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Sample records for monocytic cell migration

  1. Curcumin modulates endothelial permeability and monocyte transendothelial migration by affecting endothelial cell dynamics.

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    Monfoulet, Laurent-Emmanuel; Mercier, Sylvie; Bayle, Dominique; Tamaian, Radu; Barber-Chamoux, Nicolas; Morand, Christine; Milenkovic, Dragan

    2017-11-01

    Curcumin is a phenolic compound that exhibits beneficial properties for cardiometabolic health. We previously showed that curcumin reduced the infiltration of immune cells into the vascular wall and prevented atherosclerosis development in mice. This study aimed to investigate the effect of curcumin on monocyte adhesion and transendothelial migration (TEM) and to decipher the underlying mechanisms of these actions. Human umbilical vein endothelial cells (HUVECs) were exposed to curcumin (0.5-1μM) for 3h prior to their activation by Tumor Necrosis Factor alpha (TNF-α). Endothelial permeability, monocyte adhesion and transendothelial migration assays were conducted under static condition and shear stress that mimics blood flow. We further investigated the impact of curcumin on signaling pathways and on the expression of genes using macroarrays. Pre-exposure of endothelial cells to curcumin reduced monocyte adhesion and their transendothelial migration in both static and shear stress conditions. Curcumin also prevented changes in both endothelial permeability and the area of HUVECs when induced by TNF-α. We showed that curcumin modulated the expression of 15 genes involved in the control of cytoskeleton and endothelial junction dynamic. Finally, we showed that curcumin inhibited NF-κB signaling likely through an antagonist interplay with several kinases as suggested by molecular docking analysis. Our findings demonstrate the ability of curcumin to reduce monocyte TEM through a multimodal regulation of the endothelial cell dynamics with a potential benefit on the vascular endothelial function barrier. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Polarization of migrating monocytic cells is independent of PI 3-kinase activity.

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    Silvia Volpe

    Full Text Available BACKGROUND: Migration of mammalian cells is a complex cell type and environment specific process. Migrating hematopoietic cells assume a rapid amoeboid like movement when exposed to gradients of chemoattractants. The underlying signaling mechanisms remain controversial with respect to localization and distribution of chemotactic receptors within the plasma membrane and the role of PI 3-kinase activity in cell polarization. METHODOLOGY/PRINCIPAL FINDINGS: We present a novel model for the investigation of human leukocyte migration. Monocytic THP-1 cells transfected with the alpha(2A-adrenoceptor (alpha(2AAR display comparable signal transduction responses, such as calcium mobilization, MAP-kinase activation and chemotaxis, to the noradrenaline homologue UK 14'304 as when stimulated with CCL2, which binds to the endogenous chemokine receptor CCR2. Time-lapse video microscopy reveals that chemotactic receptors remain evenly distributed over the plasma membrane and that their internalization is not required for migration. Measurements of intramolecular fluorescence resonance energy transfer (FRET of alpha(2AAR-YFP/CFP suggest a uniform activation of the receptors over the entire plasma membrane. Nevertheless, PI 3-kinase activation is confined to the leading edge. When reverting the gradient of chemoattractant by moving the dispensing micropipette, polarized monocytes--in contrast to neutrophils--rapidly flip their polarization axis by developing a new leading edge at the previous posterior side. Flipping of the polarization axis is accompanied by re-localization of PI-3-kinase activity to the new leading edge. However, reversal of the polarization axis occurs in the absence of PI 3-kinase activation. CONCLUSIONS/SIGNIFICANCE: Accumulation and internalization of chemotactic receptors at the leading edge is dispensable for cell migration. Furthermore, uniformly distributed receptors allow the cells to rapidly reorient and adapt to changes in the

  3. Zinc oxide nanoparticles induce migration and adhesion of monocytes to endothelial cells and accelerate foam cell formation

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Yuka; Tada-Oikawa, Saeko [Graduate School of Regional Innovation Studies, Mie University, Tsu (Japan); Ichihara, Gaku [Department of Occupational and Environmental Health, Nagoya University Graduate School of Medicine, Nagoya (Japan); Yabata, Masayuki; Izuoka, Kiyora [Graduate School of Regional Innovation Studies, Mie University, Tsu (Japan); Suzuki, Masako; Sakai, Kiyoshi [Nagoya City Public Health Research Institute, Nagoya (Japan); Ichihara, Sahoko, E-mail: saho@gene.mie-u.ac.jp [Graduate School of Regional Innovation Studies, Mie University, Tsu (Japan)

    2014-07-01

    Metal oxide nanoparticles are widely used in industry, cosmetics, and biomedicine. However, the effects of exposure to these nanoparticles on the cardiovascular system remain unknown. The present study investigated the effects of nanosized TiO{sub 2} and ZnO particles on the migration and adhesion of monocytes, which are essential processes in atherosclerogenesis, using an in vitro set-up of human umbilical vein endothelial cells (HUVECs) and human monocytic leukemia cells (THP-1). We also examined the effects of exposure to nanosized metal oxide particles on macrophage cholesterol uptake and foam cell formation. The 16-hour exposure to ZnO particles increased the level of monocyte chemotactic protein-1 (MCP-1) and induced the migration of THP-1 monocyte mediated by increased MCP-1. Exposure to ZnO particles also induced adhesion of THP-1 cells to HUVECs. Moreover, exposure to ZnO particles, but not TiO{sub 2} particles, upregulated the expression of membrane scavenger receptors of modified LDL and increased cholesterol uptake in THP-1 monocytes/macrophages. In the present study, we found that exposure to ZnO particles increased macrophage cholesterol uptake, which was mediated by an upregulation of membrane scavenger receptors of modified LDL. These results suggest that nanosized ZnO particles could potentially enhance atherosclerogenesis and accelerate foam cell formation. - Highlights: • Effects of metal oxide nanoparticles on foam cell formation were investigated. • Exposure to ZnO nanoparticles induced migration and adhesion of monocytes. • Exposure to ZnO nanoparticles increased macrophage cholesterol uptake. • Expression of membrane scavenger receptors of modified LDL was also increased. • These effects were not observed after exposure to TiO{sub 2} nanoparticles.

  4. Intravital live cell triggered imaging system reveals monocyte patrolling and macrophage migration in atherosclerotic arteries

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    McArdle, Sara; Chodaczek, Grzegorz; Ray, Nilanjan; Ley, Klaus

    2015-02-01

    Intravital multiphoton imaging of arteries is technically challenging because the artery expands with every heartbeat, causing severe motion artifacts. To study leukocyte activity in atherosclerosis, we developed the intravital live cell triggered imaging system (ILTIS). This system implements cardiac triggered acquisition as well as frame selection and image registration algorithms to produce stable movies of myeloid cell movement in atherosclerotic arteries in live mice. To minimize tissue damage, no mechanical stabilization is used and the artery is allowed to expand freely. ILTIS performs multicolor high frame-rate two-dimensional imaging and full-thickness three-dimensional imaging of beating arteries in live mice. The external carotid artery and its branches (superior thyroid and ascending pharyngeal arteries) were developed as a surgically accessible and reliable model of atherosclerosis. We use ILTIS to demonstrate Cx3cr1GFP monocytes patrolling the lumen of atherosclerotic arteries. Additionally, we developed a new reporter mouse (Apoe-/-Cx3cr1GFP/+Cd11cYFP) to image GFP+ and GFP+YFP+ macrophages "dancing on the spot" and YFP+ macrophages migrating within intimal plaque. ILTIS will be helpful to answer pertinent open questions in the field, including monocyte recruitment and transmigration, macrophage and dendritic cell activity, and motion of other immune cells.

  5. Cell proliferation and migration are modulated by Cdk-1-phosphorylated endothelial-monocyte activating polypeptide II.

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    Margaret A Schwarz

    Full Text Available Endothelial-Monocyte Activating Polypeptide (EMAP II is a secreted protein with well-established anti-angiogenic activities. Intracellular EMAP II expression is increased during fetal development at epithelial/mesenchymal boundaries and in pathophysiologic fibroproliferative cells of bronchopulmonary dysplasia, emphysema, and scar fibroblast tissue following myocardial ischemia. Precise function and regulation of intracellular EMAP II, however, has not been explored to date.Here we show that high intracellular EMAP II suppresses cellular proliferation by slowing progression through the G2M cell cycle transition in epithelium and fibroblast. Furthermore, EMAP II binds to and is phosphorylated by Cdk1, and exhibits nuclear/cytoplasmic partitioning, with only nuclear EMAP II being phosphorylated. We observed that extracellular secreted EMAP II induces endothelial cell apoptosis, where as excess intracellular EMAP II facilitates epithelial and fibroblast cells migration.Our findings suggest that EMAP II has specific intracellular effects, and that this intracellular function appears to antagonize its extracellular anti-angiogenic effects during fetal development and pulmonary disease progression.

  6. Soluble vascular endothelial growth factor (VEGF) receptor-1 inhibits migration of human monocytic THP-1 cells in response to VEGF.

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    Zhu, Cansheng; Xiong, Zhaojun; Chen, Xiaohong; Lu, Zhengqi; Zhou, Guoyu; Wang, Dunjing; Bao, Jian; Hu, Xueqiang

    2011-08-01

    We aimed to investigate the regulation and contribution of vascular endothelial growth factor (VEGF) and sFlt-1(1-3) to human monocytic THP-1 migration. Ad-sFlt-1/FLAG, a recombinant adenovirus carrying the human sFlt-1(1-3) (the first three extracellular domains of FLT-1, the hVEGF receptor-1) gene, was constructed. L929 cells were infected with Ad-sFlt-1/FLAG and the expression of sFlt-1 was detected by immunofluorescent assay and ELISA. Corning(®) Transwell(®) Filter Inserts containing polyethylene terephthalate (PET) membranes with pore sizes of 3 μm were used as an experimental model to simulate THP-1 migration. Five VEGF concentrations (0, 0.1, 1, 10 and 100 ng/ml), four concentrations of sFlt-1(1-3)/FLAG expression supernatants (0.1, 1, 10 and 100 ng/ml), and monocyte chemoattractant protein-1 (MCP-1, 10 ng/ml) were used to test the ability of THP-1 cells to migrate through PET membranes. The sFlt-1(1-3) gene was successfully recombined into Ad-sFlt-1/FLAG. sFlt-1(1-3) was expressed in L929 cells transfected with Ad-sFlt-1/FLAG. THP-1 cell migration increased with increasing concentrations of VEGF, while cell migration decreased with increasing concentrations of sFlt1(1-3)/FLAG. sFlt1(1-3)/FLAG had no effect on MCP-1-induced cell migration. This study demonstrated that VEGF is able to elicit a migratory response in THP-1 cells, and that sFlt-1(1-3) is an effective inhibitor of THP-1 migration towards VEGF.

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

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    Rogier M Thurlings

    2009-11-01

    Full Text Available 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.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.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.

  8. The role and mechanism of KCa3.1 channels in human monocyte migration induced by palmitic acid.

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    Ma, Xiao-Zhen; Pang, Zheng-Da; Wang, Jun-Hong; Song, Zheng; Zhao, Li-Mei; Du, Xiao-Jun; Deng, Xiu-Ling

    2018-05-21

    Monocyte migration into diseased tissues contributes to the pathogenesis of diseases. Intermediate-conductance Ca 2+ -activated K + (K Ca 3.1) channels play an important role in cell migration. However, the role of K Ca 3.1 channels in mediating monocyte migration induced by palmitic acid (PA) is still unclear. Using cultured THP-1 cells and peripheral blood mononuclear cells from healthy subjects, we investigated the role and signaling mechanisms of K Ca 3.1 channels in mediating the migration induced by PA. Using methods of Western blotting analysis, RNA interference, cell migration assay and ELISA, we found that PA-treated monocytes exhibited increment of the protein levels of K Ca 3.1 channel and monocyte chemoattractant protein-1 (MCP-1), and the effects were reversed by co-incubation of PA with anti-TLR2/4 antibodies or by specific inhibitors of p38-MAPK, or NF-κB. In addition, PA increased monocyte migration, which was abolished by a specific K Ca 3.1 channel blocker, TRAM-34, or K Ca 3.1 small interfering RNA (siRNA). The expression and secretion of MCP-1 induced by PA was also similarly prevented by TRAM-34 and K Ca 3.1 siRNA. These results demonstrate for the first time that PA upregulates K Ca 3.1 channels through TLR2/4, p38-MAPK and NF-κB pathway to promote the expression of MCP-1, and then induce the trans-endothelial migration of monocytes. Copyright © 2018 Elsevier Inc. All rights reserved.

  9. Olopatadine Suppresses the Migration of THP-1 Monocytes Induced by S100A12 Protein

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    2006-01-01

    Full Text Available Olopatadine hydrochloride (olopatadine is an antiallergic drug with histamine H 1 receptor antagonistic activity. Recently, olopatadine has been shown to bind to S100A12 which is a member of the S100 family of calcium-binding proteins, and exerts multiple proinflammatory activities including chemotaxis for monocytes and neutrophils. In this study, we examined the possibility that the interaction of olopatadine with S100A12 inhibits the proinflammatory effects of S100A12. Pretreatment of olopatadine with S100A12 reduced migration of THP-1, a monocyte cell line, induced by S100A12 alone, but did not affect recombinant human regulated upon activation, normal T cell expressed and secreted (RANTES-induced migration. Amlexanox, which also binds to S100A12, inhibited the THP-1 migration induced by S100A12. However, ketotifen, another histamine H 1 receptor antagonist, had little effect on the activity of S100A12. These results suggest that olopatadine has a new mechanism of action, that is, suppression of the function of S100A12, in addition to histamine H 1 receptor antagonistic activity.

  10. Induction of Chemokine Secretion and Monocyte Migration by Human Choroidal Melanocytes in Response to Proinflammatory Cytokines

    DEFF Research Database (Denmark)

    Jehs, Tina; Faber, Carsten; Udsen, Maja S.

    2016-01-01

    of 10 HCM donors induced a high initial level of monocyte migration, which decreased upon stimulation with either TCM or IFN-γ and TNF-α. The supernatants from three HCM donors initially showed a low level of monocyte attraction, which increased after exposure to proinflammatory cytokines. Direct...

  11. [Peptide fragments of chemokine domain of fractalkine: effect on human monocyte migration].

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    Kukhtina, N B; Aref'eva, T I; Ruleva, N Iu; Sidorova, M V; Az'muko, A A; Bespalova, Zh D; Krasnikova, T L

    2012-01-01

    Leukocyte chemotaxis to the area of tissue damage is mediated by chemokines. According to the primary structure, chemokines are divided into four families, fractalkine (CX3CL1) is the only one member of CX3C family and the only membrane-bound chemokine. Fractalkine molecule includes the extracellular N-terminal chemokine domain, mucin-like rod, the transmembrane and the intracellular domains. In membrane-bound state fractalkine has the properties of an adhesion molecule. Chemokine domain of fractalkine (CDF) is released from cell membrane by proteolysis, and this soluble form acts as a chemoattractant for leukocytes expressing fractalkine receptor CX3CR1. Fractalkine is involved in development of a number of pathological processes caused by inflammation, and therefore a search for fractalkine inhibitors is very important. For this purpose we identified several antigenic determinants--the fragments of CDF, and the following peptides were synthesized--P41-52 H-Leu-Glu-Thr-Arg-Gln-His-Arg-Leu-Phe-Cys-Ala-Asp-NH2, P53-60 H-Pro-Lys-Glu-Gln-Trp-Val-Lys-Asp-NH2 and P60-71 H-Asp-Ala-Met-Gln-His-Leu-Asp-Arg-Gln-Ala-Ala-Ala-NH2. The peptide effects on adhesion and migration of human peripheral blood monocytes expressing fractalkine receptors were investigated. In the presence of CDF and P41-52 we observed the increased adhesion and migration of monocytes compared with spontaneous values. Peptides P53-60 and P60-71 significantly inhibited monocyte adhesion and migration stimulated by CDF. Since the chemotactic activity of chemokines was shown to be dependent on their binding to glycosaminoglycans of the cell surface and extracellular matrix, the effect ofpeptides on the interaction of CDF with heparin was analyzed by ELISA. Peptide P41-52 competed with CDF for heparin binding, while peptides P53-60 and P60-71 had no significant activity.

  12. Dopamine Increases CD14+CD16+ Monocyte Migration and Adhesion in the Context of Substance Abuse and HIV Neuropathogenesis

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    Coley, Jacqueline S.; Calderon, Tina M.; Gaskill, Peter J.; Eugenin, Eliseo A.; Berman, Joan W.

    2015-01-01

    Drug abuse is a major comorbidity of HIV infection and cognitive disorders are often more severe in the drug abusing HIV infected population. CD14+CD16+ monocytes, a mature subpopulation of peripheral blood monocytes, are key mediators of HIV neuropathogenesis. Infected CD14+CD16+ monocyte transmigration across the blood brain barrier mediates HIV entry into the brain and establishes a viral reservoir within the CNS. Despite successful antiretroviral therapy, continued influx of CD14+CD16+ monocytes, both infected and uninfected, contributes to chronic neuroinflammation and the development of HIV associated neurocognitive disorders (HAND). Drug abuse increases extracellular dopamine in the CNS. Once in the brain, CD14+CD16+ monocytes can be exposed to extracellular dopamine due to drug abuse. The direct effects of dopamine on CD14+CD16+ monocytes and their contribution to HIV neuropathogenesis are not known. In this study, we showed that CD14+CD16+ monocytes express mRNA for all five dopamine receptors by qRT-PCR and D1R, D5R and D4R surface protein by flow cytometry. Dopamine and the D1-like dopamine receptor agonist, SKF38393, increased CD14+CD16+ monocyte migration that was characterized as chemokinesis. To determine whether dopamine affected cell motility and adhesion, live cell imaging was used to monitor the accumulation of CD14+CD16+ monocytes on the surface of a tissue culture dish. Dopamine increased the number and the rate at which CD14+CD16+ monocytes in suspension settled to the dish surface. In a spreading assay, dopamine increased the area of CD14+CD16+ monocytes during the early stages of cell adhesion. In addition, adhesion assays showed that the overall total number of adherent CD14+CD16+ monocytes increased in the presence of dopamine. These data suggest that elevated extracellular dopamine in the CNS of HIV infected drug abusers contributes to HIV neuropathogenesis by increasing the accumulation of CD14+CD16+ monocytes in dopamine rich brain

  13. Soya-cerebroside, an extract of Cordyceps militaris, suppresses monocyte migration and prevents cartilage degradation in inflammatory animal models

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    Liu, Shan-Chi; Chiu, Ching-Peng; Tsai, Chun-Hao; Hung, Chun-Yin; Li, Te-Mao; Wu, Yang-Chang; Tang, Chih-Hsin

    2017-01-01

    Pathophysiological events that modulate the progression of structural changes in osteoarthritis (OA) include the secretion of inflammatory molecules, such as proinflammatory cytokines. Interleukin-1beta (IL-1β) is the prototypical inflammatory cytokine that activates OA synovial cells to release cytokines and chemokines in support of the inflammatory response. The monocyte chemoattractant protein-1 (MCP-1/CCL2) is one of the key chemokines that regulate migration and infiltration of monocytes in response to inflammation. We show in this study that IL-1β-induced MCP-1 expression and monocyte migration in OA synovial fibroblasts (OASFs) is effectively inhibited by soya-cerebroside, an extract of Cordyceps militaris. We found that soya-cerebroside up-regulated of microRNA (miR)-432 expression via inhibiting AMPK and AKT signaling pathways in OASFs. Soya-cerebroside also effectively decreased monocyte infiltration and prevented cartilage degradation in a rat inflammatory model. Our findings are the first to demonstrate that soya-cerebroside inhibits monocyte/macrophage infiltration into synoviocytes, attenuating synovial inflammation and preventing cartilage damage by reducing MCP-1 expression in vitro and in vivo. Taken together, we suggest a novel therapeutic strategy based on the use of soya-cerebroside for the management of OA. PMID:28225075

  14. CD13 is a novel mediator of monocytic/endothelial cell adhesion

    DEFF Research Database (Denmark)

    Mina-Osorio, Paola; Winnicka, Beata; O'Conor, Catherine

    2008-01-01

    During inflammation, cell surface adhesion molecules guide the adhesion and migration of circulating leukocytes across the endothelial cells lining the blood vessels to access the site of injury. The transmembrane molecule CD13 is expressed on monocytes and endothelial cells and has been shown...... to mediate homotypic cell adhesion, which may imply a role for CD13 in inflammatory monocyte trafficking. Here, we show that ligation and clustering of CD13 by mAb or viral ligands potently induce myeloid cell/endothelial adhesion in a signal transduction-dependent manner involving monocytic cytoskeletal...... rearrangement and filopodia formation. Treatment with soluble recombinant (r)CD13 blocks this CD13-dependent adhesion, and CD13 molecules from monocytic and endothelial cells are present in the same immunocomplex, suggesting a direct participation of CD13 in the adhesive interaction. This concept...

  15. Maturation of monocyte-derived dendritic cells with Toll-like receptor 3 and 7/8 ligands combined with prostaglandin E2 results in high interleukin-12 production and cell migration

    NARCIS (Netherlands)

    Boullart, A. C. Inge; Aarntzen, Erik H. J. G.; Verdijk, Pauline; Jacobs, Joannes F. M.; Schuurhuis, Danita H.; Benitez-Ribas, Daniel; Schreibelt, Gerty; van de Rakt, Mandy W. M. M.; Scharenborg, Nicole M.; de Boer, Annemiek; Kramer, Matthijs; Figdor, Carl G.; Punt, Cornelis J. A.; Adema, Gosse J.; de Vries, I. Jolanda M.

    2008-01-01

    Dendritic cells (DC) are professional antigen-presenting cells of the immune system that play a key role in regulating T cell-based immunity. In vivo, the capacity of DC to activate T cells depends on their ability to migrate to the T cell areas of lymph nodes as well as on their maturation state.

  16. Maturation of monocyte-derived dendritic cells with Toll-like receptor 3 and 7/8 ligands combined with prostaglandin E2 results in high interleukin-12 production and cell migration.

    NARCIS (Netherlands)

    Boullart, I.; Aarntzen, E.H.J.G.; Verdijk, P.; Jacobs, J.F.M.; Schuurhuis, D.H.; Benitez-Ribas, D.; Schreibelt, G.; Rakt, M.W.M.M. van de; Scharenborg, N.M.; Boer, A.J. de; Kramer, M.; Figdor, C.G.; Punt, C.J.A.; Adema, G.J.; Vries, I.J.M. de

    2008-01-01

    Dendritic cells (DC) are professional antigen-presenting cells of the immune system that play a key role in regulating T cell-based immunity. In vivo, the capacity of DC to activate T cells depends on their ability to migrate to the T cell areas of lymph nodes as well as on their maturation state.

  17. Monocytes infiltrate the pancreas via the MCP-1/CCR2 pathway and differentiate into stellate cells.

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    Kazuko Ino

    Full Text Available Recent studies have shown that monocytes possess pluripotent plasticity. We previously reported that monocytes could differentiate into hepatic stellate cells. Although stellate cells are also present in the pancreas, their origin remains unclear. An accumulation of enhanced green fluorescent protein (EGFP(+CD45(- cells was observed in the pancreases and livers of chimeric mice, which were transplanted with a single hematopoietic stem cell isolated from EGFP-transgenic mice and treated with carbon tetrachloride (CCl4. Because the vast majority of EGFP(+CD45(- cells in the pancreas expressed stellate cell-associated antigens such as vimentin, desmin, glial fibrillary acidic protein, procollagen-I, and α-smooth muscle actin, they were characterized as pancreatic stellate cells (PaSCs. EGFP(+ PaSCs were also observed in CCl4-treated mice adoptively transferred with monocytes but not with other cell lineages isolated from EGFP-transgenic mice. The expression of monocyte chemoattractant protein-1 (MCP-1 and angiotensin II (Ang II increased in the pancreas of CCl4-treated mice and their respective receptors, C-C chemokine receptor 2 (CCR2 and Ang II type 1 receptor (AT1R, were expressed on Ly6C(high monocytes isolated from EGFP-transgenic mice. We examined the effect of an AT1R antagonist, irbesartan, which is also a CCR2 antagonist, on the migration of monocytes into the pancreas. Monocytes migrated toward MCP-1 but not Ang II in vitro. Irbesartan inhibited not only their in vitro chemotaxis but also in vivo migration of adoptively transferred monocytes from peripheral blood into the pancreas. Irbesartan treatment significantly reduced the numbers of EGFP(+F4/80(+CCR2(+ monocytic cells and EGFP(+ PaSCs in the pancreas of CCl4-treated chimeric mice receiving EGFP(+ bone marrow cells. A specific CCR2 antagonist RS504393 inhibited the occurrence of EGFP(+ PaSCs in injured mice. We propose that CCR2(+ monocytes migrate into the pancreas possibly via the

  18. Lipopolysaccharide induces autotaxin expression in human monocytic THP-1 cells

    International Nuclear Information System (INIS)

    Li Song; Zhang Junjie

    2009-01-01

    Autotaxin (ATX) is a secreted enzyme with lysophospholipase D (lysoPLD) activity, which converts lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA), a bioactive phospholipid involved in numerous biological activities, including cell proliferation, differentiation, and migration. In the present study, we found that bacterial lipopolysaccharide (LPS), a well-known initiator of the inflammatory response, induced ATX expression in monocytic THP-1 cells. The activation of PKR, JNK, and p38 MAPK was required for the ATX induction. The LPS-induced ATX in THP-1 cells was characterized as the β isoform. In the presence of LPC, ATX could promote the migrations of THP-1 and Jurkat cells, which was inhibited by pertussis toxin (PTX), an inhibitor of Gi-mediated LPA receptor signaling. In summary, LPS induces ATX expression in THP-1 cells via a PKR, JNK and p38 MAPK-mediated mechanism, and the ATX induction is likely to enhance immune cell migration in proinflammatory response by regulating LPA levels in the microenvironment.

  19. Increased migration of monocytes in essential hypertension is associated with increased transient receptor potential channel canonical type 3 channels

    DEFF Research Database (Denmark)

    Zhao, Zhigang; Ni, Yinxing; Chen, Jing

    2012-01-01

    Increased transient receptor potential canonical type 3 (TRPC3) channels have been observed in patients with essential hypertension. In the present study we tested the hypothesis that increased monocyte migration is associated with increased TRPC3 expression. Monocyte migration assay was performe...

  20. Live Brugia malayi microfilariae inhibit transendothelial migration of neutrophils and monocytes.

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    Jan-Hendrik Schroeder

    Full Text Available Lymphatic filariasis is a major tropical disease caused by the parasite Brugia malayi. Microfilariae (Mf circulate in the peripheral blood for 2-3 hours in synchronisation with maximal feeding of the mosquito vector. When absent from the peripheral blood, Mf sequester in the capillaries of the lungs. Mf are therefore in close contact with vascular endothelial cells (EC and may induce EC immune function and/or wound repair mechanisms such as angiogenesis. In this study, Mf were co-cultured with human umbilical vein EC (HUVEC or human lung microvascular EC (HLMVEC and the transendothelial migration of leukocyte subsets was analysed. In addition, the protein and/or mRNA expression of chemokine, cytokine and angiogenic mediators in endothelial cells in the presence of live microfilariae were measured by a combination of cDNA arrays, protein arrays, ELISA and fluorescence antibody tests.Surprisingly, our findings indicate that Mf presence partially blocked transendothelial migration of monocytes and neutrophils, but not lymphocytes. However, Mf exposure did not result in altered vascular EC expression of key mediators of the tethering stage of extravasation, such as ICAM-1, VCAM-1 and various chemokines. To further analyse the immunological function of vascular EC in the presence of Mf, we measured the mRNA and/or protein expression of a number of pro-inflammatory mediators. We found that expression levels of the mediators tested were predominantly unaltered upon B. malayi Mf exposure. In addition, a comparison of angiogenic mediators induced by intact Mf and Wolbachia-depleted Mf revealed that even intact Mf induce the expression of remarkably few angiogenic mediators in vascular EC. Our study suggests that live microfilariae are remarkably inert in their induction and/or activation of vascular cells in their immediate local environment. Overall, this work presents important insights into the immunological function of the vascular endothelium during

  1. PECAM-1 polymorphism affects monocyte adhesion to endothelial cells.

    Science.gov (United States)

    Goodman, Reyna S; Kirton, Christopher M; Oostingh, Gertie J; Schön, Michael P; Clark, Michael R; Bradley, J Andrew; Taylor, Craig J

    2008-02-15

    Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) plays an important role in leukocyte-endothelial cell adhesion and transmigration. Single nucleotide polymorphisms of PECAM-1 encoding amino acid substitutions at positions 98 leucine/valine (L/V), 536 serine/asparagine (S/N), and 643 arginine/glycine (R/G) occur in strong genetic linkage resulting in two common haplotypes (LSR and VNG). These PECAM-1 polymorphisms are associated with graft-versus-host disease after hematopoietic stem cell transplantation and with cardiovascular disease, but whether they influence PECAM-1 function is unknown. We examined the effect of homozygous and heterozygous expression of the PECAM-1 LSR and VNG genotypes on the adhesive interactions of peripheral blood monocytes and activated endothelial cell monolayers under shear stress in a flow-based cell adhesion assay. There was no difference in monocyte adhesion between the two homozygous genotypes of PECAM-1 but when monocytes expressed both alleles in heterozygous form, firm adhesion of monocytes to endothelial cells was markedly increased. PECAM-1 polymorphism expressed in homozygous or heterozygous form by endothelial cells did not influence monocyte adhesion. This is, to our knowledge, the first demonstration that PECAM-1 genotype can alter the level of monocyte binding to endothelial cells and a demonstration that heterozygous expression of a polymorphic protein may lead to altered function.

  2. Effector CD4+ T cells recognize intravascular antigen presented by patrolling monocytes.

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    Westhorpe, Clare L V; Norman, M Ursula; Hall, Pam; Snelgrove, Sarah L; Finsterbusch, Michaela; Li, Anqi; Lo, Camden; Tan, Zhe Hao; Li, Songhui; Nilsson, Susan K; Kitching, A Richard; Hickey, Michael J

    2018-02-21

    Although effector CD4 + T cells readily respond to antigen outside the vasculature, how they respond to intravascular antigens is unknown. Here we show the process of intravascular antigen recognition using intravital multiphoton microscopy of glomeruli. CD4 + T cells undergo intravascular migration within uninflamed glomeruli. Similarly, while MHCII is not expressed by intrinsic glomerular cells, intravascular MHCII-expressing immune cells patrol glomerular capillaries, interacting with CD4 + T cells. Following intravascular deposition of antigen in glomeruli, effector CD4 + T-cell responses, including NFAT1 nuclear translocation and decreased migration, are consistent with antigen recognition. Of the MHCII + immune cells adherent in glomerular capillaries, only monocytes are retained for prolonged durations. These cells can also induce T-cell proliferation in vitro. Moreover, monocyte depletion reduces CD4 + T-cell-dependent glomerular inflammation. These findings indicate that MHCII + monocytes patrolling the glomerular microvasculature can present intravascular antigen to CD4 + T cells within glomerular capillaries, leading to antigen-dependent inflammation.

  3. The CD157-integrin partnership controls transendothelial migration and adhesion of human monocytes.

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    Lo Buono, Nicola; Parrotta, Rossella; Morone, Simona; Bovino, Paola; Nacci, Giulia; Ortolan, Erika; Horenstein, Alberto L; Inzhutova, Alona; Ferrero, Enza; Funaro, Ada

    2011-05-27

    CD157, a member of the CD38 gene family, is an NAD-metabolizing ectoenzyme and a signaling molecule whose role in polarization, migration, and diapedesis of human granulocytes has been documented; however, the molecular events underpinning this role remain to be elucidated. This study focused on the role exerted by CD157 in monocyte migration across the endothelial lining and adhesion to extracellular matrix proteins. The results demonstrated that anti-CD157 antibodies block monocyte transmigration and adhesion to fibronectin and fibrinogen but that CD157 cross-linking is sufficient to overcome the block, suggesting an active signaling role for the molecule. Consistent with this is the observation that CD157 is prevalently located within the detergent-resistant membrane microdomains to which, upon clustering, it promotes the recruitment of β(1) and β(2) integrin, which, in turn, leads to the formation of a multimolecular complex favoring signal transduction. This functional cross-talk with integrins allows CD157 to act as a receptor despite its intrinsic structural inability to do so on its own. Intracellular signals mediated by CD157 rely on the integrin/Src/FAK (focal adhesion kinase) pathway, resulting in increased activity of the MAPK/ERK1/2 and the PI3K/Akt downstream signaling pathways, which are crucial in the control of monocyte transendothelial migration. Collectively, these findings indicate that CD157 acts as a molecular organizer of signaling-competent membrane microdomains and that it forms part of a larger molecular machine ruled by integrins. The CD157-integrin partnership provides optimal adhesion and transmigration of human monocytes.

  4. The CD157-Integrin Partnership Controls Transendothelial Migration and Adhesion of Human Monocytes*

    Science.gov (United States)

    Lo Buono, Nicola; Parrotta, Rossella; Morone, Simona; Bovino, Paola; Nacci, Giulia; Ortolan, Erika; Horenstein, Alberto L.; Inzhutova, Alona; Ferrero, Enza; Funaro, Ada

    2011-01-01

    CD157, a member of the CD38 gene family, is an NAD-metabolizing ectoenzyme and a signaling molecule whose role in polarization, migration, and diapedesis of human granulocytes has been documented; however, the molecular events underpinning this role remain to be elucidated. This study focused on the role exerted by CD157 in monocyte migration across the endothelial lining and adhesion to extracellular matrix proteins. The results demonstrated that anti-CD157 antibodies block monocyte transmigration and adhesion to fibronectin and fibrinogen but that CD157 cross-linking is sufficient to overcome the block, suggesting an active signaling role for the molecule. Consistent with this is the observation that CD157 is prevalently located within the detergent-resistant membrane microdomains to which, upon clustering, it promotes the recruitment of β1 and β2 integrin, which, in turn, leads to the formation of a multimolecular complex favoring signal transduction. This functional cross-talk with integrins allows CD157 to act as a receptor despite its intrinsic structural inability to do so on its own. Intracellular signals mediated by CD157 rely on the integrin/Src/FAK (focal adhesion kinase) pathway, resulting in increased activity of the MAPK/ERK1/2 and the PI3K/Akt downstream signaling pathways, which are crucial in the control of monocyte transendothelial migration. Collectively, these findings indicate that CD157 acts as a molecular organizer of signaling-competent membrane microdomains and that it forms part of a larger molecular machine ruled by integrins. The CD157-integrin partnership provides optimal adhesion and transmigration of human monocytes. PMID:21478153

  5. 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.

  6. Adropin Contributes to Anti-Atherosclerosis by Suppressing Monocyte-Endothelial Cell Adhesion and Smooth Muscle Cell Proliferation

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    Kengo Sato

    2018-04-01

    Full Text Available Adropin, a peptide hormone expressed in liver and brain, is known to improve insulin resistance and endothelial dysfunction. Serum levels of adropin are negatively associated with the severity of coronary artery disease. However, it remains unknown whether adropin could modulate atherogenesis. We assessed the effects of adropin on inflammatory molecule expression and human THP1 monocyte adhesion in human umbilical vein endothelial cells (HUVECs, foam cell formation in THP1 monocyte-derived macrophages, and the migration and proliferation of human aortic smooth muscle cells (HASMCs in vitro and atherogenesis in Apoe−/− mice in vivo. Adropin was expressed in THP1 monocytes, their derived macrophages, HASMCs, and HUVECs. Adropin suppressed tumor necrosis factor α-induced THP1 monocyte adhesion to HUVECs, which was associated with vascular cell adhesion molecule 1 and intercellular adhesion molecule 1 downregulation in HUVECs. Adropin shifted the phenotype to anti-inflammatory M2 rather than pro-inflammatory M1 via peroxisome proliferator-activated receptor γ upregulation during monocyte differentiation into macrophages. Adropin had no significant effects on oxidized low-density lipoprotein-induced foam cell formation in macrophages. In HASMCs, adropin suppressed the migration and proliferation without inducing apoptosis via ERK1/2 and Bax downregulation and phosphoinositide 3-kinase/Akt/Bcl2 upregulation. Chronic administration of adropin to Apoe−/− mice attenuated the development of atherosclerotic lesions in the aorta, with reduced the intra-plaque monocyte/macrophage infiltration and smooth muscle cell content. Thus, adropin could serve as a novel therapeutic target in atherosclerosis and related diseases.

  7. Dexamethasone Suppresses Oxysterol-Induced Differentiation of Monocytic Cells

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    Yonghae Son

    2016-01-01

    Full Text Available Oxysterol like 27-hydroxycholesterol (27OHChol has been reported to induce differentiation of monocytic cells into a mature dendritic cell phenotype. We examined whether dexamethasone (Dx affects 27OHChol-induced differentiation using THP-1 cells. Treatment of monocytic cells with Dx resulted in almost complete inhibition of transcription and surface expression of CD80, CD83, and CD88 induced by 27OHChol. Elevated surface levels of MHC class I and II molecules induced by 27OHChol were reduced to basal levels by treatment with Dx. A decreased endocytosis ability caused by 27OHChol was recovered by Dx. We also examined effects of Dx on expression of CD molecules involved in atherosclerosis. Increased levels of surface protein and transcription of CD105, CD137, and CD166 by treatment with 27OHChol were significantly inhibited by cotreatment with Dx. These results indicate that Dx inhibits 27OHChol-induced differentiation of monocytic cells into a mature dendritic cell phenotype and expression of CD molecules whose levels are associated with atherosclerosis. In addition, we examined phosphorylation of AKT induced by 27OHChol and effect of Dx, where cotreatment with Dx inhibited the phosphorylation of AKT. The current study reports that Dx regulates oxysterol-mediated dendritic cell differentiation of monocytic cells.

  8. Regulation of ICAM-1 in Cells of the Monocyte/Macrophage System in Microgravity

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    Katrin Paulsen

    2015-01-01

    Full Text Available Cells of the immune system are highly sensitive to altered gravity, and the monocyte as well as the macrophage function is proven to be impaired under microgravity conditions. In our study, we investigated the surface expression of ICAM-1 protein and expression of ICAM-1 mRNA in cells of the monocyte/macrophage system in microgravity during clinostat, parabolic flight, sounding rocket, and orbital experiments. In murine BV-2 microglial cells, we detected a downregulation of ICAM-1 expression in clinorotation experiments and a rapid and reversible downregulation in the microgravity phase of parabolic flight experiments. In contrast, ICAM-1 expression increased in macrophage-like differentiated human U937 cells during the microgravity phase of parabolic flights and in long-term microgravity provided by a 2D clinostat or during the orbital SIMBOX/Shenzhou-8 mission. In nondifferentiated U937 cells, no effect of microgravity on ICAM-1 expression could be observed during parabolic flight experiments. We conclude that disturbed immune function in microgravity could be a consequence of ICAM-1 modulation in the monocyte/macrophage system, which in turn could have a strong impact on the interaction with T lymphocytes and cell migration. Thus, ICAM-1 can be considered as a rapid-reacting and sustained gravity-regulated molecule in mammalian cells.

  9. Cell culture plastics with immobilized interleukin-4 for monocyte differentiation

    DEFF Research Database (Denmark)

    Hansen, Morten; Hjortø, Gertrud Malene; Met, Özcan

    2011-01-01

    Standard cell culture plastic was surface modified by passive adsorption or covalent attachment of interleukin (IL)-4 and investigated for its ability to induce differentiation of human monocytes into mature dendritic cells, a process dose-dependently regulated by IL-4. Covalent attachment of IL-4...... in water instead of phosphate-buffered saline. Passively adsorbed IL-4 was observed to induce differentiation to dendritic cells, but analysis of cell culture supernatants revealed that leakage of IL-4 into solution could account for the differentiation observed. Covalent attachment resulted in bound IL-4...... at similar concentrations to the passive adsorption process, as measured by enzyme-linked immunosorbent assays, and the bound IL-4 did not leak into solution to any measurable extent during cell culture. However, covalently bound IL-4 was incapable of inducing monocyte differentiation. This may be caused...

  10. Cell culture plastics with immobilized interleukin-4 for monocyte differentiation

    DEFF Research Database (Denmark)

    Hansen, Morten; Hjortø, Gertrud Malene; Met, Ozcan

    2011-01-01

    in water instead of phosphate-buffered saline. Passively adsorbed IL-4 was observed to induce differentiation to dendritic cells, but analysis of cell culture supernatants revealed that leakage of IL-4 into solution could account for the differentiation observed. Covalent attachment resulted in bound IL-4...... at similar concentrations to the passive adsorption process, as measured by enzyme-linked immunosorbent assays, and the bound IL-4 did not leak into solution to any measurable extent during cell culture. However, covalently bound IL-4 was incapable of inducing monocyte differentiation. This may be caused...

  11. PLACENTAL SECRETORY FACTORS INFLUENCE TO THP-1 CELLS PHENOTYPE AND THP-1 CELLS TRANSENDOTHELIAL MIGRATION

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    O. I. Stepanova

    2013-01-01

    Full Text Available Decidual and placental macrophage pools are renewed due to its transendothelial monocyte migration from peripheral blood. Tissue macrophages control placental development and provide fetomaternal immunological tolerance. Preeclamptic pregnancy is accompanied by increased monocyte migration to decidual tissue and local inflammatory events. Regulatory mechanisms of monocyte recruitment to placental and decidual tissues is still unclear. Therefore we investigated the influence soluble placental factors (SPFs during the first- and third-trimester normal pregnancy, as compared to effects of these factors in preeclamptic pregnancy. We studied biological actions of SPF upon transendothelial migration of monocyte-like THP-1 cells and their phenotypic pattern. Transendothelial migration of THP-1 cells was more intensive with firsttrimester SPFs from normal pregnancy, when compared with third-trimester samples, and it was accompanied by decreased CD11a expression. SPFs from pre-eclamptic pregnancy caused an increase in transendothelial migration of THP-1 cells, as compared to SPFs from normal pregnancies, being accompanied by increased CD11b expression. The present study was supported by grants ГК №  02.740.11.0711, НШ-3594.2010.7, МД-150.2011.7 and a grant from St.-Petersburg Goverment for young scientists.

  12. Regulatory NK cells mediated between immunosuppressive monocytes and dysfunctional T cells in chronic HBV infection.

    Science.gov (United States)

    Li, Haijun; Zhai, Naicui; Wang, Zhongfeng; Song, Hongxiao; Yang, Yang; Cui, An; Li, Tianyang; Wang, Guangyi; Niu, Junqi; Crispe, Ian Nicholas; Su, Lishan; Tu, Zhengkun

    2017-09-12

    HBV infection represents a major health problem worldwide, but the immunological mechanisms by which HBV causes chronic persistent infection remain only partly understood. Recently, cell subsets with suppressive features have been recognised among monocytes and natural killer (NK) cells. Here we examine the effects of HBV on monocytes and NK cells. Monocytes and NK cells derived from chronic HBV-infected patients and healthy controls were purified and characterised for phenotype, gene expression and cytokines secretion by flow cytometry, quantitative real-time (qRT)-PCR, ELISA and western blotting. Culture and coculture of monocytes and NK cells were used to determine NK cell activation, using intracellular cytokines staining. In chronic HBV infection, monocytes express higher levels of PD-L1, HLA-E, interleukin (IL)-10 and TGF-β, and NK cells express higher levels of PD-1, CD94 and IL-10, compared with healthy individuals. HBV employs hepatitis B surface antigen (HBsAg) to induce suppressive monocytes with HLA-E, PD-L1, IL-10 and TGF-β expression via the MyD88/NFκB signalling pathway. HBV-treated monocytes induce NK cells to produce IL-10, via PD-L1 and HLA-E signals. Such NK cells inhibit autologous T cell activation. Our findings reveal an immunosuppressive cascade, in which HBV generates suppressive monocytes, which initiate regulatory NK cells differentiation resulting in T cell inhibition. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  13. Differential induction from X-irradiated human peripheral blood monocytes to dendritic cells

    International Nuclear Information System (INIS)

    Yoshino, Hironori; Takahashi, Kenji; Monzen, Satoru; Kashiwakura, Ikuo

    2008-01-01

    Dendritic cells (DCs) are a type of antigen-presenting cell which plays an essential role in the immune system. To clarify the influences of ionizing radiation on the differentiation to DCs, we focused on human peripheral blood monocytes and investigated whether X-irradiated monocytes can differentiate into DCs. The non-irradiated monocytes and 5 Gy-irradiated monocytes were induced into immature DCs (iDCs) and mature DCs (mDCs) with appropriate cytokine stimulation, and the induced cells from each monocyte expressed each DC-expressing surface antigen such as CD40, CD86 and HLA-DR. However, the expression levels of CD40 and CD86 on the iDCs derived from the 5 Gy-irradiated monocytes were higher than those of iDCs derived from non-irradiated monocytes. Furthermore, the mDCs derived from 5 Gy-irradiated monocytes had significantly less ability to stimulate allogeneic T cells in comparison to the mDCs derived from non-irradiated monocytes. There were no significant differences in the phagocytotic activity of the iDCs and cytokines detected in the supernatants conditioned by the DCs from the non-irradiated and irradiated monocytes. These results suggest that human monocytes which are exposed to ionizing radiation can thus differentiate into DCs, but there is a tendency that X-irradiation leads to an impairment of the function of DCs. (author)

  14. Activation of the canonical Wnt/β-catenin pathway enhances monocyte adhesion to endothelial cells

    International Nuclear Information System (INIS)

    Lee, Dong Kun; Nathan Grantham, R.; Trachte, Aaron L.; Mannion, John D.; Wilson, Colleen L.

    2006-01-01

    Monocyte adhesion to vascular endothelium has been reported to be one of the early processes in the development of atherosclerosis. In an attempt to develop strategies to prevent or delay atherosclerosis progression, we analyzed effects of the Wnt/β-catenin signaling pathway on monocyte adhesion to various human endothelial cells. Adhesion of fluorescein-labeled monocytes to various human endothelial cells was analyzed under a fluorescent microscope. Unlike sodium chloride, lithium chloride enhanced monocyte adhesion to endothelial cells in a dose-dependent manner. We further demonstrated that inhibitors for glycogen synthase kinase (GSK)-3β or proteosome enhanced monocyte-endothelial cell adhesion. Results of semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) indicated that activation of Wnt/β-catenin pathway did not change expression levels of mRNA for adhesion molecules. In conclusion, the canonical Wnt/β-catenin pathway enhanced monocyte-endothelial cell adhesion without changing expression levels of adhesion molecules

  15. Effects of Platelets on Platelet Concentrate Product on the Activation of Human Peripheral Blood Monocyte Cells

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    N Sadat Razavi Hoseini

    2016-02-01

    Full Text Available Introduction: Monocytes can interact with platelets due to their surface molecules such as P-selectin glycoprotein ligand-1 (PSGL-1, and form monocyte-platelet complex. In the present study, the effects of platelets interaction of platelet concentrates (PCs and peripheral blood monocytes were investigated in vitro as a model to predict the probable interactions of these cells and consequently activation of monocytes. Methods: In this experimental study, units of whole blood and PCs were prepared from Tehran Blood Transfusion Center. After isolation of monocytes from the whole blood, these cells were treated with PC- derived platelets. The activation of monocytes was assessed before and after treatment by the analysis of the respiratory burst of monocytes using dihydrorhodamine 123 (DHR-123. The study data were analyzed using the non-parametric test of Wilcoxon. Results: The purity of monocytes was determined as 86.1±2 using NycoPrep method. The respiratory burst of monocytes was increased after exposure with platelets. In fact, the difference was significant when platelets were used on the 5th day of storage (P=0.001. Conclusions: The study findings revealed that platelets have an efficient capacity to stimulate and activate monocytes. The possible involvement of molecules in the interaction of platelet-monocyte demand to be further studied in future.

  16. Monocyte enrichment from leukapheresis products by using the Elutra cell separator.

    Science.gov (United States)

    Kim, Sinyoung; Kim, Hyun Ok; Baek, Eun-Jung; Choi, Youjeong; Kim, Han-Soo; Lee, Min-Geul

    2007-12-01

    Dendritic cells (DCs), used in clinical trials for cancer immunotherapy, require processing on an expanded scale to conform to current good manufacturing practice guidelines. This study evaluated a large-scale monocyte enrichment procedure with a commercially available cell separator (Elutra, Gambro BCT) and analyzed the capacity of enriched monocytes to differentiate into DCs. Mononuclear cells were collected in two patients with malignant melanoma and seven healthy donors by leukapheresis. Continuous-counterflow elutriation with the Elutra was performed to enrich and purify monocytes from leukapheresis products. Purity and recovery of enriched monocytes were analyzed by flow cytometry. DCs were generated from the elutriated monocytes and characterized by phenotypic surface marker and stimulatory capacity in an allogeneic mixed lymphocyte reaction. In the leukapheresis products, the total MNC count was 7.3 x 10(9) +/- 0.7 x 10(9) and the mean percentage of CD14+ monocytes was 16.5 +/- 3.8 percent, which increased to 68.9 +/- 7.4 percent after elutriation with the Elutra. The mean monocyte recovery was 94.3 percent. Elutriated monocytes were successfully cultured into phenotypically and functionally mature DCs. These results indicate that the Elutra cell separator allows for fast and easy enrichment of monocytes within a closed system. Furthermore, these monocytes can be differentiated into functionally mature DCs. Compared to plastic adherence and immunomagnetic selection methods, the elutriation procedure is inexpensive, efficient, and very effective.

  17. Monocytic leukemias.

    Science.gov (United States)

    Shaw, M T

    1980-05-01

    The monocytic leukemias may be subdivided into acute monocytic leukemia, acute myelomonocytic leukemia, and subacute and chronic myelomonocytic leukemia. The clinical features of acute monocytic and acute myelomonocytic leukemias are similar and are manifestations of bone marrow failure. Gingival hypertrophy and skin infiltration are more frequent in acute monocytic leukemia. Cytomorphologically the blast cells in acute monocytic leukemia may be undifferentiated or differentiated, whereas in the acute myelomonocytic variety there are mixed populations of monocytic and myeloblastic cells. Cytochemical characteristics include strongly positive reactions for nonspecific esterase, inhibited by fluoride. The functional characteristics of acute monocytic and acute myelomonocytic cells resemble those of monocytes and include glass adherence and phagocytoses, the presence of Fc receptors for IgG and C'3, and the production of colony stimulating activity. Subacute and chronic myelomonocytic leukemias are insidious and slowly progressive diseases characterized by anemia and peripheral blood monocytosis. Atypical monocytes called paramyeloid cells are characteristic. The drugs used in the treatment of acute monocytic and acute myelomonocytic leukemias include cytosine arabinoside, the anthracyclines, and VP 16-213. Drug therapy in subacute and chronic myelomonocytic leukemias is not usually indicated, although VP 16-213 has been claimed to be effective.

  18. Transcript and protein analysis reveals better survival skills of monocyte-derived dendritic cells compared to monocytes during oxidative stress.

    Directory of Open Access Journals (Sweden)

    Ilse Van Brussel

    Full Text Available BACKGROUND: Dendritic cells (DCs, professional antigen-presenting cells with the unique ability to initiate primary T-cell responses, are present in atherosclerotic lesions where they are exposed to oxidative stress that generates cytotoxic reactive oxygen species (ROS. A large body of evidence indicates that cell death is a major modulating factor of atherogenesis. We examined antioxidant defence systems of human monocyte-derived (moDCs and monocytes in response to oxidative stress. METHODS: Oxidative stress was induced by addition of tertiary-butylhydroperoxide (tert-BHP, 30 min. Cellular responses were evaluated using flow cytometry and confocal live cell imaging (both using 5-(and-6-chloromethyl-2,7-dichlorodihydrofluorescein diacetate, CM-H(2DCFDA. Viability was assessed by the neutral red assay. Total RNA was extracted for a PCR profiler array. Five genes were selected for confirmation by Taqman gene expression assays, and by immunoblotting or immunohistochemistry for protein levels. RESULTS: Tert-BHP increased CM-H(2DCFDA fluorescence and caused cell death. Interestingly, all processes occurred more slowly in moDCs than in monocytes. The mRNA profiler array showed more than 2-fold differential expression of 32 oxidative stress-related genes in unstimulated moDCs, including peroxiredoxin-2 (PRDX2, an enzyme reducing hydrogen peroxide and lipid peroxides. PRDX2 upregulation was confirmed by Taqman assays, immunoblotting and immunohistochemistry. Silencing PRDX2 in moDCs by means of siRNA significantly increased CM-DCF fluorescence and cell death upon tert-BHP-stimulation. CONCLUSIONS: Our results indicate that moDCs exhibit higher intracellular antioxidant capacities, making them better equipped to resist oxidative stress than monocytes. Upregulation of PRDX2 is involved in the neutralization of ROS in moDCs. Taken together, this points to better survival skills of DCs in oxidative stress environments, such as atherosclerotic plaques.

  19. Elevated levels of homocysteine increase IL-6 production in monocytic Mono Mac 6 cells

    NARCIS (Netherlands)

    van Aken, B. E.; Jansen, J.; van Deventer, S. J.; Reitsma, P. H.

    2000-01-01

    Hyperhomocysteinemia is a risk factor for atherosclerosis and thrombosis. The aim of this study was to analyze if exposure of monocytic cells to increased levels of homocysteine (HCY) induces the accumulation of inflammatory mediators. Interleukin (IL)-6 production by monocytic cell line Mono Mac 6

  20. Phenotype and Function of CD209+ Bovine Blood Dendritic Cells, Monocyte-Derived-Dendritic Cells and Monocyte-Derived Macrophages.

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    Kun Taek Park

    Full Text Available Phylogenic comparisons of the mononuclear phagocyte system (MPS of humans and mice demonstrate phenotypic divergence of dendritic cell (DC subsets that play similar roles in innate and adaptive immunity. Although differing in phenotype, DC can be classified into four groups according to ontogeny and function: conventional DC (cDC1 and cDC2, plasmacytoid DC (pDC, and monocyte derived DC (MoDC. DC of Artiodactyla (pigs and ruminants can also be sub-classified using this system, allowing direct functional and phenotypic comparison of MoDC and other DC subsets trafficking in blood (bDC. Because of the high volume of blood collections required to study DC, cattle offer the best opportunity to further our understanding of bDC and MoDC function in an outbred large animal species. As reported here, phenotyping DC using a monoclonal antibody (mAb to CD209 revealed CD209 is expressed on the major myeloid population of DC present in blood and MoDC, providing a phenotypic link between these two subsets. Additionally, the present study demonstrates that CD209 is also expressed on monocyte derived macrophages (MoΦ. Functional analysis revealed each of these populations can take up and process antigens (Ags, present them to CD4 and CD8 T cells, and elicit a T-cell recall response. Thus, bDC, MoDC, and MoΦ pulsed with pathogens or candidate vaccine antigens can be used to study factors that modulate DC-driven T-cell priming and differentiation ex vivo.

  1. Effects of TNF-alpha on Endothelial Cell Collective Migration

    Science.gov (United States)

    Chen, Desu; Wu, Di; Helim Aranda-Espinoza, Jose; Losert, Wolfgang

    2013-03-01

    Tumor necrosis factor (TNF-alpha) is a small cell-signaling protein usually released by monocytes and macrophages during an inflammatory response. Previous work had shown the effects of TNF-alpha on single cell morphology, migration, and biomechanical properties. However, the effect on collective migrations remains unexplored. In this work, we have created scratches on monolayers of human umbilical endothelial cells (HUVECs) treated with 25ng/mL TNF-alpha on glass substrates. The wound healing like processes were imaged with phase contrast microscopy. Quantitative analysis of the collective migration of cells treated with TNF-alpha indicates that these cells maintain their persistent motion and alignment better than untreated cells. In addition, the collective migration was characterized by measuring the amount of non-affine deformations of the wound healing monolayer. We found a lower mean non-affinity and narrower distribution of non-affinities upon TNF-alpha stimulation. These results suggest that TNF-alpha introduces a higher degree of organized cell collective migration.

  2. Gamma interferon augments Fc gamma receptor-mediated dengue virus infection of human monocytic cells.

    OpenAIRE

    Kontny, U; Kurane, I; Ennis, F A

    1988-01-01

    It has been reported that anti-dengue antibodies at subneutralizing concentrations augment dengue virus infection of monocytic cells. This is due to the increased uptake of dengue virus in the form of virus-antibody complexes by cells via Fc gamma receptors. We analyzed the effects of recombinant human gamma interferon (rIFN-gamma) on dengue virus infection of human monocytic cells. U937 cells, a human monocytic cell line, were infected with dengue virus in the form of virus-antibody complexe...

  3. Presence of estrogen receptors in human myeloid monocytic cells (THP-1 cell line).

    Science.gov (United States)

    Cutolo, M; Villaggio, B; Bisso, A; Sulli, A; Coviello, D; Dayer, J M

    2001-01-01

    To test THP-1 cells for the presence of estrogen receptors (ER) since studies have demonstrated in vivo and in vitro, the influence of estrogens on cells involved in immune response (i.e. macrophages), and since it has been demonstrated that human myeloid monocytic THP-1 cells acquire phenotypic and functional macrophage-like features after incubation with several cytokines or pharmacological agents. Stimulation of THP-1 cells with phorbol myristate acetate (PMA) to prompt their differentiation into macrophage-like cells and evaluation of the possible induction of ER. The expression of ER was analyzed by immunocytochemical assay, reverse transcriptase polymerase chain reaction (RT-PCR) and Western blot analysis. After stimulation by PMA, the human myeloid monocytic THP-1 cells showed the presence of ER, together with markers of monocytic cell differentiation such as CD68, CD54 and HLA-DR. Estrogen effects may be exerted directly through ER on monocytes/macrophages. PMA-treated THP-1 cells may constitute a useful in vitro model to determine the effects of estrogens on macrophage-like cells and their implications in the inflammatory and immune processes.

  4. Memory T Cell Migration

    OpenAIRE

    Qianqian eZhang; Qianqian eZhang; Fadi G. Lakkis

    2015-01-01

    Immunological memory is a key feature of adaptive immunity. It provides the organism with long-lived and robust protection against infection. In organ transplantation, memory T cells pose a significant threat by causing allograft rejection that is generally resistant to immunosuppressive therapy. Therefore, a more thorough understanding of memory T cell biology is needed to improve the survival of transplanted organs without compromising the host’s ability to fight infections. This review...

  5. IL-4 induces cAMP and cGMP in human monocytic cells

    Directory of Open Access Journals (Sweden)

    B. Dugas

    1995-01-01

    Full Text Available Human monocytes, preincubated with IFN-γ respond to IL-4 by a cGMP increase through activation of an inducible NO synthase. Here, IL-4 was found to induce an accumulation of cGMP (1 – 3 min and cAMP (20 – 25 min in unstimulated monocytes. This was impaired with NOS inhibitors, but also with EGTA and calcium/calmodulin inhibitors. These results suggest that: (1 IL-4 may stimulate different NOS isoforms in resting and IFN-γ activated monocytes, and (2 cAMP accumulation may be partially dependent on the NO pathway. By RT-PCR, a type III constitutive NOS mRNA was detected in U937 monocytic cells. IL-4 also increased the [Ca2+]i in these cells. Different NOS may thus be expressed in monocytic cells depending on their differentiation and the signals they receive.

  6. DYSFUNCTION OF MONOCYTES AND DENDRITIC CELLS IN PATIENTS WITH PREMATURE OVARIAN FAILURE

    NARCIS (Netherlands)

    HOEK, A; VAN KASTEREN, Y; DE HAAN-MEULMAN, M; SCHOEMAKER, J; DREXHAGE, HA

    1993-01-01

    PROBLEM: Due to the presence of ovarian antibodies it has been suggested that premature ovarian failure (POF) belongs to the autoimmune endocrinopathies. Monocytes and the monocyte-derived dendritic cells play a prominent role in the initial stages of endocrine autoimmune reactions: the accumulation

  7. Autocrine CCL19 blocks dendritic cell migration toward weak gradients of CCL21

    DEFF Research Database (Denmark)

    Hansen, Morten; Met, Özcan; Larsen, Niels Bent

    2016-01-01

    Background aims. Maturation of dendritic cells (DCs) induces their homing from peripheral to lymphatic tissues guided by CCL21. However, in vitro matured human monocyte-derived DC cancer vaccines injected intradermally migrate poorly to lymph nodes (LNs). In vitro maturation protocols generate DCs...

  8. Generation of dendritic cells for immunotherapy is minimally impaired by granulocytes in the monocyte preparation.

    Science.gov (United States)

    ten Brinke, Anja; Karsten, Miriam L; Dieker, Miranda C; Zwaginga, Jaap Jan; Vrielink, Hans; Marieke van Ham, S

    2006-01-01

    The growing number of clinical studies, using monocyte-derived DC therapy, requires protocols where a sufficient number of dendritic cell (DCs) are produced according to current Good Manufacturing Practice guidelines. Therefore, a closed culture system for the generation of DCs is inevitable. One cost-effective way to isolate monocytes directly from leukapheresis material in a closed system is by elutriation with the Elutra cell separation system. In the Elutra, granulocytes co-purify with the monocytes. Therefore, we studied if and to what extent the presence of granulocytes in a monocyte product affects the generation of mature DCs. The presence of up to 16% granulocytes in the monocyte product had no significant effects on the quality of the DCs formed. The presence of higher granulocyte percentages, however, gradually altered DC quality. In this respect, the presence of higher number of granulocytes induced significant lower migratory capacity of the DCs and lower expression levels of CD80, CD40 and CD86. No effects were observed on the DC yield, cytokine production or the stimulatory capacity of the DCs in MLR. In conclusion, the presence of 20-30% granulocytes in a monocyte product has no major influence on the quality of the DCs generated from monocytes. Therefore, the Elutra is a suitable closed system apparatus to separate monocytes from other blood components for the generation of DCs, even from leukapheresis material which contains a high number of granulocytes.

  9. Inhibition of the differentiation of monocyte-derived dendritic cells by human gingival fibroblasts.

    Directory of Open Access Journals (Sweden)

    Sylvie Séguier

    Full Text Available We investigated whether gingival fibroblasts (GFs can modulate the differentiation and/or maturation of monocyte-derived dendritic cells (DCs and analyzed soluble factors that may be involved in this immune modulation. Experiments were performed using human monocytes in co-culture with human GFs in Transwell® chambers or using monocyte cultures treated with conditioned media (CM from GFs of four donors. The four CM and supernatants from cell culture were assayed by ELISA for cytokines involved in the differentiation of dendritic cells, such as IL-6, VEGF, TGFβ1, IL-13 and IL-10. The maturation of monocyte-derived DCs induced by LPS in presence of CM was also studied. Cell surface phenotype markers were analyzed by flow cytometry. In co-cultures, GFs inhibited the differentiation of monocyte-derived DCs and the strength of this blockade correlated with the GF/monocyte ratio. Conditioned media from GFs showed similar effects, suggesting the involvement of soluble factors produced by GFs. This inhibition was associated with a lower stimulatory activity in MLR of DCs generated with GFs or its CM. Neutralizing antibodies against IL-6 and VEGF significantly (P<0.05 inhibited the inhibitory effect of CM on the differentiation of monocytes-derived DCs and in a dose dependent manner. Our data suggest that IL-6 is the main factor responsible for the inhibition of DCs differentiation mediated by GFs but that VEGF is also involved and constitutes an additional mechanism.

  10. Palmitate-induced inflammatory pathways in human adipose microvascular endothelial cells promote monocyte adhesion and impair insulin transcytosis.

    Science.gov (United States)

    Pillon, Nicolas J; Azizi, Paymon M; Li, Yujin E; Liu, Jun; Wang, Changsen; Chan, Kenny L; Hopperton, Kathryn E; Bazinet, Richard P; Heit, Bryan; Bilan, Philip J; Lee, Warren L; Klip, Amira

    2015-07-01

    Obesity is associated with inflammation and immune cell recruitment to adipose tissue, muscle and intima of atherosclerotic blood vessels. Obesity and hyperlipidemia are also associated with tissue insulin resistance and can compromise insulin delivery to muscle. The muscle/fat microvascular endothelium mediates insulin delivery and facilitates monocyte transmigration, yet its contribution to the consequences of hyperlipidemia is poorly understood. Using primary endothelial cells from human adipose tissue microvasculature (HAMEC), we investigated the effects of physiological levels of fatty acids on endothelial inflammation and function. Expression of cytokines and adhesion molecules was measured by RT-qPCR. Signaling pathways were evaluated by pharmacological manipulation and immunoblotting. Surface expression of adhesion molecules was determined by immunohistochemistry. THP1 monocyte interaction with HAMEC was measured by cell adhesion and migration across transwells. Insulin transcytosis was measured by total internal reflection fluorescence microscopy. Palmitate, but not palmitoleate, elevated the expression of IL-6, IL-8, TLR2 (Toll-like receptor 2), and intercellular adhesion molecule 1 (ICAM-1). HAMEC had markedly low fatty acid uptake and oxidation, and CD36 inhibition did not reverse the palmitate-induced expression of adhesion molecules, suggesting that inflammation did not arise from palmitate uptake/metabolism. Instead, inhibition of TLR4 to NF-κB signaling blunted palmitate-induced ICAM-1 expression. Importantly, palmitate-induced surface expression of ICAM-1 promoted monocyte binding and transmigration. Conversely, palmitate reduced insulin transcytosis, an effect reversed by TLR4 inhibition. In summary, palmitate activates inflammatory pathways in primary microvascular endothelial cells, impairing insulin transport and increasing monocyte transmigration. This behavior may contribute in vivo to reduced tissue insulin action and enhanced tissue

  11. Monocyte function is severely impaired by the fluorochrome calcein acetomethylester

    International Nuclear Information System (INIS)

    Czepluch, Frauke S.; Olieslagers, Serve J.F.; Waltenberger, Johannes

    2007-01-01

    For rapid chemotaxis quantification, cell prelabelling is often performed with the fluorochrome calcein acetomethylester (calcein AM). We investigated whether calcein AM-prelabelling is reliable for monocyte migration analysis. Human monocytes were either preexposed to calcein AM or unlabelled. Monocyte migration towards the potent chemoattractants transforming growth factor-β1 (TGF-β1) and N-formyl-Methionin-Leucin-Phenylalanin (fMLP) was assessed using a 48-well micro-chemotaxis chamber. For quantification, cells were visualized by light microscopy and counted. Surprisingly, random migration of calcein AM-prelabelled cells was significantly impaired compared to the unlabelled control. Accordingly, monocyte chemotaxis towards either TGF-β1 or fMLP dramatically declined. Adherence of calcein AM-labelled monocytes on plastic was also significantly decreased compared to control cells. As adhesion is regarded as an essential component of monocyte migration, the reduced migration observed in calcein AM-labelled monocytes might be explained by a fluorochrome-induced adhesion defect. Therefore, use of the fluorochrome calcein AM cannot be recommended for functional testing of monocytes

  12. Stimulated monocyte IL-6 secretion predicts survival of patients with head and neck squamous cell carcinoma

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    Olofsson Jan

    2008-01-01

    Full Text Available Abstract Background This study was performed in order to determine whether monocyte in vitro function is associated with presence, stage and prognosis of head and neck squamous cell carcinoma (HNSCC disease. Methods Prospective study describing outcome, after at least five years observation, of patients treated for HNSCC disease in relation to their monocyte function. Sixty-five patients with newly diagnosed HNSCC and eighteen control patients were studied. Monocyte responsiveness was assessed by measuring levels of monocyte in vitro interleukin (IL-6 and monocyte chemotactic peptide (MCP-1 secretion after 24 hours of endotoxin stimulation in cultures supplied either with 20% autologous serum (AS or serum free medium (SFM. Survival, and if relevant, cause of death, was determined at least 5 years following primary diagnosis. Results All patients, as a group, had higher in vitro monocyte responsiveness in terms of IL-6 (AS (t = 2.03; p t = 2.49; p in vitro monocyte IL-6 endotoxin responsiveness under the SFM condition was associated with decreased survival rate (Hazard ratio (HR = 2.27; Confidence interval (CI = 1.05–4.88; p p p Conclusion In HNSCC patients, changed monocyte in vitro response to endotoxin, as measured by increased IL-6 (SFM and decreased MCP-1 (AS responsiveness, are negative prognostic factors.

  13. Stem cell migration after irradiation

    International Nuclear Information System (INIS)

    Nothdurft, W.; Fliedner, T.M.

    1979-01-01

    The survival rate of irradiated rodents could be significantly improved by shielding only the small parts of hemopoietic tissues during the course of irradiation. The populations of circulating stem cells in adult organisms are considered to be of some importance for the homeostasis between the many sites of blood cell formation and for the necessary flexibility of hemopoietic response in the face of fluctuating demands. Pluripotent stem cells are migrating through peripheral blood as has been shown for several mammalian species. Under steady state conditions, the exchange of stem cells between the different sites of blood cell formation appears to be restricted. Their presence in blood and the fact that they are in balance with the extravascular stem cell pool may well be of significance for the surveilance of the integrity of local stem cell populations. Any decrease of stem cell population in blood below a critical size results in the rapid immigration of circulating stem cells in order to restore local stem cell pool size. Blood stem cells are involved in the regeneration after whole-body irradiation if the stem cell population in bone marrows is reduced to less than 10% of the normal state. In the animals subjected to partial-body irradiation, the circulating stem cells appear to be the only source for the repopulation of the heavily irradiated, aplastic sites of hemopoietic organs. (Yamashita, S.)

  14. Endogenous pyrogen production by human blood monocytes stimulated by staphylococcal cell wall components.

    OpenAIRE

    Oken, M M; Peterson, P K; Wilkinson, B J

    1981-01-01

    To determine the properties of Staphylococcus aureus contributing to its pyrogenicity, we compared, in human monocytes, endogenous pyrogen production stimulated by heat-killed S. aureus with that stimulated by purified S. aureus cell walls or by particulate peptidoglycan prepared from the same strain. Peptidoglycan, but not the purified cell wall preparation, was found comparable to S. aureus as an endogenous pyrogen stimulus. This finding was associated with a more effective monocyte phagocy...

  15. Differential effects of malignant mesothelioma cells on THP-1 monocytes and macrophages.

    Science.gov (United States)

    Izzi, Valerio; Chiurchiù, Valerio; D'Aquilio, Fabiola; Palumbo, Camilla; Tresoldi, Ilaria; Modesti, Andrea; Baldini, Patrizia M

    2009-02-01

    Malignant mesothelioma (MM) is a highly fatal tumor arising from inner body membranes, whose extensive growth is facilitated by its week immunogenicity and by its ability to blunt the immune response which should arise from the huge mass of leukocytes typically infiltrating this tumor. It has been reported that the inflammatory infiltrate found in MM tissues is characterized by a high prevalence of macrophages. Thus, in this work we evaluated the ability of human MM cells to modulate the inflammatory phenotype of human THP-1 monocytes and macrophages, a widely used in vitro model of monocyte/macrophage differentiation. Furthermore, we tested the hypothesis that the exposure to MM cells could alter the differentiation of THP-1 monocytes favoring the development of alternatively activated, tumor-supporting macrophages. Our data prove for the first time that MM cells can polarize monocytes towards an altered inflammatory phenotype and macrophages towards an immunosuppressive phenotype. Moreover, we demonstrate that monocytes cocultivated with MM cells 'keep a memory' of their encounter with the tumor which influences their differentiation to macrophages. On the whole, we provide evidence that MM cells exert distinct, cell-specific effects on monocytes and macrophages. The thorough characterization of such effects may be of a crucial importance for the rational design of new immunotherapeutic protocols.

  16. Endogenous pyrogen production by human blood monocytes stimulated by staphylococcal cell wall components.

    Science.gov (United States)

    Oken, M M; Peterson, P K; Wilkinson, B J

    1981-01-01

    To determine the properties of Staphylococcus aureus contributing to its pyrogenicity, we compared, in human monocytes, endogenous pyrogen production stimulated by heat-killed S. aureus with that stimulated by purified S. aureus cell walls or by particulate peptidoglycan prepared from the same strain. Peptidoglycan, but not the purified cell wall preparation, was found comparable to S. aureus as an endogenous pyrogen stimulus. This finding was associated with a more effective monocyte phagocytosis of S. aureus and peptidoglycan as compared with that of purified cell walls. Lysostaphin digestion of peptidoglycan markedly reduced its pyrogenicity. To test whether the chemical composition of the ingested particles is important, latex particles were tested as possible stimuli for monocyte endogenous pyrogen release. Although 40 to 68% of monocytes ingested latex particles during the first hour, there was no evidence of endogenous pyrogen activity in the supernatant even when supernatants equivalent to 5.2 X 10(6) monocytes were tested. This study demonstrates that the pyrogenic moiety of the S. aureus cell wall resides in the peptidoglycan component. Phagocytosis is not in itself a pyrogenic stimulus, but rather serves as an effective mechanism to bring about contact between the chemical stimulus and the monocyte.

  17. Proangiogenic hematopoietic cells of monocytic origin: roles in vascular regeneration and pathogenic processes of systemic sclerosis.

    Science.gov (United States)

    Yamaguchi, Yukie; Kuwana, Masataka

    2013-02-01

    New blood vessel formation is critical, not only for organ development and tissue regeneration, but also for various pathologic processes, such as tumor development and vasculopathy. The maintenance of the postnatal vascular system requires constant remodeling, which occurs through angiogenesis, vasculogenesis, and arteriogenesis. Vasculogenesis is mediated by the de novo differentiation of mature endothelial cells from endothelial progenitor cells (EPCs). Early studies provided evidence that bone marrow-derived CD14⁺ monocytes can serve as a subset of EPCs because of their expression of endothelial markers and ability to promote neovascularization in vitro and in vivo. However, the current consensus is that monocytic cells do not give rise to endothelial cells in vivo, but function as support cells, by promoting vascular formation and repair through their immediate recruitment to the site of vascular injury, secretion of proangiogenic factors, and differentiation into mural cells. These monocytes that function in a supporting role in vascular repair are now termed monocytic pro-angiogenic hematopoietic cells (PHCs). Systemic sclerosis (SSc) is a multisystem connective tissue disease characterized by excessive fibrosis and microvasculopathy, along with poor vascular formation and repair. We recently showed that in patients with SSc, circulating monocytic PHCs increase dramatically and have enhanced angiogenic potency. These effects may be induced in response to defective vascular repair machinery. Since CD14⁺ monocytes can also differentiate into fibroblast-like cells that produce extracellular matrix proteins, here we propose a new hypothesis that aberrant monocytic PHCs, once mobilized into circulation, may also contribute to the fibrotic process of SSc.

  18. Platelet-derived stromal cell-derived factor-1 is required for the transformation of circulating monocytes into multipotential cells.

    Directory of Open Access Journals (Sweden)

    Noriyuki Seta

    Full Text Available BACKGROUND: We previously described a primitive cell population derived from human circulating CD14(+ monocytes, named monocyte-derived multipotential cells (MOMCs, which are capable of differentiating into mesenchymal and endothelial lineages. To generate MOMCs in vitro, monocytes are required to bind to fibronectin and be exposed to soluble factor(s derived from circulating CD14(- cells. The present study was conducted to identify factors that induce MOMC differentiation. METHODS: We cultured CD14(+ monocytes on fibronectin in the presence or absence of platelets, CD14(- peripheral blood mononuclear cells, platelet-conditioned medium, or candidate MOMC differentiation factors. The transformation of monocytes into MOMCs was assessed by the presence of spindle-shaped adherent cells, CD34 expression, and the potential to differentiate in vitro into mesenchymal and endothelial lineages. RESULTS: The presence of platelets or platelet-conditioned medium was required to generate MOMCs from monocytes. A screening of candidate platelet-derived soluble factors identified stromal cell-derived factor (SDF-1 as a requirement for generating MOMCs. Blocking an interaction between SDF-1 and its receptor CXCR4 inhibited MOMC generation, further confirming SDF-1's critical role in this process. Finally, circulating MOMC precursors were found to reside in the CD14(+CXCR4(high cell population. CONCLUSION: The interaction of SDF-1 with CXCR4 is essential for the transformation of circulating monocytes into MOMCs.

  19. Stimulated monocyte IL-6 secretion predicts survival of patients with head and neck squamous cell carcinoma

    International Nuclear Information System (INIS)

    Heimdal, John-Helge; Kross, Kenneth; Klementsen, Beate; Olofsson, Jan; Aarstad, Hans Jørgen

    2008-01-01

    This study was performed in order to determine whether monocyte in vitro function is associated with presence, stage and prognosis of head and neck squamous cell carcinoma (HNSCC) disease. Prospective study describing outcome, after at least five years observation, of patients treated for HNSCC disease in relation to their monocyte function. Sixty-five patients with newly diagnosed HNSCC and eighteen control patients were studied. Monocyte responsiveness was assessed by measuring levels of monocyte in vitro interleukin (IL)-6 and monocyte chemotactic peptide (MCP)-1 secretion after 24 hours of endotoxin stimulation in cultures supplied either with 20% autologous serum (AS) or serum free medium (SFM). Survival, and if relevant, cause of death, was determined at least 5 years following primary diagnosis. All patients, as a group, had higher in vitro monocyte responsiveness in terms of IL-6 (AS) (t = 2.03; p < 0.05) and MCP-1 (SFM) (t = 2.49; p < 0.05) compared to controls. Increased in vitro monocyte IL-6 endotoxin responsiveness under the SFM condition was associated with decreased survival rate (Hazard ratio (HR) = 2.27; Confidence interval (CI) = 1.05–4.88; p < 0.05). The predictive value of monocyte responsiveness, as measured by IL-6, was also retained when adjusted for age, gender and disease stage of patients (HR = 2.67; CI = 1.03–6.92; p < 0.05). With respect to MCP-1, low endotoxin-stimulated responsiveness (AS), analysed by Kaplan-Meier method, predicted decreased survival (χ = 4.0; p < 0.05). In HNSCC patients, changed monocyte in vitro response to endotoxin, as measured by increased IL-6 (SFM) and decreased MCP-1 (AS) responsiveness, are negative prognostic factors

  20. Decreased glucose uptake by hyperglycemia is regulated by different mechanisms in human cancer cells and monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Chae Kyun; Chung, June Key; Lee, Yong Jin; Hong, Mee Kyoung; Jeong, Jae Min; Lee, Dong Soo; Lee, Myung Chul [College of Medicine, Seoul National Univ., Seoul (Korea, Republic of)

    2002-04-01

    To clarify the difference in glucose uptake between human cancer cells and monocytes, we studied ({sup 18}F) fluorodeoxyglucose (FDG) uptake in three human colon cancer cell lines (SNU-C2A, SNU-C4, SNU-C5), one human lung cancer cell line (NCI-H522), and human peripheral blood monocytes. The FDG uptake of both cancer cells and monocytes was increased in glucose-free medium, but decreased in the medium containing 16.7 mM glucose (hyperglycemic). The level of Glut1 mRNA decreased in human colon cancer cells and NCI-H522 under hyperglycemic condition. Glut1 protein expression was also decreased in the four human cancer cell lines under hyperglycemic condition, whereas it was consistently undetectable in monocytes. SNU-C2A, SNU-C4 and NCI-H522 showed a similar level of hexokinase activity (7.5-10.8 mU/mg), while SNU-C5 and moncytes showed lower range of hexokinase activity (4.3-6.5 mU/mg). These data suggest that glucose uptake is regulated by different mechanisms in human cancer cells and monocytes.

  1. Decreased glucose uptake by hyperglycemia is regulated by different mechanisms in human cancer cells and monocytes

    International Nuclear Information System (INIS)

    Kim, Chae Kyun; Chung, June Key; Lee, Yong Jin; Hong, Mee Kyoung; Jeong, Jae Min; Lee, Dong Soo; Lee, Myung Chul

    2002-01-01

    To clarify the difference in glucose uptake between human cancer cells and monocytes, we studied ( 18 F) fluorodeoxyglucose (FDG) uptake in three human colon cancer cell lines (SNU-C2A, SNU-C4, SNU-C5), one human lung cancer cell line (NCI-H522), and human peripheral blood monocytes. The FDG uptake of both cancer cells and monocytes was increased in glucose-free medium, but decreased in the medium containing 16.7 mM glucose (hyperglycemic). The level of Glut1 mRNA decreased in human colon cancer cells and NCI-H522 under hyperglycemic condition. Glut1 protein expression was also decreased in the four human cancer cell lines under hyperglycemic condition, whereas it was consistently undetectable in monocytes. SNU-C2A, SNU-C4 and NCI-H522 showed a similar level of hexokinase activity (7.5-10.8 mU/mg), while SNU-C5 and moncytes showed lower range of hexokinase activity (4.3-6.5 mU/mg). These data suggest that glucose uptake is regulated by different mechanisms in human cancer cells and monocytes

  2. A rapid crosstalk of human gammadelta T cells and monocytes drives the acute inflammation in bacterial infections.

    Directory of Open Access Journals (Sweden)

    Matthias Eberl

    2009-02-01

    Full Text Available Vgamma9/Vdelta2 T cells are a minor subset of T cells in human blood and differ from other T cells by their immediate responsiveness to microbes. We previously demonstrated that the primary target for Vgamma9/Vdelta2 T cells is (E-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP, an essential metabolite produced by a large range of pathogens. Here we wished to study the consequence of this unique responsiveness in microbial infection. The majority of peripheral Vgamma9/Vdelta2 T cells shares migration properties with circulating monocytes, which explains the presence of these two distinct blood cell types in the inflammatory infiltrate at sites of infection and suggests that they synergize in anti-microbial immune responses. Our present findings demonstrate a rapid and HMB-PP-dependent crosstalk between Vgamma9/Vdelta2 T cells and autologous monocytes that results in the immediate production of inflammatory mediators including the cytokines interleukin (IL-6, interferon (IFN-gamma, tumor necrosis factor (TNF-alpha, and oncostatin M (OSM; the chemokines CCL2, CXCL8, and CXCL10; and TNF-related apoptosis-inducing ligand (TRAIL. Moreover, under these co-culture conditions monocytes differentiate within 18 hours into inflammatory dendritic cells (DCs with antigen-presenting functions. Addition of further microbial stimuli (lipopolysaccharide, peptidoglycan induces CCR7 and enables these inflammatory DCs to trigger the generation of CD4(+ effector alphabeta T cells expressing IFN-gamma and/or IL-17. Importantly, our in vitro model replicates the responsiveness to microbes of effluent cells from peritoneal dialysis (PD patients and translates directly to episodes of acute PD-associated bacterial peritonitis, where Vgamma9/Vdelta2 T cell numbers and soluble inflammatory mediators are elevated in patients infected with HMB-PP-producing pathogens. Collectively, these findings suggest a direct link between invading pathogens, microbe

  3. Systemic T Cells Immunosuppression of Glioma Stem Cell-Derived Exosomes Is Mediated by Monocytic Myeloid-Derived Suppressor Cells.

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    Rossana Domenis

    Full Text Available A major contributing factor to glioma development and progression is its ability to evade the immune system. Nano-meter sized vesicles, exosomes, secreted by glioma-stem cells (GSC can act as mediators of intercellular communication to promote tumor immune escape. Here, we investigated the immunomodulatory properties of GCS-derived exosomes on different peripheral immune cell populations. Healthy donor peripheral blood mononuclear cells (PBMCs stimulated with anti-CD3, anti-CD28 and IL-2, were treated with GSC-derived exosomes. Phenotypic characterization, cell proliferation, Th1/Th2 cytokine secretion and intracellular cytokine production were analysed by distinguishing among effector T cells, regulatory T cells and monocytes. In unfractionated PBMCs, GSC-derived exosomes inhibited T cell activation (CD25 and CD69 expression, proliferation and Th1 cytokine production, and did not affect cell viability or regulatory T-cell suppression ability. Furthermore, exosomes were able to enhance proliferation of purified CD4+ T cells. In PBMCs culture, glioma-derived exosomes directly promoted IL-10 and arginase-1 production and downregulation of HLA-DR by unstimulated CD14+ monocytic cells, that displayed an immunophenotype resembling that of monocytic myeloid-derived suppressor cells (Mo-MDSCs. Importantly, the removal of CD14+ monocytic cell fraction from PBMCs restored T-cell proliferation. The same results were observed with exosomes purified from plasma of glioblastoma patients. Our results indicate that glioma-derived exosomes suppress T-cell immune response by acting on monocyte maturation rather than on direct interaction with T cells. Selective targeting of Mo-MDSC to treat glioma should be considered with regard to how immune cells allow the acquirement of effector functions and therefore counteracting tumor progression.

  4. Modelling collective cell migration of neural crest.

    Science.gov (United States)

    Szabó, András; Mayor, Roberto

    2016-10-01

    Collective cell migration has emerged in the recent decade as an important phenomenon in cell and developmental biology and can be defined as the coordinated and cooperative movement of groups of cells. Most studies concentrate on tightly connected epithelial tissues, even though collective migration does not require a constant physical contact. Movement of mesenchymal cells is more independent, making their emergent collective behaviour less intuitive and therefore lending importance to computational modelling. Here we focus on such modelling efforts that aim to understand the collective migration of neural crest cells, a mesenchymal embryonic population that migrates large distances as a group during early vertebrate development. By comparing different models of neural crest migration, we emphasize the similarity and complementary nature of these approaches and suggest a future direction for the field. The principles derived from neural crest modelling could aid understanding the collective migration of other mesenchymal cell types. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. Do migrating cells need a nucleus?

    Science.gov (United States)

    Hawkins, Rhoda J

    2018-03-05

    How the nucleus affects cell polarity and migration is unclear. In this issue, Graham et al. (2018. J. Cell Biol. https://doi.org/10.1083/jcb.201706097) show that enucleated cells polarize and migrate in two but not three dimensions and propose that the nucleus is a necessary component of the molecular clutch regulating normal mechanical responses. © 2018 Hawkins.

  6. Collective cell migration during inflammatory response

    Science.gov (United States)

    Wu, Di; Stroka, Kimberly; Aranda-Espinoza, Helim

    2012-02-01

    Wound scratch healing assays of endothelial cell monolayers is a simple model to study collective cell migration as a function of biological signals. A signal of particular interest is the immune response, which after initial wounding in vivo causes the release of various inflammatory factors such as tumor necrosis alpha (TNF-α). TNF-α is an innate inflammatory cytokine that can induce cell growth, cell necrosis, and change cell morphology. We studied the effects of TNF-α on collective cell migration using the wound healing assays and measured several migration metrics, such as rate of scratch closure, velocities of leading edge and bulk cells, closure index, and velocity correlation functions between migrating cells. We observed that TNF-α alters all migratory metrics as a function of the size of the scratch and TNF-α content. The changes observed in migration correlate with actin reorganization upon TNF-α exposure.

  7. Soluble fibrin inhibits monocyte adherence and cytotoxicity against tumor cells: implications for cancer metastasis

    Directory of Open Access Journals (Sweden)

    Patel Shonak

    2006-08-01

    Full Text Available Abstract Background Soluble fibrin (sFn is a marker for disseminated intravascular coagulation and may have prognostic significance, especially in metastasis. However, a role for sFn in the etiology of metastatic cancer growth has not been extensively studied. We have reported that sFn cross-linked platelet binding to tumor cells via the major platelet fibrin receptor αIIbβ3, and tumor cell CD54 (ICAM-1, which is the receptor for two of the leukocyte β2 integrins (αLβ2 and aMβ2. We hypothesized that sFn may also affect leukocyte adherence, recognition, and killing of tumor cells. Furthermore, in a rat experimental metastasis model sFn pre-treatment of tumor cells enhanced metastasis by over 60% compared to untreated cells. Other studies have shown that fibrin(ogen binds to the monocyte integrin αMβ2. This study therefore sought to investigate the effect of sFn on β2 integrin mediated monocyte adherence and killing of tumor cells. Methods The role of sFn in monocyte adherence and cytotoxicity against tumor cells was initially studied using static microplate adherence and cytotoxicity assays, and under physiologically relevant flow conditions in a microscope perfusion incubator system. Blocking studies were performed using monoclonal antibodies specific for β2 integrins and CD54, and specific peptides which inhibit sFn binding to these receptors. Results Enhancement of monocyte/tumor cell adherence was observed when only one cell type was bound to sFn, but profound inhibition was observed when sFn was bound to both monocytes and tumor cells. This effect was also reflected in the pattern of monocyte cytotoxicity. Studies using monoclonal blocking antibodies and specific blocking peptides (which did not affect normal coagulation showed that the predominant mechanism of fibrin inhibition is via its binding to αMβ2 on monocytes, and to CD54 on both leukocytes and tumor cells. Conclusion sFn inhibits monocyte adherence and cytotoxicity of

  8. Protective properties of artichoke (Cynara scolymus) against oxidative stress induced in cultured endothelial cells and monocytes.

    Science.gov (United States)

    Zapolska-Downar, Danuta; Zapolski-Downar, Andrzej; Naruszewicz, Marek; Siennicka, Aldona; Krasnodebska, Barbara; Kołdziej, Blanka

    2002-11-01

    It is currently believed that oxidative stress and inflammation play a significant role in atherogenesis. Artichoke extract exhibits hypolipemic properties and contains numerous active substances with antioxidant properties in vitro. We have studied the influence of aqueous and ethanolic extracts from artichoke on intracellular oxidative stress stimulated by inflammatory mediators (TNFalpha and LPS) and ox-LDL in endothelial cells and monocytes. Oxidative stress which reflects the intracellular production of reactive oxygen species (ROS) was followed by measuring the oxidation of 2', 7'-dichlorofluorescin (DCFH) to 2', 7'-dichlorofluorescein (DCF). Agueous and ethanolic extracts from artichoke were found to inhibit basal and stimulated ROS production in endothelial cells and monocytes in dose dependent manner. In endothelial cells, the ethanolic extract (50 microg/ml) reduced ox-LDL-induced intracellular ROS production by 60% (partichoke extracts have marked protective properties against oxidative stress induced by inflammatory mediators and ox-LDL in cultured endothelial cells and monocytes.

  9. Comparative analysis of signature genes in PRRSV-infected porcine monocyte-derived cells at differential activation statuses

    Science.gov (United States)

    Activation statuses of monocytic cells are critically important for antiviral immunity. Devastating viruses like porcine reproductive and respiratory syndrome virus (PRRSV) are capable of directly infecting these cells, subverting host immunity. Monocyte-derived DCs (mDCs) are major target cells in ...

  10. Monocyte-Derived Signals Activate Human Natural Killer Cells in Response to Leishmania Parasites

    Science.gov (United States)

    Messlinger, Helena; Sebald, Heidi; Heger, Lukas; Dudziak, Diana; Bogdan, Christian; Schleicher, Ulrike

    2018-01-01

    Activated natural killer (NK) cells release interferon (IFN)-γ, which is crucial for the control of intracellular pathogens such as Leishmania. In contrast to experimental murine leishmaniasis, the human NK cell response to Leishmania is still poorly characterized. Here, we investigated the interaction of human blood NK cells with promastigotes of different Leishmania species (Leishmania major, Leishmania mexicana, Leishmania infantum, and Leishmania donovani). When peripheral blood mononuclear cells or purified NK cells and monocytes (all derived from healthy blood donors from Germany without a history of leishmaniasis) were exposed to promastigotes, NK cells showed increased surface expression of the activation marker CD69. The extent of this effect varied depending on the Leishmania species; differences between dermotropic and viscerotropic L. infantum strains were not observed. Upregulation of CD69 required direct contact between monocytes and Leishmania and was partly inhibitable by anti-interleukin (IL)-18. Unexpectedly, IL-18 was undetectable in most of the supernatants (SNs) of monocyte/parasite cocultures. Confocal fluorescence microscopy of non-permeabilized cells revealed that Leishmania-infected monocytes trans-presented IL-18 to NK cells. Native, but not heat-treated SNs of monocyte/Leishmania cocultures also induced CD69 on NK cells, indicating the involvement of a soluble heat-labile factor other than IL-18. A role for the NK cell-activating cytokines IL-1β, IL-2, IL-12, IL-15, IL-21, and IFN-α/β was excluded. The increase of CD69 was not paralleled by NK cell IFN-γ production or enhanced cytotoxicity. However, prior exposure of NK cells to Leishmania parasites synergistically increased their IFN-γ release in response to IL-12, which was dependent on endogenous IL-18. CD1c+ dendritic cells were identified as possible source of Leishmania-induced IL-12. Finally, we observed that direct contact between Leishmania and NK cells reduced the

  11. Monocyte-Derived Signals Activate Human Natural Killer Cells in Response to Leishmania Parasites

    Directory of Open Access Journals (Sweden)

    Helena Messlinger

    2018-01-01

    Full Text Available Activated natural killer (NK cells release interferon (IFN-γ, which is crucial for the control of intracellular pathogens such as Leishmania. In contrast to experimental murine leishmaniasis, the human NK cell response to Leishmania is still poorly characterized. Here, we investigated the interaction of human blood NK cells with promastigotes of different Leishmania species (Leishmania major, Leishmania mexicana, Leishmania infantum, and Leishmania donovani. When peripheral blood mononuclear cells or purified NK cells and monocytes (all derived from healthy blood donors from Germany without a history of leishmaniasis were exposed to promastigotes, NK cells showed increased surface expression of the activation marker CD69. The extent of this effect varied depending on the Leishmania species; differences between dermotropic and viscerotropic L. infantum strains were not observed. Upregulation of CD69 required direct contact between monocytes and Leishmania and was partly inhibitable by anti-interleukin (IL-18. Unexpectedly, IL-18 was undetectable in most of the supernatants (SNs of monocyte/parasite cocultures. Confocal fluorescence microscopy of non-permeabilized cells revealed that Leishmania-infected monocytes trans-presented IL-18 to NK cells. Native, but not heat-treated SNs of monocyte/Leishmania cocultures also induced CD69 on NK cells, indicating the involvement of a soluble heat-labile factor other than IL-18. A role for the NK cell-activating cytokines IL-1β, IL-2, IL-12, IL-15, IL-21, and IFN-α/β was excluded. The increase of CD69 was not paralleled by NK cell IFN-γ production or enhanced cytotoxicity. However, prior exposure of NK cells to Leishmania parasites synergistically increased their IFN-γ release in response to IL-12, which was dependent on endogenous IL-18. CD1c+ dendritic cells were identified as possible source of Leishmania-induced IL-12. Finally, we observed that direct contact between Leishmania and NK cells

  12. Comparative analysis of signature genes in porcine reproductive and respiratory syndrome virus (PRRSV)-infected porcine monocyte-derived dendritic cells at differential activation statuses

    Science.gov (United States)

    Activation statuses of monocytic cells, e.g. monocytes, macrophages and dendritic cells (DCs), are critically important for antiviral immunity. In particular, some devastating viruses, including porcine reproductive and respiratory syndrome virus (PRRSV), are capable of directly infecting these cell...

  13. Dendritic cell, monocyte and T cell activation and response to glatiramer acetate in multiple sclerosis

    DEFF Research Database (Denmark)

    Sellebjerg, F; Hesse, D; Limborg, S

    2012-01-01

    , monocytes and dendritic cells (DC) in relation to disease activity in MS patients treated with GA. Methods: Flow cytometry was used to study the activation of CD4+ T cells and T cell subsets (CD25high and CD26high cells), monocytes and DCs in a cross-sectional study of 39 untreated and 29 GA-treated MS......Background: Treatment with glatiramer acetate (GA) modestly decreases disease activity in multiple sclerosis (MS). The mechanism of action is incompletely understood and differences in the response to treatment between individuals may exist. Objective: To study the activation of CD4+ T cells...... (Bonferroni-corrected p=0.0005). The hazard ratio of relapse was 1.32 (95% confidence interval 1.05–1.64) per 1% increase in CD40+ DCs. Patients treated with GA had fewer CD4+ T cells expressing surface markers associated with T helper type 1 effector responses and more CD4+ T cells expressing surface markers...

  14. CD14+ monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    International Nuclear Information System (INIS)

    Wang, Ding; Chen, Ke; Du, Wei Ting; Han, Zhi-Bo; Ren, He; Chi, Ying

    2010-01-01

    Here, the effect of CD14 + monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-γ (IFN-γ) secretion capacities of CD4 + and CD8 + T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E 2 (PGE 2 ) as an important soluble mediator. CD14 + monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1β, either exogenously added or produced by CD14 + monocytes in culture, could trigger expression of high levels of PGE 2 by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE 2 expression, but also reversed the promotional effect of CD14 + monocytes and partially restored CD4 + and CD8 + T cell proliferation and IFN-γ secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  15. Enhancement of proinflammatory and procoagulant responses to silica particles by monocyte-endothelial cell interactions

    Directory of Open Access Journals (Sweden)

    Liu Xin

    2012-09-01

    Full Text Available Abstract Background Inorganic particles, such as drug carriers or contrast agents, are often introduced into the vascular system. Many key components of the in vivo vascular environment include monocyte-endothelial cell interactions, which are important in the initiation of cardiovascular disease. To better understand the effect of particles on vascular function, the present study explored the direct biological effects of particles on human umbilical vein endothelial cells (HUVECs and monocytes (THP-1 cells. In addition, the integrated effects and possible mechanism of particle-mediated monocyte-endothelial cell interactions were investigated using a coculture model of HUVECs and THP-1 cells. Fe3O4 and SiO2 particles were chosen as the test materials in the present study. Results The cell viability data from an MTS assay showed that exposure to Fe3O4 or SiO2 particles at concentrations of 200 μg/mL and above significantly decreased the cell viability of HUVECs, but no significant loss in viability was observed in the THP-1 cells. TEM images indicated that with the accumulation of SiO2 particles in the cells, the size, structure and morphology of the lysosomes significantly changed in HUVECs, whereas the lysosomes of THP-1 cells were not altered. Our results showed that reactive oxygen species (ROS generation; the production of interleukin (IL-6, IL-8, monocyte chemoattractant protein 1 (MCP-1, tumor necrosis factor (TNF-α and IL-1β; and the expression of CD106, CD62E and tissue factor in HUVECs and monocytes were significantly enhanced to a greater degree in the SiO2-particle-activated cocultures compared with the individual cell types alone. In contrast, exposure to Fe3O4 particles had no impact on the activation of monocytes or endothelial cells in monoculture or coculture. Moreover, using treatment with the supernatants of SiO2-particle-stimulated monocytes or HUVECs, we found that the enhancement of proinflammatory response by SiO2

  16. Multiscale Cues Drive Collective Cell Migration

    Science.gov (United States)

    Nam, Ki-Hwan; Kim, Peter; Wood, David K.; Kwon, Sunghoon; Provenzano, Paolo P.; Kim, Deok-Ho

    2016-07-01

    To investigate complex biophysical relationships driving directed cell migration, we developed a biomimetic platform that allows perturbation of microscale geometric constraints with concomitant nanoscale contact guidance architectures. This permits us to elucidate the influence, and parse out the relative contribution, of multiscale features, and define how these physical inputs are jointly processed with oncogenic signaling. We demonstrate that collective cell migration is profoundly enhanced by the addition of contract guidance cues when not otherwise constrained. However, while nanoscale cues promoted migration in all cases, microscale directed migration cues are dominant as the geometric constraint narrows, a behavior that is well explained by stochastic diffusion anisotropy modeling. Further, oncogene activation (i.e. mutant PIK3CA) resulted in profoundly increased migration where extracellular multiscale directed migration cues and intrinsic signaling synergistically conspire to greatly outperform normal cells or any extracellular guidance cues in isolation.

  17. Heterogeneity of Bovine Peripheral Blood Monocytes

    Directory of Open Access Journals (Sweden)

    Jamal Hussen

    2017-12-01

    Full Text Available Peripheral blood monocytes of several species can be divided into different subpopulations with distinct phenotypic and functional properties. Herein, we aim at reviewing published work regarding the heterogeneity of the recently characterized bovine monocyte subsets. As the heterogeneity of human blood monocytes was widely studied and reviewed, this work focuses on comparing bovine monocyte subsets with their human counterparts regarding their phenotype, adhesion and migration properties, inflammatory and antimicrobial functions, and their ability to interact with neutrophilic granulocytes. In addition, the differentiation of monocyte subsets into functionally polarized macrophages is discussed. Regarding phenotype and distribution in blood, bovine monocyte subsets share similarities with their human counterparts. However, many functional differences exist between monocyte subsets from the two species. In contrast to their pro-inflammatory functions in human, bovine non-classical monocytes show the lowest phagocytosis and reactive oxygen species generation capacity, an absent ability to produce the pro-inflammatory cytokine IL-1β after inflammasome activation, and do not have a role in the early recruitment of neutrophils into inflamed tissues. Classical and intermediate monocytes of both species also differ in their response toward major monocyte-attracting chemokines (CCL2 and CCL5 and neutrophil degranulation products (DGP in vitro. Such differences between homologous monocyte subsets also extend to the development of monocyte-derived macrophages under the influence of chemokines like CCL5 and neutrophil DGP. Whereas the latter induce the differentiation of M1-polarized macrophages in human, bovine monocyte-derived macrophages develop a mixed M1/M2 macrophage phenotype. Although only a few bovine clinical trials analyzed the correlation between changes in monocyte composition and disease, they suggest that functional differences between

  18. Imatinib and Nilotinib Off-Target Effects on Human NK Cells, Monocytes, and M2 Macrophages.

    Science.gov (United States)

    Bellora, Francesca; Dondero, Alessandra; Corrias, Maria Valeria; Casu, Beatrice; Regis, Stefano; Caliendo, Fabio; Moretta, Alessandro; Cazzola, Mario; Elena, Chiara; Vinti, Luciana; Locatelli, Franco; Bottino, Cristina; Castriconi, Roberta

    2017-08-15

    Tyrosine kinase inhibitors (TKIs) are used in the clinical management of hematological neoplasms. Moreover, in solid tumors such as stage 4 neuroblastomas (NB), imatinib showed benefits that might depend on both on-target and immunological off-target effects. We investigated the effects of imatinib and nilotinib on human NK cells, monocytes, and macrophages. High numbers of monocytes died upon exposure to TKI concentrations similar to those achieved in patients. Conversely, NK cells were highly resistant to the TKI cytotoxic effect, were properly activated by immunostimulatory cytokines, and degranulated in the presence of NB cells. In NB, neither drug reduced the expression of ligands for activating NK receptors or upregulated that of HLA class I, B7-H3, PD-L1, and PD-L2, molecules that might limit NK cell function. Interestingly, TKIs modulated the chemokine receptor repertoire of immune cells. Acting at the transcriptional level, they increased the surface expression of CXCR4, an effect observed also in NK cells and monocytes of patients receiving imatinib for chronic myeloid leukemia. Moreover, TKIs reduced the expression of CXCR3 (in NK cells) and CCR1 (in monocytes). Monocytes also decreased the expression of M-CSFR, and low numbers of cells underwent differentiation toward macrophages. M0 and M2 macrophages were highly resistant to TKIs and maintained their phenotypic and functional characteristics. Importantly, also in the presence of TKIs, the M2 immunosuppressive polarization was reverted by TLR engagement, and M1-oriented macrophages fully activated autologous NK cells. Our results contribute to better interpreting the off-target efficacy of TKIs in tumors and to envisaging strategies aimed at facilitating antitumor immune responses. Copyright © 2017 by The American Association of Immunologists, Inc.

  19. Migration of Cells in a Social Context

    DEFF Research Database (Denmark)

    Vedel, Søren; Tay, Savas; Johnston, Darius M.

    2013-01-01

    In multicellular organisms and complex ecosystems, cells migrate in a social context. While this is essential for the basic processes of life such as embryonic development, wound healing and unregulated migration furthermore is implicated in diseases such as cancer, the influence of neighboring...... cells on the individual remains poorly understood. Previous work on isolated cells has revealed a stereotypical migratory behavior, however many aspects of the migration characteristics of cells in populations remained unknown exactly because of this lack of characterization of neighbour-cell influence....... We quantified1 the migration of thousands of individual cells in their population context using time-lapse microscopy, microfluidic cell culture and automated image analysis, and discovered a much richer dynamics in the social context, with significant variations in directionality, displacement...

  20. Immunoregulatory adherent cells in human tuberculosis: radiation-sensitive antigen-specific suppression by monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kleinhenz, M.E.; Ellner, J.J.

    1985-07-01

    In human tuberculosis, adherent mononuclear cells (AMC) selectively depress in vitro responses to the mycobacterial antigen tuberculin purified protein derivative (PPD). The phenotype of this antigen-specific adherent suppressor cell was characterized by examining the functional activity of adherent cells after selective depletion of sheep erythrocyte-rosetting T cells or OKM1-reactive monocytes. Adherent cell suppression was studied in the (/sup 3/H)thymidine-incorporation microculture assay by using T cells rigorously depleted of T cells with surface receptors for the Fc portion of IgG (T gamma cells) as antigen-responsive cells. PPD-induced (/sup 3/H)thymidine incorporation by these non gamma T cells was uniformly reduced (mean, 42% +/- 10% (SD)) when autologous AMC were added to non gamma T cells at a ratio of 1:2. Antigen-specific suppression by AMC was not altered by depletion of sheep erythrocyte-rosetting T cells or treatment with indomethacin. However, AMC treated with OKM1 and complement or gamma irradiation (1,500 rads) no longer suppressed tuberculin responses in vitro. These studies identify the antigen-specific adherent suppressor cell in tuberculosis as an OKM1-reactive, non-erythrocyte-rosetting monocyte. The radiosensitivity of this monocyte immunoregulatory function may facilitate its further definition.

  1. Immunoregulatory adherent cells in human tuberculosis: radiation-sensitive antigen-specific suppression by monocytes

    International Nuclear Information System (INIS)

    Kleinhenz, M.E.; Ellner, J.J.

    1985-01-01

    In human tuberculosis, adherent mononuclear cells (AMC) selectively depress in vitro responses to the mycobacterial antigen tuberculin purified protein derivative (PPD). The phenotype of this antigen-specific adherent suppressor cell was characterized by examining the functional activity of adherent cells after selective depletion of sheep erythrocyte-rosetting T cells or OKM1-reactive monocytes. Adherent cell suppression was studied in the [ 3 H]thymidine-incorporation microculture assay by using T cells rigorously depleted of T cells with surface receptors for the Fc portion of IgG (T gamma cells) as antigen-responsive cells. PPD-induced [ 3 H]thymidine incorporation by these non gamma T cells was uniformly reduced (mean, 42% +/- 10% [SD]) when autologous AMC were added to non gamma T cells at a ratio of 1:2. Antigen-specific suppression by AMC was not altered by depletion of sheep erythrocyte-rosetting T cells or treatment with indomethacin. However, AMC treated with OKM1 and complement or gamma irradiation (1,500 rads) no longer suppressed tuberculin responses in vitro. These studies identify the antigen-specific adherent suppressor cell in tuberculosis as an OKM1-reactive, non-erythrocyte-rosetting monocyte. The radiosensitivity of this monocyte immunoregulatory function may facilitate its further definition

  2. Oral Wild-Type Salmonella Typhi Challenge Induces Activation of Circulating Monocytes and Dendritic Cells in Individuals Who Develop Typhoid Disease.

    Directory of Open Access Journals (Sweden)

    Franklin R Toapanta

    2015-06-01

    Full Text Available A new human oral challenge model with wild-type Salmonella Typhi (S. Typhi was recently developed. In this model, ingestion of 104 CFU of Salmonella resulted in 65% of subjects developing typhoid fever (referred here as typhoid diagnosis -TD- 5-10 days post-challenge. TD criteria included meeting clinical (oral temperature ≥38°C for ≥12 h and/or microbiological (S. Typhi bacteremia endpoints. One of the first lines of defense against pathogens are the cells of the innate immune system (e.g., monocytes, dendritic cells -DCs-. Various changes in circulating monocytes and DCs have been described in the murine S. Typhimurium model; however, whether similar changes are present in humans remains to be explored. To address these questions, a subset of volunteers (5 TD and 3 who did not develop typhoid despite oral challenge -NoTD- were evaluated for changes in circulating monocytes and DCs. Expression of CD38 and CD40 were upregulated in monocytes and DCs in TD volunteers during the disease days (TD-0h to TD-96h. Moreover, integrin α4β7, a gut homing molecule, was upregulated on monocytes but not DCs. CD21 upregulation was only identified in DCs. These changes were not observed among NoTD volunteers despite the same oral challenge. Moreover, monocytes and DCs from NoTD volunteers showed increased binding to S. Typhi one day after challenge. These monocytes showed phosphorylation of p38MAPK, NFkB and Erk1/2 upon stimulation with S. Typhi-LPS-QDot micelles. In contrast, monocytes from TD volunteers showed only a moderate increase in S. Typhi binding 48 h and 96 h post-TD, and only Erk1/2 phosphorylation. This is the first study to describe different activation and migration profiles, as well as differential signaling patterns, in monocytes and DCs which relate directly to the clinical outcome following oral challenge with wild type S. Typhi.

  3. Protective role of klotho protein on epithelial cells upon co-culture with activated or senescent monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Mytych, Jennifer, E-mail: jennifermytych@gmail.com [Institute of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland); Centre of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland); Wos, Izabela; Solek, Przemyslaw; Koziorowski, Marek [Institute of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland); Centre of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland)

    2017-01-15

    Monocytes ensure proper functioning and maintenance of epithelial cells, while good condition of monocytes is a key factor of these interactions. Although, it was shown that in some circumstances, a population of altered monocytes may appear, there is no data regarding their effect on epithelial cells. In this study, using direct co-culture model with LPS-activated and Dox-induced senescent THP-1 monocytes, we reported for the first time ROS-induced DNA damage, reduced metabolic activity, proliferation inhibition and cell cycle arrest followed by p16-, p21- and p27-mediated DNA damage response pathways activation, premature senescence and apoptosis induction in HeLa cells. Also, we show that klotho protein possessing anti-aging and anti-inflammatory characteristics reduced cytotoxic and genotoxic events by inhibition of insulin/IGF-IR and downregulation of TRF1 and TRF2 proteins. Therefore, klotho protein could be considered as a protective factor against changes caused by altered monocytes in epithelial cells. - Highlights: • Activated and senescent THP-1 monocytes induced cyto- and genotoxicity in HeLa cells. • Altered monocytes provoked oxidative and nitrosative stress-induced DNA damage. • DNA damage activated DDR pathways and lead to premature senescence and apoptosis. • Klotho reduced ROS/RNS-mediated toxicity through insulin/IGF-IR pathway inhibition. • Klotho protects HeLa cells from cyto- and genotoxicity induced by altered monocytes.

  4. Protective role of klotho protein on epithelial cells upon co-culture with activated or senescent monocytes

    International Nuclear Information System (INIS)

    Mytych, Jennifer; Wos, Izabela; Solek, Przemyslaw; Koziorowski, Marek

    2017-01-01

    Monocytes ensure proper functioning and maintenance of epithelial cells, while good condition of monocytes is a key factor of these interactions. Although, it was shown that in some circumstances, a population of altered monocytes may appear, there is no data regarding their effect on epithelial cells. In this study, using direct co-culture model with LPS-activated and Dox-induced senescent THP-1 monocytes, we reported for the first time ROS-induced DNA damage, reduced metabolic activity, proliferation inhibition and cell cycle arrest followed by p16-, p21- and p27-mediated DNA damage response pathways activation, premature senescence and apoptosis induction in HeLa cells. Also, we show that klotho protein possessing anti-aging and anti-inflammatory characteristics reduced cytotoxic and genotoxic events by inhibition of insulin/IGF-IR and downregulation of TRF1 and TRF2 proteins. Therefore, klotho protein could be considered as a protective factor against changes caused by altered monocytes in epithelial cells. - Highlights: • Activated and senescent THP-1 monocytes induced cyto- and genotoxicity in HeLa cells. • Altered monocytes provoked oxidative and nitrosative stress-induced DNA damage. • DNA damage activated DDR pathways and lead to premature senescence and apoptosis. • Klotho reduced ROS/RNS-mediated toxicity through insulin/IGF-IR pathway inhibition. • Klotho protects HeLa cells from cyto- and genotoxicity induced by altered monocytes.

  5. Histamine type I (H1) receptor radioligand binding studies on normal T cell subsets, B cells, and monocytes

    International Nuclear Information System (INIS)

    Cameron, W.; Doyle, K.; Rocklin, R.E.

    1986-01-01

    A single, specific binding site for [ 3 H]pyrilamine on normal human T helper, T suppressor, B cells, and monocytes was documented. The binding of the radioligand to its receptor is reversible with cold H 1 antagonist, saturates at 40 to 60 nM, and binding equilibrium is achieved in 2 to 4 min. Using a computer program (Ligand), the authors calculated the dissociation constants, binding capacities, and numbers of receptors per cell for each of the different cell types. Monocytes were found to have the highest affinity for [ 3 H]pyrilamine, followed by T helper cells, B cells and T suppressor cells (K/sub D/ = 44.6 +/- 49.4 nM). T suppressor cells were found to express the higher number of H 1 receptors per cell followed by B cells, T helper cells, and monocytes. The binding affinity for [ 3 H]pyrilamine increased over a 48-hr period, whereas the number of receptors per T cell was essentially unchanged. In contrast, T cells stimulated with Con A or PHA were shown to have a greater than fourfold increase in the number of receptors per cell, whereas the binding affinity for [ 3 H]pyrilamine decreased over the 48-hr period. Although the function of H 1 receptors on T cells, B cells, and monocytes has not been completely defined, this receptor has the potential of playing an important role in the modulating the immune response

  6. A Customizable Chamber for Measuring Cell Migration.

    Science.gov (United States)

    Chowdhury, Aniqa N; Vo, Huu Tri; Olang, Sharon; Mappus, Elliott; Peterson, Brian; Hlavac, Nora; Harvey, Tyler; Dean, Delphine

    2017-03-12

    Cell migration is a vital part of immune responses, growth, and wound healing. Cell migration is a complex process that involves interactions between cells, the extracellular matrix, and soluble and non-soluble chemical factors (e.g., chemoattractants). Standard methods for measuring the migration of cells, such as the Boyden chamber assay, work by counting cells on either side of a divider. These techniques are easy to use; however, they offer little geometric modification for different applications. In contrast, microfluidic devices can be used to observe cell migration with customizable concentration gradients of soluble factors 1 , 2 . However, methods for making microfluidics based assays can be difficult to learn. Here, we describe an easy method for creating cell culture chambers to measure cell migration in response to chemical concentration gradients. Our cell migration chamber method can create different linear concentration gradients in order to study cell migration for a variety of applications. This method is relatively easy to use and is typically performed by undergraduate students. The microchannel chamber was created by placing an acrylic insert in the shape of the final microchannel chamber well into a Petri dish. After this, poly(dimethylsiloxane) (PDMS) was poured on top of the insert. The PDMS was allowed to harden and then the insert was removed. This allowed for the creation of wells in any desired shape or size. Cells may be subsequently added to the microchannel chamber, and soluble agents can be added to one of the wells by soaking an agarose block in the desired agent. The agarose block is added to one of the wells, and time-lapse images can be taken of the microchannel chamber in order to quantify cell migration. Variations to this method can be made for a given application, making this method highly customizable.

  7. Static high-gradient magnetic fields affect the functionality of monocytic cells

    Czech Academy of Sciences Publication Activity Database

    Syrovets, T.; Schmidt, Z.; Buechele, B.; Zablotskyy, Vitaliy A.; Dejneka, Alexandr; Dempsey, N.; Simmet, T.

    2014-01-01

    Roč. 28, č. 1 (2014), s. 1-2 ISSN 0892-6638 Institutional support: RVO:68378271 Keywords : static high-gradient * magnet ic fields * affect the functionality * monocytic cells Subject RIV: BM - Solid Matter Physics ; Magnet ism OBOR OECD: Condensed matter physics (including formerly solid state physics, supercond.)

  8. Development and characterization of a bovine monocyte-derived macrophage cell line

    Science.gov (United States)

    Monocytes circulate in the blood, and later differentiate into macrophages in the tissues. They are components of the innate arm of the immune response and are one of the first lines of defense again invading pathogens. However, they also serve as host cells for intracellular pathogens such as Mycob...

  9. Generation of dendritic cells for immunotherapy is minimally impaired by granulocytes in the monocyte preparation

    NARCIS (Netherlands)

    ten Brinke, Anja; Karsten, Miriam L.; Dieker, Miranda C.; Zwaginga, Jaap Jan; Vrielink, Hans; van Ham, S. Marieke

    2006-01-01

    The growing number of clinical studies, using monocyte-derived DC therapy, requires protocols where a sufficient number of dendritic cell (DCs) are produced according to current Good Manufacturing Practice guidelines. Therefore, a closed culture system for the generation of DCs is inevitable. One

  10. SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

    International Nuclear Information System (INIS)

    Bonifati, Serena; Daly, Michele B.; St Gelais, Corine; Kim, Sun Hee; Hollenbaugh, Joseph A.; Shepard, Caitlin; Kennedy, Edward M.; Kim, Dong-Hyun; Schinazi, Raymond F.; Kim, Baek; Wu, Li

    2016-01-01

    SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G_1/G_0 phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

  11. SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

    Energy Technology Data Exchange (ETDEWEB)

    Bonifati, Serena [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States); Daly, Michele B. [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); St Gelais, Corine; Kim, Sun Hee [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States); Hollenbaugh, Joseph A.; Shepard, Caitlin [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Kennedy, Edward M. [Department of Molecular Genetics and Microbiology, Duke University, Durham, NC (United States); Kim, Dong-Hyun [Department of Pharmacy, School of Pharmacy, Kyung-Hee University, Seoul (Korea, Republic of); Schinazi, Raymond F. [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Kim, Baek, E-mail: baek.kim@emory.edu [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Department of Pharmacy, School of Pharmacy, Kyung-Hee University, Seoul (Korea, Republic of); Wu, Li, E-mail: wu.840@osu.edu [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States)

    2016-08-15

    SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G{sub 1}/G{sub 0} phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

  12. Human neutrophils facilitate tumor cell transendothelial migration.

    LENUS (Irish Health Repository)

    Wu, Q D

    2012-02-03

    Tumor cell extravasation plays a key role in tumor metastasis. However, the precise mechanisms by which tumor cells migrate through normal vascular endothelium remain unclear. In this study, using an in vitro transendothelial migration model, we show that human polymorphonuclear neutrophils (PMN) assist the human breast tumor cell line MDA-MB-231 to cross the endothelial barrier. We found that tumor-conditioned medium (TCM) downregulated PMN cytocidal function, delayed PMN apoptosis, and concomitantly upregulated PMN adhesion molecule expression. These PMN treated with TCM attached to tumor cells and facilitated tumor cell migration through different endothelial monolayers. In contrast, MDA-MB-231 cells alone did not transmigrate. FACScan analysis revealed that these tumor cells expressed high levels of intercellular adhesion molecule-1 (ICAM-1) but did not express CD11a, CD11b, or CD18. Blockage of CD11b and CD18 on PMN and of ICAM-1 on MDA-MB-231 cells significantly attenuated TCM-treated, PMN-mediated tumor cell migration. These tumor cells still possessed the ability to proliferate after PMN-assisted transmigration. These results indicate that TCM-treated PMN may serve as a carrier to assist tumor cell transendothelial migration and suggest that tumor cells can exploit PMN and alter their function to facilitate their extravasation.

  13. Oxidative stress induces monocyte necrosis with enrichment of cell-bound albumin and overexpression of endoplasmic reticulum and mitochondrial chaperones.

    Directory of Open Access Journals (Sweden)

    Haiping Tang

    Full Text Available In the present study, monocytes were treated with 5-azacytidine (azacytidine, gossypol or hydrogen peroxide to induce cell death through oxidative stress. A shift from apoptotic to necrotic cell death occurred when monocytes were treated with 100 µM azacytidine for more than 12 hours. Necrotic monocytes exhibited characteristics, including enrichment of cell-bound albumin and up-regulation of endoplasmic reticulum (ER- and mitochondrial-specific chaperones to protect mitochondrial integrity, which were not observed in other necrotic cells, including HUH-7, A2780, A549 and HOC1a. Our results show that the cell-bound albumin originates in the culture medium rather than from monocyte-derived hepatocytes, and that HSP60 is a potential binding partner of the cell-bound albumin. Proteomic analysis shows that HSP60 and protein disulfide isomerase are the most abundant up-regulated mitochondrial and ER-chaperones, and that both HSP60 and calreticulin are ubiquitinated in necrotic monocytes. In contrast, expression levels of the cytosolic chaperones HSP90 and HSP71 were down-regulated in the azacytidine-treated monocytes, concomitant with an increase in the levels of these chaperones in the cell culture medium. Collectively, our results demonstrates that chaperones from different organelles behave differently in necrotic monocytes, ER- and mitochondrial chaperones being retained and cytosolic and nuclear chaperones being released into the cell culture medium through the ruptured cell membrane. HSP60 may serve as a new target for development of myeloid leukemia treatment.

  14. Lipopolysaccharide-induced expression of cell surface receptors and cell activation of neutrophils and monocytes in whole human blood

    Directory of Open Access Journals (Sweden)

    N.E. Gomes

    2010-09-01

    Full Text Available Lipopolysaccharide (LPS activates neutrophils and monocytes, inducing a wide array of biological activities. LPS rough (R and smooth (S forms signal through Toll-like receptor 4 (TLR4, but differ in their requirement for CD14. Since the R-form LPS can interact with TLR4 independent of CD14 and the differential expression of CD14 on neutrophils and monocytes, we used the S-form LPS from Salmonella abortus equi and the R-form LPS from Salmonella minnesota mutants to evaluate LPS-induced activation of human neutrophils and monocytes in whole blood from healthy volunteers. Expression of cell surface receptors and reactive oxygen species (ROS and nitric oxide (NO generation were measured by flow cytometry in whole blood monocytes and neutrophils. The oxidative burst was quantified by measuring the oxidation of 2',7'-dichlorofluorescein diacetate and the NO production was quantified by measuring the oxidation of 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate. A small increase of TLR4 expression by monocytes was observed after 6 h of LPS stimulation. Monocyte CD14 modulation by LPS was biphasic, with an initial 30% increase followed by a 40% decrease in expression after 6 h of incubation. Expression of CD11b was rapidly up-regulated, doubling after 5 min on monocytes, while down-regulation of CXCR2 was observed on neutrophils, reaching a 50% reduction after 6 h. LPS induced low production of ROS and NO. This study shows a complex LPS-induced cell surface receptor modulation on human monocytes and neutrophils, with up- and down-regulation depending on the receptor. R- and S-form LPS activate human neutrophils similarly, despite the low CD14 expression, if the stimulation occurs in whole blood.

  15. Entropy measures of collective cell migration

    Science.gov (United States)

    Whitby, Ariadne; Parrinello, Simona; Faisal, Aldo

    2015-03-01

    Collective cell migration is a critical process during tissue formation and repair. To this end there is a need to develop tools to quantitatively measure the dynamics of collective cell migration obtained from microscopy data. Drawing on statistical physics we use entropy of velocity fields derived from dense optic flow to quantitatively measure collective migration. Using peripheral nerve repair after injury as experimental system, we study how Schwann cells, guided by fibroblasts, migrate in cord-like structures across the cut, paving a highway for neurons. This process of emergence of organised behaviour is key for successful repair, yet the emergence of leader cells and transition from a random to ordered state is not understood. We find fibroblasts induce correlated directionality in migrating Schwann cells as measured by a decrease in the entropy of motion vector. We show our method is robust with respect to image resolution in time and space, giving a principled assessment of how various molecular mechanisms affect macroscopic features of collective cell migration. Finally, the generality of our method allows us to process both simulated cell movement and microscopic data, enabling principled fitting and comparison of in silico to in vitro. ICCS, Imperial College London & MRC Clinical Sciences Centre.

  16. Raman spectroscopy of individual monocytes reveals that single-beam optical trapping of mononuclear cells occurs by their nucleus

    International Nuclear Information System (INIS)

    Fore, Samantha; Chan, James; Taylor, Douglas; Huser, Thomas

    2011-01-01

    We show that laser tweezers Raman spectroscopy of eukaryotic cells with a significantly larger diameter than the tight focus of a single-beam laser trap leads to optical trapping of the cell by its optically densest part, i.e. typically the cell's nucleus. Raman spectra of individual optically trapped monocytes are compared with location-specific Raman spectra of monocytes adhered to a substrate. When the cell's nucleus is stained with a fluorescent live cell stain, the Raman spectrum of the DNA-specific stain is observed only in the nucleus of individual monocytes. Optically trapped monocytes display the same behavior. We also show that the Raman spectra of individual monocytes exhibit the characteristic Raman signature of cells that have not yet fully differentiated and that individual primary monocytes can be distinguished from transformed monocytes based on their Raman spectra. This work provides further evidence that laser tweezers Raman spectroscopy of individual cells provides meaningful biochemical information in an entirely non-destructive fashion that permits discerning differences between cell types and cellular activity

  17. ADMA induces monocyte adhesion via activation of chemokine receptors in cultured THP-1 cells.

    Science.gov (United States)

    Chen, Meifang; Li, Yuanjian; Yang, Tianlun; Wang, Yongjin; Bai, Yongping; Xie, Xiumei

    2008-08-01

    Asymmetric dimethylarginine (ADMA), an endogenous NOS inhibitor, is also an important inflammatory factor contributing to the development of atherosclerosis (AS). The present study was to test the effect of ADMA on angiotensin (Ang) II-induced monocytic adhesion. Human monocytoid cells (THP-1) or isolated peripheral blood monocyte cells (PBMCs) were incubated with Ang II (10(-6)M) or exogenous ADMA (30 microM) for 4 or 24h in the absence or presence of losartan or antioxidant PDTC. In cultured THP-1 cells, Ang II (10(-6)M) for 24h elevated the level of ADMA in the medium, upregulated the protein expression of protein arginine methyltransferase (PRMT) and decreased the activity of dimethylarginine dimethylaminohydrolase (DDAH). Both of Ang II and ADMA increased monocytic adhesion to human umbilical vein endothelial cells (HUVECs), elevated the levels of monocyte chemoattractant protein (MCP)-1, interleukin (IL)-8 and tumor necrosis factor (TNF)-alpha and upregulated CCR(2) and CXCR(2) mRNA expression, concomitantly with increase in reactive oxygen species (ROS) generation and activation of nuclear factor (NF)-kappaB. Pretreatment with losartan (10 microM) or PDTC (10 microM) abolished the effects mediated by Ang II or ADMA. In isolated PBMCs from healthy individuals, ADMA upregulated the expression of CXCR(2) mRNA, which was attenuated by losartan (10 microM), however, ADMA had no effect on surface protein expression of CCR(2). The present results suggest that ADMA may be involved in monocytic adhesion induced by Ang II via activation of chemokine receptors by ROS/NF-kappaB pathway.

  18. Monocyte galactose/N-acetylgalactosamine-specific C-type lectin receptor stimulant immunotherapy of an experimental glioma. Part 1: stimulatory effects on blood monocytes and monocyte-derived cells of the brain

    Directory of Open Access Journals (Sweden)

    Kushchayev SV

    2012-09-01

    Full Text Available Sergiy V Kushchayev,1 Tejas Sankar,1 Laura L Eggink,4,5 Yevgeniya S Kushchayeva,5 Philip C Wiener,1,5 J Kenneth Hoober,5,6 Jennifer Eschbacher,3 Ruolan Liu,2 Fu-Dong Shi,2 Mohammed G Abdelwahab,4 Adrienne C Scheck,4 Mark C Preul11Neurosurgery Research Laboratory, 2Neuroimmunology Laboratory, 3Department of Pathology, 4Neurooncology Research, Barrow Neurological Institute, St Joseph's Hospital and Medical Center, Phoenix, 5School of Life Sciences, Arizona State University, Tempe, 6Susavion Biosciences, Inc, Tempe, AZ, USAObjectives: Immunotherapy with immunostimulants is an attractive therapy against gliomas. C-type lectin receptors specific for galactose/N-acetylgalactosamine (GCLR regulate cellular differentiation, recognition, and trafficking of monocyte-derived cells. A peptide mimetic of GCLR ligands (GCLRP was used to activate blood monocytes and populations of myeloid-derived cells against a murine glioblastoma.Methods: The ability of GCLRP to stimulate phagocytosis by human microglia and monocyte-derived cells of the brain (MDCB isolated from a human glioblastoma was initially assessed in vitro. Induction of activation markers on blood monocytes was assayed by flow cytometry after administration of GCLRP to naive mice. C57BL/6 mice underwent stereotactic intracranial implantation of GL261 glioma cells and were randomized for tumor size by magnetic resonance imaging, which was also used to assess increase in tumor size. Brain tumor tissues were analyzed using flow cytometry, histology, and enzyme-linked immunosorbent assay with respect to tumor, peritumoral area, and contralateral hemisphere regions.Results: GCLRP exhibited strong stimulatory effect on MDCBs and blood monocytes in vitro and in vivo. GCLRP was associated with an increased percentage of precursors of dendritic cells in the blood (P = 0.003, which differentiated into patrolling macrophages in tumoral (P = 0.001 and peritumoral areas (P = 0.04, rather than into dendritic cells

  19. Effects of HIV infection and ART on phenotype and function of circulating monocytes, natural killer, and innate lymphoid cells.

    Science.gov (United States)

    Nabatanzi, Rose; Cose, Stephen; Joloba, Moses; Jones, Sarah Rowland; Nakanjako, Damalie

    2018-03-15

    HIV infection causes upregulation of markers of inflammation, immune activation and apoptosis of host adaptive, and innate immune cells particularly monocytes, natural killer (NK) and innate lymphoid cells (ILCs). Although antiretroviral therapy (ART) restores CD4 T-cell counts, the persistent aberrant activation of monocytes, NK and ILCs observed likely contributes to the incomplete recovery of T-cell effector functions. A better understanding of the effects of HIV infection and ART on the phenotype and function of circulating monocytes, NK, and ILCs is required to guide development of novel therapeutic interventions to optimize immune recovery.

  20. Anti-CD20 B-cell depletion enhances monocyte reactivity in neuroimmunological disorders

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    Hohlfeld Reinhard

    2011-10-01

    Full Text Available Abstract Background Clinical trials evaluating anti-CD20-mediated B-cell depletion in multiple sclerosis (MS and neuromyelitis optica (NMO generated encouraging results. Our recent studies in the MS model experimental autoimmune encephalomyelitis (EAE attributed clinical benefit to extinction of activated B-cells, but cautioned that depletion of naïve B-cells may be undesirable. We elucidated the regulatory role of un-activated B-cells in EAE and investigated whether anti-CD20 may collaterally diminish regulatory B-cell properties in treatment of neuroimmunological disorders. Methods Myelin oligodendrocyte glycoprotein (MOG peptide-immunized C57Bl/6 mice were depleted of B-cells. Functional consequences for regulatory T-cells (Treg and cytokine production of CD11b+ antigen presenting cells (APC were assessed. Peripheral blood mononuclear cells from 22 patients receiving anti-CD20 and 23 untreated neuroimmunological patients were evaluated for frequencies of B-cells, T-cells and monocytes; monocytic reactivity was determined by TNF-production and expression of signalling lymphocytic activation molecule (SLAM. Results We observed that EAE-exacerbation upon depletion of un-activated B-cells closely correlated with an enhanced production of pro-inflammatory TNF by CD11b+ APC. Paralleling this pre-clinical finding, anti-CD20 treatment of human neuroimmunological disorders increased the relative frequency of monocytes and accentuated pro-inflammatory monocyte function; when reactivated ex vivo, a higher frequency of monocytes from B-cell depleted patients produced TNF and expressed the activation marker SLAM. Conclusions These data suggest that in neuroimmunological disorders, pro-inflammatory APC activity is controlled by a subset of B-cells which is eliminated concomitantly upon anti-CD20 treatment. While this observation does not conflict with the general concept of B-cell depletion in human autoimmunity, it implies that its safety and

  1. Robust and highly-efficient differentiation of functional monocytic cells from human pluripotent stem cells under serum- and feeder cell-free conditions.

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    Masakatsu D Yanagimachi

    Full Text Available Monocytic lineage cells (monocytes, macrophages and dendritic cells play important roles in immune responses and are involved in various pathological conditions. The development of monocytic cells from human embryonic stem cells (ESCs and induced pluripotent stem cells (iPSCs is of particular interest because it provides an unlimited cell source for clinical application and basic research on disease pathology. Although the methods for monocytic cell differentiation from ESCs/iPSCs using embryonic body or feeder co-culture systems have already been established, these methods depend on the use of xenogeneic materials and, therefore, have a relatively poor-reproducibility. Here, we established a robust and highly-efficient method to differentiate functional monocytic cells from ESCs/iPSCs under serum- and feeder cell-free conditions. This method produced 1.3 × 10(6 ± 0.3 × 10(6 floating monocytes from approximately 30 clusters of ESCs/iPSCs 5-6 times per course of differentiation. Such monocytes could be differentiated into functional macrophages and dendritic cells. This method should be useful for regenerative medicine, disease-specific iPSC studies and drug discovery.

  2. Radiation-Induced Thymidine Phosphorylase Upregulation in Rectal Cancer Is Mediated by Tumor-Associated Macrophages by Monocyte Chemoattractant Protein-1 From Cancer Cells

    International Nuclear Information System (INIS)

    Kim, Tae-Dong; Li Ge; Song, Kyoung-Sub; Kim, Jin-Man; Kim, Jun-Sang; Kim, Jong-Seok; Yun, Eun-Jin; Park, Jong-Il; Park, Hae-Duck; Hwang, Byung-Doo; Lim, Kyu; Yoon, Wan-Hee

    2009-01-01

    Purpose: The mechanisms of thymidine phosphorylase (TP) regulation induced by radiation therapy (XRT) in various tumors are poorly understood. We investigated the effect and mechanisms of preoperative XRT on TP expression in rectal cancer tissues. Methods and Materials: TP expression and CD68 and monocyte chemoattractant protein-1 (MCP-1) levels in rectal cancer tissues and cancer cell lines were evaluated before and after XRT in Western blotting, immunohistochemistry, enzyme-linked immunoassay, and reverse transcription-polymerase chain reaction studies. Isolated peripheral blood monocytes were used in the study of chemotaxis under the influence of MCP-1 released by irradiated colon cancer cells. Results: Expression of TP was significantly elevated by 9 Gy of XRT in most rectal cancer tissues but not by higher doses of XRT. In keeping with the close correlation of the increase in both TP expression and the number of tumor-associated macrophages (TAMs), anti-TP immunoreactivity was found in the CD68-positive TAMs and not the neoplastic cells. Expression of MCP-1 was increased in most cases after XRT, and this increase was strongly correlated with TP expression. However, this increase in MCP-1 expression occurred in tumor cells and not stromal cells. The XRT upregulated MCP-1 mRNA and also triggered the release of MCP-1 protein from cultured colon cancer cells. The supernatant of irradiated colon cancer cells showed strong chemotactic activity for monocyte migration, but this activity was completely abolished by neutralizing antibody. Conclusions: Use of XRT induces MCP-1 expression in cancer cells, which causes circulating monocytes to be recruited into TAMs, which then upregulate TP expression in rectal cancer tissues

  3. CD14{sup +} monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ding, E-mail: qqhewd@gmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Chen, Ke, E-mail: chenke_59@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Du, Wei Ting, E-mail: duwtpumc@yahoo.com.cn [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); Han, Zhi-Bo, E-mail: zhibohan@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Ren, He, E-mail: knifesharp2000@hotmail.com [National Engineering Research Center of Cell Products, AmCellGene Co. Ltd, TEDA, Tianjin (China); Chi, Ying, E-mail: caizhuying@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); and others

    2010-09-10

    Here, the effect of CD14{sup +} monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-{gamma} (IFN-{gamma}) secretion capacities of CD4{sup +} and CD8{sup +} T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E{sub 2} (PGE{sub 2}) as an important soluble mediator. CD14{sup +} monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1{beta}, either exogenously added or produced by CD14{sup +} monocytes in culture, could trigger expression of high levels of PGE{sub 2} by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE{sub 2} expression, but also reversed the promotional effect of CD14{sup +} monocytes and partially restored CD4{sup +} and CD8{sup +} T cell proliferation and IFN-{gamma} secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  4. ASIC proteins regulate smooth muscle cell migration.

    Science.gov (United States)

    Grifoni, Samira C; Jernigan, Nikki L; Hamilton, Gina; Drummond, Heather A

    2008-03-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated that Epithelial Na(+)Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration; however, the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence individual ASIC expression and determine the importance of ASIC proteins in wound healing and chemotaxis (PDGF-bb)-initiated migration. We found ASIC1, ASIC2, and ASIC3, but not ASIC4, expression in A10 cells. ASIC1, ASIC2, and ASIC3 siRNA molecules significantly suppressed expression of their respective proteins compared to non-targeting siRNA (RISC) transfected controls by 63%, 44%, and 55%, respectively. Wound healing was inhibited by 10, 20, and 26% compared to RISC controls following suppression of ASIC1, ASIC2, and ASIC3, respectively. Chemotactic migration was inhibited by 30% and 45%, respectively, following suppression of ASIC1 and ASIC3. ASIC2 suppression produced a small, but significant, increase in chemotactic migration (4%). Our data indicate that ASIC expression is required for normal migration and may suggest a novel role for ASIC proteins in cellular migration.

  5. Human β-Defensin 3 Reduces TNF-α-Induced Inflammation and Monocyte Adhesion in Human Umbilical Vein Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Tianying Bian

    2017-01-01

    Full Text Available The aim of this study was to investigate the role of human β-defensin 3 (hBD3 in the initiation stage of atherosclerosis with human umbilical vein endothelial cells (HUVECs triggered by tumor necrosis factor- (TNF- α. The effects of hBD3 on TNF-α-induced endothelial injury and inflammatory response were evaluated. Our data revealed that first, hBD3 reduced the production of interleukin-6 (IL-6, IL-8, monocyte chemoattractant protein-1 (MCP-1, and macrophage migration inhibitory factor (MIF in HUVECs in a dose-dependent manner. In addition, hBD3 significantly prevented intracellular reactive oxygen species (ROS production by HUVECs. Second, western blot analysis demonstrated that hBD3 dose-dependently suppressed the protein levels of intracellular adhesion molecule-1 (ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1 in TNF-α-induced HUVECs. As a result, hBD3 inhibited monocyte adhesion to TNF-α-treated endothelial cells. Additionally, hBD3 suppressed TNF-α-induced F-actin reorganization in HUVECs. Third, hBD3 markedly inhibited NF-κB activation by decreasing the phosphorylation of IKK-α/β, IκB, and p65 subunit within 30 min. Moreover, the phosphorylation of p38 and c-Jun N-terminal protein kinase (JNK in the mitogen-activated protein kinase (MAPK pathway were also inhibited by hBD3 in HUVECs. In conclusion, hBD3 exerts anti-inflammatory and antioxidative effects in endothelial cells in response to TNF-α by inhibiting NF-κB and MAPK signaling.

  6. Distinct Upstream Role of Type I IFN Signaling in Hematopoietic Stem Cell-Derived and Epithelial Resident Cells for Concerted Recruitment of Ly-6Chi Monocytes and NK Cells via CCL2-CCL3 Cascade.

    Directory of Open Access Journals (Sweden)

    Erdenebileg Uyangaa

    Full Text Available Type I interferon (IFN-I-dependent orchestrated mobilization of innate cells in inflamed tissues is believed to play a critical role in controlling replication and CNS-invasion of herpes simplex virus (HSV. However, the crucial regulators and cell populations that are affected by IFN-I to establish the early environment of innate cells in HSV-infected mucosal tissues are largely unknown. Here, we found that IFN-I signaling promoted the differentiation of CCL2-producing Ly-6Chi monocytes and IFN-γ/granzyme B-producing NK cells, whereas deficiency of IFN-I signaling induced Ly-6Clo monocytes producing CXCL1 and CXCL2. More interestingly, recruitment of Ly-6Chi monocytes preceded that of NK cells with the levels peaked at 24 h post-infection in IFN-I-dependent manner, which was kinetically associated with the CCL2-CCL3 cascade response. Early Ly-6Chi monocyte recruitment was governed by CCL2 produced from hematopoietic stem cell (HSC-derived leukocytes, whereas NK cell recruitment predominantly depended on CC chemokines produced by resident epithelial cells. Also, IFN-I signaling in HSC-derived leukocytes appeared to suppress Ly-6Ghi neutrophil recruitment to ameliorate immunopathology. Finally, tissue resident CD11bhiF4/80hi macrophages and CD11chiEpCAM+ dendritic cells appeared to produce initial CCL2 for migration-based self-amplification of early infiltrated Ly-6Chi monocytes upon stimulation by IFN-I produced from infected epithelial cells. Ultimately, these results decipher a detailed IFN-I-dependent pathway that establishes orchestrated mobilization of Ly-6Chi monocytes and NK cells through CCL2-CCL3 cascade response of HSC-derived leukocytes and epithelium-resident cells. Therefore, this cascade response of resident-to-hematopoietic-to-resident cells that drives cytokine-to-chemokine-to-cytokine production to recruit orchestrated innate cells is critical for attenuation of HSV replication in inflamed tissues.

  7. Frontline Science: ATF3 is responsible for the inhibition of TNF-α release and the impaired migration of acute ethanol-exposed monocytes and macrophages.

    Science.gov (United States)

    Hu, Chaojie; Meng, Xiaoming; Huang, Cheng; Shen, Chenlin; Li, Jun

    2017-03-01

    Binge drinking represses host innate immunity and leads to a high risk of infection. Acute EtOH-pretreated macrophages exhibit a decreased production of proinflammatory mediators in response to LPS. ATF3 is induced and counter-regulates the LPS/TLR4 inflammatory cascade. Here, we investigated the potential role of ATF3 in LPS tolerance in acute ethanol-pretreated macrophages. We found that there was an inverse correlation between ATF3 and LPS-induced TNF-α production in acute ethanol-pretreated murine monocytes and macrophages. The knockdown of ATF3 attenuated the inhibitory effects of acute ethanol treatment on LPS-induced TNF-α production. Furthermore, ChIP assays and co-IP demonstrated that ATF3, together with HDAC1, negatively modulated the transcription of TNF-α. In binge-drinking mice challenged with LPS, an up-regulation of ATF3 and HDAC1 and a concomitant decrease in TNF-α were observed. Given that HDAC1 was concomitantly induced in acute ethanol-exposed monocytes and macrophages, we used the HDACi TSA or silenced HDAC1 to explore the role of HDAC1 in acute ethanol-treated macrophages. Our results revealed that TSA treatment and HDAC1 knockdown prevented acute ethanol-induced ATF3 expression and the inhibition of TNF-α transcription. These data indicated a dual role for HDAC1 in acute ethanol-induced LPS tolerance. Furthermore, we showed that the induction of ATF3 led to the impaired migration of BM monocytes and macrophages. Overall, we present a novel role for ATF3 in the inhibition of LPS-induced TNF-α and in the impairment of monocyte and macrophage migration. © Society for Leukocyte Biology.

  8. HIV/SIV infection primes monocytes and dendritic cells for apoptosis.

    Directory of Open Access Journals (Sweden)

    Mireille Laforge

    2011-06-01

    Full Text Available Subversion or exacerbation of antigen-presenting cells (APC death modulates host/pathogen equilibrium. We demonstrated during in vitro differentiation of monocyte-derived macrophages and monocyte-derived dendritic cells (DCs that HIV sensitizes the cells to undergo apoptosis in response to TRAIL and FasL, respectively. In addition, we found that HIV-1 increased the levels of pro-apoptotic Bax and Bak molecules and decreased the levels of anti-apoptotic Mcl-1 and FLIP proteins. To assess the relevance of these observations in the context of an experimental model of HIV infection, we investigated the death of APC during pathogenic SIV-infection in rhesus macaques (RMs. We demonstrated increased apoptosis, during the acute phase, of both peripheral blood DCs and monocytes (CD14(+ from SIV(+RMs, associated with a dysregulation in the balance of pro- and anti-apoptotic molecules. Caspase-inhibitor and death receptors antagonists prevented apoptosis of APCs from SIV(+RMs. Furthermore, increased levels of FasL in the sera of pathogenic SIV(+RMs were detected, compared to non-pathogenic SIV infection of African green monkey. We suggest that inappropriate apoptosis of antigen-presenting cells may contribute to dysregulation of cellular immunity early in the process of HIV/SIV infection.

  9. Orf virus IL-10 reduces monocyte, dendritic cell and mast cell recruitment to inflamed skin.

    Science.gov (United States)

    Bennett, Jared R; Lateef, Zabeen; Fleming, Stephen B; Mercer, Andrew A; Wise, Lyn M

    2016-02-02

    Orf virus (ORFV) is a zoonotic parapoxvirus that causes pustular dermatitis of sheep, and occasionally humans. Despite causing sustained infections, ORFV induces only a transient increase in pro-inflammatory signalling and the trafficking of innate immune cells within the skin seems to be impaired. An explanation for this tempered response to ORFV infection may lie in its expression of a homolog of the anti-inflammatory cytokine, interleukin (IL)-10. Using a murine model in which inflammation was induced by bacterial lipopolysaccharide, we examined the effects of the ORFV-IL-10 protein on immune cell trafficking to and from the skin. ORFV-IL-10 limited the recruitment of blood-derived Gr-1(int)/CD11b(int) monocytes, CD11c(+ve)/MHC-II(+ve) dendritic cells and c-kit(+ve)/FcεR1(+ve) mature mast cells into inflamed skin. ORFV-IL-10 also suppressed the activation of CD11c(+ve)/MHC-II(+ve) dendritic cells within the skin, reducing their trafficking to the draining lymph node. These findings suggest that expression of IL-10 by ORFV may contribute to the impaired trafficking of innate immune cells within infected skin. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Edaravone attenuates monocyte adhesion to endothelial cells induced by oxidized low-density lipoprotein.

    Science.gov (United States)

    Li, Zhijuan; Cheng, Jianxin; Wang, Liping

    2015-10-30

    Oxidized low-density lipoprotein (oxLDL) plays a vital role in recruitment of monocytes to endothelial cells, which is important during early stages of atherosclerosis development. Edaravone, a potent and novel scavenger of free radicals inhibiting hydroxyl radicals, has been clinically used to reduce the neuronal damage following ischemic stroke. In the present study, Edaravone was revealed to markedly reduce oxLDL-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). The inhibitory mechanism of Edaravone was associated with suppression of the chemokine MCP-1 and adhesion molecule VCAM-1 and ICAM-1 expression. In addition, luciferase reporter assay results revealed that administration of Edaravone attenuated the increase in NF-κB transcriptional activity induced by oxLDL. Notably, it's also shown that Edaravone treatment blocked oxLDL induced p65 nuclear translocation in HUVECs. Results indicate that Edaravone negatively regulates endothelial inflammation. Copyright © 2015. Published by Elsevier Inc.

  11. Macrophage Activation Mechanisms in Human Monocytic Cell Line-derived Macrophages.

    Science.gov (United States)

    Sumiya, Yu; Ishikawa, Mami; Inoue, Takahiro; Inui, Toshio; Kuchiike, Daisuke; Kubo, Kentaro; Uto, Yoshihiro; Nishikata, Takahito

    2015-08-01

    Although the mechanisms of macrophage activation are important for cancer immunotherapy, they are poorly understood. Recently, easy and robust assay systems for assessing the macrophage-activating factor (MAF) using monocytic cell line-derived macrophages were established. Gene-expression profiles of U937- and THP-1-derived macrophages were compared using gene expression microarray analysis and their responses against several MAFs were examined by in vitro experiments. Activated states of these macrophages could not be assigned to a specific sub-type but showed, however, different unique characteristics. The unique of monocytic cell line-derived macrophages could provide clues to understand the activation mechanism of macrophages and, therefore, help to develop effective cancer immunotherapy with MAFs. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  12. Characterization of monocyte-derived dendritic cells maturated with IFN-alpha

    DEFF Research Database (Denmark)

    Svane, I M; Nikolajsen, K; Walter, M R

    2006-01-01

    Dendritic cells (DC) are promising candidates for cancer immunotherapy. These cells can be generated from peripheral blood monocytes cultured with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). In order to obtain full functional capacity, maturation is required......, maturation with IFN-alpha has only a small effect on induction of autologous T-cell stimulatory capacity of the DC. However, an increase in DC allogeneic T-cell stimulatory capacity was observed. These data suggest that IFN-alpha has a potential as a maturation agent used in DC-based cancer vaccine trials...

  13. HCMV Displays a Unique Transcriptome of Immunomodulatory Genes in Primary Monocyte-Derived Cell Types.

    Directory of Open Access Journals (Sweden)

    Ellen Van Damme

    Full Text Available Human cytomegalovirus (HCMV is a betaherpesvirus which rarely presents problems in healthy individuals, yet may result in severe morbidity in immunocompromised patients and in immune-naïve neonates. HCMV has a large 235 kb genome with a coding capacity of at least 165 open reading frames (ORFs. This large genome allows complex gene regulation resulting in different sets of transcripts during lytic and latent infection. While latent virus mainly resides within monocytes and CD34+ progenitor cells, reactivation to lytic infection is driven by differentiation towards terminally differentiated myeloid dendritic cells and macrophages. Consequently, it has been suggested that macrophages and dendritic cells contribute to viral spread in vivo. Thus far only limited knowledge is available on the expression of HCMV genes in terminally differentiated myeloid primary cells and whether or not the virus exhibits a different set of lytic genes in primary cells compared with lytic infection in NHDF fibroblasts. To address these questions, we used Illumina next generation sequencing to determine the HCMV transcriptome in macrophages and dendritic cells during lytic infection and compared it to the transcriptome in NHDF fibroblasts. Here, we demonstrate unique expression profiles in macrophages and dendritic cells which significantly differ from the transcriptome in fibroblasts mainly by modulating the expression of viral transcripts involved in immune modulation, cell tropism and viral spread. In a head to head comparison between macrophages and dendritic cells, we observed that factors involved in viral spread and virion composition are differentially regulated suggesting that the plasticity of the virion facilitates the infection of surrounding cells. Taken together, this study provides the full transcript expression analysis of lytic HCMV genes in monocyte-derived type 1 and type 2 macrophages as well as in monocyte-derived dendritic cells. Thereby

  14. Tailored HIV-1 vectors for genetic modification of primary human dendritic cells and monocytes.

    Science.gov (United States)

    Durand, Stéphanie; Nguyen, Xuan-Nhi; Turpin, Jocelyn; Cordeil, Stephanie; Nazaret, Nicolas; Croze, Séverine; Mahieux, Renaud; Lachuer, Joël; Legras-Lachuer, Catherine; Cimarelli, Andrea

    2013-01-01

    Monocyte-derived dendritic cells (MDDCs) play a key role in the regulation of the immune system and are the target of numerous gene therapy applications. The genetic modification of MDDCs is possible with human immunodeficiency virus type 1 (HIV-1)-derived lentiviral vectors (LVs) but requires high viral doses to bypass their natural resistance to viral infection, and this in turn affects their physiological properties. To date, a single viral protein is able to counter this restrictive phenotype, Vpx, a protein derived from members of the HIV-2/simian immunodeficiency virus SM lineage that counters at least two restriction factors present in myeloid cells. By tagging Vpx with a short heterologous membrane-targeting domain, we have obtained HIV-1 LVs incorporating high levels of this protein (HIV-1-Src-Vpx). These vectors efficiently transduce differentiated MDDCs and monocytes either as previously purified populations or as populations within unsorted peripheral blood mononuclear cells (PBMCs). In addition, these vectors can be efficiently pseudotyped with receptor-specific envelopes, further restricting their cellular tropism almost uniquely to MDDCs. Compared to conventional HIV-1 LVs, these novel vectors allow for an efficient genetic modification of MDDCs and, more importantly, do not cause their maturation or affect their survival, which are unwanted side effects of the transduction process. This study describes HIV-1-Src-Vpx LVs as a novel potent tool for the genetic modification of differentiated MDDCs and of circulating monocyte precursors with strong potential for a wide range of gene therapy applications.

  15. Monocyte chemotactic protein-1, RANTES and macrophage migration inhibitory factor levels in gingival crevicular fluid of metabolic syndrome patients with gingivitis.

    Science.gov (United States)

    Gürkan, Ali; Eren, Gülnihal; Çetinkalp, Şevki; Akçay, Yasemin Delen; Emingil, Gülnur; Atilla, Gül

    2016-09-01

    The aim of the present study was to determine gingival crevicular fluid (GCF) levels of monocyte chemotactic protein-1 (MCP-1), regulated on activation, normal T-cell expressed and secreted protein (RANTES) and macrophage migration inhibitory factor (MIF) in metabolic syndrome patients with gingivitis. Twenty metabolic syndrome patients with gingivitis (MSG), 20 MetS patients with clinically healthy periodontium (MSH), 20 systemically healthy subjects with gingivitis and 20 subjects who were both systemically and periodontally healthy were included. Periodontal and systemical parameters were recorded. GCF MCP-1, RANTES and MIF levels were assayed by enzyme-linked immunosorbent assay method. MSG and MSH groups had elevated blood pressure, triglyceride, waist circumference and fasting glucose values in comparison to gingivitis and healthy groups (Pgingivitis groups when compared to those of the MSH and healthy groups (Pgingivitis group had higher MCP-1, RANTES and MIF levels compared to the healthy group (P=0.011, P=0.0001, P=0.011 respectively). The RANTES level of MSG group was significantly higher than those of the gingivitis group (P=0.01), but MCP-1 and MIF levels were similar in the MSG and gingivitis groups (P>0.05). Elevated levels of GCF RANTES in MetS patients with gingivitis might associate with the presence of increased gingival inflammation by MetS. Low-grade systemic inflammation associated with MetS and adipose tissue-derived RANTES might lead to altered GCF RANTES levels in the presence of gingival inflammation. Copyright © 2016. Published by Elsevier Ltd.

  16. GM-CSF Monocyte-Derived Cells and Langerhans Cells As Part of the Dendritic Cell Family

    Directory of Open Access Journals (Sweden)

    Manfred B. Lutz

    2017-10-01

    Full Text Available Dendritic cells (DCs and macrophages (Mph share many characteristics as components of the innate immune system. The criteria to classify the multitude of subsets within the mononuclear phagocyte system are currently phenotype, ontogeny, transcription patterns, epigenetic adaptations, and function. More recently, ontogenetic, transcriptional, and proteomic research approaches uncovered major developmental differences between Flt3L-dependent conventional DCs as compared with Mphs and monocyte-derived DCs (MoDCs, the latter mainly generated in vitro from murine bone marrow-derived DCs (BM-DCs or human CD14+ peripheral blood monocytes. Conversely, in vitro GM-CSF-dependent monocyte-derived Mphs largely resemble MoDCs whereas tissue-resident Mphs show a common embryonic origin from yolk sac and fetal liver with Langerhans cells (LCs. The novel ontogenetic findings opened discussions on the terminology of DCs versus Mphs. Here, we bring forward arguments to facilitate definitions of BM-DCs, MoDCs, and LCs. We propose a group model of terminology for all DC subsets that attempts to encompass both ontogeny and function.

  17. p53/PUMA expression in human pulmonary fibroblasts mediates cell activation and migration in silicosis.

    Science.gov (United States)

    Wang, Wei; Liu, Haijun; Dai, Xiaoniu; Fang, Shencun; Wang, Xingang; Zhang, Yingming; Yao, Honghong; Zhang, Xilong; Chao, Jie

    2015-11-18

    Phagocytosis of SiO2 into the lung causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Clinical evidence has indicated that the activation of alveolar macrophages by SiO2 produces rapid and sustained inflammation characterized by the generation of monocyte chemotactic protein 1, which, in turn, induces fibrosis. However, the details of events downstream of monocyte chemotactic protein 1 activity in pulmonary fibroblasts remain unclear. Here, to elucidate the role of p53 in fibrosis induced by silica, both the upstream molecular mechanisms and the functional effects on cell proliferation and migration were investigated. Experiments using primary cultured adult human pulmonary fibroblasts led to the following results: 1) SiO2 treatment resulted in a rapid and sustained increase in p53 and PUMA protein levels; 2) the MAPK and PI3K pathways were involved in the SiO2-induced alteration of p53 and PUMA expression; and 3) RNA interference targeting p53 and PUMA prevented the SiO2-induced increases in fibroblast activation and migration. Our study elucidated a link between SiO2-induced p53/PUMA expression in fibroblasts and cell migration, thereby providing novel insight into the potential use of p53/PUMA in the development of novel therapeutic strategies for silicosis treatment.

  18. ASIC PROTEINS REGULATE SMOOTH MUSCLE CELL MIGRATION

    OpenAIRE

    Grifoni, Samira C.; Jernigan, Nikki L.; Hamilton, Gina; Drummond, Heather A.

    2007-01-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated Epithelial Na+ Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration, however the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence indi...

  19. Edaravone attenuates monocyte adhesion to endothelial cells induced by oxidized low-density lipoprotein

    International Nuclear Information System (INIS)

    Li, Zhijuan; Cheng, Jianxin; Wang, Liping

    2015-01-01

    Oxidized low-density lipoprotein (oxLDL) plays a vital role in recruitment of monocytes to endothelial cells, which is important during early stages of atherosclerosis development. Edaravone, a potent and novel scavenger of free radicals inhibiting hydroxyl radicals, has been clinically used to reduce the neuronal damage following ischemic stroke. In the present study, Edaravone was revealed to markedly reduce oxLDL-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). The inhibitory mechanism of Edaravone was associated with suppression of the chemokine MCP-1 and adhesion molecule VCAM-1 and ICAM-1 expression. In addition, luciferase reporter assay results revealed that administration of Edaravone attenuated the increase in NF-κB transcriptional activity induced by oxLDL. Notably, it's also shown that Edaravone treatment blocked oxLDL induced p65 nuclear translocation in HUVECs. Results indicate that Edaravone negatively regulates endothelial inflammation. - Highlights: • Edaravone reduces oxLDL-induced monocyte adhesion to HUVECs. • Edaravone attenuates oxLDL-induced expression of MCP-1, VCAM-1, and ICAM-1. • Edaravone reduces NF-κB transcriptional activity and p65 nuclear translocation.

  20. Edaravone attenuates monocyte adhesion to endothelial cells induced by oxidized low-density lipoprotein

    Energy Technology Data Exchange (ETDEWEB)

    Li, Zhijuan, E-mail: zjlee038@163.com; Cheng, Jianxin; Wang, Liping

    2015-10-30

    Oxidized low-density lipoprotein (oxLDL) plays a vital role in recruitment of monocytes to endothelial cells, which is important during early stages of atherosclerosis development. Edaravone, a potent and novel scavenger of free radicals inhibiting hydroxyl radicals, has been clinically used to reduce the neuronal damage following ischemic stroke. In the present study, Edaravone was revealed to markedly reduce oxLDL-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). The inhibitory mechanism of Edaravone was associated with suppression of the chemokine MCP-1 and adhesion molecule VCAM-1 and ICAM-1 expression. In addition, luciferase reporter assay results revealed that administration of Edaravone attenuated the increase in NF-κB transcriptional activity induced by oxLDL. Notably, it's also shown that Edaravone treatment blocked oxLDL induced p65 nuclear translocation in HUVECs. Results indicate that Edaravone negatively regulates endothelial inflammation. - Highlights: • Edaravone reduces oxLDL-induced monocyte adhesion to HUVECs. • Edaravone attenuates oxLDL-induced expression of MCP-1, VCAM-1, and ICAM-1. • Edaravone reduces NF-κB transcriptional activity and p65 nuclear translocation.

  1. Efficient elutriation of monocytes within a closed system (Elutra) for clinical-scale generation of dendritic cells.

    Science.gov (United States)

    Berger, Thomas G; Strasser, Erwin; Smith, Richard; Carste, Curt; Schuler-Thurner, Beatrice; Kaempgen, Eckhart; Schuler, Gerold

    2005-03-01

    Dendritic cells (DC) are promising tools for the immunotherapy of cancer. The induction of tumor-specific T cells and clinical regressions have already been observed in early phase I/II vaccination trials. As DC vaccination is now facing trials with larger patient collectives it becomes increasingly important to obtain large numbers of cells suitable for therapeutic applications under labor- and cost-effective conditions. We describe here a procedure that uses a novel cell separator (Elutra, Gambro BCT) to enrich monocytes from an entire apheresis product within one hour. Cells are separated on the basis of size and to a lesser extent density, by elutriation in a 40-ml conical chamber. The total monocyte recovery following elutriation (n = 6) was 98.53% (+/-8.07%), the recovery in the monocyte-rich fraction 75.45% (+/-11.31%), and the mean purity 82.95% (+/-6.01%). These monocytes can be cultured either in conventional culture dishes or in closed cell culture bags and differentiated, by using GM-CSF+IL-4 followed by a maturation cocktail composed of IL-1beta+IL-6+TNF-alpha+PGE2, into fully mature DC. The Elutra separator allows for fast and easy enrichment of monocytes within a closed system. Subsequently, elutriated monocytes can be successfully cultured into phenotypically and functionally mature DC for immunotherapeutic approaches. The method neither requires a density gradient step to enrich PBMC from leucapheresis products nor does it apply (xenogeneic) antibodies to target monocytes. Isolation of monocytes with Elutra may greatly facilitate future DC-based vaccination approaches.

  2. Cell migration induced by Leishmania (Leishmania) amazonensis, Leishmania (Leishmania) major and Leishmania (Viannia) braziliensis into the peritoneal cavity of BALB/c mice

    OpenAIRE

    DT Wakimoto; KV Gaspareto; TGV Silveira; MVC Lonardoni; SMA Aristides

    2010-01-01

    In American cutaneous leishmaniasis, the initial infection phase is characterized by recruitment of neutrophils and monocytes. The migration of these cells in response to the presence of Leishmania in the peritoneum of affected animals remains unclear. The objective of this study was to investigate cell migration to the peritoneum of BALB/c mice after infection with Leishmania (Leishmania) amazonensis, Leishmania (Viannia) braziliensis and Leishmania (Leishmania) major. Initially, Leishmania ...

  3. Plasticity of Cell Migration In Vivo and In Silico

    NARCIS (Netherlands)

    Boekhorst, V. Te; Preziosi, L.; Friedl, P.

    2016-01-01

    Cell migration results from stepwise mechanical and chemical interactions between cells and their extracellular environment. Mechanistic principles that determine single-cell and collective migration modes and their interconversions depend upon the polarization, adhesion, deformability,

  4. Plasticity of cell migration: a multiscale tuning model.

    NARCIS (Netherlands)

    Friedl, P.H.A.; Wolf, K. van der

    2010-01-01

    Cell migration underlies tissue formation, maintenance, and regeneration as well as pathological conditions such as cancer invasion. Structural and molecular determinants of both tissue environment and cell behavior define whether cells migrate individually (through amoeboid or mesenchymal modes) or

  5. Transcriptome Analysis of Circulating Immune Cell Subsets Highlight the Role of Monocytes in Zaire Ebola Virus Makona Pathogenesis

    Directory of Open Access Journals (Sweden)

    Andrea R. Menicucci

    2017-10-01

    Full Text Available Existing models of Ebola virus disease (EVD suggest antigen-presenting cells are initial targets of Zaire ebolavirus (ZEBOV. In vitro studies have shown that ZEBOV infection of monocytes and macrophages results in the production of inflammatory mediators, which may cause lymphocyte apoptosis. However, these findings have not been corroborated by in vivo studies. In this study, we report the first longitudinal analysis of transcriptional changes in purified monocytes, T-cells, and B-cells isolated from cynomolgus macaques following infection with ZEBOV-Makona. Our data reveal monocytes as one of the major immune cell subsets that supports ZEBOV replication in vivo. In addition, we report a marked increase in the transcription of genes involved in inflammation, coagulation, and vascular disease within monocytes, suggesting that monocytes contribute to EVD manifestations. Further, genes important for antigen presentation and regulation of immunity were downregulated, potentially subverting development of adaptive immunity. In contrast, lymphocytes, which do not support ZEBOV replication, showed transcriptional changes limited to a small number of interferon-stimulated genes (ISGs and a failure to upregulate genes associated with an antiviral effector immune response. Collectively, these data suggest that ZEBOV-infected monocytes play a significant role in ZEBOV-Makona pathogenesis and strategies to suppress virus replication or modify innate responses to infection in these cells should be a priority for therapeutic intervention.

  6. Identification of a Monocyte Receptor on Herpesvirus-Infected Endothelial Cells

    Science.gov (United States)

    Etingin, Orli R.; Silverstein, Roy L.; Hajjar, David P.

    1991-08-01

    The adhesion of circulating blood cells to vascular endothelium may be an initial step in atherosclerosis, inflammation, and wound healing. One mechanism for promoting cell-cell adhesion involves the expression of adhesion molecules on the surface of the target cell. Herpes simplex virus infection of endothelium induces arterial injury and has been implicated in the development of human atherosclerosis. We now demonstrate that HSV-infected endothelial cells express the adhesion molecule GMP140 and that this requires cell surface expression of HSV glycoprotein C and local thrombin generation. Monocyte adhesion to HSV-infected endothelial cells was completely inhibited by anti-GMP140 antibodies but not by antibodies to other adhesion molecules such as VCAM and ELAM-1. The induction of GMP140 expression on HSV-infected endothelium may be an important pathophysiological mechanism in virus-induced cell injury and inflammation.

  7. Interleukin 20 regulates dendritic cell migration and expression of co-stimulatory molecules

    DEFF Research Database (Denmark)

    Bech, Rikke; Jalilian, Babak; Agger, Ralf

    2016-01-01

    BACKGROUND: Psoriasis is an inflammatory disease characterized by leukocyte skin infiltration. Interestingly, recent works suggest that the migration of dendritic cells (DCs) is abnormal in psoriatic skin. DCs have significant role in regulating the function of T lymphocytes, at least in part...... influenced by the local environment of cytokines. In psoriatic skin lesions the expression of IL-20 is highly up-regulated. It is unclear if this cytokine has any influence on DCs. METHODS: Here, we investigated the influence of IL-20 in monocyte-derived dendritic cell (MDDCs) in vitro. This work addressed...

  8. Impact of jamming on collective cell migration

    Science.gov (United States)

    Nnetu, Kenechukwu David; Knorr, Melanie; Pawlizak, Steve; Fuhs, Thomas; Zink, Mareike; KäS, Josef A.

    2012-02-01

    Multi-cellular migration plays an important role in physiological processes such as embryogenesis, cancer metastasis and tissue repair. During migration, single cells undergo cycles of extension, adhesion and retraction resulting in morphological changes. In a confluent monolayer, there are inter-cellular interactions and crowding, however, the impact of these interactions on the dynamics and elasticity of the monolayer at the multi-cellular and single cell level is not well understood. Here we study the dynamics of a confluent epithelial monolayer by simultaneously measuring cell motion at the multi-cellular and single cell level for various cell densities and tensile elasticity. At the multi-cellular level, the system exhibited spatial kinetic transitions from isotropic to anisotropic migration on long times and the velocity of the monolayer decreased with increasing cell density. Moreover, the dynamics was spatially and temporally heterogeneous. Interestingly, the dynamics was also heterogeneous in wound-healing assays and the correlation length was fitted by compressed exponential. On the single cell scale, we observed transient caging effects with increasing cage rearrangement times as the system age due to an increase in density. Also, the density dependent elastic modulus of the monolayer scaled as a weak power law. Together, these findings suggest that caging effects at the single cell level initiates a slow and heterogeneous dynamics at the multi-cellular level which is similar to the glassy dynamics of deformable colloidal systems.

  9. Peripherally administered nanoparticles target monocytic myeloid cells, secondary lymphoid organs and tumors in mice.

    Directory of Open Access Journals (Sweden)

    Iraklis C Kourtis

    Full Text Available Nanoparticles have been extensively developed for therapeutic and diagnostic applications. While the focus of nanoparticle trafficking in vivo has traditionally been on drug delivery and organ-level biodistribution and clearance, recent work in cancer biology and infectious disease suggests that targeting different cells within a given organ can substantially affect the quality of the immunological response. Here, we examine the cell-level biodistribution kinetics after administering ultrasmall Pluronic-stabilized poly(propylene sulfide nanoparticles in the mouse. These nanoparticles depend on lymphatic drainage to reach the lymph nodes and blood, and then enter the spleen rather than the liver, where they interact with monocytes, macrophages and myeloid dendritic cells. They were more readily taken up into lymphatics after intradermal (i.d. compared to intramuscular administration, leading to ∼50% increased bioavailability in blood. When administered i.d., their distribution favored antigen-presenting cells, with especially strong targeting to myeloid cells. In tumor-bearing mice, the monocytic and the polymorphonuclear myeloid-derived suppressor cell compartments were efficiently and preferentially targeted, rendering this nanoparticulate formulation potentially useful for reversing the highly suppressive activity of these cells in the tumor stroma.

  10. Peripherally administered nanoparticles target monocytic myeloid cells, secondary lymphoid organs and tumors in mice.

    Science.gov (United States)

    Kourtis, Iraklis C; Hirosue, Sachiko; de Titta, Alexandre; Kontos, Stephan; Stegmann, Toon; Hubbell, Jeffrey A; Swartz, Melody A

    2013-01-01

    Nanoparticles have been extensively developed for therapeutic and diagnostic applications. While the focus of nanoparticle trafficking in vivo has traditionally been on drug delivery and organ-level biodistribution and clearance, recent work in cancer biology and infectious disease suggests that targeting different cells within a given organ can substantially affect the quality of the immunological response. Here, we examine the cell-level biodistribution kinetics after administering ultrasmall Pluronic-stabilized poly(propylene sulfide) nanoparticles in the mouse. These nanoparticles depend on lymphatic drainage to reach the lymph nodes and blood, and then enter the spleen rather than the liver, where they interact with monocytes, macrophages and myeloid dendritic cells. They were more readily taken up into lymphatics after intradermal (i.d.) compared to intramuscular administration, leading to ∼50% increased bioavailability in blood. When administered i.d., their distribution favored antigen-presenting cells, with especially strong targeting to myeloid cells. In tumor-bearing mice, the monocytic and the polymorphonuclear myeloid-derived suppressor cell compartments were efficiently and preferentially targeted, rendering this nanoparticulate formulation potentially useful for reversing the highly suppressive activity of these cells in the tumor stroma.

  11. CML/CD36 accelerates atherosclerotic progression via inhibiting foam cell migration.

    Science.gov (United States)

    Xu, Suining; Li, Lihua; Yan, Jinchuan; Ye, Fei; Shao, Chen; Sun, Zhen; Bao, Zhengyang; Dai, Zhiyin; Zhu, Jie; Jing, Lele; Wang, Zhongqun

    2018-01-01

    Among the various complications of type 2 diabetes mellitus, atherosclerosis causes the highest disability and morbidity. A multitude of macrophage-derived foam cells are retained in atherosclerotic plaques resulting not only from recruitment of monocytes into lesions but also from a reduced rate of macrophage migration from lesions. Nε-carboxymethyl-Lysine (CML), an advanced glycation end product, is responsible for most complications of diabetes. This study was designed to investigate the mechanism of CML/CD36 accelerating atherosclerotic progression via inhibiting foam cell migration. In vivo study and in vitro study were performed. For the in vivo investigation, CML/CD36 accelerated atherosclerotic progression via promoting the accumulation of macrophage-derived foam cells in aorta and inhibited macrophage-derived foam cells in aorta migrating to the para-aorta lymph node of diabetic apoE -/- mice. For the in vitro investigation, CML/CD36 inhibited RAW264.7-derived foam cell migration through NOX-derived ROS, FAK phosphorylation, Arp2/3 complex activation and F-actin polymerization. Thus, we concluded that CML/CD36 inhibited foam cells of plaque migrating to para-aorta lymph nodes, accelerating atherosclerotic progression. The corresponding mechanism may be via free cholesterol, ROS generation, p-FAK, Arp2/3, F-actin polymerization. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  12. Synthesis of complement factor B by the human monocyte U-937 cell line. Augmentation by immunostimulatory agents

    International Nuclear Information System (INIS)

    Minta, J.O.

    1988-01-01

    The human pre-monocytic cell line U-937 was shown to synthesize and to secrete increasing amounts of factor B in short term cultures in serum-free medium containing BSA. The kinetics of factor B production were higher on day 2 than on days 1 and 3. The production of factor B was reversibly inhibited by cycloheximide, indicating de novo synthesis. Metabolic labeling with [ 35 S]-methionine and SDS-PAGE analysis revealed that both intracellular and secreted factor B were single-chain proteins with similar m.w. (90,000), which co-migrated with purified plasma factor B. Incubation of U-937 cells with the immunostimulants PMA, LPS, IFN-gamma, and IL-1 resulted in a dose-dependent augmentation of factor B production. A 24-h exposure to IL-1 was shown to be required for maximal stimulation. A combination of suboptimal doses of LPS and IFN-gamma was shown to exert a synergistic effect on factor B production. The U-937 cell line is thus a valuable model for the study of the regulation of the factor B gene expression

  13. Migration of acute lymphoblastic leukemia cells into human bone marrow stroma.

    Science.gov (United States)

    Makrynikola, V; Bianchi, A; Bradstock, K; Gottlieb, D; Hewson, J

    1994-10-01

    Most cases of acute lymphoblastic leukemia (ALL) arise from malignant transformation of B-cell precursors in the bone marrow. Recent studies have shown that normal and leukemic B-cell precursors bind to bone marrow stromal cells through the beta-1 integrins VLA-4 and VLA-5, thereby exposing early lymphoid cells to regulatory cytokines. It has been recently reported that the pre-B cell line NALM-6 is capable of migrating under layers of murine stromal cells in vitro (Miyake et al. J Cell Biol 1992;119:653-662). We have further analyzed leukemic cell motility using human bone marrow fibroblasts (BMF) as a stromal layer. The precursor-B ALL cell line NALM-6 rapidly adhered to BMF, and underwent migration or tunneling into BMF layers within 5 h, as demonstrated by light and electron microscopy, and confirmed by a chromium-labeling assay. Migration was also observed with the precursor-B ALL lines Reh and KM-3, with a T leukemia line RPMI-8402, the monocytic line U937, and the mature B line Daudi. In contrast, mature B (Raji), myeloid (K562, HL-60), and T lines (CCRF-CEM, MOLT-4) did not migrate. When cases of leukemia were analyzed, BMF migration was largely confined to precursor-B ALL, occurring in eight of 13 cases tested. Of other types of leukemia, migration was observed in one of four cases of T-ALL, but no evidence was seen in six acute myeloid leukemias and two patients with chronic lymphocytic leukemia. Only minimal migration into BMF was observed with purified sorted CD10+ CD19+ early B cells from normal adult marrow, while normal mature B lymphocytes from peripheral blood did not migrate. ALL migration was inhibited by monoclonal antibodies to the beta sub-unit of the VLA integrin family, and by a combination of antibodies to VLA-4 and VLA-5. Partial inhibition was also observed when leukemic cells were incubated with antibodies to VLA-4, VLA-5, or VLA-6 alone. In contrast, treatment of stromal cells with antibodies to vascular cell adhesion molecule or

  14. Prognostic significance of peripheral monocyte count in patients with extranodal natural killer/T-cell lymphoma

    International Nuclear Information System (INIS)

    Huang, Jia-Jia; Li, Zhi-Ming; Li, Ya-Jun; Xia, Yi; Wang, Yu; Wei, Wen-Xiao; Zhu, Ying-Jie; Lin, Tong-Yu; Huang, Hui-Qiang; Jiang, Wen-Qi

    2013-01-01

    Extranodal natural killer/T-cell lymphoma (ENKL) has heterogeneous clinical manifestations and prognosis. This study aims to evaluate the prognostic impact of absolute monocyte count (AMC) in ENKL, and provide some immunologically relevant information for better risk stratification in patients with ENKL. Retrospective data from 163 patients newly diagnosed with ENKL were analyzed. The absolute monocyte count (AMC) at diagnosis was analyzed as continuous and dichotomized variables. Independent prognostic factors of survival were determined by Cox regression analysis. The AMC at diagnosis were related to overall survival (OS) and progression-free survival (PFS) in patients with ENKL. Multivariate analysis identified AMC as independent prognostic factors of survival, independent of International Prognostic Index (IPI) and Korean prognostic index (KPI). The prognostic index incorporating AMC and absolute lymphocyte count (ALC), another surrogate factor of immune status, could be used to stratify all 163 patients with ENKL into different prognostic groups. For patients who received chemotherapy followed by radiotherapy (102 cases), the three AMC/ALC index categories identified patients with significantly different survivals. When superimposed on IPI or KPI categories, the AMC/ALC index was better able to identify high-risk patients in the low-risk IPI or KPI category. The baseline peripheral monocyte count is shown to be an effective prognostic indicator of survival in ENKL patients. The prognostic index related to tumor microenvironment might be helpful to identify high-risk patients with ENKL

  15. Prognostic significance of peripheral monocyte count in patients with extranodal natural killer/T-cell lymphoma.

    Science.gov (United States)

    Huang, Jia-Jia; Li, Ya-Jun; Xia, Yi; Wang, Yu; Wei, Wen-Xiao; Zhu, Ying-Jie; Lin, Tong-Yu; Huang, Hui-Qiang; Jiang, Wen-Qi; Li, Zhi-Ming

    2013-05-03

    Extranodal natural killer/T-cell lymphoma (ENKL) has heterogeneous clinical manifestations and prognosis. This study aims to evaluate the prognostic impact of absolute monocyte count (AMC) in ENKL, and provide some immunologically relevant information for better risk stratification in patients with ENKL. Retrospective data from 163 patients newly diagnosed with ENKL were analyzed. The absolute monocyte count (AMC) at diagnosis was analyzed as continuous and dichotomized variables. Independent prognostic factors of survival were determined by Cox regression analysis. The AMC at diagnosis were related to overall survival (OS) and progression-free survival (PFS) in patients with ENKL. Multivariate analysis identified AMC as independent prognostic factors of survival, independent of International Prognostic Index (IPI) and Korean prognostic index (KPI). The prognostic index incorporating AMC and absolute lymphocyte count (ALC), another surrogate factor of immune status, could be used to stratify all 163 patients with ENKL into different prognostic groups. For patients who received chemotherapy followed by radiotherapy (102 cases), the three AMC/ALC index categories identified patients with significantly different survivals. When superimposed on IPI or KPI categories, the AMC/ALC index was better able to identify high-risk patients in the low-risk IPI or KPI category. The baseline peripheral monocyte count is shown to be an effective prognostic indicator of survival in ENKL patients. The prognostic index related to tumor microenvironment might be helpful to identify high-risk patients with ENKL.

  16. A human coronavirus responsible for the common cold massively kills dendritic cells but not monocytes.

    Science.gov (United States)

    Mesel-Lemoine, Mariana; Millet, Jean; Vidalain, Pierre-Olivier; Law, Helen; Vabret, Astrid; Lorin, Valérie; Escriou, Nicolas; Albert, Matthew L; Nal, Béatrice; Tangy, Frédéric

    2012-07-01

    Human coronaviruses are associated with upper respiratory tract infections that occasionally spread to the lungs and other organs. Although airway epithelial cells represent an important target for infection, the respiratory epithelium is also composed of an elaborate network of dendritic cells (DCs) that are essential sentinels of the immune system, sensing pathogens and presenting foreign antigens to T lymphocytes. In this report, we show that in vitro infection by human coronavirus 229E (HCoV-229E) induces massive cytopathic effects in DCs, including the formation of large syncytia and cell death within only few hours. In contrast, monocytes are much more resistant to infection and cytopathic effects despite similar expression levels of CD13, the membrane receptor for HCoV-229E. While the differentiation of monocytes into DCs in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4 requires 5 days, only 24 h are sufficient for these cytokines to sensitize monocytes to cell death and cytopathic effects when infected by HCoV-229E. Cell death induced by HCoV-229E is independent of TRAIL, FasL, tumor necrosis factor alpha, and caspase activity, indicating that viral replication is directly responsible for the observed cytopathic effects. The consequence of DC death at the early stage of HCoV-229E infection may have an impact on the early control of viral dissemination and on the establishment of long-lasting immune memory, since people can be reinfected multiple times by HCoV-229E.

  17. Modeling collective cell migration in geometric confinement

    Science.gov (United States)

    Tarle, Victoria; Gauquelin, Estelle; Vedula, S. R. K.; D'Alessandro, Joseph; Lim, C. T.; Ladoux, Benoit; Gov, Nir S.

    2017-06-01

    Monolayer expansion has generated great interest as a model system to study collective cell migration. During such an expansion the culture front often develops ‘fingers’, which we have recently modeled using a proposed feedback between the curvature of the monolayer’s leading edge and the outward motility of the edge cells. We show that this model is able to explain the puzzling observed increase of collective cellular migration speed of a monolayer expanding into thin stripes, as well as describe the behavior within different confining geometries that were recently observed in experiments. These comparisons give support to the model and emphasize the role played by the edge cells and the edge shape during collective cell motion.

  18. Stem cell migration - Methods and protocols

    Directory of Open Access Journals (Sweden)

    Carlo Alberto Redi

    2012-03-01

    Full Text Available The trafficking of stem cells is something unconsciously clear to any biologists (e.g., developmental biologists and physicians (e.g., all those taking care of hematopoietic and bone diseases and traumas; neverthless it is a phenomenon coming out as a hot topic just in these last years. Likely, the difficulties to track stem cells migration in vivo and the understanding of the elusive homing signals matching the circulating stem cells properties that makes these cells to stop and to start multiplication and differentiation....

  19. Migration of cells in a social context

    DEFF Research Database (Denmark)

    Vedel, Søren; Tay, Savas; Johnston, Darius M

    2013-01-01

    In multicellular organisms and complex ecosystems, cells migrate in a social context. Whereas this is essential for the basic processes of life, the influence of neighboring cells on the individual remains poorly understood. Previous work on isolated cells has observed a stereotypical migratory...... behavior characterized by short-time directional persistence with long-time random movement. We discovered a much richer dynamic in the social context, with significant variations in directionality, displacement, and speed, which are all modulated by local cell density. We developed a mathematical model...

  20. Monocyte-derived dendritic cells are essential for CD8+ T cell activation and anti-tumor responses after local immunotherapy

    Directory of Open Access Journals (Sweden)

    Sabine eKuhn

    2015-11-01

    Full Text Available Tumors harbor several populations of dendritic cells with the ability to prime tumor-specific T cells. However, these T cells mostly fail to differentiate into armed effectors and are unable to control tumor growth. We have previously shown that treatment with immunostimulatory agents at the tumor site can activate anti-tumor immune responses, and is associated with the appearance of a population of monocyte-derived dendritic cells in the tumor and tumor-draining lymph node. Here we use dendritic cell or monocyte depletion and monocyte transfer to show that these monocyte-derived dendritic cells are critical to the activation of anti-tumor immune responses. Treatment with the immunostimulatory agents Monosodium Urate crystals and Mycobacterium smegmatis induced the accumulation of monocytes in the draining lymph node, their upregulation of CD11c and MHCII, and expression of iNOS, TNFα and IL12p40. Blocking monocyte entry into the lymph node and tumor through neutralization of the chemokine CCL2 or inhibition of Colony Stimulating Factor-1 receptor signaling prevented the generation of monocyte-derived dendritic cells, the infiltration of tumor-specific T cells into the tumor, and anti-tumor responses. In a reciprocal fashion, monocytes transferred into mice depleted of CD11c+ cells were sufficient to rescue CD8+ T cell priming in lymph node and delay tumor growth. Thus monocytes exposed to the appropriate conditions become powerful activators of tumor-specific CD8+ T cells and anti-tumor immunity.

  1. Cell migration during heart regeneration in zebrafish.

    Science.gov (United States)

    Tahara, Naoyuki; Brush, Michael; Kawakami, Yasuhiko

    2016-07-01

    Zebrafish possess the remarkable ability to regenerate injured hearts as adults, which contrasts the very limited ability in mammals. Although very limited, mammalian hearts do in fact have measurable levels of cardiomyocyte regeneration. Therefore, elucidating mechanisms of zebrafish heart regeneration would provide information of naturally occurring regeneration to potentially apply to mammalian studies, in addition to addressing this biologically interesting phenomenon in itself. Studies over the past 13 years have identified processes and mechanisms of heart regeneration in zebrafish. After heart injury, pre-existing cardiomyocytes dedifferentiate, enter the cell cycle, and repair the injured myocardium. This process requires interaction with epicardial cells, endocardial cells, and vascular endothelial cells. Epicardial cells envelope the heart, while endocardial cells make up the inner lining of the heart. They provide paracrine signals to cardiomyocytes to regenerate the injured myocardium, which is vascularized during heart regeneration. In addition, accumulating results suggest that local migration of these major cardiac cell types have roles in heart regeneration. In this review, we summarize the characteristics of various heart injury methods used in the research community and regeneration of the major cardiac cell types. Then, we discuss local migration of these cardiac cell types and immune cells during heart regeneration. Developmental Dynamics 245:774-787, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  2. Primary Cilia, Signaling Networks and Cell Migration

    DEFF Research Database (Denmark)

    Veland, Iben Rønn

    Primary cilia are microtubule-based, sensory organelles that emerge from the centrosomal mother centriole to project from the surface of most quiescent cells in the human body. Ciliary entry is a tightly controlled process, involving diffusion barriers and gating complexes that maintain a unique...... this controls directional cell migration as a physiological response. The ciliary pocket is a membrane invagination with elevated activity of clathrin-dependent endocytosis (CDE). In paper I, we show that the primary cilium regulates TGF-β signaling and the ciliary pocket is a compartment for CDE...... on formation of the primary cilium and CDE at the pocket region. The ciliary protein Inversin functions as a molecular switch between canonical and non-canonical Wnt signaling. In paper II, we show that Inversin and the primary cilium control Wnt signaling and are required for polarization and cell migration...

  3. Paxillin: a crossroad in pathological cell migration

    Directory of Open Access Journals (Sweden)

    Ana María López-Colomé

    2017-02-01

    Full Text Available Abstract Paxilllin is a multifunctional and multidomain focal adhesion adapter protein which serves an important scaffolding role at focal adhesions by recruiting structural and signaling molecules involved in cell movement and migration, when phosphorylated on specific Tyr and Ser residues. Upon integrin engagement with extracellular matrix, paxillin is phosphorylated at Tyr31, Tyr118, Ser188, and Ser190, activating numerous signaling cascades which promote cell migration, indicating that the regulation of adhesion dynamics is under the control of a complex display of signaling mechanisms. Among them, paxillin disassembly from focal adhesions induced by extracellular regulated kinase (ERK-mediated phosphorylation of serines 106, 231, and 290 as well as the binding of the phosphatase PEST to paxillin have been shown to play a key role in cell migration. Paxillin also coordinates the spatiotemporal activation of signaling molecules, including Cdc42, Rac1, and RhoA GTPases, by recruiting GEFs, GAPs, and GITs to focal adhesions. As a major participant in the regulation of cell movement, paxillin plays distinct roles in specific tissues and developmental stages and is involved in immune response, epithelial morphogenesis, and embryonic development. Importantly, paxillin is also an essential player in pathological conditions including oxidative stress, inflammation, endothelial cell barrier dysfunction, and cancer development and metastasis.

  4. Human monocytes undergo excessive apoptosis following temozolomide activating the ATM/ATR pathway while dendritic cells and macrophages are resistant.

    Directory of Open Access Journals (Sweden)

    Martina Bauer

    Full Text Available Immunodeficiency is a severe therapy-limiting side effect of anticancer chemotherapy resulting from sensitivity of immunocompetent cells to DNA damaging agents. A central role in the immune system is played by monocytes that differentiate into macrophages and dendritic cells (DCs. In this study we compared human monocytes isolated from peripheral blood and cytokine matured macrophages and DCs derived from them and assessed the mechanism of toxicity of the DNA methylating anticancer drug temozolomide (TMZ in these cell populations. We observed that monocytes, but not DCs and macrophages, were highly sensitive to the killing effect of TMZ. Studies on DNA damage and repair revealed that the initial DNA incision was efficient in monocytes while the re-ligation step of base excision repair (BER can not be accomplished, resulting in an accumulation of DNA single-strand breaks (SSBs. Furthermore, monocytes accumulated DNA double-strand breaks (DSBs following TMZ treatment, while DCs and macrophages were able to repair DSBs. Monocytes lack the DNA repair proteins XRCC1, ligase IIIα and PARP-1 whose expression is restored during differentiation into macrophages and DCs following treatment with GM-CSF and GM-CSF plus IL-4, respectively. These proteins play a key role both in BER and DSB repair by B-NHEJ, which explains the accumulation of DNA breaks in monocytes following TMZ treatment. Although TMZ provoked an upregulation of XRCC1 and ligase IIIα, BER was not enhanced likely because PARP-1 was not upregulated. Accordingly, inhibition of PARP-1 did not sensitize monocytes, but monocyte-derived DCs in which strong PARP activation was observed. TMZ induced in monocytes the DNA damage response pathways ATM-Chk2 and ATR-Chk1 resulting in p53 activation. Finally, upon activation of the Fas-receptor and the mitochondrial pathway apoptosis was executed in a caspase-dependent manner. The downregulation of DNA repair in monocytes, resulting in their selective

  5. Drug induced increases in CNS dopamine alter monocyte, macrophage and T cell functions: implications for HAND

    Science.gov (United States)

    Gaskill, Peter J.; Calderon, Tina M.; Coley, Jacqueline S.; Berman, Joan W.

    2013-01-01

    Central nervous system (CNS) complications resulting from HIV infection remain a major public health problem as individuals live longer due to the success of combined antiretroviral therapy (cART). As many as 70% of HIV infected people have HIV associated neurocognitive disorders (HAND). Many HIV infected individuals abuse drugs, such as cocaine, heroin or methamphetamine, that may be important cofactors in the development of HIV CNS disease. Despite different mechanisms of action, all drugs of abuse increase extracellular dopamine in the CNS. The effects of dopamine on HIV neuropathogenesis are not well understood, and drug induced increases in CNS dopamine may be a common mechanism by which different types of drugs of abuse impact the development of HAND. Monocytes and macrophages are central to HIV infection of the CNS and to HAND. While T cells have not been shown to be a major factor in HIV-associated neuropathogenesis, studies indicate that T cells may play a larger role in the development of HAND in HIV infected drug abusers. Drug induced increases in CNS dopamine may dysregulate functions of, or increase HIV infection in, monocytes, macrophages and T cells in the brain. Thus, characterizing the effects of dopamine on these cells is important for understanding the mechanisms that mediate the development of HAND in drug abusers. PMID:23456305

  6. Interstitial cell migration: integrin-dependent and alternative adhesion mechanisms.

    NARCIS (Netherlands)

    Schmidt, S.; Friedl, P.H.A.

    2010-01-01

    Adhesion and migration are integrated cell functions that build, maintain and remodel the multicellular organism. In migrating cells, integrins are the main transmembrane receptors that provide dynamic interactions between extracellular ligands and actin cytoskeleton and signalling machineries. In

  7. Abnormalities in iNKT cells are associated with impaired ability of monocytes to produce IL-10 and suppress T-cell proliferation in sarcoidosis.

    Science.gov (United States)

    Crawshaw, Anjali; Kendrick, Yvonne R; McMichael, Andrew J; Ho, Ling-Pei

    2014-07-01

    Sarcoidosis is a multisystem granulomatous disorder characterized by marked T-cell expansion of T helper 1 (Th1) cells. The cause of T-cell overactivity is unknown. We hypothesized that interleukin-10 (IL-10) production by a yet undefined cell type might be defective, resulting in loss of regulation of T-cell activity. Focusing on IL-10-producing monocytes, we first showed that monocytes isolated from the peripheral blood of corticosteroid-naïve sarcoidosis patients (n = 51) produced less IL-10 compared to controls, and were less able to suppress T-cell proliferation. In addition, monocytic IL-10 production correlated negatively with disease activity score. As invariant natural killer T (iNKT) cells are known to both interact with monocytes and be reduced in sarcoidosis patients, we then asked whether iNKT-specific defects might be responsible for this reduced IL-10 production. We found that greater numbers of circulating iNKT cells was associated with higher IL-10 production. Moreover, iNKT cells enhanced monocytic IL-10 production in vitro. Defective IL-10 production and T-cell suppression by sarcoidosis monocytes could be restored following their coculture with iNKT cells, in a CD1d- and cell contact-dependent process. We suggest that reduced iNKT-cell numbers in sarcoidosis may lead to impaired monocytic IL-10 production and unchecked T-cell expansion in sarcoidosis. These findings provide fresh insight into the mechanism of sarcoidosis disease, and interaction between iNKT cells and monocytes. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Platelets Inhibit Migration of Canine Osteosarcoma Cells.

    Science.gov (United States)

    Bulla, S C; Badial, P R; Silva, R C; Lunsford, K; Bulla, C

    2017-01-01

    The interaction between platelets and tumour cells is important for tumour growth and metastasis. Thrombocytopenia or antiplatelet treatment negatively impact on cancer metastasis, demonstrating potentially important roles for platelets in tumour progression. To our knowledge, there is no information regarding the role of platelets in cancer progression in dogs. This study was designed to test whether canine platelets affected the migratory behaviour of three canine osteosarcoma cell lines and to give insights of molecular mechanisms. Intact platelets, platelet lysate and platelet releasate inhibited the migration of canine osteosarcoma cell lines. Addition of blood leucocytes to the platelet samples did not alter the inhibitory effect on migration. Platelet treatment also significantly downregulated the transcriptional levels of SNAI2 and TWIST1 genes. The interaction between canine platelets or molecules released during platelet activation and these tumour cell lines inhibits their migration, which suggests that canine platelets might antagonize metastasis of canine osteosarcoma. This effect is probably due to, at least in part, downregulation of genes related to epithelial-mesenchymal transition. Copyright © 2016. Published by Elsevier Ltd.

  9. Substrate Curvature Regulates Cell Migration -A Computational Study

    Science.gov (United States)

    He, Xiuxiu; Jiang, Yi

    Cell migration in host microenvironment is essential to cancer etiology, progression and metastasis. Cellular processes of adhesion, cytoskeletal polymerization, contraction, and matrix remodeling act in concert to regulate cell migration, while local extracellular matrix architecture modulate these processes. In this work we study how stromal microenvironment with native and cell-derived curvature at micron-meter scale regulate cell motility pattern. We developed a 3D model of single cell migration on a curved substrate. Mathematical analysis of cell morphological adaption to the cell-substrate interface shows that cell migration on convex surfaces deforms more than on concave surfaces. Both analytical and simulation results show that curved surfaces regulate the cell motile force for cell's protruding front through force balance with focal adhesion and cell contraction. We also found that cell migration on concave substrates is more persistent. These results offer a novel biomechanical explanation to substrate curvature regulation of cell migration. NIH 1U01CA143069.

  10. EFFECTS OF SECRETABLE PLACENTAL FACTORS UPON SECRETION OF CYTOKINES BY THP-1 MONOCYTE-LIKE CELLS

    Directory of Open Access Journals (Sweden)

    Ya. S. Onokhina

    2013-01-01

    Full Text Available Abstract. Мonocytes in feto-placental circulation are exposed to factors secreted by placental tissue. These factors influence monocyte functions in pregnancy. In present study, an in vitro model (monocyte-like THP-1 cells was used for assessing effects of soluble placental factors obtained from women with physiological pregnancies, or preeclampsia cases. The following effects of placental factors were revealed: increased secretion of VEGF by THP-1 cells along with decreased secretion of IL-6, IL-8 and MCP-1 under the influence of placental factors from the I. trimester of pregnancy in comparison with III. trimester. Secretion of IL-6 and MCP-1 by THP-1 cells was increased, and secretion of soluble TNFRII was decreased upon co-cultivation with soluble placental factors from the women with preeclampsia, as compared with placental products from physiological pregnancies.The work is supported by grants ГК № 02.740.11.0711 from Ministry of Education and Science, and НШ-3594.2010.7 grant from the President of Russian Federation.

  11. Stimulation of the Angiotensin II AT2 Receptor is Anti-inflammatory in Human Lipopolysaccharide-Activated Monocytic Cells

    DEFF Research Database (Denmark)

    Menk, Mario; Graw, Jan Adriaan; von Haefen, Clarissa

    2015-01-01

    and the translational level over course of time. Treatment with C21 attenuated the expression of TNFα, IL-6, and IL-10 after LPS challenge in both cell lines in a time- and dose-dependent manner. We conclude that selective AT2 receptor stimulation acts anti-inflammatory in human monocytes. Modulation of cytokine......Recently, AT2 receptors have been discovered on the surface of human immunocompetent cells such as monocytes. Data on regulative properties of this receptor on the cellular immune response are poor. We hypothesized that direct stimulation of the AT2 receptor mediates anti-inflammatory responses...... in these cells. Human monocytic THP-1 and U937 cells were stimulated with lipopolysaccharide (LPS) and the selective AT2 receptor agonist Compound 21 (C21). Expression of pro- and anti-inflammatory cytokines IL-6, IL-10, tumor necrosis factor-α (TNFα), and IL-1β were analyzed on both the transcriptional...

  12. Role of ATM in bystander signaling between human monocytes and lung adenocarcinoma cells.

    Science.gov (United States)

    Ghosh, Somnath; Ghosh, Anu; Krishna, Malini

    2015-12-01

    The response of a cell or tissue to ionizing radiation is mediated by direct damage to cellular components and indirect damage mediated by radiolysis of water. Radiation affects both irradiated cells and the surrounding cells and tissues. The radiation-induced bystander effect is defined by the presence of biological effects in cells that were not themselves in the field of irradiation. To establish the contribution of the bystander effect in the survival of the neighboring cells, lung carcinoma A549 cells were exposed to gamma-irradiation, 2Gy. The medium from the irradiated cells was transferred to non-irradiated A549 cells. Irradiated A549 cells as well as non-irradiated A549 cells cultured in the presence of medium from irradiated cells showed decrease in survival and increase in γ-H2AX and p-ATM foci, indicating a bystander effect. Bystander signaling was also observed between different cell types. Phorbol-12-myristate-13-acetate (PMA)-stimulated and gamma-irradiated U937 (human monocyte) cells induced a bystander response in non-irradiated A549 (lung carcinoma) cells as shown by decreased survival and increased γ-H2AX and p-ATM foci. Non-stimulated and/or irradiated U937 cells did not induce such effects in non-irradiated A549 cells. Since ATM protein was activated in irradiated cells as well as bystander cells, it was of interest to understand its role in bystander effect. Suppression of ATM with siRNA in A549 cells completely inhibited bystander effect in bystander A549 cells. On the other hand suppression of ATM with siRNA in PMA stimulated U937 cells caused only a partial inhibition of bystander effect in bystander A549 cells. These results indicate that apart from ATM, some additional factor may be involved in bystander effect between different cell types. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Lung cells support osteosarcoma cell migration and survival.

    Science.gov (United States)

    Yu, Shibing; Fourman, Mitchell Stephen; Mahjoub, Adel; Mandell, Jonathan Brendan; Crasto, Jared Anthony; Greco, Nicholas Giuseppe; Weiss, Kurt Richard

    2017-01-25

    Osteosarcoma (OS) is the most common primary bone tumor, with a propensity to metastasize to the lungs. Five-year survival for metastatic OS is below 30%, and has not improved for several decades despite the introduction of multi-agent chemotherapy. Understanding OS cell migration to the lungs requires an evaluation of the lung microenvironment. Here we utilized an in vitro lung cell and OS cell co-culture model to explore the interactions between OS and lung cells, hypothesizing that lung cells would promote OS cell migration and survival. The impact of a novel anti-OS chemotherapy on OS migration and survival in the lung microenvironment was also examined. Three human OS cell lines (SJSA-1, Saos-2, U-2) and two human lung cell lines (HULEC-5a, MRC-5) were cultured according to American Type Culture Collection recommendations. Human lung cell lines were cultured in growth medium for 72 h to create conditioned media. OS proliferation was evaluated in lung co-culture and conditioned media microenvironment, with a murine fibroblast cell line (NIH-3 T3) in fresh growth medium as controls. Migration and invasion were measured using a real-time cell analysis system. Real-time PCR was utilized to probe for Aldehyde Dehydrogenase (ALDH1) expression. Osteosarcoma cells were also transduced with a lentivirus encoding for GFP to permit morphologic analysis with fluorescence microscopy. The anti-OS efficacy of Disulfiram, an ALDH-inhibitor previously shown to inhibit OS cell proliferation and metastasis in vitro, was evaluated in each microenvironment. Lung-cell conditioned medium promoted osteosarcoma cell migration, with a significantly higher attractive effect on all three osteosarcoma cell lines compared to basic growth medium, 10% serum containing medium, and NIH-3 T3 conditioned medium (p cell conditioned medium induced cell morphologic changes, as demonstrated with GFP-labeled cells. OS cells cultured in lung cell conditioned medium had increased alkaline

  14. Migration of cochlear lateral wall cells.

    Science.gov (United States)

    Dunaway, George; Mhaskar, Yashanad; Armour, Gary; Whitworth, Craig; Rybak, Leonard

    2003-03-01

    The role of apoptosis and proliferation in maintenance of cochlear lateral wall cells was examined. The methods employed for detection of apoptosis were the Hoechst fluorescence stain and TUNEL (TdT-mediated dUTP-biotin nick-end-labeling) assay, and proliferations were 5-bromo-2'-deoxyuridine (BrdU) incorporation and presence of the proliferating cell nuclear antigen. The incidence of apoptosis in the strial marginal cell was 50% greater (32.9+/-3.7%) than strial intermediate and basal cells but similar to spiral ligament cells. Although division of marginal strial cells was rarely detected, a significant number of proliferating cells in the remaining stria vascularis and spiral ligament were observed. These data implied that replacement of marginal cells arose elsewhere and could be followed by a BrdU-deoxythymidine pulse-chase study. At 2 h post injection, nuclear BrdU in marginal cells was not detected; however, by 24 h post injection, 20-25% of marginal cell nuclei were BrdU-positive. These observations are consistent with the hypothesis that marginal cells were replaced by underlying cells. Cell migration appears to be an important mechanism for preserving the function and structure of the stria vascularis.

  15. Collective cell migration: Implications for wound healing and cancer invasion

    Directory of Open Access Journals (Sweden)

    Li Li

    2013-07-01

    Full Text Available During embryonic morphogenesis, wound repair and cancer invasion, cells often migrate collectively via tight cell-cell junctions, a process named collective migration. During such migration, cells move as coherent groups, large cell sheets, strands or tubes rather than individually. One unexpected finding regarding collective cell migration is that being a "multicellular structure" enables cells to better respond to chemical and physical cues, when compared with isolated cells. This is important because epithelial cells heal wounds via the migration of large sheets of cells with tight intercellular connections. Recent studies have gained some mechanistic insights that will benefit the clinical understanding of wound healing in general. In this review, we will briefly introduce the role of collective cell migration in wound healing, regeneration and cancer invasion and discuss its underlying mechanisms as well as implications for wound healing.

  16. iNKT Cell Emigration out of the Lung Vasculature Requires Neutrophils and Monocyte-Derived Dendritic Cells in Inflammation

    Directory of Open Access Journals (Sweden)

    Ajitha Thanabalasuriar

    2016-09-01

    Full Text Available iNKT cells are a subset of innate T cells that recognize glycolipids presented on CD1d molecules and protect against bacterial infections, including S. pneumoniae. Using lung intravital imaging, we examined the behavior and mechanism of pulmonary iNKT cell activation in response to the specific iNKT cell ligand α-galactosylceramide or S. pneumoniae infection. In untreated mice, the major fraction of iNKT cells resided in the vasculature, but a small critical population resided in the extravascular space in proximity to monocyte-derived DCs. Administration of either α-GalCer or S. pneumoniae induced CD1d-dependent rapid recruitment of neutrophils out of the vasculature. The neutrophils guided iNKT cells from the lung vasculature via CCL17. Depletion of monocyte-derived DCs abrogated both the neutrophil and subsequent iNKT cell extravasation. Moreover, impairing iNKT cell recruitment by blocking CCL17 increased susceptibility to S. pneumoniae infection, suggesting a critical role for the influx of iNKT cells in host defense.

  17. Captopril increases the intensity of monocyte infection by Trypanosoma cruzi and induces human T helper type 17 cells.

    Science.gov (United States)

    Coelho dos Santos, J S; Menezes, C A S; Villani, F N A; Magalhães, L M D; Scharfstein, J; Gollob, K J; Dutra, W O

    2010-12-01

    The anti-hypertensive drug captopril is used commonly to reduce blood pressure of patients with severe forms of Chagas disease, a cardiomyopathy caused by chronic infection with the intracellular protozoan Trypanosoma cruzi. Captopril acts by inhibiting angiotensin-converting enzyme (ACE), the vasopressor metallopeptidase that generates angiotensin II and promotes the degradation of bradykinin (BK). Recent studies in mice models of Chagas disease indicated that captopril can potentiate the T helper type 1 (Th1)-directing natural adjuvant property of BK. Equipped with kinin-releasing cysteine proteases, T. cruzi trypomastigotes were shown previously to invade non-professional phagocytic cells, such as human endothelial cells and murine cardiomyocytes, through the signalling of G protein-coupled bradykinin receptors (B(2) KR). Monocytes are also parasitized by T. cruzi and these cells are known to be important for the host immune response during infection. Here we showed that captopril increases the intensity of T. cruzi infection of human monocytes in vitro. The increased parasitism was accompanied by up-regulated expression of ACE in human monocytes. While T. cruzi infection increased the expression of interleukin (IL)-10 by monocytes significantly, compared to uninfected cells, T. cruzi infection in association with captopril down-modulated IL-10 expression by the monocytes. Surprisingly, studies with peripheral blood mononuclear cells revealed that addition of the ACE inhibitor in association with T. cruzi increased expression of IL-17 by CD4(+) T cells in a B(2) KR-dependent manner. Collectively, our results suggest that captopril might interfere with host-parasite equilibrium by enhancing infection of monocytes, decreasing the expression of the modulatory cytokine IL-10, while guiding development of the proinflammatory Th17 subset. © 2010 The Authors. Clinical and Experimental Immunology © 2010 British Society for Immunology.

  18. Cell shape dynamics: from waves to migration.

    Directory of Open Access Journals (Sweden)

    Meghan K Driscoll

    Full Text Available We observe and quantify wave-like characteristics of amoeboid migration. Using the amoeba Dictyostelium discoideum, a model system for the study of chemotaxis, we demonstrate that cell shape changes in a wave-like manner. Cells have regions of high boundary curvature that propagate from the leading edge toward the back, usually along alternating sides of the cell. Curvature waves are easily seen in cells that do not adhere to a surface, such as cells that are electrostatically repelled from surfaces or cells that extend over the edge of micro-fabricated cliffs. Without surface contact, curvature waves travel from the leading edge to the back of a cell at -35 µm/min. Non-adherent myosin II null cells do not exhibit these curvature waves. At the leading edge of adherent cells, curvature waves are associated with protrusive activity. Like regions of high curvature, protrusive activity travels along the boundary in a wave-like manner. Upon contact with a surface, the protrusions stop moving relative to the surface, and the boundary shape thus reflects the history of protrusive motion. The wave-like character of protrusions provides a plausible mechanism for the zig-zagging of pseudopods and for the ability of cells both to swim in viscous fluids and to navigate complex three dimensional topography.

  19. Gas6 Promotes Inflammatory (CCR2hiCX3CR1lo) Monocyte Recruitment in Venous Thrombosis.

    Science.gov (United States)

    Laurance, Sandrine; Bertin, François-René; Ebrahimian, Talin; Kassim, Yusra; Rys, Ryan N; Lehoux, Stéphanie; Lemarié, Catherine A; Blostein, Mark D

    2017-07-01

    Coagulation and inflammation are inter-related. Gas6 (growth arrest-specific 6) promotes venous thrombosis and participates to inflammation through endothelial-innate immune cell interactions. Innate immune cells can provide the initiating stimulus for venous thrombus development. We hypothesize that Gas6 promotes monocyte recruitment during venous thrombosis. Deep venous thrombosis was induced in wild-type and Gas6-deficient (-/-) mice using 5% FeCl 3 and flow reduction in the inferior vena cava. Total monocyte depletion was achieved by injection of clodronate before deep venous thrombosis. Inflammatory monocytes were depleted using an anti-C-C chemokine receptor type 2 (CCR2) antibody. Similarly, injection of an anti-chemokine ligand 2 (CCL2) antibody induced CCL2 depletion. Flow cytometry and immunofluorescence were used to characterize the monocytes recruited to the thrombus. In vivo, absence of Gas6 was associated with a reduction of monocyte recruitment in both deep venous thrombosis models. Global monocyte depletion by clodronate leads to smaller thrombi in wild-type mice. Compared with wild type, the thrombi from Gas6 -/- mice contain less inflammatory (CCR2 hi CX 3 CR1 lo ) monocytes, consistent with a Gas6-dependent recruitment of this monocyte subset. Correspondingly, selective depletion of CCR2 hi CX 3 CR1 lo monocytes reduced the formation of venous thrombi in wild-type mice demonstrating a predominant role of the inflammatory monocytes in thrombosis. In vitro, the expression of both CCR2 and CCL2 were Gas6 dependent in monocytes and endothelial cells, respectively, impacting monocyte migration. Moreover, Gas6-dependent CCL2 expression and monocyte migration were mediated via JNK (c-Jun N-terminal kinase). This study demonstrates that Gas6 specifically promotes the recruitment of inflammatory CCR2 hi CX 3 CR1 lo monocytes through the regulation of both CCR2 and CCL2 during deep venous thrombosis. © 2017 American Heart Association, Inc.

  20. Expression of NK cell and monocyte receptors in critically ill patients - potential biomarkers of sepsis

    DEFF Research Database (Denmark)

    Kjaergaard, A G; Nielsen, Jeppe Sylvest; Tønnesen, Else

    2015-01-01

    UNLABELLED: Sepsis is characterized by activation of both the innate and adaptive immune systems as a response to infection. During sepsis, the expression of surface receptors expressed on immune competent cells, such as NKG2D and NKp30 on NK cells and TLR4 and CD14 on monocytes, is partly...... regulated by pro- and anti-inflammatory mediators. In this observational study, we aimed to explore whether the expression of these receptors could be used as diagnostic and/or prognostic biomarkers in sepsis. Patients with severe sepsis or septic shock (n = 21) were compared with critically ill non...... were higher in the septic patients compared with the non-septic patients (P sepsis...

  1. Phenotypic and functional modulation of porcine monocyte-derived ...

    African Journals Online (AJOL)

    Jane

    2011-08-08

    Aug 8, 2011 ... monocyte-derived dendritic cells for foot-and-mouth disease virus. Hai-yan Shen1# ... tissues, to migrate to secondary lymphoid organs and to provide the ... innate and adaptive immune responses mentioned earlier led us to ...

  2. Speeding up pyrogenicity testing: Identification of suitable cell components and readout parameters for an accelerated monocyte activation test (MAT).

    Science.gov (United States)

    Stoppelkamp, Sandra; Würschum, Noriana; Stang, Katharina; Löder, Jasmin; Avci-Adali, Meltem; Toliashvili, Leila; Schlensak, Christian; Wendel, Hans Peter; Fennrich, Stefan

    2017-02-01

    Pyrogen testing represents a crucial safety measure for parental drugs and medical devices, especially in direct contact with blood or liquor. The European Pharmacopoeia regulates these quality control measures for parenterals. Since 2010, the monocyte activation test (MAT) has been an accepted pyrogen test that can be performed with different human monocytic cell sources: whole blood, isolated monocytic cells or monocytic cell lines with IL1β, IL6, or TNFα as readout cytokines. In the present study, we examined the three different cell sources and cytokine readout parameters with the scope of accelerating the assay time. We could show that despite all cell types being able to detect pyrogens, primary cells were more sensitive than the monocytic cell line. Quantitative real-time PCR revealed IL6 mRNA transcripts having the largest change in Ct-values upon LPS-stimulation compared to IL1β and TNFα, but quantification was unreliable. IL6 protein secretion from whole blood or peripheral blood mononuclear cells (PBMC) was also best suited for an accelerated assay with a larger linear range and higher signal-to-noise ratios upon LPS-stimulation. The unique combination with propan-2-ol or a temperature increase could additionally increase the cytokine production for earlier detection in PBMC. The increased incubation temperature could finally increase not only responses to lipopolysaccharides (LPS) but also other pyrogens by up to 13-fold. Therefore, pyrogen detection can be accelerated considerably by using isolated primary blood cells with an increased incubation temperature and IL6 as readout. These results could expedite assay time and thus help to promote further acceptance of the MAT. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  3. The Fc-receptor III of cultured human monocytes. Structural similarity with FcRIII of natural killer cells and role in the extracellular lysis of sensitized erythrocytes

    NARCIS (Netherlands)

    Klaassen, R. J.; Ouwehand, W. H.; Huizinga, T. W.; Engelfriet, C. P.; von dem Borne, A. E.

    1990-01-01

    FcRIII is not present on peripheral blood monocytes, but becomes expressed upon culturing and can be demonstrated on tissue macrophages. We studied the expression of FcRIII of cultured monocytes in detail and compared its structure with FcRIII of neutrophils and NK cells. The cell density of FcRIII

  4. Macrophage migration inhibitory factor stimulated by Helicobacter pylori increases proliferation of gastric epithelial cells

    Science.gov (United States)

    Xia, Harry Hua-Xiang; Lam, Shiu Kum; Chan, Annie O.O.; Lin, Marie Chia Mi; Kung, Hsiang Fu; Ogura, Keiji; Berg, Douglas E.; Wong, Benjamin C. Y.

    2005-01-01

    AIM: Helicobacter pylori (H pylori) is associated with increased gastric inflammatory and epithelial expression of macrophage migration inhibitory factor (MIF) and gastric epithelial cell proliferation. This study aimed at determining whether H pylori directly stimulates release of MIF in monocytes, whether the cag pathogenicity island (PAI) is involved for this function, and whether MIF stimulated by H pylori increases gastric epithelial cell proliferation in vitro. METHODS: A cytotoxic wild-type H pylori strain (TN2)and its three isogenic mutants (TN2△cag, TN2△cagA and TN2△cagE) were co-cultured with cells of a human monocyte cell line, THP-1, for 24 h at different organism/cell ratios. MIF in the supernatants was measured by an ELISA. Cells of a human gastric cancer cell line, MKN45, were then co-cultured with the supernatants, with and without monoclonal anti-MIF antibody for 24 h. The cells were further incubated for 12 h after addition of 3H-thymidine, and the levels of incorporation of 3H-thymidine were measured with a liquid scintillation counter. RESULTS: The wild-type strain and the isogenic mutants, TN2△cagA and TN2△cagE, increased MIF release at organism/cell ratios of 200/1 and 400/1, but not at the ratios of 50/1 and 100/1. However, the mutant TN2△cag did not increase the release of MIF at any of the four ratios. 3H-thymidine readings for MKN-45 cells were significantly increased with supernatants derived from the wild-type strain and the mutants TN2△cagA and TN2△cagE, but not from the mutant TN2△cag. Moreover, in the presence of monoclonal anti-MIF antibody, the stimulatory effects of the wild-type strain on cell proliferation disappeared. CONCLUSION: H pylori stimulates MIF release in monocytes, likely through its cag PAI, but not related to cagA or cagE. H pylori-stimulated monocyte culture supernatant increases gastric cell proliferation, which is blocked by anti-MIF antibody, suggesting that MIF plays an important role in H

  5. Indirect induction of endothelial cell injury by PU- or PTFE-mediated activation of monocytes.

    Science.gov (United States)

    Liu, Xin; Xue, Yang; Sun, Jiao

    2010-01-01

    Polyurethanes (PUs) and polytetrafluoroethylene (PTFE) are widely used for making cardiovascular devices, but thrombus formation on the surfaces of these devices is inevitable. Since endothelial injury can lead to thrombosis, most of the studies on PUs or PTFE focused on their damage to endothelial cells. However, few studies have attempted to clarify whether the use of foreign objects as biomaterials can cause endothelial injury by activating the innate immune system. In this study, we aimed to investigate the roles of PU- or PTFE-stimulated immune cells in endothelial-cell injury. First, monocytes (THP-1 cells) were stimulated with PU or PTFE for 24 h and, subsequently, human umbilical vein endothelial cells (HUVECs) were treated with the supernatants of the stimulated cells for 24 h. We measured the generation of intracellular reactive oxygen species (ROS) from THP-1 cells treated with PU and PTFE for 24 h, meanwhile hydrogen dioxide (H(2)O(2)), tumor necrosis factor (TNF)-α and interleukin (IL)-1β in the supernatants were also detected. Then, we assessed the apoptosis rate of the HUVECs and determined the expression of NO, inducible nitric oxide synthase (iNOS), and apoptosis-related proteins (p53, Bax, Bcl-2) in the HUVECs. The results showed that large amounts of ROS and low levels of pro-inflammatory cytokines (TNF-α and IL-1β) were produced by the stimulated THP-1 cells. After culturing with the supernatants of the PU- or PTFE-stimulated THP-1 cells, the apoptosis rate, NO production and expression of iNOS, p53 and Bax in the HUVECs were up-regulated, while Bcl-2 expression was down-regulated. In conclusion, the release of ROS by PU- or PTFE-treated THP-1 cells may induce iNOS expression and cause apoptosis in HUVECs via the p53, Bax and Bcl-2 proteins. These data provide the interesting finding that endothelial injury in the process of biomaterial-induced thrombosis can be initiated through the release of soluble mediators by monocytes.

  6. Human monocytes undergo functional re-programming during differentiation to dendritic cell mediated by human extravillous trophoblasts.

    Science.gov (United States)

    Zhao, Lei; Shao, Qianqian; Zhang, Yun; Zhang, Lin; He, Ying; Wang, Lijie; Kong, Beihua; Qu, Xun

    2016-02-09

    Maternal immune adaptation is required for a successful pregnancy to avoid rejection of the fetal-placental unit. Dendritic cells within the decidual microenvironment lock in a tolerogenic profile. However, how these tolerogenic DCs are induced and the underlying mechanisms are largely unknown. In this study, we show that human extravillous trophoblasts redirect the monocyte-to-DC transition and induce regulatory dendritic cells. DCs differentiated from blood monocytes in the presence of human extravillous trophoblast cell line HTR-8/SVneo displayed a DC-SIGN(+)CD14(+)CD1a(-) phenotype, similar with decidual DCs. HTR8-conditioned DCs were unable to develop a fully mature phenotype in response to LPS, and altered the cytokine secretory profile significantly. Functionally, conditioned DCs poorly induced the proliferation and activation of allogeneic T cells, whereas promoted CD4(+)CD25(+)Foxp3(+) Treg cells generation. Furthermore, the supernatant from DC and HTR-8/SVneo coculture system contained significant high amount of M-CSF and MCP-1. Using neutralizing antibodies, we discussed the role of M-CSF and MCP-1 during monocyte-to-DCs differentiation mediated by extravillous trophoblasts. Our data indicate that human extravillous trophoblasts play an important role in modulating the monocyte-to-DC differentiation through M-CSF and MCP-1, which facilitate the establishment of a tolerogenic microenvironment at the maternal-fetal interface.

  7. Proteoglycan biosynthesis in murine monocytic leukemic (M1) cells before and after differentiation

    International Nuclear Information System (INIS)

    McQuillan, D.J.; Yanagishita, M.; Hascall, V.C.; Bickel, M.

    1989-01-01

    Murine monocytic leukemic (M1) cells were cultured in the presence of [ 3 H]glucosamine and [ 35 S]sulfate. Labeled proteoglycans were purified by anion exchange chromatography and characterized by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with chemical and enzymatic degradation. M1 cells synthesize a single predominant species of proteoglycan which distributes almost equally between the cell and medium after 17 h labeling. The cell-associated proteoglycan has an overall size of about 135 kDa and contains three to five chondroitin sulfate chains (28-31 kDa each) attached to a chondroitinase-generated core protein of 28 kDa. The synthesis and subsequent secretion of this proteoglycan was enhanced 4-5-fold in cells induced to differentiate into macrophages. This was not a phenomenon of arrest in the G0/G1 stage of the cell cycle, since density inhibited undifferentiated cells arrested at this stage did not increase proteoglycan synthesis. The chondroitin sulfate chains contained exclusively chondroitin 4- and 6-sulfate; however, the ratio of these two disaccharides differed between the medium- and cell-associated proteoglycans, and changed during progression of the cells into a fully differentiated phenotype. Pulse-chase kinetics indicate the presence of two distinct pools of proteoglycan; one that is secreted very rapidly from the cell after a approximately 1-h lag, and a second pool that is turned over in the cell with a half-time of approximately 3.5 h. Subtle differences in the glycosylation patterns of the medium- and cell-associated species are consistent with synthesis of two pools. Papain digestion suggests that the chondroitin sulfate chains are clustered on a small protease resistant peptide. The data suggest that this proteoglycan is similar to the serglycin proteoglycan family

  8. Total white blood cell counts and LPS-induced TNF alpha production by monocytes of pregnant, pseudopregnant and cyclic rats

    NARCIS (Netherlands)

    Faas, MM; Moes, H; van der Schaaf, G; de Leij, LFMH; Heineman, MJ

    Pregnancy in the rat may be associated with an activated innate immune system. Therefore, we investigated monocyte function as well as total white blood cell (WBC) counts during the follicular phase of the ovarian cycle, pregnancy and pseudopregnancy in the rat. Rats were equipped with a permanent

  9. Total white blood cell counts and LPS-induced TNF alpha production by monocytes of pregnant, pseudopregnant and cyclic rats

    NARCIS (Netherlands)

    Faas, M. M.; Moes, H.; van der Schaaf, G.; de Leij, L. F. M. H.; Heineman, M. J.

    2003-01-01

    Pregnancy in the rat may be associated with an activated innate immune system. Therefore, we investigated monocyte function as well as total white blood cell (WBC) counts during the follicular phase of the ovarian cycle, pregnancy and pseudopregnancy in the rat. Rats were equipped with a permanent

  10. The role of granulocyte macrophage colony stimulating factor (GM-CSF) in radiation-induced tumor cell migration.

    Science.gov (United States)

    Vilalta, Marta; Brune, Jourdan; Rafat, Marjan; Soto, Luis; Graves, Edward E

    2018-03-13

    Recently it has been observed in preclinical models that that radiation enhances the recruitment of circulating tumor cells to primary tumors, and results in tumor regrowth after treatment. This process may have implications for clinical radiotherapy, which improves control of a number of tumor types but which, despite continued dose escalation and aggressive fractionation, is unable to fully prevent local recurrences. By irradiating a single tumor within an animal bearing multiple lesions, we observed an increase in tumor cell migration to irradiated and unirradiated sites, suggesting a systemic component to this process. Previous work has identified the cytokine GM-CSF, produced by tumor cells following irradiation, as a key effector of this process. We evaluated the ability of systemic injections of a PEGylated form of GM-CSF to stimulate tumor cell migration. While increases in invasion and migration were observed for tumor cells in a transwell assay, we found that daily injections of PEG-GM-CSF to tumor-bearing animals did not increase migration of cells to tumors, despite the anticipated changes in circulating levels of granulocytes and monocytes produced by this treatment. Combination of PEG-GM-CSF treatment with radiation also did not increase tumor cell migration. These findings suggest that clinical use of GM-CSF to treat neutropenia in cancer patients will not have negative effects on the aggressiveness of residual cancer cells. However, further work is needed to characterize the mechanism by which GM-CSF facilitates systemic recruitment of trafficking tumor cells to tumors.

  11. The Effect of Aggregatibacter actinomycetemcomitans Leukotoxin on Cell-to-Cell Communication in Human Monocytes

    OpenAIRE

    Hussein, Bejyan; Tamali, Ali

    2014-01-01

    Periodontitis is a complex and multifactorial dental disease and without proper treatment it eventually leads to the loss of teeth. Individuals with a higher number of Aggregatibacter actinomycetemcomitans in their micro-flora have a greater risk of developing periodontitis and the bacterium is associated with local aggressive periodontitis that affects younger populations. Aa produces a toxic virulent factor named Leukotoxin-A that can activate monocytes by a cellular chain reaction resultin...

  12. Functional Defects in Type 3 Innate Lymphoid Cells and Classical Monocytes in a Patient with Hyper-IgE Syndrome.

    Science.gov (United States)

    Chang, Yuna; Kang, Sung-Yoon; Kim, Jihyun; Kang, Hye-Ryun; Kim, Hye Young

    2017-10-01

    Hyper-IgE syndrome (HIES) is a very rare primary immune deficiency characterized by elevated serum IgE levels, recurrent bacterial infections, chronic dermatitis, and connective tissue abnormalities. Autosomal dominant (AD) HIES involves a mutation in signal transducer and activator of transcription 3 (STAT3) that leads to an impaired T H 17 response. STAT3 signaling is also involved in the function of RORγt + type 3 innate lymphoid cells (ILC3s) and RORγt + T H 17 cells. The aim of this study was to investigate the role of innate immune cells such as innate lymphoid cells (ILCs), granulocytes, and monocytes in a patient with HIES. Peripheral blood mononuclear cells (PBMCs) from a patient with HIES and three age-matched healthy controls were obtained for the analysis of the innate and adaptive immune cells. The frequencies of ILCs in PBMCs were lower in the patient with HIES than in the controls. Moreover, granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-17A produced by ILC3s in PBMCs were lower in the patient with HIES than the controls. Compared with the controls, classical monocytes (CD14 + CD16 low ), which have a high antimicrobial capability, were also lower in the patient with HIES, while non-classical monocytes (CD14 low CD16 + ) as well as intermediate monocytes (CD14 + CD16 intermediate ) were higher. Taken together, these results indicate that the impaired immune defense against pathogenic microbes in the patient with HIES might be partially explained by functional defects in ILC3s and inflammatory monocytes.

  13. Differential migration and proliferation of geometrical ensembles of cell clusters

    International Nuclear Information System (INIS)

    Kumar, Girish; Chen, Bo; Co, Carlos C.; Ho, Chia-Chi

    2011-01-01

    Differential cell migration and growth drives the organization of specific tissue forms and plays a critical role in embryonic development, tissue morphogenesis, and tumor invasion. Localized gradients of soluble factors and extracellular matrix have been shown to modulate cell migration and proliferation. Here we show that in addition to these factors, initial tissue geometry can feedback to generate differential proliferation, cell polarity, and migration patterns. We apply layer by layer polyelectrolyte assembly to confine multicellular organization and subsequently release cells to demonstrate the spatial patterns of cell migration and growth. The cell shapes, spreading areas, and cell-cell contacts are influenced strongly by the confining geometry. Cells within geometric ensembles are morphologically polarized. Symmetry breaking was observed for cells on the circular pattern and cells migrate toward the corners and in the direction parallel to the longest dimension of the geometric shapes. This migration pattern is disrupted when actomyosin based tension was inhibited. Cells near the edge or corner of geometric shapes proliferate while cells within do not. Regions of higher rate of cell migration corresponded to regions of concentrated growth. These findings demonstrate that multicellular organization can result in spatial patterns of migration and proliferation.

  14. Monocytic and granulocytic myeloid derived suppressor cells differentially regulate spatiotemporal tumour plasticity during metastatic cascade.

    Science.gov (United States)

    Ouzounova, Maria; Lee, Eunmi; Piranlioglu, Raziye; El Andaloussi, Abdeljabar; Kolhe, Ravindra; Demirci, Mehmet F; Marasco, Daniela; Asm, Iskander; Chadli, Ahmed; Hassan, Khaled A; Thangaraju, Muthusamy; Zhou, Gang; Arbab, Ali S; Cowell, John K; Korkaya, Hasan

    2017-04-06

    It is widely accepted that dynamic and reversible tumour cell plasticity is required for metastasis, however, in vivo steps and molecular mechanisms are poorly elucidated. We demonstrate here that monocytic (mMDSC) and granulocytic (gMDSC) subsets of myeloid-derived suppressor cells infiltrate in the primary tumour and distant organs with different time kinetics and regulate spatiotemporal tumour plasticity. Using co-culture experiments and mouse transcriptome analyses in syngeneic mouse models, we provide evidence that tumour-infiltrated mMDSCs facilitate tumour cell dissemination from the primary site by inducing EMT/CSC phenotype. In contrast, pulmonary gMDSC infiltrates support the metastatic growth by reverting EMT/CSC phenotype and promoting tumour cell proliferation. Furthermore, lung-derived gMDSCs isolated from tumour-bearing animals enhance metastatic growth of already disseminated tumour cells. MDSC-induced 'metastatic gene signature' derived from murine syngeneic model predicts poor patient survival in the majority of human solid tumours. Thus spatiotemporal MDSC infiltration may have clinical implications in tumour progression.

  15. cGAS Senses Human Cytomegalovirus and Induces Type I Interferon Responses in Human Monocyte-Derived Cells.

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    Jennifer Paijo

    2016-04-01

    Full Text Available Human cytomegalovirus (HCMV infections of healthy individuals are mostly unnoticed and result in viral latency. However, HCMV can also cause devastating disease, e.g., upon reactivation in immunocompromised patients. Yet, little is known about human immune cell sensing of DNA-encoded HCMV. Recent studies indicated that during viral infection the cyclic GMP/AMP synthase (cGAS senses cytosolic DNA and catalyzes formation of the cyclic di-nucleotide cGAMP, which triggers stimulator of interferon genes (STING and thus induces antiviral type I interferon (IFN-I responses. We found that plasmacytoid dendritic cells (pDC as well as monocyte-derived DC and macrophages constitutively expressed cGAS and STING. HCMV infection further induced cGAS, whereas STING expression was only moderately affected. Although pDC expressed particularly high levels of cGAS, and the cGAS/STING axis was functional down-stream of STING, as indicated by IFN-I induction upon synthetic cGAMP treatment, pDC were not susceptible to HCMV infection and mounted IFN-I responses in a TLR9-dependent manner. Conversely, HCMV infected monocyte-derived cells synthesized abundant cGAMP levels that preceded IFN-I production and that correlated with the extent of infection. CRISPR/Cas9- or siRNA-mediated cGAS ablation in monocytic THP-1 cells and primary monocyte-derived cells, respectively, impeded induction of IFN-I responses following HCMV infection. Thus, cGAS is a key sensor of HCMV for IFN-I induction in primary human monocyte-derived DC and macrophages.

  16. Transfecting Human Monocytes with RNA.

    Science.gov (United States)

    Dannull, Jens; Nair, Smita K

    2016-01-01

    Targeting monocytes as a delivery system for drugs or nucleic acids, and thereby harnessing their natural tissue-infiltrating capacity, has become an area of intense investigation in both basic and clinical research. Herein we describe an efficient method to deliver mRNA (messenger RNA) or siRNA (small interfering RNA) into human monocytes by electroporation. This method can be applied in the laboratory to monocytes isolated via magnetic bead-based techniques, or in a clinical setting using monocytes that were collected via counterflow centrifugation elutriation using the Elutra(®) Cell Separation System. We further demonstrate that electroporation of monocytes with RNA represents a robust and highly relevant approach to modify monocytes for cell-based therapies. Last, the procedure described can readily be adapted to monocytes from different species, hence facilitating research in animal models.

  17. Evaluating the Effects of Cytomegalovirus Glycoprotein B on the Maturation and Function of Monocyte-derived dendritic cells

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    Afsson shariat

    2015-11-01

    Full Text Available Background & Objectives: Interaction of cytomegalovirus glycoprotein B with toll-like receptors of dendritic cells leads to early signaling and innate immune responses. The aim of this study is to evaluate the effects of cytomegalovirus glycoprotein B on the maturation and function of monocyte-derived dendritic cells in treated groups in comparison with control groups. Materials & Methods: Blood samples were taken from 5 healthy volunteers. Following the generation of monocyte-derived dendritic cells on the fifth day of cell culture, half of the immature dendritic cells were treated with cytomegalovirus glycoprotein B, and the rest of them were induced to mature dendritic untreated cells and were used as the control group. The maturation and function of dendritic cells were evaluated in these two groups. Results: The gene expression level of toll-like receptor-4 significantly increased in the group treated with glycoprotein B (p < 0.05, whereas there were no significant differences in the expression rates of CD83, CD86, CD1a, and HLA-DR and the secretion of IL-23 from monocyte-derived dendritic cells between the treated groups and the controls. Conclusion: The increase in the gene expression of toll-like receptor-4 in monocyte-derived dendritic cells treated with cytomegalovirus glycoprotein B showed that cell contact is required to elicit cellular antiviral response and toll-like receptor activation. Thus, it is critical to recognize the viral and cellular determinants of the immune system in order to develop new therapeutic strategies against cytomegalovirus.

  18. Combination of Cobe AutoPBSC and Gambro Elutra as a platform for monocyte enrichment in dendritic cell (DC) therapy: clinical study.

    Science.gov (United States)

    Chen, Ying; Hoecker, Paul; Zeng, Jia; Dettke, Markus

    2008-01-01

    Monocytes are a common source for generating dendritic cells (DCs). The aim of the present study was to evaluate the efficiency of a platform for monocyte collection and enrichment in a clinical setting. The platform was based on the combination of two semiautomated devices; the Cobe Spectra Auto PBSC for mononuclear cells (MNC) collection followed by counterflow elutriation for monocyte enrichment (Gambro BCT Elutra). Twenty-four patients with various types of epithelial cancer participated in the study. MNC collections were first performed as large volume leukapheresis (LVL). Subsequently, MNC products were processed with an elutriation system for monocyte isolation. LVL resulted in the collection of MNC at a median of 8.1 x 10(9) cells, containing of 31.4% monocytes. A similar efficacy was also shown in patients with lower peripheral blood counts. Elutriation of the MNC product with the Cobe Elutra device resulted in the enrichment of monocytes at a median of 2.7 x 10(9) cells, with a recovery of 80.2% and a purity of 90.7%. These monocytes were then successfully developed into DCs for clinical therapy after in vitro manipulation. These data suggest that the combination of the Cobe Spectra Auto PBSC and the Gambro BCT Elutra is an effective platform for monocyte enrichment in clinical practice according to GCP standards and GMP guidelines, and can be easily implemented in the clinical routine under current DC protocols. Copyright 2008 Wiley-Liss, Inc.

  19. Targeting Toll-like receptor 7/8 enhances uptake of apoptotic leukemic cells by monocyte-derived dendritic cells but interferes with subsequent cytokine-induced maturation.

    Science.gov (United States)

    van den Ancker, Willemijn; van Luijn, Marvin M; Ruben, Jurjen M; Westers, Theresia M; Bontkes, Hetty J; Ossenkoppele, Gert J; de Gruijl, Tanja D; van de Loosdrecht, Arjan A

    2011-01-01

    Therapeutic vaccination with dendritic cells (DC) is an emerging investigational therapy for eradication of minimal residual disease in acute myeloid leukemia. Various strategies are being explored in manufacturing DC vaccines ex vivo, e.g., monocyte-derived DC (MoDC) loaded with leukemia-associated antigens (LAA). However, the optimal source of LAA and the choice of DC-activating stimuli are still not well defined. Here, loading with leukemic cell preparations (harboring both unknown and known LAA) was explored in combination with a DC maturation-inducing cytokine cocktail (CC; IL-1β, IL-6, TNF-α, and PGE(2)) and Toll-like receptor ligands (TLR-L) to optimize uptake. Since heat shock induced apoptotic blasts were more efficiently taken up than lysates, we focused on uptake of apoptotic leukemic cells. Uptake of apoptotic blast was further enhanced by the TLR7/8-L R848 (20-30%); in contrast, CC-induced maturation inhibited uptake. CC, and to a lesser extent R848, enhanced the ability of MoDC to migrate and stimulate T cells. Furthermore, class II-associated invariant chain peptide expression was down-modulated after R848- or CC-induced maturation, indicating enhanced processing and presentation of antigenic peptides. To improve both uptake and maturation, leukemic cells and MoDC were co-incubated with R848 for 24 h followed by addition of CC. However, this approach interfered with CC-mediated MoDC maturation as indicated by diminished migratory and T cell stimulatory capacity, and the absence of IL-12 production. Taken together, our data demonstrate that even though R848 improved uptake of apoptotic leukemic cells, the sequential use of R848 and CC is counter-indicated due to its adverse effects on MoDC maturation.

  20. Directional Cell Migration in Response to Repeated Substratum Stretching

    Science.gov (United States)

    Okimura, Chika; Iwadate, Yoshiaki

    2017-10-01

    Crawling migration plays an essential role in a variety of biological phenomena, including development, wound healing, and immune system function. Migration properties such as anterior-posterior polarity, directionality, and velocity are regulated not only by the reception of a chemoattractant but also by sensing mechanical inputs from the external environment. In this review, we describe the mechanical response of migrating cells, particularly under repeated stretching of the elastic substratum, highlighting the fact that there appear to be two independent mechanosensing systems that generate the polarity needed for migration. Cells that have no stress fibers, such as Dictyostelium cells and neutrophil-like differentiated HL-60 cells, migrate perpendicular to the stretching direction via myosin II localization. Cells that do possess stress fibers, however, such as fish keratocytes, migrate parallel to the stretching via a stress-fiber-dependent process.

  1. Lutein Inhibits the Migration of Retinal Pigment Epithelial Cells via Cytosolic and Mitochondrial Akt Pathways (Lutein Inhibits RPE Cells Migration

    Directory of Open Access Journals (Sweden)

    Ching-Chieh Su

    2014-08-01

    Full Text Available During the course of proliferative vitreoretinopathy (PVR, the retinal pigment epithelium (RPE cells will de-differentiate, proliferate, and migrate onto the surfaces of the sensory retina. Several studies have shown that platelet-derived growth factor (PDGF can induce migration of RPE cells via an Akt-related pathway. In this study, the effect of lutein on PDGF-BB-induced RPE cells migration was examined using transwell migration assays and Western blot analyses. We found that both phosphorylation of Akt and mitochondrial translocation of Akt in RPE cells induced by PDGF-BB stimulation were suppressed by lutein. Furthermore, the increased migration observed in RPE cells with overexpressed mitochondrial Akt could also be suppressed by lutein. Our results demonstrate that lutein can inhibit PDGF-BB induced RPE cells migration through the inhibition of both cytoplasmic and mitochondrial Akt activation.

  2. The role of monocytes and monocyte-derived dendritic cells in type 1 diabetes mellitus and autoimmune thyroid disease

    NARCIS (Netherlands)

    W.K. Lam-Tse

    2003-01-01

    textabstractType 1 diabetes mellitus (DM1) and autoimmune thyroid disease (AITD) are organ specific autoimmune diseases in which the immune system is directed against the ß cells and the thyrocytes respectively. The etio-pathogenesis of organ-specific or endocrine autoimmune diseases is complex,

  3. Endothelial-monocyte activating polypeptide II alters fibronectin based endothelial cell adhesion and matrix assembly via alpha5 beta1 integrin

    International Nuclear Information System (INIS)

    Schwarz, Margaret A.; Zheng, Hiahua; Liu, Jie; Corbett, Siobhan; Schwarz, Roderich E.

    2005-01-01

    Mature Endothelial-Monocyte Activating Polypeptide (mEMAP) II functions as a potent antiangiogenic peptide. Although the anti-tumor effect of mEMAP II has been described, little is known regarding its mechanism of action. Observations that mEMAP II induced apoptosis only in a subset of migrating and proliferating endothelial cells (EC) suggests a targeted effect on cells engaged in angiogenic activities which are known to rely upon cell adhesion and migration. Indeed, we demonstrate that mEMAP II inhibited fibronectin (FN) dependent microvascular EC (MEC) adhesion and spreading and we show that this depends upon the alpha5 beta1 integrin. Immunofluorescence analysis demonstrated that mEMAP II-dependent blockade of FN-alpha5 beta1 interactions was associated with disassembly of both actin stress fiber networks and FN matrix. These findings suggest that mEMAP II blocks MEC adhesion and spreading on fibronectin, via a direct interaction with the integrin alpha5 beta1, thus implicating that alpha5 integrin may be a mediator of mEMAP II's antiangiogenic function

  4. Association of Canine Osteosarcoma and Monocyte Phenotype and Chemotactic Function.

    Science.gov (United States)

    Tuohy, J L; Lascelles, B D X; Griffith, E H; Fogle, J E

    2016-07-01

    Monocytes/macrophages are likely key cells in immune modulation in dogs with osteosarcoma (OSA). Increased peripheral monocyte counts are negatively correlated with shorter disease-free intervals in dogs with OSA. Understanding the monocyte/macrophage's modulatory role in dogs with OSA can direct further studies in immunotherapy development for OSA. That OSA evades the immune response by down-regulating monocyte chemokine receptor expression and migratory function, and suppresses host immune responses. Eighteen dogs with OSA that have not received definitive treatment and 14 healthy age-matched controls Clinical study-expression of peripheral blood monocyte cell surface receptors, monocyte mRNA expression and cytokine secretion, monocyte chemotaxis, and survival were compared between clinical dogs with OSA and healthy control dogs. Cell surface expression of multiple chemokine receptors is significantly down-regulated in peripheral blood monocytes of dogs with OSA. The percentage expression of CCR2 (median 58%, range 2-94%) and CXCR2 expression (median 54%, range 2-92%) was higher in control dogs compared to dogs with OSA (CCR2 median 29%, range 3-45%, P = 0.0006; CXCR2 median 23%, range 0.2-52%, P = 0.0007). Prostaglandin E2 (PGE2 ) (OSA, median 347.36 pg/mL, range 103.4-1268.5; control, 136.23 pg/mL, range 69.93-542.6, P = .04) and tumor necrosis factor-alpha (TNF-α) (P = .02) levels are increased in OSA monocyte culture supernatants compared to controls. Peripheral blood monocytes of dogs with OSA exhibit decreased chemotactic function when compared to control dogs (OSA, median 1.2 directed to random migration, range 0.8-1.25; control, 1.6, range of 0.9-1.8, P = .018). Dogs with OSA have decreased monocyte chemokine receptor expression and monocyte chemotaxis, potential mechanisms by which OSA might evade the immune response. Reversal of monocyte dysfunction using immunotherapy could improve survival in dogs with OSA. Copyright © 2016 The Authors. Journal of

  5. Chemoresistance of human monocyte-derived dendritic cells is regulated by IL-17A.

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    Selma Olsson Åkefeldt

    Full Text Available Dendritic cells initiate adaptive immune responses, leading either to control cancer by effector T cells or to exacerbate cancer by regulatory T cells that inhibit IFN-γ-mediated Th1-type response. Dendritic cells can also induce Th17-type immunity, mediated by IL-17A. However, the controversial role of this cytokine in cancer requires further investigations. We generated dendritic cells from peripheral blood monocytes to investigate lifespan, phenotype and chemoresistance of dendritic cells, treated with IL-17A with or without IFN-γ. Studying the expression of Bcl-2 family members, we demonstrated that dendritic cells constitutively express one pro-survival Bcl-2 member: MCL1. Immature dendritic cells were CD40(lowHLADR(low CD1a(+ MCL1(+, did not express CD14, CD68 or BCL2A1, and displayed a short 2-day lifespan. IL-17A-treated DC exhibited a semi-mature (CD40(high HLADR(low pre-M2 (CCL22(+ CD206(+ CD163(+ IL1RN(+ IL-10(- CXCL10(- IL-12(- mixed (CD1a(+ CD14+ CD68(+ macrophage-dendritic cell phenotype. They efficiently exerted mannose receptor-mediated endocytosis and did not produce superoxide anions, in the absence of TLR engagement. Interestingly, IL-17A promoted a long-term survival of dendritic cells, beyond 12 days, that correlated to BCL2A1 induction, a pro-survival Bcl-2 family member. BCL2A1 transcription was activated by NF-κB, downstream of IL-17A transduction. Thus, immature dendritic cells only express MCL1, whereas IL-17A-treated dendritic cells concomitantly expressed two pro-survival Bcl-2 family members: MCL1 and BCL2A1. These latter developed chemoresistance to 11 of the 17 chemotherapy agents tested. However, high doses of either vinblastine or cytarabine decreased MCL1 expression and induced dendritic cell death. When IL-17A is produced in vivo, administration of anti-IL-17A biotherapy may impair dendritic cell survival by targeting BCL2A1 expression. Consequently, depending on the effector or regulatory role of dendritic

  6. Leader Cells Define Directionality of Trunk, but Not Cranial, Neural Crest Cell Migration

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    Jo Richardson

    2016-05-01

    Full Text Available Collective cell migration is fundamental for life and a hallmark of cancer. Neural crest (NC cells migrate collectively, but the mechanisms governing this process remain controversial. Previous analyses in Xenopus indicate that cranial NC (CNC cells are a homogeneous population relying on cell-cell interactions for directional migration, while chick embryo analyses suggest a heterogeneous population with leader cells instructing directionality. Our data in chick and zebrafish embryos show that CNC cells do not require leader cells for migration and all cells present similar migratory capacities. In contrast, laser ablation of trunk NC (TNC cells shows that leader cells direct movement and cell-cell contacts are required for migration. Moreover, leader and follower identities are acquired before the initiation of migration and remain fixed thereafter. Thus, two distinct mechanisms establish the directionality of CNC cells and TNC cells. This implies the existence of multiple molecular mechanisms for collective cell migration.

  7. Establishing porcine monocyte-derived macrophage and dendritic cell systems for studying the interaction with PRRSV-1

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    Helen eSingleton

    2016-06-01

    Full Text Available Monocyte-derived macrophages (MoMØ and monocyte-derived dendritic cells (MoDC are two model systems well established in human and rodent systems that can be used to study the interaction of pathogens with host cells. Porcine reproductive and respiratory syndrome virus (PRRSV is known to infect myeloid cells, such as macrophages (MØ and dendritic cells (DC. Therefore, this study aimed to establish systems for the differentiation and characterization of MoMØ and MoDC for subsequent infection with PRRSV-1. M-CSF differentiated monocyte-derived macrophages (MoMØ were stimulated with activators for classical (M1 or alternative (M2 activation. GM-CSF and IL-4 generated monocyte-derived dendritic cells (MoDC were activated with the well established maturation cocktail containing PAMPs and cytokines. In addition, MoMØ and MoDC were treated with dexamethasone and IL-10, which are known immuno-suppressive reagents. Cells were characterized by morphology, phenotype and function and porcine MØ subsets highlighted some divergence from described human counterparts, while MoDC, appeared more similar to mouse and human DCs. The infection with PRRSV-1 strain Lena demonstrated different replication kinetics between MoMØ and MoDC and within subsets of each cell type. While MoMØ susceptibility was significantly increased by dexamethasone and IL-10 with an accompanying increase in CD163/CD169 expression, MoDC supported only a minimal replication of PRRSV These findings underline the high variability in the susceptibility of porcine myeloid cells towards PRRSV-1 infection.

  8. Human bladder uroepithelial cells synergize with monocytes to promote IL-10 synthesis and other cytokine responses to uropathogenic Escherichia coli.

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    Benjamin L Duell

    Full Text Available Urinary tract infections are a major source of morbidity for women and the elderly, with Uropathogenic Escherichia coli (UPEC being the most prevalent causative pathogen. Studies in recent years have defined a key anti-inflammatory role for Interleukin-10 (IL-10 in urinary tract infection mediated by UPEC and other uropathogens. We investigated the nature of the IL-10-producing interactions between UPEC and host cells by utilising a novel co-culture model that incorporated lymphocytes, mononuclear and uroepithelial cells in histotypic proportions. This co-culture model demonstrated synergistic IL-10 production effects between monocytes and uroepithelial cells following infection with UPEC. Membrane inserts were used to separate the monocyte and uroepithelial cell types during infection and revealed two synergistic IL-10 production effects based on contact-dependent and soluble interactions. Analysis of a comprehensive set of immunologically relevant biomarkers in monocyte-uroepithelial cell co-cultures highlighted that multiple cytokine, chemokine and signalling factors were also produced in a synergistic or antagonistic fashion. These results demonstrate that IL-10 responses to UPEC occur via multiple interactions between several cells types, implying a complex role for infection-related IL-10 during UTI. Development and application of the co-culture model described in this study is thus useful to define the degree of contact dependency of biomarker production to UPEC, and highlights the relevance of histotypic co-cultures in studying complex host-pathogen interactions.

  9. Follow-the-leader cell migration requires biased cell-cell contact and local microenvironmental signals

    Science.gov (United States)

    Wynn, Michelle L.; Rupp, Paul; Trainor, Paul A.; Schnell, Santiago; Kulesa, Paul M.

    2013-06-01

    Directed cell migration often involves at least two types of cell motility that include multicellular streaming and chain migration. However, what is unclear is how cell contact dynamics and the distinct microenvironments through which cells travel influence the selection of one migratory mode or the other. The embryonic and highly invasive neural crest (NC) are an excellent model system to study this question since NC cells have been observed in vivo to display both of these types of cell motility. Here, we present data from tissue transplantation experiments in chick and in silico modeling that test our hypothesis that cell contact dynamics with each other and the microenvironment promote and sustain either multicellular stream or chain migration. We show that when premigratory cranial NC cells (at the pre-otic level) are transplanted into a more caudal region in the head (at the post-otic level), cells alter their characteristic stream behavior and migrate in chains. Similarly, post-otic NC cells migrate in streams after transplantation into the pre-otic hindbrain, suggesting that local microenvironmental signals dictate the mode of NC cell migration. Simulations of an agent-based model (ABM) that integrates the NC cell behavioral data predict that chain migration critically depends on the interplay of biased cell-cell contact and local microenvironment signals. Together, this integrated modeling and experimental approach suggests new experiments and offers a powerful tool to examine mechanisms that underlie complex cell migration patterns.

  10. Multi-cellular logistics of collective cell migration.

    Directory of Open Access Journals (Sweden)

    Masataka Yamao

    Full Text Available During development, the formation of biological networks (such as organs and neuronal networks is controlled by multicellular transportation phenomena based on cell migration. In multi-cellular systems, cellular locomotion is restricted by physical interactions with other cells in a crowded space, similar to passengers pushing others out of their way on a packed train. The motion of individual cells is intrinsically stochastic and may be viewed as a type of random walk. However, this walk takes place in a noisy environment because the cell interacts with its randomly moving neighbors. Despite this randomness and complexity, development is highly orchestrated and precisely regulated, following genetic (and even epigenetic blueprints. Although individual cell migration has long been studied, the manner in which stochasticity affects multi-cellular transportation within the precisely controlled process of development remains largely unknown. To explore the general principles underlying multicellular migration, we focus on the migration of neural crest cells, which migrate collectively and form streams. We introduce a mechanical model of multi-cellular migration. Simulations based on the model show that the migration mode depends on the relative strengths of the noise from migratory and non-migratory cells. Strong noise from migratory cells and weak noise from surrounding cells causes "collective migration," whereas strong noise from non-migratory cells causes "dispersive migration." Moreover, our theoretical analyses reveal that migratory cells attract each other over long distances, even without direct mechanical contacts. This effective interaction depends on the stochasticity of the migratory and non-migratory cells. On the basis of these findings, we propose that stochastic behavior at the single-cell level works effectively and precisely to achieve collective migration in multi-cellular systems.

  11. Analysis of primary cilia in directional cell migration in fibroblasts

    DEFF Research Database (Denmark)

    Christensen, Søren Tvorup; Veland, Iben; Schwab, Albrecht

    2013-01-01

    summarize selected methods in analyzing ciliary function in directional cell migration, including immunofluorescence microscopy, scratch assay, and chemotaxis assay by micropipette addition of PDGFRα ligands to cultures of fibroblasts. These methods should be useful not only in studying cell migration....... In particular, platelet-derived growth factor receptor alpha (PDGFRα) is compartmentalized to the primary cilium to activate signaling pathways that regulate reorganization of the cytoskeleton required for lamellipodium formation and directional migration in the presence of a specific ligand gradient. We...

  12. Integrins in cell migration – the actin connection

    OpenAIRE

    Vicente-Manzanares, Miguel; Choi, Colin Kiwon; Horwitz, Alan Rick

    2008-01-01

    The connection between integrins and actin is driving the field of cell migration in new directions. Integrins and actin are coupled through a physical linkage, which provides traction for migration. Recent studies show the importance of this linkage in regulating adhesion organization and development. Actin polymerization orchestrates adhesion assembly near the leading edge of a migrating cell, and the dynamic cross-linking of actin filaments promotes adhesion maturat...

  13. Robotic Patterning a Superhydrophobic Surface for Collective Cell Migration Screening.

    Science.gov (United States)

    Pang, Yonggang; Yang, Jing; Hui, Zhixin; Grottkau, Brian E

    2018-04-01

    Collective cell migration, in which cells migrate as a group, is fundamental in many biological and pathological processes. There is increasing interest in studying the collective cell migration in high throughput. Cell scratching, insertion blocker, and gel-dissolving techniques are some methodologies used previously. However, these methods have the drawbacks of cell damage, substrate surface alteration, limitation in medium exchange, and solvent interference. The superhydrophobic surface, on which the water contact angle is greater than 150 degrees, has been recently utilized to generate patterned arrays. Independent cell culture areas can be generated on a substrate that functions the same as a conventional multiple well plate. However, so far there has been no report on superhydrophobic patterning for the study of cell migration. In this study, we report on the successful development of a robotically patterned superhydrophobic array for studying collective cell migration in high throughput. The array was developed on a rectangular single-well cell culture plate consisting of hydrophilic flat microwells separated by the superhydrophobic surface. The manufacturing process is robotic and includes patterning discrete protective masks to the substrate using 3D printing, robotic spray coating of silica nanoparticles, robotic mask removal, robotic mini silicone blocker patterning, automatic cell seeding, and liquid handling. Compared with a standard 96-well plate, our system increases the throughput by 2.25-fold and generates a cell-free area in each well non-destructively. Our system also demonstrates higher efficiency than conventional way of liquid handling using microwell plates, and shorter processing time than manual operating in migration assays. The superhydrophobic surface had no negative impact on cell viability. Using our system, we studied the collective migration of human umbilical vein endothelial cells and cancer cells using assays of endpoint

  14. 3D cancer cell migration in a confined matrix

    Science.gov (United States)

    Alobaidi, Amani; Sun, Bo

    Cancer cell migration is widely studied in 2D motion, which does not mimic the invasion processes in vivo. More recently, 3D cell migration studies have been performed. The ability of cancer cells to migrate within the extracellular matrix depends on the physical and biochemical features of the extracellular matrix. We present a model of cell motility in confined matrix geometry. The aim of the study is to study cancer migration in collagen matrix, as a soft tissue, to investigate their motility within the confined and surrounding collagen environment. Different collagen concentrations have been used to show the ability of these cancer cells to move through such a complex structure by measuring Cancer cell migration velocity as well as the displacement. Graduate student physics department.

  15. Podosomes, But Not the Maturation Status, Determine the Protease-Dependent 3D Migration in Human Dendritic Cells.

    Science.gov (United States)

    Cougoule, Céline; Lastrucci, Claire; Guiet, Romain; Mascarau, Rémi; Meunier, Etienne; Lugo-Villarino, Geanncarlo; Neyrolles, Olivier; Poincloux, Renaud; Maridonneau-Parini, Isabelle

    2018-01-01

    Dendritic cells (DC) are professional Antigen-Presenting Cells scattered throughout antigen-exposed tissues and draining lymph nodes, and survey the body for pathogens. Their ability to migrate through tissues, a 3D environment, is essential for an effective immune response. Upon infection, recognition of Pathogen-Associated Molecular Patterns (PAMP) by Toll-like receptors (TLR) triggers DC maturation. Mature DC (mDC) essentially use the protease-independent, ROCK-dependent amoeboid mode in vivo , or in collagen matrices in vitro . However, the mechanisms of 3D migration used by human immature DC (iDC) are still poorly characterized. Here, we reveal that human monocyte-derived DC are able to use two migration modes in 3D. In porous matrices of fibrillar collagen I, iDC adopted the amoeboid migration mode. In dense matrices of gelled collagen I or Matrigel, iDC used the protease-dependent, ROCK-independent mesenchymal migration mode. Upon TLR4 activation by LPS, mDC-LPS lose the capacity to form podosomes and degrade the matrix along with impaired mesenchymal migration. TLR2 activation by Pam 3 CSK 4 resulted in DC maturation, podosome maintenance, and efficient mesenchymal migration. Under all these conditions, when DC used the mesenchymal mode in dense matrices, they formed 3D podosomes at the tip of cell protrusions. Using PGE 2 , known to disrupt podosomes in DC, we observed that the cells remained in an immature status and the mesenchymal migration mode was abolished. We also observed that, while CCL5 (attractant of iDC) enhanced both amoeboid and mesenchymal migration of iDC, CCL19 and CCL21 (attractants of mDC) only enhanced mDC-LPS amoeboid migration without triggering mesenchymal migration. Finally, we examined the migration of iDC in tumor cell spheroids, a tissue-like 3D environment. We observed that iDC infiltrated spheroids of tumor cells using both migration modes. Altogether, these results demonstrate that human DC adopt the mesenchymal mode to

  16. Expansion of monocytic myeloid-derived suppressor cells in endometriosis patients: A pilot study.

    Science.gov (United States)

    Chen, Haiwen; Qin, Shuang; Lei, Aihua; Li, Xing; Gao, Qi; Dong, Jingyin; Xiao, Qing; Zhou, Jie

    2017-06-01

    Endometriosis is a chronic inflammation disease and is closely associated with immune dysregulation. Myeloid-derived suppressor cells (MDSCs) are a negative regulator of the immune system. The aim of this study was to evaluate the possible role of MDSCs in endometriosis patients. We collected the peripheral blood and peritoneal fluid from endometriosis patients and controls and analyzed M-MDSCs level using specific monoclonal antibodies recognizing HLA-DR, CD33, CD11b, CD14 markers by flow cytometry. We found that there existed abnormal expansion of monocytic MDSCs (M-MDSCs) (HLA-DR -/low CD33 + CD11b + CD14 + ) in peripheral blood and peritoneal fluid of patients with endometriosis. Functional studies revealed that M-MDSCs from endometriosis patients significantly suppressed T-cell responses and produced high level of reactive oxygen species (ROS). The elevation of M-MDSCs from endometriosis patients may contribute to the disease progression. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Collective cell migration in morphogenesis, regeneration and cancer.

    NARCIS (Netherlands)

    Friedl, P.H.A.; Gilmour, D.

    2009-01-01

    The collective migration of cells as a cohesive group is a hallmark of the tissue remodelling events that underlie embryonic morphogenesis, wound repair and cancer invasion. In such migration, cells move as sheets, strands, clusters or ducts rather than individually, and use similar actin- and

  18. Single-cell characterization of in vitro migration and interaction dynamics of T cells expanded with IL-2 and IL-7

    Directory of Open Access Journals (Sweden)

    Johanna Maria Tauriainen

    2015-04-01

    Full Text Available T cells are pivotal in the immune defense against cancers and infectious agents. To mount an effector response against cancer cells, T cells need to migrate to the cancer-site, engage in contacts with cancer cells and perform their effector functions. Adoptive T cell therapy is an effective strategy as treatment of complications such as relapse or opportunistic infections after hematopoietic stem cell transplantations. This requires a sufficient amount of cells that are able to expand and respond to tumor or viral antigens. The cytokines interleukin (IL-2 and IL-7 drive T cell differentiation, proliferation and survival and are commonly used to expand T cells ex vivo. Here, we have used microchip-based live-cell imaging to follow the migration of individual T cells, their interactions with allogeneic monocytes, cell division and apoptosis for extended periods of time; something that cannot be achieved by commonly used methods. Our data indicate that cells grown in IL-7 + IL-2 had similar migration and contact dynamics as cells grown in IL-2 alone. However, the addition of IL-7 decreased cell death creating a more viable cell population, which should be beneficial when preparing cells for immunotherapy.

  19. Automated migration analysis based on cell texture: method & reliability

    Directory of Open Access Journals (Sweden)

    Chittenden Thomas W

    2005-03-01

    Full Text Available Abstract Background In this paper, we present and validate a way to measure automatically the extent of cell migration based on automated examination of a series of digital photographs. It was designed specifically to identify the impact of Second Hand Smoke (SHS on endothelial cell migration but has broader applications. The analysis has two stages: (1 preprocessing of image texture, and (2 migration analysis. Results The output is a graphic overlay that indicates the front lines of cell migration superimposed on each original image, with automated reporting of the distance traversed vs. time. Expert preference compares to manual placement of leading edge shows complete equivalence of automated vs. manual leading edge definition for cell migration measurement. Conclusion Our method is indistinguishable from careful manual determinations of cell front lines, with the advantages of full automation, objectivity, and speed.

  20. THP-1 monocytes but not macrophages as a potential alternative for CD34+ dendritic cells to identify chemical skin sensitizers

    International Nuclear Information System (INIS)

    Lambrechts, Nathalie; Verstraelen, Sandra; Lodewyckx, Hanne; Felicio, Ana; Hooyberghs, Jef; Witters, Hilda; Tendeloo, Viggo van; Cauwenberge, Paul van; Nelissen, Inge; Heuvel, Rosette van den; Schoeters, Greet

    2009-01-01

    Early detection of the sensitizing potential of chemicals is an emerging issue for chemical, pharmaceutical and cosmetic industries. In our institute, an in vitro classification model for prediction of chemical-induced skin sensitization based on gene expression signatures in human CD34 + progenitor-derived dendritic cells (DC) has been developed. This primary cell model is able to closely mimic the induction phase of sensitization by Langerhans cells in the skin, but it has drawbacks, such as the availability of cord blood. The aim of this study was to investigate whether human in vitro cultured THP-1 monocytes or macrophages display a similar expression profile for 13 predictive gene markers previously identified in DC and whether they also possess a discriminating capacity towards skin sensitizers and non-sensitizers based on these marker genes. To this end, the cell models were exposed to 5 skin sensitizers (ammonium hexachloroplatinate IV, 1-chloro-2,4-dinitrobenzene, eugenol, para-phenylenediamine, and tetramethylthiuram disulfide) and 5 non-sensitizers (L-glutamic acid, methyl salicylate, sodium dodecyl sulfate, tributyltin chloride, and zinc sulfate) for 6, 10, and 24 h, and mRNA expression of the 13 genes was analyzed using real-time RT-PCR. The transcriptional response of 7 out of 13 genes in THP-1 monocytes was significantly correlated with DC, whereas only 2 out of 13 genes in THP-1 macrophages. After a cross-validation of a discriminant analysis of the gene expression profiles in the THP-1 monocytes, this cell model demonstrated to also have a capacity to distinguish skin sensitizers from non-sensitizers. However, the DC model was superior to the monocyte model for discrimination of (non-)sensitizing chemicals.

  1. Microgravity modifies protein kinase C isoform translocation in the human monocytic cell line U937 and human peripheral blood T-cells

    Science.gov (United States)

    Hatton, Jason P.; Gaubert, Francois; Cazenave, Jean-Pierre; Schmitt, Didier; Hashemi, B. B. (Principal Investigator); Hughes-Fulford, M. (Principal Investigator)

    2002-01-01

    Individual protein kinase C (PKC) isoforms fulfill distinct roles in the regulation of the commitment to differentiation, cell cycle arrest, and apoptosis in both monocytes and T-cells. The human monocyte like cell line U937 and T-cells were exposed to microgravity, during spaceflight and the translocation (a critical step in PKC signaling) of individual isoforms to cell particulate fraction examined. PKC activating phorbol esters induced a rapid translocation of several PKC isoforms to the particulate fraction of U937 monocytes under terrestrial gravity (1 g) conditions in the laboratory. In microgravity, the translocation of PKC beta II, delta, and epsilon in response to phorbol esters was reduced in microgravity compared to 1 g, but was enhanced in weak hypergravity (1.4 g). All isoforms showed a net increase in particulate PKC following phorbol ester stimulation, except PKC delta which showed a net decrease in microgravity. In T-cells, phorbol ester induced translocation of PKC delta was reduced in microgravity, compared to 1 g, while PKC beta II translocation was not significantly different at the two g-levels. These data show that microgravity differentially alters the translocation of individual PKC isoforms in monocytes and T-cells, thus providing a partial explanation for the modifications previously observed in the activation of these cell types under microgravity.

  2. CD14+CD16+ monocytes are the main target of Zika virus infection in peripheral blood mononuclear cells in a paediatric study in Nicaragua.

    Science.gov (United States)

    Michlmayr, Daniela; Andrade, Paulina; Gonzalez, Karla; Balmaseda, Angel; Harris, Eva

    2017-11-01

    The recent Zika pandemic in the Americas is linked to congenital birth defects and Guillain-Barré syndrome. White blood cells (WBCs) play an important role in host immune responses early in arboviral infection. Infected WBCs can also function as 'Trojan horses' and carry viruses into immune-sheltered spaces, including the placenta, testes and brain. Therefore, defining which WBCs are permissive to Zika virus (ZIKV) is critical. Here, we analyse ZIKV infectivity of peripheral blood mononuclear cells (PBMCs) in vitro and from Nicaraguan Zika patients and show CD14 + CD16 + monocytes are the main target of infection, with ZIKV replication detected in some dendritic cells. The frequency of CD14 + monocytes was significantly decreased, while the CD14 + CD16 + monocyte population was significantly expanded during ZIKV infection compared to uninfected controls. Viral RNA was detected in PBMCs from all patients, but in serum from only a subset, suggesting PBMCs may be a reservoir for ZIKV. In Zika patients, the frequency of infected cells was lower but the percentage of infected CD14 + CD16 + monocytes was significantly higher compared to dengue cases. The gene expression profile in monocytes isolated from ZIKV- and dengue virus-infected patients was comparable, except for significant differences in interferon-γ, CXCL12, XCL1, interleukin-6 and interleukin-10 levels. Thus, our study provides a detailed picture of the innate immune profile of ZIKV infection and highlights the important role of monocytes, and CD14 + CD16 + monocytes in particular.

  3. Neuropeptide Y induces potent migration of human immature dendritic cells and promotes a Th2 polarization.

    Science.gov (United States)

    Buttari, Brigitta; Profumo, Elisabetta; Domenici, Giacomo; Tagliani, Angela; Ippoliti, Flora; Bonini, Sergio; Businaro, Rita; Elenkov, Ilia; Riganò, Rachele

    2014-07-01

    Neuropeptide Y (NPY), a major autonomic nervous system and stress mediator, is emerging as an important regulator of inflammation, implicated in autoimmunity, asthma, atherosclerosis, and cancer. Yet the role of NPY in regulating phenotype and functions of dendritic cells (DCs), the professional antigen-presenting cells, remains undefined. Here we investigated whether NPY could induce DCs to migrate, mature, and polarize naive T lymphocytes. We found that NPY induced a dose-dependent migration of human monocyte-derived immature DCs through the engagement of NPY Y1 receptor and the activation of ERK and p38 mitogen-activated protein kinases. NPY promoted DC adhesion to endothelial cells and transendothelial migration. It failed to induce phenotypic DC maturation, whereas it conferred a T helper 2 (Th2) polarizing profile to DCs through the up-regulation of interleukin (IL)-6 and IL-10 production. Thus, during an immune/inflammatory response NPY may exert proinflammatory effects through the recruitment of immature DCs, but it may exert antiinflammatory effects by promoting a Th2 polarization. Locally, at inflammatory sites, cell recruitment could be amplified in conditions of intense acute, chronic, or cold stress. Thus, altered or amplified signaling through the NPY-NPY-Y1 receptor-DC axis may have implications for the development of inflammatory conditions.-Buttari, B., Profumo, E., Domenici, G., Tagliani, A., Ippoliti, F., Bonini, S., Businaro, R., Elenkov, I., Riganò, R. Neuropeptide Y induces potent migration of human immature dendritic cells and promotes a Th2 polarization. © FASEB.

  4. Delivery of TLR7 agonist to monocytes and dendritic cells by DCIR targeted liposomes induces robust production of anti-cancer cytokines

    DEFF Research Database (Denmark)

    Klauber, Thomas Christopher Bogh; Laursen, Janne Marie; Zucker, Daniel

    2017-01-01

    Tumor immune escape is today recognized as an important cancer hallmark and is therefore a major focus area in cancer therapy. Monocytes and dendritic cells (DCs), which are central to creating a robust anti-tumor immune response and establishing an anti-tumorigenic microenvironment, are directly...... targeted by the tumor escape mechanisms to develop immunosuppressive phenotypes. Providing activated monocytes and DCs to the tumor tissue is therefore an attractive way to break the tumor-derived immune suppression and reinstate cancer immune surveillance. To activate monocytes and DCs with high...... as their immune activating potential in blood-derived monocytes, myeloid DCs (mDCs), and plasmacytoid DCs (pDCs). Monocytes and mDCs were targeted with high specificity over lymphocytes, and exhibited potent TLR7-specific secretion of the anti-cancer cytokines IL-12p70, IFN-α 2a, and IFN-γ. This delivery system...

  5. Characteristics of meniscus progenitor cells migrated from injured meniscus.

    Science.gov (United States)

    Seol, Dongrim; Zhou, Cheng; Brouillette, Marc J; Song, Ino; Yu, Yin; Choe, Hyeong Hun; Lehman, Abigail D; Jang, Kee W; Fredericks, Douglas C; Laughlin, Barbara J; Martin, James A

    2017-09-01

    Serious meniscus injuries seldom heal and increase the risk for knee osteoarthritis; thus, there is a need to develop new reparative therapies. In that regard, stimulating tissue regeneration by autologous stem/progenitor cells has emerged as a promising new strategy. We showed previously that migratory chondrogenic progenitor cells (CPCs) were recruited to injured cartilage, where they showed a capability in situ tissue repair. Here, we tested the hypothesis that the meniscus contains a similar population of regenerative cells. Explant studies revealed that migrating cells were mainly confined to the red zone in normal menisci: However, these cells were capable of repopulating defects made in the white zone. In vivo, migrating cell numbers increased dramatically in damaged meniscus. Relative to non-migrating meniscus cells, migrating cells were more clonogenic, overexpressed progenitor cell markers, and included a larger side population. Gene expression profiling showed that the migrating population was more similar to CPCs than other meniscus cells. Finally, migrating cells equaled CPCs in chondrogenic potential, indicating a capacity for repair of the cartilaginous white zone of the meniscus. These findings demonstrate that, much as in articular cartilage, injuries to the meniscus mobilize an intrinsic progenitor cell population with strong reparative potential. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1966-1972, 2017. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  6. Cell Migration in 1D and 2D Nanofiber Microenvironments.

    Science.gov (United States)

    Estabridis, Horacio M; Jana, Aniket; Nain, Amrinder; Odde, David J

    2018-03-01

    Understanding how cells migrate in fibrous environments is important in wound healing, immune function, and cancer progression. A key question is how fiber orientation and network geometry influence cell movement. Here we describe a quantitative, modeling-based approach toward identifying the mechanisms by which cells migrate in fibrous geometries having well controlled orientation. Specifically, U251 glioblastoma cells were seeded onto non-electrospinning Spinneret based tunable engineering parameters fiber substrates that consist of networks of suspended 400 nm diameter nanofibers. Cells were classified based on the local fiber geometry and cell migration dynamics observed by light microscopy. Cells were found in three distinct geometries: adhering two a single fiber, adhering to two parallel fibers, and adhering to a network of orthogonal fibers. Cells adhering to a single fiber or two parallel fibers can only move in one dimension along the fiber axis, whereas cells on a network of orthogonal fibers can move in two dimensions. We found that cells move faster and more persistently in 1D geometries than in 2D, with cell migration being faster on parallel fibers than on single fibers. To explain these behaviors mechanistically, we simulated cell migration in the three different geometries using a motor-clutch based model for cell traction forces. Using nearly identical parameter sets for each of the three cases, we found that the simulated cells naturally replicated the reduced migration in 2D relative to 1D geometries. In addition, the modestly faster 1D migration on parallel fibers relative to single fibers was captured using a correspondingly modest increase in the number of clutches to reflect increased surface area of adhesion on parallel fibers. Overall, the integrated modeling and experimental analysis shows that cell migration in response to varying fibrous geometries can be explained by a simple mechanical readout of geometry via a motor-clutch mechanism.

  7. Silk Film Topography Directs Collective Epithelial Cell Migration

    Science.gov (United States)

    Rosenblatt, Mark I.

    2012-01-01

    The following study provides new insight into how surface topography dictates directed collective epithelial cell sheet growth through the guidance of individual cell movement. Collective cell behavior of migrating human corneal limbal-epithelial cell sheets were studied on highly biocompatible flat and micro-patterned silk film surfaces. The silk film edge topography guided the migratory direction of individual cells making up the collective epithelial sheet, which resulted in a 75% increase in total culture elongation. This was due to a 3-fold decrease in cell sheet migration rate efficiency for movement perpendicular to the topography edge. Individual cell migration direction is preferred in the parallel approach to the edge topography where localization of cytoskeletal proteins to the topography’s edge region is reduced, which results in the directed growth of the collective epithelial sheet. Findings indicate customized biomaterial surfaces may be created to direct both the migration rate and direction of tissue epithelialization. PMID:23185573

  8. Sialoadhesin expressed on IFN-induced monocytes binds HIV-1 and enhances infectivity.

    Directory of Open Access Journals (Sweden)

    Hans Rempel

    2008-04-01

    Full Text Available HIV-1 infection dysregulates the immune system and alters gene expression in circulating monocytes. Differential gene expression analysis of CD14(+ monocytes from subjects infected with HIV-1 revealed increased expression of sialoadhesin (Sn, CD169, Siglec 1, a cell adhesion molecule first described in a subset of macrophages activated in chronic inflammatory diseases.We analyzed sialoadhesin expression on CD14(+ monocytes by flow cytometry and found significantly higher expression in subjects with elevated viral loads compared to subjects with undetectable viral loads. In cultured CD14(+ monocytes isolated from healthy individuals, sialoadhesin expression was induced by interferon-alpha and interferon-gamma but not tumor necrosis factor-alpha. Using a stringent binding assay, sialoadhesin-expressing monocytes adsorbed HIV-1 through interaction with the sialic acid residues on the viral envelope glycoprotein gp120. Furthermore, monocytes expressing sialoadhesin facilitated HIV-1 trans infection of permissive cells, which occurred in the absence of monocyte self-infection.Increased sialoadhesin expression on CD14(+ monocytes occurred in response to HIV-1 infection with maximum expression associated with high viral load. We show that interferons induce sialoadhesin in primary CD14(+ monocytes, which is consistent with an antiviral response during viremia. Our findings suggest that circulating sialoadhesin-expressing monocytes are capable of binding HIV-1 and effectively delivering virus to target cells thereby enhancing the distribution of HIV-1. Sialoadhesin could disseminate HIV-1 to viral reservoirs during monocyte immunosurveillance or migration to sites of inflammation and then facilitate HIV-1 infection of permissive cells.

  9. Cooperation between Monocyte-Derived Cells and Lymphoid Cells in the Acute Response to a Bacterial Lung Pathogen.

    Directory of Open Access Journals (Sweden)

    Andrew S Brown

    2016-06-01

    Full Text Available Legionella pneumophila is the causative agent of Legionnaires' disease, a potentially fatal lung infection. Alveolar macrophages support intracellular replication of L. pneumophila, however the contributions of other immune cell types to bacterial killing during infection are unclear. Here, we used recently described methods to characterise the major inflammatory cells in lung after acute respiratory infection of mice with L. pneumophila. We observed that the numbers of alveolar macrophages rapidly decreased after infection coincident with a rapid infiltration of the lung by monocyte-derived cells (MC, which, together with neutrophils, became the dominant inflammatory cells associated with the bacteria. Using mice in which the ability of MC to infiltrate tissues is impaired it was found that MC were required for bacterial clearance and were the major source of IL12. IL12 was needed to induce IFNγ production by lymphoid cells including NK cells, memory T cells, NKT cells and γδ T cells. Memory T cells that produced IFNγ appeared to be circulating effector/memory T cells that infiltrated the lung after infection. IFNγ production by memory T cells was stimulated in an antigen-independent fashion and could effectively clear bacteria from the lung indicating that memory T cells are an important contributor to innate bacterial defence. We also determined that a major function of IFNγ was to stimulate bactericidal activity of MC. On the other hand, neutrophils did not require IFNγ to kill bacteria and alveolar macrophages remained poorly bactericidal even in the presence of IFNγ. This work has revealed a cooperative innate immune circuit between lymphoid cells and MC that combats acute L. pneumophila infection and defines a specific role for IFNγ in anti-bacterial immunity.

  10. Microbial products activate monocytic cells through detergent-resistant membrane microdomains.

    Science.gov (United States)

    Epelman, Slava; Berenger, Byron; Stack, Danuta; Neely, Graham G; Ma, Ling Ling; Mody, Christopher H

    2008-12-01

    Patients with cystic fibrosis suffer recurrent pulmonary infections that are characterized by an overactive yet ineffective and destructive inflammatory response that is associated with respiratory infections by Pseudomonas aeruginosa, a pathogen that produces a number of phlogistic molecules. To better understand this process, we used exoenzyme S (ExoS), one of the key P. aeruginosa-secreted exoproducts, which is known to stimulate cells via the Toll-like receptor (TLR) pathway. We found that ExoS induced proinflammatory cytokine production via the NF-kappaB, Erk1/2, and Src kinase pathways. Because Src kinases are concentrated within cholesterol-containing, detergent-resistant membrane microdomains (DRM) (also called lipid rafts) and DRM act as signaling platforms and amplifiers on the surface of cells, we addressed the role of DRM in ExoS signaling. ExoS bound directly to a subset of DRM and induced the phosphorylation of multiple proteins within DRM, including Src kinases. Disruption of DRM by cholesterol extraction prevented NF-kappaB and Erk 1/2 activation and TNF-alpha production in response to ExoS. Activation of monocytic cells by other TLR and Nod-like receptor agonists, such as lipoteichoic acid, lipopolysaccharide, and peptidoglycan, were also dependent on DRM, and disruption prevented TNF-alpha production. Disruption of DRM did not prevent ExoS binding but did release the Src kinase, Lyn, from the DRM fraction into the detergent-soluble fraction, a site in which Src kinases are not active. These studies show that ExoS, a TLR agonist, requires direct binding to DRM for optimal signaling, which suggests that DRM are possible therapeutic targets in cystic fibrosis.

  11. Collective cell migration drives morphogenesis of the kidney nephron.

    Directory of Open Access Journals (Sweden)

    Aleksandr Vasilyev

    2009-01-01

    Full Text Available Tissue organization in epithelial organs is achieved during development by the combined processes of cell differentiation and morphogenetic cell movements. In the kidney, the nephron is the functional organ unit. Each nephron is an epithelial tubule that is subdivided into discrete segments with specific transport functions. Little is known about how nephron segments are defined or how segments acquire their distinctive morphology and cell shape. Using live, in vivo cell imaging of the forming zebrafish pronephric nephron, we found that the migration of fully differentiated epithelial cells accounts for both the final position of nephron segment boundaries and the characteristic convolution of the proximal tubule. Pronephric cells maintain adherens junctions and polarized apical brush border membranes while they migrate collectively. Individual tubule cells exhibit basal membrane protrusions in the direction of movement and appear to establish transient, phosphorylated Focal Adhesion Kinase-positive adhesions to the basement membrane. Cell migration continued in the presence of camptothecin, indicating that cell division does not drive migration. Lengthening of the nephron was, however, accompanied by an increase in tubule cell number, specifically in the most distal, ret1-positive nephron segment. The initiation of cell migration coincided with the onset of fluid flow in the pronephros. Complete blockade of pronephric fluid flow prevented cell migration and proximal nephron convolution. Selective blockade of proximal, filtration-driven fluid flow shifted the position of tubule convolution distally and revealed a role for cilia-driven fluid flow in persistent migration of distal nephron cells. We conclude that nephron morphogenesis is driven by fluid flow-dependent, collective epithelial cell migration within the confines of the tubule basement membrane. Our results establish intimate links between nephron function, fluid flow, and morphogenesis.

  12. Signaling through three chemokine receptors triggers the migration of transplanted neural precursor cells in a model of multiple sclerosis.

    Science.gov (United States)

    Cohen, Mikhal E; Fainstein, Nina; Lavon, Iris; Ben-Hur, Tamir

    2014-09-01

    Multiple sclerosis (MS) is a multifocal disease, and precursor cells need to migrate into the multiple lesions in order to exert their therapeutic effects. Therefore, cell migration is a crucial element in regenerative processes in MS, dictating the route of delivery, when cell transplantation is considered. We have previously shown that inflammation triggers migration of multi-potential neural precursor cells (NPCs) into the white matter of experimental autoimmune encephalomyelitis (EAE) rodents, a widely used model of MS. Here we investigated the molecular basis of this attraction. NPCs were grown from E13 embryonic mouse brains and transplanted into the lateral cerebral ventricles of EAE mice. Transplanted NPC migration was directed by three tissue-derived chemokines. Stromal cell-derived factor-1α, monocyte chemo-attractant protein-1 and hepatocyte growth factor were expressed in the EAE brain and specifically in microglia and astrocytes. Their cognate receptors, CXCR4, CCR2 or c-Met were constitutively expressed on NPCs. Selective blockage of CXCR4, CCR2 or c-Met partially inhibited NPC migration in EAE brains. Blocking all three receptors had an additive effect and resulted in profound inhibition of NPC migration, as compared to extensive migration of control NPCs. The inflammation-triggered NPC migration into white matter tracts was dependent on a motile NPC phenotype. Specifically, depriving NPCs from epidermal growth factor (EGF) prevented the induction of glial commitment and a motile phenotype (as indicated by an in vitro motility assay), hampering their response to neuroinflammation. In conclusion, signaling via three chemokine systems accounts for most of the inflammation-induced, tissue-derived attraction of transplanted NPCs into white matter tracts during EAE. Copyright © 2014. Published by Elsevier B.V.

  13. Equine Herpesvirus Type 1 Enhances Viral Replication in CD172a+ Monocytic Cells upon Adhesion to Endothelial Cells.

    Science.gov (United States)

    Laval, Kathlyn; Favoreel, Herman W; Poelaert, Katrien C K; Van Cleemput, Jolien; Nauwynck, Hans J

    2015-11-01

    Equine herpesvirus type 1 (EHV-1) is a main cause of respiratory disease, abortion, and encephalomyelopathy in horses. Monocytic cells (CD172a(+)) are the main carrier cells of EHV-1 during primary infection and are proposed to serve as a "Trojan horse" to facilitate the dissemination of EHV-1 to target organs. However, the mechanism by which EHV-1 is transferred from CD172a(+) cells to endothelial cells (EC) remains unclear. The aim of this study was to investigate EHV-1 transmission between these two cell types. We hypothesized that EHV-1 employs specific strategies to promote the adhesion of infected CD172a(+) cells to EC to facilitate EHV-1 spread. Here, we demonstrated that EHV-1 infection of CD172a(+) cells resulted in a 3- to 5-fold increase in adhesion to EC. Antibody blocking experiments indicated that α4β1, αLβ2, and αVβ3 integrins mediated adhesion of infected CD172a(+) cells to EC. We showed that integrin-mediated phosphatidylinositol 3-kinase (PI3K) and ERK/MAPK signaling pathways were involved in EHV-1-induced CD172a(+) cell adhesion at early times of infection. EHV-1 replication was enhanced in adherent CD172a(+) cells, which correlates with the production of tumor necrosis factor alpha (TNF-α). In the presence of neutralizing antibodies, approximately 20% of infected CD172a(+) cells transferred cytoplasmic material to uninfected EC and 0.01% of infected CD172a(+) cells transmitted infectious virus to neighboring cells. Our results demonstrated that EHV-1 infection induces adhesion of CD172a(+) cells to EC, which enhances viral replication, but that transfer of viral material from CD172a(+) cells to EC is a very specific and rare event. These findings give new insights into the complex pathogenesis of EHV-1. Equine herpesvirus type 1 (EHV-1) is a highly prevalent pathogen worldwide, causing frequent outbreaks of abortion and myeloencephalopathy, even in vaccinated horses. After primary replication in the respiratory tract, EHV-1 disseminates

  14. Effect of β-agonist on the dexamethasone-induced expression of aromatase by the human monocyte cells

    Directory of Open Access Journals (Sweden)

    Masatada Watanabe

    2017-02-01

    Full Text Available Emerging evidence suggests that sex steroids are important for human skin health. In particular, estrogen improves skin thickness, elasticity and moisture of older women. The major source of circulating estrogen is the ovary; however, local estrogen synthesis and secretion have important roles in, for example, bone metabolism and breast cancer development. We hypothesized that infiltrated peripheral monocytes are one of the sources of estrogen in skin tissues. We also hypothesized that, during atopic dermatitis under stress, a decline in the hypothalamus–pituitary–adrenal axis (HPA and facilitation of the (hypothalamus–sympathetic–adrenomedullary system (SAM attenuates estrogen secretion from monocytes. Based on this hypothesis, we tested aromatase expression in the human peripheral monocyte-derived cell line THP-1 in response to the synthetic glucocorticoid dexamethasone (Dex, the synthetic β-agonist isoproterenol (Iso and the β-antagonist propranolol (Pro. Dex mimics glucocorticoid secreted during excitation of the HPA, and Iso mimics catecholamine secreted during excitation of the SAM. We found that aromatase activity and the CYP19A1 gene transcript were both upregulated in THP-1 cells in the presence of Dex. Addition of Iso induced their downregulation and further addition of Pro rescued aromatase expression. These results may suggest that attenuation of estrogen secretion from peripheral monocytes could be a part of the pathology of stress-caused deterioration of atopic dermatitis. Further examination using an in vitro human skin model including THP-1 cells might be a valuable tool for investigating the therapeutic efficacy and mechanism of estrogen treatment for skin health.

  15. Angiopoietin 2 stimulates TIE2-expressing monocytes to suppress T cell activation and to promote regulatory T cell expansion.

    Science.gov (United States)

    Coffelt, Seth B; Chen, Yung-Yi; Muthana, Munitta; Welford, Abigail F; Tal, Andrea O; Scholz, Alexander; Plate, Karl H; Reiss, Yvonne; Murdoch, Craig; De Palma, Michele; Lewis, Claire E

    2011-04-01

    Angiopoietin 2 (ANGPT2) is a proangiogenic cytokine whose expression is often upregulated by endothelial cells in tumors. Expression of its receptor, TIE2, defines a highly proangiogenic subpopulation of myeloid cells in circulation and tumors called TIE2-expressing monocytes/macrophages (TEMs). Genetic depletion of TEMs markedly reduces tumor angiogenesis in various tumor models, emphasizing their essential role in driving tumor progression. Previously, we demonstrated that ANGPT2 augments the expression of various proangiogenic genes, the potent immunosuppressive cytokine, IL-10, and a chemokine for regulatory T cells (Tregs), CCL17 by TEMs in vitro. We now show that TEMs also express higher levels of IL-10 than TIE2(-) macrophages in tumors and that ANGPT2-stimulated release of IL-10 by TEMs suppresses T cell proliferation, increases the ratio of CD4(+) T cells to CD8(+) T cells, and promotes the expansion of CD4(+)CD25(high)FOXP3(+) Tregs. Furthermore, syngeneic murine tumors expressing high levels of ANGPT2 contained not only high numbers of TEMs but also increased numbers of Tregs, whereas genetic depletion of tumor TEMs resulted in a marked reduction in the frequency of Tregs in tumors. Taken together, our data suggest that ANGPT2-stimulated TEMs represent a novel, potent immunosuppressive force in tumors.

  16. Infection Rate and Tissue Localization of Murine IL-12p40-Producing Monocyte-Derived CD103+ Lung Dendritic Cells during Pulmonary Tuberculosis

    Science.gov (United States)

    Leepiyasakulchai, Chaniya; Taher, Chato; Chuquimia, Olga D.; Mazurek, Jolanta; Söderberg-Naucler, Cecilia; Fernández, Carmen; Sköld, Markus

    2013-01-01

    Non-hematopoietic cells, including lung epithelial cells, influence host immune responses. By co-culturing primary alveolar epithelial cells and monocytes from naïve donor mice, we show that alveolar epithelial cells support monocyte survival and differentiation in vitro, suggesting a role for non-hematopoietic cells in monocyte differentiation during the steady state in vivo. CD103+ dendritic cells (αE-DC) are present at mucosal surfaces. Using a murine primary monocyte adoptive transfer model, we demonstrate that αE-DC in the lungs and pulmonary lymph nodes are monocyte-derived during pulmonary tuberculosis. The tissue localization may influence the functional potential of αE-DC that accumulate in Mycobacterium tuberculosis-infected lungs. Here, we confirm the localization of αE-DC in uninfected mice beneath the bronchial epithelial cell layer and near the vascular wall, and show that αE-DC have a similar distribution in the lungs during pulmonary tuberculosis and are detected in the bronchoalveolar lavage fluid from infected mice. Lung DC can be targeted by M. tuberculosis in vivo and play a role in bacterial dissemination to the draining lymph node. In contrast to other DC subsets, only a fraction of lung αE-DC are infected with the bacterium. We also show that virulent M. tuberculosis does not significantly alter cell surface expression levels of MHC class II on infected cells in vivo and that αE-DC contain the highest frequency of IL-12p40+ cells among the myeloid cell subsets in infected lungs. Our results support a model in which inflammatory monocytes are recruited into the M. tuberculosis-infected lung tissue and, depending on which non-hematopoietic cells they interact with, differentiate along different paths to give rise to multiple monocyte-derived cells, including DC with a distinctive αE-DC phenotype. PMID:23861965

  17. Infection rate and tissue localization of murine IL-12p40-producing monocyte-derived CD103(+) lung dendritic cells during pulmonary tuberculosis.

    Science.gov (United States)

    Leepiyasakulchai, Chaniya; Taher, Chato; Chuquimia, Olga D; Mazurek, Jolanta; Söderberg-Naucler, Cecilia; Fernández, Carmen; Sköld, Markus

    2013-01-01

    Non-hematopoietic cells, including lung epithelial cells, influence host immune responses. By co-culturing primary alveolar epithelial cells and monocytes from naïve donor mice, we show that alveolar epithelial cells support monocyte survival and differentiation in vitro, suggesting a role for non-hematopoietic cells in monocyte differentiation during the steady state in vivo. CD103(+) dendritic cells (αE-DC) are present at mucosal surfaces. Using a murine primary monocyte adoptive transfer model, we demonstrate that αE-DC in the lungs and pulmonary lymph nodes are monocyte-derived during pulmonary tuberculosis. The tissue localization may influence the functional potential of αE-DC that accumulate in Mycobacterium tuberculosis-infected lungs. Here, we confirm the localization of αE-DC in uninfected mice beneath the bronchial epithelial cell layer and near the vascular wall, and show that αE-DC have a similar distribution in the lungs during pulmonary tuberculosis and are detected in the bronchoalveolar lavage fluid from infected mice. Lung DC can be targeted by M. tuberculosis in vivo and play a role in bacterial dissemination to the draining lymph node. In contrast to other DC subsets, only a fraction of lung αE-DC are infected with the bacterium. We also show that virulent M. tuberculosis does not significantly alter cell surface expression levels of MHC class II on infected cells in vivo and that αE-DC contain the highest frequency of IL-12p40(+) cells among the myeloid cell subsets in infected lungs. Our results support a model in which inflammatory monocytes are recruited into the M. tuberculosis-infected lung tissue and, depending on which non-hematopoietic cells they interact with, differentiate along different paths to give rise to multiple monocyte-derived cells, including DC with a distinctive αE-DC phenotype.

  18. Protein kinase Cepsilon is important for migration of neuroblastoma cells

    International Nuclear Information System (INIS)

    Stensman, Helena; Larsson, Christer

    2008-01-01

    Migration is important for the metastatic capacity and thus for the malignancy of cancer cells. There is limited knowledge on regulatory factors that promote the migration of neuroblastoma cells. This study investigates the hypothesis that protein kinase C (PKC) isoforms regulate neuroblastoma cell motility. PKC isoforms were downregulated with siRNA or modulated with activators and inhibitors. Migration was analyzed with scratch and transwell assays. Protein phosphorylation and expression levels were measured with Western blot. Stimulation with 12-O-tetradecanoylphorbol-13-acetate (TPA) induced migration of SK-N-BE(2)C neuroblastoma cells. Treatment with the general protein kinase C (PKC) inhibitor GF109203X and the inhibitor of classical isoforms Gö6976 inhibited migration while an inhibitor of PKCβ isoforms did not have an effect. Downregulation of PKCε, but not of PKCα or PKCδ, with siRNA led to a suppression of both basal and TPA-stimulated migration. Experiments using PD98059 and LY294002, inhibitors of the Erk and phosphatidylinositol 3-kinase (PI3K) pathways, respectively, showed that PI3K is not necessary for TPA-induced migration. The Erk pathway might be involved in TPA-induced migration but not in migration driven by PKCε. TPA induced phosphorylation of the PKC substrate myristoylated alanine-rich C kinase substrate (MARCKS) which was suppressed by the PKC inhibitors. Treatment with siRNA oligonucleotides against different PKC isoforms before stimulation with TPA did not influence the phosphorylation of MARCKS. PKCε is important for migration of SK-N-BE(2)C neuroblastoma cells. Neither the Erk pathway nor MARCKS are critical downstream targets of PKCε but they may be involved in TPA-mediated migration

  19. Using Single-Protein Tracking to Study Cell Migration.

    Science.gov (United States)

    Orré, Thomas; Mehidi, Amine; Massou, Sophie; Rossier, Olivier; Giannone, Grégory

    2018-01-01

    To get a complete understanding of cell migration, it is critical to study its orchestration at the molecular level. Since the recent developments in single-molecule imaging, it is now possible to study molecular phenomena at the single-molecule level inside living cells. In this chapter, we describe how such approaches have been and can be used to decipher molecular mechanisms involved in cell migration.

  20. Insulin promotes cell migration by regulating PSA-NCAM

    Energy Technology Data Exchange (ETDEWEB)

    Monzo, Hector J.; Coppieters, Natacha [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Park, Thomas I.H. [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Dieriks, Birger V.; Faull, Richard L.M. [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Dragunow, Mike [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Curtis, Maurice A., E-mail: m.curtis@auckland.ac.nz [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand)

    2017-06-01

    Cellular interactions with the extracellular environment are modulated by cell surface polysialic acid (PSA) carried by the neural cell adhesion molecule (NCAM). PSA-NCAM is involved in cellular processes such as differentiation, plasticity, and migration, and is elevated in Alzheimer's disease as well as in metastatic tumour cells. Our previous work demonstrated that insulin enhances the abundance of cell surface PSA by inhibiting PSA-NCAM endocytosis. In the present study we have identified a mechanism for insulin-dependent inhibition of PSA-NCAM turnover affecting cell migration. Insulin enhanced the phosphorylation of the focal adhesion kinase leading to dissociation of αv-integrin/PSA-NCAM clusters, and promoted cell migration. Our results show that αv-integrin plays a key role in the PSA-NCAM turnover process. αv-integrin knockdown stopped PSA-NCAM from being endocytosed, and αv-integrin/PSA-NCAM clusters co-labelled intracellularly with Rab5, altogether indicating a role for αv-integrin as a carrier for PSA-NCAM during internalisation. Furthermore, inhibition of p-FAK caused dissociation of αv-integrin/PSA-NCAM clusters and counteracted the insulin-induced accumulation of PSA at the cell surface and cell migration was impaired. Our data reveal a functional association between the insulin/p-FAK-dependent regulation of PSA-NCAM turnover and cell migration through the extracellular matrix. Most importantly, they identify a novel mechanism for insulin-stimulated cell migration. - Highlights: • Insulin modulates PSA-NCAM turnover through upregulation of p-FAK. • P-FAK modulates αv-integrin/PSA-NCAM clustering. • αv-integrin acts as a carrier for PSA-NCAM endocytosis. • Cell migration is promoted by cell surface PSA. • Insulin promotes PSA-dependent migration in vitro.

  1. Insulin promotes cell migration by regulating PSA-NCAM

    International Nuclear Information System (INIS)

    Monzo, Hector J.; Coppieters, Natacha; Park, Thomas I.H.; Dieriks, Birger V.; Faull, Richard L.M.; Dragunow, Mike; Curtis, Maurice A.

    2017-01-01

    Cellular interactions with the extracellular environment are modulated by cell surface polysialic acid (PSA) carried by the neural cell adhesion molecule (NCAM). PSA-NCAM is involved in cellular processes such as differentiation, plasticity, and migration, and is elevated in Alzheimer's disease as well as in metastatic tumour cells. Our previous work demonstrated that insulin enhances the abundance of cell surface PSA by inhibiting PSA-NCAM endocytosis. In the present study we have identified a mechanism for insulin-dependent inhibition of PSA-NCAM turnover affecting cell migration. Insulin enhanced the phosphorylation of the focal adhesion kinase leading to dissociation of αv-integrin/PSA-NCAM clusters, and promoted cell migration. Our results show that αv-integrin plays a key role in the PSA-NCAM turnover process. αv-integrin knockdown stopped PSA-NCAM from being endocytosed, and αv-integrin/PSA-NCAM clusters co-labelled intracellularly with Rab5, altogether indicating a role for αv-integrin as a carrier for PSA-NCAM during internalisation. Furthermore, inhibition of p-FAK caused dissociation of αv-integrin/PSA-NCAM clusters and counteracted the insulin-induced accumulation of PSA at the cell surface and cell migration was impaired. Our data reveal a functional association between the insulin/p-FAK-dependent regulation of PSA-NCAM turnover and cell migration through the extracellular matrix. Most importantly, they identify a novel mechanism for insulin-stimulated cell migration. - Highlights: • Insulin modulates PSA-NCAM turnover through upregulation of p-FAK. • P-FAK modulates αv-integrin/PSA-NCAM clustering. • αv-integrin acts as a carrier for PSA-NCAM endocytosis. • Cell migration is promoted by cell surface PSA. • Insulin promotes PSA-dependent migration in vitro.

  2. Skin-resident CD4+ T cells protect against Leishmania major by recruiting and activating inflammatory monocytes

    Science.gov (United States)

    Glennie, Nelson D.; Volk, Susan W.

    2017-01-01

    Tissue-resident memory T cells are required for establishing protective immunity against a variety of different pathogens, although the mechanisms mediating protection by CD4+ resident memory T cells are still being defined. In this study we addressed this issue with a population of protective skin-resident, IFNγ-producing CD4+ memory T cells generated following Leishmania major infection. We previously found that resident memory T cells recruit circulating effector T cells to enhance immunity. Here we show that resident memory CD4+ T cells mediate the delayed-hypersensitivity response observed in immune mice and provide protection without circulating T cells. This protection occurs rapidly after challenge, and requires the recruitment and activation of inflammatory monocytes, which limit parasites by production of both reactive oxygen species and nitric oxide. Overall, these data highlight a novel role for tissue-resident memory cells in recruiting and activating inflammatory monocytes, and underscore the central role that skin-resident T cells play in immunity to cutaneous leishmaniasis. PMID:28419151

  3. [Linezolid-induced Apoptosis through Mitochondrial Damage and Role of Superoxide Dismutase-1 in Human Monocytic Cell Line U937].

    Science.gov (United States)

    Fujii, Satoshi; Muraoka, Sanae; Miyamoto, Atsushi; Sakurai, Koichi

    2018-01-01

     Cytopenia is a major adverse event associated with linezolid therapy. The objective of this study was to examine whether the cytotoxicity of linezolid to eukaryotic cells was associated with mitochondrial dysfunction and apoptosis-like cell death in human leukemic monocyte lymphoma cell line U937. Apoptosis-like cell death was clearly observed when cells were incubated with linezolid, depending on the duration and linezolid concentration. Mitochondrial membrane potential of cells treated with linezolid collapsed in a short period of time, but the number of mitochondria did not decrease. Cytotoxicity of linezolid was relieved by the knockdown of superoxide dismutase-1 in U937 cells. On the other hand, no autophagy was observed in cells treated with linezolid. These results suggest that mitochondrial damages would be linked to the induction of apoptosis in U937 cells treated with linezolid and that its mechanism does not involve autophagy.

  4. Src Induces Podoplanin Expression to Promote Cell Migration*

    Science.gov (United States)

    Shen, Yongquan; Chen, Chen-Shan; Ichikawa, Hitoshi; Goldberg, Gary S.

    2010-01-01

    Nontransformed cells can force tumor cells to assume a normal morphology and phenotype by the process of contact normalization. Transformed cells must escape this process to become invasive and malignant. However, mechanisms underlying contact normalization have not been elucidated. Here, we have identified genes that are affected by contact normalization of Src-transformed cells. Tumor cells must migrate to become invasive and malignant. Src must phosphorylate the adaptor protein Cas (Crk-associated substrate) to promote tumor cell motility. We report here that Src utilizes Cas to induce podoplanin (Pdpn) expression to promote tumor cell migration. Pdpn is a membrane-bound extracellular glycoprotein that associates with endogenous ligands to promote tumor cell migration leading to cancer invasion and metastasis. In fact, Pdpn expression accounted for a major part of the increased migration seen in Src-transformed cells. Moreover, nontransformed cells suppressed Pdpn expression in adjacent Src-transformed cells. Of >39,000 genes, Pdpn was one of only 23 genes found to be induced by transforming Src activity and suppressed by contact normalization of Src-transformed cells. In addition, we found 16 genes suppressed by Src and induced by contact normalization. These genes encode growth factor receptors, adaptor proteins, and products that have not yet been annotated and may play important roles in tumor cell growth and migration. PMID:20123990

  5. Bone marrow chimeric mice reveal a role for CX₃CR1 in maintenance of the monocyte-derived cell population in the olfactory neuroepithelium.

    Science.gov (United States)

    Vukovic, Jana; Blomster, Linda V; Chinnery, Holly R; Weninger, Wolfgang; Jung, Steffen; McMenamin, Paul G; Ruitenberg, Marc J

    2010-10-01

    Macrophages in the olfactory neuroepithelium are thought to play major roles in tissue homeostasis and repair. However, little information is available at present about possible heterogeneity of these monocyte-derived cells, their turnover rates, and the role of chemokine receptors in this process. To start addressing these issues, this study used Cx₃cr1(gfp) mice, in which the gene sequence for eGFP was knocked into the CX₃CR1 gene locus in the mutant allele. Using neuroepithelial whole-mounts from Cx₃cr1(gfp/+) mice, we show that eGFP(+) cells of monocytic origin are distributed in a loose network throughout this tissue and can be subdivided further into two immunophenotypically distinct subsets based on MHC-II glycoprotein expression. BM chimeric mice were created using Cx₃cr1(gfp/+) donors to investigate turnover of macrophages (and other monocyte-derived cells) in the olfactory neuroepithelium. Our data indicate that the monocyte-derived cell population in the olfactory neuroepithelium is actively replenished by circulating monocytes and under the experimental conditions, completely turned over within 6 months. Transplantation of Cx₃cr1(gfp/gfp) (i.e., CX₃CR1-deficient) BM partially impaired the replenishment process and resulted in an overall decline of the total monocyte-derived cell number in the olfactory epithelium. Interestingly, replenishment of the CD68(low)MHC-II(+) subset appeared minimally affected by CX₃CR1 deficiency. Taken together, the established baseline data about heterogeneity of monocyte-derived cells, their replenishment rates, and the role of CX₃CR1 provide a solid basis to further examine the importance of different monocyte subsets for neuroregeneration at this unique frontier with the external environment.

  6. Human platelet lysate is a successful alternative serum supplement for propagation of monocyte-derived dendritic cells.

    Science.gov (United States)

    Švajger, Urban

    2017-04-01

    Clinical protocols for dendritic cell (DC) generation from monocytes require the use of animal serum-free supplements. Serum-free media can also require up to 1% of serum supplementation. In addition, recommendations based on the 3Rs (Refinement, Reduction, Replacement) principle also recommend the use of non-animal sera in in vitro studies. The aim of this study was to explore the potential use of platelet lysate (PL) for generation of optimally differentiated DCs from monocytes. Cells were isolated from buffy coats from healthy volunteers using immunomagnetic selection. DCs were differentiated in RPMI1640 supplemented with either 10% fetal bovine serum (FBS), 10% AB serum or 10% PL with the addition of granulocyte monocyte colony stimulating factor and interleukin-4. Generated DCs were assessed for their morphology, viability, endocytotic capacity, surface phenotype (immature, mature and tolerogenic DCs) and activation of important signaling pathways. DC function was evaluated on the basis of their allostimulatory capacity, cytokine profile and ability to induce different T-helper subsets. DCs generated with PL displayed normal viability, morphology and endocytotic capacity. Their differentiation and maturation phenotype was comparable to FBS-cultured DCs. They showed functional plasticity and up-regulated tolerogenic markers in response to their environment. PL-cultured mature DCs displayed unhindered allostimulatory potential and the capacity to induce Th1 responses. The use of PL allowed for activation of crucial signaling proteins associated with DC differentiation and maturation. This study demonstrates for the first time that human PL represents a successful alternative to FBS in differentiation of DCs from monocytes. DCs display the major phenotypic and functional characteristics compared with existing culture protocols. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  7. A pilgrim's progress: Seeking meaning in primordial germ cell migration.

    Science.gov (United States)

    Cantú, Andrea V; Laird, Diana J

    2017-10-01

    Comparative studies of primordial germ cell (PGC) development across organisms in many phyla reveal surprising diversity in the route of migration, timing and underlying molecular mechanisms, suggesting that the process of migration itself is conserved. However, beyond the perfunctory transport of cellular precursors to their later arising home of the gonads, does PGC migration serve a function? Here we propose that the process of migration plays an additional role in quality control, by eliminating PGCs incapable of completing migration as well as through mechanisms that favor PGCs capable of responding appropriately to migration cues. Focusing on PGCs in mice, we explore evidence for a selective capacity of migration, considering the tandem regulation of proliferation and migration, cell-intrinsic and extrinsic control, the potential for tumors derived from failed PGC migrants, the potential mechanisms by which migratory PGCs vary in their cellular behaviors, and corresponding effects on development. We discuss the implications of a selective role of PGC migration for in vitro gametogenesis. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  8. Macrophage-derived microvesicles promote proliferation and migration of Schwann cell on peripheral nerve repair

    Energy Technology Data Exchange (ETDEWEB)

    Zhan, Chuan, E-mail: zhchuansy@163.com; Ma, Cheng-bin; Yuan, Hong-mou; Cao, Bao-yuan; Zhu, Jia-jun

    2015-12-04

    Background: Macrophages have been implicated in peripheral nerve regeneration. However, whether macrophages-derived microvesicles (MVs) are involved in this process remains unknown. In the present study, the effects of macrophages-derived MVs on proliferation and migration of Schwann cells (SCs) were evaluated in both in vitro and in vivo. Methods: Human monocytic leukaemia cell line (THP-1) was successfully driven to M1 and M2 phenotypes by delivery of either IFN-γ or IL-4, respectively. SCs incubated with M1 or M2 macrophages-derived MVs, the cell migration and proliferation were assessed, and expression levels of nerve growth factor (NGF) and Laminin were measured. A rat model of sciatic nerve was established and the effects of macrophages-derived MVs on nerve regeneration were investigated. Results: M2-derived MVs elevated migration, proliferation, NFG and Laminin protein levels of SCs compared with M1-or M0-derived MVs. The relative expression levels of miR-223 were also increased in M2 macrophages and M2-derived MVs. Transfected M2 macrophages with miR-223 inhibitor then co-incubated with SCs, an inhibition of cell migration and proliferation and a down-regulated levels of NFG and Laminin protein expression were observed. In vivo, M2-derived MVs significantly increased the infiltration and axon number of SCs. Conclusion: M2-derived MVs promoted proliferation and migration of SCs in vitro and in vivo, which provided a therapeutic strategy for nerve regeneration. - Highlights: • M2 macrophages-derived MVs elevated migration and proliferation of SCs. • M2 macrophages-derived MVs up-regulated NFG and Laminin expression of SCs. • MiR-223 expression was increased in M2 macrophages-derived MVs. • MiR-223 inhibitor reduced migration and proliferation of SCs co-incubated with MVs. • MiR-223 inhibitor down-regulated NFG and Laminin levels of SCs co-incubated with MVs.

  9. The thioredoxin system in breast cancer cell invasion and migration

    Directory of Open Access Journals (Sweden)

    Maneet Bhatia

    2016-08-01

    Full Text Available Metastasis is the most life threatening aspect of breast cancer. It is a multi-step process involving invasion and migration of primary tumor cells with a subsequent colonization of these cells at a secondary location. The aim of the present study was to investigate the role of thioredoxin (Trx1 in the invasion and migration of breast cancer cells and to assess the strength of the association between high levels of Trx1 and thioredoxin reductase (TrxR1 expression with breast cancer patient survival. Our results indicate that the expression of both Trx1 and TrxR1 are statistically significantly increased in breast cancer patient cells compared with paired normal breast tissue from the same patient. Over-expression of Trx1 in MDA-MB-231 breast cancer cell lines enhanced cell invasion in in vitro assays while expression of a redox inactive mutant form of Trx1 (designated 1SS or the antisense mRNA inhibited cell invasion. Addition of exogenous Trx1 also enhanced cell invasion, while addition of a specific monoclonal antibody that inhibits Trx1 redox function decreased cell invasion. Over-expression of intracellular Trx1 did not increase cell migration but expression of intracellular 1SS inhibited migration. Addition of exogenous Trx1 enhanced cell migration while 1SS had no effect. Treatment with auranofin inhibited TrxR activity, cell migration and clonogenic activity of MDA-MB-231 cells, while increasing reactive oxygen species (ROS levels. Analysis of 25 independent cohorts with 5910 patients showed that Trx1 and TrxR1 were both associated with a poor patient prognosis in terms of overall survival, distant metastasis free survival and disease free survival. Therefore, targeting the Trx system with auranofin or other specific inhibitors may provide improved breast cancer patient outcomes through inhibition of cancer invasion and migration.

  10. Maturation of human dendritic cells by monocyte-conditioned medium is dependent upon trace amounts of lipopolysaccharide inducing tumour necrosis factor alpha

    DEFF Research Database (Denmark)

    Nersting, Jacob; Svenson, Morten; Andersen, Vagn

    2003-01-01

    We investigated the ability of monocyte-conditioned medium (MCM), generated by monocytes cultured on plastic-immobilised immunoglobulin, to stimulate maturation of human monocyte-derived dendritic cells (DC). Earlier reports suggest that MCM is a strong inducer of irreversible DC maturation......, whereas we find, that adding a small amount of lipopolysaccharide (LPS) to the MCM-generating cultures is required for the production of a DC-stimulatory MCM. Moreover, compared with addition of LPS directly to the DC cultures, stimulation via MCM cultures increases by several fold the DC...

  11. The effect of short-chain fatty acids on human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Nastasi, Claudia; Candela, Marco; Bonefeld, Charlotte Menné

    2015-01-01

    negligible effects, while both butyrate and propionate strongly modulated gene expression in both immature and mature human monocyte-derived DC. An Ingenuity pathway analysis based on the differentially expressed genes suggested that propionate and butyrate modulate leukocyte trafficking, as SCFA strongly......The gut microbiota is essential for human health and plays an important role in the pathogenesis of several diseases. Short-chain fatty acids (SCFA), such as acetate, butyrate and propionate, are end-products of microbial fermentation of macronutrients that distribute systemically via the blood....... The aim of this study was to investigate the transcriptional response of immature and LPS-matured human monocyte-derived DC to SCFA. Our data revealed distinct effects exerted by each individual SCFA on gene expression in human monocyte-derived DC, especially in the mature ones. Acetate only exerted...

  12. Embryonic cell-cell adhesion: a key player in collective neural crest migration.

    Science.gov (United States)

    Barriga, Elias H; Mayor, Roberto

    2015-01-01

    Cell migration is essential for morphogenesis, adult tissue remodeling, wound healing, and cancer cell migration. Cells can migrate as individuals or groups. When cells migrate in groups, cell-cell interactions are crucial in order to promote the coordinated behavior, essential for collective migration. Interestingly, recent evidence has shown that cell-cell interactions are also important for establishing and maintaining the directionality of these migratory events. We focus on neural crest cells, as they possess extraordinary migratory capabilities that allow them to migrate and colonize tissues all over the embryo. Neural crest cells undergo an epithelial-to-mesenchymal transition at the same time than perform directional collective migration. Cell-cell adhesion has been shown to be an important source of planar cell polarity and cell coordination during collective movement. We also review molecular mechanisms underlying cadherin turnover, showing how the modulation and dynamics of cell-cell adhesions are crucial in order to maintain tissue integrity and collective migration in vivo. We conclude that cell-cell adhesion during embryo development cannot be considered as simple passive resistance to force, but rather participates in signaling events that determine important cell behaviors required for cell migration. © 2015 Elsevier Inc. All rights reserved.

  13. Acute serum amyloid A induces migration, angiogenesis, and inflammation in synovial cells in vitro and in a human rheumatoid arthritis/SCID mouse chimera model.

    LENUS (Irish Health Repository)

    Connolly, Mary

    2010-06-01

    Serum amyloid A (A-SAA), an acute-phase protein with cytokine-like properties, is expressed at sites of inflammation. This study investigated the effects of A-SAA on chemokine-regulated migration and angiogenesis using rheumatoid arthritis (RA) cells and whole-tissue explants in vitro, ex vivo, and in vivo. A-SAA levels were measured by real-time PCR and ELISA. IL-8 and MCP-1 expression was examined in RA synovial fibroblasts, human microvascular endothelial cells, and RA synovial explants by ELISA. Neutrophil transendothelial cell migration, cell adhesion, invasion, and migration were examined using transwell leukocyte\\/monocyte migration assays, invasion assays, and adhesion assays with or without anti-MCP-1\\/anti-IL-8. NF-kappaB was examined using a specific inhibitor and Western blotting. An RA synovial\\/SCID mouse chimera model was used to examine the effects of A-SAA on cell migration, proliferation, and angiogenesis in vivo. High expression of A-SAA was demonstrated in RA patients (p < 0.05). A-SAA induced chemokine expression in a time- and dose-dependent manner (p < 0.05). Blockade with anti-scavenger receptor class B member 1 and lipoxin A4 (A-SAA receptors) significantly reduced chemokine expression in RA synovial tissue explants (p < 0.05). A-SAA induced cell invasion, neutrophil-transendothelial cell migration, monocyte migration, and adhesion (all p < 0.05), effects that were blocked by anti-IL-8 or anti-MCP-1. A-SAA-induced chemokine expression was mediated through NF-kappaB in RA explants (p < 0.05). Finally, in the RA synovial\\/SCID mouse chimera model, we demonstrated for the first time in vivo that A-SAA directly induces monocyte migration from the murine circulation into RA synovial grafts, synovial cell proliferation, and angiogenesis (p < 0.05). A-SAA promotes cell migrational mechanisms and angiogenesis critical to RA pathogenesis.

  14. Long-Term Live Cell Imaging of Cell Migration: Effects of Pathogenic Fungi on Human Epithelial Cell Migration.

    Science.gov (United States)

    Wöllert, Torsten; Langford, George M

    2016-01-01

    Long-term live cell imaging was used in this study to determine the responses of human epithelial cells to pathogenic biofilms formed by Candida albicans. Epithelial cells of the skin represent the front line of defense against invasive pathogens such as C. albicans but under certain circumstances, especially when the host's immune system is compromised, the skin barrier is breached. The mechanisms by which the fungal pathogen penetrates the skin and invade the deeper layers are not fully understood. In this study we used keratinocytes grown in culture as an in vitro model system to determine changes in host cell migration and the actin cytoskeleton in response to virulence factors produced by biofilms of pathogenic C. albicans. It is clear that changes in epithelial cell migration are part of the response to virulence factors secreted by biofilms of C. albicans and the actin cytoskeleton is the downstream effector that mediates cell migration. Our goal is to understand the mechanism by which virulence factors hijack the signaling pathways of the actin cytoskeleton to alter cell migration and thereby invade host tissues. To understand the dynamic changes of the actin cytoskeleton during infection, we used long-term live cell imaging to obtain spatial and temporal information of actin filament dynamics and to identify signal transduction pathways that regulate the actin cytoskeleton and its associated proteins. Long-term live cell imaging was achieved using a high resolution, multi-mode epifluorescence microscope equipped with specialized light sources, high-speed cameras with high sensitivity detectors, and specific biocompatible fluorescent markers. In addition to the multi-mode epifluorescence microscope, a spinning disk confocal long-term live cell imaging system (Olympus CV1000) equipped with a stage incubator to create a stable in vitro environment for long-term real-time and time-lapse microscopy was used. Detailed descriptions of these two long-term live

  15. Terbinafine stimulates the pro-inflammatory responses in human monocytic THP-1 cells through an ERK signaling pathway.

    Science.gov (United States)

    Mizuno, Katsuhiko; Fukami, Tatsuki; Toyoda, Yasuyuki; Nakajima, Miki; Yokoi, Tsuyoshi

    2010-10-23

    Oral antifungal terbinafine has been reported to cause liver injury with inflammatory responses in a small percentage of patients. However the underlying mechanism remains unknown. To examine the inflammatory reactions, we investigated whether terbinafine and other antifungal drugs increase the release of pro-inflammatory cytokines using human monocytic cells. Dose- and time-dependent changes in the mRNA expression levels and the release of interleukin (IL)-8 and tumor necrosis factor (TNF)α from human monocytic THP-1 and HL-60 cells with antifungal drugs were measured. Effects of terbinafine on the phosphorylation of extracellular signal-regulated kinase (ERK)1/2, p38 mitogen-activated protein (MAP) kinase and c-Jun N-terminal kinase (JNK)1/2 were investigated. The release of IL-8 and TNFα from THP-1 and HL-60 cells was significantly increased by treatment with terbinafine but not by fluconazole, suggesting that terbinafine can stimulate monocytes and increase the pro-inflammatory cytokine release. Terbinafine also significantly increased the phosphorylation of ERK1/2 and p38 MAP kinase in THP-1 cells. Pretreatment with a MAP kinase/ERK kinase (MEK)1/2 inhibitor U0126 significantly suppressed the increase of IL-8 and TNFα levels by terbinafine treatment in THP-1 cells, but p38 MAPK inhibitor SB203580 did not. These results suggested that an ERK1/2 pathway plays an important role in the release of IL-8 and TNFα in THP-1 cells treated with terbinafine. The release of inflammatory mediators by terbinafine might be one of the mechanisms underlying immune-mediated liver injury. This in vitro method may be useful to predict adverse inflammatory reactions that lead to drug-induced liver injury. Copyright © 2010 Elsevier Inc. All rights reserved.

  16. CD16+ Monocyte Subsets Are Increased in Large Abdominal Aortic Aneurysms and Are Differentially Related with Circulating and Cell-Associated Biochemical and Inflammatory Biomarkers

    Directory of Open Access Journals (Sweden)

    Giorgio Ghigliotti

    2013-01-01

    Full Text Available Proinflammatory components are present in abdominal aortic aneurysm (AAA. Circulating monocytes display heterogeneity, and three subsets have been identified, based on the differential expression for CD14 and CD16 receptors: CD14+CD16-, classical, CD14+CD16+, intermediate and CD14dim CD16+, non-classical monocytes. Increased proinflammatory CD16+ monocytes with high expression of CD143 are present in CKD patients. D-dimer is increased in AAA patients, and might contribute to the pro-inflammatory response associated to circulating monocytes. We aimed to investigate the frequency of CD14+CD16+, CD14dim CD16+ monocytes and monocyte CD143 expression in AAA patients, and their relationship with D-dimer, eGFR and other inflammatory parameters. Blood from 74 AAA patients and 30 healthy controls was analyzed to determine the frequency of CD14+, CD16+, CD14dim CD16+ monocytes and the monocyte CD143 expression by means of flow-cytometry. AAA patients had expanded CD16+ SUPsets (CD14+CD16+: 7.66 ± 0.31% vs 5.42 ± 0.27%; CD14dim CD16+: 7.43 ± 0.48% vs 5.54 ± 0.38%, AAA vs controls, mean ± SE, both p<0.05. CD14+ CD16+ cells were associated to D-dimer and age, and to reduced eGFR. CD14dim CD16+ cells were associated to uric acid, surface CD143, and reduced count of total leukocytes and neutrophils. Within AAA patients, the two CD16+ supsets and the monocyte CD143 expression display different relationships with D-dimer, parameters of renal function and circulating biochemical and inflammatory biomarkers.

  17. Modulation of monocytic leukemia cell function and survival by high gradient magnetic fields and mathematical modeling studies.

    Science.gov (United States)

    Zablotskii, Vitalii; Syrovets, Tatiana; Schmidt, Zoe W; Dejneka, Alexandr; Simmet, Thomas

    2014-03-01

    The influence of spatially modulated high gradient magnetic fields on cellular functions of human THP-1 leukemia cells is studied. We demonstrate that arrays of high-gradient micrometer-sized magnets induce i) cell swelling, ii) prolonged increased ROS production, and iii) inhibit cell proliferation, and iv) elicit apoptosis of THP-1 monocytic leukemia cells in the absence of chemical or biological agents. Mathematical modeling indicates that mechanical stress exerted on the cells by high magnetic gradient forces is responsible for triggering cell swelling and formation of reactive oxygen species followed by apoptosis. We discuss physical aspects of controlling cell functions by focused magnetic gradient forces, i.e. by a noninvasive and nondestructive physical approach. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. SOX15 regulates proliferation and migration of endometrial cancer cells.

    Science.gov (United States)

    Rui, Xiaohui; Xu, Yun; Jiang, Xiping; Guo, Caixia; Jiang, Jingting

    2017-10-31

    The study aimed to investigate the effects of Sry-like high mobility group box 15 ( SOX15 ) on proliferation and migration of endometrial cancer (EC) cells. Immunohistochemistry (IHC) was applied to determine the expression of SOX15 in EC tissues and adjacent tissues. We used cell transfection method to construct the HEC-1-A and Ishikawa cell lines with stable overexpression and low expression SOX15 Reverse-transcription quantitative real-time PCR (RT-qPCR) and Western blot were performed to examine expression of SOX15 mRNA and SOX15 protein, respectively. By conducting a series of cell proliferation assay and migration assay, we analyzed the influence of SOX15 overexpression or low expression on EC cell proliferation and migration. The expression of SOX15 mRNA and protein in EC tissues was significantly lower than that in adjacent tissues. After lentivirus-transfecting SOX15 , the expression level of SOX15 mRNA and protein was significantly increased in cells of SOX15 group, and decreased in sh- SOX15 group. Overexpression of SOX15 could suppress cell proliferation, while down-regulation of SOX15 increased cell proliferation. Flow cytometry results indicated that overexpression of SOX15 induced the ratio of cell-cycle arrest in G 1 stage. In addition, Transwell migration assay results showed that SOX15 overexpression significantly inhibited cell migration, and also down-regulation of SOX15 promoted the migration. As a whole, SOX15 could regulate the proliferation and migration of EC cells and up- regulation of SOX15 could be valuable for EC treatment. © 2017 The Author(s).

  19. Isolation of human monocytes by double gradient centrifugation and their differentiation to macrophages in teflon-coated cell culture bags.

    Science.gov (United States)

    Menck, Kerstin; Behme, Daniel; Pantke, Mathias; Reiling, Norbert; Binder, Claudia; Pukrop, Tobias; Klemm, Florian

    2014-09-09

    Human macrophages are involved in a plethora of pathologic processes ranging from infectious diseases to cancer. Thus they pose a valuable tool to understand the underlying mechanisms of these diseases. We therefore present a straightforward protocol for the isolation of human monocytes from buffy coats, followed by a differentiation procedure which results in high macrophage yields. The technique relies mostly on commonly available lab equipment and thus provides a cost and time effective way to obtain large quantities of human macrophages. Briefly, buffy coats from healthy blood donors are subjected to a double density gradient centrifugation to harvest monocytes from the peripheral blood. These monocytes are then cultured in fluorinated ethylene propylene (FEP) Teflon-coated cell culture bags in the presence of macrophage colony-stimulating factor (M-CSF). The differentiated macrophages can be easily harvested and used for subsequent studies and functional assays. Important methods for quality control and validation of the isolation and differentiation steps will be highlighted within the protocol. In summary, the protocol described here enables scientists to routinely and reproducibly isolate human macrophages without the need for cost intensive tools. Furthermore, disease models can be studied in a syngeneic human system circumventing the use of murine macrophages.

  20. Activated human mast cells induce LOX-1-specific scavenger receptor expression in human monocyte-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Mervi Alanne-Kinnunen

    Full Text Available Activated mast cells in atherosclerotic lesions degranulate and release bioactive compounds capable of regulating atherogenesis. Here we examined the ability of activated human primary mast cells to regulate the expression of the major scavenger receptors in cultured human primary monocyte-derived macrophages (HMDMs.Components released by immunologically activated human primary mast cells induced a transient expression of lectin-like oxidized LDL receptor (LOX-1 mRNA in HMDMs, while the expression of two other scavenger receptors, MSR1 and CD36, remained unaffected. The LOX-1-inducing secretory components were identified as histamine, tumor necrosis factor alpha (TNF-α, and transforming growth factor beta (TGF-β1, which exhibited a synergistic effect on LOX-1 mRNA expression. Histamine induced a transient expression of LOX-1 protein. Mast cell -induced increase in LOX-1 expression was not associated with increased uptake of oxidized LDL by the macrophages.Mast cell-derived histamine, TNF-α, and TGF-β1 act in concert to induce a transient increase in LOX-1 expression in human primary monocyte-derived macrophages. The LOX-1-inducing activity potentially endows mast cells a hitherto unrecognized role in the regulation of innate immune reactions in atherogenesis.

  1. Cancer Cell-derived Exosomes Induce Mitogen-activated Protein Kinase-dependent Monocyte Survival by Transport of Functional Receptor Tyrosine Kinases*

    Science.gov (United States)

    Song, Xiao; Ding, Yanping; Liu, Gang; Yang, Xiao; Zhao, Ruifang; Zhang, Yinlong; Zhao, Xiao; Anderson, Gregory J.; Nie, Guangjun

    2016-01-01

    Tumor-associated macrophages (TAM) play pivotal roles in cancer initiation and progression. Monocytes, the precursors of TAMs, normally undergo spontaneous apoptosis within 2 days, but can subsist in the inflammatory tumor microenvironment for continuous survival and generation of sufficient TAMs. The mechanisms underlying tumor-driving monocyte survival remain obscure. Here we report that cancer cell-derived exosomes were crucial mediators for monocyte survival in the inflammatory niche. Analysis of the survival-promoting molecules in monocytes revealed that cancer cell-derived exosomes activated Ras and extracellular signal-regulated kinases in the mitogen-activated protein kinase (MAPK) pathway, resulting in the prevention of caspase cleavage. Phosphorylated receptor tyrosine kinases (RTKs), such as phosphorylated epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER-2), were abundantly expressed in cancer cell-derived exosomes. Knock-out of EGFR or/and HER-2, or alternatively, inhibitors against their phosphorylation significantly disturbed the exosome-mediated activation of the MAPK pathway, inhibition of caspase cleavage, and increase in survival rate in monocytes. Moreover, the deprived survival-stimulating activity of exosomes due to null expression of EGFR and HER-2 could be restored by activation of another RTK, insulin receptor. Overall, our study uncovered a mechanism of tumor-associated monocyte survival and demonstrated that cancer cell-derived exosomes can stimulate the MAPK pathway in monocytes through transport of functional RTKs, leading to inactivation of apoptosis-related caspases. This work provides insights into the long sought question on monocyte survival prior to formation of plentiful TAMs in the tumor microenvironment. PMID:26895960

  2. Cancer Cell-derived Exosomes Induce Mitogen-activated Protein Kinase-dependent Monocyte Survival by Transport of Functional Receptor Tyrosine Kinases.

    Science.gov (United States)

    Song, Xiao; Ding, Yanping; Liu, Gang; Yang, Xiao; Zhao, Ruifang; Zhang, Yinlong; Zhao, Xiao; Anderson, Gregory J; Nie, Guangjun

    2016-04-15

    Tumor-associated macrophages (TAM) play pivotal roles in cancer initiation and progression. Monocytes, the precursors of TAMs, normally undergo spontaneous apoptosis within 2 days, but can subsist in the inflammatory tumor microenvironment for continuous survival and generation of sufficient TAMs. The mechanisms underlying tumor-driving monocyte survival remain obscure. Here we report that cancer cell-derived exosomes were crucial mediators for monocyte survival in the inflammatory niche. Analysis of the survival-promoting molecules in monocytes revealed that cancer cell-derived exosomes activated Ras and extracellular signal-regulated kinases in the mitogen-activated protein kinase (MAPK) pathway, resulting in the prevention of caspase cleavage. Phosphorylated receptor tyrosine kinases (RTKs), such as phosphorylated epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER-2), were abundantly expressed in cancer cell-derived exosomes. Knock-out of EGFR or/and HER-2, or alternatively, inhibitors against their phosphorylation significantly disturbed the exosome-mediated activation of the MAPK pathway, inhibition of caspase cleavage, and increase in survival rate in monocytes. Moreover, the deprived survival-stimulating activity of exosomes due to null expression of EGFR and HER-2 could be restored by activation of another RTK, insulin receptor. Overall, our study uncovered a mechanism of tumor-associated monocyte survival and demonstrated that cancer cell-derived exosomes can stimulate the MAPK pathway in monocytes through transport of functional RTKs, leading to inactivation of apoptosis-related caspases. This work provides insights into the long sought question on monocyte survival prior to formation of plentiful TAMs in the tumor microenvironment. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Gradient biomaterials and their influences on cell migration

    Science.gov (United States)

    Wu, Jindan; Mao, Zhengwei; Tan, Huaping; Han, Lulu; Ren, Tanchen; Gao, Changyou

    2012-01-01

    Cell migration participates in a variety of physiological and pathological processes such as embryonic development, cancer metastasis, blood vessel formation and remoulding, tissue regeneration, immune surveillance and inflammation. The cells specifically migrate to destiny sites induced by the gradually varying concentration (gradient) of soluble signal factors and the ligands bound with the extracellular matrix in the body during a wound healing process. Therefore, regulation of the cell migration behaviours is of paramount importance in regenerative medicine. One important way is to create a microenvironment that mimics the in vivo cellular and tissue complexity by incorporating physical, chemical and biological signal gradients into engineered biomaterials. In this review, the gradients existing in vivo and their influences on cell migration are briefly described. Recent developments in the fabrication of gradient biomaterials for controlling cellular behaviours, especially the cell migration, are summarized, highlighting the importance of the intrinsic driving mechanism for tissue regeneration and the design principle of complicated and advanced tissue regenerative materials. The potential uses of the gradient biomaterials in regenerative medicine are introduced. The current and future trends in gradient biomaterials and programmed cell migration in terms of the long-term goals of tissue regeneration are prospected. PMID:23741610

  4. The role of monocytes and T cells in 1,25-dihydroxyvitamin D3 mediated inhibition of B cell function in vitro

    DEFF Research Database (Denmark)

    Müller, K; Heilmann, C; Poulsen, L K

    1991-01-01

    1,25-Dihydroxyvitamin D3 (1,25-(OH)2D3) inhibits immunoglobulin production by human mononuclear cells (MNC) in vitro. The present study was undertaken to evaluate the role of T cells and monocytes in 1,25-(OH)2D3 induced suppression of B cell functions. The synthetic vitamin D3 analogue MC 903...... was examined in parallel. 1,25-(OH)2D3 and MC 903 showed a dose-related inhibition of IgM, IgG and IgA plaque-forming cells in poke-weed mitogen (PWM) activated cultures of MNC. This effect was most likely mediated through impairment of T cell and monocyte functions. First, the inhibitory effect was seen after...

  5. Immature dendritic cells generated from cryopreserved human monocytes show impaired ability to respond to LPS and to induce allogeneic lymphocyte proliferation.

    Directory of Open Access Journals (Sweden)

    Guilherme Ferreira Silveira

    Full Text Available Dendritic cells play a key role in the immune system, in the sensing of foreign antigens and triggering of an adaptive immune response. Cryopreservation of human monocytes was investigated to understand its effect on differentiation into immature monocyte-derived dendritic cells (imdDCs, the response to inflammatory stimuli and the ability to induce allogeneic lymphocyte proliferation. Cryopreserved (crp-monocytes were able to differentiate into imdDCs, albeit to a lesser extent than freshly (frh-obtained monocytes. Furthermore, crp-imdDCs had lower rates of maturation and cytokine/chemokine secretion in response to LPS than frh-imdDCs. Lower expression of Toll-like receptor 4 (at 24 and 48 h and higher susceptibility to apoptosis in crp-imdDCs than in fresh cells would account for the impaired maturation and cytokine/chemokine secretion observed. A mixed leukocyte reaction showed that lymphocyte proliferation was lower with crp-imdDCs than with frh-imdDCs. These findings suggested that the source of monocytes used to generate human imdDCs could influence the accuracy of results observed in studies of the immune response to pathogens, lymphocyte activation, vaccination and antigen sensing. It is not always possible to work with freshly isolated monocytes but the possible effects of freezing/thawing on the biology and responsiveness of imdDCs should be taken into account.

  6. Emergent patterns of collective cell migration under tubular confinement.

    Science.gov (United States)

    Xi, Wang; Sonam, Surabhi; Beng Saw, Thuan; Ladoux, Benoit; Teck Lim, Chwee

    2017-11-15

    Collective epithelial behaviors are essential for the development of lumens in organs. However, conventional assays of planar systems fail to replicate cell cohorts of tubular structures that advance in concerted ways on out-of-plane curved and confined surfaces, such as ductal elongation in vivo. Here, we mimic such coordinated tissue migration by forming lumens of epithelial cell sheets inside microtubes of 1-10 cell lengths in diameter. We show that these cell tubes reproduce the physiological apical-basal polarity, and have actin alignment, cell orientation, tissue organization, and migration modes that depend on the extent of tubular confinement and/or curvature. In contrast to flat constraint, the cell sheets in a highly constricted smaller microtube demonstrate slow motion with periodic relaxation, but fast overall movement in large microtubes. Altogether, our findings provide insights into the emerging migratory modes for epithelial migration and growth under tubular confinement, which are reminiscent of the in vivo scenario.

  7. Coordinate viral induction of tumor necrosis factor α and interferon β in human B cells and monocytes

    International Nuclear Information System (INIS)

    Goldfeld, A.E.; Maniatis, T.

    1989-01-01

    Human tumor necrosis factor α (TNF-α) gene expression can be induced primarily in cells of the monocyte/macrophage lineage by a variety of inducers, including lipopolysaccharide, phorbol esters such as phorbol 12-myristate 13-acetate, and virus or synthetic double-stranded RNA [poly(I)·poly(C)]. In this paper the authors show that the TNF-α gene also responds to virus and phorbol 12-myristate 13-acetate in B lymphocytes and that virus is the most potent inducer of TNF-α mRNA in both monocyte and B-cell lines. In addition, they show that viral infection coinduces the expression of TNF-α and interferon β mRNA and that viral induction of both genes is blocked by the kinase inhibitor 2-aminopurine. Inhibition of protein synthesis with cycloheximide had no effect on mRNA expression of the genes in one of three cell lines tested (U937) but blocked the viral induction of both genes in another (Namalwa). Thus, the regulatory factors required for mRNA induction of both genes are present prior to the addition of virus in U937 but not in Namalwa cells. However, in a third cell line (JY), cycloheximide blocked viral induction of the interferon β gene but not the TNF-α gene. Taken together, these observations suggest that viral induction of TNF-α and interferon β gene expression may involve overlapping pathways with both common and distinct regulatory factors

  8. Interleukin 17 receptor A modulates monocyte subsets and macrophage generation in vivo.

    Directory of Open Access Journals (Sweden)

    Shuwang Ge

    Full Text Available Interleukin (IL-17A signaling via Interleukin 17 receptor A (Il17ra contributes to the inflammatory host response by inducing recruitment of innate immune cells, but also plays a role in homeostatic neutrophilic granulocyte regulation. Monocytes, the other main innate immune cell, have a longer life span and can pursue multiple differentiation pathways towards tissue macrophages. Monocytes are divided into two subpopulations by expression of the Ly6C/Gr1 surface marker in mice. We here investigated the role of Il17ra in monocyte homeostasis and macrophage generation. In Il17ra(-/- and in mixed bone marrow chimeric wt/Il17ra(-/- mice, the concentrations of circulating Il17ra(-/- Gr1(low monocytes were significantly decreased compared to wt cells. Pulmonary, splenic and resident peritoneal Il17ra(-/- macrophages were significantly fewer than of wt origin. Bone marrow progenitor and monocyte numbers were equal, but the proportion of Il17ra(-/- Gr1(low monocytes was already decreased at bone marrow level. After monocyte depletion, initial Gr1(high and Gr1(low monocyte regeneration of Il17ra(-/- and wt cells was very similar. However, Il17ra(-/- Gr1(low counts were not sustained. After labeling with either fluorescent beads or BrdU, Il17ra(-/- Gr1(high monocyte transition to Gr1(low cells was not detectable unlike wt cells. Monocyte recruitment in acute peritonitis, which is known to be largely due to Gr1(high cell migration, was unaffected in an identical environment. Unilateral ureteral obstruction induces a less acute inflammatory and fibrotic kidney injury. Compared to wt cells in the same environment, Il17ra(-/- macrophage accumulation in the kidney was decreased. In the absence of Il17ra on all myeloid cells, renal fibrosis was significantly attenuated. Our data show that Il17ra modulates Gr1(low monocyte counts and suggest defective Gr1(high to Gr1(low monocyte transition as an underlying mechanism. Lack of Il17ra altered homeostatic tissue

  9. Use of an adaptable cell culture kit for performing lymphocyte and monocyte cell cultures in microgravity

    Science.gov (United States)

    Hatton, J. P.; Lewis, M. L.; Roquefeuil, S. B.; Chaput, D.; Cazenave, J. P.; Schmitt, D. A.

    1998-01-01

    The results of experiments performed in recent years on board facilities such as the Space Shuttle/Spacelab have demonstrated that many cell systems, ranging from simple bacteria to mammalian cells, are sensitive to the microgravity environment, suggesting gravity affects fundamental cellular processes. However, performing well-controlled experiments aboard spacecraft offers unique challenges to the cell biologist. Although systems such as the European 'Biorack' provide generic experiment facilities including an incubator, on-board 1-g reference centrifuge, and contained area for manipulations, the experimenter must still establish a system for performing cell culture experiments that is compatible with the constraints of spaceflight. Two different cell culture kits developed by the French Space Agency, CNES, were recently used to perform a series of experiments during four flights of the 'Biorack' facility aboard the Space Shuttle. The first unit, Generic Cell Activation Kit 1 (GCAK-1), contains six separate culture units per cassette, each consisting of a culture chamber, activator chamber, filtration system (permitting separation of cells from supernatant in-flight), injection port, and supernatant collection chamber. The second unit (GCAK-2) also contains six separate culture units, including a culture, activator, and fixation chambers. Both hardware units permit relatively complex cell culture manipulations without extensive use of spacecraft resources (crew time, volume, mass, power), or the need for excessive safety measures. Possible operations include stimulation of cultures with activators, separation of cells from supernatant, fixation/lysis, manipulation of radiolabelled reagents, and medium exchange. Investigations performed aboard the Space Shuttle in six different experiments used Jurkat, purified T-cells or U937 cells, the results of which are reported separately. We report here the behaviour of Jurkat and U937 cells in the GCAK hardware in ground

  10. ER Alpha Rapid Signaling Is Required for Estrogen Induced Proliferation and Migration of Vascular Endothelial Cells.

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    Qing Lu

    Full Text Available Estrogen promotes the proliferation and migration of vascular endothelial cells (ECs, which likely underlies its ability to accelerate re-endothelialization and reduce adverse remodeling after vascular injury. In previous studies, we have shown that the protective effects of E2 (the active endogenous form of estrogen in vascular injury require the estrogen receptor alpha (ERα. ERα transduces the effects of estrogen via a classical DNA binding, "genomic" signaling pathway and via a more recently-described "rapid" signaling pathway that is mediated by a subset of ERα localized to the cell membrane. However, which of these pathways mediates the effects of estrogen on endothelial cells is poorly understood. Here we identify a triple point mutant version of ERα (KRR ERα that is specifically defective in rapid signaling, but is competent to regulate transcription through the "genomic" pathway. We find that in ECs expressing wild type ERα, E2 regulates many genes involved in cell migration and proliferation, promotes EC migration and proliferation, and also blocks the adhesion of monocytes to ECs. ECs expressing KRR mutant ERα, however, lack all of these responses. These observations establish KRR ERα as a novel tool that could greatly facilitate future studies into the vascular and non-vascular functions of ERα rapid signaling. Further, they support that rapid signaling through ERα is essential for many of the transcriptional and physiological responses of ECs to E2, and that ERα rapid signaling in ECs, in vivo, may be critical for the vasculoprotective and anti-inflammatory effects of estrogen.

  11. Stem cell factor supports migration in canine mesenchymal stem cells.

    Science.gov (United States)

    Enciso, Nathaly; Ostronoff, Luciana L K; Mejías, Guillermo; León, Leticia G; Fermín, María Luisa; Merino, Elena; Fragio, Cristina; Avedillo, Luis; Tejero, Concepción

    2018-03-01

    Adult Mesenchymal Stem Cells (MSC) are cells that can be defined as multipotent cells able to differentiate into diverse lineages, under appropriate conditions. These cells have been widely used in regenerative medicine, both in preclinical and clinical settings. Initially discovered in bone marrow, MSC can now be isolated from a wide spectrum of adult and foetal tissues. Studies to evaluate the therapeutic potential of these cells are based on their ability to arrive to damaged tissues. In this paper we have done a comparative study analyzing proliferation, surface markers and OCT4, SOX9, RUNX2, PPARG genes expression in MSC cells from Bone marrow (BMMSC) and Adipose tissue (ASC). We also analyzed the role of Stem Cell Factor (SCF) on MSC proliferation and on ASCs metalloproteinases MMP-2, MMP-9 secretion. Healthy dogs were used as BMMSC donors, and ASC were collected from omentum during elective ovariohysterectomy surgery. Both cell types were cultured in IMDM medium with or without SCF, 10% Dog Serum (DS), and incubated at 38 °C with 5% CO2. Growth of BMMSCs and ASCs was exponential until 25-30 days. Flow citometry of MSCs revealed positive results for CD90 and negative for CD34, CD45 and MCH-II. Genes were evaluated by RT-PCR and metalloproteinases by zymografy. Our findings indicate morphological and immunological similarities as well as expression of genes from both origins on analyzed cells. Furthermore, SCF did not affect proliferation of MSCs, however it up-regulated MMP-2 and MMP-9 secretion in ASCs. These results suggest that metalloproteinases are possibly essential molecules pivoting migration.

  12. Multiscale mechanisms of cell migration during development: theory and experiment.

    Science.gov (United States)

    McLennan, Rebecca; Dyson, Louise; Prather, Katherine W; Morrison, Jason A; Baker, Ruth E; Maini, Philip K; Kulesa, Paul M

    2012-08-01

    Long-distance cell migration is an important feature of embryonic development, adult morphogenesis and cancer, yet the mechanisms that drive subpopulations of cells to distinct targets are poorly understood. Here, we use the embryonic neural crest (NC) in tandem with theoretical studies to evaluate model mechanisms of long-distance cell migration. We find that a simple chemotaxis model is insufficient to explain our experimental data. Instead, model simulations predict that NC cell migration requires leading cells to respond to long-range guidance signals and trailing cells to short-range cues in order to maintain a directed, multicellular stream. Experiments confirm differences in leading versus trailing NC cell subpopulations, manifested in unique cell orientation and gene expression patterns that respond to non-linear tissue growth of the migratory domain. Ablation experiments that delete the trailing NC cell subpopulation reveal that leading NC cells distribute all along the migratory pathway and develop a leading/trailing cellular orientation and gene expression profile that is predicted by model simulations. Transplantation experiments and model predictions that move trailing NC cells to the migratory front, or vice versa, reveal that cells adopt a gene expression profile and cell behaviors corresponding to the new position within the migratory stream. These results offer a mechanistic model in which leading cells create and respond to a cell-induced chemotactic gradient and transmit guidance information to trailing cells that use short-range signals to move in a directional manner.

  13. Generation of dendritic cells from human bone marrow mononuclear cells: advantages for clinical application in comparison to peripheral blood monocyte derived cells.

    Science.gov (United States)

    Bai, L; Feuerer, M; Beckhove, P; Umansky, V; Schirrmacher, V

    2002-02-01

    Dendritic cells (DCs) currently used for vaccination in clinical studies to induce immunity against malignant cells are normally generated from peripheral blood-derived monocytes. Here we studied conditions for the generation of DCs from unseparated human bone marrow (BM) mononuclear cells and compared them functionally with DCs from blood. The two types of DCs, from bone marrow (BM-DC) and peripheral blood (BL-DC), were generated in parallel from the same normal healthy donors by culturing in serum-free X-VIVO 20 medium containing GM-CSF and IL-4, and then the phenotypes and functions were compared. BM-DC generation occurred in 14 days and involved proliferative expansion from CD34 stem cells and differentiation while BL-DC generation occurred in 7 days from CD14 monocytes and involved only differentiation. A 7- to 25-fold higher number of DCs could be obtained from BM than from blood. BM-DC had similar phenotypes as BL-DC. The capacity to stimulate MLR reactivity in allogeneic T lymphocytes was higher with BM-DC than that with BL-DC. Also, the capacity to stimulate autologous memory T cell responses to tetanus toxoid (TT) or tuberculin (PPD) was higher with BM-DC than with BL-DC. These results suggest that BM-DC as produced here may be a very economic and useful source of professional antigen-presenting cells for anti-tumor immunotherapeutic protocols.

  14. Similarities and differences between helminth parasites and cancer cell lines in shaping human monocytes: Insights into parallel mechanisms of immune evasion.

    Directory of Open Access Journals (Sweden)

    Prakash Babu Narasimhan

    2018-04-01

    Full Text Available A number of features at the host-parasite interface are reminiscent of those that are also observed at the host-tumor interface. Both cancer cells and parasites establish a tissue microenvironment that allows for immune evasion and may reflect functional alterations of various innate cells. Here, we investigated how the phenotype and function of human monocytes is altered by exposure to cancer cell lines and if these functional and phenotypic alterations parallel those induced by exposure to helminth parasites. Thus, human monocytes were exposed to three different cancer cell lines (breast, ovarian, or glioblastoma or to live microfilariae (mf of Brugia malayi-a causative agent of lymphatic filariasis. After 2 days of co-culture, monocytes exposed to cancer cell lines showed markedly upregulated expression of M1-associated (TNF-α, IL-1β, M2-associated (CCL13, CD206, Mreg-associated (IL-10, TGF-β, and angiogenesis associated (MMP9, VEGF genes. Similar to cancer cell lines, but less dramatically, mf altered the mRNA expression of IL-1β, CCL13, TGM2 and MMP9. When surface expression of the inhibitory ligands PDL1 and PDL2 was assessed, monocytes exposed to both cancer cell lines and to live mf significantly upregulated PDL1 and PDL2 expression. In contrast to exposure to mf, exposure to cancer cell lines increased the phagocytic ability of monocytes and reduced their ability to induce T cell proliferation and to expand Granzyme A+ CD8+ T cells. Our data suggest that despite the fact that helminth parasites and cancer cell lines are extraordinarily disparate, they share the ability to alter the phenotype of human monocytes.

  15. Similarities and differences between helminth parasites and cancer cell lines in shaping human monocytes: Insights into parallel mechanisms of immune evasion.

    Science.gov (United States)

    Narasimhan, Prakash Babu; Akabas, Leor; Tariq, Sameha; Huda, Naureen; Bennuru, Sasisekhar; Sabzevari, Helen; Hofmeister, Robert; Nutman, Thomas B; Tolouei Semnani, Roshanak

    2018-04-01

    A number of features at the host-parasite interface are reminiscent of those that are also observed at the host-tumor interface. Both cancer cells and parasites establish a tissue microenvironment that allows for immune evasion and may reflect functional alterations of various innate cells. Here, we investigated how the phenotype and function of human monocytes is altered by exposure to cancer cell lines and if these functional and phenotypic alterations parallel those induced by exposure to helminth parasites. Thus, human monocytes were exposed to three different cancer cell lines (breast, ovarian, or glioblastoma) or to live microfilariae (mf) of Brugia malayi-a causative agent of lymphatic filariasis. After 2 days of co-culture, monocytes exposed to cancer cell lines showed markedly upregulated expression of M1-associated (TNF-α, IL-1β), M2-associated (CCL13, CD206), Mreg-associated (IL-10, TGF-β), and angiogenesis associated (MMP9, VEGF) genes. Similar to cancer cell lines, but less dramatically, mf altered the mRNA expression of IL-1β, CCL13, TGM2 and MMP9. When surface expression of the inhibitory ligands PDL1 and PDL2 was assessed, monocytes exposed to both cancer cell lines and to live mf significantly upregulated PDL1 and PDL2 expression. In contrast to exposure to mf, exposure to cancer cell lines increased the phagocytic ability of monocytes and reduced their ability to induce T cell proliferation and to expand Granzyme A+ CD8+ T cells. Our data suggest that despite the fact that helminth parasites and cancer cell lines are extraordinarily disparate, they share the ability to alter the phenotype of human monocytes.

  16. Monocytic myeloid-derived suppressor cells as prognostic factor in chronic myeloid leukaemia patients treated with dasatinib.

    Science.gov (United States)

    Giallongo, Cesarina; Parrinello, Nunziatina L; La Cava, Piera; Camiolo, Giuseppina; Romano, Alessandra; Scalia, Marina; Stagno, Fabio; Palumbo, Giuseppe A; Avola, Roberto; Li Volti, Giovanni; Tibullo, Daniele; Di Raimondo, Francesco

    2018-02-01

    Myeloid suppressor cells are a heterogeneous group of myeloid cells that are increased in patients with chronic myeloid leukaemia (CML) inducing T cell tolerance. In this study, we found that therapy with tyrosine kinase inhibitors (TKI) decreased the percentage of granulocytic MDSC, but only patients treated with dasatinib showed a significant reduction in the monocytic subset (M-MDSC). Moreover, a positive correlation was observed between number of persistent M-MDSC and the value of major molecular response in dasatinib-treated patients. Serum and exosomes from patients with CML induced conversion of monocytes from healthy volunteers into immunosuppressive M-MDSC, suggesting a bidirectional crosstalk between CML cells and MDSC. Overall, we identified M-MDSC as prognostic factors in patients treated with dasatinib. It might be of interest to understand whether MDSC may be a candidate predictive markers of relapse risk following TKI discontinuation, suggesting their potential significance as practice of precision medicine. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  17. High-dose vitamin D in Addison's disease regulates T-cells and monocytes: A pilot trial.

    Science.gov (United States)

    Penna-Martinez, Marissa; Filmann, Natalie; Bogdanou, Dimitra; Shoghi, Firouzeh; Huenecke, Sabine; Schubert, Ralf; Herrmann, Eva; Koehl, Ulrike; Husebye, Eystein S; Badenhoop, Klaus

    2018-05-01

    On the basis of the immunomodulatory actions of vitamin D (VD), we investigated the effects of high-dose VD therapy over a 3 mo period on the immune response in patients with Addison's disease (AD). This randomized, controlled, crossover trial included 13 patients with AD who received either cholecalciferol (4000 IU/d) for 3 mo followed by 3 mo placebo oil or the sequential alternative placebo followed by verum. Glucocorticoid replacement doses remained stable. The primary outcome measures were changes in 25-hydroxyvitamin D3 (25(OH)D 3 ) levels and immune cells including T helper cells (Th; CD3 + CD4 + ), late-activated Th cells (CD3 + CD4 + HLA-DR + ), regulatory T cells (CD3 + CD4 + CD25 bright CD127 dim/neg ), cytotoxic T cells (Tc; CD3 + CD8 + ), late-activated Tc cells (CD3 + CD8 + HLA-DR + ), and monocytes. The explorative analysis included the correlation of changes with VD-related gene polymorphisms and 21-hydroxylase antibody titers. Ten of 13 patients (77%) were VD deficient. Median 25(OH)D 3 concentrations increased significantly to 41.5 ng/ml (median changes: 19.95 ng/ml; P = 0.0005) after 3 mo of cholecalciferol treatment. Within the T-cells, only the late-activated Th (median changes: 1.6%; P = 0.02) and late-activated Tc cells (median changes: 4.05%; P = 0.03) decreased, whereas monocytes (median changes: 1.05%; P = 0.008) increased after VD therapy. T-cell changes were associated with two polymorphisms (CYP27B1-rs108770012 and VDR-rs10735810), but no changes in the 21-hydroxylase antibody titers were observed. Three months of treatment with cholecalciferol achieved sufficient 25(OH)D 3 levels and can regulate late-activated T-cells and monocytes in patients with AD. Explorative analysis revealed potential genetic contributions. This pilot trial provides novel insights about immunomodulation in AD. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Effects of Cigarette Smoke Condensate on Oxidative Stress, Apoptotic Cell Death, and HIV Replication in Human Monocytic Cells.

    Directory of Open Access Journals (Sweden)

    Pss Rao

    Full Text Available While cigarette smoking is prevalent amongst HIV-infected patients, the effects of cigarette smoke constituents in cells of myeloid lineage are poorly known. Recently, we have shown that nicotine induces oxidative stress through cytochrome P450 (CYP 2A6-mediated pathway in U937 monocytic cells. The present study was designed to examine the effect of cigarette smoke condensate (CSC, which contains majority of tobacco constituents, on oxidative stress, cytotoxicity, expression of CYP1A1, and/or HIV-1 replication in HIV-infected (U1 and uninfected U937 cells. The effects of CSC on induction of CYP1 enzymes in HIV-infected primary macrophages were also analyzed. The results showed that the CSC-mediated increase in production of reactive oxygen species (ROS in U937 cells is dose- and time-dependent. Moreover, CSC treatment was found to induce cytotoxicity in U937 cells through the apoptotic pathway via activation of caspase-3. Importantly, pretreatment with vitamin C blocked the CSC-mediated production of ROS and induction of caspase-3 activity. In U1 cells, acute treatment of CSC increased ROS production at 6H (>2-fold and both ROS (>2 fold and HIV-1 replication (>3-fold after chronic treatment. The CSC mediated effects were associated with robust induction in the expression of CYP1A1 mRNA upon acute CSC treatment of U937 and U1 cells (>20-fold, and upon chronic CSC treatment to U1 cells (>30-fold. In addition, the CYP1A1 induction in U937 cells was mediated through the aromatic hydrocarbon receptor pathway. Lastly, CSC, which is known to increase viral replication in primary macrophages, was also found to induce CYP1 enzymes in HIV-infected primary macrophages. While mRNA levels of both CYP1A1 and CYP1B1 were elevated following CSC treatment, only CYP1B1 protein levels were increased in HIV-infected primary macrophages. In conclusion, these results suggest a possible association between oxidative stress, CYP1 expression, and viral replication in

  19. Hematopoietic stem cell migration and proliferation after Partial body irradiation

    International Nuclear Information System (INIS)

    Murata, Takashi; Utsumi, Makoto; Hotta, Tomomitsu; Yamada, Hideo

    1983-01-01

    Stem cell migration in hematopoietic recovery after partial body irradiation was investigated with special emphasis on the comparative roles of the bone marrow and the spleen. The number of CFU-S in circulation declined rapidly and reached zero within a day after irradiation, thereafter it increased gradually. This finding suggests the presence of two different phases of stem cell migration. One is a rapid migrating phase in which stem cells are released rapidly within a day after irradiation, and the other is a slow migrating phase. The result of split doses of local body irradiation experiments implicated a role for the spleen distinct from that of the bone marrow in the preferential distribution of stem cells early after irradiation. The cell kinetic study showed that the proliferation of CFU-S occurred actively in irradiated bone marrow and the spleens as compared to that in unirradiated control. But on Day 7 and on Day 10 after irradiation, the proliferation of CFU-S in shielded bone marrow did not occur as actively as those in irradiated areas. The results of our present studies suggest that the spleen is not only the storage pools of migrating stem cells but also the main site of active proliferation of CFU-S in the early period of hematopoietic regeneration. (author)

  20. Multidisciplinary approaches to understanding collective cell migration in developmental biology.

    Science.gov (United States)

    Schumacher, Linus J; Kulesa, Paul M; McLennan, Rebecca; Baker, Ruth E; Maini, Philip K

    2016-06-01

    Mathematical models are becoming increasingly integrated with experimental efforts in the study of biological systems. Collective cell migration in developmental biology is a particularly fruitful application area for the development of theoretical models to predict the behaviour of complex multicellular systems with many interacting parts. In this context, mathematical models provide a tool to assess the consistency of experimental observations with testable mechanistic hypotheses. In this review, we showcase examples from recent years of multidisciplinary investigations of neural crest cell migration. The neural crest model system has been used to study how collective migration of cell populations is shaped by cell-cell interactions, cell-environmental interactions and heterogeneity between cells. The wide range of emergent behaviours exhibited by neural crest cells in different embryonal locations and in different organisms helps us chart out the spectrum of collective cell migration. At the same time, this diversity in migratory characteristics highlights the need to reconcile or unify the array of currently hypothesized mechanisms through the next generation of experimental data and generalized theoretical descriptions. © 2016 The Authors.

  1. Increased C-C chemokine receptor 2 gene expression in monocytes of severe obstructive sleep apnea patients and under intermittent hypoxia.

    Science.gov (United States)

    Chuang, Li-Pang; Chen, Ning-Hung; Lin, Shih-Wei; Chang, Ying-Ling; Liao, Hsiang-Ruei; Lin, Yu-Sheng; Chao, I-Ju; Lin, Yuling; Pang, Jong-Hwei S

    2014-01-01

    Obstructive sleep apnea (OSA) is known to be a risk factor of coronary artery disease. The chemotaxis and adhesion of monocytes to the endothelium in the early atherosclerosis is important. This study aimed to investigate the effect of intermittent hypoxia, the hallmark of OSA, on the chemotaxis and adhesion of monocytes. Peripheral blood was sampled from 54 adults enrolled for suspected OSA. RNA was prepared from the isolated monocytes for the analysis of C-C chemokine receptor 2 (CCR2). The effect of intermittent hypoxia on the regulation and function of CCR2 was investigated on THP-1 monocytic cells and monocytes. The mRNA and protein expression levels were investigated by RT/real-time PCR and western blot analysis, respectively. Transwell filter migration assay and cell adhesion assay were performed to study the chemotaxis and adhesion of monocytes. Monocytic CCR2 gene expression was found to be increased in severe OSA patients and higher levels were detected after sleep. Intermittent hypoxia increased the CCR2 expression in THP-1 monocytic cells even in the presence of TNF-α and CRP. Intermittent hypoxia also promoted the MCP-1-mediated chemotaxis and adhesion of monocytes to endothelial cells. Furthermore, inhibitor for p42/44 MAPK or p38 MAPK suppressed the activation of monocytic CCR2 expression by intermittent hypoxia. This is the first study to demonstrate the increase of CCR2 gene expression in monocytes of severe OSA patients. Monocytic CCR2 gene expression can be induced under intermittent hypoxia which contributes to the chemotaxis and adhesion of monocytes.

  2. Migrating glioma cells express stem cell markers and give rise to new tumors upon xenografting

    DEFF Research Database (Denmark)

    Munthe, Sune; Sørensen, Mia D; Thomassen, Mads

    2016-01-01

    Glioblastoma (GBM) is the most frequent and malignant brain tumor with an overall survival of only 14.6 months. Although these tumors are treated with surgery, radiation and chemotherapy, recurrence is inevitable. A critical population of tumor cells in terms of therapy, the so-called cancer stem......-like phenotype is currently lacking. In the present study, the aim was to characterize the phenotype of migrating tumor cells using a novel migration assay based on serum-free stem cell medium and patient-derived spheroid cultures. The results showed pronounced migration of five different GBM spheroid cultures......-related genes and the HOX-gene list in migrating cells compared to spheroids. Determination of GBM molecular subtypes revealed that subtypes of spheroids and migrating cells were identical. In conclusion, migrating tumor cells preserve expression of stem cell markers and functional CSC characteristics. Since...

  3. Laser-photophoretic migration and fractionation of human blood cells

    Energy Technology Data Exchange (ETDEWEB)

    Monjushiro, Hideaki; Tanahashi, Yuko; Watarai, Hitoshi, E-mail: watarai@chem.sci.osaka-u.ac.jp

    2013-05-13

    Graphical abstract: -- Highlights: •RBCs were migrated faster than WBCs and blood pellets by laser photophoresis. •Photophoretic efficiency of RBC and WBC was simulated by the Mie scattering theory. •Spontaneous orientation of RBC parallel to the migration direction was elucidated. •Laser photophoretic separation of RBC and WBC was possible in a tip flow system. -- Abstract: Laser photophoretic migration behavior of human blood cells in saline solution was investigated under the irradiation of Nd:YAG laser beam (532 nm) in the absence and the presence of the flow in a fused silica capillary. Red blood cells (RBC) were migrated faster than white blood cells (WBC) and blood pellets to the direction of propagation of laser light. The observed photophoretic velocity of RBC was about 11 times faster than those of others. This was understood from the larger photophoretic efficiency of RBC than that of WBC, which was simulated based on the Mie scattering theory. Furthermore, it was found that, during the photophoretic migration, RBCs spontaneously orientated parallel to the migration direction so as to reduce the drag force. Finally, it was demonstrated that RBC and WBC were separated in a micro-channel flow system by the laser photophoresis.

  4. Laser-photophoretic migration and fractionation of human blood cells

    International Nuclear Information System (INIS)

    Monjushiro, Hideaki; Tanahashi, Yuko; Watarai, Hitoshi

    2013-01-01

    Graphical abstract: -- Highlights: •RBCs were migrated faster than WBCs and blood pellets by laser photophoresis. •Photophoretic efficiency of RBC and WBC was simulated by the Mie scattering theory. •Spontaneous orientation of RBC parallel to the migration direction was elucidated. •Laser photophoretic separation of RBC and WBC was possible in a tip flow system. -- Abstract: Laser photophoretic migration behavior of human blood cells in saline solution was investigated under the irradiation of Nd:YAG laser beam (532 nm) in the absence and the presence of the flow in a fused silica capillary. Red blood cells (RBC) were migrated faster than white blood cells (WBC) and blood pellets to the direction of propagation of laser light. The observed photophoretic velocity of RBC was about 11 times faster than those of others. This was understood from the larger photophoretic efficiency of RBC than that of WBC, which was simulated based on the Mie scattering theory. Furthermore, it was found that, during the photophoretic migration, RBCs spontaneously orientated parallel to the migration direction so as to reduce the drag force. Finally, it was demonstrated that RBC and WBC were separated in a micro-channel flow system by the laser photophoresis

  5. Low Doses of Curcuma longa Modulates Cell Migration and Cell-Cell Adhesion.

    Science.gov (United States)

    de Campos, Paloma Santos; Matte, Bibiana Franzen; Diel, Leonardo Francisco; Jesus, Luciano Henrique; Bernardi, Lisiane; Alves, Alessandro Menna; Rados, Pantelis Varvaki; Lamers, Marcelo Lazzaron

    2017-09-01

    Cell invasion and metastasis are involved in clinical failures in cancer treatment, and both events require the acquisition of a migratory behavior by tumor cells. Curcumin is a promising natural product with anti-proliferative activity, but its effects on cell migration are still unclear. We evaluated the effects of curcumin on the proliferation, apoptosis, migration, and cell-cell adhesion of keratinocyte, oral squamous cell carcinoma (OSCC), and fibroblast cell lines, as well as in a xenograft model of OSCC. Curcumin (2 μM) decreased cell proliferation in cell lines with mesenchymal characteristics, while cell death was detected only at 50 μM. We observed that highly migratory cells showed a decrease on migration speed and directionality when treated with 2 or 5 μM of curcumin (50% and 40%, respectively, p curcumin dose dependently decreased cell-cell adhesion, especially on tumor-derived spheroids. Also, in a xenograft model with patient-derived OSCC cells, the administration of curcumin decreased tumor growth and aggressiveness when compared with untreated tumors, indicating the potential antitumor effect in oral cancer. These results suggest that lower doses of curcumin can influence several steps involved in tumorigenesis, including migration properties, suggesting a possible use in cancer therapy. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  6. Synthetic antimicrobial and LPS-neutralising peptides suppress inflammatory and immune responses in skin cells and promote keratinocyte migration.

    Science.gov (United States)

    Pfalzgraff, Anja; Heinbockel, Lena; Su, Qi; Gutsmann, Thomas; Brandenburg, Klaus; Weindl, Günther

    2016-08-11

    The stagnation in the development of new antibiotics and the concomitant high increase of resistant bacteria emphasize the urgent need for new therapeutic options. Antimicrobial peptides are promising agents for the treatment of bacterial infections and recent studies indicate that Pep19-2.5, a synthetic anti-lipopolysaccharide (LPS) peptide (SALP), efficiently neutralises pathogenicity factors of Gram-negative (LPS) and Gram-positive (lipoprotein/-peptide, LP) bacteria and protects against sepsis. Here, we investigated the potential of Pep19-2.5 and the structurally related compound Pep19-4LF for their therapeutic application in bacterial skin infections. SALPs inhibited LP-induced phosphorylation of NF-κB p65 and p38 MAPK and reduced cytokine release and gene expression in primary human keratinocytes and dermal fibroblasts. In LPS-stimulated human monocyte-derived dendritic cells and Langerhans-like cells, the peptides blocked IL-6 secretion, downregulated expression of maturation markers and inhibited dendritic cell migration. Both SALPs showed a low cytotoxicity in all investigated cell types. Furthermore, SALPs markedly promoted cell migration via EGFR transactivation and ERK1/2 phosphorylation and accelerated artificial wound closure in keratinocytes. Peptide-induced keratinocyte migration was mediated by purinergic receptors and metalloproteases. In contrast, SALPs did not affect proliferation of keratinocytes. Conclusively, our data suggest a novel therapeutic target for the treatment of patients with acute and chronic skin infections.

  7. Automated Tracking of Cell Migration with Rapid Data Analysis.

    Science.gov (United States)

    DuChez, Brian J

    2017-09-01

    Cell migration is essential for many biological processes including development, wound healing, and metastasis. However, studying cell migration often requires the time-consuming and labor-intensive task of manually tracking cells. To accelerate the task of obtaining coordinate positions of migrating cells, we have developed a graphical user interface (GUI) capable of automating the tracking of fluorescently labeled nuclei. This GUI provides an intuitive user interface that makes automated tracking accessible to researchers with no image-processing experience or familiarity with particle-tracking approaches. Using this GUI, users can interactively determine a minimum of four parameters to identify fluorescently labeled cells and automate acquisition of cell trajectories. Additional features allow for batch processing of numerous time-lapse images, curation of unwanted tracks, and subsequent statistical analysis of tracked cells. Statistical outputs allow users to evaluate migratory phenotypes, including cell speed, distance, displacement, and persistence, as well as measures of directional movement, such as forward migration index (FMI) and angular displacement. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

  8. RNA of Enterococcus faecalis Strain EC-12 Is a Major Component Inducing Interleukin-12 Production from Human Monocytic Cells.

    Directory of Open Access Journals (Sweden)

    Ryoichiro Nishibayashi

    Full Text Available Interleukin-12 (IL-12 is an important cytokine for the immunomodulatory effects of lactic acid bacteria (LAB. Using murine immune cells, we previously reported that the RNA of Enterococcus faecalis EC-12, a LAB strain exerting probiotic-like beneficial effects, is the major IL-12-inducing immunogenic component. However, it was recently revealed that bacterial RNA can be a ligand for Toll-like receptor (TLR 13, which is only expressed in mice. Because TLR13 is not expressed in humans, the immuno-stimulatory and -modulatory effects of LAB RNA in human cells should be augmented excluding TLR13 contribution. In experiment 1 of this study, the role of LAB RNA in IL-12 induction in human immune cells was studied using three LAB strains, E.faecalis EC-12, Lactobacillus gasseri JCM5344, and Bifidobacterium breve JCM1192. RNase A treatment of heat-killed LAB significantly decreased the IL-12 production of human peripheral blood mononuclear cells on stimulation, while RNase III treatment revealed virtually no effects. Further, IL-12 production against heat-killed E. faecalis EC-12 was abolished by depleting monocytes. These results demonstrated that single stranded RNA (ssRNA of LAB is a strong inducer of IL-12 production from human monocytes. In experiment 2, major receptor for ssRNA of E. faecalis EC-12 was identified using THP-1 cells, a human monocytic cell line. The type of RNA molecules of E. faecalis EC-12 responsible for IL-12 induction was also identified. IL-12 production induced by the total RNA of E. faecalis EC-12 was significantly reduced by the treatment of siRNA for TLR8 but not for TLR7. Furthermore, both 23S and 16S rRNA, but not mRNA, of E. faecalis EC-12 markedly induced IL-12 production from THP-1 cells. These results suggested that the recognition of ssRNA of E. faecalis EC-12 was mediated by TLR8 and that rRNA was the RNA molecule that exhibited IL-12-inducing ability in human cells.

  9. Outside-in HLA class I signaling regulates ICAM-1 clustering and endothelial cell-monocyte interactions via mTOR in transplant antibody-mediated rejection.

    Science.gov (United States)

    Salehi, Sahar; Sosa, Rebecca A; Jin, Yi-Ping; Kageyama, Shoichi; Fishbein, Michael C; Rozengurt, Enrique; Kupiec-Weglinski, Jerzy W; Reed, Elaine F

    2018-05-01

    Antibody-mediated rejection (AMR) resulting in transplant allograft vasculopathy (TAV) is the major obstacle for long-term survival of solid organ transplants. AMR is caused by donor-specific antibodies to HLA, which contribute to TAV by initiating outside-in signaling transduction pathways that elicit monocyte recruitment to activated endothelium. Mechanistic target of rapamycin (mTOR) inhibitors can attenuate TAV; therefore, we sought to understand the mechanistic underpinnings of mTOR signaling in HLA class I Ab-mediated endothelial cell activation and monocyte recruitment. We used an in vitro model to assess monocyte binding to HLA I Ab-activated endothelial cells and found mTOR inhibition reduced ezrin/radixin/moesin (ERM) phosphorylation, intercellular adhesion molecule 1 (ICAM-1) clustering, and monocyte firm adhesion to HLA I Ab-activated endothelium. Further, in a mouse model of AMR, in which C57BL/6. RAG1 -/- recipients of BALB/c cardiac allografts were passively transferred with donor-specific MHC I antibodies, mTOR inhibition significantly reduced vascular injury, ERM phosphorylation, and macrophage infiltration of the allograft. Taken together, these studies indicate mTOR inhibition suppresses ERM phosphorylation in endothelial cells, which impedes ICAM-1 clustering in response to HLA class I Ab and prevents macrophage infiltration into cardiac allografts. These findings indicate a novel therapeutic application for mTOR inhibitors to disrupt endothelial cell-monocyte interactions during AMR. © 2017 The American Society of Transplantation and the American Society of Transplant Surgeons.

  10. Follow-the-leader cell migration requires biased cell–cell contact and local microenvironmental signals

    International Nuclear Information System (INIS)

    Wynn, Michelle L; Rupp, Paul; Trainor, Paul A; Kulesa, Paul M; Schnell, Santiago

    2013-01-01

    Directed cell migration often involves at least two types of cell motility that include multicellular streaming and chain migration. However, what is unclear is how cell contact dynamics and the distinct microenvironments through which cells travel influence the selection of one migratory mode or the other. The embryonic and highly invasive neural crest (NC) are an excellent model system to study this question since NC cells have been observed in vivo to display both of these types of cell motility. Here, we present data from tissue transplantation experiments in chick and in silico modeling that test our hypothesis that cell contact dynamics with each other and the microenvironment promote and sustain either multicellular stream or chain migration. We show that when premigratory cranial NC cells (at the pre-otic level) are transplanted into a more caudal region in the head (at the post-otic level), cells alter their characteristic stream behavior and migrate in chains. Similarly, post-otic NC cells migrate in streams after transplantation into the pre-otic hindbrain, suggesting that local microenvironmental signals dictate the mode of NC cell migration. Simulations of an agent-based model (ABM) that integrates the NC cell behavioral data predict that chain migration critically depends on the interplay of biased cell–cell contact and local microenvironment signals. Together, this integrated modeling and experimental approach suggests new experiments and offers a powerful tool to examine mechanisms that underlie complex cell migration patterns. (paper)

  11. [Effect of DOT1L gene silence on proliferation of acute monocytic leukemia cell line THP-1].

    Science.gov (United States)

    Zhang, Yu-Juan; Li, Hua-Wen; Chang, Guo-Qiang; Zhang, Hong-Ju; Wang, Jian; Lin, Ya-Ni; Zhou, Jia-Xi; Li, Qing-Hua; Pang, Tian-Xiang

    2013-08-01

    This study was aimed to investigate the influence of short hairpin RNA (shRNA) on proliferation of human leukemia cell line THP-1. The shRNA targeting the site 732-752 of DOT1L mRNA was designed and chemically synthesized, then a single-vector lentiviral, tet-inducible shRNA-DOT1L system (Plko-Tet-On) was generated. Thereafter, the THP-1 cells with lentivirus were infected to create stable cell line with regulatable shRNA expression. The expression of DOT1L in the THP-1 cell line was assayed by RT-PCR. Effect of shRNA-DOT1L on the proliferation of THP-1 cells was detected with MTT method,and the change of colony forming potential of THP-1 cells was analyzed by colony forming unit test. Cell cycle distribution was tested by flow cytometry. The results indicated that the expression of DOT1L was statistically lower than that in the control groups. The proliferation and colony forming capacity of THP-1 cells were significantly inhibited. The percentage of cells at G0/G1 phase increased in THP-1/shRNA cells treated with Dox while the percentage of cells at S phase significantly decreased as compared with that in the control group. It is concluded that the shRNA targeting DOT1L can effectively inhibit the proliferation of acute monocytic leukemia cell line THP-1.

  12. Lipopolysaccharide (LPS) stimulates fresh human monocytes to lyse actinomycin D-treated WEHI-164 target cells via increased secretion of a monokine similar to tumor necrosis factor

    International Nuclear Information System (INIS)

    Chen, A.R.; McKinnon, K.P.; Koren, H.S.

    1985-01-01

    The effects of lipopolysaccharide (LPS) on tumoricidal activity of human monocytes freshly isolated from peripheral blood were studied. Actinomycin D-treated WEHI-164 cells were used as targets because they are NK insensitive and are lysed rapidly by monocytes in 6-hr 51 Cr-release assays. Monocytes exhibited significant spontaneous activity without endotoxin. Monocytes either pretreated for 1 hr with LPS or assayed in the presence of LPS exhibited 100- to 1000-fold increased cytolytic activity. Cytolytic activity was heat labile and trypsin sensitive, and was recovered from Sepharose S-200 columns in a single peak with an apparent m.w. between 25,000 and 40,000. Actinomycin D or cycloheximide treatment of monocytes before the addition of LPS inhibited cytolytic monokine production. Cytolytic monokine activity was practically neutralized by specific rabbit antisera to human tumor necrosis factor (TNF). It was concluded that, although fresh human monocytes exhibit spontaneous tumoricidal activity, LPS is a potent activating agent. Its stimulatory effects depend on new transcription and translation and are mediated by enhanced secretion of a cytolytic monokine similar to TNF

  13. The effects of acoustic vibration on fibroblast cell migration.

    Science.gov (United States)

    Mohammed, Taybia; Murphy, Mark F; Lilley, Francis; Burton, David R; Bezombes, Frederic

    2016-12-01

    Cells are known to interact and respond to external mechanical cues and recent work has shown that application of mechanical stimulation, delivered via acoustic vibration, can be used to control complex cell behaviours. Fibroblast cells are known to respond to physical cues generated in the extracellular matrix and it is thought that such cues are important regulators of the wound healing process. Many conditions are associated with poor wound healing, so there is need for treatments/interventions, which can help accelerate the wound healing process. The primary aim of this research was to investigate the effects of mechanical stimulation upon the migratory and morphological properties of two different fibroblast cells namely; human lung fibroblast cells (LL24) and subcutaneous areolar/adipose mouse fibroblast cells (L929). Using a speaker-based system, the effects of mechanical stimulation (0-1600Hz for 5min) on the mean cell migration distance (μm) and actin organisation was investigated. The results show that 100Hz acoustic vibration enhanced cell migration for both cell lines whereas acoustic vibration above 100Hz was found to decrease cell migration in a frequency dependent manner. Mechanical stimulation was also found to promote changes to the morphology of both cell lines, particularly the formation of lamellipodia and filopodia. Overall lamellipodia was the most prominent actin structure displayed by the lung cell (LL24), whereas filopodia was the most prominent actin feature displayed by the fibroblast derived from subcutaneous areolar/adipose tissue. Mechanical stimulation at all the frequencies used here was found not to affect cell viability. These results suggest that low-frequency acoustic vibration may be used as a tool to manipulate the mechanosensitivity of cells to promote cell migration. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. A human monocytic NF-κB fluorescent reporter cell line for detection of microbial contaminants in biological samples.

    Directory of Open Access Journals (Sweden)

    Claire Battin

    Full Text Available Sensing of pathogens by innate immune cells is essential for the initiation of appropriate immune responses. Toll-like receptors (TLRs, which are highly sensitive for various structurally and evolutionary conserved molecules derived from microbes have a prominent role in this process. TLR engagement results in the activation of the transcription factor NF-κB, which induces the expression of cytokines and other inflammatory mediators. The exquisite sensitivity of TLR signalling can be exploited for the detection of bacteria and microbial contaminants in tissue cultures and in protein preparations. Here we describe a cellular reporter system for the detection of TLR ligands in biological samples. The well-characterized human monocytic THP-1 cell line was chosen as host for an NF-ᴋB-inducible enhanced green fluorescent protein reporter gene. We studied the sensitivity of the resultant reporter cells for a variety of microbial components and observed a strong reactivity towards TLR1/2 and TLR2/6 ligands. Mycoplasma lipoproteins are potent TLR2/6 agonists and we demonstrate that our reporter cells can be used as reliable and robust detection system for mycoplasma contaminations in cell cultures. In addition, a TLR4-sensitive subline of our reporters was engineered, and probed with recombinant proteins expressed in different host systems. Bacterially expressed but not mammalian expressed proteins induced strong reporter activity. We also tested proteins expressed in an E. coli strain engineered to lack TLR4 agonists. Such preparations also induced reporter activation in THP-1 cells highlighting the importance of testing recombinant protein preparations for microbial contaminations beyond endotoxins. Our results demonstrate the usefulness of monocytic reporter cells for high-throughput screening for microbial contaminations in diverse biological samples, including tissue culture supernatants and recombinant protein preparations. Fluorescent reporter

  15. Stimulation of monocytes by placental microparticles involves Toll-like receptors and nuclear factor kappa-light-chain-enhancer of activated B cells

    Directory of Open Access Journals (Sweden)

    Marianne Simone Joerger-Messerli

    2014-04-01

    Full Text Available Human pregnancy is accompanied by a mild systemic inflammatory response, which includes the activation of monocytes circulating in maternal blood. This response is exaggerated in preeclampsia, a placental-dependent disorder specific to human pregnancies. We and others showed that placental syncytiotrophoblast membrane microparticles (STBM generated in vitro from normal placentas stimulated peripheral blood monocytes, which suggests a contribution of STBM to the systemic maternal inflammation. Here, we analyzed the inflammatory potential of STBM prepared from preeclamptic placentas on primary monocytes and investigated the mode of action in vitro.STBM generated in vitro by placental villous explants of normal or preeclamptic placentas were co-incubated with human peripheral blood monocytes. In some cases, inhibitors of specific cellular functions or signaling pathways were used. The analysis of the monocytic response was performed by flow cytometry, enzyme-linked immunoassays, real-time PCR and fluorescence microscopy.STBM derived from preeclamptic placentas up-regulated the cell surface expression of CD54, and stimulated the secretion of the pro-inflammatory interleukin (IL-6 and IL-8 in a similar, dose-dependent manner as did STBM prepared from normal placentas. STBM bound to the cell surface of monocytes, but phagocytosis was not necessary for activation. STBM-induced cytokine secretion was impaired in the presence of inhibitors of toll-like receptor (TLR signaling or when nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB activation was blocked.Our results suggest that the inflammatory reaction in monocytes may be initiated by the interaction of STBM with TLRs, which in turn signal through NF-κB to mediate the transcription of genes coding for pro-inflammatory factors.

  16. Inflammatory Ly6Chigh Monocytes Protect against Candidiasis through IL-15-Driven NK Cell/Neutrophil Activation.

    Science.gov (United States)

    Domínguez-Andrés, Jorge; Feo-Lucas, Lidia; Minguito de la Escalera, María; González, Leticia; López-Bravo, María; Ardavín, Carlos

    2017-06-20

    Neutrophils play a crucial role in defense against systemic candidiasis, a disease associated with a high mortality rate in patients receiving immunosuppressive therapy, although the early immune mechanisms that boost the candidacidal activity of neutrophils remain to be defined in depth. Here, we used a murine model of systemic candidiasis to explore the role of inflammatory Ly6C high monocytes in NK cell-mediated neutrophil activation during the innate immune response against C. albicans. We found that efficient anti-Candida immunity required a collaborative response between the spleen and kidney, which relied on type I interferon-dependent IL-15 production by spleen inflammatory Ly6C high monocytes to drive efficient activation and GM-CSF release by spleen NK cells; this in turn was necessary to boost the Candida killing potential of kidney neutrophils. Our findings unveil a role for IL-15 as a critical mediator in defense against systemic candidiasis and hold promise for the design of IL-15-based antifungal immunotherapies. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. 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.

  18. Immunomodulatory effects of testosterone evaluated in all-trans retinoic acid differentiated HL-60 cells, granulocytes, and monocytes

    DEFF Research Database (Denmark)

    Boje, Alex; Moesby, Lise; Timm, Michael

    2012-01-01

    The sex hormones are known to affect innate immunity in humans. In this study we evaluated the immunomodulatory effects of testosterone in a model system comprising of all-trans retinoic acid differentiated HL-60 cells, and confirmed the results in human granulocytes and monocytes. Results showed...... that testosterone at pharmacological doses reduced the production of interleukin-8 and reactive oxygen species from differentiated HL-60 cells in a concentration dependent manner without affecting phagocytosis. The cells were stimulated with zymosan, lipopolysaccharide, or Bacillus subtilis. At the highest...... concentration of testosterone (120 µM), interleukin-8 secretion was reduced 42-80%, and production of reactive oxygen species was reduced 32-46%. Flutamide, an antagonist of the classical intracellular androgen receptor, was unable to antagonize the immunosuppressive effect of testosterone. We further...

  19. The distribution of IL-13 receptor alpha1 expression on B cells, T cells and monocytes and its regulation by IL-13 and IL-4.

    Science.gov (United States)

    Graber, P; Gretener, D; Herren, S; Aubry, J P; Elson, G; Poudrier, J; Lecoanet-Henchoz, S; Alouani, S; Losberger, C; Bonnefoy, J Y; Kosco-Vilbois, M H; Gauchat, J F

    1998-12-01

    To study the expression of IL-13 receptor alpha1 (IL-13Ralpha1), specific monoclonal antibodies (mAb) were generated. Surface expression of the IL-13Ralpha1 on B cells, monocytes and T cells was assessed by flow cytometry using these specific mAb. Among tonsillar B cells, the expression was the highest on the IgD+ CD38- B cell subpopulation which is believed to represent naive B cells. Expression was also detectable on a large fraction of the IgD-CD38- B cells but not on CD38+ B cells. Activation under conditions which promote B cell Ig class switching up-regulated the expression of the receptor. However, the same stimuli had an opposite effect for IL-13Ralpha1 expression levels on monocytes. While IL-13Ralpha1 mRNA was clearly detectable in T cell preparations, no surface expression was detected. However, permeabilization of the T cells showed a clear intracellular expression of the receptor. A soluble form of the receptor was immunoprecipitated from the supernatant of activated peripheral T cells, suggesting that T cell IL-13Ralpha1 might have functions unrelated to the capacity to form a type II IL-4/IL-13R with IL-4Ralpha.

  20. Performance comparison of two microarray platforms to assess differential gene expression in human monocyte and macrophage cells

    Directory of Open Access Journals (Sweden)

    Montalescot Gilles

    2008-06-01

    Full Text Available Abstract Background In this study we assessed the respective ability of Affymetrix and Illumina microarray methodologies to answer a relevant biological question, namely the change in gene expression between resting monocytes and macrophages derived from these monocytes. Five RNA samples for each type of cell were hybridized to the two platforms in parallel. In addition, a reference list of differentially expressed genes (DEG was generated from a larger number of hybridizations (mRNA from 86 individuals using the RNG/MRC two-color platform. Results Our results show an important overlap of the Illumina and Affymetrix DEG lists. In addition, more than 70% of the genes in these lists were also present in the reference list. Overall the two platforms had very similar performance in terms of biological significance, evaluated by the presence in the DEG lists of an excess of genes belonging to Gene Ontology (GO categories relevant for the biology of monocytes and macrophages. Our results support the conclusion of the MicroArray Quality Control (MAQC project that the criteria used to constitute the DEG lists strongly influence the degree of concordance among platforms. However the importance of prioritizing genes by magnitude of effect (fold change rather than statistical significance (p-value to enhance cross-platform reproducibility recommended by the MAQC authors was not supported by our data. Conclusion Functional analysis based on GO enrichment demonstrates that the 2 compared technologies delivered very similar results and identified most of the relevant GO categories enriched in the reference list.

  1. Aging increases microglial proliferation, delays cell migration, and decreases cortical neurogenesis after focal cerebral ischemia.

    Science.gov (United States)

    Moraga, Ana; Pradillo, Jesús M; García-Culebras, Alicia; Palma-Tortosa, Sara; Ballesteros, Ivan; Hernández-Jiménez, Macarena; Moro, María A; Lizasoain, Ignacio

    2015-05-10

    Aging is not just a risk factor of stroke, but it has also been associated with poor recovery. It is known that stroke-induced neurogenesis is reduced but maintained in the aged brain. However, there is no consensus on how neurogenesis is affected after stroke in aged animals. Our objective is to determine the role of aging on the process of neurogenesis after stroke. We have studied neurogenesis by analyzing proliferation, migration, and formation of new neurons, as well as inflammatory parameters, in a model of cerebral ischemia induced by permanent occlusion of the middle cerebral artery in young- (2 to 3 months) and middle-aged mice (13 to 14 months). Aging increased both microglial proliferation, as shown by a higher number of BrdU(+) cells and BrdU/Iba1(+) cells in the ischemic boundary and neutrophil infiltration. Interestingly, aging increased the number of M1 monocytes and N1 neutrophils, consistent with pro-inflammatory phenotypes when compared with the alternative M2 and N2 phenotypes. Aging also inhibited (subventricular zone) SVZ cell proliferation by decreasing both the number of astrocyte-like type-B (prominin-1(+)/epidermal growth factor receptor (EGFR)(+)/nestin(+)/glial fibrillary acidic protein (GFAP)(+) cells) and type-C cells (prominin-1(+)/EGFR(+)/nestin(-)/Mash1(+) cells), and not affecting apoptosis, 1 day after stroke. Aging also inhibited migration of neuroblasts (DCX(+) cells), as indicated by an accumulation of neuroblasts at migratory zones 14 days after injury; consistently, aged mice presented a smaller number of differentiated interneurons (NeuN(+)/BrdU(+) and GAD67(+) cells) in the peri-infarct cortical area 14 days after stroke. Our data confirm that stroke-induced neurogenesis is maintained but reduced in aged animals. Importantly, we now demonstrate that aging not only inhibits proliferation of specific SVZ cell subtypes but also blocks migration of neuroblasts to the damaged area and decreases the number of new interneurons in

  2. Cell Migration According to Shape of Graphene Oxide Micropatterns

    Directory of Open Access Journals (Sweden)

    Sung Eun Kim

    2016-10-01

    Full Text Available Photolithography is a unique process that can effectively manufacture micro/nano-sized patterns on various substrates. On the other hand, the meniscus-dragging deposition (MDD process can produce a uniform surface of the substrate. Graphene oxide (GO is the oxidized form of graphene that has high hydrophilicity and protein absorption. It is widely used in biomedical fields such as drug delivery, regenerative medicine, and tissue engineering. Herein, we fabricated uniform GO micropatterns via MDD and photolithography. The physicochemical properties of the GO micropatterns were characterized by atomic force microscopy (AFM, scanning electron microscopy (SEM, and Raman spectroscopy. Furthermore, cell migration on the GO micropatterns was investigated, and the difference in cell migration on triangle and square GO micropatterns was examined for their effects on cell migration. Our results demonstrated that the GO micropatterns with a desired shape can be finely fabricated via MDD and photolithography. Moreover, it was revealed that the shape of GO micropatterns plays a crucial role in cell migration distance, speed, and directionality. Therefore, our findings suggest that the GO micropatterns can serve as a promising biofunctional platform and cell-guiding substrate for applications to bioelectric devices, cell-on-a-chip, and tissue engineering scaffolds.

  3. Notch1-Dll4 signaling and mechanical force regulate leader cell formation during collective cell migration

    OpenAIRE

    Riahi, Reza; Sun, Jian; Wang, Shue; Long, Min; Zhang, Donna D.; Wong, Pak Kin

    2015-01-01

    At the onset of collective cell migration, a subset of cells within an initially homogenous population acquires a distinct “leader” phenotype with characteristic morphology and motility. However, the factors driving leader cell formation as well as the mechanisms regulating leader cell density during the migration process remain to be determined. Here, we use single cell gene expression analysis and computational modeling to show that leader cell identity is dynamically regulated by Dll4 sign...

  4. Vaccine adjuvant MF59 promotes the intranodal differentiation of antigen-loaded and activated monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Rossella Cioncada

    Full Text Available MF59 is an oil-in-water emulsion adjuvant approved for human influenza vaccination in European Union. The mode of action of MF59 is not fully elucidated yet, but results from several years of investigation indicate that MF59 establishes an immunocompetent environment at injection site which promotes recruitment of immune cells, including antigen presenting cells (APCs, that are facilitated to engulf antigen and transport it to draining lymph node (dLN where the antigen is accumulated. In vitro studies showed that MF59 promotes the differentiation of monocytes to dendritic cells (Mo-DCs. Since after immunization with MF59, monocytes are rapidly recruited both at the injection site and in dLN and appear to have a morphological change toward a DC-like phenotype, we asked whether MF59 could play a role in inducing differentiation of Mo-DC in vivo. To address this question we immunized mice with the auto-fluorescent protein Phycoerythrin (PE as model antigen, in presence or absence of MF59. We measured the APC phenotype and their antigen uptake within dLNs, the antigen distribution within the dLN compartments and the humoral response to PE. In addition, using Ovalbumin as model antigen, we measured the capacity of dLN APCs to induce antigen-specific CD4 T cell proliferation. Here, we show, for the first time, that MF59 promotes differentiation of Mo-DCs within dLNs from intranodal recruited monocytes and we suggest that this differentiation could take place in the medullary compartment of the LN. In addition we show that the Mo-DC subset represents the major source of antigen-loaded and activated APCs within the dLN when immunizing with MF59. Interestingly, this finding correlates with the enhanced triggering of antigen-specific CD4 T cell response induced by LN APCs. This study therefore demonstrates that MF59 is able to promote an immunocompetent environment also directly within the dLN, offering a novel insight on the mechanism of action of

  5. Microparticle Shedding by Erythrocytes, Monocytes and Vascular Smooth Muscular Cells Is Reduced by Aspirin in Diabetic Patients.

    Science.gov (United States)

    Chiva-Blanch, Gemma; Suades, Rosa; Padró, Teresa; Vilahur, Gemma; Peña, Esther; Ybarra, Juan; Pou, Jose M; Badimon, Lina

    2016-07-01

    Diabetes mellitus is associated with an enhanced risk for cardiovascular disease and its prevalence is increasing. Diabetes induces metabolic stress on blood and vascular cells, promoting platelet activation and vascular dysfunction. The level of vascular cell activation can be measured by the number and phenotype of microparticles found in the circulation. The aim of this study was to investigate the effect of a platelet-inhibitory dose of aspirin on the number and type of microparticles shed to the circulation. Forty-three diabetic patients were enrolled in the study and received a daily dose of 100mg of aspirin for 10 days to cover the average platelet life-span in the circulation. Before and after the intervention period, circulating microparticles were characterized and quantified by flow cytometry. Type 1 diabetic patients had about twice the number of tissue factor-positive circulating microparticles (derived both from platelets and monocytes) and endothelial-derived E-selectin positive microparticles than type 2 diabetic patients. Aspirin therapy significantly inhibited platelets since cyclooxygenase 1 derived thromboxane generation levels were reduced by 99%. Microparticles derived from erythrocytes, activated monocytes, and smooth muscle cells were significantly reduced after 10 days of aspirin administration. These results indicate that: a) vascular and blood cells in type 1 diabetic patients are exposed to more sustained stress shown by their specific microparticle origin and levels; b) aspirin therapy inhibits vascular wall cell activation and microparticle shedding, and c) the effects of aspirin are similar in type 1 and 2 diabetes. Copyright © 2016 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

  6. Evaluating the Use of Monocytes with a Degradable Polyurethane for Vascular Tissue Regeneration

    Science.gov (United States)

    Battiston, Kyle Giovanni

    Monocytes are one of the first cell types present following the implantation of a biomaterial or tissue engineered construct. Depending on the monocyte activation state supported by the biomaterial, monocytes and their derived macrophages (MDMs) can act as positive contributors to tissue regeneration and wound healing, or conversely promote a chronic inflammatory response that leads to fibrous encapsulation and implant rejection. A degradable polar hydrophobic iconic polyurethane (D-PHI) has been shown to reduce pro-inflammatory monocyte/macrophage response compared to tissue culture polystyrene (TCPS), a substrate routinely used for in vitro culture of cells, as well as poly(lactide- co-glycolide) (PLGA), a standard synthetic biodegradable biomaterial in the tissue engineering field. D-PHI has also shown properties suitable for use in a vascular tissue engineering context. In order to understand the mechanism through which D-PHI attenuates pro-inflammatory monocyte response, this thesis investigated the ability of D-PHI to modulate interactions with adsorbed serum proteins and the properties of D-PHI that were important for this activity. D-PHI was shown to regulate protein adsorption in a manner that produced divergent monocyte responses compared to TCPS and PLGA when coated with the serum proteins alpha2-macroglobulin or immunoglobulin G (IgG). In the case of IgG, D-PHI was shown to reduce pro-inflammatory binding site exposure as a function of the material's polar, hydrophobic, and ionic character. Due to the favourable monocyte activation state supported by D-PHI, and the importance of monocytes/macrophages in regulating the response of tissue-specific cell types in vivo, the ability of a D-PHI-stimulated monocyte/macrophage activation state to contribute to modulating the response of vascular smooth muscle cells (VSMCs) in a vascular tissue engineering context was investigated. D-PHI- stimulated monocytes promoted VSMC growth and migration through biomolecule

  7. Monoclonal antibody OKM5 inhibits the in vitro binding of Plasmodium falciparum-infected erythrocytes to monocytes, endothelial, and C32 melanoma cells

    International Nuclear Information System (INIS)

    Barnwell, J.W.; Ockenhouse, C.F.; Knowles, D.M. II

    1985-01-01

    Plasmodium falciparum-infected erythrocytes bind in vitro to human endothelial cells, monocytes, and a certain melanoma cell line. Evidence suggests that this interaction is mediated by similar mechanisms which lead to the sequestration of parasitized erythrocytes in vivo through their attachment to endothelial cells of small blood vessels. They show here the monoclonal antibody OKM5, previously shown to react with the membranes of endothelial cells, monocyte,s and platelets, also reacts with the C32 melanoma cell line which also binds P. falciparum-infected erythrocytes. At relatively low concentrations, OKM5 inhibits and reverses the in vitro adherence of infected erythrocytes to target cells. As with monocytes, OKM5 antibody recognizes an 125 I-labeled protein of approximately 88 Kd on the surface of C32 melanoma cells. It seems likely, therefore, that the 88 Kd polypeptide plays a role in cytoadherence, possibly as the receptor or part of a receptor for a ligand on the surface of infected erythrocytes

  8. Short-lived, transitory cell-cell interactions foster migration-dependent aggregation.

    Directory of Open Access Journals (Sweden)

    Melissa D Pope

    Full Text Available During embryonic development, motile cells aggregate into cohesive groups, which give rise to tissues and organs. The role of cell migration in regulating aggregation is unclear. The current paradigm for aggregation is based on an equilibrium model of differential cell adhesivity to neighboring cells versus the underlying substratum. In many biological contexts, however, dynamics is critical. Here, we provide evidence that multicellular aggregation dynamics involves both local adhesive interactions and transport by cell migration. Using time-lapse video microscopy, we quantified the duration of cell-cell contacts among migrating cells that collided and adhered to another cell. This lifetime of cell-cell interactions exhibited a monotonic decreasing dependence on substratum adhesivity. Parallel quantitative measurements of cell migration speed revealed that across the tested range of adhesive substrata, the mean time needed for cells to migrate and encounter another cell was greater than the mean adhesion lifetime, suggesting that aggregation dynamics may depend on cell motility instead of the local differential adhesivity of cells. Consistent with this hypothesis, aggregate size exhibited a biphasic dependence on substratum adhesivity, matching the trend we observed for cell migration speed. Our findings suggest a new role for cell motility, alongside differential adhesion, in regulating developmental aggregation events and motivate new design principles for tuning aggregation dynamics in tissue engineering applications.

  9. Identification of proangiogenic TIE2-expressing monocytes (TEMs) in human peripheral blood and cancer.

    Science.gov (United States)

    Venneri, Mary Anna; De Palma, Michele; Ponzoni, Maurilio; Pucci, Ferdinando; Scielzo, Cristina; Zonari, Erika; Mazzieri, Roberta; Doglioni, Claudio; Naldini, Luigi

    2007-06-15

    Tumor-infiltrating myeloid cells, including tumor-associated macrophages (TAMs), have been implicated in tumor progression. We recently described a lineage of mouse monocytes characterized by expression of the Tie2 angiopoietin receptor and required for the vascularization and growth of several tumor models. Here, we report that TIE2 expression in human blood identifies a subset of monocytes distinct from classical inflammatory monocytes and comprised within the less abundant "resident" population. These TIE2-expressing monocytes (TEMs) accounted for 2% to 7% of blood mononuclear cells in healthy donors and were distinct from rare circulating endothelial cells and progenitors. In human cancer patients, TEMs were observed in the blood and, intriguingly, within the tumors, where they represented the main monocyte population distinct from TAMs. Conversely, TEMs were hardly detected in nonneoplastic tissues. In vitro, TEMs migrated toward angiopoietin-2, a TIE2 ligand released by activated endothelial cells and angiogenic vessels, suggesting a homing mechanism for TEMs to tumors. Purified human TEMs, but not TEM-depleted monocytes, markedly promoted angiogenesis in xenotransplanted human tumors, suggesting a potentially critical role of TEMs in human cancer progression. Human TEMs may provide a novel, biologically relevant marker of angiogenesis and represent a previously unrecognized target of cancer therapy.

  10. Plectin deficiency in liver cancer cells promotes cell migration and sensitivity to sorafenib treatment.

    Science.gov (United States)

    Cheng, Chiung-Chi; Chao, Wei-Ting; Liao, Chen-Chun; Tseng, Yu-Hui; Lai, Yen-Chang Clark; Lai, Yih-Shyong; Hsu, Yung-Hsiang; Liu, Yi-Hsiang

    2018-01-02

    Plectin involved in activation of kinases in cell signaling pathway and plays important role in cell morphology and migration. Plectin knockdown promotes cell migration by activating focal adhesion kinase and Rac1-GTPase activity in liver cells. Sorafenib is a multi-targeting tyrosine kinase inhibitor that improves patient survival on hepatocellular carcinoma. The aim of this study is to investigate the correlation between the expression of plectin and cell migration as well as the sensitivity of hepatoma cell lines exposing to sorafenib. Hepatoma cell lines PLC/PRF/5 and HepG2 were used to examine the level of plectin expression and cell migration in comparison with Chang liver cell line. In addition, sensitivity of the 3 cell lines to sorafenib treatment was also measured. Expression of plectin was lower in PLC/PRF/5 and HepG2 hepatoma cells than that of Chang liver cells whereas HepG2 and PLC/PRF/5 cells exhibit higher rate of cell migration in trans-well migration assay. Immunohistofluorecent staining on E-cadherin revealed the highest rate of collective cell migration in HepG2 cells and the lowest was found in Chang liver cells. Likewise, HepG2 cell line was most sensitive to sorafenib treatment and Chang liver cells exhibited the least sensitivity. The drug sensitivity to sorafenib treatment showed inverse correlation with the expression of plectin. We suggest that plectin deficiency and increased E-cadherin in hepatoma cells were associated with higher rates of cell motility, collective cell migration as well as higher drug sensitivity to sorafenib treatment.

  11. Role of laminin receptor in tumor cell migration

    DEFF Research Database (Denmark)

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

    1987-01-01

    Polyclonal antisera were made against biochemically purified laminin receptor protein as well as against synthetic peptides deduced from a complementary DNA clone corresponding to the COOH-terminal end of the laminin receptor (U.M. Wewer et al., Proc. Natl. Acad. Sci. USA, 83: 7137-7141, 1986......). 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...

  12. Th1/M1 conversion to Th2/M2 responses in models of inflammation lacking cell death stimulates maturation of monocyte precursors to fibroblasts

    Directory of Open Access Journals (Sweden)

    JoAnn eTrial

    2013-09-01

    Full Text Available We have demonstrated that cardiac fibrosis arises from the differentiation of monocyte-derived fibroblasts. We present here evidence that this process requires sequential Th1 and Th2 induction promoting analogous M1 (classically activated and M2 (alternatively activated macrophage polarity. Our models are 1 mice subjected to daily repetitive ischemia reperfusion (I/R without infarction and 2 the in vitro transmigration of human mononuclear leukocytes through human cardiac microvascular endothelium. In the mouse heart, leukocytes entered after I/R in response to monocyte chemoattractant protein-1 (MCP-1 which is the major cytokine induced by this protocol. Monocytes within the heart then differentiated into fibroblasts making collagen while bearing the markers of M2 macrophages. T cells were seen in these hearts as well as in the human heart with cardiomyopathy. In the in vitro model, transmigration of the leukocytes was likewise induced by MCP-1 and some monocytes matured into fibroblasts bearing M2 markers. In this model, the MCP-1 stimulus induced a transient Th1 and M1 response that developed into a predominately Th2 and M2 response. An increase in the Th2 product IL-13 was present in both the human and the mouse models, consistent with its known role in fibrosis. In these simplified models, in which there is no cell death to stimulate an anti-inflammatory response, there is nonetheless a resolution of inflammation enabling a profibrotic environment. This induces the maturation of monocyte precursors into fibroblasts.

  13. Umbilical Cord-derived Mesenchymal Stem Cells Instruct Monocytes Towards an IL10-producing Phenotype by Secreting IL6 and HGF

    Science.gov (United States)

    Deng, Yinan; Zhang, Yingcai; Ye, Linsen; Zhang, Tong; Cheng, Jintao; Chen, Guihua; Zhang, Qi; Yang, Yang

    2016-01-01

    Human UC-MSCs are regarded as an attractive alternative to BM-MSCs for clinical applications due to their easy preparation, higher proliferation and lower immunogenicity. However, the mechanisms underlying immune suppression by UC-MSCs are still unclear. We studied the mechanism of inhibition by UC-MSCs during the differentiation of monocytes into DCs and focused on the specific source and the role of the involved cytokines. We found that UC-MSCs suppressed monocyte differentiation into DCs and instructed monocytes towards other cell types, with clear decreases in the expression of co-stimulatory molecules, in the secretion of inflammatory factors and in allostimulatory capacity. IL6, HGF and IL10 might be involved in this process because they were detected at higher levels in a coculture system. UC-MSCs produce IL-6 and HGF, and neutralization of IL-6 and HGF reversed the suppressive effect of UC-MSCs. IL10 was not produced by UC-MSCs but was exclusively produced by monocytes after exposure to UC-MSCs, IL-6 or HGF. In summary, we found that the UC-MSC-mediated inhibitory effect was dependent on IL6 and HGF secreted by UC-MSCs and that this effect induced monocyte-derived cells to produce IL10, which might indirectly strengthen the suppressive effect of UC-MSCs. PMID:27917866

  14. Umbilical Cord-derived Mesenchymal Stem Cells Instruct Monocytes Towards an IL10-producing Phenotype by Secreting IL6 and HGF.

    Science.gov (United States)

    Deng, Yinan; Zhang, Yingcai; Ye, Linsen; Zhang, Tong; Cheng, Jintao; Chen, Guihua; Zhang, Qi; Yang, Yang

    2016-12-05

    Human UC-MSCs are regarded as an attractive alternative to BM-MSCs for clinical applications due to their easy preparation, higher proliferation and lower immunogenicity. However, the mechanisms underlying immune suppression by UC-MSCs are still unclear. We studied the mechanism of inhibition by UC-MSCs during the differentiation of monocytes into DCs and focused on the specific source and the role of the involved cytokines. We found that UC-MSCs suppressed monocyte differentiation into DCs and instructed monocytes towards other cell types, with clear decreases in the expression of co-stimulatory molecules, in the secretion of inflammatory factors and in allostimulatory capacity. IL6, HGF and IL10 might be involved in this process because they were detected at higher levels in a coculture system. UC-MSCs produce IL-6 and HGF, and neutralization of IL-6 and HGF reversed the suppressive effect of UC-MSCs. IL10 was not produced by UC-MSCs but was exclusively produced by monocytes after exposure to UC-MSCs, IL-6 or HGF. In summary, we found that the UC-MSC-mediated inhibitory effect was dependent on IL6 and HGF secreted by UC-MSCs and that this effect induced monocyte-derived cells to produce IL10, which might indirectly strengthen the suppressive effect of UC-MSCs.

  15. L-Cysteine in vitro can restore cellular glutathione and inhibits the expression of cell adhesion molecules in G6PD-deficient monocytes.

    Science.gov (United States)

    Parsanathan, Rajesh; Jain, Sushil K

    2018-04-06

    L-Cysteine is a precursor of glutathione (GSH), a potent physiological antioxidant. Excess glucose-6-phosphate dehydrogenase (G6PD) deficiency in African Americans and low levels of L-cysteine diet in Hispanics can contributes to GSH deficiency and oxidative stress. Oxidative stress and monocyte adhesion was considered to be an initial event in the progression of vascular dysfunction and atherosclerosis. However, no previous study has investigated the contribution of GSH/G6PD deficiency to the expression of monocyte adhesion molecules. Using human U937 monocytes, this study examined the effect of GSH/G6PD deficiency and L-cysteine supplementation on monocyte adhesion molecules. G6PD/GSH deficiency induced by either siRNA or inhibitors (6AN/BSO, respectively) significantly (p adhesion molecules (ICAM-1, VCAM-1, SELL, ITGB1 and 2); NADPH oxidase (NOX), reactive oxygen species (ROS) and MCP-1 were upregulated, and decreases in levels of GSH, and nitric oxide were observed. The expression of ICAM-1 and VCAM-1 mRNA levels increased in high glucose, MCP-1 or TNF-α-treated G6PD-deficient compared to G6PD-normal cells. L-Cysteine treatment significantly (p adhesion molecules. Thus, GSH/G6PD deficiency increases susceptibility to monocyte adhesion processes, whereas L-cysteine supplementation can restore cellular GSH/G6PD and attenuates NOX activity and expression of cell adhesion molecules.

  16. Reciprocal control of cell proliferation and migration

    Directory of Open Access Journals (Sweden)

    De Donatis Alina

    2010-09-01

    Full Text Available Abstract In adult tissue the quiescent state of a single cell is maintained by the steady state conditions of its own microenvironment for what concern both cell-cell as well as cell-ECM interaction and soluble factors concentration. Physiological or pathological conditions can alter this quiescent state through an imbalance of both soluble and insoluble factors that can trigger a cellular phenotypic response. The kind of cellular response depends by many factors but one of the most important is the concentration of soluble cytokines sensed by the target cell. In addition, due to the intrinsic plasticity of many cellular types, every single cell is able, in response to the same stimulus, to rapidly switch phenotype supporting minimal changes of microenviromental cytokines concentration. Wound healing is a typical condition in which epithelial, endothelial as well as mesenchymal cells are firstly subjected to activation of their motility in order to repopulate the damaged region and then they show a strong proliferative response in order to successfully complete the wound repair process. This schema constitute the leitmotif of many other physiological or pathological conditions such as development vasculogenesis/angiogenesis as well as cancer outgrowth and metastasis. Our review focuses on the molecular mechanisms that control the starting and, eventually, the switching of cellular phenotypic outcome in response to changes in the symmetry of the extracellular environment.

  17. Supernatants from oral epithelial cells and gingival fibroblasts modulate human immunodeficiency virus type 1 promoter activation induced by periodontopathogens in monocytes/macrophages.

    Science.gov (United States)

    González, O A; Ebersole, J L; Huang, C B

    2010-04-01

    Bacterial and host cell products during coinfections of Human Immunodeficiency Virus type 1-positive (HIV-1(+)) patients regulate HIV-1 recrudescence in latently infected cells (e.g. T cells, monocytes/macrophages), impacting highly active antiretroviral therapy (HAART) failure and progression of acquired immunodeficiency syndrome. A high frequency of oral opportunistic infections (e.g. periodontitis) in HIV-1(+) patients has been demonstrated; however, their potential to impact HIV-1 exacerbation is unclear. We sought to determine the ability of supernatants derived from oral epithelial cells (OKF4) and human gingival fibroblasts (Gin-4) challenged with periodontal pathogens, to modulate the HIV-1 promoter activation in monocytes/macrophages. BF24 monocytes/macrophages transfected with the HIV-1 promoter driving the expression of chloramphenicol acetyltransferase (CAT) were stimulated with Porphyromonas gingivalis, Fusobacterium nucleatum, or Treponema denticola in the presence of supernatants from OKF4 or Gin4 cells either unstimulated or previously pulsed with bacteria. CAT levels were determined by enzyme-linked immunosorbent assay and cytokine production was evaluated by Luminex beadlyte assays. OKF4 and Gin4 supernatants enhanced HIV-1 promoter activation particularly related to F. nucleatum challenge. An additive effect was observed in HIV-1 promoter activation when monocytes/macrophages were simultaneously stimulated with gingival cell supernatants and bacterial extracts. OKF4 cells produced higher levels of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukins -6 and -8 in response to F. nucleatum and P. gingivalis. Preincubation of OKF4 supernatants with anti-GM-CSF reduced the additive effect in periodontopathogen-induced HIV-1 promoter activation. These results suggest that soluble mediators produced by gingival resident cells in response to periodontopathogens could contribute to HIV-1 promoter activation in monocytes

  18. Functional role of CD11c+ monocytes in atherogenesis associated with hypercholesterolemia

    Science.gov (United States)

    Monocyte activation and migration into the arterial wall are key events in atherogenesis associated with hypercholesterolemia. CD11c/CD18, a beta2 integrin expressed on human monocytes and a subset of mouse monocytes, has been shown to play a distinct role in human monocyte adhesion on endothelial c...

  19. An automated cell-counting algorithm for fluorescently-stained cells in migration assays

    Directory of Open Access Journals (Sweden)

    Novielli Nicole M

    2011-10-01

    Full Text Available Abstract A cell-counting algorithm, developed in Matlab®, was created to efficiently count migrated fluorescently-stained cells on membranes from migration assays. At each concentration of cells used (10,000, and 100,000 cells, images were acquired at 2.5 ×, 5 ×, and 10 × objective magnifications. Automated cell counts strongly correlated to manual counts (r2 = 0.99, P

  20. Migrastatin analogues inhibit canine mammary cancer cell migration and invasion.

    Directory of Open Access Journals (Sweden)

    Kinga Majchrzak

    Full Text Available BACKGROUND: Cancer spread to other organs is the main cause of death of oncological patients. Migration of cancer cells from a primary tumour is the crucial step in the complex process of metastasis, therefore blocking this process is currently the main treatment strategy. Metastasis inhibitors derived from natural products, such as, migrastatin, are very promising anticancer agents. Thus, the aim of our study was to investigate the effect of six migrastatin analogues (MGSTA-1 to 6 on migration and invasion of canine mammary adenocarcinoma cell lines isolated from primary tumours and their metastases to the lungs. Canine mammary tumours constitute a valuable tool for studying multiple aspect of human cancer. RESULTS: OUR RESULTS SHOWED THAT TWO OF SIX FULLY SYNTHETIC ANALOGUES OF MIGRASTATIN: MGSTA-5 and MGSTA-6 were potent inhibitors of canine mammary cancer cells migration and invasion. These data were obtained using the wound healing test, as well as trans-well migration and invasion assays. Furthermore, the treatment of cancer cells with the most effective compound (MGSTA-6 disturbed binding between filamentous F-actin and fascin1. Confocal microscopy analyses revealed that treatment with MGSTA-6 increased the presence of unbound fascin1 and reduced co-localization of F-actin and fascin1 in canine cancer cells. Most likely, actin filaments were not cross-linked by fascin1 and did not generate the typical filopodial architecture of actin filaments in response to the activity of MGSTA-6. Thus, administration of MGSTA-6 results in decreased formation of filopodia protrusions and stress fibres in canine mammary cancer cells, causing inhibition of cancer migration and invasion. CONCLUSION: Two synthetic migrastatin analogues (MGSTA-5 and MGSTA-6 were shown to be promising compounds for inhibition of cancer metastasis. They may have beneficial therapeutic effects in cancer therapy in dogs, especially in combination with other anticancer drugs

  1. Molecular mechanisms governing primordial germ cell migration in zebrafish

    NARCIS (Netherlands)

    Doitsidou, M.

    2005-01-01

    In most sexually reproducing organisms primordial germ cells (pGCs) are specified early in development in places that are distinct from the region where the somatic part of the gonad develops. From their places of specification they have to migrate towards the site where they associate with somatic

  2. Emerging role for nuclear rotation and orientation in cell migration

    Czech Academy of Sciences Publication Activity Database

    Maninová, Miloslava; Iwanicki, M. P.; Vomastek, Tomáš

    2014-01-01

    Roč. 8, č. 1 (2014), s. 42-48 ISSN 1933-6918 R&D Projects: GA ČR GA204/09/0614 Grant - others:Marie Cúrie EU FP7(BE) 231086 Institutional support: RVO:61388971 Keywords : cell polarity * actin * migration * microtubules Subject RIV: EE - Microbiology, Virology Impact factor: 4.505, year: 2014

  3. STRIPAK components determine mode of cancer cell migration and metastasis

    DEFF Research Database (Denmark)

    Madsen, Chris D; Hooper, Steven; Tozluoglu, Melda

    2015-01-01

    demonstrate that co-localization of contractile activity and actin-plasma membrane linkage reduces cell speed on planar surfaces, but favours migration in confined environments similar to those observed in vivo. We further show that FAM40B mutations found in human tumours uncouple it from PP2A and enable...

  4. URG11 Regulates Prostate Cancer Cell Proliferation, Migration, and Invasion

    Directory of Open Access Journals (Sweden)

    Bin Pan

    2018-01-01

    Full Text Available Upregulated gene 11 (URG11, a new gene upregulated by hepatitis B virus X protein, is involved in the development and progression of several tumors, including liver, stomach, lung, and colon cancers. However, the role of URG11 in prostate cancer remains yet to be elucidated. By determined expression in human prostate cancer tissues, URG11 was found significantly upregulated and positively correlated with the severity of prostate cancer, compared with that in benign prostatic hyperplasia tissues. Further, the mRNA and protein levels of URG11 were significantly upregulated in human prostate cancer cell lines (DU145, PC3, and LNCaP, compared with human prostate epithelial cell line (RWPE-1. Moreover, by the application of siRNA against URG11, the proliferation, migration, and invasion of prostate cancer cells were markedly inhibited. Genetic knockdown of URG11 also induced cell cycle arrest at G1/S phase, induced apoptosis, and decreased the expression level of β-catenin in prostate cancer cells. Overexpression of URG11 promoted the expression of β-catenin, the growth, the migration, and invasion ability of prostate cancer cells. Taken together, this study reveals that URG11 is critical for the proliferation, migration, and invasion in prostate cancer cells, providing the evidence of URG11 to be a novel potential therapeutic target of prostate cancer.

  5. Endogenous electric fields as guiding cue for cell migration

    Science.gov (United States)

    Funk, Richard H. W.

    2015-01-01

    This review covers two topics: (1) “membrane potential of low magnitude and related electric fields (bioelectricity)” and (2) “cell migration under the guiding cue of electric fields (EF).”Membrane potentials for this “bioelectricity” arise from the segregation of charges by special molecular machines (pumps, transporters, ion channels) situated within the plasma membrane of each cell type (including eukaryotic non-neural animal cells). The arising patterns of ion gradients direct many cell- and molecular biological processes such as embryogenesis, wound healing, regeneration. Furthermore, EF are important as guiding cues for cell migration and are often overriding chemical or topographic cues. In osteoblasts, for instance, the directional information of EF is captured by charged transporters on the cell membrane and transferred into signaling mechanisms that modulate the cytoskeleton and motor proteins. This results in a persistent directional migration along an EF guiding cue. As an outlook, we discuss questions concerning the fluctuation of EF and the frequencies and mapping of the “electric” interior of the cell. Another exciting topic for further research is the modeling of field concepts for such distant, non-chemical cellular interactions. PMID:26029113

  6. Impact of cell shape on cell migration behavior on elastic substrate

    International Nuclear Information System (INIS)

    Zhong Yuan; Ji Baohua

    2013-01-01

    Cell shape is known to have profound effects on a number of cell behaviors. In this paper we have studied its role in cell migration through modeling the effect of cell shape on the cell traction force distribution, the traction force dependent stability of cell adhesion and the matrix rigidity dependent traction force formation. To quantify the driving force of cell migration, a new parameter called the motility factor, that takes account of the effect of cell shape, matrix rigidity and dynamic stability of cell adhesion, is proposed. We showed that the motility factor depends on the matrix rigidity in a biphasic manner, which is consistent with the experimental observations of the biphasic dependence of cell migration speed on the matrix rigidity. We showed that the cell shape plays a pivotal role in the cell migration behavior by regulating the traction force at the cell front and rear. The larger the cell polarity, the larger the motility factor is. The keratocyte-like shape has a larger motility factor than the fibroblast-like shape, which explains why keratocyte has a much higher migration speed. The motility factor might be an appropriate parameter for a quantitative description of the driving force of cell migration. (paper)

  7. Alcohol and cannabinoids differentially affect HIV infection and function of human monocyte-derived dendritic cells (MDDC

    Directory of Open Access Journals (Sweden)

    Marisela eAgudelo

    2015-12-01

    Full Text Available During human immunodeficiency virus (HIV infection, alcohol has been known to induce inflammation while cannabinoids have been shown to have an anti-inflammatory role. For instance cannabinoids have been shown to reduce susceptibility to HIV-1 infection and attenuate HIV replication in macrophages. Recently, we demonstrated that alcohol induces cannabinoid receptors and regulates cytokine production by monocyte-derived dendritic cells (MDDC. However, the ability of alcohol and cannabinoids to alter MDDC function during HIV infection has not been clearly elucidated yet. In order to study the potential impact of alcohol and cannabinoids on differentiated MDDC infected with HIV, monocytes were cultured for 7 days with GM-CSF and IL-4, differentiated MDDC were infected with HIV-1Ba-L and treated with EtOH (0.1 and 0.2%, THC (5 and 10 uM, or JWH-015 (5 and 10 uM for 4-7 days. HIV infection of MDDC was confirmed by p24 and Long Terminal Repeats (LTR estimation. MDDC endocytosis assay and cytokine array profiles were measured to investigate the effects of HIV and substances of abuse on MDDC function. Our results show the HIV+EtOH treated MDDC had the highest levels of p24 production and expression when compared with the HIV positive controls and the cannabinoid treated cells. Although both cannabinoids, THC and JWH-015 had lower levels of p24 production and expression, the HIV+JWH-015 treated MDDC had the lowest levels of p24 when compared to the HIV+THC treated cells. In addition, MDDC endocytic function and cytokine production were also differentially altered after alcohol and cannabinoid treatments. Our results show a differential effect of alcohol and cannabinoids, which may provide insights into the divergent inflammatory role of alcohol and cannabinoids to modulate MDDC function in the context of HIV infection.

  8. Dengue Virus-Infected Dendritic Cells, but Not Monocytes, Activate Natural Killer Cells through a Contact-Dependent Mechanism Involving Adhesion Molecules.

    Science.gov (United States)

    Costa, Vivian Vasconcelos; Ye, Weijian; Chen, Qingfeng; Teixeira, Mauro Martins; Preiser, Peter; Ooi, Eng Eong; Chen, Jianzhu

    2017-08-01

    Natural killer (NK) cells play a protective role against dengue virus (DENV) infection, but the cellular and molecular mechanisms are not fully understood. Using an optimized humanized mouse model, we show that human NK cells, through the secretion of gamma interferon (IFN-γ), are critical in the early defense against DENV infection. Depletion of NK cells or neutralization of IFN-γ leads to increased viremia and more severe thrombocytopenia and liver damage in humanized mice. In vitro studies using autologous human NK cells show that DENV-infected monocyte-derived dendritic cells (MDDCs), but not monocytes, activate NK cells in a contact-dependent manner, resulting in upregulation of CD69 and CD25 and secretion of IFN-γ. Blocking adhesion molecules (LFA-1, DNAM-1, CD2, and 2β4) on NK cells abolishes NK cell activation, IFN-γ secretion, and the control of DENV replication. NK cells activated by infected MDDCs also inhibit DENV infection in monocytes. These findings show the essential role of human NK cells in protection against acute DENV infection in vivo , identify adhesion molecules and dendritic cells required for NK cell activation, and delineate the sequence of events for NK cell activation and protection against DENV infection. IMPORTANCE Dengue is a mosquito-transmitted viral disease with a range of symptoms, from mild fever to life-threatening dengue hemorrhagic fever. The diverse disease manifestation is thought to result from a complex interplay between viral and host factors. Using mice engrafted with a human immune system, we show that human NK cells inhibit virus infection through secretion of the cytokine gamma interferon and reduce disease pathogenesis, including depletion of platelets and liver damage. During a natural infection, DENV initially infects dendritic cells in the skin. We find that NK cells interact with infected dendritic cells through physical contact mediated by adhesion molecules and become activated before they can control

  9. Suppression of NRF2–ARE activity sensitizes chemotherapeutic agent-induced cytotoxicity in human acute monocytic leukemia cells

    International Nuclear Information System (INIS)

    Peng, Hui; Wang, Huihui; Xue, Peng; Hou, Yongyong; Dong, Jian; Zhou, Tong; Qu, Weidong; Peng, Shuangqing; Li, Jin; Carmichael, Paul L.; Nelson, Bud; Clewell, Rebecca; Zhang, Qiang; Andersen, Melvin E.; Pi, Jingbo

    2016-01-01

    Nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of the antioxidant response element (ARE)-dependent transcription, plays a pivotal role in chemical detoxification in normal and tumor cells. Consistent with previous findings that NRF2–ARE contributes to chemotherapeutic resistance of cancer cells, we found that stable knockdown of NRF2 by lentiviral shRNA in human acute monocytic leukemia (AML) THP-1 cells enhanced the cytotoxicity of several chemotherapeutic agents, including arsenic trioxide (As 2 O 3 ), etoposide and doxorubicin. Using an ARE-luciferase reporter expressed in several human and mouse cells, we identified a set of compounds, including isonicotinic acid amides, isoniazid and ethionamide, that inhibited NRF2–ARE activity. Treatment of THP-1 cells with ethionamide, for instance, significantly reduced mRNA expression of multiple ARE-driven genes under either basal or As 2 O 3 -challenged conditions. As determined by cell viability and cell cycle, suppression of NRF2–ARE by ethionamide also significantly enhanced susceptibility of THP-1 and U937 cells to As 2 O 3 -induced cytotoxicity. In THP-1 cells, the sensitizing effect of ethionamide on As 2 O 3 -induced cytotoxicity was highly dependent on NRF2. To our knowledge, the present study is the first to demonstrate that ethionamide suppresses NRF2–ARE signaling and disrupts the transcriptional network of the antioxidant response in AML cells, leading to sensitization to chemotherapeutic agents. - Highlights: • Identification of novel inhibitors of ARE-dependent transcription • Suppression of NRF2–ARE sensitizes THP-1 cells to chemotherapy. • Ethionamide suppresses ARE-dependent transcriptional activity. • Ethionamide and isoniazid increase the cytotoxicity of As 2 O 3 in AML cells. • Sensitization of THP-1 cells to As 2 O 3 toxicity by ethionamide is NRF2-dependent.

  10. Complement-Mediated Enhancement of Monocyte Adhesion to Endothelial Cells by HLA Antibodies, and Blockade by a Specific Inhibitor of the Classical Complement Cascade, TNT003

    Science.gov (United States)

    Valenzuela, Nicole M.; Thomas, Kimberly A.; Mulder, Arend; Parry, Graham C.; Panicker, Sandip; Reed, Elaine F.

    2017-01-01

    Background Antibody-mediated rejection (AMR) of most solid organs is characterized by evidence of complement activation and/or intragraft macrophages (C4d + and CD68+ biopsies). We previously demonstrated that crosslinking of HLA I by antibodies triggered endothelial activation and monocyte adhesion. We hypothesized that activation of the classical complement pathway at the endothelial cell surface by HLA antibodies would enhance monocyte adhesion through soluble split product generation, in parallel with direct endothelial activation downstream of HLA signaling. Methods Primary human aortic endothelial cells (HAEC) were stimulated with HLA class I antibodies in the presence of intact human serum complement. C3a and C5a generation, endothelial P-selectin expression, and adhesion of human primary and immortalized monocytes (Mono Mac 6) were measured. Alternatively, HAEC or monocytes were directly stimulated with purified C3a or C5a. Classical complement activation was inhibited by pretreatment of complement with an anti-C1s antibody (TNT003). Results Treatment of HAEC with HLA antibody and human complement increased the formation of C3a and C5a. Monocyte recruitment by human HLA antibodies was enhanced in the presence of intact human serum complement or purified C3a or C5a. Specific inhibition of the classical complement pathway using TNT003 or C1q-depleted serum significantly reduced adhesion of monocytes in the presence of human complement. Conclusions Despite persistent endothelial viability in the presence of HLA antibodies and complement, upstream complement anaphylatoxin production exacerbates endothelial exocytosis and leukocyte recruitment. Upstream inhibition of classical complement may be therapeutic to dampen mononuclear cell recruitment and endothelial activation characteristic of microvascular inflammation during AMR. PMID:28640789

  11. Untangling cell tracks: Quantifying cell migration by time lapse image data analysis.

    Science.gov (United States)

    Svensson, Carl-Magnus; Medyukhina, Anna; Belyaev, Ivan; Al-Zaben, Naim; Figge, Marc Thilo

    2018-03-01

    Automated microscopy has given researchers access to great amounts of live cell imaging data from in vitro and in vivo experiments. Much focus has been put on extracting cell tracks from such data using a plethora of segmentation and tracking algorithms, but further analysis is normally required to draw biologically relevant conclusions. Such relevant conclusions may be whether the migration is directed or not, whether the population has homogeneous or heterogeneous migration patterns. This review focuses on the analysis of cell migration data that are extracted from time lapse images. We discuss a range of measures and models used to analyze cell tracks independent of the biological system or the way the tracks were obtained. For single-cell migration, we focus on measures and models giving examples of biological systems where they have been applied, for example, migration of bacteria, fibroblasts, and immune cells. For collective migration, we describe the model systems wound healing, neural crest migration, and Drosophila gastrulation and discuss methods for cell migration within these systems. We also discuss the role of the extracellular matrix and subsequent differences between track analysis in vitro and in vivo. Besides methods and measures, we are putting special focus on the need for openly available data and code, as well as a lack of common vocabulary in cell track analysis. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

  12. Efficient, long term production of monocyte-derived macrophages from human pluripotent stem cells under partly-defined and fully-defined conditions.

    Directory of Open Access Journals (Sweden)

    Bonnie van Wilgenburg

    Full Text Available Human macrophages are specialised hosts for HIV-1, dengue virus, Leishmania and Mycobacterium tuberculosis. Yet macrophage research is hampered by lack of appropriate cell models for modelling infection by these human pathogens, because available myeloid cell lines are, by definition, not terminally differentiated like tissue macrophages. We describe here a method for deriving monocytes and macrophages from human Pluripotent Stem Cells which improves on previously published protocols in that it uses entirely defined, feeder- and serum-free culture conditions and produces very consistent, pure, high yields across both human Embryonic Stem Cell (hESC and multiple human induced Pluripotent Stem Cell (hiPSC lines over time periods of up to one year. Cumulatively, up to ∼3×10(7 monocytes can be harvested per 6-well plate. The monocytes produced are most closely similar to the major blood monocyte (CD14(+, CD16(low, CD163(+. Differentiation with M-CSF produces macrophages that are highly phagocytic, HIV-1-infectable, and upon activation produce a pro-inflammatory cytokine profile similar to blood monocyte-derived macrophages. Macrophages are notoriously hard to genetically manipulate, as they recognise foreign nucleic acids; the lentivector system described here overcomes this, as pluripotent stem cells can be relatively simply genetically manipulated for efficient transgene expression in the differentiated cells, surmounting issues of transgene silencing. Overall, the method we describe here is an efficient, effective, scalable system for the reproducible production and genetic modification of human macrophages, facilitating the interrogation of human macrophage biology.

  13. Circulating microparticles in acute diabetic Charcot foot exhibit a high content of inflammatory cytokines, and support monocyte-to-osteoclast cell induction.

    Science.gov (United States)

    Pasquier, Jennifer; Thomas, Binitha; Hoarau-Véchot, Jessica; Odeh, Tala; Robay, Amal; Chidiac, Omar; Dargham, Soha R; Turjoman, Rebal; Halama, Anna; Fakhro, Khalid; Menzies, Robert; Jayyousi, Amin; Zirie, Mahmoud; Al Suwaidi, Jassim; Rafii, Arash; Malik, Rayaz A; Talal, Talal; Abi Khalil, Charbel

    2017-11-27

    Circulating microparticles (MPs) are major mediators in cardiovascular complications of type 2 diabetes (T2D); however, their contribution to Charcot foot (CF) disease is not known. Here, we purified and assessed the origin, concentration and content of circulating MPs from 33 individuals: 11 with T2D and acute CF, 11 T2D patients with equivalent neuropathy and 11 non-diabetic controls. First, we demonstrated that there were no differences in the distribution of MPs of endothelial, platelet origin among the 3 groups. However, MPs from leukocytes and monocytes origin were increased in CF patients. Moreover, we demonstrated that monocytes-derived MPs originated more frequently from intermediate and non-classical monocytes in CF patients. Five cytokines (G-CSF, GM-CSF, IL-1-ra, IL-2 and IL-16) were significantly increased in MPs from acute CF patients. Applying ingenuity pathways analysis, we found that those cytokines interacted well and induced the activation of pathways that are involved in osteoclast formation. Further, we treated THP-1 monocytes and monocytes sorted from healthy patients with CF-derived MPs during their differentiation into osteoclasts, which increased their differentiation into multinucleated osteoclast-like cells. Altogether, our study suggests that circulating MPs in CF disease have a high content of inflammatory cytokines and could increase osteoclast differentiation in vitro.

  14. Migration Phenotype of Brain-Cancer Cells Predicts Patient Outcomes

    Directory of Open Access Journals (Sweden)

    Chris L. Smith

    2016-06-01

    Full Text Available Glioblastoma multiforme is a heterogeneous and infiltrative cancer with dismal prognosis. Studying the migratory behavior of tumor-derived cell populations can be informative, but it places a high premium on the precision of in vitro methods and the relevance of in vivo conditions. In particular, the analysis of 2D cell migration may not reflect invasion into 3D extracellular matrices in vivo. Here, we describe a method that allows time-resolved studies of primary cell migration with single-cell resolution on a fibrillar surface that closely mimics in vivo 3D migration. We used this platform to screen 14 patient-derived glioblastoma samples. We observed that the migratory phenotype of a subset of cells in response to platelet-derived growth factor was highly predictive of tumor location and recurrence in the clinic. Therefore, migratory phenotypic classifiers analyzed at the single-cell level in a patient-specific way can provide high diagnostic and prognostic value for invasive cancers.

  15. Bleb Expansion in Migrating Cells Depends on Supply of Membrane from Cell Surface Invaginations.

    Science.gov (United States)

    Goudarzi, Mohammad; Tarbashevich, Katsiaryna; Mildner, Karina; Begemann, Isabell; Garcia, Jamie; Paksa, Azadeh; Reichman-Fried, Michal; Mahabaleshwar, Harsha; Blaser, Heiko; Hartwig, Johannes; Zeuschner, Dagmar; Galic, Milos; Bagnat, Michel; Betz, Timo; Raz, Erez

    2017-12-04

    Cell migration is essential for morphogenesis, organ formation, and homeostasis, with relevance for clinical conditions. The migration of primordial germ cells (PGCs) is a useful model for studying this process in the context of the developing embryo. Zebrafish PGC migration depends on the formation of cellular protrusions in form of blebs, a type of protrusion found in various cell types. Here we report on the mechanisms allowing the inflation of the membrane during bleb formation. We show that the rapid expansion of the protrusion depends on membrane invaginations that are localized preferentially at the cell front. The formation of these invaginations requires the function of Cdc42, and their unfolding allows bleb inflation and dynamic cell-shape changes performed by migrating cells. Inhibiting the formation and release of the invaginations strongly interfered with bleb formation, cell motility, and the ability of the cells to reach their target. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Evidence for tension-based regulation of Drosophila MAL and SRF during invasive cell migration.

    Science.gov (United States)

    Somogyi, Kálmán; Rørth, Pernille

    2004-07-01

    Cells migrating through a tissue exert force via their cytoskeleton and are themselves subject to tension, but the effects of physical forces on cell behavior in vivo are poorly understood. Border cell migration during Drosophila oogenesis is a useful model for invasive cell movement. We report that this migration requires the activity of the transcriptional factor serum response factor (SRF) and its cofactor MAL-D and present evidence that nuclear accumulation of MAL-D is induced by cell stretching. Border cells that cannot migrate lack nuclear MAL-D but can accumulate it if they are pulled by other migrating cells. Like mammalian MAL, MAL-D also responds to activated Diaphanous, which affects actin dynamics. MAL-D/SRF activity is required to build a robust actin cytoskeleton in the migrating cells; mutant cells break apart when initiating migration. Thus, tension-induced MAL-D activity may provide a feedback mechanism for enhancing cytoskeletal strength during invasive migration.

  17. Expression of adhesion molecules, monocyte interactions and oxidative stress in human endothelial cells exposed to wood smoke and diesel exhaust particulate matter

    DEFF Research Database (Denmark)

    Forchhammer, Lykke; Loft, Steffen; Roursgaard, Martin

    2012-01-01

    Toxicological effects of wood smoke particles are less investigated than traffic-related combustion particles. We investigated the effect of wood smoke particles, generated by smouldering combustion conditions, on human umbilical endothelial cells (HUVECs) co-cultured with or without monocytic TH...

  18. Induction of autophagy is essential for monocyte-macrophage differentiation

    OpenAIRE

    Zhang, Yan; Morgan, Michael J.; Chen, Kun; Choksi, Swati; Liu, Zheng-gang

    2012-01-01

    Monocytes are programmed to undergo apoptosis in the absence of stimulation. Stimuli that promote monocyte-macrophage differentiation not only cause cellular changes, but also prevent the default apoptosis of monocytes. In the present study, we demonstrate that autophagy is induced when monocytes are triggered to differentiate and that the induction of autophagy is pivotal for the survival and differentiation of monocytes. We also show that inhibition of autophagy results in apoptosis of cell...

  19. Niche matters: The comparison between bone marrow stem cells and endometrial stem cells and stromal fibroblasts reveal distinct migration and cytokine profiles in response to inflammatory stimulus.

    Directory of Open Access Journals (Sweden)

    Masuma Khatun

    Full Text Available Intrinsic inflammatory characteristics play a pivotal role in stem cell recruitment and homing through migration where the subsequent change in niche has been shown to alter these characteristics. The bone marrow mesenchymal stem cells (bmMSCs have been demonstrated to migrate to the endometrium contributing to the stem cell reservoir and regeneration of endometrial tissue. Thus, the aim of the present study was to compare the inflammation-driven migration and cytokine secretion profile of human bmMSCs to endometrial mesenchymal stem cells (eMSCs and endometrial fibroblasts (eSFs.The bmMSCs were isolated from bone marrow aspirates through culturing, whereas eMSCs and eSFs were FACS-isolated. All cell types were tested for their surface marker, proliferation profiles and migration properties towards serum and inflammatory attractants. The cytokine/chemokine secretion profile of 35 targets was analysed in each cell type at basal level along with lipopolysaccharide (LPS-induced state.Both stem cell types, bmMSCs and eMSCs, presented with similar stem cell surface marker profiles as well as possessed high proliferation and migration potential compared to eSFs. In multiplex assays, the secretion of 16 cytokine targets was detected and LPS stimulation expanded the cytokine secretion pattern by triggering the secretion of several targets. The bmMSCs exhibited higher cytokine secretion of vascular endothelial growth factor (VEGF-A, stromal cell-derived factor-1 alpha (SDF-1α, interleukin-1 receptor antagonist (IL-1RA, IL-6, interferon-gamma inducible protein (IP-10, monocyte chemoattractant protein (MCP-1, macrophage inflammatory protein (MIP1α and RANTES compared to eMSCs and/or eSFs after stimulation with LPS. The basal IL-8 secretion was higher in both endometrial cell types compared to bmMSCs.Our results highlight that similar to bmMSCs, the eMSCs possess high migration activity while the differentiation process towards stromal fibroblasts seemed

  20. Gliadin fragments promote migration of dendritic cells

    Czech Academy of Sciences Publication Activity Database

    Chládková, Barbara; Kamanová, Jana; Palová-Jelínková, Lenka; Cinová, Jana; Šebo, Peter; Tučková, Ludmila

    2011-01-01

    Roč. 15, č. 4 (2011), 938-948 ISSN 1582-1838 R&D Projects: GA ČR GA310/07/0414; GA ČR GD310/08/H077; GA ČR GA310/08/0447; GA AV ČR IAA500200801; GA AV ČR IAA500200914 Institutional research plan: CEZ:AV0Z50200510 Keywords : celiac disease * gliadin * dendritic cell Subject RIV: EC - Immunology Impact factor: 4.125, year: 2011

  1. Role of the extracellular matrix during neural crest cell migration.

    Science.gov (United States)

    Perris, R; Perissinotto, D

    2000-07-01

    Once specified to become neural crest (NC), cells occupying the dorsal portion of the neural tube disrupt their cadherin-mediated cell-cell contacts, acquire motile properties, and embark upon an extensive migration through the embryo to reach their ultimate phenotype-specific sites. The understanding of how this movement is regulated is still rather fragmentary due to the complexity of the cellular and molecular interactions involved. An additional intricate aspect of the regulation of NC cell movement is that the timings, modes and patterns of NC cell migration are intimately associated with the concomitant phenotypic diversification that cells undergo during their migratory phase and the fact that these changes modulate the way that moving cells interact with their microenvironment. To date, two interplaying mechanisms appear central for the guidance of the migrating NC cells through the embryo: one involves secreted signalling molecules acting through their cognate protein kinase/phosphatase-type receptors and the other is contributed by the multivalent interactions of the cells with their surrounding extracellular matrix (ECM). The latter ones seem fundamental in light of the central morphogenetic role played by the intracellular signals transduced through the cytoskeleton upon integrin ligation, and the convergence of these signalling cascades with those triggered by cadherins, survival/growth factor receptors, gap junctional communications, and stretch-activated calcium channels. The elucidation of the importance of the ECM during NC cell movement is presently favoured by the augmenting knowledge about the macromolecular structure of the specific ECM assembled during NC development and the functional assaying of its individual constituents via molecular and genetic manipulations. Collectively, these data propose that NC cell migration may be governed by time- and space-dependent alterations in the expression of inhibitory ECM components; the relative ratio

  2. Immortalized porcine mesenchymal cells derived from nasal mucosa, lungs, lymph nodes, spleen and bone marrow retain their stemness properties and trigger the expression of siglec-1 in co-cultured blood monocytic cells.

    Science.gov (United States)

    Garba, Abubakar; Desmarets, Lowiese M B; Acar, Delphine D; Devriendt, Bert; Nauwynck, Hans J

    2017-01-01

    Mesenchymal stromal cells have been isolated from different sources. They are multipotent cells capable of differentiating into many different cell types, including osteocytes, chondrocytes and adipocytes. They possess a therapeutic potential in the management of immune disorders and the repair of damaged tissues. Previous work in our laboratory showed an increase of the percentages of CD172a+, CD14+, CD163+, Siglec-1+, CD4+ and CD8+ hematopoietic cells, when co-cultured with immortalized mesenchymal cells derived from bone marrow. The present work aimed to demonstrate the stemness properties of SV40-immortalized mesenchymal cells derived from nasal mucosa, lungs, spleen, lymph nodes and red bone marrow and their immunomodulatory effect on blood monocytes. Mesenchymal cells from nasal mucosa, lungs, spleen, lymph nodes and red bone marrow were isolated and successfully immortalized using simian virus 40 large T antigen (SV40LT) and later, co-cultured with blood monocytes, in order to examine their differentiation stage (expression of Siglec-1). Flow cytometric analysis revealed that the five mesenchymal cell lines were positive for mesenchymal cell markers CD105, CD44, CD90 and CD29, but lacked the expression of myeloid cell markers CD16 and CD11b. Growth analysis of the cells demonstrated that bone marrow derived-mesenchymal cells proliferated faster compared with those derived from the other tissues. All five mesenchymal cell lines co-cultured with blood monocytes for 1, 2 and 7 days triggered the expression of siglec-1 in the monocytes. In contrast, no siglec-1+ cells were observed in monocyte cultures without mesenchymal cell lines. Mesenchymal cells isolated from nasal mucosa, lungs, spleen, lymph nodes and bone marrow were successfully immortalized and these cell lines retained their stemness properties and displayed immunomodulatory effects on blood monocytes.

  3. How Do Cells Make Decisions: Engineering Micro- and Nanoenvironments for Cell Migration

    Directory of Open Access Journals (Sweden)

    Siti Hawa Ngalim

    2010-01-01

    Full Text Available Cell migration contributes to cancer metastasis and involves cell adhesion to the extracellular matrix (ECM, force generation through the cell's cytoskeletal, and finally cell detachment. Both adhesive cues from the ECM and soluble cues from neighbouring cells and tissue trigger intracellular signalling pathways that are essential for cell migration. While the machinery of many signalling pathways is relatively well understood, how hierarchies of different and conflicting signals are established is a new area of cellular cancer research. We examine the recent advances in microfabrication, microfluidics, and nanotechnology that can be utilized to engineer micro- and nanoscaled cellular environments. Controlling both adhesive and soluble cues for migration may allow us to decipher how cells become motile, choose the direction for migration, and how oncogenic transformations influences these decision-making processes.

  4. Anandamide inhibits adhesion and migration of breast cancer cells

    International Nuclear Information System (INIS)

    Grimaldi, Claudia; Pisanti, Simona; Laezza, Chiara; Malfitano, Anna Maria; Santoro, Antonietta; Vitale, Mario; Caruso, Maria Gabriella; Notarnicola, Maria; Iacuzzo, Irma; Portella, Giuseppe; Di Marzo, Vincenzo; Bifulco, Maurizio

    2006-01-01

    The endocannabinoid system regulates cell proliferation in human breast cancer cells. We reasoned that stimulation of cannabinoid CB 1 receptors could induce a non-invasive phenotype in breast mtastatic cells. In a model of metastatic spreading in vivo, the metabolically stable anandamide analogue, 2-methyl-2'-F-anandamide (Met-F-AEA), significantly reduced the number and dimension of metastatic nodes, this effect being antagonized by the selective CB 1 antagonist SR141716A. In MDA-MB-231 cells, a highly invasive human breast cancer cell line, and in TSA-E1 cells, a murine breast cancer cell line, Met-F-AEA inhibited adhesion and migration on type IV collagen in vitro without modifying integrin expression: both these effects were antagonized by SR141716A. In order to understand the molecular mechanism involved in these processes, we analyzed the phosphorylation of FAK and Src, two tyrosine kinases involved in migration and adhesion. In Met-F-AEA-treated cells, we observed a decreased tyrosine phosphorylation of both FAK and Src, this effect being attenuated by SR141716A. We propose that CB 1 receptor agonists inhibit tumor cell invasion and metastasis by modulating FAK phosphorylation, and that CB 1 receptor activation might represent a novel therapeutic strategy to slow down the growth of breast carcinoma and to inhibit its metastatic diffusion in vivo

  5. A model for cell type localization in the migrating slug of ...

    Indian Academy of Sciences (India)

    PRAKASH

    . Localization of the three major cell types within the migrating slug stage is a dynamic process (Sternfeld 1992;. A model for cell type localization in the migrating slug of Dictyostelium discoideum based on differential chemotactic sensitivity to ...

  6. Neutrophil to lymphocyte with monocyte to lymphocyte ratio and white blood cell count in prediction of lung cancer

    Directory of Open Access Journals (Sweden)

    Thang Thanh Phan

    2018-04-01

    Full Text Available Background Lung cancer is the most common cause of cancer deaths in both sexes, while it is very difficult for screenings and early detection. Aims This study aims to clarify the role of systematic inflammation markers, including white blood cell (WBC, neutrophil (NEU, monocyte (MONO, platelet (PLT, neutrophil to lymphocyte ratio (NLR, monocyte to lymphocyte ratio (MLR and platelet to lymphocyte ratio (PLR in prediction of lung cancer. Methods A case-control study was conducted on 1,315 primary lung cancer patients and 1,315 healthy adults with matched age and gender at Cho Ray hospital. NLR, MLR and PLR were calculated by using neutrophil, lymphocyte, monocyte and platelet count which were recalled from laboratory database. With 600 cases in the derivation set, the logistic regression with univariate analysis was used to identify the impacted marker, then developing the optimal prediction model for lung cancer by logistic regression with multivariate method. The diagnostic values of optimal model consisting of sensitivity (Sen, specificity (Spe, positive predictive value (PPV, negative predictive value (NPV and the area under the ROC curve (AUC value were extracted and verified on all data, in validation set. Results The median values of WBC, NEU, MONO, PLT, NLR, MLR and PLR in lung cancer were not significantly difference between histological subtypes and clinical stages (p > 0.05, but higher than the values in control group (p < 0.01. Multivariates analysis shows that NLR, MLR and WBC were three parameters that have the significant impact of the optimal prediction model (p < 0.01. The AUC value, sensitivity and specificity of the optimal model for lung cancer detection were 0.881, 73.5 per cent (95 per cent CI:70.3–76.6 and 87.7 per cent (95 per centCI:85.2–89.9, respectively. Whereas, the PPV and NPV values of prediction model were 85.7 per cent (95 per cent CI:82.8–88.2 and 76.8 (95 per centCI:73.9–79.5, respectively. Among three

  7. Allogeneic transplantation of programmable cells of monocytic origin (PCMO) improves angiogenesis and tissue recovery in critical limb ischemia (CLI): a translational approach.

    Science.gov (United States)

    Berndt, Rouven; Hummitzsch, Lars; Heß, Katharina; Albrecht, Martin; Zitta, Karina; Rusch, Rene; Sarras, Beke; Bayer, Andreas; Cremer, Jochen; Faendrich, Fred; Groß, Justus

    2018-04-27

    Employing growth factor-induced partial reprogramming in vitro, peripheral human blood monocytes can acquire a state of plasticity along with expression of various markers of pluripotency. These so-called programmable cells of monocytic origin (PCMO) hold great promise in regenerative therapies. The aim of this translational study was to explore and exploit the functional properties of PCMO for allogeneic cell transplantation therapy in critical limb ischemia (CLI). Using our previously described differentiation protocol, murine and human monocytes were differentiated into PCMO. We examined paracrine secretion of pro-angiogenic and tissue recovery-associated proteins under hypoxia and induction of angiogenesis by PCMO in vitro. Allogeneic cell transplantation of PCMO was performed in a hind limb ischemia mouse model in comparison to cell transplantation of native monocytes and a placebo group. Moreover, we analyzed retrospectively four healing attempts with PCMO in patients with peripheral artery disease (PAD; Rutherford classification, stage 5 and 6). Statistical analysis was performed by using one-way ANOVA, Tukey's test or the Student's t test, p < 0.05. Cell culture experiments revealed good resilience of PCMO under hypoxia, enhanced paracrine release of pro-angiogenic and tissue recovery-associated proteins and induction of angiogenesis in vitro by PCMO. Animal experiments demonstrated significantly enhanced SO 2 saturation, blood flow, neoangiogenesis and tissue recovery after treatment with PCMO compared to treatment with native monocytes and placebo. Finally, first therapeutic application of PCMO in humans demonstrated increased vascular collaterals and improved wound healing in patients with chronic CLI without exaggerated immune response, malignant processes or extended infection after 12 months. In all patients minor and/or major amputations of the lower extremity could be avoided. In summary, PCMO improve angiogenesis and tissue recovery in chronic

  8. Effective clinical-scale production of dendritic cell vaccines by monocyte elutriation directly in medium, subsequent culture in bags and final antigen loading using peptides or RNA transfection.

    Science.gov (United States)

    Erdmann, Michael; Dörrie, Jan; Schaft, Niels; Strasser, Erwin; Hendelmeier, Martin; Kämpgen, Eckhart; Schuler, Gerold; Schuler-Thurner, Beatrice

    2007-09-01

    Dendritic cell (DC) vaccination approaches are advancing fast into the clinic. The major obstacle for further improvement is the current lack of a simple functionally "closed" system to generate standardized monocyte-derived (mo) DC vaccines. Here, we significantly optimized the use of the Elutra counterflow elutriation system to enrich monocytic DC precursors by (1) developing an algorithm to avoid red blood cell debulking and associated monocyte loss before elutriation, and (2) by elutriation directly in culture medium rather than phosphate-buffered saline. Upon elutriation the bags containing the collected monocytes are simply transferred into the incubator to generate DC progeny as the final "open" washing step is no longer required. Elutriation resulted in significantly more (> or = 2-fold) and purer DC than the standard gradient centrifugation/adherence-based monocyte enrichment, whereas morphology, maturation markers, viability, migratory capacity, and T cell stimulatory capacity were identical. Subsequently, we compared RNA transfection, as this is an increasingly used approach to load DC with antigen. Elutra-derived and adherence-derived DC could be electroporated with similar, high efficiency (on average >85% green fluorescence protein positive), and appeared also equal in antigen expression kinetics. Both Elutra-derived and adherence-derived DC, when loaded with the MelanA peptide or electroporated with MelanA RNA, showed a high T cell stimulation capacity, that is, priming of MelanA-specific CD8+ T cells. Our optimized Elutra-based procedure is straightforward, clearly superior to the standard gradient centrifugation/plastic adherence protocol, and now allows the generation of large numbers of peptide-loaded or RNA-transfected DC in a functionally closed system.

  9. Multiple modes of proepicardial cell migration require heartbeat.

    Science.gov (United States)

    Plavicki, Jessica S; Hofsteen, Peter; Yue, Monica S; Lanham, Kevin A; Peterson, Richard E; Heideman, Warren

    2014-05-15

    The outermost layer of the vertebrate heart, the epicardium, forms from a cluster of progenitor cells termed the proepicardium (PE). PE cells migrate onto the myocardium to give rise to the epicardium. Impaired epicardial development has been associated with defects in valve development, cardiomyocyte proliferation and alignment, cardiac conduction system maturation and adult heart regeneration. Zebrafish are an excellent model for studying cardiac development and regeneration; however, little is known about how the zebrafish epicardium forms. We report that PE migration occurs through multiple mechanisms and that the zebrafish epicardium is composed of a heterogeneous population of cells. Heterogeneity is first observed within the PE and persists through epicardium formation. Using in vivo imaging, histology and confocal microscopy, we show that PE cells migrate through a cellular bridge that forms between the pericardial mesothelium and the heart. We also observed the formation of PE aggregates on the pericardial surface, which were released into the pericardial cavity. It was previously reported that heartbeat-induced pericardiac fluid advections are necessary for PE cluster formation and subsequent epicardium development. We manipulated heartbeat genetically and pharmacologically and found that PE clusters clearly form in the absence of heartbeat. However, when heartbeat was inhibited the PE failed to migrate to the myocardium and the epicardium did not form. We isolated and cultured hearts with only a few epicardial progenitor cells and found a complete epicardial layer formed. However, pharmacologically inhibiting contraction in culture prevented epicardium formation. Furthermore, we isolated control and silent heart (sih) morpholino (MO) injected hearts prior to epicardium formation (60 hpf) and co-cultured these hearts with "donor" hearts that had an epicardium forming (108 hpf). Epicardial cells from donor hearts migrated on to control but not sih MO

  10. Glatiramer acetate (GA) prevents TNF-α-induced monocyte adhesion to primary endothelial cells through interfering with the NF-κB pathway

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Guoqian; Zhang, Xueyan; Su, Zhendong; Li, Xueqi, E-mail: xueqili075@yeah.net

    2015-01-30

    Highlights: • GA inhibited TNF-α-induced binding of monocytes to endothelial cells. • GA inhibited the induction of adhesion molecules MCP-1, VCAM-1 and E-selectin. • GA inhibits NF-κB p65 nuclear translocation and transcriptional activity. • GA inhibits TNF-α-induced IκBα degradation. - Abstract: Pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α) is considered to be the major one contributing to the process of development of endothelial dysfunction. Exposure to TNF-α induces the expression of a number of proinflammatory chemokines, such as monocyte chemotactic protein-1 (MCP-1), and adhesion molecules, including vascular adhesion molecule-1 (VCAM-1) and E-selectin, which mediate the interaction of invading monocytes with vascular endothelial cells. Glatiramer acetate (GA) is a licensed clinical drug for treating patients suffering from multiple sclerosis (MS). The effects of GA in vascular disease have not shown before. In this study, we found that GA significantly inhibited TNF-α-induced binding of monocytes to endothelial cells. Mechanistically, we found that GA ameliorated the upregulation of MCP-1, VCAM-1, and E-selectin induced by TNF-α. Notably, this process is mediated by inhibiting the nuclear translocation and activation of NF-κB. Our results also indicate that GA pretreatment attenuates the up-regulation of COX-2 and iNOS. These data suggest that GA might have a potential benefit in therapeutic endothelial dysfunction related diseases.

  11. Candida albicans Targets a Lipid Raft/Dectin-1 Platform to Enter Human Monocytes and Induce Antigen Specific T Cell Responses.

    Directory of Open Access Journals (Sweden)

    Valeria de Turris

    Full Text Available Several pathogens have been described to enter host cells via cholesterol-enriched membrane lipid raft microdomains. We found that disruption of lipid rafts by the cholesterol-extracting agent methyl-β-cyclodextrin or by the cholesterol-binding antifungal drug Amphotericin B strongly impairs the uptake of the fungal pathogen Candida albicans by human monocytes, suggesting a role of raft microdomains in the phagocytosis of the fungus. Time lapse confocal imaging indicated that Dectin-1, the C-type lectin receptor that recognizes Candida albicans cell wall-associated β-glucan, is recruited to lipid rafts upon Candida albicans uptake by monocytes, supporting the notion that lipid rafts act as an entry platform. Interestingly disruption of lipid raft integrity and interference with fungus uptake do not alter cytokine production by monocytes in response to Candida albicans but drastically dampen fungus specific T cell response. In conclusion, these data suggest that monocyte lipid rafts play a crucial role in the innate and adaptive immune responses to Candida albicans in humans and highlight a new and unexpected immunomodulatory function of the antifungal drug Amphotericin B.

  12. The origin and migration of primordial germ cells in sturgeons.

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    Taiju Saito

    Full Text Available Primordial germ cells (PGCs arise elsewhere in the embryo and migrate into developing gonadal ridges during embryonic development. In several model animals, formation and migration patterns of PGCs have been studied, and it is known that these patterns vary. Sturgeons (genus Acipenser have great potential for comparative and evolutionary studies of development. Sturgeons belong to the super class Actinoptergii, and their developmental pattern is similar to that of amphibians, although their phylogenetic position is an out-group to teleost fishes. Here, we reveal an injection technique for sturgeon eggs allowing visualization of germplasm and PGCs. Using this technique, we demonstrate that the PGCs are generated at the vegetal pole of the egg and they migrate on the yolky cell mass toward the gonadal ridge. We also provide evidence showing that PGCs are specified by inheritance of maternally supplied germplasm. Furthermore, we demonstrate that the migratory mechanism is well-conserved between sturgeon and other remotely related teleosts, such as goldfish, by a single PGCs transplantation (SPT assay. The mode of PGCs specification in sturgeon is similar to that of anurans, but the migration pattern resembles that of teleosts.

  13. Inhibition of Rho-associated kinases disturbs the collective cell migration of stratified TE-10 cells

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    Taro Mikami

    2015-01-01

    Full Text Available BACKGROUND: The collective cell migration of stratified epithelial cells is considered to be an important phenomenon in wound healing, development, and cancer invasion; however, little is known about the mechanisms involved. Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs in cell migration is controversial and might be cell-type dependent. Here, we report the development of a novel modified scratch assay that was used to observe the collective cell migration of stratified TE-10 cells derived from a human esophageal cancer specimen. RESULTS: Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet. The leading edge of cells in the cell sheet formed a simple layer and moved forward regularly; these rows were followed by the stratified epithelium. ROCK inhibitors and ROCK small interfering RNAs (siRNAs disturbed not only the collective migration of the leading edge of this cell sheet, but also the stratified layer in the rear. In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion. CONCLUSIONS: The data presented in this study suggest that ROCKs play an important role in mediating the collective migration of TE-10 cell sheets. In addition, differences between the effects of siRNAs targeting either RhoA or ROCKs suggested that distinct mechanisms regulate the collective cell migration in the simple epithelium of the wound edge versus the stratified layer of the epithelium.

  14. Mib1 contributes to persistent directional cell migration by regulating the Ctnnd1-Rac1 pathway.

    Science.gov (United States)

    Mizoguchi, Takamasa; Ikeda, Shoko; Watanabe, Saori; Sugawara, Michiko; Itoh, Motoyuki

    2017-10-31

    Persistent directional cell migration is involved in animal development and diseases. The small GTPase Rac1 is involved in F-actin and focal adhesion dynamics. Local Rac1 activity is required for persistent directional migration, whereas global, hyperactivated Rac1 enhances random cell migration. Therefore, precise control of Rac1 activity is important for proper directional cell migration. However, the molecular mechanism underlying the regulation of Rac1 activity in persistent directional cell migration is not fully understood. Here, we show that the ubiquitin ligase mind bomb 1 (Mib1) is involved in persistent directional cell migration. We found that knockdown of MIB1 led to an increase in random cell migration in HeLa cells in a wound-closure assay. Furthermore, we explored novel Mib1 substrates for cell migration and found that Mib1 ubiquitinates Ctnnd1. Mib1-mediated ubiquitination of Ctnnd1 K547 attenuated Rac1 activation in cultured cells. In addition, we found that posterior lateral line primordium cells in the zebrafish mib1 ta52b mutant showed increased random migration and loss of directional F-actin-based protrusion formation. Knockdown of Ctnnd1 partially rescued posterior lateral line primordium cell migration defects in the mib1 ta52b mutant. Taken together, our data suggest that Mib1 plays an important role in cell migration and that persistent directional cell migration is regulated, at least in part, by the Mib1-Ctnnd1-Rac1 pathway. Published under the PNAS license.

  15. Molecular aspects of tumor cell migration and invasion

    Directory of Open Access Journals (Sweden)

    Giuseppina Bozzuto

    2010-03-01

    Full Text Available Cell migration and invasion are crucial steps in many physiological events. However, they are also implicated in the physiopathology of many diseases, such as cancer. To spread through the tissues, tumor cells use mechanisms that involve several molecular actors: adhesion receptor families, receptor tyrosine kinases, cytoskeleton proteins, adapter and signalling proteins interplay in a complex scenario. The balance of cellular signals for proliferation and survival responses also regulates migratory behaviours of tumor cells. To complicate the scene of crime drug resistance players can interfere thus worsening this delicate situation. The complete understanding of this molecular jungle is an impossible mission: some molecular aspects are reviewed in this paper.

  16. Nifedipine promotes the proliferation and migration of breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Dong-Qing Guo

    Full Text Available Nifedipine is widely used as a calcium channel blocker (CCB to treat angina and hypertension,but it is controversial with respect the risk of stimulation of cancers. In this study, we demonstrated that nifedipine promoted the proliferation and migration of breast cancer cells both invivo and invitro. However, verapamil, another calcium channel blocker, didn't exert the similar effects. Nifedipine and high concentration KCl failed to alter the [Ca2+]i in MDA-MB-231 cells, suggesting that such nifedipine effect was not related with calcium channel. Moreover, nifedipine decreased miRNA-524-5p, resulting in the up-regulation of brain protein I3 (BRI3. Erk pathway was consequently activated and led to the proliferation and migration of breast cancer cells. Silencing BRI3 reversed the promoting effect of nifedipine on the breast cancer. In a summary, nifedipine stimulated the proliferation and migration of breast cancer cells via the axis of miRNA-524-5p-BRI3-Erk pathway independently of its calcium channel-blocking activity. Our findings highlight that nifedipine but not verapamil is conducive for breast cancer growth and metastasis, urging that the caution should be taken in clinic to prescribe nifedipine to women who suffering both hypertension and breast cancer, and hypertension with a tendency in breast cancers.

  17. Aquaporin 2 promotes cell migration and epithelial morphogenesis.

    Science.gov (United States)

    Chen, Ying; Rice, William; Gu, Zhizhan; Li, Jian; Huang, Jianmin; Brenner, Michael B; Van Hoek, Alfred; Xiong, Jianping; Gundersen, Gregg G; Norman, Jim C; Hsu, Victor W; Fenton, Robert A; Brown, Dennis; Lu, Hua A Jenny

    2012-09-01

    The aquaporin 2 (AQP2) water channel, expressed in kidney collecting ducts, contributes critically to water homeostasis in mammals. Animals lacking or having significantly reduced levels of AQP2, however, have not only urinary concentrating abnormalities but also renal tubular defects that lead to neonatal mortality from renal failure. Here, we show that AQP2 is not only a water channel but also an integrin-binding membrane protein that promotes cell migration and epithelial morphogenesis. AQP2 expression modulates the trafficking and internalization of integrin β1, facilitating its turnover at focal adhesions. In vitro, disturbing the interaction between AQP2 and integrin β1 by mutating the RGD motif led to reduced endocytosis, retention of integrin β1 at the cell surface, and defective cell migration and tubulogenesis. Similarly, in vivo, AQP2-null mice exhibited significant retention of integrin β1 at the basolateral membrane and had tubular abnormalities. In summary, these data suggest that the water channel AQP2 interacts with integrins to promote renal epithelial cell migration, contributing to the structural and functional integrity of the mammalian kidney.

  18. Haloperidol Abrogates Matrix Metalloproteinase-9 Expression by Inhibition of NF-κB Activation in Stimulated Human Monocytic Cells

    Directory of Open Access Journals (Sweden)

    Yueh-Lun Lee

    2018-01-01

    Full Text Available Much evidence has indicated that matrix metalloproteinases (MMPs participate in the progression of neuroinflammatory disorders. The present study was undertaken to investigate the inhibitory effect and mechanism of the antipsychotic haloperidol on MMP activation in the stimulated THP-1 monocytic cells. Haloperidol exerted a strong inhibition on tumor necrosis factor- (TNF- α-induced MMP-9 gelatinolysis of THP-1 cells. A concentration-dependent inhibitory effect of haloperidol was observed in TNF-α-induced protein and mRNA expression of MMP-9. On the other hand, haloperidol slightly affected cell viability and tissue inhibition of metalloproteinase-1 levels. It significantly inhibited the degradation of inhibitor-κB-α (IκBα in activated cells. Moreover, it suppressed activated nuclear factor-κB (NF-κB detected by a mobility shift assay, NF-κB reporter gene, and chromatin immunoprecipitation analyses. Consistent with NF-κB inhibition, haloperidol exerted a strong inhibition of lipopolysaccharide- (LPS- induced MMP-9 gelatinolysis but not of transforming growth factor-β1-induced MMP-2. In in vivo studies, administration of haloperidol significantly attenuated LPS-induced intracerebral MMP-9 activation of the brain homogenate and the in situ in C57BL/6 mice. In conclusion, the selective anti-MMP-9 activation of haloperidol could possibly involve the inhibition of the NF-κB signal pathway. Hence, it was found that haloperidol treatment may represent a bystander of anti-MMP actions for its conventional psychotherapy.

  19. Angiotensin II facilitates breast cancer cell migration and metastasis.

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    Sylvie Rodrigues-Ferreira

    Full Text Available Breast cancer metastasis is a leading cause of death by malignancy in women worldwide. Efforts are being made to further characterize the rate-limiting steps of cancer metastasis, i.e. extravasation of circulating tumor cells and colonization of secondary organs. In this study, we investigated whether angiotensin II, a major vasoactive peptide both produced locally and released in the bloodstream, may trigger activating signals that contribute to cancer cell extravasation and metastasis. We used an experimental in vivo model of cancer metastasis in which bioluminescent breast tumor cells (D3H2LN were injected intra-cardiacally into nude mice in order to recapitulate the late and essential steps of metastatic dissemination. Real-time intravital imaging studies revealed that angiotensin II accelerates the formation of metastatic foci at secondary sites. Pre-treatment of cancer cells with the peptide increases the number of mice with metastases, as well as the number and size of metastases per mouse. In vitro, angiotensin II contributes to each sequential step of cancer metastasis by promoting cancer cell adhesion to endothelial cells, trans-endothelial migration and tumor cell migration across extracellular matrix. At the molecular level, a total of 102 genes differentially expressed following angiotensin II pre-treatment were identified by comparative DNA microarray. Angiotensin II regulates two groups of connected genes related to its precursor angiotensinogen. Among those, up-regulated MMP2/MMP9 and ICAM1 stand at the crossroad of a network of genes involved in cell adhesion, migration and invasion. Our data suggest that targeting angiotensin II production or action may represent a valuable therapeutic option to prevent metastatic progression of invasive breast tumors.

  20. Angiotensin Converting Enzyme Regulates Cell Proliferation and Migration.

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    Erika Costa de Alvarenga

    Full Text Available The angiotensin-I converting enzyme (ACE plays a central role in the renin-angiotensin system, acting by converting the hormone angiotensin-I to the active peptide angiotensin-II (Ang-II. More recently, ACE was shown to act as a receptor for Ang-II, and its expression level was demonstrated to be higher in melanoma cells compared to their normal counterparts. However, the function that ACE plays as an Ang-II receptor in melanoma cells has not been defined yet.Therefore, our aim was to examine the role of ACE in tumor cell proliferation and migration.We found that upon binding to ACE, Ang-II internalizes with a faster onset compared to the binding of Ang-II to its classical AT1 receptor. We also found that the complex Ang-II/ACE translocates to the nucleus, through a clathrin-mediated process, triggering a transient nuclear Ca2+ signal. In silico studies revealed a possible interaction site between ACE and phospholipase C (PLC, and experimental results in CHO cells, demonstrated that the β3 isoform of PLC is the one involved in the Ca2+ signals induced by Ang-II/ACE interaction. Further studies in melanoma cells (TM-5 showed that Ang-II induced cell proliferation through ACE activation, an event that could be inhibited either by ACE inhibitor (Lisinopril or by the silencing of ACE. In addition, we found that stimulation of ACE by Ang-II caused the melanoma cells to migrate, at least in part due to decreased vinculin expression, a focal adhesion structural protein.ACE activation regulates melanoma cell proliferation and migration.

  1. Role of human pulmonary fibroblast-derived MCP-1 in cell activation and migration in experimental silicosis

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xueting [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Fang, Shencun [Nine Department of Respiratory Medicine, Nanjing Chest Hospital, Nanjing, Jiangsu 210029 (China); Liu, Haijun [Neurobiology Laboratory, New Drug Screening Centre, China Pharmaceutical University, Nanjing, Jiangsu 210009 (China); Wang, Xingang; Dai, Xiaoniu; Yin, Qing; Yun, Tianwei [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Wang, Wei; Zhang, Yingming [Nine Department of Respiratory Medicine, Nanjing Chest Hospital, Nanjing, Jiangsu 210029 (China); Liao, Hong [Neurobiology Laboratory, New Drug Screening Centre, China Pharmaceutical University, Nanjing, Jiangsu 210009 (China); Zhang, Wei [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Yao, Honghong [Department of Pharmacology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Chao, Jie, E-mail: chaojie@seu.edu.cn [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China)

    2015-10-15

    Background: Silicosis is a systemic disease caused by inhaling silicon dioxide (SiO{sub 2}). Phagocytosis of SiO{sub 2} in the lung initiates an inflammatory cascade that results in fibroblast proliferation and migration and subsequent fibrosis. Clinical evidence indicates that the activation of alveolar macrophages by SiO{sub 2} produces rapid and sustained inflammation that is characterized by the generation of monocyte chemotactic protein 1 (MCP-1), which induces fibrosis. Pulmonary fibroblast-derived MCP-1 may play a critical role in fibroblast proliferation and migration. Methods and results: Experiments using primary cultured adult human pulmonary fibroblasts (HPF-a) demonstrated the following results: 1) SiO{sub 2} treatment resulted in the rapid and sustained induction of MCP-1 as well as the elevation of the CC chemokine receptor type 2 (CCR2) protein levels; 2) pretreatment of HPF-a with RS-102895, a specific CCR2 inhibitor, abolished the SiO{sub 2}-induced increase in cell activation and migration in both 2D and 3D culture systems; and 3) RNA interference targeting CCR2 prevented the SiO{sub 2}-induced increase in cell migration. Conclusion: These data demonstrated that the up-regulation of pulmonary fibroblast-derived MCP-1 is involved in pulmonary fibroblast migration induced by SiO{sub 2}. CCR2 was also up-regulated in response to SiO{sub 2}, and this up-regulation facilitated the effect of MCP-1 on fibroblasts. Our study deciphered the link between fibroblast-derived MCP-1 and SiO{sub 2}-induced cell migration. This finding provides novel insight into the potential of MCP-1 in the development of novel therapeutic strategies for silicosis. - Highlights: • Role of pulmonary fibroblast-derived MCP-1 in experimental silicosis was studied. • SiO{sub 2} induced MCP-1 release from cultured human pulmonary fibroblast (HPF-a). • SiO{sub 2} directly activated HPF-a via the MCP-1/CCR2 pathway. • SiO{sub 2} increased HPF-a migration in both 2D and 3D

  2. Role of human pulmonary fibroblast-derived MCP-1 in cell activation and migration in experimental silicosis

    International Nuclear Information System (INIS)

    Liu, Xueting; Fang, Shencun; Liu, Haijun; Wang, Xingang; Dai, Xiaoniu; Yin, Qing; Yun, Tianwei; Wang, Wei; Zhang, Yingming; Liao, Hong; Zhang, Wei; Yao, Honghong; Chao, Jie

    2015-01-01

    Background: Silicosis is a systemic disease caused by inhaling silicon dioxide (SiO 2 ). Phagocytosis of SiO 2 in the lung initiates an inflammatory cascade that results in fibroblast proliferation and migration and subsequent fibrosis. Clinical evidence indicates that the activation of alveolar macrophages by SiO 2 produces rapid and sustained inflammation that is characterized by the generation of monocyte chemotactic protein 1 (MCP-1), which induces fibrosis. Pulmonary fibroblast-derived MCP-1 may play a critical role in fibroblast proliferation and migration. Methods and results: Experiments using primary cultured adult human pulmonary fibroblasts (HPF-a) demonstrated the following results: 1) SiO 2 treatment resulted in the rapid and sustained induction of MCP-1 as well as the elevation of the CC chemokine receptor type 2 (CCR2) protein levels; 2) pretreatment of HPF-a with RS-102895, a specific CCR2 inhibitor, abolished the SiO 2 -induced increase in cell activation and migration in both 2D and 3D culture systems; and 3) RNA interference targeting CCR2 prevented the SiO 2 -induced increase in cell migration. Conclusion: These data demonstrated that the up-regulation of pulmonary fibroblast-derived MCP-1 is involved in pulmonary fibroblast migration induced by SiO 2 . CCR2 was also up-regulated in response to SiO 2 , and this up-regulation facilitated the effect of MCP-1 on fibroblasts. Our study deciphered the link between fibroblast-derived MCP-1 and SiO 2 -induced cell migration. This finding provides novel insight into the potential of MCP-1 in the development of novel therapeutic strategies for silicosis. - Highlights: • Role of pulmonary fibroblast-derived MCP-1 in experimental silicosis was studied. • SiO 2 induced MCP-1 release from cultured human pulmonary fibroblast (HPF-a). • SiO 2 directly activated HPF-a via the MCP-1/CCR2 pathway. • SiO 2 increased HPF-a migration in both 2D and 3D model via the MCP-1/CCR2 pathway. • RNA-i of MCP-1/CCR2

  3. In vitro effects of monophthalates on cytokine expression in the monocytic cell line THP-1 and in peripheral blood mononuclear cells from allergic and non-allergic donors

    DEFF Research Database (Denmark)

    Glue, C; Millner, A; Bødtger, Uffe

    2002-01-01

    It has recently been shown that plasticizers are present in indoor air dust, which may lead to human exposure via the inhalation route. Moreover, studies have indicated that plasticizers may possess adjuvant effects increasing the health damaging potential of allergens. The aim of this study...... was to investigate the in vitro effect of metabolites of phthalate plastisizers, such as whether an adjuvant effect is paralleled by changes of the cytokine expression in the monocytic cell line THP-1 and in peripheral blood mononuclear cells (PBMCs) from allergics and non-allergics. The toxicity monitored by cell...... viability was determined by incubating THP-1 cells with a 10-fold dilution series of monophthalates for 24 h. At different points in time cytokine expression (IL-1beta, IL-6, IL-12alpha (p35)) in THP-1 cells incubated with non-toxic concentrations of monophthalate (2-20 microg/ml)+/-LPS (1 microg/ml) were...

  4. Migration and Tissue Tropism of Innate Lymphoid Cells

    Science.gov (United States)

    Kim, Chang H.; Hashimoto-Hill, Seika; Kim, Myunghoo

    2016-01-01

    Innate lymphoid cell (ILCs) subsets differentially populate various barrier and non-barrier tissues, where they play important roles in tissue homeostasis and tissue-specific responses to pathogen attack. Recent findings have provided insight into the molecular mechanisms that guide ILC migration into peripheral tissues, revealing common features among different ILC subsets as well as important distinctions. Recent studies have also highlighted the impact of tissue-specific cues on ILC migration, and the importance of the local immunological milieu. We review these findings here and discuss how the migratory patterns and tissue tropism of different ILC subsets relate to the development and differentiation of these cells, and to ILC-mediated tissue-specific regulation of innate and adaptive immune responses. In this context we outline open questions and important areas of future research. PMID:26708278

  5. Systems Biology Reveals Cigarette Smoke-Induced Concentration-Dependent Direct and Indirect Mechanisms That Promote Monocyte-Endothelial Cell Adhesion.

    Science.gov (United States)

    Poussin, Carine; Laurent, Alexandra; Peitsch, Manuel C; Hoeng, Julia; De Leon, Hector

    2015-10-01

    Cigarette smoke (CS) affects the adhesion of monocytes to endothelial cells, a critical step in atherogenesis. Using an in vitro adhesion assay together with innovative computational systems biology approaches to analyze omics data, our study aimed at investigating CS-induced mechanisms by which monocyte-endothelial cell adhesion is promoted. Primary human coronary artery endothelial cells (HCAECs) were treated for 4 h with (1) conditioned media of human monocytic Mono Mac-6 (MM6) cells preincubated with low or high concentrations of aqueous CS extract (sbPBS) from reference cigarette 3R4F for 2 h (indirect treatment, I), (2) unconditioned media similarly prepared without MM6 cells (direct treatment, D), or (3) freshly generated sbPBS (fresh direct treatment, FD). sbPBS promoted MM6 cells-HCAECs adhesion following I and FD, but not D. In I, the effect was mediated at a low concentration through activation of vascular inflammation processes promoted in HCAECs by a paracrine effect of the soluble mediators secreted by sbPBS-treated MM6 cells. Tumor necrosis factor α (TNFα), a major inducer, was actually shed by unstable CS compound-activated TNFα-converting enzyme. In FD, the effect was triggered at a high concentration that also induced some toxicity. This effect was mediated through an yet unknown mechanism associated with a stress damage response promoted in HCAECs by unstable CS compounds present in freshly generated sbPBS, which had decayed in D unconditioned media. Aqueous CS extract directly and indirectly promotes monocytic cell-endothelial cell adhesion in vitro via distinct concentration-dependent mechanisms. © The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. A hot water extract of Curcuma longa inhibits adhesion molecule protein expression and monocyte adhesion to TNF-α-stimulated human endothelial cells.

    Science.gov (United States)

    Kawasaki, Kengo; Muroyama, Koutarou; Yamamoto, Norio; Murosaki, Shinji

    2015-01-01

    The recruitment of arterial leukocytes to endothelial cells is an important step in the progression of various inflammatory diseases. Therefore, its modulation is thought to be a prospective target for the prevention or treatment of such diseases. Adhesion molecules on endothelial cells are induced by proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), and contribute to the recruitment of leukocytes. In the present study, we investigated the effect of hot water extract of Curcuma longa (WEC) on the protein expression of adhesion molecules, monocyte adhesion induced by TNF-α in human umbilical vascular endothelial cells (HUVECs). Treatment of HUVECs with WEC significantly suppressed both TNF-α-induced protein expression of adhesion molecules and monocyte adhesion. WEC also suppressed phosphorylation and degradation of nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα) induced by TNF-α in HUVECs, suggesting that WEC inhibits the NF-κB signaling pathway.

  7. Mesenchymal Stem Cells Induce Expression of CD73 in Human Monocytes In Vitro and in a Swine Model of Myocardial Infarction In Vivo

    Directory of Open Access Journals (Sweden)

    Marta Monguió-Tortajada

    2017-11-01

    Full Text Available The ectoenzymes CD39 and CD73 regulate the purinergic signaling through the hydrolysis of adenosine triphosphate (ATP/ADP to AMP and to adenosine (Ado, respectively. This shifts the pro-inflammatory milieu induced by extracellular ATP to the anti-inflammatory regulation by Ado. Mesenchymal stem cells (MSCs have potent immunomodulatory capabilities, including monocyte modulation toward an anti-inflammatory phenotype aiding tissue repair. In vitro, we observed that human cardiac adipose tissue-derived MSCs (cATMSCs and umbilical cord MSCs similarly polarize monocytes toward a regulatory M2 phenotype, which maintained the expression of CD39 and induced expression of CD73 in a cell contact dependent fashion, correlating with increased functional activity. In addition, the local treatment with porcine cATMSCs using an engineered bioactive graft promoted the in vivo CD73 expression on host monocytes in a swine model of myocardial infarction. Our results suggest the upregulation of ectonucleotidases on MSC-conditioned monocytes as an effective mechanism to amplify the long-lasting immunomodulatory and healing effects of MSCs delivery.

  8. CD155/PVR plays a key role in cell motility during tumor cell invasion and migration

    International Nuclear Information System (INIS)

    Sloan, Kevin E; Ilag, Leodevico L; Jay, Daniel G; Eustace, Brenda K; Stewart, Jean K; Zehetmeier, Carol; Torella, Claudia; Simeone, Marina; Roy, Jennifer E; Unger, Christine; Louis, David N

    2004-01-01

    Invasion is an important early step of cancer metastasis that is not well understood. Developing therapeutics to limit metastasis requires the identification and validation of candidate proteins necessary for invasion and migration. We developed a functional proteomic screen to identify mediators of tumor cell invasion. This screen couples Fluorophore Assisted Light Inactivation (FALI) to a scFv antibody library to systematically inactivate surface proteins expressed by human fibrosarcoma cells followed by a high-throughput assessment of transwell invasion. Using this screen, we have identified CD155 (the poliovirus receptor) as a mediator of tumor cell invasion through its role in migration. Knockdown of CD155 by FALI or by RNAi resulted in a significant decrease in transwell migration of HT1080 fibrosarcoma cells towards a serum chemoattractant. CD155 was found to be highly expressed in multiple cancer cell lines and primary tumors including glioblastoma (GBM). Knockdown of CD155 also decreased migration of U87MG GBM cells. CD155 is recruited to the leading edge of migrating cells where it colocalizes with actin and αv-integrin, known mediators of motility and adhesion. Knockdown of CD155 also altered cellular morphology, resulting in cells that were larger and more elongated than controls when plated on a Matrigel substrate. These results implicate a role for CD155 in mediating tumor cell invasion and migration and suggest that CD155 may contribute to tumorigenesis

  9. Complement 5a Enhances Hepatic Metastases of Colon Cancer via Monocyte Chemoattractant Protein-1-mediated Inflammatory Cell Infiltration.

    Science.gov (United States)

    Piao, Chunmei; Cai, Lun; Qiu, Shulan; Jia, Lixin; Song, Wenchao; Du, Jie

    2015-04-24

    Complement 5a (C5a), a potent immune mediator generated by complement activation, promotes tumor growth; however, its role in tumor metastasis remains unclear. We demonstrate that C5a contributes to tumor metastases by modulating tumor inflammation in hepatic metastases of colon cancer. Colon cancer cell lines generate C5a under serum-free conditions, and C5a levels increase over time in a murine syngeneic colon cancer hepatic metastasis model. Furthermore, in the absence of C5a receptor or upon pharmacological inhibition of C5a production with an anti-C5 monoclonal antibody, tumor metastasis is severely impaired. A lack of C5a receptor in colon cancer metastatic foci reduces the infiltration of macrophages, neutrophils, and dendritic cells, and the role for C5a receptor on these cells were further verified by bone marrow transplantation experiments. Moreover, C5a signaling increases the expression of the chemokine monocyte chemoattractant protein-1 and the anti-inflammatory molecules arginase-1, interleukin 10, and transforming growth factor β, but is inversely correlated with the expression of pro-inflammatory molecules, which suggests a mechanism for the role of C5a in the inflammatory microenvironment required for tumor metastasis. Our results indicate a new and potentially promising therapeutic application of complement C5a inhibitor for the treatment of malignant tumors. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Complement 5a Enhances Hepatic Metastases of Colon Cancer via Monocyte Chemoattractant Protein-1-mediated Inflammatory Cell Infiltration*

    Science.gov (United States)

    Piao, Chunmei; Cai, Lun; Qiu, Shulan; Jia, Lixin; Song, Wenchao; Du, Jie

    2015-01-01

    Complement 5a (C5a), a potent immune mediator generated by complement activation, promotes tumor growth; however, its role in tumor metastasis remains unclear. We demonstrate that C5a contributes to tumor metastases by modulating tumor inflammation in hepatic metastases of colon cancer. Colon cancer cell lines generate C5a under serum-free conditions, and C5a levels increase over time in a murine syngeneic colon cancer hepatic metastasis model. Furthermore, in the absence of C5a receptor or upon pharmacological inhibition of C5a production with an anti-C5 monoclonal antibody, tumor metastasis is severely impaired. A lack of C5a receptor in colon cancer metastatic foci reduces the infiltration of macrophages, neutrophils, and dendritic cells, and the role for C5a receptor on these cells were further verified by bone marrow transplantation experiments. Moreover, C5a signaling increases the expression of the chemokine monocyte chemoattractant protein-1 and the anti-inflammatory molecules arginase-1, interleukin 10, and transforming growth factor β, but is inversely correlated with the expression of pro-inflammatory molecules, which suggests a mechanism for the role of C5a in the inflammatory microenvironment required for tumor metastasis. Our results indicate a new and potentially promising therapeutic application of complement C5a inhibitor for the treatment of malignant tumors. PMID:25739439

  11. Genomic Profiling of a Human Leukemic Monocytic Cell-Line (THP-1 Exposed to Alpha Particle Radiation

    Directory of Open Access Journals (Sweden)

    Vinita Chauhan

    2012-01-01

    Full Text Available This study examined alpha (α- particle radiation effects on global changes in gene expression in human leukemic monocytic cells (THP-1 for the purposes of mining for candidate biomarkers that could be used for the development of a biological assessment tool. THP-1 cells were exposed to α-particle radiation at a dose range of 0 to 1.5 Gy. Twenty-four hours and three days after exposure gene expression was monitored using microarray technology. A total of 16 genes were dose responsive and classified as early onset due to their expression 24 h after exposure. Forty-eight transcripts were dose responsive and classified as late-onset as they were expressed 72 h after exposure. Among these genes, 6 genes were time and dose responsive and validated further using alternate technology. These transcripts were upregulated and associated with biological processes related to immune function, organelle stability and cell signalling/communication. This panel of genes merits further validation to determine if they are strong candidate biomarkers indicative of α-particle exposure.

  12. The radioactive labeling of monocytes

    International Nuclear Information System (INIS)

    Ensing, G.J.

    1985-01-01

    With the aim of studying a possible relationship between circulating monocytes and Sternberg-Reed cells investigations were started on the specific labeling of monocytes. In this thesis the literature on the pertinent data has been reviewed and a series of experiments on the monocyte labeling procedure has been described. The principles of cell labeling with radioactive compounds were discussed. 1. Total separation of the particular cell population to be labeled and subsequent labeling with a non-specific radiopharmaceutical. 2. Specific cell labeling in a mixture of cell types based on a well defined affinity of the cell under study for the radiopharmaceutical used. Next the radionuclides that can be used for cell labeling purposes were discussed with special attention for 111 In and its chelates. The principles of radiodosimetry were also discussed shortly. This section was focussed on the radiation dose the labeled cells receive because of the intracellular localized radioactivity. The radiation burden is high in comparison to amounts of radiation known to affect cell viability. A newly developed method for labeling monocytes specifically by phagocytosis of 111 In-Fe-colloid without apparent loss of cells was described in detail. (Auth.)

  13. Adhesion and migration of cells responding to microtopography.

    Science.gov (United States)

    Estévez, Maruxa; Martínez, Elena; Yarwood, Stephen J; Dalby, Matthew J; Samitier, Josep

    2015-05-01

    It is known that cells respond strongly to microtopography. However, cellular mechanisms of response are unclear. Here, we study wild-type fibroblasts responding to 25 µm(2) posts and compare their response to that of FAK(-/-) fibroblasts and fibroblasts with PMA treatment to stimulate protein kinase C (PKC) and the small g-protein Rac. FAK knockout cells modulated adhesion number and size in a similar way to cells on topography; that is, they used more, smaller adhesions, but migration was almost completely stalled demonstrating the importance of FAK signaling in contact guidance and adhesion turnover. Little similarity, however, was observed to PKC stimulated cells and cells on the topography. Interestingly, with PKC stimulation the cell nuclei became highly deformable bringing focus on these surfaces to the study of metastasis. Surfaces that aid the study of cellular migration are important in developing understanding of mechanisms of wound healing and repair in aligned tissues such as ligament and tendon. © 2014 Wiley Periodicals, Inc.

  14. From cell differentiation to cell collectives : Bacillus subtilis uses division of labor to migrate

    NARCIS (Netherlands)

    van Gestel, Jordi; Vlamakis, Hera; Kolter, Roberto

    The organization of cells, emerging from cell-cell interactions, can give rise to collective properties. These properties are adaptive when together cells can face environmental challenges that they separately cannot. One particular challenge that is important for microorganisms is migration. In

  15. Comparison of alpha-Type-1 polarizing and standard dendritic cell cytokine cocktail for maturation of therapeutic monocyte-derived dendritic cell preparations from cancer patients

    DEFF Research Database (Denmark)

    Trepiakas, Redas; Pedersen, Anders Elm; Met, Ozcan

    2008-01-01

    The current "gold standard" for generation of dendritic cell (DC) used in DC-based cancer vaccine studies is maturation of monocyte-derived DCs with tumor necrosis factor-alpha (TNF-alpha)/IL-1beta/IL-6 and prostaglandin E(2) (PGE(2)). Recently, a protocol for producing so-called alpha-Type-1...... polarized dendritic cells (alphaDC1) in serum-free medium was published based on maturation of monocyte-derived DCs with TNF-alpha/IL-1-beta/polyinosinic:polycytidylic acid (poly-I:C)/interferon (IFN)-alpha and IFN-gamma. This DC maturation cocktail was described to fulfill the criteria for optimal DC......-regulation of inhibitory molecules such as PD-L1, ILT2, ILT3 as compared to sDC. Although alphaDC1 matured DCs secreted more IL-12p70 and IL-23 these DCs had lower or similar stimulatory capacity compared to sDCs when used as stimulating cells in mixed lymphocyte reaction (MLR) or for induction of autologous influenza...

  16. Suppression of NRF2–ARE activity sensitizes chemotherapeutic agent-induced cytotoxicity in human acute monocytic leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Hui [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Institute of Disease Control and Prevention, Academy of Military Medical Sciences, Beijing (China); Wang, Huihui [School of Public Health, China Medical University, 77 Puhe Road, Shenyang North New Area, Shenyang (China); Xue, Peng [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Key Laboratory of the Public Health Safety, Ministry of Education, School of Public Health, Fudan University, Shanghai (China); Hou, Yongyong [School of Public Health, China Medical University, 77 Puhe Road, Shenyang North New Area, Shenyang (China); Dong, Jian [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Institute of Biology and Medicine, Wuhan University of Science and Technology, Wuhan (China); Zhou, Tong [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Qu, Weidong [Key Laboratory of the Public Health Safety, Ministry of Education, School of Public Health, Fudan University, Shanghai (China); Peng, Shuangqing [Institute of Disease Control and Prevention, Academy of Military Medical Sciences, Beijing (China); Li, Jin; Carmichael, Paul L. [Unilever, Safety & Environmental Assurance Centre, Colworth Science Park, Sharnbrook, Bedfordshire MK44 1LQ (United Kingdom); Nelson, Bud; Clewell, Rebecca; Zhang, Qiang; Andersen, Melvin E. [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Pi, Jingbo, E-mail: jpi@mail.cmu.edu.cn [School of Public Health, China Medical University, 77 Puhe Road, Shenyang North New Area, Shenyang (China); The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States)

    2016-02-01

    Nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of the antioxidant response element (ARE)-dependent transcription, plays a pivotal role in chemical detoxification in normal and tumor cells. Consistent with previous findings that NRF2–ARE contributes to chemotherapeutic resistance of cancer cells, we found that stable knockdown of NRF2 by lentiviral shRNA in human acute monocytic leukemia (AML) THP-1 cells enhanced the cytotoxicity of several chemotherapeutic agents, including arsenic trioxide (As{sub 2}O{sub 3}), etoposide and doxorubicin. Using an ARE-luciferase reporter expressed in several human and mouse cells, we identified a set of compounds, including isonicotinic acid amides, isoniazid and ethionamide, that inhibited NRF2–ARE activity. Treatment of THP-1 cells with ethionamide, for instance, significantly reduced mRNA expression of multiple ARE-driven genes under either basal or As{sub 2}O{sub 3}-challenged conditions. As determined by cell viability and cell cycle, suppression of NRF2–ARE by ethionamide also significantly enhanced susceptibility of THP-1 and U937 cells to As{sub 2}O{sub 3}-induced cytotoxicity. In THP-1 cells, the sensitizing effect of ethionamide on As{sub 2}O{sub 3}-induced cytotoxicity was highly dependent on NRF2. To our knowledge, the present study is the first to demonstrate that ethionamide suppresses NRF2–ARE signaling and disrupts the transcriptional network of the antioxidant response in AML cells, leading to sensitization to chemotherapeutic agents. - Highlights: • Identification of novel inhibitors of ARE-dependent transcription • Suppression of NRF2–ARE sensitizes THP-1 cells to chemotherapy. • Ethionamide suppresses ARE-dependent transcriptional activity. • Ethionamide and isoniazid increase the cytotoxicity of As{sub 2}O{sub 3} in AML cells. • Sensitization of THP-1 cells to As{sub 2}O{sub 3} toxicity by ethionamide is NRF2-dependent.

  17. Natural killer cell signal integration balances synapse symmetry and migration.

    Directory of Open Access Journals (Sweden)

    Fiona J Culley

    2009-07-01

    Full Text Available Natural killer (NK cells discern the health of other cells by recognising the balance of activating and inhibitory ligands expressed by each target cell. However, how the integration of activating and inhibitory signals relates to formation of the NK cell immune synapse remains a central question in our understanding of NK cell recognition. Here we report that ligation of LFA-1 on NK cells induced asymmetrical cell spreading and migration. In contrast, ligation of the activating receptor NKG2D induced symmetrical spreading of ruffled lamellipodia encompassing a dynamic ring of f-actin, concurrent with polarization towards a target cell and a "stop" signal. Ligation of both LFA-1 and NKG2D together resulted in symmetrical spreading but co-ligation of inhibitory receptors reverted NK cells to an asymmetrical migratory configuration leading to inhibitory synapses being smaller and more rapidly disassembled. Using micropatterned activating and inhibitory ligands, signals were found to be continuously and locally integrated during spreading. Together, these data demonstrate that NK cells spread to form large, stable, symmetrical synapses if activating signals dominate, whereas asymmetrical migratory "kinapses" are favoured if inhibitory signals dominate. This clarifies how the integration of activating and inhibitory receptor signals is translated to an appropriate NK cell response.

  18. A dynamic real time in vivo and static ex vivo analysis of granulomonocytic cell migration in the collagen-induced arthritis model.

    Directory of Open Access Journals (Sweden)

    Ruth Byrne

    Full Text Available Neutrophilic granulocytes and monocytes (granulomonocytic cells; GMC drive the inflammatory process at the earliest stages of rheumatoid arthritis (RA. The migratory behavior and functional properties of GMC within the synovial tissue are, however, only incompletely characterized. Here we have analyzed GMC in the murine collagen-induced arthritis (CIA model of RA using multi-photon real time in vivo microscopy together with ex vivo analysis of GMC in tissue sections.GMC were abundant as soon as clinical arthritis was apparent. GMC were motile and migrated randomly through the synovial tissue. In addition, we observed the frequent formation of cell clusters consisting of both neutrophilic granulocytes and monocytes that actively contributed to the inflammatory process of arthritis. Treatment of animals with a single dose of prednisolone reduced the mean velocity of cell migration and diminished the overall immigration of GMC.In summary, our study shows that the combined application of real time in vivo microscopy together with elaborate static post-mortem analysis of GMC enables the description of dynamic migratory characteristics of GMC together with their precise location in a complex anatomical environment. Moreover, this approach is sensitive enough to detect subtle therapeutic effects within a very short period of time.

  19. Exit Strategies: S1P Signaling and T Cell Migration.

    Science.gov (United States)

    Baeyens, Audrey; Fang, Victoria; Chen, Cynthia; Schwab, Susan R

    2015-12-01

    Whereas the role of sphingosine 1-phosphate receptor 1 (S1PR1) in T cell egress and the regulation of S1P gradients between lymphoid organs and circulatory fluids in homeostasis are increasingly well understood, much remains to be learned about S1P signaling and distribution during an immune response. Recent data suggest that the role of S1PR1 in directing cells from tissues into circulatory fluids is reprised again and again, particularly in guiding activated T cells from non-lymphoid tissues into lymphatics. Conversely, S1P receptor 2 (S1PR2), which antagonizes migration towards chemokines, confines cells within tissues. Here we review the current understanding of the roles of S1P signaling in activated T cell migration. In this context, we outline open questions, particularly regarding the shape of S1P gradients in different tissues in homeostasis and inflammation, and discuss recent strategies to measure S1P. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Regulation of tumor necrosis factor gene expression by ionizing radiation in human myeloid leukemia cells and peripheral blood monocytes

    International Nuclear Information System (INIS)

    Sherman, M.L.; Datta, R.; Hallahan, D.E.; Weichselbaum, R.R.; Kufe, D.W.

    1991-01-01

    Previous studies have demonstrated that ionizing radiation induces the expression of certain cytokines, such as TNF alpha/cachectin. However, there is presently no available information regarding the molecular mechanisms responsible for the regulation of cytokine gene expression by ionizing radiation. In this report, we describe the regulation of the TNF gene by ionizing radiation in human myeloid leukemia cells. The increase in TNF transcripts by x rays was both time- and dose-dependent as determined by Northern blot analysis. Similar findings were obtained in human peripheral blood monocytes. Transcriptional run-on analyses have demonstrated that ionizing radiation stimulates the rate of TNF gene transcription. Furthermore, induction of TNF mRNA was increased in the absence of protein synthesis. In contrast, ionizing radiation had little effect on the half-life of TNF transcripts. These findings indicate that the increase in TNF mRNA observed after irradiation is regulated by transcriptional mechanisms and suggest that production of this cytokine by myeloid cells may play a role in the pathophysiologic effects of ionizing radiation

  1. The frequency of tumor-infiltrating Tie-2-expressing monocytes in renal cell carcinoma: its relationship to angiogenesis and progression.

    Science.gov (United States)

    Ji, Jindong; Zhang, Guangbo; Sun, Bo; Yuan, Hexing; Huang, Yuhua; Zhang, Jianglei; Wei, Xuedong; Zhang, Xuefeng; Hou, Jianquan

    2013-10-01

    To examine the frequency of tumor-infiltrating Tie-2-expressing monocytes (TEMs) in renal cell carcinoma (RCC) and its association with microvessel density (MVD) and other clinical-pathologic features. This study enrolled 65 consecutive patients with RCC treated with radical nephrectomy. The frequency of tumor-infiltrating TEMs, which was defined as CD14(+) Tie-2(+) cells, was assessed using flow cytometry. MVD was measured by immunohistochemistry using anti-CD34 antibody. The association between clinicopathologic parameters, MVD, and the frequency of tumor-infiltrating TEMs in RCC was assessed. High frequency of tumor-infiltrating TEMs was significantly associated with advanced stage (P = .018), positive lymph nodes (P = .013), high grade (P = .019), and metastases (P = .006). Correlation analysis revealed that the frequency of TEMs was positively correlated with MVD. Our findings revealed a significant association between prognostic tumor features, MVD, and the frequency of tumor-infiltrating TEMs in RCC and indicated that TEMs may play an important role in angiogenesis and progression of RCC. Copyright © 2013 Elsevier Inc. All rights reserved.

  2. CD11c(hi) Dendritic Cells Regulate Ly-6C(hi) Monocyte Differentiation to Preserve Immune-privileged CNS in Lethal Neuroinflammation.

    Science.gov (United States)

    Kim, Jin Hyoung; Choi, Jin Young; Kim, Seong Bum; Uyangaa, Erdenebelig; Patil, Ajit Mahadev; Han, Young Woo; Park, Sang-Youel; Lee, John Hwa; Kim, Koanhoi; Eo, Seong Kug

    2015-12-02

    Although the roles of dendritic cells (DCs) in adaptive defense have been defined well, the contribution of DCs to T cell-independent innate defense and subsequent neuroimmunopathology in immune-privileged CNS upon infection with neurotropic viruses has not been completely defined. Notably, DC roles in regulating innate CD11b(+)Ly-6C(hi) monocyte functions during neuroinflammation have not yet been addressed. Using selective ablation of CD11c(hi)PDCA-1(int/lo) DCs without alteration in CD11c(int)PDCA-1(hi) plasmacytoid DC number, we found that CD11c(hi) DCs are essential to control neuroinflammation caused by infection with neurotropic Japanese encephalitis virus, through early and increased infiltration of CD11b(+)Ly-6C(hi) monocytes and higher expression of CC chemokines. More interestingly, selective CD11c(hi) DC ablation provided altered differentiation and function of infiltrated CD11b(+)Ly-6C(hi) monocytes in the CNS through Flt3-L and GM-CSF, which was closely associated with severely enhanced neuroinflammation. Furthermore, CD11b(+)Ly-6C(hi) monocytes generated in CD11c(hi) DC-ablated environment had a deleterious rather than protective role during neuroinflammation, and were more quickly recruited into inflamed CNS, depending on CCR2, thereby exacerbating neuroinflammation via enhanced supply of virus from the periphery. Therefore, our data demonstrate that CD11c(hi) DCs provide a critical and unexpected role to preserve the immune-privileged CNS in lethal neuroinflammation via regulating the differentiation, function, and trafficking of CD11b(+)Ly-6C(hi) monocytes.

  3. Impact of tumor cell cytoskeleton organization on invasiveness and migration: a microchannel-based approach.

    Directory of Open Access Journals (Sweden)

    Claudio G Rolli

    2010-01-01

    Full Text Available Cell migration is a fundamental feature of the interaction of cells with their surrounding. The cell's stiffness and ability to deform itself are two major characteristics that rule migration behavior especially in three-dimensional tissue. We simulate this situation making use of a micro-fabricated migration chip to test the active invasive behavior of pancreatic cancer cells (Panc-1 into narrow channels. At a channel width of 7 microm cell migration through the channels was significantly impeded due to size exclusion. A striking increase in cell invasiveness was observed once the cells were treated with the bioactive lipid sphingosylphosphorylcholine (SPC that leads to a reorganization of the cell's keratin network, an enhancement of the cell's deformability, and also an increase in the cell's migration speed on flat surfaces. The migration speed of the highly deformed cells inside the channels was three times higher than of cells on flat substrates but was not affected upon SPC treatment. Cells inside the channels migrated predominantly by smooth sliding while maintaining constant cell length. In contrast, cells on adhesion mediating narrow lines moved in a stepwise way, characterized by fluctuations in cell length. Taken together, with our migration chip we demonstrate that the dimensionality of the environment strongly affects the migration phenotype and we suggest that the spatial cytoskeletal keratin organization correlates with the tumor cell's invasive potential.

  4. 5-hydroxytryptamine modulates migration, cytokine and chemokine release and T-cell priming capacity of dendritic cells in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Tobias Müller

    Full Text Available Beside its well described role in the central and peripheral nervous system 5-hydroxytryptamine (5-HT, commonly known as serotonin, is also a potent immuno-modulator. Serotoninergic receptors (5-HTR are expressed by a broad range of inflammatory cell types, including dendritic cells (DCs. In this study, we aimed to further characterize the immuno-biological properties of serotoninergic receptors on human monocyte-derived DCs. 5-HT was able to induce oriented migration in immature but not in LPS-matured DCs via activation of 5-HTR(1 and 5-HTR(2 receptor subtypes. Accordingly, 5-HT also increased migration of pulmonary DCs to draining lymph nodes in vivo. By binding to 5-HTR(3, 5-HTR(4 and 5-HTR(7 receptors, 5-HT up-regulated production of the pro-inflammatory cytokine IL-6. Additionally, 5-HT influenced chemokine release by human monocyte-derived DCs: production of the potent Th1 chemoattractant IP-10/CXCL10 was inhibited in mature DCs, whereas CCL22/MDC secretion was up-regulated in both immature and mature DCs. Furthermore, DCs matured in the presence of 5-HT switched to a high IL-10 and low IL-12p70 secreting phenotype. Consistently, 5-HT favoured the outcome of a Th2 immune response both in vitro and in vivo. In summary, our study shows that 5-HT is a potent regulator of human dendritic cell function, and that targeting serotoninergic receptors might be a promising approach for the treatment of inflammatory disorders.

  5. Epigenetic changes in T-cell and monocyte signatures and production of neurotoxic cytokines in ALS patients.

    Science.gov (United States)

    Lam, Larry; Chin, Lydia; Halder, Ramesh C; Sagong, Bien; Famenini, Sam; Sayre, James; Montoya, Dennis; Rubbi, Liudmilla; Pellegrini, Matteo; Fiala, Milan

    2016-10-01

    We have investigated transcriptional and epigenetic differences in peripheral blood mononuclear cells (PBMCs) of monozygotic female twins discordant in the diagnosis of amyotrophic lateral sclerosis (ALS). Exploring DNA methylation differences by reduced representation bisulfite sequencing (RRBS), we determined that, over time, the ALS twin developed higher abundances of the CD14 macrophages and lower abundances of T cells compared to the non-ALS twin. Higher macrophage signature in the ALS twin was also shown by RNA sequencing (RNA-seq). Moreover, the twins differed in the methylome at loci near several genes, including EGFR and TNFRSF11A, and in the pathways related to the tretinoin and H3K27me3 markers. We also tested cytokine production by PBMCs. The ALS twin's PBMCs spontaneously produced IL-6 and TNF-α, whereas PBMCs of the healthy twin produced these cytokines only when stimulated by superoxide dismutase (SOD)-1. These results and flow cytometric detection of CD45 and CD127 suggest the presence of memory T cells in both twins, but effector T cells only in the ALS twin. The ALS twin's PBMC supernatants, but not the healthy twin's, were toxic to rat cortical neurons, and this toxicity was strongly inhibited by an IL-6 receptor antibody (tocilizumab) and less well by TNF-α and IL-1β antibodies. The putative neurotoxicity of IL-6 and TNF-α is in agreement with a high expression of these cytokines on infiltrating macrophages in the ALS spinal cord. We hypothesize that higher macrophage abundance and increased neurotoxic cytokines have a fundamental role in the phenotype and treatment of certain individuals with ALS.-Lam, L., Chin, L., Halder, R. C., Sagong, B., Famenini, S., Sayre, J., Montoya, D., Rubbi L., Pellegrini, M., Fiala, M. Epigenetic changes in T-cell and monocyte signatures and production of neurotoxic cytokines in ALS patients. © FASEB.

  6. Involvement of mitogen-activated protein kinases and NFκB in LPS-induced CD40 expression on human monocytic cells

    International Nuclear Information System (INIS)

    Wu Weidong; Alexis, Neil E.; Chen Xian; Bromberg, Philip A.; Peden, David B.

    2008-01-01

    CD40 is a costimulatory molecule linking innate and adaptive immune responses to bacterial stimuli, as well as a critical regulator of functions of other costimulatory molecules. The mechanisms regulating lipopolysaccharide (LPS)-induced CD40 expression have not been adequately characterized in human monocytic cells. In this study we used a human monocytic cell line, THP-1, to investigate the possible mechanisms of CD40 expression following LPS exposure. Exposure to LPS resulted in a dose- and time-dependent increase in CD40 expression. Further studies using immunoblotting and pharmacological inhibitors revealed that mitogen-activated protein kinases (MAPKs) and NFκB were activated by LPS exposure and involved in LPS-induced CD40 expression. Activation of MAPKs was not responsible for LPS-induced NFκB activation. TLR4 was expressed on THP-1 cells and pretreatment of cells with a Toll-like receptor 4 (TLR4) neutralizing antibody (HTA125) significantly blunted LPS-induced MAPK and NFκB activation and ensuing CD40 expression. Additional studies with murine macrophages expressing wild type and mutated TLR4 showed that TLR4 was implicated in LPS-induced ERK and NFκB activation, and CD40 expression. Moreover, blockage of MAPK and NFκB activation inhibited LPS-induced TLR4 expression. In summary, LPS-induced CD40 expression in monocytic cells involves MAPKs and NFκB

  7. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    International Nuclear Information System (INIS)

    Zhang, Fenxi; Hong, Yan; Liang, Wenmei; Ren, Tongming; Jing, Suhua; Lin, Juntang

    2012-01-01

    Highlights: ► Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). ► Presence of SCs dramatically increased proliferation and migration of UCMSCs. ► Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of “nurse” cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  8. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Fenxi, E-mail: fxzhang0824@gmail.com [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Hong, Yan; Liang, Wenmei [Department of Histology and Embryology, Guiyang Medical University, Guizhou 550004, People' s Republic of China (China); Ren, Tongming [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Jing, Suhua [ICU Center, The Third Hospital of Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Lin, Juntang [Stem Cell Center, Xinxiang Medical University, Henan 453003, People' s Republic of China (China)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). Black-Right-Pointing-Pointer Presence of SCs dramatically increased proliferation and migration of UCMSCs. Black-Right-Pointing-Pointer Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of 'nurse' cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  9. CSF-1R as an inhibitor of apoptosis and promoter of proliferation, migration and invasion of canine mammary cancer cells

    Science.gov (United States)

    2013-01-01

    Background Tumor-associated macrophages (TAMs) have high impact on the cancer development because they can facilitate matrix invasion, angiogenesis, and tumor cell motility. It gives cancer cells the capacity to invade normal tissues and metastasize. The signaling of colony-stimulating factor-1 receptor (CSF-1R) which is an important regulator of proliferation and differentiation of monocytes and macrophages regulates most of the tissue macrophages. However, CSF-1R is expressed also in breast epithelial tissue during some physiological stages i.g.: pregnancy and lactation. Its expression has been also detected in various cancers. Our previous study has showed the expression of CSF-1R in all examined canine mammary tumors. Moreover, it strongly correlated with grade of malignancy and ability to metastasis. This study was therefore designed to characterize the role of CSF-1R in canine mammary cancer cells proliferation, apoptosis, migration, and invasion. As far as we know, the study presented hereby is a pioneering experiment in this field of veterinary medicine. Results We showed that csf-1r silencing significantly increased apoptosis (Annexin V test), decreased proliferation (measured as Ki67 expression) and decreased migration (“wound healing” assay) of canine mammary cancer cells. Treatment of these cells with CSF-1 caused opposite effect. Moreover, csf-1r knock-down changed growth characteristics of highly invasive cell lines on Matrigel matrix, and significantly decreased the ability of these cells to invade matrix. CSF-1 treatment increased invasion of cancer cells. Conclusion The evidence of the expression and functional role of the CSF-1R in canine mammary cancer cells indicate that CSF-1R targeting may be a good therapeutic approach. PMID:23561040

  10. Cell tracking and therapy evaluation of bone marrow monocytes and stromal cells using SPECT and CMR in a canine model of myocardial infarction

    Directory of Open Access Journals (Sweden)

    Merrifield Peter

    2009-04-01

    Full Text Available Abstract Background The clinical application of stem cell therapy for myocardial infarction will require the development of methods to monitor treatment and pre-clinical assessment in a large animal model, to determine its effectiveness and the optimum cell population, route of delivery, timing, and flow milieu. Objectives To establish a model for a in vivo tracking to monitor cell engraftment after autologous transplantation and b concurrent measurement of infarct evolution and remodeling. Methods We evaluated 22 dogs (8 sham controls, 7 treated with autologous bone marrow monocytes, and 7 with stromal cells using both imaging of 111Indium-tropolone labeled cells and late gadolinium enhancement CMR for up to12 weeks after a 3 hour coronary occlusion. Hearts were also examined using immunohistochemistry for capillary density and presence of PKH26 labeled cells. Results In vivo Indium imaging demonstrated an effective biological clearance half-life from the injection site of ~5 days. CMR demonstrated a pattern of progressive infarct shrinkage over 12 weeks, ranging from 67–88% of baseline values with monocytes producing a significant treatment effect. Relative infarct shrinkage was similar through to 6 weeks in all groups, following which the treatment effect was manifest. There was a trend towards an increase in capillary density with cell treatment. Conclusion This multi-modality approach will allow determination of the success and persistence of engraftment, and a correlation of this with infarct size shrinkage, regional function, and left ventricular remodeling. There were overall no major treatment effects with this particular model of transplantation immediately post-infarct.

  11. Collagen attachment to the substrate controls cell clustering through migration

    International Nuclear Information System (INIS)

    Hou, Yue; Rodriguez, Laura Lara; Wang, Juan; Schneider, Ian C

    2014-01-01

    Cell clustering and scattering play important roles in cancer progression and tissue engineering. While the extracellular matrix (ECM) is known to control cell clustering, much of the quantitative work has focused on the analysis of clustering between cells with strong cell–cell junctions. Much less is known about how the ECM regulates cells with weak cell–cell contact. Clustering characteristics were quantified in rat adenocarcinoma cells, which form clusters on physically adsorbed collagen substrates, but not on covalently attached collagen substrates. Covalently attaching collagen inhibited desorption of collagen from the surface. While changes in proliferation rate could not explain differences seen in the clustering, changes in cell motility could. Cells plated under conditions that resulted in more clustering had a lower persistence time and slower migration rate than those under conditions that resulted in less clustering. Understanding how the ECM regulates clustering will not only impact the fundamental understanding of cancer progression, but also will guide the design of tissue engineered constructs that allow for the clustering or dissemination of cells throughout the construct. (paper)

  12. Functional characterization and phenotypic monitoring of human hematopoietic stem cell expansion and differentiation of monocytes and macrophages by whole-cell mass spectrometry

    Directory of Open Access Journals (Sweden)

    Guido Vogel

    2018-01-01

    Full Text Available The different facets of macrophages allow them to play distinct roles in tissue homeostasis, tissue repair and in response to infections. Individuals displaying dysregulated macrophage functions are proposed to be prone to inflammatory disorders or infections. However, this being a cause or a consequence of the pathology remains often unclear. In this context, we isolated and expanded CD34+ HSCs from healthy blood donors and derived them into CD14+ myeloid progenitors which were further enriched and differentiated into macrophages. Aiming for a comprehensive phenotypic profiling, we generated whole-cell mass spectrometry (WCMS fingerprints of cell samples collected along the different stages of the differentiation process to build a predictive model using a linear discriminant analysis based on principal components. Through the capacity of the model to accurately predict sample's identity of a validation set, we demonstrate that WCMS profiles obtained from bona fide blood monocytes and respectively derived macrophages mirror profiles obtained from equivalent HSC derivatives. Finally, HSC-derived macrophage functionalities were assessed by quantifying cytokine and chemokine responses to a TLR agonist in a 34-plex luminex assay and by measuring their capacity to phagocytise mycobacteria. These functional read-outs could not discriminate blood monocytes-derived from HSC-derived macrophages. To conclude, we propose that this method opens new avenues to distinguish the impact of human genetics on the dysregulated biological properties of macrophages in pathological conditions.

  13. Fc receptors for mouse IgG1 on human monocytes: polymorphism and role in antibody-induced T cell proliferation.

    Science.gov (United States)

    Tax, W J; Hermes, F F; Willems, R W; Capel, P J; Koene, R A

    1984-09-01

    In previous studies, it was shown that there is polymorphism in the mitogenic effect of mouse IgG1 monoclonal antibodies against the T3 antigen of human T cells. This polymorphism implies that IgG1 anti-T3 antibodies are not mitogenic for T cells from 30% of healthy individuals. The present results demonstrate that this polymorphism is caused by polymorphism of an Fc receptor for mouse IgG1, present on human monocytes. The Fc receptor for murine IgG1 could be detected by a newly developed rosetting assay on monocytes from all individuals responsive to the mitogenic effect of IgG1 anti-T3 antibodies. This Fc receptor was not detectable on monocytes from those individuals exhibiting no mitogenic responses to IgG1 anti-T3 monoclonal antibodies. Cross-linking of T3 antigens appears to be essential for antibody-induced mitosis of T cells, because mononuclear cells that did not proliferate in response to WT 31 (an IgG1 antibody against T3 antigen) showed a proliferative response to Sepharose beads coated with WT 31. The Fc receptor--if functionally present--may be involved in the cross-linking of T3 antigens through anti-T3 antibodies. Further evidence for the involvement of this Fc receptor in antibody-induced T cell proliferation was provided by inhibition studies. Immune complexes containing IgG1 antibodies were able to inhibit the proliferative response to IgG1 anti-T3 antibodies. This inhibition by immune complexes appears to be mediated through the monocyte Fc receptor for mouse IgG1. These findings are important for the interpretation of previously described inhibitory effects of anti-T cell monoclonal antibodies on T cell proliferation, and show that such inhibitory effects may be monocyte-mediated (via immune complexes) rather than caused by a direct involvement of the respective T cell antigens in T cell mitosis. The Fc receptor for mouse IgG1 plays a role in antibody-induced T cell proliferation. Its polymorphism may have important implications for the

  14. Investigating the Role of Surface Materials and Three Dimensional Architecture on In Vitro Differentiation of Porcine Monocyte-Derived Dendritic Cells.

    Science.gov (United States)

    Hartmann, Sofie Bruun; Mohanty, Soumyaranjan; Skovgaard, Kerstin; Brogaard, Louise; Flagstad, Frederikke Bjergvang; Emnéus, Jenny; Wolff, Anders; Summerfield, Artur; Jungersen, Gregers

    2016-01-01

    In vitro generation of dendritic-like cells through differentiation of peripheral blood monocytes is typically done using two-dimensional polystyrene culture plates. In the process of optimising cell culture techniques, engineers have developed fluidic micro-devises usually manufactured in materials other than polystyrene and applying three-dimensional structures more similar to the in vivo environment. Polydimethylsiloxane (PDMS) is an often used polymer for lab-on-a-chip devices but not much is known about the effect of changing the culture surface material from polystyrene to PDMS. In the present study the differentiation of porcine monocytes to monocyte-derived dendritic cells (moDCs) was investigated using CD172apos pig blood monocytes stimulated with GM-CSF and IL-4. Monocytes were cultured on surfaces made of two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS and carbonised three-dimensional PDMS. Cells cultured conventionally (on two-dimensional polystyrene) differentiated into moDCs as expected. Interestingly, gene expression of a wide range of cytokines, chemokines, and pattern recognition receptors was influenced by culture surface material and architecture. Distinct clustering of cells, based on similar expression patterns of 46 genes of interest, was seen for cells isolated from two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS. Changing the material from polystyrene to PDMS resulted in cells with expression patterns usually associated with macrophage expression (upregulation of CD163 and downregulation of CD1a, FLT3, LAMP3 and BATF3). However, this was purely based on gene expression level, and no functional assays were included in this study which would be necessary in order to classify the cells as being macrophages. When changing to three-dimensional culture the cells became increasingly activated in terms of IL6, IL8, IL10 and CCR5 gene expression. Further stimulation with LPS resulted

  15. Investigating the Role of Surface Materials and Three Dimensional Architecture on In Vitro Differentiation of Porcine Monocyte-Derived Dendritic Cells.

    Directory of Open Access Journals (Sweden)

    Sofie Bruun Hartmann

    Full Text Available In vitro generation of dendritic-like cells through differentiation of peripheral blood monocytes is typically done using two-dimensional polystyrene culture plates. In the process of optimising cell culture techniques, engineers have developed fluidic micro-devises usually manufactured in materials other than polystyrene and applying three-dimensional structures more similar to the in vivo environment. Polydimethylsiloxane (PDMS is an often used polymer for lab-on-a-chip devices but not much is known about the effect of changing the culture surface material from polystyrene to PDMS. In the present study the differentiation of porcine monocytes to monocyte-derived dendritic cells (moDCs was investigated using CD172apos pig blood monocytes stimulated with GM-CSF and IL-4. Monocytes were cultured on surfaces made of two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS and carbonised three-dimensional PDMS. Cells cultured conventionally (on two-dimensional polystyrene differentiated into moDCs as expected. Interestingly, gene expression of a wide range of cytokines, chemokines, and pattern recognition receptors was influenced by culture surface material and architecture. Distinct clustering of cells, based on similar expression patterns of 46 genes of interest, was seen for cells isolated from two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS. Changing the material from polystyrene to PDMS resulted in cells with expression patterns usually associated with macrophage expression (upregulation of CD163 and downregulation of CD1a, FLT3, LAMP3 and BATF3. However, this was purely based on gene expression level, and no functional assays were included in this study which would be necessary in order to classify the cells as being macrophages. When changing to three-dimensional culture the cells became increasingly activated in terms of IL6, IL8, IL10 and CCR5 gene expression. Further stimulation

  16. Alveolar macrophage-epithelial cell interaction following exposure to atmospheric particles induces the release of mediators involved in monocyte mobilization and recruitment

    Directory of Open Access Journals (Sweden)

    Mukae Hiroshi

    2005-08-01

    Full Text Available Abstract Background Studies from our laboratory have shown that human alveolar macrophages (AM and bronchial epithelial cells (HBEC exposed to ambient particles (PM10 in vitro increase their production of inflammatory mediators and that supernatants from PM10-exposed cells shorten the transit time of monocytes through the bone marrow and promote their release into the circulation. Methods The present study concerns co-culture of AM and HBEC exposed to PM10 (EHC-93 and the production of mediators involved in monocyte kinetics measured at both the mRNA and protein levels. The experiments were also designed to determine the role of the adhesive interaction between these cells via the intercellular adhesion molecule (ICAM-1 in the production of these mediators. Results AM/HBEC co-cultures exposed to 100 μg/ml of PM10 for 2 or 24 h increased their levels of granulocyte-macrophage colony-stimulating factor (GM-CSF, M-CSF, macrophage inflammatory protein (MIP-1β, monocyte chemotactic protein (MCP-1, interleukin (IL-6 and ICAM-1 mRNA, compared to exposed AM or HBEC mono-cultures, or control non-exposed co-cultures. The levels of GM-CSF, M-CSF, MIP-1β and IL-6 increased in co-cultured supernatants collected after 24 h exposure compared to control cells (p 10-induced increase in co-culture mRNA expression. Conclusion We conclude that an ICAM-1 independent interaction between AM and HBEC, lung cells that process inhaled particles, increases the production and release of mediators that enhance bone marrow turnover of monocytes and their recruitment into tissues. We speculate that this interaction amplifies PM10-induced lung inflammation and contributes to both the pulmonary and systemic morbidity associated with exposure to air pollution.

  17. Temporal cascade of inflammatory cytokines and cell-type populations in monocyte chemotactic protein-1 (MCP-1)-mediated aneurysm healing.

    Science.gov (United States)

    Hoh, Brian L; Fazal, Hanain Z; Hourani, Siham; Li, Mengchen; Lin, Li; Hosaka, Koji

    2018-03-01

    We have previously shown that monocyte chemotactic protein-1 (MCP-1) promotes aneurysm healing. To determine the temporal cascade and durability of aneurysm healing. Murine carotid aneurysms were treated with MCP-1-releasing or poly(lactic-co-glycolic) acid (PLGA)-only coils. Aneurysm healing was assessed by quantitative measurements of intraluminal tissue ingrowth on 5 μm sections by blinded observers. Aneurysm healing occurred in stages characteristic of normal wound healing. The 1st stage (day 3) was characterized by a spike in neutrophils and T cells. The 2nd stage (week 1) was characterized by an influx of macrophages and CD45+ cells significantly greater with MCP-1 than with PLGA (p<0.05). The third stage (week 2-3) was characterized by proliferation of smooth muscle cells and fibroblasts (greater with MCP-1 than with PLGA, p<0.05). The fourth stage (3-6 months) was characterized by leveling off of smooth muscle cells and fibroblasts. M1 macrophages were greater at week 1, whereas M2 macrophages were greater at weeks 2 and 3 with MCP-1 than with PLGA. Interleukin 6 was present early and increased through week 2 (p<0.05 compared with PLGA) then decreased and leveled off through 6 months. Tumour necrosis factor α was present early and remained constant through 6 months. MCP-1 and PLGA treatment had similar rates of tissue ingrowth at early time points, but MCP-1 had a significantly greater tissue ingrowth at week 3 (p<0.05), which persisted for 6 months. The sequential cascade is consistent with an inflammatory model of injury, repair, and remodeling. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  18. Uptake of silver nanoparticles by monocytic THP-1 cells depends on particle size and presence of serum proteins

    Energy Technology Data Exchange (ETDEWEB)

    Kettler, Katja, E-mail: K.Kettler@science.ru.nl [Radboud University Nijmegen, Department of Environmental Science (Netherlands); Giannakou, Christina; Jong, Wim H. de [National Institute for Public Health and the Environment (RIVM) (Netherlands); Hendriks, A. Jan [Radboud University Nijmegen, Department of Environmental Science (Netherlands); Krystek, Petra [Philips Innovation Services (Netherlands)

    2016-09-15

    Human health risks by silver nanoparticle (AgNP) exposure are likely to increase due to the increasing number of NP-containing products and demonstrated adverse effects in various cell lines. Unfortunately, results from (toxicity) studies are often based on exposure dose and are often measured only at a fixed time point. NP uptake kinetics and the time-dependent internal cellular concentration are often not considered. Macrophages are the first line of defense against invading foreign agents including NPs. How macrophages deal with the particles is essential for potential toxicity of the NPs. However, there is a considerable lack of uptake studies of particles in the nanometer range and macrophage-like cells. Therefore, uptake rates were determined over 24 h for three different AgNPs sizes (20, 50 and 75 nm) in medium with and without fetal calf serum. Non-toxic concentrations of 10 ng Ag/mL for monocytic THP-1 cells, representing realistic exposure concentration for short-term exposures, were chosen. The uptake of Ag was higher in medium without fetal calf serum and showed increasing uptake for decreasing NP sizes, both on NP mass and on number basis. Internal cellular concentrations reached roughly 32/10 %, 25/18 % and 21/15 % of the nominal concentration in the absence of fetal calf serum/with fetal calf serum for 20-, 50- and 75-nm NPs, respectively. Our research shows that uptake kinetics in macrophages differ for various NP sizes. To increase the understanding of the mechanism of NP toxicity in cells, the process of uptake (timing) should be considered.

  19. Probing Leader Cells in Endothelial Collective Migration by Plasma Lithography Geometric Confinement.

    Science.gov (United States)

    Yang, Yongliang; Jamilpour, Nima; Yao, Baoyin; Dean, Zachary S; Riahi, Reza; Wong, Pak Kin

    2016-03-03

    When blood vessels are injured, leader cells emerge in the endothelium to heal the wound and restore the vasculature integrity. The characteristics of leader cells during endothelial collective migration under diverse physiological conditions, however, are poorly understood. Here we investigate the regulation and function of endothelial leader cells by plasma lithography geometric confinement generated. Endothelial leader cells display an aggressive phenotype, connect to follower cells via peripheral actin cables and discontinuous adherens junctions, and lead migrating clusters near the leading edge. Time-lapse microscopy, immunostaining, and particle image velocimetry reveal that the density of leader cells and the speed of migrating clusters are tightly regulated in a wide range of geometric patterns. By challenging the cells with converging, diverging and competing patterns, we show that the density of leader cells correlates with the size and coherence of the migrating clusters. Collectively, our data provide evidence that leader cells control endothelial collective migration by regualting the migrating clusters.

  20. Hsc70 regulates cell surface ASIC2 expression and vascular smooth muscle cell migration.

    Science.gov (United States)

    Grifoni, Samira C; McKey, Susan E; Drummond, Heather A

    2008-05-01

    Recent studies suggest members of the degenerin (DEG)/epithelial Na(+) channel (ENaC)/acid-sensing ion channel (ASIC) protein family play an important role in vascular smooth muscle cell (VSMC) migration. In a previous investigation, we found suppression of a certain DEG/ENaC/ASIC member, ASIC2, increased VSMC chemotactic migration, raising the possibility that ASIC2 may play an inhibitory role. Because ASIC2 protein was retained in the cytoplasm, we reasoned increasing surface expression of ASIC2 might unmask the inhibitory role of ASIC2 in VSMC migration so we could test the hypothesis that ASIC2 inhibits VSMC migration. Therefore, we used the chemical chaperone glycerol to enhance ASIC2 expression. Glycerol 1) increased cytoplasm ASIC2 expression, 2) permitted detection of ASIC2 at the cell surface, and 3) inhibited platelet-derived growth factor (PDGF)-bb mediated VSMC migration. Furthermore, ASIC2 silencing completely abolished the inhibitory effect of glycerol on migration, suggesting upregulation of ASIC2 is responsible for glycerol-induced inhibition of VSMC migration. Because other investigators have shown that glycerol regulates ENaC/ASIC via interactions with a certain heat shock protein, heat shock protein 70 (Hsc70), we wanted to determine the importance of Hsc70 on ASIC2 expression in VSMCs. We found that Hsc70 silencing increases ASIC2 cell surface expression and inhibits VSMC migration, which is abolished by cosilencing ASIC2. These data demonstrate that Hsc70 inhibits ASIC2 expression, and, when the inhibitory effect of Hsc70 is removed, ASIC2 expression increases, resulting in reduced VSMC migration. Because VSMC migration contributes to vasculogenesis and remodeling following vascular injury, our findings raise the possibility that ASIC2-Hsc70 interactions may play a role in these processes.

  1. Arecoline inhibits endothelial cell growth and migration and the attachment to mononuclear cells

    Directory of Open Access Journals (Sweden)

    Shuei-Kuen Tseng

    2014-09-01

    Conclusion: Arecoline impaired vascular endothelial cells by inhibiting their growth and migration and their adhesion to U937 mononuclear cells. These results reveal that arecoline may contribute to the pathogenesis of oral submucous fibrosis and cardiovascular diseases by affecting endothelial cell function in BQ chewers.

  2. Intermediate Monocytes but Not TIE2-Expressing Monocytes Are a Sensitive Diagnostic Indicator for Colorectal Cancer

    Science.gov (United States)

    Schauer, Dominic; Starlinger, Patrick; Reiter, Christian; Jahn, Nikolaus; Zajc, Philipp; Buchberger, Elisabeth; Bachleitner-Hofmann, Thomas; Bergmann, Michael; Stift, Anton; Gruenberger, Thomas; Brostjan, Christine

    2012-01-01

    We have conducted the first study to determine the diagnostic potential of the CD14++CD16+ intermediate monocytes as compared to the pro-angiogenic subset of CD14++CD16+TIE2+ TIE2-expressing monocytes (TEMs) in cancer. These monocyte populations were investigated by flow cytometry in healthy volunteers (N = 32) and in colorectal carcinoma patients with localized (N = 24) or metastatic (N = 37) disease. We further determined blood levels of cytokines associated with monocyte regulation. The results revealed the intermediate monocyte subset to be significantly elevated in colorectal cancer patients and to show the highest frequencies in localized disease. Multivariate regression analysis identified intermediate monocytes as a significant independent variable in cancer prediction. With a cut-off value at 0.37% (intermediate monocytes of total leukocytes) the diagnostic sensitivity and specificity ranged at 69% and 81%, respectively. In contrast, TEM levels were elevated in localized cancer but did not differ significantly between groups and none of the cytokines correlated with monocyte subpopulations. Of interest, in vitro analyses supported the observation that intermediate monocytes were more potently induced by primary as opposed to metastatic cancer cells which may relate to the immunosuppressive milieu established in the advanced stage of metastatic disease. In conclusion, intermediate monocytes as compared to TIE2-expressing monocytes are a more sensitive diagnostic indicator of colorectal cancer. PMID:22973451

  3. Intermediate monocytes but not TIE2-expressing monocytes are a sensitive diagnostic indicator for colorectal cancer.

    Directory of Open Access Journals (Sweden)

    Dominic Schauer

    Full Text Available We have conducted the first study to determine the diagnostic potential of the CD14++CD16+ intermediate monocytes as compared to the pro-angiogenic subset of CD14++CD16+TIE2+ TIE2-expressing monocytes (TEMs in cancer. These monocyte populations were investigated by flow cytometry in healthy volunteers (N = 32 and in colorectal carcinoma patients with localized (N = 24 or metastatic (N = 37 disease. We further determined blood levels of cytokines associated with monocyte regulation. The results revealed the intermediate monocyte subset to be significantly elevated in colorectal cancer patients and to show the highest frequencies in localized disease. Multivariate regression analysis identified intermediate monocytes as a significant independent variable in cancer prediction. With a cut-off value at 0.37% (intermediate monocytes of total leukocytes the diagnostic sensitivity and specificity ranged at 69% and 81%, respectively. In contrast, TEM levels were elevated in localized cancer but did not differ significantly between groups and none of the cytokines correlated with monocyte subpopulations. Of interest, in vitro analyses supported the observation that intermediate monocytes were more potently induced by primary as opposed to metastatic cancer cells which may relate to the immunosuppressive milieu established in the advanced stage of metastatic disease. In conclusion, intermediate monocytes as compared to TIE2-expressing monocytes are a more sensitive diagnostic indicator of colorectal cancer.

  4. Resveratrol blocks interleukin-18-EMMPRIN cross-regulation and smooth muscle cell migration

    OpenAIRE

    Venkatesan, Balachandar; Valente, Anthony J.; Reddy, Venkatapuram Seenu; Siwik, Deborah A.; Chandrasekar, Bysani

    2009-01-01

    Vascular smooth muscle cell (SMC) migration is an important mechanism in atherogenesis and postangioplasty arterial remodeling. Previously, we demonstrated that the proinflammatory cytokine interleukin (IL)-18 is a potent inducer of SMC migration. Since extracellular matrix metalloproteinase inducer (EMMPRIN) stimulates ECM degradation and facilitates cell migration, we investigated whether IL-18 and EMMPRIN regulate each other's expression, whether their cross talk induces SMC migration, and...

  5. Electroporated Antigen-Encoding mRNA Is Not a Danger Signal to Human Mature Monocyte-Derived Dendritic Cells

    Directory of Open Access Journals (Sweden)

    Stefanie Hoyer

    2015-01-01

    Full Text Available For therapeutic cancer vaccination, the adoptive transfer of mRNA-electroporated dendritic cells (DCs is frequently performed, usually with monocyte-derived, cytokine-matured DCs (moDCs. However, DCs are rich in danger-sensing receptors which could recognize the exogenously delivered mRNA and induce DC activation, hence influencing the DCs’ immunogenicity. Therefore, we examined whether electroporation of mRNA with a proper cap and a poly-A tail of at least 64 adenosines had any influence on cocktail-matured moDCs. We used 16 different RNAs, encoding tumor antigens (MelanA, NRAS, BRAF, GNAQ, GNA11, and WT1, and variants thereof. None of those RNAs induced changes in the expression of CD25, CD40, CD83, CD86, and CD70 or the secretion of the cytokines IL-8, IL-6, and TNFα of more than 1.5-fold compared to the control condition, while an mRNA encoding an NF-κB-activation protein as positive control induced massive secretion of the cytokines. To determine whether mRNA electroporation had any effect on the whole transcriptome of the DCs, we performed microarray analyses of DCs of 6 different donors. None of 60,000 probes was significantly different between mock-electroporated DCs and MelanA-transfected DCs. Hence, we conclude that no transcriptional programs were induced within cocktail-matured DCs by electroporation of single tumor-antigen-encoding mRNAs.

  6. 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.

  7. ERK-dependent and -independent pathways trigger human neural progenitor cell migration

    International Nuclear Information System (INIS)

    Moors, Michaela; Cline, Jason E.; Abel, Josef; Fritsche, Ellen

    2007-01-01

    Besides differentiation and apoptosis, cell migration is a basic process in brain development in which neural cells migrate several centimeters within the developing brain before reaching their proper positions and forming the right connections. For identifying signaling events that control neural migration and are therefore potential targets of chemicals to disturb normal brain development, we developed a human neurosphere-based migration assay based on normal human neural progenitor (NHNP) cells, in which the distance is measured that cells wander over time. Applying this assay, we investigated the role of the extracellular signal-regulated kinases 1 and 2 (ERK1/2) in the regulation of NHNP cell migration. Exposure to model substances like ethanol or phorbol 12-myristate 13-acetate (PMA) revealed a correlation between ERK1/2 activation and cell migration. The participation of phospho-(P-) ERK1/2 was confirmed by exposure of the cells to the MEK inhibitor PD98059, which directly prohibits ERK1/2 phosphorylation and inhibited cell migration. We identified protein kinase C (PKC) and epidermal growth factor receptor (EGFR) as upstream signaling kinases governing ERK1/2 activation, thereby controlling NHNP cell migration. Additionally, treatments with src kinase inhibitors led to a diminished cell migration without affecting ERK1/2 phosphorylation. Based on these results, we postulate that migration of NHNP cells is controlled via ERK1/2-dependent and -independent pathways

  8. Coxiella burnetii Nine Mile II proteins modulate gene expression of monocytic host cells during infection

    Directory of Open Access Journals (Sweden)

    Shaw Edward I

    2010-09-01

    Full Text Available Abstract Background Coxiella burnetii is an intracellular bacterial pathogen that causes acute and chronic disease in humans. Bacterial replication occurs within enlarged parasitophorous vacuoles (PV of eukaryotic cells, the biogenesis and maintenance of which is dependent on C. burnetii protein synthesis. These observations suggest that C. burnetii actively subverts host cell processes, however little is known about the cellular biology mechanisms manipulated by the pathogen during infection. Here, we examined host cell gene expression changes specifically induced by C. burnetii proteins during infection. Results We have identified 36 host cell genes that are specifically regulated when de novo C. burnetii protein synthesis occurs during infection using comparative microarray analysis. Two parallel sets of infected and uninfected THP-1 cells were grown for 48 h followed by the addition of chloramphenicol (CAM to 10 μg/ml in one set. Total RNA was harvested at 72 hpi from all conditions, and microarrays performed using Phalanx Human OneArray™ slides. A total of 784 (mock treated and 901 (CAM treated THP-1 genes were up or down regulated ≥2 fold in the C. burnetii infected vs. uninfected cell sets, respectively. Comparisons between the complementary data sets (using >0 fold, eliminated the common gene expression changes. A stringent comparison (≥2 fold between the separate microarrays revealed 36 host cell genes modulated by C. burnetii protein synthesis. Ontological analysis of these genes identified the innate immune response, cell death and proliferation, vesicle trafficking and development, lipid homeostasis, and cytoskeletal organization as predominant cellular functions modulated by C. burnetii protein synthesis. Conclusions Collectively, these data indicate that C. burnetii proteins actively regulate the expression of specific host cell genes and pathways. This is in addition to host cell genes that respond to the presence of the

  9. Cell migration through connective tissue in 3-D

    Science.gov (United States)

    Fabry, Ben

    2008-03-01

    A prerequisite for metastasis formation is the ability of tumor cells to invade and migrate through connective tissue. Four key components endow tumor cells with this ability: secretion of matrix-degrading enzymes, firm but temporary adhesion onto connective tissue fibers, contractile force generation, and rapid remodeling of cytoskeletal structures. Cell adhesion, contraction, and cytoskeletal remodeling are biomechanical parameter that can be measured on single cells using a panel of biophysical methods. We use 2-D and 3-D traction microscopy to measure contractile forces; magnetic tweezer microrheology to estimate adhesion strengths, cytoskeletal stiffness and molecular turn-over rates; and nanoscale particle tracking to measure cytoskeletal remodeling. On a wide range of tumor cell lines we could show that cell invasiveness correlates with increased expression of integrin adhesion receptors, increased contractile force generation, and increased speed of cytoskeletal reorganization. Each of those biomechanical parameters, however, varied considerably between cell lines of similar invasivity, suggesting that tumor cells employ multiple invasion strategies that cannot be unambiguously characterized using a single assay.

  10. Seminal plasma induces global transcriptomic changes associated with cell migration, proliferation and viability in endometrial epithelial cells and stromal fibroblasts.

    Science.gov (United States)

    Chen, Joseph C; Johnson, Brittni A; Erikson, David W; Piltonen, Terhi T; Barragan, Fatima; Chu, Simon; Kohgadai, Nargis; Irwin, Juan C; Greene, Warner C; Giudice, Linda C; Roan, Nadia R

    2014-06-01

    How does seminal plasma (SP) affect the transcriptome of human primary endometrial epithelial cells (eEC) and stromal fibroblasts (eSF)? Exposure of eEC and eSF to SP in vitro increases expression of genes and secreted proteins associated with cellular migration, proliferation, viability and inhibition of cell death. Studies in both humans and animals suggest that SP can access and induce physiological changes in the upper female reproductive tract (FRT), which may participate in promoting reproductive success. This is a cross sectional study involving control samples versus treatment. SP (pooled from twenty donors) was first tested for dose- and time-dependent cytotoxic effects on eEC and eSF (n = 4). As exposure of eEC or eSF to 1% SP for 6 h proved to be non-toxic, a second set of eEC/eSF samples (n = 4) was treated under these conditions for transcriptome, protein and functional analysis. With a third set of samples (n = 3), we further compared the transcriptional response of the cells to SP versus fresh semen. eEC and eSF were isolated from endometrial biopsies from women of reproductive age undergoing benign gynecologic procedures and maintained in vitro. RNA was isolated and processed for microarray studies to analyze global transcriptomic changes. Secreted factors in conditioned media from SP-treated cells were analyzed by Luminex and for the ability to stimulate migration of CD14+ monocytes and CD4+ T cells. Pathway identifications were determined using the Z-scoring system in Ingenuity Pathways Analysis (Z scores ≥|1.5|). SP induced transcriptomic changes (P reproductive success, female reproductive health and susceptibility to sexually transmitted diseases. The gene list provided by the transcriptome analysis reported here should prove a valuable resource for understanding the response of the upper FRT to SP exposure. This project was supported by NIH AI083050-04 (W.C.G./L.C.G.); NIH U54HD 055764 (L.C.G.); NIH 1F32HD074423-02 (J.C.C.); DOD W81XWH-11

  11. Collective cell migration without proliferation: density determines cell velocity and wave velocity

    Science.gov (United States)

    Tlili, Sham; Gauquelin, Estelle; Li, Brigitte; Cardoso, Olivier; Ladoux, Benoît; Delanoë-Ayari, Hélène; Graner, François

    2018-05-01

    Collective cell migration contributes to embryogenesis, wound healing and tumour metastasis. Cell monolayer migration experiments help in understanding what determines the movement of cells far from the leading edge. Inhibiting cell proliferation limits cell density increase and prevents jamming; we observe long-duration migration and quantify space-time characteristics of the velocity profile over large length scales and time scales. Velocity waves propagate backwards and their frequency depends only on cell density at the moving front. Both cell average velocity and wave velocity increase linearly with the cell effective radius regardless of the distance to the front. Inhibiting lamellipodia decreases cell velocity while waves either disappear or have a lower frequency. Our model combines conservation laws, monolayer mechanical properties and a phenomenological coupling between strain and polarity: advancing cells pull on their followers, which then become polarized. With reasonable values of parameters, this model agrees with several of our experimental observations. Together, our experiments and model disantangle the respective contributions of active velocity and of proliferation in monolayer migration, explain how cells maintain their polarity far from the moving front, and highlight the importance of strain-polarity coupling and density in long-range information propagation.

  12. The effect of cytosolic extract of Alternaria aternata fungus on Monocyte-derived dendritic cell maturation and T-lymphocyte polarization in the presence of myelin basic protein

    Directory of Open Access Journals (Sweden)

    Loghmanni A

    2013-03-01

    Full Text Available Background: Multiple Sclerosis (MS is an autoimmune disease with impairment in function of central nervous system. Macrophages and dendritic cells play important roles in alleviating or progression of the disease. These cells can cause inflammation and damage to the myelin of nerve cells by realizing of harmful substances when these cells get matured. We studied the effect of Alternaria alternata extract on maturation of monocyte- derived dendritic cell (modc and T-cell responses in the presence of Myelin Basic Protein (MBP as a laboratory model of multiple sclerosis (MS. The purpose of this study is suitable dendritic cells production for usage in MS immunotherapy.Methods: For this study plastic adherent monocytes were cultured with granulocyte/ macrophage- colony stimulating factor (GM-CSF and interleukin -4 for converting these cells to modc and pulsed with MBP and matured in the presence of monocyte-conditioned medium (MCM in control group and MCM + Alternaria alternata extract in treatment groups. Anti-CD14, anti-CD83, anti-human leukocyte antigen-DR (anti HLA-DR monoclonal antibody were carried out for phenotyping. Autologos T cell responses and cytokine production were evaluated.Results: The results showed that the expression of CD14 decreased and CD83, HLA-DR increased in treatment groups in comparison with control groups. The production amount of IL-10 overcame IL-12 and in T cell the production of cytokines, IL-17 and Interferon-γ (IFN-γ decreased and IL-4 was increased (P<0.05. These effects escalated with increasing of dosage from 50 to 100 (mg/ml (P<0.001.Conclusion: Alternaria alternata extract can cause maturation of MBP-pulsed modc and skewing of T- lymphocyte toward Th2 and thereby can evolve into a new strategy in immunotherapy of MS.

  13. Mesenchymal Stem/Multipotent Stromal Cells from Human Decidua Basalis Reduce Endothelial Cell Activation.

    Science.gov (United States)

    Alshabibi, Manal A; Al Huqail, Al Joharah; Khatlani, Tanvir; Abomaray, Fawaz M; Alaskar, Ahmed S; Alawad, Abdullah O; Kalionis, Bill; Abumaree, Mohamed Hassan

    2017-09-15

    Recently, we reported the isolation and characterization of mesenchymal stem cells from the decidua basalis of human placenta (DBMSCs). These cells express a unique combination of molecules involved in many important cellular functions, which make them good candidates for cell-based therapies. The endothelium is a highly specialized, metabolically active interface between blood and the underlying tissues. Inflammatory factors stimulate the endothelium to undergo a change to a proinflammatory and procoagulant state (ie, endothelial cell activation). An initial response to endothelial cell activation is monocyte adhesion. Activation typically involves increased proliferation and enhanced expression of adhesion and inflammatory markers by endothelial cells. Sustained endothelial cell activation leads to a type of damage to the body associated with inflammatory diseases, such as atherosclerosis. In this study, we examined the ability of DBMSCs to protect endothelial cells from activation through monocyte adhesion, by modulating endothelial proliferation, migration, adhesion, and inflammatory marker expression. Endothelial cells were cocultured with DBMSCs, monocytes, monocyte-pretreated with DBMSCs and DBMSC-pretreated with monocytes were also evaluated. Monocyte adhesion to endothelial cells was examined following treatment with DBMSCs. Expression of endothelial cell adhesion and inflammatory markers was also analyzed. The interaction between DBMSCs and monocytes reduced endothelial cell proliferation and monocyte adhesion to endothelial cells. In contrast, endothelial cell migration increased in response to DBMSCs and monocytes. Endothelial cell expression of adhesion and inflammatory molecules was reduced by DBMSCs and DBMSC-pretreated with monocytes. The mechanism of reduced endothelial proliferation involved enhanced phosphorylation of the tumor suppressor protein p53. Our study shows for the first time that DBMSCs protect endothelial cells from activation by

  14. Peripherally Administered Nanoparticles Target Monocytic Myeloid Cells, Secondary Lymphoid Organs and Tumors in Mice

    OpenAIRE

    Kourtis, Iraklis C.; Hirosue, Sachiko; de Titta, Alexandre; Kontos, Stephan; Stegmann, Toon; Hubbell, Jeffrey A.; Swartz, Melody A.

    2013-01-01

    Nanoparticles have been extensively developed for therapeutic and diagnostic applications. While the focus of nanoparticle trafficking in vivo has traditionally been on drug delivery and organ-level biodistribution and clearance, recent work in cancer biology and infectious disease suggests that targeting different cells within a given organ can substantially affect the quality of the immunological response. Here, we examine the cell-level biodistribution kinetics after administering ultrasma...

  15. Indirubin inhibits cell proliferation, migration, invasion and angiogenesis in tumor-derived endothelial cells

    Directory of Open Access Journals (Sweden)

    Li Z

    2018-05-01

    Full Text Available Zhuohong Li, Chaofu Zhu, Baiping An, Yu Chen, Xiuyun He, Lin Qian, Lan Lan, Shijie Li Department of Oncology, The Affiliated Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, China Purpose: Hepatocellular carcinoma is one of the most predominant malignancies with high fatality rate and its incidence is rising at an alarming rate because of its resistance to radio- and chemotherapy. Indirubin is the major active anti-tumor ingredient of a traditional Chinese herbal medicine. The present study aimed to analyze the effects of indirubin on cell proliferation, migration, invasion, and angiogenesis of tumor-derived endothelial cells (Td-EC. Methods: Td-EC were derived from human umbilical vein endothelial cells (HUVEC by treating HUVEC with the conditioned medium of human liver cancer cell line HepG2. Cell proliferation, migration, invasion, and angiogenesis were assessed by MTT, wound healing, in vitro cell invasion, and in vitro tube formation assay. Results: Td-EC were successfully obtained from HUVEC cultured with 50% culture supernatant from serum-starved HepG2 cells. Indirubin significantly inhibited Td-EC proliferation in a dose- and time-dependent manner. Indirubin also inhibited Td-EC migration, invasion, and angiogenesis. However, indirubin’s effects were weaker on HUVEC than Td-EC. Conclusion: Indirubin significantly inhibited Td-EC proliferation, migration, invasion, and angiogenesis. Keywords: indirubin, Td-EC, proliferation, migration, invasion, angiogenesis

  16. Depletion of CD11c⁺ cells in the CD11c.DTR model drives expansion of unique CD64⁺ Ly6C⁺ monocytes that are poised to release TNF-α.

    Science.gov (United States)

    Sivakumaran, Shivajanani; Henderson, Stephen; Ward, Sophie; Sousa, Pedro Santos E; Manzo, Teresa; Zhang, Lei; Conlan, Thomas; Means, Terry K; D'Aveni, Maud; Hermine, Olivier; Rubio, Marie-Thérèse; Chakraverty, Ronjon; Bennett, Clare L

    2016-01-01

    Dendritic cells (DCs) play a vital role in innate and adaptive immunities. Inducible depletion of CD11c(+) DCs engineered to express a high-affinity diphtheria toxin receptor has been a powerful tool to dissect DC function in vivo. However, despite reports showing that loss of DCs induces transient monocytosis, the monocyte population that emerges and the potential impact of monocytes on studies of DC function have not been investigated. We found that depletion of CD11c(+) cells from CD11c.DTR mice induced the expansion of a variant CD64(+) Ly6C(+) monocyte population in the spleen and blood that was distinct from conventional monocytes. Expansion of CD64(+) Ly6C(+) monocytes was independent of mobilization from the BM via CCR2 but required the cytokine, G-CSF. Indeed, this population was also expanded upon exposure to exogenous G-CSF in the absence of DC depletion. CD64(+) Ly6C(+) monocytes were characterized by upregulation of innate signaling apparatus despite the absence of inflammation, and an increased capacity to produce TNF-α following LPS stimulation. Thus, depletion of CD11c(+) cells induces expansion of a unique CD64(+) Ly6C(+) monocyte population poised to synthesize TNF-α. This finding will require consideration in experiments using depletion strategies to test the role of CD11c(+) DCs in immunity. © 2015 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. TLR4-mediated expression of Mac-1 in monocytes plays a pivotal role in monocyte adhesion to vascular endothelium.

    Directory of Open Access Journals (Sweden)

    Seung Jin Lee

    Full Text Available Toll-like receptor 4 (TLR4 is known to mediate monocyte adhesion to endothelial cells, however, its role on the expression of monocyte adhesion molecules is unclear. In the present study, we investigated the role of TLR4 on the expression of monocyte adhesion molecules, and determined the functional role of TLR4-induced adhesion molecules on monocyte adhesion to endothelial cells. When THP-1 monocytes were stimulated with Kdo2-Lipid A (KLA, a specific TLR4 agonist, Mac-1 expression was markedly increased in association with an increased adhesion of monocytes to endothelial cells. These were attenuated by anti-Mac-1 antibody, suggesting a functional role of TLR4-induced Mac-1 on monocyte adhesion to endothelial cells. In monocytes treated with MK886, a 5-lipoxygenase (LO inhibitor, both Mac-1 expression and monocyte adhesion to endothelial cells induced by KLA were markedly attenuated. Moreover, KLA increased the expression of mRNA and protein of 5-LO, suggesting a pivotal role of 5-LO on these processes. In in vivo studies, KLA increased monocyte adhesion to aortic endothelium of wild-type (WT mice, which was attenuated in WT mice treated with anti-Mac-1 antibody as well as in TLR4-deficient mice. Taken together, TLR4-mediated expression of Mac-1 in monocytes plays a pivotal role on monocyte adhesion to vascular endothelium, leading to increased foam cell formation in the development of atherosclerosis.

  18. Basics elements for modelling the dynamics of cell migration in cell culture

    International Nuclear Information System (INIS)

    FarIas, Ro; Vidal, Cs; Rapacioli, M; Flores, V

    2007-01-01

    This paper introduces some basic elements for modelling the dynamics of cell migration activity over a bi-dimensional substratum. A square matrix, representing the substratum, is implemented in order to generate virtual cells with an initial random uniform distribution, with the ability to freely move within the matrix and to interact with each others by mean of adhesive forces. Two different conditions were examined: A) cells can freely move and after contacting with another cell they both completely inhibit their migration; B) cells that come into contact have the ability to rotate respect to each other without losing their contacts and retaining the ability to move together but at a slower rate, being the decrease in the rate of movement proportional to the number of contacting cells. The dynamics of the migration process in these two conditions was evaluated by recording the evolution of several parameters as a function of time. Minor modifications in some parameters (mobility, intensity of cell-cell and cell-substratum adhesiveness) significantly change the dynamics and the final result of the virtual migrating cells

  19. Epstein Barr virus Latent Membrane Protein-1 enhances dendritic cell therapy lymph node migration, activation, and IL-12 secretion.

    Directory of Open Access Journals (Sweden)

    James M Termini

    Full Text Available Dendritic cells (DC are a promising cell type for cancer vaccines due to their high immunostimulatory capacity. However, improper maturation of DC prior to treatment may account for the limited efficacy of DC vaccine clinical trials. Latent Membrane Protein-1 (LMP1 of Epstein-Barr virus was examined for its ability to mature and activate DC as a gene-based molecular adjuvant for DC vaccines. DC were transduced with an adenovirus 5 vector (Ad5 expressing LMP1 under the control of a Tet-inducible promoter. Ad5-LMP1 was found to mature and activate both human and mouse DC. LMP1 enhanced in vitro migration of DC toward CCL19, as well as in vivo migration of DC to the inguinal lymph nodes of mice following intradermal injection. LMP1-transduced DC increased T cell proliferation in a Pmel-1 adoptive transfer model and enhanced survival in B16-F10 melanoma models. LMP1-DC also enhanced protection in a vaccinia-Gag viral challenge assay. LMP1 induced high levels of IL-12p70 secretion in mouse DC when compared to standard maturation protocols. Importantly, LMP1-transduced human DC retained the capacity to secrete IL-12p70 and TNF in response to DC restimulation. In contrast, DC matured with Monocyte Conditioned Media-Mimic cocktail (Mimic were impaired in IL-12p70 secretion following restimulation. Overall, LMP1 matured and activated DC, induced migration to the lymph node, and generated high levels of IL-12p70 in a murine model. We propose LMP1 as a promising molecular adjuvant for DC vaccines.

  20. Activation of the aryl hydrocarbon receptor affects activation and function of human monocyte-derived dendritic cells.

    Science.gov (United States)

    Wang, C; Ye, Z; Kijlstra, A; Zhou, Y; Yang, P

    2014-08-01

    Aryl hydrocarbon receptor (AhR) is well known for mediating the toxic effects of dioxin-containing pollutants, but has also been shown to be involved in the natural regulation of the immune response. In this study, we investigated the effect of AhR activation by its endogenous ligands 6-formylindolo[3,2-b]carbazole (FICZ) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) on the differentiation, maturation and function of monocyte-derived DCs in Behçet's disease (BD) patients. In this study, we showed that AhR activation by FICZ and ITE down-regulated the expression of co-stimulatory molecules including human leucocyte antigen D-related (HLA-DR), CD80 and CD86, while it had no effect on the expression of CD83 and CD40 on DCs derived from BD patients and normal controls. Lipopolysaccharide (LPS)-treated dendritic cells (DCs) from active BD patients showed a higher level of interleukin (IL)-1β, IL-6, IL-23 and tumour necrosis factor (TNF)-α production. FICZ or ITE significantly inhibited the production of IL-1β, IL-6, IL-23 and TNF-α, but induced IL-10 production by DCs derived from active BD patients and normal controls. FICZ or ITE-treated DCs significantly inhibited the T helper type 17 (Th17) and Th1 cell response. Activation of AhR either by FICZ or ITE inhibits DC differentiation, maturation and function. Further studies are needed to investigate whether manipulation of the AhR pathway may be used to treat BD or other autoimmune diseases. © 2014 British Society for Immunology.

  1. Modulation of monocytic leukemia cell function and survival by highgradient magnetic fields and mathematical modeling studies

    Czech Academy of Sciences Publication Activity Database

    Zablotskyy, Vitaliy A.; Syrovets, T.; Schmidt, Z.W.; Dejneka, Alexandr; Simmet, T.

    2014-01-01

    Roč. 35, č. 10 (2014), s. 3164-3171 ISSN 0142-9612 Grant - others:AV ČR(CZ) M100101219 Institutional support: RVO:68378271 Keywords : magnetic field * cell proliferation * leukemia * apoptosis * modeling Subject RIV: BO - Biophysics Impact factor: 8.557, year: 2014

  2. CD16(+) monocytes with smooth muscle cell characteristics are reduced in human renal chronic transplant dysfunction

    NARCIS (Netherlands)

    Boersema, M.; van den Born, Joost; van Ark, J.; Harms, Geertruida; Seelen, M. A.; van Dijk, M. C. R. F.; van Goor, H.; Navis, G. J.; Popa, E. R.; Hillebrands, J. L.

    In chronic transplant dysfunction (CTD), persistent (allo)immune-mediated inflammation eventually leads to tissue remodeling including neointima formation in intragraft arteries. We previously showed that recipient-derived neointimal alpha-SMA(+) smooth muscle-like cells are present in human renal

  3. Bipolar Role for Myelo-Monocytic Cells in Autoimmune Diseases and Psychiatric Disorders

    NARCIS (Netherlands)

    W. Beumer (Wouter)

    2013-01-01

    markdownabstract__Abstract__ The immune system is a complex system of tissue with cells and messenger molecules interacting to protect an organism against pathogens. Autoimmunity is the failure of the immune system to recognize its own constituent parts as harmless self and therefore it leads to

  4. Functional relevance of protein glycosylation to the pro-inflammatory effects of extracellular matrix metalloproteinase inducer (EMMPRIN) on monocytes/macrophages.

    Science.gov (United States)

    Ge, Heng; Yuan, Wei; Liu, Jidong; He, Qing; Ding, Song; Pu, Jun; He, Ben

    2015-01-01

    Extracellular matrix metalloproteinase inducer (EMMPRIN) is an important pro-inflammatory protein involved in the cellular functions of monocytes/macrophages. We have hypothesized that high-level heterogeneousness of protein glycosylation of EMMPRIN may have functional relevance to its biological effects and affect the inflammatory activity of monocytes/macrophages. The glycosylation patterns of EMMPRIN expressed by monocytes/macrophages (THP-1 cells) in response to different extracellular stimuli were observed, and the structures of different glycosylation forms were identified. After the purification of highly- and less-glycosylated proteins respectively, the impacts of different glycosylation forms on the pro-inflammatory effects of EMMPRIN were examined in various aspects, such as cell adhesion to endothelial cells, cell migrations, cytokine expression, and activation of inflammatory signalling pathway. 1) It was mainly the highly-glycosylated form of EMMPRIN (HG-EMMPRIN) that increased after being exposed to inflammatory signals (PMA and H2O2). 2) Glycosylation of EMMPRIN in monocytes/macrophages led to N-linked-glycans being added to the protein, with the HG form containing complex-type glycans and the less-glycosylated form (LG) the simple type. 3) Only the HG-EMMPRIN but not the LG-EMMPRIN exhibited pro-inflammatory effects and stimulated inflammatory activities of the monocytes/macrophages (i.e., activation of ERK1/2 and NF-κB pathway, enhanced monocyte-endothelium adhesion, cell migration and matrix metalloproteinase -9 expression). Post-transcriptional glycosylation represents an important mechanism that determines the biological effects of EMMPRIN in monocytes/macrophages. Glycosylation of EMMPRIN may serve as a potential target for regulating the inflammatory activities of monocytes/macrophages.

  5. Functional relevance of protein glycosylation to the pro-inflammatory effects of extracellular matrix metalloproteinase inducer (EMMPRIN on monocytes/macrophages.

    Directory of Open Access Journals (Sweden)

    Heng Ge

    Full Text Available Extracellular matrix metalloproteinase inducer (EMMPRIN is an important pro-inflammatory protein involved in the cellular functions of monocytes/macrophages. We have hypothesized that high-level heterogeneousness of protein glycosylation of EMMPRIN may have functional relevance to its biological effects and affect the inflammatory activity of monocytes/macrophages.The glycosylation patterns of EMMPRIN expressed by monocytes/macrophages (THP-1 cells in response to different extracellular stimuli were observed, and the structures of different glycosylation forms were identified. After the purification of highly- and less-glycosylated proteins respectively, the impacts of different glycosylation forms on the pro-inflammatory effects of EMMPRIN were examined in various aspects, such as cell adhesion to endothelial cells, cell migrations, cytokine expression, and activation of inflammatory signalling pathway.1 It was mainly the highly-glycosylated form of EMMPRIN (HG-EMMPRIN that increased after being exposed to inflammatory signals (PMA and H2O2. 2 Glycosylation of EMMPRIN in monocytes/macrophages led to N-linked-glycans being added to the protein, with the HG form containing complex-type glycans and the less-glycosylated form (LG the simple type. 3 Only the HG-EMMPRIN but not the LG-EMMPRIN exhibited pro-inflammatory effects and stimulated inflammatory activities of the monocytes/macrophages (i.e., activation of ERK1/2 and NF-κB pathway, enhanced monocyte-endothelium adhesion, cell migration and matrix metalloproteinase -9 expression.Post-transcriptional glycosylation represents an important mechanism that determines the biological effects of EMMPRIN in monocytes/macrophages. Glycosylation of EMMPRIN may serve as a potential target for regulating the inflammatory activities of monocytes/macrophages.

  6. [Expression of ICAT and Wnt signaling-related proteins in the monocytic differentiation of HL-60 cells induced by a new steroidal drug NSC67657].

    Science.gov (United States)

    Wang, J S; Wang, W J; Wang, T; Zhang, Y

    2016-04-01

    To investigate the expression of mRNA and proteins of β-catenin, TCF-4 (ICAT) and Wnt signaling pathway-related genes in the monocytic differentiation of acute myeloid leukemia HL-60 cells induced by a new steroidal drug NSC67657. Wright's staining and α-NBE staining were used to observe the differentiation of HL-60 cells after 5 days of 10 μmol/L NSC67657 treatment. Flow cytometry (FCM) was used to detect the differentiation and cell cycles. The expressions of mRNA and proteins of ICAT and Wnt signaling pathway-related factors, including β-catenin, TCF-4, c-myc, cyclin D1 and TCF-1 before and after differentiation, were detected by RT-PCR and Western blot. Morphological observation showed that NSC67657 induced monocytic differentiation of HL-60 cells. At 5 days after 10 μmol/L NSC67657 treatment, the number of CD14(+) HL-60 cells was (94.37±2.84)%, significantly higher than the (1.31±0.09)% in control group (Pcells were of (18.76±0.98)%, significantly lower than that of (34.38±2.61) % in the control group (Pprotein, and down-regulated the expression of β-catenin mRNA and protin (Pprotein and nuclear protein in the HL-60 cells (P>0.05 for all). The target genes of Wnt signaling pathway, including c-myc, cyclinD1 and TCF-1 mRNA and proteins in the HL-60 cells were significantly down-regulated after NSC67657 treatment (Pcells, and down-regulates the expression of β-catenin and target genes of Wnt signaling pathway. These results indicate that Wnt signaling pathway may be directly or indirectly involved in the monocytic differentiation process of HL-60 cells.

  7. Abnormal expression of leiomyoma cytoskeletal proteins involved in cell migration.

    Science.gov (United States)

    Ura, Blendi; Scrimin, Federica; Arrigoni, Giorgio; Athanasakis, Emmanouil; Aloisio, Michelangelo; Monasta, Lorenzo; Ricci, Giuseppe

    2016-05-01

    Uterine leiomyomas are monoclonal tumors. Several factors are involved in the neoplastic transformation of the myometrium. In our study we focused on dysregulated cytoskeletal proteins in the leiomyoma as compared to the myometrium. Paired tissue samples of ten leiomyomas and adjacent myometria were obtained and analyzed by two‑dimensional gel electrophoresis (2-DE). Mass spectrometry was used for protein identification, and western blotting for 2-DE data validation. The values of ten cytoskeletal proteins were found to be significantly different: eight proteins were upregulated in the leiomyoma and two proteins were downregulated. Three of the upregulated proteins (myosin regulatory light polypeptide 9, four and a half LIM domains protein 1 and LIM and SH3 domain protein 1) are involved in cell migration, while downregulated protein transgelin is involved in replicative senescence. Myosin regulatory light polypeptide 9 (MYL9) was further validated by western blotting because it is considered to be a cell migration marker in several cancers and could play a key role in leiomyoma development. Our data demonstrate significant alterations in the expression of cytoskeletal proteins involved in leiomyoma growth. A better understanding of the involvement of cytoskeletal proteins in leiomyoma pathogenesis may contribute to the identification of new therapeutic targets and the development of new pharmacological approaches.

  8. Indole-3-carbinol induces G1 cell cycle arrest and apoptosis through aryl hydrocarbon receptor in THP-1 monocytic cell line.

    Science.gov (United States)

    Mohammadi, Saeed; Seyedhosseini, Fakhri Sadat; Behnampour, Nasser; Yazdani, Yaghoub

    2017-10-01

    The role of aryl hydrocarbon receptor (AhR) in carcinogenesis has been studied recently. Indole-3-carbinol (I3C) is an AhR agonist and a potential anticancer agent. Here, we investigated the effects of I3C on cell cycle progression and apoptosis through activation of AhR on THP-1 acute myeloid leukemia (AML) cell line. MTT viability assay was used to measure the cytotoxic effects of I3C on THP-1 cells. Apoptosis and cell cycle assays were investigated using flow cytometry. Real time RT-PCR was conducted to measure the alterations in the expression of AhR gene, key genes associated with AhR activation (IL1β and CYP1A1) and major genes involved in cell cycle regulation and apoptosis including P27, P21, CDK2, P53, BCL2 and FasR. Our findings revealed that I3C inhibits the proliferation of THP-1 cells in a dose- and time-dependent manner with minimal toxicity over normal monocytes. The AhR target genes (CYP1A1, IL1β) were overexpressed upon I3C treatment (p cycle arrest was also observed using flow cytometry. G1-acting cell cycle genes (P21, P27 and P53) were overexpressed (p cycle arrest in a dose- and time-dependent manner. Therefore, AhR could be targeted as a novel treatment possibility in AML.

  9. Cytoglobin inhibits migration through PI3K/AKT/mTOR pathway in fibroblast cells.

    Science.gov (United States)

    Demirci, Selami; Doğan, Ayşegül; Apdik, Hüseyin; Tuysuz, Emre Can; Gulluoglu, Sukru; Bayrak, Omer Faruk; Şahin, Fikrettin

    2018-01-01

    Cell proliferation and migration are crucial in many physiological processes including development, cancer, tissue repair, and wound healing. Cell migration is regulated by several signaling molecules. Identification of genes related to cell migration is required to understand molecular mechanism of non-healing chronic wounds which is a major concern in clinics. In the current study, the role of cytoglobin (CYGB) gene in fıbroblast cell migration and proliferation was described. L929 mouse fibroblast cells were transduced with lentiviral particles for CYGB and GFP, and analyzed for cell proliferation and migration ability. Fibroblast cells overexpressing CYGB displayed decreased cell proliferation, colony formation capacity, and cell migration. Phosphorylation levels of mTOR and two downstream effectors S6 and 4E-BP1 which take part in PI3K/AKT/mTOR signaling declined in CYGB-overexpressing cells. Microarray analysis indicated that CYGB overexpression leads to downregulation of cell proliferation, migration, and tumor growth associated genes in L929 cell line. This study demonstrated the role of CYGB in fibroblast cell motility and proliferation. CYGB could be a promising candidate for further studies as a potential target for diseases related to cell migration such as cancer and chronic wound treatment.

  10. Effect of 60Co γ-ray irradiation on cytoskeleton of human peripheral blood monocytes with whole mount cell electron microscopy in vitro

    International Nuclear Information System (INIS)

    Chen Xiaomei; Guo Yuhua; Yin Zhiwei

    1992-01-01

    Whole mount cell electron microscopy was used in combination with selective extraction to prepare cytoskeletal framework. Cytoskeleton prepared by Triton X-100 treatment of human peripheral blood monocytes appeared on electron microscopy as a highly organized and interconnected three-dimensional matrix of different fibrous elements. By three-dimensional visualization of Triton X-100 resistant cytoskeletons it was demonstrated that different doses of 60 Co γ-rays caused distinctive and reproducible alterations of the cytoskeleton of intact human peripheral blood monocytes in vitro. The alterations were similar to those caused by cytochalasin B and by colchicine. From these observations and other workers'studies, it is presumed that 60 Co γ-ray irradiation may inhibit cytoplasmic microtubule and microfilament assembling

  11. Ewing's sarcoma of bone tumor cells produces MCSF that stimulates monocyte proliferation in a novel mouse model of Ewing's sarcoma of bone.

    Science.gov (United States)

    Margulies, B S; DeBoyace, S D; Damron, T A; Allen, M J

    2015-10-01

    Ewing's sarcoma of bone is a primary childhood malignancy of bone that is treated with X-radiation therapy in combination with surgical excision and chemotherapy. To better study Ewing's sarcoma of bone we developed a novel model of primary Ewing's sarcoma of bone and then treated animals with X-radiation therapy. We identified that uncontrolled tumor resulted in lytic bone destruction while X-radiation therapy decreased lytic bone destruction and increased limb-length asymmetry, a common, crippling complication of X-radiation therapy. Osteoclasts were indentified adjacent to the tumor, however, we were unable to detect RANK-ligand in the Ewing's tumor cells in vitro, which lead us to investigate alternate mechanisms for osteoclast formation. Ewing's sarcoma tumor cells and archival Ewing's sarcoma of bone tumor biopsy samples were shown to express MCSF, which could promote osteoclast formation. Increased monocyte numbers were detected in peripheral blood and spleen in animals with untreated Ewing's sarcoma tumor while monocyte number in animals treated with x-radiation had normal numbers of monocytes. Our data suggest that our Ewing's sarcoma of bone model will be useful in the study Ewing's sarcoma tumor progression in parallel with the effects of chemotherapy and X-radiation therapy. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Ewing's Sarcoma of Bone Tumor Cells Produce MCSF that Stimulates Monocyte Proliferation in a Novel Mouse Model of Ewing's Sarcoma of Bone

    Science.gov (United States)

    Margulies, BS; DeBoyace, SD; Damron, TA; Allen, MJ

    2015-01-01

    Ewing's sarcoma of bone is a primary childhood malignancy of bone that is treated with X-radiation therapy in combination with surgical excision and chemotherapy. To better study Ewing's sarcoma of bone we developed a novel model of primary Ewing's sarcoma of bone and then treated animals with X-radiation therapy. We identified that uncontrolled tumor resulted in lytic bone destruction while X-radiation therapy decreased lytic bone destruction and increased limb-length asymmetry, a common, crippling complication of X-radiation therapy. Osteoclasts were indentified adjacent to the tumor, however, we were unable to detect RANK-ligand in the Ewing's tumor cells in vitro, which lead us to investigate alternate mechanisms for osteoclast formation. Ewing's sarcoma tumor cells and archival Ewing's sarcoma of bone tumor biopsy samples were shown to express MCSF, which could promote osteoclast formation. Increased monocyte numbers were detected in peripheral blood and spleen in animals with untreated Ewing's sarcoma tumor while monocyte number in animals treated with x-radiation had normal numbers of monocytes. Our data suggest that our Ewing's sarcoma of bone model will be useful in the study Ewing's sarcoma tumor progression in parallel with the effects of chemotherapy and X-radiation therapy. PMID:26051470

  13. HLA class I antibodies trigger increased adherence of monocytes to endothelial cells by eliciting an increase in endothelial P-selectin and, depending on subclass, by engaging FcγRs.

    Science.gov (United States)

    Valenzuela, Nicole M; Mulder, Arend; Reed, Elaine F

    2013-06-15

    Ab-mediated rejection (AMR) of solid organ transplants is characterized by intragraft macrophages. It is incompletely understood how donor-specific Ab binding to graft endothelium promotes monocyte adhesion, and what, if any, contribution is made by the Fc region of the Ab. We investigated the mechanisms underlying monocyte recruitment by HLA class I (HLA I) Ab-activated endothelium. We used a panel of murine mAbs of different subclasses to crosslink HLA I on human aortic, venous, and microvascular endothelial cells and measured the binding of human monocytic cell lines and peripheral blood monocytes. Both anti-HLA I murine (m)IgG1 and mIgG2a induced endothelial P-selectin, which was required for monocyte adhesion to endothelium irrespective of subclass. mIgG2a but not mIgG1 could bind human FcγRs. Accordingly, HLA I mIgG2a but not mIgG1 treatment of endothelial cells significantly augmented recruitment, predominantly through FcγRI, and, to a lesser extent, FcγRIIa. Moreover, HLA I mIgG2a promoted firm adhesion of monocytes to ICAM-1 through Mac-1, which may explain the prominence of monocytes during AMR. We confirmed these observations using human HLA allele-specific mAbs and IgG purified from transplant patient sera. HLA I Abs universally elicit endothelial exocytosis leading to monocyte adherence, implying that P-selectin is a putative therapeutic target to prevent macrophage infiltration during AMR. Importantly, the subclass of donor-specific Ab may influence its pathogenesis. These results imply that human IgG1 and human IgG3 should have a greater capacity to trigger monocyte infiltration into the graft than IgG2 or IgG4 due to enhancement by FcγR interactions.

  14. HLA class I antibodies trigger increased adherence of monocytes to endothelial cells by eliciting an increase in endothelial P-selectin and, depending on subclass, by engaging FcγRs1

    Science.gov (United States)

    Valenzuela, Nicole M; Mulder, Arend; Reed, Elaine F

    2013-01-01

    Antibody-mediated rejection of solid organ transplants is characterized by intragraft macrophages. It is incompletely understood how donor specific antibody binding to graft endothelium promotes monocyte adhesion, and what, if any, contribution is made by the Fc region of the antibody. We investigated the mechanisms underlying monocyte recruitment by HLA class I antibody-activated endothelium. We used a panel of murine monoclonal antibodies of different subclasses to crosslink HLA I on human aortic, venous and microvascular endothelial cells, and measured the binding of human monocytic cell lines and peripheral blood monocytes. Both anti-HLA I murine IgG1 and mIgG2a induced endothelial P-selectin, which was required for monocyte adhesion to endothelium irrespective of subclass. Mouse IgG2a but not mIgG1 could bind human FcγRs. Accordingly, HLA I mIgG2a but not mIgG1 treatment of endothelial cells significantly augmented recruitment, predominantly through FcγRI, and, to a lesser extent, FcγRIIa. Moreover, HLA I mIgG2a promoted firm adhesion of monocytes to ICAM-1 through Mac-1, which may explain the prominence of monocytes during antibody mediated rejection. We confirmed these observations using human HLA allele specific monoclonal antibodies and IgG purified from transplant patient sera. HLA I antibodies universally elicit endothelial exocytosis leading to monocyte adherence, implying that P-selectin is a putative therapeutic target to prevent macrophage infiltration during antibody-mediated rejection. Importantly, the subclass of donor specific antibody may influence its pathogenesis. These results imply that hIgG1 and hIgG3 should have a greater capacity to trigger monocyte infiltration into the graft than IgG2 or IgG4 due to enhancement by FcγR interactions. PMID:23690477

  15. Polarization Affects Airway Epithelial Conditioning of Monocyte-Derived Dendritic Cells

    DEFF Research Database (Denmark)

    Papazian, Dick; Chhoden, Tashi; Arge, Maria

    2015-01-01

    were allowed to polarize on filter inserts, and MDDCs were allowed to adhere to the epithelial basal side. In an optimized setup, the cell application was reversed, and the culture conditions were modified to preserve cellular polarization and integrity. These two parameters were crucial for the MDDCs....... In conclusion, we determined that AEC conditioning favoring cellular integrity leads to a tolerogenic MDDC phenotype, which is likely to be important in regulating immune responses against commonly inhaled allergens....

  16. Ganoderma lucidum polysaccharides in human monocytic leukemia cells: from gene expression to network construction

    Directory of Open Access Journals (Sweden)

    Ou Chern-Han

    2007-11-01

    Full Text Available Abstract Background Ganoderma lucidum has been widely used as a herbal medicine for promoting health and longevity in China and other Asian countries. Polysaccharide extracts from Ganoderma lucidum have been reported to exhibit immuno-modulating and anti-tumor activities. In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-α. This gave rise to our investigation on how F3 stimulates immuno-modulating or anti-tumor effects in human leukemia THP-1 cells. Results Here, we integrated time-course DNA microarray analysis, quantitative PCR assays, and bioinformatics methods to study the F3-induced effects in THP-1 cells. Significantly disturbed pathways induced by F3 were identified with statistical analysis on microarray data. The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment. Based on time-course gene expression measurements of the identified pathway, we reconstructed a plausible regulatory network of the involved genes using reverse-engineering computational approach. Conclusion Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

  17. Ganoderma lucidum polysaccharides in human monocytic leukemia cells: from gene expression to network construction.

    Science.gov (United States)

    Cheng, Kun-Chieh; Huang, Hsuan-Cheng; Chen, Jenn-Han; Hsu, Jia-Wei; Cheng, Hsu-Chieh; Ou, Chern-Han; Yang, Wen-Bin; Chen, Shui-Tein; Wong, Chi-Huey; Juan, Hsueh-Fen

    2007-11-09

    Ganoderma lucidum has been widely used as a herbal medicine for promoting health and longevity in China and other Asian countries. Polysaccharide extracts from Ganoderma lucidum have been reported to exhibit immuno-modulating and anti-tumor activities. In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-alpha. This gave rise to our investigation on how F3 stimulates immuno-modulating or anti-tumor effects in human leukemia THP-1 cells. Here, we integrated time-course DNA microarray analysis, quantitative PCR assays, and bioinformatics methods to study the F3-induced effects in THP-1 cells. Significantly disturbed pathways induced by F3 were identified with statistical analysis on microarray data. The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment. Based on time-course gene expression measurements of the identified pathway, we reconstructed a plausible regulatory network of the involved genes using reverse-engineering computational approach. Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

  18. Mammary collective cell migration involves transient loss of epithelial features and individual cell migration within the epithelium

    Science.gov (United States)

    Ewald, Andrew J.; Huebner, Robert J.; Palsdottir, Hildur; Lee, Jessie K.; Perez, Melissa J.; Jorgens, Danielle M.; Tauscher, Andrew N.; Cheung, Kevin J.; Werb, Zena; Auer, Manfred

    2012-01-01

    Normal mammary morphogenesis involves transitions between simple and multilayered epithelial organizations. We used electron microscopy and molecular markers to determine whether intercellular junctions and apico-basal polarity were maintained in the multilayered epithelium. We found that multilayered elongating ducts had polarized apical and basal tissue surfaces both in three-dimensional culture and in vivo. However, individual cells were only polarized on surfaces in contact with the lumen or extracellular matrix. The basolateral marker scribble and the apical marker atypical protein kinase C zeta localized to all interior cell membranes, whereas PAR3 displayed a cytoplasmic localization, suggesting that the apico-basal polarity was incomplete. Despite membrane localization of E-cadherin and β-catenin, we did not observe a defined zonula adherens connecting interior cells. Instead, interior cells were connected through desmosomes and exhibited complex interdigitating membrane protrusions. Single-cell labeling revealed that individual cells were both protrusive and migratory within the epithelial multilayer. Inhibition of Rho kinase (ROCK) further reduced intercellular adhesion on apical and lateral surfaces but did not disrupt basal tissue organization. Following morphogenesis, segregated membrane domains were re-established and junctional complexes re-formed. We observed similar epithelial organization during mammary morphogenesis in organotypic culture and in vivo. We conclude that mammary epithelial morphogenesis involves a reversible, spatially limited, reduction in polarity and intercellular junctions and active individualistic cell migration. Our data suggest that reductions in polarity and adhesion during breast cancer progression might reflect partial recapitulation of a normal developmental program. PMID:22344263

  19. The effects of dietary fatty acids on the postprandial triglyceride-rich lipoprotein/apoB48 receptor axis in human monocyte/macrophage cells.

    Science.gov (United States)

    Varela, Lourdes M; Ortega-Gomez, Almudena; Lopez, Sergio; Abia, Rocio; Muriana, Francisco J G; Bermudez, Beatriz

    2013-12-01

    Intestinally produced triglyceride-rich lipoproteins (TRL) play an important role in the progression of atherosclerosis. In this study, we investigated the relevance of monounsaturated fatty acid (MUFA) and saturated fatty acid (SFA) in postprandial TRL in affecting the transcriptional activity of the apolipoprotein-B48 receptor (ApoB48R) and its functionality in human monocyte/macrophage cells. Healthy male volunteers were administered four standardized high-fat meals containing butter, high-palmitic sunflower oil, olive oil (ROO) or a mixture of vegetable and fish oils (50 g/m(2) body surface area) to obtain a panel of postprandial TRL with gradual MUFA oleic acid-to-SFA palmitic acid ratios. The increase in this ratio was linearly associated with a decrease of ApoB48R up-regulation and lipid accumulation in THP-1 and primary monocytes. ApoB48R mRNA levels and intracellular triglycerides were also lower in the monocytes from volunteers after the ingestion of the ROO meal when compared to the ingestion of the butter meal. In THP-1 macrophages, the increase in the MUFA oleic acid-to-SFA palmitic acid ratio in the postprandial TRL was linearly correlated with an increase in ApoB48R down-regulation and a decrease in lipid accumulation. We also revealed that the nuclear receptor transcription factors PPARα, PPARβ/δ, and PPARγ and the PPAR-RXR transcriptional complex were involved in sensing the proportion of MUFA oleic acid and SFA palmitic acid, and these were also involved in adjusting the transcriptional activity of ApoB48R. The results of this study support the notion that MUFA-rich dietary fats may prevent excessive lipid accumulation in monocyte/macrophage cells by targeting the postprandial TRL/ApoB48R axis. © 2013.

  20. Cell intrinsic modulation of Wnt signaling controls neuroblast migration in C. elegans

    NARCIS (Netherlands)

    Mentink, Remco A; Middelkoop, Teije C; Rella, Lorenzo; Ji, Ni; Tang, Chung Yin; Betist, Marco C; van Oudenaarden, Alexander; Korswagen, Hendrik C

    2014-01-01

    Members of the Wnt family of secreted signaling proteins are key regulators of cell migration and axon guidance. In the nematode C. elegans, the migration of the QR neuroblast descendants requires multiple Wnt ligands and receptors. We found that the migration of the QR descendants is divided into

  1. Pluripotency gene expression and growth control in cultures of peripheral blood monocytes during their conversion into programmable cells of monocytic origin (PCMO: evidence for a regulatory role of autocrine activin and TGF-β.

    Directory of Open Access Journals (Sweden)

    Hendrik Ungefroren

    Full Text Available Previous studies have shown that peripheral blood monocytes can be converted in vitro to a stem cell-like cell termed PCMO as evidenced by the re-expression of pluripotency-associated genes, transient proliferation, and the ability to adopt the phenotype of hepatocytes and insulin-producing cells upon tissue-specific differentiation. However, the regulatory interactions between cultured cells governing pluripotency and mitotic activity have remained elusive. Here we asked whether activin(s and TGF-β(s, are involved in PCMO generation. De novo proliferation of PCMO was higher under adherent vs. suspended culture conditions as revealed by the appearance of a subset of Ki67-positive monocytes and correlated with down-regulation of p21WAF1 beyond day 2 of culture. Realtime-PCR analysis showed that PCMO express ActRIIA, ALK4, TβRII, ALK5 as well as TGF-β1 and the βA subunit of activin. Interestingly, expression of ActRIIA and ALK4, and activin A levels in the culture supernatants increased until day 4 of culture, while levels of total and active TGF-β1 strongly declined. PCMO responded to both growth factors in an autocrine fashion with intracellular signaling as evidenced by a rise in the levels of phospho-Smad2 and a drop in those of phospho-Smad3. Stimulation of PCMO with recombinant activins (A, B, AB and TGF-β1 induced phosphorylation of Smad2 but not Smad3. Inhibition of autocrine activin signaling by either SB431542 or follistatin reduced both Smad2 activation and Oct4A/Nanog upregulation. Inhibition of autocrine TGF-β signaling by either SB431542 or anti-TGF-β antibody reduced Smad3 activation and strongly increased the number of Ki67-positive cells. Furthermore, anti-TGF-β antibody moderately enhanced Oct4A/Nanog express