proliferation

    Directory of Open Access Journals (Sweden)

    Nikolina Jovancevic

    2017-11-01

    Full Text Available The odorant receptor 51E2 (OR51E2, which is well-characterized in prostate cancer cells and epidermal pigment cells, was identified for the first time as the most highly expressed OR in human fetal and adult retinal pigment epithelial (RPE cells. Immunofluorescence staining and Western blot analysis revealed OR51E2 localization throughout the cytosol and in the plasma membrane. Additionally, immunohistochemical staining of diverse layers of the eye showed that the expression of OR51E2 is restricted to the pigment cells of the RPE and choroid. The results of Ca2+-imaging experiments demonstrate that activation of OR51E2 triggers a Ca2+ dependent signal pathway in RPE cells. Downstream signaling of OR51E2 involves the activation of adenylyl cyclase, ERK1/2 and AKT. The activity of these protein kinases likely accounts for the demonstrated increase in the migration and proliferation of RPE cells upon stimulation with the OR51E2 ligand β-ionone. These findings suggest that OR51E2 is involved in the regulation of RPE cell growth. Thus, OR51E2 represents a potential target for the treatment of proliferative disorders.

  17. Towards a correct description of zooplankton feeding in models: taking into account food-mediated unsynchronized vertical migration.

    Science.gov (United States)

    Morozov, A Yu; Arashkevich, E G

    2010-01-21

    Complex nature of foraging behaviour of zooplankton makes it difficult to describe adequately zooplankton grazing in models with vertical space. In mean-field models (based on systems of PDEs or coupled ODEs), zooplankton feeding at a given depth is normally computed as the product of the local functional response and the zooplankton density at this depth. Such simplification is often at odds with field observations which show the absence of clear relationship between intake rates of organisms and the ambient food density. The observed discrepancy is generic and is often caused by fast non-synchronous vertical migration of organisms with different nutrition status. In this paper, we suggest a simple way of incorporating unsynchronized short-term vertical migration of zooplankton into the mean-field modelling framework. We compute grazing of zooplankton in each layer depending on feeding activity of organisms in the layer. We take into account grazing impact of animals which are in the active phase of foraging cycle at the given moment of time but neglect the impact of animals which are in the non-active phase of the cycle (e.g. digesting food). Unsynchronized vertical migration determines the vertical distribution of actively feeding animals in layers depending on vertical distribution of food. In this paper, we compare two generic plankton models: (i) a model based on 'classical' grazing approach and (ii) a model incorporating food-mediated unsynchronized vertical migration of zooplankton. We show that including unsynchronized food-mediated migration would make the behaviour of a plankton model more realistic. This would imply a significant enhancement of ecosystem's stability and some additional mechanisms of regulation of algal blooms. In the system with food-mediated unsynchronized vertical migration, the control of phytoplankton by herbivorous becomes possible even for very large concentrations of nutrients in the water (formally, when the system's carrying

  18. Pharmacological significance of the interplay between angiotensin receptors: MAS receptors as putative final mediators of the effects elicited by angiotensin AT1 receptors antagonists.

    Science.gov (United States)

    Pernomian, Larissa; Pernomian, Laena; Gomes, Mayara S; da Silva, Carlos H T P

    2015-12-15

    The interplay between angiotensin AT1 receptors and MAS receptors relies on several inward regulatory mechanisms from renin-angiotensin system (RAS) including the functional crosstalk between angiotensin II and angiotensin-(1-7), the competitive AT1 antagonism exhibited by angiotensin-(1-7), the antagonist feature assigned to AT1/MAS heterodimerization on AT1 signaling and the AT1-mediated downregulation of angiotensin-converting enzyme 2 (ACE2). Recently, such interplay has acquired an important significance to RAS Pharmacology since a few studies have supporting strong evidences that MAS receptors mediate the effects elicited by AT1 antagonists. The present Perspective provides an overview of the regulatory mechanisms involving AT1 and MAS receptors, their significance to RAS Pharmacology and the future directions on the interplay between angiotensin receptors. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Epidermal growth factor receptor signaling mediates aldosterone-induced profibrotic responses in kidney

    Energy Technology Data Exchange (ETDEWEB)

    Sheng, Lili; Yang, Min; Ding, Wei [Department of Nephrology, Shanghai Fifth People' s Hospital, Fudan University, Shanghai 200240 (China); Zhang, Minmin [Department of Nephrology, Shanghai Huashan Hospital, Fudan University, Shanghai 200240 (China); Niu, Jianying [Department of Nephrology, Shanghai Fifth People' s Hospital, Fudan University, Shanghai 200240 (China); Qiao, Zhongdong [School of Life Science and Biotechnology, Shanghai Jiaotong University, Shanghai 200240 (China); Gu, Yong, E-mail: yonggu@vip.163.com [Department of Nephrology, Shanghai Fifth People' s Hospital, Fudan University, Shanghai 200240 (China); Department of Nephrology, Shanghai Huashan Hospital, Fudan University, Shanghai 200240 (China)

    2016-08-01

    Aldosterone has been recognized as a risk factor for the development of chronic kidney disease (CKD). Studies have indicated that enhanced activation of epidermal growth factor receptor (EGFR) is associated with the development and progression of renal fibrosis. But if EGFR is involved in aldosterone-induced renal fibrosis is less investigated. In the present study, we examined the effect of erlotinib, an inhibitor of EGFR tyrosine kinase activity, on the progression of aldosterone-induced renal profibrotic responses in a murine model underwent uninephrectomy. Erlotinib-treated rats exhibited relieved structural lesion comparing with rats treated with aldosterone alone, as characterized by glomerular hypertrophy, mesangial cell proliferation and expansion. Also, erlotinib inhibited the expression of TGF-β, α-SMA and mesangial matrix proteins such as collagen Ⅳ and fibronectin. In cultured mesangial cells, inhibition of EGFR also abrogated aldosterone-induced expression of extracellular matrix proteins, cell proliferation and migration. We also demonstrated that aldosterone induced the phosphorylation of EGFR through generation of ROS. And the activation of EGFR resulted in the phosphorylation of ERK1/2, leading to the activation of profibrotic pathways. Taken together, we concluded that aldosterone-mediated tissue fibrosis relies on ROS induced EGFR/ERK activation, highlighting EGFR as a potential therapeutic target for modulating renal fibrosis. - Highlights: • EGFR was involved in aldosterone-induced renal profibrotic responses. • Aldosterone-induced EGFR activation was mediated by MR-dependent ROS generation. • EGFR activated the MAPK/ERK1/2 signaling to promote renal fibrosis.

  20. Activation of membrane estrogen receptors attenuates opioid receptor-like1 receptor-mediated antinociception via an ERK-dependent non-genomic mechanism.

    Science.gov (United States)

    Small, K M; Nag, S; Mokha, S S

    2013-01-01

    To our knowledge, the present data are the first to demonstrate that activation of membrane estrogen receptors (mERs) abolishes opioid receptor-like 1 (ORL1) receptor-mediated analgesia via extracellular signal-regulated kinase (ERK)-dependent non-genomic mechanisms. Estrogen was shown previously to both attenuate ORL1-mediated antinociception and down-regulate the ORL1 gene expression. The present study investigated whether non-genomic mechanisms contribute to estrogen-induced attenuation of ORL1-mediated antinociception by the mERs GPR30, Gq-coupled mER, ERα, and ERβ. E2BSA [β-estradiol-6-(O-carboxymethyl)oxime: bovine serum albumin] (0.5mM), a membrane impermeant analog of estradiol, injected intrathecally immediately prior to orphanin FQ (OFQ;10 nmol), the endogenous ligand for the ORL1 receptor, abolished OFQ's antinociceptive effect in both male and ovariectomized (OVX) female rats, assessed using the heat-induced tail-flick assay. This effect was not altered by protein synthesis inhibitor, anisomycin (125 μg), given intrathecally 15 min prior to E2BSA and OFQ. Intrathecal application of selective receptor agonists permitted the relative contributions of various estrogen receptors in mediating this blockade of the antinociceptive response of OFQ. Activation of GPR30, Gq-mER, ERα, but not ERβ abolished ORL1-mediated antinociception in males and OVX females. E2BSA produced a parallel and significant increase in the phosphorylation of ERK 2 only in OVX females, and pre-treatment with MEK/ERK 1/2 inhibitor, U0126 (10 μg), blocked the mER-mediated abolition of ORL1-mediated antinociception in OVX females. Taken together, the data are consistent with the interpretations that mER activation attenuates ORL1-mediated antinociception through a non-genomic, ERK 2-dependent mechanism in females. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

  1. EGFRvIII-specific chimeric antigen receptor T cells migrate to and kill tumor deposits infiltrating the brain parenchyma in an invasive xenograft model of glioblastoma.

    Science.gov (United States)

    Miao, Hongsheng; Choi, Bryan D; Suryadevara, Carter M; Sanchez-Perez, Luis; Yang, Shicheng; De Leon, Gabriel; Sayour, Elias J; McLendon, Roger; Herndon, James E; Healy, Patrick; Archer, Gary E; Bigner, Darell D; Johnson, Laura A; Sampson, John H

    2014-01-01

    Glioblastoma (GBM) is the most common primary malignant brain tumor in adults and is uniformly lethal. T-cell-based immunotherapy offers a promising platform for treatment given its potential to specifically target tumor tissue while sparing the normal brain. However, the diffuse and infiltrative nature of these tumors in the brain parenchyma may pose an exceptional hurdle to successful immunotherapy in patients. Areas of invasive tumor are thought to reside behind an intact blood brain barrier, isolating them from effective immunosurveillance and thereby predisposing the development of "immunologically silent" tumor peninsulas. Therefore, it remains unclear if adoptively transferred T cells can migrate to and mediate regression in areas of invasive GBM. One barrier has been the lack of a preclinical mouse model that accurately recapitulates the growth patterns of human GBM in vivo. Here, we demonstrate that D-270 MG xenografts exhibit the classical features of GBM and produce the diffuse and invasive tumors seen in patients. Using this model, we designed experiments to assess whether T cells expressing third-generation chimeric antigen receptors (CARs) targeting the tumor-specific mutation of the epidermal growth factor receptor, EGFRvIII, would localize to and treat invasive intracerebral GBM. EGFRvIII-targeted CAR (EGFRvIII+ CAR) T cells demonstrated in vitro EGFRvIII antigen-specific recognition and reactivity to the D-270 MG cell line, which naturally expresses EGFRvIII. Moreover, when administered systemically, EGFRvIII+ CAR T cells localized to areas of invasive tumor, suppressed tumor growth, and enhanced survival of mice with established intracranial D-270 MG tumors. Together, these data demonstrate that systemically administered T cells are capable of migrating to the invasive edges of GBM to mediate antitumor efficacy and tumor regression.

  2. srGAP1 mediates the migration inhibition effect of Slit2-Robo1 in colorectal cancer

    OpenAIRE

    Feng, Yuyang; Feng, Lei; Yu, Di; Zou, Jian; Huang, Zhaohui

    2016-01-01

    Background The neuronal guidance molecule Slit2 plays suppressive role in tumorigenesis and progression. We previously showed that Slit2-Robo1 inhibit cell migration in colorectal cancer (CRC). However, little is known about its downstream effectors in CRC. This study tries to identify whether the Slit-Robo Rho GTPase activating protein 1 (srGAP1) could mediate the inhibitory effect of Slit2-Robo1 on CRC cell migration. Methods The protein expression of srGAP1 in clinical CRC tissues was test...

  3. Inhibition of pattern recognition receptor-mediated inflammation by bioactive phytochemicals

    Science.gov (United States)

    Emerging evidence reveals that pattern-recognition receptors (PRRs), Toll-like receptors (TLRs) and Nucleotide-binding oligomerization domain proteins (NODs) mediate both infection-induced and sterile inflammation by recognizing pathogen-associated molecular patterns (PAMPs) and endogenous molecules...

  4. Notch signaling mediates granulocyte-macrophage colony-stimulating factor priming-induced transendothelial migration of human eosinophils.

    Science.gov (United States)

    Liu, L Y; Wang, H; Xenakis, J J; Spencer, L A

    2015-07-01

    Priming with cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) enhances eosinophil migration and exacerbates the excessive accumulation of eosinophils within the bronchial mucosa of asthmatics. However, mechanisms that drive GM-CSF priming are incompletely understood. Notch signaling is an evolutionarily conserved pathway that regulates cellular processes, including migration, by integrating exogenous and cell-intrinsic cues. This study investigates the hypothesis that the priming-induced enhanced migration of human eosinophils requires the Notch signaling pathway. Using pan Notch inhibitors and newly developed human antibodies that specifically neutralize Notch receptor 1 activation, we investigated a role for Notch signaling in GM-CSF-primed transmigration of human blood eosinophils in vitro and in the airway accumulation of mouse eosinophils in vivo. Notch receptor 1 was constitutively active in freshly isolated human blood eosinophils, and inhibition of Notch signaling or specific blockade of Notch receptor 1 activation during GM-CSF priming impaired priming-enhanced eosinophil transendothelial migration in vitro. Inclusion of Notch signaling inhibitors during priming was associated with diminished ERK phosphorylation, and ERK-MAPK activation was required for GM-CSF priming-induced transmigration. In vivo in mice, eosinophil accumulation within allergic airways was impaired following systemic treatment with Notch inhibitor, or adoptive transfer of eosinophils treated ex vivo with Notch inhibitor. These data identify Notch signaling as an intrinsic pathway central to GM-CSF priming-induced eosinophil tissue migration. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  5. Orexin receptor-1 mediates brown fat developmental differentiation

    OpenAIRE

    Sellayah, Dyan; Sikder, Devanjan

    2012-01-01

    Orexin A (OX) is a small excitatory neuropeptide hormone that stimulates feeding, wakefulness and energy expenditure via a pair of G-coupled protein receptors, namely orexin receptor-1 (OXR1) and orexin receptor-2 (OXR2). OX-deficient mice are sensitive to obesity despite being hypophagic. The obesogenic effect of OX-deletion is due to brown adipose tissue (BAT) dysfunction, a defect that originates during fetal growth. Brown preadipocytes in OX-null mice display undifferentiated histological...

  6. Receptor mediated agglutination of human spermatozoa by spermagglutinating factor isolated from Staphylococcus aureus.

    Science.gov (United States)

    Kaur, Siftjit; Prabha, Vijay; Sarwal, Abha

    2010-12-01

    We examined spermagglutinating factor isolated from Staphylococcus aureus for evidence of receptor mediated agglutination of human spermatozoa. Binding to spermatozoa by spermagglutinating factor isolated from S. aureus with a high degree of specificity indicates receptor-ligand interaction. To examine this interaction we isolated and purified the ligand and the receptor. To assess receptor mediated agglutination of spermatozoa further we blocked spermagglutination induced by spermagglutinating factor in the presence of receptor. Spermagglutinating factor induced spermagglutination was competitively inhibited by adding purified receptor, indicating that sperm agglutinating factor isolated from S. aureus attaches to specific receptors on human spermatozoa. The spermagglutinating factor receptor was a protein with a molecular weight of approximately 57 kDa. Spermagglutinating factor induced spermagglutination and at higher concentrations had a spermicidal effect, which was inhibited by introducing the receptor. As observed on scanning electron microscopy studies, incubating spermatozoa with spermagglutinating factor showed profound morphological alterations. However, spermatozoa with normal morphology were noted when incubated with spermagglutinating factor in the presence of receptor, indicating that morphological alterations may account for spermatozoa agglutination by spermagglutinating factor. Results suggest that spermagglutinating factor isolated from S. aureus may bind specifically to sperm surface receptor sites before causing spermagglutination. Copyright © 2010 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  7. Menthol enhances phasic and tonic GABAA receptor-mediated currents in midbrain periaqueductal grey neurons

    Science.gov (United States)

    Lau, Benjamin K; Karim, Shafinaz; Goodchild, Ann K; Vaughan, Christopher W; Drew, Geoffrey M

    2014-01-01

    Background and Purpose Menthol, a naturally occurring compound in the essential oil of mint leaves, is used for its medicinal, sensory and fragrant properties. Menthol acts via transient receptor potential (TRPM8 and TRPA1) channels and as a positive allosteric modulator of recombinant GABAA receptors. Here, we examined the actions of menthol on GABAA receptor-mediated currents in intact midbrain slices. Experimental Approach Whole-cell voltage-clamp recordings were made from periaqueductal grey (PAG) neurons in midbrain slices from rats to determine the effects of menthol on GABAA receptor-mediated phasic IPSCs and tonic currents. Key Results Menthol (150–750 μM) produced a concentration-dependent prolongation of spontaneous GABAA receptor-mediated IPSCs, but not non-NMDA receptor-mediated EPSCs throughout the PAG. Menthol actions were unaffected by TRPM8 and TRPA1 antagonists, tetrodotoxin and the benzodiazepine antagonist, flumazenil. Menthol also enhanced a tonic current, which was sensitive to the GABAA receptor antagonists, picrotoxin (100 μM), bicuculline (30 μM) and Zn2+ (100 μM), but unaffected by gabazine (10 μM) and a GABAC receptor antagonist, 1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid hydrate (TPMPA; 50 μM). In addition, menthol potentiated currents induced by the extrasynaptic GABAA receptor agonist THIP/gaboxadol (10 μM). Conclusions and Implications These results suggest that menthol positively modulates both synaptic and extrasynaptic populations of GABAA receptors in native PAG neurons. The development of agents that potentiate GABAA-mediated tonic currents and phasic IPSCs in a manner similar to menthol could provide a basis for novel GABAA-related pharmacotherapies. PMID:24460753

  8. Chemically primed bone-marrow derived mesenchymal stem cells show enhanced expression of chemokine receptors contributed to their migration capability

    Directory of Open Access Journals (Sweden)

    Hamid Reza Bidkhori

    2016-01-01

    Full Text Available Objective(s:The limited homing potential of bone-marrow-derived mesenchymal stem cells (BM-MSC is the key obstacle in MSC-based therapy. It is believed that chemokines and chemokine receptor interactions play key roles in cellular processes associated with migration. Meanwhile, MSCs express a low level of distinct chemokine receptors and they even lose these receptors on their surface after a few passages which influence their therapeutic applications negatively. This study investigated whether treatment of BM-MSCs with hypoxia-mimicking agents would increase expression of some chemokine receptors and cell migration. Materials and Methods: BM-MSCs were treated at passage 2 for our gene expression profiling. All qPCR experiments were performed by SYBR Green method in CFX-96 Bio-Rad Real-Time PCR. The Boyden chamber assay was utilized to investigate BM-MSC homing. Results:Possible approaches to increasing the expression level of chemokine receptors by different hypoxia-mimicking agents such as valproic acid (VPA, CoCl2, and desferrioxamine (DFX are described. Results show DFX efficiently up-regulate the CXCR7 and CXCR4 gene expression while VPA increase only the CXCR7 gene expression and no significant change in expression level of CXCR4 and the CXCR7 gene was detectable by CoCl2 treatment. Chemotaxis assay results show that pre-treatment with DFX, VPA, and Cocl2 enhances significantly the migration ability of BM-MSCs compared with the untreated control group and DFX treatment accelerates MSCs homing significantly with a higher rate than VPA and Cocl2 treatments. Conclusion: Our data supports the notion that pretreatment of MSC with VPA and DFX improves the efficiency of MSC therapy by triggering homing regulatory signaling pathways.

  9. Activation of sphingosine kinase by muscarinic receptors enhances NO-mediated and attenuates EDHF-mediated vasorelaxation

    NARCIS (Netherlands)

    Mulders, Arthur C. M.; Mathy, Marie-Jeanne; Meyer Zu Heringdorf, Dagmar; ter Braak, Michael; Hajji, Najat; Olthof, Dominique C.; Michel, Martin C.; Alewijnse, Astrid E.; Peters, Stephan L. M.

    2009-01-01

    Local formation of the sphingomyelin metabolite sphingosine-1-phosphate (S1P) within the vascular wall has been shown to modulate vascular reactivity. In this study we investigated whether sphingosine kinase, the enzyme responsible for S1P synthesis, plays a role in muscarinic receptor-mediated NO

  10. The binding of NCAM to FGFR1 induces a specific cellular response mediated by receptor trafficking

    DEFF Research Database (Denmark)

    Francavilla, Chiara; Cattaneo, Paola; Berezin, Vladimir

    2009-01-01

    different from that elicited by FGF-2. In contrast to FGF-induced degradation of endocytic FGFR1, NCAM promotes the stabilization of the receptor, which is recycled to the cell surface in a Rab11- and Src-dependent manner. In turn, FGFR1 recycling is required for NCAM-induced sustained activation of various...... effectors. Furthermore, NCAM, but not FGF-2, promotes cell migration, and this response depends on FGFR1 recycling and sustained Src activation. Our results implicate NCAM as a nonconventional ligand for FGFR1 that exerts a peculiar control on the intracellular trafficking of the receptor, resulting...... in a specific cellular response. Besides introducing a further level of complexity in the regulation of FGFR1 function, our findings highlight the link of FGFR recycling with sustained signaling and cell migration and the critical role of these events in dictating the cellular response evoked by receptor...

  11. Expression and function of CB1 receptor in the rat striatum: localization and effects on D1 and D2 dopamine receptor-mediated motor behaviors.

    Science.gov (United States)

    Martín, Ana Belén; Fernandez-Espejo, Emilio; Ferrer, Belén; Gorriti, Miguel Angel; Bilbao, Ainhoa; Navarro, Miguel; Rodriguez de Fonseca, Fernando; Moratalla, Rosario

    2008-06-01

    Cannabinoid CB1 receptors are densely expressed on striatal projection neurons expressing dopamine D1 or D2 receptors. However, the specific neuronal distribution of CB1 receptors within the striatum is not known. Previous research has established that the endocannabinoid system controls facilitation of behavior by dopamine D2 receptors, but it is not clear if endocannabinoids also modulate D1 receptor-mediated motor behavior. In the present study, we show that cannabinoid CB1 receptor mRNA is present in striatonigral neurons expressing substance P and dopamine D1 receptors, as well as in striatopallidal neurons expressing enkephalin and dopamine D2 receptors. We explored the functional relevance of the interaction between dopamine D1 and D2 receptors and cannabinoid CB1 receptors with behavioral pharmacology experiments. Potentiation of endogenous cannabinoid signaling by the uptake blocker AM404 blocked dopamine D1 receptor-mediated grooming and D2 receptor-mediated oral stereotypies. In addition, contralateral turning induced by unilateral intrastriatal infusion of D1 receptor agonists is counteracted by AM404 and potentiated by the cannabinoid antagonist SR141716A. These results indicate that the endocannabinoid system negatively modulates D1 receptor-mediated behaviors in addition to its previously described effect on dopamine D2 receptor-mediated behaviors. The effect of AM404 on grooming behavior was absent in dopamine D1 receptor knockout mice, demonstrating its dependence on D1 receptors. This study indicates that the endocannabinoid system is a relevant negative modulator of both dopamine D1 and D2 receptor-mediated behaviors, a finding that may contribute to our understanding of basal ganglia motor disorders.

  12. Regulation of C-Type Lectin Receptor-Mediated Antifungal Immunity

    Directory of Open Access Journals (Sweden)

    Juan Tang

    2018-02-01

    Full Text Available Of all the pathogen recognition receptor families, C-type lectin receptor (CLR-induced intracellular signal cascades are indispensable for the initiation and regulation of antifungal immunity. Ongoing experiments over the last decade have elicited diverse CLR functions and novel regulatory mechanisms of CLR-mediated-signaling pathways. In this review, we highlight novel insights in antifungal innate and adaptive-protective immunity mediated by CLRs and discuss the potential therapeutic strategies against fungal infection based on targeting the mediators in the host immune system.

  13. Ethanol inhibits epileptiform activity and NMDA receptor-mediated synaptic transmission in rat amygdaloid slices

    Energy Technology Data Exchange (ETDEWEB)

    Gean, P.W. (Cheng-Kung Univ., Tainan (Taiwan))

    1992-02-26

    The effect of ethanol on the epileptiform activity induced by Mg{sup ++}-free solution was studied in rat amygdalar slices using intracellular recording techniques. The spontaneous and evoked epileptiform discharges consisting of an initial burst followed by afterdischarges were observed 20-30 min after switching to Mg{sup ++}-free medium. Superfusion with ethanol reversibly reduced the duration of spontaneous and evoked bursting discharges in a concentration-dependent manner. Synaptic response mediated by N-methyl-D-aspartate (NMDA) receptor activation was isolated by application of a solution containing the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and either in Mg{sup ++}-free solution or in the presence of 50 {mu}M bicuculline. Application of ethanol reversibly suppressed the duration of NMDA receptor-mediated synaptic response. These results suggest that intoxicating concentrations of ethanol possess anticonvulsant activity through blocking the NMDA receptor-mediated synaptic excitation.

  14. A Collapsin Response Mediator Protein 2 Isoform Controls Myosin II-Mediated Cell Migration and Matrix Assembly by Trapping ROCK II

    Science.gov (United States)

    Morgan-Fisher, Marie; Wait, Robin; Couchman, John R.; Wewer, Ulla M.

    2012-01-01

    Collapsin response mediator protein 2 (CRMP-2) is known as a regulator of neuronal polarity and differentiation through microtubule assembly and trafficking. Here, we show that CRMP-2 is ubiquitously expressed and a splice variant (CRMP-2L), which is expressed mainly in epithelial cells among nonneuronal cells, regulates myosin II-mediated cellular functions, including cell migration. While the CRMP-2 short form (CRMP-2S) is recognized as a substrate of the Rho-GTP downstream kinase ROCK in neuronal cells, a CRMP-2 complex containing 2L not only bound the catalytic domain of ROCK II through two binding domains but also trapped and inhibited the kinase. CRMP-2L protein levels profoundly affected haptotactic migration and the actin-myosin cytoskeleton of carcinoma cells as well as nontransformed epithelial cell migration in a ROCK activity-dependent manner. Moreover, the ectopic expression of CRMP-2L but not -2S inhibited fibronectin matrix assembly in fibroblasts. Underlying these responses, CRMP-2L regulated the kinase activity of ROCK II but not ROCK I, independent of GTP-RhoA levels. This study provides a new insight into CRMP-2 as a controller of myosin II-mediated cellular functions through the inhibition of ROCK II in nonneuronal cells. PMID:22431514

  15. Thyroid hormone receptor inhibits hepatoma cell migration through transcriptional activation of Dickkopf 4

    Energy Technology Data Exchange (ETDEWEB)

    Chi, Hsiang-Cheng; Liao, Chen-Hsin [Department of Biochemistry, School of Medicine, Chang-Gung University, Taoyuan 333, Taiwan, ROC (China); Huang, Ya-Hui [Medical Research Central, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan, ROC (China); Wu, Sheng-Ming; Tsai, Chung-Ying; Liao, Chia-Jung; Tseng, Yi-Hsin; Lin, Yang-Hsiang; Chen, Cheng-Yi; Chung, I-Hsiao; Wu, Tzu-I [Department of Biochemistry, School of Medicine, Chang-Gung University, Taoyuan 333, Taiwan, ROC (China); Chen, Wei-Jan [First Cardiovascular Division, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan, ROC (China); Lin, Kwang-Huei, E-mail: khlin@mail.cgu.edu.tw [Department of Biochemistry, School of Medicine, Chang-Gung University, Taoyuan 333, Taiwan, ROC (China)

    2013-09-13

    Highlights: •T{sub 3} affects DKK4 mRNA and protein expression in HepG2-TR cells. •Regulation of DKK4 by T{sub 3} is at transcriptional level. •DKK4 overexpression suppresses hepatoma cell metastasis. -- Abstract: Triiodothyronine (T{sub 3}) is a potent form of thyroid hormone mediates several physiological processes including cellular growth, development, and differentiation via binding to the nuclear thyroid hormone receptor (TR). Recent studies have demonstrated critical roles of T{sub 3}/TR in tumor progression. Moreover, long-term hypothyroidism appears to be associated with the incidence of human hepatocellular carcinoma (HCC), independent of other major HCC risk factors. Dickkopf (DKK) 4, a secreted protein that antagonizes the canonical Wnt signaling pathway, is induced by T{sub 3} at both mRNA and protein levels in HCC cell lines. However, the mechanism underlying T{sub 3}-mediated regulation of DKK4 remains unknown. In the present study, the 5′ promoter region of DKK4 was serially deleted, and the reporter assay performed to localize the T{sub 3} response element (TRE). Consequently, we identified an atypical direct repeat TRE between nucleotides −1645 and −1629 conferring T{sub 3} responsiveness to the DKK4 gene. This region was further validated using chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA). Stable DKK4 overexpression in SK-Hep-1 cells suppressed cell invasion and metastatic potential, both in vivo andin vitro, via reduction of matrix metalloproteinase-2 (MMP-2) expression. Our findings collectively suggest that DKK4 upregulated by T{sub 3}/TR antagonizes the Wnt signal pathway to suppress tumor cell progression, thus providing new insights into the molecular mechanism underlying thyroid hormone activity in HCC.

  16. Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Yuan, E-mail: yuan.xu@ki.se; Cardell, Lars-Olaf

    2014-02-15

    Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B{sub 2} receptor agonist) and des-Arg{sup 9}-bradykinin- (selective B{sub 1} receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE{sub 2}. The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg{sup 9}-bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B{sub 2} receptors, but not those on B{sub 1}. Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in

  17. Effect of TNF-like weak inducer of apoptosis and its receptor on migration of hepatic stellate cells

    Directory of Open Access Journals (Sweden)

    SU Min

    2018-01-01

    Full Text Available Objective To investigate the effect of TNF-like weak inducer of apoptosis (TWAEK and its receptor fibroblast growth factor-inducible 14 (Fn14 on the migration of hepatic stellate cells and the possible mechanism. Methods The human hepatic stellate cell line LX-2 cells were treated with TWEAK or Fn14 specific small interfering RNA (Fn14 siRNA+TWEAK. Transwell chamber was used to observe the migration of hepatic stellate cells, and real-time PCR and Western blot were used to measure the expression of matrix metalloproteinase-9 (MMP9. The independent samples t-test was used for comparison of continuous data between two groups; a one-way analysis of variance was used for comparison between multiple groups, and the least significant difference t-test was used for further comparison between two groups. Results Compared with normal LX-2 cells, the TWEAK group had a significant increase in the migration of LX-2 cells (105±8 vs 164±17, t=5.287,P<0.01, and compared with the negative control group, the Fn14 siRNA+TWEAK group had a significant reduction in the number of migrated cells (122±9 vs 58±7, t=9.836, P<0.01. When LX-2 cells were treated with TWEAK, the mRNA and protein expression of MMP9 increased in a time-dependent manner (both P<0.05, while the Fn14 siRNA+TWEAK group had significant reductions in the mRNA and protein expression of MMP9 compared with the TWEAK group (t=5.358, P<0.01. Conclusion TWEAK and its receptor Fn14 can promote the migration of hepatic stellate cells by upregulating MMP9, and blockade of this pathway may become a potential target for the treatment of liver fibrosis.

  18. Cellular mechanisms of the 5-HT7 receptor-mediated signaling

    Directory of Open Access Journals (Sweden)

    Daria eGuseva

    2014-10-01

    Full Text Available Serotonin (5-hydroxytryptamine or 5-HT is an important neurotransmitter regulating a wide range of physiological and pathological functions via activation of heterogeneously expressed 5-HT receptors. The 5-HT7 receptor is one of the most recently described members of the 5-HT receptor family. Functionally, 5-HT7 receptor is associated with a number of physiological and pathological responses, including serotonin-induced phase shifting of the circadian rhythm, control of memory as well as locomotor and exploratory activity. A large body of evidence indicates involvement of the 5-HT7 receptor in anxiety and depression, and recent studies suggest that 5-HT7 receptor can be highly relevant for the treatment of major depressive disorders. The 5-HT7 receptor is coupled to the stimulatory Gs-protein, and receptor stimulation results in activation of adenylyl cyclase (AC leading to a rise of cAMP concentration. In addition, this receptor is coupled to the G12-protein to activate small GTPases of the Rho family. This review focuses on molecular mechanisms responsible for the 5-HT7 receptor-mediated signaling. We provide detailed overview of signaling cascades controlled and regulated by the 5-HT7 receptor and discuss the functional impact of 5-HT7 receptor for the regulation of different cellular and subcellular processes.

  19. Netrin-1 Mediates Neuronal Survival Through PIKE-L Interaction With the Dependence Receptor UNC5B

    Science.gov (United States)

    Tang, Xiaoling; Jang, Sung-Wuk; Okada, Masashi; Chan, Chi-Bun; Feng, Yue; Liu, Yu; Luo, Shi-Wen; Hong, Yan; Rama, Nicolas; Xiong, Wen-cheng; Mehlen, Patrick; Ye, Keqiang

    2010-01-01

    Netrins, a family of secreted molecules, play critical roles in axon guidance and cell migration during neuronal development 1,2. In addition to its role as a chemotropic molecule, netrin-1 also acts as a survival factor 3–7. Both UNC5 (i.e. UNC5A, B, C or D) and DCC are transmembrane receptors for netrin-18,9. In the absence of netrin-1, DCC and UNC5 act as dependence receptors and trigger apoptosis 3,6,10. However, how netrin-1 suppresses the apoptotic activity of the receptors remains elusive. Here, we show that netrin-1 induces interaction of UNC5B with the brain specific GTPase PIKE-L. This interaction triggers activation of PI 3-kinase signaling, prevents UNC5B’s pro-apoptotic activity and enhances neuronal survival. Moreover, this process tightly relies on Fyn as PIKE-L is tyrosine phosphorylated in response to netrin-1 and the netrin-1-mediated interaction of UNC5B with PIKE-L is inhibited in Fyn null mice. Thus, PIKE-L acts as a downstream survival effector for netrin-1 through UNC5B in the nervous system. PMID:18469807

  20. M2-like macrophages are responsible for collagen degradation through a mannose receptor-mediated pathway

    DEFF Research Database (Denmark)

    Madsen, Daniel H; Leonard, Daniel; Masedunskas, Andrius

    2013-01-01

    Tissue remodeling processes critically depend on the timely removal and remodeling of preexisting collagen scaffolds. Nevertheless, many aspects related to the turnover of this abundant extracellular matrix component in vivo are still incompletely understood. We therefore took advantage of recent...... of the collagen receptors mannose receptor (Mrc1) and urokinase plasminogen activator receptor-associated protein (Endo180 and Mrc2) impaired this intracellular collagen degradation pathway. This study demonstrates the importance of receptor-mediated cellular uptake to collagen turnover in vivo and identifies...

  1. Aquaporin-8 mediates human esophageal cancer Eca-109 cell migration via the EGFR-Erk1/2 pathway.

    Science.gov (United States)

    Chang, Heng; Shi, Yong-Hua; Talaf, Tuo-Kan; Lin, Chen

    2014-01-01

    Abnormal expression of aquaporins (AQPs) has been reported in several human cancers. Epidermal growth factor receptor (EGFR)-extracellular signal-regulated kinases1/2 (ERK1/2) are associated with tumorigenesis and cancer progression and may upregulate AQPs expression. In this study, we investigated acquaporin-8 expression and signaling via epidermal growth factor receptor-extracellular signal-regulated kinases1/2 in human esophageal cancer Eca-109 cells by western blot, immunofluorescence and wound healing (scratch) assays. Our results showed that epidermal growth factor (EGF) induced both Eca-109 migration and AQP8 expression. Wound healing results showed that cell migration was increased by 1.23-1.10-fold at 24 h and 48 h after EGF treatment. AQP8 expression was significantly increased (1.19-fold) at 48 h after EGF treatment in Eca-109. The EGFR kinase inhibitor, PD153035, blocked EGF-induced AQP8 expression and cell migration. AQP8 expression was decreased from 3.65-fold (EGF-treated) to 0.55-fold (PD153035-treated) in Eca-109. Furthermore, the MEK [MAPK (mitogen-activated protein kinase)/Erk1/2]/Erk1/2 inhibitor U0126 also inhibited EGF-induced AQP8 expression and cell migration. AQP8 expression was decreased from 3.92-fold (EGF-treated) to 1.38-fold (U0126-treated) in Eca-109. In conclusions, EGF induces AQP8 expression and cell migration in Eca-109 cells via the EGFR/Erk1/2 signal transduction pathway.

  2. CSF-1 receptor signalling is governed by pre-requisite EHD1 mediated receptor display on the macrophage cell surface.

    Science.gov (United States)

    Cypher, Luke R; Bielecki, Timothy Alan; Huang, Lu; An, Wei; Iseka, Fany; Tom, Eric; Storck, Matthew D; Hoppe, Adam D; Band, Vimla; Band, Hamid

    2016-09-01

    Colony stimulating factor-1 receptor (CSF-1R), a receptor tyrosine kinase (RTK), is the master regulator of macrophage biology. CSF-1 can bind CSF-1R resulting in receptor activation and signalling essential for macrophage functions such as proliferation, differentiation, survival, polarization, phagocytosis, cytokine secretion, and motility. CSF-1R activation can only occur after the receptor is presented on the macrophage cell surface. This process is reliant upon the underlying macrophage receptor trafficking machinery. However, the mechanistic details governing this process are incompletely understood. C-terminal Eps15 Homology Domain-containing (EHD) proteins have recently emerged as key regulators of receptor trafficking but have not yet been studied in the context of macrophage CSF-1R signalling. In this manuscript, we utilize primary bone-marrow derived macrophages (BMDMs) to reveal a novel function of EHD1 as a regulator of CSF-1R abundance on the cell surface. We report that EHD1-knockout (EHD1-KO) macrophages cell surface and total CSF-1R levels are significantly decreased. The decline in CSF-1R levels corresponds with reduced downstream macrophage functions such as cell proliferation, migration, and spreading. In EHD1-KO macrophages, transport of newly synthesized CSF-1R to the macrophage cell surface was reduced and was associated with the shunting of the receptor to the lysosome, which resulted in receptor degradation. These findings reveal a novel and functionally important role for EHD1 in governing CSF-1R signalling via regulation of anterograde transport of CSF-1R to the macrophage cell surface. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. G Protein-Coupled Receptors: Extranuclear Mediators for the Non-Genomic Actions of Steroids

    Directory of Open Access Journals (Sweden)

    Chen Wang

    2014-09-01

    Full Text Available Steroids hormones possess two distinct actions, a delayed genomic effect and a rapid non-genomic effect. Rapid steroid-triggered signaling is mediated by specific receptors localized most often to the plasma membrane. The nature of these receptors is of great interest and accumulated data suggest that G protein-coupled receptors (GPCRs are appealing candidates. Increasing evidence regarding the interaction between steroids and specific membrane proteins, as well as the involvement of G protein and corresponding downstream signaling, have led to identification of physiologically relevant GPCRs as steroid extranuclear receptors. Examples include G protein-coupled receptor 30 (GPR30 for estrogen, membrane progestin receptor for progesterone, G protein-coupled receptor family C group 6 member A (GPRC6A and zinc transporter member 9 (ZIP9 for androgen, and trace amine associated receptor 1 (TAAR1 for thyroid hormone. These receptor-mediated biological effects have been extended to reproductive development, cardiovascular function, neuroendocrinology and cancer pathophysiology. However, although great progress have been achieved, there are still important questions that need to be answered, including the identities of GPCRs responsible for the remaining steroids (e.g., glucocorticoid, the structural basis of steroids and GPCRs’ interaction and the integration of extranuclear and nuclear signaling to the final physiological function. Here, we reviewed the several significant developments in this field and highlighted a hypothesis that attempts to explain the general interaction between steroids and GPCRs.

  4. Migration of dendritic cell based cancer vaccines: in vivo veritas?

    NARCIS (Netherlands)

    Adema, Gosse J.; de Vries, I. Jolanda M.; Punt, Cornelis J. A.; Figdor, Carl G.

    2005-01-01

    Ex vivo generated cancer vaccines based on dendritic cells (DCs) are currently applied in the clinic. The migration of DCs from the tissues to the lymph nodes is tightly controlled and involves many different mediators and their receptors. A recent study demonstrated that the rate of migration of

  5. ApoE Receptor 2 Mediation of Trophoblast Dysfunction and Pregnancy Complications Induced by Antiphospholipid Antibodies in Mice.

    Science.gov (United States)

    Ulrich, Victoria; Gelber, Shari E; Vukelic, Milena; Sacharidou, Anastasia; Herz, Joachim; Urbanus, Rolf T; de Groot, Philip G; Natale, David R; Harihara, Anirudha; Redecha, Patricia; Abrahams, Vikki M; Shaul, Philip W; Salmon, Jane E; Mineo, Chieko

    2016-03-01

    Pregnancies in women with the antiphospholipid syndrome (APS) are frequently complicated by fetal loss and intrauterine growth restriction (IUGR). How circulating antiphospholipid antibodies (aPL) cause pregnancy complications in APS is poorly understood. We sought to determine whether the low-density lipoprotein receptor family member apolipoprotein E receptor 2 (ApoER2) mediates trophoblast dysfunction and pregnancy complications induced by aPL. Placental and trophoblast ApoER2 expression was evaluated by immunohistochemistry and immunoblotting. Normal human IgG and aPL were purified from healthy individuals and APS patients, respectively. The role of ApoER2 in aPL-induced changes in trophoblast proliferation and migration and in kinase activation was assessed using RNA interference in HTR-8/SVneo cells. The participation of ApoER2 in aPL-induced pregnancy loss and IUGR was evaluated in pregnant ApoER2(+/+) and ApoER2(-/-) mice injected with aPL or normal human IgG. We found that ApoER2 is abundant in human and mouse placental trophoblasts and in multiple trophoblast-derived cell lines, including HTR-8/SVneo cells. ApoER2 and its interaction with the cell surface protein β2 -glycoprotein I were required for aPL-induced inhibition of cultured trophoblast proliferation and migration. In parallel, aPL antagonism of Akt kinase activation by epidermal growth factor in trophoblasts was mediated by ApoER2. Furthermore, in a murine passive-transfer model of pregnancy complications of APS, ApoER2(-/-) mice were protected from both aPL-induced fetal loss and aPL-induced IUGR. ApoER2 plays a major role in the attenuation of trophoblast function by aPL, and the receptor mediates aPL-induced pregnancy complications in vivo in mice. ApoER2-directed interventions can now potentially be developed to combat the pregnancy complications associated with APS. © 2015, American College of Rheumatology.

  6. srGAP1 mediates the migration inhibition effect of Slit2-Robo1 in colorectal cancer.

    Science.gov (United States)

    Feng, Yuyang; Feng, Lei; Yu, Di; Zou, Jian; Huang, Zhaohui

    2016-12-07

    The neuronal guidance molecule Slit2 plays suppressive role in tumorigenesis and progression. We previously showed that Slit2-Robo1 inhibit cell migration in colorectal cancer (CRC). However, little is known about its downstream effectors in CRC. This study tries to identify whether the Slit-Robo Rho GTPase activating protein 1 (srGAP1) could mediate the inhibitory effect of Slit2-Robo1 on CRC cell migration. The protein expression of srGAP1 in clinical CRC tissues was tested by immunohistochemistry staining. Conditioned medium was prepared from HEK293 cells stably expressing Slit2-myc, Robo1-HA or RoboN (a soluble extracellular domain of Robo1). Immunoprecipitation (IP) was applied to check the interaction between Robo1 and srGAP1, and immunofluorescence (IF) was used to observe the subcellular localization of Robo1 and srGAP1. Small GTPase pull-down assay was used to determine the activity of Cdc42. A modified wound healing assay was performed to detect cell migration. The protein expression of srGAP1 was remarkably decreased in 47.5% of CRC tissues compared with adjacent noncancerous tissues, and the decreased srGAP1 expression was associated with lymphatic invasion, poor tumor differentiation, high TNM stage, and poor survival (P Robo1-interacting protein and exhibited similar dynamic subcellular distribution after Slit2 treatment in CRC cells. Small GTPase pull-down assay and migration assay indicated that Slit2-Robo1 signaling inhibited Cdc42 activity and CRC cell motility through srGAP1. Downregulation of srGAP1 in CRC was associated with tumor progression and poor prognosis. srGAP1 is an important downstream molecule of Slit2 signalling in CRC, and mediates the anti-migration function of Slit2 by inhibiting Cdc42.

  7. Blockade of nicotinic and muscarinic receptors facilitates spontaneous migration of human peripheral granulocytes: failure in cystic fibrosis.

    Science.gov (United States)

    Wessler, Ignaz; Neumann, Stefanie; Razen, Michael; Zepp, Fred; Kirkpatrick, Charles James

    2012-11-27

    Circulating leucocytes express muscarinic (m) and nicotinic (n) receptors and synthesize acetylcholine (ACh) regulating various cell functions. Leucocytes from patients with cystic fibrosis contain less ACh; therefore it was tested whether the regulation of cellular functions like migration differed from healthy volunteers. Peripheral blood (10-20 ml) was used, leucocytes were isolated by Ficoll® gradient and the commercial MIGRATEST® combined with flow cytometric analysis was applied (pore size 3 μm). In the absence of test substances 4900±1800 (n=10) leucocytes migrated within a time period of 2 h. In the presence of tubocurarine (TC, 30 μM) the cell number increased to 7500±2700 [n=10] corresponding to an increase of 162±20% (mean of individual experiments; pGranulocytes isolated from patients with cystic fibrosis did not respond (2h migration) to 30 μM TC (control: 5180±1400 cells [n=10]; TC: 5800±1400 [n=10]). Also in the presence of atropine (1 μM) and TC (30 μM) a significant effect was not detected (5800±1300 [n=10]). Auto-paracrine acetylcholine limits the migration of unstimulated peripheral granulocytes. This effect is impaired in cystic fibrosis most likely because of a reduced endogenous cholinergic tone. Copyright © 2012 Elsevier Inc. All rights reserved.

  8. Orexin receptor-1 mediates brown fat developmental differentiation

    Science.gov (United States)

    Sellayah, Dyan; Sikder, Devanjan

    2012-01-01

    Orexin A (OX) is a small excitatory neuropeptide hormone that stimulates feeding, wakefulness and energy expenditure via a pair of G-coupled protein receptors, namely orexin receptor-1 (OXR1) and orexin receptor-2 (OXR2). OX-deficient mice are sensitive to obesity despite being hypophagic. The obesogenic effect of OX-deletion is due to brown adipose tissue (BAT) dysfunction, a defect that originates during fetal growth. Brown preadipocytes in OX-null mice display undifferentiated histological appearance and fail to support both diet- and cold-induced thermogenesis. We show that the OXR1-null mice phenocopies the differentiation defect observed in the ligand-null mice indicating that OXR1 relays OX’s differentiation and thermogenic function. Consistent with this, OX fails to induce differentiation in cultured OXR1-null preadipocytes. Thus, OX signaling via OXR1 constitutes an important thermoregulatory mechanism that defends against cold and obesity. PMID:23700511

  9. Muscadine grape skin extract can antagonize Snail-cathepsin L-mediated invasion, migration and osteoclastogenesis in prostate and breast cancer cells.

    Science.gov (United States)

    Burton, Liza J; Smith, Basil A; Smith, Bethany N; Loyd, Quentin; Nagappan, Peri; McKeithen, Danielle; Wilder, Catera L; Platt, Manu O; Hudson, Tamaro; Odero-Marah, Valerie A

    2015-09-01

    To develop new and effective chemopreventive agents against bone metastasis, we assessed the effects of muscadine grape skin extract (MSKE), whose main bioactive component is anthocyanin, on bone turnover, using prostate and breast cancer cell models overexpressing Snail transcription factor. MSKE has been shown previously to promote apoptosis in prostate cancer cells without affecting normal prostate epithelial cells. Snail is overexpressed in prostate and breast cancer, and is associated with increased invasion, migration and bone turnover/osteoclastogenesis. Cathepsin L (CatL) is a cysteine cathepsin protease that is overexpressed in cancer and involved in bone turnover. Snail overexpression in prostate (LNCaP, ARCaP-E) and breast (MCF-7) cancer cells led to increased CatL expression/activity and phosphorylated STAT-3 (pSTAT-3), compared to Neo vector controls, while the reverse was observed in C4-2 (the aggressive subline of LNCaP) cells with Snail knockdown. Moreover, CatL expression was higher in prostate and breast tumor tissue compared to normal tissue. MSKE decreased Snail and pSTAT3 expression, and abrogated Snail-mediated CatL activity, migration and invasion. Additionally, Snail overexpression promoted osteoclastogenesis, which was significantly inhibited by the MSKE as effectively as Z-FY-CHO, a CatL-specific inhibitor, or osteoprotegerin, a receptor activator of nuclear factor kappa B ligand (RANKL) antagonist. Overall, these novel findings suggest that Snail regulation of CatL may occur via STAT-3 signaling and can be antagonized by MSKE, leading to decreased cell invasion, migration and bone turnover. Therefore, inhibition using a natural product such as MSKE could potentially be a promising bioactive compound for bone metastatic cancer. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  10. A novel monoclonal antibody to human laminin α5 chain strongly inhibits integrin-mediated cell adhesion and migration on laminins 511 and 521.

    Directory of Open Access Journals (Sweden)

    Zenebech Wondimu

    Full Text Available Laminins, a large family of αβγ heterotrimeric proteins mainly found in basement membranes, are strong promoters of adhesion and migration of multiple cell types, such as tumor and immune cells, via several integrin receptors. Among laminin α (LMα chains, α5 displays the widest tissue distribution in adult life and is synthesized by most cell types. Here, we have generated and characterized five novel monoclonal antibodies (mAbs to the human LMα5 chain to further study the biological relevance of α5 laminins, such as laminins 511 (α5β1γ1 and 521 (α5β2γ1. As detected by ELISA, immunohistochemistry, immunoprecipitation and Western blotting, each antibody displayed unique properties when compared to mAb 4C7, the prototype LMα5 antibody. Of greatest interest, mAb 8G9, but not any other antibody, strongly inhibited α3β1/α6β1 integrin-mediated adhesion and migration of glioma, melanoma, and carcinoma cells on laminin-511 and, together with mAb 4C7, on laminin-521. Accordingly, mAb 8G9 abolished the interaction of soluble α3β1 integrin with immobilized laminins 511 and 521. Binding of mAb 8G9 to laminin-511 was unaffected by the other mAbs to the LMα5 chain but largely hindered by mAb 4E10 to a LMβ1 chain epitope near the globular domain of laminin-511. Thus, mAb 8G9 defines a novel epitope localized at or near the integrin-binding globular domain of the LMα5 chain, which is essential for cell adhesion and migration, and identifies a potential therapeutic target in malignant and inflammatory diseases.

  11. Dynamics of the actin cytoskeleton mediates receptor cross talk: An emerging concept in tuning receptor signaling

    Science.gov (United States)

    Mattila, Pieta K.; Batista, Facundo D.

    2016-01-01

    Recent evidence implicates the actin cytoskeleton in the control of receptor signaling. This may be of particular importance in the context of immune receptors, such as the B cell receptor, where dysregulated signaling can result in autoimmunity and malignancy. Here, we discuss the role of the actin cytoskeleton in controlling receptor compartmentalization, dynamics, and clustering as a means to regulate receptor signaling through controlling the interactions with protein partners. We propose that the actin cytoskeleton is a point of integration for receptor cross talk through modulation of protein dynamics and clustering. We discuss the implication of this cross talk via the cytoskeleton for both ligand-induced and low-level constitutive (tonic) signaling necessary for immune cell survival. PMID:26833785

  12. Myosin-II-Mediated Directional Migration of Dictyostelium Cells in Response to Cyclic Stretching of Substratum

    Science.gov (United States)

    Iwadate, Yoshiaki; Okimura, Chika; Sato, Katsuya; Nakashima, Yuta; Tsujioka, Masatsune; Minami, Kazuyuki

    2013-01-01

    Living cells are constantly subjected to various mechanical stimulations, such as shear flow, osmotic pressure, and hardness of substratum. They must sense the mechanical aspects of their environment and respond appropriately for proper cell function. Cells adhering to substrata must receive and respond to mechanical stimuli from the substrata to decide their shape and/or migrating direction. In response to cyclic stretching of the elastic substratum, intracellular stress fibers in fibroblasts and endothelial, osteosarcoma, and smooth muscle cells are rearranged perpendicular to the stretching direction, and the shape of those cells becomes extended in this new direction. In the case of migrating Dictyostelium cells, cyclic stretching regulates the direction of migration, and not the shape, of the cell. The cells migrate in a direction perpendicular to that of the stretching. However, the molecular mechanisms that induce the directional migration remain unknown. Here, using a microstretching device, we recorded green fluorescent protein (GFP)-myosin-II dynamics in Dictyostelium cells on an elastic substratum under cyclic stretching. Repeated stretching induced myosin II localization equally on both stretching sides in the cells. Although myosin-II-null cells migrated randomly, myosin-II-null cells expressing a variant of myosin II that cannot hydrolyze ATP migrated perpendicular to the stretching. These results indicate that Dictyostelium cells accumulate myosin II at the portion of the cell where a large strain is received and migrate in a direction other than that of the portion where myosin II accumulated. This polarity generation for migration does not require the contraction of actomyosin. PMID:23442953

  13. Experimental Cannabinoid 2 Receptor-Mediated Immune Modulation in Sepsis

    Directory of Open Access Journals (Sweden)

    J. Sardinha

    2014-01-01

    Full Text Available Sepsis is a complex condition that results from a dysregulated immune system in response to a systemic infection. Current treatments lack effectiveness in reducing the incidence and mortality associated with this disease. The endocannabinoid system offers great promise in managing sepsis pathogenesis due to its unique characteristics. The present study explored the effect of modulating the CB2 receptor pathway in an acute sepsis mouse model. Endotoxemia was induced by intravenous injection of lipopolysaccharide (LPS in mice and intestinal microcirculation was assessed through intravital microscopy. We found that HU308 (CB2 receptor agonist reduced the number of adherent leukocytes in submucosal venules but did not restore muscular and mucosal villi FCD in endotoxemic mice. AM630 (CB2 receptor antagonist maintained the level of adherent leukocytes induced by LPS but further reduced muscular and mucosal villi FCD. URB597 (FAAH inhibitor and JZL184 (MAGL inhibitor both reduced the number of adherent leukocytes in submucosal venules but did not restore the mucosal villi FCD. Using various compounds we have shown different mechanisms of activating CB2 receptors to reduce leukocyte endothelial interactions in order to prevent further inflammatory damage during sepsis.

  14. Fas receptor-mediated apoptosis : a clinical application?

    NARCIS (Netherlands)

    Timmer, T; de Vries, EGE; de Jong, S

    Fas is a membrane protein belonging to the death receptor family. Cross-linking of Fas by its ligand, FasL, or agonistic anti-Fas antibodies, induces apoptosis of cells expressing Fas on the membrane by triggering a cascade of caspases. Since many different tumours express Fas on their membrane,

  15. Androgen Receptor-Mediated Escape Mechanism from Androgen Ablation Therapy

    Science.gov (United States)

    2008-10-31

    except that no salmon sperm DNA was used as blocking reagent. The immunoprecipitated DNA and un- enriched input DNA were treated with RNase A and purified...A, Wang Y, Suzuki K, Mirosevich J, et al. (2005) Foxa1 and Foxa2 interact with the androgen receptor to regulate prostate and epididymal genes

  16. Nuclear Receptor Cofactors in PPARγ-Mediated Adipogenesis and Adipocyte Energy Metabolism

    Directory of Open Access Journals (Sweden)

    Emily Powell

    2007-01-01

    Full Text Available Transcriptional cofactors are integral to the proper function and regulation of nuclear receptors. Members of the peroxisome proliferator-activated receptor (PPAR family of nuclear receptors are involved in the regulation of lipid and carbohydrate metabolism. They modulate gene transcription in response to a wide variety of ligands, a process that is mediated by transcriptional coactivators and corepressors. The mechanisms by which these cofactors mediate transcriptional regulation of nuclear receptor function are still being elucidated. The rapidly increasing array of cofactors has brought into focus the need for a clear understanding of how these cofactors interact in ligand- and cell-specific manners. This review highlights the differential effects of the assorted cofactors regulating the transcriptional action of PPARγ and summarizes the recent advances in understanding the physiological functions of corepressors and coactivators.

  17. Migration ability and Toll-like receptor expression of human mesenchymal stem cells improves significantly after three-dimensional culture.

    Science.gov (United States)

    Zhou, Panpan; Liu, Zilin; Li, Xue; Zhang, Bing; Wang, Xiaoyuan; Lan, Jing; Shi, Qing; Li, Dong; Ju, Xiuli

    2017-09-16

    While the conventional two-dimensional (2D) culture protocol is well accepted for the culture of mesenchymal stem cells (MSCs), this method fails to recapitulate the in vivo native three-dimensional (3D) cellular microenvironment, and may result in phenotypic changes, and homing and migration capacity impairments. MSC preparation in 3D culture systems has been considered an attractive preparatory and delivery method recently. We seeded human umbilical cord-derived MSCs (hUCMSCs) in a 3D culture system with porcine acellular dermal matrix (PADM), and investigated the phenotypic changes, the expression changes of some important receptors, including Toll-like receptors (TLRs) and C-X-C chemokine receptor type 4 (CXCR4) when hUCMSCs were transferred from 2D to 3D systems, as well as the alterations in in vivo homing and migration potential. It was found that the percentage of CD105-positive cells decreased significantly, whereas that of CD34- and CD271-positive cells increased significantly in 3D culture, compared to that in 2D culture. The mRNA and protein expression levels of TLR2, TLR3, TLR4, TLR6, and CXCR4 in hUCMSCs were increased significantly upon culturing with PADM for 3 days, compared to the levels in 2D culture. The numbers of migratory 3D hUCMSCs in the heart, liver, spleen, and bone marrow were significantly greater than the numbers of 2D hUCMSCs, and the worst migration occurred in 3D + AMD3100 (CXCR4 antagonist) hUCMSCs. These results suggested that 3D culture of hUCMSCs with PADM could alter the phenotypic characteristics of hUCMSCs, increase their TLR and CXCR4 expression levels, and promote their migratory and homing capacity in which CXCR4 plays an important role. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. (-)-Delta9-tetrahydrocannabinol antagonizes the peripheral cannabinoid receptor-mediated inhibition of adenylyl cyclase.

    Science.gov (United States)

    Bayewitch, M; Rhee, M H; Avidor-Reiss, T; Breuer, A; Mechoulam, R; Vogel, Z

    1996-04-26

    (-)-Delta9-Tetrahydrocannabinol ((-)-Delta9-THC) is the major active psychotropic component of the marijuana plant, Cannabis sativa. The membrane proteins that have been found to bind this material or its derivatives have been called the cannabinoid receptors. Two GTP-binding protein-coupled cannabinoid receptors have been cloned. CB1 or the neuronal cannabinoid receptor is found mostly in neuronal cells and tissues while CB2 or the peripheral cannabinoid receptor has been detected in spleen and in several cells of the immune system. It has previously been shown that activation of CB1 or CB2 receptors by cannabinoid agonists inhibits adenylyl cyclase activity. Utilizing Chinese hamster ovary cells and COS cells transfected with the cannabinoid receptors we report that (-)-Delta9-THC binds to both receptors with similar affinity. However, in contrast to its capacity to serve as an agonist for the CB1 receptor, (-)-Delta9-THC was only able to induce a very slight inhibition of adenylyl cyclase at the CB2 receptor. Morever, (-)-Delta9-THC antagonizes the agonist-induced inhibition of adenylyl cyclase mediated by CB2. Therefore, we conclude that (-)-Delta9-THC constitutes a weak antagonist for the CB2 receptor.

  19. Androgen receptor as a mediator and biomarker of radioresistance in triple-negative breast cancer.

    Science.gov (United States)

    Speers, Corey; Zhao, Shuang G; Chandler, Ben; Liu, Meilan; Wilder-Romans, Kari; Olsen, Eric; Nyati, Shyam; Ritter, Cassandra; Alluri, Prasanna G; Kothari, Vishal; Hayes, Daniel F; Lawrence, Theodore S; Spratt, Daniel E; Wahl, Daniel R; Pierce, Lori J; Feng, Felix Y

    2017-01-01

    receptor-negative triple-negative breast cancer or estrogen-receptor-positive, androgen receptor-negative breast cancer cell lines. androgen receptor inhibition with MDV3100 significantly radiosensitized triple-negative breast cancer xenografts in mouse models and markedly delayed tumor doubling/tripling time and tumor weight. Radiosensitization was at least partially dependent on impaired dsDNA break repair mediated by DNA protein kinase catalytic subunit. Our results implicate androgen receptor as a mediator of radioresistance in breast cancer and identify androgen receptor inhibition as a potentially effective strategy for the treatment of androgen receptor-positive radioresistant tumors.

  20. Glucocorticoid receptor-mediated induction of glutamine synthetase in skeletal muscle cells in vitro

    Science.gov (United States)

    Max, Stephen R.; Thomas, John W.; Banner, Carl; Vitkovic, Ljubisa; Konagaya, Masaaki

    1987-01-01

    The regulation by glucocorticoids of glutamine synthetase in L6 muscle cells in culture is studied. Glutamine synthetase activity was strikingly enhanced by dexamethasone. The dexamethasone-mediated induction of glutamine synthetase activity was blocked by RU38486, a glucocorticoid antagonist, indicating the involvement of intracellular glucocorticoid receptors in the induction process. RU38486 alone was without effect. Northern blot analysis revealed that dexamethasone-mediated enhancement of glutamine synthetase activity involves increased levels of glutamine synthetase mRNA. Glucocorticoids regulate the expression of glutamine synthetase mRNA in cultured muscle cells via interaction with intracellular receptors. Such regulation may be relevant to control of glutamine production by muscle.

  1. Stimulation of accumbal GABAA receptors inhibits delta2-, but not delta1-, opioid receptor-mediated dopamine efflux in the nucleus accumbens of freely moving rats.

    Science.gov (United States)

    Aono, Yuri; Kiguchi, Yuri; Watanabe, Yuriko; Waddington, John L; Saigusa, Tadashi

    2017-11-15

    The nucleus accumbens contains delta-opioid receptors that may reduce inhibitory neurotransmission. Reduction in GABAA receptor-mediated inhibition of accumbal dopamine release due to delta-opioid receptor activation should be suppressed by stimulating accumbal GABAA receptors. As delta-opioid receptors are divided into delta2- and delta1-opioid receptors, we analysed the effects of the GABAA receptor agonist muscimol on delta2- and delta1-opioid receptor-mediated accumbal dopamine efflux in freely moving rats using in vivo microdialysis. Drugs were administered intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 25-50min infusions. The delta2-opioid receptor agonist deltorphin II (25.0nmol)- and delta1-opioid receptor agonist DPDPE (5.0nmol)-induced increases in dopamine efflux were inhibited by the delta2-opioid receptor antagonist naltriben (1.5nmol) and the delta1-opioid receptor antagonist BNTX (150.0pmol), respectively. Muscimol (250.0pmol) inhibited deltorphin II (25.0nmol)-induced dopamine efflux. The GABAA receptor antagonist bicuculline (50.0pmol), which failed to affect deltorphin II (25.0nmol)-induced dopamine efflux, counteracted the inhibitory effect of muscimol on deltorphin II-induced dopamine efflux. Neither muscimol (250.0pmol) nor bicuculline (50.0 and 500.0pmol) altered DPDPE (5.0nmol)-induced dopamine efflux. The present results show that reduction in accumbal GABAA receptor-mediated inhibition of dopaminergic activity is necessary to produce delta2-opioid receptor-induced increase in accumbal dopamine efflux. This study indicates that activation of delta2- but not delta1-opioid receptors on the cell bodies and/or terminals of accumbal GABAergic interneurons inhibits GABA release and, accordingly, decreases GABAA receptor-mediated inhibition of dopaminergic terminals, resulting in enhanced accumbal dopamine efflux. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Comparison of lentiviral and sleeping beauty mediated αβ T cell receptor gene transfer.

    Science.gov (United States)

    Field, Anne-Christine; Vink, Conrad; Gabriel, Richard; Al-Subki, Roua; Schmidt, Manfred; Goulden, Nicholas; Stauss, Hans; Thrasher, Adrian; Morris, Emma; Qasim, Waseem

    2013-01-01

    Transfer of tumour antigen-specific receptors to T cells requires efficient delivery and integration of transgenes, and currently most clinical studies are using gamma retroviral or lentiviral systems. Whilst important proof-of-principle data has been generated for both chimeric antigen receptors and αβ T cell receptors, the current platforms are costly, time-consuming and relatively inflexible. Alternative, more cost-effective, Sleeping Beauty transposon-based plasmid systems could offer a pathway to accelerated clinical testing of a more diverse repertoire of recombinant high affinity T cell receptors. Nucleofection of hyperactive SB100X transposase-mediated stable transposition of an optimised murine-human chimeric T cell receptor specific for Wilm's tumour antigen from a Sleeping Beauty transposon plasmid. Whilst transfer efficiency was lower than that mediated by lentiviral transduction, cells could be readily enriched and expanded, and mediated effective target cells lysis in vitro and in vivo. Integration sites of transposed TCR genes in primary T cells were almost randomly distributed, contrasting the predilection of lentiviral vectors for transcriptionally active sites. The results support exploitation of the Sleeping Beauty plasmid based system as a flexible and adaptable platform for accelerated, early-phase assessment of T cell receptor gene therapies.

  3. Comparison of lentiviral and sleeping beauty mediated αβ T cell receptor gene transfer.

    Directory of Open Access Journals (Sweden)

    Anne-Christine Field

    Full Text Available Transfer of tumour antigen-specific receptors to T cells requires efficient delivery and integration of transgenes, and currently most clinical studies are using gamma retroviral or lentiviral systems. Whilst important proof-of-principle data has been generated for both chimeric antigen receptors and αβ T cell receptors, the current platforms are costly, time-consuming and relatively inflexible. Alternative, more cost-effective, Sleeping Beauty transposon-based plasmid systems could offer a pathway to accelerated clinical testing of a more diverse repertoire of recombinant high affinity T cell receptors. Nucleofection of hyperactive SB100X transposase-mediated stable transposition of an optimised murine-human chimeric T cell receptor specific for Wilm's tumour antigen from a Sleeping Beauty transposon plasmid. Whilst transfer efficiency was lower than that mediated by lentiviral transduction, cells could be readily enriched and expanded, and mediated effective target cells lysis in vitro and in vivo. Integration sites of transposed TCR genes in primary T cells were almost randomly distributed, contrasting the predilection of lentiviral vectors for transcriptionally active sites. The results support exploitation of the Sleeping Beauty plasmid based system as a flexible and adaptable platform for accelerated, early-phase assessment of T cell receptor gene therapies.

  4. Cyclic strain-mediated regulation of vascular endothelial cell migration and tube formation.

    Science.gov (United States)

    Von Offenberg Sweeney, Nicholas; Cummins, Philip M; Cotter, Eoin J; Fitzpatrick, Paul A; Birney, Yvonne A; Redmond, Eileen M; Cahill, Paul A

    2005-04-08

    Hemodynamic forces exerted by blood flow (cyclic strain, shear stress) affect the initiation and progression of angiogenesis; however, the precise signaling mechanism(s) involved are unknown. In this study, we examine the role of cyclic strain in regulating bovine aortic endothelial cell (BAEC) migration and tube formation, indices of angiogenesis. Considering their well-documented mechanosensitivity, functional inter-dependence, and involvement in angiogenesis, we hypothesized roles for matrix metalloproteinases (MMP-2/9), RGD-dependent integrins, and urokinase plasminogen activator (uPA) in this process. BAECs were exposed to equibiaxial cyclic strain (5% strain, 1Hz for 24h) before their migration and tube formation was assessed by transwell migration and collagen gel tube formation assays, respectively. In response to strain, both migration and tube formation were increased by 1.83+/-0.1- and 1.84+/-0.1-fold, respectively. Pertussis toxin, a Gi-protein inhibitor, decreased strain-induced migration by 45.7+/-32% and tube formation by 69.8+/-13%, whilst protein tyrosine kinase (PTK) inhibition with genistein had no effect. siRNA-directed attenuation of endothelial MMP-9 (but not MMP-2) expression/activity decreased strain-induced migration and tube formation by 98.6+/-41% and 40.7+/-31%, respectively. Finally, integrin blockade with cRGD peptide and siRNA-directed attenuation of uPA expression reduced strain-induced tube formation by 85.7+/-15% and 84.7+/-31%, respectively, whilst having no effect on migration. Cyclic strain promotes BAEC migration and tube formation in a Gi-protein-dependent PTK-independent manner. Moreover, we demonstrate for the first time a putative role for MMP-9 in both strain-induced events, whilst RGD-dependent integrins and uPA appear only to be involved in strain-induced tube formation.

  5. Suppression of Thyroid Hormone Receptor-Mediated Transcription ...

    African Journals Online (AJOL)

    The effects of methamidophos on cerebellar Purkinje cell dendritogenesis, granule cell neurite morphology and synaptic plasticity are currently under investigation. Taken together, our results show that methamidophos can potentially disrupt TR–mediated gene expression, suggesting that methamidophos may interfere with ...

  6. Nonmuscle myosin heavy chain IIA mediates integrin LFA-1 de-adhesion during T lymphocyte migration.

    Science.gov (United States)

    Morin, Nicole A; Oakes, Patrick W; Hyun, Young-Min; Lee, Dooyoung; Chin, Y Eugene; Chin, Eugene Y; King, Michael R; Springer, Timothy A; Shimaoka, Motomu; Tang, Jay X; Reichner, Jonathan S; Kim, Minsoo

    2008-01-21

    Precise spatial and temporal regulation of cell adhesion and de-adhesion is critical for dynamic lymphocyte migration. Although a great deal of information has been learned about integrin lymphocyte function-associated antigen (LFA)-1 adhesion, the mechanism that regulates efficient LFA-1 de-adhesion from intercellular adhesion molecule (ICAM)-1 during T lymphocyte migration is unknown. Here, we show that nonmuscle myosin heavy chain IIA (MyH9) is recruited to LFA-1 at the uropod of migrating T lymphocytes, and inhibition of the association of MyH9 with LFA-1 results in extreme uropod elongation, defective tail detachment, and decreased lymphocyte migration on ICAM-1, without affecting LFA-1 activation by chemokine CXCL-12. This defect was reversed by a small molecule antagonist that inhibits both LFA-1 affinity and avidity regulation, but not by an antagonist that inhibits only affinity regulation. Total internal reflection fluorescence microscopy of the contact zone between migrating T lymphocytes and ICAM-1 substrate revealed that inactive LFA-1 is selectively localized to the posterior of polarized T lymphocytes, whereas active LFA-1 is localized to their anterior. Thus, during T lymphocyte migration, uropodal adhesion depends on LFA-1 avidity, where MyH9 serves as a key mechanical link between LFA-1 and the cytoskeleton that is critical for LFA-1 de-adhesion.

  7. Role of amyloid β protein receptors in mediating synaptic plasticity.

    Science.gov (United States)

    Li, Yu; Sun, Zhongqing; Cao, Qiaoyu; Chen, Meiwan; Luo, Huanmin; Lin, Xi; Xiao, Fei

    2017-04-01

    There are few diseases in modern biomedicine that have garnered as much scientific interest and public concern as Alzheimer's disease (AD). The amyloid hypothesis has become the dominant model of AD pathogenesis; however, the details of the hypothesis are changing over time. Recently, given the increasing recognition, subtle effects of amyloid β protein (Aβ) on synaptic efficacy may be critical to AD progression. Synaptic plasticity is the important neurochemical foundation of learning and memory. Recent studies have identified that soluble Aβ oligomers combine with certain receptors to impair synaptic plasticity in AD, which advanced the amyloid hypothesis. The aim of the present review was to summarize the role of Aβ-relevant receptors in regulating synaptic plasticity and their downstream signaling cascades, which may provide novel insights into the understanding of the pathogenesis of AD and the development of therapeutic strategies to slow down the progression of AD-associated memory decline in the early stages.

  8. An histidine covalent receptor/butenolide complex mediates strigolactone perception

    Science.gov (United States)

    Badet-Denisot, Marie-Ange; Pillot, Jean-Paul; Cornu, David; Le Caer, Jean-Pierre; Burger, Marco; Pelissier, Frank; Retailleau, Pascal; Turnbull, Colin; Bonhomme, Sandrine; Chory, Joanne; Rameau, Catherine; Boyer, François-Didier

    2016-01-01

    Strigolactone plant hormones control plant architecture and are key players in both symbiotic and parasitic interactions. They contain an ABC tricyclic lactone connected to a butenolide group, the D-ring. The DWARF14 (D14) strigolactone receptor belongs to the superfamily of α/β-hydrolases and is known to hydrolyze the bond between the ABC lactone and the D-ring. Here we characterize the binding and catalytic functions of RAMOSUS3 (RMS3), the pea (Pisum sativum) ortholog of rice (Oryza sativa) D14 strigolactone receptor. Using novel profluorescent probes with strigolactone-like bioactivity, we show that RMS3 acts as a single-turnover enzyme that explains its apparent low enzymatic rate. We further demonstrate the formation of a covalent RMS3/D-ring complex, essential for bioactivity, in which the D-ring is attached to Histidine 247 of the catalytic triad. These results reveal an undescribed mechanism of plant hormone reception where the receptor performs an irreversible enzymatic reaction to generate its own ligand. PMID:27479744

  9. Phosphoinositide 3-kinase mediated signaling in lobster olfactory receptor neurons

    Science.gov (United States)

    Corey, Elizabeth A.; Bobkov, Yuriy; Pezier, Adeline; Ache, Barry W.

    2010-01-01

    In vertebrates and some invertebrates, odorant molecules bind to G protein-coupled receptors (GPCRs) on olfactory receptor neurons (ORNs) to initiate signal transduction. Phosphoinositide 3-kinase (PI3K) activity has been implicated physiologically in olfactory signal transduction, suggesting a potential role for a GPCR-activated class I PI3K. Using isoform-specific antibodies, we identified a protein in the olfactory signal transduction compartment of lobster ORNs that is antigenically similar to mammalian PI3Kγ and cloned a gene for a PI3K with amino acid homology with PI3Kβ. The lobster olfactory PI3K co-immunoprecipitates with the G protein α and β subunits, and an odorant-evoked increase in phosphatidylinositol (3,4,5)-trisphosphate can be detected in the signal transduction compartment of the ORNs. PI3Kγ and β isoform-specific inhibitors reduce the odorant-evoked output of lobster ORNs in vivo. Collectively, these findings provide evidence that PI3K is indeed activated by odorant receptors in lobster ORNs and further support the potential involvement of G protein activated PI3K signaling in olfactory transduction. PMID:20132480

  10. Src activates Abl to augment Robo1 expression in order to promote tumor cell migration

    OpenAIRE

    Khusial, P. Raaj; Vadla, Bhaskar; Krishnan, Harini; Ramlall, Trudy F.; Shen, Yongquan; Ichikawa, Hitoshi; Geng, Jian-Guo; Goldberg, Gary S.

    2010-01-01

    Cell migration is an essential step in cancer invasion and metastasis. A number of orchestrated cellular events involving tyrosine kinases and signaling receptors enable cancer cells to dislodge from primary tumors and colonize elsewhere in the body. For example, activation of the Src and Abl kinases can mediate events that promote tumor cell migration. Also, activation of the Robo1 receptor can induce tumor cell migration. However, while the importance of Src, Abl, and Robo1 in cell migratio...

  11. Functional profile of the binary brain corticosteroid receptor system: mediating, multitasking, coordinating, integrating.

    Science.gov (United States)

    de Kloet, E R

    2013-11-05

    This contribution is focused on the action of the naturally occurring corticosteroids, cortisol and corticosterone, which are secreted from the adrenals in hourly pulses and after stress with the goal to maintain resilience and health. To achieve this goal the action of the corticosteroids displays an impressive diversity, because it is cell-specific and context-dependent in coordinating the individual's response to changing environments. These diverse actions of corticosterone are mediated by mineralocorticoid- and glucocorticoid-receptors that operate as a binary system in concert with neurotransmitter and neuropeptide signals to activate and inhibit stress reactions, respectively. Classically MR and GR are gene transcription factors, but recently these receptors appear to mediate also rapid non-genomic actions on excitatory neurotransmission suggesting that they integrate functions over time. Hence the balance of receptor-mediated actions is crucial for homeostasis. This balanced function of mineralo- and glucocorticoid-receptors can be altered epigenetically by a history of traumatic (early) life events and the experience of repeated stressors as well as by predisposing genetic variants in signaling pathways of these receptors. One of these variants, mineralocorticoid receptor haplotype 2, is associated with dispositional optimism in appraisal of environmental challenges. Imbalance in receptor-mediated corticosterone actions was found to leave a genomic signature highlighting the role of master switches such as cAMP response element-binding protein and mammalian target of rapamycin to compromise health, and to promote vulnerability to disease. Diabetic encephalopathy is a pathology of imbalanced corticosterone action, which can be corrected in its pre-stage by a brief treatment with the antiglucocorticoid mifepristone. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Participation of the cell polarity protein PALS1 to T-cell receptor-mediated NF-κB activation.

    Science.gov (United States)

    Carvalho, Gabrielle; Poalas, Konstantinos; Demian, Catherine; Hatchi, Emeline; Vazquez, Aimé; Bidère, Nicolas

    2011-03-30

    Beside their established function in shaping cell architecture, some cell polarity proteins were proposed to participate to lymphocyte migration, homing, scanning, as well as activation following antigen receptor stimulation. Although PALS1 is a central component of the cell polarity network, its expression and function in lymphocytes remains unknown. Here we investigated whether PALS1 is present in T cells and whether it contributes to T Cell-Receptor (TCR)-mediated activation. By combining RT-PCR and immunoblot assays, we found that PALS1 is constitutively expressed in human T lymphocytes as well as in Jurkat T cells. siRNA-based knockdown of PALS1 hampered TCR-induced activation and optimal proliferation of lymphocyte. We further provide evidence that PALS1 depletion selectively hindered TCR-driven activation of the transcription factor NF-κB. The cell polarity protein PALS1 is expressed in T lymphocytes and participates to the optimal activation of NF-κB following TCR stimulation.

  13. Participation of the cell polarity protein PALS1 to T-cell receptor-mediated NF-κB activation.

    Directory of Open Access Journals (Sweden)

    Gabrielle Carvalho

    Full Text Available BACKGROUND: Beside their established function in shaping cell architecture, some cell polarity proteins were proposed to participate to lymphocyte migration, homing, scanning, as well as activation following antigen receptor stimulation. Although PALS1 is a central component of the cell polarity network, its expression and function in lymphocytes remains unknown. Here we investigated whether PALS1 is present in T cells and whether it contributes to T Cell-Receptor (TCR-mediated activation. METHODOLOGY/PRINCIPAL FINDINGS: By combining RT-PCR and immunoblot assays, we found that PALS1 is constitutively expressed in human T lymphocytes as well as in Jurkat T cells. siRNA-based knockdown of PALS1 hampered TCR-induced activation and optimal proliferation of lymphocyte. We further provide evidence that PALS1 depletion selectively hindered TCR-driven activation of the transcription factor NF-κB. CONCLUSIONS: The cell polarity protein PALS1 is expressed in T lymphocytes and participates to the optimal activation of NF-κB following TCR stimulation.

  14. Angiogenic peptide (AG)-30/5C activates human keratinocytes to produce cytokines/chemokines and to migrate and proliferate via MrgX receptors.

    Science.gov (United States)

    Kiatsurayanon, Chanisa; Niyonsaba, François; Chieosilapatham, Panjit; Okumura, Ko; Ikeda, Shigaku; Ogawa, Hideoki

    2016-09-01

    In addition to their antimicrobial activities, antimicrobial peptides, also known as host defense peptides (HDPs) activate keratinocytes; promote wound healing; and improve the skin barrier. AG-30/5C is a novel angiogenic HDP that activates various functions of fibroblasts and endothelial cells, including cytokine/chemokine production and wound healing. To investigate whether AG-30/5C activates human keratinocytes and to examine the underlying mechanisms. Production of cytokines/chemokines was assessed by ELISA. Expression of Mas-related G-protein coupled receptors X (MrgXs) in keratinocytes was determined by real-time PCR and Western blot. MAPK and NF-κB activation was analysed by Western blot. Cell migration was assessed by chemotaxis microchamber and in vitro wound closure assay, whereas cell proliferation was analysed using an XTT assay. We found that AG-30/5C was more efficient than its parent peptide AG-30 in increasing the production of various cytokines/chemokines and promoting keratinocyte migration and proliferation. Furthermore, MrgX3 and MrgX4 receptors were constitutively expressed in keratinocytes at higher levels than MrgX1 and MrgX2, and were up-regulated upon stimulation with TLR ligands. Because MrgX3 and MrgX4 siRNAs suppressed AG-30/5C-mediated cytokine/chemokine production, keratinocyte migration and proliferation, we propose that AG-30/5C utilizes these MrgXs to stimulate keratinocytes. In addition, AG-30/5C-induced activation of keratinocytes was controlled by MAPK and NF-κB pathways, as evidenced by the inhibitory effects of ERK-, JNK-, p38- and NF-κB-specific inhibitors. Indeed, we confirmed that AG-30/5C enhanced phosphorylation of MAPKs and IκB. Our findings provide novel evidence that AG-30/5C may be a useful therapeutic agent for wound healing by activating human keratinocytes. Copyright © 2016 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  15. Nuclear receptor corepressor-dependent repression of peroxisome-proliferator-activated receptor delta-mediated transactivation

    DEFF Research Database (Denmark)

    Krogsdam, Anne-M; Nielsen, Curt A F; Neve, Søren

    2002-01-01

    delta-RXR alpha heterodimer bound to an acyl-CoA oxidase (ACO)-type peroxisome-proliferator response element recruited a glutathione S-transferase-NCoR fusion protein in a ligand-independent manner. Contrasting with most other nuclear receptors, PPAR delta was found to interact equally well......The nuclear receptor corepressor (NCoR) was isolated as a peroxisome-proliferator-activated receptor (PPAR) delta interacting protein using the yeast two-hybrid system. NCoR interacted strongly with the ligand-binding domain of PPAR delta, whereas interactions with the ligand-binding domains...

  16. Angiomotin is a novel component of cadherin-11/β-catenin/p120 complex and is critical for cadherin-11-mediated cell migration

    Science.gov (United States)

    Ortiz, Angelica; Lee, Yu-Chen; Yu, Guoyu; Liu, Hsuan-Chen; Lin, Song-Chang; Bilen, Melmet Asim; Cho, Hyojin; Yu-Lee, Li-Yuan; Lin, Sue-Hwa

    2015-01-01

    Loss of E-cadherin and up-regulation of mesenchymal cadherins, a hallmark of the epithelial–mesenchymal transition, contributes to migration and dissemination of cancer cells. Expression of human cadherin-11 (Cad11), also known as osteoblast cadherin, in prostate cancer increases the migration of prostate cancer cells. How Cad11 mediates cell migration is unknown. Using the human Cad11 cytoplasmic domain in pulldown assays, we identified human angiomotin (Amot), known to be involved in cell polarity, migration, and Hippo pathway, as a component of the Cad11 protein complex. Deletion analysis showed that the last C-terminal 10 amino acids in Cad11 cytoplasmic domain are required for Amot binding. Further, Cad11 preferentially interacts with Amot-p80 than Amot-p130 isoform and binds directly to the middle domain of Amot-p80. Cad11-Amot interaction affects Cad11-mediated cell migration, but not homophilic adhesion, as deletion of Amot binding motif of Cad11 (Cad11-ΔAmot) did not abolish Cad11-mediated cell–cell adhesion of mouse L cells, but significantly reduced Cad11-mediated cell migration of human C4-2B4 and PC3-mm2 prostate cancer cells and human HEK293T cells. Together, our studies identified Amot-p80 as a novel component of the Cad11 complex and demonstrated that Amot-p80 is critical for Cad11-mediated cell migration.—Ortiz, A., Lee, Y.-C., Yu, G., Liu, H.-C., Lin, S.-C., Bilen, M. A., Cho, H., Yu-Lee, L.-Y., Lin, S.-H. Angiomotin is a novel component of cadherin-11/β-catenin/p120 complex and is critical for cadherin-11-mediated cell migration. PMID:25466890

  17. Internal wave-mediated shading causes frequent vertical migrations in fishes

    KAUST Repository

    Kaartvedt, Stein

    2012-04-25

    We provide evidence that internal waves cause frequent vertical migrations (FVM) in fishes. Acoustic data from the Benguela Current revealed that pelagic scattering layers of fish below ~140 m moved in opposite phases to internal waves, ascending ~20 m towards the wave trough and descending from the wave crest. At the trough, the downward displacement of upper waters and the upward migration of fish created an overlapping zone. Near-bottom fish correspondingly left the benthic boundary zone at the wave trough, ascending into an acoustic scattering layer likely consisting of zooplankton and then descending to the benthic boundary zone at the wave crest. We suggest that this vertical fish migration is a response to fluctuations in light intensity of 3 to 4 orders of magnitude caused by shading from a turbid surface layer that had chlorophyll a values of 3 to 4 mg m−3 and varied in thickness from ~15 to 50 m at a temporal scale corresponding to the internal wave period (30 min). This migration frequency thus is much higher than that of the common and widespread light-associated diel vertical migration. Vertical movements affect prey encounters, growth, and survival. We hypothesize that FVM increase the likelihood of prey encounters and the time for safe visual foraging among planktivorous fish, thereby contributing to efficient trophic transfer in major upwelling areas.

  18. Knockdown of EphB1 receptor decreases medulloblastoma cell growth and migration and increases cellular radiosensitization.

    Science.gov (United States)

    Bhatia, Shilpa; Baig, Nimrah A; Timofeeva, Olga; Pasquale, Elena B; Hirsch, Kellen; MacDonald, Tobey J; Dritschilo, Anatoly; Lee, Yi Chien; Henkemeyer, Mark; Rood, Brian; Jung, Mira; Wang, Xiao-Jing; Kool, Marcel; Rodriguez, Olga; Albanese, Chris; Karam, Sana D

    2015-04-20

    The expression of members of the Eph family of receptor tyrosine kinases and their ephrin ligands is frequently dysregulated in medulloblastomas. We assessed the expression and functional role of EphB1 in medulloblastoma cell lines and engineered mouse models. mRNA and protein expression profiling showed expression of EphB1 receptor in the human medulloblastoma cell lines DAOY and UW228. EphB1 downregulation reduced cell growth and viability, decreased the expression of important cell cycle regulators, and increased the percentage of cells in G1 phase of the cell cycle. It also modulated the expression of proliferation, and cell survival markers. In addition, EphB1 knockdown in DAOY cells resulted in significant decrease in migration, which correlated with decreased β1-integrin expression and levels of phosphorylated Src. Furthermore, EphB1 knockdown enhanced cellular radiosensitization of medulloblastoma cells in culture and in a genetically engineered mouse medulloblastoma model. Using genetically engineered mouse models, we established that genetic loss of EphB1 resulted in a significant delay in tumor recurrence following irradiation compared to EphB1-expressing control tumors. Taken together, our findings establish that EphB1 plays a key role in medulloblastoma cell growth, viability, migration, and radiation sensitivity, making EphB1 a promising therapeutic target.

  19. Receptor targeting of hemoglobin mediated by the haptoglobins

    DEFF Research Database (Denmark)

    Nielsen, Marianne Jensby; Moestrup, Søren Kragh

    2009-01-01

    -scavenging effects associated with "free" hemoglobin, and, furthermore, elicits an anti-inflammatory response. In the late primate evolution, haptoglobin variants with distinct functions have arisen, including haptoglobin polymers and the haptoglobin-related protein. The latter associates with a subspecies of high...... the parasite. In conclusion, variant human homologous hemoglobin-binding proteins that collectively may be designated the haptoglobins have diverted from the haptoglobin gene. On hemoglobin and receptor interaction, these haptoglobins contribute to different biologic events that go beyond simple removal from...

  20. Androgen Receptor-Mediated Escape Mechanisms from Androgen Ablation Therapy

    Science.gov (United States)

    2005-10-01

    p. 132-138. 7. Han , G., et al., Mutation of the androgen receptor causes oncogenic transformation of the prostate. Proc Natl Acad Sci U S A, 2005. 102...the confor- (1997) 306-360. mational state of the AR may recruit specific TPR- [9] K. Richter, J. Buchner , Hsp90: chaperoning signal transduc-tion...Biol. 42 connection, Trends Biochem. Sci. 22 (1997) 87-92. (1992) 803-812.[19] 2) D.F. 812. S[34] U. Jakob, J. Buchner , Assisting spontaneity: the role

  1. Muscarinic receptor expression and receptor-mediated detrusor contraction: comparison of juvenile and adult porcine tissue

    NARCIS (Netherlands)

    Wuest, Melinda; Eichhorn, Birgit; Braeter, Manfred; Strugala, Gerhard; Michel, Martin C.; Ravens, Ursula

    2008-01-01

    Urinary bladder function is known to mature during fetal and postnatal development, including changes in neurotransmitter regulation of detrusor contraction. However, only few experimental data are available about muscarinic receptor antagonist function in the urinary bladder from young animals. In

  2. N-arachidonoyl glycine, an abundant endogenous lipid, potently drives directed cellular migration through GPR18, the putative abnormal cannabidiol receptor

    Directory of Open Access Journals (Sweden)

    Vogel Zvi

    2010-03-01

    Full Text Available Abstract Background Microglia provide continuous immune surveillance of the CNS and upon activation rapidly change phenotype to express receptors that respond to chemoattractants during CNS damage or infection. These activated microglia undergo directed migration towards affected tissue. Importantly, the molecular species of chemoattractant encountered determines if microglia respond with pro- or anti-inflammatory behaviour, yet the signaling molecules that trigger migration remain poorly understood. The endogenous cannabinoid system regulates microglial migration via CB2 receptors and an as yet unidentified GPCR termed the 'abnormal cannabidiol' (Abn-CBD receptor. Abn-CBD is a synthetic isomer of the phytocannabinoid cannabidiol (CBD and is inactive at CB1 or CB2 receptors, but functions as a selective agonist at this Gi/o-coupled GPCR. N-arachidonoyl glycine (NAGly is an endogenous metabolite of the endocannabinoid anandamide and acts as an efficacious agonist at GPR18. Here, we investigate the relationship between NAGly, Abn-CBD, the unidentified 'Abn-CBD' receptor, GPR18, and BV-2 microglial migration. Results Using Boyden chamber migration experiments, yellow tetrazolium (MTT conversion, In-cell Western, qPCR and immunocytochemistry we show that NAGly, at sub-nanomolar concentrations, and Abn-CBD potently drive cellular migration in both BV-2 microglia and HEK293-GPR18 transfected cells, but neither induce migration in HEK-GPR55 or non-transfected HEK293 wildtype cells. Migration effects are blocked or attenuated in both systems by the 'Abn-CBD' receptor antagonist O-1918, and low efficacy agonists N-arachidonoyl-serine and cannabidiol. NAGly promotes proliferation and activation of MAP kinases in BV-2 microglia and HEK293-GPR18 cells at low nanomolar concentrations - cellular responses correlated with microglial migration. Additionally, BV-2 cells show GPR18 immunocytochemical staining and abundant GPR18 mRNA. qPCR demonstrates that

  3. Hypersensitivity Induced by Activation of Spinal Cord PAR2 Receptors Is Partially Mediated by TRPV1 Receptors.

    Directory of Open Access Journals (Sweden)

    Petra Mrozkova

    Full Text Available Protease-activated receptors 2 (PAR2 and transient receptor potential vanilloid 1 (TRPV1 receptors in the peripheral nerve endings are implicated in the development of increased sensitivity to mechanical and thermal stimuli, especially during inflammatory states. Both PAR2 and TRPV1 receptors are co-expressed in nociceptive dorsal root ganglion (DRG neurons on their peripheral endings and also on presynaptic endings in the spinal cord dorsal horn. However, the modulation of nociceptive synaptic transmission in the superficial dorsal horn after activation of PAR2 and their functional coupling with TRPV1 is not clear. To investigate the role of spinal PAR2 activation on nociceptive modulation, intrathecal drug application was used in behavioural experiments and patch-clamp recordings of spontaneous, miniature and dorsal root stimulation-evoked excitatory postsynaptic currents (sEPSCs, mEPSCs, eEPSCs were performed on superficial dorsal horn neurons in acute rat spinal cord slices. Intrathecal application of PAR2 activating peptide SLIGKV-NH2 induced thermal hyperalgesia, which was prevented by pretreatment with TRPV1 antagonist SB 366791 and was reduced by protein kinases inhibitor staurosporine. Patch-clamp experiments revealed robust decrease of mEPSC frequency (62.8 ± 4.9%, increase of sEPSC frequency (127.0 ± 5.9% and eEPSC amplitude (126.9 ± 12.0% in dorsal horn neurons after acute SLIGKV-NH2 application. All these EPSC changes, induced by PAR2 activation, were prevented by SB 366791 and staurosporine pretreatment. Our results demonstrate an important role of spinal PAR2 receptors in modulation of nociceptive transmission in the spinal cord dorsal horn at least partially mediated by activation of presynaptic TRPV1 receptors. The functional coupling between the PAR2 and TRPV1 receptors on the central branches of DRG neurons may be important especially during different pathological states when it may enhance pain perception.

  4. Hypersensitivity Induced by Activation of Spinal Cord PAR2 Receptors Is Partially Mediated by TRPV1 Receptors.

    Science.gov (United States)

    Mrozkova, Petra; Spicarova, Diana; Palecek, Jiri

    2016-01-01

    Protease-activated receptors 2 (PAR2) and transient receptor potential vanilloid 1 (TRPV1) receptors in the peripheral nerve endings are implicated in the development of increased sensitivity to mechanical and thermal stimuli, especially during inflammatory states. Both PAR2 and TRPV1 receptors are co-expressed in nociceptive dorsal root ganglion (DRG) neurons on their peripheral endings and also on presynaptic endings in the spinal cord dorsal horn. However, the modulation of nociceptive synaptic transmission in the superficial dorsal horn after activation of PAR2 and their functional coupling with TRPV1 is not clear. To investigate the role of spinal PAR2 activation on nociceptive modulation, intrathecal drug application was used in behavioural experiments and patch-clamp recordings of spontaneous, miniature and dorsal root stimulation-evoked excitatory postsynaptic currents (sEPSCs, mEPSCs, eEPSCs) were performed on superficial dorsal horn neurons in acute rat spinal cord slices. Intrathecal application of PAR2 activating peptide SLIGKV-NH2 induced thermal hyperalgesia, which was prevented by pretreatment with TRPV1 antagonist SB 366791 and was reduced by protein kinases inhibitor staurosporine. Patch-clamp experiments revealed robust decrease of mEPSC frequency (62.8 ± 4.9%), increase of sEPSC frequency (127.0 ± 5.9%) and eEPSC amplitude (126.9 ± 12.0%) in dorsal horn neurons after acute SLIGKV-NH2 application. All these EPSC changes, induced by PAR2 activation, were prevented by SB 366791 and staurosporine pretreatment. Our results demonstrate an important role of spinal PAR2 receptors in modulation of nociceptive transmission in the spinal cord dorsal horn at least partially mediated by activation of presynaptic TRPV1 receptors. The functional coupling between the PAR2 and TRPV1 receptors on the central branches of DRG neurons may be important especially during different pathological states when it may enhance pain perception.

  5. Characterization of the muscarinic receptor subtype mediating vasodilation in the rat perfused mesenteric vascular bed preparation

    NARCIS (Netherlands)

    Hendriks, M. G.; Pfaffendorf, M.; van Zwieten, P. A.

    1992-01-01

    1. The nature of the muscarinic receptor subtype mediating endothelium-dependent vascular relaxation was investigated in the perfused mesenteric vascular bed preparation which is a model for resistance vessels. 2. After methoxamine-induced vasoconstriction the vessels were dilated with

  6. REST mediates androgen receptor actions on gene repression and predicts early recurrence of prostate cancer

    DEFF Research Database (Denmark)

    Svensson, Charlotte; Ceder, Jens; Iglesias Gato, Diego

    2014-01-01

    The androgen receptor (AR) is a key regulator of prostate tumorgenesis through actions that are not fully understood. We identified the repressor element (RE)-1 silencing transcription factor (REST) as a mediator of AR actions on gene repression. Chromatin immunoprecipitation showed that AR binds...

  7. MicroRNA-219 modulates NMDA receptor-mediated neurobehavioral dysfunction

    DEFF Research Database (Denmark)

    Kocerha, Jannet; Faghihi, Mohammad Ali; Lopez-Toledano, Miguel A

    2009-01-01

    N-methyl-D-aspartate (NMDA) glutamate receptors are regulators of fast neurotransmission and synaptic plasticity in the brain. Disruption of NMDA-mediated glutamate signaling has been linked to behavioral deficits displayed in psychiatric disorders such as schizophrenia. Recently, noncoding RNA...

  8. In Vivo Tracking of Chemokine Receptor CXCR4-Engineered Mesenchymal Stem Cell Migration by Optical Molecular Imaging

    Directory of Open Access Journals (Sweden)

    Senthilkumar Kalimuthu

    2017-01-01

    Full Text Available CXCR4, the stromal cell-derived factor-1 receptor, plays an important role in the migration of hematopoietic progenitor/stem cells to injured and inflamed areas. Noninvasive cell tracking methods could be useful for monitoring cell fate. Therefore, in this study, we evaluated the efficacy of an intravenous infusion of genetically engineered mesenchymal stem cells (MSCs overexpressing CXC chemokine receptor 4 (CXCR4 to home to the tumor, by optical imaging. We constructed a retroviral vector containing CXCR with dual reporter genes, eGFP and Fluc2, under the control of an EF1α promoter (pBABE-EF1α-CXCR4-eGFP-IRES-Fluc2. We also developed an eGFP-Fluc2 construct in the Retro-X retroviral vector (Retro-X-eGFP-Fluc2. MSCs were transduced with retroviruses to generate CXCR4-overexpressing MSCs (MSC-CXCR4/Fluc2 and MSCs (MSC/Fluc2. CXCR4 mRNA and protein expression was confirmed by RT-PCR and Western blotting, respectively, and it was higher in MSC-CXCR4/Fluc2 than in naive MSCs. eGFP expression was confirmed by confocal microscopy. The transfected MSC-CXCR4/Fluc2 cells showed higher migratory capacity than naive MSCs observed in Transwell migration assay. The in vivo migration of CXCR4-overexpressing MSCs to MDAMB231/Rluc tumor model by BLI imaging was also confirmed. Intravenous delivery of genetically modified MSCs overexpressing CXCR4 with a Fluc2 reporter gene may be a useful, noninvasive BLI imaging tool for tracking cell fate.

  9. Glucocorticoid receptor, but not mineralocorticoid receptor, mediates cortisol regulation of epidermal ionocyte development and ion transport in zebrafish (danio rerio.

    Directory of Open Access Journals (Sweden)

    Shelly Abad Cruz

    Full Text Available Cortisol is the major endogenous glucocorticoid (GC both in human and fish, mediated by corticosteroid receptors. Due to the absence of aldosterone production in teleost fish, cortisol is also traditionally accepted to function as mineralocorticoid (MC; but whether it acts through the glucocorticoid receptor (GR or the mineralocorticoid receptor (MR remains a subject of debate. Here, we used loss-of-function and rescue assays to determine whether cortisol affects zebrafish epidermal ionocyte development and function via the GR and/or the MR. GR knockdown morphants displayed a significant decrease in the major ionocytes, namely Na(+-K(+-ATPase-rich cells (NaRCs and H(+-ATPase-rich cells (HRCs, as well as other cells, including epidermal stem cells (ESCs, keratinocytes, and mucus cells; conversely, cell numbers were unaffected in MR knockdown morphants. In agreement, GR morphants, but not MR morphants, exhibited decreased NaRC-mediated Ca(2+ uptake and HRC-mediated H(+ secretion. Rescue via GR capped mRNA injection or exogenous cortisol incubation normalized the number of epidermal ionocytes in GR morphants. We also provide evidence for GR localization in epidermal cells. At the transcript level, GR mRNA is ubiquitously expressed in gill sections and present in both NaRCs and HRCs, supporting the knockdown and functional assay results in embryo. Altogether, we have provided solid molecular evidence that GR is indeed present on ionocytes, where it mediates the effects of cortisol on ionocyte development and function. Hence, cortisol-GR axis performs the roles of both GC and MC in zebrafish skin and gills.

  10. AMP-activated protein kinase activation mediates CCL3-induced cell migration and matrix metalloproteinase-2 expression in human chondrosarcoma

    Science.gov (United States)

    2013-01-01

    Chemokine (C-C motif) ligand 3 (CCL3), also known as macrophage inflammatory protein-1α, is a cytokine involved in inflammation and activation of polymorphonuclear leukocytes. CCL3 has been detected in infiltrating cells and tumor cells. Chondrosarcoma is a highly malignant tumor that causes distant metastasis. However, the effect of CCL3 on human chondrosarcoma metastasis is still unknown. Here, we found that CCL3 increased cellular migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells. Pre-treatment of cells with the MMP-2 inhibitor or transfection with MMP-2 specific siRNA abolished CCL3-induced cell migration. CCL3 has been reported to exert its effects through activation of its specific receptor, CC chemokine receptor 5 (CCR5). The CCR5 and AMP-activated protein kinase (AMPK) inhibitor or siRNA also attenuated CCL3-upregulated cell motility and MMP-2 expression. CCL3-induced expression of MMP-2 and migration were also inhibited by specific inhibitors, and inactive mutants of AMPK, p38 mitogen activated protein kinase (p38 or p38-MAPK), and nuclear factor κB (NF-κB) cascades. On the other hand, CCL3 treatment demonstrably activated AMPK, p38, and NF-κB signaling pathways. Furthermore, the expression levels of CCL3, CCR5, and MMP-2 were correlated in human chondrosarcoma specimens. Taken together, our results indicate that CCL3 enhances the migratory ability of human chondrosarcoma cells by increasing MMP-2 expression via the CCR5, AMPK, p38, and NF-κB pathways. PMID:24047437

  11. IL-8 Is Involved in Estrogen-Related Receptor α-Regulated Proliferation and Migration of Colorectal Cancer Cells.

    Science.gov (United States)

    Ding, Sijuan; Tang, Zhaohui; Jiang, Yongjun; Huang, Honglin; Luo, Pengfei; Qing, Bohua; Zhang, Siyuan; Tang, Ruoting

    2017-10-09

    Studies revealed that estrogenic signals were involved in the development of colorectal cancer (CRC), while the roles of estrogen related receptor (ERR) on the progression of CRC have not been well illustrated. Its roles on the development of CRC were investigated. The expression of ERRα/β/γ in CRC cells were measured. The effects of ERRα on cell proliferation, migration and expression of cytokines were investigated accordingly. Our data revealed that the expression of ERRα, while not ERRβ or ERRγ, was significantly increased in CRC cells and clinical CRC tissues. Both the inverse agonist of ERRα (XCT-790) and si-ERRα can inhibit the proliferation of CRC cells. XCT-790 treatment can also suppress the wound healing and in vitro migration of CRC cells. Cytokine assays showed that XCT-790 can significantly decrease the expression of interleukin-8 (IL-8), while not IL-4, IL-6, IL-8, IL-9, IL-10, IL-18, IFN-γ, or TGF-β, in CRC cells. Over expression of ERRα increased the expression of IL-8. Luciferase assay showed XCT-790 decreased the promoter activity of IL-8. XCT-790 can increase the decay of IL-8 mRNA in SW480 cells. The recombinant IL-8 (rIL-8) can rescue XCT-790 induced suppression of proliferation and migration of CRC cells. XCT-790 can decrease the phosphorylation of ERK1/2 and STAT3, two downstream signal molecules of IL-8, in CRC cells. While rIL-8 can markedly attenuate XCT-790 induced dephosphorylation of ERK1/2 and STAT3. Our data showed that ERRα can trigger the proliferation and migration of CRC cells via up regulation of IL-8. Therefor targeted inhibition of ERRα/IL-8 might be a potential approach for CRC treatment and drug development.

  12. Adult mouse subventricular zone stem and progenitor cells are sessile and epidermal growth factor receptor negatively regulates neuroblast migration.

    Directory of Open Access Journals (Sweden)

    Yongsoo Kim

    2009-12-01

    Full Text Available The adult subventricular zone (SVZ contains stem and progenitor cells that generate neuroblasts throughout life. Although it is well accepted that SVZ neuroblasts are migratory, recent evidence suggests their progenitor cells may also exhibit motility. Since stem and progenitor cells are proliferative and multipotential, if they were also able to move would have important implications for SVZ neurogenesis and its potential for repair.We studied whether SVZ stem and/or progenitor cells are motile in transgenic GFP+ slices with two photon time lapse microscopy and post hoc immunohistochemistry. We found that stem and progenitor cells; mGFAP-GFP+ cells, bright nestin-GFP+ cells and Mash1+ cells were stationary in the SVZ and rostral migratory stream (RMS. In our search for motile progenitor cells, we uncovered a population of motile betaIII-tubulin+ neuroblasts that expressed low levels of epidermal growth factor receptor (EGFr. This was intriguing since EGFr drives proliferation in the SVZ and affects migration in other systems. Thus we examined the potential role of EGFr in modulating SVZ migration. Interestingly, EGFr(low neuroblasts moved slower and in more tortuous patterns than EGFr-negative neuroblasts. We next questioned whether EGFr stimulation affects SVZ cell migration by imaging Gad65-GFP+ neuroblasts in the presence of transforming growth factor alpha (TGF-alpha, an EGFr-selective agonist. Indeed, acute exposure to TGF-alpha decreased the percentage of motile cells by approximately 40%.In summary, the present study directly shows that SVZ stem and progenitor cells are static, that EGFr is retained on some neuroblasts, and that EGFr stimulation negatively regulates migration. This result suggests an additional role for EGFr signaling in the SVZ.

  13. Hippocampus NMDA receptors selectively mediate latent extinction of place learning.

    Science.gov (United States)

    Goodman, Jarid; Gabriele, Amanda; Packard, Mark G

    2016-09-01

    Extinction of maze learning may be achieved with or without the animal performing the previously acquired response. In typical "response extinction," animals are given the opportunity to make the previously acquired approach response toward the goal location of the maze without reinforcement. In "latent extinction," animals are not given the opportunity to make the previously acquired response and instead are confined to the previous goal location without reinforcement. Previous evidence indicates that the effectiveness of these protocols may depend on the type of memory being extinguished. Thus, one aim of the present study was to further examine the effectiveness of response and latent extinction protocols across dorsolateral striatum (DLS)-dependent response learning and hippocampus-dependent place learning tasks. In addition, previous neural inactivation experiments indicate a selective role for the hippocampus in latent extinction, but have not investigated the precise neurotransmitter mechanisms involved. Thus, the present study also examined whether latent extinction of place learning might depend on NMDA receptor activity in the hippocampus. In experiment 1, adult male Long-Evans rats were trained in a response learning task in a water plus-maze, in which animals were reinforced to make a consistent body-turn response to reach an invisible escape platform. Results indicated that response extinction, but not latent extinction, was effective at extinguishing memory in the response learning task. In experiment 2, rats were trained in a place learning task, in which animals were reinforced to approach a consistent spatial location containing the hidden escape platform. In experiment 2, animals also received intra-hippocampal infusions of the NMDA receptor antagonist 2-amino-5-phosphopentanoic acid (AP5; 5.0 or 7.5 ug/0.5 µg) or saline vehicle immediately before response or latent extinction training. Results indicated that both extinction protocols were

  14. Dopamine receptor-mediated regulation of neuronal “clock” gene expression

    Science.gov (United States)

    Imbesi, Marta; Yildiz, Sevim; Arslan, Ahmet Dirim; Sharma, Rajiv; Manev, Hari; Uz, Tolga

    2009-01-01

    Using transgenic mice model (i.e., “clock” knockouts), clock transcription factors have been suggested as critical regulators of dopaminergic behaviors induced by drugs of abuse. Moreover, it has been shown that systemic administration of psychostimulants, such as cocaine and methamphetamine regulate the striatal expression of clock genes. However, it is not known whether dopamine receptors mediate these regulatory effects of psychostimulants at the cellular level. Primary striatal neurons in culture express dopamine receptors as well as clock genes and have been successfully used in studying dopamine receptor functioning. Therefore, we investigated the role of dopamine receptors on neuronal clock gene expression in this model using specific receptor agonists. We found an inhibitory effect on the expression of mClock and mPer1 genes with the D2-class (i.e., D2/D3) receptor agonist quinpirole. We also found a generalized stimulatory effect on the expression of clock genes mPer1, mClock, mNPAS2, and mBmal1 with the D1-class (i.e., D1) receptor agonist SKF38393. Further, we tested whether systemic administration of dopamine receptor agonists causes similar changes in striatal clock gene expression in vivo. We found quinpirole-induced alterations in mPER1 protein levels in the mouse striatum (i.e., rhythm shift). Collectively, our results indicate that the DA receptor system may mediate psychostimulant-induced changes in clock gene expression. Using striatal neurons in culture as a model, further research is needed to better understand how dopamine signaling modulates the expression dynamics of clock genes (i.e., intracellular signaling pathways) and thereby influences neuronal gene expression, neuronal transmission, and brain functioning. PMID:19017537

  15. Vascular endothelin ET(B) receptor-mediated contraction requires phosphorylation of ERK1/2 proteins

    DEFF Research Database (Denmark)

    Luo, Guogang; Jamali, Roya; Cao, Yong-Xiao

    2006-01-01

    In cardiovascular diseases, endothelin type B (ET(B)) receptors in arterial smooth muscle cells are upregulated. The present study revealed that organ culture of rat mesenteric artery segments enhanced endothelin ET(B) receptor-mediated contraction paralleled with increase in the receptor m...

  16. Characterization of murine melanocortin receptors mediating adipocyte lipolysis and examination of signalling pathways involved

    DEFF Research Database (Denmark)

    Møller, Cathrine Laustrup; Raun, Kirsten; Jacobsen, Marianne Lambert

    2011-01-01

    The melanocortin receptors (MCRs) belong to the G-protein coupled receptors (family A). So far, 5 different subtypes have been described (MC1R-MC5R) and of these MC2R and MC5R have been proposed to act directly in adipocytes and regulate lipolysis in rodents. Using ACTH and a-melanocyte stimulating...... hormone (a-MSH) generated from proopiomelanocortin (POMC), as well as synthetic MSH analogues to stimulate lipolysis in murine 3T3-L1 adipocytes it is shown that MC2R and MC5R are lipolytic mediators in differentiated 3T3-L1 adipocytes. Involvement of cAMP, phosphorylated extracellular signal...... is mediated through MC5R in a cAMP independent manner. Finally, we identify essential differences in MCR mediated lipolysis when using 3T3-L1 cells compared to primary adipocytes....

  17. miR-300 promotes proliferation and EMT-mediated colorectal cancer migration and invasion by targeting p53.

    Science.gov (United States)

    Wang, Lin; Yu, Peiwu

    2016-12-01

    p53 mutations in tumors can induce the loss of wild-type tumor-suppressing p53 function, which results in the increase in proliferation, migration and invasion ability in cancer cells. Studies have shown that the expression of p53 is regulated by several microRNAs (miRNAs). In the present study, we found that miR-300 and p53 were significantly increased in colorectal cancer (CRC) tissues when compared with levels noted in adjacent colorectal tissues. Both miR-300 and p53 were significantly correlated with lymphatic metastasis and TNM stage. Both miR-300 and p53 promoted CRC cell (SW480 and HT29) proliferation, migration, and invasion, respectively, in vitro. In addition, we found that miR-300 is a direct positive regulator of p53 through binding to the binding site in the 3'UTR of the p53 gene in human CRC cells. Moreover, both miR-300 and p53 induced CRC cell epithelial‑mesenchymal transition (EMT) respectively. Taken together, we demonstrated that miR-300 promoted proliferation and EMT-mediated CRC migration and invasion by targeting p53. These findings provide a new theoretical basis and potential therapeutic targets, and thus lays the foundation for exploring the pathogenesis of CRC.

  18. The gastrin and cholecystokinin receptors mediated signaling network: a scaffold for data analysis and new hypotheses on regulatory mechanisms.

    Science.gov (United States)

    Tripathi, Sushil; Flobak, Åsmund; Chawla, Konika; Baudot, Anaïs; Bruland, Torunn; Thommesen, Liv; Kuiper, Martin; Lægreid, Astrid

    2015-07-24

    The gastrointestinal peptide hormones cholecystokinin and gastrin exert their biological functions via cholecystokinin receptors CCK1R and CCK2R respectively. Gastrin, a central regulator of gastric acid secretion, is involved in growth and differentiation of gastric and colonic mucosa, and there is evidence that it is pro-carcinogenic. Cholecystokinin is implicated in digestion, appetite control and body weight regulation, and may play a role in several digestive disorders. We performed a detailed analysis of the literature reporting experimental evidence on signaling pathways triggered by CCK1R and CCK2R, in order to create a comprehensive map of gastrin and cholecystokinin-mediated intracellular signaling cascades. The resulting signaling map captures 413 reactions involving 530 molecular species, and incorporates the currently available knowledge into one integrated signaling network. The decomposition of the signaling map into sub-networks revealed 18 modules that represent higher-level structures of the signaling map. These modules allow a more compact mapping of intracellular signaling reactions to known cell behavioral outcomes such as proliferation, migration and apoptosis. The integration of large-scale protein-protein interaction data to this literature-based signaling map in combination with topological analyses allowed us to identify 70 proteins able to increase the compactness of the map. These proteins represent experimentally testable hypotheses for gaining new knowledge on gastrin- and cholecystokinin receptor signaling. The CCKR map is freely available both in a downloadable, machine-readable SBML-compatible format and as a web resource through PAYAO ( http://sblab.celldesigner.org:18080/Payao11/bin/). We have demonstrated how a literature-based CCKR signaling map together with its protein interaction extensions can be analyzed to generate new hypotheses on molecular mechanisms involved in gastrin- and cholecystokinin-mediated regulation of

  19. The mediating effects of parental and peer pressure on the migration intentions of graduates in Nanjing

    NARCIS (Netherlands)

    Cui, C.; Geertman, S.C.M.; Hooimeijer, P.

    2016-01-01

    This paper analyses the migration intentions of university graduates using the Theory of Planned Behaviour not just to unravel their intention but also to uncover how subjective perceptions enter the decision-making process. The results suggest that perceived parental and peer pressures have strong

  20. Effectiveness of artificial bark flaps in mediating migration of late-instar gypsy moth larvae

    Science.gov (United States)

    Michael L. McManus; Harvey R. Smith

    1984-01-01

    Field studies demonstrated that migrating larval instars of the gypsy moth, Lymantria dispar (L.), preferred resting locations placed on tree boles at 4.6 m over those placed at 1.5 and 3 m. More larvae were found beneath bark flaps than beneath flaps of hard black plastic.

  1. Calpain2 mediates Rab5-driven focal adhesion disassembly and cell migration.

    Science.gov (United States)

    Mendoza, Pablo A; Silva, Patricio; Díaz, Jorge; Arriagada, Cecilia; Canales, Jimena; Cerda, Oscar; Torres, Vicente A

    2017-11-03

    The early endosome protein Rab5 was recently shown to promote cell migration by enhancing focal adhesion disassembly through mechanisms that remain elusive. Focal adhesion disassembly is associated to proteolysis of talin, in a process that requires calpain2. Since calpain2 has been found at vesicles and endosomal compartments, we hypothesized that Rab5 stimulates calpain2 activity, leading to enhanced focal adhesion disassembly in migrating cells. We observed that calpain2 co-localizes with EEA1-positive early endosomes and co-immunoprecipitates with EEA1 and Rab5 in A549 lung carcinoma cells undergoing spreading, whereas Rab5 knock-down decreased the accumulation of calpain2 at early endosomal-enriched fractions. In addition, Rab5 silencing decreased calpain2 activity, as shown by cleavage of the fluorogenic substrate tBOC-LM-CMAC and the endogenous substrate talin. Accordingly, Rab5 promoted focal adhesion disassembly in a calpain2-dependent manner, as expression of GFP-Rab5 accelerated focal adhesion disassembly in nocodazole-synchronized cells, whereas pharmacological inhibition of calpain2 with N-acetyl-Leu-Leu-Met prevented both focal adhesion disassembly and cell migration induced by Rab5. In summary, these data uncover Rab5 as a novel regulator of calpain2 activity and focal adhesion proteolysis leading to cell migration.

  2. Activation of Toll-like receptor-9 promotes cellular migration via up-regulating MMP-2 expression in oral squamous cell carcinoma.

    Directory of Open Access Journals (Sweden)

    Min Ruan

    Full Text Available PURPOSE: Activation of Toll like receptors (TLRs signaling has been implicated in promoting malignant cell invasion and metastatic potential. Previously we demonstrated that increased TLR-9 expression predicted poor survival in oral cancer patients. The objective of this study is to further investigate the roles and potential molecular mechanisms of TLR-9 signaling in human oral cancer cell invasion. METHODS: Cell migration, invasion and protein expression were detected by wound healing assay, Transwell chambers model and western blot. The secretion and activity levels of metalloproteinases-2/9 were quantified by ELISA and Gelatin zymography. EMSA and ChIP assays were employed to detect the activity of AP-1signal pathway. TLR-9 siRNA transfection was used to regulate the expression and activity of TLR-9 in oral cancer cell line HB cells. RESULT: The results of both wound healing assay and in vitro Transwell assay revealed that activation of TLR-9 induced dose- and time- dependent migration and invasion of HB cells. An increased expression, secretion and activity of MMP-2 were observed upon the treatment of CpG-ODN. The TLR-9 signaling-mediated MMP-2 expression appeared to be a consequence of AP-1 activation, because that their DNA binding activity was enhanced by CpG-ODN treatment. All these influences were efficiently repressed by the knockdown of TLR-9 through siRNA or pretreatment of an AP-1 inhibitor. CONCLUSION: Activation of TLR-9 signaling could promote human oral cancer HB cells invasion with the induction of MMP-2 presentation by attenuating AP-1 binding activity, suggesting a novel anti-metastatic application for TLR-9 targeted therapy in oral cancer in the future.

  3. Adipocyte glucocorticoid receptors mediate fat-to-brain signaling.

    Science.gov (United States)

    de Kloet, Annette D; Krause, Eric G; Solomon, Matia B; Flak, Jonathan N; Scott, Karen A; Kim, Dong-Hoon; Myers, Brent; Ulrich-Lai, Yvonne M; Woods, Stephen C; Seeley, Randy J; Herman, James P

    2015-06-01

    Stress-related (e.g., depression) and metabolic pathologies (e.g., obesity) are important and often co-morbid public health concerns. Here we identify a connection between peripheral glucocorticoid receptor (GR) signaling originating in fat with the brain control of both stress and metabolism. Mice with reduced adipocyte GR hypersecrete glucocorticoids following acute psychogenic stress and are resistant to diet-induced obesity. This hypersecretion gives rise to deficits in responsiveness to exogenous glucocorticoids, consistent with reduced negative feedback via adipocytes. Increased stress reactivity occurs in the context of elevated hypothalamic expression of hypothalamic-pituitary-adrenal (HPA) axis-excitatory neuropeptides and in the absence of altered adrenal sensitivity, consistent with a central cite of action. Our results identify a novel mechanism whereby activation of the adipocyte GR promotes peripheral energy storage while inhibiting the HPA axis, and provide functional evidence for a fat-to-brain regulatory feedback network that serves to regulate not just homeostatic energy balance but also responses to psychogenic stimuli. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Bile acid-induced arrhythmia is mediated by muscarinic M2 receptors in neonatal rat cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Siti H Sheikh Abdul Kadir

    Full Text Available BACKGROUND: Intrahepatic cholestasis of pregnancy (ICP is a common disease affecting up to 5% of pregnancies and which can cause fetal arrhythmia and sudden intrauterine death. We previously demonstrated that bile acid taurocholate (TC, which is raised in the bloodstream of ICP, can acutely alter the rate and rhythm of contraction and induce abnormal calcium destabilization in cultured neonatal rat cardiomyocytes (NRCM. Apart from their hepatic functions bile acids are ubiquitous signalling molecules with diverse systemic effects mediated by either the nuclear receptor FXR or by a recently discovered G-protein coupled receptor TGR5. We aim to investigate the mechanism of bile-acid induced arrhythmogenic effects in an in-vitro model of the fetal heart. METHODS AND RESULTS: Levels of bile acid transporters and nuclear receptor FXR were studied by quantitative real time PCR, western blot and immunostaining, which showed low levels of expression. We did not observe functional involvement of the canonical receptors FXR and TGR5. Instead, we found that TC binds to the muscarinic M(2 receptor in NRCM and serves as a partial agonist of this receptor in terms of inhibitory effect on intracellular cAMP and negative chronotropic response. Pharmacological inhibition and siRNA-knockdown of the M(2 receptor completely abolished the negative effect of TC on contraction, calcium transient amplitude and synchronisation in NRCM clusters. CONCLUSION: We conclude that in NRCM the TC-induced arrhythmia is mediated by the partial agonism at the M(2 receptor. This mechanism might serve as a promising new therapeutic target for fetal arrhythmia.

  5. 5-HT2 receptors mediate functional modulation of GABAa receptors and inhibitory synaptic transmissions in human iPS-derived neurons.

    Science.gov (United States)

    Wang, Haitao; Hu, Lingli; Liu, Chunhua; Su, Zhenghui; Wang, Lihui; Pan, Guangjin; Guo, Yiping; He, Jufang

    2016-02-03

    Neural progenitors differentiated from induced pluripotent stem cells (iPS) hold potentials for treating neurological diseases. Serotonin has potent effects on neuronal functions through multiple receptors, underlying a variety of neural disorders. Glutamate and GABA receptors have been proven functional in neurons differentiated from iPS, however, little is known about 5-HT receptor-mediated modulation in such neuronal networks. In the present study, human iPS were differentiated into cells possessing featured physiological properties of cortical neurons. Whole-cell patch-clamp recording was used to examine the involvement of 5-HT2 receptors in functional modulation of GABAergic synaptic transmission. We found that serotonin and DOI (a selective agonist of 5-HT2A/C receptor) reversibly reduced GABA-activated currents, and this 5-HT2A/C receptor mediated inhibition required G protein, PLC, PKC, and Ca(2+) signaling. Serotonin increased the frequency of miniature inhibitory postsynaptic currents (mIPSCs), which could be mimicked by α-methylserotonin, a 5-HT2 receptor agonist. In contrast, DOI reduced both frequency and amplitude of mIPSCs. These findings suggested that in iPS-derived human neurons serotonin postsynaptically reduced GABAa receptor function through 5-HT2A/C receptors, but presynaptically other 5-HT2 receptors counteracted the action of 5-HT2A/C receptors. Functional expression of serotonin receptors in human iPS-derived neurons provides a pre-requisite for their normal behaviors after grafting.

  6. AMPA receptor mediated excitotoxicity in neocortical neurons is developmentally regulated and dependent upon receptor desensitization

    DEFF Research Database (Denmark)

    Jensen, J B; Schousboe, A; Pickering, D S

    1998-01-01

    was blocked was seen as early as 5 DIV since 10 microM MK-801 did not completely block the response whereas 10 microM NBQX did. The 2,3-benzodiazepine GYKI compounds, which have been reported to be selective non-competitive AMPA receptor antagonists, were here observed to block the AMPA toxicity...

  7. Lebestatin, a disintegrin from Macrovipera venom, inhibits integrin-mediated cell adhesion, migration and angiogenesis.

    Science.gov (United States)

    Olfa, Kallech-Ziri; José, Luis; Salma, Daoud; Amine, Bazaa; Najet, Srairi Abid; Nicolas, Andreotti; Maxime, Lehmann; Raoudha, Zouari; Kamel, Mabrouk; Jacques, Marvaldi; Jean-Marc, Sabatier; Mohamed, El Ayeb; Naziha, Marrakchi

    2005-12-01

    Lebestatin, a new member of the lysine-threonine-serine (KTS)-disintegrin family, was purified to homogeneity from Tunisian snake (Macrovipera lebetina) venom. It is a single-chain polypeptide composed of 41 amino acids. The amino-acid sequence of lebestatin shows that it displays a pattern of cysteines similar to other short disintegrins, but contains the sequence KTS rather than RGD in its integrin-binding loop. Lebestatin presents a high homology with obtustatin and viperistatin. Lebestatin interacts specifically with the alpha1beta1 integrin. It was thus able to inhibit both adhesion and migration of PC12 and alpha1beta1 integrin-expressing CHO cells (CHO-alpha1) to type I and IV collagens. This disintegrin also affected adhesion and migration of endothelial cells and exhibited an anti-angiogenic effect in vivo when using the 8-day-old embryo chick chorioallantoic membrane model.

  8. Gap Junction–mediated Cell–Cell Communication Modulates Mouse Neural Crest Migration

    OpenAIRE

    Huang, G.Y.; Cooper, E.S.; Waldo, K.; Kirby, M L; Gilula, N B; Lo, C.W.

    1998-01-01

    Previous studies showed that conotruncal heart malformations can arise with the increase or decrease in α1 connexin function in neural crest cells. To elucidate the possible basis for the quantitative requirement for α1 connexin gap junctions in cardiac development, a neural crest outgrowth culture system was used to examine migration of neural crest cells derived from CMV43 transgenic embryos overexpressing α1 connexins, and from α1 connexin knockout (KO) mice and FC transgenic mice expressi...

  9. Habitat-mediated timing of migration in polar bears: an individual perspective.

    Science.gov (United States)

    Cherry, Seth G; Derocher, Andrew E; Lunn, Nicholas J

    2016-07-01

    Migration phenology is largely determined by how animals respond to seasonal changes in environmental conditions. Our perception of the relationship between migratory behavior and environmental cues can vary depending on the spatial scale at which these interactions are measured. Understanding the behavioral mechanisms behind population-scale movements requires knowledge of how individuals respond to local cues. We show how time-to-event models can be used to predict what factors are associated with the timing of an individual's migratory behavior using data from GPS collared polar bears (Ursus maritimus) that move seasonally between sea ice and terrestrial habitats. We found the concentration of sea ice that bears experience at a local level, along with the duration of exposure to these conditions, was most associated with individual migration timing. Our results corroborate studies that assume thresholds of >50% sea ice concentration are necessary for suitable polar bear habitat; however, continued periods (e.g., days to weeks) of exposure to suboptimal ice concentrations during seasonal melting were required before the proportion of bears migrating to land increased substantially. Time-to-event models are advantageous for examining individual movement patterns because they account for the idea that animals make decisions based on an accumulation of knowledge from the landscapes they move through and not simply the environment they are exposed to at the time of a decision. Understanding the migration behavior of polar bears moving between terrestrial and marine habitat, at multiple spatiotemporal scales, will be a major aspect of quantifying observed and potential demographic responses to climate-induced environmental changes.

  10. Stress Signals, Mediated by Membranous Glucocorticoid Receptor, Activate PLC/PKC/GSK-3β/β-catenin Pathway to Inhibit Wound Closure.

    Science.gov (United States)

    Jozic, Ivan; Vukelic, Sasa; Stojadinovic, Olivera; Liang, Liang; Ramirez, Horacio A; Pastar, Irena; Tomic Canic, Marjana

    2017-05-01

    Glucocorticoids (GCs), key mediators of stress signals, are also potent wound healing inhibitors. To understand how stress signals inhibit wound healing, we investigated the role of membranous glucocorticoid receptor (mbGR) by using cell-impermeable BSA-conjugated dexamethasone. We found that mbGR inhibits keratinocyte migration and wound closure by activating a Wnt-like phospholipase (PLC)/ protein kinase C (PKC) signaling cascade. Rapid activation of mbGR/PLC/PKC further leads to activation of known biomarkers of nonhealing found in patients, β-catenin and c-myc. Conversely, a selective inhibitor of PKC, calphostin C, blocks mbGR/PKC pathway, and rescues GC-mediated inhibition of keratinocyte migration in vitro and accelerates wound epithelialization of human wounds ex vivo. This novel signaling mechanism may have a major impact on understanding how stress response via GC signaling regulates homeostasis and its role in development and treatments of skin diseases, including wound healing. To test tissue specificity of this nongenomic signaling mechanism, we tested retinal and bronchial human epithelial cells and fibroblasts. We found that mbGR/PLC/PKC signaling cascade exists in all cell types tested, suggesting a more general role. The discovery of this nongenomic signaling pathway, in which glucocorticoids activate Wnt pathway via mbGR, provides new insights into how stress-mediated signals may activate growth signals in various epithelial and mesenchymal tissues. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  11. Retinoids enhance glucocorticoid-induced apoptosis of T cells by facilitating glucocorticoid receptor-mediated transcription

    Science.gov (United States)

    Tóth, K; Sarang, Z; Scholtz, B; Brázda, P; Ghyselinck, N; Chambon, P; Fésüs, L; Szondy, Z

    2011-01-01

    Glucocorticoid-induced apoptosis of thymocytes is one of the first recognized forms of programmed cell death. It was shown to require gene activation induced by the glucocorticoid receptor (GR) translocated into the nucleus following ligand binding. In addition, the necessity of the glucocorticoid-induced, but transcription-independent phosphorylation of phosphatidylinositol-specific phospholipase C (PI-PLC) has also been shown. Here we report that retinoic acids, physiological ligands for the nuclear retinoid receptors, enhance glucocorticoid-induced death of mouse thymocytes both in vitro and in vivo. The effect is mediated by retinoic acid receptor (RAR) alpha/retinoid X receptor (RXR) heterodimers, and occurs when both RARα and RXR are ligated by retinoic acids. We show that the ligated RARα/RXR interacts with the ligated GR, resulting in an enhanced transcriptional activity of the GR. The mechanism through which this interaction promotes GR-mediated transcription does not require DNA binding of the retinoid receptors and does not alter the phosphorylation status of Ser232, known to regulate the transcriptional activity of GR. Phosphorylation of PI-PLC was not affected. Besides thymocytes, retinoids also promoted glucocorticoid-induced apoptosis of various T-cell lines, suggesting that they could be used in the therapy of glucocorticoid-sensitive T-cell malignancies. PMID:21072052

  12. Enhanced nicotinic receptor mediated relaxations in gastroesophageal muscle fibers from Barrett's esophagus patients.

    Science.gov (United States)

    Miller, L S; Vegesna, A K; Braverman, A S; Barbe, M F; Ruggieri, M R

    2014-03-01

    Increased nicotinic receptor mediated relaxation in the gastroesophageal antireflux barrier may be involved in the pathophysiology of reflux. This study is designed to determine whether the defects we previously identified in gastroesophageal reflux disease patients in- vivo are due to abnormalities of the gastric sling, gastric clasp, or lower esophageal circular (LEC) muscle fibers. Muscle strips from whole stomachs and esophagi were obtained from 16 normal donors and 15 donors with histologically proven Barrett's esophagus. Contractile and relaxant responses of gastric sling, gastric clasp, or LEC fibers were determined to increasing concentrations of carbachol and to nicotine after inducing maximal contraction to bethanechol. Muscarinic receptor density was measured using subtype selective immunoprecipitation. Barrett's esophagus gastric sling and LEC fibers have decreased carbachol-induced contractions. Barrett's esophagus sling fibers have decreased M2 -muscarinic receptors and LEC fibers have decreased M3 receptors. Relaxations of all three fiber types are greater in Barrett's esophagus specimens to both high carbachol concentrations and to nicotine following bethanechol precontraction. The maximal response to bethanechol is greater in Barrett esophagus sling and LEC fibers. The increased contractile response to bethanechol in Barrett's specimens indicates that the defect is likely not due to the smooth muscle itself. The enhanced nicotinic receptor mediated response may be involved in greater relaxation of the muscles within the high-pressure zone of the gastroesophageal junction during transient lower esophageal sphincter relaxations and during deglutitive inhibition and may be involved in the pathophysiology of gastroesophageal reflux disease. © 2013 John Wiley & Sons Ltd.

  13. Chronic psychoemotional stress impairs cannabinoid-receptor-mediated control of GABA transmission in the striatum.

    Science.gov (United States)

    Rossi, Silvia; De Chiara, Valentina; Musella, Alessandra; Kusayanagi, Hajime; Mataluni, Giorgia; Bernardi, Giorgio; Usiello, Alessandro; Centonze, Diego

    2008-07-16

    Exposure to stressful events has a myriad of consequences in animals and in humans, and triggers synaptic adaptations in many brain areas. Stress might also alter cannabinoid-receptor-mediated transmission in the brain, but no physiological study has addressed this issue so far. In the present study, we found that social defeat stress, induced in mice by exposure to aggression, altered cannabinoid CB(1)-receptor-mediated control of synaptic transmission in the striatum. In fact, the presynaptic inhibition of GABAergic IPSCs induced by the cannabinoid CB(1) receptor agonist HU210 [(6aR)-trans-3-(1,1-dimethylheptyl)-6a,7,10,10a-tetrahydro-1-hydroxy-6,6-dimethyl-6H-dibenzo[b,d]pyran-9-methanol] was reduced after a single stressful episode and fully abolished after 3 and 7 d of stress exposure. Repeated psychoemotional stress also impaired the sensitivity of GABA synapses to endocannabinoids mobilized by group I metabotropic glutamate receptor stimulation, whereas the cannabinoid CB(1)-mediated control of glutamate transmission was unaffected by repeated exposure to an aggressor. Corticosteroids released in response to the activation of the hypothalamic-pituitary-adrenal axis played a major role in the synaptic defects observed in stressed animals, because these alterations were fully prevented by pharmacological blockade of glucocorticoid receptors and were mimicked by corticosterone injections. The recovery of stress-induced synaptic defects was favored when stressed mice were given access to a running wheel or to sucrose consumption, which function as potent natural rewards. A similar rescuing effect was obtained by a single injection of cocaine, a psychostimulant with strong rewarding properties. Targeting cannabinoid CB(1) receptors or endocannabinoid metabolism might be a valuable option to treat stress-associated neuropsychiatric conditions.

  14. Receptor-Mediated Drug Delivery to Macrophages in Chemotherapy of Leishmaniasis

    Science.gov (United States)

    Mukhopadhyay, Amitabha; Chaudhuri, Gautam; Arora, Sunil K.; Sehgal, Shobha; Basu, Sandip K.

    1989-05-01

    Methotrexate coupled to maleylated bovine serum albumin was taken up efficiently through the ``scavenger'' receptors present on macrophages and led to selective killing of intracellular Leishmania mexicana amazonensis amastigotes in cultured hamster peritoneal macrophages. The drug conjugate was nearly 100 times as effective as free methotrexate in eliminating the intracellular parasites. Furthermore, in a model of experimental cutaneous leishmaniasis in hamsters, the drug conjugate brought about more than 90% reduction in the size of footpad lesions within 11 days. In contrast, the free drug at a similar concentration did not significantly affect lesion size. These studies demonstrate the potential of receptor-mediated drug delivery in the therapy of macrophage-associated diseases.

  15. Prostaglandin receptor EP3 regulates cell proliferation and migration with impact on survival of endometrial cancer patients.

    Science.gov (United States)

    Zhu, Junyan; Trillsch, Fabian; Mayr, Doris; Kuhn, Christina; Rahmeh, Martina; Hofmann, Simone; Vogel, Marianne; Mahner, Sven; Jeschke, Udo; von Schönfeldt, Viktoria

    2018-01-02

    Prostaglandin E2 (PGE2) receptor 3 (EP3) regulates tumor cell proliferation, migration, and invasion in numerous cancers. The role of EP3 as a prognostic biomarker in endometrial cancer remains unclear. The primary aim of this study was to analyze the prognostic significance of EP3 expression in endometrial cancer. We analyzed the EP3 expression of 140 endometrial carcinoma patients by immunohistochemistry. RL95-2 endometrial cancer cell line was chosen from four endometrial cancer cell lines (RL95-2, Ishikawa, HEC-1-A, and HEC-1-B) according to EP3 expression level. Treated with PGE2 and EP3 antagonist, RL95-2 cells were investigated by MTT, BrdU, and wound healing assay for functional assessment of EP3. EP3 staining differed significantly according to WHO tumor grading in both whole cohort (p = 0.01) and the subgroup of endometrioid carcinoma (p = 0.01). Patients with high EP3 expression in their respective tumors had impaired progression-free survival as well as overall survival in both cohorts above. EP3 expression in the overall cohort was identified as an independent prognostic marker for progression-free survival (HR 1.014, 95%CI 1.003-1.024, p = 0.01) when adjusted for age, stage, grading, and recurrence. Treatment with EP3 antagonists induced upregulation of estrogen receptor β and decreased activity of Ras and led to attenuated proliferation and migration of RL95-2 cells. EP3 seems to play a crucial role in endometrial cancer progression. In the context of limited systemic treatment options for endometrial cancer, this explorative analysis identifies EP3 as a potential target for diagnostic workup and therapy.

  16. Chromatin organization regulated by EZH2-mediated H3K27me3 is required for OPN-induced migration of bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Liu, Lingling; Luo, Qing; Sun, Jinghui; Ju, Yang; Morita, Yasuyuki; Song, Guanbin

    2018-03-01

    Osteopontin (OPN) is a chemokine-like extracellular matrix-associated protein involved in the migration of bone marrow-derived mesenchymal stem cells (BMSCs). An increasing number of studies have found that chromatin organization may affect cellular migration. However, whether OPN regulates chromatin organization is not understood, nor are the underlying molecular mechanisms. In this study, we investigated the link between chromatin organization and BMSC migration and demonstrated that OPN-mediated BMSC migration leads to elevated levels of heterochromatin marker histone H3 lysine 27 trimethylation (H3K27me3) through the methyltransferase EZH2. The expression of EZH2 reorganizes the chromatin structure of BMSCs. Pharmacological inhibition or depletion of EZH2 blocks BMSC migration. Moreover, using an atomic force microscope (AFM), we found that chromatin decondensation alters the mechanical properties of the nucleus. In addition, inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) signals represses OPN-promoted chromatin condensation and cell migration. Thus, our results identify a mechanism by which ERK1/2 signalling drives specific chromatin modifications in BMSCs, which alters chromatin organization and thereby enables OPN-mediated BMSC migration. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. bFGF-Regulating MAPKs Are Involved in High Glucose-Mediated ROS Production and Delay of Vascular Endothelial Cell Migration.

    Directory of Open Access Journals (Sweden)

    Zhong Xin Zhu

    Full Text Available High blood sugar is a symptom of diabetes mellitus (DM. Vascular endothelial cells (VECs directly contact the blood and are damaged when blood sugar levels are high. However, the molecular mechanism underlying this process remains elusive. To analyze the effects of DM on migration, we simulated DM by applying high glucose (HG to the human VEC. HG delayed cell migration and induced phosphorylation of MAPKs (JNK and ERK. By contrast, in presence of bFGF, cell migration was promoted and MAPK phosphorylation levels were reduced. Furthermore, treatment with JNK and ERK inhibitors rescued HG-mediated delay of cell migration. Molecular and cell biological studies demonstrated that HG increased ROS production, whereas treatment with bFGF or JNK/ERK inhibitors blocked HG-induced ROS accumulation. Addition of MnTMPyP, a ROS scavenger, reduced HG-induced ROS production and accelerated cell migration, suggesting that the influence of HG on bFGF-MAPK signaling causes accumulation of ROS, which in turn regulate cell migration. This is the first study to elucidate the molecular mechanism of HG-mediated VEC migration; these findings could facilitate the development of novel therapies for DM.

  18. Independent regulation of α1 and α2 adrenergic receptor-mediated vasoconstriction in vivo.

    Science.gov (United States)

    Muszkat, Mordechai; Kurnik, Daniel; Sofowora, Gbenga G; Wood, Alastair J J; Stein, C Michael

    2011-02-01

    Vascular α1 and α2 adrenergic receptors mediate vasoconstriction and are major determinants of peripheral vascular tone. There is a wide variability in vasoconstrictor sensitivity to α1 and α2 adrenergic receptor agonists among individuals. In previous studies, this variability was not explained by identified α1 and α2 adrenergic receptor genetic variants. Thus, we hypothesized that adrenergic vasoconstrictor sensitivity is determined by shared constrictor mechanisms downstream of the individual receptors and that α1 and α2 adrenergic receptor-mediated vasoconstrictor sensitivity would therefore be correlated. Dorsal hand vein responses to increasing doses of the α1 adrenergic receptor agonist phenylephrine (12-12 000 ng/min) and the α2 adrenergic receptor agonist dexmedetomidine (0.01-100 ng/min) were measured in healthy individuals using a linear variable differential transformer. From individual dose-response curves, we calculated the dose of phenylephrine and dexmedetomidine that produced 50% (ED50) of maximum venoconstriction (Emax) for each patient. We examined the correlation between phenylephrine and dexmedetomidine ED50 and Emax before and after adjustment for covariates (age, sex, ethnicity, BMI, blood pressure, heart rate, and baseline plasma norepinephrine concentrations). In 62 patients (36 men, 34 African-American, 28 whites), the median ED50 for dexmedetomidine was 1.32 ng/min [interquartile range (IQR) 0.45-5.37 ng/min] and for phenylephrine 177.8 ng/min (IQR 40.7-436.5 ng/min). The Emax for phenylephrine was 90.8% (82.2-99.6%) and for dexmedetomidine 80.0% (64.7-95.2%). There was no correlation between individual sensitivities (ED50) to phenylephrine and dexmedetomidine, before and after adjustment for covariates (P > 0.30). Both phenylephrine and dexmedetomidine produce strong venoconstriction in the dorsal hand vein; however, there is no significant correlation between vascular sensitivity to an α1 and α2 adrenergic receptor agonist

  19. Safety Pharmacology assessment of drugs with biased 5-HT(2B) receptor agonism mediating cardiac valvulopathy.

    Science.gov (United States)

    Cavero, Icilio; Guillon, Jean-Michel

    2014-01-01

    The rhythmic opening and tightly closing of cardiac valve leaflets are cardiac cyclic events imposing to blood a unidirectional course along the vascular tree. Drugs with 5-HT2B agonism properties can seriously compromise this biological function critical for hemodynamic efficiency as their intrinsic pro-fibrotic effects can, with time, make valvular coaptation blood regurgitant. Cardiac valve anatomy, physiology and pathology as well as 5-HT2B receptor properties (coupling, effects mediated, biased agonism) are briefly exposed. Approaches to unveil 5-HT2B receptor liability of drug candidates are detailed. In silico computational models can rapidly probe molecules for chemical signatures associated with 5-HT2B receptor affinity. In vitro radioligand competition assays allow quantifying receptor binding capacity (Ki, IC50), the pharmacological nature (agonism, antagonism) of which can be ascertained from cytosolic second messenger (inositol phosphates, Ca(++), MAPK2) changes. Potencies calculated from the latter data may exhibit variability as they are dependent upon the readout measured and the experimental conditions (e.g., receptor density level of cell material expressing human 5-HT2B receptors). The in vivo valvulopathy effects of 5-HT2B receptor agonists can be assessed by echocardiographic measurements and valve histology in rats chronically treated with the candidate drug. Finally, safety margins derived from from nonclinical and clinical data are evaluated in terms of the readout, usefulness and scientific reliability. The Safety Pharmacology toolbox for detecting possible 5-HT2B receptor agonism liabilities of candidate drugs requires meticulous optimization and validation of all its (in silico, in vitro and in vivo) components to perfect its human predictability power. In particular, since 5-HT2B receptor agonism is biased in nature, the most predictive readout(s) of valvular liability should be identified and prioritized in keeping with best scientific

  20. Protein kinase C α enhances migration of breast cancer cells through FOXC2-mediated repression of p120-catenin.

    Science.gov (United States)

    Pham, Thao N D; Perez White, Bethany E; Zhao, Huiping; Mortazavi, Fariborz; Tonetti, Debra A

    2017-12-07

    Despite recent advances in the diagnosis and treatment of breast cancer, metastasis remains the main cause of death. Since migration of tumor cells is considered a prerequisite for tumor cell invasion and metastasis, a pressing goal in tumor biology has been to elucidate factors regulating their migratory activity. Protein kinase C alpha (PKCα) is a serine-threonine protein kinase implicated in cancer metastasis and associated with poor prognosis in breast cancer patients. In this study, we set out to define the signaling axis mediated by PKCα to promote breast cancer cell migration. Oncomine™ overexpression analysis was used to probe for PRKCA (PKCα) and FOXC2 expression in mRNA datasets. The heat map of PRKCA, FOXC2, and CTNND1 were obtained from the UC Santa Cruz platform. Survival data were obtained by PROGgene and available at http://www.compbio.iupui.edu/proggene . Markers for EMT and adherens junction were assessed by Western blotting and quantitative polymerase chain reaction. Effects of PKCα and FOXC2 on migration and invasion were assessed in vitro by transwell migration and invasion assays respectively. Cellular localization of E-cadherin and p120-catenin was determined by immunofluorescent staining. Promoter activity of p120-catenin was determined by dual luciferase assay using a previously validated p120-catenin reporter construct. Interaction between FOXC2 and p120-catenin promoter was verified by chromatin immunoprecipitation assay. We determined that PKCα expression is necessary to maintain the migratory and invasive phenotype of both endocrine resistant and triple negative breast cancer cell lines. FOXC2 acts as a transcriptional repressor downstream of PKCα, and represses p120-catenin expression. Consequently, loss of p120-catenin leads to destabilization of E-cadherin at the adherens junction. Inhibition of either PKCα or FOXC2 is sufficient to rescue p120-catenin expression and trigger relocalization of p120-catenin and E-cadherin to

  1. Aspirin Inhibits Platelet-Derived Sphingosine-1-Phosphate Induced Endothelial Cell Migration.

    Science.gov (United States)

    Polzin, Amin; Knoop, Betül; Böhm, Andreas; Dannenberg, Lisa; Zurek, Mark; Zeus, Tobias; Kelm, Malte; Levkau, Bodo; Rauch, Bernhard H

    2018-01-01

    Aspirin plays a crucial role in the prevention of cardiovascular diseases. We previously described that aspirin has effects beyond inhibition of platelet aggregation, as it inhibited thrombin-mediated release of sphingosine-1-phosphate (S1P) from human platelets. S1P is a bioactive lipid with important functions on inflammation and apoptosis. In endothelial cells (EC), S1P is a key regulator of cell migration. In this study, we aimed to analyze the effects of aspirin on platelet-induced EC migration. Human umbilical EC migration was measured by Boyden chamber assay. EC migration was induced by platelet supernatants of thrombin receptor-activating peptide-1 (AP1) stimulated platelets. To investigate the S1P receptor subtype that promotes EC migration, specific inhibitors of S1P receptor subtypes were applied. S1P induced EC migration in a concentration-dependent manner. EC migration induced by AP1-stimulated platelet supernatants was reduced by aspirin. S1P1 receptor inhibition almost completely abolished EC migration induced by activated platelets. The inhibition of S1P2 or S1P3 receptor had no effect. Aspirin inhibits EC migration induced by activated platelets that is in part due to S1P and mediated by the endothelial S1P1 receptor. The clinical significance of this novel mechanism of aspirin action has to be investigated in future studies. © 2017 S. Karger AG, Basel.

  2. Gastrin-Releasing Peptide Receptor Mediates Activation of the Epidermal Growth Factor Receptor in Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Sufi Mary Thomas

    2005-04-01

    Full Text Available Gastrin-releasing peptide receptor (GRPR and the epidermal growth factor receptor (EGFR are expressed in several cancers including non-small cell lung carcinoma (NSCLC. Here we demonstrate the activation of EGFR by the GRPR ligand, gastrin-releasing peptide (GRP, in NSCLC cells. GRP induced rapid activation of p44/42 MAPK in lung cancer cells through EGFR. GRP-mediated activation of MAPK in NSCLC cells was abrogated by pretreatment with the anti-EGFR-neutralizing antibody, C225. Pretreatment of NSCLC cells with neutralizing antibodies to the EGFR ligands, TGF-α or HB-EGF, also decreased GRP-mediated MAPK activation. On matrix metalloproteinase (MMP inhibition, GRP failed to activate MAPK in NSCLC cells. EGF and GRP both stimulated NSCLC proliferation, and inhibition of either EGFR or GRPR resulted in cell death. Combining a GRPR antagonist with the EGFR tyrosine kinase inhibitor, gefitinib, resulted in additive cytotoxic effects. Additive effects were seen at gefitinib concentrations from 1 to 18μM, encompassing the ID50 values of both gefitinib-sensitive and gefitinib-resistant NSCLC cell lines. Because a major effect of GRPR appears to be promoting the release of EGFR ligand, this study suggests that a greater inhibition of cell proliferation may occur by abrogating EGFR ligand release in consort with inhibition of EGFR.

  3. Exosomal signaling during hypoxia mediates microvascular endothelial cell migration and vasculogenesis.

    Directory of Open Access Journals (Sweden)

    Carlos Salomon

    Full Text Available Vasculogenesis and angiogenesis are critical processes in fetal circulation and placental vasculature development. Placental mesenchymal stem cells (pMSC are known to release paracrine factors (some of which are contained within exosomes that promote angiogenesis and cell migration. The aims of this study were: to determine the effects of oxygen tension on the release of exosomes from pMSC; and to establish the effects of pMSC-derived exosomes on the migration and angiogenic tube formation of placental microvascular endothelial cells (hPMEC. pMSC were isolated from placental villi (8-12 weeks of gestation, n = 6 and cultured under an atmosphere of 1%, 3% or 8% O2. Cell-conditioned media were collected and exosomes (exo-pMSC isolated by differential and buoyant density centrifugation. The dose effect (5-20 µg exosomal protein/ml of pMSC-derived exosomes on hPMEC migration and tube formation were established using a real-time, live-cell imaging system (Incucyte™. The exosome pellet was resuspended in PBS and protein content was established by mass spectrometry (MS. Protein function and canonical pathways were identified using the PANTHER program and Ingenuity Pathway Analysis, respectively. Exo-pMSC were identified, by electron microscopy, as spherical vesicles, with a typical cup-shape and diameters around of 100 nm and positive for exosome markers: CD63, CD9 and CD81. Under hypoxic conditions (1% and 3% O2 exo-pMSC released increased by 3.3 and 6.7 folds, respectively, when compared to the controls (8% O2; p<0.01. Exo-pMSC increased hPMEC migration by 1.6 fold compared to the control (p<0.05 and increased hPMEC tube formation by 7.2 fold (p<0.05. MS analysis identified 390 different proteins involved in cytoskeleton organization, development, immunomodulatory, and cell-to-cell communication. The data obtained support the hypothesis that pMSC-derived exosomes may contribute to placental vascular adaptation to low oxygen tension under both

  4. Silencing Mediator of Retinoid and Thyroid Hormone Receptors (SMRT) regulates glucocorticoid action in adipocytes.

    Science.gov (United States)

    Emont, Margo P; Mantis, Stelios; Kahn, Jonathan H; Landeche, Michael; Han, Xuan; Sargis, Robert M; Cohen, Ronald N

    2015-05-15

    Local modulation of glucocorticoid action in adipocytes regulates adiposity and systemic insulin sensitivity. However, the specific cofactors that mediate glucocorticoid receptor (GR) action in adipocytes remain unclear. Here we show that the silencing mediator of retinoid and thyroid hormone receptors (SMRT) is recruited to GR in adipocytes and regulates ligand-dependent GR function. Decreased SMRT expression in adipocytes in vivo increases expression of glucocorticoid-responsive genes. Moreover, adipocytes with decreased SMRT expression exhibit altered glucocorticoid regulation of lipolysis. We conclude that SMRT regulates the metabolic functions of GR in adipocytes in vivo. Modulation of GR-SMRT interactions in adipocytes represents a novel approach to control the local degree of glucocorticoid action and thus influence adipocyte metabolic function. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  5. The Role of PPAR Receptors and Leukotriene B4 Receptors in Mediating the Effects of LY293111 in Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    Thomas E. Adrian

    2008-01-01

    Full Text Available Pancreatic cancer is a devastating disease in which current therapies are inadequate. Separate lines of research have identified the 5-lipoxygenase/leukotriene B4 receptor pathway and the PPAR pathway as potential targets for prevention or treatment of this disease. LY293111 was originally designed as a potent leukotriene B4 receptor antagonist for treatment of inflammatory conditions. LY293111 was also known to have inhibitory effects on 5-lipoxygenase, which is upstream of the production of leukotrienes. LY293111 was shown to have potent anticancer effects in pancreatic cancer and several other solid malignancies, where it caused cell cycle arrest and marked apoptosis. Subsequently, it came to light that LY293111 exhibited PPAR agonist activity in addition to its effects on the 5-lipoxygenase pathway. This raises the question of which of the two targets is of greatest importance with regard to the anticancer effects of this agent. The evidence to date is not conclusive, but suggests that the effects of LY293111 may be mediated by both LTB4 receptors and PPAR.

  6. Crosstalk Between Leptin Receptor and IGF-IR in Breast Cancer: A Potential Mediator of Chemoresistance

    Science.gov (United States)

    2011-04-01

    patients, regardless of menopausal status or age (2). Although the exact molecular mechanisms by which obesity influences cancer biology are unknown...Leptin Receptor and IGF-IR in Breast Cancer: A Potential Mediator of Chemoresistance Dr. Rita Nahta Emory University Atlanta, GA 30322 Obesity ...is a major risk factor for breast cancer, and is associated with reduced treatment response and reduced overall survival. The obesity -associated

  7. An Impermeant Ganetespib Analog Inhibits Extracellular Hsp90-Mediated Cancer Cell Migration that Involves Lysyl Oxidase 2-like Protein

    Directory of Open Access Journals (Sweden)

    Jessica McCready

    2014-04-01

    Full Text Available Extracellular Hsp90 (eHsp90 activates a number of client proteins outside of cancer cells required for migration and invasion. Therefore, eHsp90 may serve as a novel target for anti-metastatic drugs as its inhibition using impermeant Hsp90 inhibitors would not affect the numerous vital intracellular Hsp90 functions in normal cells. While some eHsp90 clients are known, it is important to establish other proteins that act outside the cell to validate eHsp90 as a drug target to limit cancer spread. Using mass spectrometry we identified two precursor proteins Galectin 3 binding protein (G3BP and Lysyl oxidase 2-like protein (LOXL2 that associate with eHsp90 in MDA-MB231 breast cancer cell conditioned media and confirmed that LOXL2 binds to eHsp90 in immunoprecipitates. We introduce a novel impermeant Hsp90 inhibitor STA-12-7191 derived from ganetespib and show that it is markedly less toxic to cells and can inhibit cancer cell migration in a dose dependent manner. We used STA-12-7191 to test if LOXL2 and G3BP are potential eHsp90 clients. We showed that while LOXL2 can increase wound healing and compensate for STA-12-7191-mediated inhibition of wound closure, addition of G3BP had no affect on this assay. These findings support of role for LOXL2 in eHsp90 stimulated cancer cell migration and provide preliminary evidence for the use of STA-12-7191 to inhibit eHsp90 to limit cancer invasion.

  8. An Impermeant Ganetespib Analog Inhibits Extracellular Hsp90-Mediated Cancer Cell Migration that Involves Lysyl Oxidase 2-like Protein

    Energy Technology Data Exchange (ETDEWEB)

    McCready, Jessica [Department of Natural Sciences, Assumption College, Worcester, MA 01609 (United States); Wong, Daniel S. [Department of Developmental Molecular and Chemical Biology, Tufts University School of Medicine, Boston, MA 02111 (United States); Cell and Molecular Physiology Program, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, MA 02111 (United States); Burlison, Joseph A.; Ying, Weiwen [Synta Pharmaceuticals, Lexington, MA 02421 (United States); Jay, Daniel G., E-mail: daniel.jay@tufts.edu [Department of Developmental Molecular and Chemical Biology, Tufts University School of Medicine, Boston, MA 02111 (United States); Cell and Molecular Physiology Program, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, MA 02111 (United States)

    2014-04-30

    Extracellular Hsp90 (eHsp90) activates a number of client proteins outside of cancer cells required for migration and invasion. Therefore, eHsp90 may serve as a novel target for anti-metastatic drugs as its inhibition using impermeant Hsp90 inhibitors would not affect the numerous vital intracellular Hsp90 functions in normal cells. While some eHsp90 clients are known, it is important to establish other proteins that act outside the cell to validate eHsp90 as a drug target to limit cancer spread. Using mass spectrometry we identified two precursor proteins Galectin 3 binding protein (G3BP) and Lysyl oxidase 2-like protein (LOXL2) that associate with eHsp90 in MDA-MB231 breast cancer cell conditioned media and confirmed that LOXL2 binds to eHsp90 in immunoprecipitates. We introduce a novel impermeant Hsp90 inhibitor STA-12-7191 derived from ganetespib and show that it is markedly less toxic to cells and can inhibit cancer cell migration in a dose dependent manner. We used STA-12-7191 to test if LOXL2 and G3BP are potential eHsp90 clients. We showed that while LOXL2 can increase wound healing and compensate for STA-12-7191-mediated inhibition of wound closure, addition of G3BP had no affect on this assay. These findings support of role for LOXL2 in eHsp90 stimulated cancer cell migration and provide preliminary evidence for the use of STA-12-7191 to inhibit eHsp90 to limit cancer invasion.

  9. Lentivirus-Mediated RNA Interference Targeting RhoA Slacks the Migration, Proliferation, and Myelin Formation of Schwann Cells.

    Science.gov (United States)

    Wen, Jinkun; Qian, Changhui; Pan, Mengjie; Wang, Xianghai; Li, Yuanyuan; Lu, Yanmeng; Zhou, Zhitao; Yan, Qing; Li, Lixia; Liu, Zhongying; Wu, Wutian; Guo, Jiasong

    2017-03-01

    RhoA, a member of Rho GTPases family, is known to play an important role in remodeling actin cytoskeleton. During the development of the peripheral nervous system (PNS), Schwann cells undergo proliferation, migration, and radial sorting and finally wrap the related axons compactly to form myelin sheath. All these processes involve actin cytoskeletal remodeling. However, the role of RhoA on Schwann cell during development is still unclear. To address this question, we first used a lentiviral vector-mediated short hairpin (sh) RNA targeting RhoA to knock down the expression of RhoA in the cultured Schwann cells in vitro. Effects of RhoA on Schwann cell proliferation and migration were examined by BrdU assay and transwell assay, respectively. Results of the present study indicated that downregulated RhoA expression in cultured Schwann cells significantly slacked the cells' capabilities of migration and proliferation. Then, we investigated the role of RhoA in the developing rat sciatic nerves. Immunohistology and Western blotting showed that RhoA was mainly expressed in Schwann cells in the sciatic nerves and was peaked at 2 weeks postnatal then kept in low level up to 8 weeks. In the subjected rats whose sciatic nerves were microinjected with lentiviral vectors at postnatal 3 days, we found that the lentiviruses mainly transfected Schwann cells, and the RhoA expression in the transfected Schwann cells was significantly knocked down. Four weeks after lentivirus microinjection, immunohistology and transmission electron microscopy illustrated that RhoA knockdown resulted in hypomyelination and significant decrease of the thickness of myelin in the transfected area. Overall data of current study suggested that RhoA plays a critical role in Schwann cell biology and is essential for myelination in developing peripheral nerve.

  10. EZH2-mediated epigenetic silencing of TIMP2 promotes ovarian cancer migration and invasion.

    Science.gov (United States)

    Yi, Xiaoqing; Guo, Jianfeng; Guo, Jing; Sun, Si; Yang, Ping; Wang, Junjie; Li, Yuan; Xie, Lisha; Cai, Jing; Wang, Zehua

    2017-06-15

    Enhancer of zeste homolog 2 (EZH2) is often increased in malignant tumors and is involved in metastasis. EZH2 silences gene expression by tri-methylating the lysine 27 residue of histone H3 (H3K27me3). However, the mechanism underlying EZH2 promotion of ovarian cancer metastasis remains elusive. Here, we showed that EZH2 is up-regulated in ovarian cancer and is associated with tumor metastasis and poor survival by mRNA sequencing and microarray results from databases. Tissue microarray and immunohistochemistry results revealed that EZH2 was negatively correlated with the expression of tissue inhibitor of metalloproteinases 2 (TIMP2). EZH2 overexpression inhibited TIMP2 expression and promoted proteolytic activities of matrix metalloproteinases 2 and 9 and vice versa. EZH2 promoted ovarian cancer invasion and migration, which could be largely reversed by TIMP2 down-regulation in vitro and in vivo. Both H3K27me3 inhibition and demethylation could reduce methylation of the TIMP2 promoter and finally reactivate TIMP2 transcription. The presence of EZH2 and H3K27me3 at the TIMP2 promoter was confirmed by chromatin immunoprecipitation. H3K27me3 and DNA methyltransferases at the promoter were significantly increased by EZH2 overexpression. These results suggest that EZH2 inhibits TIMP2 expression via H3K27me3 and DNA methylation, which relieve the repression of MMP and facilitate ovarian cancer invasion and migration.

  11. Expression of CD74, the receptor for macrophage migration inhibitory factor, in non-small cell lung cancer.

    Science.gov (United States)

    McClelland, Marc; Zhao, Liujian; Carskadon, Shannon; Arenberg, Douglas

    2009-02-01

    Macrophage migration inhibitory factor (MIF) is a multifunctional cytokine that is overexpressed in lung cancer. The MIF receptor was recently discovered and found to be the invariant chain of the HLA class II molecule, CD74. We hypothesized that the expression of this receptor-ligand pair in lung cancer is associated with the angiogenic activity and level of CXC chemokine expression in human specimens of non-small cell lung cancer. We, therefore, performed immunolocalization of CD74 and compared it with the localization of MIF in non-small cell lung cancer to determine their respective locations, as well as the relationship between the co-expression of MIF-CD74 and angiogenic CXC chemokines with tumor angiogenesis. We found intense CD74 expression by immunohistochemistry in 57 of 70 tumors with minimal to no staining in the remaining 13 tumors. Comparing the localization of CD74 with its putative ligand, MIF, we found that CD74 and MIF were co-expressed in tumors in close proximity, and that co-expression of the MIF-CD74 pair was associated with both higher levels of tumor-associated angiogenic CXC chemokines (ie, the ELR score) and greater vascularity compared with tumors in which MIF-CD74 co-expression was not present. We also found that MIF induced angiogenic CXC chemokine expression in an autocrine manner in vitro, a function that was specifically inhibited by antibodies to CD74.

  12. Expansion of Sphingosine Kinase and Sphingosine-1-Phosphate Receptor Function in Normal and Cancer Cells: From Membrane Restructuring to Mediation of Estrogen Signaling and Stem Cell Programming.

    Science.gov (United States)

    Sukocheva, Olga A

    2018-01-31

    Sphingolipids, sphingolipid metabolizing enzymes, and their receptors network are being recognized as part of the signaling mechanisms, which govern breast cancer cell growth, migration, and survival during chemotherapy treatment. Approximately 70% of breast cancers are estrogen receptor (ER) positive and, thus, rely on estrogen signaling. Estrogen activates an intracellular network composed of many cytoplasmic and nuclear mediators. Some estrogen effects can be mediated by sphingolipids. Estrogen activates sphingosine kinase 1 (SphK1) and amplifies the intracellular concentration of sphingosine-1-phosphate (S1P) in breast cancer cells during stimulation of proliferation and survival. Specifically, Estrogen activates S1P receptors (S1PR) and induces growth factor receptor transactivation. SphK, S1P, and S1PR expression are causally associated with endocrine resistance and progression to advanced tumor stages in ER-positive breast cancers in vivo . Recently, the network of SphK/S1PR was shown to promote the development of ER-negative cancers and breast cancer stem cells, as well as stimulating angiogenesis. Novel findings confirm and broaden our knowledge about the cross-talk between sphingolipids and estrogen network in normal and malignant cells. Current S1PRs therapeutic inhibition was indicated as a promising chemotherapy approach in non-responsive and advanced malignancies. Considering that sphingolipid signaling has a prominent role in terminally differentiated cells, the impact should be considered when designing specific SphK/S1PR inhibitors. This study analyzes the dynamic of the transformation of sphingolipid axis during a transition from normal to pathological condition on the level of the whole organism. The sphingolipid-based mediation and facilitation of global effects of estrogen were critically accented as a bridging mechanism that should be explored in cancer prevention.

  13. Expansion of Sphingosine Kinase and Sphingosine-1-Phosphate Receptor Function in Normal and Cancer Cells: From Membrane Restructuring to Mediation of Estrogen Signaling and Stem Cell Programming

    Directory of Open Access Journals (Sweden)

    Olga A. Sukocheva

    2018-01-01

    Full Text Available Sphingolipids, sphingolipid metabolizing enzymes, and their receptors network are being recognized as part of the signaling mechanisms, which govern breast cancer cell growth, migration, and survival during chemotherapy treatment. Approximately 70% of breast cancers are estrogen receptor (ER positive and, thus, rely on estrogen signaling. Estrogen activates an intracellular network composed of many cytoplasmic and nuclear mediators. Some estrogen effects can be mediated by sphingolipids. Estrogen activates sphingosine kinase 1 (SphK1 and amplifies the intracellular concentration of sphingosine-1-phosphate (S1P in breast cancer cells during stimulation of proliferation and survival. Specifically, Estrogen activates S1P receptors (S1PR and induces growth factor receptor transactivation. SphK, S1P, and S1PR expression are causally associated with endocrine resistance and progression to advanced tumor stages in ER-positive breast cancers in vivo. Recently, the network of SphK/S1PR was shown to promote the development of ER-negative cancers and breast cancer stem cells, as well as stimulating angiogenesis. Novel findings confirm and broaden our knowledge about the cross-talk between sphingolipids and estrogen network in normal and malignant cells. Current S1PRs therapeutic inhibition was indicated as a promising chemotherapy approach in non-responsive and advanced malignancies. Considering that sphingolipid signaling has a prominent role in terminally differentiated cells, the impact should be considered when designing specific SphK/S1PR inhibitors. This study analyzes the dynamic of the transformation of sphingolipid axis during a transition from normal to pathological condition on the level of the whole organism. The sphingolipid-based mediation and facilitation of global effects of estrogen were critically accented as a bridging mechanism that should be explored in cancer prevention.

  14. The miR-199-dynamin regulatory axis controls receptor-mediated endocytosis.

    Science.gov (United States)

    Aranda, Juan F; Canfrán-Duque, Alberto; Goedeke, Leigh; Suárez, Yajaira; Fernández-Hernando, Carlos

    2015-09-01

    Small non-coding RNAs (microRNAs) are important regulators of gene expression that modulate many physiological processes; however, their role in regulating intracellular transport remains largely unknown. Intriguingly, we found that the dynamin (DNM) genes, a GTPase family of proteins responsible for endocytosis in eukaryotic cells, encode the conserved miR-199a and miR-199b family of miRNAs within their intronic sequences. Here, we demonstrate that miR-199a and miR-199b regulate endocytic transport by controlling the expression of important mediators of endocytosis such as clathrin heavy chain (CLTC), Rab5A, low-density lipoprotein receptor (LDLR) and caveolin-1 (Cav-1). Importantly, miR-199a-5p and miR-199b-5p overexpression markedly inhibits CLTC, Rab5A, LDLR and Cav-1 expression, thus preventing receptor-mediated endocytosis in human cell lines (Huh7 and HeLa). Of note, miR-199a-5p inhibition increases target gene expression and receptor-mediated endocytosis. Taken together, our work identifies a new mechanism by which microRNAs regulate intracellular trafficking. In particular, we demonstrate that the DNM, miR-199a-5p and miR-199b-5p genes act as a bifunctional locus that regulates endocytosis, thus adding an unexpected layer of complexity in the regulation of intracellular trafficking. © 2015. Published by The Company of Biologists Ltd.

  15. APP regulates NGF receptor trafficking and NGF-mediated neuronal differentiation and survival.

    Directory of Open Access Journals (Sweden)

    Yun-wu Zhang

    Full Text Available β-Amyloid precursor protein (APP is a key factor in Alzheimer's disease (AD but its physiological function is largely undetermined. APP has been found to regulate retrograde transport of nerve growth factor (NGF, which plays a crucial role in mediating neuronal survival and differentiation. Herein, we reveal the mechanism underlying APP-mediated NGF trafficking, by demonstrating a direct interaction between APP and the two NGF receptors, TrkA and p75NTR. Downregulation of APP leads to reduced cell surface levels of TrkA/p75NTR and increased endocytosis of TrkA/p75NTR and NGF. In addition, APP-deficient cells manifest defects in neurite outgrowth and are more susceptible to Aβ-induced neuronal death at physiological levels of NGF. However, APP-deficient cells show better responses to NGF-stimulated differentiation and survival than control cells. This may be attributed to increased receptor endocytosis and enhanced activation of Akt and MAPK upon NGF stimulation in APP-deficient cells. Together, our results suggest that APP mediates endocytosis of NGF receptors through direct interaction, thereby regulating endocytosis of NGF and NGF-induced downstream signaling pathways for neuronal survival and differentiation.

  16. Receptor mediated uptake of paclitaxel from a synthetic high density lipoprotein nanocarrier.

    Science.gov (United States)

    Mooberry, Linda K; Nair, Maya; Paranjape, Sulabha; McConathy, Walter J; Lacko, Andras G

    2010-01-01

    The purpose of these studies was to determine the mechanism(s) whereby paclitaxel (PTX), is taken up by cancer cells, once encapsulated into synthetic/reconstituted high density lipoprotein (rHDL). The uptake of PTX was found to be facilitated by the scavenger receptor type B-1 (SR-B1) when drug-loaded rHDL particles were incubated with cells that express the SRB1 receptor. Studies with double-labeled, PTX containing rHDL nanoparticles showed that prostate cancer (PC-3) cells incorporated PTX primarily via a selective (SR-B1 type) uptake mechanism. In the presence of a 10-fold excess of plasma HDL, PTX uptake decreased to 30% of the control. These findings suggest that the incorporation of lipophilic drugs by cancer cells from rHDL nanoparticles is facilitated by a receptor mediated (SR-B1) mechanism.

  17. Upregulation of endothelin ETB receptor-mediated vasoconstriction in rat coronary artery after organ culture

    DEFF Research Database (Denmark)

    Eskesen, Karen; Edvinsson, Lars

    2006-01-01

    descending coronary arteries isolated from hearts of rats as response to application of the selective endothelin ET(B) receptor agonist, Sarafotoxin 6c and endothelin-1. In segments cultured 1 day in serum free Dulbecco's Modified Eagle's Medium, Sarafotoxin 6c induced a concentration dependent contraction......The aim of this study was to examine if endothelin ET(B) receptor-mediated contraction occurred in isolated segments of rat coronary arteries during organ culture. Presence of contractile endothelin ET(B) receptors was studied by measuring the change in isometric tension in rings of left anterior...... after culturing could be eliminated with addition of the transcriptional blocker, actinomycin D, to the culture medium or be significantly attenuated by application of the translational inhibitor, cycloheximide. The vasoconstrictor effect of endothelin-1 or to depolarisation by high K...

  18. Transferrin protein nanospheres: a nanoplatform for receptor-mediated cancer cell labeling and gene delivery

    Science.gov (United States)

    McDonald, Michael A.; Spurlin, Tighe A.; Tona, Alessandro; Elliott, John T.; Halter, Michael; Plant, Anne L.

    2010-02-01

    This paper presents preliminary results on the use of transferrin protein nanospheres (TfpNS) for targeting cancer cells in vitro. Protein nanospheres represent an easily prepared and modifiable nanoplatform for receptor-specific targeting, molecular imaging and gene delivery. Rhodamine B isothiocyanate conjugated TfpNS (RBITC-TfpNS) show significantly enhanced uptake in vitro in SK-MEL-28 human malignant melanoma cells known to overexpress transferrin receptors compared to controls. RBITCTfpNS labeling of the cancer cells is due to transferrin receptor-mediated uptake, as demonstrated by competitive inhibition with native transferrin. Initial fluorescence microscopy studies indicate GFP plasmid can be transfected into melanoma cells via GFP plasmid encapsulated by TfpNS.

  19. Inflammatory Mediators and Insulin Resistance in Obesity: Role of Nuclear Receptor Signaling in Macrophages

    Directory of Open Access Journals (Sweden)

    Lucía Fuentes

    2010-01-01

    Full Text Available Visceral obesity is coupled to a general low-grade chronic inflammatory state characterized by macrophage activation and inflammatory cytokine production, leading to insulin resistance (IR. The balance between proinflammatory M1 and antiinflammatory M2 macrophage phenotypes within visceral adipose tissue appears to be crucially involved in the development of obesity-associated IR and consequent metabolic abnormalities. The ligand-dependent transcription factors peroxisome proliferator activated receptors (PPARs have recently been implicated in the determination of the M1/M2 phenotype. Liver X receptors (LXRs, which form another subgroup of the nuclear receptor superfamily, are also important regulators of proinflammatory cytokine production in macrophages. Disregulation of macrophage-mediated inflammation by PPARs and LXRs therefore underlies the development of IR. This review summarizes the role of PPAR and LXR signaling in macrophages and current knowledge about the impact of these actions in the manifestation of IR and obesity comorbidities such as liver steatosis and diabetic osteopenia.

  20. 25(OH) vitamin D suppresses macrophage adhesion and migration by downregulation of ER stress and scavenger receptor A1 in type 2 diabetes.

    Science.gov (United States)

    Riek, Amy E; Oh, Jisu; Darwech, Isra; Moynihan, Clare E; Bruchas, Robin R; Bernal-Mizrachi, Carlos

    2014-10-01

    Cardiovascular disease (CVD) is the leading cause of mortality in patients with type 2 diabetes mellitus (T2DM). Vitamin D deficiency is not only more prevalent in diabetics but also doubles the risk of developing CVD. However, it is unknown whether 25-hydroxy vitamin D [25(OH)D3] replacement slows monocyte adhesion and migration, critical mechanisms involved in atherosclerosis progression. In this study, monocytes from vitamin D-deficient diabetic patients were cultured either in the patient's serum or in vitamin D-deficient media with or without 25(OH)D3 treatment. Adding 25(OH)D3 to monocytes cultured in vitamin D-deficient serum or media decreased monocyte adhesion to fibronectin and migration stimulated by monocyte chemotactic protein 1 (MCP-1). Accordingly, 25(OH)D3 decreased adhesion marker β1- and β2-integrin expression and migration receptor chemokine (C-C motif) receptor 2 (CCR2) expression. 25(OH)D3 treatment downregulated monocyte endoplasmic reticulum (ER) stress and scavenger receptor class A, type 1 (SR-A1) expression. The absence of SR-A1 prevented the increased macrophage adhesion and migration induced by vitamin D deficiency. Moreover, the absence of SR-A1 prevented the induction of adhesion and migration and expression of their associated membrane receptors by Thapsigargin, an ER stress inducer. These results identify cellular activation of monocyte/macrophage vitamin D signaling through 25(OH)D3 as a potential mechanism that could modulate adhesion and migration in diabetic subjects. This article is part of a Special Issue entitled '16th Vitamin D Workshop'. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. Structural Basis for Receptor-Mediated Selective Autophagy of Aminopeptidase I Aggregates

    Directory of Open Access Journals (Sweden)

    Akinori Yamasaki

    2016-06-01

    Full Text Available Selective autophagy mediates the degradation of various cargoes, including protein aggregates and organelles, thereby contributing to cellular homeostasis. Cargo receptors ensure selectivity by tethering specific cargo to lipidated Atg8 at the isolation membrane. However, little is known about the structural requirements underlying receptor-mediated cargo recognition. Here, we report structural, biochemical, and cell biological analysis of the major selective cargo protein in budding yeast, aminopeptidase I (Ape1, and its complex with the receptor Atg19. The Ape1 propeptide has a trimeric coiled-coil structure, which tethers dodecameric Ape1 bodies together to form large aggregates. Atg19 disassembles the propeptide trimer and forms a 2:1 heterotrimer, which not only blankets the Ape1 aggregates but also regulates their size. These receptor activities may promote elongation of the isolation membrane along the aggregate surface, enabling sequestration of the cargo with high specificity.

  2. Expression of insulin receptor (IR) A and B isoforms, IGF-IR, and IR/IGF-IR hybrid receptors in vascular smooth muscle cells and their role in cell migration in atherosclerosis.

    Science.gov (United States)

    Beneit, N; Fernández-García, C E; Martín-Ventura, J L; Perdomo, L; Escribano, Ó; Michel, J B; García-Gómez, G; Fernández, S; Díaz-Castroverde, S; Egido, J; Gómez-Hernández, A; Benito, M

    2016-12-01

    Abnormal proliferation and migration of vascular smooth muscle cells (VSMCs) is a major contributor to the development of atherosclerotic process. In a previous work, we demonstrated that the insulin receptor isoform A (IRA) and its association with the insulin-like growth factor-I receptor (IGF-IR) confer a proliferative advantage to VSMCs. However, the role of IR and IGF-IR in VSMC migration remains poorly understood. Wound healing assays were performed in VSMCs bearing IR (IRLoxP+/+ VSMCs), or not (IR-/- VSMCs), expressing IRA (IRA VSMCs) or expressing IRB (IRB VSMCs). To study the role of IR isoforms and IGF-IR in experimental atherosclerosis, we used ApoE-/- mice at 8, 12, 18 and 24 weeks of age. Finally, we analyzed the mRNA expression of total IR, IRB isoform, IGF-IR and IGFs by qRT-PCR in the medial layer of human aortas. IGF-I strongly induced migration of the four cell lines through IGF-IR. In contrast, insulin and IGF-II only caused a significant increase of IRA VSMC migration which might be favored by the formation of IRA/IGF-IR receptors. Additionally, a specific IGF-IR inhibitor, picropodophyllin, completely abolished insulin- and IGF-II-induced migration in IRB, but not in IRA VSMCs. A significant increase of IRA and IGF-IR, and VSMC migration were observed in fibrous plaques from 24-week-old ApoE-/- mice. Finally, we observed a marked increase of IGF-IR, IGF-I and IGF-II in media from fatty streaks as compared with both healthy aortas and fibrolipidic lesions, favoring the ability of medial VSMCs to migrate into the intima. Our data suggest that overexpression of IGF-IR or IRA isoform, as homodimers or as part of IRA/IGF-IR hybrid receptors, confers a stronger migratory capability to VSMCs as might occur in early stages of atherosclerotic process.

  3. The G-Protein-Coupled Chemoattractant Receptor Fpr2 Exacerbates High Glucose-Mediated Proinflammatory Responses of Müller Glial Cells

    Directory of Open Access Journals (Sweden)

    Ying Yu

    2017-12-01

    Full Text Available In proliferative diabetic retinopathy (PDR, activated Müller glial cells (MGCs exhibit increased motility and a fibroblast-like proliferation phenotype that contribute to the formation of fibrovascular membrane. In this study, we investigated the capacity of high glucose (HG to regulate the expression of cell surface receptors that may participate in the proinflammatory responses of MGCs. We found that MGCs express a G-protein coupled chemoattractant receptor formyl peptide receptor 2 (Fpr2 and fibroblast growth factor receptor 1 (FGFR1, which mediated MGC migration and proliferation in response to corresponding ligands. HG upregulated Fpr2 through an NF-κB pathway in MGCs, increased the activation of MAPKs coupled to Fpr2 and FGFR1, which also further enhanced the production of vascular endothelial growth factor by MGCs in the presence of HG. In vivo, Fpr2 was more highly expressed by retina MGCs of diabetic mice and the human counterpart FPR2 was detected in the retina MGCs in fibrovascular membrane of PDR patients. To support the potential pathological relevance of Fpr2, an endogenous Fpr2 agonist cathelin-related antimicrobial peptide was detected in mouse MGCs and the retina, which was upregulated by HG. These results suggest that Fpr2, together with FGFR1, may actively participate in the pathogenesis of PDR thus may be considered as one of the potential therapeutic targets.

  4. Steroid Hormone Receptors as Potential Mediators of the Clinical Effects of Dutasteride: A Prospective, Randomized, Double-Blind Study

    National Research Council Canada - National Science Library

    Alonso, João C. C; Reis, Leonardo O; Garcia, Patrick V; Ferreira, Ubirajara; Matheus, Wagner E; Simões, Fabiano A; Rejowski, Ronald F; Alonso-Vale, Maria Isabel C; Fávaro, Wagner J

    2017-01-01

    This study characterizes the clinical and morphofunctional effects of a 5α-reductase inhibitor on steroid hormone receptors in normal human prostate tissue, as potential mediators of the clinical effects of dutasteride...

  5. Laser induced breakdown detection for the assessment of colloid mediated radionuclide migration

    CERN Document Server

    Walther, C; Hauser, W; Kim, J I; Scherbaum, F J

    2002-01-01

    Colloids play an important role in the transport of pollutants in the environment. Harmful substances can undergo transport over large distances if bound to colloids in aqueous surrounding. One important example is the migration of Pu(IV) at unexpectedly high rates over several miles at a Nevada nuclear detonation test site. For long term safety assessments of radioactive waste repositories, it is hence crucial to know about the amount, size distribution and chemical composition of colloids in the ground water. Standard methods (e.g. light scattering) can be applied for high concentrations and large sizes of particles. Colloids smaller than 50 nm, however, are detected with very low efficiency. Laser induced breakdown detection (LIBD) can fill this gap. A new instrumentation is presented, which as compared to previous instruments, opens up a much wider operational dynamic range, now covering three orders of magnitude in size (5-1000 nm) and seven orders of magnitude in particle concentration (1 ppt - several ...

  6. Death Receptor-Mediated Cell Death and Proinflammatory Signaling in Nonalcoholic SteatohepatitisSummary

    Directory of Open Access Journals (Sweden)

    Petra Hirsova

    2015-01-01

    Full Text Available Nonalcoholic fatty liver disease (NAFLD is becoming a public health problem worldwide. A subset of patients develop an inflammatory disease, nonalcoholic steatohepatitis (NASH, characterized by steatosis, hepatocellular death, macrophage and neutrophil accumulation, and varying stages of fibrosis. Hepatocyte cell death triggers the cellular inflammatory response, therefore reducing cell death may be salutary in the steatohepatitis disease process. Recently, a better understanding of hepatocyte apoptosis in NASH has been obtained and new information regarding other cell death modes such as necroptosis and pyroptosis has been reported. Hepatocyte lipotoxicity is often triggered by death receptors. In addition to causing apoptosis, death receptors have been shown to mediate proinflammatory signaling, suggesting that apoptosis in this context is not an immunologically silent process. Here, we review recent developments in our understanding of hepatocyte cell death by death receptors and its mechanistic link to inflammation in NASH. We emphasize how proapoptotic signaling by death receptors may induce the release of proinflammatory extracellular vesicles, thereby recruiting and activating macrophages and promoting the steatohepatitis process. Potential therapeutic strategies are discussed based on this evolving information. Keywords: Apoptosis, Caspase Inhibitor, Cell Death, Death Receptors, Exosomes, Extracellular Vesicles, Fibrosis, Inflammation, Inflammasome, Microvesicles, Necroptosis, Pyroptosis

  7. Serotonin(2) receptors mediate respiratory recovery after cervical spinal cord hemisection in adult rats.

    Science.gov (United States)

    Zhou, S Y; Basura, G J; Goshgarian, H G

    2001-12-01

    The aim of the present study was to specifically investigate the involvement of serotonin [5-hydroxytryptamine (5-HT(2))] receptors in 5-HT-mediated respiratory recovery after cervical hemisection. Experiments were conducted on C(2) spinal cord-hemisected, anesthetized (chloral hydrate, 400 mg/kg ip), vagotomized, pancuronium- paralyzed, and artificially ventilated female Sprague-Dawley rats in which CO(2) levels were monitored and maintained. Twenty-four hours after spinal hemisection, the ipsilateral phrenic nerve displayed no respiratory-related activity indicative of a functionally complete hemisection. Intravenous administration of the 5-HT(2A/2C)-receptor agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI) induced respiratory-related activity in the phrenic nerve ipsilateral to hemisection under conditions in which CO(2) was maintained at constant levels and augmented the activity induced under conditions of hypercapnia. The effects of DOI were found to be dose dependent, and the recovery of activity could be maintained for up to 2 h after a single injection. DOI-induced recovery was attenuated by the 5-HT(2)-receptor antagonist ketanserin but not with the 5-HT(2C)-receptor antagonist RS-102221, suggesting that 5-HT(2A) and not necessarily 5-HT(2C) receptors may be involved in the induction of respiratory recovery after cervical spinal cord injury.

  8. Multivalent ligand-receptor-mediated interaction of small filled vesicles with a cellular membrane

    Science.gov (United States)

    Zhdanov, Vladimir P.

    2017-07-01

    The ligand-receptor-mediated contacts of small sub-100-nm-sized lipid vesicles (or nanoparticles) with the cellular membrane are of interest in the contexts of cell-to-cell communication, endocytosis of membrane-coated virions, and drug (RNA) delivery. In all these cases, the interior of vesicles is filled by biologically relevant content. Despite the diversity of such systems, the corresponding ligand-receptor interaction possesses universal features. One of them is that the vesicle-membrane contacts can be accompanied by the redistribution of ligands and receptors between the contact and contact-free regions. In particular, the concentrations of ligands and receptors may become appreciably higher in the contact regions and their composition may there be different compared to that in the suspended state in the solution. A statistical model presented herein describes the corresponding distribution of various ligands and receptors and allows one to calculate the related change of the free energy with variation of the vesicle-engulfment extent. The results obtained are used to clarify the necessary conditions for the vesicle-assisted pathway of drug delivery.

  9. Glucocorticoid receptor mediated suppression of natural killer cell activity: identification of associated deacetylase and corepressor molecules.

    Science.gov (United States)

    Bush, Kristin A; Krukowski, Karen; Eddy, Justin L; Janusek, Linda Witek; Mathews, Herbert L

    2012-01-01

    Physical and psychological stressors reduce natural killer cell function. This reduction in cellular function results from stress-induced release of glucocorticoids. Glucocorticoids act upon natural killer cells to deacetylate and transrepress immune response genes through epigenetic processes. However, other than the glucocorticoid receptor, the proteins that participate in this process are not well described in natural killer cells. The purpose of this study was to identify the proteins associated with the glucocorticoid receptor that are likely epigenetic participants in this process. Treatment of natural killer cells with the synthetic glucocorticoid, dexamethasone, produced a significant time dependent reduction in natural killer cell activity as early as 8h post treatment. This reduction in natural killer cell activity was preceded by nuclear localization of the glucocorticoid receptor with histone deacetylase 1 and the corepressor, SMRT. Other class I histone deacetylases were not associated with the glucocorticoid receptor nor was the corepressor NCoR. These results demonstrate histone deacetylase 1 and SMRT to associate with the ligand activated glucocorticoid receptor within the nuclei of natural killer cells and to be the likely participants in the histone deacetylation and transrepression that accompanies glucocorticoid mediated reductions in natural killer cell function. Copyright © 2012 Elsevier Inc. All rights reserved.

  10. Ganglioside mediate the interaction between Nogo receptor 1 and LINGO-1.

    Science.gov (United States)

    Saha, Nayanendu; Kolev, Momchil V; Semavina, Mariya; Himanen, Juha; Nikolov, Dimitar B

    2011-09-16

    Upon spinal cord injury, the myelin inhibitors, including the myelin-associated glycoprotein (MAG), Nogo-A and the oligodendrocyte myelin glycoprotein (OMgp), bind to and signal via a single neuronal receptor/co-receptor complex comprising of Nogo receptor 1(NgR1)/LINGO-1 and p75 or TROY, impeding regeneration of injured axons. We employed a cell-free system to study the binding of NgR1 to its co-receptors and the myelin inhibitor Nogo-A, and show that gangliosides mediate the interaction of NgR1 with LINGO-1. Solid phase binding assays demonstrate that the sialic acid moieties of gangliosides and the stalk of NgR1 are the principal determinants of these molecular interactions. Moreover, the tripartite complex comprising of NgR1, LINGO-1 and ganglioside exhibits stronger binding to Nogo-A (Nogo-54) in the presence of p75, suggesting the gangliosides modulate the myelin inhibitor-receptor signaling. Copyright © 2011 Elsevier Inc. All rights reserved.

  11. Emotion dysregulation mediates the relationship between trauma exposure, post-migration living difficulties and psychological outcomes in traumatized refugees.

    Science.gov (United States)

    Nickerson, Angela; Bryant, Richard A; Schnyder, Ulrich; Schick, Matthis; Mueller, Julia; Morina, Naser

    2015-03-01

    While emotion dysregulation represents an important mechanism underpinning psychological responses to trauma, little research has investigated this in refugees. In the current study, we examined the mediating role of emotion dysregulation in the relationship between refugee experiences (trauma and living difficulties) and psychological outcomes. Participants were 134 traumatized treatment-seeking refugees who completed measures indexing trauma exposure, post-migration living difficulties, difficulties in emotion regulation, posttraumatic stress disorder, depression, and explosive anger. Findings revealed distinctive patterns of emotion dysregulation associated with each of these psychological disorders. Results also indicated that emotion regulation difficulties mediated the association between both trauma and psychological symptoms, and living difficulties and psychological symptoms. Limitations include a cross-sectional design and the use of measures that had not been validated across all cultural groups in this study. These findings underscore the key role of emotion dysregulation in psychological responses of refugees, and highlight potential directions for treatment interventions for traumatized refugees. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. CD26-mediated regulation of periostin expression contributes to migration and invasion of malignant pleural mesothelioma cells

    Energy Technology Data Exchange (ETDEWEB)

    Komiya, Eriko [Department of Therapy Development and Innovation for Immune Disorders and Cancers, Graduate School of Medicine, Juntendo University, 2-1-1, Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Ohnuma, Kei, E-mail: kohnuma@juntendo.ac.jp [Department of Therapy Development and Innovation for Immune Disorders and Cancers, Graduate School of Medicine, Juntendo University, 2-1-1, Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Yamazaki, Hiroto; Hatano, Ryo; Iwata, Satoshi; Okamoto, Toshihiro [Department of Therapy Development and Innovation for Immune Disorders and Cancers, Graduate School of Medicine, Juntendo University, 2-1-1, Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Dang, Nam H. [Division of Hematology/Oncology, University of Florida, 1600 SW Archer Road, Box 100278, Room MSB M410A, Gainesville, FL 32610 (United States); Yamada, Taketo [Department of Pathology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582 (Japan); Morimoto, Chikao [Department of Therapy Development and Innovation for Immune Disorders and Cancers, Graduate School of Medicine, Juntendo University, 2-1-1, Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan)

    2014-05-16

    Highlights: • CD26-expressing MPM cells upregulate production of periostin. • The intracytoplasmic region of CD26 mediates the upregulation of periostin. • CD26 expression leads to nuclear translocation of Twist1 via phosphorylation of Src. • Secreted periostin enhances migration and invasion of MPM cells. - Abstract: Malignant pleural mesothelioma (MPM) is an aggressive malignancy arising from mesothelial lining of pleura. It is generally associated with a history of asbestos exposure and has a very poor prognosis, partly due to the lack of a precise understanding of the molecular mechanisms associated with its malignant behavior. In the present study, we expanded on our previous studies on the enhanced motility and increased CD26 expression in MPM cells, with a particular focus on integrin adhesion molecules. We found that expression of CD26 upregulates periostin secretion by MPM cells, leading to enhanced MPM cell migratory and invasive activity. Moreover, we showed that upregulation of periostin expression results from the nuclear translocation of the basic helix-loop-helix transcription factor Twist1, a process that is mediated by CD26-associated activation of Src phosphorylation. While providing new and profound insights into the molecular mechanisms involved in MPM biology, these findings may also lead to the development of novel therapeutic strategies for MPM.

  13. Perceived discrimination and religiosity as potential mediating factors between migration and depressive symptoms: a transnational study of an indigenous Mayan population.

    Science.gov (United States)

    Salgado, Hugo; Haviland, Isa; Hernandez, Marcella; Lozano, Diana; Osoria, Ruby; Keyes, David; Kang, Eastern; Zúñiga, María Luisa

    2014-06-01

    Evidence suggests that in the US perceived discrimination among migrants of Mexican origin is associated with depressive symptoms. Factors that confer resilience, such as religiosity, could serve as a mediating factor in the context of migration stressors. We hypothesized that migration is associated with higher depressive symptoms and that discrimination and religiosity would mediate this relationship in a binational (US and Mexican) sample of indigenous Mexican migrants. We applied path analysis modeling to test our hypotheses with a sample of 650 individuals (n = 583 in Mexico; n = 67 in US). Results indicated that migration experience and current US residence were associated with perceived discrimination, which in turn were associated with a higher risk for depressive symptoms. Among women not living in the US, religiosity was associated with lower perceived discrimination. Discrimination is pervasive among male and female transnational and domestic migrants and religiosity may serve as a protective factor against discrimination for some women.

  14. IL-4 type 1 receptor signaling up-regulates KCNN4 expression, and increases the KCa3.1 current and its contribution to migration of alternative-activated microglia

    Directory of Open Access Journals (Sweden)

    Roger eFerreira

    2014-07-01

    Full Text Available The Ca2+-activated K+ channel, KCa3.1 (KCNN4/IK1/SK4, contributes to ‘classical’, pro-inflammatory activation of microglia, and KCa3.1 blockers have improved the outcome in several rodent models of CNS damage. For instance, blocking KCa3.1 with TRAM-34 rescued retinal ganglion neurons after optic nerve damage in vivo and, reduced p38 MAP kinase activation, production of reactive oxygen and nitrogen species, and neurotoxicity by microglia in vitro. In pursuing the therapeutic potential of KCa3.1 blockers, it is crucial to assess KCa3.1 contributions to other microglial functions and activation states, especially the IL-4-induced ‘alternative’ activation state that can counteract pro-inflammatory states. We recently found that IL-4 increases microglia migration—a crucial function in the healthy and damaged CNS—and that KCa3.1 contributes to P2Y2 receptor-stimulated migration. Here, we discovered that KCa3.1 is greatly increased in alternative-activated rat microglia and then contributes to an enhanced migratory capacity. IL-4 up-regulated KCNN4 mRNA (by 6 hr and greatly increased the KCa3.1 current by 1 day, and this required de novo protein synthesis. The increase in current was sustained for at least 6 days. IL-4 increased microglial migration and this was reversed by blocking KCa3.1 with TRAM-34. A panel of inhibitors of signal-transduction mediators was used to analyze contributions of IL-4-related signaling pathways. Induction of KCNN4 mRNA and KCa3.1 current was mediated specifically through IL-4 binding to the type I receptor and, surprisingly, it required JAK3, Ras/MEK/ERK signaling and the transcription factor, AP-1, rather than JAK2, STAT6 or PI3K. The same receptor subtype and pathway were required for the enhanced KCa3.1-dependent migration. In providing the first direct signaling link between an IL-4 receptor, expression and roles of an ion channel, this study also highlights the potential importance of KCa3.1 in

  15. Receptor-mediated enhancement of beta adrenergic drug activity by ascorbate in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Patrick F Dillon

    Full Text Available RATIONALE: Previous in vitro research demonstrated that ascorbate enhances potency and duration of activity of agonists binding to alpha 1 adrenergic and histamine receptors. OBJECTIVES: Extending this work to beta 2 adrenergic systems in vitro and in vivo. METHODS: Ultraviolet spectroscopy was used to study ascorbate binding to adrenergic receptor preparations and peptides. Force transduction studies on acetylcholine-contracted trachealis preparations from pigs and guinea pigs measured the effect of ascorbate on relaxation due to submaximal doses of beta adrenergic agonists. The effect of inhaled albuterol with and without ascorbate was tested on horses with heaves and sheep with carbachol-induced bronchoconstriction. MEASUREMENTS: Binding constants for ascorbate binding to beta adrenergic receptor were derived from concentration-dependent spectral shifts. Dose- dependence curves were obtained for the relaxation of pre-contracted trachealis preparations due to beta agonists in the presence and absence of varied ascorbate. Tachyphylaxis and fade were also measured. Dose response curves were determined for the effect of albuterol plus-and-minus ascorbate on airway resistance in horses and sheep. MAIN RESULTS: Ascorbate binds to the beta 2 adrenergic receptor at physiological concentrations. The receptor recycles dehydroascorbate. Physiological and supra-physiological concentrations of ascorbate enhance submaximal epinephrine and isoproterenol relaxation of trachealis, producing a 3-10-fold increase in sensitivity, preventing tachyphylaxis, and reversing fade. In vivo, ascorbate improves albuterol's effect on heaves and produces a 10-fold enhancement of albuterol activity in "asthmatic" sheep. CONCLUSIONS: Ascorbate enhances beta-adrenergic activity via a novel receptor-mediated mechanism; increases potency and duration of beta adrenergic agonists effective in asthma and COPD; prevents tachyphylaxis; and reverses fade. These novel effects are

  16. Use of plasma C-reactive protein, procalcitonin, neutrophils,macrophage migration inhibitory factor, soluble urokinase-type plasminogen activator receptor, and soluble triggering receptor expressed on myeloid cells-1 in combination to diagnose infections: a prospective study

    DEFF Research Database (Denmark)

    Kofoed, Kristian; Andersen, Ove; Kronborg, Gitte

    2007-01-01

    department and department of infectious diseases at a university hospital. A multiplex immunoassay measuring soluble urokinase-type plasminogen activator (suPAR) and soluble triggering receptor expressed on myeloid cells (sTREM)-1 and macrophage migration inhibitory factor (MIF) was used in parallel...

  17. Prosurvival NMDA 2A receptor signaling mediates postconditioning neuroprotection in the hippocampus.

    Science.gov (United States)

    Zhang, Xi; Zhang, Quanguang; Tu, Jingyi; Zhu, Ying; Yang, Fang; Liu, Bin; Brann, Darrell; Wang, Ruimin

    2015-03-01

    Ischemic postconditioning (Post C), which involves administration of a brief ischemia after the initial ischemic event, has been demonstrated to be strongly neuroprotective against global cerebral ischemia (GCI) and to improve cognitive outcome. To enhance understanding of the underlying mechanisms, the current study examined the role of NMDA receptors in mediating the beneficial effects of Post C (3 min ischemia) administered 2 days after GCI in adult male rats. The results revealed that Post C was strongly neuroprotective against GCI, and that this effect was blocked by administration of the NMDA receptor antagonist MK-801. Further work revealed that the NR2A-type NMDA receptors mediate the Post C beneficial effects as administration of a NR2A-preferring antagonist (NVP-A) blocked Post C neuroprotection and cognitive enhancement, while administration of a NR2B-preferring antagonist (Ro25) was without effect. Post C significantly up-regulated NR2A levels and phosphorylation of NR2A in the hippocampal CA1 region after Post C. Post C also increased Ca(2+) influx and activation/phosphorylation of CamKIIα at Thr(286), effects that were NR2A mediated as they were blocked by NVP-A. Phosphorylation of ERK and CREB was also increased by Post C, as were two downstream CREB-dependent prosurvival factors, brain derived neurotropic factor (BDNF) and Bcl2, effects that were blocked by the NR2A antagonist, NVP-A. Taken as a whole, the current study provides evidence that NR2A-activation and downstream prosurvival signaling is a critical mediator of Post C-induced neuroprotection and cognitive enhancement following GCI. © 2014 Wiley Periodicals, Inc.

  18. CHOLINERGIC CONTRACTION OF THE GUINEA-PIG LUNG STRIP IS MEDIATED BY 9USCARINIC M(2)-LIKE RECEPTORS

    NARCIS (Netherlands)

    ROFFEL, AF; ELZINGA, CRS; ZAAGSMA, J

    1993-01-01

    The muscarinic receptor subtype mediating contraction of the guinea pig lung strip preparation was investigated and compared with that in guinea pig tracheal and human peripheral airway (small bronchi) smooth muscle preparations, using a number of subtype selective muscarinic receptor antagonists.

  19. Human umbilical cord perivascular cells exhibited enhanced migration capacity towards hepatocellular carcinoma in comparison with bone marrow mesenchymal stromal cells: a role for autocrine motility factor receptor.

    Science.gov (United States)

    Bayo, Juan; Fiore, Esteban; Aquino, Jorge B; Malvicini, Mariana; Rizzo, Manglio; Peixoto, Estanislao; Alaniz, Laura; Piccioni, Flavia; Bolontrade, Marcela; Podhajcer, Osvaldo; Garcia, Mariana G; Mazzolini, Guillermo

    2014-01-01

    Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. Unfortunately, the incidence and mortality associated with HCC are increasing. Therefore, new therapeutic strategies are urgently needed and the use of mesenchymal stromal cells (MSCs) as carrier of therapeutic genes is emerging as a promising option. Different sources of MSCs are being studied for cell therapy and bone marrow-derived cells are the most extensively explored; however, birth associated-tissues represent a very promising source. The aim of this work was to compare the in vitro and in vivo migration capacity between bone marrow MSCs (BM-MSCs) and human umbilical cord perivascular cells (HUCPVCs) towards HCC. We observed that HUCPVCs presented higher in vitro and in vivo migration towards factors released by HCC. The expression of autocrine motility factor (AMF) receptor, genes related with the availability of the receptor on the cell surface (caveolin-1 and -2) and metalloproteinase 3, induced by the receptor activation and important for cell migration, was increased in HUCPVCs. The chemotactic response towards recombinant AMF was increased in HUCPVCs compared to BM-MSCs, and its inhibition in the conditioned medium from HCC induced higher decrease in HUCPVC migration than in BM-MSC. Our results indicate that HUCPVCs could be a useful cellular source to deliver therapeutic genes to HCC.

  20. Human Umbilical Cord Perivascular Cells Exhibited Enhanced Migration Capacity towards Hepatocellular Carcinoma in Comparison with Bone Marrow Mesenchymal Stromal Cells: A Role for Autocrine Motility Factor Receptor

    Directory of Open Access Journals (Sweden)

    Juan Bayo

    2014-01-01

    Full Text Available Hepatocellular carcinoma (HCC is the third cause of cancer-related death worldwide. Unfortunately, the incidence and mortality associated with HCC are increasing. Therefore, new therapeutic strategies are urgently needed and the use of mesenchymal stromal cells (MSCs as carrier of therapeutic genes is emerging as a promising option. Different sources of MSCs are being studied for cell therapy and bone marrow-derived cells are the most extensively explored; however, birth associated-tissues represent a very promising source. The aim of this work was to compare the in vitro and in vivo migration capacity between bone marrow MSCs (BM-MSCs and human umbilical cord perivascular cells (HUCPVCs towards HCC. We observed that HUCPVCs presented higher in vitro and in vivo migration towards factors released by HCC. The expression of autocrine motility factor (AMF receptor, genes related with the availability of the receptor on the cell surface (caveolin-1 and -2 and metalloproteinase 3, induced by the receptor activation and important for cell migration, was increased in HUCPVCs. The chemotactic response towards recombinant AMF was increased in HUCPVCs compared to BM-MSCs, and its inhibition in the conditioned medium from HCC induced higher decrease in HUCPVC migration than in BM-MSC. Our results indicate that HUCPVCs could be a useful cellular source to deliver therapeutic genes to HCC.

  1. Anorectic actions of prolactin-releasing peptide are mediated by corticotropin-releasing hormone receptors.

    Science.gov (United States)

    Lawrence, Catherine B; Liu, Yong-Ling; Stock, Michael J; Luckman, Simon M

    2004-01-01

    Prolactin-releasing peptide (PrRP) reduces food intake and body weight and modifies body temperature when administered centrally in rats, suggesting a role in energy homeostasis. However, the mediators of PrRP's actions are unknown. The present study, therefore, first examined the possible involvement of the anorectic neuropeptides corticotropin-releasing hormone (CRH) and the melanocortins (e.g., alpha-melanocyte-stimulating hormone) in PrRP's effects on food intake and core body temperature and, second, determined if PrRP affects energy expenditure by measuring oxygen consumption (Vo2). Intracerebroventricular injection of PrRP (4 nmol) to 24-h-fasted male Sprague-Dawley rats decreased food intake and modified body temperature. Blockade of central CRH receptors by intracerebroventricular coadministration of the CRH receptor antagonist astressin (20 microg) reversed the PrRP-induced reduction in feeding. However, astressin's effect on PrRP-induced changes in body temperature was complicated because the antagonist itself caused a slight rise in body temperature. In contrast, intracerebroventricular coadministration of the melanocortin receptor-3/4 antagonist SHU-9119 (0.1 nmol) had no effect on any of PrRP's actions. Finally, intracerebroventricular injection of PrRP (4 nmol) caused a significantly greater Vo2 over a 3-h test period compared with vehicle-treated rats. These results show that the anorectic actions of PrRP are mediated by central CRH receptors but not by melanocortin receptors-3/4 and that PrRP can modify Vo2.

  2. Differential fibroblast growth factor 8 (FGF8-mediated autoregulation of its cognate receptors, Fgfr1 and Fgfr3, in neuronal cell lines.

    Directory of Open Access Journals (Sweden)

    Natasha N Mott

    2010-04-01

    Full Text Available Fibroblast growth factors (FGFs mediate a vast range of CNS developmental processes including neural induction, proliferation, migration, and cell survival. Despite the critical role of FGF signaling for normal CNS development, few reports describe the mechanisms that regulate FGF receptor gene expression in the brain. We tested whether FGF8 could autoregulate two of its cognate receptors, Fgfr1 and Fgfr3, in three murine cell lines with different lineages: fibroblast-derived cells (3T3 cells, neuronal cells derived from hippocampus (HT-22 cells, and neuroendocrine cells derived from hypothalamic gonadotropin-releasing hormone (GnRH neurons (GT1-7 cells. GnRH is produced by neurons in the hypothalamus and is absolutely required for reproductive competence in vertebrate animals. Several lines of evidence strongly suggest that Fgf8 is critical for normal development of the GnRH system, therefore, the GT1-7 cells provided us with an additional endpoint, Gnrh gene expression and promoter activity, to assess potential downstream consequences of FGF8-induced modulation of FGF receptor levels. Results from this study suggest that the autoregulation of its cognate receptor represents a common downstream effect of FGF8. Further, we show that Fgfr1 and Fgfr3 are differentially regulated within the same cell type, implicating these two receptors in different biological roles. Moreover, Fgfr1 and Fgfr3 are differentially regulated among different cell types, suggesting such autoregulation occurs in a cell type-specific fashion. Lastly, we demonstrate that FGF8b decreases Gnrh promoter activity and gene expression, possibly reflecting a downstream consequence of altered FGF receptor populations. Together, our data bring forth the possibility that, in addition to the FGF synexpression group, autoregulation of FGFR expression by FGF8 represents a mechanism by which FGF8 could fine-tune its regulatory actions.

  3. Ligation of human Fc receptor like-2 by monoclonal antibodies down-regulates B-cell receptor-mediated signalling

    Science.gov (United States)

    Shabani, Mahdi; Bayat, Ali Ahmad; Jeddi-Tehrani, Mahmood; Rabbani, Hodjatallah; Hojjat-Farsangi, Mohammad; Ulivieri, Cristina; Amirghofran, Zahra; Baldari, Cosima Tatiana; Shokri, Fazel

    2014-01-01

    B-cell antigen receptor (BCR) signalling and its regulation through negative and positive regulators are critical for balancing B-cell response and function. Human Fc receptor like-2 (FCRL2), a member of the newly identified FCRL family, could influence B-cell signalling due to possession of both immunoreceptor tyrosine-based activation and inhibitory motifs (ITAM and ITIM). Since the natural ligand of FCRL2 has not been identified, we generated FCRL2-specific monoclonal antibodies (mAbs) and employed them to investigate the influence of FCRL2 stimulation on BCR signalling in an FCRL2-expressing B-cell line. Two anti-FCRL2 mAb-producing hybridoma clones (5A7-E7 and 3D8-G8) were selected. None of the mAbs displayed any cross-reactivity with the other members of the FCRL family including recombinant FCRL1, -3, -4 and -5, as tested by FACS and ELISA techniques. Engagement of the FCRL2 by these mAbs resulted in significant inhibition of BCR signalling mediators such as calcium mobilization and phosphorylation of the mitogen-activated protein kinases Erk, p38 and Jnk. These findings indicate that the FCRL2 ITIM motifs are functional and the anti-FCRL2 mAbs may mimic the natural ligand of FCRL2 by induction of inhibitory signals in B cells. PMID:24797767

  4. Comparative study on transcriptional activity of 17 parabens mediated by estrogen receptor α and β and androgen receptor.

    Science.gov (United States)

    Watanabe, Yoko; Kojima, Hiroyuki; Takeuchi, Shinji; Uramaru, Naoto; Ohta, Shigeru; Kitamura, Shigeyuki

    2013-07-01

    The structure-activity relationships of parabens which are widely used as preservatives for transcriptional activities mediated by human estrogen receptor α (hERα), hERβ and androgen receptor (hAR) were investigated. Fourteen of 17 parabens exhibited hERα and/or hERβ agonistic activity at concentrations of ≤ 1 × 10(-5)M, whereas none of the 17 parabens showed AR agonistic or antagonistic activity. Among 12 parabens with linear alkyl chains ranging in length from C₁ to C₁₂, heptylparaben (C₇) and pentylparaben (C₅) showed the most potent ERα and ERβ agonistic activity in the order of 10(-7)M and 10(-8)M, respectively, and the activities decreased in a stepwise manner as the alkyl chain was shortened to C₁ or lengthened to C₁₂. Most parabens showing estrogenic activity exhibited ERβ-agonistic activity at lower concentrations than those inducing ERα-agonistic activity. The estrogenic activity of butylparaben was markedly decreased by incubation with rat liver microsomes, and the decrease of activity was blocked by a carboxylesterase inhibitor. These results indicate that parabens are selective agonists for ERβ over ERα; their interactions with ERα/β are dependent on the size and bulkiness of the alkyl groups; and they are metabolized by carboxylesterases, leading to attenuation of their estrogenic activity. Copyright © 2013 Elsevier Ltd. All rights reserved.

  5. Zinc-induced Neurotoxicity Mediated by Transient Receptor Potential Melastatin 7 Channels*

    Science.gov (United States)

    Inoue, Koichi; Branigan, Deborah; Xiong, Zhi-Gang

    2010-01-01

    Transient receptor potential melastatin 7 (TRPM7) channels are novel Ca2+-permeable non-selective cation channels ubiquitously expressed. Activation of TRPM7 channels has been shown to be involved in cellular Mg2+ homeostasis, diseases caused by abnormal magnesium absorption, and in Ca2+-mediated neuronal injury under ischemic conditions. Here we show strong evidence suggesting that TRPM7 channels also play an important role in cellular Zn2+ homeostasis and in Zn2+-mediated neuronal injury. Using a combination of fluorescent Zn2+ imaging, small interfering RNA, pharmacological analysis, and cell injury assays, we show that activation of TRPM7 channels augmented Zn2+-induced injury of cultured mouse cortical neurons. The Zn2+-mediated neurotoxicity was inhibited by nonspecific TRPM7 blockers Gd3+ or 2-aminoethoxydiphenyl borate, and by knockdown of TRPM7 channels with small interfering RNA. In addition, Zn2+-mediated neuronal injury under oxygen-glucose deprivation conditions was also diminished by silencing TRPM7. Furthermore, we show that overexpression of TRPM7 channels in HEK293 cells increased intracellular Zn2+ accumulation and Zn2+-induced cell injury, while silencing TRPM7 by small interfering RNA attenuated the Zn2+-mediated cell toxicity. Thus, TRPM7 channels may represent a novel target for neurological disorders where Zn2+ toxicity plays an important role. PMID:20048154

  6. Zinc-induced neurotoxicity mediated by transient receptor potential melastatin 7 channels.

    Science.gov (United States)

    Inoue, Koichi; Branigan, Deborah; Xiong, Zhi-Gang

    2010-03-05

    Transient receptor potential melastatin 7 (TRPM7) channels are novel Ca(2+)-permeable non-selective cation channels ubiquitously expressed. Activation of TRPM7 channels has been shown to be involved in cellular Mg(2+) homeostasis, diseases caused by abnormal magnesium absorption, and in Ca(2+)-mediated neuronal injury under ischemic conditions. Here we show strong evidence suggesting that TRPM7 channels also play an important role in cellular Zn(2+) homeostasis and in Zn(2+)-mediated neuronal injury. Using a combination of fluorescent Zn(2+) imaging, small interfering RNA, pharmacological analysis, and cell injury assays, we show that activation of TRPM7 channels augmented Zn(2+)-induced injury of cultured mouse cortical neurons. The Zn(2+)-mediated neurotoxicity was inhibited by nonspecific TRPM7 blockers Gd(3+) or 2-aminoethoxydiphenyl borate, and by knockdown of TRPM7 channels with small interfering RNA. In addition, Zn(2+)-mediated neuronal injury under oxygen-glucose deprivation conditions was also diminished by silencing TRPM7. Furthermore, we show that overexpression of TRPM7 channels in HEK293 cells increased intracellular Zn(2+) accumulation and Zn(2+)-induced cell injury, while silencing TRPM7 by small interfering RNA attenuated the Zn(2+)-mediated cell toxicity. Thus, TRPM7 channels may represent a novel target for neurological disorders where Zn(2+) toxicity plays an important role.

  7. Glutamate Receptors within the Mesolimbic Dopamine System Mediate Alcohol Relapse Behavior.

    Science.gov (United States)

    Eisenhardt, Manuela; Leixner, Sarah; Luján, Rafael; Spanagel, Rainer; Bilbao, Ainhoa

    2015-11-25

    Glutamatergic input within the mesolimbic dopamine (DA) pathway plays a critical role in the development of addictive behavior. Although this is well established for some drugs of abuse, it is not known whether glutamate receptors within the mesolimbic system are involved in mediating the addictive properties of chronic alcohol use. Here we evaluated the contribution of mesolimbic NMDARs and AMPARs in mediating alcohol-seeking responses induced by environmental stimuli and relapse behavior using four inducible mutant mouse lines lacking the glutamate receptor genes Grin1 or Gria1 in either DA transporter (DAT) or D1R-expressing neurons. We first demonstrate the lack of GluN1 or GluA1 in either DAT- or D1R-expressing neurons in our mutant mouse lines by colocalization studies. We then show that GluN1 and GluA1 receptor subunits within these neuronal subpopulations mediate the alcohol deprivation effect, while having no impact on context- plus cue-induced reinstatement of alcohol-seeking behavior. We further validated these results pharmacologically by demonstrating similar reductions in the alcohol deprivation effect after infusion of the NMDAR antagonist memantine into the nucleus accumbens and ventral tegmental area of control mice, and a rescue of the mutant phenotype via pharmacological potentiation of AMPAR activity using aniracetam. In conclusion, dopamine neurons as well as D1R-expressing medium spiny neurons and their glutamatergic inputs via NMDARs and AMPARs act in concert to influence relapse responses. These results provide a neuroanatomical and molecular substrate for relapse behavior and emphasize the importance of glutamatergic drugs in modulating relapse behavior. Here we provide genetic and pharmacological evidence that glutamate receptors within the mesolimbic dopamine system play an essential role in alcohol relapse. Using various inducible and site-specific transgenic mouse models and pharmacological validation experiments, we show that critical

  8. Immunomodulatory activity of commonly used drugs on Fc-receptor-mediated human natural killer cell activation.

    Science.gov (United States)

    Theorell, Jakob; Gustavsson, Anna-Lena; Tesi, Bianca; Sigmundsson, Kristmundur; Ljunggren, Hans-Gustaf; Lundbäck, Thomas; Bryceson, Yenan T

    2014-06-01

    Natural killer (NK) cells mediate defense against neoplastic as well as infected cells. Yet, how their effector functions are affected by the large variety of pharmacological compounds commonly in use has not been investigated systematically. Here, we screened 1,200 in-use or previously approved drugs for their biological effect on freshly isolated human peripheral blood-derived NK cells. Mimicking antibody-dependent cellular cytotoxicity (ADCC), known to be important in antibody-based immunotherapies against, e.g., human malignancies, the cells were stimulated by Fc-receptor (CD16) engagement. Cellular responses were assessed by flow cytometry. Fifty-six compounds that significantly inhibited and twelve that enhanced one or more of the readouts of adhesion, exocytosis, and chemokine production were identified and confirmed as hits. Among the confirmed inhibitors, 80 % could be assigned to one of seven major pharmacological classes. These classes were β2-adrenergic agonists, prostaglandins, phosphodiesterase-4 inhibitors, Ca(2+)-channel blockers, histamine H1-receptor antagonists, serotonin/dopamine receptor antagonists, and topoisomerase inhibitors that displayed distinct inhibitory patterns on NK cell responses. Among observed enhancers, interestingly, two ergosterol synthesis inhibitors were identified that specifically promoted exocytosis. In summary, these results provide a comprehensive knowledge base of the effect known drugs have on NK cells. More specifically, they provide an overview of drugs that may modulate NK cell-mediated ADCC in the context of clinical immunotherapies.

  9. Purinergic receptor antagonists inhibit odorant-mediated CREB phosphorylation in sustentacular cells of mouse olfactory epithelium.

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2012-02-01

    BACKGROUND: Extracellular nucleotides have long been known to play neuromodulatory roles and to be involved in intercellular signalling. In the olfactory system, ATP is released by olfactory neurons, and exogenous ATP can evoke an increase in intracellular calcium concentration in sustentacular cells, the nonneuronal supporting cells of the olfactory epithelium. Here we investigate the hypothesis that olfactory neurons communicate with sustentacular cells via extracellular ATP and purinergic receptor activation. RESULTS: Here we show that exposure of mice to a mixture of odorants induced a significant increase in the levels of the transcription factor CREB phosphorylated at Ser-133 in the nuclei of both olfactory sensory neurons and sustentacular cells. This activation was dependent on adenylyl cyclase III-mediated olfactory signaling and on activation of P2Y purinergic receptors on sustentacular cells. Purinergic receptor antagonists inhibited odorant-dependent CREB phosphorylation specifically in the nuclei of the sustentacular cells. CONCLUSION: Our results point to a possible role for extracellular nucleotides in mediating intercellular communication between the neurons and sustentacular cells of the olfactory epithelium in response to odorant exposure. Maintenance of extracellular ionic gradients and metabolism of noxious chemicals by sustentacular cells may therefore be regulated in an odorant-dependent manner by olfactory sensory neurons.

  10. Regulation of ENaC-Mediated Sodium Reabsorption by Peroxisome Proliferator-Activated Receptors

    Directory of Open Access Journals (Sweden)

    Tengis S. Pavlov

    2010-01-01

    Full Text Available Peroxisome proliferator-activated receptors (PPARs are members of a steroid hormone receptor superfamily that responds to changes in lipid and glucose homeostasis. Peroxisomal proliferator-activated receptor subtype γ (PPARγ has received much attention as the target for antidiabetic drugs, as well as its role in responding to endogenous compounds such as prostaglandin J2. However, thiazolidinediones (TZDs, the synthetic agonists of the PPARγ are tightly associated with fluid retention and edema, as potentially serious side effects. The epithelial sodium channel (ENaC represents the rate limiting step for sodium absorption in the renal collecting duct. Consequently, ENaC is a central effector impacting systemic blood volume and pressure. The role of PPARγ agonists on ENaC activity remains controversial. While PPARγ agonists were shown to stimulate ENaC-mediated renal salt absorption, probably via Serum- and Glucocorticoid-Regulated Kinase 1 (SGK1, other studies reported that PPARγ agonist-induced fluid retention is independent of ENaC activity. The current paper provides new insights into the control and function of ENaC and ENaC-mediated sodium transport as well as several other epithelial channels/transporters by PPARs and particularly PPARγ. The potential contribution of arachidonic acid (AA metabolites in PPAR-dependent mechanisms is also discussed.

  11. Purinergic receptor antagonists inhibit odorant-mediated CREB phosphorylation in sustentacular cells of mouse olfactory epithelium

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2011-08-22

    Abstract Background Extracellular nucleotides have long been known to play neuromodulatory roles and to be involved in intercellular signalling. In the olfactory system, ATP is released by olfactory neurons, and exogenous ATP can evoke an increase in intracellular calcium concentration in sustentacular cells, the nonneuronal supporting cells of the olfactory epithelium. Here we investigate the hypothesis that olfactory neurons communicate with sustentacular cells via extracellular ATP and purinergic receptor activation. Results Here we show that exposure of mice to a mixture of odorants induced a significant increase in the levels of the transcription factor CREB phosphorylated at Ser-133 in the nuclei of both olfactory sensory neurons and sustentacular cells. This activation was dependent on adenylyl cyclase III-mediated olfactory signaling and on activation of P2Y purinergic receptors on sustentacular cells. Purinergic receptor antagonists inhibited odorant-dependent CREB phosphorylation specifically in the nuclei of the sustentacular cells. Conclusion Our results point to a possible role for extracellular nucleotides in mediating intercellular communication between the neurons and sustentacular cells of the olfactory epithelium in response to odorant exposure. Maintenance of extracellular ionic gradients and metabolism of noxious chemicals by sustentacular cells may therefore be regulated in an odorant-dependent manner by olfactory sensory neurons.

  12. SR-B1 Is a Silica Receptor that Mediates Canonical Inflammasome Activation

    Directory of Open Access Journals (Sweden)

    Misato Tsugita

    2017-01-01

    Full Text Available The inhalation of silica dust is associated with fibrosis and lung cancer, which are triggered by macrophage inflammatory responses; however, how macrophages recognize silica remains largely unknown. Here, we identify by functional expression cloning the class B scavenger receptor SR-B1 as a silica receptor. Through an extracellular α-helix, both mouse and human SR-B1 specifically recognized amorphous and crystalline silica, but not titanium dioxide nanoparticles, latex nanoparticles, or monosodium urate crystals, although all particles exhibited negative surface potentials. Genetic deletion of SR-B1 and masking of SR-B1 by monoclonal antibodies showed that SR-B1-mediated recognition of silica is associated with caspase-1-mediated inflammatory responses in mouse macrophages and human peripheral blood monocytes. Furthermore, SR-B1 was involved in silica-induced pulmonary inflammation in mice. These results indicate that SR-B1 is a silica receptor associated with canonical inflammasome activation.

  13. Mannose Receptor Mediates the Immune Response to Ganoderma atrum Polysaccharides in Macrophages.

    Science.gov (United States)

    Li, Wen-Juan; Tang, Xiao-Fang; Shuai, Xiao-Xue; Jiang, Cheng-Jia; Liu, Xiang; Wang, Le-Feng; Yao, Yu-Fei; Nie, Shao-Ping; Xie, Ming-Yong

    2017-01-18

    The ability of mannose receptor (MR) to recognize the carbohydrate structures is well-established. Here, we reported that MR was crucial for the immune response to a Ganoderma atrum polysaccharide (PSG-1), as evidenced by elevation of MR in association with increase of phagocytosis and concentrations of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in normal macrophages. Elevation of MR triggered by PSG-1 also led to control lipopolysaccharide (LPS)-triggered inflammatory response via the increase of interleukin-10 (IL-10) and inhibition of phagocytosis and IL-1β. Anti-MR antibody partly attenuated PSG-1-mediated anti-inflammatory responses, while it could not affect TNF-α secretion, suggesting that another receptor was involved in PSG-1-triggered immunomodulatory effects. MR and toll-like receptor (TLR)4 coordinated the influences on the TLR4-mediated signaling cascade by the nuclear factor-κB (NF-κB) pathway in LPS-stimulated macrophages subjected to PSG-1. Collectively, immune response to PSG-1 required recognition by MR in macrophages. The NF-κB pathway served as a central role for the coordination of MR and TLR4 to elicit immune response to PSG-1.

  14. SR-B1 Is a Silica Receptor that Mediates Canonical Inflammasome Activation.

    Science.gov (United States)

    Tsugita, Misato; Morimoto, Nobuyuki; Tashiro, Manabu; Kinoshita, Kengo; Nakayama, Masafumi

    2017-01-31

    The inhalation of silica dust is associated with fibrosis and lung cancer, which are triggered by macrophage inflammatory responses; however, how macrophages recognize silica remains largely unknown. Here, we identify by functional expression cloning the class B scavenger receptor SR-B1 as a silica receptor. Through an extracellular α-helix, both mouse and human SR-B1 specifically recognized amorphous and crystalline silica, but not titanium dioxide nanoparticles, latex nanoparticles, or monosodium urate crystals, although all particles exhibited negative surface potentials. Genetic deletion of SR-B1 and masking of SR-B1 by monoclonal antibodies showed that SR-B1-mediated recognition of silica is associated with caspase-1-mediated inflammatory responses in mouse macrophages and human peripheral blood monocytes. Furthermore, SR-B1 was involved in silica-induced pulmonary inflammation in mice. These results indicate that SR-B1 is a silica receptor associated with canonical inflammasome activation. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  15. Functional characterization of GABAA receptor-mediated modulation of cortical neuron network activity in microelectrode array recordings

    DEFF Research Database (Denmark)

    Bader, Benjamin M; Steder, Anne; Klein, Anders Bue

    2017-01-01

    The numerous γ-aminobutyric acid type A receptor (GABAAR) subtypes are differentially expressed and mediate distinct functions at neuronal level. In this study we have investigated GABAAR-mediated modulation of the spontaneous activity patterns of primary neuronal networks from murine frontal...

  16. PCSK9-mediated degradation of the LDL receptor generates a 17 kDa C-terminal LDL receptor fragment.

    Science.gov (United States)

    Tveten, Kristian; Strøm, Thea Bismo; Berge, Knut Erik; Leren, Trond P

    2013-06-01

    Proprotein convertase subtilisin/kexin type 9 (PCSK9) binds to the LDL receptor (LDLR) at the cell surface and reroutes the internalized LDLR to intracellular degradation. In this study, we have shown that PCSK9-mediated degradation of the full-length 160 kDa LDLR generates a 17 kDa C-terminal LDLR fragment. This fragment was not generated from mutant LDLRs resistant to PCSK9-mediated degradation or when degradation was prevented by chemicals such as ammonium chloride or the cysteine cathepsin inhibitor E64d. The observation that the 17 kDa fragment was only detected when the cells were cultured in the presence of the γ-secretase inhibitor DAPT indicates that this 17 kDa fragment undergoes γ-secretase cleavage within the transmembrane domain. The failure to detect the complementary 143 kDa ectodomain fragment is likely to be due to its rapid degradation in the endosomal lumen. The 17 kDa C-terminal LDLR fragment was also generated from a Class 5 mutant LDLR undergoing intracellular degradation. Thus, one may speculate that an LDLR with bound PCSK9 and a Class 5 LDLR with bound LDL are degraded by a similar mechanism that could involve ectodomain cleavage in the endosome.

  17. The Viral KSHV Chemokine vMIP-II Inhibits the Migration of Naive and Activated Human NK Cells by Antagonizing Two Distinct Chemokine Receptors

    Science.gov (United States)

    Yamin, Rachel; Kaynan, Noa S.; Glasner, Ariella; Vitenshtein, Alon; Tsukerman, Pinchas; Bauman, Yoav; Ophir, Yael; Elias, Shlomo; Bar-On, Yotam; Gur, Chamutal; Mandelboim, Ofer

    2013-01-01

    Natural killer (NK) cells are innate immune cells able to rapidly kill virus-infected and tumor cells. Two NK cell populations are found in the blood; the majority (90%) expresses the CD16 receptor and also express the CD56 protein in intermediate levels (CD56Dim CD16Pos) while the remaining 10% are CD16 negative and express CD56 in high levels (CD56Bright CD16Neg). NK cells also reside in some tissues and traffic to various infected organs through the usage of different chemokines and chemokine receptors. Kaposi's sarcoma-associated herpesvirus (KSHV) is a human virus that has developed numerous sophisticated and versatile strategies to escape the attack of immune cells such as NK cells. Here, we investigate whether the KSHV derived cytokine (vIL-6) and chemokines (vMIP-I, vMIP-II, vMIP-III) affect NK cell activity. Using transwell migration assays, KSHV infected cells, as well as fusion and recombinant proteins, we show that out of the four cytokine/chemokines encoded by KSHV, vMIP-II is the only one that binds to the majority of NK cells, affecting their migration. We demonstrate that vMIP-II binds to two different receptors, CX3CR1 and CCR5, expressed by naïve CD56Dim CD16Pos NK cells and activated NK cells, respectively. Furthermore, we show that the binding of vMIP-II to CX3CR1 and CCR5 blocks the binding of the natural ligands of these receptors, Fractalkine (Fck) and RANTES, respectively. Finally, we show that vMIP-II inhibits the migration of naïve and activated NK cells towards Fck and RANTES. Thus, we present here a novel mechanism in which KSHV uses a unique protein that antagonizes the activity of two distinct chemokine receptors to inhibit the migration of naïve and activated NK cells. PMID:23966863

  18. The aryl hydrocarbon receptor mediates raloxifene-induced apoptosis in estrogen receptor-negative hepatoma and breast cancer cells.

    Science.gov (United States)

    O'Donnell, E F; Koch, D C; Bisson, W H; Jang, H S; Kolluri, S K

    2014-01-30

    Identification of new molecular targets for the treatment of breast cancer is an important clinical goal, especially for triple-negative breast cancer, which is refractory to existing targeted treatments. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor known primarily as the mediator of dioxin toxicity. However, the AhR can also inhibit cellular proliferation in a ligand-dependent manner and act as a tumor suppressor in mice, and thus may be a potential anticancer target. To investigate the AhR as an anticancer target, we conducted a small molecule screen to discover novel AhR ligands with anticancer properties. We identified raloxifene, a selective estrogen receptor (ER) modulator currently used in the clinic for prevention of ER-positive breast cancer and osteoporosis in post-menopausal women, as an AhR activator. Raloxifene directly bound the AhR and induced apoptosis in ER-negative mouse and human hepatoma cells in an AhR-dependent manner, indicating that the AhR is a molecular target of raloxifene and mediates raloxifene-induced apoptosis in the absence of ER. Raloxifene selectively induced apoptosis of triple-negative MDA-MB-231 breast cancer cells compared with non-transformed mammary epithelial cells via the AhR. Combined with recent data showing that raloxifene inhibits triple-negative breast cancer xenografts in vivo (Int J Oncol. 43(3):785-92, 2013), our results support the possibility of repurposing of raloxifene as an AhR-targeted therapeutic for triple-negative breast cancer patients. To this end, we also evaluated the role of AhR expression on survival of patients diagnosed with breast cancer. We found that higher expression of the AhR is significantly associated with increased overall survival and distant metastasis-free survival in both hormone-dependent (ER-positive) and hormone-independent (ER and progesterone receptor (PR)-negative) breast cancers. Together, our data strongly support the possibility of using the Ah

  19. Activation of G-Protein-Coupled Estrogen Receptor Inhibits the Migration of Human Nonsmall Cell Lung Cancer Cells via IKK-β/NF-κB Signals.

    Science.gov (United States)

    Zhu, Guangfa; Huang, Yan; Wu, Chunting; Wei, Dong; Shi, Yingxin

    2016-08-01

    Estrogen signals have been suggested to modulate the progression and metastasis of nonsmall cell lung cancer (NSCLC), which is one of the leading causes of cancer deaths worldwide. While there are limited data concerning the roles and effects of G-protein-coupled estrogen receptor (GPER) on the progression of NSCLC, our present study reveals that the expression of GPER in NSCLC cells is obviously greater than that in lung fibroblast cell line MRC-5. Activation of GPER via its specific agonist G-1 decreases the in vitro motility of A549 and H358 cells and the expression of matrix metalloproteinase 2 (MMP-2) and MMP-9. Further, G-1 treatment can rapidly decrease the phosphorylation, nuclear translocation, and promoter activities of NF-κB in NSCLC cells. BAY 11-7082, the inhibitor of NF-κB, also inhibits the expression of MMP-2/9, while overexpression of p65 significantly attenuates G-1-induced downregulation of MMP-2/9. It suggests that inhibition of NF-κB mediates G-1-induced MMP-2/9 downregulation. G-1 treatment significantly down regulates the phosphorylation of IκB kinase β (IKK-β) and IκBα, while not IKK-α, in both 549 and H358 cells. ACHP, the specific inhibitor of IKK-β, can reinforce G-1-induced MMP-2/9 downregulation and invasion suppression of A549 cells. Collectively, our results suggest that activation of GPER can inhibit the migration of human NSCLC cells via suppression of IKK-β/NF-κB signals. These findings will help to better understand the roles and mechanisms of GPER as a potential therapy target for NSCLC patients.

  20. PAC₁ receptors mediate positive chronotropic responses to PACAP-27 and VIP in isolated mouse atria.

    Science.gov (United States)

    Hoover, Donald B; Girard, Beatrice M; Hoover, Jeffrey L; Parsons, Rodney L

    2013-08-05

    PACAP and VIP have prominent effects on cardiac function in several species, but little is known about their influence on the murine heart. Accordingly, we evaluated the expression of PACAP/VIP receptors in mouse heart and the response of isolated atria to peptide agonists. Quantitative PCR demonstrated that PAC₁, VPAC₁, and VPAC₂ receptor mRNAs are present throughout the mouse heart. Expression of all three receptor transcripts was low, PAC₁ being the lowest. No regional differences in expression were detected for individual receptor mRNAs after normalization to L32. Pharmacological effects of PACAP-27, VIP, and the selective PAC₁ agonist maxadilan were evaluated in isolated, spontaneously beating atria from C57BL/6 mice of either sex. Incremental additions of PACAP-27 at 1 min intervals caused a concentration-dependent tachycardia with a logEC₅₀=-9.08 ± 0.15 M (n=7) and a maximum of 96.3 ± 5.9% above baseline heart rate. VIP and maxadilan also caused tachycardia but their potencies were about two orders of magnitude less. Increasing the dosing interval to 5 min caused a leftward shift of the concentration-response curve to maxadilan but no changes in the curves for PACAP-27 or VIP. Under this condition, neither the potency nor the efficacy of maxadilan differed from those of PACAP-27. Neither PACAP-27 nor maxadilan caused tachyphylaxis, and maximal responses to maxadilan were maintained for at least 2 h. We conclude that all three VIP/PACAP family receptors are expressed by mouse cardiac tissue, but only PAC₁ receptors mediate positive chronotropic responses to PACAP-27 and VIP. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. Heterologous activation of protein kinase C stimulates phosphorylation of delta-opioid receptor at serine 344, resulting in beta-arrestin- and clathrin-mediated receptor internalization

    DEFF Research Database (Denmark)

    Xiang, B; Yu, G H; Guo, J

    2001-01-01

    The purpose of the current study is to investigate the effect of opioid-independent, heterologous activation of protein kinase C (PKC) on the responsiveness of opioid receptor and the underlying molecular mechanisms. Our result showed that removing the C terminus of delta opioid receptor (DOR......, and ionomycin resulted in DOR internalization that required phosphorylation of Ser-344. Expression of dominant negative beta-arrestin and hypertonic sucrose treatment blocked PMA-induced DOR internalization, suggesting that PKC mediates DOR internalization via a beta-arrestin- and clathrin-dependent mechanism...... phosphorylation could inhibit PKC-catalyzed heterologous DOR phosphorylation and subsequent internalization. These data demonstrate that the responsiveness of opioid receptor is regulated by both PKC and GRK through agonist-dependent and agonist-independent mechanisms and PKC-mediated receptor phosphorylation...

  2. RNA interference-mediated targeting of DKK1 gene expression in Ishikawa endometrial carcinoma cells causes increased tumor cell invasion and migration.

    Science.gov (United States)

    Yi, Nuo; Liao, Qin-Ping; Li, Zhen-Hua; Xie, Bao-Jiang; Hu, Yu-Hong; Yi, Wei; Liu, Min

    2013-09-01

    The Wnt signaling pathway plays an essential role in tumor invasion and migration. DKK1 functions as an important inhibitor of the pathway and represents a promising target for cancer therapy. The aim of the present study was to determine the role of DKK1 in endometrial carcinoma (EC) cell invasion and migration using RNA interference (RNAi) technology. Ishikawa EC cells were transfected at high efficiency with specific DKK1 siRNA. RT-PCR and western blot analysis were used to determine the mRNA and protein levels of DKK1, β-catenin and metalloproteinase 14 (MMP14) in siRNA-treated and -untreated cells. In addition, the invasion and migration of the EC cells were detected by invasion and migration assays. Transient transfection of DKK1 siRNA significantly inhibited the mRNA and protein levels of DKK1. Markedly increased cell invasion and migration was observed following treatment with DKK1 siRNA when compared with the negative control siRNA-treated and siRNA-untreated cells. The knockdown of DKK1 also elevated the mRNA and protein levels of β-catenin and MMP14 involved in the Wnt signaling pathway, indicating that targeting this gene may promote intracellular Wnt signal transduction and thus, accelerate EC cell invasion and migration in vitro. The RNAi-mediated targeting of DKK1 gene expression in Ishikawa EC cells resulted in increased tumor cell invasion and migration. DKK1 was identified as an inhibitor of EC cell invasion and migration via its novel role in the Wnt signaling pathway. Targeting DKK1 may therefore represent an effective anti-invasion and -migration strategy for the treatment of EC.

  3. Rabenosyn-5 defines the fate of the transferrin receptor following clathrin-mediated endocytosis.

    Science.gov (United States)

    Navaroli, Deanna M; Bellvé, Karl D; Standley, Clive; Lifshitz, Lawrence M; Cardia, James; Lambright, David; Leonard, Deborah; Fogarty, Kevin E; Corvera, Silvia

    2012-02-21

    Cell surface receptors and other proteins internalize through diverse mechanisms at the plasma membrane and are sorted to different destinations. Different subpopulations of early endosomes have been described, raising the question of whether different internalization mechanisms deliver cargo into different subsets of early endosomes. To address this fundamental question, we developed a microscopy platform to detect the precise position of endosomes relative to the plasma membrane during the uptake of ligands. Axial resolution is maximized by concurrently applied total internal reflection fluorescence and epifluorescence-structured light. We found that transferrin receptors are delivered selectively from clathrin-coated pits on the plasma membrane into a specific subpopulation of endosomes enriched in the multivalent Rab GTPase and phosphoinositide-binding protein Rabenosyn-5. Depletion of Rabenosyn-5, but not of other early endosomal proteins such as early endosome antigen 1, resulted in impaired transferrin uptake and lysosomal degradation of transferrin receptors. These studies reveal a critical role for Rabenosyn-5 in determining the fate of transferrin receptors internalized by clathrin-mediated endocytosis and, more broadly, a mechanism whereby the delivery of cargo from the plasma membrane into specific early endosome subpopulations is required for its appropriate intracellular traffic.

  4. Ultraslow Water-Mediated Transmembrane Interactions Regulate the Activation of A2A Adenosine Receptor

    Science.gov (United States)

    Lee, Yoonji; Kim, Songmi; Choi, Sun; Hyeon, Changbong

    2016-09-01

    Water molecules inside G-protein coupled receptor have recently been spotlighted in a series of crystal structures. To decipher the dynamics and functional roles of internal waters in GPCR activity, we studied A$_{\\text{2A}}$ adenosine receptor using $\\mu$sec-molecular dynamics simulations. Our study finds that the amount of water flux across the transmembrane (TM) domain varies depending on the receptor state, and that the water molecules of the TM channel in the active state flow three times slower than those in the inactive state. Depending on the location in solvent-protein interface as well as the receptor state, the average residence time of water in each residue varies from $\\sim\\mathcal{O}(10^2)$ psec to $\\sim\\mathcal{O}(10^2)$ nsec. Especially, water molecules, exhibiting ultraslow relaxation ($\\sim\\mathcal{O}(10^2)$ nsec) in the active state, are found around the microswitch residues that are considered activity hotspots for GPCR function. A continuous allosteric network spanning the TM domain, arising from water-mediated contacts, is unique in the active state, underscoring the importance of slow waters in the GPCR activation.

  5. The Fc and not CD4 Receptor Mediates Antibody Enhancement of HIV Infection in Human Cells

    Science.gov (United States)

    Homsy, Jacques; Meyer, Mia; Tateno, Masatoshi; Clarkson, Sarah; Levy, Jay A.

    1989-06-01

    Antibodies that enhance human immunodeficiency virus (HIV) infectivity have been found in the blood of infected individuals and in infected or immunized animals. These findings raise serious concern for the development of a safe vaccine against acquired immunodeficiency syndrome. To address the in vivo relevance and mechanism of this phenomenon, antibody-dependent enhancement of HIV infectivity in peripheral blood macrophages, lymphocytes, and human fibroblastoid cells was studied. Neither Leu3a, a monoclonal antibody directed against the CD4 receptor, nor soluble recombinant CD4 even at high concentrations prevented this enhancement. The addition of monoclonal antibody to the Fc receptor III (anti-FcRIII), but not of antibodies that react with FcRI or FcRII, inhibited HIV type 1 and HIV type 2 enhancement in peripheral blood macrophages. Although enhancement of HIV infection in CD4+ lymphocytes could not be blocked by anti-FcRIII, it was inhibited by the addition of human immunoglobulin G aggregates. The results indicate that the FcRIII receptor on human macrophages and possibly another Fc receptor on human CD4+ lymphocytes mediate antibody-dependent enhancement of HIV infectivity and that this phenomenon proceeds through a mechanism independent of the CD4 protein.

  6. Targeting of liposomes to cells bearing nerve growth factor receptors mediated by biotinylated NGF

    Energy Technology Data Exchange (ETDEWEB)

    Rosenberg, M.B.

    1986-01-01

    Previous studies of liposome targeting have concentrated on immunological systems, the use of ligand-receptor interactions has received little attention. The protein hormone beta-nerve growth factor (NGF) was modified by biotinylation via carboxyl group substitution (C-bio-NGF) under reaction conditions that yielded an average of 3 biotin additions per NGF subunit. NGF was also biotinylated through amino group substitution to produce derivatives with ratios of 1, 2 and 4 biotin moieties per NGF subunit (N-bio-NGF). These derivatives were compared with native NGF for their ability to compete with /sup 125/I-NGF for binding to NGF receptors on rat pheochromocytoma (PC 12) cells at 4/sup 0/C. C-bio-NGF was as effective as native NGF in binding to NGF receptors, while N-bio-NGF containing 1 biotin per NGF subunit was only 28% as active in binding as native NGF. C-bio-NGF, but not N-bio-NGF, mediated the specific binding of /sup 125/I-streptavidin to PC12 cells. Biocytin-NGF, a derivative of C-bio-NGF with an extended spacer chain, was also synthesized and retained full biological and receptor binding activities. C-bio-NGF and biocytin-NGF were as effective as native NGF in a bioassay involving induction of neurite outgrowth from PC12 cells.

  7. Structural insight into antibody-mediated antagonism of the Glucagon-like peptide-1 Receptor

    Science.gov (United States)

    Hennen, Stephanie; Kodra, János T.; Soroka, Vladyslav; Krogh, Berit O.; Wu, Xiaoai; Kaastrup, Peter; Ørskov, Cathrine; Rønn, Sif G.; Schluckebier, Gerd; Barbateskovic, Silvia; Gandhi, Prafull S.; Reedtz-Runge, Steffen

    2016-01-01

    The Glucagon-like peptide-1 receptor (GLP-1R) is a member of the class B G protein-coupled receptor (GPCR) family and a well-established target for the treatment of type 2 diabetes. The N-terminal extracellular domain (ECD) of GLP-1R is important for GLP-1 binding and the crystal structure of the GLP-1/ECD complex was reported previously. The first structure of a class B GPCR transmembrane (TM) domain was solved recently, but the full length receptor structure is still not well understood. Here we describe the molecular details of antibody-mediated antagonism of the GLP-1R using both in vitro pharmacology and x-ray crystallography. We showed that the antibody Fab fragment (Fab 3F52) blocked the GLP-1 binding site of the ECD directly and thereby acts as a competitive antagonist of native GLP-1. Interestingly, Fab 3F52 also blocked a short peptide agonist believed to engage primarily the transmembrane and extracellular loop region of GLP-1R, whereas functionality of an allosteric small-molecule agonist was not inhibited. This study has implications for the structural understanding of the GLP-1R and related class B GPCRs, which is important for the development of new and improved therapeutics targeting these receptors. PMID:27196125

  8. Gut feelings of safety: tolerance to the microbiota mediated by innate immune receptors.

    Science.gov (United States)

    Swiatczak, Bartlomiej; Cohen, Irun R

    2015-10-01

    To enable microbial colonization of the gut mucosa, the intestinal immune system must not only react to danger signals but also recognize cues that indicate safety. Recognition of safety, paradoxically, is mediated by the same environmental sensors that are involved in signaling danger. Indeed, in addition to their well-established role in inducing inflammation in response to stress signals, pattern recognition receptors and a variety of metabolic sensors also promote gut-microbiota symbiosis by responding to "microbial symbiosis factors", "resolution-associated molecular patterns", markers of energy extraction and other signals indicating the absence of pathogenic infection and tissue damage. Here we focus on how the paradoxical roles of immune receptors and other environmental sensors define the microbiota signature of an individual. © 2015 The Societies and Wiley Publishing Asia Pty Ltd.

  9. Advanced Glycation End-Products and Their Receptor-Mediated Roles: Inflammation and Oxidative Stress

    Directory of Open Access Journals (Sweden)

    Parisa Younessi

    2011-09-01

    Full Text Available Glycation is a protein modification, which results in a change in a protein structure. Glycation is believed to be the etiology of various age-related diseases such as diabetes mellitus and Alzheimer’s disease (AD. Activation of microglia and resident macrophages in the brain by glycated proteins with subsequent oxidative stress and cytokine release may be an important factor in the progression of AD. It is also suggested that interaction between an advanced glycation end product (AGE and its receptor (RAGE results in glial activation as well as cytokine release and reactive oxygen species release. The use of antioxidants, receptor mediated compounds and reactive oxygen species scavenging enzyme produce an opportunity to intervene with AGE-RAGE signaling pathway, and thereby to slow down the progression of aging-related diseases

  10. β-Arrestin regulation of myosin light chain phosphorylation promotes AT1aR-mediated cell contraction and migration.

    Directory of Open Access Journals (Sweden)

    Elie Simard

    Full Text Available Over the last decade, it has been established that G-protein-coupled receptors (GPCRs signal not only through canonical G-protein-mediated mechanisms, but also through the ubiquitous cellular scaffolds β-arrestin-1 and β-arrestin-2. Previous studies have implicated β-arrestins as regulators of actin reorganization in response to GPCR stimulation while also being required for membrane protrusion events that accompany cellular motility. One of the most critical events in the active movement of cells is the cyclic phosphorylation and activation of myosin light chain (MLC, which is required for cellular contraction and movement. We have identified the myosin light chain phosphatase Targeting Subunit (MYPT-1 as a binding partner of the β-arrestins and found that β-arrestins play a role in regulating the turnover of phosphorylated myosin light chain. In response to stimulation of the angiotensin Type 1a Receptor (AT1aR, MLC phosphorylation is induced quickly and potently. We have found that β-arrestin-2 facilitates dephosphorylation of MLC, while, in a reciprocal fashion, β-arrestin 1 limits dephosphorylation of MLC. Intriguingly, loss of either β-arrestin-1 or 2 blocks phospho-MLC turnover and causes a decrease in the contraction of cells as monitored by atomic force microscopy (AFM. Furthermore, by employing the β-arrestin biased ligand [Sar(1,Ile(4,Ile(8]-Ang, we demonstrate that AT1aR-mediated cellular motility involves a β-arrestin dependent component. This suggests that the reciprocal regulation of MLC phosphorylation status by β-arrestins-1 and 2 causes turnover in the phosphorylation status of MLC that is required for cell contractility and subsequent chemotaxic motility.

  11. Remodeling of purinergic receptor-mediated Ca2+ signaling as a consequence of EGF-induced epithelial-mesenchymal transition in breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Felicity M Davis

    Full Text Available BACKGROUND: The microenvironment plays a pivotal role in tumor cell proliferation, survival and migration. Invasive cancer cells face a new set of environmental challenges as they breach the basement membrane and colonize distant organs during the process of metastasis. Phenotypic switching, such as that which occurs during epithelial-mesenchymal transition (EMT, may be associated with a remodeling of cell surface receptors and thus altered responses to signals from the tumor microenvironment. METHODOLOGY/PRINCIPAL FINDINGS: We assessed changes in intracellular Ca(2+ in cells loaded with Fluo-4 AM using a fluorometric imaging plate reader (FLIPR(TETRA and observed significant changes in the potency of ATP (EC(50 0.175 µM (-EGF versus 1.731 µM (+EGF, P<0.05, and the nature of the ATP-induced Ca(2+ transient, corresponding with a 10-fold increase in the mesenchymal marker vimentin (P<0.05. We observed no change in the sensitivity to PAR2-mediated Ca(2+ signaling, indicating that these alterations are not simply a consequence of changes in global Ca(2+ homeostasis. To determine whether changes in ATP-mediated Ca(2+ signaling are preceded by alterations in the transcriptional profile of purinergic receptors, we analyzed the expression of a panel of P2X ionotropic and P2Y metabotropic purinergic receptors using real-time RT-PCR and found significant and specific alterations in the suite of ATP-activated purinergic receptors during EGF-induced EMT in breast cancer cells. Our studies are the first to show that P2X(5 ionotropic receptors are enriched in the mesenchymal phenotype and that silencing of P2X(5 leads to a significant reduction (25%, P<0.05 in EGF-induced vimentin protein expression. CONCLUSIONS: The acquisition of a new suite of cell surface purinergic receptors is a feature of EGF-mediated EMT in MDA-MB-468 breast cancer cells. Such changes may impart advantageous phenotypic traits and represent a novel mechanism for the targeting of

  12. Scavenger receptors mediate the role of SUMO and Ftz-f1 in Drosophila steroidogenesis.

    Directory of Open Access Journals (Sweden)

    Ana Talamillo

    2013-04-01

    Full Text Available SUMOylation participates in ecdysteroid biosynthesis at the onset of metamorphosis in Drosophila melanogaster. Silencing the Drosophila SUMO homologue smt3 in the prothoracic gland leads to reduced lipid content, low ecdysone titers, and a block in the larval-pupal transition. Here we show that the SR-BI family of Scavenger Receptors mediates SUMO functions. Reduced levels of Snmp1 compromise lipid uptake in the prothoracic gland. In addition, overexpression of Snmp1 is able to recover lipid droplet levels in the smt3 knockdown prothoracic gland cells. Snmp1 expression depends on Ftz-f1 (an NR5A-type orphan nuclear receptor, the expression of which, in turn, depends on SUMO. Furthermore, we show by in vitro and in vivo experiments that Ftz-f1 is SUMOylated. RNAi-mediated knockdown of ftz-f1 phenocopies that of smt3 at the larval to pupal transition, thus Ftz-f1 is an interesting candidate to mediate some of the functions of SUMO at the onset of metamorphosis. Additionally, we demonstrate that the role of SUMOylation, Ftz-f1, and the Scavenger Receptors in lipid capture and mobilization is conserved in other steroidogenic tissues such as the follicle cells of the ovary. smt3 knockdown, as well as ftz-f1 or Scavenger knockdown, depleted the lipid content of the follicle cells, which could be rescued by Snmp1 overexpression. Therefore, our data provide new insights into the regulation of metamorphosis via lipid homeostasis, showing that Drosophila Smt3, Ftz-f1, and SR-BIs are part of a general mechanism for uptake of lipids such as cholesterol, required during development in steroidogenic tissues.

  13. Baicalein mediates inhibition of migration and invasiveness of skin carcinoma through Ezrin in A431 cells

    Directory of Open Access Journals (Sweden)

    Wu Bin

    2011-12-01

    inhibited by baicalein. Conclusions Baicalein reduces the migration and invasiveness of A431 cells through the inhibition of Ezrin expression, which leads to the suppression of tumor metastasis.

  14. Oxidative stress augments toll-like receptor 8 mediated neutrophilic responses in healthy subjects

    Directory of Open Access Journals (Sweden)

    Matsunaga Kazuto

    2009-06-01

    Full Text Available Abstract Background Excessive oxidative stress has been reported to be generated in inflamed tissues and contribute to the pathogenesis of inflammatory lung diseases, exacerbations of which induced by viral infections are associated with toll-like receptor (TLR activation. Among these receptors, TLR8 has been reported as a key receptor that recognizes single-strand RNA virus. However, it remains unknown whether TLR8 signaling is potentiated by oxidative stress. The aim of this study is to examine whether oxidative stress modulates TLR8 signaling in vitro. Methods Human peripheral blood neutrophils were obtained from healthy non-smokers and stimulated with TLR 7/8 agonist imidazoquinoline resiquimod (R848 in the presence or absence of hydrogen peroxide (H2O2. Neutrophilic responses including cytokine release, superoxide production and chemotaxis were examined, and the signal transduction was also analyzed. Results Activation of TLR8, but not TLR7, augmented IL-8 release. The R848-augmented IL-8 release was significantly potentiated by pretreatment with H2O2 (p L-cysteine reversed this potentiation. The combination of H2O2 and R848 significantly potentiated NF-kB phosphorylation and IkBα degradation. The H2O2-potentiated IL-8 release was suppressed by MG-132, a proteosome inhibitor, and by dexamethasone. The expressions of TLR8, myeloid differentiation primary response gene 88 (MyD88, and tumor necrosis factor receptor-associated factor 6 (TRAF6 were not affected by H2O2. Conclusion TLR8-mediated neutrophilic responses were markedly potentiated by oxidative stress, and the potentiation was mediated by enhanced NF-kB activation. These results suggest that oxidative stress might potentiate the neutrophilic inflammation during viral infection.

  15. Differential modulation of AMPA receptor mediated currents by Evans Blue in postnatal rat hippocampal neurones

    OpenAIRE

    Schürmann, Britta; Wu, Xueqing; Dietzel, Irmgard D; Leßmann, Volkmar

    1997-01-01

    The modulation of non-N-methyl-D-aspartate (NMDA) receptor-mediated whole cell currents and of glutamatergic synaptic transmission by purified Evans Blue (EB) was investigated in rat cultured postnatal hippocampal neurones by use of patch clamp recordings and a fast drug application system.Three different groups of neurones could be distinguished with respect to the type of modulation obtained with 10 μM EB: EB was either a predominant inhibitor of desensitization (13% of the neurones), a pre...

  16. Med1 subunit of the mediator complex in nuclear receptor-regulated energy metabolism, liver regeneration, and hepatocarcinogenesis.

    Science.gov (United States)

    Jia, Yuzhi; Viswakarma, Navin; Reddy, Janardan K

    2014-01-01

    Several nuclear receptors regulate diverse metabolic functions that impact on critical biological processes, such as development, differentiation, cellular regeneration, and neoplastic conversion. In the liver, some members of the nuclear receptor family, such as peroxisome proliferator-activated receptors (PPARs), constitutive androstane receptor (CAR), farnesoid X receptor (FXR), liver X receptor (LXR), pregnane X receptor (PXR), glucocorticoid receptor (GR), and others, regulate energy homeostasis, the formation and excretion of bile acids, and detoxification of xenobiotics. Excess energy burning resulting from increases in fatty acid oxidation systems in liver generates reactive oxygen species, and the resulting oxidative damage influences liver regeneration and liver tumor development. These nuclear receptors are important sensors of exogenous activators as well as receptor-specific endogenous ligands. In this regard, gene knockout mouse models revealed that some lipid-metabolizing enzymes generate PPARα-activating ligands, while others such as ACOX1 (fatty acyl-CoA oxidase1) inactivate these endogenous PPARα activators. In the absence of ACOX1, the unmetabolized ACOX1 substrates cause sustained activation of PPARα, and the resulting increase in energy burning leads to hepatocarcinogenesis. Ligand-activated nuclear receptors recruit the multisubunit Mediator complex for RNA polymerase II-dependent gene transcription. Evidence indicates that the Med1 subunit of the Mediator is essential for PPARα, PPARγ, CAR, and GR signaling in liver. Med1 null hepatocytes fail to respond to PPARα activators in that these cells do not show induction of peroxisome proliferation and increases in fatty acid oxidation enzymes. Med1-deficient hepatocytes show no increase in cell proliferation and do not give rise to liver tumors. Identification of nuclear receptor-specific coactivators and Mediator subunits should further our understanding of the complexities of metabolic

  17. IL-17 receptor signaling and T helper 17-mediated autoimmune demyelinating disease.

    Science.gov (United States)

    Zepp, Jarod; Wu, Ling; Li, Xiaoxia

    2011-05-01

    Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS). Experimental autoimmune encephalomyelitis (EAE) is widely used to dissect molecular mechanisms of MS and to develop new therapeutic strategies. The T helper 17 (Th17) subset of CD4 T cells plays a crucial role in the development of EAE. IL-17, a cytokine produced by Th17 cells, participates in EAE pathogenesis through induction of inflammatory gene expression in target cells. Recent work has shown that Act1, a U-box E3 ubiquitin ligase, is recruited to IL-17 receptor (IL-17R) upon IL-17 stimulation and is required for IL-17-mediated signaling. Here, we review the molecular and cellular mechanisms by which IL-17 and Act1-mediated signaling contribute to EAE. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. The Expression of Leptin, Estrogen Receptors, and Vitellogenin mRNAs in Migrating Female Chum Salmon, : The Effects of Hypo-osmotic Environmental Changes

    Directory of Open Access Journals (Sweden)

    Young Jae Choi

    2014-04-01

    Full Text Available Leptin plays an important role in energy homeostasis and reproductive function in fish, especially in reproduction. Migrating fish, such as salmonoids, are affected by external environmental factors, and salinity changes are a particularly important influence on spawning migrations. The aim of this study was to test whether changes in salinity affect the expression of leptin, estrogen receptors (ERs, and vitellogenin (VTG in chum salmon (Oncorhynchus keta. The expression and activity of leptin, the expression of ERs and VTG, and the levels of estradiol-17β and cortisol increased after the fish were transferred to FW, demonstrating that changes in salinity stimulate the HPG axis in migrating female chum salmon. These findings reveal details about the role of elevated leptin levels and sex steroid hormones in stimulating sexual maturation and reproduction in response to salinity changes in chum salmon.

  19. Epithelial estrogen receptor 1 intrinsically mediates squamous differentiation in the mouse vagina.

    Science.gov (United States)

    Miyagawa, Shinichi; Iguchi, Taisen

    2015-10-20

    Estrogen-mediated actions in female reproductive organs are tightly regulated, mainly through estrogen receptor 1 (ESR1). The mouse vaginal epithelium cyclically exhibits cell proliferation and differentiation in response to estrogen and provides a unique model for analyzing the homeostasis of stratified squamous epithelia. To address the role of ESR1-mediated tissue events during homeostasis, we analyzed mice with a vaginal epithelium-specific knockout of Esr1 driven by keratin 5-Cre (K5-Esr1KO). We show here that loss of epithelial ESR1 in the vagina resulted in aberrant epithelial cell proliferation in the suprabasal cell layers and led to failure of keratinized differentiation. Gene expression analysis showed that several known estrogen target genes, including erbB growth factor ligands, were not induced by estrogen in the K5-Esr1KO mouse vagina. Organ culture experiments revealed that the addition of erbB growth factor ligands, such as amphiregulin, could activate keratinized differentiation in the absence of epithelial ESR1. Thus, epithelial ESR1 integrates estrogen and growth factor signaling to mediate regulation of cell proliferation in squamous differentiation, and our results provide new insights into estrogen-mediated homeostasis in female reproductive organs.

  20. [Programmed necrosis mediated by receptor-interacting protein 3: a new target for liver disease research].

    Science.gov (United States)

    Zhang, J; Jing, Y; Li, Y N; Zhou, L; Wang, B M

    2016-09-20

    Hepatocyte death mainly includes apoptosis and necrosis and is a critical process in the pathophysiological mechanism of liver injury caused by various reasons. Recent studies have shown that key regulatory molecules in the inhibition of apoptosis such as caspase cannot be used as targets for inhibiting disease progression in clinical practice. In recent years, programmed necrosis mediated by receptor-interacting protein 3(RIP3)becomes a new hot research topic. It not only plays an important role in inducing inflammatory response, but also is closely regulated by intracellular signal factors, and it is a type of active cell death which can be interfered with. Compared with apoptosis, programmed necrosis is accompanied by the release of various inflammatory factors, which significantly affects local immune microenvironment. RIP3-mediated programmed necrosis has been taken seriously in many diseases. Although its mechanism of action in liver disease remains unclear, the results of recent studies confirmed its important role in the development of liver disease. This article reviews the research advances in the role of RIP3-mediated programmed necrosis signaling pathway in liver disease of various causes and investigates the possibility of RIP3-mediated programmed necrosis as a new target in the treatment of liver disease.

  1. Characterization of the 5-hydroxytryptamine receptors mediating contraction in the pig isolated intravesical ureter

    Science.gov (United States)

    Hernández, Medardo; Barahona, María Victoria; Simonsen, Ulf; Recio, Paz; Rivera, Luis; Martínez, Ana Cristina; García-Sacristán, Albino; Orensanz, Luis M; Prieto, Dolores

    2003-01-01

    This study was designed to investigate the effect of 5-hydroxytryptamine (5-HT) and to characterize the 5-HT receptors involved in 5-HT responses in the pig intravesical ureter. 5-HT (0.01–10 μM) concentration-dependently increased the tone of intravesical ureteral strips, whereas the increases in phasic contractions were concentration-independent. The 5-HT2 receptor agonist α-methyl 5-HT, mimicked the effect on tone whereas weak or no response was obtained with 5-CT, 8-OH-DPAT, m-chlorophenylbiguanide and RS 67333, 5-HT1, 5-HT1A, 5-HT3 and 5-HT4 receptor agonists, respectively. 5-HT did not induce relaxation of U46619-contracted ureteral preparations. Pargyline (100 μM), a monoaminooxidase A/B activity inhibitor, produced leftward displacements of the concentration-response curves for 5-HT. 5-HT-induced tone was reduced by the 5-HT2 and 5-HT2A receptor antagonists ritanserine (0.1 μM) and spiperone (0.2 μM), respectively. However, 5-HT contraction was not antagonized by cyanopindolol (2 μM), SDZ–SER 082 (1 μM), Y-25130 (1 μM) and GR 113808 (0.1 μM), which are respectively, 5-HT1A/1B, 5-HT2B/2C, 5-HT3, and 5-HT4 selective receptor antagonists. Removal of the urothelium did not modify 5-HT-induced contractions. Blockade of neuronal voltage-activated sodium channels, α-adrenergic receptors and adrenergic neurotransmission with tetrodotoxin (1 μM), phentolamine (0.3 μM) and guanethidine (10 μM), respectively, reduced the contractions to 5-HT. However, physostigmine (1 μM), atropine (0.1 μM) and suramin (30 μM), inhibitors of cholinesterase activity, muscarinic- and purinergic P2-receptors, respectively, failed to modify the contractions to 5-HT. These results suggest that 5-HT increases the tone of the pig intravesical ureter through 5-HT2A receptors located at the smooth muscle. Part of the 5-HT contraction is indirectly mediated via noradrenaline release from sympathetic nerves. PMID:12522083

  2. Interleukin-1 receptor-associated kinase-1 plays an essential role for Toll-like receptor (TLR)7- and TLR9-mediated interferon-α induction

    OpenAIRE

    Uematsu, Satoshi; Sato, Shintaro; Yamamoto, Masahiro; Hirotani, Tomonori; Kato, Hiroki; Takeshita, Fumihiko; Matsuda, Michiyuki; Coban, Cevayir; Ishii, Ken J.; Kawai, Taro; Takeuchi, Osamu; Akira, Shizuo

    2005-01-01

    Toll-like receptors (TLRs) recognize microbial pathogens and trigger innate immune responses. Among TLR family members, TLR7, TLR8, and TLR9 induce interferon (IFN)-α in plasmacytoid dendritic cells (pDCs). This induction requires the formation of a complex consisting of the adaptor MyD88, tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) and IFN regulatory factor (IRF) 7. Here we show an essential role of IL-1 receptor-associated kinase (IRAK)-1 in TLR7- and TLR9-mediated IRF7...

  3. NFIB Mediates BRN2 Driven Melanoma Cell Migration and Invasion Through Regulation of EZH2 and MITF

    Directory of Open Access Journals (Sweden)

    Mitchell E. Fane

    2017-02-01

    Full Text Available While invasion and metastasis of tumour cells are the principle factor responsible for cancer related deaths, the mechanisms governing the process remain poorly defined. Moreover, phenotypic divergence of sub-populations of tumour cells is known to underpin alternative behaviors linked to tumour progression such as proliferation, survival and invasion. In the context of melanoma, heterogeneity between two transcription factors, BRN2 and MITF, has been associated with phenotypic switching between predominantly invasive and proliferative behaviors respectively. Epigenetic changes, in response to external cues, have been proposed to underpin this process, however the mechanism by which the phenotypic switch occurs is unclear. Here we report the identification of the NFIB transcription factor as a novel downstream effector of BRN2 function in melanoma cells linked to the migratory and invasive characteristics of these cells. Furthermore, the function of NFIB appears to drive an invasive phenotype through an epigenetic mechanism achieved via the upregulation of the polycomb group protein EZH2. A notable target of NFIB mediated up-regulation of EZH2 is decreased MITF expression, which further promotes a less proliferative, more invasive phenotype. Together our data reveal that NFIB has the ability to promote dynamic changes in the chromatin state of melanoma cells to facilitate migration, invasion and metastasis.

  4. Calcium sensing receptor as a novel mediator of adipose tissue dysfunction: mechanisms and potential clinical implications

    Directory of Open Access Journals (Sweden)

    Roberto Bravo

    2016-09-01

    Full Text Available Obesity is currently a serious worldwide public health problem, reaching pandemic levels. For decades, dietary and behavioral approaches have failed to prevent this disease from expanding, and health authorities are challenged by the elevated prevalence of co-morbid conditions. Understanding how obesity-associated diseases develop from a basic science approach is recognized as an urgent task to face this growing problem. White adipose tissue is an active endocrine organ, with a crucial influence on whole-body homeostasis. White adipose tissue dysfunction plays a key role linking obesity with its associated diseases such as type 2 diabetes mellitus, cardiovascular disease and some cancers. Among the regulators of white adipose tissue physiology, the calcium-sensing receptor has arisen as a potential mediator of white adipose tissue dysfunction. Expression of the receptor has been described in human preadipocytes, adipocytes, and the human adipose cell lines LS14 and SW872. The evidence suggests that calcium-sensing receptor activation in the visceral (i.e. unhealthy white adipose tissue is associated with an increased proliferation of adipose progenitor cells and elevated adipocyte differentiation. In addition, exposure of adipose cells to calcium-sensing receptor activators in vitro elevates proinflammatory cytokine expression and secretion. An increased proinflammatory environment in white adipose tissue plays a key role in the development of white adipose tissue dysfunction that leads to peripheral organ fat deposition and insulin resistance, among other consequences. We propose that calcium-sensing receptor may be one relevant therapeutic target in the struggle to confront the health consequences of the current worldwide obesity pandemic.

  5. The Arabidopsis flagellin receptor FLS2 mediates the perception of Xanthomonas Ax21 secreted peptides.

    Science.gov (United States)

    Danna, Cristian H; Millet, Yves A; Koller, Teresa; Han, Sang-Wook; Bent, Andrew F; Ronald, Pamela C; Ausubel, Frederick M

    2011-05-31

    Detection of microbes by plants relies in part on an array of pattern-recognition receptors that recognize conserved microbial signatures, so-called "microbe-associated molecular patterns." The Arabidopsis thaliana receptor-like kinase FLS2 is the pattern-recognition receptor for bacterial flagellin. Similarly to FLS2, the rice transmembrane protein XA21 is the receptor for the sulfated form of the Xanthomonas oryzae pv. oryzae secreted protein Ax21. Here we show that Ax21-derived peptides activate Arabidopsis immunity, triggering responses similar to those elicited by flagellin, including an oxidative burst, induction of defense-response genes, and enhanced resistance to bacterial pathogens. To identify Arabidopsis Xa21 functional homologs, we used a reverse genetics approach to screen T-DNA insertion mutants corresponding to all 47 of the Arabidopsis genes encoding non-RD kinases belonging to the interleukin-1 receptor-associated kinase (IRAK) family. Surprisingly, among all of these mutant lines, only fls2 mutants exhibited a significant loss of response to Ax21-derived peptides. Ax21 peptides also failed to activate defense-related responses in an fls2-24 mutant that does not bind Flg22. Moreover, a Flg22Δ2 variant of Flg22 that binds to FLS2 but does not activate FLS2-mediated signaling suppressed Ax21-derived peptide signaling, indicating mutually exclusive perception of Flg22 or Ax21 peptides by FLS2. The data indicate that FLS2 functions beyond flagellin perception to detect other microbe-associated molecular patterns.

  6. Dopamine Receptor Mediation of the Exploratory/Hyperactivity Effects of Modafinil

    Science.gov (United States)

    Young, Jared W; Kooistra, Klaas; Geyer, Mark A

    2011-01-01

    Modafinil (2-((diphenylmethyl)sulfinyl)acetamide) is described as an atypical stimulant and is a putative cognition enhancer for schizophrenia, but the precise mechanisms of action remain unclear. Receptor knockout (KO) mice offer an opportunity to identify receptors that contribute to a drug-induced effect. Here we examined the effects of modafinil on exploration in C57BL/6J mice, in dopamine drd1, drd2, drd3, and drd4 wild-type (WT), heterozygous (HT), and KO mice, and in 129/SJ mice pretreated with the drd1 antagonist SCH23390 using a cross-species test paradigm based on the behavioral pattern monitor. Modafinil increased activity, specific exploration (rearing), and the smoothness of locomotor paths (reduced spatial d) in C57BL/6J and 129/SJ mice (increased holepoking was also observed in these mice). These behavioral profiles are similar to that produced by the dopamine transporter inhibitor GBR12909. Modafinil was ineffective at increasing activity in male drd1 KOs, rearing in female drd1 KOs, or reducing spatial d in all drd1 KOs, but produced similar effects in drd1 WT and HT mice as in C57BL/6J mice. Neither dopamine drd2 nor drd3 mutants attenuated modafinil-induced effects. Drd4 mutants exhibited a genotype dose-dependent attenuation of modafinil-induced increases in specific exploration. Furthermore, the drd1 KO effects were largely supported by the SCH23390 study. Thus, the dopamine drd1 receptor appears to exert a primary role in modafinil-induced effects on spontaneous exploration, whereas the dopamine drd4 receptor appears to be important for specific exploration. The modafinil-induced alterations in exploratory behavior may reflect increased synaptic dopamine and secondary actions mediated by dopamine drd1 and drd4 receptors. PMID:21412225

  7. Anti-CD22-mediated change of adhesion molecule expression and migrational behavior of B cells from patients with SLE in comparison to healthy donors

    OpenAIRE

    Blaßfeld, Daniela Anneliese Karin

    2013-01-01

    Epratuzumab, a humanized monoclonal antibody targeting CD22 has reduced disease activity in SLE patients in clinical studies and represents a potential new therapeutic agent for the treatment of SLE. Further, a reduction in peripheral blood B lymphocytes has been described after epratuzumab treatment. Therefore this dissertation analyzes the influence of epratuzumab on expression and functionality of receptors of B cell migration in vitro using fluorescence activated cell sorting and a functi...

  8. Receptor-mediated binding and uptake of GnRH agonist and antagonist by pituitary cells

    Energy Technology Data Exchange (ETDEWEB)

    Jennes, L.; Stumpf, W.E.; Conn, P.M.

    1984-01-01

    The intracellular pathway of an enzyme resistant GnRH agonist (D- Lys6 -GnRH) conjugated to ferritin or to colloidal gold was followed in cultured pituitary cells. After an initial uniform distribution over the cell surface of gonadotropes, the electrondense marker was internalized, either individually or in small groups. After longer incubation times, the marker appeared in the lysosomal compartment and the Golgi apparatus, where it could be found in the vesicular as well as cisternal portion. In addition, the receptor-mediated endocytosis of the GnRH antagonist D-p-Glu1-D-Phe2-D-Trp3-D- Lys6 -GnRH was studied by light and electron microscopic autoradiography after 30 and 60 min of incubation to ensure uptake. At both time points, in in vitro as well as in vivo studies, silver grains were localized over cytoplasmic organelles of castration cells, including dilated endoplasmic reticulum, lysosomes, and clear vesicles. No consistent association with cell nuclei, mitochondria, or secretory vesicles could be observed. The results suggest that both agonist and antagonist are binding selectively to the plasma membrane of gonadotropes and subsequently are taken up via receptor-mediated endocytosis for degradation or possible action on synthetic processes.

  9. Receptor-mediated endocytosis of lysozyme in renal proximal tubules of the frog Rana temporaria

    Directory of Open Access Journals (Sweden)

    E.V. Seliverstova

    2015-04-01

    Full Text Available The mechanism of protein reabsorption in the kidney of lower vertebrates remains insufficiently investigated in spite of raising interest to the amphibian and fish kidneys as a useful model for physiological and pathophysiological examinations. In the present study, we examined the renal tubular uptake and the internalization rote of lysozyme after its intravenous injection in the wintering frog Rana temporaria using immunohisto- and immunocytochemistry and specific markers for some endocytic compartments. The distinct expression of megalin and cubilin in the proximal tubule cells of lysozyme-injected frogs was revealed whereas kidney tissue of control animals showed no positive immunoreactivity. Lysozyme was detected in the apical endocytic compartment of the tubular cells and colocalized with clathrin 10 min after injection. After 20 min, lysozyme was located in the subapical compartment negative to clathrin (endosomes, and intracellular trafficking of lysozyme was coincided with the distribution of megalin and cubilin. However, internalized protein was retained in the endosomes and did not reach lysosomes within 30 min after treatment that may indicate the inhibition of intracellular trafficking in hibernating frogs. For the first time, we provided the evidence that lysozyme is filtered through the glomeruli and absorbed by receptor-mediated clathrin-dependent endocytosis in the frog proximal tubule cells. Thus, the protein uptake in the amphibian mesonephros is mediated by megalin and cubilin that confirms a critical role of endocytic receptors in the renal reabsorption of proteins in amphibians as in mammals.

  10. Receptor mediated endocytosis of vicilin in Callosobruchus maculatus (Coleoptera: Chrysomelidae) larval midgut epithelial cells.

    Science.gov (United States)

    Kunz, Daniele; Oliveira, Gabriel B; Uchôa, Adriana F; Samuels, Richard I; Macedo, Maria Lígia R; Silva, Carlos P

    2017-08-01

    The transport of proteins across the intestinal epithelium of insects is still not well understood. There is evidence that vicilin, a major storage protein of cowpea seeds (Vigna unguiculata), is internalized in larvae of the seed-beetle Callosobruchus maculatus. It has been reported that this vicilin interacts with proteins present in the microvillar membranes of columnar cells along the digestive tract of the larvae. In the present work, we studied the cellular pathway involved in endocytosis of vicilin in larval C. maculatus by employing ex vivo experiments. In the ex vivo approach, we incubated FITC-labelled vicilin with isolated midgut wholemounts in the absence or in the presence of endocytosis inhibitors. The fate of labelled or non-labelled globulins was monitored by confocal microscopy and fluorescence measurement. Our results suggest that the internalization of vicilins is due to receptor-mediated endocytosis. Here we report the identity of a microvillar vicilin-binding protein that was purified using affinity chromatography on a vicilin-sepharose column. The putative vicilin receptor showed high homology to proteins with the CRAL-TRIO domain, specifically the Sec14 superfamily member α-tocopherol transfer protein. The precise mechanism involved in vicilin internalization was defined through the use of specific inhibitors of the endocytosis pathway. The inhibitors filipin III and nystatin significantly inhibited the endocytosis of vicilin, while chlorpromazine and phenylarsine oxide had a much lower effect on endocytosis, suggesting that the endocytic pathway is predominantly mediated by caveolin. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Blockade of cannabinoid 1 receptor improves GLP-1R mediated insulin secretion in mice.

    Science.gov (United States)

    González-Mariscal, Isabel; Krzysik-Walker, Susan M; Kim, Wook; Rouse, Michael; Egan, Josephine M

    2016-03-05

    The cannabinoid 1 receptor (CB1) is an important regulator of energy metabolism. Reports of in vivo and in vitro studies give conflicting results regarding its role in insulin secretion, possibly due to circulatory factors, such as incretins. We hypothesized that this receptor may be a regulator of the entero-insular axis. We found that despite lower food consumption and lower body weight postprandial GLP-1 plasma concentrations were increased in CB1(-/-) mice compared to CB1(+/+) mice administered a standard diet or high fat/sugar diet. Upon exogenous GLP-1 treatment, CB1(-/-) mice had increased glucose-stimulated insulin secretion. In mouse insulinoma cells, cannabinoids reduced GLP-1R-mediated intracellular cAMP accumulation and subsequent insulin secretion. Importantly, such effects were also evident in human islets, and were prevented by pharmacologic blockade of CB1. Collectively, these findings suggest a novel mechanism in which endocannabinoids are negative modulators of incretin-mediated insulin secretion. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  12. 5-HT2A receptor is the predominant receptor mediating contraction of the isolated porcine distal ureter to 5-HT in young and old animals.

    Science.gov (United States)

    Lim, Iris; Chess-Williams, Russ; Sellers, Donna

    2017-11-07

    Isolated ureteral strips develop spontaneous phasic contractile activity which is enhanced by 5-hydroytryptamine (5-HT). The aim of this study was to identify the receptor subtype mediating these responses and to determine whether responses to 5-HT change with age. The frequency of contractions and the overall contractile activity (measured as the area under the curve, AUC) were recorded in strips of porcine distal ureter isolated from young (3 months) and old (2 years) pigs. Responses to 5-HT were examined in the absence and presence of selective 5-HT receptor subtype antagonists. Tissues from the younger animals elicited larger contractile responses to 5-HT (5885 ± 335g-1 s) than tissues from the older animals (2787 ± 317g-1 s, P receptor antagonist ketanserin (10-100nM) elicited rightward shifts of 5-HT concentration-response curves, antagonising AUC and frequency responses with high affinity in tissues from both age groups (pKD values 8.4-8.8). The slopes of the corresponding Schild plots were not significantly different from unity, suggesting a competitive antagonism at a single receptor, except for frequency responses to 5-HT in the older animals. Antagonists selective for other 5-HT receptor subtypes: methiothepin (non-selective), RS-10221 (5-HT2C), ondansetron (5-HT3), GR-113808 (5-HT4), SB699551 (5-HT5), SB399885 (5-HT6), SB269970 (5-HT7)) had no effect on 5-HT-induced responses. The results suggest that the 5-HT2A receptor subtype is the predominant receptor mediating 5-HT responses in ureteral tissues, being the sole mediator of responses in tissues from young animals, but with another receptor subtype also playing a minor role in the older animals. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Adenosine inhibits renin release from juxtaglomerular cells via an A1 receptor-TRPC-mediated pathway

    Science.gov (United States)

    Ortiz-Capisano, M. Cecilia; Atchison, Douglas K.; Harding, Pamela; Lasley, Robert D.

    2013-01-01

    Renin is synthesized and released from juxtaglomerular (JG) cells. Adenosine inhibits renin release via an adenosine A1 receptor (A1R) calcium-mediated pathway. How this occurs is unknown. In cardiomyocytes, adenosine increases intracellular calcium via transient receptor potential canonical (TRPC) channels. We hypothesized that adenosine inhibits renin release via A1R activation, opening TRPC channels. However, higher concentrations of adenosine may stimulate renin release through A2R activation. Using primary cultures of isolated mouse JG cells, immunolabeling demonstrated renin and A1R in JG cells, but not A2R subtypes, although RT-PCR indicated the presence of mRNA of both A2AR and A2BR. Incubating JG cells with increasing concentrations of adenosine decreased renin release. Different concentrations of the adenosine receptor agonist N-ethylcarboxamide adenosine (NECA) did not change renin. Activating A1R with 0.5 μM N6-cyclohexyladenosine (CHA) decreased basal renin release from 0.22 ± 0.05 to 0.14 ± 0.03 μg of angiotensin I generated per milliliter of sample per hour of incubation (AngI/ml/mg prot) (P renin. Reducing extracellular calcium with EGTA increased renin release (0.35 ± 0.08 μg AngI/ml/mg prot; P renin inhibition by CHA (0.28 ± 0.06 μg AngI/ml/mg prot; P renin release by 55%, and blocked the inhibitory effect of CHA. Repeating these experiments in JG cells from A1R knockout mice using CHA or NECA demonstrated no effect on renin release. However, RT-PCR showed mRNA from TRPC isoforms 3 and 6 in isolated JG cells. Adding the TRPC blocker SKF-96365 reversed CHA-mediated inhibition of renin release. Thus A1R activation results in a calcium-dependent inhibition of renin release via TRPC-mediated calcium entry, but A2 receptors do not regulate renin release. PMID:23884142

  14. Transient receptor potential vanilloid type 1 is vital for (-)-epigallocatechin-3-gallate mediated activation of endothelial nitric oxide synthase.

    Science.gov (United States)

    Guo, Bei-Chia; Wei, Jeng; Su, Kuo-Hui; Chiang, An-Na; Zhao, Jin-Feng; Chen, Hsiang-Ying; Shyue, Song-Kun; Lee, Tzong-Shyuan

    2015-04-01

    Epigallocatechin-3-gallate (EGCG), the most abundant catechin of green tea, has beneficial effects on physiological functions of endothelial cells (ECs), yet the detailed mechanisms are not fully understood. In this study, we investigated the role of transient receptor potential vanilloid type 1 (TRPV1), a ligand-gated nonselective calcium channel, in EGCG-mediated endothelial nitric oxide (NO) synthase (eNOS) activation and angiogenesis. In ECs, treatment with EGCG time-dependently increased the intracellular level of Ca(2+) . Removal of extracellular calcium (Ca(2+) ) by EGTA or EDTA or inhibition of TRPV1 by capsazepine or SB366791 abrogated EGCG-increased intracellular Ca(2+) level in ECs or TRPV1-transfected HEK293 cells. Additionally, EGCG increased the phsophorylation of eNOS at Ser635 and Ser1179, Akt at Ser473, calmodulin-dependent protein kinase II (CaMKII) at Thr286 and AMP-activated protein kinase (AMPK) at Thr172, all abolished by the TRPV1 antagonist capsazepine. EGCG-induced NO production was diminished by pretreatment with LY294002 (an Akt inhibitor), KN62 (a CaMKII inhibitor), and compound C (an AMPK inhibitor). Moreover, blocking TRPV1 activation prevented EGCG-induced EC proliferation, migration, and tube formation, as well as angiogenesis in Matrigel plugs in mice. EGCG may trigger activation of TRPV1-Ca(2+) signaling, which leads to phosphorylation of Akt, AMPK, and CaMKII; eNOS activation; NO production; and, ultimately, angiogenesis in ECs. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. PHP14 regulates hepatic stellate cells migration in liver fibrosis via mediating TGF-β1 signaling to PI3Kγ/AKT/Rac1 pathway.

    Science.gov (United States)

    Xu, Anjian; Li, Yanmeng; Zhao, Wenshan; Hou, Fei; Li, Xiaojin; Sun, Lan; Chen, Wei; Yang, Aiting; Wu, Shanna; Zhang, Bei; Yao, Jingyi; Wang, Huan; Huang, Jian

    2018-02-01

    Hepatic fibrosis is characterized by the activation of hepatic stellate cells (HSCs). Migration of the activated HSCs to the site of injury is one of the key characteristics during the wound healing process. We have previously demonstrated that 14 kDa phosphohistidine phosphatase (PHP14) is involved in migration and lamellipodia formation of HSCs. However, the role of PHP14 in liver fibrosis remains unknown. In this study, we first assessed PHP14 expression and distribution in liver fibrotic tissues using western blot, immunohistochemistry, and double immunofluorescence staining. Next, we investigated the role of PHP14 in liver fibrosis and, more specifically, the migration of HSCs by Transwell assay and 3D collagen matrices assay. Finally, we explored the possible molecular mechanisms of the effects of PHP14 on these processes. Our results show that the PHP14 expression is up-regulated in fibrotic liver and mainly in HSCs. Importantly, TGF-β1 can induce PHP14 expression in HSCs accompanied with the activation of HSCs. Consistent with the previous study, PHP14 promotes HSCs migration, especially, promotes 3D floating collagen matrices contraction but inhibits stressed-released matrices contraction. Mechanistically, the PI3Kγ/AKT/Rac1 pathway is involved in migration regulated by PHP14. Moreover, PHP14 specifically mediates the TGF-β1 signaling to PI3Kγ/AKT pathway and regulates HSC migration, and thus participates in liver fibrosis. Our study identified the role of PHP14 in liver fibrosis, particularly HSC migration, and suggested a novel mediator of transducting TGF-β1 signaling to PI3Kγ/AKT/Rac1 pathway. PHP14 is up-regulated in fibrotic liver and activated hepatic stellate cells. The expression of PHP14 is induced by TGF-β1. The migration of hepatic stellate cells is regulated by PHP14. PHP14 is a mediator of TGF-β1 signaling to PI3Kγ/AKT/Rac1 pathway in hepatic stellate cells.

  16. The Melanocortin 3 Receptor: A Novel Mediator of Exercise-Induced Inflammation Reduction in Postmenopausal Women?

    Directory of Open Access Journals (Sweden)

    Tara M. Henagan

    2011-01-01

    Full Text Available The purpose of this study was to determine whether resistance exercise training-induced reductions in inflammation are mediated via melanocortin 3 receptor expression in obese (BMI 32.7±3.7 women (65.6±2.8 yrs randomized to either a control (=11 or resistance training group (=12. The resistance trained group performed resistance training 3 days/week for 12 weeks. Resting blood samples were collected before and after the training intervention in both resistance trained and control groups. Resistance training upregulated melanocortin 3 receptor mRNA by 16-fold (=.035 and decreased monocyte count, without changing leukocyte number, body composition, or body weight. Resistance trained individuals exhibited increased sensitivity to inflammatory stimuli, whereas control individuals exhibited no change. While there was no change in whole blood tumor necrosis factor alpha mRNA between the groups, whole blood interleukin 10 mRNA was higher in the resistance trained group following the intervention period. In summary, it appears that resistance training may modulate melanocortin 3 receptor expression, providing a possible mechanism for the anti-inflammatory effects of exercise training.

  17. Nuclear receptor co-repressor SMRT regulates mitochondrial oxidative metabolism and mediates aging related metabolic deterioration

    Science.gov (United States)

    Reilly, Shannon M.; Bhargava, Prerna; Liu, Sihao; Gangl, Matthew R.; Gorgun, Cem; Nofsinger, Russell R.; Evans, Ronald M.; Qi, Lu; Hu, Frank B.; Lee, Chih-Hao

    2010-01-01

    Summary The transcriptional co-repressor SMRT utilizes two major receptor interacting domains (RID1 and RID2) to mediate nuclear receptor (NR) signaling through epigenetic modification. The physiological significance of such interaction remains unclear. We find SMRT expression and its occupancy on peroxisome proliferator-activated receptor (PPAR) target gene promoters are increased with age in major metabolic tissues. Genetic manipulations to selectively disable RID1 (SMRTmRID1) demonstrate that shifting SMRT repression to RID2-associated NRs, notably PPARs, causes premature aging and related metabolic diseases accompanied by reduced mitochondrial function and anti-oxidant gene expression. SMRTmRID1 cells exhibit increased susceptibility to oxidative damage, which could be rescued by PPAR activation or anti-oxidant treatment. In concert, several human SMRT gene polymorphisms are found to nominally associate with type 2 diabetes and adiponectin levels. These data uncover a role for SMRT in mitochondrial oxidative metabolism and the aging process, which may serve as a drug target to improve health span. PMID:21109196

  18. Nuclear receptor corepressor SMRT regulates mitochondrial oxidative metabolism and mediates aging-related metabolic deterioration.

    Science.gov (United States)

    Reilly, Shannon M; Bhargava, Prerna; Liu, Sihao; Gangl, Matthew R; Gorgun, Cem; Nofsinger, Russell R; Evans, Ronald M; Qi, Lu; Hu, Frank B; Lee, Chih-Hao

    2010-12-01

    The transcriptional corepressor SMRT utilizes two major receptor-interacting domains (RID1 and RID2) to mediate nuclear receptor (NR) signaling through epigenetic modification. The physiological significance of such interaction remains unclear. We find SMRT expression and its occupancy on peroxisome proliferator-activated receptor (PPAR) target gene promoters are increased with age in major metabolic tissues. Genetic manipulations to selectively disable RID1 (SMRT(mRID1)) demonstrate that shifting SMRT repression to RID2-associated NRs, notably PPARs, causes premature aging and related metabolic diseases accompanied by reduced mitochondrial function and antioxidant gene expression. SMRT(mRID1) cells exhibit increased susceptibility to oxidative damage, which could be rescued by PPAR activation or antioxidant treatment. In concert, several human Smrt gene polymorphisms are found to nominally associate with type 2 diabetes and adiponectin levels. These data uncover a role for SMRT in mitochondrial oxidative metabolism and the aging process, which may serve as a drug target to improve health span. Copyright © 2010 Elsevier Inc. All rights reserved.

  19. Peptides in receptor-mediated radiotherapy : from design to the clinical application in cancers.

    Directory of Open Access Journals (Sweden)

    Catherine eLOZZA

    2013-09-01

    Full Text Available Short peptides can show high affinity for specific receptors overexpressed on tumor cells. Some of these are already used in cancerology as diagnostic tools and others are in clinical trials for therapeutic applications. Therefore, peptides exhibit great potential as a diagnostic tool but also as an alternative or an additional antitumoral approach upon the covalent attachment of a therapeutic moiety such as a radionuclide or a cytotoxic drug. The chemistry offers flexibility to graft onto the targeting peptide either fluorine or iodine directly, or metallic radionuclides through appropriate chelating agent. Since short peptides are straightforward to synthesize, there is an opportunity to further improve existing peptides or to design new ones for clinical applications. However, several considerations have to be taken into account to optimize the recognition properties of the targeting peptide to its receptor, to improve its stability in the biological fluids and its residence in the body, or to increase its overall therapeutic effect. In this review, we highlight the different aspects which need to be considered for the development of an efficient peptide receptor-mediated radionuclide therapy in different neoplasms.

  20. Inhibition of insulin/IGF-1 receptor signaling protects from mitochondria-mediated kidney failure.

    Science.gov (United States)

    Ising, Christina; Koehler, Sybille; Brähler, Sebastian; Merkwirth, Carsten; Höhne, Martin; Baris, Olivier R; Hagmann, Henning; Kann, Martin; Fabretti, Francesca; Dafinger, Claudia; Bloch, Wilhelm; Schermer, Bernhard; Linkermann, Andreas; Brüning, Jens C; Kurschat, Christine E; Müller, Roman-Ulrich; Wiesner, Rudolf J; Langer, Thomas; Benzing, Thomas; Brinkkoetter, Paul Thomas

    2015-03-01

    Mitochondrial dysfunction and alterations in energy metabolism have been implicated in a variety of human diseases. Mitochondrial fusion is essential for maintenance of mitochondrial function and requires the prohibitin ring complex subunit prohibitin-2 (PHB2) at the mitochondrial inner membrane. Here, we provide a link between PHB2 deficiency and hyperactive insulin/IGF-1 signaling. Deletion of PHB2 in podocytes of mice, terminally differentiated cells at the kidney filtration barrier, caused progressive proteinuria, kidney failure, and death of the animals and resulted in hyperphosphorylation of S6 ribosomal protein (S6RP), a known mediator of the mTOR signaling pathway. Inhibition of the insulin/IGF-1 signaling system through genetic deletion of the insulin receptor alone or in combination with the IGF-1 receptor or treatment with rapamycin prevented hyperphosphorylation of S6RP without affecting the mitochondrial structural defect, alleviated renal disease, and delayed the onset of kidney failure in PHB2-deficient animals. Evidently, perturbation of insulin/IGF-1 receptor signaling contributes to tissue damage in mitochondrial disease, which may allow therapeutic intervention against a wide spectrum of diseases. © 2015 The Authors. Published under the terms of the CC BY 4.0 license.

  1. Mental health consequences of international migration for Vietnamese Americans and the mediating effects of physical health and social networks: results from a natural experiment approach.

    Science.gov (United States)

    Fu, Hongyun; VanLandingham, Mark J

    2012-05-01

    Although the existing literature on immigrant mental health is extensive, major substantive and methodological gaps remain. Substantively, there is little population-based research that focuses on the mental health consequences of migration for Vietnamese Americans. More generally, although a wide range of mental health problems among immigrants has been identified, the potential causal or mediating mechanisms underlying these problems remain elusive. This latter substantive shortcoming is related to a key methodological challenge involving the potentially confounding effects of selection on migration-related outcomes. This article addresses these challenges by employing a "natural experiment" design, involving comparisons among three population-based samples of Vietnamese immigrants, never-leavers, and returnees (N=709). Data were collected in Ho Chi Minh City and in New Orleans between 2003 and 2005. The study investigates the long-term impact of international migration on Vietnamese mental health, and the potential mediating effects of social networks and physical health on these migration-related outcomes. The results reveal both mental health advantages and disadvantages among Vietnamese immigrants relative to the two groups of Vietnamese nationals. Selection can be ruled out for some of these differences, and both social networks and physical health are found to play important explanatory roles.

  2. Hepatitis B virus X protein promotes interleukin-7 receptor expression via NF-κB and Notch1 pathway to facilitate proliferation and migration of hepatitis B virus-related hepatoma cells

    Directory of Open Access Journals (Sweden)

    Fanyun Kong

    2016-11-01

    Full Text Available Abstract Background Interleukin-7 receptor (IL-7R is involved in the abnormal function of solid tumors, but the role and regulatory mechanisms of IL-7R in HBV-related hepatocellular carcinoma (HCC are still unclear. Methods Gene and protein expression levels of IL-7R were examined in hepatoma cells transfected with hepatitis B virus (HBV plasmids and in hepatoma cells transfected with the multifunctional nonstructural protein X (HBX. The expression of HBX and IL-7R was measured by immunohistochemical analysis in HBV-related HCC tissues. The role of NF-κB and Notch1 pathways in HBX-mediated expression of IL-7R in hepatoma cells was examined. Activation of IL-7R downstream of intracellular signaling proteins AKT, JNK, STAT5, and the associated molecules CyclinD1 and matrix metalloproteinase-9 (MMP-9, was assessed in HBX-positive cells with or without treatment with IL-7R short hairpin RNA (shRNA. Additionally, the role of IL-7R in HBX-mediated proliferation and migration of hepatoma cells was investigated. Results The expression of IL-7R was increased in hepatoma cells transfected with HBV plasmids; HBX was responsible for the HBV-mediated upregulation of IL-7R. Compared to adjacent tissues, the expression of HBX and IL-7R was increased in HBV-related HCC tissues. Additionally, the relative expression levels of HBX were associated with IL-7R in HBV-related HCC tissues. The activation of NF-κB pathways and expression of Notch1 were increased in hepatoma cells transfected with HBX, and inhibition of NF-κB and Notch1 pathways significantly decreased HBX-mediated expression of IL-7R. The activation of AKT and JNK and the expression of CyclinD1 and MMP-9 were increased in HBX-positive cells. When cells were treated with IL-7R shRNA, the activation of AKT and JNK, as well as the expression of CyclinD1 and MMP-9, were significantly inhibited. Additionally, IL-7R was responsible for HBX-induced proliferation and migration ability of hepatoma cells

  3. Enhanced nicotinic acetylcholine receptor-mediated [3H]norepinephrine release from neonatal rat hypothalamus.

    Science.gov (United States)

    O'Leary, K T; Leslie, F M

    2006-01-01

    Nicotinic acetylcholine receptor (nAChR)-evoked release of norepinephrine (NE) has been demonstrated in a number of brain regions that receive sole noradrenergic innervation from the locus coeruleus (LC). Many of these structures display enhanced nicotine-stimulated NE release in the neonate. We have examined the hypothalamus in order to determine if this region, which receives NE projections from both the LC and medullary catecholaminergic nuclei, also demonstrates maturational changes in nAChR-mediated NE release. Quantification of radiolabeled-NE release from rat hypothalamus slices by a maximally effective dose of nicotine revealed a peak response during the first postnatal week. This was followed by a decrease at postnatal day (P) 14, and a second peak at P21. Thereafter, release was equivalent to that observed at P14. Comparison of the pharmacological properties of nAChRs mediating NE release in neonatal (P7) and mature hypothalamus suggested involvement of different nAChR subtypes at the two ages. Using the selective toxin, DSP-4, nAChR-mediated NE release in the neonatal hypothalamus was shown to be from LC terminals. Our findings demonstrate an early sensitivity of hypothalamic LC terminals to nAChR regulation that may be associated with development of systems controlling critical homeostatic functions such as stress, feeding and cardiovascular regulation.

  4. The arginine vasopressin V1b receptor gene and prosociality: Mediation role of emotional empathy.

    Science.gov (United States)

    Wu, Nan; Shang, Siyuan; Su, Yanjie

    2015-09-01

    The vasopressin V1b receptor (AVPR1B) gene has been shown to be closely associated with bipolar disorder and depression. However, whether it relates to positive social outcomes, such as empathy and prosocial behavior, remains unknown. This study explored the possible role of the AVPR1B gene rs28373064 in empathy and prosociality. A total of 256 men, who were genetically unrelated, non-clinical ethnic Han Chinese college students, participated in the study. Prosociality was tested by measuring the prosocial tendencies of cognitive and emotional empathy using the Interpersonal Reactivity Index (IRI). The single nucleotide polymorphism (SNP), rs28373064, was genotyped using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. The results suggest that the AVPR1B gene rs28373064 is linked to emotional empathy and prosociality. The mediation analysis indicated that the effect of the AVPR1B gene on prosociality might be mediated by emotional empathy. This study demonstrated the link between the AVPR1B gene and prosociality and provided evidence that emotional empathy might mediate the relation between the AVPR1B gene and prosociality. © 2015 The Institute of Psychology, Chinese Academy of Sciences and Wiley Publishing Asia Pty Ltd.

  5. Vacuolar Sorting Receptor-Mediated Trafficking of Soluble Vacuolar Proteins in Plant Cells

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    Hyangju Kang

    2014-08-01

    Full Text Available Vacuoles are one of the most prominent organelles in plant cells, and they play various important roles, such as degradation of waste materials, storage of ions and metabolites, and maintaining turgor. During the past two decades, numerous advances have been made in understanding how proteins are specifically delivered to the vacuole. One of the most crucial steps in this process is specific sorting of soluble vacuolar proteins. Vacuolar sorting receptors (VSRs, which are type I membrane proteins, are involved in the sorting and packaging of soluble vacuolar proteins into transport vesicles with the help of various accessory proteins. To date, large amounts of data have led to the development of two different models describing VSR-mediated vacuolar trafficking that are radically different in multiple ways, particularly regarding the location of cargo binding to, and release from, the VSR and the types of carriers utilized. In this review, we summarize current literature aimed at elucidating VSR-mediated vacuolar trafficking and compare the two models with respect to the sorting signals of vacuolar proteins, as well as the molecular machinery involved in VSR-mediated vacuolar trafficking and its action mechanisms.

  6. Loss of progesterone receptor-mediated actions induce preterm cellular and structural remodeling of the cervix and premature birth.

    Science.gov (United States)

    Yellon, Steven M; Dobyns, Abigail E; Beck, Hailey L; Kurtzman, James T; Garfield, Robert E; Kirby, Michael A

    2013-01-01

    A decline in serum progesterone or antagonism of progesterone receptor function results in preterm labor and birth. Whether characteristics of premature remodeling of the cervix after antiprogestins or ovariectomy are similar to that at term was the focus of the present study. Groups of pregnant rats were treated with vehicle, a progesterone receptor antagonist (onapristone or mifepristone), or ovariectomized on day 17 postbreeding. As expected, controls given vehicle delivered at term while rats delivered preterm after progesterone receptor antagonist treatment or ovariectomy. Similar to the cervix before term, the preterm cervix of progesterone receptor antagonist-treated rats was characterized by reduced cell nuclei density, decreased collagen content and structure, as well as a greater presence of macrophages per unit area. Thus, loss of nuclear progesterone receptor-mediated actions promoted structural remodeling of the cervix, increased census of resident macrophages, and preterm birth much like that found in the cervix at term. In contrast to the progesterone receptor antagonist-induced advance in characteristics associated with remodeling, ovariectomy-induced loss of systemic progesterone did not affect hypertrophy, extracellular collagen, or macrophage numbers in the cervix. Thus, the structure and macrophage census in the cervix appear sufficient for premature ripening and birth to occur well before term. With progesterone receptors predominantly localized on cells other than macrophages, the findings suggest that interactions between cells may facilitate the loss of progesterone receptor-mediated actions as part of a final common mechanism that remodels the cervix in certain etiologies of preterm and with parturition at term.

  7. Crystal structure of the urokinase receptor in a ligand-free form

    DEFF Research Database (Denmark)

    Xu, Xiang; Gårdsvoll, Henrik; Yuan, Cai

    2012-01-01

    The urokinase receptor urokinase-type plasminogen activator receptor (uPAR) is a surface receptor capable of not only focalizing urokinase-type plasminogen activator (uPA)-mediated fibrinolysis to the pericellular micro-environment but also promoting cell migration and chemotaxis. Consistent with...

  8. MDMA-Induced Dissociative State not Mediated by the 5-HT2A Receptor

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    Drew J. Puxty

    2017-07-01

    Full Text Available Previous research has shown that a single dose of MDMA induce a dissociative state, by elevating feelings of depersonalization and derealization. Typically, it is assumed that action on the 5-HT2A receptor is the mechanism underlying these psychedelic experiences. In addition, other studies have shown associations between dissociative states and biological parameters (heart rate, cortisol, which are elevated by MDMA. In order to investigate the role of the 5-HT2 receptor in the MDMA-induced dissociative state and the association with biological parameters, a placebo-controlled within-subject study was conducted including a single oral dose of MDMA (75 mg, combined with placebo or a single oral dose of the 5-HT2 receptor blocker ketanserin (40 mg. Twenty healthy recreational MDMA users filled out a dissociative states scale (CADSS 90 min after treatments, which was preceded and followed by assessment of a number of biological parameters (cortisol levels, heart rate, MDMA blood concentrations. Findings showed that MDMA induced a dissociative state but this effect was not counteracted by pre-treatment with ketanserin. Heart rate was the only biological parameter that correlated with the MDMA-induced dissociative state, but an absence of correlation between these measures when participants were pretreated with ketanserin suggests an absence of directional effects of heart rate on dissociative state. It is suggested that the 5-HT2 receptor does not mediate the dissociative effects caused by a single dose of MDMA. Further research is needed to determine the exact neurobiology underlying this effect and whether these effects contribute to the therapeutic potential of MDMA.

  9. Bradykinin and adenosine receptors mediate desflurane induced postconditioning in human myocardium: role of reactive oxygen species

    Directory of Open Access Journals (Sweden)

    Gérard Jean-Louis

    2010-07-01

    Full Text Available Abstract Background Desflurane during early reperfusion has been shown to postcondition human myocardium, in vitro. We investigated the role of adenosine and bradykinin receptors, and generation of radical oxygen species in desflurane-induced postconditioning in human myocardium. Methods We recorded isometric contraction of human right atrial trabeculae hanged in an oxygenated Tyrode's solution (34 degrees Celsius, stimulation frequency 1 Hz. After a 30-min hypoxic period, desflurane 6% was administered during the first 5 min of reoxygenation. Desflurane was administered alone or with pretreatment of N-mercaptopropionylglycine, a reactive oxygen species scavenger, 8-(p-Sulfophenyltheophylline, an adenosine receptor antagonist, HOE140, a selective B2 bradykinin receptor antagonist. In separate groups, adenosine and bradykinin were administered during the first minutes of reoxygenation alone or in presence of N-mercaptopropionylglycine. The force of contraction of trabeculae was recorded continuously. Developed force at the end of a 60-min reoxygenation period was compared (mean ± standard deviation between the groups by a variance analysis and post hoc test. Results Desflurane 6% (84 ± 6% of baseline enhanced the recovery of force after 60-min of reoxygenation as compared to control group (51 ± 8% of baseline, P N-mercaptopropionylglycine (54 ± 3% of baseline, 8-(p-Sulfophenyltheophylline (62 ± 9% of baseline, HOE140 (58 ± 6% of baseline abolished desflurane-induced postconditioning. Adenosine (80 ± 9% of baseline and bradykinin (83 ± 4% of baseline induced postconditioning (P vs control, N-mercaptopropionylglycine abolished the beneficial effects of adenosine and bradykinin (54 ± 8 and 58 ± 5% of baseline, respectively. Conclusions In vitro, desflurane-induced postconditioning depends on reactive oxygen species production, activation of adenosine and bradykinin B2 receptors. And, the cardioprotective effect of adenosine and bradykinin

  10. Schwann Cell Migration Induced by Earthworm Extract via Activation of PAs and MMP2/9 Mediated through ERK1/2 and p38

    Science.gov (United States)

    Chang, Yung-Ming; Shih, Ying-Ting; Chen, Yueh-Sheng; Liu, Chien-Liang; Fang, Wen-Kuei; Tsai, Chang-Hai; Tsai, Fuu-Jen; Kuo, Wei-Wen; Lai, Tung-Yuan; Huang, Chih-Yang

    2011-01-01

    The earthworm, which has stasis removal and wound-healing functions, is a widely used Chinese herbal medicine in China. Schwann cell migration is critical for the regeneration of injured nerves. Schwann cells provide an essentially supportive activity for neuron regeneration. However, the molecular migration mechanisms induced by earthworms in Schwann cells remain unclear. Here, we investigate the roles of MAPK (ERK1/2, JNK and p38) pathways for earthworm-induced matrix-degrading proteolytic enzyme (PAs and MMP2/9) production in Schwann cells. Moreover, earthworm induced phosphorylation of ERK1/2 and p38, but not JNK, activate the downstream signaling expression of PAs and MMPs in a time-dependent manner. Earthworm-stimulated ERK1/2 and p38 phosphorylation was attenuated by pretreatment with U0126 and SB203580, resulting in migration and uPA-related signal pathway inhibition. The results were confirmed using small interfering ERK1/2 and p38 RNA. These results demonstrated that earthworms can stimulate Schwann cell migration and up-regulate PAs and MMP2/9 expression mediated through the MAPK pathways, ERK1/2 and p38. Taken together, our data suggests the MAPKs (ERK1/2, p38)-, PAs (uPA, tPA)-, MMP (MMP2, MMP9) signaling pathway of Schwann cells regulated by earthworms might play a major role in Schwann cell migration and nerve regeneration. PMID:19808845

  11. Distinctive Modulation of Dopamine Release in the Nucleus Accumbens Shell Mediated by Dopamine and Acetylcholine Receptors.

    Science.gov (United States)

    Shin, Jung Hoon; Adrover, Martin F; Alvarez, Veronica A

    2017-11-15

    Nucleus accumbens (NAc) shell shows unique dopamine (DA) signals in vivo and plays a unique role in DA-dependent behaviors such as reward-motivated learning and the response to drugs of abuse. A disynaptic mechanism for DA release was reported and shown to require synchronized firing of cholinergic interneurons (CINs) and activation of nicotinic acetylcholine (ACh) receptors (nAChRs) in DA neuron (DAN) axons. The properties of this disynaptic mechanism of DA transmission are not well understood in the NAc shell. In this study, in vitro fast-scan cyclic voltammetry was used to examine the modulation of DA transmission evoked by CINs firing in the shell of mice and compared with other striatal regions. We found that DA signals in the shell displayed significant degree of summation in response to train stimulation of CINs, contrary to core and dorsal striatum. The summation was amplified by a D2-like receptor antagonist and experiments with mice with targeted deletion of D2 receptors to DANs or CINs revealed that D2 receptors in CINs mediate a fast inhibition observed within 100 ms of the first pulse, whereas D2 autoreceptors in DAN terminals are engaged in a slower inhibition that peaks at ∼500 ms. ACh also contributes to the use-dependent inhibition of DA release through muscarinic receptors only in the shell, where higher activity of acetylcholinesterase minimizes nAChR desensitization and promotes summation. These findings show that DA signals are modulated differentially by endogenous DA and ACh in the shell, which may underlie the unique features of shell DA signals in vivoSIGNIFICANCE STATEMENT The present study reports that dopamine (DA) release evoked by activation of cholinergic interneurons displays a high degree of summation in the shell and shows unique modulation by endogenous DA and acetylcholine. Desensitization of nicotinic receptors, which is a prevailing mechanism for use-dependent inhibition in the nucleus accumbens core and dorsal striatum, is

  12. The BH3-only protein bid does not mediate death-receptor-induced liver injury in obstructive cholestasis.

    Science.gov (United States)

    Nalapareddy, Padmavathi devi; Schüngel, Sven; Hong, Ji-Young; Manns, Michael P; Jaeschke, Hartmut; Vogel, Arndt

    2009-09-01

    The accumulation of bile acids during obstructive cholestasis causes liver injury and fibrosis, which is at least partly mediated by the death receptors Tumor necrosis factor-related apoptosis-inducing ligand, Tumor necrosis factor-alpha, and Fas. The BH3-interacting domain death agonist Bid is a critical mediator of death receptor-induced apoptosis in hepatocytes. Our aim for this study was, therefore, to elucidate whether Bid also mediates death receptor-induced liver injury in obstructive cholestasis. Overall, survival and various aspects of liver injury were analyzed in wild-type and Bid(-/-) mice after bile duct ligation (BDL), a commonly used model to study obstructive cholestasis in mice. Liver injury was examined at 3, 7, and 14 days after BDL. Loss of Bid did not affect the number of bile infarcts, serum aspartate aminotransferase values, or animal survival. Processing of procaspase-3 and procaspase-9, and caspase-3 enzyme activities, were not detectable in either group, and Bid(-/-) mice displayed the same pattern of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling positive hepatocytes as wild-type controls following BDL. In contrast to Fas-receptor deficient lpr mice, hepatic fibrosis and the inflammatory response was not affected by loss of Bid. Together, these data suggest that Bid is not a downstream target of death receptors in obstructive cholestasis and does not significantly contribute to bile acid induced liver injury and fibrosis.

  13. Posttranslational Modification of HOIP Blocks Toll-Like Receptor 4-Mediated Linear-Ubiquitin-Chain Formation.

    Science.gov (United States)

    Bowman, James; Rodgers, Mary A; Shi, Mude; Amatya, Rina; Hostager, Bruce; Iwai, Kazuhiro; Gao, Shou-Jiang; Jung, Jae U

    2015-11-17

    Linear ubiquitination is an atypical posttranslational modification catalyzed by the linear-ubiquitin-chain assembly complex (LUBAC), containing HOIP, HOIL-1L, and Sharpin. LUBAC facilitates NF-κB activation and inflammation upon receptor stimulation by ligating linear ubiquitin chains to critical signaling molecules. Indeed, linear-ubiquitination-dependent signaling is essential to prevent pyogenic bacterial infections that can lead to death. While linear ubiquitination is essential for intracellular receptor signaling upon microbial infection, this response must be measured and stopped to avoid tissue damage and autoimmunity. While LUBAC is activated upon bacterial stimulation, the mechanisms regulating LUBAC activity in response to bacterial stimuli have remained elusive. We demonstrate that LUBAC activity itself is downregulated through ubiquitination, specifically, ubiquitination of the catalytic subunit HOIP at the carboxyl-terminal lysine 1056. Ubiquitination of Lys1056 dynamically altered HOIP conformation, resulting in the suppression of its catalytic activity. Consequently, HOIP Lys1056-to-Arg mutation led not only to persistent LUBAC activity but also to prolonged NF-κB activation induced by bacterial lipopolysaccharide-mediated Toll-like receptor 4 (TLR4) stimulation, whereas it showed no effect on NF-κB activation induced by CD40 stimulation. This study describes a novel posttranslational regulation of LUBAC-mediated linear ubiquitination that is critical for specifically directing TLR4-mediated NF-κB activation. Posttranslational modification of proteins enables cells to respond quickly to infections and immune stimuli in a tightly controlled manner. Specifically, covalent modification of proteins with the small protein ubiquitin is essential for cells to initiate and terminate immune signaling in response to bacterial and viral infection. This process is controlled by ubiquitin ligase enzymes, which themselves must be regulated to prevent

  14. Presynaptic CRF1 Receptors Mediate the Ethanol Enhancement of GABAergic Transmission in the Mouse Central Amygdala

    Directory of Open Access Journals (Sweden)

    Zhiguo Nie

    2009-01-01

    Full Text Available Corticotropin-releasing factor (CRF is a 41-amino-acid neuropeptide involved in stress responses initiated from several brain areas, including the amygdala formation. Research shows a strong relationship between stress, brain CRF, and excessive alcohol consumption. Behavioral studies suggest that the central amygdala (CeA is significantly involved in alcohol reward and dependence. We recently reported that the ethanol augmentation of GABAergic synaptic transmission in rat CeA involves CRF1 receptors, because both CRF and ethanol significantly enhanced the amplitude of evoked GABAergic inhibitory postsynaptic currents (IPSCs in CeA neurons from wild-type (WT and CRF2 knockout (KO mice, but not in neurons of CRF1 KO mice. The present study extends these findings using selective CRF receptor ligands, gene KO models, and miniature IPSC (mIPSC analysis to assess further a presynaptic role for the CRF receptors in mediating ethanol effects in the CeA. In whole-cell patch recordings of pharmacologically isolated GABAAergic IPSCs from slices of mouse CeA, both CRF and ethanol augmented evoked IPSCs in a concentration-dependent manner, with low EC50s. A CRF1 (but not CRF2 KO construct and the CRF1-selective nonpeptide antagonist NIH-3 (LWH-63 blocked the augmenting effect of both CRF and ethanol on evoked IPSCs. Furthermore, the new selective CRF1 agonist stressin1, but not the CRF2 agonist urocortin 3, also increased evoked IPSC amplitudes. Both CRF and ethanol decreased paired-pulse facilitation (PPF of evoked IPSCs and significantly enhanced the frequency, but not the amplitude, of spontaneous miniature GABAergic mIPSCs in CeA neurons of WT mice, suggesting a presynaptic site of action. The PPF effect of ethanol was abolished in CeA neurons of CRF1 KO mice. The CRF1 antagonist NIH-3 blocked the CRF- and ethanol-induced enhancement of mIPSC frequency in CeA neurons. These data indicate that presynaptic CRF1 receptors play a critical role in permitting

  15. Enhancement of D1 dopamine receptor-mediated locomotor stimulation in M4 muscarinic acetylcholine receptor knockout mice

    OpenAIRE

    Gomeza, Jesus; Zhang, Lu; Kostenis, Evi; Felder, Christian; Bymaster, Frank; Brodkin, Jesse; Shannon, Harlan; Xia, Bing; Deng, Chu-Xia; Wess, Jürgen

    1999-01-01

    Muscarinic acetylcholine receptors (M1–M5) regulate many key functions of the central and peripheral nervous system. Primarily because of the lack of receptor subtype-selective ligands, the precise physiological roles of the individual muscarinic receptor subtypes remain to be elucidated. Interestingly, the M4 receptor subtype is expressed abundantly in the striatum and various other forebrain regions. To study its potential role in the regulation of locomotor activity and other central funct...

  16. Endothelin potentiates TRPV1 via ETA receptor-mediated activation of protein kinase C

    Directory of Open Access Journals (Sweden)

    Furkert Jens

    2007-11-01

    Full Text Available Abstract Background Endothelin-1 (ET-1 both stimulates nociceptors and sensitizes them to noxious stimuli, an effect probably mediated by the ETA receptor (ETAR expressed in sensory neurons. The cellular mechanisms of this ET-1-mediated effect are only poorly understood. TRPV1, the heat-, pH- and capsaicin-sensitive cation channel already known to be modulated by a number of cellular mediators released in response to noxious stimuli and during inflammation, is a potential target for the action of ET-1. Results We studied the effects of ET-1 on TRPV1 in sensory neurons from the dorsal root ganglion (DRG and in HEK293 cells coexpressing TRPV1 and the ETAR. Specific 125I-ET-1 binding sites (817 ± 92 fmol/mg were detected in membrane preparations of DRG with an ETAR/ETBR ratio of 60:40. In an immunofluorescence analysis, coexpression of TRPV1 and the ETAR was found in a subpopulation of primary sensory neurons. ET-1 strongly potentiated capsaicin-induced TRPV1 currents in some neurons, and in HEK293 cells co-expressing TRPV1 and the ETAR. Weaker potentiation was observed in HEK293 cells coexpressing TRPV1 and the ETBR. ETAR activation also increased responses to low pH and heat. In HEK293 cells, strong potentiation of TRPV1 like that induced by ET-1 via the ETAR could be induced by PKC activation, but not with activators of the adenylyl cyclase or the PKA pathway. Furthermore, inhibition of PKC with bisindolylmaleimide X (BIM X or mutation of the PKC phosphorylation site S800 completely prevented ETAR-mediated potentiation. Conclusion We conclude that ET-1 potentiates TRPV1 by a PKC-dependent mechanism and that this could play a major role in the algogenic and hyperalgesic effects of ET-1 described in previous studies.

  17. Induction of retinoic acid receptor β mediates growth inhibition in retinoid resistant human colon carcinoma cells

    OpenAIRE

    Nicke, B; Riecken, E; Rosewicz, S

    1999-01-01

    BACKGROUND—The molecular mechanisms underlying the differential sensitivity of human colon carcinoma cells to retinoid mediated growth inhibition are poorly understood.
AIM—To identify the intracellular mechanisms responsible for resistance against retinoid mediated growth inhibition in human colon carcinoma cells.
METHODS—Anchorage independent growth of the human colon carcinoma cell lines HT29 and LoVo was determined by a human tumour clonogenic assay. Retinoid receptor expression was evalu...

  18. Tamoxifen resistance and metastasis of human breast cancer cells were mediated by the membrane-associated estrogen receptor ER-α36 signaling in vitro.

    Science.gov (United States)

    Gu, Wenwen; Dong, Nian; Wang, Peng; Shi, Changgen; Yang, Jun; Wang, Jian

    2017-04-01

    The drug resistance and tumor metastasis have been the main obstacles for the longer-term therapeutic effects of tamoxifen (TAM) on estrogen receptor-positive (ER+) breast cancer, but the mechanisms underlying the TAM resistance are still unclear. Here, we demonstrated that the membrane-associated estrogen receptor ER-α36 signaling, but not the G protein-coupled estrogen receptor 1 (GPER1) signaling, might be involved in the TAM resistance and metastasis of breast cancer cells. In this study, a model of ER+ breast cancer cell MCF-7 that involves the up-regulated expression of ER-α36 and unchanged expression of ER-α66 and GPER1 was established via the removal of insulin from the cell culture medium. The mechanism of TAM resistance in the ER+ breast cancer cell line MCF-7 was investigated, and the results showed that the stimulating effect of insulin on susceptibility of MCF-7 to TAM was mediated by ER-α36 and that the expression level of ER-α36 in TAM-resistant MCF-7 cells was also significantly increased. Both TAM and estradiol (E2) could promote the migration of triple negative (ER-α66-/PR-/HER2-) and ER-α36+/GPER1+ breast cancer cells MDA-MB-231. The migration of MDA-MB-231 cells was inhibited by the down-regulated intracellular expression of ER-α36 by transient transfection of specific small interfering RNA, whereas no effect of GPER1 down-regulation was observed. Meanwhile, the effect of TAM on the migration of ER-α36-down-regulated MDA-MB-231 cells was also reduced. Furthermore, it was found that TAM enhanced the distribution of integrin β1 on the cell surface but did not affect the expression of integrin β1 in MDA-MB-231 cells. Collectively, these data suggested that ER-α36 signaling might play critical roles in acquired and de novo TAM resistance and metastasis of breast cancer, and ER-α36 might present a potential biomarker of TAM resistance in the clinical diagnosis and treatment of ER+ breast cancer.

  19. Enzyme induction and histopathology elucidate aryl hydrocarbon receptor-mediated versus non-aryl hydrocarbon receptor-mediated effects of Aroclor 1268 in American mink (Neovison vison).

    Science.gov (United States)

    Folland, William R; Newsted, John L; Fitzgerald, Scott D; Fuchsman, Phyllis C; Bradley, Patrick W; Kern, John; Kannan, Kurunthachalam; Zwiernik, Matthew J

    2016-03-01

    Polychlorinated biphenyl (PCB) concentrations reported in preferred prey and blubber of bottlenose dolphins from the Turtle-Brunswick River estuary (Georgia, USA) suggest the potential for adverse effects. However, PCBs in Turtle-Brunswick River estuary dolphins are primarily derived from Aroclor 1268, and predicting toxic effects of Aroclor 1268 is uncertain because of the mixture's unique composition and associated physiochemical characteristics. These differences suggest that toxicity benchmarks for other PCB mixtures may not be relevant to dolphins exposed to Aroclor 1268. American mink (Neovison vison) were used as a surrogate model for cetaceans to characterize mechanisms of action associated with Aroclor 1268 exposure. Mink share similarities in phylogeny and life history with cetaceans and are characteristically sensitive to PCBs, making them an attractive surrogate species for marine mammals in ecotoxicity studies. Adult female mink and a subsequent F1 generation were exposed to Aroclor 1268 through diet, and effects on enzyme induction, histopathology, thyroid hormone regulation, hematology, organ weights, and body condition index were compared to a negative control and a 3,3',4,4',5-pentachlorobiphenyl (PCB 126)-positive control. Aroclor 1268 dietary exposure concentrations ranged from 1.8 µg/g wet weight to 29 µg/g wet weight. Anemia, hypothyroidism, and hepatomegaly were observed in mink exposed to Aroclor 1268 beyond various dietary thresholds. Cytochrome P450 induction and squamous epithelial proliferation jaw lesions were low in Aroclor 1268 treatments relative to the positive control. Differences in enzyme induction and the development of squamous epithelial proliferation jaw lesions between Aroclor 1268 treatments and the positive control, coupled with effects observed in Aroclor 1268 treatments not observed in the positive control, indicate that mechanisms additional to the aryl hydrocarbon receptor-mediated pathway are associated with

  20. Astrocytes Regulate GLP-1 Receptor-Mediated Effects on Energy Balance.

    Science.gov (United States)

    Reiner, David J; Mietlicki-Baase, Elizabeth G; McGrath, Lauren E; Zimmer, Derek J; Bence, Kendra K; Sousa, Gregory L; Konanur, Vaibhav R; Krawczyk, Joanna; Burk, David H; Kanoski, Scott E; Hermann, Gerlinda E; Rogers, Richard C; Hayes, Matthew R

    2016-03-23

    Astrocytes are well established modulators of extracellular glutamate, but their direct influence on energy balance-relevant behaviors is largely understudied. As the anorectic effects of glucagon-like peptide-1 receptor (GLP-1R) agonists are partly mediated by central modulation of glutamatergic signaling, we tested the hypothesis that astrocytic GLP-1R signaling regulates energy balance in rats. Central or peripheral administration of a fluorophore-labeled GLP-1R agonist, exendin-4, localizes within astrocytes and neurons in the nucleus tractus solitarius (NTS), a hindbrain nucleus critical for energy balance control. This effect is mediated by GLP-1R, as the uptake of systemically administered fluorophore-tagged exendin-4 was blocked by central pretreatment with the competitive GLP-1R antagonist exendin-(9-39). Ex vivo analyses show prolonged exendin-4-induced activation (live cell calcium signaling) of NTS astrocytes and neurons; these effects are also attenuated by exendin-(9-39), indicating mediation by the GLP-1R. In vitro analyses show that the application of GLP-1R agonists increases cAMP levels in astrocytes. Immunohistochemical analyses reveal that endogenous GLP-1 axons form close synaptic apposition with NTS astrocytes. Finally, pharmacological inhibition of NTS astrocytes attenuates the anorectic and body weight-suppressive effects of intra-NTS GLP-1R activation. Collectively, data demonstrate a role for NTS astrocytic GLP-1R signaling in energy balance control. Glucagon-like peptide-1 receptor (GLP-1R) agonists reduce food intake and are approved by the Food and Drug Administration for the treatment of obesity, but the cellular mechanisms underlying the anorectic effects of GLP-1 require further investigation. Astrocytes represent a major cellular population in the CNS that regulates neurotransmission, yet the role of astrocytes in mediating energy balance is largely unstudied. The current data provide novel evidence that astrocytes within the NTS

  1. Inhibition of non-small cell lung cancer cell migration by grape seed proanthocyanidins is mediated through the inhibition of nitric oxide, guanylate cyclase, and ERK1/2.

    Science.gov (United States)

    Punathil, Thejass; Katiyar, Santosh K

    2009-03-01

    Tumor cell migration is considered as a major event in the metastatic cascade. Here we examined the effect of grape seed proanthocyanidins (GSPs) on migration capacity and signaling mechanisms using nonsmall cell human lung cancer cells. Using in vitro migration assay, we found that treatment of A549 and H1299 cells with GSPs resulted in concentration-dependent inhibition of migration of these cells. The migration capacity of cells was reduced in presence of N(G)-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase. GSPs suppressed the elevated levels of endogenous NO/NOS in A549 and H1299 cells and blocked the migration promoting capacity of L-arginine. Treatment with guanylate cyclase (GC) inhibitor 1-H-[1,2,4]oxadiaxolo[4,3-a]quinolalin-1-one (ODQ) reduced the migration of A549 cells whereas additional presence of 8-bromoguanosine 3'5'-cyclic monophosphate (8-Br-cGMP, cGMP analogue) restored the migration of these cells, suggesting a role for GC in migration of A549 cells. GSPs reduced the elevated levels of cGMP in cancer cells and also blocked the migration restoring activity of 8-Br-cGMP. The mitogen-activated protein kinase kinase (MAPKK) inhibitor, UO126, inhibited the migration of A549 cells, indicating a role for MAPKK in the migration. Additionally, UO126 and ODQ inhibited the migration restoring effects of L-arginine in L-NAME-treated cells, suggesting the involvement of cGMP and MAPK pathways in NO-mediated migration. GSPs inhibited L-arginine and 8-Br-cGMP-induced activation of ERK1/2 in A549 cells. Together, these results indicate sequential inhibition of NO/NOS, GC, and MAPK pathways by GSPs in mediating the inhibitory signals for cell migration, an essential step in invasion and metastasis. (c) 2008 Wiley-Liss, Inc.

  2. Sphingosine-1-phosphate Receptor Agonism Reduces Bordetella pertussis–mediated Lung Pathology

    Science.gov (United States)

    Skerry, Ciaran; Scanlon, Karen; Rosen, Hugh; Carbonetti, Nicholas H.

    2015-01-01

    Recent pertussis resurgence represents a major public health concern. Currently, there are no effective treatments for critical pertussis in infants. Recent data have demonstrated the potential of sphingosine-1-phosphate receptor (S1PR) agonism in the treatment of infectious diseases. We used the murine Bordetella pertussis model to test the hypothesis that treatment with S1PR agonist AAL-R reduces pulmonary inflammation during infection. AAL-R treatment resulted in reduced expression of inflammatory cytokines and chemokines and attenuated lung pathology in infected mice. These results demonstrate a role for sphingosine-1-phosphate (S1P) signaling in B. pertussis–mediated pathology and highlight the possibility of host-targeted therapy for pertussis. PMID:25538274

  3. PEG-coated gold nanoparticles attenuate β-adrenergic receptor-mediated cardiac hypertrophy.

    Science.gov (United States)

    Qiao, Yuhui; Zhu, Baoling; Tian, Aiju; Li, Zijian

    2017-01-01

    Gold nanoparticles (AuNPs) are widely used as a drug delivery vehicle, which can accumulate in the heart through blood circulation. Therefore, it is very important to understand the effect of AuNPs on the heart, especially under pathological conditions. In this study, we found that PEG-coated AuNPs attenuate β-adrenergic receptor (β-AR)-mediated acute cardiac hypertrophy and inflammation. However, both isoproterenol, a non-selective β-AR agonist, and AuNPs did not induce cardiac function change or cardiac fibrosis. AuNPs exerted an anti-cardiac hypertrophy effect by decreasing β 1 -AR expression and its downstream ERK1/2 hypertrophic pathway. Our results indicated that AuNPs might be safe and have the potential to be used as multi-functional materials (drug carrier systems and anti-cardiac hypertrophy agents).

  4. Toll-Like Receptor-3 Mediates HIV-1-Induced Interleukin-6 Expression in the Human Brain Endothelium via TAK1 and JNK Pathways: Implications for Viral Neuropathogenesis.

    Science.gov (United States)

    Bhargavan, Biju; Kanmogne, Georgette D

    2017-11-11

    HIV-1-associated neurocognitive disorders (HAND) is associated with blood-brain-barrier (BBB) inflammation, and inflammation involves toll-like receptors (TLRs) signaling. It is not known whether primary human brain microvascular endothelial cells (HBMEC), the major BBB component, express TLRs or whether TLRs are involved in BBB dysfunction and HAND. We demonstrate that HBMEC express TLR3, 4, 5, 7, 9, and 10, and TLR3 was the most abundant. HIV-1 and TLR3 activation increased endothelial TLR3 transcription and expression. HIV-1-positive human subjects showed significantly higher TLR3 expression in brain tissues and blood vessels, with higher TLR3 levels in subjects with HAND. HIV-1 and TLR3 activation increased endothelial IL6 expression by 6-to-127-fold (P < 0.001), activated c-jun(serine-63) and SAPK/JNK(Thr183/Tyr185). HIV-1 upregulated IL6 through interleukin-1 receptor-associated-kinase (IRAK)-1/4/TAK1/JNK pathways, via ATP-dependent JNK activation. TLR3 activation upregulated IL6 through TAK1/JNK pathways, via ATP-dependent or -independent JNK activation. HIV-1 and TLR3 activation also upregulated transcription factors associated with IL6 and TAK1/JNK pathways (Jun, CEBPA, STAT1). Blocking TLR3 activation prevented HIV-1- and TLR3 ligands-induced upregulation of these transcription factors, prevented IL6 transcription and expression, c-jun and JNK activation. HIV-1 and TLR3 ligands significantly increased monocytes adhesion and migration through the BBB, and decreased endothelial claudin-5 expression. Blocking TLR3 and JNK activation prevented HIV-1- and TLR3 ligands-induced claudin-5 downregulation, monocytes adhesion and transendothelial migration. These data suggest that viral immune recognition via endothelial TLR3 is involved in endothelial inflammation and BBB dysfunction in HIV/AIDS and HAND. Our data provides novel insights into the molecular basis of these HIV-1- and TLR3-mediated effects.

  5. Metabotropic glutamate receptors mediate lipopolysaccharide-induced fever and sickness behavior.

    Science.gov (United States)

    Weiland, Tracey J; Anthony-Harvey-Beavis, Debra; Voudouris, Nicholas J; Kent, Stephen

    2006-05-01

    Several mechanisms have been proposed for neuroimmune communication supporting the sickness syndrome (fever, anorexia, inactivity, and cachexia) following infection. We examined the role of glutamate as a neurochemical intermediary of sickness behavior induced by intraperitoneal lipopolysaccharide (LPS). Mice implanted with biotelemetry devices capable of detecting body temperature (Tb) were administered LPS (50 or 500 microg/kg i.p., serotype 0111:B4) with or without i.p. pretreatment with vehicle or broad-spectrum antagonists selective for N-methyl-d-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic (AMPA)/kainite, or metabotropic glutamate (mGlu) receptors. While NMDA and AMPA/kainate receptor antagonism failed to attenuate LPS-induced sickness behavior, antagonism of metabotropic receptors with l(+)-AP3 reduced the febrile (0-11h: control: 37.32+/-0.16 degrees C, l(+)-AP3: 36.66+/-0.27), anorexic (control: -87+/-5%, l(+)-AP3: 48+/-12% scotophase food intake), and cachexic (control: -8.9+/-0.4%, l(+)-AP3: -6.1+/-1.3% body weight) effects of 500 microg/kg LPS, and produced a biphasic Tb effect in response to 50 microg/kg LPS (1h: -0.90+/-0.26; 6h: 1.78+/-0.35 degrees C relative to baseline). At this dose the Tb of l(+)-AP3-treated mice was 1.18 degrees C lower than controls 2h post-injection, and 0.68 degrees C greater that controls 8h post-injection. These results suggest a role for mGlu receptors in mediating fever, anorexia, and cachexia possibly via activation of extra-vagal pathways, since the attenuating effect of l(+)-AP3 increased with increasing dosages of LPS. Given the critical role ascribed to mGlu receptors in neurotransmitter release and astrocytic processes, it is possible that these observations reflect an l(+)-AP3-induced attenuation of these systems.

  6. The TM2 6′ Position of GABAA Receptors Mediates Alcohol Inhibition

    Science.gov (United States)

    Howard, Rebecca J.; Trudell, James R.; Harris, R. Adron

    2012-01-01

    Ionotropic GABAA receptors (GABAARs), which mediate inhibitory neurotransmission in the central nervous system, are implicated in the behavioral effects of alcohol and alcoholism. Site-directed mutagenesis studies support the presence of discrete molecular sites involved in alcohol enhancement and, more recently, inhibition of GABAARs. We used Xenopus laevis oocytes to investigate the 6′ position in the second transmembrane region of GABAARs as a site influencing alcohol inhibition. We asked whether modification of the 6′ position by substitution with larger residues or methanethiol labeling [using methyl methanethiosulfonate (MMTS)] of a substituted cysteine, reduced GABA action and/or blocked further inhibition by alcohols. Labeling of the 6′ position in either α2 or β2 subunits reduced responses to GABA. In addition, methanol and ethanol potentiation increased after MMTS labeling or substitution with tryptophan or methionine, consistent with elimination of an inhibitory site for these alcohols. Specific alcohols, but not the anesthetic etomidate, competed with MMTS labeling at the 6′ position. We verified a role for the 6′ position in previously tested α2β2 as well as more physiologically relevant α2β2γ2s GABAARs. Finally, we built a novel molecular model based on the invertebrate glutamate-gated chloride channel receptor, a GABAAR homolog, revealing that the 6′ position residue faces the channel pore, and modification of this residue alters volume and polarity of the pore-facing cavity in this region. These results indicate that the 6′ positions in both α2 and β2 GABAAR subunits mediate inhibition by short-chain alcohols, which is consistent with the presence of multiple counteracting sites of action for alcohols on ligand-gated ion channels. PMID:22072732

  7. Cytoskeleton-related regulation of primary cilia shortening mediated by melanin-concentrating hormone receptor 1.

    Science.gov (United States)

    Tomoshige, Sakura; Kobayashi, Yuki; Hosoba, Kosuke; Hamamoto, Akie; Miyamoto, Tatsuo; Saito, Yumiko

    2017-11-01

    Primary cilia are specialized microtubule-based organelles. Their importance is highlighted by the gamut of ciliary diseases associated with various syndromes including diabetes and obesity. Primary cilia serve as signaling hubs through selective interactions with ion channels and conventional G-protein-coupled receptors (GPCRs). Melanin-concentrating hormone (MCH) receptor 1 (MCHR1), a key regulator of feeding, is selectively expressed in neuronal primary cilia in distinct regions of the mouse brain. We previously found that MCH acts on ciliary MCHR1 and induces cilia shortening through a Gi/o-dependent Akt pathway with no cell cycle progression. Many factors can participate in cilia length control. However, the mechanisms for how these molecules are relocated and coordinated to activate cilia shortening are poorly understood. In the present study, we investigated the role of cytoskeletal dynamics in regulating MCH-induced cilia shortening using clonal MCHR1-expressing hTERT-RPE1 cells. Pharmacological and biochemical approaches showed that cilia shortening mediated by MCH was associated with increased soluble cytosolic tubulin without changing the total tubulin amount. Enhanced F-actin fiber intensity was also observed in MCH-treated cells. The actions of various pharmacological agents revealed that coordinated actin machinery, especially actin polymerization, was required for MCHR1-mediated cilia shortening. A recent report indicated the existence of actin-regulated machinery for cilia shortening through GPCR agonist-dependent ectosome release. However, our live-cell imaging experiments showed that MCH progressively elicited cilia shortening without exclusion of fluorescence-positive material from the tip. Short cilia phenotypes have been associated with various metabolic disorders. Thus, the present findings may contribute toward better understanding of how the cytoskeleton is involved in the GPCR ligand-triggered cilia shortening with cell mechanical

  8. Enterococcus faecalis Gelatinase Mediates Intestinal Permeability via Protease-Activated Receptor 2

    Science.gov (United States)

    Maharshak, Nitsan; Huh, Eun Young; Paiboonrungruang, Chorlada; Shanahan, Michael; Thurlow, Lance; Herzog, Jeremy; Djukic, Zorka; Orlando, Roy; Pawlinski, Rafal; Ellermann, Melissa; Borst, Luke; Patel, Siten; Dotan, Iris; Sartor, Ryan B.

    2015-01-01

    Microbial protease-mediated disruption of the intestinal epithelium is a potential mechanism whereby a dysbiotic enteric microbiota can lead to disease. This mechanism was investigated using the colitogenic, protease-secreting enteric microbe Enterococcus faecalis. Caco-2 and T-84 epithelial cell monolayers and the mouse colonic epithelium were exposed to concentrated conditioned media (CCM) from E. faecalis V583 and E. faecalis lacking the gelatinase gene (gelE). The flux of fluorescein isothiocyanate (FITC)-labeled dextran across monolayers or the mouse epithelium following exposure to CCM from parental or mutant E. faecalis strains indicated paracellular permeability. A protease-activated receptor 2 (PAR2) antagonist and PAR2-deficient (PAR2−/−) mice were used to investigate the role of this receptor in E. faecalis-induced permeability. Gelatinase (GelE) purified from E. faecalis V583 was used to confirm the ability of this protease to induce epithelial cell permeability and activate PAR2. The protease-mediated permeability of colonic epithelia from wild-type (WT) and PAR2−/− mice by fecal supernatants from ulcerative colitis patients was assessed. Secreted E. faecalis proteins induced permeability in epithelial cell monolayers, which was reduced in the absence of gelE or by blocking PAR2 activity. Secreted E. faecalis proteins induced permeability in the colonic epithelia of WT mice that was absent in tissues from PAR2−/− mice. Purified GelE confirmed the ability of this protease to induce epithelial cell permeability via PAR2 activation. Fecal supernatants from ulcerative colitis patients induced permeability in the colonic epithelia of WT mice that was reduced in tissues from PAR2−/− mice. Our investigations demonstrate that GelE from E. faecalis can regulate enteric epithelial permeability via PAR2. PMID:25916983

  9. Hepatic ZIP14-mediated Zinc Transport Contributes to Endosomal Insulin Receptor Trafficking and Glucose Metabolism.

    Science.gov (United States)

    Aydemir, Tolunay Beker; Troche, Catalina; Kim, Min-Hyun; Cousins, Robert J

    2016-11-11

    Zinc influences signaling pathways through controlled targeted zinc transport. Zinc transporter Zip14 KO mice display a phenotype that includes impaired intestinal barrier function with low grade chronic inflammation, hyperinsulinemia, and increased body fat, which are signatures of diet-induced diabetes (type 2 diabetes) and obesity in humans. Hyperglycemia in type 2 diabetes and obesity is caused by insulin resistance. Insulin resistance results in inhibition of glucose uptake by liver and other peripheral tissues, principally adipose and muscle and with concurrently higher hepatic glucose production. Therefore, modulation of hepatic glucose metabolism is an important target for antidiabetic treatment approaches. We demonstrate that during glucose uptake, cell surface abundance of zinc transporter ZIP14 and mediated zinc transport increases. Zinc is distributed to multiple sites in hepatocytes through sequential translocation of ZIP14 from plasma membrane to early and late endosomes. Endosomes from Zip14 KO mice were zinc-deficient because activities of the zinc-dependent insulin-degrading proteases insulin-degrading enzyme and cathepsin D were impaired; hence insulin receptor activity increased. Transient increases in cytosolic zinc levels are concurrent with glucose uptake and suppression of glycogen synthesis. In contrast, Zip14 KO mice exhibited greater hepatic glycogen synthesis and impaired gluconeogenesis and glycolysis related to low cytosolic zinc levels. We can conclude that ZIP14-mediated zinc transport contributes to regulation of endosomal insulin receptor activity and glucose homeostasis in hepatocytes. Therefore, modulation of ZIP14 transport activity presents a new target for management of diabetes and other glucose-related disorders. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Prostaglandin E2 potentiation of P2X3 receptor mediated currents in dorsal root ganglion neurons

    Directory of Open Access Journals (Sweden)

    Huang Li-Yen

    2007-08-01

    Full Text Available Abstract Prostaglandin E2 (PGE2 is a well-known inflammatory mediator that enhances the excitability of DRG neurons. Homomeric P2X3 and heteromeric P2X2/3 receptors are abundantly expressed in dorsal root ganglia (DRG neurons and participate in the transmission of nociceptive signals. The interaction between PGE2 and P2X3 receptors has not been well delineated. We studied the actions of PGE2 on ATP-activated currents in dissociated DRG neurons under voltage-clamp conditions. PGE2 had no effects on P2X2/3 receptor-mediated responses, but significantly potentiated fast-inactivating ATP currents mediated by homomeric P2X3 receptors. PGE2 exerted its action by activating EP3 receptors. To study the mechanism underlying the action of PGE2, we found that the adenylyl cyclase activator, forskolin and the membrane-permeable cAMP analogue, 8-Br-cAMP increased ATP currents, mimicking the effect of PGE2. In addition, forskolin occluded the enhancement produced by PGE2. The protein kinase A (PKA inhibitors, H89 and PKA-I blocked the PGE2 effect. In contrast, the PKC inhibitor, bisindolymaleimide (Bis did not change the potentiating action of PGE2. We further showed that PGE2 enhanced α,β-meATP-induced allodynia and hyperalgesia and the enhancement was blocked by H89. These observations suggest that PGE2 binds to EP3 receptors, resulting in the activation of cAMP/PKA signaling pathway and leading to an enhancement of P2X3 homomeric receptor-mediated ATP responses in DRG neurons.

  11. ADF/cofilin-mediated actin dynamics regulate AMPA receptor trafficking during synaptic plasticity.

    Science.gov (United States)

    Gu, Jiaping; Lee, Chi Wai; Fan, Yanjie; Komlos, Daniel; Tang, Xin; Sun, Chicheng; Yu, Kuai; Hartzell, H Criss; Chen, Gong; Bamburg, James R; Zheng, James Q

    2010-10-01

    Dendritic spines undergo actin-based growth and shrinkage during synaptic plasticity, in which the actin depolymerizing factor (ADF)/cofilin family of actin-associated proteins are important. Elevated ADF/cofilin activities often lead to reduced spine size and immature spine morphology but can also enhance synaptic potentiation in some cases. Thus, ADF/cofilin may have distinct effects on postsynaptic structure and function. We found that ADF/cofilin-mediated actin dynamics regulated AMPA receptor (AMPAR) trafficking during synaptic potentiation, which was distinct from actin's structural role in spine morphology. Specifically, elevated ADF/cofilin activity markedly enhanced surface addition of AMPARs after chemically induced long-term potentiation (LTP), whereas inhibition of ADF/cofilin abolished AMPAR addition. We found that chemically induced LTP elicited a temporal sequence of ADF/cofilin dephosphorylation and phosphorylation that underlies AMPAR trafficking and spine enlargement. These findings suggest that temporally regulated ADF/cofilin activities function in postsynaptic modifications of receptor number and spine size during synaptic plasticity.

  12. Astrocytic β2-adrenergic receptors mediate hippocampal long-term memory consolidation

    KAUST Repository

    Gao, Virginia

    2016-07-12

    Emotionally relevant experiences form strong and long-lasting memories by critically engaging the stress hormone/neurotransmitter noradrenaline, which mediates and modulates the consolidation of these memories. Noradrenaline acts through adrenergic receptors (ARs), of which β2- Adrenergic receptors (βARs) are of particular importance. The differential anatomical and cellular distribution of βAR subtypes in the brain suggests that they play distinct roles in memory processing, although much about their specific contributions and mechanisms of action remains to be understood. Here we show that astrocytic rather than neuronal β2ARs in the hippocampus play a key role in the consolidation of a fear-based contextual memory. These hippocampal β2ARs, but not β1ARs, are coupled to the training-dependent release of lactate from astrocytes, which is necessary for long- Term memory formation and for underlying molecular changes. This key metabolic role of astrocytic β2ARs may represent a novel target mechanism for stress-related psychopathologies and neurodegeneration.

  13. Histamine 1 Receptor Blockade Enhances Eosinophil-Mediated Clearance of Adult Filarial Worms

    Science.gov (United States)

    Fox, Ellen Mueller; Morris, Christopher P.; Hübner, Marc P.; Mitre, Edward

    2015-01-01

    Filariae are tissue-invasive nematodes that cause diseases such as elephantiasis and river blindness. The goal of this study was to characterize the role of histamine during Litomosoides sigmodontis infection of BALB/c mice, a murine model of filariasis. Time course studies demonstrated that while expression of histidine decarboxylase mRNA increases throughout 12 weeks of infection, serum levels of histamine exhibit two peaks—one 30 minutes after primary infection and one 8 weeks later. Interestingly, mice treated with fexofenadine, a histamine receptor 1 inhibitor, demonstrated significantly reduced worm burden in infected mice compared to untreated infected controls. Although fexofenadine-treated mice had decreased antigen-specific IgE levels as well as lower splenocyte IL-5 and IFNγ production, they exhibited a greater than fourfold rise in eosinophil numbers at the tissue site where adult L. sigmodontis worms reside. Fexofenadine-mediated clearance of L. sigmodontis worms was dependent on host eosinophils, as fexofenadine did not decrease worm burdens in eosinophil-deficient dblGATA mice. These findings suggest that histamine release induced by tissue invasive helminths may aid parasite survival by diminishing eosinophilic responses. Further, these results raise the possibility that combining H1 receptor inhibitors with current anthelmintics may improve treatment efficacy for filariae and other tissue-invasive helminths. PMID:26204515

  14. P2X7 receptor-mediated analgesia in cancer-induced bone pain

    DEFF Research Database (Denmark)

    Falk, Sarah; D. Schwab, Samantha; Frøsig-Jørgensen, Majbrit

    2015-01-01

    for cancer-induced bone pain. The P2X7 receptor (P2X7R) is involved in a variety of cellular functions and has been linked to both inflammatory and neuropathic pain. Here we study the analgesic potential of P2X7 receptor antagonism in a rat model of cancer-induced bone pain. In cancer-bearing animals, the P2...... to low intensity or electrical stimulation. In contrast, A839977 had no effect on the tested parameters in naïve or sham animals. In awake animals, 40mg/kg A839977 (i.p) significantly reduced both early and late stage pain behavior. In contrast, no effect was observed in sham or vehicle-treated animals....... The results suggest that the P2X7R is involved in the mechanisms of cancer-induced bone pain, and that P2X7R antagonism might be a useful analgesic target. No effect was observed in sham or naïve animals, indicating that the P2X7R-mediated effect is state-dependent, and might therefore be an advantageous...

  15. Transient Receptor Potential Melastatin-3 (TRPM3) Mediates Nociceptive-Like Responses in Hydra vulgaris.

    Science.gov (United States)

    Malafoglia, Valentina; Traversetti, Lorenzo; Del Grosso, Floriano; Scalici, Massimiliano; Lauro, Filomena; Russo, Valeria; Persichini, Tiziana; Salvemini, Daniela; Mollace, Vincenzo; Fini, Massimo; Raffaeli, William; Muscoli, Carolina; Colasanti, Marco

    2016-01-01

    The ability of mammals to feel noxious stimuli lies in a heterogeneous group of primary somatosensory neurons termed nociceptors, which express specific membrane receptors, such as the Transient Receptor Potential (TRP) family. Here, we show that one of the most important nociceptive-like pathways is conserved in the freshwater coelenterate Hydra vulgaris, the most primitive organism possessing a nervous system. In particular, we found that H. vulgaris expresses TRPM3, a nociceptor calcium channel involved in the detection of noxious heat in mammals. Furthermore, we detected that both heat shock and TRPM3 specific agonist (i.e., pregnenolone sulfate) induce the modulation of the heat shock protein 70 (HSP70) and the nitric oxide synthase (NOS), two genes activated by TRP-mediated heat painful stimuli in mammals. As expected, these effects are inhibited by a TRPM3 antagonist (i.e., mefenamic acid). Interestingly, the TRPM3 agonist and heat shock also induce the expression of nuclear transcription erythroid 2-related factor (Nrf2) and superoxide dismutase (SOD), known markers of oxidative stress; noteworthy gene expression was also inhibited by the TRPM3 antagonist. As a whole, our results demonstrate the presence of conserved molecular oxidative/nociceptive-like pathways at the primordial level of the animal kingdom.

  16. Human Milk Components Modulate Toll-Like Receptor-Mediated Inflammation.

    Science.gov (United States)

    He, YingYing; Lawlor, Nathan T; Newburg, David S

    2016-01-01

    Toll-like receptor (TLR) signaling is central to innate immunity. Aberrant expression of TLRs is found in neonatal inflammatory diseases. Several bioactive components of human milk modulate TLR expression and signaling pathways, including soluble toll-like receptors (sTLRs), soluble cluster of differentiation (sCD) 14, glycoproteins, small peptides, and oligosaccharides. Some milk components, such as sialyl (α2,3) lactose and lacto-N-fucopentaose III, are reported to increase TLR signaling; under some circumstances this might contribute toward immunologic balance. Human milk on the whole is strongly anti-inflammatory, and contains abundant components that depress TLR signaling pathways: sTLR2 and sCD14 inhibit TLR2 signaling; sCD14, lactadherin, lactoferrin, and 2'-fucosyllactose attenuate TLR4 signaling; 3'-galactosyllactose inhibits TLR3 signaling, and β-defensin 2 inhibits TLR7 signaling. Feeding human milk to neonates decreases their risk of sepsis and necrotizing enterocolitis. Thus, the TLR regulatory components found in human milk hold promise as benign oral prophylactic and therapeutic treatments for the many gastrointestinal inflammatory disorders mediated by abnormal TLR signaling. © 2016 American Society for Nutrition.

  17. Implications of astrocytes in mediating the protective effects of Selective Estrogen Receptor Modulators upon brain damage

    Directory of Open Access Journals (Sweden)

    George E. Barreto

    2015-04-01

    Full Text Available Selective Estrogen Receptor Modulators (SERMs are steroidal or non-steroidal compounds that are already used in clinical practice for the treatment of breast cancer, osteoporosis and menopausal symptoms. While SERMs actions in the breast, bone, and uterus have been well characterized, their actions in the brain are less well understood. Previous works have demonstrated the beneficial effects of SERMs in different chronic neurodegenerative diseases like Alzheimer, Parkinson’s disease and Multiple sclerosis, as well as acute degeneration as stroke and traumatic brain injury. Moreover, these compounds exhibit similar protective actions as those of estradiol in the Central Nervous System, overt any secondary effect. For these reasons, in the past few years, there has been a growing interest in the neuroprotective effects exerted directly or indirectly by SERMs in the SNC. In this context, astrocytes play an important role in the maintenance of brain metabolism, and antioxidant support to neurons, thus indicating that better protection of astrocytes are an important asset targeting neuronal protection. Moreover, various clinical and experimental studies have reported that astrocytes are essential for the neuroprotective effects of SERMs during neuronal injuries, as these cells express different estrogen receptors in cell membrane, demonstrating that part of SERMs effects upon injury may be mediated by astrocytes. The present work highlights the current evidence on the protective mechanisms of SERMs, such as tamoxifen and raloxifene, in the SNC, and their modulation of astrocytic properties as promising therapeutic targets during brain damage.

  18. Purinergic A2b Receptor Activation by Extracellular Cues Affects Positioning of the Centrosome and Nucleus and Causes Reduced Cell Migration.

    Science.gov (United States)

    Ou, Young; Chan, Gordon; Zuo, Jeremy; Rattner, Jerome B; van der Hoorn, Frans A

    2016-07-15

    The tight, relative positioning of the nucleus and centrosome in mammalian cells is important for the regulation of cell migration. Under pathophysiological conditions, the purinergic A2b receptor can regulate cell motility, but the underlying mechanism remains unknown. Expression of A2b, normally low, is increased in tissues experiencing adverse physiological conditions, including hypoxia and inflammation. ATP is released from such cells. We investigated whether extracellular cues can regulate centrosome-nucleus positioning and cell migration. We discovered that hypoxia as well as extracellular ATP cause a reversible increase in the distance between the centrosome and nucleus and reduced cell motility. We uncovered the underlying pathway: both treatments act through the A2b receptor and specifically activate the Epac1/RapGef3 pathway. We show that cells lacking A2b do not respond in this manner to hypoxia or ATP but transfection of A2b restores this response, that Epac1 is critically involved, and that Rap1B is important for the relative positioning of the centrosome and nucleus. Our results represent, to our knowledge, the first report demonstrating that pathophysiological conditions can impact the distance between the centrosome and nucleus. Furthermore, we identify the A2b receptor as a central player in this process. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Ionotropic glutamate receptors mediate inducible defense in the water flea Daphnia pulex.

    Directory of Open Access Journals (Sweden)

    Hitoshi Miyakawa

    Full Text Available Phenotypic plasticity is the ability held in many organisms to produce different phenotypes with a given genome in response to environmental stimuli, such as temperature, nutrition and various biological interactions. It seems likely that environmental signals induce a variety of mechanistic responses that influence ontogenetic processes. Inducible defenses, in which prey animals alter their morphology, behavior and/or other traits to help protect against direct or latent predation threats, are among the most striking examples of phenotypic plasticity. The freshwater microcrustacean Daphnia pulex forms tooth-like defensive structures, "neckteeth," in response to chemical cues or signals, referred to as "kairomones," in this case released from phantom midge larvae, a predator of D. pulex. To identify factors involved in the reception and/or transmission of a kairomone, we used microarray analysis to identify genes up-regulated following a short period of exposure to the midge kairomone. In addition to identifying differentially expressed genes of unknown function, we also found significant up-regulation of genes encoding ionotropic glutamate receptors, which are known to be involved in neurotransmission in many animal species. Specific antagonists of these receptors strongly inhibit the formation of neckteeth in D. pulex, although agonists did not induce neckteeth by themselves, indicating that ionotropic glutamate receptors are necessary but not sufficient for early steps of neckteeth formation in D. pulex. Moreover, using co-exposure of D. pulex to antagonists and juvenile hormone (JH, which physiologically mediates neckteeth formation, we found evidence suggesting that the inhibitory effect of antagonists is not due to direct inhibition of JH synthesis/secretion. Our findings not only provide a candidate molecule required for the inducible defense response in D. pulex, but also will contribute to the understanding of complex mechanisms

  20. Amygdala opioid receptors mediate the electroacupuncture-induced deterioration of sleep disruptions in epilepsy rats.

    Science.gov (United States)

    Yi, Pei-Lu; Lu, Chin-Yu; Cheng, Chiung-Hsiang; Tsai, Yi-Fong; Lin, Chung-Tien; Chang, Fang-Chia

    2013-11-12

    Clinical and experimental evidence demonstrates that sleep and epilepsy reciprocally affect each other. Previous studies indicated that epilepsy alters sleep homeostasis; in contrast, sleep disturbance deteriorates epilepsy. If a therapy possesses both epilepsy suppression and sleep improvement, it would be the priority choice for seizure control. Effects of acupuncture of Feng-Chi (GB20) acupoints on epilepsy suppression and insomnia treatment have been documented in the ancient Chinese literature, Lingshu Jing (Classic of the Miraculous Pivot). Therefore, this study was designed to investigate the effect of electroacupuncture (EA) stimulation of bilateral Feng-Chi acupoints on sleep disruptions in rats with focal epilepsy. Our result indicates that administration of pilocarpine into the left central nucleus of amygdala (CeA) induced focal epilepsy and decreased both rapid eye movement (REM) sleep and non-REM (NREM) sleep. High-frequency (100 Hz) EA stimulation of bilateral Feng-Chi acupoints, in which a 30-min EA stimulation was performed before the dark period of the light:dark cycle in three consecutive days, further deteriorated pilocarpine-induced sleep disruptions. The EA-induced exacerbation of sleep disruption was blocked by microinjection of naloxone, μ- (naloxonazine), κ- (nor-binaltorphimine) or δ-receptor antagonists (natrindole) into the CeA, suggesting the involvement of amygdaloid opioid receptors. The present study suggests that high-frequency (100 Hz) EA stimulation of bilateral Feng-Chi acupoints exhibits no benefit in improving pilocarpine-induced sleep disruptions; in contrast, EA further deteriorated sleep disturbances. Opioid receptors in the CeA mediated EA-induced exacerbation of sleep disruptions in epileptic rats.

  1. The adhesion G protein-coupled receptor G2 (ADGRG2/GPR64) constitutively activates SRE and NFκB and is involved in cell adhesion and migration

    DEFF Research Database (Denmark)

    Cornelia Peeters, Miriam; Fokkelman, Michiel; Boogaard, Bob

    2015-01-01

    Adhesion G protein-coupled receptors (ADGRs) are believed to be activated by auto-proteolytic cleavage of their very large extracellular N-terminal domains normally acting as a negative regulator of the intrinsically constitutively active seven transmembrane domain. ADGRG2 (or GPR64) which...... intracellular signal transduction. Knockdown of ADGRG2 by siRNA in the highly motile breast cancer cell lines Hs578T and MDA-MB-231 resulted in a strong reduction in cell adhesion and subsequent cell migration which was associated with a selective reduction in RelB, an NFκB family member. It is concluded...... that the adhesion GPCR ADGRG2 is critically involved in the adhesion and migration of certain breast cancer cells through mechanisms including a non-canonical NFkB pathway and that ADGRG2 could be a target for treatment of certain types of cancer....

  2. Human cytomegalovirus chemokine receptor US28 induces migration of cells on a CX3CL1-presenting surface

    DEFF Research Database (Denmark)

    Hjortø, Gertrud M; Kiilerich-Pedersen, Katrine; Selmeczi, David

    2013-01-01

    -activated endothelial cells, causing redirection of CX3CR1-expressing leukocytes in the blood to sites of infection. Here, we used stable transfected cell lines to examine how US28 expression affects cell migration on immobilized full-length CX3CL1, to model how HCMV-infected leukocytes interact with inflamed...... the migration of cells upon binding to the same ligand. Overall, this indicates that infected cells probably move more than uninfected cells in inflamed tissues with high CX3CL1 expression, with soluble chemokines affecting the final migration....

  3. Neurotrophin receptor p75NTR mediates Huntington’s disease–associated synaptic and memory dysfunction

    Science.gov (United States)

    Brito, Verónica; Giralt, Albert; Enriquez-Barreto, Lilian; Puigdellívol, Mar; Suelves, Nuria; Zamora-Moratalla, Alfonsa; Ballesteros, Jesús J.; Martín, Eduardo D.; Dominguez-Iturza, Nuria; Morales, Miguel; Alberch, Jordi; Ginés, Sílvia

    2014-01-01

    Learning and memory deficits are early clinical manifestations of Huntington’s disease (HD). These cognitive impairments have been mainly associated with frontostriatal HD pathology; however, compelling evidence provided by several HD murine models suggests that the hippocampus may contribute to synaptic deficits and memory dysfunction in HD. The neurotrophin receptor p75NTR negatively regulates spine density, which is associated with learning and memory; therefore, we explored whether disturbed p75NTR function in the hippocampus could contribute to synaptic dysfunction and memory deficits in HD. Here, we determined that levels of p75NTR are markedly increased in the hippocampus of 2 distinct mouse models of HD and in HD patients. Normalization of p75NTR levels in HD mutant mice heterozygous for p75NTR prevented memory and synaptic plasticity deficits and ameliorated dendritic spine abnormalities, likely through normalization of the activity of the GTPase RhoA. Moreover, viral-mediated overexpression of p75NTR in the hippocampus of WT mice reproduced HD learning and memory deficits, while knockdown of p75NTR in the hippocampus of HD mice prevented cognitive decline. Together, these findings provide evidence of hippocampus-associated memory deficits in HD and demonstrate that p75NTR mediates synaptic, learning, and memory dysfunction in HD. PMID:25180603

  4. AMPA receptor-induced local brain-derived neurotrophic factor signaling mediates motor recovery after stroke.

    Science.gov (United States)

    Clarkson, Andrew N; Overman, Justine J; Zhong, Sheng; Mueller, Rudolf; Lynch, Gary; Carmichael, S Thomas

    2011-03-09

    Stroke is the leading cause of adult disability. Recovery after stroke shares similar molecular and cellular properties with learning and memory. A main component of learning-induced plasticity involves signaling through AMPA receptors (AMPARs). We systematically tested the role of AMPAR function in motor recovery in a mouse model of focal stroke. AMPAR function controls functional recovery beginning 5 d after the stroke. Positive allosteric modulators of AMPARs enhance recovery of limb control when administered after a delay from the stroke. Conversely, AMPAR antagonists impair motor recovery. The contributions of AMPARs to recovery are mediated by release of brain-derived neurotrophic factor (BDNF) in periinfarct cortex, as blocking local BDNF function in periinfarct cortex blocks AMPAR-mediated recovery and prevents the normal pattern of motor recovery. In contrast to a delayed AMPAR role in motor recovery, early administration of AMPAR agonists after stroke increases stroke damage. These findings indicate that the role of glutamate signaling through the AMPAR changes over time in stroke: early potentiation of AMPAR signaling worsens stroke damage, whereas later potentiation of the same signaling system improves functional recovery.

  5. Reduction of decoy receptor 3 enhances TRAIL-mediated apoptosis in pancreatic cancer.

    Directory of Open Access Journals (Sweden)

    Wei Wang

    Full Text Available Most human pancreatic cancer cells are resistant to tumor necrosis factor (TNF-related apoptosis-inducing ligand (TRAIL-mediated apoptosis. However, the mechanisms by which pancreatic cancer cells utilize their extracellular molecules to counteract the proapoptotic signaling mediated by the TNF family are largely unknown. In this study, we demonstrate for the first time that DcR3, a secreted decoy receptor that malignant pancreatic cancer cells express at a high level, acts as an extracellular antiapoptotic molecule by binding to TRAIL and counteracting its death-promoting function. The reduction of DcR3 with siRNA unmasked TRAIL and greatly enhanced TRAIL-induced apoptosis. Gemcitabine, a first-line drug for pancreatic cancer, also reduced the level of DcR3. The addition of DcR3 siRNA further enhanced gemcitabine-induced apoptosis. Notably, our in vivo study demonstrated that the therapeutic effect of gemcitabine could be enhanced via further reduction of DcR3, suggesting that downregulation of DcR3 in tumor cells could tip the balance of pancreatic cells towards apoptosis and potentially serve as a new strategy for pancreatic cancer therapy.

  6. Chemokine receptor Ccr1 drives neutrophil-mediated kidney immunopathology and mortality in invasive candidiasis.

    Science.gov (United States)

    Lionakis, Michail S; Fischer, Brett G; Lim, Jean K; Swamydas, Muthulekha; Wan, Wuzhou; Richard Lee, Chyi-Chia; Cohen, Jeffrey I; Scheinberg, Phillip; Gao, Ji-Liang; Murphy, Philip M

    2012-01-01

    Invasive candidiasis is the 4(th) leading cause of nosocomial bloodstream infection in the US with mortality that exceeds 40% despite administration of antifungal therapy; neutropenia is a major risk factor for poor outcome after invasive candidiasis. In a fatal mouse model of invasive candidiasis that mimics human bloodstream-derived invasive candidiasis, the most highly infected organ is the kidney and neutrophils are the major cellular mediators of host defense; however, factors regulating neutrophil recruitment have not been previously defined. Here we show that mice lacking chemokine receptor Ccr1, which is widely expressed on leukocytes, had selectively impaired accumulation of neutrophils in the kidney limited to the late phase of the time course of the model; surprisingly, this was associated with improved renal function and survival without affecting tissue fungal burden. Consistent with this, neutrophils from wild-type mice in blood and kidney switched from Ccr1(lo) to Ccr1(high) at late time-points post-infection, when Ccr1 ligands were produced at high levels in the kidney and were chemotactic for kidney neutrophils ex vivo. Further, when a 1∶1 mixture of Ccr1(+/+) and Ccr1(-/-) donor neutrophils was adoptively transferred intravenously into Candida-infected Ccr1(+/+) recipient mice, neutrophil trafficking into the kidney was significantly skewed toward Ccr1(+/+) cells. Thus, neutrophil Ccr1 amplifies late renal immunopathology and increases mortality in invasive candidiasis by mediating excessive recruitment of neutrophils from the blood to the target organ.

  7. [Lentiviral vector-mediated RNA interfere gene Nogo receptor to repair spinal cord injury].

    Science.gov (United States)

    Lü, Bi-tao; Yuan, Wen; Xu, Sheng-ming

    2010-10-15

    To evaluate the effects of lentiviral vector-mediated RNA interfere gene Nogo receptor (NgR) of rat cortical neurons in repairing spinal cord injury. The recombinant-lentiviral vector with small inferring RNA siNgR199 which had been constructed was transfected into rat cortical neuron cells in vitro in 3 multiplicity of infection (MOI). The infection rate was determined with fluorescent microscope, and the target gene was detected by PCR analysis. Then, the recombinant was injected into the cortical motor area of the rats with severe spinal cord injury, and the saline was also injected into other rats with severe spinal cord injury as a match control. The functional recovery of the rats' hindlimb was assessed using BBB score and the nerve fiber of the injured region was observed by nerve tracing. The rate of recombinant infecting rat cortical neuron in vitro exceeded 99%. PCR analysis confirmed that the effect of lentiviral vector-mediated RNA interfering gene NgR of rat cortical neurons in vitro was 61%. Although all rats with spinal cord injury were observed to have the hindlimb functional recovery, these rats injected with recombinant had better hindlimb functional recovery than others showing by more BBB score (P vector with siNgR199 which had been constructed is able to promote the growth of nerve fiber and the functional recovery of the rats' hindlimb.

  8. Chemokine receptor Ccr1 drives neutrophil-mediated kidney immunopathology and mortality in invasive candidiasis.

    Directory of Open Access Journals (Sweden)

    Michail S Lionakis

    Full Text Available Invasive candidiasis is the 4(th leading cause of nosocomial bloodstream infection in the US with mortality that exceeds 40% despite administration of antifungal therapy; neutropenia is a major risk factor for poor outcome after invasive candidiasis. In a fatal mouse model of invasive candidiasis that mimics human bloodstream-derived invasive candidiasis, the most highly infected organ is the kidney and neutrophils are the major cellular mediators of host defense; however, factors regulating neutrophil recruitment have not been previously defined. Here we show that mice lacking chemokine receptor Ccr1, which is widely expressed on leukocytes, had selectively impaired accumulation of neutrophils in the kidney limited to the late phase of the time course of the model; surprisingly, this was associated with improved renal function and survival without affecting tissue fungal burden. Consistent with this, neutrophils from wild-type mice in blood and kidney switched from Ccr1(lo to Ccr1(high at late time-points post-infection, when Ccr1 ligands were produced at high levels in the kidney and were chemotactic for kidney neutrophils ex vivo. Further, when a 1∶1 mixture of Ccr1(+/+ and Ccr1(-/- donor neutrophils was adoptively transferred intravenously into Candida-infected Ccr1(+/+ recipient mice, neutrophil trafficking into the kidney was significantly skewed toward Ccr1(+/+ cells. Thus, neutrophil Ccr1 amplifies late renal immunopathology and increases mortality in invasive candidiasis by mediating excessive recruitment of neutrophils from the blood to the target organ.

  9. Extrasynaptic NMDA receptors couple preferentially to excitotoxicity via calpain-mediated cleavage of STEP.

    Science.gov (United States)

    Xu, Jian; Kurup, Pradeep; Zhang, Yongfang; Goebel-Goody, Susan M; Wu, Peter H; Hawasli, Ammar H; Baum, Matthew L; Bibb, James A; Lombroso, Paul J

    2009-07-22

    NMDA receptor (NMDAR)-mediated excitotoxicity plays an important role in several CNS disorders, including epilepsy, stroke, and ischemia. Here we demonstrate the involvement of striatal-enriched protein tyrosine phosphatase (STEP) in this critical process. STEP(61) is an alternatively spliced member of the family that is present in postsynaptic terminals. In an apparent paradox, STEP(61) regulates extracellular signal-regulated kinase 1/2 (ERK1/2) and p38, two proteins with opposing functions; activated p38 promotes cell death, whereas activated ERK1/2 promotes cell survival. We found that synaptic stimulation of NMDARs promoted STEP(61) ubiquitination and degradation, concomitant with ERK1/2 activation. In contrast, extrasynaptic stimulation of NMDARs invoked calpain-mediated proteolysis of STEP(61), producing the truncated cleavage product STEP(33) and activation of p38. The calpain cleavage site on STEP was mapped to the kinase interacting motif, a domain required for substrate binding. As a result, STEP(33) neither interacts with nor dephosphorylates STEP substrates. A synthetic peptide spanning the calpain cleavage site efficiently reduced STEP(61) degradation and attenuated p38 activation and cell death in slice models. Furthermore, this peptide was neuroprotective when neurons were subjected to excitotoxicity or cortical slices were exposed to ischemic conditions. These findings suggest a novel mechanism by which differential NMDAR stimulation regulates STEP(61) to promote either ERK1/2 or p38 activation and identifies calpain cleavage of STEP(61) as a valid target for the development of neuroprotective therapy.

  10. Evodiamine Induces Transient Receptor Potential Vanilloid-1-Mediated Protective Autophagy in U87-MG Astrocytes

    Directory of Open Access Journals (Sweden)

    Ann-Jeng Liu

    2013-01-01

    Full Text Available Cerebral ischemia is a leading cause of mortality and morbidity worldwide, which results in cognitive and motor dysfunction, neurodegenerative diseases, and death. Evodiamine (Evo is extracted from Evodia rutaecarpa Bentham, a plant widely used in Chinese herbal medicine, which possesses variable biological abilities, such as anticancer, anti-inflammation, antiobesity, anti-Alzheimer’s disease, antimetastatic, antianoxic, and antinociceptive functions. But the effect of Evo on ischemic stroke is unclear. Increasing data suggest that activation of autophagy, an adaptive response to environmental stresses, could protect neurons from ischemia-induced cell death. In this study, we found that Evo induced autophagy in U87-MG astrocytes. A scavenger of extracellular calcium and an antagonist of transient receptor potential vanilloid-1 (TRPV-1 decreased the percentage of autophagy accompanied by an increase in apoptosis, suggesting that Evo may induce calcium-mediated protective autophagy resulting from an influx of extracellular calcium. The same phenomena were also confirmed by a small interfering RNA technique to knock down the expression of TRPV1. Finally, Evo-induced c-Jun N-terminal kinases (JNK activation was reduced by a TRPV1 antagonist, indicating that Evo-induced autophagy may occur through a calcium/c-Jun N-terminal kinase (JNK pathway. Collectively, Evo induced an influx of extracellular calcium, which led to JNK-mediated protective autophagy, and this provides a new option for ischemic stroke treatment.

  11. Evodiamine Induces Transient Receptor Potential Vanilloid-1-Mediated Protective Autophagy in U87-MG Astrocytes

    Science.gov (United States)

    Liu, Ann-Jeng; Wang, Sheng-Hao; Hou, Sz-Ying; Chiu, Wen-Ta; Hsiao, Sheng-Huang; Chen, Thay-Hsiung

    2013-01-01

    Cerebral ischemia is a leading cause of mortality and morbidity worldwide, which results in cognitive and motor dysfunction, neurodegenerative diseases, and death. Evodiamine (Evo) is extracted from Evodia rutaecarpa Bentham, a plant widely used in Chinese herbal medicine, which possesses variable biological abilities, such as anticancer, anti-inflammation, antiobesity, anti-Alzheimer's disease, antimetastatic, antianoxic, and antinociceptive functions. But the effect of Evo on ischemic stroke is unclear. Increasing data suggest that activation of autophagy, an adaptive response to environmental stresses, could protect neurons from ischemia-induced cell death. In this study, we found that Evo induced autophagy in U87-MG astrocytes. A scavenger of extracellular calcium and an antagonist of transient receptor potential vanilloid-1 (TRPV-1) decreased the percentage of autophagy accompanied by an increase in apoptosis, suggesting that Evo may induce calcium-mediated protective autophagy resulting from an influx of extracellular calcium. The same phenomena were also confirmed by a small interfering RNA technique to knock down the expression of TRPV1. Finally, Evo-induced c-Jun N-terminal kinases (JNK) activation was reduced by a TRPV1 antagonist, indicating that Evo-induced autophagy may occur through a calcium/c-Jun N-terminal kinase (JNK) pathway. Collectively, Evo induced an influx of extracellular calcium, which led to JNK-mediated protective autophagy, and this provides a new option for ischemic stroke treatment. PMID:24454492

  12. Novel method demonstrates differential ligand activation and phosphatase-mediated deactivation of insulin receptor tyrosine-specific phosphorylation.

    Science.gov (United States)

    Cieniewicz, Anne M; Cooper, Philip R; McGehee, Jennifer; Lingham, Russell B; Kihm, Anthony J

    2016-08-01

    Insulin receptor signaling is a complex cascade leading to a multitude of intracellular functional responses. Three natural ligands, insulin, IGF1 and IGF2, are each capable of binding with different affinities to the insulin receptor, and result in variable biological responses. However, it is likely these affinity differences alone cannot completely explain the myriad of diverse cellular outcomes. Ligand binding initiates activation of a signaling cascade resulting in phosphorylation of the IR itself and other intracellular proteins. The direct catalytic activity along with the temporally coordinated assembly of signaling proteins is critical for insulin receptor signaling. We hypothesized that determining differential phosphorylation among individual tyrosine sites activated by ligand binding or dephosphorylation by phosphatases could provide valuable insight into insulin receptor signaling. Here, we present a sensitive, novel immunoassay adapted from Meso Scale Discovery technology to quantitatively measure changes in site-specific phosphorylation levels on endogenous insulin receptors from HuH7 cells. We identified insulin receptor phosphorylation patterns generated upon differential ligand activation and phosphatase-mediated deactivation. The data demonstrate that insulin, IGF1 and IGF2 elicit different insulin receptor phosphorylation kinetics and potencies that translate to downstream signaling. Furthermore, we show that insulin receptor deactivation, regulated by tyrosine phosphatases, occurs distinctively across specific tyrosine residues. In summary, we present a novel, quantitative and high-throughput assay that has uncovered differential ligand activation and site-specific deactivation of the insulin receptor. These results may help elucidate some of the insulin signaling mechanisms, discriminate ligand activity and contribute to a better understanding of insulin receptor signaling. We propose this methodology as a powerful approach to characterize

  13. Biased G protein-coupled receptor agonism mediates Neu1 sialidase and matrix metalloproteinase-9 crosstalk to induce transactivation of insulin receptor signaling.

    Science.gov (United States)

    Haxho, Fiona; Haq, Sabah; Szewczuk, Myron R

    2017-12-24

    G protein-coupled receptors (GPCR) can participate in a number of signaling pathways, and this property led to the concept of biased GPCR agonism. Agonists, antagonists and allosteric modulators can bind to GPCRs in different ways, creating unique conformations that differentially modulate signaling through one or more G proteins. A unique neuromedin B (NMBR) GPCR-signaling platform controlling mammalian neuraminidase-1 (Neu1) and matrix metalloproteinase-9 (MMP9) crosstalk has been reported in the activation of the insulin receptor (IR) through the modification of the IR glycosylation. Here, we propose that there exists a biased GPCR agonism as small diffusible molecules in the activation of Neu1-mediated insulin receptor signaling. GPCR agonists bombesin, bradykinin, angiotensin I and angiotensin II significantly and dose-dependently induce Neu1 sialidase activity and IR activation in human IR-expressing rat hepatoma cell lines (HTC-IR), in the absence of insulin. Furthermore, the GPCR agonist-induced Neu1 sialidase activity could be specifically blocked by the NMBR inhibitor, BIM-23127. Protein expression analyses showed that these GPCR agonists significantly induced phosphorylation of IRβ and insulin receptor substrate-1 (IRS1). Among these, angiotensin II was the most potent GPCR agonist capable of promoting IRβ phosphorylation in HTC-IR cells. Interestingly, treatment with BIM-23127 and Neu1 inhibitor oseltamivir phosphate were able to block GPCR agonist-induced IR activation in HTC cells in vitro. Additionally, we found that angiotensin II receptor (type I) exists in a multimeric receptor complex with Neu1, IRβ and NMBR in naïve (unstimulated) and stimulated HTC-IR cells with insulin, bradykinin, angiotensin I and angiotensin II. This complex suggests a molecular link regulating the interaction and signaling mechanism between these molecules on the cell surface. These findings uncover a biased GPCR agonist-induced IR transactivation signaling axis

  14. Suppression of TLR4-mediated inflammatory response by macrophage class A scavenger receptor (CD204)

    Energy Technology Data Exchange (ETDEWEB)

    Ohnishi, Koji; Komohara, Yoshihiro; Fujiwara, Yukio; Takemura, Kenichi [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Lei, XiaoFeng [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Department of Biochemistry, Showa University School of Medicine, Tokyo (Japan); Nakagawa, Takenobu [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Sakashita, Naomi [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Department of Human Pathology, Institute of Health Biosciences, The University of Tokushima, Tokushima (Japan); Takeya, Motohiro, E-mail: takeya@kumamoto-u.ac.jp [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan)

    2011-08-05

    Highlights: {yields} We focused on the interaction between SR-A and TLR4 signaling in this study. {yields} SR-A deletion promoted NF{kappa}B activation in macrophages in septic model mouse. {yields} SR-A suppresses both MyD88-dependent and -independent TLR4 signaling in vitro. {yields} SR-A clears LPS binding to TLR4 which resulting in the suppression of TLR4 signals. -- Abstract: The class A scavenger receptor (SR-A, CD204), one of the principal receptors expressed on macrophages, has been found to regulate inflammatory response and attenuate septic endotoxemia. However, the detailed mechanism of this process has not yet been well characterized. To clarify the regulative mechanisms of lipopolysaccharide (LPS)-induced macrophage activation by SR-A, we evaluated the activation of Toll-like receptor 4 (TLR4)-mediated signaling molecules in SR-A-deficient (SR-A{sup -/-}) macrophages. In a septic shock model, the blood levels of tumor necrosis factor (TNF)-{alpha}, interleukin (IL)-6 and interferon (IFN)-{beta} were significantly increased in SR-A{sup -/-} mice compared to wild-type mice, and elevated nuclear factor kappa B (NF{kappa}B) activation was detected in SR-A{sup -/-} macrophages. SR-A deletion increased the production of pro-inflammatory cytokines, and the phosphorylation of mitogen-activated protein kinase (MAPK) and NF{kappa}B in vitro. SR-A deletion also promoted the nuclear translocation of NF{kappa}B and IFN regulatory factor (IRF)-3. In addition, a competitive binding assay with acetylated low-density lipoprotein, an SR-A-specific ligand, and anti-SR-A antibody induced significant activation of TLR4-mediated signaling molecules in wild-type macrophages but not in SR-A{sup -/-} macrophages. These results suggest that SR-A suppresses the macrophage activation by inhibiting the binding of LPS to TLR4 in a competitive manner and it plays a pivotal role in the regulation of the LPS-induced inflammatory response.

  15. Calcium dependency of the AT1-receptor mediated effects in the rat portal vein: influence of calcium antagonists

    NARCIS (Netherlands)

    Zhang, J. S.; van Meel, J. C.; Pfaffendorf, M.; Zhang, J.; van Zwieten, P. A.

    1994-01-01

    The calcium dependency of AT1-receptor mediated contractions was studied in isolated rat portal vein preparations. The spontaneous phasic contractile force of the rat portal vein was increased (ED50 = 1.76 mmol/l) and the frequency of contractions decreased by raising the extracellular calcium

  16. Food-associated estrogenic compounds induce estrogen receptor-mediated luciferase gene expression in transgenic male mice

    NARCIS (Netherlands)

    Veld, ter M.G.R.; Zawadzka, E.; Berg, van den J.H.J.; Saag, van der P.T.; Rietjens, I.M.C.M.; Murk, A.J.

    2008-01-01

    The present paper aims at clarifying to what extent seven food-associated compounds, shown before to be estrogenic in vitro, can induce estrogenic effects in male mice with an estrogen receptor (ER)-mediated luciferase (luc) reporter gene system. The luc induction was determined in different tissues

  17. Food-associated estrogenic compounds induce estrogen receptor-mediated luciferase gene expression in transgenic male mice.

    NARCIS (Netherlands)

    ter Veld, M.G.R.; Zawadzka, E.; van den Berg, J.H.J.; van der Saag, P.T.; Rietjens, I.M.C.M.; Murk, A.J.

    2008-01-01

    The present paper aims at clarifying to what extent seven food-associated compounds, shown before to be estrogenic in vitro, can induce estrogenic effects in male mice with an estrogen receptor (ER)-mediated luciferase (luc) reporter gene system. The luc induction was determined in different tissues

  18. siRNA-mediated silencing of bFGF gene inhibits the proliferation, migration, and invasion of human pituitary adenoma cells.

    Science.gov (United States)

    Zhou, Kai; Fan, Yan-Dong; Duysenbi, Serick; Wu, Peng-Fei; Feng, Zhao-Hai; Qian, Zheng; Zhang, Ting-Rong

    2017-06-01

    Human pituitary adenoma is one of the most common intracranial tumors with an incidence as high as 16.7%. Recent evidence has hinted a relationship between growth factors of pituitary or hypothalamic origin and proliferation of human pituitary adenoma cells. This study explores the effects of small interfering RNA-mediated silencing of basic fibroblast growth factor gene on the proliferation, migration, and invasion of human pituitary adenoma cells. Human pituitary adenoma tissues were collected to obtain human pituitary adenoma cells. The basic fibroblast growth factor silencing interference plasmid was constructed, and the human pituitary adenoma cells were transfected and assigned as basic fibroblast growth factor-small interfering RNA, negative control-small interfering RNA, and blank groups. The quantitative real-time polymerase chain reaction and Western blotting were carried out to detect the expression of basic fibroblast growth factor, pituitary tumor transforming gene, vascular endothelial growth factor, cluster of differentiation 147, and matrix metalloproteinase 9. Cell Counting Kit-8 assay was conducted to observe cell proliferation at 0, 24, 48, and 72 h. Flow cytometry was used to determine cell cycle. Transwell and scratch test were applied to detect the invasion and migration of pituitary adenoma cells. Protein kinase C activity was detected. In comparison with the blank group, the basic fibroblast growth factor-small interfering RNA group showed reduced messenger RNA and protein expression of basic fibroblast growth factor, reduced cell viability at 24, 48, and 72 h, increased cells in G0/G1 stage, declined cells in S and G2/M stages, decreased number of cell migration, shortened migrating distance, reduced protein kinase C activity, and decreased expression of pituitary tumor transforming gene, vascular endothelial growth factor, cluster of differentiation 147, and matrix metalloproteinase 9. However, the negative control-small interfering

  19. The phosphatase calcineurin PP2BAβ mediates part of mineralocorticoid receptor transcriptional activity.

    Science.gov (United States)

    Seiferth, Anja; Ruhs, Stefanie; Mildenberger, Sigrid; Gekle, Michael; Grossmann, Claudia

    2012-06-01

    Recently it was shown that the mineralocorticoid receptor (MR) may exert part of its transcriptional activity by mediation of calcineurin (PP2B). Here we investigated the mechanism of interaction of MR with calcineurin and provide a new MR signaling pathway with potential physiological and pathophysiological relevance. MR → calcineurin crosstalk was assessed in a heterologous expression system (human embryonic kidney cells), which provides the opportunity for detailed mechanistic investigation. SiRNA knockdown experiments show that activated MR, but not GR, reduces CREB- and enhances NFaT-mediated transcriptional activation via the catalytic calcineurin subunit PP2BAβ but not via PP2BAα. Altered PP2BAβ expression, elevated cytosolic Ca(2+), activation of mitogen-activated kinase [p38, extracellular signal-regulated kinase (ERK) 1/2], or protein kinase C do not seem to be involved, whereas inhibition of the chaperone heat-shock protein 90 (HSP90) abrogated the effect of MR. Coimmunoprecipitation indicates the existence of protein complexes harboring MR and PP2BAβ independent of MR activation but dependent on HSP90. Activated MR alters the subcellular distribution of PP2BAβ, enhancing its nuclear fraction, and reduces mRNA expression of the endogenous inhibitor CAIN (calcineurin inhibitor) but not of RCAN1 (regulator of calcineurin). Overall, transcriptional relevant MR → calcineurin crosstalk occurs via the catalytic subunit PP2BAβ, enables glucocorticoid response element-independent genomic signaling of MR, and is of potential pathophysiological relevance. Mechanistically, the crosstalk results from HSP90-mediated cytosolic protein complex formation, altered subcellular distribution, and altered endogenous inhibitor expression.

  20. NMDA Receptors Mediate Stimulus-Timing-Dependent Plasticity and Neural Synchrony in the Dorsal Cochlear Nucleus.

    Science.gov (United States)

    Stefanescu, Roxana A; Shore, Susan E

    2015-01-01

    Auditory information relayed by auditory nerve fibers and somatosensory information relayed by granule cell parallel fibers converge on the fusiform cells (FCs) of the dorsal cochlear nucleus, the first brain station of the auditory pathway. In vitro, parallel fiber synapses on FCs exhibit spike-timing-dependent plasticity with Hebbian learning rules, partially mediated by the NMDA receptor (NMDAr). Well-timed bimodal auditory-somatosensory stimulation, in vivo equivalent of spike-timing-dependent plasticity, can induce stimulus-timing-dependent plasticity (StTDP) of the FCs spontaneous and tone-evoked firing rates. In healthy guinea pigs, the resulting distribution of StTDP learning rules across a FC neural population is dominated by a Hebbian profile while anti-Hebbian, suppressive and enhancing LRs are less frequent. In this study, we investigate in vivo, the NMDAr contribution to FC baseline activity and long term plasticity. We find that blocking the NMDAr decreases the synchronization of FC- spontaneous activity and mediates differential modulation of FC rate-level functions such that low, and high threshold units are more likely to increase, and decrease, respectively, their maximum amplitudes. Three significant alterations in mean learning-rule profiles were identified: transitions from an initial Hebbian profile towards (1) an anti-Hebbian; (2) a suppressive profile; and (3) transitions from an anti-Hebbian to a Hebbian profile. FC units preserving their learning rules showed instead, NMDAr-dependent plasticity to unimodal acoustic stimulation, with persistent depression of tone-evoked responses changing to persistent enhancement following the NMDAr antagonist. These results reveal a crucial role of the NMDAr in mediating FC baseline activity and long-term plasticity which have important implications for signal processing and auditory pathologies related to maladaptive plasticity of dorsal cochlear nucleus circuitry.

  1. Terminalia chebula Fructus Inhibits Migration and Proliferation of Vascular Smooth Muscle Cells and Production of Inflammatory Mediators in RAW 264.7

    Directory of Open Access Journals (Sweden)

    Hyun-Ho Lee

    2015-01-01

    Full Text Available Pathogenesis of atherosclerosis and neointima formation after angioplasty involves vascular smooth muscle cells (VSMCs migration and proliferation followed by inflammatory responses mediated by recruited macrophages in the neointima. Terminalia chebula is widely used traditional medicine in Asia for its beneficial effects against cancer, diabetes, and bacterial infection. The study was designed to determine whether Terminalia chebula fructus water extract (TFW suppresses VSMC migration and proliferation and inflammatory mediators production in macrophage (RAW 264.7. Our results showed that TFW possessed strong antioxidative effects in 1,1-diphenyl-2-picryl hydrazyl (DPPH scavenging and lipid peroxidation assays. In addition, TFW reduced nitric oxide (NO production, inducible nitric oxide synthase (iNOS, and cyclooxygenase-2 (COX-2 expression in RAW 264.7 cells. Also, TFW inhibited platelet-derived growth factor (PDGF-BB induced VSMC migration as determined by wound healing and Boyden chamber assays. The antimigratory effect of TFW was due to its inhibitory effect on metalloproteinase-9 (MMP-9 expression, focal adhesion kinase (FAK activation, and Rho-family of small GTPases (Cdc42 and RhoA expression in VSMCs. Furthermore, TFW suppressed PDGF-BB induced VSMC proliferation by downregulation of mitogen activated protein kinases (MAPKs signaling molecules. These results suggest that TFW could be a beneficial resource in the prevention of atherosclerosis.

  2. The depletion of ATM inhibits colon cancer proliferation and migration via B56γ2-mediated Chk1/p53/CD44 cascades.

    Science.gov (United States)

    Liu, Rui; Tang, Jiajia; Ding, Chaodong; Liang, Weicheng; Zhang, Li; Chen, Tianke; Xiong, Yan; Dai, Xiaowei; Li, Wenfeng; Xu, Yunsheng; Hu, Jin; Lu, Liting; Liao, Wanqin; Lu, Xincheng

    2017-04-01

    Ataxia-telangiectasia mutated (ATM) protein kinase is a major guardian of genomic stability, and its well-established function in cancer is tumor suppression. Here, we report an oncogenic role of ATM. Using two isogenic sets of human colon cancer cell lines that differed only in their ATM status, we demonstrated that ATM deficiency significantly inhibits cancer cell proliferation, migration, and invasion. The tumor-suppressive function of ATM depletion is not modulated by the compensatory activation of ATR, but it is associated with B56γ2-mediated Chk1/p53/CD44 signaling pathways. Under normal growth conditions, the depletion of ATM prevents B56γ2 ubiquitination and degradation, which activates PP2A-mediated Chk1/p53/p21 signaling pathways, leading to senescence and cell cycle arrest. CD44 was validated as a novel ATM target based on its ability to rescue cell migration and invasion defects in ATM-depleted cells. The activation of p53 induced by ATM depletion suppresses CD44 transcription, thus resulting in epithelial-mesenchymal transition (EMT) and cell migration suppression. Our study suggests that ATM has tumorigenic potential in post-formed colon neoplasia, and it supports ATM as an appealing target for improving cancer therapy. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Sphingosine-1-phosphate/sphingosine-1-phosphate receptor 1 signaling is required for migration of naive human T cells from the thymus to the periphery.

    Science.gov (United States)

    Resop, Rachel S; Douaisi, Marc; Craft, Joshua; Jachimowski, Loes C M; Blom, Bianca; Uittenbogaart, Christel H

    2016-08-01

    The mechanisms that govern the egress of mature thymocytes from the human thymus to the periphery remain understudied yet are of utmost importance to the field of basic immunology, as well as T-cell reconstitution in various immunodeficiencies. We examined the expression and function of sphingosine-1-phosphate (S1P) receptors in human thymocyte egress. We aimed to determine whether S1P receptors (S1P-Rs) play a role in mature human thymocyte egress and to identify the thymocyte population or populations that express S1P-Rs and respond to S1P by migrating across a concentration gradient. Human thymocytes were exposed to S1P in Transwell plate migration assays coupled to flow cytometry to evaluate the response to S1P of thymocytes at different stages of maturation. Constitutive S1P-R expression was quantified by means of real-time PCR in sorted thymocyte subsets and flow cytometry. S1P-R1 and Kruppel-like factor 2 expression were monitored after S1P exposure by using flow cytometry and quantitative PCR. S1P-R1 was the prevalent S1P receptor on mature human thymocytes (CD3(hi)CD27(+)CD69(-)), the population that also demonstrated the greatest response to S1P in migration assays. Pretreatment with FTY720, an S1P-R1 nonselective modulator significantly reduced migration and suggested a role for S1P-R2 in retaining thymocytes in the tissue. Lastly, surface S1P-R1 expression, as well S1PR1 and Kruppel-like factor 2 (KLF2) transcripts, were significantly decreased in mature thymocytes on exposure to S1P. Mature human thymocytes rely on S1P-R1 to migrate toward S1P. Taken in the context of murine work demonstrating that S1P is required for thymocyte egress to the periphery, our data highlight a new key chemokine for human thymocyte egress. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  4. A new CysLT1 and CysLT2 receptors-mediated anaphylaxis guinea pig model.

    Science.gov (United States)

    Sekioka, Tomohiko; Kadode, Michiaki; Osakada, Noriko; Fujita, Manabu; Matsumura, Naoya; Yamaura, Yoshiyuki; Nakade, Shinji; Nabe, Takeshi; Kawabata, Kazuhito

    2017-04-01

    Although the effectiveness of CysLT1 receptor antagonists on asthma has been clinically established, the effects of CysLT2 receptor antagonists are still unclear. The purpose of this study was to develop a new CysLT1 and CysLT2 receptors-mediated anaphylaxis guinea pig model using S-hexyl GSH, a γ-glutamyl transpeptidase (GTP) inhibitor, to suppress conversion of LTC4 to LTD4. Actively sensitized guinea pigs were challenged with OVA in the absence or presence of S-hexyl GSH, and survival rate following anaphylactic response was monitored. OVA-induced fatal anaphylaxis in the absence of S-hexyl GSH was almost completely inhibited by montelukast, a CysLT1 receptor antagonist, but not by the CysLT2 receptor antagonist BayCysLT2RA. However, under treatment with S-hexyl-GSH, the inhibitory effect of motelukast was dramatically diminished, whereas that of BayCysLT2RA was markedly increased. The dual CysLT1/2 receptor antagonist ONO-6950 effectively inhibited anaphylactic response in both S-hexyl GSH-treated and non-treated animals. LC/MS/MS analysis revealed that S-hexyl GSH treatment actually inhibited LTC4 metabolism in the blood and lung tissues. Using S-hexyl GSH, we developed a novel CysLT1 and CysLT2 receptors-mediated anaphylaxis guinea pig model that can be useful for not only screening both CysLT2 and CysLT1/2 receptors antagonists, but also for functional analysis of CysLT2 receptors. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. A role of periaqueductal grey NR2B-containing NMDA receptor in mediating persistent inflammatory pain

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    Yang Qi

    2009-12-01

    Full Text Available Abstract The midbrain periaqueductal grey (PAG is a structure known for its roles in pain transmission and modulation. Noxious stimuli potentiate the glutamate synaptic transmission and enhance glutamate NMDA receptor expression in the PAG. However, little is known about roles of NMDA receptor subunits in the PAG in processing the persistent inflammatory pain. The present study was undertaken to investigate NR2A- and NR2B-containing NMDA receptors in the PAG and their modulation to the peripheral painful inflammation. Noxious stimuli induced by hind-paw injection of complete Freund's adjuvant (CFA caused up-regulation of NR2B-containing NMDA receptors in the PAG, while NR2A-containing NMDA receptors were not altered. Whole-cell patch-clamp recordings revealed that NMDA receptor mediated mEPSCs were increased significantly in the PAG synapse during the chronic phases of inflammatory pain in mice. PAG local infusion of Ro 25-6981, an NR2B antagonist, notably prolonged the paw withdrawal latency to thermal radian heat stimuli bilaterally in rats. Hyperoside (Hyp, one of the flavonoids compound isolated from Rhododendron ponticum L., significantly reversed up-regulation of NR2B-containing NMDA receptors in the PAG and exhibited analgesic activities against persistent inflammatory stimuli in mice. Our findings provide strong evidence that up-regulation of NR2B-containing NMDA receptors in the PAG involves in the modulation to the peripheral persistent inflammatory pain.

  6. Leukotriene D4 receptor-mediated hydrolysis of phosphoinositide and mobilization of calcium in sheep tracheal smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Mong, S.; Miller, J.; Wu, H.L.; Crooke, S.T.

    1988-02-01

    A sheep tracheal smooth muscle primary culture cell system was developed to characterize leukotriene D4 (LTD4) receptor-mediated biochemical and pharmacological effects. (/sup 3/H)LTD4 binding to the enriched plasma membrane receptor was specific, stereoselective and saturable. LTE4 and high affinity receptor antagonists bound to the receptors with a rank-order potency that was expected from previous smooth muscle contraction studies. In the (/sup 3/H)myoinositol labeled cells, LTD4 and LTE4 induced phosphoinositide hydrolysis. The biosynthesis of (/sup 3/H)inositol-trisphosphate was rapid and the induction of biosynthesis of (/sup 3/H)inositol-monophosphate by LTs was stereoselective and specific and was inhibited specifically by a receptor antagonist, SKF 104353. In the fura-2 loaded smooth muscle cells, LTD4 and LTE4 induced transient intracellular Ca++ mobilization. The fura-2/Ca++ transient was stereoselective and specific and was inhibited by receptor antagonist, SKF 104353. These results suggest that the cultured sheep tracheal smooth muscle cells have plasma membrane receptors for LTD4. These receptors were coupled to a phospholipase C that, when activated by agonists, induced hydrolysis of inositol containing phospholipids. The hydrolysis products, e.g. diacylglycerol and inositol-trisphosphate, may serve as intracellular messengers that trigger or contribute to the contractile effect in sheep tracheal smooth muscle.

  7. NR2B-NMDA receptor mediated modulation of the tyrosine phosphatase STEP regulates glutamate induced neuronal cell death

    Science.gov (United States)

    Poddar, Ranjana; Deb, Ishani; Mukherjee, Saibal; Paul, Surojit

    2011-01-01

    The present study examines the role of a neuron-specific tyrosine phosphatase (STEP) in excitotoxic cell death. Our findings demonstrate that p38 MAPK, a stress-activated kinase that is known to play a role in the etiology of excitotoxic cell death is a substrate of STEP. Glutamate-mediated NMDA receptor stimulation leads to rapid but transient activation of p38 MAPK, which is primarily dependent on NR2A-NMDA receptor activation. Conversely, activation of NR2B-NMDA receptors leads to dephosphorylation and subsequent activation of STEP, which in turn leads to inactivation of p38 MAPK. Thus during transient NMDA receptor stimulation, increases in STEP activity appears to limit the duration of activation of p38 MAPK and improves neuronal survival. However, if NR2B-NMDA receptor stimulation is sustained, protective effects of STEP activation are lost, as these stimuli cause significant degradation of active STEP, leading to secondary activation of p38 MAP kinase. Consistent with this observation, a cell transducible TAT-STEP peptide that constitutively binds to p38 MAPK attenuated neuronal cell death caused by sustained NMDA receptor stimulation. The findings imply that the activation and levels of STEP are dependent on the duration and magnitude of NR2B-NMDA receptor stimulation and STEP serves as a modulator of NMDA receptor dependent neuronal injury, through its regulation of p38 MAPK. PMID:21029094

  8. FYVE1/FREE1 Interacts with the PYL4 ABA Receptor and Mediates its Delivery to the Vacuolar Degradation Pathway.

    Science.gov (United States)

    Belda-Palazon, Borja; Rodriguez, Lesia; Fernandez, Maria A; Castillo, Mari-Cruz; Anderson, Erin A; Gao, Caiji; González-Guzmán, Miguel; Peirats-Llobet, Marta; Zhao, Qiong; De Winne, Nancy; Gevaert, Kris; De Jaeger, Geert; Jiang, Liwen; Leon, Jose; Mullen, Robert T; Rodriguez, Pedro L

    2016-08-05

    Recently, we described the ubiquitylation of PYL4 and PYR1 by the RING E3 ubiquitin ligase RSL1 at the plasma membrane of Arabidopsis thaliana. This suggested that ubiquitylated ABA receptors might be targeted to the vacuolar degradation pathway because such ubiquitylation is usually an internalization signal for the endocytic route. Here, we show that FYVE1 (previously termed FREE1), a recently described component of the endosomal sorting complex required for transport (ESCRT) machinery, interacted with RSL1-receptor complexes and recruited PYL4 to endosomal compartments. Although the ESCRT pathway has been assumed to be reserved for integral membrane proteins, we show the involvement of this pathway in the degradation of ABA receptors, which can be associated with membranes but are not integral membrane proteins. Knock-down fyve1 alleles are hypersensitive to ABA, illustrating the biological relevance of the ESCRT pathway for the modulation of ABA signaling. In addition, fyve1 mutants are impaired in the targeting of ABA receptors for vacuolar degradation, leading to increased accumulation of PYL4 and an enhanced response to ABA. Pharmacological and genetic approaches revealed a dynamic turnover of ABA receptors from the plasma membrane to the endosomal/vacuolar degradation pathway, which was mediated by FYVE1 and was dependent on RSL1. This process involves clathrin-mediated endocytosis and trafficking of PYL4 through the ESCRT pathway, which helps to regulate the turnover of ABA receptors and attenuate ABA signaling. © 2016 American Society of Plant Biologists. All rights reserved.

  9. A peptide antagonist of the ErbB1 receptor inhibits receptor activation, tumor cell growth and migration in vitro and xenograft tumor growth in vivo

    DEFF Research Database (Denmark)

    Xu, Ruodan; Povlsen, Gro Klitgaard; Soroka, Vladislav

    2010-01-01

    The epidermal growth factor family of receptor tyrosine kinases (ErbBs) plays essential roles in tumorigenesis and cancer disease progression, and therefore has become an attractive target for structure-based drug design. ErbB receptors are activated by ligand-induced homo- and heterodimerization...... lung cancer cell line A549. The Inherbin3 peptide may be a useful tool for investigating the mechanisms of ErbB receptor homo- and heterodimerization. Moreover, the here described biological effects of Inherbin3 suggest that peptide-based targeting of ErbB receptor dimerization is a promising anti-cancer...

  10. IL-1β Induces MMP-9-Dependent Brain Astrocytic Migration via Transactivation of PDGF Receptor/NADPH Oxidase 2-Derived Reactive Oxygen Species Signals.

    Science.gov (United States)

    Yang, Chuen-Mao; Hsieh, Hsi-Lung; Yu, Ping-Hsien; Lin, Chih-Chung; Liu, Shiau-Wen

    2015-08-01

    Matrix metalloproteinase-9 (MMP-9) plays a crucial role in pathological processes of brain inflammation, injury, and neurodegeneration. Moreover, cytokines such as interleukin-1β (IL-1β) induce expression of several inflammatory mediators in brain astrocytes, which may be important for brain inflammatory disorders. Recent studies have implicated that increased oxidative stress may contribute to the brain injury and inflammation. However, whether IL-1β-induced MMP-9 expression mediated through oxidative stress remains unclear. Therefore, we investigated the role of redox signals in IL-1β-induced MMP-9 expression in rat brain astrocytes (RBA-1 cells). Herein, we first demonstrated that reactive oxygen species (ROS) play a crucial role in ILβ-induced MMP-9 expression by zymography, real-time PCR, and ROS staining in cultured RBA-1 cells. Next, IL-1β-induced MMP-9 expression is mediated through a c-Src-mediated transactivation of PDGFR/PI3K/Akt cascade linking to p47(phox)/NADPH oxidase 2 (Nox2)/ROS signaling pathway. Nox2-dependent ROS generation led to activation of MAPKs and the downstream transcription factors NF-κB and AP-1 (i.e., ATF2), which enhanced MMP-9 promoter activity, and thereby turned on transcription of MMP-9 gene. Functionally, IL-1β-induced MMP-9 expression promoted astrocytic migration. These results demonstrated that in RBA-1 cells, activation of NF-κB and AP-1 (ATF2) by the c-Src/PDGFR/PI3K/Akt-mediated Nox2/ROS/MAPKs signals is required for upregulation of MMP-9 and cell migration enhanced by IL-1β.

  11. GABA(A) receptor pi (GABRP) stimulates basal-like breast cancer cell migration through activation of extracellular-regulated kinase 1/2 (ERK1/2).

    Science.gov (United States)

    Sizemore, Gina M; Sizemore, Steven T; Seachrist, Darcie D; Keri, Ruth A

    2014-08-29

    Breast cancer is a heterogeneous disease comprised of distinct subtypes predictive of patient outcome. Tumors of the basal-like subtype have a poor prognosis due to inherent aggressiveness and the lack of targeted therapeutics. Basal-like tumors typically lack estrogen receptor-α, progesterone receptor and HER2/ERBB2, or in other words they are triple negative (TN). Continued evaluation of basal-like breast cancer (BLBC) biology is essential to identify novel therapeutic targets. Expression of the pi subunit of the GABA(A) receptor (GABRP) is associated with the BLBC/TN subtype, and herein, we reveal its expression also correlates with metastases to the brain and poorer patient outcome. GABRP expression in breast cancer cell lines also demonstrates a significant correlation with the basal-like subtype suggesting that GABRP functions in the initiation and/or progression of basal-like tumors. To address this postulate, we stably silenced GABRP in two BLBC cell lines, HCC1187 and HCC70 cells. Decreased GABRP reduces in vitro tumorigenic potential and migration concurrent with alterations in the cytoskeleton, specifically diminished cellular protrusions and expression of the BLBC-associated cytokeratins, KRT5, KRT6B, KRT14, and KRT17. Silencing GABRP also decreases phosphorylation of extracellular regulated kinase 1/2 (ERK1/2) in both cell lines and selective inhibition of ERK1/2 similarly decreases the basal-like cytokeratins as well as migration. Combined, these data reveal a GABRP-ERK1/2-cytokeratin axis that maintains the migratory phenotype of basal-like breast cancer. GABRP is a component of a cell surface receptor, thus, these findings suggest that targeting this new signaling axis may have therapeutic potential in BLBC. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Src Is a Potential Therapeutic Target in Endocrine-Resistant Breast Cancer Exhibiting Low Estrogen Receptor-Mediated Transactivation.

    Directory of Open Access Journals (Sweden)

    Stephanie K Guest

    Full Text Available Despite the effectiveness of endocrine therapies in estrogen receptor positive (ER+ breast cancer, approximately 40% of patients relapse. Previously, we identified the Focal-adhesion kinase canonical pathway as a major contributor of resistance to estrogen deprivation and cellular-sarcoma kinase (c-src as a dominant gene in this pathway. Dasatinib, a pan-src inhibitor, has recently been used in clinical trials to treat ER+ patients but has shown mixed success. In the following study, using isogenic cell line models, we provide a potential explanation for these findings and suggest a sub-group that may benefit. A panel of isogenic cell lines modelling resistance to aromatase inhibitors (LTED and tamoxifen (TAMR were assessed for response to dasatinib ± endocrine therapy. Dasatinib caused a dose-dependent decrease in proliferation in MCF7-TAMR cells and resensitized them to tamoxifen and fulvestrant but not in HCC1428-TAMR. In contrast, in estrogen-deprived conditions, dasatinib increased the proliferation rate of parental-MCF7 cells and had no effect on MCF7-LTED or HCC1428-LTED. Treatment with dasatinib caused a decrease in src-phosphorylation and inhibition of downstream pathways, including AKT and ERK1/2 in all cell lines tested, but only the MCF7-TAMR showed a concomitant decrease in markers of cell cycle progression. Inhibition of src also caused a significant decrease in cell migration in both MCF7-LTED and MCF7-TAMR cells. Finally, we showed that, in MCF7-TAMR cells, in contrast to tamoxifen sensitive cell lines, ER is expressed throughout the cell rather than being restricted to the nucleus and that treatment with dasatinib resulted in nuclear shuttling of ER, which was associated with an increase in ER-mediated transcription. These data suggest that src has differential effects in endocrine-resistant cell lines, particularly in tamoxifen resistant models, with low ER genomic activity, providing further evidence of the importance of

  13. Evidence of a Cell Surface Role for Hsp90 Complex Proteins Mediating Neuroblast Migration in the Subventricular Zone

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    Leo M. Miyakoshi

    2017-05-01

    Full Text Available In most mammalian brains, the subventricular zone (SVZ is a germinative layer that maintains neurogenic activity throughout adulthood. Neuronal precursors arising from this region migrate through the rostral migratory stream (RMS and reach the olfactory bulbs where they differentiate and integrate into the local circuitry. Recently, studies have shown that heat shock proteins have an important role in cancer cell migration and blocking Hsp90 function was shown to hinder cell migration in the developing cerebellum. In this work, we hypothesize that chaperone complexes may have an important function regulating migration of neuronal precursors from the subventricular zone. Proteins from the Hsp90 complex are present in the postnatal SVZ as well as in the RMS. Using an in vitro SVZ explant model, we have demonstrated the expression of Hsp90 and Hop/STI1 by migrating neuroblasts. Treatment with antibodies against Hsp90 and co-chaperone Hop/STI1, as well as Hsp90 and Hsp70 inhibitors hinder neuroblast chain migration. Time-lapse videomicroscopy analysis revealed that cell motility and average migratory speed was decreased after exposure to both antibodies and inhibitors. Antibodies recognizing Hsp90, Hsp70, and Hop/STI1 were found bound to the membranes of cells from primary SVZ cultures and biotinylation assays demonstrated that Hsp70 and Hop/STI1 could be found on the external leaflet of neuroblast membranes. The latter could also be detected in conditioned medium samples obtained from cultivated SVZ cells. Our results suggest that chaperones Hsp90, Hsp70, and co-chaperone Hop/STI1, components of the Hsp90 complex, regulate SVZ neuroblast migration in a concerted manner through an extracellular mechanism.

  14. Kinetic analysis of receptor-mediated endocytosis of G-CSF derivative, nartograstim, in rat bone marrow cells.

    Science.gov (United States)

    Kuwabara, T; Kobayashi, S; Sugiyama, Y

    1996-07-01

    To elucidate the mechanism of the receptor-mediated clearance of granulocyte colony-stimulating factor (G-CSF), we performed kinetic analyses of the receptor-mediated endocytosis (RME) processes using a human G-CSF derivative, nartograstim (NTG), and isolated rat bone marrow cells. The first-order rate constants involved in RME processes were obtained by computerized model fitting of the time courses of the ligand-receptor complex on both the cell surface and in the cell interior and the degradation products in the medium in the pulse-chase experiment. They were also calculated based on a kinetic model involving the ligand concentration dependence of the initial binding rate, the steady-state degradation rate, and the steady-state amounts of ligand on both the cell surface and in the interior. The rate constants for the RME processes after receptor binding determined in the different experiments were similar, that is, the half-times for the dissociation, internalization, and degradation of the ligand-receptor complex were 770, 10-30, and 20 min, respectively. However, the association constant obtained by measuring the initial binding was fivefold greater than that calculated under steady-state conditions. These kinetic analyses support the hypothesis that the internalization of the receptor may be accelerated by ligand binding, causing downregulation of the receptor on the cell surface. These overall kinetic analyses based on steady-state and non-steady-state data of the RME processes clarify the dynamics of the interaction between NTG and its receptor.

  15. Loss of progesterone receptor-mediated actions induce preterm cellular and structural remodeling of the cervix and premature birth.

    Directory of Open Access Journals (Sweden)

    Steven M Yellon

    Full Text Available A decline in serum progesterone or antagonism of progesterone receptor function results in preterm labor and birth. Whether characteristics of premature remodeling of the cervix after antiprogestins or ovariectomy are similar to that at term was the focus of the present study. Groups of pregnant rats were treated with vehicle, a progesterone receptor antagonist (onapristone or mifepristone, or ovariectomized on day 17 postbreeding. As expected, controls given vehicle delivered at term while rats delivered preterm after progesterone receptor antagonist treatment or ovariectomy. Similar to the cervix before term, the preterm cervix of progesterone receptor antagonist-treated rats was characterized by reduced cell nuclei density, decreased collagen content and structure, as well as a greater presence of macrophages per unit area. Thus, loss of nuclear progesterone receptor-mediated actions promoted structural remodeling of the cervix, increased census of resident macrophages, and preterm birth much like that found in the cervix at term. In contrast to the progesterone receptor antagonist-induced advance in characteristics associated with remodeling, ovariectomy-induced loss of systemic progesterone did not affect hypertrophy, extracellular collagen, or macrophage numbers in the cervix. Thus, the structure and macrophage census in the cervix appear sufficient for premature ripening and birth to occur well before term. With progesterone receptors predominantly localized on cells other than macrophages, the findings suggest that interactions between cells may facilitate the loss of progesterone receptor-mediated actions as part of a final common mechanism that remodels the cervix in certain etiologies of preterm and with parturition at term.

  16. 6-Phosphogluconate dehydrogenase regulates tumor cell migration in vitro by regulating receptor tyrosine kinase c-Met

    Energy Technology Data Exchange (ETDEWEB)

    Chan, Barden, E-mail: cchan@bidmc.harvard.edu [Division of Interdisciplinary Medicine and Biotechnology, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215 (United States); VanderLaan, Paul A. [Department of Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215 (United States); Sukhatme, Vikas P. [Division of Interdisciplinary Medicine and Biotechnology, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215 (United States)

    2013-09-20

    Highlights: •Expression of 6PGD positively correlates with advancing stage of lung carcinoma. •Knockdown of 6PGD by shRNA potently inhibits c-Met tyrosine phosphorylation. •Exogenous HGF fails to restore c-Met phosphorylation in cells with 6PGD knocked down. •6PGD knockdown results in inhibition of cell migration in vitro. •Constitutively active TPR-cMet significantly restores migration of cells without 6PGD. -- Abstract: 6-Phosphogluconate dehydrogenase (6PGD) is the third enzyme in the oxidative pentose phosphate pathway (PPP). Recently, we reported that knockdown of 6PGD inhibited lung tumor growth in vitro and in a xenograft model in mice. In this study, we continued to examine the functional role of 6PGD in cancer. We show that 6PGD expression positively correlates with advancing stage of lung carcinoma. In search of functional signals related to 6PGD, we discovered that knockdown of 6PGD significantly inhibited phosphorylation of c-Met at tyrosine residues known to be critical for activity. This downregulation of c-Met phosphorylation correlated with inhibition of cell migration in vitro. Overexpression of a constitutively active c-Met specifically rescued the migration but not proliferation phenotype of 6PGD knockdown. Therefore, 6PGD appears to be required for efficient c-Met signaling and migration of tumor cells in vitro.

  17. Differential inhibitory effects of CysLT(1 receptor antagonists on P2Y(6 receptor-mediated signaling and ion transport in human bronchial epithelia.

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    Wendy Ka-hoi Lau

    Full Text Available BACKGROUND: Cysteinyl leukotriene (CysLT is one of the proinflammatory mediators released by the bronchi during inflammation. CysLTs exert their biological effects via specific G-protein-coupled receptors. CysLT(1 receptor antagonists are available for clinical use for the treatment of asthma. Recently, crosstalk between CysLT(1 and P2Y(6 receptors has been delineated. P2Y receptors are expressed in apical and/or basolateral membranes of virtually all polarized epithelia to control the transport of fluid and electrolytes. Previous research suggests that CysLT(1 receptor antagonists inhibit the effects of nucleotides acting at P2Y receptors. However, the detailed molecular mechanism underlying the inhibition remains unresolved. METHODOLOGY/PRINCIPAL FINDINGS: In this study, western blot analysis confirmed that both CysLT(1 and P2Y(6 receptors were expressed in the human bronchial epithelial cell line 16HBE14o-. All three CysLT(1 antagonists inhibited the uridine diphosphate (UDP-evoked I(SC, but only montelukast inhibited the UDP-evoked [Ca(2+](i increase. In the presence of forskolin or 8-bromoadenosine 3'5' cyclic monophosphate (8-Br-cAMP, the UDP-induced I(SC was potentiated but was reduced by pranlukast and zafirlukast but not montelukast. Pranlukast inhibited the UDP-evoked I(SC potentiated by an Epac activator, 8-(4-Chlorophenylthio-2'-O-methyladenosine-3',5'-cyclic monophosphate (8-CPT-2'-O-Me-cAMP, while montelukast and zafirlukast had no such effect. Pranlukast inhibited the real-time increase in cAMP changes activated by 8-CPT-2'-O-Me-cAMP as monitored by fluorescence resonance energy transfer imaging. Zafirlukast inhibited the UDP-induced I(SC potentiated by N(6-Phenyladenosine-3',5'-cyclic monophosphorothioate, Sp-isomer (Sp-6-Phe-cAMP; a PKA activator and UDP-activated PKA activity. CONCLUSIONS/SIGNIFICANCE: In summary, our data strongly suggest for the first time that in human airway epithelia, the three specific CysLT(1 receptor

  18. Ciliary neurotrophic factor mediates dopamine D2 receptor-induced CNS neurogenesis in adult mice.

    Science.gov (United States)

    Yang, Peng; Arnold, Sheila A; Habas, Agata; Hetman, Michal; Hagg, Theo

    2008-02-27

    Neurogenesis continues in the adult forebrain subventricular zone (SVZ) and the dentate gyrus of the hippocampal formation. Degeneration of dopaminergic projections in Parkinson's disease and animals reduces, whereas ciliary neurotrophic factor (CNTF) promotes, neurogenesis. We tested whether the dopaminergic system promotes neurogenesis through CNTF. Astrocytes of the SVZ and dentate gyrus expressed CNTF and were close to dopaminergic terminals. Dopaminergic denervation in adult mice reduced CNTF mRNA by approximately 60%, whereas systemic treatment with the D2 agonist quinpirole increased CNTF mRNA in the SVZ and hippocampal formation, and in cultured astrocytes by 1.5-5 fold. The effect of quinpirole in vitro was blocked by the D2 antagonist eticlopride and did not cause astroglial proliferation or hypertrophy. Systemic quinpirole injections increased proliferation in wild-type mice by approximately 25-75% but not in CNTF-/- littermates or in the SVZ of mice infused with CNTF antibodies. Quinpirole increased the number of neuroblasts in wild-type but not in CNTF-/- littermates. Neurogenesis was reduced by approximately 20% in CNTF-/- mice, confirming the endogenous role of CNTF. Nigrostriatal denervation did not affect SVZ proliferation in CNTF-/- mice, suggesting that the dopaminergic innervation normally regulates neurogenesis through CNTF. Quinpirole acted on postsynaptic receptors as it reversed the reduced proliferation seen after dopaminergic denervation in wild-type mice. Thus, CNTF mediates dopaminergic innervation- and D2 receptor-induced neurogenesis in the adult forebrain. Because CNTF is predominantly expressed in the nervous system, this mechanism and the ability to pharmacologically modulate it have implications for Parkinson's disease and cell-replacement therapies for other disorders.

  19. Caveolae are essential for angiotensin II type 1 receptor-mediated ANP secretion.

    Science.gov (United States)

    Oh, Young-Bin; Gao, Shan; Lim, Jung Min; Kim, Hyung Tae; Park, Byung-Hyun; Kim, Suhn Hee

    2011-07-01

    Caveolae may act as mechanosensors and function as binding sites for calcium ions. The intracaveolar localization of atrial natriuretic peptide (ANP) derived from the direct interaction of atrial granules with caveolae has been demonstrated. The aim of this study was to define the effect of caveolae on ANP secretion induced by stretch and angiotensin II. The isolated perfused beating atria from Sprague-Dawley rats were used. To disrupt caveolae, 10mM methyl-β-cyclodextrin (MbCD) was applied for 1h and the number of caveoli were markedly decreased. MbCD increased basal ANP secretion and atrial diastolic pressure. The molecular profile of ANP in perfusate from control atria showed mainly one major peak corresponded to synthetic ANP whereas that from MbCD-treated atria showed two major immunoreactive peaks corresponded to synthetic rat ANP and proANP. High atrial stretch induced by elevating the height of outflow catheter from 5 cm H₂O to 7.5 cm H₂O increased atrial contractility and ANP secretion. The response of ANP secretion to high stretch was attenuated in MbCD-pretreated atria. Pretreatment with MbCD abolished angiotensin II-induced suppression and losartan-induced stimulation of ANP secretion. However, the effect of angiotenisin (1-7) on ANP secretion was not altered by MbCD treatment. The expression of angiotensin II type 1 receptor protein was reduced by MbCD treatment. These data suggest that caveolae are essential for angiotensin II type 1 receptor-mediated ANP secretion and relate to the processing of proANP. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Rapid effects of phytoestrogens on human colonic smooth muscle are mediated by oestrogen receptor beta.

    LENUS (Irish Health Repository)

    Hogan, A M

    2012-02-01

    Epidemiological studies have correlated consumption of dietary phytoestrogens with beneficial effects on colon, breast and prostate cancers. Genomic and non-genomic mechanisms are responsible for anti-carcinogenic effects but, until now, the effect on human colon was assumed to be passive and remote. No direct effect on human colonic smooth muscle has previously been described. Institutional research board approval was granted. Histologically normal colon was obtained from the proximal resection margin of colorectal carcinoma specimens. Circular smooth muscle strips were microdissected and suspended under 1g of tension in organ baths containing oxygenated Krebs solution at 37 degrees C. After an equilibration period, tissues were exposed to diarylpropionitrile (DPN) (ER beta agonist) and 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole (PPT) (ER alpha agonist) or to the synthetic phytoestrogen compounds genistein (n=8), daidzein (n=8), fisetin (n=8) and quercetin (n=8) in the presence or absence of fulvestrant (oestrogen receptor antagonist). Mechanism of action was investigated by inhibition of downstream pathways. The cholinergic agonist carbachol was used to induce contractile activity. Tension was recorded isometrically. Phytoestrogens inhibit carbachol-induced colonic contractility. In keeping with a non-genomic, rapid onset direct action, the effect was within minutes, reversible and similar to previously described actions of 17 beta oestradiol. No effect was seen in the presence of fulvestrant indicating receptor modulation. While the DPN exerted inhibitory effects, PPT did not. The effect appears to be reliant on a p38\\/mitogen activated protein kinase mediated induction of nitric oxide production in colonic smooth muscle. The present data set provides the first description of a direct effect of genistein, daidzein, fisetin and quercetin on human colonic smooth muscle. The presence of ER in colonic smooth muscle has been functionally proven and the beta

  1. Nicotinic receptors and lurasidone-mediated reversal of phencyclidine-induced deficit in novel object recognition.

    Science.gov (United States)

    Miyauchi, Masanori; Neugebauer, Nichole M; Oyamada, Yoshihiro; Meltzer, Herbert Y

    2016-03-15

    Enhancement of cholinergic function via nicotinic acetylcholine (ACh) receptor (nAChR) agonism is a potential approach for the treatment of cognitive impairment associated with schizophrenia (CIAS). Some atypical antipsychotic drugs (AAPDs) enhance ACh release in rodent brain, indirectly stimulating these receptors. Here, we elucidate which nAChR subtypes mediate novel object recognition (NOR) in normal rats and contribute to the ability of the AAPD, lurasidone, to improve the NOR deficit in sub-chronic (sc) phencyclidine (PCP)-treated rats, a model for CIAS. The ability of lurasidone and nAChR ligands to reverse the scPCP-induced deficit in NOR was assessed in female, Long-Evans rats. The broad acting nAChR antagonist, mecamylamine (MEC), induced a NOR deficit in normal rats. The NOR deficit secondary to scPCP was reversed by either selective α4β2* nAChR agonism (A-85380) or α7 nAChRs agonism (PNU-282987); these effects were blocked by DHβE and MLA, selective antagonists of α4β2* and α7 nAChR, respectively. The ability of lurasidone to reverse the scPCP-induced NOR deficit was blocked by MEC, but not MLA or DHβE. However, sub-effective doses (SED) of either A-85380 or PNU-282987 potentiated the ability of SED lurasidone to reverse the scPCP-induced NOR deficit. These results identify both α4β2* and α7 nAChRs as candidates for enhancing the ability of lurasidone and other AAPDs, which increase the release of ACh, to improve CIAS. Copyright © 2016. Published by Elsevier B.V.

  2. The Mannose Receptor in Regulation of Helminth-Mediated Host Immunity

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    Irma van Die

    2017-11-01

    Full Text Available Infection with parasitic helminths affects humanity and animal welfare. Parasitic helminths have the capacity to modulate host immune responses to promote their survival in infected hosts, often for a long time leading to chronic infections. In contrast to many infectious microbes, however, the helminths are able to induce immune responses that show positive bystander effects such as the protection to several immune disorders, including multiple sclerosis, inflammatory bowel disease, and allergies. They generally promote the generation of a tolerogenic immune microenvironment including the induction of type 2 (Th2 responses and a sub-population of alternatively activated macrophages. It is proposed that this anti-inflammatory response enables helminths to survive in their hosts and protects the host from excessive pathology arising from infection with these large pathogens. In any case, there is an urgent need to enhance understanding of how helminths beneficially modulate inflammatory reactions, to identify the molecules involved and to promote approaches to exploit this knowledge for future therapeutic interventions. Evidence is increasing that C-type lectins play an important role in driving helminth-mediated immune responses. C-type lectins belong to a large family of calcium-dependent receptors with broad glycan specificity. They are abundantly present on immune cells, such as dendritic cells and macrophages, which are essential in shaping host immune responses. Here, we will focus on the role of the C-type lectin macrophage mannose receptor (MR in helminth–host interactions, which is a critically understudied area in the field of helminth immunobiology. We give an overview of the structural aspects of the MR including its glycan specificity, and the functional implications of the MR in helminth–host interactions focusing on a few selected helminth species.

  3. Polyclonal HER2-specific antibodies induced by vaccination mediate receptor internalization and degradation in tumor cells

    Science.gov (United States)

    2012-01-01

    Introduction Sustained HER2 signaling at the cell surface is an oncogenic mechanism in a significant proportion of breast cancers. While clinically effective therapies targeting HER2 such as mAbs and tyrosine kinase inhibitors exist, tumors overexpressing HER2 eventually progress despite treatment. Thus, abrogation of persistent HER2 expression at the plasma membrane to synergize with current approaches may represent a novel therapeutic strategy. Methods We generated polyclonal anti-HER2 antibodies (HER2-VIA) by vaccinating mice with an adenovirus expressing human HER2, and assessed their signaling effects in vitro and anti-tumor effects in a xenograft model. In addition, we studied the signaling effects of human HER2-specific antibodies induced by vaccinating breast cancer patients with a HER2 protein vaccine. Results HER2-VIA bound HER2 at the plasma membrane, initially activating the downstream kinases extracellular signal-regulated protein kinase 1/2 and Akt, but subsequently inducing receptor internalization in clathrin-coated pits in a HER2 kinase-independent manner, followed by ubiquitination and degradation of HER2 into a 130 kDa fragment phosphorylated at tyrosine residues 1,221/1,222 and 1,248. Following vaccination of breast cancer patients with the HER2 protein vaccine, HER2-specific antibodies were detectable and these antibodies bound to cell surface-expressed HER2 and inhibited HER2 signaling through blocking tyrosine 877 phosphorylation of HER2. In contrast to the murine antibodies, human anti-HER2 antibodies induced by protein vaccination did not mediate receptor internalization and degradation. Conclusion These data provide new insight into HER2 trafficking at the plasma membrane and the changes induced by polyclonal HER2-specific antibodies. The reduction of HER2 membrane expression and HER2 signaling by polyclonal antibodies induced by adenoviral HER2 vaccines supports human clinical trials with this strategy for those breast cancer patients

  4. Formation of a Protein Corona on Silver Nanoparticles Mediates Cellular Toxicity via Scavenger Receptors

    Science.gov (United States)

    Shannahan, Jonathan H.; Podila, Ramakrishna; Aldossari, Abdullah A.; Emerson, Hilary; Powell, Brian A.; Ke, Pu Chun; Rao, Apparao M.; Brown, Jared M.

    2015-01-01

    Addition of a protein corona (PC) or protein adsorption layer on the surface of nanomaterials following their introduction into physiological environments may modify their activity, bio-distribution, cellular uptake, clearance, and toxicity. We hypothesize that silver nanoparticles (AgNPs) will associate with proteins common to human serum and cell culture media forming a PC that will impact cell activation and cytotoxicity. Furthermore, the role of scavenger receptor BI (SR-BI) in mediating this toxicity was evaluated. Citrate-suspended 20 nm AgNPs were incubated with human serum albumin (HSA), bovine serum albumin (BSA), high-density lipoprotein (HDL), or water (control) to form a PC. AgNPs associated with each protein (HSA, BSA, and HDL) forming PCs as assessed by electron microscopy, hyperspectral analysis, ζ-potential, and hydrodynamic size. Addition of the PC decreased uptake of AgNPs by rat lung epithelial and rat aortic endothelial cells. Hyperspectral analysis demonstrated a loss of the AgNP PC following internalization. Cells demonstrated concentration-dependent cytotoxicity following exposure to AgNPs with or without PCs (0, 6.25, 12.5, 25 or 50 μg/ml). All PC-coated AgNPs were found to activate cells by inducing IL-6 mRNA expression. A small molecule SR-BI inhibitor was utilized to determine the role of SR-BI in the observed effects. Pretreatment with the SR-BI inhibitor decreased internalization of AgNPs with or without PCs, and reduced both cytotoxicity and IL-6 mRNA expression. This study characterizes the formation of a PC on AgNPs and demonstrates its influence on cytotoxicity and cell activation through a cell surface receptor. PMID:25326241

  5. Kainate receptor-mediated modulation of hippocampal fast spiking interneurons in a rat model of schizophrenia.

    Directory of Open Access Journals (Sweden)

    Barbara Gisabella

    Full Text Available Kainate receptor (KAR subunits are believed to be involved in abnormal GABAergic neurotransmission in the hippocampus (HIPP in schizophrenia (SZ and bipolar disorder. Postmortem studies have shown changes in the expression of the GluR5/6 subunits of KARs in the stratum oriens (SO of sectors CA2/3, where the basolateral amygdala (BLA sends a robust projection. Previous work using a rat model of SZ demonstrated that BLA activation leads to electrophysiological changes in fast-spiking interneurons in SO of CA2/3. The present study explores KAR modulation of interneurons in CA2/3 in response to BLA activation. Intrinsic firing properties of these interneurons through KAR-mediated activity were measured with patch-clamp recordings from rats that received 15 days of picrotoxin infusion into the BLA. Chronic BLA activation induced changes in the firing properties of CA2/3 interneurons associated with modifications in the function of KARs. Specifically, the responsiveness of these interneurons to activation of KARs was diminished in picrotoxin-treated rats, while the after-hyperpolarization (AHP amplitude was increased. In addition, we tested blockers of KAR subunits which have been shown to have altered gene expression in SO sector CA2/3 of SZ subjects. The GluR5 antagonist UBP296 further decreased AP frequency and increased AHP amplitude in picrotoxin-treated rats. Application of the GluR6/7 antagonist NS102 suggested that activation of GluR6/7 KARs may be required to maintain the high firing rates in SO interneurons in the presence of KA. Moreover, the GluR6/7 KAR-mediated signaling may be suppressed in PICRO-treated rats. Our findings indicate that glutamatergic activity from the BLA may modulate the firing properties of CA2/3 interneurons through GluR5 and GluR6/7 KARs. These receptors are expressed in GABAergic interneurons and play a key role in the synchronization of gamma oscillations. Modulation of interneuronal activity through KARs in

  6. Wnt co-receptors Lrp5 and Lrp6 differentially mediate Wnt3a signaling in osteoblasts.

    Directory of Open Access Journals (Sweden)

    Aimy Sebastian

    Full Text Available Wnt3a is a major regulator of bone metabolism however, very few of its target genes are known in bone. Wnt3a preferentially signals through transmembrane receptors Frizzled and co-receptors Lrp5/6 to activate the canonical signaling pathway. Previous studies have shown that the canonical Wnt co-receptors Lrp5 and Lrp6 also play an essential role in normal postnatal bone homeostasis, yet, very little is known about specific contributions by these co-receptors in Wnt3a-dependent signaling. We used high-throughput sequencing technology to identify target genes regulated by Wnt3a in osteoblasts and to elucidate the role of Lrp5 and Lrp6 in mediating Wnt3a signaling. Our study identified 782 genes regulated by Wnt3a in primary calvarial osteoblasts. Wnt3a up-regulated the expression of several key regulators of osteoblast proliferation/ early stages of differentiation while inhibiting genes expressed in later stages of osteoblastogenesis. We also found that Lrp6 is the key mediator of Wnt3a signaling in osteoblasts and Lrp5 played a less significant role in mediating Wnt3a signaling.

  7. Activation of the sympathetic nervous system mediates hypophagic and anxiety-like effects of CB₁ receptor blockade.

    Science.gov (United States)

    Bellocchio, Luigi; Soria-Gómez, Edgar; Quarta, Carmelo; Metna-Laurent, Mathilde; Cardinal, Pierre; Binder, Elke; Cannich, Astrid; Delamarre, Anna; Häring, Martin; Martín-Fontecha, Mar; Vega, David; Leste-Lasserre, Thierry; Bartsch, Dusan; Monory, Krisztina; Lutz, Beat; Chaouloff, Francis; Pagotto, Uberto; Guzman, Manuel; Cota, Daniela; Marsicano, Giovanni

    2013-03-19

    Complex interactions between periphery and the brain regulate food intake in mammals. Cannabinoid type-1 (CB1) receptor antagonists are potent hypophagic agents, but the sites where this acute action is exerted and the underlying mechanisms are not fully elucidated. To dissect the mechanisms underlying the hypophagic effect of CB1 receptor blockade, we combined the acute injection of the CB1 receptor antagonist rimonabant with the use of conditional CB1-knockout mice, as well as with pharmacological modulation of different central and peripheral circuits. Fasting/refeeding experiments revealed that CB1 receptor signaling in many specific brain neurons is dispensable for the acute hypophagic effects of rimonabant. CB1 receptor antagonist-induced hypophagia was fully abolished by peripheral blockade of β-adrenergic transmission, suggesting that this effect is mediated by increased activity of the sympathetic nervous system. Consistently, we found that rimonabant increases gastrointestinal metabolism via increased peripheral β-adrenergic receptor signaling in peripheral organs, including the gastrointestinal tract. Blockade of both visceral afferents and glutamatergic transmission in the nucleus tractus solitarii abolished rimonabant-induced hypophagia. Importantly, these mechanisms were specifically triggered by lipid-deprivation, revealing a nutrient-specific component acutely regulated by CB1 receptor blockade. Finally, peripheral blockade of sympathetic neurotransmission also blunted central effects of CB1 receptor blockade, such as fear responses and anxiety-like behaviors. These data demonstrate that, independently of their site of origin, important effects of CB1 receptor blockade are expressed via activation of peripheral sympathetic activity. Thus, CB1 receptors modulate bidirectional circuits between the periphery and the brain to regulate feeding and other behaviors.

  8. shRNA-mediated downregulation of α-N-Acetylgalactosaminidase inhibits migration and invasion of cancer cell lines

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    Ehsan Saburi

    2017-09-01

    Full Text Available Objective(s: Extracellular matrix (ECM is composed of many kinds of glycoproteins containing glycosaminoglycans (GAGs moiety. The research was conducted based on the N-Acetylgalactosamine (GalNAc degradation of ECM components by α-N-acetylgalactosaminidase (Nagalase which facilitates migration and invasion of cancer cells. This study aims to investigate the effects of Naga-shRNA downregulation on migration and invasion of cancer cell lines. Materials and Methods: In this study, MCF-7 cell line (human mammary carcinoma cell line and A2780 (human ovarian carcinoma cell line were used. The level of normalized Naga expression and Nagalase protein were evaluated by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay/western blotting, respectively. Migration and invasion were determined using transwell assays, and statistical analysis was carried out by ANOVA test. Results: Response to transduction by shRNA compared to the control group, migrative and invasive properties of the transfected cells were significantly inhibited. Conclusion: These results indicate that Nagalase may have an important role in migration and invasion of cancer cells and can be considered as a candidate for further studies.

  9. Methods of solving rapid binding target-mediated drug disposition model for two drugs competing for the same receptor.

    Science.gov (United States)

    Yan, Xiaoyu; Chen, Yang; Krzyzanski, Wojciech

    2012-10-01

    The target-mediated drug disposition (TMDD) model has been adopted to describe pharmacokinetics for two drugs competing for the same receptor. A rapid binding assumption introduces total receptor and total drug concentrations while free drug concentrations C (A) and C (B) are calculated from the equilibrium (Gaddum) equations. The Gaddum equations are polynomials in C (A) and C (B) of second degree that have explicit solutions involving complex numbers. The aim of this study was to develop numerical methods to solve the rapid binding TMDD model for two drugs competing for the same receptor that can be implemented in pharmacokinetic software. Algebra, calculus, and computer simulations were used to develop algorithms and investigate properties of solutions to the TMDD model with two drugs competitively binding to the same receptor. A general rapid binding approximation of the TMDD model for two drugs competing for the same receptor has been proposed. The explicit solutions to the equilibrium equations employ complex numbers, which cannot be easily solved by pharmacokinetic software. Numerical bisection algorithm and differential representation were developed to solve the system instead of obtaining an explicit solution. The numerical solutions were validated by MATLAB 7.2 solver for polynomial roots. The applicability of these algorithms was demonstrated by simulating concentration-time profiles resulting from exogenous and endogenous IgG competing for the neonatal Fc receptor (FcRn), and darbepoetin competing with endogenous erythropoietin for the erythropoietin receptor. These models were implemented in ADAPT 5 and Phoenix WinNonlin 6.0, respectively.

  10. NOD1 cooperates with TLR2 to enhance T cell receptor-mediated activation in CD8 T cells.

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    Blandine C Mercier

    Full Text Available Pattern recognition receptors (PRR, like Toll-like receptors (TLR and NOD-like receptors (NLR, are involved in the detection of microbial infections and tissue damage by cells of the innate immune system. Recently, we and others have demonstrated that TLR2 can additionally function as a costimulatory receptor on CD8 T cells. Here, we establish that the intracytosolic receptor NOD1 is expressed and functional in CD8 T cells. We show that C12-iEDAP, a synthetic ligand for NOD1, has a direct impact on both murine and human CD8 T cells, increasing proliferation and effector functions of cells activated via their T cell receptor (TCR. This effect is dependent on the adaptor molecule RIP2 and is associated with an increased activation of the NF-κB, JNK and p38 signaling pathways. Furthermore, we demonstrate that NOD1 stimulation can cooperate with TLR2 engagement on CD8 T cells to enhance TCR-mediated activation. Altogether our results indicate that NOD1 might function as an alternative costimulatory receptor in CD8 T cells. Our study provides new insights into the function of NLR in T cells and extends to NOD1 the recent concept that PRR stimulation can directly control T cell functions.

  11. Embryonic GABA(B receptor blockade alters cell migration, adult hypothalamic structure, and anxiety- and depression-like behaviors sex specifically in mice.

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    Matthew S Stratton

    Full Text Available Neurons of the paraventricular nucleus of the hypothalamus (PVN regulate the hypothalamic- pituitary-adrenal (HPA axis and the autonomic nervous system. Females lacking functional GABA(B receptors because of a genetic disruption of the R1 subunit have altered cellular characteristics in and around the PVN at birth. The genetic disruption precluded appropriate assessments of physiology or behavior in adulthood. The current study was conducted to test the long term impact of a temporally restricting pharmacological blockade of the GABA(B receptor to a 7-day critical period (E11-E17 during embryonic development. Experiments tested the role of GABA(B receptor signaling in fetal development of the PVN and later adult capacities for adult stress related behaviors and physiology. In organotypic slices containing fetal PVN, there was a female specific, 52% increase in cell movement speeds with GABA(B receptor antagonist treatment that was consistent with a sex-dependent lateral displacement of cells in vivo following 7 days of fetal exposure to GABA(B receptor antagonist. Anxiety-like and depression-like behaviors, open-field activity, and HPA mediated responses to restraint stress were measured in adult offspring of mothers treated with GABA(B receptor antagonist. Embryonic exposure to GABA(B receptor antagonist resulted in reduced HPA axis activation following restraint stress and reduced depression-like behaviors. There was also increased anxiety-like behavior selectively in females and hyperactivity in males. A sex dependent response to disruptions of GABA(B receptor signaling was identified for PVN formation and key aspects of physiology and behavior. These changes correspond to sex specific prevalence in similar human disorders, namely anxiety disorders and hyperactivity.

  12. The contribution of IL-6 to beta 3 adrenergic receptor mediated adipose tissue remodeling

    Science.gov (United States)

    Buzelle, Samyra L; MacPherson, Rebecca E K; Peppler, Willem T; Castellani, Laura; Wright, David C

    2015-01-01

    The chronic activation of beta 3 adrenergic receptors results in marked alterations in adipose tissue morphology and metabolism, including increases in mitochondrial content and the expression of enzymes involved in lipogenesis and glyceroneogenesis. Acute treatment with CL 316,243, a beta 3 adrenergic agonist, induces the expression of interleukin 6. Interestingly, IL-6 has been shown to induce mitochondrial genes in cultured adipocytes. Therefore, the purpose of this paper was to examine the role of interleukin 6 in mediating the in vivo effects of CL 316,243 in white adipose tissue. Circulating IL-6, and markers of IL-6 signaling in white adipose tissue were increased 4 h following a single injection of CL 316,243 in C57BL6/J mice. Once daily injections of CL 316,243 for 5 days increased the protein content of a number of mitochondrial proteins including CORE1, Cytochrome C, PDH, MCAD, and Citrate Synthase to a similar extent in adipose tissue from WT and IL-6−/− mice. Conversely, CL 316,243-induced increases in COXIV and phosphorylated AMPK were attenuated in IL-6−/− mice. Likewise, the slight, but significant, CL 316,243-induced increases in ATGL, PEPCK, and PPARγ, were reduced or absent in adipose tissue IL-6−/− mice. The attenuated response to CL 316,243 in white adipose tissue in IL-6−/− mice was associated with reductions in whole-body oxygen consumption and energy expenditure in the light phase. Our findings suggest that IL-6 plays a limited role in CL 316,243-mediated adipose tissue remodeling. PMID:25713332

  13. Kinin B1 Receptor Promotes Neurogenic Hypertension Through Activation of Centrally Mediated Mechanisms.

    Science.gov (United States)

    Sriramula, Srinivas; Lazartigues, Eric

    2017-12-01

    Hypertension is associated with increased activity of the kallikrein-kinin system. Kinin B1 receptor (B1R) activation leads to vasoconstriction and inflammation. Despite evidence supporting a role for the B1R in blood pressure regulation, the mechanisms by which B1R could alter autonomic function and participate in the pathogenesis of hypertension remain unidentified. We sought to explore whether B1R-mediated inflammation contributes to hypertension and investigate the molecular mechanisms involved. In this study, we tested the hypothesis that activation of B1R in the brain is involved in the pathogenesis of hypertension, using the deoxycorticosterone acetate-salt model of neurogenic hypertension in wild-type and B1R knockout mice. Deoxycorticosterone acetate-salt treatment in wild-type mice led to significant increases in B1R mRNA and protein levels and bradykinin levels, enhanced gene expression of carboxypeptidase N supporting an increase in the B1R ligand, associated with enhanced blood pressure, inflammation, sympathoexcitation, autonomic dysfunction, and impaired baroreflex sensitivity, whereas these changes were blunted or prevented in B1R knockout mice. B1R stimulation was further shown to involve activation of the ASK1-JNK-ERK1/2 and NF-κB pathways in the brain. To dismiss potential developmental alterations in knockout mice, we further used B1R blockade selectively in the brain of wild-type mice. Supporting the central origin of this mechanism, intracerebroventricular infusion of a specific B1R antagonist, attenuated the deoxycorticosterone acetate-salt-induced increase in blood pressure in wild-type mice. Our data provide the first evidence of a central role for B1R-mediated inflammatory pathways in the pathogenesis of deoxycorticosterone acetate-salt hypertension and offer novel insights into possible B1R-targeted therapies for the treatment of neurogenic hypertension. © 2017 American Heart Association, Inc.

  14. Adolescent chronic mild stress alters hippocampal CB1 receptor-mediated excitatory neurotransmission and plasticity.

    Science.gov (United States)

    Reich, C G; Mihalik, G R; Iskander, A N; Seckler, J C; Weiss, M S

    2013-12-03

    Endocannabinoids (eCBs) are involved in the stress response and alterations in eCB signaling may contribute to the etiology of mood disorders. Exposure to chronic mild stress (CMS), a model of depression, produces downregulation of the cannabinoid 1 (CB1) receptor in the hippocampus of male rats. However, it is unknown how this stress-induced change in CB1 levels affects eCB-mediated neurotransmission. In vitro, field potential recordings from CMS-exposed (21-days) rats were performed to assess the effects of stress on eCB-regulated glutamatergic neurotransmission in/on hippocampal area CA1. We observed that application of the CB1 agonist, WIN 55,212-5 (1 μM), in stress animals resulted in a ∼135% increase in excitatory neurotransmission, whereas CB1 activation in non-stress animals leads to a ∼30% decrease. However, during blockade of GABA(A) neurotransmission with picrotoxin, CB1 activation yielded a ∼35% decrease in stress animals. These findings indicate that CMS does not directly affect glutamatergic neurotransmission. Rather, CMS sensitizes CB1 function on GABAergic terminals, leading to less inhibition and an increase in excitatory neurotransmission. This finding is reinforced in that induction of weak long-term-potentiation (LTP) is enhanced in CMS-exposed animals compared to controls and this enhancement is CB1-dependent. Lastly, we observed that the LTP-blocking property of WIN 55,212-5 shifts from being glutamate-dependent in non-stress animals to being GABA-dependent in stress animals. These results effectively demonstrate that CMS significantly alters hippocampal eCB-mediated neurotransmission and synaptic plasticity. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

  15. Intra-vermis H4 receptor agonist impairs performance in anxiety- and fear-mediated models.

    Science.gov (United States)

    Fernandes, C E M; Serafim, K R; Gianlorenco, A C L; Mattioli, R

    2017-10-01

    The neural histaminergic system modulates cognitive performance in various animal models. However, little is known about the effects of the H4 histaminergic receptor in the central nervous system. The purpose of this study was to investigate the effect of histaminergic H4 agonist VUF-8430 microinjection into the cerebellar vermis on the consolidation of emotional memory in mice subjected to the elevated plus maze (EPM) and inhibitory avoidance task (IAT). All experiments were performed on two consecutive days: exposure (T1 and D1) and 24h after, which we called re-exposure (T2 and D2). The animals received saline (SAL) or VUF (0.15 nmol; 0.49 nmol; 1.48 nmol/0.1μl) administered post-exposure. Experiment 1 was conducted in the EPM, and the animals were free to explore the maze for 5min. In T1, immediately after exposure, the pharmacological treatment was given; in T2, there was only re-exposure to the EPM. Experiment 2 involved the IAT, and the pharmacological treatment was provided post-D1; in D2, the animals were only re-exposed to the IAT. In Experiment 1, increased open arm exploration (% open arm entries and% open arms time) for 0.49 and 1.48nmol of VUF were recorded in T2 compared to T1. In Experiment 2, a significant decrease in consolidation latency was recorded for the group that received 1.48nmol of VUF compared to the SAL group in D2. These results indicate that a 1.48nmol VUF microinjection into the cerebellar vermis impaired performance in both models, even though one model was anxiety-mediated (EPM) and the other was fear-mediated (IAT). Copyright © 2017 Elsevier Inc. All rights reserved.

  16. TRPV1-mediated calcium signal couples with cannabinoid receptors and sodium-calcium exchangers in rat odontoblasts.

    Science.gov (United States)

    Tsumura, Maki; Sobhan, Ubaidus; Muramatsu, Takashi; Sato, Masaki; Ichikawa, Hideki; Sahara, Yoshinori; Tazaki, Masakazu; Shibukawa, Yoshiyuki

    2012-08-01

    Odontoblasts are involved in the transduction of stimuli applied to exposed dentin. Although expression of thermo/mechano/osmo-sensitive transient receptor potential (TRP) channels has been demonstrated, the properties of TRP vanilloid 1 (TRPV1)-mediated signaling remain to be clarified. We investigated physiological and pharmacological properties of TRPV1 and its functional coupling with cannabinoid (CB) receptors and Na(+)-Ca(2+) exchangers (NCXs) in odontoblasts. Anandamide (AEA), capsaicin (CAP), resiniferatoxin (RF) or low-pH evoked Ca(2+) influx. This influx was inhibited by capsazepine (CPZ). Delay in time-to-activation of TRPV1 channels was observed between application of AEA or CAP and increase in [Ca(2+)](i). In the absence of extracellular Ca(2+), however, an immediate increase in [Ca(2+)](i) was observed on administration of extracellular Ca(2+), followed by activation of TRPV1 channels. Intracellular application of CAP elicited inward current via opening of TRPV1 channels faster than extracellular application. With extracellular RF application, no time delay was observed in either increase in [Ca(2+)](i) or inward current, indicating that agonist binding sites are located on both extra- and intracellular domains. KB-R7943, an NCX inhibitor, yielded an increase in the decay time constant during TRPV1-mediated Ca(2+) entry. Increase in [Ca(2+)](i) by CB receptor agonist, 2-arachidonylglycerol, was inhibited by CB1 receptor antagonist or CPZ, as well as by adenylyl cyclase inhibitor. These results showed that TRPV1-mediated Ca(2+) entry functionally couples with CB1 receptor activation via cAMP signaling. Increased [Ca(2+)](i) by TRPV1 activation was extruded by NCXs. Taken together, this suggests that cAMP-mediated CB1-TRPV1 crosstalk and TRPV1-NCX coupling play an important role in driving cellular functions following transduction of external stimuli to odontoblasts. Copyright © 2012 Elsevier Ltd. All rights reserved.

  17. Identification of a novel antagonist of the ErbB1 receptor capable of inhibiting migration of human glioblastoma cells

    DEFF Research Database (Denmark)

    Staberg, Mikkel; Riemer, Christian; Xu, Ruodan

    2013-01-01

    BACKGROUND: Receptors of the ErbB family are involved in the development of various cancers, and the inhibition of these receptors represents an attractive therapeutic concept. Upon ligand binding, ErbB receptors become activated as homo- or heterodimers, leading to the activation of downstream......B1 targeting peptide, termed Herfin-1, was designed based on a model of the tertiary structure of the EGF-EGFR ternary complex. The binding kinetics of this peptide were determined employing surface plasmon resonance analyses. ErbB1-4 expression and phosphorylation in human glioblastoma cell lines U....... Additionally, Herfin-1 was found to increase neurite outgrowth in cerebellar granule neurons, likely through the inhibition of a sustained weak ErbB1 activation. CONCLUSIONS: Targeting the ErbB1 receptor dimerization interface is a promising strategy to inhibit receptor activation in ErbB1-expressing glioma...

  18. 3-Mercaptopyruvate Sulfurtransferase, Not Cystathionine β-Synthase Nor Cystathionine γ-Lyase, Mediates Hypoxia-Induced Migration of Vascular Endothelial Cells.

    Science.gov (United States)

    Tao, Beibei; Wang, Rui; Sun, Chen; Zhu, Yichun

    2017-01-01

    Hypoxia-induced angiogenesis is a common phenomenon in many physiological and patho-physiological processes. However, the potential differential roles of three hydrogen sulfide producing systems cystathionine γ-lyase (CSE)/H2S, cystathionine β-synthase (CBS)/H2S, and 3-mercaptopyruvate sulfurtransferase (MPST)/H2S in hypoxia-induced angiogenesis are still unknown. We found that minor hypoxia (10% oxygen) significantly increased the migration of vascular endothelial cells while hypoxia (8% oxygen) significantly inhibited cell migration. The present study was performed using cells cultured in 10% oxygen. RNA interference was used to block the endogenous generation of hydrogen sulfide by CSE, CBS, or MPST in a vascular endothelial cell migration model in both normoxia and hypoxia. The results showed that CBS had a promoting effect on the migration of vascular endothelial cells cultured in both normoxic and hypoxic conditions. In contrast, CSE had an inhibitory effect on cell migration. MPST had a promoting effect on the migration of vascular endothelial cells cultured in hypoxia; however, it had no effect on the cells cultured in normoxia. Importantly, it was found that the hypoxia-induced increase in vascular endothelial cell migration was mediated by MPST, but not CSE or CBS. The western blot analyses showed that hypoxia significantly increased MPST protein levels, decreased CSE protein levels and did not change CBS levels, suggesting that these three hydrogen sulfide-producing systems respond differently to hypoxic conditions. Interestingly, MPST protein levels were elevated by hypoxia in a bi-phasic manner and MPST mRNA levels increased later than the first stage elevation of the protein levels, implying that the expression of MPST induced by hypoxia was also regulated at a post-transcriptional level. RNA pull-down assay showed that some candidate RNA binding proteins, such as nucleolin and Annexin A2, were dissociated from the 3'-UTR of MPST mRNA in hypoxia

  19. Luteolin inhibits human prostate tumor growth by suppressing vascular endothelial growth factor receptor 2-mediated angiogenesis

    National Research Council Canada - National Science Library

    Pratheeshkumar, Poyil; Son, Young-Ok; Budhraja, Amit; Wang, Xin; Ding, Songze; Wang, Lei; Hitron, Andrew; Lee, Jeong-Chae; Kim, Donghern; Divya, Sasidharan Padmaja; Chen, Gang; Zhang, Zhuo; Luo, Jia; Shi, Xianglin

    2012-01-01

    .... In vitro studies using rat aortic ring assay showed that luteolin at non-toxic concentrations significantly inhibited microvessel sprouting and proliferation, migration, invasion and tube formation...

  20. Opioid receptors mediate the acquisition, but not the expression of mitragynine-induced conditioned place preference in rats.

    Science.gov (United States)

    Yusoff, Nurul H M; Mansor, Sharif M; Müller, Christian P; Hassan, Zurina

    2017-08-14

    Mitragynine is the main psychoactive ingredient of the herbal drug preparation Kratom (Ketum), derived from the plant Mitragyna speciosa. Kratom is a widely abused drug in Southeast Asian and has a psychostimulant profile at low-medium doses, while high doses have opioidergic effects. Mitragynine was shown to possess opiate receptor affinity. However, its role in the behavioural effects of mitragynine is unclear. Here we asked whether the reinforcing effects of mitragynine are mediated by opiate receptors using a conditioned place preference (CPP) paradigm in rats. In the first experiment we tested the effects of the opiate receptor antagonist naloxone (0.1, 0.3 and 1.0mg/kg) on the acquisition of mitragynine (10mg/kg)-induced CPP. In the second experiment, we tested the involvement of opiate receptors in the expression of mitragynine-induced CPP in rats. We found that naloxone suppresses the acquisition of mitragynine-induced CPP. This effect was already evident at a dose of naloxone (0.1mg/kg) which, by itself, had no conditioned place aversion (CPA) effect. Higher doses of naloxone induced a CPA and blocked mitragynine-induced CPP. In contrast, naloxone had no effect on the expression of mitragynine-induced CPP. These findings suggest that the acquisition, but not the expression of the reinforcing effects of mitragynine is mediated by opiate receptors. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. {delta}-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    Energy Technology Data Exchange (ETDEWEB)

    Heiss, Anika; Ammer, Hermann [Institute of Pharmacology, Toxicology and Pharmacy Ludwig-Maximilians-University of Munich Koeniginstrasse 16 80539 Muenchen Federal Republic of Germany (Germany); Eisinger, Daniela A., E-mail: eisinger@pharmtox.vetmed.uni-muenchen.de [Institute of Pharmacology, Toxicology and Pharmacy Ludwig-Maximilians-University of Munich Koeniginstrasse 16 80539 Muenchen Federal Republic of Germany (Germany)

    2009-07-15

    {delta}-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen{sup 2,5}]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory G{sub i/o} proteins, because pre-treatment with pertussis toxin, but not over-expression of the G{sub q/11} scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the G{beta}{gamma} scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  2. The Role of Phragmites australis in Mediating Inland Salt Marsh Migration in a Mid-Atlantic Estuary

    Science.gov (United States)

    Smith, Joseph A. M.

    2013-01-01

    Many sea level rise adaptation plans emphasize the protection of adjacent uplands to allow for inland salt marsh migration, but little empirical information exists on this process. Using aerial photos from 1930 and 2006 of Delaware Estuary coastal habitats in New Jersey, I documented the rate of coastal forest retreat and the rate of inland salt marsh migration across 101.1 km of undeveloped salt marsh and forest ecotone. Over this time, the amount of forest edge at this ecotone nearly doubled. In addition, the average amount of forest retreat was 141.2 m while the amount of salt marsh inland migration was 41.9 m. Variation in forest retreat within the study area was influenced by variation in slope. The lag between the amount of forest retreat and salt marsh migration is accounted for by the presence of Phragmites australis which occupies the forest and salt marsh ecotone. Phragmites expands from this edge into forest dieback areas, and the ability of salt marsh to move inland and displace Phragmites is likely influenced by salinity at both an estuary-wide scale and at the scale of local subwatersheds. Inland movement of salt marsh is lowest at lower salinity areas further away from the mouth of the estuary and closer to local heads of tide. These results allow for better p