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Sample records for cell s-nitrosothiol formation

  1. Breath formate is a marker of airway S-nitrosothiol depletion in severe asthma.

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    Roby Greenwald

    Full Text Available BACKGROUND: Children with severe asthma have poor symptom control and elevated markers of airway oxidative and nitrosative stress. Paradoxically, they have decreased airway levels of S-nitrosothiols (SNOs, a class of endogenous airway smooth muscle relaxants. This deficiency results from increased activity of an enzyme that both reduces SNOs to ammonia and oxidizes formaldehyde to formic acid, a volatile carboxylic acid that is more easily detected in exhaled breath condensate (EBC than SNOs. We therefore hypothesize that depletion of airway SNOs is related to asthma pathology, and breath formate concentration may be a proxy measure of SNO catabolism. METHODS AND FINDINGS: We collected EBC samples from children and adolescents, including 38 with severe asthma, 46 with mild-to-moderate asthma and 16 healthy adolescent controls, and the concentration of ionic constituents was quantified using ion chromatography. The concentrations of EBC components with volatile conjugates were log-normally distributed. Formate was the principal ion that displayed a significant difference between asthma status classifications. The mean EBC formate concentration was 40% higher in samples collected from all asthmatics than from healthy controls (mean = 5.7 microM, mean+/-standard deviation = 3.1-10.3 microM vs. 4.0, 2.8-5.8 microM, p = 0.05. EBC formate was higher in severe asthmatics than in mild-to-moderate asthmatics (6.8, 3.7-12.3 microM vs. 4.9, 2.8-8.7 microM, p = 0.012. In addition, formate concentration was negatively correlated with methacholine PC(20 (r = -0.39, p = 0.002, asthmatics only, and positively correlated with the NO-derived ion nitrite (r = 0.46, p<0.0001 as well as with total serum IgE (r = 0.28, p = 0.016, asthmatics only. Furthermore, formate was not significantly correlated with other volatile organic acids nor with inhaled corticosteroid dose. CONCLUSIONS: We conclude that EBC formate concentration is significantly higher in the breath of

  2. Gastric S-nitrosothiol formation drives the antihypertensive effects of oral sodium nitrite and nitrate in a rat model of renovascular hypertension.

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    Pinheiro, Lucas C; Amaral, Jefferson H; Ferreira, Graziele C; Portella, Rafael L; Ceron, Carla S; Montenegro, Marcelo F; Toledo, Jose Carlos; Tanus-Santos, Jose E

    2015-10-01

    Many effects of nitrite and nitrate are attributed to increased circulating concentrations of nitrite, ultimately converted into nitric oxide (NO(•)) in the circulation or in tissues by mechanisms associated with nitrite reductase activity. However, nitrite generates NO(•) , nitrous anhydride, and other nitrosating species at low pH, and these reactions promote S-nitrosothiol formation when nitrites are in the stomach. We hypothesized that the antihypertensive effects of orally administered nitrite or nitrate involve the formation of S-nitrosothiols, and that those effects depend on gastric pH. The chronic effects of oral nitrite or nitrate were studied in two-kidney, one-clip (2K1C) hypertensive rats treated with omeprazole (or vehicle). Oral nitrite lowered blood pressure and increased plasma S-nitrosothiol concentrations independently of circulating nitrite levels. Increasing gastric pH with omeprazole did not affect the increases in plasma nitrite and nitrate levels found after treatment with nitrite. However, treatment with omeprazole severely attenuated the increases in plasma S-nitrosothiol concentrations and completely blunted the antihypertensive effects of nitrite. Confirming these findings, very similar results were found with oral nitrate. To further confirm the role of gastric S-nitrosothiol formation, we studied the effects of oral nitrite in hypertensive rats treated with the glutathione synthase inhibitor buthionine sulfoximine (BSO) to induce partial thiol depletion. BSO treatment attenuated the increases in S-nitrosothiol concentrations and antihypertensive effects of oral nitrite. These data show that gastric S-nitrosothiol formation drives the antihypertensive effects of oral nitrite or nitrate and has major implications, particularly to patients taking proton pump inhibitors. PMID:26159506

  3. S-nitrosothiols regulate cell-surface pH buffering by airway epithelial cells during the human immune response to rhinovirus.

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    Carraro, Silvia; Doherty, Joseph; Zaman, Khalequz; Gainov, Iain; Turner, Ronald; Vaughan, John; Hunt, John F; Márquez, Javier; Gaston, Benjamin

    2006-05-01

    Human rhinovirus infection is a common trigger for asthma exacerbations. Asthma exacerbations and rhinovirus infections are both associated with markedly decreased pH and ammonium levels in exhaled breath condensates. This observation is thought to be related, in part, to decreased activity of airway epithelial glutaminase. We studied whether direct rhinovirus infection and/or the host immune response to the infection decreased airway epithelial cell surface pH in vitro. Interferon-gamma and tumor necrosis factor-alpha, but not direct rhinovirus infection, decreased pH, an effect partly associated with decreased ammonium concentrations. This effect was 1) prevented by nitric oxide synthase inhibition; 2) independent of cyclic GMP; 3) associated with an increase in endogenous airway epithelial cell S-nitrosothiol concentration; 4) mimicked by the exogenous S-nitrosothiol, S-nitroso-N-acetyl cysteine; and 5) independent of glutaminase expression and activity. We then confirmed that decreased epithelial pH inhibits human rhinovirus replication in airway epithelial cells. These data suggest that a nitric oxide synthase-dependent host response to viral infection mediated by S-nitrosothiols, rather than direct infection itself, plays a role in decreased airway surface pH during human rhinovirus infection. This host immune response may serve to protect the lower airways from direct infection in the normal host. In patients with asthma, however, this fall in pH could be associated with the increased mucus production, augmented inflammatory cell degranulation, bronchoconstriction, and cough characteristic of an asthma exacerbation. PMID:16603595

  4. Differential regulation of metabolism by nitric oxide and S-nitrosothiols in endothelial cells

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    Diers, Anne R.; Broniowska, Katarzyna A.; Darley-Usmar, Victor M.; Hogg, Neil

    2011-01-01

    S-nitrosation of thiols in key proteins in cell signaling pathways is thought to be an important contributor to nitric oxide (NO)-dependent control of vascular (patho)physiology. Multiple metabolic enzymes are targets of both NO and S-nitrosation, including those involved in glycolysis and oxidative phosphorylation. Thus it is important to understand how these metabolic pathways are integrated by NO-dependent mechanisms. Here, we compared the effects of NO and S-nitrosation on both glycolysis...

  5. Selective trapping of SNO-BSA and GSNO by benzenesulfinic acid sodium salt: mechanistic study of thiosulphonate formation and feasibility as a protein S-nitrosothiol detection strategy

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    Reeves, Benjamin D.; Hilmer, Jonathan K.; Mellmann, Lisa; Hartzheim, Myra; Poffenberger, Kevin; Johnson, Keith; Joshi, Neelambari; Singel, David J.; Grieco, Paul A.

    2013-01-01

    The conversion of S-nitrosothiols to thiosulphonates by reaction with the sodium salt of benzenesulfinic acid (PhSO2Na) has been examined in detail with the exemplary substrates S-nitrosoglutathione (GSNO) and S-nitrosylated bovine serum albumin (SNO-BSA). The reaction stoichiometry (2:1, PhSO2Na:RSNO) and the rate law (first order in both PhSO2Na and RSNO) have been determined under mild acidic conditions (pH 4.0). The products have been identified as the corresponding thiosulphonates (GSSO2Ph and BSA-SSO2Ph) along with PhSO2NHOH obtained in a 1:1 ratio. GSH, GSSG, and BSA were unreactive to PhSO2Na. PMID:24187391

  6. Conversion of nitrite to nitric oxide at zinc via S-nitrosothiols.

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    Cardenas, Allan Jay P; Abelman, Rebecca; Warren, Timothy H

    2014-01-01

    Nitrite is an important reservoir of nitric oxide activity in the plasma and cells. Using a biomimetic model, we demonstrate the conversion of zinc-bound nitrite in the tris(pyrazolyl)borate complex (iPr2)TpZn(NO2) to the corresponding S-nitrosothiol RSNO and zinc thiolate (iPr2)TpZn-SR via reaction with thiols H-SR. Decomposition of the S-nitrosothiol formed releases nitric oxide gas. PMID:24217415

  7. S-Nitrosothiol Signaling in Respiratory Biology

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    Gaston, Benjamin; Singel, David; Doctor, Allan; Stamler, Jonathan S.

    2006-01-01

    Genetic and biochemical data demonstrate a pivotal role for S-nitrosothiols (SNOs) in mediating the actions of nitric oxide synthases (NOSs). SNOs serve to convey NO bioactivity and to regulate protein function. This understanding is of immediate interest to the pulmonary clinical and research communities. This article reviews the following: (1) biochemical and cellular evidence that SNOs in amino acids, peptides, and proteins elicit NOS-dependent signaling in the respiratory system and (2) s...

  8. Pulmonary alveolar epithelial uptake of S-nitrosothiols is regulated by L-type amino acid transporter.

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    Granillo, Olivia M; Brahmajothi, Mulugu V; Li, Sheng; Whorton, A Richard; Mason, S Nicholas; McMahon, Timothy J; Auten, Richard L

    2008-07-01

    Nitric oxide (NO) effects are often mediated via S-nitrosothiol (SNO) formation; SNO uptake has recently been shown to be mediated in some cell types via system L-type amino acid transporters (LAT-1, 2). Inhaled NO therapy may exert some biological effects via SNO formation. We therefore sought to determine if pulmonary epithelial SNO uptake depended on LAT or peptide transporter 2 (PEPT2). Both LAT-1 and PEPT2 proteins were detected by immunoblot and immunocytochemistry in L2 cells and rat lung. We tested SNO uptake through the transporters by exposing rat alveolar epithelial cells (L2 and type II) to RSNOs: S-nitrosoglutathione, S-nitrosocysteinylglycine (SNO-Cys-Gly), S-nitrosocysteine (CSNO), and to NO donor diethylamine NONOate (DEA-NONOate). SNO was detected in cell lysates by ozone chemiluminescence. NO uptake was detected by fluorescence in alveolar epithelial cells loaded with 4-amino-5-methylamino-2',7'-difluorofluorescein (DAF-FM) diacetate cultured in submersion and exposed to RSNOs and DEA NONOate. Addition of L-Cys but not D-Cys to RSNOs or DEA NONOate increased SNO and DAF-FM signal that was inhibited by coincubation with LAT competitors. Incubation of cells with PEPT2 substrate SNO-Cys-Gly showed no increase in SNO or DAF-FM signal unless incubated with L-Cys. This was unaffected by PEPT2 inhibition. We conclude that RSNOs (thionitrites, S-nitrosothiols) and NO enter alveolar epithelial cells predominantly by S-nitrosation of L-Cys, which is then imported through LAT.

  9. Nitrosopersulfide (SSNO− accounts for sustained NO bioactivity of S-nitrosothiols following reaction with sulfide

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    Miriam M. Cortese-Krott

    2014-01-01

    Full Text Available Sulfide salts are known to promote the release of nitric oxide (NO from S-nitrosothiols and potentiate their vasorelaxant activity, but much of the cross-talk between hydrogen sulfide and NO is believed to occur via functional interactions of cell regulatory elements such as phosphodiesterases. Using RFL-6 cells as an NO reporter system we sought to investigate whether sulfide can also modulate nitrosothiol-mediated soluble guanylyl cyclase (sGC activation following direct chemical interaction. We find a U-shaped dose response relationship where low sulfide concentrations attenuate sGC stimulation by S-nitrosopenicillamine (SNAP and cyclic GMP levels are restored at equimolar ratios. Similar results are observed when intracellular sulfide levels are raised by pre-incubation with the sulfide donor, GYY4137. The outcome of direct sulfide/nitrosothiol interactions also critically depends on molar reactant ratios and is accompanied by oxygen consumption. With sulfide in excess, a ‘yellow compound’ accumulates that is indistinguishable from the product of solid-phase transnitrosation of either hydrosulfide or hydrodisulfide and assigned to be nitrosopersulfide (perthionitrite, SSNO−; λmax 412 nm in aqueous buffers, pH 7.4; 448 nm in DMF. Time-resolved chemiluminescence and UV–visible spectroscopy analyses suggest that its generation is preceded by formation of the short-lived NO-donor, thionitrite (SNO−. In contrast to the latter, SSNO− is rather stable at physiological pH and generates both NO and polysulfides on decomposition, resulting in sustained potentiation of SNAP-induced sGC stimulation. Thus, sulfide reacts with nitrosothiols to form multiple bioactive products; SSNO− rather than SNO− may account for some of the longer-lived effects of nitrosothiols and contribute to sulfide and NO signaling.

  10. Quantification and Localization of S-Nitrosothiols (SNOs) in Higher Plants.

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    Barroso, Juan B; Valderrama, Raquel; Carreras, Alfonso; Chaki, Mounira; Begara-Morales, Juan C; Sánchez-Calvo, Beatriz; Corpas, Francisco J

    2016-01-01

    S-nitrosothiols (SNOs) are a family of molecules produced by the reaction of nitric oxide (NO) with -SH thiol groups present in the cysteine residues of proteins and peptides caused by a posttranslational modification (PTM) known as S-nitrosylation (strictly speaking S-nitrosation) that can affect the cellular function of proteins. These molecules are a relatively more stable form of NO and consequently can act as a major intracellular NO reservoir and, in some cases, as a long-distance NO signal. Additionally, SNOs can be transferred between small peptides and protein thiol groups through S-transnitrosylation mechanisms. Thus, detection and cellular localization of SNOs in plant cells can be useful tools to determine how these molecules are modulated under physiological and adverse conditions and to determine their importance as a mechanism for regulating different biochemical pathways. Using a highly sensitive chemiluminescence ozone technique and a specific fluorescence probe (Alexa Fluor 488 Hg-link phenylmercury), the methods described in this chapter enable us to determine SNOs in an nM range as well as their cellular distribution in the tissues of different plant species. PMID:27094417

  11. Decomposition of S-Nitrosothiols Induced by UV and Sunlight

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    Manoj M. Veleeparampil

    2009-01-01

    Full Text Available Photochemical release of nitric oxide (NO from the S-nitroso derivatives of glutathione, L-cysteine, N-acetyl-L-cysteine, L-cysteinemethylester, D,L-penicillamine, N-acetyl-D,L-penicillamine, and N-acetylcysteamine has been investigated at neutral and acidic pH. The release of NO from RSNO is one of the key reactions that could be utilized in photodynamic therapy. The UV-VIS and HPLC analyses have shown that under argon saturated conditions, disulfide (RSSR is the major product of UV as well as sunlight induced decomposition. While in aerated conditions, nitirite—the end product of the oxidation of NO—was also observed along with disulfide. The formation of thiyl radical as the intermediate was reconfirmed by laser flash photolysis. The initial rate of formation of NO was on the order of 10−10dm3mol−1s−1. The quantum yields of these reactions were in the range of 0.2–0.8. The high quantum yields observed in the photo induced release of NO from RSNO using both UV and sunlight demonstrate the potential application of these reactions in photodynamic therapy.

  12. S-nitrosothiol tethered polymer hexagons: synthesis, characterisation and antibacterial effect.

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    Priya, S; Nithya, R; Berchmans, Sheela

    2014-01-01

    In this work, we portray a new controlled nitric oxide (NO) delivery platform by grafting S-nitrosothiol derived from cysteine into the polymeric backbone of poly(vinyl methyl ether-co-maleic anhydride). Nitrosothiols (RSNO's) are linked to the polymeric backbone through solvent displacement method. By adjusting solvent polarity, materials of different shapes and sizes varying between μm and nm are prepared. More often our method of preparation resulted in hexagonally shaped polymeric materials. The structure and RSNO conjugation analysis was investigated using scanning electron microscopy (SEM), FT-IR, UV-Vis spectroscopy and thermogravimetric analysis (TGA). Bactericidal efficacy of nitric oxide releasing polymer hexagons, a novel antibacterial agent is demonstrated against Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. Confocal microscopic studies revealed the enhanced bactericidal effect of polymer hexagons via membrane destruction. Results suggest that this biocompatible NO releasing RSNO conjugated polymer hexagons could be potentially useful for antimicrobial applications.

  13. S-Nitrosothiols modulate G protein-coupled receptor signaling in a reversible and highly receptor-specific manner

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    Mönkkönen Kati S

    2005-04-01

    Full Text Available Abstract Background Recent studies indicate that the G protein-coupled receptor (GPCR signaling machinery can serve as a direct target of reactive oxygen species, including nitric oxide (NO and S-nitrosothiols (RSNOs. To gain a broader view into the way that receptor-dependent G protein activation – an early step in signal transduction – might be affected by RSNOs, we have studied several receptors coupling to the Gi family of G proteins in their native cellular environment using the powerful functional approach of [35S]GTPγS autoradiography with brain cryostat sections in combination with classical G protein activation assays. Results We demonstrate that RSNOs, like S-nitrosoglutathione (GSNO and S-nitrosocysteine (CysNO, can modulate GPCR signaling via reversible, thiol-sensitive mechanisms probably involving S-nitrosylation. RSNOs are capable of very targeted regulation, as they potentiate the signaling of some receptors (exemplified by the M2/M4 muscarinic cholinergic receptors, inhibit others (P2Y12 purinergic, LPA1lysophosphatidic acid, and cannabinoid CB1 receptors, but may only marginally affect signaling of others, such as adenosine A1, μ-opioid, and opiate related receptors. Amplification of M2/M4 muscarinic responses is explained by an accelerated rate of guanine nucleotide exchange, as well as an increased number of high-affinity [35S]GTPγS binding sites available for the agonist-activated receptor. GSNO amplified human M4 receptor signaling also under heterologous expression in CHO cells, but the effect diminished with increasing constitutive receptor activity. RSNOs markedly inhibited P2Y12 receptor signaling in native tissues (rat brain and human platelets, but failed to affect human P2Y12 receptor signaling under heterologous expression in CHO cells, indicating that the native cellular signaling partners, rather than the P2Y12 receptor protein, act as a molecular target for this action. Conclusion These in vitro studies

  14. S-nitrosothiol repletion by an inhaled gas regulates pulmonary function

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    Moya, Martin P.; Gow, Andrew J.; McMahon, Timothy J.; Toone, Eric J.; Cheifetz, Ira M.; Goldberg, Ronald N.; Stamler, Jonathan S.

    2001-05-01

    NO synthases are widely distributed in the lung and are extensively involved in the control of airway and vascular homeostasis. It is recognized, however, that the O2-rich environment of the lung may predispose NO toward toxicity. These Janus faces of NO are manifest in recent clinical trials with inhaled NO gas, which has shown therapeutic benefit in some patient populations but increased morbidity in others. In the airways and circulation of humans, most NO bioactivity is packaged in the form of S-nitrosothiols (SNOs), which are relatively resistant to toxic reactions with O2/O. This finding has led to the proposition that channeling of NO into SNOs may provide a natural defense against lung toxicity. The means to selectively manipulate the SNO pool, however, has not been previously possible. Here we report on a gas, O-nitrosoethanol (ENO), which does not react with O2 or release NO and which markedly increases the concentration of indigenous species of SNO within airway lining fluid. Inhalation of ENO provided immediate relief from hypoxic pulmonary vasoconstriction without affecting systemic hemodynamics. Further, in a porcine model of lung injury, there was no rebound in cardiopulmonary hemodynamics or fall in oxygenation on stopping the drug (as seen with NO gas), and additionally ENO protected against a decline in cardiac output. Our data suggest that SNOs within the lung serve in matching ventilation to perfusion, and can be manipulated for therapeutic gain. Thus, ENO may be of particular benefit to patients with pulmonary hypertension, hypoxemia, and/or right heart failure, and may offer a new therapeutic approach in disorders such as asthma and cystic fibrosis, where the airways may be depleted of SNOs.

  15. Formation of Vascular S-Nitrosothiols and Plasma Nitrates/Nitrites Following Inhalation of Diesel Emissions

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    Knuckles, Travis L.; Buntz, Jennifer G.; Paffett, Michael; Channell, Meghan; Harmon, Molly; Cherng, Tom; Lucas, Selita N.; Jacob D. McDonald; Kanagy, Nancy L.; Matthew J Campen

    2011-01-01

    Epidemiological studies have associated traffic-related airborne pollution with adverse cardiovascular outcomes. Nitric oxide (NO) is a common component of fresh diesel and gasoline engine emissions that rapidly transforms both in the atmosphere and once inhaled. Because of this rapid transformation, limited information is available in terms of potential human exposures and adverse health effects. Young rats were exposed to whole diesel emissions (DE) adjusted to 300 µg/m3 of particulate matt...

  16. Enhanced flow injection analysis for measurements of S-nitrosothiols species in biological samples using highly selective amperometric nitric oxide sensor

    Institute of Scientific and Technical Information of China (English)

    Chun Cui Huang; Hui Bo Shao

    2012-01-01

    A highly selective nitric oxide (NO) sensor is fabricated and applied to devise an enhanced flow injection analysis (FIA) system for S-nitrosothiols (RSNOs) measurement in biological samples.The NO sensor is prepared using a polytetrafluoroethylene (PTFE) gas-permeable membrane loaded with Teflon AF(R) solution,a copolymer of tetrafluoroethylene and 2,2-bis(trifluoroethylene)-4,5-difluoro-1,3-dioxole,to improve selectivity.This method is much simpler and possesses good performance over a wide range of RSNOs concentrations.Standard deviation for three parallel measurements of blood plasma is 4.0%.The use of the gas sensing configuration as the detector enhances selectivity of the FIA measurement vs.using less selective electrochemical detectors that do not use PTFE/Teflon type outer membranes.

  17. Evidence that the nitrergic neurotransmitter and endothelium-derived relaxing factor might be S-nitrosothiols in the mouse corpus cavernosum.

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    Büyükafşar K

    1999-10-01

    Full Text Available The effects of thimerosal, a sulfhydryl oxidizing agent on nitrergic, endothelium-dependent and -independent relaxations were investigated to examine the possibility that the nitrergic neurotransmitter and endothelium-derived relaxing factor (EDRF could be S-nitrosothiol or free nitric oxide (NO in the isolated mouse corpus cavernosum. Thimerosal (5 x 10(-6-2 x 10(-5 M inhibited or almost abolished electrical field stimulation--(EFS, 30V, 0.5 ms, 15 sec, 1, 2, 4, 8, 16 Hz, acetylcholine--(ACh, 5 x 10(-8-1.25 x 10(-6 M, glyceryl trinitrate--(GTN, 3 x 10(-7-3 x 10(-6 M, and S-nitrosoglutathione--(GSNO, 5 x 10(-6-1.25 x 10(-4 M induced relaxations. Thiomerosal inhibition seems to be specific to L-arginine NO pathways since it had no effect on acidified sodium nitrite--(10(-4-5 x 10(-4 M, photoactivated sodium nitrite--(2 x 10(-4 M, isoprenaline--(10(-6 M, or papaverine--(10(-4 M elicited relaxations. Moreover, the inhibitory effect of thimerosal on the nitrergic, ACh- or GTN-induced relaxations were partly reversed by sulfhydryl-containing compounds, L-cysteine (10(-3 M, dithiothreitol (10(-3 M, or glutathione (10(-3 M. However L-methionine (10(-3 M, which contains a methyl group on the sulphur atom, failed to restore the thimerosal inhibition. Thimerosal did not change the contraction produced by 10(-4 M NG-nitro-L-arginine methyl ester. These findings indicate that the nitrergic neurotransmitter as well as EDRF may not be free NO but NO-transferring molecules, probably S-nitrosothiols, in the mouse corpus cavernosum.

  18. S-nitrosothiols dilate the mesenteric artery more potently than the femoral artery by a cGMP and L-type calcium channel-dependent mechanism.

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    Liu, Taiming; Schroeder, Hobe J; Zhang, Meijuan; Wilson, Sean M; Terry, Michael H; Longo, Lawrence D; Power, Gordon G; Blood, Arlin B

    2016-08-31

    S-nitrosothiols (SNOs) are metabolites of NO with potent vasodilatory activity. Our previous studies in sheep indicated that intra-arterially infused SNOs dilate the mesenteric vasculature more than the femoral vasculature. We hypothesized that the mesenteric artery is more responsive to SNO-mediated vasodilation, and investigated various steps along the NO/cGMP pathway to determine the mechanism for this difference. In anesthetized adult sheep, we monitored the conductance of mesenteric and femoral arteries during infusion of S-nitroso-l-cysteine (L-cysNO), and found mesenteric vascular conductance increased (137 ± 3%) significantly more than femoral conductance (26 ± 25%). Similar results were found in wire myography studies of isolated sheep mesenteric and femoral arteries. Vasodilation by SNOs was attenuated in both vessel types by the presence of ODQ (sGC inhibitor), and both YC-1 (sGC agonist) and 8-Br-cGMP (cGMP analog) mediated more potent relaxation in mesenteric arteries than femoral arteries. The vasodilatory difference between mesenteric and femoral arteries was eliminated by antagonists of either protein kinase G or L-type Ca(2+) channels. Western immunoblots showed a larger L-type Ca(2+)/sGC abundance ratio in mesenteric arteries than in femoral arteries. Fetal sheep mesenteric arteries were more responsive to SNOs than adult mesenteric arteries, and had a greater L-Ca(2+)/sGC ratio (p = 0.047 and r = -0.906 for correlation between Emax and L-Ca(2+)/sGC). These results suggest that mesenteric arteries, especially those in fetus, are more responsive to SNO-mediated vasodilation than femoral arteries due to a greater role of the L-type calcium channel in the NO/cGMP pathway.

  19. S-nitrosothiols dilate the mesenteric artery more potently than the femoral artery by a cGMP and L-type calcium channel-dependent mechanism.

    Science.gov (United States)

    Liu, Taiming; Schroeder, Hobe J; Zhang, Meijuan; Wilson, Sean M; Terry, Michael H; Longo, Lawrence D; Power, Gordon G; Blood, Arlin B

    2016-08-31

    S-nitrosothiols (SNOs) are metabolites of NO with potent vasodilatory activity. Our previous studies in sheep indicated that intra-arterially infused SNOs dilate the mesenteric vasculature more than the femoral vasculature. We hypothesized that the mesenteric artery is more responsive to SNO-mediated vasodilation, and investigated various steps along the NO/cGMP pathway to determine the mechanism for this difference. In anesthetized adult sheep, we monitored the conductance of mesenteric and femoral arteries during infusion of S-nitroso-l-cysteine (L-cysNO), and found mesenteric vascular conductance increased (137 ± 3%) significantly more than femoral conductance (26 ± 25%). Similar results were found in wire myography studies of isolated sheep mesenteric and femoral arteries. Vasodilation by SNOs was attenuated in both vessel types by the presence of ODQ (sGC inhibitor), and both YC-1 (sGC agonist) and 8-Br-cGMP (cGMP analog) mediated more potent relaxation in mesenteric arteries than femoral arteries. The vasodilatory difference between mesenteric and femoral arteries was eliminated by antagonists of either protein kinase G or L-type Ca(2+) channels. Western immunoblots showed a larger L-type Ca(2+)/sGC abundance ratio in mesenteric arteries than in femoral arteries. Fetal sheep mesenteric arteries were more responsive to SNOs than adult mesenteric arteries, and had a greater L-Ca(2+)/sGC ratio (p = 0.047 and r = -0.906 for correlation between Emax and L-Ca(2+)/sGC). These results suggest that mesenteric arteries, especially those in fetus, are more responsive to SNO-mediated vasodilation than femoral arteries due to a greater role of the L-type calcium channel in the NO/cGMP pathway. PMID:27235767

  20. Direct formate fuel cells: A review

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    An, L.; Chen, R.

    2016-07-01

    Direct formate fuel cells (DFFC), which convert the chemical energy stored in formate directly into electricity, are recently attracting more attention, primarily because of the use of the carbon-neutral fuel and the low-cost electrocatalytic and membrane materials. As an emerging energy technology, the DFFC has made a rapid progress in recent years (currently, the state-of-the-art power density is 591 mW cm-2 at 60 °C). This article provides a review of past research on the development of this type of fuel cell, including the working principle, mechanisms and materials of the electrocatalytic oxidation of formate, singe-cell designs and performance, as well as innovative system designs. In addition, future perspectives with regard to the development of this fuel cell system are also highlighted.

  1. Formation and separation of root border cells.

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    Driouich, Azeddine; Durand, Caroline; Vicré-Gibouin, Maïté

    2007-01-01

    Plant roots release a large number of border cells into the rhizosphere, which are believed to play a key role in root development and health. The formation and loss of these cells from the root cap region is a developmentally regulated process that is also controlled by phytohormones and environmental factors. The separation of border cells involves the complete dissociation of individual cells from each other and from root tissue. This process requires the activity of cell wall-degrading enzymes that solubilize the cell wall connections between cells. We present and discuss the solubilization process with an emphasis on pectin-degrading enzymes as well as the recently discovered root border-like cells of Arabidopsis thaliana.

  2. Organic Tandem Solar Cells: Design and Formation

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    Chen, Chun-Chao

    polyelectrolyte layer functioning as the surface dipole formation layer to provide better electrical contact with the photoactive layer. Due to the effectiveness of the conjugated polyelectrolyte layer, performance improvement was also observed. Furthermore, other issues regarding the semi-transparent tandem solar cells (e.g., photocurrent matching, exterior color tuning, and transparency tuning) are all explored to optimize best performance. In Chapter 5 and 6, the architectures of double- and triple-junction tandem solar cells are explored. Theoretically, triple-junction tandem solar cells with three photoactive absorbers with cascaded energy bandgaps have the potential to achieve higher performance, in comparison with double-junction tandem solar cells. Such expectations can be ascribed to the minimized carrier thermalization loss and further improved light absorption. However, the design of triple-junction solar cells often involves sophisticated multiple layer deposition as well as substantial optimization. Therefore, there is a lack of successful demonstrations of triple-junction solar cells outperforming the double-junction counterparts. To solve the incompatible issues related to the layer deposition in the fabrication, we proposed a novel architecture of inverted-structure tandem solar cells with newly designed interconnecting layers. Our design of interconnecting layers does not only focus on maintaining the orthogonal solution processing advantages, but also provides an excellent compatibility in the energy level alignment to allow different absorber materials to be used. Furthermore, we also explored the light management inside the double- and triple-junction tandem solar cells. The study of light management was carried out through optical simulation method based transfer matrix formalism. The intention is to obtain a balanced photocurrent output from each subcells inside the tandem solar cell, thus the minimal recombination loss at the contact of interconnecting

  3. Comparison of Cell formation techniques in Cellular manufacturing using three cell formation algorithms

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    Prabhat Kumar Giri

    2016-01-01

    Full Text Available In the present era of globalization and competitive market, cellular manufacturing has become a vital tool for meeting the challenges of improving productivity, which is the way to sustain growth. Getting best results of cellular manufacturing depends on the formation of the machine cells and part families. This paper examines advantages of ART method of cell formation over array based clustering algorithms, namely ROC-2 and DCA. The comparison and evaluation of the cell formation methods has been carried out in the study. The most appropriate approach is selected and used to form the cellular manufacturing system. The comparison and evaluation is done on the basis of performance measure as grouping efficiency and improvements over the existing cellular manufacturing system is presented.

  4. Formative cell divisions: principal determinants of plant morphogenesis.

    Science.gov (United States)

    Smolarkiewicz, Michalina; Dhonukshe, Pankaj

    2013-03-01

    Formative cell divisions utilizing precise rotations of cell division planes generate and spatially place asymmetric daughters to produce different cell layers. Therefore, by shaping tissues and organs, formative cell divisions dictate multicellular morphogenesis. In animal formative cell divisions, the orientation of the mitotic spindle and cell division planes relies on intrinsic and extrinsic cortical polarity cues. Plants lack known key players from animals, and cell division planes are determined prior to the mitotic spindle stage. Therefore, it appears that plants have evolved specialized mechanisms to execute formative cell divisions. Despite their profound influence on plant architecture, molecular players and cellular mechanisms regulating formative divisions in plants are not well understood. This is because formative cell divisions in plants have been difficult to track owing to their submerged positions and imprecise timings of occurrence. However, by identifying a spatiotemporally inducible cell division plane switch system applicable for advanced microscopy techniques, recent studies have begun to uncover molecular modules and mechanisms for formative cell divisions. The identified molecular modules comprise developmentally triggered transcriptional cascades feeding onto microtubule regulators that now allow dissection of the hierarchy of the events at better spatiotemporal resolutions. Here, we survey the current advances in understanding of formative cell divisions in plants in the context of embryogenesis, stem cell functionality and post-embryonic organ formation. PMID:23248201

  5. Radiation induced formation of giant cells (Saccharomyces uvarum). Pt. 1

    International Nuclear Information System (INIS)

    X-irradiated yeast cells (Saccharomyces uvarum) grown in liquid media stop mitosis and form giant cells. Chitin ring formation, being a prerequisite for cell separation, was studied by fluorescence microscopy using Calcofluor White, a chitin specific dye. Experiments with inhibitors of DNA synthesis (hydroxyurea) and chitin synthesis (polyoxin D) demonstrate chitin ring formation to be dependent on DNA synthesis, whereas bud formation is independent of DNA synthesis and chitin ring formation respectively. Basing on these results the formation of X-ray induced giant cells implies one DNA replication which in turn induces the formation of only one chitin ring between mother cell and giant bud. Obviously no septum can be formed. Thus cell separation does not occur, but the bud already formed, produces another bud demonstrating that bud formation itself is independent of DNA synthesis. (orig.)

  6. Regulatory T cell identity: formation and maintenance

    OpenAIRE

    Li, Xudong; Zheng, Ye

    2015-01-01

    T regulatory (Treg) cells are central to the maintenance of immune homeostasis. The transcription factor Foxp3 is essential for specifying the Treg cell lineage during development, and continued expression of Foxp3 in mature Treg cells is necessary for suppressive function. Treg cells can lose Foxp3 expression under certain conditions, and this is associated with autoimmune pathology. Here we review recent insights into the mechanisms that maintain Treg cell stability and function, and place ...

  7. Development of confocal immunofluorescence FRET microscopy to Investigate eNOS and GSNOR localization and interaction in pulmonary endothelial cells

    Science.gov (United States)

    Rehman, Shagufta; Brown-Steinke, Kathleen; Palmer, Lisa; Periasamy, Ammasi

    2015-03-01

    Confocal FRET microscopy is a widely used technique for studying protein-protein interactions in live or fixed cells. Endothelial nitric oxide synthase (eNOS) and S-nitrosoglutathione reductase (GSNOR) are enzymes involved in regulating the bioavailability of S-nitrosothiols (SNOs) in the pulmonary endothelium and have roles in the development of pulmonary arterial hypertension. Labeling of endogenous proteins to better understand a disease process can be challenging. We have used immunofluorescence to detect endogenous eNOS and GSNOR in primary pulmonary endothelial cells to co-localize these proteins as well as to study their interaction by FRET. The challenge has been in selecting the right immunofluorescence labeling condition, right antibody, the right blocking reagent, the right FRET pair and eliminating cross-reactivity of secondary antibodies. We have used Alexa488 and Alexa568 as a FRET pair. After a series of optimizations, the data from Confocal Laser Scanning Microscopy (CLSM) demonstrate co-localization of eNOS and GSNOR in the perinuclear region of the pulmonary endothelial cell primarily within the cis-Golgi with lower levels of co-localization seen within the trans-Golgi. FRET studies demonstrate, for the first time, interaction between eNOS and GSNOR in both murine and bovine pulmonary endothelial cells. Further characterization of eNOSGSNOR interaction and the subcellular location of this interaction will provide mechanistic insight into the importance of S-nitrosothiol signaling in pulmonary biology, physiology and pathology.

  8. Formation and cultivation of medaka primordial germ cells.

    Science.gov (United States)

    Li, Zhendong; Li, Mingyou; Hong, Ni; Yi, Meisheng; Hong, Yunhan

    2014-07-01

    Primordial germ cell (PGC) formation is pivotal for fertility. Mammalian PGCs are epigenetically induced without the need for maternal factors and can also be derived in culture from pluripotent stem cells. In egg-laying animals such as Drosophila and zebrafish, PGCs are specified by maternal germ plasm factors without the need for inducing factors. In these organisms, PGC formation and cultivation in vitro from indeterminate embryonic cells have not been possible. Here, we report PGC formation and cultivation in vitro from blastomeres dissociated from midblastula embryos (MBEs) of the fish medaka (Oryzias latipes). PGCs were identified by using germ-cell-specific green fluorescent protein (GFP) expression from a transgene under the control of the vasa promoter. Embryo perturbation was exploited to study PGC formation in vivo, and dissociated MBE cells were cultivated under various conditions to study PGC formation in vitro. Perturbation of somatic development did not prevent PGC formation in live embryos. Dissociated MBE blastomeres formed PGCs in the absence of normal somatic structures and of known inducing factors. Most importantly, under culture conditions conducive to stem cell derivation, some dissociated MBE blastomeres produced GFP-positive PGC-like cells. These GFP-positive cells contained genuine PGCs, as they expressed PGC markers and migrated into the embryonic gonad to generate germline chimeras. Our data thus provide evidence for PGC preformation in medaka and demonstrate, for the first time, that PGC formation and derivation can be obtained in culture from early embryos of medaka as a lower vertebrate model.

  9. Cell Competition Drives the Formation of Metastatic Tumors in a Drosophila Model of Epithelial Tumor Formation

    DEFF Research Database (Denmark)

    Eichenlaub, Teresa; Cohen, Stephen M; Herranz, Héctor

    2016-01-01

    Cell competition is a homeostatic process in which proliferating cells compete for survival. Elimination of otherwise normal healthy cells through competition is important during development and has recently been shown to contribute to maintaining tissue health during organismal aging. The...... mechanisms that allow for ongoing cell competition during adult life could, in principle, contribute to tumorigenesis. However, direct evidence supporting this hypothesis has been lacking. Here, we provide evidence that cell competition drives tumor formation in a Drosophila model of epithelial cancer. Cells...... Septin family protein Peanut. Cytokinesis failure due to downregulation of Peanut is required for tumorigenesis. This study provides evidence that the cellular mechanisms that drive cell competition during normal tissue growth can be co-opted to drive tumor formation and metastasis. Analogous mechanisms...

  10. The Role of Plant Hormones in Nematode Feeding Cell Formation

    NARCIS (Netherlands)

    Goverse, A.; Bird, D.

    2011-01-01

    In this Chapter, we discuss recent advances in the role of plant hormones in the molecular mechanisms underlying feeding cell formation both by cyst (CN) and root-knot nematodes (RKN). Phytohormones are small signalling molecules that regulate plant growth and development, including the formation of

  11. Suppression of T cell-induced osteoclast formation

    Energy Technology Data Exchange (ETDEWEB)

    Karieb, Sahar; Fox, Simon W., E-mail: Simon.fox@plymouth.ac.uk

    2013-07-12

    Highlights: •Genistein and coumestrol prevent activated T cell induced osteoclast formation. •Anti-TNF neutralising antibodies prevent the pro-osteoclastic effect of activated T cells. •Phytoestrogens inhibit T cell derived TNF alpha and inflammatory cytokine production. •Phytoestrogens have a broader range of anti-osteoclastic actions than other anti-resorptives. -- Abstract: Inhibition of T cell derived cytokine production could help suppress osteoclast differentiation in inflammatory skeletal disorders. Bisphosphonates are typically prescribed to prevent inflammatory bone loss but are not tolerated by all patients and are associated with an increased risk of osteonecrosis of the jaw. In light of this other anti-resorptives such as phytoestrogens are being considered. However the effect of phytoestrogens on T cell-induced osteoclast formation is unclear. The effect of genistein and coumestrol on activated T cell-induced osteoclastogenesis and cytokine production was therefore examined. Concentrations of genistein and coumestrol (10{sup −7} M) previously shown to directly inhibit osteoclast formation also suppressed the formation of TRAP positive osteoclast induced by con A activated T cells, which was dependent on inhibition of T cell derived TNF-α. While both reduced osteoclast formation their mechanism of action differed. The anti-osteoclastic effect of coumestrol was associated with a dual effect on con A induced T cell proliferation and activation; 10{sup −7} M coumestrol significantly reducing T cell number (0.36) and TNF-α (0.47), IL-1β (0.23) and IL-6 (0.35) expression, whereas genistein (10{sup −7} M) had no effect on T cell number but a more pronounced effect on T cell differentiation reducing expression of TNF-α (0.49), IL-1β (0.52), IL-6 (0.71) and RANKL (0.71). Phytoestrogens therefore prevent the pro-osteoclastic action of T cells suggesting they may have a role in the control of inflammatory bone loss.

  12. Nanopore formation in neuroblastoma cells following ultrashort electric pulse exposure

    Science.gov (United States)

    Roth, Caleb C.; Payne, Jason A.; Wilmink, Gerald J.; Ibey, Bennett L.

    2011-03-01

    Ultrashort or nanosecond electrical pulses (USEP) cause repairable damage to the plasma membranes of cells through formation of nanopores. These nanopores are able to pass small ions such as sodium, calcium, and potassium, but remain impermeable to larger molecules like trypan blue and propidium iodide. What remains uncertain is whether generation of nanopores by ultrashort electrical pulses can inhibit action potentials in excitable cells. In this paper, we explored the sensitivity of excitable cells to USEP using Calcium Green AM 1 ester fluorescence to measure calcium uptake indicative of nanopore formation in the plasma membrane. We determined the threshold for nanopore formation in neuroblastoma cells for three pulse parameters (amplitude, pulse width, and pulse number). Measurement of such thresholds will guide future studies to determine if USEP can inhibit action potentials without causing irreversible membrane damage.

  13. Asymmetric spindle pole formation in CPAP-depleted mitotic cells.

    Science.gov (United States)

    Lee, Miseon; Chang, Jaerak; Chang, Sunghoe; Lee, Kyung S; Rhee, Kunsoo

    2014-02-21

    CPAP is an essential component for centriole formation. Here, we report that CPAP is also critical for symmetric spindle pole formation during mitosis. We observed that pericentriolar material between the mitotic spindle poles were asymmetrically distributed in CPAP-depleted cells even with intact numbers of centrioles. The length of procentrioles was slightly reduced by CPAP depletion, but the length of mother centrioles was not affected. Surprisingly, the young mother centrioles of the CPAP-depleted cells are not fully matured, as evidenced by the absence of distal and subdistal appendage proteins. We propose that the selective absence of centriolar appendages at the young mother centrioles may be responsible for asymmetric spindle pole formation in CPAP-depleted cells. Our results suggest that the neural stem cells with CPAP mutations might form asymmetric spindle poles, which results in premature initiation of differentiation.

  14. Formation of photovoltaic modules based on polycrystalline solar cells

    OpenAIRE

    L.A. Dobrzański; A. Drygała; A. Januszka

    2009-01-01

    Purpose: The main aim of the paper is formation of photovoltaic modules and analysis of their main electric parameters.Design/methodology/approach: Photovoltaic modules were produced from four polycrystalline silicon solar cells, that were cut and next joined in series. Soft soldering technique and copper-tin strip were used for joining cells.Findings: In order to provide useful power for any application, the individual solar cells must be connected together to give the appropriate current an...

  15. Laminin 5 regulates polycystic kidney cell proliferation and cyst formation.

    Science.gov (United States)

    Joly, Dominique; Berissi, Sophie; Bertrand, Amélie; Strehl, Laetitia; Patey, Natacha; Knebelmann, Bertrand

    2006-09-29

    Renal cyst formation is the hallmark of autosomal dominant polycystic kidney disease (ADPKD). ADPKD cyst-lining cells have an increased proliferation rate and are surrounded by an abnormal extracellular matrix (ECM). We have previously shown that Laminin 5 (Ln-5, a alpha(3)beta(3)gamma(2) trimer) is aberrantly expressed in the pericystic ECM of ADPKD kidneys. We report that ADPKD cells in primary cultures produce and secrete Ln-5 that is incorporated to the pericystic ECM in an in vitro model of cystogenesis. In monolayers, purified Ln-5 induces ERK activation and proliferation of ADPKD cells, whereas upon epidermal growth factor stimulation blocking endogenously produced Ln-5 with anti-gamma(2) chain antibody reduces the sustained ERK activation and inhibits proliferation. In three-dimensional gel culture, addition of purified Ln-5 stimulates cell proliferation and cyst formation, whereas blocking endogenous Ln-5 strongly inhibits cyst formation. Ligation of alpha(6)beta(4) integrin, a major Ln-5 receptor aberrantly expressed by ADPKD cells, induces beta(4) integrin phosphorylation, ERK activation, cell proliferation, and cyst formation. These findings indicate that Ln-5 is an important regulator of ADPKD cell proliferation and cystogenesis and suggest that Ln-5 gamma(2) chain and Ln-5-alpha(6)beta(4) integrin interaction both contribute to these phenotypic changes. PMID:16870608

  16. A role for amyloid in cell aggregation and biofilm formation.

    Directory of Open Access Journals (Sweden)

    Melissa C Garcia

    Full Text Available Cell adhesion molecules in Saccharomyces cerevisiae and Candida albicans contain amyloid-forming sequences that are highly conserved. We have now used site-specific mutagenesis and specific peptide perturbants to explore amyloid-dependent activity in the Candida albicans adhesin Als5p. A V326N substitution in the amyloid-forming region conserved secondary structure and ligand binding, but abrogated formation of amyloid fibrils in soluble Als5p and reduced cell surface thioflavin T fluorescence. When displayed on the cell surface, Als5p with this substitution prevented formation of adhesion nanodomains and formation of large cellular aggregates and model biofilms. In addition, amyloid nanodomains were regulated by exogenous peptides. An amyloid-forming homologous peptide rescued aggregation and biofilm activity of Als5p(V326N cells, and V326N substitution peptide inhibited aggregation and biofilm activity in Als5p(WT cells. Therefore, specific site mutation, inhibition by anti-amyloid peturbants, and sequence-specificity of pro-amyloid and anti-amyloid peptides showed that amyloid formation is essential for nanodomain formation and activation.

  17. Formation of a cylindrical bridge in cell division

    Science.gov (United States)

    Citron, Daniel; Schmidt, Laura E.; Reichl, Elizabeth; Ren, Yixin; Robinson, Douglas; Zhang, Wendy W.

    2007-11-01

    In nature, the shape transition associated with the division of a mother cell into two daughter cells proceeds via a variety of routes. In the cylinder-thinning route, which has been observed in Dictyostelium and most animal cells, the mother cell first forms a broad bridge-like region, also known as a furrow, between two daughter cells. The furrow then rapidly evolves into a cylindrical bridge, which thins and eventually severs the mother cell into two. The fundamental mechanism underlying this division route is not understood. Recent experiments on Dictyostelium found that, while the cylinder-thinning route persists even when key actin cross-linking proteins are missing, it is disrupted by the removal of force-generating myosin-II proteins. Other measurements revealed that mutant cells lacking myosin-II have a much more uniform tension over the cell surface than wild-type cells. This suggests that tension variation may be important. Here we use a fluid model, previously shown to reproduce the thinning dynamics [Zhang & Robinson, PNAS 102, 7186 (2005)], to test this idea. Consistent with the experiments, the model shows that the cylinder formation process occurs regardless of the exact viscoelastic properties of the cell. In contrast to the experiments, a tension variation in the model hinders, rather then expedites, the cylinder formation.

  18. Aggregation of Red Blood Cells: From Rouleaux to Clot Formation

    CERN Document Server

    Wagner, C; Svetina, S

    2013-01-01

    Red blood cells are known to form aggregates in the form of rouleaux. This aggregation process is believed to be reversible, but there is still no full understanding on the binding mechanism. There are at least two competing models, based either on bridging or on depletion. We review recent experimental results on the single cell level and theoretical analyses of the depletion model and of the influence of the cell shape on the binding strength. Another important aggregation mechanism is caused by activation of platelets. This leads to clot formation which is life saving in the case of wound healing but also a major cause of death in the case of a thrombus induced stroke. We review historical and recent results on the participation of red blood cells in clot formation.

  19. Ancestral vascular lumen formation via basal cell surfaces.

    Directory of Open Access Journals (Sweden)

    Tomás Kucera

    Full Text Available The cardiovascular system of bilaterians developed from a common ancestor. However, no endothelial cells exist in invertebrates demonstrating that primitive cardiovascular tubes do not require this vertebrate-specific cell type in order to form. This raises the question of how cardiovascular tubes form in invertebrates? Here we discovered that in the invertebrate cephalochordate amphioxus, the basement membranes of endoderm and mesoderm line the lumen of the major vessels, namely aorta and heart. During amphioxus development a laminin-containing extracellular matrix (ECM was found to fill the space between the basal cell surfaces of endoderm and mesoderm along their anterior-posterior (A-P axes. Blood cells appear in this ECM-filled tubular space, coincident with the development of a vascular lumen. To get insight into the underlying cellular mechanism, we induced vessels in vitro with a cell polarity similar to the vessels of amphioxus. We show that basal cell surfaces can form a vascular lumen filled with ECM, and that phagocytotic blood cells can clear this luminal ECM to generate a patent vascular lumen. Therefore, our experiments suggest a mechanism of blood vessel formation via basal cell surfaces in amphioxus and possibly in other invertebrates that do not have any endothelial cells. In addition, a comparison between amphioxus and mouse shows that endothelial cells physically separate the basement membranes from the vascular lumen, suggesting that endothelial cells create cardiovascular tubes with a cell polarity of epithelial tubes in vertebrates and mammals.

  20. A Review of Cell Formation from Perspective of Objective Function

    Institute of Scientific and Technical Information of China (English)

    WANG Xiaoqing; TANG Jiafu

    2006-01-01

    The initial and significant step in the design of a cellular manufacturing system is cell formation (CF). CF problem is proposed in this paper as a decision problem that determines to manufacture specified types of part in a manufacturing plant which machines and their associated parts are grouped together to form cell in a way that a concerned objective is optimized. For describing CF problem clearly, this paper firstly presents a review of cell formation problem from the view points of objective function. The CF problems are classified into three categories, which are cost oriented, flexibility oriented and grouping efficiency oriented CF problems. Then, the paper presents a comprehensive conceptual mathematical formulation describing the general cost problem and a decision variable for comprehensive describing routing flexibility and two trade-off questions in grouping efficiency issues. Finally, based on the review and discussion, the paper proposes five directions for future research in the CF field.

  1. Role of polarized cell divisions in zebrafish neural tube formation.

    Science.gov (United States)

    Clarke, Jon

    2009-04-01

    Development of epithelial cell polarity and morphogenesis of a central lumen are essential prerequisites for the formation of the vertebrate neural tube. In teleost fish embryos this first involves the formation of a solid neural rod structure that then undergoes a process of cavitation to form a lumen. This process is initiated from a neural plate that has a distinct organization compared to other vertebrates, and involves complex cell intercalations and rearrangements. A key element is a mode of polarized cell division that generates daughters with mirror-image apico-basal polarity. These mirror-symmetric divisions have powerful morphogenetic influence because when they occur in ectopic locations they orchestrate the development of ectopic apical and basal specializations and the development of ectopic neural tubes.

  2. Effect of supercooling and cell volume on intracellular ice formation.

    Science.gov (United States)

    Prickett, Richelle C; Marquez-Curtis, Leah A; Elliott, Janet A W; McGann, Locksley E

    2015-04-01

    Intracellular ice formation (IIF) has been linked to death of cells cryopreserved in suspension. It has been assumed that cells can be supercooled by 2 to 10°C before IIF occurs, but measurements of the degree of supercooling that cells can tolerate are often confounded by changing extracellular temperature and solutions of different osmolality (which affect the cell volume). The purpose of this study was to examine how the incidence of IIF in the absence of cryoprotectants is affected by the degree of supercooling and cell volume. Human umbilical vein endothelial cells were suspended in isotonic (300 mOsm) and hypertonic (∼600 to 700 mOsm) solutions and exposed to supercooling ranging from 2 to 10°C before extracellular ice was nucleated. The number of cells undergoing IIF was examined in a cryostage (based on the darkening of cells upon intracellular freezing ("flashing")) as a function of the degree of supercooling, and cell survival post-thaw was assessed using a membrane integrity assay. We found that while the incidence of IIF increased with supercooling in both isotonic and hypertonic solutions, it was higher in the isotonic solution at any given degree of supercooling. Since cells in hypertonic solution were shrunken due to water efflux, we hypothesized that the difference in IIF behavior could be attributed to the decreased volume of cells in the hypertonic solution. Our results confirm that cells with a smaller diameter before extracellular ice nucleation have a decreased probability of IIF and suggest that cell volume could play a more significant role in the incidence of IIF than the extracellular ice nucleation temperature. PMID:25707695

  3. A hydrodynamic microchip for formation of continuous cell chains

    Science.gov (United States)

    Khoshmanesh, Khashayar; Zhang, Wei; Tang, Shi-Yang; Nasabi, Mahyar; Soffe, Rebecca; Tovar-Lopez, Francisco J.; Rajadas, Jayakumar; Mitchell, Arnan

    2014-05-01

    Here, we demonstrate the unique features of a hydrodynamic based microchip for creating continuous chains of model yeast cells. The system consists of a disk shaped microfluidic structure, containing narrow orifices that connect the main channel to an array of spoke channels. Negative pressure provided by a syringe pump draws fluid from the main channel through the narrow orifices. After cleaning process, a thin layer of water is left between the glass substrate and the polydimethylsiloxane microchip, enabling leakage beneath the channel walls. A mechanical clamp is used to adjust the operation of the microchip. Relaxing the clamp allows leakage of liquid beneath the walls in a controllable fashion, leading to formation of a long cell chain evenly distributed along the channel wall. The unique features of the microchip are demonstrated by creating long chains of yeast cells and model 15 μm polystyrene particles along the side wall and analysing the hydrogen peroxide induced death of patterned cells.

  4. Unequal distribution of plastids during generative cell formation in Impatiens.

    Science.gov (United States)

    van Went, J L

    1984-07-01

    This paper describes the unequal distribution of plastids in the developing microspores of Impatiens walleriana and Impatiens glandulifera which leads to the exclusion of plastids from the generative cell. During the development from young microspore to the onset of mitosis a change in the organization of the cytoplasm and distribution of organelles is gradually established. This includes the formation of vacuoles at the poles of the elongate-shaped microspores, the movement of the nucleus to a position near the microspore wall in the central part of the cell, and the accumulation of the plastids to a position near the wall at the opposite side of the cell. In Impatiens walleriana, the accumulated plastids are separated from each other by ER cisterns, and some mitochondria are also accumulated. In both Impatiens species, the portion of the microspore in which the generative cell will be formed is completely devoid of plastids at the time mitosis starts. PMID:24257638

  5. A peptide antagonist disrupts NK cell inhibitory synapse formation.

    Science.gov (United States)

    Borhis, Gwenoline; Ahmed, Parvin S; Mbiribindi, Bérénice; Naiyer, Mohammed M; Davis, Daniel M; Purbhoo, Marco A; Khakoo, Salim I

    2013-03-15

    Productive engagement of MHC class I by inhibitory NK cell receptors depends on the peptide bound by the MHC class I molecule. Peptide:MHC complexes that bind weakly to killer cell Ig-like receptors (KIRs) can antagonize the inhibition mediated by high-affinity peptide:MHC complexes and cause NK cell activation. We show that low-affinity peptide:MHC complexes stall inhibitory signaling at the step of Src homology protein tyrosine phosphatase 1 recruitment and do not go on to form the KIR microclusters induced by high-affinity peptide:MHC, which are associated with Vav dephosphorylation and downstream signaling. Furthermore, the low-affinity peptide:MHC complexes prevented the formation of KIR microclusters by high-affinity peptide:MHC. Thus, peptide antagonism of NK cells is an active phenomenon of inhibitory synapse disruption.

  6. Aroma formation by immobilized yeast cells in fermentation processes.

    Science.gov (United States)

    Nedović, V; Gibson, B; Mantzouridou, T F; Bugarski, B; Djordjević, V; Kalušević, A; Paraskevopoulou, A; Sandell, M; Šmogrovičová, D; Yilmaztekin, M

    2015-01-01

    Immobilized cell technology has shown a significant promotional effect on the fermentation of alcoholic beverages such as beer, wine and cider. However, genetic, morphological and physiological alterations occurring in immobilized yeast cells impact on aroma formation during fermentation processes. The focus of this review is exploitation of existing knowledge on the biochemistry and the biological role of flavour production in yeast for the biotechnological production of aroma compounds of industrial importance, by means of immobilized yeast. Various types of carrier materials and immobilization methods proposed for application in beer, wine, fruit wine, cider and mead production are presented. Engineering aspects with special emphasis on immobilized cell bioreactor design, operation and scale-up potential are also discussed. Ultimately, examples of products with improved quality properties within the alcoholic beverages are addressed, together with identification and description of the future perspectives and scope for cell immobilization in fermentation processes.

  7. Cell colony formation induced by Xenopus egg extract as a marker for improvement of cloned blastocyst formation in pig

    DEFF Research Database (Denmark)

    Liu, Ying; Østrup, Olga; Li, Juan;

    2011-01-01

    method based on the colony formation of cells after extract treatment, and subsequent in vitro cloning efficiency using treated cells as chromatin donors. Porcine fetal fibroblasts were treated with each batch of extract, and cultured in embryonic stem cell (ES) medium for 12 days. The number of forming...... colonies in treated cells was counted on Day 7 after extract treatment and significant variability was detected between different batches of extract. Similarly, when using cells from colonies at Days 7 to 8 after treatment for handmade cloning, increased blastocyst formation rates were observed after...... the cells were treated with a batch showing higher colony formation. In conclusion, assessment of cell colony formation may be used as selection marker for Xenopus egg extract used for pretreatment of donor cells prior to cloning....

  8. Blockade of mast cell activation reduces cutaneous scar formation.

    Science.gov (United States)

    Chen, Lin; Schrementi, Megan E; Ranzer, Matthew J; Wilgus, Traci A; DiPietro, Luisa A

    2014-01-01

    Damage to the skin initiates a cascade of well-orchestrated events that ultimately leads to repair of the wound. The inflammatory response is key to wound healing both through preventing infection and stimulating proliferation and remodeling of the skin. Mast cells within the tissue are one of the first immune cells to respond to trauma, and upon activation they release pro-inflammatory molecules to initiate recruitment of leukocytes and promote a vascular response in the tissue. Additionally, mast cells stimulate collagen synthesis by dermal fibroblasts, suggesting they may also influence scar formation. To examine the contribution of mast cells in tissue repair, we determined the effects the mast cell inhibitor, disodium cromoglycate (DSCG), on several parameters of dermal repair including, inflammation, re-epithelialization, collagen fiber organization, collagen ultrastructure, scar width and wound breaking strength. Mice treated with DSCG had significantly reduced levels of the inflammatory cytokines IL-1α, IL-1β, and CXCL1. Although DSCG treatment reduced the production of inflammatory mediators, the rate of re-epithelialization was not affected. Compared to control, inhibition of mast cell activity caused a significant decrease in scar width along with accelerated collagen re-organization. Despite the reduced scar width, DSCG treatment did not affect the breaking strength of the healed tissue. Tryptase β1 exclusively produced by mast cells was found to increase significantly in the course of wound healing. However, DSCG treatment did not change its level in the wounds. These results indicate that blockade of mast cell activation reduces scar formation and inflammation without further weakening the healed wound. PMID:24465509

  9. Blockade of mast cell activation reduces cutaneous scar formation.

    Science.gov (United States)

    Chen, Lin; Schrementi, Megan E; Ranzer, Matthew J; Wilgus, Traci A; DiPietro, Luisa A

    2014-01-01

    Damage to the skin initiates a cascade of well-orchestrated events that ultimately leads to repair of the wound. The inflammatory response is key to wound healing both through preventing infection and stimulating proliferation and remodeling of the skin. Mast cells within the tissue are one of the first immune cells to respond to trauma, and upon activation they release pro-inflammatory molecules to initiate recruitment of leukocytes and promote a vascular response in the tissue. Additionally, mast cells stimulate collagen synthesis by dermal fibroblasts, suggesting they may also influence scar formation. To examine the contribution of mast cells in tissue repair, we determined the effects the mast cell inhibitor, disodium cromoglycate (DSCG), on several parameters of dermal repair including, inflammation, re-epithelialization, collagen fiber organization, collagen ultrastructure, scar width and wound breaking strength. Mice treated with DSCG had significantly reduced levels of the inflammatory cytokines IL-1α, IL-1β, and CXCL1. Although DSCG treatment reduced the production of inflammatory mediators, the rate of re-epithelialization was not affected. Compared to control, inhibition of mast cell activity caused a significant decrease in scar width along with accelerated collagen re-organization. Despite the reduced scar width, DSCG treatment did not affect the breaking strength of the healed tissue. Tryptase β1 exclusively produced by mast cells was found to increase significantly in the course of wound healing. However, DSCG treatment did not change its level in the wounds. These results indicate that blockade of mast cell activation reduces scar formation and inflammation without further weakening the healed wound.

  10. Blockade of mast cell activation reduces cutaneous scar formation.

    Directory of Open Access Journals (Sweden)

    Lin Chen

    Full Text Available Damage to the skin initiates a cascade of well-orchestrated events that ultimately leads to repair of the wound. The inflammatory response is key to wound healing both through preventing infection and stimulating proliferation and remodeling of the skin. Mast cells within the tissue are one of the first immune cells to respond to trauma, and upon activation they release pro-inflammatory molecules to initiate recruitment of leukocytes and promote a vascular response in the tissue. Additionally, mast cells stimulate collagen synthesis by dermal fibroblasts, suggesting they may also influence scar formation. To examine the contribution of mast cells in tissue repair, we determined the effects the mast cell inhibitor, disodium cromoglycate (DSCG, on several parameters of dermal repair including, inflammation, re-epithelialization, collagen fiber organization, collagen ultrastructure, scar width and wound breaking strength. Mice treated with DSCG had significantly reduced levels of the inflammatory cytokines IL-1α, IL-1β, and CXCL1. Although DSCG treatment reduced the production of inflammatory mediators, the rate of re-epithelialization was not affected. Compared to control, inhibition of mast cell activity caused a significant decrease in scar width along with accelerated collagen re-organization. Despite the reduced scar width, DSCG treatment did not affect the breaking strength of the healed tissue. Tryptase β1 exclusively produced by mast cells was found to increase significantly in the course of wound healing. However, DSCG treatment did not change its level in the wounds. These results indicate that blockade of mast cell activation reduces scar formation and inflammation without further weakening the healed wound.

  11. Vesicle Size Regulates Nanotube Formation in the Cell

    OpenAIRE

    Qian Peter Su; Wanqing Du; Qinghua Ji; Boxin Xue; Dong Jiang; Yueyao Zhu; Jizhong Lou; Li Yu; Yujie Sun

    2016-01-01

    Intracellular membrane nanotube formation and its dynamics play important roles for cargo transportation and organelle biogenesis. Regarding the regulation mechanisms, while much attention has been paid on the lipid composition and its associated protein molecules, effects of the vesicle size has not been studied in the cell. Giant unilamellar vesicles (GUVs) are often used for in vitro membrane deformation studies, but they are much larger than most intracellular vesicles and the in vitro st...

  12. Biological Characteristics of Foam Cell Formation in Smooth Muscle Cells Derived from Bone Marrow Stem Cells

    Directory of Open Access Journals (Sweden)

    Pengke Yan, Chenglai Xia, Caiwen Duan, Shihuang Li, Zhengrong Mei

    2011-01-01

    Full Text Available Bone marrow mesenchymal stem cells (BMSC can differentiate into diverse cell types, including adipogenic, osteogenic, chondrogenic and myogenic lineages. There are lots of BMSC accumulated in atherosclerosis vessels and differentiate into VSMC. However, it is unclear whether VSMC originated from BMSC (BMSC-SMC could remodel the vessel in new tunica intima or promote the pathogenesis of atherosclerosis. In this study, BMSC were differentiated into VSMC in response to the transforming growth factor β (TGF-β and shown to express a number of VSMC markers, such as α-smooth muscle actin (α-SMA and smooth muscle myosin heavy chain1 (SM-MHC1. BMSC-SMC became foam cells after treatment with 80 mg/L ox-LDL for 72 hours. Ox-LDL could upregulate scavenger receptor class A (SR-A but downregulate the ATP-binding cassette transporter A1 (ABCA1 and caveolin-1 protein expression, suggesting that modulating relative protein activity contributes to smooth muscle foam cell formation in BMSC-SMC. Furthermore, we found that BMSC-SMC have some biological characteristics that are similar to VSMC, such as the ability of proliferation and secretion of extracellular matrix, but, at the same time, retain some biological characteristics of BMSC, such as a high level of migration. These results suggest that BMSC-SMC could be induced to foam cells and be involved in the development of atherosclerosis.

  13. Evagination of cells controls bio-silica formation and maturation during spicule formation in sponges.

    Directory of Open Access Journals (Sweden)

    Xiaohong Wang

    Full Text Available The enzymatic-silicatein mediated formation of the skeletal elements, the spicules of siliceous sponges starts intracellularly and is completed extracellularly. With Suberites domuncula we show that the axial growth of the spicules proceeds in three phases: (I formation of an axial canal; (II evagination of a cell process into the axial canal, and (III assembly of the axial filament composed of silicatein. During these phases the core part of the spicule is synthesized. Silicatein and its substrate silicate are stored in silicasomes, found both inside and outside of the cellular extension within the axial canal, as well as all around the spicule. The membranes of the silicasomes are interspersed by pores of ≈ 2 nm that are likely associated with aquaporin channels which are implicated in the hardening of the initial bio-silica products formed by silicatein. We can summarize the sequence of events that govern spicule formation as follows: differential GENETIC READOUT (of silicatein → FRACTAL ASSOCIATION of the silicateins → EVAGINATION of cells by hydro-mechanical forces into the axial canal → and finally PROCESSIVE BIO-SILICA POLYCONDENSATION around the axial canal. We termed this process, occurring sequentially or in parallel, BIO-INORGANIC SELF-ORGANIZATION.

  14. CELL FORMATION IN GROUP TECHNOLOGY: A SIMILARITY ORDER CLUSTERING APPROACH

    Directory of Open Access Journals (Sweden)

    Godfrey C. Onwubolu

    2012-01-01

    Full Text Available Grouping parts into families which can be produced by a cluster of machine cells is the cornerstone of cellular manufacturing, which in turn is the building block for flexible manufacturing systems. Cellular manufacturing is a group technology (GT concept that has recently attracted the attention of manufacturing firms operating under jobshop environment to consider redesigning their manufacturing systems so as to take advantage of increased throughput, reduction in work-in-progress, set-up time, and lead times; leading to product quality and customer satisfaction. The paper presents a generalised approach for machine cell formation from a jobshop using similarity order clustering technique for preliminary cell grouping and considering machine utilisation for the design of nonintergrouping material handling using the single-pass heuristic. The work addresses the shortcomings of cellular manufacturing systems design and implementations which ignore machine utilisations, group sizes and intergroup moves.

  15. Hydroxyl radical formation in phagocytic cells of the rat.

    Science.gov (United States)

    Drath, D B; Karnovsky, M L; Huber, G L

    1979-01-01

    Polymorphonuclear leukocytes (PMN) and macrophages, harvested from the peritoneum and lung, release superoxide (O-.2) and hydrogen peroxide (H2O2) during phagocytosis. These two agents are thought to react with each other to produce a highly active oxidative substance known as hydroxyl radical (OH.). We present evidence suggesting that these radicals are generated by phagocytic cells of the rat. Our findings are based upon an assay where ethylene gas is generated from methional by the action of this radical. Ethylene generation was shown to be inhibited by superoxide dismutase, catalase, and scavengers of OH.. Of the cells examined, PMN generated the most ethylene from methional, exhibiting a fourfold increase during phagocytosis. Pulmonary and peritoneal macrophages caused smaller amounts of this gas to be formed. Regardless of cell type, an intact cell was required for ethylene generation. Zymosan appeared to be the most effective particle for all cells in ethylene formation from methional, although opsonization was critical only for PMN. Ethylene generation was dependent on cell concentration to an extent and increased with time. PMID:222719

  16. [A new technic for LE cell formation. Preliminary report].

    Science.gov (United States)

    Gómez-Estrada, H; Fernández-Quintero, P; Toro-Pérez, A; Arellando-Blanco, J; Ochoa Díaz-López, H; Tapia-Arizmendi, G

    1976-01-01

    In special designed vials using coverslides glued to segments of polyethilene test tubes of 0.15 ml of fibrin free blood were incubated during 45 min to 37 degrees C in a humid chamber. Blood samples were obtained from patients with disseminated Lupus erythematosus (DLE) and normal subjects (N). Adhered polymorpho-nuclear cells (PNC) to glass were washed with Hank's solution; immediately 0.15 ml of DLE or N serum containing 5X10(6) lymphocytes (L) were added to culture cells and incubated at 37 degrees C during 30 min. Lymphocytes were previously incubated at 37 degrees C during 30 min with either N serum or DLE serum. Thereafter the segment of polyethilene test tube was detached from coverslides and cells attached to glass was washed with Hank's solution and stained with Wright solution. PMN of DLE and N in presence of L of DLE and N incubated with fresh serum of DLE formed 5 to 15% of LE cells. Determining factor for LE cells formation is the serum of DLE. Slides contained only PMN and LE cells which make easy the observation of results. All possible combinations with PMN and DLE serum and N serum allowed inclusion of several negative or positive control groups.

  17. Vesicle Size Regulates Nanotube Formation in the Cell

    Science.gov (United States)

    Su, Qian Peter; Du, Wanqing; Ji, Qinghua; Xue, Boxin; Jiang, Dong; Zhu, Yueyao; Lou, Jizhong; Yu, Li; Sun, Yujie

    2016-01-01

    Intracellular membrane nanotube formation and its dynamics play important roles for cargo transportation and organelle biogenesis. Regarding the regulation mechanisms, while much attention has been paid on the lipid composition and its associated protein molecules, effects of the vesicle size has not been studied in the cell. Giant unilamellar vesicles (GUVs) are often used for in vitro membrane deformation studies, but they are much larger than most intracellular vesicles and the in vitro studies also lack physiological relevance. Here, we use lysosomes and autolysosomes, whose sizes range between 100 nm and 1 μm, as model systems to study the size effects on nanotube formation both in vivo and in vitro. Single molecule observations indicate that driven by kinesin motors, small vesicles (100–200 nm) are mainly transported along the tracks while a remarkable portion of large vesicles (500–1000 nm) form nanotubes. This size effect is further confirmed by in vitro reconstitution assays on liposomes and purified lysosomes and autolysosomes. We also apply Atomic Force Microscopy (AFM) to measure the initiation force for nanotube formation. These results suggest that the size-dependence may be one of the mechanisms for cells to regulate cellular processes involving membrane-deformation, such as the timing of tubulation-mediated vesicle recycling. PMID:27052881

  18. Vesicle Size Regulates Nanotube Formation in the Cell.

    Science.gov (United States)

    Su, Qian Peter; Du, Wanqing; Ji, Qinghua; Xue, Boxin; Jiang, Dong; Zhu, Yueyao; Lou, Jizhong; Yu, Li; Sun, Yujie

    2016-04-07

    Intracellular membrane nanotube formation and its dynamics play important roles for cargo transportation and organelle biogenesis. Regarding the regulation mechanisms, while much attention has been paid on the lipid composition and its associated protein molecules, effects of the vesicle size has not been studied in the cell. Giant unilamellar vesicles (GUVs) are often used for in vitro membrane deformation studies, but they are much larger than most intracellular vesicles and the in vitro studies also lack physiological relevance. Here, we use lysosomes and autolysosomes, whose sizes range between 100 nm and 1 μm, as model systems to study the size effects on nanotube formation both in vivo and in vitro. Single molecule observations indicate that driven by kinesin motors, small vesicles (100-200 nm) are mainly transported along the tracks while a remarkable portion of large vesicles (500-1000 nm) form nanotubes. This size effect is further confirmed by in vitro reconstitution assays on liposomes and purified lysosomes and autolysosomes. We also apply Atomic Force Microscopy (AFM) to measure the initiation force for nanotube formation. These results suggest that the size-dependence may be one of the mechanisms for cells to regulate cellular processes involving membrane-deformation, such as the timing of tubulation-mediated vesicle recycling.

  19. Cell-cycle dependent micronucleus formation and mitotic disturbances induced by 5-azacytidine in mammalian cells

    OpenAIRE

    Stopper, Helga; Körber, C.; Schiffmann, D; Caspary, W J

    2012-01-01

    5-Azacytidine was originally developed to treat human myelogenous leukemia. However, interest in this compound has expanded because of reports of its ability to affect cell differentiation and to alter eukaryotic gene expression. In an ongoing attempt to understand the biochemical effects of this compound, we examined the effects of 5-azacytidine on mitosis and on micronucleus formation in mammalian cells. In L5178Y mouse cells, 5-azacytidine induced micronuclei at concentrations at which we ...

  20. Notch1-Dll4 signalling and mechanical force regulate leader cell formation during collective cell migration.

    Science.gov (United States)

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

    2015-03-13

    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 the 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 modelling to show that the leader cell identity is dynamically regulated by Dll4 signalling through both Notch1 and cellular stress in a migrating epithelium. Time-lapse microscopy reveals that Dll4 is induced in leader cells after the creation of the cell-free region and leader cells are regulated via Notch1-Dll4 lateral inhibition. Furthermore, mechanical stress inhibits Dll4 expression and leader cell formation in the monolayer. Collectively, our findings suggest that a reduction of mechanical force near the boundary promotes Notch1-Dll4 signalling to dynamically regulate the density of leader cells during collective cell migration.

  1. Dynamic Cell Formation based on Multi-objective Optimization Model

    Directory of Open Access Journals (Sweden)

    Guozhu Jia

    2013-08-01

    Full Text Available In this paper, a multi-objective model is proposed to address the dynamic cellular manufacturing (DCM formation problem. This model considers four conflicting objectives: relocation cost, machine utilization, material handling cost and maintenance cost. The model also considers the situation that some machines could be shared by more than one cell at the same period. A genetic algorithm is applied to get the solution of this mathematical model. Three numerical examples are simulated to evaluate the validity of this model.  

  2. EVOLUTIONARY ALGORITHMS WITH PREFERENCE FOR MANUFACTURING CELLS FORMATION

    Institute of Scientific and Technical Information of China (English)

    WANG Jianwei; WEI Xiaopeng; LI Rui

    2008-01-01

    Due to the combinatorial nature of cell formation problem and the characteristics of multi-objective and multi-constrain, a novel method of evolutionary algorithm with preference is proposed. The analytic hierarchy process (AHP) is adopted to determine scientifically the weights of the sub-objective functions. The satisfaction of constraints is considered as a new objective, the ratio of the population which doesn't satisfy all constraints is assigned as the weight of new objective. In addition, the self-adaptation of weights is applied in order to converge more easily towards the feasible domain. Therefore, both features multi-criteria and constrains are dealt with simultaneously. Finally, an example is selected from the literature to evaluate the performance of the proposed approach. The results validate the effectiveness of the proposed method in designing the manufacturing cells.

  3. Molecular Mechanisms for Vascular Development and Secondary Cell Wall Formation

    Science.gov (United States)

    Yang, Jung Hyun; Wang, Huanzhong

    2016-01-01

    Vascular tissues are important for transporting water and nutrients throughout the plant and as physical support of upright growth. The primary constituents of vascular tissues, xylem, and phloem, are derived from the meristematic vascular procambium and cambium. Xylem cells develop secondary cell walls (SCWs) that form the largest part of plant lignocellulosic biomass that serve as a renewable feedstock for biofuel production. For the last decade, research on vascular development and SCW biosynthesis has seen rapid progress due to the importance of these processes to plant biology and to the biofuel industry. Plant hormones, transcriptional regulators and peptide signaling regulate procambium/cambium proliferation, vascular patterning, and xylem differentiation. Transcriptional regulatory pathways play a pivot role in SCW biosynthesis. Although most of these discoveries are derived from research in Arabidopsis, many genes have shown conserved functions in biofuel feedstock species. Here, we review the recent advances in our understanding of vascular development and SCW formation and discuss potential biotechnological uses. PMID:27047525

  4. Formation of photovoltaic modules based on polycrystalline solar cells

    Directory of Open Access Journals (Sweden)

    L. A. Dobrzański

    2009-12-01

    Full Text Available Purpose: The main aim of the paper is formation of photovoltaic modules and analysis of their main electric parameters.Design/methodology/approach: Photovoltaic modules were produced from four polycrystalline silicon solar cells, that were cut and next joined in series. Soft soldering technique and copper-tin strip were used for joining cells.Findings: In order to provide useful power for any application, the individual solar cells must be connected together to give the appropriate current and voltage levels. Taking this fact into account the analysis of photovoltaic module construction was performed.Research limitations/implications: The main goal of the research is to show the practical application of solar cells. Two photovoltaic modules were assembled and their basic electric properties were analysed. It was shown that they may be successively applied as an alternative energy source.Practical implications: Photovoltaic modules are irreplaceable in areas which are far away from power network. Simply photovoltaic module can supply small device without any problem.Originality/value: The produced photovoltaic modules and photovoltaic systems confirm the utility of solar energy in every place where the sun radiation is available. Because of exhaust conventional energy sources like coal or earth gas, new renewable sources of energy (sunlight, wind are more and more often used. It brings huge ecological benefits.

  5. Segrosome Complex Formation during DNA Trafficking in Bacterial Cell Division.

    Science.gov (United States)

    Oliva, María A

    2016-01-01

    Bacterial extrachromosomal DNAs often contribute to virulence in pathogenic organisms or facilitate adaptation to particular environments. The transmission of genetic information from one generation to the next requires sufficient partitioning of DNA molecules to ensure that at least one copy reaches each side of the division plane and is inherited by the daughter cells. Segregation of the bacterial chromosome occurs during or after replication and probably involves a strategy in which several protein complexes participate to modify the folding pattern and distribution first of the origin domain and then of the rest of the chromosome. Low-copy number plasmids rely on specialized partitioning systems, which in some cases use a mechanism that show striking similarity to eukaryotic DNA segregation. Overall, there have been multiple systems implicated in the dynamic transport of DNA cargo to a new cellular position during the cell cycle but most seem to share a common initial DNA partitioning step, involving the formation of a nucleoprotein complex called the segrosome. The particular features and complex topologies of individual segrosomes depend on both the nature of the DNA binding protein involved and on the recognized centromeric DNA sequence, both of which vary across systems. The combination of in vivo and in vitro approaches, with structural biology has significantly furthered our understanding of the mechanisms underlying DNA trafficking in bacteria. Here, I discuss recent advances and the molecular details of the DNA segregation machinery, focusing on the formation of the segrosome complex. PMID:27668216

  6. Bioluminescence Reporter Gene Imaging Characterize Human Embryonic Stem Cell-Derived Teratoma Formation

    OpenAIRE

    Su, Weijun; Zhou, Manqian; Zheng, Yizhou; Fan, Yan; HAN, ZHONGCHAO; Kong, Deling; Wu, Joseph C.; Xiang, Rong; Li, Zongjin

    2011-01-01

    Human embryonic stem (hES) cells are capable of differentiation into virtually all cell types and hold tremendous potential as cell sources for regenerative therapies. However, teratoma formation can be the main obstacle for hES cells therapy. In order to understand the biology and physiology of hES cells teratoma formation, we investigated the angiogenic process within teratomas and characterized teratoma cells. In this study, hES cells transduced with double fusion reporter gene that consis...

  7. Mipu1 Overexpression Protects Macrophages from oxLDL-Induced Foam Cell Formation and Cell Apoptosis

    OpenAIRE

    Qu, Shun-Lin; Fan, Wen-Jing; Zhang, Chi; Guo, Fang; Han, Dan; Pan, Wen-Jun; Li, Wei; Feng, Da-Ming; JIANG, ZHI-SHENG

    2014-01-01

    Mipu1 (myocardial ischemic preconditioning upregulated protein 1) is a novel N-terminal Kruppel-associated box (KRAB)/C2H2 zinc finger superfamily protein, that displays a powerful effect in protecting H9c2 cells from oxidative stress-induced cell apoptosis. The present study aims to investigate the effect of Mipu1 overexpression on oxidized low-density lipoprotein (oxLDL)-induced foam cell formation, cell apoptosis, and its possible mechanisms. New Zealand healthy rabbits were used to establ...

  8. The contribution of cell-cell signaling and motility to bacterial biofilm formation

    DEFF Research Database (Denmark)

    Shrout, Joshua D; Tolker-Nielsen, Tim; Givskov, Michael;

    2011-01-01

    Many bacteria grow attached to a surface as biofilms. Several factors dictate biofilm formation, including responses by the colonizing bacteria to their environment. Here we review how bacteria use cell-cell signaling (also called quorum sensing) and motility during biofilm formation. Specifically...... gene expression important to the production of polysaccharides, rhamnolipid, and other virulence factors. Surface motility affects the assembly and architecture of biofilms, and some aspects of motility are also influenced by quorum sensing. While some genes and their function are specific to P....... aeruginosa, many aspects of biofilm development can be used as a model system to understand how bacteria differentially colonize surfaces....

  9. Macropinocytosis contributes to the macrophage foam cell formation in RAW264.7 cells

    Institute of Scientific and Technical Information of China (English)

    Wenqi Yao; Ke Li; Kan Liao

    2009-01-01

    The key event in the atherosclerosis development is the lipids uptake by macrophage and the formation of foam cell in subendothelial arterial space. Besides the uptake of modified low-density lipoprotein (LDL) by scavenger receptor-mediated endocytosis, macrophages possess constitutive macropinocytosis, which is capable of taking up a large quantity of solute. Macrophage foam cell formation could be induced in RAW264.7 cells by increasing the serum concentration in the culture medium. Foam cell formation induced by serum could be blocked by phosphoinositide 3-kinase inhibi-tor, LY294002 or wortmannin, which inhibited macro-pinocytosis but not receptor-mediated endocytosis. Further analysis indicated that macropinocytosis took place at the gangliosides-enriched membrane area. Cholesterol depletion by β-methylcyclodextrin-blocked macropinocytosis without affecting scavenger receptor-mediated endocytosis of modified LDLs. These results suggested that macropinocytosis might be one of the important mechanisms for lipid uptake in macrophage. And it made significant contribution to the lipid accumulation and foam cell formation.

  10. Stem Cell-Soluble Signals Enhance Multilumen Formation in SMG Cell Clusters.

    Science.gov (United States)

    Maruyama, C L M; Leigh, N J; Nelson, J W; McCall, A D; Mellas, R E; Lei, P; Andreadis, S T; Baker, O J

    2015-11-01

    Saliva plays a major role in maintaining oral health. Patients with salivary hypofunction exhibit difficulty in chewing and swallowing foods, tooth decay, periodontal disease, and microbial infections. At this time, treatments for hyposalivation are limited to medications (e.g., muscarinic receptor agonists: pilocarpine and cevimeline) that induce saliva secretion from residual acinar cells as well as artificial salivary substitutes. Therefore, advancement of restorative treatments is necessary to improve the quality of life in these patients. Our previous studies indicated that salivary cells are able to form polarized 3-dimensional structures when grown on growth factor-reduced Matrigel. This basement membrane is rich in laminin-III (L1), which plays a critical role in salivary gland formation. Mitotically inactive feeder layers have been used previously to support the growth of many different cell types, as they provide factors necessary for cell growth and organization. The goal of this study was to improve salivary gland cell differentiation in primary cultures by using a combination of L1 and a feeder layer of human hair follicle-derived mesenchymal stem cells (hHF-MSCs). Our results indicated that the direct contact of mouse submandibular (mSMG) cell clusters and hHF-MSCs was not required for mSMG cells to form acinar and ductal structures. However, the hHF-MSC conditioned medium enhanced cell organization and multilumen formation, indicating that soluble signals secreted by hHF-MSCs play a role in promoting these features. PMID:26285810

  11. ZAP-70 kinase regulates HIV cell-to-cell spread and virological synapse formation

    OpenAIRE

    Sol-Foulon, Nathalie; Sourisseau, Marion; Porrot, Françoise; Thoulouze, Maria-Isabel; Trouillet, Céline; Nobile, Cinzia; Blanchet, Fabien; Di Bartolo, Vincenzo; Noraz, Nelly; Taylor, Naomi; Alcover, Andres; Hivroz, Claire; Schwartz, Olivier

    2007-01-01

    HIV efficiently spreads in lymphocytes, likely through virological synapses (VSs). These cell–cell junctions share some characteristics with immunological synapses, but cellular proteins required for their constitution remain poorly characterized. We have examined here the role of ZAP-70, a key kinase regulating T-cell activation and immunological synapse formation, in HIV replication. In lymphocytes deficient for ZAP-70, or expressing a kinase-dead mutant of the protein, HIV replication was ...

  12. DICER Regulates the Formation and Maintenance of Cell-Cell Junctions in the Mouse Seminiferous Epithelium.

    Science.gov (United States)

    Korhonen, Hanna Maria; Yadav, Ram Prakash; Da Ros, Matteo; Chalmel, Frédéric; Zimmermann, Céline; Toppari, Jorma; Nef, Serge; Kotaja, Noora

    2015-12-01

    The endonuclease DICER that processes micro-RNAs and small interfering RNAs is essential for normal spermatogenesis and male fertility. We previously showed that the deletion of Dicer1 gene in postnatal spermatogonia in mice using Ngn3 promoter-driven Cre expression caused severe defects in the morphogenesis of haploid spermatid to mature spermatozoon, including problems in cell polarization and nuclear elongation. In this study, we further analyzed the same mouse model and revealed that absence of functional DICER in differentiating male germ cells induces disorganization of the cell-cell junctions in the seminiferous epithelium. We detected discontinuous and irregular apical ectoplasmic specializations between elongating spermatids and Sertoli cells. The defective anchoring of spermatids to Sertoli cells caused a premature release of spermatids into the lumen. Our findings may help also explain the abnormal elongation process of remaining spermatids because these junctions and the correct positioning of germ cells in the epithelium are critically important for the progression of spermiogenesis. Interestingly, cell adhesion-related genes were generally upregulated in Dicer1 knockout germ cells. Claudin 5 ( Cldn5 ) was among the most upregulated genes and we show that the polarized localization of CLAUDIN5 in the apical ectoplasmic specializations was lost in Dicer1 knockout spermatids. Our results suggest that DICER-dependent pathways control the formation and organization of cell-cell junctions in the seminiferous epithelium via the regulation of cell adhesion-related genes. PMID:26510868

  13. RNA-binding IMPs promote cell adhesion and invadopodia formation

    DEFF Research Database (Denmark)

    Vikesaa, Jonas; Hansen, Thomas V O; Jønson, Lars;

    2006-01-01

    Oncofetal RNA-binding IMPs have been implicated in mRNA localization, nuclear export, turnover and translational control. To depict the cellular actions of IMPs, we performed a loss-of-function analysis, which showed that IMPs are necessary for proper cell adhesion, cytoplasmic spreading...... and invadopodia formation. Loss of IMPs was associated with a coordinate downregulation of mRNAs encoding extracellular matrix and adhesion proteins. The transcripts were present in IMP RNP granules, implying that IMPs were directly involved in the post-transcriptional control of the transcripts. In particular......, we show that a 5.0 kb CD44 mRNA contained multiple IMP-binding sites in its 3'UTR, and following IMP depletion this species became unstable. Direct knockdown of the CD44 transcript mimicked the effect of IMPs on invadopodia, and we infer that CD44 mRNA stabilization may be involved in IMP...

  14. Black Silicon formation using dry etching for solar cells applications

    Energy Technology Data Exchange (ETDEWEB)

    Murias, D. [Instituto Nacional de Astrofisica, Optica y Electronica, INAOE, Puebla (Mexico); Reyes-Betanzo, C., E-mail: creyes@inaoep.mx [Instituto Nacional de Astrofisica, Optica y Electronica, INAOE, Puebla (Mexico); Moreno, M.; Torres, A.; Itzmoyotl, A. [Instituto Nacional de Astrofisica, Optica y Electronica, INAOE, Puebla (Mexico); Ambrosio, R.; Soriano, M. [Universidad Autonoma de Ciudad Juarez, Chihuahua (Mexico); Lucas, J. [Instituto Tecnologico de Tehuacan, Puebla (Mexico); Cabarrocas, P. Roca i [Laboratoire de Physique des Interfaces et des Couches Minces, Ecole Polytechnique, CNRS, Palaiseau (France)

    2012-09-20

    A study on the formation of Black Silicon on crystalline silicon surface using SF{sub 6}/O{sub 2} and SF{sub 6}/O{sub 2}/CH{sub 4} based plasmas in a reactive ion etching (RIE) system is presented. The effect of the RF power, chamber pressure, process time, gas flow rates, and gas mixtures on the texture of silicon surface has been analyzed. Completely Black Silicon surfaces containing pyramid like structures have been obtained, using an optimized mask-free plasma process. Moreover, the Black Silicon surfaces have demonstrated average values of 1% and 4% for specular and diffuse reflectance respectively, feature that is suitable for the fabrication of low cost solar cells.

  15. Myosin-II dependent cell contractility contributes to spontaneous nodule formation of mesothelioma cells

    CERN Document Server

    Tárnoki-Zách, Julia; Méhes, Elod; Paku, Sándor; Neufeld, Zoltán; Hegedus, Balázs; Döme, Balázs; Czirok, Andras

    2015-01-01

    We demonstrate that characteristic nodules emerge in cultures of several malignant pleural mesothelioma (MPM) cell lines. Instead of excessive local cell proliferation, the nodules arise by Myosin II-driven cell contractility. The aggregation process can be prevented or reversed by suitable pharmacological inhibitors of acto-myosin contractility. A cell-resolved elasto-plastic model of the multicellular patterning process indicates that the morphology and size of the nodules as well as the speed of their formation is determined by the mechanical tension cells exert on their neighbors, and the stability of cell-substrate adhesion complexes. A linear stability analysis of a homogenous, self-tensioned Maxwell fluid indicates the unconditional presence of a patterning instability.

  16. Human germ cell formation in xenotransplants of induced pluripotent stem cells carrying X chromosome aneuploidies.

    Science.gov (United States)

    Dominguez, Antonia A; Chiang, H Rosaria; Sukhwani, Meena; Orwig, Kyle E; Reijo Pera, Renee A

    2014-09-22

    Turner syndrome is caused by complete or partial loss of the second sex chromosome and is characterized by spontaneous fetal loss in >90% of conceptions. Survivors possess an array of somatic and germline clinical characteristics. Induced pluripotent stem cells (iPSCs) offer an opportunity for insight into genetic requirements of the X chromosome linked to Turner syndrome. We derived iPSCs from Turner syndrome and control individuals and examined germ cell development as a function of X chromosome composition. We demonstrate that two X chromosomes are not necessary for reprogramming or maintenance of pluripotency and that there are minimal differences in gene expression, at the single cell level, linked to X chromosome aneuploidies. Formation of germ cells, as assessed in vivo through a murine xenotransplantation model, indicated that undifferentiated iPSCs, independent of X chromosome composition, are capable of forming germ-cell-like cells (GCLCs) in vivo. In combination with clinical data regarding infertility in women with X chromosome aneuploidies, results suggest that two intact X chromosomes are not required for human germ cell formation, qualitatively or quantitatively, but rather are likely to be required for maintenance of human germ cells to adulthood.

  17. An improved alkaline direct formate paper microfluidic fuel cell.

    Science.gov (United States)

    Galvan, Vicente; Domalaon, Kryls; Tang, Catherine; Sotez, Samantha; Mendez, Alex; Jalali-Heravi, Mehdi; Purohit, Krutarth; Pham, Linda; Haan, John; Gomez, Frank A

    2016-02-01

    Paper-based microfluidic fuel cells (MFCs) are a potential replacement for traditional FCs and batteries due to their low cost, portability, and simplicity to operate. In MFCs, separate solutions of fuel and oxidant migrate through paper due to capillary action and laminar flow and, upon contact with each other and catalyst, produce electricity. In the present work, we describe an improved microfluidic paper-based direct formate FC (DFFC) employing formate and hydrogen peroxide as the anode fuel and cathode oxidant, respectively. The dimensions of the lateral column, current collectors, and cathode were optimized. A maximum power density of 2.53 mW/cm(2) was achieved with a DFFC of surface area 3.0 cm(2) , steel mesh as current collector, 5% carbon to paint mass ratio for cathode electrode and, 30% hydrogen peroxide. The longevity of the MFC's detailed herein is greater than eight hours with continuous flow of streams. In a series configuration, the MFCs generate sufficient energy to power light-emitting diodes and a handheld calculator.

  18. An improved alkaline direct formate paper microfluidic fuel cell.

    Science.gov (United States)

    Galvan, Vicente; Domalaon, Kryls; Tang, Catherine; Sotez, Samantha; Mendez, Alex; Jalali-Heravi, Mehdi; Purohit, Krutarth; Pham, Linda; Haan, John; Gomez, Frank A

    2016-02-01

    Paper-based microfluidic fuel cells (MFCs) are a potential replacement for traditional FCs and batteries due to their low cost, portability, and simplicity to operate. In MFCs, separate solutions of fuel and oxidant migrate through paper due to capillary action and laminar flow and, upon contact with each other and catalyst, produce electricity. In the present work, we describe an improved microfluidic paper-based direct formate FC (DFFC) employing formate and hydrogen peroxide as the anode fuel and cathode oxidant, respectively. The dimensions of the lateral column, current collectors, and cathode were optimized. A maximum power density of 2.53 mW/cm(2) was achieved with a DFFC of surface area 3.0 cm(2) , steel mesh as current collector, 5% carbon to paint mass ratio for cathode electrode and, 30% hydrogen peroxide. The longevity of the MFC's detailed herein is greater than eight hours with continuous flow of streams. In a series configuration, the MFCs generate sufficient energy to power light-emitting diodes and a handheld calculator. PMID:26572774

  19. Micronucleus formation induced by dielectric barrier discharge plasma exposure in brain cancer cells

    Science.gov (United States)

    Kaushik, Nagendra K.; Uhm, Hansup; Ha Choi, Eun

    2012-02-01

    Induction of micronucleus formation (cytogenetic damage) in brain cancer cells upon exposure of dielectric barrier discharge plasma has been investigated. We have investigated the influence of exposure and incubation times on T98G brain cancer cells by using growth kinetic, clonogenic, and micronucleus formation assay. We found that micronucleus formation rate directly depends on the plasma exposure time. It is also shown that colony formation capacity of cells has been inhibited by the treatment of plasma at all doses. Cell death and micronucleus formation are shown to be significantly elevated by 120 and 240 s exposure of dielectric barrier discharge plasma.

  20. In vitro formation of β cell pseudoislets using islet-derived endothelial cells.

    Directory of Open Access Journals (Sweden)

    Michael G Spelios

    Full Text Available β cell pseudoislets (PIs are used for the in vitro study of β-cells in a three-dimensional (3-D configuration. Current methods of PI induction require unique culture conditions and extensive mechanical manipulations. Here we report a novel co-culture system consisting of high passage β-cells and islet-derived endothelial cells (iECs that results in a rapid and spontaneous formation of free-floating PIs. PI structures were formed as early as 72 h following co-culture setup and were preserved for more than 14 d. These PIs, composed solely of β-cells, were similar in size to that of native islets and showed an increased percentage of proinsulin-positive cells, increased insulin gene expression in response to glucose stimulation, and restored glucose-stimulated insulin secretion when compared to β-cells cultured as monolayers. Key extracellular matrix proteins that were absent in β-cells cultured alone were deposited by iECs on PIs and were found in and around the PIs. iEC-induced PIs are a readily available tool for examining β cell function in a native 3-D configuration and can be used for examining β-cell/iEC interactions in vitro.

  1. Mind bomb 1 is required for pancreatic ß-cell formation

    DEFF Research Database (Denmark)

    Horn, Signe; Kobberup, Sune; Jørgensen, Mette C;

    2012-01-01

    the insulin producing ß-cells. However, signals that regulate proximodistal (P-D) patterning and thus formation of ß-cell progenitors are unknown. Here we show that Mind bomb 1 (Mib1) is required for correct P-D patterning of the developing pancreas and ß-cell formation. We found that endoderm...

  2. Analysis of torsional barrier height of HSNO as the simplest S-nitrosothiol

    Indian Academy of Sciences (India)

    Reza Fazaeli; Mohammad Solimannejad; Abdolvahab Seif

    2013-07-01

    Torsional barrier height of thionitrous acid is analysed with Gaussian-2(G2), quadratic complete basis set (CBS-Q) and DFT-B3LYP/CBS-Q (CBS-QB3) methods. In agreement with purely intuitive arguments, it was determined that the cis to trans barrier height is nearly 5.7-6.3 kcal mol-1. In addition, the stability of rotation as function of competing dissociation pathways and also result of natural bond orbital analysis are discussed.

  3. Lateral inhibition-induced pattern formation controlled by the size and geometry of the cell.

    Science.gov (United States)

    Seirin Lee, Sungrim

    2016-09-01

    Pattern formation in development biology is one of the fundamental processes by which cells change their functions. It is based on the communication of cells via intra- and intercellular dynamics of biochemicals. Thus, the cell is directly involved in biochemical interactions. However, many theoretical approaches describing biochemical pattern formation have usually neglected the cell's role or have simplified the subcellular process without considering cellular aspects despite the cell being the environment where biochemicals interact. On the other hand, recent experimental observations suggest that a change in the physical conditions of cell-to-cell contact can result in a change in cell fate and tissue patterning in a lateral inhibition system. Here we develop a mathematical model by which biochemical dynamics can be directly observed with explicitly expressed cell structure and geometry in higher dimensions, and reconsider pattern formation by lateral inhibition of the Notch-Delta signaling pathway. We explore how the physical characteristic of cell, such as cell geometry or size, influences the biochemical pattern formation in a multi-cellular system. Our results suggest that a property based on cell geometry can be a novel mechanism for symmetry breaking inducing cell asymmetry. We show that cell volume can critically influence cell fate determination and pattern formation at the tissue level, and the surface area of the cell-to-cell contact can directly affect the spatial range of patterning. PMID:27229622

  4. Lateral inhibition-induced pattern formation controlled by the size and geometry of the cell.

    Science.gov (United States)

    Seirin Lee, Sungrim

    2016-09-01

    Pattern formation in development biology is one of the fundamental processes by which cells change their functions. It is based on the communication of cells via intra- and intercellular dynamics of biochemicals. Thus, the cell is directly involved in biochemical interactions. However, many theoretical approaches describing biochemical pattern formation have usually neglected the cell's role or have simplified the subcellular process without considering cellular aspects despite the cell being the environment where biochemicals interact. On the other hand, recent experimental observations suggest that a change in the physical conditions of cell-to-cell contact can result in a change in cell fate and tissue patterning in a lateral inhibition system. Here we develop a mathematical model by which biochemical dynamics can be directly observed with explicitly expressed cell structure and geometry in higher dimensions, and reconsider pattern formation by lateral inhibition of the Notch-Delta signaling pathway. We explore how the physical characteristic of cell, such as cell geometry or size, influences the biochemical pattern formation in a multi-cellular system. Our results suggest that a property based on cell geometry can be a novel mechanism for symmetry breaking inducing cell asymmetry. We show that cell volume can critically influence cell fate determination and pattern formation at the tissue level, and the surface area of the cell-to-cell contact can directly affect the spatial range of patterning.

  5. Tudor and its domains: germ cell formation from a Tudor perspective

    Institute of Scientific and Technical Information of China (English)

    Travis THOMSON; Paul LASKO

    2005-01-01

    In many metazoan species, germ cell formation requires the germ plasm, a specialized cytoplasm which often contains electron dense structures. Genes required for germ cell formation in Drosophila have been isolated predominantly in screens for maternal-effect mutations. One such gene is tudor (tud); without proper tud function germ cell formation does not occur. Unlike other genes involved in Drosophila germ cell specification tud is dispensable for other somatic functions such as abdominal patterning. It is not known how TUD contributes at a molecular level to germ cell formation but in tud mutants, polar granule formation is severely compromised, and mitochondrially encoded ribosomal RNAs do not localize to the polar granule. TUD is composed of 11 repeats of the protein motif called the Tudor domain. There are similar proteins to TUD in the germ line of other metazoan species including mice. Probable vertebrate orthologues of Drosophila genes involved in germ cell specification will be discussed.

  6. Myotube formation is affected by adipogenic lineage cells in a cell-to-cell contact-independent manner

    Energy Technology Data Exchange (ETDEWEB)

    Takegahara, Yuki; Yamanouchi, Keitaro, E-mail: akeita@mail.ecc.u-tokyo.ac.jp; Nakamura, Katsuyuki; Nakano, Shin-ichi; Nishihara, Masugi

    2014-05-15

    Intramuscular adipose tissue (IMAT) formation is observed in some pathological conditions such as Duchenne muscular dystrophy (DMD) and sarcopenia. Several studies have suggested that IMAT formation is not only negatively correlated with skeletal muscle mass but also causes decreased muscle contraction in sarcopenia. In the present study, we examined w hether adipocytes affect myogenesis. For this purpose, skeletal muscle progenitor cells were transfected with siRNA of PPARγ (siPPARγ) in an attempt to inhibit adipogenesis. Myosin heavy chain (MHC)-positive myotube formation was promoted in cells transfected with siPPARγ compared to that of cells transfected with control siRNA. To determine whether direct cell-to-cell contact between adipocytes and myoblasts is a prerequisite for adipocytes to affect myogenesis, skeletal muscle progenitor cells were cocultured with pre- or mature adipocytes in a Transwell coculture system. MHC-positive myotube formation was inhibited when skeletal muscle progenitor cells were cocultured with mature adipocytes, but was promoted when they were cocultured with preadipocytes. Similar effects were observed when pre- or mature adipocyte-conditioned medium was used. These results indicate that preadipocytes play an important role in maintaining skeletal muscle mass by promoting myogenesis; once differentiated, the resulting mature adipocytes negatively affect myogenesis, leading to the muscle deterioration observed in skeletal muscle pathologies. - Highlights: • We examined the effects of pre- and mature adipocytes on myogenesis in vitro. • Preadipocytes and mature adipocytes affect myoblast fusion. • Preadipocytes play an important role in maintaining skeletal muscle mass. • Mature adipocytes lead to muscle deterioration observed in skeletal muscle pathologies.

  7. Systemic mesenchymal stem cell administration enhances bone formation in fracture repair but not load-induced bone formation

    Directory of Open Access Journals (Sweden)

    AE Rapp

    2015-01-01

    Full Text Available Mesenchymal stem cells (MSC were shown to support bone regeneration, when they were locally transplanted into poorly healing fractures. The benefit of systemic MSC transplantation is currently less evident. There is consensus that systemically applied MSC are recruited to the site of injury, but it is debated whether they actually support bone formation. Furthermore, the question arises as to whether circulating MSC are recruited only in case of injury or whether they also participate in mechanically induced bone formation. To answer these questions we injected green fluorescent protein (GFP-labelled MSC into C57BL/6J mice, which were subjected either to a femur osteotomy or to non-invasive mechanical ulna loading to induce bone formation. We detected GFP-labelled MSC in the early (day 10 and late fracture callus (day 21 by immunohistochemistry. Stromal cell-derived factor 1 (SDF-1 or CXCL-12, a key chemokine for stem cell attraction, was strongly expressed by virtually all cells near the osteotomy – indicating that SDF-1 may mediate cell migration to the site of injury. We found no differences in SDF-1 expression between the groups. Micro-computed tomography (µCT revealed significantly more bone in the callus of the MSC treated mice compared to untreated controls. The bending stiffness of callus was not significantly altered after MSC-application. In contrast, we failed to detect GFP-labelled MSC in the ulna after non-invasive mechanical loading. Histomorphometry and µCT revealed a significant load-induced increase in bone formation; however, no further increase was found after MSC administration. Concluding, our results suggest that systemically administered MSC are recruited and support bone formation only in case of injury but not in mechanically induced bone formation.

  8. Cutting edge: Bcl6-interacting corepressor contributes to germinal center T follicular helper cell formation and B cell helper function.

    Science.gov (United States)

    Yang, Jessica A; Tubo, Noah J; Gearhart, Micah D; Bardwell, Vivian J; Jenkins, Marc K

    2015-06-15

    CD4(+) germinal center (GC)-T follicular helper (Tfh) cells help B cells become long-lived plasma cells and memory cells. The transcriptional repressor Bcl6 plays a key role in GC-Tfh formation by inhibiting the expression of genes that promote differentiation into other lineages. We determined whether BCOR, a component of a Polycomb repressive complex that interacts with the Bcl6 BTB domain, influences GC-Tfh differentiation. T cell-targeted BCOR deficiency led to a substantial loss of peptide:MHC class II-specific GC-Tfh cells following Listeria monocytogenes infection and a 2-fold decrease following immunization with a peptide in CFA. The reduction in GC-Tfh cells was associated with diminished plasma cell and GC B cell formation. Thus, T cell-expressed BCOR is critical for optimal GC-Tfh cell differentiation and humoral immunity. PMID:25964495

  9. Critical role of mast cell chymase in mouse abdominal aortic aneurysm formation

    DEFF Research Database (Denmark)

    Sun, J; Zhang, J; Lindholt, Jes S.;

    2009-01-01

    Mast cell chymase may participate in the pathogenesis of human abdominal aortic aneurysm (AAA), yet a direct contribution of this serine protease to AAA formation remains unknown.......Mast cell chymase may participate in the pathogenesis of human abdominal aortic aneurysm (AAA), yet a direct contribution of this serine protease to AAA formation remains unknown....

  10. Mipu1 overexpression protects macrophages from oxLDL-induced foam cell formation and cell apoptosis.

    Science.gov (United States)

    Qu, Shun-Lin; Fan, Wen-Jing; Zhang, Chi; Guo, Fang; Han, Dan; Pan, Wen-Jun; Li, Wei; Feng, Da-Ming; Jiang, Zhi-Sheng

    2014-12-01

    Mipu1 (myocardial ischemic preconditioning upregulated protein 1) is a novel N-terminal Kruppel-associated box (KRAB)/C2H2 zinc finger superfamily protein, that displays a powerful effect in protecting H9c2 cells from oxidative stress-induced cell apoptosis. The present study aims to investigate the effect of Mipu1 overexpression on oxidized low-density lipoprotein (oxLDL)-induced foam cell formation, cell apoptosis, and its possible mechanisms. New Zealand healthy rabbits were used to establish atherosclerosis model, and serum levels of triglycerides, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol were detected by an automatic biochemical analyzer. Sudan IV staining was used to detect atherosclerotic lesions. The RAW264.7 macrophage cell line was selected as the experimental material. Oil red O staining, high-performance liquid chromatography, and Dil-labeled lipoprotein were used to detect cholesterol accumulation qualitatively and quantitatively, respectively. Flow cytometry was used to determine cell apoptosis. Real-time quantitative polymerase chain reaction (PCR) was used to detect the mRNA expression of the main proteins that are associated with the transport of cholesterol, such as ABCA1, ABCG1, SR-BI, and CD36. Western blot analysis was used to detect the protein expression of Mipu1. There were atherosclerotic lesions in the high-fat diet group with Sudan IV staining. High-fat diet decreased Mipu1 expression and increased CD36 expression significantly at the 10th week compared with standard-diet rabbits. Mipu1 overexpression decreased oxLDL-induced cholesterol accumulation, oxLDL uptake, cell apoptosis, and cleaved caspase-3. Mipu1 overexpression inhibited the oxLDL-induced CD36 mRNA and protein expression, but it did not significantly inhibit the mRNA expression of ABCA1, ABCG1, and SR-BI. Mipu1 overexpression inhibits oxLDL-induced foam cell formation and cell apoptosis. Mipu1 overexpression reduces the

  11. 3D in vitro cell culture models of tube formation

    NARCIS (Netherlands)

    Zegers, M.M.P.

    2014-01-01

    Building the complex architecture of tubular organs is a highly dynamic process that involves cell migration, polarization, shape changes, adhesion to neighboring cells and the extracellular matrix, physicochemical characteristics of the extracellular matrix and reciprocal signaling with the mesench

  12. An Approach to Determining the Optimal Cell Number of Manufacturing Cell Formation

    Directory of Open Access Journals (Sweden)

    Jianwei Wang

    2013-04-01

    Full Text Available An approach to determining the optimal cell number of manufacturing cell formation is presented. Firstly, the difference of weighting exponent, cluster center and metrics how to have an impact upon the clustering results and membership function are studied. Secondly, a method to determine the optimal m value is given. Two-order partial derivative of the objective function for FCM is calculated, and the variational weighting exponent m is obtained that can prevent the parameter from being the unique value and play an important role in the process of fuzzy clustering. Moreover, in order to avoid a single validity index can not assess correctly, partition coefficient (PC, classification entropy (CE, Fukuyama and Sugeno (FS and Xie and Beni (XB are considered as multi-performance indexes to evaluate the cluster validity, and then an optimal number c is chosen based on these validity measures. Finally, test exampls are given to illustrate the validity of the proposed approach.

  13. Single cell motility and trail formation in populations of microglia

    Science.gov (United States)

    Lee, Kyoung Jin

    2009-03-01

    Microglia are a special type of glia cell in brain that has immune responses. They constitute about 20 % of the total glia population within the brain. Compared to other glia cells, microglia are very motile, constantly moving to destroy pathogens and to remove dead neurons. While doing so, they exhibit interesting body shapes, have cell-to-cell communications, and have chemotatic responses to each other. Interestingly, our recent in vitro studies show that their unusual motile behaviors can self-organize to form trails, similar to those in populations of ants. We have studied the changes in the physical properties of these trails by varying the cell population density and by changing the degree of spatial inhomogeneities (``pathogens''). Our experimental observations can be quite faithfully reproduced by a simple mathematical model involving many motile cells whose mechanical motion are driven by actin polymerization and depolymerization process within the individual cell body and by external chemical gradients.

  14. Effects of irradiated biodegradable polymer in endothelial cell monolayer formation

    Energy Technology Data Exchange (ETDEWEB)

    Arbeitman, Claudia R.; Grosso, Mariela F. del [CONICET – Consejo Nacional de Investigaciones Científicas y Técnicas (Argentina); Gerencia de Investigación y Aplicaciones, TANDAR-CNEA (Argentina); Behar, Moni [Instituto de Física, UFRGS, Porto Alegre, RS (Brazil); García Bermúdez, Gerardo, E-mail: ggb@tandar.cnea.gov.ar [CONICET – Consejo Nacional de Investigaciones Científicas y Técnicas (Argentina); Gerencia de Investigación y Aplicaciones, TANDAR-CNEA (Argentina); Escuela de Ciencia y Tecnología, UNSAM (Argentina)

    2013-11-01

    In this work we study cell adhesion, proliferation and cell morphology of endothelial cell cultured on poly-L-lactide acid (PLLA) modified by heavy ion irradiation. Thin films of PLLA samples were irradiated with sulfur (S) at energies of 75 MeV and gold (Au) at 18 MeV ion-beams. Ion beams were provided by the Tandar (Buenos Aires, Argentina) and Tandetron (Porto Alegre, Brazil) accelerators, respectively. The growth of a monolayer of bovine aortic endothelial cells (BAEC) onto unirradiated and irradiated surfaces has been studied by in vitro techniques in static culture. Cell viability and proliferation increased on modified substrates. But the results on unirradiated samples, indicate cell death (necrosis/apoptosis) with the consequent decrease in proliferation. We analyzed the correlation between irradiation parameters and cell metabolism and morphology.

  15. Formate: an Energy Storage and Transport Bridge between Carbon Dioxide and a Formate Fuel Cell in a Single Device.

    Science.gov (United States)

    Vo, Tracy; Purohit, Krutarth; Nguyen, Christopher; Biggs, Brenna; Mayoral, Salvador; Haan, John L

    2015-11-01

    We demonstrate the first device to our knowledge that uses a solar panel to power the electrochemical reduction of dissolved carbon dioxide (carbonate) into formate that is then used in the same device to operate a direct formate fuel cell (DFFC). The electrochemical reduction of carbonate is carried out on a Sn electrode in a reservoir that maintains a constant carbon balance between carbonate and formate. The electron-rich formate species is converted by the DFFC into electrical energy through electron release. The product of DFFC operation is the electron-deficient carbonate species that diffuses back to the reservoir bulk. It is possible to continuously charge the device using alternative energy (e.g., solar) to convert carbonate to formate for on-demand use in the DFFC; the intermittent nature of alternative energy makes this an attractive design. In this work, we demonstrate a proof-of-concept device that performs reduction of carbonate, storage of formate, and operation of a DFFC.

  16. Calcium Signalling Triggered by NAADP in T Cells Determines Cell Shape and Motility During Immune Synapse Formation

    Science.gov (United States)

    Nebel, Merle; Zhang, Bo; Odoardi, Francesca; Flügel, Alexander; Potter, Barry V. L.; Guse, Andreas H.

    2016-01-01

    Nicotinic acid adenine dinucleotide phosphate (NAADP) has been implicated as an initial Ca2+ trigger in T cell Ca2+ signalling, but its role in formation of the immune synapse in CD4+ effector T cells has not been analysed. CD4+ T cells are activated by the interaction with peptide-MHCII complexes on the surface of antigen-presenting cells. Establishing a two-cell system including primary rat CD4+ T cells specific for myelin basic protein and rat astrocytes enabled us to mirror this activation process in vitro and to analyse Ca2+ signalling, cell shape changes and motility in T cells during formation and maintenance of the immune synapse. After immune synapse formation, T cells showed strong, antigen-dependent increases in free cytosolic calcium concentration ([Ca2+]i). Analysis of cell shape and motility revealed rounding and immobilization of T cells depending on the amplitude of the Ca2+ signal. NAADP-antagonist BZ194 effectively blocked Ca2+ signals in T cells evoked by the interaction with antigen-presenting astrocytes. BZ194 reduced the percentage of T cells showing high Ca2+ signals thereby supporting the proposed trigger function of NAADP for global Ca2+ signalling. Taken together, the NAADP signalling pathway is further confirmed as a promising target for specific pharmacological intervention to modulate T cell activation. PMID:27747143

  17. Stem cells catalyze cartilage formation by neonatal articular chondrocytes in 3D biomimetic hydrogels

    Science.gov (United States)

    Lai, Janice H.; Kajiyama, Glen; Smith, Robert Lane; Maloney, William; Yang, Fan

    2013-12-01

    Cartilage loss is a leading cause of disability among adults and effective therapy remains elusive. Neonatal chondrocytes (NChons) are an attractive allogeneic cell source for cartilage repair, but their clinical translation has been hindered by scarce donor availability. Here we examine the potential for catalyzing cartilage tissue formation using a minimal number of NChons by co-culturing them with adipose-derived stem cells (ADSCs) in 3D hydrogels. Using three different co-culture models, we demonstrated that the effects of co-culture on cartilage tissue formation are dependent on the intercellular distance and cell distribution in 3D. Unexpectedly, increasing ADSC ratio in mixed co-culture led to increased synergy between NChons and ADSCs, and resulted in the formation of large neocartilage nodules. This work raises the potential of utilizing stem cells to catalyze tissue formation by neonatal chondrocytes via paracrine signaling, and highlights the importance of controlling cell distribution in 3D matrices to achieve optimal synergy.

  18. Nicotinate-Curcumin Impedes Foam Cell Formation from THP-1 Cells through Restoring Autophagy Flux.

    Science.gov (United States)

    Gu, Hong-Feng; Li, Hai-Zhe; Tang, Ya-Ling; Tang, Xiao-Qing; Zheng, Xi-Long; Liao, Duan-Fang

    2016-01-01

    Our previous studies have indicated that a novel curcumin derivate nicotinate-curcumin (NC) has beneficial effects on the prevention of atherosclerosis, but the precise mechanisms are not fully understood. Given that autophagy regulates lipid metabolism, the present study was designed to investigate whether NC decreases foam cell formation through restoring autophagy flux in oxidized low-density lipoprotein (ox-LDL)-treated THP-1 cells. Our results showed that ox-LDL (100 μg/ml) was accumulated in THP-1 cells and impaired autophagy flux. Ox-LDL-induced impairment of autophagy was enhanced by treatment with the autophagy inhibitor chloroquine (CQ) and rescued by the autophagy inducer rapamycin. The aggregation of ox-LDL was increased by CQ, but decreased by rapamycin. In addition, colocalization of lipid droplets with LC3-II was remarkably reduced in ox-LDL group. In contrast, NC (10 μM) rescued the impaired autophagy flux by significantly increasing level of LC3-II, the number of autophagolysosomes, and the degradation of p62 in ox-LDL-treated THP-1 cells. Inhibition of the PI3K-Akt-mTOR signaling was required for NC-rescued autophagy flux. Notably, our results showed that NC remarkably promoted the colocalization of lipid droplets with autophagolysosomes, increased efflux of cholesterol, and reduced ox-LDL accumulation in THP-1 cells. However, treatment with 3-methyladenine (3-MA) or CQ reduced the protective effects of NC on lipid accumulation. Collectively, the findings suggest that NC decreases lipid accumulation in THP-1 cells through restoring autophagy flux, and further implicate that NC may be a potential therapeutic reagent to reverse atherosclerosis.

  19. Nicotinate-Curcumin Impedes Foam Cell Formation from THP-1 Cells through Restoring Autophagy Flux.

    Directory of Open Access Journals (Sweden)

    Hong-Feng Gu

    Full Text Available Our previous studies have indicated that a novel curcumin derivate nicotinate-curcumin (NC has beneficial effects on the prevention of atherosclerosis, but the precise mechanisms are not fully understood. Given that autophagy regulates lipid metabolism, the present study was designed to investigate whether NC decreases foam cell formation through restoring autophagy flux in oxidized low-density lipoprotein (ox-LDL-treated THP-1 cells. Our results showed that ox-LDL (100 μg/ml was accumulated in THP-1 cells and impaired autophagy flux. Ox-LDL-induced impairment of autophagy was enhanced by treatment with the autophagy inhibitor chloroquine (CQ and rescued by the autophagy inducer rapamycin. The aggregation of ox-LDL was increased by CQ, but decreased by rapamycin. In addition, colocalization of lipid droplets with LC3-II was remarkably reduced in ox-LDL group. In contrast, NC (10 μM rescued the impaired autophagy flux by significantly increasing level of LC3-II, the number of autophagolysosomes, and the degradation of p62 in ox-LDL-treated THP-1 cells. Inhibition of the PI3K-Akt-mTOR signaling was required for NC-rescued autophagy flux. Notably, our results showed that NC remarkably promoted the colocalization of lipid droplets with autophagolysosomes, increased efflux of cholesterol, and reduced ox-LDL accumulation in THP-1 cells. However, treatment with 3-methyladenine (3-MA or CQ reduced the protective effects of NC on lipid accumulation. Collectively, the findings suggest that NC decreases lipid accumulation in THP-1 cells through restoring autophagy flux, and further implicate that NC may be a potential therapeutic reagent to reverse atherosclerosis.

  20. Lgr5(+ve) stem/progenitor cells contribute to nephron formation during kidney development

    NARCIS (Netherlands)

    Barker, N.; Rookmaaker, M.B.; Kujala, P.; Ng, A.; Leushacke, M.; Snippert, H.; van de Wetering, M.; Tan, S.; van Es, J.H.; Huch, M.; Poulsom, R.; Verhaar, M.C.; Peters, P.J.; Clevers, H.

    2012-01-01

    Multipotent stem cells and their lineage-restricted progeny drive nephron formation within the developing kidney. Here, we document expression of the adult stem cell marker Lgr5 in the developing kidney and assess the stem/progenitor identity of Lgr5(+ve) cells via in vivo lineage tracing. The appea

  1. Lgr5(+ve) Stem/Progenitor Cells Contribute to Nephron Formation during Kidney Development

    NARCIS (Netherlands)

    Barker, Nick; Rookmaaker, Maarten B.; Kujala, Pekka; Ng, Annie; Leushacke, Marc; Snippert, Hugo; van de Wetering, Marc; Tan, Shawna; Van Es, Johan H.; Huch, Meritxell; Poulsom, Richard; Verhaar, Marianne C.; Peters, Peter J.; Clevers, Hans

    2012-01-01

    Multipotent stem cells and their lineage-restricted progeny drive nephron formation within the developing kidney. Here, we document expression of the adult stem cell marker Lgr5 in the developing kidney and assess the stem/progenitor identity of Lgr5(+ve) cells via in vivo lineage tracing. The appea

  2. Methanofullerene elongated nanostructure formation for enhanced organic solar cells

    Energy Technology Data Exchange (ETDEWEB)

    Reyes-Reyes, M. [Instituto de Investigacion en Comunicacion Optica, Universidad Autonoma de San Luis Potosi, Alvaro Obregon 64, San Luis Potosi (Mexico)], E-mail: reyesm@cactus.iico.uaslp.mx; Lopez-Sandoval, R. [Instituto Potosino de Investigacion Cientifica y Tecnologica, Camino a la presa San Jose 2055, CP 78216. San Luis Potosi (Mexico); Arenas-Alatorre, J. [Instituto de Fisica, UNAM, Apartado Postal 20-364, 01000, Mexico, D.F. (Mexico); Garibay-Alonso, R. [Instituto Potosino de Investigacion Cientifica y Tecnologica, Camino a la presa San Jose 2055, CP 78216. San Luis Potosi (Mexico); Carroll, D.L. [Center for Nanotechnology and Molecular Materials, Department of Physics. Wake Forest University, Winston-Salem NC 27109 (United States); Lastras-Martinez, A. [Instituto de Investigacion en Comunicacion Optica, Universidad Autonoma de San Luis Potosi, Alvaro Obregon 64, San Luis Potosi (Mexico)

    2007-11-01

    Using transmission electron microscopy (TEM) and Z-contrast imaging we have demonstrated elongated nanostructure formation of fullerene derivative [6,6]-phenyl-C61-butyric acid methyl ester (PCBM) within an organic host through annealing. The annealing provides an enhanced mobility of the PCBM molecules and, with good initial dispersion, allows for the formation of exaggerated grain growth within the polymer host. We have assembled these nanostructures within the regioregular conjugated polymer poly(3-hexylthiophene) (P3HT). This PCBM elongated nanostructure formation maybe responsible for the very high efficiencies observed, at very low loadings of PCBM (1:0.6, polymer to PCBM), in annealed photovoltaics. Moreover, our high resolution TEM and electron energy loss spectroscopy studies clearly show that the PCBM crystals remain crystalline and are unaffected by the 200-keV electron beam.

  3. Methanofullerene elongated nanostructure formation for enhanced organic solar cells

    International Nuclear Information System (INIS)

    Using transmission electron microscopy (TEM) and Z-contrast imaging we have demonstrated elongated nanostructure formation of fullerene derivative [6,6]-phenyl-C61-butyric acid methyl ester (PCBM) within an organic host through annealing. The annealing provides an enhanced mobility of the PCBM molecules and, with good initial dispersion, allows for the formation of exaggerated grain growth within the polymer host. We have assembled these nanostructures within the regioregular conjugated polymer poly(3-hexylthiophene) (P3HT). This PCBM elongated nanostructure formation maybe responsible for the very high efficiencies observed, at very low loadings of PCBM (1:0.6, polymer to PCBM), in annealed photovoltaics. Moreover, our high resolution TEM and electron energy loss spectroscopy studies clearly show that the PCBM crystals remain crystalline and are unaffected by the 200-keV electron beam

  4. Formation and action of oxygen activated species in cell cultures

    International Nuclear Information System (INIS)

    The differences of hydrogen peroxide sensibility of mammal cell lineages (man, mouse, chinese hamster) in culture are studied. The cellular survival and the frequency of DNA induced breaks by hydrogen peroxide are analysed. The efficiency of elimination of DNA breaks by cells is determined. The possible relation between the cell capacity of repair and its survival to hydrogen peroxide action is also discussed. (M.A.)

  5. Thrombospondin 1 promotes synaptic formation in bone marrow-derived neuron-like cells

    Institute of Scientific and Technical Information of China (English)

    Yun Huang; Mingnan Lu; Weitao Guo; Rong Zeng; Bin Wang; Huaibo Wang

    2013-01-01

    In this study, a combination of growth factors was used to induce bone marrow mesenchymal stem cells differentiation into neuron-like cells, in a broader attempt to observe the role of thrombospondin 1 in synapse formation. Results showed that there was no significant difference in the differentiation rate of neuron-like cells between bone marrow mesenchymal stem cells with thrombospondin induction and those without. However, the cell shape was more complex and the neurites were dendritic, with unipolar, bipolar or multipolar morphologies, after induction with thrombospondin 1. The induced cells were similar in morphology to normal neurites. Immunohistochemical staining showed that the number of positive cells for postsynaptic density protein 95 and synaptophysin 1 protein was significantly increased after induction with thrombospondin 1. These findings indicate that thrombospondin 1 promotes synapse formation in neuron-like cells that are differentiated from bone marrow mesenchymal stem cells.

  6. Endothelial cell motility, coordination and pattern formation during vasculogenesis.

    Science.gov (United States)

    Czirok, Andras

    2013-01-01

    How vascular networks assemble is a fundamental problem of developmental biology that also has medical importance. To explain the organizational principles behind vascular patterning, we must understand how can tissue level structures be controlled through cell behavior patterns like motility and adhesion that, in turn, are determined by biochemical signal transduction processes? We discuss the various ideas that have been proposed as mechanisms for vascular network assembly: cell motility guided by extracellular matrix alignment (contact guidance), chemotaxis guided by paracrine and autocrine morphogens, and multicellular sprouting guided by cell-cell contacts. All of these processes yield emergent patterns, thus endothelial cells can form an interconnected structure autonomously, without guidance from an external pre-pattern.

  7. Laser-based techniques for living cell pattern formation

    Science.gov (United States)

    Hopp, Béla; Smausz, Tomi; Papdi, Bence; Bor, Zsolt; Szabó, András; Kolozsvári, Lajos; Fotakis, Costas; Nógrádi, Antal

    2008-10-01

    In the production of biosensors or artificial tissues a basic step is the immobilization of living cells along the required pattern. In this paper the ability of some promising laser-based methods to influence the interaction between cells and various surfaces is presented. In the first set of experiments laser-induced patterned photochemical modification of polymer foils was used to achieve guided adherence and growth of cells to the modified areas: (a) Polytetrafluoroethylene was irradiated with ArF excimer laser ( λ=193 nm, FWHM=20 ns, F=9 mJ/cm2) in presence of triethylene tetramine liquid photoreagent; (b) a thin carbon layer was produced by KrF excimer laser ( λ=248 nm, FWHM=30 ns, F=35 mJ/cm2) irradiation on polyimide surface to influence the cell adherence. It was found that the incorporation of amine groups in the PTFE polymer chain instead of the fluorine atoms can both promote and prevent the adherence of living cells (depending on the applied cell types) on the treated surfaces, while the laser generated carbon layer on polyimide surface did not effectively improve adherence. Our attempts to influence the cell adherence by morphological modifications created by ArF laser irradiation onto polyethylene terephtalate surface showed a surface roughness dependence. This method was effective only when the Ra roughness parameter of the developed structure did not exceed the 0.1 micrometer value. Pulsed laser deposition with femtosecond KrF excimer lasers ( F=2.2 J/cm2) was effectively used to deposit structured thin films from biomaterials (endothelial cell growth supplement and collagen embedded in starch matrix) to promote the adherence and growth of cells. These results present evidence that some surface can be successfully altered to induce guided cell growth.

  8. A Mathematical Programming Model for Cell Formation Problem with Machine Replication

    Directory of Open Access Journals (Sweden)

    Reza Raminfar

    2013-01-01

    Full Text Available Cell formation (CF is a crucial aspect in the design of cellular manufacturing (CM systems. This paper develops a comprehensive mathematical programming model for the cell formation problem, where product demands, cell size limits, sequence of operations, multiple units of identical machines, machine capacity, or machine cost are all considered. In this model, the intercell moves are restricted to be unidirectional from one cell to the downstream cells, without backtracking. The proposed model is investigated through several numerical examples. To evaluate the solution quality of the proposed model, it is compared with some well-known cell formation methods from the literature, by using group capability index (GCI as a performance measure. The results and comparisons indicate that the proposed model produces solution with a higher performance.

  9. A Theoretical Model of Jigsaw-Puzzle Pattern Formation by Plant Leaf Epidermal Cells.

    Science.gov (United States)

    Higaki, Takumi; Kutsuna, Natsumaro; Akita, Kae; Takigawa-Imamura, Hisako; Yoshimura, Kenji; Miura, Takashi

    2016-04-01

    Plant leaf epidermal cells exhibit a jigsaw puzzle-like pattern that is generated by interdigitation of the cell wall during leaf development. The contribution of two ROP GTPases, ROP2 and ROP6, to the cytoskeletal dynamics that regulate epidermal cell wall interdigitation has already been examined; however, how interactions between these molecules result in pattern formation remains to be elucidated. Here, we propose a simple interface equation model that incorporates both the cell wall remodeling activity of ROP GTPases and the diffusible signaling molecules by which they are regulated. This model successfully reproduces pattern formation observed in vivo, and explains the counterintuitive experimental results of decreased cellulose production and increased thickness. Our model also reproduces the dynamics of three-way cell wall junctions. Therefore, this model provides a possible mechanism for cell wall interdigitation formation in vivo.

  10. The direct formate fuel cell with an alkaline anion exchange membrane

    Science.gov (United States)

    Bartrom, Amy M.; Haan, John L.

    2012-09-01

    We demonstrate for the first time an operating Direct Formate Fuel Cell employing formate salts as the anode fuel, air or oxygen as the oxidant, a polymer anion exchange membrane, and metal catalysts at the anode and cathode. Operation of the DFFC at 60 °C using 1 M KOOCH and 2 M KOH as the anode fuel and electrolyte and oxygen gas at the cathode produces 144 mW cm-2 of peak power density, 181 mA cm-2 current density at 0.6 V, and an open circuit voltage of 0.931 V. This performance is competitive with alkaline Direct Liquid Fuel Cells (DLFCs) previously reported in the literature and demonstrates that formate fuel is a legitimate contender with alcohol fuels for alkaline DLFCs. A survey of the literature shows that a formate-oxygen fuel cell has a high theoretical potential, and the safe, renewable formate fuel does not poison the anode catalyst.

  11. Mechanical Model of Geometric Cell and Topological Algorithm for Cell Dynamics from Single-Cell to Formation of Monolayered Tissues with Pattern

    KAUST Repository

    Kachalo, Sëma

    2015-05-14

    Geometric and mechanical properties of individual cells and interactions among neighboring cells are the basis of formation of tissue patterns. Understanding the complex interplay of cells is essential for gaining insight into embryogenesis, tissue development, and other emerging behavior. Here we describe a cell model and an efficient geometric algorithm for studying the dynamic process of tissue formation in 2D (e.g. epithelial tissues). Our approach improves upon previous methods by incorporating properties of individual cells as well as detailed description of the dynamic growth process, with all topological changes accounted for. Cell size, shape, and division plane orientation are modeled realistically. In addition, cell birth, cell growth, cell shrinkage, cell death, cell division, cell collision, and cell rearrangements are now fully accounted for. Different models of cell-cell interactions, such as lateral inhibition during the process of growth, can be studied in detail. Cellular pattern formation for monolayered tissues from arbitrary initial conditions, including that of a single cell, can also be studied in detail. Computational efficiency is achieved through the employment of a special data structure that ensures access to neighboring cells in constant time, without additional space requirement. We have successfully generated tissues consisting of more than 20,000 cells starting from 2 cells within 1 hour. We show that our model can be used to study embryogenesis, tissue fusion, and cell apoptosis. We give detailed study of the classical developmental process of bristle formation on the epidermis of D. melanogaster and the fundamental problem of homeostatic size control in epithelial tissues. Simulation results reveal significant roles of solubility of secreted factors in both the bristle formation and the homeostatic control of tissue size. Our method can be used to study broad problems in monolayered tissue formation. Our software is publicly

  12. Matrix elasticity of void-forming hydrogels controls transplanted-stem-cell-mediated bone formation

    Science.gov (United States)

    Huebsch, Nathaniel; Lippens, Evi; Lee, Kangwon; Mehta, Manav; Koshy, Sandeep T.; Darnell, Max C.; Desai, Rajiv M.; Madl, Christopher M.; Xu, Maria; Zhao, Xuanhe; Chaudhuri, Ovijit; Verbeke, Catia; Kim, Woo Seob; Alim, Karen; Mammoto, Akiko; Ingber, Donald E.; Duda, Georg N.; Mooney, David J.

    2015-12-01

    The effectiveness of stem cell therapies has been hampered by cell death and limited control over fate. These problems can be partially circumvented by using macroporous biomaterials that improve the survival of transplanted stem cells and provide molecular cues to direct cell phenotype. Stem cell behaviour can also be controlled in vitro by manipulating the elasticity of both porous and non-porous materials, yet translation to therapeutic processes in vivo remains elusive. Here, by developing injectable, void-forming hydrogels that decouple pore formation from elasticity, we show that mesenchymal stem cell (MSC) osteogenesis in vitro, and cell deployment in vitro and in vivo, can be controlled by modifying, respectively, the hydrogel’s elastic modulus or its chemistry. When the hydrogels were used to transplant MSCs, the hydrogel’s elasticity regulated bone regeneration, with optimal bone formation at 60 kPa. Our findings show that biophysical cues can be harnessed to direct therapeutic stem cell behaviours in situ.

  13. Antibody to a molecular marker of cell position inhibits synapse formation in retina.

    OpenAIRE

    Trisler, D.; Bekenstein, J; Daniels, M P

    1986-01-01

    A topographic gradient of TOP molecules in retina can be used to identify neuron position. Antibody to TOP from hybridoma cells that were injected into in vivo embryo eyes diffused into the retina and bound in a topographic gradient of [antibody.TOP] ([Ab.TOP]) complexes. Synapse formation in retina was inhibited in the presence of anti-TOP antibody. This suggests that TOP is involved in synapse formation and that recognition of position by neurons is necessary for normal synapse formation.

  14. Midbody remnant licenses primary cilia formation in epithelial cells.

    Science.gov (United States)

    Ott, Carolyn M

    2016-08-01

    Tethered midbody remnants dancing across apical microvilli, encountering the centrosome, and beckoning forth a cilium-who would have guessed this is how polarized epithelial cells coordinate the end of mitosis and the beginning of ciliogenesis? New evidence from Bernabé-Rubio et al. (2016. J. Cell Biol http://dx.doi.org/10.1083/jcb.201601020) supports this emerging model. PMID:27482049

  15. Blockade of Mast Cell Activation Reduces Cutaneous Scar Formation

    OpenAIRE

    Lin Chen; Megan E Schrementi; Ranzer, Matthew J.; Wilgus, Traci A.; Luisa A DiPietro

    2014-01-01

    Damage to the skin initiates a cascade of well-orchestrated events that ultimately leads to repair of the wound. The inflammatory response is key to wound healing both through preventing infection and stimulating proliferation and remodeling of the skin. Mast cells within the tissue are one of the first immune cells to respond to trauma, and upon activation they release pro-inflammatory molecules to initiate recruitment of leukocytes and promote a vascular response in the tissue. Additionally...

  16. Ancestral vascular lumen formation via basal cell surfaces

    OpenAIRE

    Tomás Kucera; Boris Strilić; Kathrin Regener; Michael Schubert; Vincent Laudet; Eckhard Lammert

    2015-01-01

    The cardiovascular system of bilaterians developed from a common ancestor. However, no endothelial cells exist in invertebrates demonstrating that primitive cardiovascular tubes do not require this vertebrate-specific cell type in order to form. This raises the question of how cardiovascular tubes form in invertebrates? Here we discovered that in the invertebrate cephalochordate amphioxus, the basement membranes of endoderm and mesoderm line the lumen of the major vessels, namely aorta and he...

  17. Hypertrophic scar formation is associated with an increased number of epidermal Langerhans cells.

    NARCIS (Netherlands)

    Niessen, F.B.; Schalkwijk, J.; Vos, H.; Timens, W.

    2004-01-01

    The exact pathogenesis of hypertrophic scar and keloid formation is still unknown and a good therapy to prevent or treat these scars is lacking. Because immunological processes seem to be important in excessive scar formation, immunological cells and parameters were studied in a standardized breast

  18. Hypertrophic scar formation is associated with an increased number of epidermal Langerhans cells

    NARCIS (Netherlands)

    Niessen, FB; Schalkwijk, J; Vos, H; Timens, W

    2004-01-01

    The exact pathogenesis of hypertrophic scar and keloid formation is still unknown and a good therapy to prevent or treat these scars is lacking. Because immunological processes seem to be important in excessive scar formation, immunological cells and parameters were studied in a standardized breast

  19. Melanoma Cell Expression of CD200 Inhibits Tumor Formation and Lung Metastasis via Inhibition of Myeloid Cell Functions

    OpenAIRE

    Talebian, Fatemeh; Liu, Jin-Qing; Liu, Zhenzhen; Khattabi, Mazin; He, Yukai; Ganju, Ramesh; Bai, Xue-feng

    2012-01-01

    CD200 is a cell surface glycoprotein that functions through engaging CD200 receptor on cells of the myeloid lineage and inhibits their functions. Expression of CD200 has been implicated in a variety of human cancer cells including melanoma cells and has been thought to play a protumor role. To investigate the role of cancer cell expression of CD200 in tumor formation and metastasis, we generated CD200-positive and CD200-negative B16 melanoma cells. Subcutaneous injection of CD200-positive B16...

  20. Melanoma cell expression of CD200 inhibits tumor formation and lung metastasis via inhibition of myeloid cell functions.

    Directory of Open Access Journals (Sweden)

    Fatemeh Talebian

    Full Text Available CD200 is a cell surface glycoprotein that functions through engaging CD200 receptor on cells of the myeloid lineage and inhibits their functions. Expression of CD200 has been implicated in a variety of human cancer cells including melanoma cells and has been thought to play a protumor role. To investigate the role of cancer cell expression of CD200 in tumor formation and metastasis, we generated CD200-positive and CD200-negative B16 melanoma cells. Subcutaneous injection of CD200-positive B16 melanoma cells inhibited tumor formation and growth in C57BL/6 mice but not in Rag1⁻/⁻C57BL/6 mice. However, i.v. injection of CD200-positive B16 melanoma cells dramatically inhibited tumor foci formation in the lungs of both C57BL/6 and Rag1⁻/⁻C57BL6 mice. Flow cytometry analysis revealed higher expression of CD200R in Gr1⁺ myeloid cells in the lung than in peripheral myeloid cells. Depletion of Gr1⁺ cells or stimulation of CD200R with an agonistic antibody in vivo dramatically inhibited tumor foci formation in the lungs. In addition, treatment with tumor antigen specific CD4 or CD8 T cells or their combination yielded a survival advantage for CD200 positive tumor bearing mice over mice bearing CD200-negative tumors. Taken together, we have revealed a novel role for CD200-CD200R interaction in inhibiting tumor formation and metastasis. Targeting CD200R may represent a novel approach for cancer immunotherapy.

  1. Mirror-symmetric microtubule assembly and cell interactions drive lumen formation in the zebrafish neural rod

    OpenAIRE

    Buckley, Clare E.; Ren, Xiaoyun; Ward, Laura C; Girdler, Gemma C; Araya, Claudio; Green, Mary J; Clark, Brian S.; Link, Brian A.; Clarke, Jonathan D. W.

    2012-01-01

    By analysing the cellular and subcellular events that occur in the centre of the developing zebrafish neural rod, we have uncovered a novel mechanism of cell polarisation during lumen formation. Cells from each side of the neural rod interdigitate across the tissue midline. This is necessary for localisation of apical junctional proteins to the region where cells intersect the tissue midline. Cells assemble a mirror-symmetric microtubule cytoskeleton around the tissue midline, which is necess...

  2. MECHANICAL VIBRATION INHIBITS OSTEOCLAST FORMATION BY REDUCING DC-STAMP RECEPTOR EXPRESSION IN OSTEOCLAST PRECURSOR CELLS

    OpenAIRE

    Kulkarni, R. N.; P. A. Voglewede; Liu, D.

    2013-01-01

    It is well known that physical inactivity leads to loss of muscle mass, but it also causes bone loss. Mechanistically, osteoclastogenesis and bone resorption have recently been shown to be regulated by vibration. However, the underlying mechanism behind the inhibition of osteoclast formation is yet unknown. Therefore, we investigated whether mechanical vibration of osteoclast precursor cells affects osteoclast formation by the involvement of fusion-related molecules such as dendritic cell-spe...

  3. Fibrinogen-Induced Streptococcus mutans Biofilm Formation and Adherence to Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Telma Blanca Lombardo Bedran

    2013-01-01

    Full Text Available Streptococcus mutans, the predominant bacterial species associated with dental caries, can enter the bloodstream and cause infective endocarditis. The aim of this study was to investigate S. mutans biofilm formation and adherence to endothelial cells induced by human fibrinogen. The putative mechanism by which biofilm formation is induced as well as the impact of fibrinogen on S. mutans resistance to penicillin was also evaluated. Bovine plasma dose dependently induced biofilm formation by S. mutans. Of the various plasma proteins tested, only fibrinogen promoted the formation of biofilm in a dose-dependent manner. Scanning electron microscopy observations revealed the presence of complex aggregates of bacterial cells firmly attached to the polystyrene support. S. mutans in biofilms induced by the presence of fibrinogen was markedly resistant to the bactericidal effect of penicillin. Fibrinogen also significantly increased the adherence of S. mutans to endothelial cells. Neither S. mutans cells nor culture supernatants converted fibrinogen into fibrin. However, fibrinogen is specifically bound to the cell surface of S. mutans and may act as a bridging molecule to mediate biofilm formation. In conclusion, our study identified a new mechanism promoting S. mutans biofilm formation and adherence to endothelial cells which may contribute to infective endocarditis.

  4. Collagen-IV supported embryoid bodies formation and differentiation from buffalo (Bubalus bubalis) embryonic stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Taru Sharma, G., E-mail: gts553@gmail.com [Reproductive Physiology Laboratory, Division of Physiology and Climatology, Indian Veterinary Research Institute, Izatnagar-243 122, Bareilly, U.P. (India); Dubey, Pawan K.; Verma, Om Prakash; Pratheesh, M.D.; Nath, Amar; Sai Kumar, G. [Reproductive Physiology Laboratory, Division of Physiology and Climatology, Indian Veterinary Research Institute, Izatnagar-243 122, Bareilly, U.P. (India)

    2012-08-03

    Graphical abstract: EBs formation, characterization and expression of germinal layers marker genes of in vivo developed teratoma using four different types of extracellular matrices. Highlights: Black-Right-Pointing-Pointer Collagen-IV matrix is found cytocompatible for EBs formation and differentiation. Black-Right-Pointing-Pointer Established 3D microenvironment for ES cells development and differentiation into three germ layers. Black-Right-Pointing-Pointer Collagen-IV may be useful as promising candidate for ES cells based therapeutic applications. -- Abstract: Embryoid bodies (EBs) are used as in vitro model to study early extraembryonic tissue formation and differentiation. In this study, a novel method using three dimensional extracellular matrices for in vitro generation of EBs from buffalo embryonic stem (ES) cells and its differentiation potential by teratoma formation was successfully established. In vitro derived inner cell masses (ICMs) of hatched buffalo blastocyst were cultured on buffalo fetal fibroblast feeder layer for primary cell colony formation. For generation of EBs, pluripotent ES cells were seeded onto four different types of extracellular matrices viz; collagen-IV, laminin, fibronectin and matrigel using undifferentiating ES cell culture medium. After 5 days of culture, ESCs gradually grew into aggregates and formed simple EBs having circular structures. Twenty-six days later, they formed cystic EBs over collagen matrix with higher EBs formation and greater proliferation rate as compared to other extracellular matrices. Studies involving histological observations, fluorescence microscopy and RT-PCR analysis of the in vivo developed teratoma revealed that presence of all the three germ layer derivatives viz. ectoderm (NCAM), mesoderm (Flk-1) and endoderm (AFP). In conclusion, the method described here demonstrates a simple and cost-effective way of generating EBs from buffalo ES cells. Collagen-IV matrix was found cytocompatible as it

  5. Biophysical Properties of Scaffolds Modulate Human Blood Vessel Formation from Circulating Endothelial Colony-Forming Cells

    Science.gov (United States)

    Critser, Paul J.; Yoder, Mervin C.

    A functional vascular system forms early in development and is continually remodeled throughout the life of the organism. Impairment to the regeneration or repair of this system leads to tissue ischemia, dysfunction, and disease. The process of vascular formation and remodeling is complex, relying on local microenvironmental cues, cytokine signaling, and multiple cell types to function properly. Tissue engineering strategies have attempted to exploit these mechanisms to develop functional vascular networks for the generation of artificial tissues and therapeutic strategies to restore tissue homeostasis. The success of these strategies requires the isolation of appropriate progenitor cell sources which are straightforward to obtain, display high proliferative potential, and demonstrate an ability to form functional vessels. Several populations are of interest including endothelial colony-forming cells, a subpopulation of endothelial progenitor cells. Additionally, the development of scaffolds to deliver and support progenitor cell survival and function is crucial for the formation of functional vascular networks. The composition and biophysical properties of these scaffolds have been shown to modulate endothelial cell behavior and vessel formation. However, further investigation is needed to better understand how these mechanical properties and biophysical properties impact vessel formation. Additionally, several other cell populations are involved in neoangiogenesis and formation of tissue parenchyma and an understanding of the potential impact of these cell populations on the biophysical properties of scaffolds will also be needed to advance these strategies. This chapter examines how the biophysical properties of matrix scaffolds can influence vessel formation and remodeling and, in particular, the impact on in vivo human endothelial progenitor cell vessel formation.

  6. Collagen-IV supported embryoid bodies formation and differentiation from buffalo (Bubalus bubalis) embryonic stem cells

    International Nuclear Information System (INIS)

    Graphical abstract: EBs formation, characterization and expression of germinal layers marker genes of in vivo developed teratoma using four different types of extracellular matrices. Highlights: ► Collagen-IV matrix is found cytocompatible for EBs formation and differentiation. ► Established 3D microenvironment for ES cells development and differentiation into three germ layers. ► Collagen-IV may be useful as promising candidate for ES cells based therapeutic applications. -- Abstract: Embryoid bodies (EBs) are used as in vitro model to study early extraembryonic tissue formation and differentiation. In this study, a novel method using three dimensional extracellular matrices for in vitro generation of EBs from buffalo embryonic stem (ES) cells and its differentiation potential by teratoma formation was successfully established. In vitro derived inner cell masses (ICMs) of hatched buffalo blastocyst were cultured on buffalo fetal fibroblast feeder layer for primary cell colony formation. For generation of EBs, pluripotent ES cells were seeded onto four different types of extracellular matrices viz; collagen-IV, laminin, fibronectin and matrigel using undifferentiating ES cell culture medium. After 5 days of culture, ESCs gradually grew into aggregates and formed simple EBs having circular structures. Twenty-six days later, they formed cystic EBs over collagen matrix with higher EBs formation and greater proliferation rate as compared to other extracellular matrices. Studies involving histological observations, fluorescence microscopy and RT-PCR analysis of the in vivo developed teratoma revealed that presence of all the three germ layer derivatives viz. ectoderm (NCAM), mesoderm (Flk-1) and endoderm (AFP). In conclusion, the method described here demonstrates a simple and cost-effective way of generating EBs from buffalo ES cells. Collagen-IV matrix was found cytocompatible as it supported buffalo EBs formation, their subsequent differentiation could prove to

  7. Raman scattering evidence of hydrohalite formation on frozen yeast cells

    CERN Document Server

    Okotrub, K A

    2012-01-01

    We studied yeast cells in physiological solution during freezing by Raman microspectroscopy technique. The purpose was to find out the origin of a sharp peak near ~3430 cm^-1 in Raman spectrum of frozen mammalian cells, observed earlier (J. Dong et al, Biophys. J., 99 (2010) 2453), which presumably could be used as an indicator of intracellar ice appearance. We have shown that this line (actually doublet of 3408 and 3425 cm^-1) corresponds to Raman spectrum of hydrohalite (NaCl-2H2O), which is formed as the result of the eutectic crystallization of the liquid solution around the cells. We also show that the spatial distribution of hydrohalite in the sample significantly depends on the cooling rate. At lower cooling rate (1{\\deg}C/min), products of eutectic crystallization form layer on the cell surface which thickness varies for different cells and can reach ~1 {\\mu}m in thickness. At higher cooling rate (20{\\deg}C/min), the hydrohalite distribution appears more homogeneous, in the sample, and the eutectic cr...

  8. Biofilm formation on polystyrene in detached vs. planktonic cells of polyhydroxyalkanoate-accumulating Halomonas venusta.

    Science.gov (United States)

    Berlanga, Mercedes; Domènech, Òscar; Guerrero, Ricardo

    2014-12-01

    Biofilm development is characterized by distinct stages of initial attachment, microcolony formation and maturation (sessile cells), and final detachment (dispersal of new, planktonic cells). In this work we examined the influence of polyhydroxyalkanoate (PHA) accumulation on bacterial surface properties and biofilm formation on polystyrene in detached vs. planktonic cells of an environmental strain isolated from microbial mats, Halomonas venusta MAT28. This strain was cultured either in an artificial biofilm in which the cells were immobilized on alginate beads (sessile) or as free-swimming (planktonic) cells. For the two modes of growth, conditions allowing or preventing PHA accumulation were established. Cells detached from alginate beads and their planktonic counterparts were used to study cell surface properties and cellular adhesion on polystyrene. Detached cells showed a slightly higher affinity than planktonic cells for chloroform (Lewis-acid) and a greater hydrophobicity (affinity for hexadecane and hexane). Those surface characteristics of the detached cells may explain their better adhesion on polystyrene compared to planktonic cells. Adhesion to polystyrene was not significantly different between H. venusta cells that had accumulated PHA vs. those that did not. These observations suggest that the surface properties of detached cells clearly differ from those of planktonic cells and that for at least the first 48 h after detachment from alginate beads H. venusta retained the capacity of sessile cells to adhere to polystyrene and to form a biofilm. PMID:26421734

  9. Cdc42 is not essential for filopodium formation, directed migration, cell polarization, and mitosis in fibroblastoid cells

    DEFF Research Database (Denmark)

    Czuchra, Aleksandra; Wu, Xunwei; Meyer, Hannelore;

    2005-01-01

    of Cdc42 did not affect filopodium or lamellipodium formation and had no significant influence on the speed of directed migration nor on mitosis. Cdc42-deficient cells displayed a more elongated cell shape and had a reduced area. Furthermore, directionality during migration and reorientation of the Golgi...

  10. High-Throughput Single-Cell Derived Sphere Formation for Cancer Stem-Like Cell Identification and Analysis

    Science.gov (United States)

    Chen, Yu-Chih; Ingram, Patrick N.; Fouladdel, Shamileh; McDermott, Sean P.; Azizi, Ebrahim; Wicha, Max S.; Yoon, Euisik

    2016-06-01

    Considerable evidence suggests that many malignancies are driven by a cellular compartment that displays stem cell properties. Cancer stem-like cells (CSCs) can be identified by expression of cell surface markers or enzymatic activity, but these methods are limited by phenotypic heterogeneity and plasticity of CSCs. An alternative phenotypic methodology based on in-vitro sphere formation has been developed, but it is typically labor-intensive and low-throughput. In this work, we present a 1,024-microchamber microfluidic platform for single-cell derived sphere formation. Utilizing a hydrodynamic capturing scheme, more than 70% of the microchambers capture only one cell, allowing for monitoring of sphere formation from heterogeneous cancer cell populations for identification of CSCs. Single-cell derived spheres can be retrieved and dissociated for single-cell analysis using a custom 96-gene panel to probe heterogeneity within the clonal CSC spheres. This microfluidic platform provides reliable and high-throughput sphere formation for CSC identification and downstream clonal analysis.

  11. Ectopic bone formation in bone marrow stem cell seeded calcium phosphate scaffolds as compared to autograft and (cell seeded allograft

    Directory of Open Access Journals (Sweden)

    J O Eniwumide

    2007-08-01

    Full Text Available Improvements to current therapeutic strategies are needed for the treatment of skeletal defects. Bone tissue engineering offers potential advantages to these strategies. In this study, ectopic bone formation in a range of scaffolds was assessed. Vital autograft and devitalised allograft served as controls and the experimental groups comprised autologous bone marrow derived stem cell seeded allograft, biphasic calcium phosphate (BCP and tricalcium phosphate (TCP, respectively. All implants were implanted in the back muscle of adult Dutch milk goats for 12 weeks. Micro-computed tomography (µCT analysis and histomorphometry was performed to evaluate and quantify ectopic bone formation. In good agreement, both µCT and histomorphometric analysis demonstrated a significant increase in bone formation by cell-seeded calcium phosphate scaffolds as compared to the autograft, allograft and cell-seeded allograft implants. An extensive resorption of the autograft, allograft and cell-seeded allograft implants was observed by histology and confirmed by histomorphometry. Cell-seeded TCP implants also showed distinct signs of degradation with histomorphometry and µCT, while the degradation of the cell-seeded BCP implants was negligible. These results indicate that cell-seeded calcium phosphate scaffolds are superior to autograft, allograft or cell-seeded allograft in terms of bone formation at ectopic implantation sites. In addition, the usefulness of µCT for the efficient and non-destructive analysis of mineralised bone and calcium phosphate scaffold was demonstrated.

  12. Rap1 integrates tissue polarity, lumen formation, and tumorigenicpotential in human breast epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Itoh, Masahiko; Nelson, Celeste M.; Myers, Connie A.; Bissell,Mina J.

    2006-09-29

    Maintenance of apico-basal polarity in normal breast epithelial acini requires a balance between cell proliferation, cell death, and proper cell-cell and cell-extracellular matrix signaling. Aberrations in any of these processes can disrupt tissue architecture and initiate tumor formation. Here we show that the small GTPase Rap1 is a crucial element in organizing acinar structure and inducing lumen formation. Rap1 activity in malignant HMT-3522 T4-2 cells is appreciably higher than in S1 cells, their non-malignant counterparts. Expression of dominant-negative Rap1 resulted in phenotypic reversion of T4-2 cells, led to formation of acinar structures with correct apico-basal polarity, and dramatically reduced tumor incidence despite the persistence of genomic abnormalities. The resulting acini contained prominent central lumina not observed when other reverting agents were used. Conversely, expression of dominant-active Rap1 in T4-2 cells inhibited phenotypic reversion and led to increased invasiveness and tumorigenicity. Thus, Rap1 acts as a central regulator of breast architecture, with normal levels of activation instructing apical polarity during acinar morphogenesis, and increased activation inducing tumor formation and progression to malignancy.

  13. 3D in vitro cell culture models of tube formation.

    Science.gov (United States)

    Zegers, Mirjam M

    2014-07-01

    Building the complex architecture of tubular organs is a highly dynamic process that involves cell migration, polarization, shape changes, adhesion to neighboring cells and the extracellular matrix, physicochemical characteristics of the extracellular matrix and reciprocal signaling with the mesenchyme. Understanding these processes in vivo has been challenging as they take place over extended time periods deep within the developing organism. Here, I will discuss 3D in vitro models that have been crucial to understand many of the molecular and cellular mechanisms and key concepts underlying branching morphogenesis in vivo. PMID:24613912

  14. Investigation of Contact Formation during Silicon Solar Cell Production

    Science.gov (United States)

    Mojrová, Barbora

    2016-05-01

    This article deals with the investigation of the influence of sintering conditions on the formation process of screen printed contacts on passivated boron doped P+ emitters. The experiment was focused on measuring of resistance changes of two thick film pastes during firing processes with different conditions. Two different temperature profiles were compared at an atmospheric concentration of O2. The influence of the O2 concentration on resistance was investigated for one profile. A rapid thermal processing furnace modified for in-situ resistance measurements was used. The change of resistance was measured simultaneously with the temperature.

  15. Rapid Formation of Cell Aggregates and Spheroids Induced by a "Smart" Boronic Acid Copolymer.

    Science.gov (United States)

    Amaral, Adérito J R; Pasparakis, George

    2016-09-01

    Cell surface engineering has emerged as a powerful approach to forming cell aggregates/spheroids and cell-biomaterial ensembles with significant uses in tissue engineering and cell therapeutics. Herein, we demonstrate that cell membrane remodeling with a thermoresponsive boronic acid copolymer induces the rapid formation of spheroids using either cancer or cardiac cell lines under conventional cell culture conditions at minute concentrations. It is shown that the formation of well-defined spheroids is accelerated by at least 24 h compared to non-polymer-treated controls, and, more importantly, the polymer allows for fine control of the aggregation kinetics owing to its stimulus response to temperature and glucose content. On the basis of its simplicity and effectiveness to promote cellular aggregation, this platform holds promise in three-dimensional tissue/tumor modeling and tissue engineering applications. PMID:27571512

  16. Mesenchymal Stem Cell (MSC) Aggregate Formation in vivo

    Science.gov (United States)

    Bartosh, Thomas J.; Ylostalo, Joni H.

    2016-01-01

    Human mesenchymal stem/progenitor cells (MSCs) isolated from various adult tissues show remarkable therapeutic potential and are being employed in clinical trials for the treatment of numerous diseases (Prockop et al., 2010). While routes of cell administration vary, profound beneficial effects of MSCs in animal models have been observed following intraperitoneal injections of the cells (Roddy et al., 2011). Similar to MSC spheres formed in culture under conditions where attachment to plastic is not permitted (Bartosh et al., 2010), MSCs injected into the peritoneum of mice spontaneously aggregate into 3D sphere-like structures (Bartosh et al., 2013). During the process of sphere assembly and compaction, MSCs upregulate expression of numerous therapeutic anti-inflammatory and immune modulatory factors. Here we describe the method we previously used for the generation of human bone marrow-derived MSC aggregates/spheres in vivo (Bartosh et al., 2013). By tagging the MSCs with green fluorescent protein (GFP), the aggregates formed can be easily visualized, collected and analyzed for changes in cellular properties and interactions with host immune cells.

  17. Mobilization of regulatory T cells in response to carotid injury does not influence subsequent neointima formation.

    Directory of Open Access Journals (Sweden)

    Amit Saxena

    Full Text Available AIM: T cells have been attributed an important role in modulating repair responses following vascular injury. The aim of this study was to investigate the role of different T cell subsets in this context. METHODS AND RESULTS: A non-obstructive collar was introduced to inflict carotid artery injury in mice and subsequent activation of immune cells in draining lymph nodes and spleen were studied by flow cytometry. Carotid artery injury of wild type mice was associated with mobilization of both Th1 type CD4(+IFNγ(+ and regulatory CD4(+CD25(+FoxP3(+ T cells in draining lymph nodes. Studies using FoxP3-green fluorescent protein (GFP transgenic C57/Bl6 mice demonstrated scattered presence of regulatory T cells in the adventitial tissue of injured arteries as well as a massive emigration of regulatory T cells from the spleen in response to carotid injury. However, deletion of antigen presentation to CD4+ T cells (H2(0 mice, as well as deletion of regulatory T cells (through treatment with blocking anti-CD25 antibodies, did not affect neointima formation. Also deletion of antigen presentation to CD8(+ T cells (Tap1(0 mice was without effect on carotid collar-induced neointima formation. CONCLUSION: The results demonstrate that carotid artery injury is associated with mobilization of regulatory T cells. Depletion of regulatory T cells does not, however, influence the subsequent repair processes leading to the formation of a neointima. The results also demonstrate that lack of CD8(+ T cells does not influence neointima formation in presence of functional CD4(+ T cells and B cells.

  18. Neocarzinostatin-induced Rad51 nuclear focus formation is cell cycle regulated and aberrant in AT cells

    International Nuclear Information System (INIS)

    DNA double-stranded breaks are the most detrimental form of DNA damage and, if not repaired properly, may lead to an accumulation of chromosomal aberrations and eventually tumorigenesis. Proteins of the Rad51/Rad52 epitasis group are crucial for the recombinational repair of DNA double-stranded breaks, whereas the Rad50/NBS1/Mre11 nuclease complex is involved in both the recombinational and the end-joining repair of DNA double-stranded breaks. Herein, we demonstrate that the chemotherapeutic enediyne antibiotic neocarzinostatin induced Rad51, but not NBS1, nuclear focus formation in a cell- cycle-dependent manner. Furthermore, neocarzinostatin-induced Rad51 foci formation revealed a slower kinetic change in AT cells, but not in wild-type or NBS cells. In summary, our results suggest that neocarzinostatin induces Rad51 focus formation through an ATM- and cell-cycle-dependent, but NBS1-independent, pathway

  19. BCL6 interacting corepressor contributes to germinal center T follicular helper cell formation and B cell helper function

    OpenAIRE

    Yang, Jessica A.; Tubo, Noah J.; Gearhart, Micah D.; Bardwell, Vivian J.; Jenkins, Marc K.

    2015-01-01

    CD4+ germinal center (GC) T follicular helper (GC-Tfh) cells help B cells become long-lived plasma cells and memory cells. The transcriptional repressor BCL6 plays a key role in GC-Tfh formation by inhibiting the expression of genes that promote differentiation into other lineages. We determined whether BCOR, a component of a Polycomb repressive complex that interacts with the BCL6 BTB domain, influences GC-Tfh differentiation. T cell-targeted BCOR deficiency led to a substantial loss of pept...

  20. An integrated approach for the cell formation and layout design in cellular manufacturing systems

    NARCIS (Netherlands)

    Javadi, Babak; Jolai, Fariborz; Slomp, Jannes; Rabbani, Masoud; Tavakkoli-Moghaddam, Reza

    2013-01-01

    In this paper, a comprehensive model is presented for cell formation and layout design in cellular manufacturing systems (CMS). The proposed model incorporates an extensive coverage of important operational features and especially layout design aspects to determine optimal cell configuration and Int

  1. Study of budding yeast colony formation and its characterizations by using circular granular cell

    Science.gov (United States)

    Aprianti, D.; Haryanto, F.; Purqon, A.; Khotimah, S. N.; Viridi, S.

    2016-03-01

    Budding yeast can exhibit colony formation in solid substrate. The colony of pathogenic budding yeast can colonize various surfaces of the human body and medical devices. Furthermore, it can form biofilm that resists drug effective therapy. The formation of the colony is affected by the interaction between cells and with its growth media. The cell budding pattern holds an important role in colony expansion. To study this colony growth, the molecular dynamic method was chosen to simulate the interaction between budding yeast cells. Every cell was modelled by circular granular cells, which can grow and produce buds. Cohesion force, contact force, and Stokes force govern this model to mimic the interaction between cells and with the growth substrate. Characterization was determined by the maximum (L max) and minimum (L min) distances between two cells within the colony and whether two lines that connect the two cells in the maximum and minimum distances intersect each other. Therefore, it can be recognized the colony shape in circular, oval, and irregular shapes. Simulation resulted that colony formation are mostly in oval shape with little branch. It also shows that greater cohesion strength obtains more compact colony formation.

  2. Arabidopsis R-SNARE proteins VAMP721 and VAMP722 are required for cell plate formation.

    Directory of Open Access Journals (Sweden)

    Liang Zhang

    Full Text Available BACKGROUND: Cell plate formation during plant cytokinesis is facilitated by SNARE complex-mediated vesicle fusion at the cell-division plane. However, our knowledge regarding R-SNARE components of membrane fusion machinery for cell plate formation remains quite limited. METHODOLOGY/PRINCIPAL FINDINGS: We report the in vivo function of Arabidopsis VAMP721 and VAMP722, two closely sequence-related R-SNAREs, in cell plate formation. Double homozygous vamp721vamp722 mutant seedlings showed lethal dwarf phenotypes and were characterized by rudimentary roots, cotyledons and hypocotyls. Furthermore, cell wall stubs and incomplete cytokinesis were frequently observed in vamp721vamp722 seedlings. Confocal images revealed that green fluorescent protein-tagged VAMP721 and VAMP722 were preferentially localized to the expanding cell plates in dividing cells. Drug treatments and co-localization analyses demonstrated that punctuate organelles labeled with VAMP721 and VAMP722 represented early endosomes overlapped with VHA-a1-labeled TGN, which were distinct from Golgi stacks and prevacuolar compartments. In addition, protein traffic to the plasma membrane, but not to the vacuole, was severely disrupted in vamp721vamp722 seedlings by subcellular localization of marker proteins. CONCLUSION/SIGNIFICANCE: These observations suggest that VAMP721 and VAMP722 are involved in secretory trafficking to the plasma membrane via TGN/early endosomal compartment, which contributes substantially to cell plate formation during plant cytokinesis.

  3. Methods for studying biofilm formation: flow cells and confocal laser scanning microscopy

    DEFF Research Database (Denmark)

    Tolker-Nielsen, Tim; Sternberg, Claus

    2014-01-01

    In this chapter methods for growing and analyzing biofilms under hydrodynamic conditions in flow cells are described. Use of flow cells allows for direct microscopic investigation of biofilm formation. The flow in these chambers is essentially laminar, which means that the biofilms can be grown u...

  4. NRP1 Regulates CDC42 Activation to Promote Filopodia Formation in Endothelial Tip Cells

    Directory of Open Access Journals (Sweden)

    Alessandro Fantin

    2015-06-01

    Full Text Available Sprouting blood vessels are led by filopodia-studded endothelial tip cells that respond to angiogenic signals. Mosaic lineage tracing previously revealed that NRP1 is essential for tip cell function, although its mechanistic role in tip cells remains poorly defined. Here, we show that NRP1 is dispensable for genetic tip cell identity. Instead, we find that NRP1 is essential to form the filopodial bursts that distinguish tip cells morphologically from neighboring stalk cells, because it enables the extracellular matrix (ECM-induced activation of CDC42, a key regulator of filopodia formation. Accordingly, NRP1 knockdown and pharmacological CDC42 inhibition similarly impaired filopodia formation in vitro and in developing zebrafish in vivo. During mouse retinal angiogenesis, CDC42 inhibition impaired tip cell and vascular network formation, causing defects that resembled those due to loss of ECM-induced, but not VEGF-induced, NRP1 signaling. We conclude that NRP1 enables ECM-induced filopodia formation for tip cell function during sprouting angiogenesis.

  5. Lipid body formation during maturation of human mast cells.

    Science.gov (United States)

    Dichlberger, Andrea; Schlager, Stefanie; Lappalainen, Jani; Käkelä, Reijo; Hattula, Katarina; Butcher, Sarah J; Schneider, Wolfgang J; Kovanen, Petri T

    2011-12-01

    Lipid droplets, also called lipid bodies (LB) in inflammatory cells, are important cytoplasmic organelles. However, little is known about the molecular characteristics and functions of LBs in human mast cells (MC). Here, we have analyzed the genesis and components of LBs during differentiation of human peripheral blood-derived CD34(+) progenitors into connective tissue-type MCs. In our serum-free culture system, the maturing MCs, derived from 18 different donors, invariably developed triacylglycerol (TG)-rich LBs. Not known heretofore, the MCs transcribe the genes for perilipins (PLIN)1-4, but not PLIN5, and PLIN2 and PLIN3 display different degrees of LB association. Upon MC activation and ensuing degranulation, the LBs were not cosecreted with the cytoplasmic secretory granules. Exogenous arachidonic acid (AA) enhanced LB genesis in Triacsin C-sensitive fashion, and it was found to be preferentially incorporated into the TGs of LBs. The large TG-associated pool of AA in LBs likely is a major precursor for eicosanoid production by MCs. In summary, we demonstrate that cultured human MCs derived from CD34(+) progenitors in peripheral blood provide a new tool to study regulatory mechanisms involving LB functions, with particular emphasis on AA metabolism, eicosanoid biosynthesis, and subsequent release of proinflammatory lipid mediators from these cells.

  6. Mxi1 influences cyst formation in three-dimensional cell culture

    Directory of Open Access Journals (Sweden)

    Yeon Joo Yook

    2012-03-01

    Full Text Available Cyst formation is a major characteristic of ADPKD and iscaused by the abnormal proliferation of epithelial cells. Renalcyst formation disrupts renal function and induces diversecomplications. The mechanism of cyst formation is unclear.mIMCD-3 cells were established to develop simple epithelialcell cysts in 3-D culture. We confirmed previously that Mxi1plays a role in cyst formation in Mxi1-deficient mice. Cysts inMxi1 transfectanted cells were showed by collagen or mebiolgels in 3-D cell culture system. Causative genes of ADPKDwere measured by q RT-PCR. Herein, Mxi1 transfectants rarelyformed a simple epithelial cyst and induced cell death.Overexpression of Mxi1 resulted in a decrease in the PKD1,PKD2 and c-myc mRNA relating to the pathway of cystformation. These data indicate that Mxi1 influences cystformation of mIMCD-3 cells in 3-D culture and that Mxi1 maycontrol the mechanism of renal cyst formation. [BMB reports2012; 45(3: 189-193

  7. Oxygen-consuming chlor alkali cell configured to minimize peroxide formation

    Science.gov (United States)

    Chlistunoff, Jerzy B.; Lipp, Ludwig; Gottesfeld, Shimshon

    2006-08-01

    Oxygen-consuming zero gap chlor-alkali cell was configured to minimize peroxide formation. The cell included an ion-exchange membrane that divided the cell into an anode chamber including an anode and a cathode chamber including an oxygen gas diffusion cathode. The cathode included a single-piece of electrically conducting graphitized carbon cloth. Catalyst and polytetrafluoroethylene were attached to only one side of the cloth. When the cathode was positioned against the cation exchange membrane with the catalyst side away from the membrane, electrolysis of sodium chloride to chlorine and caustic (sodium hydroxide) proceeded with minimal peroxide formation.

  8. Influence of porous silicon formation on the performance of multi-crystalline silicon solar cells

    Indian Academy of Sciences (India)

    M Saad; M Naddaf

    2015-06-01

    The effect of formation of porous silicon on the performance of multi-crystalline silicon (mc-Si) solar cells is presented. Surface treatment of mc-Si solar cells was performed by electrochemical etching in HF-based solution. The effect of etching is viewed through scanning electron microscope (SEM) photographs that indicated the formation of a porous layer on the surface. Total reflection spectroscopy measurements on solar cells revealed reduced reflection after etching. In order to demonstrate the effect of this porous layer on the solar cell performance, illumination-dependent – characteristics and spectral response measurements were performed and analysed before and after etching. At all illumination intensities, short-circuit current density and open-circuit voltage values for the etched solar cell were higher than those before etching, whereas fill factor values were lower for the etched cell at high illumination intensities. An interpretation of these findings is presented.

  9. The in vitro myelin formation in neurospheres of human neural stem cells

    Institute of Scientific and Technical Information of China (English)

    杨立业; 郑佳坤; 刘相名; 惠国桢; 郭礼和

    2003-01-01

    Objective: To explore the culture conditions of human neural stem cells and to investigate the ultrastructure of neurospheres.Methods: The cells from the embryonic human cortices were mechanically dissociated. N2 medium was adapted to culture and expand the cells. The cells were identified by immunocytochemistry and EM was applied to examine the ultrastructure of neurospheres.Results: The neural stem cells from human embryonic brains were successfully cultured and formed typical neurospheres in suspension, and most of the cells expressed vimentin, which was a marker for neural progenitor cells, and the cells could differentiate into neurons, astrocytes and oligodendrocytes. In vitro myelin formation in neurospheres were observed at an early stage of culture.Conclusions: Human neural stem cells can be cultured from embryonic brains, can form the typical neurospheres in suspension in vitro and have the ability of myelinating, and may be potential source for transplantation in treating myelin disorders.

  10. Practical strategies for modulating foam cell formation and behavior.

    Science.gov (United States)

    Uitz, Elisabeth; Bahadori, Babak; McCarty, Mark F; Moghadasian, Mohammed H

    2014-10-16

    Although high density lipoprotein (HDL)-mediated reverse cholesterol transport is crucial to the prevention and reversal of atheroma, a recent meta-analysis makes evident that current pharmaceutical strategies for modulating HDL cholesterol levels lower cardiovascular risk only to the extent that they concurrently decrease low density lipoprotein (LDL) cholesterol. This corresponds well with findings of a recent Mendelian randomization analysis, in which genetic polymorphisms associated with HDL cholesterol but no other known cardiovascular risk factors failed to predict risk for myocardial infarction. Although it is still seems appropriate to search for therapies that could improve the efficiency with which HDL particles induce reverse cholesterol transport, targeting HDL cholesterol levels per se with current measures appears to be futile. It may therefore be more promising to promote reverse cholesterol transport with agents that directly target foam cells. Macrophage expression of the cholesterol transport proteins adenosine triphosphate binding cassette transporter A1, adenosine triphosphate binding cassette transporter G1, and scavenger receptor class B member 1 is transcriptionally up-regulated by activated liver X receptors (LXR), whereas nuclear factor (NF)-kappaB antagonizes their expression. Taurine, which inhibits atherogenesis in rodent studies, has just been discovered to act as a weak agonist for LXRalpha. Conversely, it may be possible to oppose NF-kappaB activation in macrophages with a range of measures. Induction of heme oxygenase-1, which can be attained with phase 2 inducer phytochemicals such as lipoic acid and green tea catechins, promotes reverse cholesterol transport in macrophages and inhibits atherogenesis in rodents, likely due to, in large part, NF-kappaB antagonism. Inhibition of macrophage nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity with the spirulina-derived bilirubin-mimetic phycocyanobilin may also oppose

  11. Quantitative comparison between microfluidic and microtiter plate formats for cell-based assays.

    Science.gov (United States)

    Yin, Huabing; Pattrick, Nicola; Zhang, Xunli; Klauke, Norbert; Cordingley, Hayley C; Haswell, Steven J; Cooper, Jonathan M

    2008-01-01

    In this paper, we compare a quantitative cell-based assay measuring the intracellular Ca2+ response to the agonist uridine 5'-triphosphate in Chinese hamster ovary cells, in both microfluidic and microtiter formats. The study demonstrates that, under appropriate hydrodynamic conditions, there is an excellent agreement between traditional well-plate assays and those obtained on-chip for both suspended immobilized cells and cultured adherent cells. We also demonstrate that the on-chip assay, using adherent cells, provides the possibility of faster screening protocols with the potential for resolving subcellular information about local Ca2+ flux.

  12. Identification of separate slow and fast muscle precursor cells in vivo, prior to somite formation.

    Science.gov (United States)

    Devoto, S H; Melançon, E; Eisen, J S; Westerfield, M

    1996-11-01

    We have examined the development of specific muscle fiber types in zebrafish axial muscle by labeling myogenic precursor cells with vital fluorescent dyes and following their subsequent differentiation and fate. Two populations of muscle precursors, medial and lateral, can be distinguished in the segmental plate by position, morphology and gene expression. The medial cells, known as adaxial cells, are large, cuboidal cells adjacent to the notochord that express myoD. Surprisingly, after somite formation, they migrate radially away from the notochord, becoming a superficial layer of muscle cells. A subset of adaxial cells develop into engrailed-expressing muscle pioneers. Adaxial cells differentiate into slow muscle fibers of the adult fish. We have named the lateral population of cells in the segmental plate, lateral presomitic cells. They are smaller, more irregularly shaped and separated from the notochord by adaxial cells; they do not express myoD until after somite formation. Lateral presomitic cells remain deep in the myotome and they differentiate into fast muscle fibers. Thus, slow and fast muscle fiber types in zebrafish axial muscle arise from distinct populations of cells in the segmental plate that develop in different cellular environments and display distinct behaviors. PMID:8951054

  13. The MP65 gene is required for cell wall integrity, adherence to epithelial cells and biofilm formation in Candida albicans

    Directory of Open Access Journals (Sweden)

    Girolamo Antonietta

    2011-05-01

    Full Text Available Abstract Background The MP65 gene of Candida albicans (orf19.1779 encodes a putative β-glucanase mannoprotein of 65 kDa, which plays a main role in a host-fungus relationship, morphogenesis and pathogenicity. In this study, we performed an extensive analysis of a mp65Δ mutant to assess the role of this protein in cell wall integrity, adherence to epithelial cells and biofilm formation. Results The mp65Δ mutant showed a high sensitivity to a range of cell wall-perturbing and degrading agents, especially Congo red, which induced morphological changes such as swelling, clumping and formation of hyphae. The mp65Δ mutant showed an activation of two MAPKs (Mkc1p and Cek1p, a high level of expression of two stress-related genes (DDR48 and SOD5, and a modulated expression of β-glucan epitopes, but no gross changes in cell wall polysaccharide composition. Interestingly, the mp65Δ mutant displayed a marked reduction in adhesion to BEC and Caco-2 cells and severe defects in biofilm formation when compared to the wild type. All of the mentioned properties were totally or partially recovered in a revertant strain, demonstrating the specificity of gene deletion. Conclusions We demonstrate that the MP65 gene of Candida albicans plays a significant role in maintaining cell wall integrity, as well as in adherence to epithelia and biofilm formation, which are major virulence attributes of this fungus.

  14. Collective motion of cells mediates segregation and pattern formation in co-cultures.

    Directory of Open Access Journals (Sweden)

    Elod Méhes

    Full Text Available Pattern formation by segregation of cell types is an important process during embryonic development. We show that an experimentally yet unexplored mechanism based on collective motility of segregating cells enhances the effects of known pattern formation mechanisms such as differential adhesion, mechanochemical interactions or cell migration directed by morphogens. To study in vitro cell segregation we use time-lapse videomicroscopy and quantitative analysis of the main features of the motion of individual cells or groups. Our observations have been extensive, typically involving the investigation of the development of patterns containing up to 200,000 cells. By either comparing keratocyte types with different collective motility characteristics or increasing cells' directional persistence by the inhibition of Rac1 GTP-ase we demonstrate that enhanced collective cell motility results in faster cell segregation leading to the formation of more extensive patterns. The growth of the characteristic scale of patterns generally follows an algebraic scaling law with exponent values up to 0.74 in the presence of collective motion, compared to significantly smaller exponents in case of diffusive motion.

  15. QHREDGS enhances tube formation, metabolism and survival of endothelial cells in collagen-chitosan hydrogels.

    Directory of Open Access Journals (Sweden)

    Jason W Miklas

    Full Text Available Cell survival in complex, vascularized tissues, has been implicated as a major bottleneck in advancement of therapies based on cardiac tissue engineering. This limitation motivates the search for small, inexpensive molecules that would simultaneously be cardio-protective and vasculogenic. Here, we present peptide sequence QHREDGS, based upon the fibrinogen-like domain of angiopoietin-1, as a prime candidate molecule. We demonstrated previously that QHREDGS improved cardiomyocyte metabolism and mitigated serum starved apoptosis. In this paper we further demonstrate the potency of QHREDGS in its ability to enhance endothelial cell survival, metabolism and tube formation. When endothelial cells were exposed to the soluble form of QHREDGS, improvements in endothelial cell barrier functionality, nitric oxide production and cell metabolism (ATP levels in serum starved conditions were found. The functionality of the peptide was then examined when conjugated to collagen-chitosan hydrogel, a potential carrier for in vivo application. The presence of the peptide in the hydrogel mitigated paclitaxel induced apoptosis of endothelial cells in a dose dependent manner. Furthermore, the peptide modified hydrogels stimulated tube-like structure formation of encapsulated endothelial cells. When integrin αvβ3 or α5β1 were antibody blocked during cell encapsulation in peptide modified hydrogels, tube formation was abolished. Therefore, the dual protective nature of the novel peptide QHREDGS may position this peptide as an appealing augmentation for collagen-chitosan hydrogels that could be used for biomaterial delivered cell therapies in the settings of myocardial infarction.

  16. QHREDGS enhances tube formation, metabolism and survival of endothelial cells in collagen-chitosan hydrogels.

    Science.gov (United States)

    Miklas, Jason W; Dallabrida, Susan M; Reis, Lewis A; Ismail, Nesreen; Rupnick, Maria; Radisic, Milica

    2013-01-01

    Cell survival in complex, vascularized tissues, has been implicated as a major bottleneck in advancement of therapies based on cardiac tissue engineering. This limitation motivates the search for small, inexpensive molecules that would simultaneously be cardio-protective and vasculogenic. Here, we present peptide sequence QHREDGS, based upon the fibrinogen-like domain of angiopoietin-1, as a prime candidate molecule. We demonstrated previously that QHREDGS improved cardiomyocyte metabolism and mitigated serum starved apoptosis. In this paper we further demonstrate the potency of QHREDGS in its ability to enhance endothelial cell survival, metabolism and tube formation. When endothelial cells were exposed to the soluble form of QHREDGS, improvements in endothelial cell barrier functionality, nitric oxide production and cell metabolism (ATP levels) in serum starved conditions were found. The functionality of the peptide was then examined when conjugated to collagen-chitosan hydrogel, a potential carrier for in vivo application. The presence of the peptide in the hydrogel mitigated paclitaxel induced apoptosis of endothelial cells in a dose dependent manner. Furthermore, the peptide modified hydrogels stimulated tube-like structure formation of encapsulated endothelial cells. When integrin αvβ3 or α5β1 were antibody blocked during cell encapsulation in peptide modified hydrogels, tube formation was abolished. Therefore, the dual protective nature of the novel peptide QHREDGS may position this peptide as an appealing augmentation for collagen-chitosan hydrogels that could be used for biomaterial delivered cell therapies in the settings of myocardial infarction. PMID:24013716

  17. Artificial induction of Sox21 regulates sensory cell formation in the embryonic chicken inner ear.

    Directory of Open Access Journals (Sweden)

    Stephen D Freeman

    Full Text Available During embryonic development, hair cells and support cells in the sensory epithelia of the inner ear derive from progenitors that express Sox2, a member of the SoxB1 family of transcription factors. Sox2 is essential for sensory specification, but high levels of Sox2 expression appear to inhibit hair cell differentiation, suggesting that factors regulating Sox2 activity could be critical for both processes. Antagonistic interactions between SoxB1 and SoxB2 factors are known to regulate cell differentiation in neural tissue, which led us to investigate the potential roles of the SoxB2 member Sox21 during chicken inner ear development. Sox21 is normally expressed by sensory progenitors within vestibular and auditory regions of the early embryonic chicken inner ear. At later stages, Sox21 is differentially expressed in the vestibular and auditory organs. Sox21 is restricted to the support cell layer of the auditory epithelium, while it is enriched in the hair cell layer of the vestibular organs. To test Sox21 function, we used two temporally distinct gain-of-function approaches. Sustained over-expression of Sox21 from early developmental stages prevented prosensory specification, and abolished the formation of both hair cells and support cells. However, later induction of Sox21 expression at the time of hair cell formation in organotypic cultures of vestibular epithelia inhibited endogenous Sox2 expression and Notch activity, and biased progenitor cells towards a hair cell fate. Interestingly, Sox21 did not promote hair cell differentiation in the immature auditory epithelium, which fits with the expression of endogenous Sox21 within mature support cells in this tissue. These results suggest that interactions among endogenous SoxB family transcription factors may regulate sensory cell formation in the inner ear, but in a context-dependent manner.

  18. Pectinous cell wall thickenings formation - A common defense strategy of plants to cope with Pb.

    Science.gov (United States)

    Krzesłowska, Magdalena; Rabęda, Irena; Basińska, Aneta; Lewandowski, Michał; Mellerowicz, Ewa J; Napieralska, Anna; Samardakiewicz, Sławomir; Woźny, Adam

    2016-07-01

    Lead, one of the most abundant and hazardous trace metals affecting living organisms, has been commonly detected in plant cell walls including some tolerant plants, mining ecotypes and hyperaccumulators. We have previously shown that in tip growing Funaria sp. protonemata cell wall is remodeled in response to lead by formation of thickenings rich in low-methylesterified pectins (pectin epitope JIM5 - JIM5-P) able to bind metal ions, which accumulate large amounts of Pb. Hence, it leads to the increase of cell wall capacity for Pb compartmentalization. Here we show that diverse plant species belonging to different phyla (Arabidopsis, hybrid aspen, star duckweed), form similar cell wall thickenings in response to Pb. These thickenings are formed in tip growing cells such as the root hairs, and in diffuse growing cells such as meristematic and root cap columella cells of root apices in hybrid aspen and Arabidopsis and in mesophyll cells in star duckweed fronds. Notably, all analyzed cell wall thickenings were abundant in JIM5-P and accumulated high amounts of Pb. In addition, the co-localization of JIM5-P and Pb commonly occurred in these cells. Hence, cell wall thickenings formed the extra compartment for Pb accumulation. In this way plant cells increased cell wall capacity for compartmentalization of this toxic metal, protecting protoplast from its toxicity. As cell wall thickenings occurred in diverse plant species and cell types differing in the type of growth we may conclude that pectinous cell wall thickenings formation is a widespread defense strategy of plants to cope with Pb. Moreover, detection of natural defense strategy, increasing plant cell walls capacity for metal accumulation, reveals a promising direction for enhancing plant efficiency in phytoremediation. PMID:27107260

  19. Pectinous cell wall thickenings formation - A common defense strategy of plants to cope with Pb.

    Science.gov (United States)

    Krzesłowska, Magdalena; Rabęda, Irena; Basińska, Aneta; Lewandowski, Michał; Mellerowicz, Ewa J; Napieralska, Anna; Samardakiewicz, Sławomir; Woźny, Adam

    2016-07-01

    Lead, one of the most abundant and hazardous trace metals affecting living organisms, has been commonly detected in plant cell walls including some tolerant plants, mining ecotypes and hyperaccumulators. We have previously shown that in tip growing Funaria sp. protonemata cell wall is remodeled in response to lead by formation of thickenings rich in low-methylesterified pectins (pectin epitope JIM5 - JIM5-P) able to bind metal ions, which accumulate large amounts of Pb. Hence, it leads to the increase of cell wall capacity for Pb compartmentalization. Here we show that diverse plant species belonging to different phyla (Arabidopsis, hybrid aspen, star duckweed), form similar cell wall thickenings in response to Pb. These thickenings are formed in tip growing cells such as the root hairs, and in diffuse growing cells such as meristematic and root cap columella cells of root apices in hybrid aspen and Arabidopsis and in mesophyll cells in star duckweed fronds. Notably, all analyzed cell wall thickenings were abundant in JIM5-P and accumulated high amounts of Pb. In addition, the co-localization of JIM5-P and Pb commonly occurred in these cells. Hence, cell wall thickenings formed the extra compartment for Pb accumulation. In this way plant cells increased cell wall capacity for compartmentalization of this toxic metal, protecting protoplast from its toxicity. As cell wall thickenings occurred in diverse plant species and cell types differing in the type of growth we may conclude that pectinous cell wall thickenings formation is a widespread defense strategy of plants to cope with Pb. Moreover, detection of natural defense strategy, increasing plant cell walls capacity for metal accumulation, reveals a promising direction for enhancing plant efficiency in phytoremediation.

  20. Bmi1 overexpression in the cerebellar granule cell lineage of mice affects cell proliferation and survival without initiating medulloblastoma formation

    Directory of Open Access Journals (Sweden)

    Hourinaz Behesti

    2013-01-01

    BMI1 is a potent inducer of neural stem cell self-renewal and neural progenitor cell proliferation during development and in adult tissue homeostasis. It is overexpressed in numerous human cancers – including medulloblastomas, in which its functional role is unclear. We generated transgenic mouse lines with targeted overexpression of Bmi1 in the cerebellar granule cell lineage, a cell type that has been shown to act as a cell of origin for medulloblastomas. Overexpression of Bmi1 in granule cell progenitors (GCPs led to a decrease in cerebellar size due to decreased GCP proliferation and repression of the expression of cyclin genes, whereas Bmi1 overexpression in postmitotic granule cells improved cell survival in response to stress by altering the expression of genes in the mitochondrial cell death pathway and of Myc and Lef-1. Although no medulloblastomas developed in ageing cohorts of transgenic mice, crosses with Trp53−/− mice resulted in a low incidence of medulloblastoma formation. Furthermore, analysis of a large collection of primary human medulloblastomas revealed that tumours with a BMI1high TP53low molecular profile are significantly enriched in Group 4 human medulloblastomas. Our data suggest that different levels and timing of Bmi1 overexpression yield distinct cellular outcomes within the same cellular lineage. Importantly, Bmi1 overexpression at the GCP stage does not induce tumour formation, suggesting that BMI1 overexpression in GCP-derived human medulloblastomas probably occurs during later stages of oncogenesis and might serve to enhance tumour cell survival.

  1. Different Effects of BORIS/CTCFL on Stemness Gene Expression, Sphere Formation and Cell Survival in Epithelial Cancer Stem Cells.

    Directory of Open Access Journals (Sweden)

    Loredana Alberti

    Full Text Available Cancer stem cells are cancer cells characterized by stem cell properties and represent a small population of tumor cells that drives tumor development, progression, metastasis and drug resistance. To date, the molecular mechanisms that generate and regulate cancer stem cells are not well defined. BORIS (Brother of Regulator of Imprinted Sites or CTCFL (CTCF-like is a DNA-binding protein that is expressed in normal tissues only in germ cells and is re-activated in tumors. Recent evidences have highlighted the correlation of BORIS/CTCFL expression with poor overall survival of different cancer patients. We have previously shown an association of BORIS-expressing cells with stemness gene expression in embryonic cancer cells. Here, we studied the role of BORIS in epithelial tumor cells. Using BORIS-molecular beacon that was already validated, we were able to show the presence of BORIS mRNA in cancer stem cell-enriched populations (side population and spheres of cervical, colon and breast tumor cells. BORIS silencing studies showed a decrease of sphere formation capacity in breast and colon tumor cells. Importantly, BORIS-silencing led to down-regulation of hTERT, stem cell (NANOG, OCT4, SOX2 and BMI1 and cancer stem cell markers (ABCG2, CD44 and ALDH1 genes. Conversely, BORIS-induction led to up-regulation of the same genes. These phenotypes were observed in cervical, colon and invasive breast tumor cells. However, a completely different behavior was observed in the non-invasive breast tumor cells (MCF7. Indeed, these cells acquired an epithelial mesenchymal transition phenotype after BORIS silencing. Our results demonstrate that BORIS is associated with cancer stem cell-enriched populations of several epithelial tumor cells and the different phenotypes depend on the origin of tumor cells.

  2. Monocytes from HIV+ individuals show impaired cholesterol efflux and increased foam cell formation after transendothelial migration

    Science.gov (United States)

    MAISA, Anna; HEARPS, Anna C.; ANGELOVICH, Thomas A.; PEREIRA, Candida F.; ZHOU, Jingling; SHI, Margaret D.Y.; PALMER, Clovis S.; MULLER, William A.; CROWE, Suzanne M.; JAWOROWSKI, Anthony

    2016-01-01

    Design HIV+ individuals have an increased risk of atherosclerosis and cardiovascular disease which is independent of antiretroviral therapy and traditional risk factors. Monocytes play a central role in the development of atherosclerosis, and HIV-related chronic inflammation and monocyte activation may contribute to increased atherosclerosis, but the mechanisms are unknown. Methods Using an in vitro model of atherosclerotic plaque formation, we measured the transendothelial migration of purified monocytes from age-matched HIV+ and uninfected donors and examined their differentiation into foam cells. Cholesterol efflux and the expression of cholesterol metabolism genes were also assessed. Results Monocytes from HIV+ individuals showed increased foam cell formation compared to controls (18.9% vs 0% respectively, p=0.004) and serum from virologically suppressed HIV+ individuals potentiated foam cell formation by monocytes from both uninfected and HIV+ donors. Plasma TNF levels were increased in HIV+ vs control donors (5.9 vs 3.5 pg/ml, p=0.02) and foam cell formation was inhibited by blocking antibodies to TNF receptors, suggesting a direct effect on monocyte differentiation to foam cells. Monocytes from virologically suppressed HIV+ donors showed impaired cholesterol efflux and decreased expression of key genes regulating cholesterol metabolism, including the cholesterol transporter ABCA1 (p=0.02). Conclusions Monocytes from HIV+ individuals show impaired cholesterol efflux and are primed for foam cell formation following trans-endothelial migration. Factors present in HIV+ serum, including elevated TNF levels, further enhance foam cell formation. The pro-atherogenic phenotype of monocytes persists in virologically suppressed HIV+ individuals and may contribute mechanistically to increased atherosclerosis in this population. PMID:26244384

  3. Formation of salivary acinar cell spheroids in vitro above a polyvinyl alcohol-coated surface.

    Science.gov (United States)

    Chen, Min-Huey; Chen, Yi-Jane; Liao, Chih-Chen; Chan, Yen-Hui; Lin, Chia-Yung; Chen, Rung-Shu; Young, Tai-Hong

    2009-09-15

    Tissue engineering of salivary glands offers the potential for future use in the treatment of patients with salivary hypofunction. Biocompatible materials that promote acinar cell aggregation and function in vitro are an essential part of salivary gland tissue engineering. In this study, rat parotid acinar cells assembled into three-dimensional aggregates above the polyvinyl alcohol (PVA)-coated surface. These aggregates developed compact acinar cell spheroids resembling in vivo physiological condition, which were different from the traditional monolayered morphology in vitro. Cells remained viable and with better functional activity in response to acetylcholine in the spheroids and could form monolayered acinar cells when they were reinoculated on tissue culture polystyrene wells. To interpret the phenomenon further, we proposed that the formation of acinar cell spheroids on the PVA is mediated by a balance between two competing forces: the interactions of cell-PVA and cell-cell. This study demonstrated the formation of functional cell spheroids above a PVA-coated surface may provide an in vitro system for investigating cell behaviors for tissue engineering of artificial salivary gland.

  4. The adaptor molecule SAP plays essential roles during invariant NKT cell cytotoxicity and lytic synapse formation.

    Science.gov (United States)

    Das, Rupali; Bassiri, Hamid; Guan, Peng; Wiener, Susan; Banerjee, Pinaki P; Zhong, Ming-Chao; Veillette, André; Orange, Jordan S; Nichols, Kim E

    2013-04-25

    The adaptor molecule signaling lymphocytic activation molecule-associated protein (SAP) plays critical roles during invariant natural killer T (iNKT) cell ontogeny. As a result, SAP-deficient humans and mice lack iNKT cells. The strict developmental requirement for SAP has made it difficult to discern its possible involvement in mature iNKT cell functions. By using temporal Cre recombinase-mediated gene deletion to ablate SAP expression after completion of iNKT cell development, we demonstrate that SAP is essential for T-cell receptor (TCR)-induced iNKT cell cytotoxicity against T-cell and B-cell leukemia targets in vitro and iNKT-cell-mediated control of T-cell leukemia growth in vivo. These findings are not restricted to the murine system: silencing RNA-mediated suppression of SAP expression in human iNKT cells also significantly impairs TCR-induced cytolysis. Mechanistic studies reveal that iNKT cell killing requires the tyrosine kinase Fyn, a known SAP-binding protein. Furthermore, SAP expression is required within iNKT cells to facilitate their interaction with T-cell targets and induce reorientation of the microtubule-organizing center to the immunologic synapse (IS). Collectively, these studies highlight a novel and essential role for SAP during iNKT cell cytotoxicity and formation of a functional IS.

  5. An improved model for nucleation-limited ice formation in living cells during freezing.

    Directory of Open Access Journals (Sweden)

    Jingru Yi

    Full Text Available Ice formation in living cells is a lethal event during freezing and its characterization is important to the development of optimal protocols for not only cryopreservation but also cryotherapy applications. Although the model for probability of ice formation (PIF in cells developed by Toner et al. has been widely used to predict nucleation-limited intracellular ice formation (IIF, our data of freezing Hela cells suggest that this model could give misleading prediction of PIF when the maximum PIF in cells during freezing is less than 1 (PIF ranges from 0 to 1. We introduce a new model to overcome this problem by incorporating a critical cell volume to modify the Toner's original model. We further reveal that this critical cell volume is dependent on the mechanisms of ice nucleation in cells during freezing, i.e., surface-catalyzed nucleation (SCN and volume-catalyzed nucleation (VCN. Taken together, the improved PIF model may be valuable for better understanding of the mechanisms of ice nucleation in cells during freezing and more accurate prediction of PIF for cryopreservation and cryotherapy applications.

  6. An improved model for nucleation-limited ice formation in living cells during freezing.

    Science.gov (United States)

    Yi, Jingru; Liang, Xin M; Zhao, Gang; He, Xiaoming

    2014-01-01

    Ice formation in living cells is a lethal event during freezing and its characterization is important to the development of optimal protocols for not only cryopreservation but also cryotherapy applications. Although the model for probability of ice formation (PIF) in cells developed by Toner et al. has been widely used to predict nucleation-limited intracellular ice formation (IIF), our data of freezing Hela cells suggest that this model could give misleading prediction of PIF when the maximum PIF in cells during freezing is less than 1 (PIF ranges from 0 to 1). We introduce a new model to overcome this problem by incorporating a critical cell volume to modify the Toner's original model. We further reveal that this critical cell volume is dependent on the mechanisms of ice nucleation in cells during freezing, i.e., surface-catalyzed nucleation (SCN) and volume-catalyzed nucleation (VCN). Taken together, the improved PIF model may be valuable for better understanding of the mechanisms of ice nucleation in cells during freezing and more accurate prediction of PIF for cryopreservation and cryotherapy applications. PMID:24852166

  7. Promotion of experimental thrombus formation by the procoagulant activity of breast cancer cells

    International Nuclear Information System (INIS)

    The routine observation of tumor emboli in the peripheral blood of patients with carcinomas raises questions about the clinical relevance of these circulating tumor cells. Thrombosis is a common clinical manifestation of cancer, and circulating tumor cells may play a pathogenetic role in this process. The presence of coagulation-associated molecules on cancer cells has been described, but the mechanisms by which circulating tumor cells augment or alter coagulation remains unclear. In this study we utilized suspensions of a metastatic adenocarcinoma cell line, MDA-MB-231, and a non-metastatic breast epithelial cell line, MCF-10A, as models of circulating tumor cells to determine the thromobogenic activity of these blood-foreign cells. In human plasma, both metastatic MDA-MB-231 cells and non-metastatic MCF-10A cells significantly enhanced clotting kinetics. The effect of MDA-MB-231 and MCF-10A cells on clotting times was cell number-dependent and inhibited by a neutralizing antibody to tissue factor (TF) as well as inhibitors of activated factor X and thrombin. Using fluorescence microscopy, we found that both MDA-MB-231 and MCF-10A cells supported the binding of fluorescently labeled thrombin. Furthermore, in a model of thrombus formation under pressure-driven flow, MDA-MB-231 and MCF-10A cells significantly decreased the time to occlusion. Our findings indicate that the presence of breast epithelial cells in blood can stimulate coagulation in a TF-dependent manner, suggesting that tumor cells that enter the circulation may promote the formation of occlusive thrombi under shear flow conditions

  8. Promotion of experimental thrombus formation by the procoagulant activity of breast cancer cells

    Science.gov (United States)

    Berny-Lang, M. A.; Aslan, J. E.; Tormoen, G. W.; Patel, I. A.; Bock, P. E.; Gruber, A.; McCarty, O. J. T.

    2011-02-01

    The routine observation of tumor emboli in the peripheral blood of patients with carcinomas raises questions about the clinical relevance of these circulating tumor cells. Thrombosis is a common clinical manifestation of cancer, and circulating tumor cells may play a pathogenetic role in this process. The presence of coagulation-associated molecules on cancer cells has been described, but the mechanisms by which circulating tumor cells augment or alter coagulation remains unclear. In this study we utilized suspensions of a metastatic adenocarcinoma cell line, MDA-MB-231, and a non-metastatic breast epithelial cell line, MCF-10A, as models of circulating tumor cells to determine the thromobogenic activity of these blood-foreign cells. In human plasma, both metastatic MDA-MB-231 cells and non-metastatic MCF-10A cells significantly enhanced clotting kinetics. The effect of MDA-MB-231 and MCF-10A cells on clotting times was cell number-dependent and inhibited by a neutralizing antibody to tissue factor (TF) as well as inhibitors of activated factor X and thrombin. Using fluorescence microscopy, we found that both MDA-MB-231 and MCF-10A cells supported the binding of fluorescently labeled thrombin. Furthermore, in a model of thrombus formation under pressure-driven flow, MDA-MB-231 and MCF-10A cells significantly decreased the time to occlusion. Our findings indicate that the presence of breast epithelial cells in blood can stimulate coagulation in a TF-dependent manner, suggesting that tumor cells that enter the circulation may promote the formation of occlusive thrombi under shear flow conditions.

  9. The formation mechanism for printed silver-contacts for silicon solar cells

    OpenAIRE

    Fields, Jeremy D; Ahmad, Md. Imteyaz; Pool, Vanessa L.; Yu, Jiafan; Van Campen, Douglas G.; Parilla, Philip A.; Toney, Michael F; van Hest, Maikel F. A. M.

    2016-01-01

    Screen-printing provides an economically attractive means for making Ag electrical contacts to Si solar cells, but the use of Ag substantiates a significant manufacturing cost, and the glass frit used in the paste to enable contact formation contains Pb. To achieve optimal electrical performance and to develop pastes with alternative, abundant and non-toxic materials, a better understanding the contact formation process during firing is required. Here, we use in situ X-ray diffraction during ...

  10. Inhibition of gold nanoparticles (AuNPs) on pathogenic biofilm formation and invasion to host cells

    OpenAIRE

    Qilin Yu; Jianrong Li; Yueqi Zhang; Yufan Wang; Lu Liu; Mingchun Li

    2016-01-01

    Owing to the growing infectious diseases caused by eukaryotic and prokaryotic pathogens, it is urgent to develop novel antimicrobial agents against clinical pathogenic infections. Biofilm formation and invasion into the host cells are vital processes during pathogenic colonization and infection. In this study, we tested the inhibitory effect of Au nanoparticles (AuNPs) on pathogenic growth, biofilm formation and invasion. Interestingly, although the synthesized AuNPs had no significant toxici...

  11. Coilin phosphomutants disrupt Cajal body formation, reduce cell proliferation and produce a distinct coilin degradation product.

    Directory of Open Access Journals (Sweden)

    Zunamys I Carrero

    Full Text Available Coilin is a nuclear phosphoprotein that accumulates in Cajal bodies (CBs. CBs participate in ribonucleoprotein and telomerase biogenesis, and are often found in cells with high transcriptional demands such as neuronal and cancer cells, but can also be observed less frequently in other cell types such as fibroblasts. Many proteins enriched within the CB are phosphorylated, but it is not clear what role this modification has on the activity of these proteins in the CB. Coilin is considered to be the CB marker protein and is essential for proper CB formation and composition in mammalian cells. In order to characterize the role of coilin phosphorylation on CB formation, we evaluated various coilin phosphomutants using transient expression. Additionally, we generated inducible coilin phosphomutant cell lines that, when used in combination with endogenous coilin knockdown, allow for the expression of the phosphomutants at physiological levels. Transient expression of all coilin phosphomutants except the phosphonull mutant (OFF significantly reduces proliferation. Interestingly, a stable cell line induced to express the coilin S489D phosphomutant displays nucleolar accumulation of the mutant and generates a N-terminal degradation product; neither of which is observed upon transient expression. A N-terminal degradation product and nucleolar localization are also observed in a stable cell line induced to express a coilin phosphonull mutant (OFF. The nucleolar localization of the S489D and OFF coilin mutants observed in the stable cell lines is decreased when endogenous coilin is reduced. Furthermore, all the phosphomutant cells lines show a significant reduction in CB formation when compared to wild-type after endogenous coilin knockdown. Cell proliferation studies on these lines reveal that only wild-type coilin and the OFF mutant are sufficient to rescue the reduction in proliferation associated with endogenous coilin depletion. These results emphasize

  12. Plant metabolism and cell wall formation in space (microgravity) and on Earth

    Science.gov (United States)

    Lewis, Norman G.

    1994-01-01

    Variations in cell wall chemistry provide vascular plants with the ability to withstand gravitational forces, as well as providing facile mechanisms for correctional responses to various gravitational stimuli, e.g., in reaction wood formation. A principal focus of our current research is to precisely and systematically dissect the essentially unknown mechanism(s) of vascular plant cell wall assembly, particularly with respect to formation of its phenolic constituents, i.e., lignins and suberins, and how gravity impacts upon these processes. Formation of these phenolic polymers is of particular interest, since it appears that elaboration of their biochemical pathways was essential for successful land adaptation. By extrapolation, we are also greatly intrigued as to how the microgravity environment impacts upon 'normal' cell wall assembly mechanisms/metabolism.

  13. Miniature fuel cell with monolithically fabricated Si electrodes - Alloy catalyst formation -

    Science.gov (United States)

    Ogura, Daiki; Suzuki, Takahiro; Katayama, Noboru; Dowaki, Kiyoshi; Hayase, Masanori

    2013-12-01

    A novel Pd-Pt catalyst formation process was proposed for reduction of Pt usage. In our miniature fuel cells, porous Pt was used as the catalyst, and the Pt usage was quite high. To reduce the Pt usage, we have attempted to deposit Pt on porous Pd by galvanic replacement, and relatively large output was demonstrated. In this study, in order to reduce more Pt usage and explore the alloy catalyst formation process, atomic layer deposition by UPD-SLRR (Under Potential Deposition - Surface Limited Redox Replacement) was applied to the Pd-Pt catalyst formation. The new process was verified at each process steps by EDS elemental analysis, and the expected spectra were obtained. Prototype cells were constructed by the new process, and cell output was raised to 420mW/cm2 by the Pd-Pt catalyst from 125mW/cm2 with Pd catalyst.

  14. Inhibition of neurosphere formation in neural stem/progenitor cells by acrylamide.

    Science.gov (United States)

    Chen, Jong-Hang; Lee, Don-Ching; Chen, Mei-Shu; Ko, Ying-Chin; Chiu, Ing-Ming

    2015-01-01

    Previous studies showed that transplantation of cultured neural stem/progenitor cells (NSPCs) could improve functional recovery for various neurological diseases. This study aims to develop a stem cell-based model for predictive toxicology of development in the neurological system after acrylamide exposure. Treatment of mouse (KT98/F1B-GFP) and human (U-1240 MG/F1B-GFP) NSPCs with 0.5 mM acrylamide resulted in the inhibition of neurosphere formation (definition of self-renewal ability in NSPCs), but not inhibition of cell proliferation. Apoptosis and differentiation of KT98 (a precursor of KT98/F1B-GFP) and KT98/F1B-GFP are not observed in acrylamide-treated neurospheres. Analysis of secondary neurosphere formation and differentiation of neurons and glia illustrated that acrylamide-treated KT98 and KT98/F1B-GFP neurospheres retain the NSPC properties, such as self-renewal and differentiation capacity. Correlation of acrylamide-inhibited neurosphere formation with cell-cell adhesion was observed in mouse NSPCs by live cell image analysis and the presence of acrylamide. Protein expression levels of cell adhesion molecules [neural cell adhesion molecule (NCAM) and N-cadherin] and extracellular signal-regulated kinases (ERK) in acrylamide-treated KT98/F1B-GFP and U-1240 MG/F1B-GFP neurospheres demonstrated that NCAM decreased and phospho-ERK (pERK) increased, whereas expression of N-cadherin remained unchanged. Analysis of AKT (protein kinase B, PKB)/β-catenin pathway showed decrease in phospho-AKT (p-AKT) and cyclin D1 expression in acrylamide-treated neurospheres of KT98/F1B-GFP. Furthermore, PD98059, an ERK phosphorylation inhibitor, attenuated acrylamide-induced ERK phosphorylation, indicating that pERK contributed to the cell proliferation, but not in neurosphere formation in mouse NSPCs. Coimmunoprecipitation results of KT98/F1B-GFP cell lysates showed that the complex of NCAM and fibroblast growth factor receptor 1 (FGFR1) is present in the neurosphere, and the

  15. Actinomyces naeslundii GroEL-dependent initial attachment and biofilm formation in a flow cell system.

    Science.gov (United States)

    Arai, Toshiaki; Ochiai, Kuniyasu; Senpuku, Hidenobu

    2015-02-01

    Actinomyces naeslundii is an early colonizer with important roles in the development of the oral biofilm. The effects of butyric acid, one of short chain fatty acids in A. naeslundii biofilm formation was observed using a flow cell system with Tryptic soy broth without dextrose and with 0.25% sucrose (TSB sucrose). Significant biofilms were established involving live and dead cells in TSB sucrose with 60mM butyric acid but not in concentrations of 6, 30, 40, and 50mM. Biofilm formation failed in 60mM sodium butyrate but biofilm level in 60mM sodium butyrate (pH4.7) adjusted with hydrochloric acid as 60mM butyric media (pH4.7) was similar to biofilm levels in 60mM butyric acid. Therefore, butyric acid and low pH are required for significant biofilm formation in the flow cell. To determine the mechanism of biofilm formation, we investigated initial A. naeslundii colonization in various conditions and effects of anti-GroEL antibody. The initial colonization was observed in the 60mM butyric acid condition and anti-GroEL antibody inhibited the initial colonization. In conclusion, we established a new biofilm formation model in which butyric acid induces GroEL-dependent initial colonization of A. naeslundii resulting in significant biofilm formation in a flow system. PMID:25555820

  16. Positioning of polarity formation by extracellular signaling during asymmetric cell division.

    Science.gov (United States)

    Seirin Lee, Sungrim

    2016-07-01

    Anterior-posterior (AP) polarity formation of cell membrane proteins plays a crucial role in determining cell asymmetry, which ultimately generates cell diversity. In Caenorhabditis elegans, a single fertilized egg cell (P0), its daughter cell (P1), and the germline precursors (P2 and P3 cells) form two exclusive domains of different PAR proteins on the membrane along the anterior-posterior axis. However, the phenomenon of polarity reversal has been observed in which the axis of asymmetric cell division of the P2 and P3 cells is formed in an opposite manner to that of the P0 and P1 cells. The extracellular signal MES-1/SRC-1 has been shown to induce polarity reversal, but the detailed mechanism remains elusive. Here, using a mathematical model, I explore the mechanism by which MES-1/SRC-1 signaling can induce polarity reversal and ultimately affect the process of polarity formation. I show that a positive correlation between SRC-1 and the on-rate of PAR-2 is the essential mechanism underlying polarity reversal, providing a mathematical basis for the orientation of cell polarity patterns. PMID:27086039

  17. Estrogen Receptor α Regulates β-Cell Formation During Pancreas Development and Following Injury.

    Science.gov (United States)

    Yuchi, Yixing; Cai, Ying; Legein, Bart; De Groef, Sofie; Leuckx, Gunter; Coppens, Violette; Van Overmeire, Eva; Staels, Willem; De Leu, Nico; Martens, Geert; Van Ginderachter, Jo A; Heimberg, Harry; Van de Casteele, Mark

    2015-09-01

    Identifying pathways for β-cell generation is essential for cell therapy in diabetes. We investigated the potential of 17β-estradiol (E2) and estrogen receptor (ER) signaling for stimulating β-cell generation during embryonic development and in the severely injured adult pancreas. E2 concentration, ER activity, and number of ERα transcripts were enhanced in the pancreas injured by partial duct ligation (PDL) along with nuclear localization of ERα in β-cells. PDL-induced proliferation of β-cells depended on aromatase activity. The activation of Neurogenin3 (Ngn3) gene expression and β-cell growth in PDL pancreas were impaired when ERα was turned off chemically or genetically (ERα(-/-)), whereas in situ delivery of E2 promoted β-cell formation. In the embryonic pancreas, β-cell replication, number of Ngn3(+) progenitor cells, and expression of key transcription factors of the endocrine lineage were decreased by ERα inactivation. The current study reveals that E2 and ERα signaling can drive β-cell replication and formation in mouse pancreas.

  18. Disinfection byproduct formation from chlorination of pure bacterial cells and pipeline biofilms.

    Science.gov (United States)

    Wang, Jun-Jian; Liu, Xin; Ng, Tsz Wai; Xiao, Jie-Wen; Chow, Alex T; Wong, Po Keung

    2013-05-15

    Disinfection byproduct (DBP) formation is commonly attributed to the reaction between natural organic matters and disinfectants, yet few have considered the contribution from disinfecting bacterial materials - the essential process of water disinfection. Here, we explored the DBP formation from chlorination and chloramination of Escherichia coli and found that most selected DBPs were detectable, including trihalomethanes, haloacetonitriles, chloral hydrate, chloropicrin, and 1,1,1-trichloro-2-propanone. A positive correlation (P = 0.08-0.09) between DBP formation and the log reduction of E. coli implied that breaking down of bacterial cells released precursors for DBP formation. As Pseudomonas aeruginosa is a dominant bacterial species in pipeline biofilms, the DBP formation potentials (DBPFPs) from its planktonic cells and biofilms were characterized. Planktonic cells formed 7-11 times greater trihalomethanes per carbon of those from biofilms but significantly lower (P disinfection of bacterial planktonic cells in source water and ex situ reaction between biofilms and residual chlorine in pipeline networks as hitherto unknown DBP sources in drinking water.

  19. Formation of reactive oxygen species in rat epithelial cells upon stimulation with fly ash

    Indian Academy of Sciences (India)

    K Voelkel; H F Krug; S Diabaté

    2003-02-01

    Fly ash was used as a model for ambient particulate matter which is under suspicion to cause adverse pulmonary health effects. The fly ash was pre-sized and contained only particles < 20 m including an ultrafine fraction (< 100 nm) that contributed 31% to the particle number. In our study, we investigated the influence of fly ash on the promotion of early inflammatory reactions like the formation of reactive oxygen species (ROS) in rat lung epithelial cells (RLE-6TN). Furthermore, we determined the formation of nitric oxide (NO). The cells show a clear dose-response relationship concerning the formation of ROS with regard to the mass of particles applied. Lipopolysaccharide (LPS) added as a co-stimulus did not increase the formation of ROS induced by fly ash. Furthermore, in LPS (0.1 g/ml) and tumour necrosis factor-alpha (TNF-alpha; 1 ng/ml) pre-treated cells no increase in reactive oxygen species comparable to fly ash alone is observable. In presence of the metal chelator, desferrioxamine (DFO), ROS formation can be significantly reduced. Neither fly ash nor LPS induced a significant NO release in RLE-6TN cells.

  20. Resveratrol inhibits myeloma cell growth, prevents osteoclast formation, and promotes osteoblast differentiation

    DEFF Research Database (Denmark)

    Boissy, Patrice; Andersen, Thomas L; Abdallah, Basem M;

    2005-01-01

    Multiple myeloma is characterized by the accumulation of clonal malignant plasma cells in the bone marrow, which stimulates bone destruction by osteoclasts and reduces bone formation by osteoblasts. In turn, the changed bone microenvironment sustains survival of myeloma cells. Therefore......, a challenge for treating multiple myeloma is discovering drugs targeting not only myeloma cells but also osteoclasts and osteoblasts. Because resveratrol (trans-3,4',5-trihydroxystilbene) is reported to display antitumor activities on a variety of human cancer cells, we investigated the effects...... of this natural compound on myeloma and bone cells. We found that resveratrol reduces dose-dependently the growth of myeloma cell lines (RPMI 8226 and OPM-2) by a mechanism involving cell apoptosis. In cultures of human primary monocytes, resveratrol inhibits dose-dependently receptor activator of nuclear factor...

  1. MUC16 provides immune protection by inhibiting synapse formation between NK and ovarian tumor cells

    Directory of Open Access Journals (Sweden)

    Migneault Martine

    2010-01-01

    Full Text Available Abstract Background Cancer cells utilize a variety of mechanisms to evade immune detection and attack. Effective immune detection largely relies on the formation of an immune synapse which requires close contact between immune cells and their targets. Here, we show that MUC16, a heavily glycosylated 3-5 million Da mucin expressed on the surface of ovarian tumor cells, inhibits the formation of immune synapses between NK cells and ovarian tumor targets. Our results indicate that MUC16-mediated inhibition of immune synapse formation is an effective mechanism employed by ovarian tumors to evade immune recognition. Results Expression of low levels of MUC16 strongly correlated with an increased number of conjugates and activating immune synapses between ovarian tumor cells and primary naïve NK cells. MUC16-knockdown ovarian tumor cells were more susceptible to lysis by primary NK cells than MUC16 expressing controls. This increased lysis was not due to differences in the expression levels of the ligands for the activating receptors DNAM-1 and NKG2D. The NK cell leukemia cell line (NKL, which does not express KIRs but are positive for DNAM-1 and NKG2D, also conjugated and lysed MUC16-knockdown cells more efficiently than MUC16 expressing controls. Tumor cells that survived the NKL challenge expressed higher levels of MUC16 indicating selective lysis of MUC16low targets. The higher csMUC16 levels on the NKL resistant tumor cells correlated with more protection from lysis as compared to target cells that were never exposed to the effectors. Conclusion MUC16, a carrier of the tumor marker CA125, has previously been shown to facilitate ovarian tumor metastasis and inhibits NK cell mediated lysis of tumor targets. Our data now demonstrates that MUC16 expressing ovarian cancer cells are protected from recognition by NK cells. The immune protection provided by MUC16 may lead to selective survival of ovarian cancer cells that are more efficient in

  2. Survivin Modulates Squamous Cell Carcinoma-Derived Stem-Like Cell Proliferation, Viability and Tumor Formation in Vivo

    Directory of Open Access Journals (Sweden)

    Roberta Lotti

    2016-01-01

    Full Text Available Squamous Cell Carcinoma-derived Stem-like Cells (SCC-SC originate from alterations in keratinocyte stem cells (KSC gene expression and sustain tumor development, invasion and recurrence. Since survivin, a KSC marker, is highly expressed in SCC-SC, we evaluate its role in SCC-SC cell growth and SCC models. Survivin silencing by siRNA decreases clonal growth of SCC keratinocytes and viability of total, rapidly adhering (RAD and non-RAD (NRAD cells from primary SCC. Similarly, survivin silencing reduces the expression of stem cell markers (OCT4, NOTCH1, CD133, β1-integrin, while it increases the level of differentiation markers (K10, involucrin. Moreover, survivin silencing improves the malignant phenotype of SCC 3D-reconstruct, as demonstrated by reduced epidermal thickness, lower Ki-67 positive cell number, and decreased expression of MMP9 and psoriasin. Furthermore, survivin depletion by siRNA in RasG12V-IκBα-derived tumors leads to smaller tumor formation characterized by lower mitotic index and reduced expression of the tumor-associated marker HIF1α, VEGF and CD51. Therefore, our results indicate survivin as a key gene in regulating SCC cancer stem cell formation and cSCC development.

  3. Survivin Modulates Squamous Cell Carcinoma-Derived Stem-Like Cell Proliferation, Viability and Tumor Formation in Vivo.

    Science.gov (United States)

    Lotti, Roberta; Palazzo, Elisabetta; Petrachi, Tiziana; Dallaglio, Katiuscia; Saltari, Annalisa; Truzzi, Francesca; Quadri, Marika; Puviani, Mario; Maiorana, Antonino; Marconi, Alessandra; Pincelli, Carlo

    2016-01-01

    Squamous Cell Carcinoma-derived Stem-like Cells (SCC-SC) originate from alterations in keratinocyte stem cells (KSC) gene expression and sustain tumor development, invasion and recurrence. Since survivin, a KSC marker, is highly expressed in SCC-SC, we evaluate its role in SCC-SC cell growth and SCC models. Survivin silencing by siRNA decreases clonal growth of SCC keratinocytes and viability of total, rapidly adhering (RAD) and non-RAD (NRAD) cells from primary SCC. Similarly, survivin silencing reduces the expression of stem cell markers (OCT4, NOTCH1, CD133, β₁-integrin), while it increases the level of differentiation markers (K10, involucrin). Moreover, survivin silencing improves the malignant phenotype of SCC 3D-reconstruct, as demonstrated by reduced epidermal thickness, lower Ki-67 positive cell number, and decreased expression of MMP9 and psoriasin. Furthermore, survivin depletion by siRNA in Ras(G12V)-IκBα-derived tumors leads to smaller tumor formation characterized by lower mitotic index and reduced expression of the tumor-associated marker HIF1α, VEGF and CD51. Therefore, our results indicate survivin as a key gene in regulating SCC cancer stem cell formation and cSCC development.

  4. Sequential Salinomycin Treatment Results in Resistance Formation through Clonal Selection of Epithelial-Like Tumor Cells

    Directory of Open Access Journals (Sweden)

    Florian Kopp

    2014-12-01

    Full Text Available Acquiring therapy resistance is one of the major obstacles in the treatment of patients with cancer. The discovery of the cancer stem cell (CSC–specific drug salinomycin raised hope for improved treatment options by targeting therapy-refractory CSCs and mesenchymal cancer cells. However, the occurrence of an acquired salinomycin resistance in tumor cells remains elusive. To study the formation of salinomycin resistance, mesenchymal breast cancer cells were sequentially treated with salinomycin in an in vitro cell culture assay, and the resulting differences in gene expression and salinomycin susceptibility were analyzed. We demonstrated that long-term salinomycin treatment of mesenchymal cancer cells resulted in salinomycin-resistant cells with elevated levels of epithelial markers, such as E-cadherin and miR-200c, a decreased migratory capability, and a higher susceptibility to the classic chemotherapeutic drug doxorubicin. The formation of salinomycin resistance through the acquisition of epithelial traits was further validated by inducing mesenchymal-epithelial transition through an overexpression of miR-200c. The transition from a mesenchymal to a more epithelial-like phenotype of salinomycin-treated tumor cells was moreover confirmed in vivo, using syngeneic and, for the first time, transgenic mouse tumor models. These results suggest that the acquisition of salinomycin resistance through the clonal selection of epithelial-like cancer cells could become exploited for improved cancer therapies by antagonizing the tumor-progressive effects of epithelial-mesenchymal transition.

  5. Synthetic niches for differentiation of human embryonic stem cells bypassing embryoid body formation.

    Science.gov (United States)

    Liu, Yarong; Fox, Victoria; Lei, Yuning; Hu, Biliang; Joo, Kye-Il; Wang, Pin

    2014-07-01

    The unique self-renewal and pluripotency features of human embryonic stem cells (hESCs) offer the potential for unlimited development of novel cell therapies. Currently, hESCs are cultured and differentiated using methods, such as monolayer culture and embryoid body (EB) formation. As such, achieving efficient differentiation into higher order structures remains a challenge, as well as maintaining cell viability during differentiation into homogeneous cell populations. Here, we describe the application of highly porous polymer scaffolds as synthetic stem cell niches. Bypassing the EB formation step, these scaffolds are capable of three-dimensional culture of undifferentiated hESCs and subsequent directed differentiation into three primary germ layers. H9 hESCs were successfully maintained and proliferated in biodegradable polymer scaffolds based on poly (lactic-co-glycolic acid) (PLGA). The results showed that cells within PLGA scaffolds retained characteristics of undifferentiated pluripotent stem cells. Moreover, the scaffolds allowed differentiation towards the lineage of interest by the addition of growth factors to the culture system. The in vivo transplantation study revealed that the scaffolds could provide a microenvironment that enabled hESCs to interact with their surroundings, thereby promoting cell differentiation. Therefore, this approach, which provides a unique culture/differentiation system for hESCs, will find its utility in various stem cell-based tissue-engineering applications.

  6. Sequential Salinomycin Treatment Results in Resistance Formation through Clonal Selection of Epithelial-Like Tumor Cells.

    Science.gov (United States)

    Kopp, Florian; Hermawan, Adam; Oak, Prajakta Shirish; Ulaganathan, Vijay Kumar; Herrmann, Annika; Elnikhely, Nefertiti; Thakur, Chitra; Xiao, Zhiguang; Knyazev, Pjotr; Ataseven, Beyhan; Savai, Rajkumar; Wagner, Ernst; Roidl, Andreas

    2014-12-01

    Acquiring therapy resistance is one of the major obstacles in the treatment of patients with cancer. The discovery of the cancer stem cell (CSC)-specific drug salinomycin raised hope for improved treatment options by targeting therapy-refractory CSCs and mesenchymal cancer cells. However, the occurrence of an acquired salinomycin resistance in tumor cells remains elusive. To study the formation of salinomycin resistance, mesenchymal breast cancer cells were sequentially treated with salinomycin in an in vitro cell culture assay, and the resulting differences in gene expression and salinomycin susceptibility were analyzed. We demonstrated that long-term salinomycin treatment of mesenchymal cancer cells resulted in salinomycin-resistant cells with elevated levels of epithelial markers, such as E-cadherin and miR-200c, a decreased migratory capability, and a higher susceptibility to the classic chemotherapeutic drug doxorubicin. The formation of salinomycin resistance through the acquisition of epithelial traits was further validated by inducing mesenchymal-epithelial transition through an overexpression of miR-200c. The transition from a mesenchymal to a more epithelial-like phenotype of salinomycin-treated tumor cells was moreover confirmed in vivo, using syngeneic and, for the first time, transgenic mouse tumor models. These results suggest that the acquisition of salinomycin resistance through the clonal selection of epithelial-like cancer cells could become exploited for improved cancer therapies by antagonizing the tumor-progressive effects of epithelial-mesenchymal transition. PMID:25500079

  7. Lumen Formation Is an Intrinsic Property of Isolated Human Pluripotent Stem Cells.

    Science.gov (United States)

    Taniguchi, Kenichiro; Shao, Yue; Townshend, Ryan F; Tsai, Yu-Hwai; DeLong, Cynthia J; Lopez, Shawn A; Gayen, Srimonta; Freddo, Andrew M; Chue, Deming J; Thomas, Dennis J; Spence, Jason R; Margolis, Benjamin; Kalantry, Sundeep; Fu, Jianping; O'Shea, K Sue; Gumucio, Deborah L

    2015-12-01

    We demonstrate that dissociated human pluripotent stem cells (PSCs) are intrinsically programmed to form lumens. PSCs form two-cell cysts with a shared apical domain within 20 hr of plating; these cysts collapse to form monolayers after 5 days. Expression of pluripotency markers is maintained throughout this time. In two-cell cysts, an apical domain, marked by EZRIN and atypical PKCζ, is surrounded by apically targeted organelles (early endosomes and Golgi). Molecularly, actin polymerization, regulated by ARP2/3 and mammalian diaphanous-related formin 1 (MDIA), promotes lumen formation, whereas actin contraction, mediated by MYOSIN-II, inhibits this process. Finally, we show that lumenal shape can be manipulated in bioengineered micro-wells. Since lumen formation is an indispensable step in early mammalian development, this system can provide a powerful model for investigation of this process in a controlled environment. Overall, our data establish that lumenogenesis is a fundamental cell biological property of human PSCs. PMID:26626176

  8. Human disc cells in monolayer vs 3D culture: cell shape, division and matrix formation

    Directory of Open Access Journals (Sweden)

    Hanley Edward N

    2000-10-01

    Full Text Available Abstract Background The relationship between cell shape, proliferation, and extracellular matrix (ECM production, important aspects of cell behavior, is examined in a little-studied cell type, the human annulus cell from the intervertebral disc, during monolayer vs three-dimensional (3D culture. Results Three experimental studies showed that cells respond specifically to culture microenvironments by changes in cell shape, mitosis and ECM production: 1 Cell passages showed extensive immunohistochemical evidence of Type I and II collagens only in 3D culture. Chondroitin sulfate and keratan sulfate were abundant in both monolayer and 3D cultures. 2 Cells showed significantly greater proliferation in monolayer in the presence of platelet-derived growth factor compared to cells in 3D. 3 Cells on Matrigel™-coated monolayer substrates became rounded and formed nodular colonies, a finding absent during monolayer growth. Conclusions The cell's in vivo interactions with the ECM can regulate shape, gene expression and other cell functions. The shape of the annulus cell changes markedly during life: the young, healthy disc contains spindle shaped cells and abundant collagen. With aging and degeneration, many cells assume a strikingly different appearance, become rounded and are surrounded by unusual accumulations of ECM products. In vitro manipulation of disc cells provides an experimental window for testing how disc cells from given individuals respond when they are grown in environments which direct cells to have either spindle- or rounded-shapes. In vitro assessment of the response of such cells to platelet-derived growth factor and to Matrigel™ showed a continued influence of cell shape even in the presence of a growth factor stimulus. These findings contribute new information to the important issue of the influence of cell shape on cell behavior.

  9. Comparative study on DBPs formation profiles of intermediate organics from hydroxyl radicals oxidation of microbial cells.

    Science.gov (United States)

    Ou, Tai-You; Wang, Gen-Shuh

    2016-05-01

    This study assessed the characteristics of disinfection byproducts (DBPs) formation from intermediate organics during UV/H2O2 treatment of activated sludge and algae cells under various reaction conditions. The DBPs including trihalomethanes (THMs), haloacetic acids (HAAs), haloketones (HKs) and haloacetonitriles (HANs) in UV/H2O2-treated and chlorinated water were measured. The results showed that both dissolved organic carbon (DOC) and dissolved organic nitrogen (DON) increased during the initial stage of UV/H2O2 treatment due to the lysis of sludge and algae cells, which enhanced the formation of both C- and N-DBPs; however, both DOC and DON decreased after longer reaction times. During the UV/H2O2 treatments, THMs formation potential (THMFP) peaked earlier than did HAAs formation potential (HAAFP). This shows that the dissolved organics released from lysis of microbial cells in the early stages of oxidation favor the production of THMs over HAAs; however, HAAs precursors increased with the oxidation time. Chlorination with bromide increased the formation of THMs and HAAs but less HKs and HANs were produced. Comparisons of normalized DBP formation potential (DBPFP) of samples collected during UV/H2O2 treatments of four different types of organic matter showed that the highest DBPFP occurred in filtered treated wastewater effluent, followed by samples of activated sludge, filtered eutrophicated pond water, and samples of algae cells. With increasing oxidation time, the dominant DBP species shifted from THMs to HAAs in the samples of activated sludge and algae cells. The DBPFP tests also showed that more HAAs were formed in biologically treated wastewater effluent, while the eutrophicated source water produced more THMs.

  10. Induction of HLA-B27 heavy chain homodimer formation after activation in dendritic cells

    OpenAIRE

    Santos, Susana G.; Lynch, Sarah; Campbell, Elaine C.; Antoniou, Antony N.; Simon J Powis

    2008-01-01

    Introduction Ankylosing spondylitis (AS) is a severe, chronic inflammatory arthritis, with a strong association to the human major histocompatibilty complex (MHC) class I allele human leucocyte antigen (HLA) B27. Disulfide-linked HLA-B27 heavy-chain homodimers have been implicated as novel structures involved in the aetiology of AS. We have studied the formation of HLA-B27 heavy-chain homodimers in human dendritic cells, which are key antigen-presenting cells and regulators of mammalian immun...

  11. Teratoma formation of human embryonic stem cells in three-dimensional perfusion culture bioreactors.

    Science.gov (United States)

    Stachelscheid, H; Wulf-Goldenberg, A; Eckert, K; Jensen, J; Edsbagge, J; Björquist, P; Rivero, M; Strehl, R; Jozefczuk, J; Prigione, A; Adjaye, J; Urbaniak, T; Bussmann, P; Zeilinger, K; Gerlach, J C

    2013-09-01

    Teratoma formation in mice is today the most stringent test for pluripotency that is available for human pluripotent cells, as chimera formation and tetraploid complementation cannot be performed with human cells. The teratoma assay could also be applied for assessing the safety of human pluripotent cell-derived cell populations intended for therapeutic applications. In our study we examined the spontaneous differentiation behaviour of human embryonic stem cells (hESCs) in a perfused 3D multi-compartment bioreactor system and compared it with differentiation of hESCs and human induced pluripotent cells (hiPSCs) cultured in vitro as embryoid bodies and in vivo in an experimental mouse model of teratoma formation. Results from biochemical, histological/immunohistological and ultrastuctural analyses revealed that hESCs cultured in bioreactors formed tissue-like structures containing derivatives of all three germ layers. Comparison with embryoid bodies and the teratomas revealed a high degree of similarity of the tissues formed in the bioreactor to these in the teratomas at the histological as well as transcriptional level, as detected by comparative whole-genome RNA expression profiling. The 3D culture system represents a novel in vitro model that permits stable long-term cultivation, spontaneous multi-lineage differentiation and tissue formation of pluripotent cells that is comparable to in vivo differentiation. Such a model is of interest, e.g. for the development of novel cell differentiation strategies. In addition, the 3D in vitro model could be used for teratoma studies and pluripotency assays in a fully defined, controlled environment, alternatively to in vivo mouse models.

  12. Insulin and glucose play a role in foam cell formation and function

    OpenAIRE

    Keller Susanna R; Huo Yuqing; Rissing Benjamin A; Miller Yury I; Jain Nitin; Shashkin Pavel N; Vandenhoff George E; Nadler Jerry L; McIntyre Thomas M

    2006-01-01

    Abstract Background Foam cell formation in diabetic patients often occurs in the presence of high insulin and glucose levels. To test whether hyperinsulinemic hyperglycemic conditions affect foam cell differentiation, we examined gene expression, cytokine production, and Akt phosphorylation in human monocyte-derived macrophages incubated with two types of oxidized low density lipoprotein (LDL), minimally modified LDL (mmLDL) and extensively oxidized LDL (OxLDL). Methods and results Using Affy...

  13. Convective cell formation and anomalous diffusion due to electromagnetic drift wave turbulence

    International Nuclear Information System (INIS)

    Convective cell formation and spectral cascade processes due to gravitational drift Alfven waves are studied using a new type of model equation. Conservation relations are derived and explosive instability is found for systems near marginal finite β stability. This instability also remains when the effects of poor as well as favorable curvature regions are included, i.e., for ballooning modes. The anomalous diffusion due to convective cells and quasi-linear effects are compared

  14. Exosomes from B cells and Dendritic cells: mechanisms of formation, secretion and targeting

    NARCIS (Netherlands)

    Buschow, S.I.

    2006-01-01

    Many cell types, including dendritic cells (DC) and B cells, secrete small vesicles called exosomes. Exosomes from immune cells are thought to have immuno-regulatory functions but their precise role remains unresolved. The aim of the studies presented in this thesis was to get more insight into the

  15. Dynamics and roles of phragmoplast microfilaments in cell plate formation during cytokinesis of tobacco BY-2 cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Yan; ZHANG WenJie; BALUSKA Frantisek; MENZEL Diedrik; REN HaiYun

    2009-01-01

    The phragmoplast is a special apparatus that functions in establishing a cell plate in dividing plant cells.It is known that microfilaments (MFs) are involved in constituting phragmoplast structure, but the dynamic distribution and role of phragmoplast MFs are far from being understood. In this study, the precise structure and dynamics of MFs during the initiation and the late lateral expansion of the phragmoplast were observed by using a tobacco BY-2 cell line stably expressing the microfilament reporter construct GFP-f ABD2. Three-dimensional imaging showed that the phragmoplast MFs were initiated by two populations of MFs emerging between the reconstituting daughter nuclei at anaphase, which migrated to the mid-zone and gave rise to two layers of microfilament arrays. FM4-64 stained vesicles accumulated and fused with the cell plate between the two populations of MFs. The two layers of microfilament arrays of phragmoplast with ends overlapped always surrounded the centrifugally expanding cell plate. Partial disruption of MFs at metaphase by low concentration of latrunculin B resulted in the inhibition of the cell plate consolidation and the blockage of cell plate lateral expansion,whereas high concentration of latrunculin B restrained the progression of the cell cycle. Treating the cell after the initiation of phragmoplast led to the cease of the expansion of the cell plate. Our observations provide new insights into the precise structure and dynamics of phragmoplast MFs during cytokinesis and suggest that dynamic phragmoplast MFs are important in cell plate formation.

  16. "Danger" conditions increase sulfamethoxazole-protein adduct formation in human antigen-presenting cells.

    Science.gov (United States)

    Lavergne, S N; Wang, H; Callan, H E; Park, B K; Naisbitt, D J

    2009-11-01

    Antigen-presenting cells (APC) are thought to play an important role in the pathogenesis of drug-induced immune reactions. Various pathological factors can activate APC and therefore influence the immune equilibrium. It is interesting that several diseases have been associated with an increased rate of drug allergy. The aim of this project was to evaluate the impact of such "danger signals" on sulfamethoxazole (SMX) metabolism in human APC (peripheral blood mononuclear cells, Epstein-Barr virus-modified B lymphocytes, monocyte-derived dendritic cells, and two cell lines). APC were incubated with SMX (100 microM-2 mM; 5 min-24 h), in the presence of pathological factors: bacterial endotoxins (lipopolysaccharide and staphylococcal enterotoxin B), flu viral proteins, cytokines [interleukin (IL)-1beta, IL-6, IL-10; tumor necrosis factor-alpha; interferon-gamma; and transforming growth factor-beta], inflammatory molecules (prostaglandin E2, human serum complement, and activated protein C), oxidants (buthionine sulfoximine and H(2)O(2)), and hyperthermia (37.5-39.5 degrees C). Adduct formation was evaluated by enzyme-linked immunosorbent assay and confocal microscopy. SMX-protein adduct formation was time- and concentration-dependent for each cell type tested, in both physiological and danger conditions. A danger environment significantly increased the formation of SMX-protein adducts and significantly shortened the delay for their detection. An additive effect was observed with a combination of danger signals. Dimedone (chemical selectively binding cysteine sulfenic acid) and antioxidants decreased both baseline and danger-enhanced SMX-adduct formation. Various enzyme inhibitors were associated with a significant decrease in SMX-adduct levels, with a pattern varying depending on the cell type and the culture conditions. These results illustrate that danger signals enhance the formation of intracellular SMX-protein adducts in human APC. These findings might be relevant

  17. Reggies/flotillins regulate E-cadherin-mediated cell contact formation by affecting EGFR trafficking.

    Science.gov (United States)

    Solis, Gonzalo P; Schrock, Yvonne; Hülsbusch, Nikola; Wiechers, Marianne; Plattner, Helmut; Stuermer, Claudia A O

    2012-05-01

    The reggie/flotillin proteins are implicated in membrane trafficking and, together with the cellular prion protein (PrP), in the recruitment of E-cadherin to cell contact sites. Here, we demonstrate that reggies, as well as PrP down-regulation, in epithelial A431 cells cause overlapping processes and abnormal formation of adherens junctions (AJs). This defect in cell adhesion results from reggie effects on Src tyrosine kinases and epidermal growth factor receptor (EGFR): loss of reggies reduces Src activation and EGFR phosphorylation at residues targeted by Src and c-cbl and leads to increased surface exposure of EGFR by blocking its internalization. The prolonged EGFR signaling at the plasma membrane enhances cell motility and macropinocytosis, by which junction-associated E-cadherin is internalized and recycled back to AJs. Accordingly, blockage of EGFR signaling or macropinocytosis in reggie-deficient cells restores normal AJ formation. Thus, by promoting EGFR internalization, reggies restrict the EGFR signaling involved in E-cadherin macropinocytosis and recycling and regulate AJ formation and dynamics and thereby cell adhesion.

  18. Biofilm formation of Salmonella serotypes in simulated meat processing environments and its relationship to cell characteristics.

    Science.gov (United States)

    Wang, Huhu; Ding, Shijie; Dong, Yang; Ye, Keping; Xu, Xinglian; Zhou, Guanghong

    2013-10-01

    Salmonella attached to meat contact surfaces encountered in meat processing facilities may serve as a source of cross-contamination. In this study, the influence of serotypes and media on biofilm formation of Salmonella was investigated in a simulated meat processing environment, and the relationships between biofilm formation and cell characteristics were also determined. All six serotypes (Salmonella enterica serotype Heidelberg, Salmonella Derby, Salmonella Agona, Salmonella Indiana, Salmonella Infantis, and Salmonella Typhimurium) can readily form biofilms on stainless steel surfaces, and the amounts of biofilms were significantly influenced by the serotypes, incubation media, and incubation time used in this study. Significant differences in cell surface hydrophobicity, autoaggregation, motility, and growth kinetic parameters were observed between individual serotypes tested. Except for growth kinetic parameters, the cell characteristics were correlated with the ability of biofilm formation incubated in tryptic soy broth, whereas no correlation with biofilm formation incubated in meat thawing-loss broth (an actual meat substrate) was found. Salmonella grown in meat thawing-loss broth showed a "cloud-shaped" morphology in the mature biofilm, whereas when grown in tryptic soy broth it had a "reticulum-shaped" appearance. Our study provides some practical information to understand the process of biofilm formation on meat processing contact surfaces.

  19. Monte Carlo study of receptor-lipid raft formation on a cell membrane

    Science.gov (United States)

    Yu-Yang, Paul; Srinivas Reddy, A.; Raychaudhuri, Subhadip

    2012-02-01

    Receptors are cell surface molecules that bind with extracellular ligand molecules leading to propagation of downstream signals and cellular activation. Even though ligand binding-induced formation of receptor-lipid rafts has been implicated in such a process, the formation mechanism of such large stable rafts is not understood. We present findings from our Monte Carlo (MC) simulations involving (i) receptor interaction with the membrane lipids and (ii) lipid-lipid interactions between raft forming lipids. We have developed a hybrid MC simulation method that combines a probabilistic MC simulation with an explicit free energy-based MC scheme. Some of the lipid-mediated interactions, such as the cholesterol-lipid interactions, are simulated in an implicit way. We examine the effect of varying attractive interactions between raft forming lipids and ligand-bound receptors and show that strong coupling between receptor-receptor and receptor-sphingolipid molecules generate raft formation similar to that observed in recent biological experiments. We study the effect of variation of receptor affinity for ligands (as happens in adaptive immune cells) on raft formation. Such affinity dependence in receptor-lipid raft formation provides insight into important problems in B cell biology.

  20. Differential survival of solitary and aggregated bacterial cells promotes aggregate formation on leaf surfaces

    Science.gov (United States)

    Monier, J.-M.; Lindow, S. E.

    2003-01-01

    The survival of individual Pseudomonas syringae cells was determined on bean leaf surfaces maintained under humid conditions or periodically exposed to desiccation stress. Cells of P. syringae strain B728a harboring a GFP marker gene were visualized by epifluorescence microscopy, either directly in situ or after recovery from leaves, and dead cells were identified as those that were stained with propidium iodide in such populations. Under moist, conducive conditions on plants, the proportion of total live cells was always high, irrespective of their aggregated state. In contrast, the proportion of the total cells that remained alive on leaves that were periodically exposed to desiccation stress decreased through time and was only ≈15% after 5 days. However, the fraction of cells in large aggregates that were alive on such plants in both condition was much higher than more solitary cells. Immediately after inoculation, cells were randomly distributed over the leaf surface and no aggregates were observed. However, a very aggregated pattern of colonization was apparent within 7 days, and >90% of the living cells were located in aggregates of 100 cells or more. Our results strongly suggest that, although conducive conditions favor aggregate formation, such cells are much more capable of tolerating environmental stresses, and the preferential survival of cells in aggregates promotes a highly clustered spatial distribution of bacteria on leaf surfaces. PMID:14665692

  1. Deletion of Brg1 causes abnormal hair cell planer polarity, hair cell anchorage, and scar formation in mouse cochlea

    Science.gov (United States)

    Jin, Yecheng; Ren, Naixia; Li, Shiwei; Fu, Xiaolong; Sun, Xiaoyang; Men, Yuqin; Xu, Zhigang; Zhang, Jian; Xie, Yue; Xia, Ming; Gao, Jiangang

    2016-01-01

    Hair cells (HCs) are mechanosensors that play crucial roles in perceiving sound, acceleration, and fluid motion. The precise architecture of the auditory epithelium and its repair after HC loss is indispensable to the function of organ of Corti (OC). In this study, we showed that Brg1 was highly expressed in auditory HCs. Specific deletion of Brg1 in postnatal HCs resulted in rapid HC degeneration and profound deafness in mice. Further experiments showed that cell-intrinsic polarity of HCs was abolished, docking of outer hair cells (OHCs) by Deiter’s cells (DCs) failed, and scar formation in the reticular lamina was deficient. We demonstrated that Brg1 ablation disrupted the Gαi/Insc/LGN and aPKC asymmetric distributions, without overt effects on the core planer cell polarity (PCP) pathway. We also demonstrated that Brg1-deficient HCs underwent apoptosis, and that leakage in the reticular lamina caused by deficient scar formation shifted the mode of OHC death from apoptosis to necrosis. Together, these data demonstrated a requirement for Brg1 activity in HC development and suggested a role for Brg1 in the proper cellular structure formation of HCs. PMID:27255603

  2. TCPs, WUSs, and WINDs: Families of transcription factors that regulate shoot meristem formation, stem cell maintenance, and somatic cell differentiation

    Directory of Open Access Journals (Sweden)

    Miho eIkeda

    2014-09-01

    Full Text Available In contrast to somatic mammalian cells, which cannot alter their fate, plant cells can dedifferentiate to form totipotent callus cells and regenerate a whole plant, following treatment with specific phytohormones. However, the regulatory mechanisms and key factors that control differentiation-dedifferentiation and cell totipotency have not been completely clarified in plants. Recently, several plant transcription factors that regulate meristem formation and dedifferentiation have been identified and include members of the TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP, WUSCHEL (WUS, and WOUND INDUCED DEDIFFERENTIATION (WIND1 families. WUS and WIND positively control plant cell totipotency, while TCP negatively controls it. Interestingly, TCP is a transcriptional activator that acts as a negative regulator of shoot meristem formation, and WUS is a transcriptional repressor that positively maintains totipotency of the stem cells of the shoot meristem. We describe here the functions of TCP, WUS and WIND transcription factors in the regulation of differentiation-dedifferentiation by positive and negative transcriptional regulators.

  3. Disturbance of the bacterial cell wall specifically interferes with biofilm formation.

    Science.gov (United States)

    Bucher, Tabitha; Oppenheimer-Shaanan, Yaara; Savidor, Alon; Bloom-Ackermann, Zohar; Kolodkin-Gal, Ilana

    2015-12-01

    In nature, bacteria communicate via chemical cues and establish complex communities referred to as biofilms, wherein cells are held together by an extracellular matrix. Much research is focusing on small molecules that manipulate and prevent biofilm assembly by modifying cellular signalling pathways. However, the bacterial cell envelope, presenting the interface between bacterial cells and their surroundings, is largely overlooked. In our study, we identified specific targets within the biosynthesis pathways of the different cell wall components (peptidoglycan, wall teichoic acids and teichuronic acids) hampering biofilm formation and the anchoring of the extracellular matrix with a minimal effect on planktonic growth. In addition, we provide convincing evidence that biofilm hampering by transglycosylation inhibitors and D-Leucine triggers a highly specific response without changing the overall protein levels within the biofilm cells or the overall levels of the extracellular matrix components. The presented results emphasize the central role of the Gram-positive cell wall in biofilm development, resistance and sustainment. PMID:26472159

  4. Disturbance of the bacterial cell wall specifically interferes with biofilm formation.

    Science.gov (United States)

    Bucher, Tabitha; Oppenheimer-Shaanan, Yaara; Savidor, Alon; Bloom-Ackermann, Zohar; Kolodkin-Gal, Ilana

    2015-12-01

    In nature, bacteria communicate via chemical cues and establish complex communities referred to as biofilms, wherein cells are held together by an extracellular matrix. Much research is focusing on small molecules that manipulate and prevent biofilm assembly by modifying cellular signalling pathways. However, the bacterial cell envelope, presenting the interface between bacterial cells and their surroundings, is largely overlooked. In our study, we identified specific targets within the biosynthesis pathways of the different cell wall components (peptidoglycan, wall teichoic acids and teichuronic acids) hampering biofilm formation and the anchoring of the extracellular matrix with a minimal effect on planktonic growth. In addition, we provide convincing evidence that biofilm hampering by transglycosylation inhibitors and D-Leucine triggers a highly specific response without changing the overall protein levels within the biofilm cells or the overall levels of the extracellular matrix components. The presented results emphasize the central role of the Gram-positive cell wall in biofilm development, resistance and sustainment.

  5. Mitotic rounding alters cell geometry to ensure efficient bipolar spindle formation.

    Science.gov (United States)

    Lancaster, Oscar M; Le Berre, Maël; Dimitracopoulos, Andrea; Bonazzi, Daria; Zlotek-Zlotkiewicz, Ewa; Picone, Remigio; Duke, Thomas; Piel, Matthieu; Baum, Buzz

    2013-05-13

    Accurate animal cell division requires precise coordination of changes in the structure of the microtubule-based spindle and the actin-based cell cortex. Here, we use a series of perturbation experiments to dissect the relative roles of actin, cortical mechanics, and cell shape in spindle formation. We find that, whereas the actin cortex is largely dispensable for rounding and timely mitotic progression in isolated cells, it is needed to drive rounding to enable unperturbed spindle morphogenesis under conditions of confinement. Using different methods to limit mitotic cell height, we show that a failure to round up causes defects in spindle assembly, pole splitting, and a delay in mitotic progression. These defects can be rescued by increasing microtubule lengths and therefore appear to be a direct consequence of the limited reach of mitotic centrosome-nucleated microtubules. These findings help to explain why most animal cells round up as they enter mitosis.

  6. Development of Large-Format Lithium-Ion Cells with Silicon Anode and Low Flammable Electrolyte

    Science.gov (United States)

    Wu, James J.; Hernandez-Lugo, D. M.; Smart, M. C.; Ratnakumar, B. V.; Miller, T. B.; Lvovich, V. F.; Lytle, J. K.

    2014-01-01

    NASA is developing safe, high energy and high capacity lithium-ion cell designs and batteries for future missions under NASAs Advanced Space Power System (ASPS) project. Advanced cell components, such as high specific capacity silicon anodes and low-flammable electrolytes have been developed for improving the cell specific energy and enhancing safety. To advance the technology readiness level, we have developed large-format flight-type hermetically sealed battery cells by incorporating high capacity silicon anodes, commercially available lithium nickel, cobalt, aluminum oxide (NCA) cathodes, and low-flammable electrolytes. In this report, we will present the performance results of these various battery cells. In addition, we will also discuss the post-test cell analysis results as well.

  7. Arginine-vasopressin stimulates the formation of phosphatidic acid in rat Leydig cells

    DEFF Research Database (Denmark)

    Nielsen, J.R.; Hansen, Harald S.; Jensen, B.

    1987-01-01

    Arginine-vasopressin (AVP) stimulated the formation of labelled phosphatidic acid (PA) in [C]arachidonic acid-prelabelled rat Leydig cells. After addition of 10 M AVP [C]arachidonoylphosphatidic acid reached a maximum within 2 min. The increase was dose-dependent (10-10 M). No change in labelling...

  8. Neurofibromin Deficient Myeloid Cells are Critical Mediators of Aneurysm Formation In Vivo

    Science.gov (United States)

    Li, Fang; Downing, Brandon D.; Smiley, Lucy C.; Mund, Julie A.; DiStasi, Matthew R.; Bessler, Waylan K.; Sarchet, Kara N.; Hinds, Daniel M.; Kamendulis, Lisa M.; Hingtgen, Cynthia M.; Case, Jamie; Clapp, D. Wade; Conway, Simon J.; Stansfield, Brian K.; Ingram, David A.

    2014-01-01

    Background Neurofibromatosis Type 1 (NF1) is a genetic disorder resulting from mutations in the NF1 tumor suppressor gene. Neurofibromin, the protein product of NF1, functions as a negative regulator of Ras activity in circulating hematopoietic and vascular wall cells, which are critical for maintaining vessel wall homeostasis. NF1 patients have evidence of chronic inflammation resulting in development of premature cardiovascular disease, including arterial aneurysms, which may manifest as sudden death. However, the molecular pathogenesis of NF1 aneurysm formation is unknown. Method and Results Utilizing an angiotensin II-induced aneurysm model, we demonstrate that heterozygous inactivation of Nf1 (Nf1+/−) enhanced aneurysm formation with myeloid cell infiltration and increased oxidative stress in the vessel wall. Using lineage-restricted transgenic mice, we show loss of a single Nf1 allele in myeloid cells is sufficient to recapitulate the Nf1+/− aneurysm phenotype in vivo. Finally, oral administration of simvastatin or the antioxidant apocynin, reduced aneurysm formation in Nf1+/− mice. Conclusion These data provide genetic and pharmacologic evidence that Nf1+/− myeloid cells are the cellular triggers for aneurysm formation in a novel model of NF1 vasculopathy and provide a potential therapeutic target. PMID:24370551

  9. Forskolin enhances in vivo bone formation by human mesenchymal stromal cells

    NARCIS (Netherlands)

    Doorn, J.; Siddappa, R.; Blitterswijk, van C.A.; Boer, de J.

    2012-01-01

    Activation of the protein kinase A (PKA) pathway with dibutyryl cyclic adenosine monophosphate (db-cAMP) was recently shown to enhance osteogenic differentiation of human mesenchymal stromal cells (hMSCs) in vitro and bone formation in vivo. The major drawback of this compound is its inhibitory effe

  10. Formation of hydrogen peroxide and nitric oxide in rat skeletal muscle cells during contractions

    DEFF Research Database (Denmark)

    Silveira, Leonardo R.; Pereira-Da-Silva, Lucia; Juel, Carsten;

    2003-01-01

    We examined intra- and extracellular H(2)O(2) and NO formation during contractions in primary rat skeletal muscle cell culture. The fluorescent probes DCFH-DA/DCFH (2,7-dichlorofluorescein-diacetate/2,7-dichlorofluorescein) and DAF-2-DA/DAF-2 (4,5-diaminofluorescein-diacetate/4,5-diaminofluoresce...

  11. Paraoxonases 1, 2, and 3, oxidative stress, and macrophage foam cell formation during atherosclerosis development.

    Science.gov (United States)

    Aviram, Michael; Rosenblat, Mira

    2004-11-01

    Paraoxonases PON1 and PON3, which are both associated in serum with HDL, protect the serum lipids from oxidation, probably as a result of their ability to hydrolyze specific oxidized lipids. The activity of HDL-associated PON1 seems to involve an activity (phospholipase A2-like activity, peroxidase-like activity, lactonase activity) which produces LPC. To study the possible role of PON1 in macrophage foam cell formation and atherogenesis we used macrophages from control mice, from PON1 knockout mice, and from PON1 transgenic mice. Furthermore, we analyzed PON1-treated macrophages and PON1-transfected cells to demonstrate the contribution of PON1 to the attenuation of macrophage cholesterol and oxidized lipid accumulation and foam cell formation. PON1 was shown to inhibit cholesterol influx [by reducing the formation of oxidized LDL (Ox-LDL), increasing the breakdown of specific oxidized lipids in Ox-LDL, and decreasing macrophage uptake of Ox-LDL]. PON1 also inhibits cholesterol biosynthesis and stimulates HDL-mediated cholesterol efflux from macrophages. PON2 and PON3 protect against oxidative stress, with PON2 acting mainly at the cellular level. Whereas serum PON1 and PON3 were inactivated under oxidative stress, macrophage PON2 expression and activity were increased under oxidative stress, probably as a compensatory mechanism against oxidative stress. Intervention to increase the paraoxonases (cellular and humoral) by dietary or pharmacological means can reduce macrophage foam cell formation and attenuate atherosclerosis development. PMID:15454271

  12. Stereotypical cell division orientation controls neural rod midline formation in zebrafish.

    Science.gov (United States)

    Quesada-Hernández, Elena; Caneparo, Luca; Schneider, Sylvia; Winkler, Sylke; Liebling, Michael; Fraser, Scott E; Heisenberg, Carl-Philipp

    2010-11-01

    The development of multicellular organisms is dependent on the tight coordination between tissue growth and morphogenesis. The stereotypical orientation of cell divisions has been proposed to be a fundamental mechanism by which proliferating and growing tissues take shape. However, the actual contribution of stereotypical division orientation (SDO) to tissue morphogenesis is unclear. In zebrafish, cell divisions with stereotypical orientation have been implicated in both body-axis elongation and neural rod formation, although there is little direct evidence for a critical function of SDO in either of these processes. Here we show that SDO is required for formation of the neural rod midline during neurulation but dispensable for elongation of the body axis during gastrulation. Our data indicate that SDO during both gastrulation and neurulation is dependent on the noncanonical Wnt receptor Frizzled 7 (Fz7) and that interfering with cell division orientation leads to severe defects in neural rod midline formation but not body-axis elongation. These findings suggest a novel function for Fz7-controlled cell division orientation in neural rod midline formation during neurulation.

  13. Inhibition of gold nanoparticles (AuNPs) on pathogenic biofilm formation and invasion to host cells

    Science.gov (United States)

    Yu, Qilin; Li, Jianrong; Zhang, Yueqi; Wang, Yufan; Liu, Lu; Li, Mingchun

    2016-01-01

    Owing to the growing infectious diseases caused by eukaryotic and prokaryotic pathogens, it is urgent to develop novel antimicrobial agents against clinical pathogenic infections. Biofilm formation and invasion into the host cells are vital processes during pathogenic colonization and infection. In this study, we tested the inhibitory effect of Au nanoparticles (AuNPs) on pathogenic growth, biofilm formation and invasion. Interestingly, although the synthesized AuNPs had no significant toxicity to the tested pathogens, Candida albicans and Pseudomonas aeruginosa, the nanoparticles strongly inhibited pathogenic biofilm formation and invasion to dental pulp stem cells (DPSCs). Further investigations revealed that AuNPs abundantly bound to the pathogen cells, which likely contributed to their inhibitory effect on biofilm formation and invasion. Moreover, treatment of AuNPs led to activation of immune response-related genes in DPSCs, which may enhance the activity of host immune system against the pathogens. Zeta potential analysis and polyethylene glycol (PEG)/polyethyleneimine (PEI) coating tests further showed that the interaction between pathogen cells and AuNPs is associated with electrostatic attractions. Our findings shed novel light on the application of nanomaterials in fighting against clinical pathogens, and imply that the traditional growth inhibition test is not the only way to evaluate the drug effect during the screening of antimicrobial agents. PMID:27220400

  14. Inhibition of gold nanoparticles (AuNPs) on pathogenic biofilm formation and invasion to host cells.

    Science.gov (United States)

    Yu, Qilin; Li, Jianrong; Zhang, Yueqi; Wang, Yufan; Liu, Lu; Li, Mingchun

    2016-01-01

    Owing to the growing infectious diseases caused by eukaryotic and prokaryotic pathogens, it is urgent to develop novel antimicrobial agents against clinical pathogenic infections. Biofilm formation and invasion into the host cells are vital processes during pathogenic colonization and infection. In this study, we tested the inhibitory effect of Au nanoparticles (AuNPs) on pathogenic growth, biofilm formation and invasion. Interestingly, although the synthesized AuNPs had no significant toxicity to the tested pathogens, Candida albicans and Pseudomonas aeruginosa, the nanoparticles strongly inhibited pathogenic biofilm formation and invasion to dental pulp stem cells (DPSCs). Further investigations revealed that AuNPs abundantly bound to the pathogen cells, which likely contributed to their inhibitory effect on biofilm formation and invasion. Moreover, treatment of AuNPs led to activation of immune response-related genes in DPSCs, which may enhance the activity of host immune system against the pathogens. Zeta potential analysis and polyethylene glycol (PEG)/polyethyleneimine (PEI) coating tests further showed that the interaction between pathogen cells and AuNPs is associated with electrostatic attractions. Our findings shed novel light on the application of nanomaterials in fighting against clinical pathogens, and imply that the traditional growth inhibition test is not the only way to evaluate the drug effect during the screening of antimicrobial agents. PMID:27220400

  15. Free Energies of Formation Measurements on Solid-State Electrochemical Cells

    Science.gov (United States)

    Rollino, J. A.; Aronson, S.

    1972-01-01

    A simple experiment is proposed that can provide the student with some insight into the chemical properties of solids. It also demonstrates the relationship between the Gibbs free energy of formation of an ionic solid and the emf of an electrochemical cell. (DF)

  16. Jin Fu Kang Oral Liquid Inhibits Lymphatic Endothelial Cells Formation and Migration

    Directory of Open Access Journals (Sweden)

    Hai-Lang He

    2016-01-01

    Full Text Available Lung cancer is the leading cause of cancer-related deaths worldwide. Jin Fu Kang (JFK, an oral liquid prescription of Chinese herbal drugs, has been clinically available for the treatment of non-small cell lung cancer (NSCLC. Lymphangiogenesis is a primary event in the process of cancer development and metastasis, and the formation and migration of lymphatic endothelial cells (LECs play a key role in the lymphangiogenesis. To assess the activity of stromal cell-derived factor-1 (SDF-1 and the coeffect of SDF-1 and vascular endothelial growth factor-C (VEGF-C on the formation and migration of LECs and clarify the inhibitory effects of JFK on the LECs, the LECs were differentiated from CD34+/VEGFR-3+ endothelial progenitor cells (EPCs, and JFK-containing serums were prepared from rats. SDF-1 and VEGF-C both induced the differentiation of CD34+/VEGFR-3+ EPCs towards LECs and enhanced the LECs migration. Couse of SDF-1 and VEGF-C displayed an additive effect on the LECs formation but not on their migration. JFK inhibited the formation and migration of LECs, and the inhibitory effects were most probably via regulation of the SDF-1/CXCR4 and VEGF-C/VEGFR-3 axes. The current finding suggested that JFK might inhibit NSCLC through antilymphangiogenesis and also provided a potential to discover antilymphangiogenesis agents from natural resources.

  17. Hypochlorite- and hypobromite-mediated radical formation and its role in cell lysis

    DEFF Research Database (Denmark)

    Hawkins, C L; Brown, B E; Davies, Michael Jonathan

    2001-01-01

    Activated leukocytes generate the potent oxidants HOCl and HOBr via the formation of H(2)O(2) and the release of peroxidase enzymes (myeloperoxidase, eosinophil peroxidase). HOCl and HOBr are potent microbiocidal agents, but excessive or misplaced production can cause tissue damage and cell lysis...

  18. Cell Stratification, Spheroid Formation and Bioscaffolds Used to Grow Cells in Three Dimensional Cultures

    Directory of Open Access Journals (Sweden)

    Hana Hrebíková

    2015-12-01

    Full Text Available The cell culture became an invaluable tool for studying cell behaviour, development, function, gene expression, toxicity of compounds and efficacy of novel drugs. Although most results were obtained from cell cultivation in two-dimensional (2D systems, in which cells are grown in a monolayer, three-dimensional (3D cultures are more promising as they correspond closely to the native arrangement of cells in living tissues. In our study, we focused on three types of 3D in vitro systems used for cultivation of one cell type. Cell morphology, their spatial distribution inside of resulting multicellular structures and changes in time were analysed with histological examination of samples harvested at different time periods. In multilayered cultures of WRL 68 hepatocytes grown on semipermeable membranes and non-passaged neurospheres generated by proliferation of neural progenitor cells, the cells were tightly apposed, showed features of cell differentiation but also cell death that was observable in short-term cultures. Biogenic scaffolds composed of extracellular matrix of the murine tibial anterior muscle were colonized with C2C12 myoblasts in vitro. The recellularized scaffolds did not reach high cell densities comparable with the former systems but supported well cell anchorage and migration without any signs of cell regression.

  19. Polyanhydride Nanovaccines Induce Germinal Center B Cell Formation and Sustained Serum Antibody Responses.

    Science.gov (United States)

    Vela Ramirez, Julia E; Tygrett, Lorraine T; Hao, Jihua; Habte, Habtom H; Cho, Michael W; Greenspan, Neil S; Waldschmidt, Thomas J; Narasimhan, Balaji

    2016-06-01

    Biodegradable polymeric nanoparticle-based subunit vaccines have shown promising characteristics by enhancing antigen presentation and inducing protective immune responses when compared with soluble protein. Specifically, polyanhydride nanoparticle-based vaccines (i.e., nanovaccines) have been shown to successfully encapsulate and release antigens, activate B and T cells, and induce both antibody- and cell-mediated immunity towards a variety of immunogens. One of the characteristics of strong thymus-dependent antibody responses is the formation of germinal centers (GC) and the generation of GC B cells, which is part of the T helper cell driven cellular response. In order to further understand the role of nanovaccines in the induction of antigen-specific immune responses, their ability to induce germinal center B cell formation and isotype switching and the effects thereof on serum antibody responses were investigated in these studies. Polyanhydride nanovaccines based on 1,6-bis(p-carboxyphenoxy)hexane and 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane were used to subcutaneously administer a viral antigen. GC B cell formation and serum antibody responses induced by the nanovaccines were compared to that induced by alum-based vaccine formulations. It was demonstrated that a single dose of polyanhydride nanovaccines resulted in the formation of robust GCs and serum antibody in comparison to that induced by the alum-based formulation. This was attributed to the sustained release of antigen provided by the nanovaccines. When administered in a multiple dose regimen, the highest post-immunization titer and GC B cell number was enhanced, and the immune response induced by the nanovaccines was further sustained. These studies provide foundational information on the mechanism of action of polyanhydride nanovaccines. PMID:27319223

  20. Retinoic acid-treated pluripotent stem cells undergoing neurogenesis present increased aneuploidy and micronuclei formation.

    Directory of Open Access Journals (Sweden)

    Rafaela C Sartore

    Full Text Available The existence of loss and gain of chromosomes, known as aneuploidy, has been previously described within the central nervous system. During development, at least one-third of neural progenitor cells (NPCs are aneuploid. Notably, aneuploid NPCs may survive and functionally integrate into the mature neural circuitry. Given the unanswered significance of this phenomenon, we tested the hypothesis that neural differentiation induced by all-trans retinoic acid (RA in pluripotent stem cells is accompanied by increased levels of aneuploidy, as previously described for cortical NPCs in vivo. In this work we used embryonal carcinoma (EC cells, embryonic stem (ES cells and induced pluripotent stem (iPS cells undergoing differentiation into NPCs. Ploidy analysis revealed a 2-fold increase in the rate of aneuploidy, with the prevalence of chromosome loss in RA primed stem cells when compared to naïve cells. In an attempt to understand the basis of neurogenic aneuploidy, micronuclei formation and survivin expression was assessed in pluripotent stem cells exposed to RA. RA increased micronuclei occurrence by almost 2-fold while decreased survivin expression by 50%, indicating possible mechanisms by which stem cells lose their chromosomes during neural differentiation. DNA fragmentation analysis demonstrated no increase in apoptosis on embryoid bodies treated with RA, indicating that cell death is not the mandatory fate of aneuploid NPCs derived from pluripotent cells. In order to exclude that the increase in aneuploidy was a spurious consequence of RA treatment, not related to neurogenesis, mouse embryonic fibroblasts were treated with RA under the same conditions and no alterations in chromosome gain or loss were observed. These findings indicate a correlation amongst neural differentiation, aneuploidy, micronuclei formation and survivin downregulation in pluripotent stem cells exposed to RA, providing evidence that somatically generated chromosomal

  1. Regulation of germinal center responses, memory B cells and plasma cell formation-an update.

    Science.gov (United States)

    Corcoran, Lynn M; Tarlinton, David M

    2016-04-01

    Progress in understanding humoral immunity has been accelerated by the powerful experimental approaches of genetics, genomics and imaging. Excellent reviews of these advances appeared in 2015 in celebration of the 50th anniversary of the discovery of B cell and T cell lineages in the chicken. Here we provide a contemporary model of B cell differentiation, highlighting recent publications illuminating germinal center (GC), memory B cell and antibody-secreting plasma cell biology. The important contributions of CD4T cells to antibody responses have been thoroughly reviewed elsewhere.

  2. Engineering the Oryza sativa cell wall with rice NAC transcription factors regulating secondary wall formation

    Directory of Open Access Journals (Sweden)

    Kouki eYoshida

    2013-10-01

    Full Text Available Plant tissues that require structural rigidity synthesize a thick, strong secondary cell wall of lignin, cellulose and hemicelluloses in a complicated bridged structure. Master regulators of secondary wall synthesis were identified in dicots, and orthologs of these regulators have been identified in monocots, but regulation of secondary cell wall formation in monocots has not been extensively studied. Here we demonstrate that the rice transcription factors SECONDARY WALL NAC DOMAIN PROTEINs (SWNs can regulate secondary wall formation in rice (Oryza sativa and are potentially useful for engineering the monocot cell wall. The OsSWN1 promoter is highly active in sclerenchymatous cells of the leaf blade and less active in xylem cells. By contrast, the OsSWN2 promoter is highly active in xylem cells and less active in sclerenchymatous cells. OsSWN2 splicing variants encode two proteins; the shorter protein (OsSWN2S has very low transcriptional activation ability, but the longer protein (OsSWN2L and OsSWN1 have strong transcriptional activation ability. In rice, expression of an OsSWN2S chimeric repressor, driven by the OsSWN2 promoter, resulted in stunted growth and para-wilting (leaf rolling and browning under normal water conditions due to impaired vascular vessels. The same OsSWN2S chimeric repressor, driven by the OsSWN1 promoter, caused a reduction of cell wall thickening in sclerenchymatous cells, a drooping leaf phenotype, reduced lignin and xylose contents and increased digestibility as forage. These data suggest that OsSWNs regulate secondary wall formation in rice and manipulation of OsSWNs may enable improvements in monocotyledonous crops for forage or biofuel applications.

  3. Engineering the Oryza sativa cell wall with rice NAC transcription factors regulating secondary wall formation.

    Science.gov (United States)

    Yoshida, Kouki; Sakamoto, Shingo; Kawai, Tetsushi; Kobayashi, Yoshinori; Sato, Kazuhito; Ichinose, Yasunori; Yaoi, Katsuro; Akiyoshi-Endo, Miho; Sato, Hiroko; Takamizo, Tadashi; Ohme-Takagi, Masaru; Mitsuda, Nobutaka

    2013-01-01

    Plant tissues that require structural rigidity synthesize a thick, strong secondary cell wall of lignin, cellulose and hemicelluloses in a complicated bridged structure. Master regulators of secondary wall synthesis were identified in dicots, and orthologs of these regulators have been identified in monocots, but regulation of secondary cell wall formation in monocots has not been extensively studied. Here we demonstrate that the rice transcription factors SECONDARY WALL NAC DOMAIN PROTEINs (SWNs) can regulate secondary wall formation in rice (Oryza sativa) and are potentially useful for engineering the monocot cell wall. The OsSWN1 promoter is highly active in sclerenchymatous cells of the leaf blade and less active in xylem cells. By contrast, the OsSWN2 promoter is highly active in xylem cells and less active in sclerenchymatous cells. OsSWN2 splicing variants encode two proteins; the shorter protein (OsSWN2S) has very low transcriptional activation ability, but the longer protein (OsSWN2L) and OsSWN1 have strong transcriptional activation ability. In rice, expression of an OsSWN2S chimeric repressor, driven by the OsSWN2 promoter, resulted in stunted growth and para-wilting (leaf rolling and browning under normal water conditions) due to impaired vascular vessels. The same OsSWN2S chimeric repressor, driven by the OsSWN1 promoter, caused a reduction of cell wall thickening in sclerenchymatous cells, a drooping leaf phenotype, reduced lignin and xylose contents and increased digestibility as forage. These data suggest that OsSWNs regulate secondary wall formation in rice and manipulation of OsSWNs may enable improvements in monocotyledonous crops for forage or biofuel applications. PMID:24098302

  4. Liver type I regulatory T cells suppress germinal center formation in HBV-tolerant mice.

    Science.gov (United States)

    Xu, Long; Yin, Wenwei; Sun, Rui; Wei, Haiming; Tian, Zhigang

    2013-10-15

    The liver plays a critical role in inducing systemic immune tolerance, for example, during limiting hypersensitivity to food allergy and in rendering acceptance of allotransplant or even hepatotropic pathogens. We investigated the unknown mechanisms of liver tolerance by using an established hepatitis B virus (HBV)-carrier mouse model, and found that these mice exhibited an antigen-specific tolerance toward peripheral HBsAg vaccination, showing unenlarged draining lymph node (DLN), lower number of germinal centers (GC), and inactivation of GC B cells and follicular T helper (Tfh) cells. Both in vivo and in vitro immune responses toward HBsAg were suppressed by mononuclear cells from HBV-carrier mice, which were CD4(+) Foxp3(-) type 1 regulatory T (Tr1)-like cells producing IL-10. Using recipient Rag1(-/-) mice, hepatic Tr1-like cells from day 7 of HBV-persistent mice acquired the ability to inhibit anti-HBV immunity 3 d earlier than splenic Tr1-like cells, implying that hepatic Tr1-like cells were generated before those in spleen. Kupffer cell depletion or IL-10 deficiency led to impairment of Tr1-like cell generation, along with breaking HBV persistence. The purified EGFP(+)CD4(+) T cells (containing Tr1-like cells) from HBV-carrier mice trafficked in higher numbers to DLN in recipient mice after HBsAg vaccination, and subsequently inactivated both Tfh cells and GC B cells via secreting IL-10, resulting in impaired GC formation and anti-HB antibody production. Thus, our results indicate Tr1-like cells migrate from the liver to the DLN and inhibit peripheral anti-HBV immunity by negatively regulating GC B cells and Tfh cells. PMID:24089450

  5. Three-Dimensional Culture Assay to Explore Cancer Cell Invasiveness and Satellite Tumor Formation.

    Science.gov (United States)

    Côté, Marie-France; Turcotte, Audrey; Doillon, Charles; Gobeil, Stephane

    2016-01-01

    Mammalian cell culture in monolayers is widely used to study various physiological and molecular processes. However, this approach to study growing cells often generates unwanted artifacts. Therefore, cell culture in a three-dimensional (3D) environment, often using extracellular matrix components, emerged as an interesting alternative due to its close similarity to the native in vivo tissue or organ. We developed a 3D cell culture system using two compartments, namely (i) a central compartment containing cancer cells embedded in a collagen gel acting as a pseudo-primary macrospherical tumor and (ii) a peripheral cell-free compartment made of a fibrin gel, i.e. an extracellular matrix component different from that used in the center, in which cancer cells can migrate (invasion front) and/or form microspherical tumors representing secondary or satellite tumors. The formation of satellite tumors in the peripheral compartment is remarkably correlated to the known aggressiveness or metastatic origin of the native tumor cells, which makes this 3D culture system unique. This cell culture approach might be considered to assess cancer cell invasiveness and motility, cell-extracellular matrix interactions and as a method to evaluate anti-cancer drug properties. PMID:27585303

  6. Formation of Nanoscale Bioimprints of Muscle Cells Using UV-Cured Spin-Coated Polymers

    Directory of Open Access Journals (Sweden)

    Fahmi Samsuri

    2009-01-01

    Full Text Available We report a nanoscale replication method suitable for biological specimens that has potential in single cell studies and in formation of 3D biocompatible scaffolds. Earlier studies using a heat-curable polydimethylsiloxane (PDMS or a UV-curable elastomer introduced Bioimprint replication to facilitate cell imaging. However, the replicating conditions for thermal polymerization are known to cause cell dehydration during curing. In this study, a UV-cured methacrylate copolymer was developed for use in creating replicas of living cells and was tested on rat muscle cells. Bioimprints of muscle cells were formed by spin coating under UV irradiation. The polymer replicas were then separated from the muscle cells and were analyzed under an Atomic Force Microscope (AFM, in tapping mode, because it has low tip-sample forces and thus will not destroy the fine structures of the imprint. The new polymer is biocompatible with higher replication resolution and has a faster curing process than other types of silicon-based organic polymers such as PDMS. High resolution images of the muscle cell imprints showed the micro-and nanostructures of the muscle cells, including cellular fibers and structures within the cell membranes. The AFM is able to image features at nanoscale resolution with the potential for recognizing abnormalities on cell membranes at early stages of disease progression.

  7. Effects of gradient magnetic force and diamagnetic torque on formation of osteoclast-like giant cell

    Energy Technology Data Exchange (ETDEWEB)

    Iwasaka, M [Department of Medical System Engineering, Graduate School of Engineering, Chiba University, Chiba 263-8522 (Japan); Ikehata, M [Railway Technology Institute, 2-8-38 Hikari-cho, Kokubunji-shi, Tokyo 185-8540 (Japan); Hirota, N [National Institute for Materials Science, Sengenl-2-1, Tsukuba 305-0047 (Japan)], E-mail: iwasaka@faculty.chiba-u.jp, E-mail: iwasaka-m@umin.ac.jp

    2009-03-01

    In bone tissue, two kinds of cells, osteoblast (OB) and osteoclast (OC), contribute to remodeling of bone. In the present study, a co-culture system of bone-forming cell (OB) and -dissolving cell (OC) was incubated in static magnetic fields of horizontal 14 T and vertical gradient 10 T. Effect of two kinds of magnetic fields was an inhibition of OC formation. Three kinds of mechanisms, magnetic orientation of OB, diamagnetic torque force acting on OC, and possible reduction of earth's gravity were discussed.

  8. Effects of gradient magnetic force and diamagnetic torque on formation of osteoclast-like giant cell

    Science.gov (United States)

    Iwasaka, M.; Ikehata, M.; Hirota, N.

    2009-03-01

    In bone tissue, two kinds of cells, osteoblast (OB) and osteoclast (OC), contribute to remodeling of bone. In the present study, a co-culture system of bone-forming cell (OB) and -dissolving cell (OC) was incubated in static magnetic fields of horizontal 14 T and vertical gradient 10 T. Effect of two kinds of magnetic fields was an inhibition of OC formation. Three kinds of mechanisms, magnetic orientation of OB, diamagnetic torque force acting on OC, and possible reduction of earth's gravity were discussed.

  9. Saltatory formation, sliding and dissolution of ER–PM junctions in migrating cancer cells

    OpenAIRE

    Dingsdale, Hayley; Okeke, Emmanuel; Awais, Muhammad; Haynes, Lee; Criddle, David N.; Sutton, Robert; Tepikin, Alexei V.

    2013-01-01

    We demonstrated three novel forms of dynamic behaviour of junctions between the ER (endoplasmic reticulum) and the PM (plasma membrane) in migrating cancer cells: saltatory formation, long-distance sliding and dissolution. The individual ER–PM junctions formed near the leading edge of migrating cells (usually within 0.5 μm of polymerized actin and close to focal adhesions) and appeared suddenly without sliding from the interior of the cell. The long distance sliding and dissolution of ER–PM j...

  10. Context and location dependence of adaptive Foxp3+ regulatory T cell formation during immunopathological conditions

    OpenAIRE

    Heiber, Joshua F.; Geiger, Terrence L.

    2012-01-01

    Circulating Foxp3+ regulatory T cells (Treg) may arise in the thymus (natural Treg, nTreg) or through the adaptive upregulation of Foxp3 after T cell activation (induced Treg, iTreg). In this brief review, we explore evidence for the formation and function of iTreg during pathologic conditions. Determining the ontogeny and function of Treg populations has relied on the use of manipulated systems in which either iTreg or nTreg are absent, or lineage tracing of T cell clones through repertoire ...

  11. Selective ablation of the androgen receptor in mouse sertoli cells affects sertoli cell maturation, barrier formation and cytoskeletal development.

    Directory of Open Access Journals (Sweden)

    Ariane Willems

    Full Text Available The observation that mice with a selective ablation of the androgen receptor (AR in Sertoli cells (SC (SCARKO mice display a complete block in meiosis supports the contention that SC play a pivotal role in the control of germ cell development by androgens. To delineate the physiological and molecular mechanism responsible for this control, we compared tubular development in pubertal SCARKO mice and littermate controls. Particular attention was paid to differences in SC maturation, SC barrier formation and cytoskeletal organization and to the molecular mediators potentially involved. Functional analysis of SC barrier development by hypertonic perfusion and lanthanum permeation techniques and immunohistochemical analysis of junction formation showed that SCARKO mice still attempt to produce a barrier separating basal and adluminal compartment but that barrier formation is delayed and defective. Defective barrier formation was accompanied by disturbances in SC nuclear maturation (immature shape, absence of prominent, tripartite nucleoli and SC polarization (aberrant positioning of SC nuclei and cytoskeletal elements such as vimentin. Quantitative RT-PCR was used to study the transcript levels of genes potentially related to the described phenomena between day 8 and 35. Differences in the expression of SC genes known to play a role in junction formation could be shown from day 8 for Cldn11, from day 15 for Cldn3 and Espn, from day 20 for Cdh2 and Jam3 and from day 35 for ZO-1. Marked differences were also noted in the transcript levels of several genes that are also related to cell adhesion and cytoskeletal dynamics but that have not yet been studied in SC (Actn3, Ank3, Anxa9, Scin, Emb, Mpzl2. It is concluded that absence of a functional AR in SC impedes the remodeling of testicular tubules expected at the onset of spermatogenesis and interferes with the creation of the specific environment needed for germ cell development.

  12. Machine-Part cell formation through visual decipherable clustering of Self Organizing Map

    CERN Document Server

    Chattopadhyay, Manojit; Dan, Pranab K; 10.1007/s00170-010-2802-4

    2011-01-01

    Machine-part cell formation is used in cellular manufacturing in order to process a large variety, quality, lower work in process levels, reducing manufacturing lead-time and customer response time while retaining flexibility for new products. This paper presents a new and novel approach for obtaining machine cells and part families. In the cellular manufacturing the fundamental problem is the formation of part families and machine cells. The present paper deals with the Self Organising Map (SOM) method an unsupervised learning algorithm in Artificial Intelligence, and has been used as a visually decipherable clustering tool of machine-part cell formation. The objective of the paper is to cluster the binary machine-part matrix through visually decipherable cluster of SOM color-coding and labelling via the SOM map nodes in such a way that the part families are processed in that machine cells. The Umatrix, component plane, principal component projection, scatter plot and histogram of SOM have been reported in t...

  13. The pesticide methoxychlor decreases myotube formation in cell culture by slowing myoblast proliferation.

    Science.gov (United States)

    Steffens, Bradley W; Batia, Lyn M; Baarson, Chad J; Choi, Chang-Kun Charles; Grow, Wade A

    2007-08-01

    We studied the effect of the estrogenic pesticide methoxychlor (MXC) on skeletal muscle development using C2C12 cell culture. Myoblast cultures were exposed to various concentrations of MXC at various times during the process of myoblast fusion into myotubes. We observed that MXC exposure decreased myotube formation. In addition, we observed myoblasts with cytoplasmic vacuoles in cultures exposed to MXC. Because cytoplasmic vacuoles can be characteristic of cell death, apoptosis assays and trypan blue exclusion assays were performed. We found no difference in the frequency of apoptosis or in the frequency of cell death for cultures exposed to MXC and untreated cultures. Collectively, these results indicate that MXC exposure decreases myotube formation without causing cell death. In contrast, when cell proliferation was assessed, untreated cultures had a myoblast proliferation rate 50% greater than cultures exposed to MXC. We conclude that MXC decreases myotube formation at least in part by slowing myoblast proliferation. Furthermore, we suggest that direct exposure to MXC could affect skeletal muscle development in animals or humans, in addition to the defects in reproductive development that have previously been reported.

  14. Toxin YafQ increases persister cell formation by reducing indole signalling.

    Science.gov (United States)

    Hu, Ying; Kwan, Brian W; Osbourne, Devon O; Benedik, Michael J; Wood, Thomas K

    2015-04-01

    Persister cells survive antibiotic and other environmental stresses by slowing metabolism. Since toxins of toxin/antitoxin (TA) systems have been postulated to be responsible for persister cell formation, we investigated the influence of toxin YafQ of the YafQ/DinJ Escherichia coli TA system on persister cell formation. Under stress, YafQ alters metabolism by cleaving transcripts with in-frame 5'-AAA-G/A-3' sites. Production of YafQ increased persister cell formation with multiple antibiotics, and by investigating changes in protein expression, we found that YafQ reduced tryptophanase levels (TnaA mRNA has 16 putative YafQ cleavage sites). Consistently, TnaA mRNA levels were also reduced by YafQ. Tryptophanase is activated in the stationary phase by the stationary-phase sigma factor RpoS, which was also reduced dramatically upon production of YafQ. Tryptophanase converts tryptophan into indole, and as expected, indole levels were reduced by the production of YafQ. Corroborating the effect of YafQ on persistence, addition of indole reduced persistence. Furthermore, persistence increased upon deleting tnaA, and persistence decreased upon adding tryptophan to the medium to increase indole levels. Also, YafQ production had a much smaller effect on persistence in a strain unable to produce indole. Therefore, YafQ increases persistence by reducing indole, and TA systems are related to cell signalling.

  15. Role of VLDL Receptor in the Process of Foam Cell Formation

    Institute of Scientific and Technical Information of China (English)

    屈伸; 吴凡; 田俊; 李映红; 王燕; 王宇哲; 宗义强

    2004-01-01

    Summary: The role of very low density lipoprotein receptor (LVLDR) in the process of foam cell formation was investigated. After the primary cultured mouse peritoneal macrophages were incubated with VLDL, β-VLDL or low density lipoprotein (LDL), respectively for 24 h and 48 h, foam cells formation was identified by oil red O staining and cellular contents of triglyceride (TG) and total cholesterol (TC) were determined. The mRNA levels of LDLR, LDLR related protein (LRP)and VLDLR were detected by semi-quantitative RT-PCR. The results demonstrated that VLDL, βVLDL and LDL could increase the contents of TG and TC in macrophages. Cells treated with VLDL or β-VLDL showed markedly increased expression of VLDLR and decreased expression of LDLR, whereas LRP was up-regulated slightly. For identifying the effect of VLDL receptor on cellular lipid accumulation, Idl-A7-VR cells, which expresses VLDLR and trace amount of LRP without functional LDLR, was used to incubate with lipoproteins for further examination. The results elucidated that the uptake of triglyceride-rich lipoprotein mediated by VLDLR plays an important role in accumulation of lipid and the formation of foam cells.

  16. PPARy phosphorylation mediated by JNK MAPK: a potential role in macrophage-derived foam cell formation

    Institute of Scientific and Technical Information of China (English)

    Ran YIN; Yu-gang DONG; Hong-lang LI

    2006-01-01

    Aim: To investigate whether oxidized low-density lipoprotein (ox-LDL) modulates peroxisome proliferator-activated receptor γ (PPARγ) activity through phosphorylation in macrophages, and the effect of PPARy phosphorylation on macrophages-derived foam cell formation. Methods: After exposing the cultured THP-1 cells to ox-LDL in the presence or absence of different mitogen-activated protein kinase (MAPK) inhibitors, PPARγ and phosphorylated PPARγ protein levels were detected by Western blot. MAPK activity was analyzed using MAP Kinase Assay Kit. Intracellular cholesterol accumulation was assessed by Oil red O staining and cholesterol oxidase enzymatic method. The Mrna level of PPARγ target gene was determined by reverse transcription-polymerase chain reaction (RT-PCR). Results: ox-LDL evaluated PPARγ phosphorylation status and subsequently decreased PPARγ target gene expression in a dose-dependent manner. Ox-LDL also induced MAPK activation. Treatment of THP-1 cells with c-Jun N-terminal kinase-, but not p38- or extracellular signal-regulated kinase-MAPK inhibitor, significantly suppressed PPARγ phosphorylation induced by ox-LDL, which in turn inhibited foam cell formation. Conclusion: In addition to its ligand-dependent activation, ox-LDL modulates PPARγ activity through phosphorylation, which is mediated by MAPK activation. PPARγ phosphorylation mediated by MAPK facilitates foam cell formation from macrophages exposed to ox-LDL.

  17. Ion bombardment induced formation of micro-craters in plant cell envelopes

    International Nuclear Information System (INIS)

    Ion beam bombardment of biological material has been recently applied for gene transfer in both plant and bacterial cells. A consistent physical mechanism for this significant result has not yet been developed. A fundamental question about the mechanism is the possible formation of pathways due to ion bombardment that are responsible for the gene transfer. We have carried out investigations of the effects of low-energy bombardment by both gaseous and metallic ion species of onion skin cells on their surface microstructure. Our experimental results reveal evidence demonstrating that the formation of micro-crater-like structures on the plant cell envelope surface is a general phenomenon consequent to ion bombardment, no matter what ion species, under certain ion beam conditions. The micro-craters are about 0.1-1 μm in size (diameter) and a few tens of nanometers in depth. The micro-crater formation process seems to be unrelated to the chemical composition of and rapid water evaporation from the cell envelope, but is associated with the special microstructure of the cell wall

  18. Vicenistatin induces early endosome-derived vacuole formation in mammalian cells.

    Science.gov (United States)

    Nishiyama, Yuko; Ohmichi, Tomohiro; Kazami, Sayaka; Iwasaki, Hiroki; Mano, Kousuke; Nagumo, Yoko; Kudo, Fumitaka; Ichikawa, Sosaku; Iwabuchi, Yoshiharu; Kanoh, Naoki; Eguchi, Tadashi; Osada, Hiroyuki; Usui, Takeo

    2016-05-01

    Homotypic fusion of early endosomes is important for efficient protein trafficking and sorting. The key controller of this process is Rab5 which regulates several effectors and PtdInsPs levels, but whose mechanisms are largely unknown. Here, we report that vicenistatin, a natural product, enhanced homotypic fusion of early endosomes and induced the formation of large vacuole-like structures in mammalian cells. Unlike YM201636, another early endosome vacuolating compound, vicenistatin did not inhibit PIKfyve activity in vitro but activated Rab5-PAS pathway in cells. Furthermore, vicenistatin increased the membrane surface fluidity of cholesterol-containing liposomes in vitro, and cholesterol deprivation from the plasma membrane stimulated vicenistatin-induced vacuolation in cells. These results suggest that vicenistatin is a novel compound that induces the formation of vacuole-like structures by activating Rab5-PAS pathway and increasing membrane fluidity. PMID:27104762

  19. The diaphanous-related formins promote protrusion formation and cell-to-cell spread of Listeria monocytogenes.

    Science.gov (United States)

    Fattouh, Ramzi; Kwon, Hyunwoo; Czuczman, Mark A; Copeland, John W; Pelletier, Laurence; Quinlan, Margot E; Muise, Aleixo M; Higgins, Darren E; Brumell, John H

    2015-04-01

    The Gram-positive bacterium Listeria monocytogenes is a facultative intracellular pathogen whose virulence depends on its ability to spread from cell to cell within an infected host. Although the actin-related protein 2/3 (Arp2/3) complex is necessary and sufficient for Listeria actin tail assembly, previous studies suggest that other actin polymerization factors, such as formins, may participate in protrusion formation. Here, we show that Arp2/3 localized to only a minor portion of the protrusion. Moreover, treatment of L. monocytogenes-infected HeLa cells with a formin FH2-domain inhibitor significantly reduced protrusion length. In addition, the Diaphanous-related formins 1-3 (mDia1-3) localized to protrusions, and knockdown of mDia1, mDia2, and mDia3 substantially decreased cell-to-cell spread of L. monocytogenes. Rho GTPases are known to be involved in formin activation. Our studies also show that knockdown of several Rho family members significantly influenced bacterial cell-to-cell spread. Collectively, these findings identify a Rho GTPase-formin network that is critically involved in the cell-to-cell spread of L. monocytogenes. PMID:25281757

  20. Mic13 Is Essential for Formation of Crista Junctions in Mammalian Cells

    Science.gov (United States)

    Anand, Ruchika; Strecker, Valentina; Urbach, Jennifer; Wittig, Ilka; Reichert, Andreas S.

    2016-01-01

    Mitochondrial cristae are connected to the inner boundary membrane via crista junctions which are implicated in the regulation of oxidative phosphorylation, apoptosis, and import of lipids and proteins. The MICOS complex determines formation of crista junctions. We performed complexome profiling and identified Mic13, also termed Qil1, as a subunit of the MICOS complex. We show that MIC13 is an inner membrane protein physically interacting with MIC60, a central subunit of the MICOS complex. Using the CRISPR/Cas method we generated the first cell line deleted for MIC13. These knockout cells show a complete loss of crista junctions demonstrating that MIC13 is strictly required for the formation of crista junctions. MIC13 is required for the assembly of MIC10, MIC26, and MIC27 into the MICOS complex. However, it is not needed for the formation of the MIC60/MIC19/MIC25 subcomplex suggesting that the latter is not sufficient for crista junction formation. MIC13 is also dispensable for assembly of respiratory chain complexes and for maintaining mitochondrial network morphology. Still, lack of MIC13 resulted in a moderate reduction of mitochondrial respiration. In summary, we show that MIC13 has a fundamental role in crista junction formation and that assembly of respiratory chain supercomplexes is independent of mitochondrial cristae shape. PMID:27479602

  1. Ice formation in PEM fuel cells operated isothermally at sub-freezing temperatures

    Energy Technology Data Exchange (ETDEWEB)

    Mukundan, Rangachary [Los Alamos National Laboratory; Luhan, Roger W [Los Alamos National Laboratory; Davey, John R [Los Alamos National Laboratory; Spendelow, Jacob S [Los Alamos National Laboratory; Borup, Rodney L [Los Alamos National Laboratory; Hussey, Daniel S [NIST; Jacobson, David L [NIST; Arif, Muhammad [NIST

    2009-01-01

    The effect of MEA and GDL structure and composition on the performance of single-PEM fuel cells operated isothermally at subfreezing temperatures is presented. The cell performance and durability are not only dependent on the MEA/GDL materials used but also on their interfaces. When a cell is operated isothermally at sub-freezing temperatures in constant current mode, the water formation due to the current density initially hydrates the membrane/ionomer and then forms ice in the catalyst layer/GDL. An increase in high frequency resistance was also observed in certain MEAs where there is a possibility of ice formation between the catalyst layer and GDL leading to a loss in contact area. The total water/ice holding capacity for any MEA was lower at lower temperatures and higher current densities. The durability of MEAs subjected to multiple isothermal starts was better for LANL prepared MEAs as compared to commercial MEAs, and cloth GDLs when compared to paper GDLs. The ice formation was monitored using high-resolution neutron radiography and was found to be concentrated near the cathode catalyst layer. However, there was significant ice formation in the GDLs especially at the higher temperature ({approx} -10 C) and lower current density (0.02 A/cm{sup 2}) operations. These results are consistent with the longer-term durability observations that show more severe degradation at the lower temperatures.

  2. Swarming and complex pattern formation in Paenibacillus vortex studied by imaging and tracking cells

    Directory of Open Access Journals (Sweden)

    Jacob Eshel

    2008-02-01

    Full Text Available Abstract Background Swarming motility allows microorganisms to move rapidly over surfaces. The Gram-positive bacterium Paenibacillus vortex exhibits advanced cooperative motility on agar plates resulting in intricate colonial patterns with geometries that are highly sensitive to the environment. The cellular mechanisms that underpin the complex multicellular organization of such a simple organism are not well understood. Results Swarming by P. vortex was studied by real-time light microscopy, by in situ scanning electron microscopy and by tracking the spread of antibiotic-resistant cells within antibiotic-sensitive colonies. When swarming, P. vortex was found to be peritrichously flagellated. Swarming by the curved cells of P. vortex occurred on an extremely wide range of media and agar concentrations (0.3 to 2.2% w/v. At high agar concentrations (> 1% w/v rotating colonies formed that could be detached from the main mass of cells by withdrawal of cells into the latter. On lower percentage agars, cells moved in an extended network composed of interconnected "snakes" with short-term collision avoidance and sensitivity to extracts from swarming cells. P. vortex formed single Petri dish-wide "supercolonies" with a colony-wide exchange of motile cells. Swarming cells were coupled by rapidly forming, reversible and non-rigid connections to form a loose raft, apparently connected via flagella. Inhibitors of swarming (p-Nitrophenylglycerol and Congo Red were identified. Mitomycin C was used to trigger filamentation without inhibiting growth or swarming; this facilitated dissection of the detail of swarming. Mitomycin C treatment resulted in malcoordinated swarming and abortive side branch formation and a strong tendency by a subpopulation of the cells to form minimal rotating aggregates of only a few cells. Conclusion P. vortex creates complex macroscopic colonies within which there is considerable reflux and movement and interaction of cells. Cell

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

    International Nuclear Information System (INIS)

    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 TiO2 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 TiO2 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 TiO2 nanoparticles

  4. 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.

  5. Adenosine formation in contracting primary rat skeletal muscle cells and endothelial cells in culture

    DEFF Research Database (Denmark)

    Hellsten, Ylva; Frandsen, Ulrik

    1997-01-01

    in the extracellular adenosine concentration (421 +/- 91 and 235 +/- 30 nmol (g protein)-1, respectively; P muscle cells (161 +/- 20 nmol (g protein)-1). The ATP concentration was lower (18%; P muscle cells...... found to have ecto-forms of several enzymes involved in nucleotide metabolism, including ATPases capable of converting extracellular ATP to ADP and AMP. 5. Adenosine added to the cell medium was taken up by muscle cells and incorporated into the adenine nucleotide pool so that after 30 min of incubation......, over 95% of the adenosine label was present in ATP, ADP and AMP. A similar extent of incorporation of adenosine into the nucleotide pool was evident in the endothelial cells. 6. The present data suggest that contracting muscle cells induce an elevation in the extracellular adenosine concentration...

  6. Differentiation of mouse embryonic stem cells into endoderm without embryoid body formation.

    Directory of Open Access Journals (Sweden)

    Peter T W Kim

    Full Text Available Pluripotent embryonic stem cells hold a great promise as an unlimited source of tissue for treatment of chronic diseases such as Type 1 diabetes. Herein, we describe a protocol using all-trans-retinoic acid, basic fibroblast growth factor and dibutyryl cAMP (DBcAMP in the absence of embryoid body formation, for differentiation of murine embryonic stem cells into definitive endoderm that may serve as pancreatic precursors. The produced cells were analyzed by quantitative PCR, immunohistochemistry and static insulin release assay for markers of trilaminar embryo, and pancreas. Differentiated cells displayed increased Sox17 and Foxa2 expression consistent with definitive endoderm production. There was minimal production of Sox7, an extraembryonic endoderm marker, and Oct4, a marker of pluripotency. There was minimal mesoderm or neuroectoderm formation based on expression levels of the markers brachyury and Sox1, respectively. Various assays revealed that the cell clusters generated by this protocol express markers of the pancreatic lineage including insulin I, insulin II, C-peptide, PDX-1, carboxypeptidase E, pan-cytokeratin, amylase, glucagon, PAX6, Ngn3 and Nkx6.1. This protocol using all-trans-retinoic acid, DBcAMP, in the absence of embryoid bodies, generated cells that have features of definitive endoderm that may serve as pancreatic endocrine precursors.

  7. Primary Phenomenon in the Network Formation of Endothelial Cells: Effect of Charge.

    Science.gov (United States)

    Arai, Shunto

    2015-12-07

    Blood vessels are essential organs that are involved in the supply of nutrients and oxygen and play an important role in regulating the body's internal environment, including pH, body temperature, and water homeostasis. Many studies have examined the formation of networks of endothelial cells. The results of these studies have revealed that vascular endothelial growth factor (VEGF) affects the interactions of these cells and modulates the network structure. Though almost all previous simulation studies have assumed that the chemoattractant VEGF is present before network formation, vascular endothelial cells secrete VEGF only after the cells bind to the substrate. This suggests VEGF is not essential for vasculogenesis especially at the early stage. Using a simple experiment, we find chain-like structures which last quite longer than it is expected, unless the energetically stable cluster should be compact. Using a purely physical model and simulation, we find that the hydrodynamic interaction retard the compaction of clusters and that the chains are stabilized through the effects of charge. The charge at the surface of the cells affect the interparticle potential, and the resulting repulsive forces prevent the chains from folding. The ions surrounding the cells may also be involved in this process.

  8. Neocartilage formation from mesenchymal stem cells grown in type II collagen-hyaluronan composite scaffolds.

    Science.gov (United States)

    Yeh, Hsi-Yi; Lin, Ting-Yu; Lin, Chen-Huan; Yen, B Linju; Tsai, Ching-Lin; Hsu, Shan-Hui

    2013-01-01

    Three-dimensional (3D) collagen type II-hyaluronan (HA) composite scaffolds (CII-HA) which mimics the extracellular environment of natural cartilage were fabricated in this study. Rheological measurements demonstrated that the incorporation of HA increased the compression modulus of the scaffolds. An initial in vitro evaluation showed that scaffolds seeded with porcine chondrocytes formed cartilaginous-like tissue after 8 weeks, and HA functioned to promote the growth of chondrocytes into scaffolds. Placenta-derived multipotent cells (PDMC) and gingival fibroblasts (GF) were seeded on tissue culture polystyrene (TCPS), CII-HA films, and small intestinal submucosa (SIS) sheets for comparing their chondrogenesis differentiation potentials with those of adipose-derived adult stem cells (ADAS) and bone marrow-derived mesenchymal stem cells (BMSC). Among different cells, PDMC showed the greatest chondrogenic differentiation potential on both CII-HA films and SIS sheets upon TGF-β3 induction, followed by GF. This was evidenced by the up-regulation of chondrogenic genes (Sox9, aggrecan, and collagen type II), which was not observed for cells grown on TCPS. This finding suggested the essential role of substrate materials in the chondrogenic differentiation of PDMC and GF. Neocartilage formation was more obvious in both PDMC and GF cells plated on CII-HA composite scaffolds vs. 8-layer SIS at 28 days in vitro. Finally, implantation of PDMC/CII-HA constructs into NOD-SCID mice confirmed the formation of tissue-engineered cartilage in vivo.

  9. Serum amyloid A stimulates macrophage foam cell formation via lectin-like oxidized low-density lipoprotein receptor 1 upregulation

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Ha Young, E-mail: hayoung@skku.edu [Department of Biological Science, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Mitochondria Hub Regulation Center, Dong-A University, Busan 602-714 (Korea, Republic of); Kim, Sang Doo [Department of Biological Science, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Baek, Suk-Hwan [Department of Biochemistry and Molecular Biology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Choi, Joon Hyuk [Department of Pathology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Cho, Kyung-Hyun [School of Biotechnology, Yeungnam University, Gyeongsan 712-749 (Korea, Republic of); Zabel, Brian A. [Palo Alto Institute for Research and Education, Veterans Affairs Hospital, Palo Alto, CA 94304 (United States); Bae, Yoe-Sik, E-mail: yoesik@skku.edu [Department of Biological Science, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Mitochondria Hub Regulation Center, Dong-A University, Busan 602-714 (Korea, Republic of); Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University, Seoul 135-710 (Korea, Republic of)

    2013-03-29

    Highlights: ► SAA induced macrophage foam cell formation. ► SAA stimulated upregulation of lectin-like oxidized low-density lipoprotein receptor 1 (LOX1). ► SAA-induced LOX1 expression and foam cell formation is mediated by JNK/NF-κB signaling. ► HDL-conjugated SAA also stimulates foam cell formation via LOX1 upregulation. ► The finding reveals a novel mechanism of action of SAA in the pathogenesis of atherosclerosis. -- Abstract: Elevated levels of serum amyloid A (SAA) is a risk factor for cardiovascular diseases, however, the role of SAA in the pathophysiology of atherosclerosis remains unclear. Here we show that SAA induced macrophage foam cell formation. SAA-stimulated foam cell formation was mediated by c-jun N-terminal kinase (JNK) signaling. Moreover, both SAA and SAA-conjugated high density lipoprotein stimulated the expression of the important scavenger receptor lectin-like oxidized low-density lipoprotein receptor 1 (LOX1) via nuclear factor-κB (NF-κB). A LOX1 antagonist carrageenan significantly blocked SAA-induced foam cell formation, indicating that SAA promotes foam cell formation via LOX1 expression. Our findings therefore suggest that SAA stimulates foam cell formation via LOX1 induction, and thus likely contributes to atherogenesis.

  10. The Diaphanous-Related Formins Promote Protrusion Formation and Cell-to-Cell Spread of Listeria monocytogenes

    OpenAIRE

    Fattouh, Ramzi; Kwon, Hyunwoo; Czuczman, Mark A.; Copeland, John W.; Pelletier, Laurence; Quinlan, Margot E.; Aleixo M Muise; Higgins, Darren E.; Brumell, John H.

    2014-01-01

    The Gram-positive bacterium Listeria monocytogenes is a facultative intracellular pathogen whose virulence depends on its ability to spread from cell to cell within an infected host. Although the actin-related protein 2/3 (Arp2/3) complex is necessary and sufficient for Listeria actin tail assembly, previous studies suggest that other actin polymerization factors, such as formins, may participate in protrusion formation. Here, we show that Arp2/3 localized to only a minor portion of the protr...

  11. Formation of multinuclear cells induced by dimethyl sulfoxide: inhibition of cytokinesis and occurrence of novel nuclear division in dictyostelium cells

    OpenAIRE

    Fukui, Y.

    1980-01-01

    Our previous studies showed that 10 percent dimethyl sulfoxide (DMSO) induces the formation of actin microfilament bundles in the cell nucleus together with the dislocation of cortical microfilaments from the plasma membrane. The present study investigated the effects of DMSO on diverse activities mediated by cellular microfilaments as the second step toward assessing potential differences between nuclear and cytoplasmic actins of dictyostelium mucoroides. DMSO was found to reversibly inhibit...

  12. Knockdown of the cell cycle inhibitor p21 enhances cartilage formation by induced pluripotent stem cells.

    Science.gov (United States)

    Diekman, Brian O; Thakore, Pratiksha I; O'Connor, Shannon K; Willard, Vincent P; Brunger, Jonathan M; Christoforou, Nicolas; Leong, Kam W; Gersbach, Charles A; Guilak, Farshid

    2015-04-01

    The limited regenerative capacity of articular cartilage contributes to progressive joint dysfunction associated with cartilage injury or osteoarthritis. Cartilage tissue engineering seeks to provide a biological substitute for repairing damaged or diseased cartilage, but requires a cell source with the capacity for extensive expansion without loss of chondrogenic potential. In this study, we hypothesized that decreased expression of the cell cycle inhibitor p21 would enhance the proliferative and chondrogenic potential of differentiated induced pluripotent stem cells (iPSCs). Murine iPSCs were directed to differentiate toward the chondrogenic lineage with an established protocol and then engineered to express a short hairpin RNA (shRNA) to reduce the expression of p21. Cells expressing the p21 shRNA demonstrated higher proliferative potential during monolayer expansion and increased synthesis of glycosaminoglycans (GAGs) in pellet cultures. Furthermore, these cells could be expanded ∼150-fold over three additional passages without a reduction in the subsequent production of GAGs, while control cells showed reduced potential for GAG synthesis with three additional passages. In pellets from extensively passaged cells, knockdown of p21 attenuated the sharp decrease in cell number that occurred in control cells, and immunohistochemical analysis showed that p21 knockdown limited the production of type I and type X collagen while maintaining synthesis of cartilage-specific type II collagen. These findings suggest that manipulating the cell cycle can augment the monolayer expansion and preserve the chondrogenic capacity of differentiated iPSCs, providing a strategy for enhancing iPSC-based cartilage tissue engineering.

  13. Cancer stem cell marker CD90 inhibits ovarian cancer formation via β3 integrin

    Science.gov (United States)

    Chen, Wei-Ching; Hsu, Hui-Ping; Li, Chung-Yen; Yang, Ya-Ju; Hung, Yu-Hsuan; Cho, Chien-Yu; Wang, Chih-Yang; Weng, Tzu-Yang; Lai, Ming-Derg

    2016-01-01

    Cancer stem cell (CSC) markers have been identified for CSC isolation and proposed as therapeutic targets in various types of cancers. CD90, one of the characterized markers in liver and gastric cancer, is shown to promote cancer formation. However, the underexpression level of CD90 in ovarian cancer cells and the evidence supporting the cellular mechanism have not been investigated. In the present study, we found that the DNA copy number of CD90 is correlated with mRNA expression in ovarian cancer tissue and the ovarian cancer patients with higher CD90 have good prognosis compared to the patients with lower CD90. Although the expression of CD90 in human ovarian cancer SKOV3 cells enhances the cell proliferation by MTT and anchorage-dependent growth assay, CD90 inhibits the anchorage-independent growth ability in vitro and tumor formation in vivo. CD90 overexpression suppresses the sphere-forming ability and ALDH activity and enhances the cell apoptosis, indicating that CD90 may reduce the cell growth by the properties of CSC and anoikis. Furthermore, CD90 reduces the expression of other CSC markers, including CD133 and CD24. The inhibition of CD133 is attenuated by the mutant CD90, which is replaced with RLE domain into RLD domain. Importantly, the CD90-regulated inhibition of CD133 expression, anchorage-independent growth and signal transduction of mTOR and AMPK are restored by the β3 integrin shRNA. Our results provide evidence that CD90 mediates the antitumor formation by interacting with β3 integrin, which provides new insight that can potentially be applied in the development of therapeutic strategies in ovarian cancer. PMID:27633757

  14. Sialic Acid on the Glycosylphosphatidylinositol Anchor Regulates PrP-mediated Cell Signaling and Prion Formation.

    Science.gov (United States)

    Bate, Clive; Nolan, William; Williams, Alun

    2016-01-01

    The prion diseases occur following the conversion of the cellular prion protein (PrP(C)) into disease-related isoforms (PrP(Sc)). In this study, the role of the glycosylphosphatidylinositol (GPI) anchor attached to PrP(C) in prion formation was examined using a cell painting technique. PrP(Sc) formation in two prion-infected neuronal cell lines (ScGT1 and ScN2a cells) and in scrapie-infected primary cortical neurons was increased following the introduction of PrP(C). In contrast, PrP(C) containing a GPI anchor from which the sialic acid had been removed (desialylated PrP(C)) was not converted to PrP(Sc). Furthermore, the presence of desialylated PrP(C) inhibited the production of PrP(Sc) within prion-infected cortical neurons and ScGT1 and ScN2a cells. The membrane rafts surrounding desialylated PrP(C) contained greater amounts of sialylated gangliosides and cholesterol than membrane rafts surrounding PrP(C). Desialylated PrP(C) was less sensitive to cholesterol depletion than PrP(C) and was not released from cells by treatment with glimepiride. The presence of desialylated PrP(C) in neurons caused the dissociation of cytoplasmic phospholipase A2 from PrP-containing membrane rafts and reduced the activation of cytoplasmic phospholipase A2. These findings show that the sialic acid moiety of the GPI attached to PrP(C) modifies local membrane microenvironments that are important in PrP-mediated cell signaling and PrP(Sc) formation. These results suggest that pharmacological modification of GPI glycosylation might constitute a novel therapeutic approach to prion diseases.

  15. Cytoprotective effect of imatinib mesylate in non-BCR-ABL-expressing cells along with autophagosome formation

    Energy Technology Data Exchange (ETDEWEB)

    Ohtomo, Tadashi [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan); Miyazawa, Keisuke, E-mail: miyazawa@tokyo-med.ac.jp [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan); Naito, Munekazu [Department of Anatomy, Tokyo Medical University, Tokyo (Japan); Moriya, Shota [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan); Kuroda, Masahiko [Department of Molecular Pathology, Tokyo Medical University, Tokyo (Japan); Itoh, Masahiro [Department of Anatomy, Tokyo Medical University, Tokyo (Japan); Tomoda, Akio [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan)

    2010-01-01

    Treatment with imatinib mesylate (IM) results in an increased viable cell number of non-BCR-ABL-expressing cell lines by inhibiting spontaneous apoptosis. Electron microscopy revealed an increase of autophagosomes in response to IM. IM attenuated the cytotoxic effect of cytosine arabinoside, as well as inhibiting cell death with serum-deprived culture. Cytoprotection with autophagosome formation by IM was observed in various leukemia and cancer cell lines as well as normal murine embryonic fibroblasts (MEFs). Complete inhibition of autophagy by knockdown of atg5 in the Tet-off atg5{sup -/-} MEF system attenuated the cytoprotective effect of IM, indicating that the effect is partially dependent on autophagy. However, cytoprotection by IM was not mediated through suppression of ROS production via mitophagy, ER stress via ribophagy, or proapoptotic function of ABL kinase. Although the target tyrosine kinase(s) of IM remains unclear, our data provide novel therapeutic possibilities of using IM for cytoprotection.

  16. Inhibiting actin depolymerization enhances osteoblast differentiation and bone formation in human stromal stem cells

    DEFF Research Database (Denmark)

    Chen, Li; Shi, Kaikai; Frary, Charles Edward;

    2015-01-01

    Remodeling of the actin cytoskeleton through actin dynamics is involved in a number of biological processes, but its role in human stromal (skeletal) stem cells (hMSCs) differentiation is poorly understood. In the present study, we demonstrated that stabilizing actin filaments by inhibiting gene...... expression of the two main actin depolymerizing factors (ADFs): Cofilin 1 (CFL1) and Destrin (DSTN) in hMSCs, enhanced cell viability and differentiation into osteoblastic cells (OB) in vitro, as well as heterotopic bone formation in vivo. Similarly, treating hMSC with Phalloidin, which is known to stabilize...... polymerized actin filaments, increased hMSCs viability and OB differentiation. Conversely, Cytocholasin D, an inhibitor of actin polymerization, reduced cell viability and inhibited OB differentiation of hMSC. At a molecular level, preventing Cofilin phosphorylation through inhibition of LIM domain kinase 1...

  17. Glioblastoma formation from cell population depleted of Prominin1-expressing cells.

    Directory of Open Access Journals (Sweden)

    Kenji Nishide

    Full Text Available Prominin1 (Prom1, also known as CD133 in human has been widely used as a marker for cancer stem cells (CSCs, which self-renew and are tumorigenic, in malignant tumors including glioblastoma multiforme (GBM. However, there is other evidence showing that Prom1-negative cancer cells also form tumors in vivo. Thus it remains controversial whether Prom1 is a bona fide marker for CSCs. To verify if Prom1-expressing cells are essential for tumorigenesis, we established a mouse line, whose Prom1-expressing cells can be eliminated conditionally by a Cre-inducible DTA gene on the Prom1 locus together with a tamoxifen-inducible CreER(TM, and generated glioma-initiating cells (GICs-LD by overexpressing both the SV40 Large T antigen and an oncogenic H-Ras(L61 in neural stem cells of the mouse line. We show here that the tamoxifen-treated GICs-LD (GICs-DTA form tumor-spheres in culture and transplantable GBM in vivo. Thus, our studies demonstrate that Prom1-expressing cells are dispensable for gliomagenesis in this mouse model.

  18. Co-delivery of cell-wall-forming enzymes in the same vesicle for coordinated fungal cell wall formation.

    Science.gov (United States)

    Schuster, Martin; Martin-Urdiroz, Magdalena; Higuchi, Yujiro; Hacker, Christian; Kilaru, Sreedhar; Gurr, Sarah J; Steinberg, Gero

    2016-01-01

    Fungal cells are surrounded by an extracellular cell wall. This complex matrix of proteins and polysaccharides protects against adverse stresses and determines the shape of fungal cells. The polysaccharides of the fungal wall include 1,3-β-glucan and chitin, which are synthesized by membrane-bound synthases at the growing cell tip. A hallmark of filamentous fungi is the class V chitin synthase, which carries a myosin-motor domain. In the corn smut fungus Ustilago maydis, the myosin-chitin synthase Mcs1 moves to the plasma membrane in secretory vesicles, being delivered by kinesin-1 and myosin-5. The myosin domain of Mcs1 enhances polar secretion by tethering vesicles at the site of exocytosis. It remains elusive, however, how other cell-wall-forming enzymes are delivered and how their activity is coordinated post secretion. Here, we show that the U. maydis class VII chitin synthase and 1,3-β-glucan synthase travel in Mcs1-containing vesicles, and that their apical secretion depends on Mcs1. Once in the plasma membrane, anchorage requires enzyme activity, which suggests co-synthesis of chitin and 1,3-β-glucan polysaccharides at sites of exocytosis. Thus, delivery of cell-wall-forming enzymes in Mcs1 vesicles ensures local foci of fungal cell wall formation. PMID:27563844

  19. A cell-regulatory mechanism involving feedback between contraction and tissue formation guides wound healing progression.

    Directory of Open Access Journals (Sweden)

    Clara Valero

    Full Text Available Wound healing is a process driven by cells. The ability of cells to sense mechanical stimuli from the extracellular matrix that surrounds them is used to regulate the forces that cells exert on the tissue. Stresses exerted by cells play a central role in wound contraction and have been broadly modelled. Traditionally, these stresses are assumed to be dependent on variables such as the extracellular matrix and cell or collagen densities. However, we postulate that cells are able to regulate the healing process through a mechanosensing mechanism regulated by the contraction that they exert. We propose that cells adjust the contraction level to determine the tissue functions regulating all main activities, such as proliferation, differentiation and matrix production. Hence, a closed-regulatory feedback loop is proposed between contraction and tissue formation. The model consists of a system of partial differential equations that simulates the evolution of fibroblasts, myofibroblasts, collagen and a generic growth factor, as well as the deformation of the extracellular matrix. This model is able to predict the wound healing outcome without requiring the addition of phenomenological laws to describe the time-dependent contraction evolution. We have reproduced two in vivo experiments to evaluate the predictive capacity of the model, and we conclude that there is feedback between the level of cell contraction and the tissue regenerated in the wound.

  20. SNCG shRNA suppressed breast cancer cell xenograft formation and growth in nude mice

    Institute of Scientific and Technical Information of China (English)

    SHEN Pei-hong; FAN Qing-xia; LI Yan-wei; ZHANG Wei; HE Xiao-kai; WANG Zhen; ZHANG Yun-han

    2011-01-01

    Background Overexpression of breast cancer-specific gene 1 (SNCG) is associated with poor prognosis in advanced breast cancer patients. This study aimed to determine the effects of SNCG knockdown in breast cancer cells by using small hairpin RNA (shRNA).Methods Four different SNCG shRNA oligonucleotides were designed and chemically synthesized to construct mammalian expression vectors. These vectors were then stably transfected into a breast cancer MCF-7 cell line to knockdown SNCG expression. After SNCG knockdown was confirmed, the stable cell lines were inoculated into nude mice. SNCG mRNA and protein expressions were analyzed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively in both the stable cell lines and xenografts.Results All four SNCG shRNA constructs significantly reduced SNCG mRNA and protein levels in MCF-7 cells, as compared to the unrelated sequence control shRNA and the liposome control mice (P<0.05). SNCG-knockdown MCF-7cells formed significantly smaller tumor masses than cells expressing the unrelated sequence control or the liposome control mice (P<0.05).Conclusion SNCG shRNA effectively suppressed breast cancer cell formation in vivo and may be a useful clinical strategy to control breast cancer.

  1. WD40-repeat proteins in plant cell wall formation: current evidence and research prospects

    Directory of Open Access Journals (Sweden)

    Gea eGuerriero

    2015-12-01

    Full Text Available The metabolic complexity of living organisms relies on supramolecular protein structures which ensure vital processes, such as signal transduction, transcription, translation and cell wall synthesis. In eukaryotes WD40-repeat (WDR proteins often function as molecular hubs mediating supramolecular interactions. WDR proteins may display a variety of interacting partners and participate in the assembly of complexes involved in distinct cellular functions. In plants, the formation of lignocellulosic biomass involves extensive synthesis of cell wall polysaccharides, a process that requires the assembly of large transmembrane enzyme complexes, intensive vesicle trafficking, interactions with the cytoskeleton, and coordinated gene expression. Because of their function as supramolecular hubs, WDR proteins could participate in each or any of these steps, although to date only few WDR proteins have been linked to the cell wall by experimental evidence. Nevertheless, several potential cell wall-related WDR proteins were recently identified using in silico aproaches, such as analyses of co-expression, interactome and conserved gene neighbourhood. Notably, some WDR genes are frequently genomic neighbours of genes coding for GT2-family polysaccharide synthases in eukaryotes, and this WDR-GT2 collinear microsynteny is detected in diverse taxa. In angiosperms, two WDR genes are collinear to cellulose synthase genes, CESAs, whereas in ascomycetous fungi several WDR genes are adjacent to chitin synthase genes, chs. In this Perspective we summarize and discuss experimental and in silico studies on the possible involvement of WDR proteins in plant cell wall formation. The prospects of biotechnological engineering for enhanced biomass production are discussed.

  2. Periostin secreted by mesenchymal stem cells supports tendon formation in an ectopic mouse model.

    Science.gov (United States)

    Noack, Sandra; Seiffart, Virginia; Willbold, Elmar; Laggies, Sandra; Winkel, Andreas; Shahab-Osterloh, Sandra; Flörkemeier, Thilo; Hertwig, Falk; Steinhoff, Christine; Nuber, Ulrike A; Gross, Gerhard; Hoffmann, Andrea

    2014-08-15

    True tendon regeneration in human patients remains a vision of musculoskeletal therapies. In comparison to other mesenchymal lineages the biology of tenogenic differentiation is barely understood. Specifically, easy and efficient protocols are lacking that might enable tendon cell and tissue differentiation based on adult (stem) cell sources. In the murine mesenchymal progenitor cell line C3H10T½, overexpression of the growth factor bone morphogenetic protein 2 (BMP2) and a constitutively active transcription factor, Smad8 L+MH2, mediates tendon cell differentiation in vitro and the formation of tendon-like tissue in vivo. We hypothesized that during this differentiation secreted factors involved in extracellular matrix formation exert a major impact on tendon development. Gene expression analyses revealed four genes encoding secreted factors that are notably upregulated: periostin, C-type lectin domain family 3 (member b), RNase A4, and follistatin-like 1. These factors have not previously been implicated in tendon biology. Among these, periostin showed a specific expression in tenocytes of adult mouse Achilles tendon and in chondrocytes within the nonmineralized fibrocartilage zone of the enthesis with the calcaneus. Overexpression of periostin alone or in combination with constitutively active BMP receptor type in human mesenchymal stem cells and subsequent implantation into ectopic sites in mice demonstrated a reproducible moderate tenogenic capacity that has not been described before. Therefore, periostin may belong to the factors contributing to the development of tenogenic tissue. PMID:24809660

  3. Intermittent high oxygen influences the formation of neural retinal tissue from human embryonic stem cells.

    Science.gov (United States)

    Gao, Lixiong; Chen, Xi; Zeng, Yuxiao; Li, Qiyou; Zou, Ting; Chen, Siyu; Wu, Qian; Fu, Caiyun; Xu, Haiwei; Yin, Zheng Qin

    2016-01-01

    The vertebrate retina is a highly multilayered nervous tissue with a large diversity of cellular components. With the development of stem cell technologies, human retinas can be generated in three-dimensional (3-D) culture in vitro. However, understanding the factors modulating key productive processes and the way that they influence development are far from clear. Oxygen, as the most essential element participating in metabolism, is a critical factor regulating organic development. In this study, using 3-D culture of human stem cells, we examined the effect of intermittent high oxygen treatment (40% O2) on the formation and cellular behavior of neural retinas (NR) in the embryonic body (EB). The volume of EB and number of proliferating cells increased significantly under 40% O2 on day 38, 50, and 62. Additionally, the ratio of PAX6+ cells within NR was significantly increased. The neural rosettes could only develop with correct apical-basal polarity under 40% O2. In addition, the generation, migration and maturation of retinal ganglion cells were enhanced under 40% O2. All of these results illustrated that 40% O2 strengthened the formation of NR in EB with characteristics similar to the in vivo state, suggesting that the hyperoxic state facilitated the retinal development in vitro. PMID:27435522

  4. H2O2-induced Leaf Cell Death and the Crosstalk of Reactive Nitric/Oxygen Species([F])

    Institute of Scientific and Technical Information of China (English)

    Yiqin Wang; Aihong Lin; Gary J.Loake; Chengcai Chu

    2013-01-01

    In plants,the chloroplast is the main reactive oxygen species (ROS) producing site under high light stress.Catalase (CAT),which decomposes hydrogen peroxide (H2O2),is one of the controlling enzymes that maintains leaf redox homeostasis.The catalase mutants with reduced leaf catalase activity from different plant species exhibit an H2O2-induced leaf cell death phenotype.This phenotype was differently affected by light intensity or photoperiod,which may be caused by plant species,leaf redox status or growth conditions.In the rice CAT mutant nitric oxide excess 1 (noe1),higher H2O2 levels induced the generation of nitric oxide (NO) and higher S-nitrosothiol (SNO) levels,suggesting that NO acts as an important endogenous mediator in H2O2-induced leaf cell death.As a free radical,NO could also react with other intracellular and extracellular targets and form a series of related molecules,collectively called reactive nitrogen species (RNS).Recent studies have revealed that both RNS and ROS are important partners in plant leaf cell death.Here,we summarize the recent progress on H2O2-induced leaf cell death and the crosstalk of RNS and ROS signals in the plant hypersensitive response (HR),leaf senescence,and other forms of leaf cell death triggered by diverse environmental conditions.

  5. Monocytes induce STAT3 activation in human mesenchymal stem cells to promote osteoblast formation.

    Directory of Open Access Journals (Sweden)

    Vicky Nicolaidou

    Full Text Available A major therapeutic challenge is how to replace bone once it is lost. Bone loss is a characteristic of chronic inflammatory and degenerative diseases such as rheumatoid arthritis and osteoporosis. Cells and cytokines of the immune system are known to regulate bone turnover by controlling the differentiation and activity of osteoclasts, the bone resorbing cells. However, less is known about the regulation of osteoblasts (OB, the bone forming cells. This study aimed to investigate whether immune cells also regulate OB differentiation. Using in vitro cell cultures of human bone marrow-derived mesenchymal stem cells (MSC, it was shown that monocytes/macrophages potently induced MSC differentiation into OBs. This was evident by increased alkaline phosphatase (ALP after 7 days and the formation of mineralised bone nodules at 21 days. This monocyte-induced osteogenic effect was mediated by cell contact with MSCs leading to the production of soluble factor(s by the monocytes. As a consequence of these interactions we observed a rapid activation of STAT3 in the MSCs. Gene profiling of STAT3 constitutively active (STAT3C infected MSCs using Illumina whole human genome arrays showed that Runx2 and ALP were up-regulated whilst DKK1 was down-regulated in response to STAT3 signalling. STAT3C also led to the up-regulation of the oncostatin M (OSM and LIF receptors. In the co-cultures, OSM that was produced by monocytes activated STAT3 in MSCs, and neutralising antibodies to OSM reduced ALP by 50%. These data indicate that OSM, in conjunction with other mediators, can drive MSC differentiation into OB. This study establishes a role for monocyte/macrophages as critical regulators of osteogenic differentiation via OSM production and the induction of STAT3 signalling in MSCs. Inducing the local activation of STAT3 in bone cells may be a valuable tool to increase bone formation in osteoporosis and arthritis, and in localised bone remodelling during fracture repair.

  6. Enhancement of hybridoma formation, clonability and cell proliferation in a nanoparticle-doped aqueous environment

    Directory of Open Access Journals (Sweden)

    Karnieli Ohad

    2008-01-01

    Full Text Available Abstract Background The isolation and production of human monoclonal antibodies is becoming an increasingly important pursuit as biopharmaceutical companies migrate their drug pipelines away from small organic molecules. As such, optimization of monoclonal antibody technologies is important, as this is becoming the new rate-limiting step for discovery and development of new pharmaceuticals. The major limitations of this system are the efficiency of isolating hybridoma clones, the process of stabilizing these clones and optimization of hybridoma cell secretion, especially for large-scale production. Many previous studies have demonstrated how perturbations in the aqueous environment can impact upon cell biology. In particular, radio frequency (RF irradiation of solutions can have dramatic effects on behavior of solutions, cells and in particular membrane proteins, although this effect decays following removal of the RF. Recently, it was shown that nanoparticle doping of RF irradiated water (NPD water produced a stabilized aqueous medium that maintained the characteristic properties of RF irradiated water for extended periods of time. Therefore, the ordering effect in water of the RF irradiation can now be studied in systems that required prolonged periods for analysis, such as eukaryotic cell culture. Since the formation of hybridoma cells involves the formation of a new membrane, a process that is affected by the surrounding aqueous environment, we tested these nanoparticle doped aqueous media formulations on hybridoma cell production. Results In this study, we tested the entire process of isolation and production of human monoclonal antibodies in NPD water as a means for further enhancing human monoclonal antibody isolation and production. Our results indicate an overall enhancement of hybridoma yield, viability, clonability and secretion. Furthermore, we have demonstrated that immortal cells proliferate faster whereas primary human fibroblasts

  7. A genetic algorithm for a bi-objective mathematical model for dynamic virtual cell formation problem

    Science.gov (United States)

    Moradgholi, Mostafa; Paydar, Mohammad Mahdi; Mahdavi, Iraj; Jouzdani, Javid

    2016-05-01

    Nowadays, with the increasing pressure of the competitive business environment and demand for diverse products, manufacturers are force to seek for solutions that reduce production costs and rise product quality. Cellular manufacturing system (CMS), as a means to this end, has been a point of attraction to both researchers and practitioners. Limitations of cell formation problem (CFP), as one of important topics in CMS, have led to the introduction of virtual CMS (VCMS). This research addresses a bi-objective dynamic virtual cell formation problem (DVCFP) with the objective of finding the optimal formation of cells, considering the material handling costs, fixed machine installation costs and variable production costs of machines and workforce. Furthermore, we consider different skills on different machines in workforce assignment in a multi-period planning horizon. The bi-objective model is transformed to a single-objective fuzzy goal programming model and to show its performance; numerical examples are solved using the LINGO software. In addition, genetic algorithm (GA) is customized to tackle large-scale instances of the problems to show the performance of the solution method.

  8. The formation mechanism for printed silver-contacts for silicon solar cells

    Science.gov (United States)

    Fields, Jeremy D.; Ahmad, Md. Imteyaz; Pool, Vanessa L.; Yu, Jiafan; van Campen, Douglas G.; Parilla, Philip A.; Toney, Michael F.; van Hest, Maikel F. A. M.

    2016-04-01

    Screen-printing provides an economically attractive means for making Ag electrical contacts to Si solar cells, but the use of Ag substantiates a significant manufacturing cost, and the glass frit used in the paste to enable contact formation contains Pb. To achieve optimal electrical performance and to develop pastes with alternative, abundant and non-toxic materials, a better understanding the contact formation process during firing is required. Here, we use in situ X-ray diffraction during firing to reveal the reaction sequence. The findings suggest that between 500 and 650 °C PbO in the frit etches the SiNx antireflective-coating on the solar cell, exposing the Si surface. Then, above 650 °C, Ag+ dissolves into the molten glass frit - key for enabling deposition of metallic Ag on the emitter surface and precipitation of Ag nanocrystals within the glass. Ultimately, this work clarifies contact formation mechanisms and suggests approaches for development of inexpensive, nontoxic solar cell contacting pastes.

  9. The formation mechanism for printed silver-contacts for silicon solar cells

    Energy Technology Data Exchange (ETDEWEB)

    Fields, Jeremy D.; Ahmad, Md. Imteyaz; Pool, Vanessa L.; Yu, Jiafan; Van Campen, Douglas G.; Parilla, Philip A.; Toney, Michael F.; van Hest, Maikel F. A. M.

    2016-04-01

    Screen-printing provides an economically attractive means for making Ag electrical contacts to Si solar cells, but the use of Ag substantiates a significant manufacturing cost, and the glass frit used in the paste to enable contact formation contains Pb. To achieve optimal electrical performance and to develop pastes with alternative, abundant, and non-toxic materials requires understanding the contact formation process during firing. Here, we use in-situ X-ray diffraction during firing to reveal the reaction sequence. The findings suggest that between 500 degrees C and 650 degrees C PbO in the frit etches the SiNx antireflective-coating on the solar cell, exposing the Si surface. Then, above 650 degrees C, Ag+ dissolves into the molten glass frit -- key for enabling deposition of metallic Ag on the emitter surface and precipitation of Ag nanocrystals within the glass. Ultimately, this work clarifies contact formation mechanisms and suggests approaches for development of inexpensive, nontoxic solar cell contacting pastes.

  10. The formation of argpyrimidine, a methylglyoxal-arginine adduct, in the nucleus of neural cells

    International Nuclear Information System (INIS)

    Methylglyoxal (MG) is an endogenous metabolite in glycolysis and forms stable adducts primarily with arginine residues of intracellular proteins. The biological role of this modification in cell function is not known. In the present study, we found that a MG-detoxification enzyme glyoxalase I (GLO1) is mainly expressed in the ventricular zone (VZ) at embryonic day 16 which neural stem and progenitor cells localize. Moreover, immunohistochemical analysis revealed that argpyrimidine, a major MG-arginine adduct, is predominantly produced in cortical plate neurons not VZ during cerebral cortex development and is exclusively located in the nucleus. Immunoblotting experiment showed that the formation of argpyrimidine occurs on some nuclear proteins of cortical neurons. To our knowledge, this is first report of the argpyrimidine formation in the nucleus of neuron. These findings suggest that GLO1, which is dominantly expressed in the embryonic VZ, reduces the intracellular level of MG and suppresses the formation of argpyrimidine in neural stem and progenitor cells. Argpyrimidine may contribute to the neural differentiation and/or the maintenance of the differentiated state via the modification of nuclear proteins.

  11. An Integrated Model for Production Planning and Cell Formation in Cellular Manufacturing Systems

    Directory of Open Access Journals (Sweden)

    Reza Raminfar

    2013-01-01

    Full Text Available Cellular manufacturing (CM is a production approach directed towards reducing costs, as well as increasing system's flexibility in today's small-to-medium lot production environment. Many structural and operational issues should be considered for a successful CM design and implementation such as cell formation (CF, production planning, and facility layout. Most researchers have addressed these issues sequentially or independently, instead of jointly optimizing a combination of these issues. In order to attain better results to ensure that the system will be capable of remaining efficient in unknown future situations, these issues should be addressed simultaneously. In this paper, a mathematical model is developed using an integrated approach for production planning and cell formation problems in a CM. A set of numerical examples are provided from existing the literature in order to test and illustrate the proposed model. In order to evaluate and verify the performance of the proposed model, it is compared with a well-known cell formation methods (rank order clustering and direct clustering analysis, using group capability index (GCI measure. The results and comparisons indicate that the proposed model has a significantly higher and satisfactory performance and it is reliable for the design and the analysis of CM systems.

  12. Efficient Parallel Sorting for Migrating Birds Optimization When Solving Machine-Part Cell Formation Problems

    Directory of Open Access Journals (Sweden)

    Ricardo Soto

    2016-01-01

    Full Text Available The Machine-Part Cell Formation Problem (MPCFP is a NP-Hard optimization problem that consists in grouping machines and parts in a set of cells, so that each cell can operate independently and the intercell movements are minimized. This problem has largely been tackled in the literature by using different techniques ranging from classic methods such as linear programming to more modern nature-inspired metaheuristics. In this paper, we present an efficient parallel version of the Migrating Birds Optimization metaheuristic for solving the MPCFP. Migrating Birds Optimization is a population metaheuristic based on the V-Flight formation of the migrating birds, which is proven to be an effective formation in energy saving. This approach is enhanced by the smart incorporation of parallel procedures that notably improve performance of the several sorting processes performed by the metaheuristic. We perform computational experiments on 1080 benchmarks resulting from the combination of 90 well-known MPCFP instances with 12 sorting configurations with and without threads. We illustrate promising results where the proposal is able to reach the global optimum in all instances, while the solving time with respect to a nonparallel approach is notably reduced.

  13. The postischemic environment differentially impacts teratoma or tumor formation after transplantation of human embryonic stem cell-derived neural progenitors

    DEFF Research Database (Denmark)

    Seminatore, Christine; Polentes, Jerome; Ellman, Ditte;

    2010-01-01

    Risk of tumorigenesis is a major obstacle to human embryonic and induced pluripotent stem cell therapy. Likely linked to the stage of differentiation of the cells at the time of implantation, formation of teratoma/tumors can also be influenced by factors released by the host tissue. We have...... analyzed the relative effects of the stage of differentiation and the postischemic environment on the formation of adverse structures by transplanted human embryonic stem cell-derived neural progenitors....

  14. Nonneuronal cells regulate synapse formation in the vestibular sensory epithelium via erbB-dependent BDNF expression

    OpenAIRE

    Gómez-Casati, Maria E; MURTIE, JOSHUA C.; Rio, Carlos; Stankovic, Konstantina; Liberman, M. Charles; Corfas, Gabriel

    2010-01-01

    Recent studies indicate that molecules released by glia can induce synapse formation. However, what induces glia to produce such signals, their identity, and their in vivo relevance remain poorly understood. Here we demonstrate that supporting cells of the vestibular organ—cells that have many characteristics of glia—promote synapse formation only when induced by neuron-derived signals. Furthermore, we identify BDNF as the synaptogenic signal produced by these nonneuronal cells. Mice in which...

  15. microRNA-150 inhibits the formation of macrophage foam cells through targeting adiponectin receptor 2.

    Science.gov (United States)

    Li, Jing; Zhang, Suhua

    2016-08-01

    Transformation of macrophages into foam cells plays a critical role in the pathogenesis of atherosclerosis. The aim of this study was to determine the expression and biological roles of microRNA (miR)-150 in the formation of macrophage foam cells and to identify its functional target(s). Exposure to 50 μg/ml oxidized low-density lipoprotein (oxLDL) led to a significant upregulation of miR-150 in THP-1 macrophages. Overexpression of miR-150 inhibited oxLDL-induced lipid accumulation in THP-1 macrophages, while knockdown of miR-150 enhanced lipid accumulation. apoA-I- and HDL-mediated cholesterol efflux was increased by 66% and 43%, respectively, in miR-150-overexpressing macrophages relative to control cells. In contrast, downregulation of miR-150 significantly reduced cholesterol efflux from oxLDL-laden macrophages. Bioinformatic analysis and luciferase reporter assay revealed adiponectin receptor 2 (AdipoR2) as a direct target of miR-150. Small interfering RNA-mediated downregulation of AdipoR2 phenocopied the effects of miR-150 overexpression, reducing lipid accumulation and facilitating cholesterol efflux in oxLDL-treated THP-1 macrophages. Knockdown of AdipoR2 induced the expression of proliferator-activated receptor gamma (PPARγ), liver X receptor alpha (LXRα), ABCA1, and ABCG1. Moreover, pharmacological inhibition of PPARγ or LXRα impaired AdipoR2 silencing-induced upregulation of ABCA1 and ABCG1. Taken together, our results indicate that miR-150 can attenuate oxLDL-induced lipid accumulation in macrophages via promotion of cholesterol efflux. The suppressive effects of miR-150 on macrophage foam cell formation are mediated through targeting of AdipoR2. Delivery of miR-150 may represent a potential approach to prevent macrophage foam cell formation in atherosclerosis. PMID:27216461

  16. Efficient retina formation requires suppression of both Activin and BMP signaling pathways in pluripotent cells

    Directory of Open Access Journals (Sweden)

    Kimberly A. Wong

    2015-03-01

    Full Text Available Retina formation requires the correct spatiotemporal patterning of key regulatory factors. While it is known that repression of several signaling pathways lead to specification of retinal fates, addition of only Noggin, a known BMP antagonist, can convert pluripotent Xenopus laevis animal cap cells to functional retinal cells. The aim of this study is to determine the intracellular molecular events that occur during this conversion. Surprisingly, blocking BMP signaling alone failed to mimic Noggin treatment. Overexpressing Noggin in pluripotent cells resulted in a concentration-dependent suppression of both Smad1 and Smad2 phosphorylation, which act downstream of BMP and Activin signaling, respectively. This caused a decrease in downstream targets: endothelial marker, xk81, and mesodermal marker, xbra. We treated pluripotent cells with dominant-negative receptors or the chemical inhibitors, dorsomorphin and SB431542, which each target either the BMP or Activin signaling pathway. We determined the effect of these treatments on retina formation using the Animal Cap Transplant (ACT assay; in which treated pluripotent cells were transplanted into the eye field of host embryos. We found that inhibition of Activin signaling, in the presence of BMP signaling inhibition, promotes efficient retinal specification in Xenopus tissue, mimicking the affect of adding Noggin alone. In whole embryos, we found that the eye field marker, rax, expanded when adding both dominant-negative Smad1 and Smad2, as did treating the cells with both dorsomorphin and SB431542. Future studies could translate these findings to a mammalian culture assay, in order to more efficiently produce retinal cells in culture.

  17. Synergistic inhibition of endothelial cell proliferation, tube formation, and sprouting by cyclosporin A and itraconazole.

    Science.gov (United States)

    Nacev, Benjamin A; Liu, Jun O

    2011-01-01

    Pathological angiogenesis contributes to a number of diseases including cancer and macular degeneration. Although angiogenesis inhibitors are available in the clinic, their efficacy against most cancers is modest due in part to the existence of alternative and compensatory signaling pathways. Given that angiogenesis is dependent on multiple growth factors and a broad signaling network in vivo, we sought to explore the potential of multidrug cocktails for angiogenesis inhibition. We have screened 741 clinical drug combinations for the synergistic inhibition of endothelial cell proliferation. We focused specifically on existing clinical drugs since the re-purposing of clinical drugs allows for a more rapid and cost effective transition to clinical studies when compared to new drug entities. Our screen identified cyclosporin A (CsA), an immunosuppressant, and itraconazole, an antifungal drug, as a synergistic pair of inhibitors of endothelial cell proliferation. In combination, the IC(50) dose of each drug is reduced by 3 to 9 fold. We also tested the ability of the combination to inhibit endothelial cell tube formation and sprouting, which are dependent on two essential processes in angiogenesis, endothelial cell migration and differentiation. We found that CsA and itraconazole synergistically inhibit tube network size and sprout formation. Lastly, we tested the combination on human foreskin fibroblast viability as well as Jurkat T cell and HeLa cell proliferation, and found that endothelial cells are selectively targeted. Thus, it is possible to combine existing clinical drugs to synergistically inhibit in vitro models of angiogenesis. This strategy may be useful in pursuing the next generation of antiangiogenesis therapy.

  18. Synergistic inhibition of endothelial cell proliferation, tube formation, and sprouting by cyclosporin A and itraconazole.

    Directory of Open Access Journals (Sweden)

    Benjamin A Nacev

    Full Text Available Pathological angiogenesis contributes to a number of diseases including cancer and macular degeneration. Although angiogenesis inhibitors are available in the clinic, their efficacy against most cancers is modest due in part to the existence of alternative and compensatory signaling pathways. Given that angiogenesis is dependent on multiple growth factors and a broad signaling network in vivo, we sought to explore the potential of multidrug cocktails for angiogenesis inhibition. We have screened 741 clinical drug combinations for the synergistic inhibition of endothelial cell proliferation. We focused specifically on existing clinical drugs since the re-purposing of clinical drugs allows for a more rapid and cost effective transition to clinical studies when compared to new drug entities. Our screen identified cyclosporin A (CsA, an immunosuppressant, and itraconazole, an antifungal drug, as a synergistic pair of inhibitors of endothelial cell proliferation. In combination, the IC(50 dose of each drug is reduced by 3 to 9 fold. We also tested the ability of the combination to inhibit endothelial cell tube formation and sprouting, which are dependent on two essential processes in angiogenesis, endothelial cell migration and differentiation. We found that CsA and itraconazole synergistically inhibit tube network size and sprout formation. Lastly, we tested the combination on human foreskin fibroblast viability as well as Jurkat T cell and HeLa cell proliferation, and found that endothelial cells are selectively targeted. Thus, it is possible to combine existing clinical drugs to synergistically inhibit in vitro models of angiogenesis. This strategy may be useful in pursuing the next generation of antiangiogenesis therapy.

  19. Effect of bioactive extruded PLA/HA composite films on focal adhesion formation of preosteoblastic cells.

    Science.gov (United States)

    Persson, Maria; Lorite, Gabriela S; Kokkonen, Hanna E; Cho, Sung-Woo; Lehenkari, Petri P; Skrifvars, Mikael; Tuukkanen, Juha

    2014-09-01

    The quality of the initial cell attachment to a biomaterial will influence any further cell function, including spreading, proliferation, differentiation and viability. Cell attachment is influenced by the material's ability to adsorb proteins, which is related to the surface chemistry and topography of the material. In this study, we incorporated hydroxyapatite (HA) particles into a poly(lactic acid) (PLA) composite and evaluated the surface structure and the effects of HA density on the initial cell attachment in vitro of murine calvarial preosteoblasts (MC3T3-EI). Scanning electron microscopy (SEM), atomic force microscopy (AFM) and infrared spectroscopy (FTIR) showed that the HA particles were successfully incorporated into the PLA matrix and located at the surface which is of importance in order to maintain the bioactive effect of the HA particles. SEM and AFM investigation revealed that the HA density (particles/area) as well as surface roughness increased with HA loading concentration (i.e. 5, 10, 15 and 20wt%), which promoted protein adsorption. Furthermore, the presence of HA on the surface enhanced cell spreading, increased the formation of actin stress fibers and significantly improved the expression of vinculin in MC3T3-E1 cells which is a key player in the regulation of cell adhesion. These results suggest the potential utility of PLA/HA composites as biomaterials for use as a bone substitute material and in tissue engineering applications. PMID:24986753

  20. Aggregate formation and suspension culture of human pluripotent stem cells and differentiated progeny.

    Science.gov (United States)

    Hookway, Tracy A; Butts, Jessica C; Lee, Emily; Tang, Hengli; McDevitt, Todd C

    2016-05-15

    Culture of human pluripotent stem cells (hPSC) as in vitro multicellular aggregates has been increasingly used as a method to model early embryonic development. Three-dimensional assemblies of hPSCs facilitate interactions between cells and their microenvironment to promote morphogenesis, analogous to the multicellular organization that accompanies embryogenesis. In this paper, we describe a method for reproducibly generating and maintaining populations of homogeneous three-dimensional hPSC aggregates using forced aggregation and rotary orbital suspension culture. We propose solutions to several challenges associated with the consistent formation and extended culture of cell spheroids generated from hPSCs and their differentiated progeny. Further, we provide examples to demonstrate how aggregation can be used as a tool to select specific subpopulations of cells to create homotypic spheroids, or as a means to introduce multiple cell types to create heterotypic tissue constructs. Finally, we demonstrate that the aggregation and rotary suspension method can be used to support culture and maintenance of hPSC-derived cell populations representing each of the three germ layers, underscoring the utility of this platform for culturing many different cell types. PMID:26658353

  1. Addition of Adipose-Derived Stem Cells to Mesenchymal Stem Cell Sheets Improves Bone Formation at an Ectopic Site

    Directory of Open Access Journals (Sweden)

    Zhifa Wang

    2016-02-01

    Full Text Available To determine the effect of adipose-derived stem cells (ADSCs added to bone marrow-derived mesenchymal stem cell (MSC sheets on bone formation at an ectopic site. We isolated MSCs and ADSCs from the same rabbits. We then prepared MSC sheets for implantation with or without ADSCs subcutaneously in the backs of severe combined immunodeficiency (SCID mice. We assessed bone formation at eight weeks after implantation by micro-computed tomography and histological analysis. In osteogenic medium, MSCs grew to form multilayer sheets containing many calcium nodules. MSC sheets without ADSCs formed bone-like tissue; although neo-bone and cartilage-like tissues were sparse and unevenly distributed by eight weeks after implantation. In comparison, MSC sheets with ADSCs promoted better bone regeneration as evidenced by the greater density of bone, increased mineral deposition, obvious formation of blood vessels, large number of interconnected ossified trabeculae and woven bone structures, and greater bone volume/total volume within the composite constructs. Our results indicate that although sheets of only MSCs have the potential to form tissue engineered bone at an ectopic site, the addition of ADSCs can significantly increase the osteogenic potential of MSC sheets. Thus, the combination of MSC sheets with ADSCs may be regarded as a promising therapeutic strategy to stimulate bone regeneration.

  2. ZnO Nanoparticles Affect Bacillus subtilis Cell Growth and Biofilm Formation.

    Directory of Open Access Journals (Sweden)

    Yi-Huang Hsueh

    Full Text Available Zinc oxide nanoparticles (ZnO NPs are an important antimicrobial additive in many industrial applications. However, mass-produced ZnO NPs are ultimately disposed of in the environment, which can threaten soil-dwelling microorganisms that play important roles in biodegradation, nutrient recycling, plant protection, and ecological balance. This study sought to understand how ZnO NPs affect Bacillus subtilis, a plant-beneficial bacterium ubiquitously found in soil. The impact of ZnO NPs on B. subtilis growth, FtsZ ring formation, cytosolic protein activity, and biofilm formation were assessed, and our results show that B. subtilis growth is inhibited by high concentrations of ZnO NPs (≥ 50 ppm, with cells exhibiting a prolonged lag phase and delayed medial FtsZ ring formation. RedoxSensor and Phag-GFP fluorescence data further show that at ZnO-NP concentrations above 50 ppm, B. subtilis reductase activity, membrane stability, and protein expression all decrease. SDS-PAGE Stains-All staining results and FT-IR data further demonstrate that ZnO NPs negatively affect exopolysaccharide production. Moreover, it was found that B. subtilis biofilm surface structures became smooth under ZnO-NP concentrations of only 5-10 ppm, with concentrations ≤ 25 ppm significantly reducing biofilm formation activity. XANES and EXAFS spectra analysis further confirmed the presence of ZnO in co-cultured B. subtilis cells, which suggests penetration of cell membranes by either ZnO NPs or toxic Zn+ ions from ionized ZnO NPs, the latter of which may be deionized to ZnO within bacterial cells. Together, these results demonstrate that ZnO NPs can affect B. subtilis viability through the inhibition of cell growth, cytosolic protein expression, and biofilm formation, and suggest that future ZnO-NP waste management strategies would do well to mitigate the potential environmental impact engendered by the disposal of these nanoparticles.

  3. Detection of S-nitrosothiol and nitrosylated proteins in Arachis hypogaea functional nodule: response of the nitrogen fixing symbiont.

    Directory of Open Access Journals (Sweden)

    Debasis Maiti

    Full Text Available To detect the presence of NO, ROS and RNS in nodules of crack entry legumes, we used Arachis hypogaea functional nodule. The response of two cognate partner rhizobia was compared towards NO and GSNO using S. meliloti and Bradyrhizobium sp NC921001. ROS, NO, nitrosothiol and bacteroids were detected by fluorescence microscopy. Redox enzymes and thiol pools were detected biochemically. Nitrosothiols were found to be present but ROS and NO were absent in A. hypogaea nodule. A number of S-nitrosylated proteins were also detected. The total thiol pool and most of the redox enzymes were low in nodule cytosolic extract but these were found to be high in the partner microorganisms indicating partner rhizobia could protect the nodule environment against the nitrosothiols. Both S. meliloti and Bradyrhizobium sp NC921001 were found to contain GSNO reductase. Interestingly, there was a marked difference in growth pattern between S. meliloti and Bradyrhizobium sp in presence of sodium nitroprusside (SNP and S-nitrosoglutathione (GSNO. Bradyrhizobium sp was found to be much more tolerant to NO donor compounds than the S. meliloti. In contrast, S. meliloti showed resistance to GSNO but was sensitive to SNP. Together our data indicate that nodule environment of crack entry legumes is different than the nodules of infection mode entry in terms of NO, ROS and RNS. Based on our biochemical characterization, we propose that exchange of redox molecules and reactive chemical species is possible between the bacteroid and nodule compartment.

  4. Mesenchymal stem cells alleviate atherosclerosis by elevating number and function of CD4(+)CD25 (+)FOXP3 (+) regulatory T-cells and inhibiting macrophage foam cell formation.

    Science.gov (United States)

    Wang, Zhi Xiao; Wang, Chong Quan; Li, Xiao Yan; Feng, Gao Ke; Zhu, Hong Ling; Ding, Yan; Jiang, Xue Jun

    2015-02-01

    Atherosclerosis is a chronic inflammatory disease characterized by the formation of plaques inside arteries, leading to narrowing and blockage. Potential therapeutic strategies include expanding the population of regulatory T-cells (Tregs) to enhance atheroprotective immunity, and inhibiting the formation of macrophage foam cells. Here, we studied the effect of bone marrow-derived mesenchymal stem cells (BM-MSCs) on atherosclerotic plaque formation in Apolipoprotein E(-/-) (ApoE-KO) mice, and elucidated the underlying mechanism. BM-MSCs isolated from 4 week-old ApoE-KO mice were evaluated by flow cytometry for expression of MSC-specific markers. Thirty eight week-old ApoE-KO mice were randomly divided into three experimental groups (n = 10 per group): 1. MSC group-received BM-MSCs intravenously; 2. Vehicle group-received DMEM; 3. Control group-did not receive any treatment. Administration of MSCs resulted in a marked decrease in the size of atherosclerotic plaques 3 months after treatment. In addition, the number and function of CD4(+)CD25(+)FOXP3(+) regulatory T-cells (Tregs) in cultured splenocytes, and the expression of FOXP3 at both mRNA and protein levels, was significantly increased in the MSC group. In vitro experiments further indicated that the formation of macrophage foam cells was inhibited by treatment with MSCs, accompanied by a significant downregulation in CD36 and scavenger receptor A (SRA). Our findings suggest that MSCs play an atheroprotective role by enhancing the number and function of Tregs and inhibiting the formation of macrophage foam cells. Hence, administration of MSCs to atherosclerotic patients might have significant clinical benefits.

  5. Sustained diacylglycerol formation from inositol phospholipids in angiotensin II-stimulated vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Griendling, K.K.; Rittenhouse, S.E.; Brock, T.A.; Ekstein, L.S.; Gimbrone, M.A. Jr.; Alexander, R.W.

    1986-05-05

    Angiotensin II acts on cultured rat aortic vascular smooth muscle cells to stimulate phospholipase C-mediated hydrolysis of membrane phosphoinositides and subsequent formation of diacylglycerol and inositol phosphates. In intact cells, angiotensin II induces a dose-dependent increase in diglyceride which is detectable after 5 s and sustained for at least 20 min. Angiotensin II (100 nM)-stimulated diglyceride formation is biphasic, peaking at 15 s (227 +/- 19% control) and at 5 min (303 +/- 23% control). Simultaneous analysis of labeled inositol phospholipids shows that at 15 s phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 4-phosphate (PIP) decline to 52 +/- 6% control and 63 +/- 5% control, respectively, while phosphatidylinositol (PI) remains unchanged. In contrast, at 5 min, PIP2 and PIP have returned toward control levels (92 +/- 2 and 82 +/- 4% control, respectively), while PI has decreased substantially (81 +/- 2% control). The calcium ionophore ionomycin (15 microM) stimulates diglyceride accumulation but does not cause PI hydrolysis. 4 beta-Phorbol 12-myristate 13-acetate, an activator of protein kinase C, inhibits early PIP and PIP2 breakdown and diglyceride formation, without inhibiting late-phase diglyceride accumulation. Thus, angiotensin II induces rapid transient breakdown of PIP and PIP2 and delayed hydrolysis of PI. The rapid attenuation of polyphosphoinositide breakdown is likely caused by a protein kinase C-mediated inhibition of PIP and PIP2 hydrolysis. While in vascular smooth muscle stimulated with angiotensin II inositol 1,4,5-trisphosphate formation is transient, diglyceride production is biphasic, suggesting that initial and sustained diglyceride formation from the phosphoinositides results from different biochemical and/or cellular processes.

  6. Lipocalin-2 inhibits osteoclast formation by suppressing the proliferation and differentiation of osteoclast lineage cells.

    Science.gov (United States)

    Kim, Hyun-Ju; Yoon, Hye-Jin; Yoon, Kyung-Ae; Gwon, Mi-Ri; Jin Seong, Sook; Suk, Kyoungho; Kim, Shin-Yoon; Yoon, Young-Ran

    2015-06-10

    Lipocalin-2 (LCN2) is a member of the lipocalin superfamily and plays a critical role in the regulation of various physiological processes, such as inflammation and obesity. In this study, we report that LCN2 negatively modulates the proliferation and differentiation of osteoclast precursors, resulting in impaired osteoclast formation. The overexpression of LCN2 in bone marrow-derived macrophages or the addition of recombinant LCN2 protein inhibits the formation of multinuclear osteoclasts. LCN2 suppresses macrophage colony-stimulating factor (M-CSF)-induced proliferation of osteoclast precursor cells without affecting their apoptotic cell death. Interestingly, LCN2 decreases the expression of the M-CSF receptor, c-Fms, and subsequently blocks its downstream signaling cascades. In addition, LCN2 inhibits RANKL-induced osteoclast differentiation and attenuates the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are important modulators in osteoclastogenesis. Mechanistically, LCN2 inhibits NF-κB signaling pathways, as demonstrated by the suppression of IκBα phosphorylation, nuclear translocation of p65, and NF-κB transcriptional activity. Thus, LCN2 is an anti-osteoclastogenic molecule that exerts its effects by retarding the proliferation and differentiation of osteoclast lineage cells.

  7. Rapid and efficient generation of neurons from human pluripotent stem cells in a multititre plate format.

    Science.gov (United States)

    Zhang, Miao; Schöler, Hans R; Greber, Boris

    2013-01-01

    Existing protocols for the generation of neurons from human pluripotent stem cells (hPSCs) are often tedious in that they are multistep procedures involving the isolation and expansion of neural precursor cells, prior to terminal differentiation. In comparison to these time-consuming approaches, we have recently found that combined inhibition of three signaling pathways, TGFβ/SMAD2, BMP/SMAD1, and FGF/ERK, promotes rapid induction of neuroectoderm from hPSCs, followed by immediate differentiation into functional neurons. Here, we have adapted our procedure to a novel multititre plate format, to further enhance its reproducibility and to make it compatible with mid-throughput applications. It comprises four days of neuroectoderm formation in floating spheres (embryoid bodies), followed by a further four days of differentiation into neurons under adherent conditions. Most cells obtained with this protocol appear to be bipolar sensory neurons. Moreover, the procedure is highly efficient, does not require particular expert skills, and is based on a simple chemically defined medium with cost-efficient small molecules. Due to these features, the procedure may serve as a useful platform for further functional investigation as well as for cell-based screening approaches requiring human sensory neurons or neurons of any type. PMID:23486189

  8. Decrease in spermidine content during logarithmic phase of cell growth delays spore formation of Bacillus subtilis.

    Science.gov (United States)

    Ishii, I; Takada, H; Terao, K; Kakegawa, T; Igarashi, K; Hirose, S

    1994-11-01

    Bacillus subtilis 168M contained a large amount of spermidine during the logarithmic phase of growth, but the amount decreased drastically during the stationary phase. The extracts, prepared from B. subtilis cells harvested in the logarithmic phase, contained activity of arginine decarboxylase (ADC) rather than the activity of ornithine decarboxylase. In the presence of alpha-difluoromethylarginine (DFMA), a specific and irreversible inhibitor of ADC, the amount of spermidine in B. subtilis during the logarithmic phase decreased to about 25% of the control cells. Under these conditions, spore formation of B. subtilis 168M delayed greatly without significant inhibition of cell growth. The decrease in spermidine content in the logarithmic phase rather than in the stationary phase was involved in the delay of sporulation. Electron microscopy of cells at 24 hrs. of culture confirmed the delay of spore formation by the decrease of spermidine content. Furthermore, the delay of sporulation was negated by the addition of spermidine. These data suggest that a large amount of spermidine existing during the logarithmic phase plays an important role in the sporulation of B. subtilis.

  9. Lipocalin-2 inhibits osteoclast formation by suppressing the proliferation and differentiation of osteoclast lineage cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hyun-Ju, E-mail: biohjk@knu.ac.kr [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Yoon, Hye-Jin [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Yoon, Kyung-Ae [Department of Orthopedic Surgery, Skeletal Diseases Genome Research Center, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Gwon, Mi-Ri; Jin Seong, Sook [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Suk, Kyoungho [Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Kim, Shin-Yoon [Department of Orthopedic Surgery, Skeletal Diseases Genome Research Center, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Yoon, Young-Ran, E-mail: yry@knu.ac.kr [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of)

    2015-06-10

    Lipocalin-2 (LCN2) is a member of the lipocalin superfamily and plays a critical role in the regulation of various physiological processes, such as inflammation and obesity. In this study, we report that LCN2 negatively modulates the proliferation and differentiation of osteoclast precursors, resulting in impaired osteoclast formation. The overexpression of LCN2 in bone marrow-derived macrophages or the addition of recombinant LCN2 protein inhibits the formation of multinuclear osteoclasts. LCN2 suppresses macrophage colony-stimulating factor (M-CSF)-induced proliferation of osteoclast precursor cells without affecting their apoptotic cell death. Interestingly, LCN2 decreases the expression of the M-CSF receptor, c-Fms, and subsequently blocks its downstream signaling cascades. In addition, LCN2 inhibits RANKL-induced osteoclast differentiation and attenuates the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are important modulators in osteoclastogenesis. Mechanistically, LCN2 inhibits NF-κB signaling pathways, as demonstrated by the suppression of IκBα phosphorylation, nuclear translocation of p65, and NF-κB transcriptional activity. Thus, LCN2 is an anti-osteoclastogenic molecule that exerts its effects by retarding the proliferation and differentiation of osteoclast lineage cells. - Highlights: • LCN2 expression is regulated during osteoclast development. • LCN2 suppresses M-CSF-mediated osteoclast precursor proliferation. • LCN2 inhibits RANKL-induced osteoclast differentiation.

  10. Lumen Formation Is an Intrinsic Property of Isolated Human Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Kenichiro Taniguchi

    2015-12-01

    Full Text Available We demonstrate that dissociated human pluripotent stem cells (PSCs are intrinsically programmed to form lumens. PSCs form two-cell cysts with a shared apical domain within 20 hr of plating; these cysts collapse to form monolayers after 5 days. Expression of pluripotency markers is maintained throughout this time. In two-cell cysts, an apical domain, marked by EZRIN and atypical PKCζ, is surrounded by apically targeted organelles (early endosomes and Golgi. Molecularly, actin polymerization, regulated by ARP2/3 and mammalian diaphanous-related formin 1 (MDIA, promotes lumen formation, whereas actin contraction, mediated by MYOSIN-II, inhibits this process. Finally, we show that lumenal shape can be manipulated in bioengineered micro-wells. Since lumen formation is an indispensable step in early mammalian development, this system can provide a powerful model for investigation of this process in a controlled environment. Overall, our data establish that lumenogenesis is a fundamental cell biological property of human PSCs.

  11. Exocytosis of macrophage lysosomes leads to digestion of apoptotic adipocytes and foam cell formation.

    Science.gov (United States)

    Haka, Abigail S; Barbosa-Lorenzi, Valéria C; Lee, Hyuek Jong; Falcone, Domenick J; Hudis, Clifford A; Dannenberg, Andrew J; Maxfield, Frederick R

    2016-06-01

    Many types of apoptotic cells are phagocytosed and digested by macrophages. Adipocytes can be hundreds of times larger than macrophages, so they are too large to be digested by conventional phagocytic processes. The nature of the interaction between macrophages and apoptotic adipocytes has not been studied in detail. We describe a cellular process, termed exophagy, that is important for macrophage clearance of dead adipocytes and adipose tissue homeostasis. Using mouse models of obesity, human tissue, and a cell culture model, we show that macrophages form hydrolytic extracellular compartments at points of contact with dead adipocytes using local actin polymerization. These compartments are acidic and contain lysosomal enzymes delivered by exocytosis. Uptake and complete degradation of adipocyte fragments, which are released by extracellular hydrolysis, leads to macrophage foam cell formation. Exophagy-mediated foam cell formation is a highly efficient means by which macrophages internalize large amounts of lipid, which may ultimately overwhelm the metabolic capacity of the macrophage. This process provides a mechanism for degradation of objects, such as dead adipocytes, that are too large to be phagocytosed by macrophages. PMID:27044658

  12. Oleic acid induces smooth muscle foam cell formation and enhances atherosclerotic lesion development via CD36

    Directory of Open Access Journals (Sweden)

    Tang Bing

    2011-04-01

    Full Text Available Abstract Background Elevated plasma free fatty acid (FFA levels have been linked to the development of atherosclerosis. However, how FFA causes atherosclerosis has not been determined. Because fatty acid translocase (FAT/CD36 is responsible for the uptake of FFA, we hypothesized that the atherogenic effects of FFA may be mediated via CD36. Results We tested this hypothesis using cultured rat aortic smooth muscle cells (SMCs treated with oleic acid (OA. We found that OA induces lipid accumulation in SMCs in a dose dependent manner. Rat aortic SMCs treated for 48 hours with OA (250 μmol/L became foam cells based on morphological (Oil Red O staining and biochemical (5 times increase in cellular triglyceride criteria. Moreover, specific inhibition of CD36 by sulfo-N-succinimidyl oleate significantly attenuated OA induced lipid accumulation and foam cell formation. To confirm these results in vivo, we used ApoE-deficient mice fed with normal chow (NC, OA diet, NC plus lipolysis inhibitor acipimox or OA plus acipimox. OA-fed mice showed increased plasma FFA levels and enhanced atherosclerotic lesions in the aortic sinus compared to the NC group (both p 5 μm2 vs. OA plus acipimox: 2.60 ± 0.10 ×105 μm2, p p Conclusions These findings suggest that OA induces smooth muscle foam cell formation and enhances atherosclerotic lesions in part though CD36. Furthermore, these findings provide a novel model for the investigation of atherosclerosis.

  13. Collagen cross-linking by adipose-derived mesenchymal stromal cells and scar-derived mesenchymal cells : Are mesenchymal stromal cells involved in scar formation?

    NARCIS (Netherlands)

    van den Bogaerdt, Antoon J.; van der Veen, Vincent C.; van Zuijlen, Paul P. M.; Reijnen, Linda; Verkerk, Michelle; Bank, Ruud A.; Middelkoop, Esther; Ulrich, Magda M. W.

    2009-01-01

    In this work, different fibroblast-like (mesenchymal) cell populations that might be involved in wound healing were characterized and their involvement in scar formation was studied by determining collagen synthesis and processing. Depending on the physical and mechanical properties of the tissues,

  14. miR-93 suppresses proliferation and colony formation of human colon cancer stem cells

    Institute of Scientific and Technical Information of China (English)

    Xiao-Feng Yu; Jian Zou; Zhi-Jun Bao; Jie Dong

    2011-01-01

    AIM: To identify differentially expressed microRNAs (miRNAs) in human colon cancer stem cells (SW1116csc) and study their function in SW1116csc proliferation.METHODS: SW1116csc were isolated from the human colon cancer cell line, SW1116 and cultured in serum free medium. A miRNA microarray was used to detect differential expression profiles of miRNAs in SW1116csc and SW1116 cells. Real-time quantitative polymerase chain reaction (PCR) was performed to verify the dif ferential expression of candidate miRNAs obtained from the microarray. Target mRNAs of differentially expressed miRNAs were predicted with target predic tion tools. miRNA expression plasmids were transfected into SW1116csc using Lipofectamine 2000 reagent. Cell proliferation curves were generated with trypan blue staining, and the colony formation rate of transfected cells was measured with the soft agar colony formation assay. Expression of target mRNAs and proteins from differentially expressed miRNAs were detected using reverse transcription (RT)-PCR and western blotting.RESULTS: Compared with expression in SW1116 cells, 35 miRNAs (including hsa-miR-192, hsa-miR-29b, hsa-miR-215, hsa-miR-194, hsa-miR-33a and hsa-miR-32) were upregulated more than 1.5-fold, and 11 miRNAs (including hsa-miR-93, hsa-miR-1231, hsa-miRPIus-F1080, hsa-miR-524-3p, hsa-miR-886-3p and hsa-miR-561) were downregulated in SW1116csc. The miRNA microarray results were further validated with quantitative RT-PCR. miR-93 was downregulated, and its predicted mRNA targets included BAMBI, CCND2, CDKN1A, HDAC8, KIF23, MAP3K9, MAP3K11, MYCN, PPARD, TLE4 and ZDHHC1. Overexpressed miR-93 sig nificantly inhibited cell proliferation and colony forma tion by SW1116CSC. Furthermore, miR-93 negatively regulated the mRNA and protein levels of HDAC8 and TLE4.CONCLUSION: Some miRNAs were differentially ex pressed during differentiation of SW1116csc into SW1116 cells. miR-93 may inhibit SW1116csc proliferation and colony formation.

  15. Influence of cell-to-cell variability on spatial pattern formation

    NARCIS (Netherlands)

    Greese, B.; Wester, K.; Bensch, R.; Ronneberger, O.; Timmer, J.; Huulskamp, M.; Fleck, C.

    2012-01-01

    Many spatial patterns in biology arise through differentiation of selected cells within a tissue, which is regulated by a genetic network. This is specified by its structure, parameterisation and the noise on its components and reactions. The latter, in particular, is not well examined because it is

  16. IL-20 activates human lymphatic endothelial cells causing cell signalling and tube formation

    DEFF Research Database (Denmark)

    Hammer, Troels; Tritsaris, Katerina; Hübschmann, Martin V;

    2009-01-01

    IL-20 is an arteriogenic cytokine that remodels collateral networks in vivo, and plays a role in cellular organization. Here, we investigate its role in lymphangiogenesis using a lymphatic endothelial cell line, hTERT-HDLEC, which expresses the lymphatic markers LYVE-1 and podoplanin. Upon stimul...

  17. Geometrical Aspects During Formation of Compact Aggregates of Red Blood Cells

    Directory of Open Access Journals (Sweden)

    Cardoso A.V.

    2002-01-01

    Full Text Available In the past forty years considerable progress has been achieved on the knowledge of human blood as a non-Newtonian shear-thinning suspension, whose initial state, that is at rest (stasis or at very low shear rates, has a gel-like internal structure which is destroyed as shear stress increases. The main goal of this communication is to describe the role of geometrical aspects during RBC (red blood cell aggregate formation, growth and compaction on naturally aggregate (porcine blood and non-aggregate (bovine blood samples. We consider how these aspects coupled with tension equilibrium are decisive to transform red cell linear roleaux to three-dimensional aggregates or clusters. Geometrical aspects are also crucial on the compaction of red blood cell aggregates. These densely packed aggregates could precipitate out of blood- either as dangerous deposits on arterial walls, or as clots which travel in suspension until they block some crucial capillary.

  18. Magnolol inhibits migration of vascular smooth muscle cells via cytoskeletal remodeling pathway to attenuate neointima formation

    Energy Technology Data Exchange (ETDEWEB)

    Karki, Rajendra [Division of Pharmacology and Toxicology, School of Pharmacy, University of Missouri-Kansas City (United States); Department of Oriental Medicine Resources, Mokpo National University (Korea, Republic of); Kim, Seong-Bin [Jeollanamdo Development Institute for Korean Traditional Medicine, Jangheung gun, Jeollanamdo (Korea, Republic of); Kim, Dong-Wook, E-mail: dbkim@mokpo.ac.kr [Department of Oriental Medicine Resources, Mokpo National University (Korea, Republic of)

    2013-12-10

    Background: Increased proliferation and migration of vascular smooth muscle cells (VSMCs) contribute importantly to the formation of both atherosclerotic and restenotic lesions. The objective of this study was to investigate the effect of magnolol on VSMC migration. Methods: The proteolytic activity of matrix metalloproteinases (MMPs) in tumor necrosis factor alpha (TNF-α) stimulated VSMCs was performed by gelatin zymography. VSMC migration was assessed by wound healing and Boyden chamber methods. Collagen induced VSMC adhesion was determined by spectrofluorimeter and stress fibers formation was evaluated by fluorescence microscope. The expression of signaling molecules involved in stress fibers formation was determined by western blot. The phosphorylation of myosin light chain (MLC20) was determined by urea-glycerol polyacrylamide gel electrophoresis. Immunohistochemistry was performed to determine the expression of β1-integrin and collagen type I in the injured carotid arteries of rats on day 35 after vascular injury. Results: VSMC migration was strongly inhibited by magnolol without affecting MMPs expression. Also, magnolol inhibited β1-integrin expression, FAK phosphorylation and RhoA and Cdc42 activation to inhibit the collagen induced stress fibers formation. Moreover, magnolol inhibited the phosphorylation of MLC20. Our in vivo results showed that magnolol inhibited β1-integrin expression, collagen type I deposition and FAK phosphorylation in injured carotid arteries without affecting MMP-2 activity. Conclusions: Magnolol inhibited VSMC migration via inhibition of cytoskeletal remodeling pathway to attenuate neointima formation. General significance: This study provides a rationale for further evaluation of magnolol for the management of atherosclerosis and restenosis. - Highlights: • Magnolol strongly inhibited migration of VSMCs. • Magnolol inhibited stress fibers formation. • MLC20 phosphorylation was also inhibited by magnolol. • Anti

  19. Effect of Genistein on vasculogenic mimicry formation by human uveal melanoma cells

    Directory of Open Access Journals (Sweden)

    Gu Haijuan

    2009-09-01

    Full Text Available Abstract Background Vasculogenic mimicry (VM was increasingly recognized as a form of aggressive melanoma acquiring blood supply. Genistein had attracted much attention as a potential anticancer agent. Therefore, we examined the effect of Genistein on VM in human uveal melanoma cells. Methods VM structure was detected by periodic acid-Schiff (PAS staining for uveal melanoma C918 cells cultured on the three-dimensional type I collagen gels after exposed to Genistein. We used reverse transcription polymerase chain reaction (RT-PCR and Western Blot analysis to examine the effect of Genistein on vascular endothelial cadherin (VE-cadherin mRNA and protein expression. The nude mice models of human uveal melanoma C918 cells were established to assess the number of VM using immunohistochemical and PAS double-staining. Results Genistein inhibited the survival of C918 cells in vitro. The ectopic model study showed that VM in tumor tissue sections were significantly reduced by Genistein in vivo. In vitro, the VM structure was found in control, 25 and 50 μM Genistein-treatment groups but not in 100 and 200 μM. RT-PCR and Western Blot showed that 100 and 200 μM concentration of Genistein could significantly decrease VE-cadherin mRNA and protein expression of C918 cells compared with control (P 0.05. Conclusion Genistein inhibits VM formation of uveal melanoma cells in vivo and in vitro. One possible underlying molecular mechanism by which Genistein could inhibit VM formation of uveal melanoma is related to down-regulation of VE-cadherin.

  20. Protein complex formation and intranuclear dynamics of NAC1 in cancer cells.

    Science.gov (United States)

    Nakayama, Naomi; Kato, Hiroaki; Sakashita, Gyosuke; Nariai, Yuko; Nakayama, Kentaro; Kyo, Satoru; Urano, Takeshi

    2016-09-15

    Nucleus accumbens-associated protein 1 (NAC1) is a cancer-related transcription regulator protein that is also involved in the pluripotency and differentiation of embryonic stem cells. NAC1 is overexpressed in various carcinomas including ovarian, cervical, breast, and pancreatic carcinomas. NAC1 knock-down was previously shown to result in the apoptosis of ovarian cancer cell lines and to rescue their sensitivity to chemotherapy, suggesting that NAC1 may be a potential therapeutic target, but protein complex formation and the dynamics of intranuclear NAC1 in cancer cells remain poorly understood. In this study, analysis of HeLa cell lysates by fast protein liquid chromatography (FPLC) on a sizing column showed that the NAC1 peak corresponded to an apparent molecular mass of 300-500 kDa, which is larger than the estimated molecular mass (58 kDa) of the protein. Furthermore, live cell photobleaching analyses with green fluorescent protein (GFP)-fused NAC1 proteins revealed the intranuclear dynamics of NAC1. Collectively our results demonstrate that NAC1 forms a protein complex to function as a transcriptional regulator in cancer cells.

  1. Protein complex formation and intranuclear dynamics of NAC1 in cancer cells.

    Science.gov (United States)

    Nakayama, Naomi; Kato, Hiroaki; Sakashita, Gyosuke; Nariai, Yuko; Nakayama, Kentaro; Kyo, Satoru; Urano, Takeshi

    2016-09-15

    Nucleus accumbens-associated protein 1 (NAC1) is a cancer-related transcription regulator protein that is also involved in the pluripotency and differentiation of embryonic stem cells. NAC1 is overexpressed in various carcinomas including ovarian, cervical, breast, and pancreatic carcinomas. NAC1 knock-down was previously shown to result in the apoptosis of ovarian cancer cell lines and to rescue their sensitivity to chemotherapy, suggesting that NAC1 may be a potential therapeutic target, but protein complex formation and the dynamics of intranuclear NAC1 in cancer cells remain poorly understood. In this study, analysis of HeLa cell lysates by fast protein liquid chromatography (FPLC) on a sizing column showed that the NAC1 peak corresponded to an apparent molecular mass of 300-500 kDa, which is larger than the estimated molecular mass (58 kDa) of the protein. Furthermore, live cell photobleaching analyses with green fluorescent protein (GFP)-fused NAC1 proteins revealed the intranuclear dynamics of NAC1. Collectively our results demonstrate that NAC1 forms a protein complex to function as a transcriptional regulator in cancer cells. PMID:27424155

  2. Lgr5+ve Stem/Progenitor Cells Contribute to Nephron Formation during Kidney Development

    Directory of Open Access Journals (Sweden)

    Nick Barker

    2012-09-01

    Full Text Available Multipotent stem cells and their lineage-restricted progeny drive nephron formation within the developing kidney. Here, we document expression of the adult stem cell marker Lgr5 in the developing kidney and assess the stem/progenitor identity of Lgr5+ve cells via in vivo lineage tracing. The appearance and localization of Lgr5+ve cells coincided with that of the S-shaped body around embryonic day 14. Lgr5 expression remained restricted to cell clusters within developing nephrons in the cortex until postnatal day 7, when expression was permanently silenced. In vivo lineage tracing identified Lgr5 as a marker of a stem/progenitor population within nascent nephrons dedicated to generating the thick ascending limb of Henle’s loop and distal convoluted tubule. The Lgr5 surface marker and experimental models described here will be invaluable for deciphering the contribution of early nephron stem cells to developmental defects and for isolating human nephron progenitors as a prerequisite to evaluating their therapeutic potential.

  3. Appressorium formation in the corn smut fungus Ustilago maydis requires a G2 cell cycle arrest.

    Science.gov (United States)

    Castanheira, Sónia; Pérez-Martín, José

    2015-01-01

    Many of the most important plant diseases are caused by fungal pathogens that form specialized cell structures to breach the leaf surface as well as to proliferate inside the plant. To initiate pathogenic development, the fungus responds to a set of inductive cues. Some of them are of extracellular nature (environmental signals) while others respond to intracellular conditions (developmental signals). These signals have to be integrated into a single response that has as a major outcome changes in the morphogenesis of the fungus. The cell cycle regulation is pivotal during these cellular differentiations, and we hypothesized that cell cycle regulation would be likely to provide control points for infection development by fungal pathogens. Although efforts have been done in various fungal systems, there is still limited information available regarding the relationship of these processes with the induction of the virulence programs. Hence, the role of fungal cell cycle regulators -which are wide conserved elements- as true virulence factors, has yet to be defined. Here we discuss the recent finding that the formation of the appressorium, a structure required for plant penetration, in the corn smut fungus Ustilago maydis seems to be incompatible with an active cell cycle and, therefore genetic circuits evolved in this fungus to arrest the cell cycle during the growth of this fungus on plant surface, before the appressorium-mediated penetration into the plant tissue.

  4. Inactivation of glutathione peroxidase activity contributes to UV-induced squamous cell carcinoma formation.

    Science.gov (United States)

    Walshe, Jennifer; Serewko-Auret, Magdalena M; Teakle, Ngari; Cameron, Sarina; Minto, Kelly; Smith, Louise; Burcham, Philip C; Russell, Terry; Strutton, Geoffrey; Griffin, Anthony; Chu, Fong-Fong; Esworthy, Stephen; Reeve, Vivienne; Saunders, Nicholas A

    2007-05-15

    Cutaneous squamous cell carcinomas (CSCC) are a common malignancy of keratinocytes that arise in sites of the skin exposed to excessive UV radiation. In the present study, we show that human SCC cell lines, preneoplastic solar keratoses (SK), and CSCC are associated with perturbations in glutathione peroxidase (GPX) activity and peroxide levels. Specifically, we found that two of three SKs and four of five CSCCs, in vivo, were associated with decreased GPX activity and all SKs and CSCCs were associated with an elevated peroxide burden. Given the association of decreased GPX activity with CSCC, we examined the basis for the GPX deficiency in the CSCCs. Our data indicated that GPX was inactivated by a post-translational mechanism and that GPX could be inactivated by increases in intracellular peroxide levels. We next tested whether the decreased peroxidase activity coupled with an elevated peroxidative burden might contribute to CSCC formation in vivo. This was tested in Gpx1(-/-) and Gpx2(-/-) mice exposed to solar-simulated UV radiation. These studies showed that Gpx2 deficiency predisposed mice to UV-induced CSCC formation. These results suggest that inactivation of GPX2 in human skin may be an early event in UV-induced SCC formation. PMID:17510403

  5. Forskolin enhances in vivo bone formation by human mesenchymal stromal cells.

    Science.gov (United States)

    Doorn, Joyce; Siddappa, Ramakrishnaiah; van Blitterswijk, Clemens A; de Boer, Jan

    2012-03-01

    Activation of the protein kinase A (PKA) pathway with dibutyryl cyclic adenosine monophosphate (db-cAMP) was recently shown to enhance osteogenic differentiation of human mesenchymal stromal cells (hMSCs) in vitro and bone formation in vivo. The major drawback of this compound is its inhibitory effect on proliferation of hMSCs. Therefore, we investigated whether fine-tuning of the dose and timing of PKA activation could enhance bone formation even further, with minimum effects on proliferation. To test this, we selected two different PKA activators (8-bromo-cAMP (8-br-cAMP) and forskolin) and compared their effects on proliferation and osteogenic differentiation with those of db-cAMP. We found that all three compounds induced alkaline phosphatase levels, bone-specific target genes, and secretion of insulin-like growth factor-1, although 8-br-cAMP induced adipogenic differentiation in long-term cultures and was thus considered unsuitable for further in vivo testing. All three compounds inhibited proliferation of hMSCs in a dose-dependent manner, with forskolin inhibiting proliferation most. The effect of forskolin on in vivo bone formation was tested by pretreating hMSCs before implantation, and we observed greater amounts of bone using forskolin than db-cAMP. Our data show forskolin to be a novel agent that can be used to increase bone formation and also suggests a role for PKA in the delicate balance between adipogenic and osteogenic differentiation. PMID:21942968

  6. alpha-Lactalbumin species variation, HAMLET formation, and tumor cell death.

    Science.gov (United States)

    Pettersson, Jenny; Mossberg, Ann-Kristin; Svanborg, Catharina

    2006-06-23

    HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a tumoricidal complex of apo alpha-lactalbumin and oleic acid, formed in casein after low pH treatment of human milk. This study examined if HAMLET-like complexes are present in casein from different species and if isolated alpha-lactalbumin from those species can form such complexes with oleic acid. Casein from human, bovine, equine, and porcine milk was separated by ion exchange chromatography and active complexes were only found in human casein. This was not explained by alpha-lactalbumin sequence variation, as purified bovine, equine, porcine, and caprine alpha-lactalbumins formed complexes with oleic acid with biological activity similar to HAMLET. We conclude that structural variation of alpha-lactalbumins does not preclude the formation of HAMLET-like complexes and that natural HAMLET formation in casein was unique to human milk, which also showed the highest oleic acid content.

  7. Stretch Injury of Human Induced Pluripotent Stem Cell Derived Neurons in a 96 Well Format

    Science.gov (United States)

    Sherman, Sydney A.; Phillips, Jack K.; Costa, J. Tighe; Cho, Frances S.; Oungoulian, Sevan R.; Finan, John D.

    2016-01-01

    Traumatic brain injury (TBI) is a major cause of mortality and morbidity with limited therapeutic options. Traumatic axonal injury (TAI) is an important component of TBI pathology. It is difficult to reproduce TAI in animal models of closed head injury, but in vitro stretch injury models reproduce clinical TAI pathology. Existing in vitro models employ primary rodent neurons or human cancer cell line cells in low throughput formats. This in vitro neuronal stretch injury model employs human induced pluripotent stem cell-derived neurons (hiPSCNs) in a 96 well format. Silicone membranes were attached to 96 well plate tops to create stretchable, culture substrates. A custom-built device was designed and validated to apply repeatable, biofidelic strains and strain rates to these plates. A high content approach was used to measure injury in a hypothesis-free manner. These measurements are shown to provide a sensitive, dose-dependent, multi-modal description of the response to mechanical insult. hiPSCNs transition from healthy to injured phenotype at approximately 35% Lagrangian strain. Continued development of this model may create novel opportunities for drug discovery and exploration of the role of human genotype in TAI pathology. PMID:27671211

  8. Functional cooperation between FACT and MCM is coordinated with cell cycle and differential complex formation

    Directory of Open Access Journals (Sweden)

    Lin Chih-Li

    2010-02-01

    Full Text Available Abstract Background Functional cooperation between FACT and the MCM helicase complex constitutes an integral step during DNA replication initiation. However, mode of regulation that underlies the proper functional interaction of FACT and MCM is poorly understood. Methods & Results Here we present evidence indicating that such interaction is coordinated with cell cycle progression and differential complex formation. We first demonstrate the existence of two distinct FACT-MCM subassemblies, FACT-MCM2/4/6/7 and FACT-MCM2/3/4/5. Both complexes possess DNA unwinding activity and are subject to cell cycle-dependent enzymatic regulation. Interestingly, analysis of functional attributes further suggests that they act at distinct, and possibly sequential, steps during origin establishment and replication initiation. Moreover, we show that the phosphorylation profile of the FACT-associated MCM4 undergoes a cell cycle-dependent change, which is directly correlated with the catalytic activity of the FACT-MCM helicase complexes. Finally, at the quaternary structure level, physical interaction between FACT and MCM complexes is generally dependent on persistent cell cycle and further stabilized upon S phase entry. Cessation of mitotic cycle destabilizes the complex formation and likely leads to compromised coordination and activities. Conclusions Together, our results correlate FACT-MCM functionally and temporally with S phase and DNA replication. They further demonstrate that enzymatic activities intrinsically important for DNA replication are tightly controlled at various levels, thereby ensuring proper progression of, as well as exit from, the cell cycle and ultimately euploid gene balance.

  9. CoCl2, a mimic of hypoxia, induces formation of polyploid giant cells with stem characteristics in colon cancer.

    Directory of Open Access Journals (Sweden)

    Laura M Lopez-Sánchez

    Full Text Available The induction of polyploidy is considered the reproductive end of cells, but there is evidence that polyploid giant cancer cells (PGCCs contribute to cell repopulation during tumor relapse. However, the role of these cells in the development, progression and response to therapy in colon cancer remains undefined. Therefore, the main objective of this study was to investigate the generation of PGCCs in colon cancer cells and identify mechanisms of formation. Treatment of HCT-116 and Caco-2 colon cancer cells with the hypoxia mimic CoCl2 induced the formation of cells with larger cell and nuclear size (PGCCs, while the cells with normal morphology were selectively eliminated. Cytometric analysis showed that CoCl2 treatment induced G2 cell cycle arrest and the generation of a polyploid cell subpopulation with increased cellular DNA content. Polyploidy of hypoxia-induced PGCCs was confirmed by FISH analysis. Furthermore, CoCl2 treatment effectively induced the stabilization of HIF-1α, the differential expression of a truncated form of p53 (p47 and decreased levels of cyclin D1, indicating molecular mechanisms associated with cell cycle arrest at G2. Generation of PGCCs also contributed to expansion of a cell subpopulation with cancer stem cells (CSCs characteristics, as indicated by colonosphere formation assays, and enhanced chemoresistance to 5-fluorouracil and oxaliplatin. In conclusion, the pharmacological induction of hypoxia in colon cancer cells causes the formation of PGCCs, the expansion of a cell subpopulation with CSC characteristics and chemoresistance. The molecular mechanisms involved, including the stabilization of HIF-1 α, the involvement of p53/p47 isoform and cell cycle arrest at G2, suggest novel targets to prevent tumor relapse and treatment failure in colon cancer.

  10. Hydroxylated polychlorinated biphenyls increase reactive oxygen species formation and induce cell death in cultured cerebellar granule cells

    International Nuclear Information System (INIS)

    Polychlorinated biphenyls (PCBs) are persistent organic pollutants that bioaccumulate in the body, however, they can be metabolized to more water-soluble products. Although they are more readily excreted than the parent compounds, some of the metabolites are still hydrophobic and may be more available to target tissues, such as the brain. They can also cross the placenta and reach a developing foetus. Much less is known about the toxicity of PCB metabolites than about the parent compounds. In the present study, we have investigated the effects of eight hydroxylated (OH) PCB congeners (2'-OH PCB 3, 4-OH PCB 14, 4-OH PCB 34, 4'-OH PCB 35, 4-OH PCB 36, 4'-OH PCB 36, 4-OH PCB 39, and 4'-OH PCB 68) on reactive oxygen species (ROS) formation and cell viability in rat cerebellar granule cells. We found that, similar to their parent compounds, OH-PCBs are potent ROS inducers with potency 4-OH PCB 14 < 4-OH PCB 36 < 4-OH PCB 34 < 4'-OH PCB 36 < 4'-OH PCB 68 < 4-OH PCB 39 < 4'-OH PCB 35. 4-OH PCB 36 was the most potent cell death inducer, and caused apoptotic or necrotic morphology depending on concentration. Inhibition of ERK1/2 kinase with U0126 reduced both cell death and ROS formation, suggesting that ERK1/2 activation is involved in OH-PCB toxicity. The results indicate that the hydroxylation of PCBs may not constitute a detoxification reaction. Since OH-PCBs like their parent compounds are retained in the body and may be more widely distributed to sensitive tissues, it is important that not only the levels of the parent compounds but also the levels of their metabolites are taken into account during risk assessment of PCBs and related compounds.

  11. Hepatic differentiation of human pluripotent stem cells in miniaturized format suitable for high-throughput screen

    Directory of Open Access Journals (Sweden)

    Arnaud Carpentier

    2016-05-01

    Full Text Available The establishment of protocols to differentiate human pluripotent stem cells (hPSCs including embryonic (ESC and induced pluripotent (iPSC stem cells into functional hepatocyte-like cells (HLCs creates new opportunities to study liver metabolism, genetic diseases and infection of hepatotropic viruses (hepatitis B and C viruses in the context of specific genetic background. While supporting efficient differentiation to HLCs, the published protocols are limited in terms of differentiation into fully mature hepatocytes and in a smaller-well format. This limitation handicaps the application of these cells to high-throughput assays. Here we describe a protocol allowing efficient and consistent hepatic differentiation of hPSCs in 384-well plates into functional hepatocyte-like cells, which remain differentiated for more than 3 weeks. This protocol affords the unique opportunity to miniaturize the hPSC-based differentiation technology and facilitates screening for molecules in modulating liver differentiation, metabolism, genetic network, and response to infection or other external stimuli.

  12. A Site-Specific Phosphorylation of the Focal Adhesion Kinase Controls the Formation of Spheroid Cell Clusters

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Gosau, Martin; Kristensen, Lars Peter;

    2014-01-01

    Human dental follicle cells (DFCs) are ectomesenchymal multipotent stem cells that form spheroid cell clusters (SCCs) under serum free medium cell culture conditions (SFM). Until today, molecular mechanisms for the formation of SCCs are unknown. In this study a quantitative phosphoproteomics...... kinase (FAK) signaling pathway. In addition to the phosphoproteomics approach we showed that a specific phosphorylation of FAK (Y397) was required for the formation of SCCs. In conclusion, this study disclosed the phosphoproteome of SCCs for the first time and showed that the FAK signaling pathway...

  13. Charge Formation, Recombination, and Sweep-Out Dynamics in Organic Solar Cells

    OpenAIRE

    Cowan, Sarah R.; Banerji, Natalie; Leong, Wei Lin; Heeger, Alan J.

    2012-01-01

    This article presents a critical discussion of the various physical processes occurring in organic bulk heterojunction (BHJ) solar cells based on recent experimental results. The investigations span from photoexcitation to charge separation, recombination, and sweep-out to the electrodes. Exciton formation and relaxation in poly[N-9?-hepta-decanyl-2,7-carbazole-alt-5,5-(4',7'-di-2-thienyl-2',1',3'-benzothiadiazole) (PCDTBT) and poly-3(hexylthiophene) (P3HT) are discussed based on a fluorescen...

  14. Regularities of the formation of comutagenesis in irradiated human somatic cells

    International Nuclear Information System (INIS)

    The pattern (regularity) of co-mutagenic effects formation in healthy individuals' irradiated cells (T-lymphocytes) under verapamil and ascorbic acid treatment is studied. It is determined that the additional action of the investigated drugs in concentrations greater than the therapeutic one potentiate the effects of low doses of ionizing radiation by 1.5 times. Ascorbic acid and verapamil show co-mutagenic effects under high-dose irradiation (2.0 Gy), increasing the chromosomal aberration level by 1.4 times, regardless of the concentration of drugs.

  15. High-Throughput Cancer Cell Sphere Formation for Characterizing the Efficacy of Photo Dynamic Therapy in 3D Cell Cultures

    Science.gov (United States)

    Chen, Yu-Chih; Lou, Xia; Zhang, Zhixiong; Ingram, Patrick; Yoon, Euisik

    2015-07-01

    Photodynamic therapy (PDT), wherein light sensitive non-toxic agents are locally and selectively activated using light, has emerged as an appealing alternative to traditional cancer chemotherapy. Yet to date, PDT efficacy has been mostly characterized using 2D cultures. Compared to 2D cultures, 3D sphere culture generates unique spatial distributions of nutrients and oxygen for the cells that better mimics the in-vivo conditions. Using a novel polyHEMA (non-adherent polymer) fabrication process, we developed a microfluidic sphere formation platform that can (1) generate 1,024 uniform (size variation cancer spheres within a 2 cm by 2 cm core area, (2) culture spheres for more than 2 weeks, and (3) allow the retrieval of spheres. Using the presented platform, we have successfully characterized the different responses in 2D and 3D cell culture to PDT. Furthermore, we investigated the treatment resistance effect in cancer cells induced by tumor associated fibroblasts (CAF). Although the CAFs can enhance the resistance to traditional chemotherapy agents, no significant difference in PDT was observed. The preliminary results suggest that the PDT can be an attractive alternative cancer therapy, which is less affected by the therapeutic resistance induced by cancer associated cells.

  16. In-situ formation of growth-factor-loaded coacervate microparticle-embedded hydrogels for directing encapsulated stem cell fate.

    Science.gov (United States)

    Jeon, Oju; Wolfson, David W; Alsberg, Eben

    2015-04-01

    The spontaneous formation of coacervate microdroplet-laden photo-crosslinked hydrogels derived from the simple mixing of oxidized, methacrylated alginate (OMA) and methacrylated gelatin (GelMA) enables simultaneous creation of drug-laden microdroplets and encapsulation of stem cells in photopolymerized coacervate hydrogels under physiological conditions. This can be utilized as a novel platform for in situ formation of localized, sustained bioactive molecule delivery to encapsulate stem cells for therapeutic applications.

  17. Galectin-4 Reduces Migration and Metastasis Formation of Pancreatic Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Ana I Belo

    Full Text Available Galectin-4 (Gal-4 is a member of the galectin family of glycan binding proteins that shows a significantly higher expression in cystic tumors of the human pancreas and in pancreatic adenocarcinomas compared to normal pancreas. However, the putative function of Gal-4 in tumor progression of pancreatic cancer is still incompletely understood. In this study the role of Gal-4 in cancer progression was investigated, using a set of defined pancreatic cancer cell lines, Pa-Tu-8988S (PaTu-S and Pa-Tu-8988T (PaTu-T, as a model. These two cell lines are derived from the same liver metastasis of a human primary pancreatic adenocarcinoma, but differ in their growth characteristics and metastatic capacity. We demonstrated that Gal-4 expression is high in PaTu-S, which shows poor migratory properties, whereas much lower Gal-4 levels are observed in the highly metastatic cell line PaTu-T. In PaTu-S, Gal-4 is found in the cytoplasm, but it is also secreted and accumulates at the membrane at sites of contact with neighboring cells. Moreover, we show that Gal-4 inhibits metastasis formation by delaying migration of pancreatic cancer cells in vitro using a scratch assay, and in vivo using zebrafish (Danio rerio as an experimental model. Our data suggest that Gal-4 may act at the cell-surface of PaTu-S as an adhesion molecule to prevent release of the tumor cells, but has in addition a cytosolic function by inhibiting migration via a yet unknown mechanism.

  18. Neocartilage formation from predifferentiated human adipose derived stem cells in vivo

    Institute of Scientific and Technical Information of China (English)

    Xiao-bing JIN; Yong-sheng SUN; Ke ZHANG; Jing WANG; Xiao-dong JU; Si-quan LOU

    2007-01-01

    Aim: To examine the chondrogenic potential of human adipose derived stem cells (hASC) induced by human transforming growth factor beta2 (hTGF beta2) in vitro, and to investigate if predifferentiated hASC can produce neocartilage in vivo. Methods: hASC were isolated from subcutaneous adipose tissue and cul-tured in pellets with the addition of hTGF beta2. Chondrogenic differentiation was assayed by RT-PCR, Western blotting, toluidine blue staining, and immuno-histochemistry staining for collagen type Ⅱ. For the in vivo study, intact induced cell pellets or the released cells embedded in alginate gel with different concentra-tions were implanted subcutaneously in nude mice. Specimens were harvested at different time points and carried with histological and immunohistochemistry ex-amination to evaluate the cartilage formation. Results: RT-PCR analysis revealed that hASC produced aggrecan and collagen type Ⅱ after 7 d of induction and continued throughout the culture period. This was also demonstrated by the Western blot analysis, positive staining of toluidine blue, and immunohistochem-istry for collagen type Ⅱ. After reseeding in the monolayer, the cells isolated from the pellets displayed a polygonal morphology compared with the primary spindle shape, hASC were released from the induced cell pellets when embedded in alginate gel (implanted cell concentration=5x106/mL or higher). They produced neocartilage after 12 weeks in vivo culture; however, intact induced cell pellets implanted subcutaneously rapidly lost their differentiated phenotype. Conclusion:Chondrogenesis of hASC in vitro can be induced by combining pellet culture and hTGF beta2 treatment. Predifferentiated hASC embedded in alginate gel have the ability of producing neocartilage in vivo.

  19. Replication independent formation of extrachromosomal circular DNA in mammalian cell-free system.

    Directory of Open Access Journals (Sweden)

    Zoya Cohen

    Full Text Available Extrachromosomal circular DNA (eccDNA is a pool of circular double stranded DNA molecules found in all eukaryotic cells and composed of repeated chromosomal sequences. It was proposed to be involved in genomic instability, aging and alternative telomere lengthening. Our study presents novel mammalian cell-free system for eccDNA generation. Using purified protein extract we show that eccDNA formation does not involve de-novo DNA synthesis suggesting that eccDNA is generated through excision of chromosomal sequences. This process is carried out by sequence-independent enzymes as human protein extract can produce mouse-specific eccDNA from high molecular weight mouse DNA, and vice versa. EccDNA production does not depend on ATP, requires residual amounts of Mg(2+ and is enhanced by double strand DNA breaks.

  20. A stochastic model for the cell formation problem considering machine reliability

    Science.gov (United States)

    Esmailnezhad, Bahman; Fattahi, Parviz; Kheirkhah, Amir Saman

    2015-03-01

    This paper presents a new mathematical model to solve cell formation problem in cellular manufacturing systems, where inter-arrival time, processing time, and machine breakdown time are probabilistic. The objective function maximizes the number of operations of each part with more arrival rate within one cell. Because a queue behind each machine; queuing theory is used to formulate the model. To solve the model, two metaheurstic algorithms such as modified particle swarm optimization and genetic algorithm are proposed. For the generation of initial solutions in these algorithms, a new heuristic method is developed, which always creates feasible solutions. Both metaheurstic algorithms are compared against global solutions obtained from Lingo software's branch and bound (B&B). Also, a statistical method will be used for comparison of solutions of two metaheurstic algorithms. The results of numerical examples indicate that considering the machine breakdown has significant effect on block structures of machine-part matrixes.

  1. Development of a cell formation heuristic by considering realistic data using principal component analysis and Taguchi's method

    Science.gov (United States)

    Kumar, Shailendra; Sharma, Rajiv Kumar

    2015-12-01

    Over the last four decades of research, numerous cell formation algorithms have been developed and tested, still this research remains of interest to this day. Appropriate manufacturing cells formation is the first step in designing a cellular manufacturing system. In cellular manufacturing, consideration to manufacturing flexibility and production-related data is vital for cell formation. The consideration to this realistic data makes cell formation problem very complex and tedious. It leads to the invention and implementation of highly advanced and complex cell formation methods. In this paper an effort has been made to develop a simple and easy to understand/implement manufacturing cell formation heuristic procedure with considerations to the number of production and manufacturing flexibility-related parameters. The heuristic minimizes inter-cellular movement cost/time. Further, the proposed heuristic is modified for the application of principal component analysis and Taguchi's method. Numerical example is explained to illustrate the approach. A refinement in the results is observed with adoption of principal component analysis and Taguchi's method.

  2. Serial imaging of human embryonic stem-cell engraftment and teratoma formation in live mouse models

    Institute of Scientific and Technical Information of China (English)

    Martin G Pomper; Holly Hammond; Xiaobing Yu; Zhaohui Ye; Catherine A Foss; Doris D Lin; James J Fox; Linzhao Cheng

    2009-01-01

    Two new types of lentiviral vectors expressing a reporter transgene encoding either firefly lueiferase (fLue) for bioluminescence imaging or the HSV1 thymidine kinase (HSV1-TK) for radiopharmaceutical-based imaging were constructed to monitor human embryonic stem cell (hESC) engraftment and proliferation in live mice after trans-plantation. The constitutive expression of either transgene did not alter the properties of hESCs in the culture. We next monitored the formation of teratomas in SCID mice to test (1) whether the gene-modified hESCs maintain their developmental pluripotency, and (2) whether sustained reporter gene expression allows noninvasive, whole-body im-aging of hESC derivatives in a live mouse model. We observed teratoma formation from both types of gene-modified cells as well as wild-type bESCs 2-4 months after inoculation. Using an optical imaging system, bioluminescence from the fLuc-transduced hESCs was easily detected in mice bearing teratomas long before palpable tumors could be de-tected. To develop a noninvasive imaging method more readily translatable to the clinic, we also utilized HSV1-TK and its specific substrate, 1-(2'-deoxy-2'-fluoro-β-D-arabinofuranosyl)-5-[125I]iodouracil ([125I]FIAU), as a reporter/ probe pair. After systemic administration, [125I]FIAU is phosphorylated only by the transgene-encoded HSV1-TK enzyme and retained within transduced (and transplanted) cells, allowing sensitive and quantitative imaging by single-photon emission computed tomography. Noninvasive imaging methods such as these may enable us to moni-tor the presence and distribution of transplanted human stem cells repetitively within live recipients over a long term through the expression of a reporter gene.

  3. Bdellovibrio bacteriovorus Inhibits Staphylococcus aureus Biofilm Formation and Invasion into Human Epithelial Cells

    Science.gov (United States)

    Monnappa, Ajay K.; Dwidar, Mohammed; Seo, Jeong Kon; Hur, Jin-Hoe; Mitchell, Robert J.

    2014-01-01

    Bdellovibrio bacteriovorus HD100 is a predatory bacterium that attacks many Gram-negative human pathogens. A serious drawback of this strain, however, is its ineffectiveness against Gram-positive strains, such as the human pathogen Staphylococcus aureus. Here we demonstrate that the extracellular proteases produced by a host-independent B. bacteriovorus (HIB) effectively degrade/inhibit the formation of S. aureus biofilms and reduce its virulence. A 10% addition of HIB supernatant caused a 75% or greater reduction in S. aureus biofilm formation as well as 75% dispersal of pre-formed biofilms. LC-MS-MS analyses identified various B. bacteriovorus proteases within the supernatant, including the serine proteases Bd2269 and Bd2321. Tests with AEBSF confirmed that serine proteases were active in the supernatant and that they impacted S. aureus biofilm formation. The supernatant also possessed a slight DNAse activity. Furthermore, treatment of planktonic S. aureus with the supernatant diminished its ability to invade MCF-10a epithelial cells by 5-fold but did not affect the MCF-10a viability. In conclusion, this study illustrates the hitherto unknown ability of B. bacteriovorus to disperse Gram-positive pathogenic biofilms and mitigate their virulence.

  4. In vitro direct osteogenesis of murine embryonic stem cells without embryoid body formation.

    Science.gov (United States)

    Hwang, Yu-Shik; Polak, Julia M; Mantalaris, Athanasios

    2008-10-01

    Embryonic stem cells (ESCs) posses the ability to self-renew and differentiate into a multitude of lineages, including the osteogenic lineage in vitro. Currently, most approaches have focused on embryonic body (EB)-mediated osteogenic differentiation, which relies on formation of all three germ layers resulting in limited yields and labour-intensive culture processes. Our study aimed at developing an efficient culture strategy resulting in the upregulated in vitro osteogenic differentiation of murine ESCs (mESCs), which completely avoided EB formation. Specifically, mESCs were cultured in HepG2 conditioned medium for 3 days and then directed into osteogenic differentiation for 21 days without prior EB formation. The mineralised bone nodules generated were characterized by Alizarin red S-staining, phenotypic alkaline phosphatase expression, time-course analysis of ALPase activity, the presence of type I collagen and osteopontin, and osteocalcin, cbfa-1/runx-2, and osterix gene expression. Our method of direct osteogenic differentiation of mESCs represents a novel and efficient approach that results in enhanced yields and could have significant applications in bone tissue engineering.

  5. Heterozygous inactivation of the Nf1 gene in myeloid cells enhances neointima formation via a rosuvastatin-sensitive cellular pathway.

    Science.gov (United States)

    Stansfield, Brian K; Bessler, Waylan K; Mali, Raghuveer; Mund, Julie A; Downing, Brandon; Li, Fang; Sarchet, Kara N; DiStasi, Matthew R; Conway, Simon J; Kapur, Reuben; Ingram, David A

    2013-03-01

    Mutations in the NF1 tumor suppressor gene cause Neurofibromatosis type 1 (NF1). Neurofibromin, the protein product of NF1, functions as a negative regulator of Ras activity. Some NF1 patients develop cardiovascular disease, which represents an underrecognized disease complication and contributes to excess morbidity and mortality. Specifically, NF1 patients develop arterial occlusion resulting in tissue ischemia and sudden death. Murine studies demonstrate that heterozygous inactivation of Nf1 (Nf1(+/-)) in bone marrow cells enhances neointima formation following arterial injury. Macrophages infiltrate Nf1(+/-) neointimas, and NF1 patients have increased circulating inflammatory monocytes in their peripheral blood. Therefore, we tested the hypothesis that heterozygous inactivation of Nf1 in myeloid cells is sufficient for neointima formation. Specific ablation of a single copy of the Nf1 gene in myeloid cells alone mobilizes a discrete pro-inflammatory murine monocyte population via a cell autonomous and gene-dosage dependent mechanism. Furthermore, lineage-restricted heterozygous inactivation of Nf1 in myeloid cells is sufficient to reproduce the enhanced neointima formation observed in Nf1(+/-) mice when compared with wild-type controls, and homozygous inactivation of Nf1 in myeloid cells amplified the degree of arterial stenosis after arterial injury. Treatment of Nf1(+/-) mice with rosuvastatin, a stain with anti-inflammatory properties, significantly reduced neointima formation when compared with control. These studies identify neurofibromin-deficient myeloid cells as critical cellular effectors of Nf1(+/-) neointima formation and propose a potential therapeutic for NF1 cardiovascular disease. PMID:23197650

  6. Enhancement of committed hematopoietic stem cell colony formation by nandrolone decanoate after sublethal whole body irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Gallicchio, V.S.; Chen, M.G.; Watts, T.D.

    1984-11-01

    The ability of an anabolic steroid, nandrolone decanoate, to increase committed topoietic stem cell (CFU-gm, CFU-e, and BFU-e) colony formation after sublethal irradiation was evaluated. Immediately after receiving whole body irradiation and on the next two days, each mouse was injected intraperitoneally with nandrolone decanoate (1.25 mg) in propylene glycol. Irradiated control mice received only propylene glycol. Compared to controls, drug-treated mice showed marked peripheral blood leukocytosis and more stable packed red cell volume. Drug-treated mice also demonstrated increased erythropoiesis, as CFU-e/BFU-e concentrations from both marrow (9% to 581%) and spleen (15% to 797%) were elevated. Granulopoiesis was increased similarly, as CFU-gm concentrations from marrow (38% to 685%) and spleen (9% to 373%) were elevated. These results demonstrate that nandrolone decanoate enhances hematopoietic stem cell recovery after sublethal whole body irradiation. This suggests that following hematopoietic suppression, nandrolone decanoate may stimulate the recovery of hematopoiesis at the stem cell level and in peripheral blood.

  7. Cells in the monkey ponto-medullary reticular formation modulate their activity with slow finger movements.

    Science.gov (United States)

    Soteropoulos, Demetris S; Williams, Elizabeth R; Baker, Stuart N

    2012-08-15

    Recent work has shown that the primate reticulospinal tract can influence spinal interneurons and motoneurons involved in control of the hand. However, demonstrating connectivity does not reveal whether reticular outputs are modulated during the control of different types of hand movement. Here, we investigated how single unit discharge in the pontomedullary reticular formation (PMRF) modulated during performance of a slow finger movement task in macaque monkeys. Two animals performed an index finger flexion–extension task to track a target presented on a computer screen; single units were recorded both from ipsilateral PMRF (115 cells) and contralateral primary motor cortex (M1, 210 cells). Cells in both areas modulated their activity with the task (M1: 87%, PMRF: 86%). Some cells (18/115 in PMRF; 96/210 in M1) received sensory input from the hand, showing a short-latency modulation in their discharge following a rapid passive extension movement of the index finger. Effects in ipsilateral electromyogram to trains of stimuli were recorded at 45 sites in the PMRF. These responses involved muscles controlling the digits in 13/45 sites (including intrinsic hand muscles, 5/45 sites). We conclude that PMRF may contribute to the control of fine finger movements, in addition to its established role in control of more proximal limb and trunk movements. This finding may be especially important in understanding functional recovery after brain lesions such as stroke. PMID:22641776

  8. Flat leaf formation realized by cell-division control and mutual recessive gene regulation.

    Science.gov (United States)

    Hayakawa, Yoshinori; Tachikawa, Masashi; Mochizuki, Atsushi

    2016-09-01

    Most of the land plants generally have dorsoventrally flat leaves, maximizing the surface area of both upper (adaxial) side and lower (abaxial) side. The former is specialized for light capturing for photosynthesis and the latter is specialized for gas exchange. From findings of molecular genetics, it has been considered that the coupled dynamics between tissue morphogenesis and gene regulation for cell identity is responsible for making flat leaves. The hypothesis claims that a flat leaf is generated under two assumptions, (i) two mutually recessive groups of genes specify adaxial and abaxial sides of a leaf, (ii) cell divisions are induced at the limited region in the leaf margin where both of two groups are expressed. We examined the plausibility and possibility of this hypothesis from the dynamical point of view. We studied a mathematical model where two processes are coupled, tissue morphogenesis induced by cell division and deformation, and dynamics of gene regulations. From the analysis of the model we found that the classically believed hypothesis is not sufficient to generate flat leaves with high probability. We examined several different modifications and revision of the model. Then we found that a simple additional rule of polarized cell division facilitates flat leaf formation. The result of our analysis gives prediction of possible mechanism, which can be easily verified in experiments. PMID:27287339

  9. The Secreted Protease PrtA Controls Cell Growth, Biofilm Formation and Pathogenicity in Xylella fastidiosa

    Science.gov (United States)

    Gouran, Hossein; Gillespie, Hyrum; Nascimento, Rafael; Chakraborty, Sandeep; Zaini, Paulo A.; Jacobson, Aaron; Phinney, Brett S.; Dolan, David; Durbin-Johnson, Blythe P.; Antonova, Elena S.; Lindow, Steven E.; Mellema, Matthew S.; Goulart, Luiz R.; Dandekar, Abhaya M.

    2016-01-01

    Pierce’s disease (PD) is a deadly disease of grapevines caused by the Gram-negative bacterium Xylella fastidiosa. Though disease symptoms were formerly attributed to bacteria blocking the plant xylem, this hypothesis is at best overly simplistic. Recently, we used a proteomic approach to characterize the secretome of X. fastidiosa, both in vitro and in planta, and identified LesA as one of the pathogenicity factors of X. fastidiosa in grapevines that leads to leaf scorching and chlorosis. Herein, we characterize another such factor encoded by PD0956, designated as an antivirulence secreted protease “PrtA” that displays a central role in controlling in vitro cell proliferation, length, motility, biofilm formation, and in planta virulence. The mutant in X. fastidiosa exhibited reduced cell length, hypermotility (and subsequent lack of biofilm formation) and hypervirulence in grapevines. These findings are supported by transcriptomic and proteomic analyses with corresponding plant infection data. Of particular interest, is the hypervirulent response in grapevines observed when X. fastidiosa is disrupted for production of PrtA, and that PD-model tobacco plants transformed to express PrtA exhibited decreased symptoms after infection by X. fastidiosa. PMID:27492542

  10. The Secreted Protease PrtA Controls Cell Growth, Biofilm Formation and Pathogenicity in Xylella fastidiosa.

    Science.gov (United States)

    Gouran, Hossein; Gillespie, Hyrum; Nascimento, Rafael; Chakraborty, Sandeep; Zaini, Paulo A; Jacobson, Aaron; Phinney, Brett S; Dolan, David; Durbin-Johnson, Blythe P; Antonova, Elena S; Lindow, Steven E; Mellema, Matthew S; Goulart, Luiz R; Dandekar, Abhaya M

    2016-01-01

    Pierce's disease (PD) is a deadly disease of grapevines caused by the Gram-negative bacterium Xylella fastidiosa. Though disease symptoms were formerly attributed to bacteria blocking the plant xylem, this hypothesis is at best overly simplistic. Recently, we used a proteomic approach to characterize the secretome of X. fastidiosa, both in vitro and in planta, and identified LesA as one of the pathogenicity factors of X. fastidiosa in grapevines that leads to leaf scorching and chlorosis. Herein, we characterize another such factor encoded by PD0956, designated as an antivirulence secreted protease "PrtA" that displays a central role in controlling in vitro cell proliferation, length, motility, biofilm formation, and in planta virulence. The mutant in X. fastidiosa exhibited reduced cell length, hypermotility (and subsequent lack of biofilm formation) and hypervirulence in grapevines. These findings are supported by transcriptomic and proteomic analyses with corresponding plant infection data. Of particular interest, is the hypervirulent response in grapevines observed when X. fastidiosa is disrupted for production of PrtA, and that PD-model tobacco plants transformed to express PrtA exhibited decreased symptoms after infection by X. fastidiosa. PMID:27492542

  11. The mycotoxin zearalenone enhances cell proliferation, colony formation and promotes cell migration in the human colon carcinoma cell line HCT116.

    Science.gov (United States)

    Abassi, Haila; Ayed-Boussema, Imen; Shirley, Sarah; Abid, Salwa; Bacha, Hassen; Micheau, Olivier

    2016-07-01

    Zearalenone (ZEN) and Aflatoxin B1 (AFB1) are fungal secondary metabolites produced by Fusarium and Aspergillus genera, respectively. These mycotoxins are found world-wide as corn and wheat contaminants. AFB1 is probably the most toxic and carcinogenic mycotoxin. It has been demonstrated to be mutagenic, genotoxic, and hepatocarcinogenic. ZEN is a non-steroidal estrogenic mycotoxin that displays hepatotoxicity, immunotoxicity and genotoxicity. Its mutagenic and carcinogenic properties have so far remained controversial and questionable. Using the colon carcinoma cell line HCT116, we will show here that ZEN, at low concentrations, enhances cell proliferation, increases colony formation and fastens cell migration after wound healing. The highest effect of ZEN was observed at a concentration 10 times lower as compared to AFB1. Our findings suggest thus that this mycotoxin exhibits carcinogenesis-like properties in HCT116 cells. PMID:27084041

  12. Formation of germline chimera Gaok chicken used circulation primordial germ cells (circulation PGCs fresh and thawed

    Directory of Open Access Journals (Sweden)

    Kostaman T

    2014-03-01

    Full Text Available Formation of germline chimeras by transfer of chicken primordial germ cells (PGCs is one of the effective techniques for preservation and regeneration of genetic resources in chickens. This study attempted to form germline chimeras of Gaok chicken buy purifying circulated PGCs of donor embryo before it is transferred to the recipient (White Leghorn chickens=WL and studied the ability of recipient embryo on survival in incubators, and hatchability. This study used 200 fertile eggs of Gaok and 90 fertile WL breed all of the eggs was incubated at 380C and 60% humidity in a portable incubator. PGCs-circulation of the blood collected Gaok embryos at stage 14-16 were taken from the dorsal aorta, and then purified by centrifugation method using nycodenz. PGCs-circulation results further purification frozen in liquid nitrogen before being transferred to the recipient embryo. The results showed that for the development of embryos transferred to the fresh circulation of PGCs-circulation as many as 25 cells can survive up to day 14, while one of the transferred of 50 and 100 cells into recipient embryos was hatched (10%. On the contrari recipient embryos that are transferred to the frozen PGCs-circulation the embryos development was shorter, and only survived until day 10th (treatment 25 cells, day 14th (treatment of 50 cells and day 17th (treatment of 100 cells. It is concluded that the amount of PGCs-circulation embryos transferred to the recipient is one factor that influence the success of the development germline chimeras.

  13. Overexpression of stress-related genes enhances cell viability and velum formation in Sherry wine yeasts.

    Science.gov (United States)

    Fierro-Risco, Jesús; Rincón, Ana María; Benítez, Tahía; Codón, Antonio C

    2013-08-01

    Flor formation and flor endurance have been related to ability by Saccharomyces cerevisiae flor yeasts to resist hostile conditions such as oxidative stress and the presence of acetaldehyde and ethanol. Ethanol and acetaldehyde toxicity give rise to formation of reactive oxygen species (ROS) and loss of cell viability. Superoxide dismutases Sod1p and Sod2p and other proteins such as Hsp12p are involved in oxidative stress tolerance. In this study, genes SOD1, SOD2, and HSP12 were overexpressed in flor yeast strains FJF206, FJF414 and B16. In the SOD1 and SOD2 transformant strains superoxide dismutases encoded by genes SOD1 and SOD2 increased their specific activity considerably as a direct result of overexpression of genes SOD1 and SOD2, indirectly, catalase, glutathione reductase, and glutathione peroxidase activities increased too. The HSP12 transformant strains showed higher levels of glutathione peroxidase and reductase activities. These transformant strains showed an increase in intracellular glutathione content, a reduction in peroxidized lipid concentration, and higher resistance to oxidative stress conditions. As a result, flor formation by these strains took place more rapidly than by their parental strains, velum being thicker and with higher percentages of viable cells. In addition, a slight decrease in ethanol and glycerol concentrations, and an increase in acetaldehyde were detected in wines matured under velum formed by transformant strains, as compared to their parental strains. In the industry, velum formed by transformant strains with increased viability may result in acceleration of both metabolism and wine aging, thus reducing time needed for wine maturation. PMID:23553032

  14. Thickness-controllable electrospun fibers promote tubular structure formation by endothelial progenitor cells

    Directory of Open Access Journals (Sweden)

    Hong JK

    2015-02-01

    Full Text Available Jong Kyu Hong,1,2 Ju Yup Bang,3 Guan Xu,4 Jun-Hee Lee,1 Yeon-Ju Kim,1 Ho-Jun Lee,5 Han Seong Kim,3 Sang-Mo Kwon1,2,6 1Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan, South Korea; 2Conversence Stem Cell Research Center, Medical Research Institute, School of Medicine, Pusan National University, Yangsan, South Korea; 3Department of Organic Material Science, Pusan National University, Geumjeong-gu, Busan, South Korea; 4Department of Radiology, School of Medicine, University of Michigan, Ann Arbor, MI, USA; 5Department of Electrical Engineering, Pusan National University, Geumjeong-gu, Busan, South Korea; 6Immunoregulatory Therapeutics Group in Brain Busan 21 Project, Department of Physiology, Pusan National University School of Medicine, Yangsan, South Korea Abstract: Controlling the thickness of an electrospun nanofibrous scaffold by altering its pore size has been shown to regulate cell behaviors such as cell infiltration into a three-dimensional (3D scaffold. This is of great importance when manufacturing tissue-engineering scaffolds using an electrospinning process. In this study, we report the development of a novel process whereby additional aluminum foil layers were applied to the accumulated electrospun fibers of an existing aluminum foil collector, effectively reducing the incidence of charge buildup. Using this process, we fabricated an electrospun scaffold with a large pore (pore size >40 µm while simultaneously controlling the thickness. We demonstrate that the large pore size triggered rapid infiltration (160 µm in 4 hours of cell culture of individual endothelial progenitor cells (EPCs and rapid cell colonization after seeding EPC spheroids. We confirmed that the 3D, but not two-dimensional, scaffold structures regulated tubular structure formation by the EPCs. Thus, incorporation of stem cells into a highly

  15. Lipid body formation plays a central role in cell fate determination during developmental differentiation of Myxococcus xanthus.

    Science.gov (United States)

    Hoiczyk, Egbert; Ring, Michael W; McHugh, Colleen A; Schwär, Gertrud; Bode, Edna; Krug, Daniel; Altmeyer, Matthias O; Lu, Jeff Zhiqiang; Bode, Helge B

    2009-10-01

    Cell differentiation is widespread during the development of multicellular organisms, but rarely observed in prokaryotes. One example of prokaryotic differentiation is the gram-negative bacterium Myxococcus xanthus. In response to starvation, this gliding bacterium initiates a complex developmental programme that results in the formation of spore-filled fruiting bodies. How the cells metabolically support the necessary complex cellular differentiation from rod-shaped vegetative cells into spherical spores is unknown. Here, we present evidence that intracellular lipid bodies provide the necessary metabolic fuel for the development of spores. Formed at the onset of starvation, these lipid bodies gradually disappear until they are completely used up by the time the cells have become mature spores. Moreover, it appears that lipid body formation in M. xanthus is an important initial step indicating cell fate during differentiation. Upon starvation, two subpopulations of cells occur: cells that form lipid bodies invariably develop into spores, while cells that do not form lipid bodies end up becoming peripheral rods, which are cells that lack signs of morphological differentiation and stay in a vegetative-like state. These data indicate that lipid bodies not only fuel cellular differentiation but that their formation represents the first known morphological sign indicating cell fate during differentiation. PMID:19788540

  16. NF-kB activity-dependent P-selectin involved in ox-LDL-induced foam cell formation in U937 cell

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yi, E-mail: wangyi2004a@126.com [Department of Cardiology, Shanghai First People' s Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200080 (China); Wang, Xiang; Sun, Minghui; Zhang, Zhenyu; Cao, Heng; Chen, Xiaoqing [Department of Cardiology, Shanghai First People' s Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200080 (China)

    2011-08-05

    Highlights: {yields} Ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. {yields} Ox-LDL induced expression of P-selectin through degradation of IkBa and augment of NF-kB activity and protein level during macrophage-derived foam cell formation. {yields} P-selectin and NF-kB may be identified as pivotal regulators of ox-LDL-induced foam cell formation. {yields} Therapy based on the inhibition of P-selectin and NF-kB may complement conventional treatments to prevent atherosclerosis. -- Abstract: Oxidized low-density lipoprotein (ox-LDL) plays a critical role in regulation of atherosclerosis. However, little is known about the role of Nuclear factor kB (NF-kB) activity-dependent P-selectin in ox-LDL-induced foam cell formation during atherosclerosis. In this study, we first investigated ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. Treatment of U937 cells with ox-LDL increased lipid accumulation as well as intracellular cholesterol content. Next, a comparative analysis of gene expression profiling using cDNA microarray and Real-time-PCR indicated that ox-LDL exposure induced, in three treated groups, an extremely marked increase in the mRNA level of P-selectin. Protein levels of P-selectin and its upstream regulators IkBa and NF-kB showed that NF-kB pathway is involved in the ox-LDL-induced foam cell formation. Finally, overexpression of NF-kB significantly accelerated, whereas, inhibition of NF-kB with siRNA remarkably attenuated ox-LDL-induced macrophage-derived foam cell formation. It was concluded that the activity of NF-kB is augmented during macrophage-derived foam cell formation. Activation of NF-kB increased, whereas, inhibition of NF-kB decreased ox-LDL-induced P-selectin expression and lipid accumulation in macrophages, suggesting ox-LDL induced expression of P-selectin through degradation of IkBa and activation of NF-kB in the

  17. Teratoma Formation by Human Embryonic Stem Cells is site-dependent and enhanced by the presence of Matrigel

    DEFF Research Database (Denmark)

    Prokhorova, Tatyana A; Harkness, Linda M; Frandsen, Ulrik;

    2008-01-01

    When implanted into immunodeficient mice, human embryonic stem cells (hESC) give rise to teratoma, tumour-like formations containing tissues belonging to all three germ layers. The ability to form teratoma is a sine qua non characteristic of pluripotent stem cells. However, limited data...... of differentiated to un-differentiated tissues was significantly decreased suggesting defective pluripotency of the cells. In conclusion, subcutaneous implantation of hESC in presence of Matrigel appears to be the most efficient, reproducible and the easiest approach for teratoma formation by hESC. Also, teratoma...

  18. Dolomitized cells within chert of the Permian Assistência Formation, Paraná Basin, Brazil

    Science.gov (United States)

    Calça, Cléber P.; Fairchild, Thomas R.; Cavalazzi, Barbara; Hachiro, Jorge; Petri, Setembrino; Huila, Manuel Fernando Gonzalez; Toma, Henrique E.; Araki, Koiti

    2016-04-01

    Dolomitic microscopic structures in the form of microspheres, "horseshoe- shaped" objects, and thin botryoidal crusts found within microfossiliferous chert within stromatolites of the Evaporite Bed (EB) of the Permian Assistência Formation, Irati Subgroup, Paraná Basin, Brazil, have been investigated by means of optical microscopy, X-ray fluorescence, scanning electron microscopy, Raman spectrometry and energy-dispersive X-ray spectrometry. The microspheres were identified as dolomitized coccoidal cyanobacteria based on similarity in size, spheroidal and paired hemispheroidal morphologies and colonial habit to co-occurring silicified organic-walled cyanobacteria embedded within the same microfabric and rock samples. The co-occurrence of dolomite, pyrite framboids, and abundant dispersed carbonaceous material and silicified cells is consistent with a hypersaline depositional environment with abundant cyanobacterial mats and elevated Mg2 +/Ca2 + ratios and reducing conditions with active anoxic microbial processes near the water-(bio)sediment interface. The abundance of extracellular polymeric substances facilitated anoxic microbial processes (sulfate reduction), providing essential conditions for possible primary microbially induced dolomitization. In most of the dolomitized cells dolomite occurs only as an external layer; in fully dolomitized cells magnesium is richest in the outermost layer. Presumably, the dolomitization process was favored by the presence of anoxic microbial degraders and negatively charged functional groups at the surface of the cyanobacterial cells. Botryoidal dolomite rims of silica-filled fenestrae formed by a similar process and inherited the botryoidal morphology of the cell as originally lining the fenestrae. Silicification interrupted the dolomitization of the largely organic biosediment, mostly by permineralization, but locally by substitution, thereby preserving not only dolomitic microspheres, but also huge numbers of structurally

  19. Nanolayer formation on titanium by phosphonated gelatin for cell adhesion and growth enhancement

    Directory of Open Access Journals (Sweden)

    Zhou X

    2015-09-01

    Full Text Available Xiaoyue Zhou,1,2,* Shin-Hye Park,1,* Hongli Mao,3 Takashi Isoshima,1 Yi Wang,2 Yoshihiro Ito1,3 1Nano Medical Engineering Laboratory, RIKEN, Wako, Saitama, Japan; 2Department of Regenerative Medicine, School of Pharmaceutical Sciences, Jilin University, Changchun, Jilin, People’s Republic of China; 3Emergent Bioengineering Materials Research Team, RIKEN Center for Emergent Matter Science, Wako, Saitama, Japan *These authors contributed equally to this work Abstract: Phosphonated gelatin was prepared for surface modification of titanium to stimulate cell functions. The modified gelatin was synthesized by coupling with 3-aminopropylphosphonic acid using water-soluble carbodiimide and characterized by 31P nuclear magnetic resonance and gel permeation chromatography. Circular dichroism revealed no differences in the conformations of unmodified and phosphonated gelatin. However, the gelation temperature was changed by the modification. Even a high concentration of modified gelatin did not form a gel at room temperature. Time-of-flight secondary ion mass spectrometry showed direct bonding between the phosphonated gelatin and the titanium surface after binding. The binding behavior of phosphonated gelatin on the titanium surface was quantitatively analyzed by a quartz crystal microbalance. Ellipsometry showed the formation of a several nanometer layer of gelatin on the surface. Contact angle measurement indicated that the modified titanium surface was hydrophobic. Enhancement of the attachment and spreading of MC-3T3L1 osteoblastic cells was observed on the phosphonated gelatin-modified titanium. These effects on cell adhesion also led to growth enhancement. Phosphonation of gelatin was effective for preparation of a cell-stimulating titanium surface. Keywords: phosphonated gelatin, surface modification, titanium, cell adhesion

  20. GABA agonist promoted formation of low affinity GABA receptors on cerebellar granule cells is restricted to early development

    DEFF Research Database (Denmark)

    Belhage, B; Hansen, G H; Schousboe, A;

    1988-01-01

    The ability of the GABA receptor agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) to promote formation of low affinity GABA receptors on cerebellar granule cells was tested using primary cultures of these neurons. Granule cells were exposed to THIP (150 microM) for 6 hr after......, respectively, 4, 7, 10 and 14 days in culture. It was found that THIP treatment of 4- and 7-day-old cultures led to formation of low affinity GABA receptors, whereas such receptors could not be detected after THIP treatment in the older cultures (10 and 14 days) in spite of the fact that these cultured granule...... cells expressed a high density of high affinity GABA receptors. It is concluded that the ability of THIP to promote formation of low affinity GABA receptors on cerebellar granule cells is restricted to an early developmental period....

  1. In vivo ectopic bone formation by devitalized mineralized stem cell carriers produced under mineralizing culture condition.

    Science.gov (United States)

    Chai, Yoke Chin; Geris, Liesbet; Bolander, Johanna; Pyka, Grzegorz; Van Bael, Simon; Luyten, Frank P; Schrooten, Jan

    2014-12-01

    Functionalization of tissue engineering scaffolds with in vitro-generated bone-like extracellular matrix (ECM) represents an effective biomimetic approach to promote osteogenic differentiation of stem cells in vitro. However, the bone-forming capacity of these constructs (seeded with or without cells) is so far not apparent. In this study, we aimed at developing a mineralizing culture condition to biofunctionalize three-dimensional (3D) porous scaffolds with highly mineralized ECM in order to produce devitalized, osteoinductive mineralized carriers for human periosteal-derived progenitors (hPDCs). For this, three medium formulations [i.e., growth medium only (BM1), with ascorbic acid (BM2), and with ascorbic acid and dexamethasone (BM3)] supplemented with calcium (Ca(2+)) and phosphate (PO4 (3-)) ions simultaneously as mineralizing source were investigated. The results showed that, besides the significant impacts on enhancing cell proliferation (the highest in BM3 condition), the formulated mineralizing media differentially regulated the osteochondro-related gene markers in a medium-dependent manner (e.g., significant upregulation of BMP2, bone sialoprotein, osteocalcin, and Wnt5a in BM2 condition). This has resulted in distinguished cell populations that were identifiable by specific gene signatures as demonstrated by the principle component analysis. Through devitalization, mineralized carriers with apatite crystal structures unique to each medium condition (by X-ray diffraction and SEM analysis) were obtained. Quantitatively, BM3 condition produced carriers with the highest mineral and collagen contents as well as human-specific VEGF proteins, followed by BM2 and BM1 conditions. Encouragingly, all mineralized carriers (after reseeded with hPDCs) induced bone formation after 8 weeks of subcutaneous implantation in nude mice models, with BM2-carriers inducing the highest bone volume, and the lowest in the BM3 condition (as quantitated by nano-computed tomography

  2. Slow cluster formation of purified human or rhesus T cells requires protein kinase C and LFA-1.

    Science.gov (United States)

    Eylar, E H; Molina, C; Báez, I; Kessler, M

    1996-03-01

    Homotropic T cell adhesion, as generally studied, consists of a rapid, transient binding process that is measured over a 15-120 min. period. Here we report a slow type of adhesion process occurring with human or rhesus T cells, purified from peripheral blood, that manifests itself by the formation of rounded, multi-layer clusters which may contain hundreds of cells. The maximal number and size of the clusters peak 1-2 days after the addition of phorbol ester, an absolute requirement. The number of clusters formed is proportional to phorbol ester concentration up to 1.25 ng/mL. Phorbol esters such as phorbol myristate acetate (PMA), phorbol dibutyrate (PDB), and 7-octylindolactam (OIL) induced optimal cluster formation at 1-13 ng/mL, levels slightly higher than that required to induce mitogenesis of purified T cells. Phorbol itself and the alpha-form of the ester were inactive. Both cluster formation and mitogenesis (stimulated by Con A or anti-CD3) are completely inhibited by staurosporin at 12.5 ng/mL. Even at 2.5 ng/mL, 74% of cluster formation was inhibited, which strongly implies a crucial role for protein kinase C. In the presence of accessory cells, T cell clusters were suppressed. Monoclonal Ab such as anti-CD3, mouse anti-CD3 followed by anti-mouse IgG, anti-CD4, anti-CD4A, anti-CD2, anti-CD8, and anti-CD45 did not induce cluster formation. None were inhibitory or stimulatory in the presence of PMA, except for anti-CD3 which enhanced cluster formation by 26%. However, anti-LFA-1 beta-chain (mouse monoclonal) completely blocked cluster formation over the range studied (63-1000 ng/mL) for both human and rhesus cells; rat anti-LFA-1 only blocked human cell adhesion. Anti LFA-1 only partially inhibited T cell mitogenesis. These results show that slow cluster formation shares the LFA-1 and phorbol ester requirements of the rapid adhesion of T cells requiring LFA-1 and ICAM-1. However, cluster occurs at a very low phorbol ester concentration, appears more

  3. The inhibition of macrophage foam cell formation by 9-cis β-carotene is driven by BCMO1 activity.

    Directory of Open Access Journals (Sweden)

    Noa Zolberg Relevy

    Full Text Available Atherosclerosis is a major cause of morbidity and mortality in developed societies, and begins when activated endothelial cells recruit monocytes and T-cells from the bloodstream into the arterial wall. Macrophages that accumulate cholesterol and other fatty materials are transformed into foam cells. Several epidemiological studies have demonstrated that a diet rich in carotenoids is associated with a reduced risk of heart disease; while previous work in our laboratory has shown that the 9-cis β-carotene rich alga Dunaliella inhibits atherogenesis in mice. The effect of 9-cis β-carotene on macrophage foam cell formation has not yet been investigated. In the present work, we sought to study whether the 9-cis β-carotene isomer, isolated from the alga Dunaliella, can inhibit macrophage foam cell formation upon its conversion to retinoids. The 9-cis β-carotene and Dunaliella lipid extract inhibited foam cell formation in the RAW264.7 cell line, similar to 9-cis retinoic acid. Furthermore, dietary enrichment with the algal powder in mice resulted in carotenoid accumulation in the peritoneal macrophages and in the inhibition of foam cell formation ex-vivo and in-vivo. We also found that the β-carotene cleavage enzyme β-carotene 15,15'-monooxygenase (BCMO1 is expressed and active in macrophages. Finally, 9-cis β-carotene, as well as the Dunaliella extract, activated the nuclear receptor RXR in hepa1-6 cells. These results indicate that dietary carotenoids, such as 9-cis β-carotene, accumulate in macrophages and can be locally cleaved by endogenous BCMO1 to form 9-cis retinoic acid and other retinoids. Subsequently, these retinoids activate the nuclear receptor RXR that, along with additional nuclear receptors, can affect various metabolic pathways, including those involved in foam cell formation and atherosclerosis.

  4. The Inhibition of Macrophage Foam Cell Formation by 9-Cis β-Carotene Is Driven by BCMO1 Activity

    Science.gov (United States)

    Zolberg Relevy, Noa; Bechor, Sapir; Harari, Ayelet; Ben-Amotz, Ami; Kamari, Yehuda; Harats, Dror; Shaish, Aviv

    2015-01-01

    Atherosclerosis is a major cause of morbidity and mortality in developed societies, and begins when activated endothelial cells recruit monocytes and T-cells from the bloodstream into the arterial wall. Macrophages that accumulate cholesterol and other fatty materials are transformed into foam cells. Several epidemiological studies have demonstrated that a diet rich in carotenoids is associated with a reduced risk of heart disease; while previous work in our laboratory has shown that the 9-cis β-carotene rich alga Dunaliella inhibits atherogenesis in mice. The effect of 9-cis β-carotene on macrophage foam cell formation has not yet been investigated. In the present work, we sought to study whether the 9-cis β-carotene isomer, isolated from the alga Dunaliella, can inhibit macrophage foam cell formation upon its conversion to retinoids. The 9-cis β-carotene and Dunaliella lipid extract inhibited foam cell formation in the RAW264.7 cell line, similar to 9-cis retinoic acid. Furthermore, dietary enrichment with the algal powder in mice resulted in carotenoid accumulation in the peritoneal macrophages and in the inhibition of foam cell formation ex-vivo and in-vivo. We also found that the β-carotene cleavage enzyme β-carotene 15,15’-monooxygenase (BCMO1) is expressed and active in macrophages. Finally, 9-cis β-carotene, as well as the Dunaliella extract, activated the nuclear receptor RXR in hepa1-6 cells. These results indicate that dietary carotenoids, such as 9-cis β-carotene, accumulate in macrophages and can be locally cleaved by endogenous BCMO1 to form 9-cis retinoic acid and other retinoids. Subsequently, these retinoids activate the nuclear receptor RXR that, along with additional nuclear receptors, can affect various metabolic pathways, including those involved in foam cell formation and atherosclerosis. PMID:25629601

  5. Vascularized bone tissue formation induced by fiber-reinforced scaffolds cultured with osteoblasts and endothelial cells.

    Science.gov (United States)

    Liu, Xinhui; Zhang, Guoping; Hou, Chuanyong; Wang, Hua; Yang, Yelin; Guan, Guoping; Dong, Wei; Gao, Hongyang; Feng, Qingling

    2013-01-01

    The repair of the damaged bone tissue caused by damage or bone disease was still a problem. Current strategies including the use of autografts and allografts have the disadvantages, namely, diseases transmission, tissue availability and donor morbidity. Bone tissue engineering has been developed and regarded as a new way of regenerating bone tissues to repair or substitute damaged or diseased ones. The main limitation in engineering in vitro tissues is the lack of a sufficient blood vessel system, the vascularization. In this paper, a new-typed hydroxyapatite/collagen composite scaffold which was reinforced by chitosan fibers and cultured with osteoblasts and endothelial cells was fabricated. General observation, histological observation, detection of the degree of vascularization, and X-ray examination had been done to learn the effect of vascularized bone repair materials on the regeneration of bone. The results show that new vessel and bone formed using implant cultured with osteoblasts and endothelial cells. Nanofiber-reinforced scaffold cultured with osteoblasts and endothelial cells can induce vascularized bone tissue formation. PMID:24369019

  6. Vascularized Bone Tissue Formation Induced by Fiber-Reinforced Scaffolds Cultured with Osteoblasts and Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Xinhui Liu

    2013-01-01

    Full Text Available The repair of the damaged bone tissue caused by damage or bone disease was still a problem. Current strategies including the use of autografts and allografts have the disadvantages, namely, diseases transmission, tissue availability and donor morbidity. Bone tissue engineering has been developed and regarded as a new way of regenerating bone tissues to repair or substitute damaged or diseased ones. The main limitation in engineering in vitro tissues is the lack of a sufficient blood vessel system, the vascularization. In this paper, a new-typed hydroxyapatite/collagen composite scaffold which was reinforced by chitosan fibers and cultured with osteoblasts and endothelial cells was fabricated. General observation, histological observation, detection of the degree of vascularization, and X-ray examination had been done to learn the effect of vascularized bone repair materials on the regeneration of bone. The results show that new vessel and bone formed using implant cultured with osteoblasts and endothelial cells. Nanofiber-reinforced scaffold cultured with osteoblasts and endothelial cells can induce vascularized bone tissue formation.

  7. Neurl4 contributes to germ cell formation and integrity in Drosophila

    Directory of Open Access Journals (Sweden)

    Jennifer Jones

    2015-08-01

    Full Text Available Primordial germ cells (PGCs form at the posterior pole of the Drosophila embryo, and then migrate to their final destination in the gonad where they will produce eggs or sperm. Studies of the different stages in this process, including assembly of germ plasm in the oocyte during oogenesis, specification of a subset of syncytial embryonic nuclei as PGCs, and migration, have been informed by genetic analyses. Mutants have defined steps in the process, and the identities of the affected genes have suggested biochemical mechanisms. Here we describe a novel PGC phenotype. When Neurl4 activity is reduced, newly formed PGCs frequently adopt irregular shapes and appear to bud off vesicles. PGC number is also reduced, an effect exacerbated by a separate role for Neurl4 in germ plasm formation during oogenesis. Like its mammalian homolog, Drosophila Neurl4 protein is concentrated in centrosomes and downregulates centrosomal protein CP110. Reducing CP110 activity suppresses the abnormal PGC morphology of Neurl4 mutants. These results extend prior analyses of Neurl4 in cultured cells, revealing a heightened requirement for Neurl4 in germ-line cells in Drosophila.

  8. A multi-cell, multi-scale model of vertebrate segmentation and somite formation.

    Directory of Open Access Journals (Sweden)

    Susan D Hester

    2011-10-01

    Full Text Available Somitogenesis, the formation of the body's primary segmental structure common to all vertebrate development, requires coordination between biological mechanisms at several scales. Explaining how these mechanisms interact across scales and how events are coordinated in space and time is necessary for a complete understanding of somitogenesis and its evolutionary flexibility. So far, mechanisms of somitogenesis have been studied independently. To test the consistency, integrability and combined explanatory power of current prevailing hypotheses, we built an integrated clock-and-wavefront model including submodels of the intracellular segmentation clock, intercellular segmentation-clock coupling via Delta/Notch signaling, an FGF8 determination front, delayed differentiation, clock-wavefront readout, and differential-cell-cell-adhesion-driven cell sorting. We identify inconsistencies between existing submodels and gaps in the current understanding of somitogenesis mechanisms, and propose novel submodels and extensions of existing submodels where necessary. For reasonable initial conditions, 2D simulations of our model robustly generate spatially and temporally regular somites, realistic dynamic morphologies and spontaneous emergence of anterior-traveling stripes of Lfng. We show that these traveling stripes are pseudo-waves rather than true propagating waves. Our model is flexible enough to generate interspecies-like variation in somite size in response to changes in the PSM growth rate and segmentation-clock period, and in the number and width of Lfng stripes in response to changes in the PSM growth rate, segmentation-clock period and PSM length.

  9. Long segmental hyperplasia of interstitial cells of Cajal with giant diverticulum formation.

    Science.gov (United States)

    Xue, Liyan; Qiu, Tian; Song, Ying; Shan, Ling; Liu, Xiuyun; Guo, Lei; Ying, Jianming; Zou, Shuangmei; Shi, Susheng; Polydorides, Alexandros D; Zhao, Xinming; Lu, Ning; Lin, Dongmei

    2013-01-01

    Sporadic gastrointestinal stromal tumors (GISTs) usually form a well-circumscribed mass. In contrast, diffuse interstitial cell of Cajal (ICC) hyperplasia along the Auerbach plexus without a discrete mass may occur in patients with germline mutations in the NF1, c-KIT or PDGFRA genes. However, sporadic, diffuse ICC hyperplasia without c-KIT or PDGFRA mutations has not been reported. We describe herein one such case, forming a giant diverticulum. A 63-year-old woman with no features of Neurofibromatosis 1 (NF1) presented with increasing abdominal pain for more than 30 years. A large, diverticulum-like mass in the ileum was resected. Microscopically, a diffuse proliferation of bland spindle cells was seen extending for 12 cm, replacing the muscularis propria and lined by intact mucosa. The spindle cells were CD117+/CD34+/DOG1+/SMA+/Desmin-/S100-. Mutation analyses did not reveal any mutations in c-KIT or PDGFRA. The lesion had two silent mutations in the NF1 gene. It is rare of the diffuse form of sporadic ICC hyperplasia showing diffuse longitudinal microscopic growth completely replacing the muscularis propria, mimicking diffuse ICC hyperplasia in hereditary GIST syndromes, but without solid components and no c-KIT or PDGFRA gene mutations. This peculiar form of sporadic ICC hyperplasia may be related to intestinal dysmotility in this ileal segment and giant diverticulum formation. PMID:24294389

  10. Effect of single-walled carbon nanotubes on tumor cells viability and formation of multicellular tumor spheroids

    Science.gov (United States)

    Yakymchuk, Olena M.; Perepelytsina, Olena M.; Dobrydnev, Alexey V.; Sydorenko, Mychailo V.

    2015-03-01

    This paper describes the impact of different concentrations of single-walled carbon nanotubes (SWCNTs) on cell viability of breast adenocarcinoma, MCF-7 line, and formation of multicellular tumor spheroids (MTS). Chemical composition and purity of nanotubes is controlled by Fourier transform infrared spectroscopy. The strength and direction of the influence of SWCNTs on the tumor cell population was assessed by cell counting and measurement of the volume of multicellular tumor spheroids. Effect of SWCNTs on the formation of multicellular spheroids was compared with the results obtained by culturing tumor cells with ultra dispersed diamonds (UDDs). Our results demonstrated that SWCNTs at concentrations ranging from 12.5 to 50 μg/ml did not have cytotoxic influence on tumor cells; instead, they had weak cytostatic effect. The increasing of SWCNTs concentration to 100 to 200 μg/ml stimulated proliferation of tumor cells, especially in suspension fractions. The result of this influence was in formation of more MTS in cell culture with SWCNTs compared with UDDs and control samples. In result, the median volume of MTS after cultivation with SWCNTs at 100 to 200 μg/ml concentrations is 3 to 5 times greater than that in samples which were incubated with the UDDs and is 2.5 times greater than that in control cultures. So, if SWCNTs reduced cell adhesion to substrate and stimulated formation of tumor cell aggregates volume near 7 · 10-3 mm3, at the same time, UDDs reduced adhesion and cohesive ability of cells and stimulated generation of cell spheroids volume no more than 4 · 10-3 mm3. Our results could be useful for the control of cell growth in three-dimensional culture.

  11. SWAP-70 controls formation of the splenic marginal zone through regulating T1B-cell differentiation.

    Science.gov (United States)

    Chopin, Michaël; Quemeneur, Laurence; Ripich, Tatsiana; Jessberger, Rolf

    2010-12-01

    T1 and T2 transitional B cells are precursors for marginal zone B cells (MZB), which surround splenic follicles. MZB are essential for marginal zone formation, are central to the innate immune response, and contribute to adaptive immunity. Differentiation, migration, and homing of MZB and their precursors remain to be fully understood. We show that SWAP-70, a RhoGTPase-interacting and F-actin-binding protein with functions in cell polarization, migration, and adhesion regulates MZB development and marginal zone formation. The percentage of MZB in spleen of Swap70(-/-) mice was reduced to about one-third of that found in WT mice. Swap70(-/-) T1 cells accumulated in integrin ligand(high) regions of the splenic red pulp and failed to efficiently develop into T2 cells. Adoptive transfer and mixed BM chimera experiments demonstrated this to be a B-cell intrinsic phenotype. T-cell-independent antibody production was not impaired, however, and thus suggests that this process does not require correct homing of MZB precursors. B-cell adhesion through α(L)β(2) and α(4)β(1) integrins was hyper-activated in vitro and on tissue sections, and S1P-stimulated chemokinesis of MZB was reduced in the absence of SWAP-70. Thus, SWAP-70 acts as a regulator of the adhesion process, particularly important for differentiation control of B-cell precursors and their contribution to splenic tissue formation.

  12. Berberine promotes the development of atherosclerosis and foam cell formation by inducing scavenger receptor A expression in macrophage

    Institute of Scientific and Technical Information of China (English)

    Ke Li; Wenqi Yao; Xiudan Zheng; Kan Liao

    2009-01-01

    Berberine is identified to lower the serum cholesterol level in human and hamster through the induction of low density lipoproteins (LDL) receptor in hepatic cells. To evaluate its potential in preventing atherosclerosis, the effect of berberine on atherosclerosis development in apolipoprotein E-deficient (apoE-/-) mice was investigated, in apoE-/-mice, berberine induced in vivo foam cell formation and promoted atherosclerosis development. The foam cell for-mation induced by berberine was also observed in mouse RAW264.7 cells, as well as in mouse and human primary macrophages. By inducing scavenger receptor A (SR-A) expression in macrophages, berberine increased the uptake of modified LDL (DiO-Ac-LDL). Berberine-induced SR-A expression was also observed in macrophage foam cells in vivo and in the cells at atherosclerotic lesion. Analysis in RAW264.7 cells indicated that berberine induced SR-A ex-pression by suppressing PTEN expression, which led to sustained Akt activation. Our results suggest that to evaluate the potential of a cholesterol-reducing compound in alleviating atherosclerosis, its effect on the cells involved in ath-erosclerosis development, such as macrophages, should also be considered. Promotion of foam cell formation could counter-balance the beneficial effect of lowering serum cholesterol.

  13. Nuclear translocation of phosphorylated STAT3 regulates VEGF-A-induced lymphatic endothelial cell migration and tube formation

    Energy Technology Data Exchange (ETDEWEB)

    Okazaki, Hideki; Tokumaru, Sho; Hanakawa, Yasushi; Shiraishi, Ken; Shirakata, Yuji; Dai, Xiuju; Yang, Lijun; Tohyama, Mikiko; Hashimoto, Koji [Department of Dermatology, Ehime University Graduate School of Medicine, Toon, Ehime 791-0295 (Japan); Sayama, Koji, E-mail: sayama@m.ehime-u.ac.jp [Department of Dermatology, Ehime University Graduate School of Medicine, Toon, Ehime 791-0295 (Japan)

    2011-09-02

    Highlights: {yields} VEGF-A enhanced lymphatic endothelial cell migration and increased tube formation. {yields} VEGF-A treated lymphatic endothelial cell showed activation of STAT3. {yields} Dominant-negative STAT3 inhibited VEGF-A-induced lymphatic endothelial cell migration and tube formation. -- Abstract: Vascular endothelial growth factor (VEGF) is an endothelial cell-specific growth factor that regulates endothelial functions, and signal transducers and activators of transcription (STATs) are known to be important during VEGF receptor signaling. The aim of this study was to determine whether STAT3 regulates VEGF-induced lymphatic endothelial cell (LEC) migration and tube formation. VEGF-A (33 ng/ml) enhanced LEC migration by 2-fold and increased tube length by 25% compared with the control, as analyzed using a Boyden chamber and Matrigel assay, respectively. Western blot analysis and immunostaining revealed that VEGF-A induced the nuclear translocation of phosphorylated STAT3 in LECs, and this translocation was blocked by the transfection of LECs with an adenovirus vector expressing a dominant-negative mutant of STAT3 (Ax-STAT3F). Transfection with Ax-STAT3F also almost completely inhibited VEGF-A-induced LEC migration and tube formation. These results indicate that STAT3 is essential for VEGF-A-induced LEC migration and tube formation and that STAT3 regulates LEC functions.

  14. Influence of Ligands on the Formation of Kesterite Thin Films for Solar Cells: A Comparative Study.

    Science.gov (United States)

    Huang, Tang Jiao; Yin, Xuesong; Tang, Chunhua; Qi, Guojun; Gong, Hao

    2016-05-10

    The preparation of solar-cell-grade Cu2 ZnSnS4 (CZTS) thin films from ligand-capped small-grained CZTS particles remains hindered by problems of phase segregation, composition non-uniformity, and in particular carbon-layer formation. Herein, through a systematic comparative study of annealed films of CZTS nanocrystals prepared using conventional oleylamine and those prepared using formamide, these problems are found to be mainly attributable to the influence of the ligands, and mechanisms are proposed. Importantly, the origin of the carbon layer in oleylamine-capped CZTS films is revealed to be the reaction between oleylamine and sulfur. This carbon layer has a very poor electrical conductivity, which can be the reason for the limited performance of such films. Fortunately, these problems can almost all be avoided by replacing oleylamine with formamide to form CZTS films. PMID:27059551

  15. A new approach for cell formation and scheduling with assembly operations and product structure

    Directory of Open Access Journals (Sweden)

    Mir Bahador Aryanezhad

    2011-01-01

    Full Text Available In this paper, a new formulation model for cellular manufacturing system (CMS design problem is proposed. The proposed model of this paper considers assembly operations and product structure so that it includes the scheduling problem with the formation of manufacturing cells, simultaneously. Since the proposed model is nonlinear, a linearization method is applied to gain optimal solution when the model is solved using direct implementation of mixed integer programming. A new genetic algorithm (GA is also proposed to solve the resulted model for large-scale problems. We examine the performance of the proposed method using the direct implementation and the proposed GA method. The results indicate that the proposed GA approach could provide efficient assembly and product structure for real-world size problems.

  16. Effect of temperature on carrier formation efficiency in organic photovoltaic cells

    Energy Technology Data Exchange (ETDEWEB)

    Moritomo, Yutaka, E-mail: moritomo.yutaka.gf@u.tsukuba.ac.jp; Yonezawa, Kouhei [Graduate School of Pure and Applied Science, University of Tsukuba, Tsukuba 305-8571 (Japan); Yasuda, Takeshi [Photovoltaic Materials Unit, National Institute for Materials Science (NIMS), Tsukuba 305-0047 (Japan)

    2014-08-18

    The internal quantum efficiency (ϕ{sub IQ}) of an organic photovoltaic cell is governed by plural processes. Here, we propose that ϕ{sub IQ} can be experimentally decomposed into carrier formation (ϕ{sub CF}) and carrier transfer (ϕ{sub CT}) efficiencies. By combining femtosecond time-resolved and electrochemical spectroscopy, we clarified the effect of temperature on ϕ{sub CF} in a regioregular poly(3-hexylthiophene) (rr-P3HT)/[6,6]-phenyl C{sub 61}-butyric acid methyl ester blend film. We found that ϕ{sub CF} (=0.55) at 80 K is the same as that (=0.55) at 300 K. The temperature insensitivity of ϕ{sub CF} indicates that the electron-hole pairs at the D/A interface are seldom subjected to coulombic binding energy.

  17. Differential regulation of macropinocytosis in macrophages by cytokines: implications for foam cell formation and atherosclerosis.

    Science.gov (United States)

    Michael, Daryn R; Ashlin, Tim G; Davies, Charlotte S; Gallagher, Hayley; Stoneman, Thomas W; Buckley, Melanie L; Ramji, Dipak P

    2013-10-01

    A key event during the formation of lipid-rich foam cells during the progression of atherosclerosis is the uptake of modified low-density lipoproteins (LDL) by macrophages in response to atherogenic mediators in the arterial intima. In addition to scavenger receptor-dependent uptake of LDL, macropinocytosis is known to facilitate the uptake of LDL through the constitutive and passive internalization of large quantities of extracellular solute. In this study we confirm the ability of macropinocytosis to facilitate the uptake of modified LDL by human macrophages and show its modulation by TGF-β, IFN-γ, IL-17A and IL-33. Furthermore we show that the TGF-β-mediated inhibition of macropinocytosis is a Smad-2/-3-independent process.

  18. Modeling the PbI2 formation in perovskite solar cells using XRD/XPS patterns

    Science.gov (United States)

    Sohrabpoor, Hamed; Elyasi, Majid; Aldosari, Marouf; Gorji, Nima E.

    2016-09-01

    The impact of prolonged irradiation and air humidity on the stability of perovskite solar cells is modeled using X-ray diffraction and X-ray photoelectron spectroscopy patterns reported in the literature. Light or air-moisture causes the formation of a thin PbI2 or oxide defective layers (in nanoscale) at the interface of perovskite/hole-transport-layer or at the junction with metallic back contact. This thin layer blocks the carrier transport/passivation at the interfaces and cause degradation of device parameters. Variation in thickness of defective layers, changes the XRD and XPS peaks. This allows detection and estimation of the type, crystallinity and thickness of the defective layer. A simple model is developed here to extract the thickness of such thin defective layers formed in nanometer scale at the back region of several perovskite devices. Based on this information, corrected energy band diagram of every device before and after degradation/aging is drawn and discussed in order to obtain insight into the carrier transport and charge collection at the barrier region. In addition, graphene contacted perovskite devices are investigated showing that honey-comb network of graphene contact reduces the effect of aging leading to formation of a thinner defective layer at the perovskite surface compared to perovskite devices with conventional inorganic contacts i.e. Au, Al.

  19. Identification of red blood cell rouleaux formation using photoacoustic ultrasound spectroscopy

    Science.gov (United States)

    Kibria, Fayruz; Hysi, Eno; Strohm, Eric M.; Kolios, Michael C.

    2014-03-01

    Red blood cell (RBC) rouleaux formation is a reversible phenomenon that occurs during low blood flow and small shearing forces in circulation. Certain pathological conditions can alter the molecular constituents of blood and properties of the RBCs leading to enhanced rouleaux formation, which results in impaired perfusion and tissue oxygenation. In this study rouleaux were artificially generated using Dextran-70 and examined using a photoacoustic (PA) microscope. Individual rouleau were irradiated with a 532 nm pulsed laser focused to a 10 μm spot size, and the resulting PA signals recorded with a 200 MHz transducer. The laser and transducer were co-aligned, with the sample positioned between them. The frequency-domain PA ultrasound spectra were calculated for rouleaux with lengths ranging from 10 to 20 μm. For the rouleaux, a single spectral minimum at 269+/-4 MHz was observed. The spectral minima were in good agreement with a theoretical thermoelastic expansion model using an infinite length cylindrical absorber, bearing a diameter equivalent to an average human RBC (7.8 μm). These results suggest that PA ultrasound spectroscopy can be potentially used as a tool for monitoring blood samples for the presence of rouleaux.

  20. Simulation of the Formation of DNA Double Strand Breaks and Chromosome Aberrations in Irradiated Cells

    Science.gov (United States)

    Plante, Ianik; Ponomarev, Artem L.; Wu, Honglu; Blattnig, Steve; George, Kerry

    2014-01-01

    The formation of DNA double-strand breaks (DSBs) and chromosome aberrations is an important consequence of ionizing radiation. To simulate DNA double-strand breaks and the formation of chromosome aberrations, we have recently merged the codes RITRACKS (Relativistic Ion Tracks) and NASARTI (NASA Radiation Track Image). The program RITRACKS is a stochastic code developed to simulate detailed event-by-event radiation track structure: [1] This code is used to calculate the dose in voxels of 20 nm, in a volume containing simulated chromosomes, [2] The number of tracks in the volume is calculated for each simulation by sampling a Poisson distribution, with the distribution parameter obtained from the irradiation dose, ion type and energy. The program NASARTI generates the chromosomes present in a cell nucleus by random walks of 20 nm, corresponding to the size of the dose voxels, [3] The generated chromosomes are located within domains which may intertwine, and [4] Each segment of the random walks corresponds to approx. 2,000 DNA base pairs. NASARTI uses pre-calculated dose at each voxel to calculate the probability of DNA damage at each random walk segment. Using the location of double-strand breaks, possible rejoining between damaged segments is evaluated. This yields various types of chromosomes aberrations, including deletions, inversions, exchanges, etc. By performing the calculations using various types of radiations, it will be possible to obtain relative biological effectiveness (RBE) values for several types of chromosome aberrations.

  1. Cell differentiation and tissue formation in the unique fruits of devil's claws (Martyniaceae).

    Science.gov (United States)

    Horbens, Melanie; Gao, Jie; Neinhuis, Christoph

    2014-06-01

    • Premise of the study: Martyniaceae are characterized by capsules with two upwardly curved, horn-shaped extensions representing morphologically specialized epizoochorous fruits. Because the capsules are assumed to cling to hooves and ankles of large mammals, fiber arrangement and tissue combinations within the endocarp ensuring proper attachment to the vector's feet during transport are of particular interest. In this first detailed anatomical investigation, the functional adaptation of the fruits and their implications for the specific dispersal mode are provided. The peculiar fiber arrangement may also be of interest for future biomimetic composite materials.• Methods: Endocarp anatomy and details of tissue differentiation were examined in fruits of Ibicella lutea and Proboscidea louisianica subsp. fragrans combining light microscopy, SEM, and x-ray microtomography analysis.• Key results: While tips of the extensions are predominantly reinforced by longitudinally oriented fibers, in the middle segment these fibers are densely packed in individual bundles entwined and separated by transversely elongated cells. Within the capsule wall, the fiber bundles are embedded in a dense mesh of transversely oriented fibers that circularly reinforce and protect the loculus. This fibrous pericarp tissue develops within few days by localized cell divisions and intrusive growth of primarily isodiametric parenchyma cells in the pistil.• Conclusions: The study allows insight into a unique and complex example of functionally driven cell growth and tissue formation. Long-horned fruits of Martyniaceae obviously are highly specialized to epizoochorous dispersal, pointing to primary vector-related seed dispersal. The highly ordered arrangement of fibers results in a great mechanical firmness.

  2. Anti-atherosclerotic potential of gossypetin via inhibiting LDL oxidation and foam cell formation

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Jing-Hsien [School of Nutrition, Chung Shan Medical University, Taichung, Taiwan (China); Department of Medical Research, Chung Shan Medical University Hospital, Taichung, Taiwan (China); Tsai, Chia-Wen [Department of Nutrition, China Medical University, Taichung, Taiwan (China); Wang, Chi-Ping [Department of Clinical Laboratory, Chung Shan Medical University Hospital, Taichung, Taiwan (China); Lin, Hui-Hsuan, E-mail: linhh@csmu.edu.tw [Department of Medical Research, Chung Shan Medical University Hospital, Taichung, Taiwan (China); School of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung, Taiwan (China)

    2013-10-15

    Gossypetin, a flavone originally isolated from Hibiscus species, has been shown to possess antioxidant, antimicrobial, and antimutagenic activities. Here, we investigated the mechanism(s) underlying the anti-atherosclerotic potential of gossypetin. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) scavenging activity assay showed that the addition of > 50 μM of gossypetin could scavenge over 50% of DPPH radicals. The inhibitory effects of gossypetin on the lipid and protein oxidation of LDL were defined by thiobarbituric acid reactive substance (TBARS) assay, the relative electrophoretic mobility (REM) of oxidized LDL (ox-LDL), and fragmentation of apoB in the Cu{sup 2+}-induced oxidation of LDL. Gossypetin showed potential in reducing ox-LDL-induced foam cell formation and intracellular lipid accumulation, and uptake ability of macrophages under non-cytotoxic concentrations. Molecular data showed that these influences of gossypetin might be mediated via peroxisome proliferator-activated receptor α (PPARα)/liver-X receptor α (LXRα)/ATP-binding cassette transporter A1 (ABCA1) and PPARγ/scavenger receptor CD36 pathways, as demonstrated by the transfection of PPARα siRNA or PPARγ expression vector. Our data implied that gossypetin regulated the PPAR signals, which in turn led to stimulation of cholesterol removal from macrophages and delay atherosclerosis. These results suggested that gossypetin potentially could be developed as an anti-atherosclerotic agent. - Highlights: • The anti-atherosclerotic effect of gossypetin in vitro was examined. • Gossypetin inhibited LDL oxidation. • Gossypetin showed potential in reducing on the formation of foam cells. • Gossypetin functions against ox-LDL through PPARa activation and PPARγ depression.

  3. Piperlongumine inhibits atherosclerotic plaque formation and vascular smooth muscle cell proliferation by suppressing PDGF receptor signaling

    Energy Technology Data Exchange (ETDEWEB)

    Son, Dong Ju [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Kim, Soo Yeon [Division of Life Science, Korea Basic Science Institute, Daejeon (Korea, Republic of); Han, Seong Su [University of Iowa Carver College of Medicine, Department of Pathology, Iowa City, IA (United States); Kim, Chan Woo [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Kumar, Sandeep [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Park, Byeoung Soo [Nanotoxtech Co., Ansan (Korea, Republic of); Lee, Sung Eun [Division of Applied Biology and Chemistry, Kyungpook National University, Daegu (Korea, Republic of); Yun, Yeo Pyo [College of Pharmacy, Chungbuk National University, Cheongju (Korea, Republic of); Jo, Hanjoong, E-mail: hjo@emory.edu [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Park, Young Hyun, E-mail: pyh012@sch.ac.kr [Department of Food Science and Nutrition, College of Natural Sciences, Soonchunhyang University, Asan (Korea, Republic of)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Anti-atherogenic effect of PL was examined using partial carotid ligation model in ApoE KO mice. Black-Right-Pointing-Pointer PL prevented atherosclerotic plaque development, VSMCs proliferation, and NF-{kappa}B activation. Black-Right-Pointing-Pointer Piperlongumine reduced vascular smooth muscle cell activation through PDGF-R{beta} and NF-{kappa}B-signaling. Black-Right-Pointing-Pointer PL may serve as a new therapeutic molecule for atherosclerosis treatment. -- Abstract: Piperlongumine (piplartine, PL) is an alkaloid found in the long pepper (Piper longum L.) and has well-documented anti-platelet aggregation, anti-inflammatory, and anti-cancer properties; however, the role of PL in prevention of atherosclerosis is unknown. We evaluated the anti-atherosclerotic potential of PL in an in vivo murine model of accelerated atherosclerosis and defined its mechanism of action in aortic vascular smooth muscle cells (VSMCs) in vitro. Local treatment with PL significantly reduced atherosclerotic plaque formation as well as proliferation and nuclear factor-kappa B (NF-{kappa}B) activation in an in vivo setting. PL treatment in VSMCs in vitro showed inhibition of migration and platelet-derived growth factor BB (PDGF-BB)-induced proliferation to the in vivo findings. We further identified that PL inhibited PDGF-BB-induced PDGF receptor beta activation and suppressed downstream signaling molecules such as phospholipase C{gamma}1, extracellular signal-regulated kinases 1 and 2 and Akt. Lastly, PL significantly attenuated activation of NF-{kappa}B-a downstream transcriptional regulator in PDGF receptor signaling, in response to PDGF-BB stimulation. In conclusion, our findings demonstrate a novel, therapeutic mechanism by which PL suppresses atherosclerosis plaque formation in vivo.

  4. Anti-atherosclerotic potential of gossypetin via inhibiting LDL oxidation and foam cell formation

    International Nuclear Information System (INIS)

    Gossypetin, a flavone originally isolated from Hibiscus species, has been shown to possess antioxidant, antimicrobial, and antimutagenic activities. Here, we investigated the mechanism(s) underlying the anti-atherosclerotic potential of gossypetin. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) scavenging activity assay showed that the addition of > 50 μM of gossypetin could scavenge over 50% of DPPH radicals. The inhibitory effects of gossypetin on the lipid and protein oxidation of LDL were defined by thiobarbituric acid reactive substance (TBARS) assay, the relative electrophoretic mobility (REM) of oxidized LDL (ox-LDL), and fragmentation of apoB in the Cu2+-induced oxidation of LDL. Gossypetin showed potential in reducing ox-LDL-induced foam cell formation and intracellular lipid accumulation, and uptake ability of macrophages under non-cytotoxic concentrations. Molecular data showed that these influences of gossypetin might be mediated via peroxisome proliferator-activated receptor α (PPARα)/liver-X receptor α (LXRα)/ATP-binding cassette transporter A1 (ABCA1) and PPARγ/scavenger receptor CD36 pathways, as demonstrated by the transfection of PPARα siRNA or PPARγ expression vector. Our data implied that gossypetin regulated the PPAR signals, which in turn led to stimulation of cholesterol removal from macrophages and delay atherosclerosis. These results suggested that gossypetin potentially could be developed as an anti-atherosclerotic agent. - Highlights: • The anti-atherosclerotic effect of gossypetin in vitro was examined. • Gossypetin inhibited LDL oxidation. • Gossypetin showed potential in reducing on the formation of foam cells. • Gossypetin functions against ox-LDL through PPARa activation and PPARγ depression

  5. In vitro formation of the anthranoid scaffold by cell-free extracts from yeast-extract-treated Cassia bicapsularis cell cultures.

    Science.gov (United States)

    Abdel-Rahman, Iman A M; Beuerle, Till; Ernst, Ludger; Abdel-Baky, Afaf M; Desoky, Ezz El-Din K; Ahmed, Amany S; Beerhues, Ludger

    2013-04-01

    The anthranoid skeleton is believed to be formed by octaketide synthase (OKS), a member of the type III polyketide synthase (PKS) superfamily. Recombinant OKSs catalyze stepwise condensation of eight acetyl units to form a linear octaketide intermediate which, however, is incorrectly folded and cyclized to give the shunt products SEK4 and SEK4b. Here we report in vitro formation of the anthranoid scaffold by cell-free extracts from yeast-extract-treated Cassia bicapsularis cell cultures. Unlike field- and in vitro-grown shoots which accumulate anthraquinones, cell cultures mainly contained tetrahydroanthracenes, formation of which was increased 2.5-fold by the addition of yeast extract. The elicitor-stimulated accumulation of tetrahydroanthracenes was preceded by an approx. 35-fold increase in OKS activity. Incubation of cell-free extracts from yeast-extract-treated cell cultures with acetyl-CoA and [2-(14)C]malonyl-CoA led to formation of torosachrysone (tetrahydroanthracene) and emodin anthrone, beside two yet unidentified products. No product formation occurred in the absence of acetyl-CoA as starter substrate. To confirm the identities of the enzymatic products, cell-free extracts were incubated with acetyl-CoA and [U-(13)C(3)]malonyl-CoA and (13)C incorporation was analyzed by ESI-MS/MS. Detection of anthranoid biosynthesis in cell-free extracts indicates in vitro cooperation of OKS with a yet unidentified factor or enzyme for octaketide cyclization.

  6. Reactive oxygen and nitrogen (ROS and RNS) species generation and cell death in tomato suspension cultures--Botrytis cinerea interaction.

    Science.gov (United States)

    Pietrowska, E; Różalska, S; Kaźmierczak, A; Nawrocka, J; Małolepsza, U

    2015-01-01

    This article reports events connected to cell survival and Botrytis cinerea infection development in cell suspension cultures of two tomato cultivars which show different levels of susceptibility to the pathogen: cv. Corindo (more susceptible) and cv. Perkoz (less susceptible). In parallel changes in reactive oxygen (ROS) and nitrogen (RNS) species generation and in S-nitrosoglutathione reductase (GSNOR) activity were studied. In vivo staining methods with acridine orange (AO) and ethidium bromide (EB) as well as fluorescent microscopy were used to assess tomato and B. cinerea cells death. The biochemical studies of ROS and RNS concentrations in plant cell extract were complemented by in vivo ROS and nitric oxide (NO) imaging using nitro blue tetrazolium (NBT), diaminobenzidine (DAB) and diaminofluorescein diacetate (DAF-DA) staining methods, and confocal microscope technique. B. cinerea infection proceeded slower in Perkoz cell cultures. It was evidenced by measuring the pathogen conidia germination and germination tube development in which nuclei revealing cell death dominated. Two different types of tomato cell death were observed: cells with necrotic nuclei dominated in Corindo whereas in Perkoz cells with characteristic of vacuolar death type prevailed. In Perkoz cells, constitutive levels of NO and S-nitrosothiols (SNO) were significantly higher and hydrogen peroxide (H₂O₂) and superoxide anion (O₂(-)) concentrations were slightly higher as compared with Corindo cells. Moreover, increases in these molecule concentrations as a result of B. cinerea inoculation were observed in both, Perkoz and Corindo cell cultures. The enzymatic GSNOR activity seems to be an important player in controlling the SNO level in tomato cells. Involvements of the studied compounds in molecular mechanisms of tomato resistance to B. cinerea are discussed in the paper. PMID:25064634

  7. Comparison of permanganate preoxidation and preozonation on algae containing water: cell integrity, characteristics, and chlorinated disinfection byproduct formation.

    Science.gov (United States)

    Xie, Pengchao; Ma, Jun; Fang, Jingyun; Guan, Yinghong; Yue, Siyang; Li, Xuchun; Chen, Liwei

    2013-12-17

    Aqueous suspensions of Microcystis aeruginosa were preoxidized with either ozone or permanganate and then subjected to chlorination under conditions simulating drinking water purification. The impacts of the two oxidants on the algal cells and on the subsequent production of dissolved organic matter and disinfection byproducts were investigated. Preozonation dramatically increased disinfection byproduct formation during chlorination, especially the formation of haloaldehydes, haloacetonitriles, and halonitromethanes. Preoxidation with permanganate had much less effect on disinfection byproduct formation. Preozonation destroyed algal cell walls and cell membranes to release intracellular organic matter (IOM), and less than 2.0% integrated cells were left after preozonation with the dosage as low as 0.4 mg/L. Preoxidation with permanganate mainly released organic matter adsorbed on the cells' surface without causing any damage to the cells' integrity, so the increase in byproduct formation was much less. More organic nitrogen and lower molecular weight precursors were produced in a dissolved phase after preozonation than permanganate preoxidation, which contributes to the significant increase of disinfection byproducts after preozonation. The results suggest that permanganate is a better choice than ozone for controlling algae derived pollutants and disinfection byproducts.

  8. Phytoplankton aggregate formation: observations of patterns and mechanisms of cell sticking and the significance of exopolymeric material

    DEFF Research Database (Denmark)

    Kiørboe, Thomas; Hansen, Jorgen L. S.

    1993-01-01

    the potential for aggregate formation. In the laboratory, we examined variation in stickiness in five species of diatoms and two species of flagellates grown in batch cultures. We also investigated the production of particulate mucus by phytoplankton cells and its role in aggregate formation, and we studied...... cause other, non-sticky particles to coagulate. This emphasizes the potential importance of diatom-derived particulate mucus for particle flocculation in the ocean....

  9. Endothelial Progenitor Cells Promote Directional Three-Dimensional Endothelial Network Formation by Secreting Vascular Endothelial Growth Factor

    OpenAIRE

    Yoshinori Abe; Yoshiyuki Ozaki; Junichi Kasuya; Kimiko Yamamoto; Joji Ando; Ryo Sudo; Mariko Ikeda; Kazuo Tanishita

    2013-01-01

    Endothelial progenitor cell (EPC) transplantation induces the formation of new blood-vessel networks to supply nutrients and oxygen, and is feasible for the treatment of ischemia and cardiovascular diseases. However, the role of EPCs as a source of proangiogenic cytokines and consequent generators of an extracellular growth factor microenvironment in three-dimensional (3D) microvessel formation is not fully understood. We focused on the contribution of EPCs as a source of proangiogenic cytoki...

  10. Biocatalytic Formation of Gold Nanoparticles Decorated with Functional Proteins inside Recombinant Escherichia coli Cells.

    Science.gov (United States)

    Hosomomi, Yukiho; Niide, Teppei; Wakabayashi, Rie; Goto, Masahiro; Kamiya, Noriho

    2016-01-01

    A novel strategy for the preparation of protein-decorated gold nanoparticles (Au NPs) was developed inside Escherichia coli cells, where an artificial oxidoreductase, composed of antibody-binding protein (pG), Bacillus stearothermophilus glycerol dehydrogenase (BsGLD) and a peptide tag with gold-binding affinity (H6C), was overexpressed in the cytoplasm. In situ formation of Au NPs was promoted by a natural electron-donating cofactor, nicotinamide adenine dinucleotide (NAD), which was regenerated to the reduced form of NADH by the catalytic activity of the fusion protein (pG-BsGLD-H6C) overexpressed in the cytoplasm of E. coli, with the concomitant addition of exogenous glycerol to the reaction system. The fusion protein was self-immobilized on Au NPs inside the E. coli cells, which was confirmed by SDS-PAGE and western blotting analyses of the resultant Au NPs. Finally, the IgG binding ability of the pG moiety displayed on Au NPs was evaluated by an enzyme-linked immunosorbent assay. PMID:26960608

  11. Insulin and glucose play a role in foam cell formation and function

    Directory of Open Access Journals (Sweden)

    Keller Susanna R

    2006-06-01

    Full Text Available Abstract Background Foam cell formation in diabetic patients often occurs in the presence of high insulin and glucose levels. To test whether hyperinsulinemic hyperglycemic conditions affect foam cell differentiation, we examined gene expression, cytokine production, and Akt phosphorylation in human monocyte-derived macrophages incubated with two types of oxidized low density lipoprotein (LDL, minimally modified LDL (mmLDL and extensively oxidized LDL (OxLDL. Methods and results Using Affymetrix GeneChip® arrays, we found that several genes directly related to insulin signaling were changed. The insulin receptor and glucose-6-phosphate dehydrogenase were upregulated by mmLDL and OxLDL, whereas insulin-induced gene 1 was significantly down-regulated. In hyperinsulinemic hyperglycemic conditions, modified LDL upregulated Akt phosphorylation and expression of the insulin-regulated aminopeptidase. The level of proinflammatory cytokines, IL-lβ, IL-12, and IL-6, and of a 5-lipoxygenase eicosanoid, 5-hydroxyeicosatetraenoic acid (5-HETE, was also increased. Conclusion These results suggest that the exposure of macrophages to modified low density lipoproteins in hyperglycemic hyperinsulinemic conditions affects insulin signaling and promotes the release of proinflammatory stimuli, such as cytokines and eicosanoids. These in turn may contribute to the development of insulin resistance.

  12. Cell-mediated BMP-2 liberation promotes bone formation in a mechanically unstable implant environment.

    Science.gov (United States)

    Hägi, Tobias T; Wu, Gang; Liu, Yuelian; Hunziker, Ernst B

    2010-05-01

    The flexible alloplastic materials that are used in bone-reconstruction surgery lack the mechanical stability that is necessary for sustained bone formation, even if this process is promoted by the application of an osteogenic agent, such as BMP-2. We hypothesize that if BMP-2 is delivered gradually, in a cell-mediated manner, to the surgical site, then the scaffolding material's lack of mechanical stability becomes a matter of indifference. Flexible discs of Ethisorb were functionalized with BMP-2, which was either adsorbed directly onto the material (rapid release kinetics) or incorporated into a calcium-phosphate coating (slow release kinetics). Unstabilized and titanium-plate-stabilized samples were implanted subcutaneously in rats and retrieved up to 14 days later for a histomorphometric analysis of bone and cartilage volumes. On day 14, the bone volume associated with titanium-plate-stabilized discs bearing an adsorbed depot of BMP-2 was 10-fold higher than that associated with their mechanically unstabilized counterparts. The bone volume associated with discs bearing a coating-incorporated depot of BMP-2 was similar in the mechanically unstabilized and titanium-plate-stabilized groups, and comparable to that associated with the titanium-plate-stabilized discs bearing an adsorbed depot of BMP-2. Hence, if an osteogenic agent is delivered in a cell-mediated manner (via coating degradation), ossification can be promoted even within a mechanically unstable environment.

  13. Autoradiographic study on the incorporation of carbon-14 labeled formate and adenine into nucleic acid in blood-forming cells

    International Nuclear Information System (INIS)

    The incorporation of [14C]formate and [8-14C]adenine into nucleic acid in blood-forming cells was studied by the autoradiographic technique. The isotopic markers were injected subcutaneously into young rats weighting from 100 to 150 g three times every 24 hours and the animals were examined 3 hours after the last injection. In the case of [14C]formate injection, erythroblasts exhibited extremely strong labeling in contrast to weaker labeling of other blood-forming cells. In the case of [14C]adenine administration, on the other hand, immature cells of the granuclocytic series as well as immature reticulum cells (proliferating cells of reticular tissue) were much more heavily labeled than were other blood-forming cells, particularly the erythroblasts which revealed weak or no labeling. By digestion or extraction of DNA, RNA or both from cells with DNase, RNase or hot 10% perchloric acid treatment, respectively, it was confirmed that the observed heavy labeling of any type of cells with either [14C]formate or [14C]adenine was due chiefly to incorporation of the radioactive materials into nuclear DNA. The present results are discussed together with the findings of earlier studies on lymphoid cells which indicate that, in certain cell types, the patterns of [3H]deoxycytidine labeling differ considerably from the corresponding patterns of [3H]deoxycytidine labeling. The present and earlier findings provide evidence to substantiate that, among blood-forming cells, there are considerable variations in the labeling patterns of nuclear DNA depending on differences in the radioactive DNA precursors used as well as in the cell types. (author)

  14. Elevated AKR1C3 expression promotes prostate cancer cell survival and prostate cell-mediated endothelial cell tube formation: implications for prostate cancer progressioan

    International Nuclear Information System (INIS)

    Aldo-keto reductase (AKR) 1C family member 3 (AKR1C3), one of four identified human AKR1C enzymes, catalyzes steroid, prostaglandin, and xenobiotic metabolism. In the prostate, AKR1C3 is up-regulated in localized and advanced prostate adenocarcinoma, and is associated with prostate cancer (PCa) aggressiveness. Here we propose a novel pathological function of AKR1C3 in tumor angiogenesis and its potential role in promoting PCa progression. To recapitulate elevated AKR1C3 expression in cancerous prostate, the human PCa PC-3 cell line was stably transfected with an AKR1C3 expression construct to establish PC3-AKR1C3 transfectants. Microarray and bioinformatics analysis were performed to identify AKR1C3-mediated pathways of activation and their potential biological consequences in PC-3 cells. Western blot analysis, reverse transcription-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and an in vitro Matrigel angiogenesis assays were applied to validate the pro-angiogenic activity of PC3-AKR1C3 transfectants identified by bioinformatics analysis. Microarray and bioinformatics analysis suggested that overexpression of AKR1C3 in PC-3 cells modulates estrogen and androgen metabolism, activates insulin-like growth factor (IGF)-1 and Akt signaling pathways, as well as promotes tumor angiogenesis and aggressiveness. Levels of IGF-1 receptor (IGF-1R) and Akt activation as well as vascular endothelial growth factor (VEGF) expression and secretion were significantly elevated in PC3-AKR1C3 transfectants in comparison to PC3-mock transfectants. PC3-AKR1C3 transfectants also promoted endothelial cell (EC) tube formation on Matrigel as compared to the AKR1C3-negative parental PC-3 cells and PC3-mock transfectants. Pre-treatment of PC3-AKR1C3 transfectants with a selective IGF-1R kinase inhibitor (AG1024) or a non-selective phosphoinositide 3-kinases (PI3K) inhibitor (LY294002) abolished ability of the cells to promote EC tube formation. Bioinformatics

  15. Elevated AKR1C3 expression promotes prostate cancer cell survival and prostate cell-mediated endothelial cell tube formation: implications for prostate cancer progressioan

    Directory of Open Access Journals (Sweden)

    Davis Jeffrey S

    2010-12-01

    Full Text Available Abstract Background Aldo-keto reductase (AKR 1C family member 3 (AKR1C3, one of four identified human AKR1C enzymes, catalyzes steroid, prostaglandin, and xenobiotic metabolism. In the prostate, AKR1C3 is up-regulated in localized and advanced prostate adenocarcinoma, and is associated with prostate cancer (PCa aggressiveness. Here we propose a novel pathological function of AKR1C3 in tumor angiogenesis and its potential role in promoting PCa progression. Methods To recapitulate elevated AKR1C3 expression in cancerous prostate, the human PCa PC-3 cell line was stably transfected with an AKR1C3 expression construct to establish PC3-AKR1C3 transfectants. Microarray and bioinformatics analysis were performed to identify AKR1C3-mediated pathways of activation and their potential biological consequences in PC-3 cells. Western blot analysis, reverse transcription-polymerase chain reaction (RT-PCR, enzyme-linked immunosorbent assay (ELISA, and an in vitro Matrigel angiogenesis assays were applied to validate the pro-angiogenic activity of PC3-AKR1C3 transfectants identified by bioinformatics analysis. Results Microarray and bioinformatics analysis suggested that overexpression of AKR1C3 in PC-3 cells modulates estrogen and androgen metabolism, activates insulin-like growth factor (IGF-1 and Akt signaling pathways, as well as promotes tumor angiogenesis and aggressiveness. Levels of IGF-1 receptor (IGF-1R and Akt activation as well as vascular endothelial growth factor (VEGF expression and secretion were significantly elevated in PC3-AKR1C3 transfectants in comparison to PC3-mock transfectants. PC3-AKR1C3 transfectants also promoted endothelial cell (EC tube formation on Matrigel as compared to the AKR1C3-negative parental PC-3 cells and PC3-mock transfectants. Pre-treatment of PC3-AKR1C3 transfectants with a selective IGF-1R kinase inhibitor (AG1024 or a non-selective phosphoinositide 3-kinases (PI3K inhibitor (LY294002 abolished ability of the cells

  16. Inhibition of pH fronts in corrosion cells due to the formation of cerium hydroxide

    International Nuclear Information System (INIS)

    The effect of cerium-based corrosion inhibitors on the pH front between the alkaline cathode and acidic anode in corrosion cells has been studied. The cerium component of these inhibitors can affect the pH front since it precipitates in an alkaline environment as cerium hydroxide, which is important since the corrosion inhibition mechanism of the cerium component is a result of its deposition as a highly electrical resistive (passivation) layer on the cathode. It is studied whether the cerium can reach the cathode when fed into the corrosion cell from an external source after the onset of corrosion. To this end a simulation model was set up that includes the Poisson–Nernst–Planck theory to describe ion transport and the Frumkin–Butler–Volmer equation to describe charge transfer at the electrodes. In this model both the self-dissociation of water and the formation of cerium hydroxide are taken into account. To support our findings experimentally a corrosion cell consisting of an aluminum and copper electrode was used, in which the pH fronts were visualized using a pH-indicator. Two types of inhibitors were used; namely, highly soluble CeCl3 and sparsely soluble cerium dibutylphosphate, Ce(dbp)3. The results show that CeCl3 can reduce the size of the alkaline region and reach the cathode to form a passivation layer, whereas the solubility in case of Ce(dbp)3 is too low to supply sufficient amounts of trivalent cerium cations to penetrate the alkaline region. This behavior can be explained by the simulation results, which reveal a threshold for the corrosion inhibitor solubility below which no passivation of the cathode occurs

  17. Formation of tRNA granules in the nucleus of heat-induced human cells

    Energy Technology Data Exchange (ETDEWEB)

    Miyagawa, Ryu [Radioisotope Center, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032 (Japan); Department of Biological Science, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8654 (Japan); Mizuno, Rie [Radioisotope Center, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032 (Japan); Watanabe, Kazunori, E-mail: watanabe@ric.u-tokyo.ac.jp [Radioisotope Center, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032 (Japan); Ijiri, Kenichi [Radioisotope Center, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032 (Japan); Department of Biological Science, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8654 (Japan)

    2012-02-03

    Highlights: Black-Right-Pointing-Pointer tRNAs are tranlocated into the nucleus in heat-induced HeLa cells. Black-Right-Pointing-Pointer tRNAs form the unique granules in the nucleus. Black-Right-Pointing-Pointer tRNA ganules overlap with nuclear stress granules. -- Abstract: The stress response, which can trigger various physiological phenomena, is important for living organisms. For instance, a number of stress-induced granules such as P-body and stress granule have been identified. These granules are formed in the cytoplasm under stress conditions and are associated with translational inhibition and mRNA decay. In the nucleus, there is a focus named nuclear stress body (nSB) that distinguishes these structures from cytoplasmic stress granules. Many splicing factors and long non-coding RNA species localize in nSBs as a result of stress. Indeed, tRNAs respond to several kinds of stress such as heat, oxidation or starvation. Although nuclear accumulation of tRNAs occurs in starved Saccharomyces cerevisiae, this phenomenon is not found in mammalian cells. We observed that initiator tRNA{sup Met} (Meti) is actively translocated into the nucleus of human cells under heat stress. During this study, we identified unique granules of Meti that overlapped with nSBs. Similarly, elongator tRNA{sup Met} was translocated into the nucleus and formed granules during heat stress. Formation of tRNA granules is closely related to the translocation ratio. Then, all tRNAs may form the specific granules.

  18. Bone marrow cells participate in tumor vessel formation that supports the growth of Ewing’s sarcoma in the lung

    OpenAIRE

    Zhou, Zhichao; Stewart, Keri Schadler; Yu, Ling; Kleinerman, Eugenie S.

    2010-01-01

    An MHC-mismatch bone marrow (BM) transplant Ewing’s sarcoma mouse model was used to investigate whether BM cells participate in the vessel formation that support Ewing’s sarcoma lung metastasis. BM cells from H-2Kb/d donor mice were transplanted into sublethally irradiated H-2Kd recipient mice. Donor BM cells were identified using the H-2Kb marker. Engraftment was confirmed by identifying the H-2Kb IL-1β-type specific polymorphism. After engraftment highly lung metastatic TC71-PM4 cells were ...

  19. Biotransport and intracellular ice formation phenomena in freezing human embryonic kidney cells (HEK293T).

    Science.gov (United States)

    Xu, Yunpeng; Zhao, Gang; Zhou, Xiaoming; Ding, Weiping; Shu, Zhiquan; Gao, Dayong

    2014-04-01

    The objective of this study is to determine the cryobiological characteristics of human embryonic kidney (HEK293T) cells. The cell membrane hydraulic conductivity (L(pg)) and the activation energy of water transport (E(Lp)) were determined in the absence/presence of cryoprotectant agent (CPA), while the nucleation rate kinetic and thermodynamic parameters (Ωo(SCN) and κo(SCN)) were determined in the absence of CPA. Since dehydration and intracellular ice formation (IIF) are two factors that may cause damage to cells during the freezing process, systematical freezing experiments were carried out at different cooling rates (5, 10, 15, 20, 30, and 60°C/min) under the commercial available cryomicroscopy (FDCS 196, Linkham, Waterfield, UK) to further explore the cryoinjury mechanism for HEK293T cells. By simultaneously fitting the water transport equation to the experimentally measured volumetric shrinkage data at 5, 10, and 15°C/min, the "combined best fit" membrane permeability parameters for HEK293T cells in both phosphate buffer saline (PBS) and CPA media (0.75M Me2SO in PBS) are determined. They are L(pg)=2.85×10(-14)m/s/Pa (0.17μm/min/atm), E(Lp)=142.91kJ/mol (34.13kcal/mol) (R(2)=0.990), and L(pg)[cpa]=2.73±0.44×10(-14)m/s/Pa (0.16±0.03μm/min/atm), E(Lp)[cpa]=152.52±27.69kJ/mol (36.42±6.61kcal/mol) (R(2)=0.993), respectively. An optimal cooling rate B(opt) (the highest cooling rate without IIF) was determined to be 14.24°C/min in the absence of CPA. Additionally, the ice nucleation parameters (Ωo(SCN) and κo(SCN)) were averaged to be 1.31±0.11×10(8)m(-2)s(-1) and 7.67±2.55×10(9)K(5) for the cooling rates 20, 30, and 60°C/min. PMID:24582893

  20. [Diverse morphological types of dormant cells and conditions for their formation in Azospirillum brasilense].

    Science.gov (United States)

    Muliukin, A L; Suzina, N E; Pogorelova, A Iu; Antoniuk, L P; Duda, V I; El'-Registan, G I

    2009-01-01

    Differences in generation of dormant forms (DF) were revealed between two strains of non-sporeforming gram-negative bacteria Azospirillum brasilense, Sp7 (non-endophytic) and Sp245 (endophytic strain). In post-stationary ageing bacterial cultures grown in a synthetic medium with a fivefold decreased initial nitrogen content, strain Sp7 formed two types of cyst-like resting cells (CRC). Strain Sp245 did not form such types of DF under the same conditions. CRC of the first type were formed in strain Sp245 only under phosphorus deficiency (C > P). The endophytic strain was also shown to form structurally differentiated cells under complete starvation, i.e. at a transfer of early stationary cultures, grown in the media with C > N unbalance, to saline solution (pH 7.2). These DF had a complex structure similar to that of azotobacter cysts. The CRC, which are generated by both azospirilla strains and belong to distinct morphological types, possessed the following major features: absence of division; specific ultrastructural organization; long-term maintenance of viability (for 4 months and more); higher heat resistance (50-60 degrees C, 10 min) as compared with vegetative cells, i.e. the important criteria for dormant prokaryotic forms. However, CRC of non-endophytic strain Sp7 had higher heat resistance (50, 55, 60 degrees C). The viability maintenance and the portion of heat-resistant cells depended on the conditions of maturation and storage of CRC populations. Long-term storage (for 4 months and more) of azospirilla DF populations at -20 degrees C was optimal for maintenance of their colony-forming ability (57% of the CFU number in stationary cultures), whereas the largest percentage of heat-resistant cells was in CRC suspensions incubated in a spent culture medium (but not in saline solution) at room temperature. The data on the intraspecies diversity of azospirilla DF demonstrate the relation between certain type DF formation to the type of interaction (non

  1. Novel Mechanisms of Compromised Lymphatic Endothelial Cell Homeostasis in Obesity: The Role of Leptin in Lymphatic Endothelial Cell Tube Formation and Proliferation.

    Directory of Open Access Journals (Sweden)

    Akinori Sato

    Full Text Available Leptin is a hormone produced by adipose tissue that regulates various physiological processes. Recent studies have shown that the level of circulating leptin is elevated in obese patients and have suggested a relationship between obesity and postoperative lymphedema. However, the mechanisms by which postoperative lymphedema develops in obese patients and the mechanisms by which leptin regulates lymphatic endothelial cell homeostasis such as tube formation and cell proliferation remain unknown. Here we report that leptin regulates tube formation and cell proliferation in human dermal lymphatic endothelial cells (HDLECs by activation of the signal transducer and activator of transcription 3 pathway, which is downstream signaling of the leptin receptor. Additionally, we found that upregulation of suppressor of cytokine signaling 3 underlies the mechanisms by which a high dose of leptin inhibits cell proliferation and tube formation. Leptin also enhanced expression of the proinflammatory cytokine IL-6 in HDLECs. Interestingly, IL-6 rescues the compromised cell proliferation and tube formation caused by treatment with a high dose of leptin in an autocrine or paracrine manner. Taken together, our findings reveal a novel mechanism by which compromised HDLECs maintain their homeostasis during inflammation mediated by leptin and IL-6. Thus, regulating the level of leptin or IL-6 may be a viable strategy to reduce the incidence of postoperative lymphedema.

  2. Novel Mechanisms of Compromised Lymphatic Endothelial Cell Homeostasis in Obesity: The Role of Leptin in Lymphatic Endothelial Cell Tube Formation and Proliferation

    Science.gov (United States)

    Kawata, Koji; Kawada, Masaya; Jitsukawa, Sumito; Yamashita, Keiji; Sato, Noriyuki; Himi, Tetsuo; Ichimiya, Shingo

    2016-01-01

    Leptin is a hormone produced by adipose tissue that regulates various physiological processes. Recent studies have shown that the level of circulating leptin is elevated in obese patients and have suggested a relationship between obesity and postoperative lymphedema. However, the mechanisms by which postoperative lymphedema develops in obese patients and the mechanisms by which leptin regulates lymphatic endothelial cell homeostasis such as tube formation and cell proliferation remain unknown. Here we report that leptin regulates tube formation and cell proliferation in human dermal lymphatic endothelial cells (HDLECs) by activation of the signal transducer and activator of transcription 3 pathway, which is downstream signaling of the leptin receptor. Additionally, we found that upregulation of suppressor of cytokine signaling 3 underlies the mechanisms by which a high dose of leptin inhibits cell proliferation and tube formation. Leptin also enhanced expression of the proinflammatory cytokine IL-6 in HDLECs. Interestingly, IL-6 rescues the compromised cell proliferation and tube formation caused by treatment with a high dose of leptin in an autocrine or paracrine manner. Taken together, our findings reveal a novel mechanism by which compromised HDLECs maintain their homeostasis during inflammation mediated by leptin and IL-6. Thus, regulating the level of leptin or IL-6 may be a viable strategy to reduce the incidence of postoperative lymphedema. PMID:27366905

  3. Robust Optimization Approach for Design for a Dynamic Cell Formation Considering Labor Utilization: Bi-objective Mathematical Mode

    Directory of Open Access Journals (Sweden)

    Hiwa Farughi

    2016-05-01

    Full Text Available In this paper, robust optimization of a bi-objective mathematical model in a dynamic cell formation problem considering labor utilization with uncertain data is carried out. The robust approach is used to reduce the effects of fluctuations of the uncertain parameters with regards to all the possible future scenarios. In this research, cost parameters of the cell formation and demand fluctuations are subject to uncertainty and a mixed-integer programming (MIP model is developed to formulate the related robust dynamic cell formation problem. Then the problem is transformed into a bi-objective linear one. The first objective function seeks to minimize relevant costs of the problem including machine procurement and relocation costs, machine variable cost, inter-cell movement and intra-cell movement costs, overtime cost and labor shifting cost between cells, machine maintenance cost, inventory, holding part cost. The second objective function seeks to minimize total man-hour deviations between cells or indeed labor utilization of the modeled.

  4. Murid Gammaherpesvirus Latency-Associated Protein M2 Promotes the Formation of Conjugates between Transformed B Lymphoma Cells and T Helper Cells.

    Directory of Open Access Journals (Sweden)

    Diana Fontinha

    Full Text Available Establishment of persistent infection in memory B cells by murid herpesvirus-4 (MuHV-4 depends on the proliferation of latently infected germinal center B cells, for which T cell help is essential. Whether the virus is capable of modulating B-T helper cell interaction for its own benefit is still unknown. Here, we investigate if the MuHV-4 latency associated M2 protein, which assembles multiprotein complexes with B cell signaling proteins, plays a role. We observed that M2 led to the upregulation of adhesion and co-stimulatory molecules in transduced B cell lines. In an MHC-II restricted OVA peptide-specific system, M2 polarized to the B-T helper contact zone. Furthermore, it promoted B cell polarization, as demonstrated by the increased proximity of the B cell microtubule organizing center to the interface. Consistent with these data, M2 promoted the formation of B-T helper cell conjugates. In an in vitro competition assay, this translated into a competitive advantage, as T cells preferentially conjugated with M2-expressing B cells. However, expression of M2 alone in B cells was not sufficient to lead to T cell activation, as it only occurred in the presence of specific peptide. Taken together, these findings support that M2 promotes the formation of B-T helper cell conjugates. In an in vivo context this may confer a competitive advantage to the infected B cell in acquisition of T cell help and initiation of a germinal center reaction, hence host colonization.

  5. Establishment of a preadipocyte cell line derived from mature adipocytes of GFP transgenic mice and formation of adipose tissue.

    Science.gov (United States)

    Nobusue, Hiroyuki; Endo, Tsuyoshi; Kano, Koichiro

    2008-06-01

    We established a preadipocyte cell line from mature adipocytes obtained from subcutaneous fat tissue of green fluorescent protein (GFP) transgenic mice. The floating top layer, containing mature adipocytes, was isolated from subcutaneous fat tissue by collagenase digestion and filtration. Fluorescence-activated cell sorting and microscopic analysis revealed that the floating cell fraction comprised a highly homogeneous adipocyte population with no adipose stromal-vascular cells. Isolated mature adipocytes dedifferentiated into fibroblast-like cells and actively proliferated in ceiling culture. In vitro studies showed that the cells could redifferentiate into mature adipocytes in an identical way to 3T3-L1 preadipocytes. No changes in the differentiation pattern were observed during the propagation of our cells. They were successfully maintained and differentiated for at least 22 passages. We named these cells dedifferentiated fat (DFAT-GFP) cells. When DFAT-GFP cells were implanted subcutaneously into C57BL/6N mice, they developed highly vascularized fat pads that morphologically resembled normal subcutaneous adipose tissue and consisted of GFP-positive cells; however, implanted 3T3-L1 cells did not have such an effect on the mice. We conclude that DFAT-GFP cells provide a model that should enable us to study the mechanisms of adipocyte differentiation and adipose tissue formation in vivo and in vitro. PMID:18386066

  6. Impact of flavonoids on matrix metalloproteinase secretion and invadopodia formation in highly invasive A431-III cancer cells.

    Directory of Open Access Journals (Sweden)

    Yo-Chuen Lin

    Full Text Available Metastasis is a major cause of mortality in cancer patients. Invadopodia are considered to be crucial structures that allow cancer cells to penetrate across the extracellular matrix (ECM by using matrix metalloproteinases (MMPs. Previously, we isolated a highly invasive A431-III subline from parental A431 cells by Boyden chamber assay. The A431-III cells possess higher invasive and migratory abilities, elevated levels of MMP-9 and an enhanced epithelial-mesenchymal transition (EMT phenotype. In this study, we discovered that A431-III cells had an increased potential to form invadopodia and an improved capacity to degrade ECM compared with the original A431 cells. We also observed enhanced phosphorylation levels of cortactin and Src in A431-III cells; these phosphorylated proteins have been reported to be the main regulators of invadopodia formation. Flavonoids, almost ubiquitously distributed in food plants and plant food products, have been documented to exhibit anti-tumor properties. Therefore, it was of much interest to explore the effects of flavonoid antioxidants on the metastatic activity of A431-III cells. Exposure of A431-III cells to two potent dietary flavonoids, namely luteolin (Lu and quercetin (Qu, caused inhibition of invadopodia formation and decrement in ECM degradation. We conclude that Lu and Qu attenuate the phosphorylation of cortactin and Src in A431-III cells. As a consequence, there ensues a disruption of invadopodia generation and the suppression of MMP secretion. These changes, in concert, bring about a reduction in metastasis.

  7. 5-azacytidine induces anoikis, inhibits mammosphere formation and reduces metalloproteinase 9 activity in MCF-7 human breast cancer cells.

    Science.gov (United States)

    Chang, Hsueh-Wei; Wang, Hui-Chun; Chen, Chiau-Yi; Hung, Ting-Wei; Hou, Ming-Feng; Yuan, Shyng-Shiou F; Huang, Chih-Jen; Tseng, Chao-Neng

    2014-01-01

    Cancer stem cells are a subset of cancer cells that initiate the growth of tumors. Low levels of cancer stem cells also exist in established cancer cell lines, and can be enriched in serum-free tumorsphere cultures. Since cancer stem cells have been reported to be resilient to common chemotherapeutic drugs in comparison to regular cancer cells, screening for compounds selectively targeting cancer stem cells may provide an effective therapeutic strategy. We found that 5-azacytidine (5-AzaC) selectively induced anoikis of MCF-7 in suspension cultures with an EC₅₀ of 8.014 µM, and effectively inhibited tumorsphere formation, as well as the migration and matrix metalloproteinases-9 (MMP-9) activity of MCF-7 cells. Furthermore, 5-AzaC and radiation collaboratively inhibited MCF-7 tumorsphere formation at clinically relevant radiation doses. Investigating the underlying mechanism may provide insight into signaling pathways crucial for cancer stem cell survival and pave the way to novel potential therapeutic targets. PMID:24633350

  8. Sequential Salinomycin Treatment Results in Resistance Formation through Clonal Selection of Epithelial-Like Tumor Cells12

    Science.gov (United States)

    Kopp, Florian; Hermawan, Adam; Oak, Prajakta Shirish; Ulaganathan, Vijay Kumar; Herrmann, Annika; Elnikhely, Nefertiti; Thakur, Chitra; Xiao, Zhiguang; Knyazev, Pjotr; Ataseven, Beyhan; Savai, Rajkumar; Wagner, Ernst; Roidl, Andreas

    2014-01-01

    Acquiring therapy resistance is one of the major obstacles in the treatment of patients with cancer. The discovery of the cancer stem cell (CSC)–specific drug salinomycin raised hope for improved treatment options by targeting therapy-refractory CSCs and mesenchymal cancer cells. However, the occurrence of an acquired salinomycin resistance in tumor cells remains elusive. To study the formation of salinomycin resistance, mesenchymal breast cancer cells were sequentially treated with salinomycin in an in vitro cell culture assay, and the resulting differences in gene expression and salinomycin susceptibility were analyzed. We demonstrated that long-term salinomycin treatment of mesenchymal cancer cells resulted in salinomycin-resistant cells with elevated levels of epithelial markers, such as E-cadherin and miR-200c, a decreased migratory capability, and a higher susceptibility to the classic chemotherapeutic drug doxorubicin. The formation of salinomycin resistance through the acquisition of epithelial traits was further validated by inducing mesenchymal-epithelial transition through an overexpression of miR-200c. The transition from a mesenchymal to a more epithelial-like phenotype of salinomycin-treated tumor cells was moreover confirmed in vivo, using syngeneic and, for the first time, transgenic mouse tumor models. These results suggest that the acquisition of salinomycin resistance through the clonal selection of epithelial-like cancer cells could become exploited for improved cancer therapies by antagonizing the tumor-progressive effects of epithelial-mesenchymal transition. PMID:25500079

  9. 5-Azacytidine Induces Anoikis, Inhibits Mammosphere Formation and Reduces Metalloproteinase 9 Activity in MCF-7 Human Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Hsueh-Wei Chang

    2014-03-01

    Full Text Available Cancer stem cells are a subset of cancer cells that initiate the growth of tumors. Low levels of cancer stem cells also exist in established cancer cell lines, and can be enriched in serum-free tumorsphere cultures. Since cancer stem cells have been reported to be resilient to common chemotherapeutic drugs in comparison to regular cancer cells, screening for compounds selectively targeting cancer stem cells may provide an effective therapeutic strategy. We found that 5-azacytidine (5-AzaC selectively induced anoikis of MCF-7 in suspension cultures with an EC50 of 8.014 µM, and effectively inhibited tumorsphere formation, as well as the migration and matrix metalloproteinases-9 (MMP-9 activity of MCF-7 cells. Furthermore, 5-AzaC and radiation collaboratively inhibited MCF-7 tumorsphere formation at clinically relevant radiation doses. Investigating the underlying mechanism may provide insight into signaling pathways crucial for cancer stem cell survival and pave the way to novel potential therapeutic targets.

  10. MicroRNA-34a inhibits osteoblast differentiation and in vivo bone formation of human stromal stem cells

    DEFF Research Database (Denmark)

    Chen, Li; Holmstrøm, Kim; Qiu, Weimin;

    2014-01-01

    of miR-34a. siRNA-mediated reduction of JAG1 expression inhibited OB differentiation. Moreover, a number of known cell cycle regulator and cell proliferation proteins, such as cyclin D1, cyclin-dependent kinase 4 and 6 (CDK4 and CDK6), E2F transcription factor three, and cell division cycle 25 homolog......Osteoblast differentiation and bone formation (osteogenesis) are regulated by transcriptional and post-transcriptional mechanisms. Recently, microRNAs (miRNAs) were identified as novel key regulators of human stromal (skeletal, mesenchymal) stem cells (hMSC) differentiation. Here, we identified mi...... effects controlling both hMSC proliferation and OB differentiation. Tissue-specific inhibition of miR-34a might be a potential novel therapeutic strategy for enhancing in vivo bone formation....

  11. Enhanced caveolin-1 expression in smooth muscle cells: Possible prelude to neointima formation

    Institute of Scientific and Technical Information of China (English)

    Jing; Huang; John; H; Wolk; Michael; H; Gewitz; James; E; Loyd; James; West; Eric; D; Austin; Rajamma; Mathew

    2015-01-01

    AIM: To study the genesis of neointima formation in pulmonary hypertension(PH), we investigated the role of caveolin-1 and related proteins. METHODS: Male Sprague Dawley rats were given monocrotaline(M, 40 mg/kg) or subjected to hypobaric hypoxia(H) to induce PH. Another group was given M and subjected to H to accelerate the disease process(M + H). Right ventricular systolic pressure, right ventricular hypertrophy, lung histology for medial hypertrophy and the presence of neointimal lesions were examined at 2 and 4 wk. The expression of caveolin-1 and its regulatory protein peroxisome proliferator-activated receptor(PPAR) γ, caveolin-2, proliferative and antiapoptotic factors(PY-STAT3, p-Erk, Bcl-x L), endothelial nitric oxide synthase(e NOS) and heat shock protein(HSP) 90 in the lungs were analyzed, and the results from M + H group were compared with the controls, M and H groups. Double immunofluorescence technique was used to identify the localization of caveolin-1 in pulmonary arteries in rat lungs and in human PH lung tissue. RESULTS: In the M + H group, PH was more severe compared with M or H group. In the 4 wk M+H group, several arteries with reduced caveolin-1 expression in endothelial layer coupled with an increased expression in smooth muscle cells(SMC), exhibited neointimal lesions. Neointima was present only in the arteries exhibiting enhanced caveolin-1 expression in SMC. Lung tissue obtained from patients with PH also revealed neointimal lesions only in the arteries exhibiting endothelial caveolin-1 loss accompanied by an increased caveolin-1 expression in SMC. Reduction in e NOS and HSP90 expression was present in the M groups(2 and 4 wk), but not in the M + H groups. In both M groups and in the M + H group at 2 wk, endothelial caveolin-1 loss was accompanied by an increase in PPARγ expression. In the M + H group at 4 wk, increase in caveolin-1 expression was accompanied by a reduction in the PPARγ expression. In the H group, there was neither a

  12. Formation of hydroxyl radical from San Joaquin Valley particles extracted in a cell-free solution

    Science.gov (United States)

    Shen, H.; Anastasio, C.

    2011-06-01

    Previous studies have suggested that the adverse health effects from ambient particulate matter (PM) are linked to the formation of reactive oxygen species (ROS) by PM. While hydroxyl radical (•OH) is the most reactive of the ROS species, there are few quantitative studies of •OH generation from PM. Here we report on •OH formation from PM collected at an urban (Fresno) and rural (Westside) site in the San Joaquin Valley (SJV) of California. We quantified •OH in PM extracts using a cell-free, phosphate-buffered saline (PBS) solution with or without 50 μM ascorbate (Asc). The results show that generally the urban Fresno PM generates much more •OH than the rural Westside PM. The presence of Asc at a physiologically relevant concentration in the extraction solution greatly enhances •OH formation from all the samples. Fine PM (PM2.5) generally makes more •OH than the corresponding coarse PM (PMcf, i.e., 2.5 to 10 μm) normalized by air volume collected, while the coarse PM typically generates more •OH normalized by PM mass. •OH production by SJV PM is reduced on average by (97 ± 6) % when the transition metal chelator desferoxamine (DSF) is added to the extraction solution, indicating a dominant role of transition metals. By measuring calibration curves of •OH generation from copper and iron, and quantifying copper and iron concentrations in our particle extracts, we find that PBS-soluble copper is primarily responsible for •OH production by the SJV PM, while iron often makes a significant contribution. Extrapolating our results to expected burdens of PM-derived •OH in human lung lining fluid suggests that typical daily PM exposures in the San Joaquin Valley are unlikely to result in a high amount of pulmonary •OH, although high PM events could produce much higher levels of •OH, which might lead to cytotoxicity.

  13. Formation of hydroxyl radical from San Joaquin Valley particles extracted in a cell-free solution

    Directory of Open Access Journals (Sweden)

    H. Shen

    2011-06-01

    Full Text Available Previous studies have suggested that the adverse health effects from ambient particulate matter (PM are linked to the formation of reactive oxygen species (ROS by PM. While hydroxyl radical (OH is the most reactive of the ROS species, there are few quantitative studies of OH generation from PM. Here we report on OH formation from PM collected at an urban (Fresno and rural (Westside site in the San Joaquin Valley (SJV of California. We quantified OH in PM extracts using a cell-free, phosphate-buffered saline (PBS solution with or without 50 μM ascorbate (Asc. The results show that generally the urban Fresno PM generates much more OH than the rural Westside PM. The presence of Asc at a physiologically relevant concentration in the extraction solution greatly enhances OH formation from all the samples. Fine PM (PM2.5 generally makes more OH than the corresponding coarse PM (PMcf, i.e., 2.5 to 10 μm normalized by air volume collected, while the coarse PM typically generates more OH normalized by PM mass. OH production by SJV PM is reduced on average by (97 ± 6 % when the transition metal chelator desferoxamine (DSF is added to the extraction solution, indicating a dominant role of transition metals. By measuring calibration curves of OH generation from copper and iron, and quantifying copper and iron concentrations in our particle extracts, we find that PBS-soluble copper is primarily responsible for OH production by the SJV PM, while iron often makes a significant contribution. Extrapolating our results to expected burdens of PM-derived OH in human lung lining fluid suggests that typical daily PM exposures in the San Joaquin Valley are unlikely to result in a high amount of pulmonary OH, although high PM events could produce much higher levels

  14. Slow Turning Lateral Vessel Bioreactor Improves Embryoid Body Formation and Cardiogenic Differentiation of Mouse Embryonic Stem Cells

    OpenAIRE

    2013-01-01

    Embryonic stem cells (ESCs) have the ability to form aggregates, which are called embryoid bodies (EBs). EBs mimic early embryonic development and are commonly produced for cardiomyogenesis. Here, we describe a method of EB formation in hydrodynamic conditions using a slow-turning lateral vessel (STLV) bioreactor and the subsequent differentiation of EBs into cardiomyocytes. EBs formed in the STLV were compared with conventional techniques, such as hanging drop (HD) or static suspension cell ...

  15. Formation mechanisms of Cu(In,Ga)Se{sub 2} solar cells prepared from electrodeposited precursors

    Energy Technology Data Exchange (ETDEWEB)

    Oliva, F., E-mail: florian.oliva@nexcis.fr [Nexcis Photovoltaic Technology, 190 Avenue Célestin Coq, 13106 Rousset (France); Ecole Nationale Supérieure des Mines, CMP-EMSE, MOC, 880 Avenue de Mimet, 13541 Gardanne (France); Broussillou, C.; Annibaliano, M.; Frederich, N.; Grand, P.P. [Nexcis Photovoltaic Technology, 190 Avenue Célestin Coq, 13106 Rousset (France); Roussy, A.; Collot, P. [Ecole Nationale Supérieure des Mines, CMP-EMSE, MOC, 880 Avenue de Mimet, 13541 Gardanne (France); Bodnar, S. [Nexcis Photovoltaic Technology, 190 Avenue Célestin Coq, 13106 Rousset (France)

    2013-05-01

    The development of low cost industrial processes is one of the key issues to make Cu(In,Ga)Se{sub 2} based solar cells reach grid-parity. Such a process is found by using a two-step technology based on the sequential electro-deposition of a metallic precursor followed by a rapid annealing. Three types of metallic precursors (two-compound systems as copper–indium, copper–gallium and three-compound system as copper–indium–gallium) have been electrodeposited on a molybdenum sputtered soda lime glass and alloyed through a low annealing temperature. Then a selenium film has been evaporated and the stack has been annealed at high temperature in a rapid thermal processing furnace. A one-step heating profile has been used from room temperature to 550 °C in less than 1 min. Samples for which the heating was stopped after different annealing times have been characterized using several techniques: X-ray fluorescence spectrometry for elemental composition, X-ray diffraction and Raman spectroscopy for phase composition, scanning electron microscopy for structural analysis and glow discharge optical emission spectroscopy for diffusion study. Preferential formation reactions of the two-compound based metallic precursors have been studied and compared with the copper–indium–gallium metallic precursor used in a two step process. A gallium free system reacts faster than a gallium-based system and presents well-formed ternary compound after a standard selenization. However, the incorporation of gallium can be improved through a longer annealing time or a higher annealing temperature. - Highlights: ► We studied formation mechanisms of Cu(In,Ga)Se{sub 2} based solar cells. ► Electrodeposited metallic precursors are selenized in a two step process. ► Three systems were analyzed: Cu–In + Se, Cu–Ga + Se and Cu–In–Ga + Se systems. ► A longer annealing time improves the incorporation of gallium. ► A higher annealing temperature improves the incorporation of

  16. TM4SF1: a tetraspanin-like protein necessary for nanopodia formation and endothelial cell migration.

    Science.gov (United States)

    Zukauskas, Andrew; Merley, Anne; Li, Dan; Ang, Lay-Hong; Sciuto, Tracey E; Salman, Samantha; Dvorak, Ann M; Dvorak, Harold F; Jaminet, Shou-Ching Shih

    2011-09-01

    Transmembrane-4-L-six-family-1 (TM4SF1) is a tetraspanin-like membrane protein that is highly and selectively expressed by cultured endothelial cells (EC) and, in vivo, by EC lining angiogenic tumor blood vessels. TM4SF1 is necessary for the formation of unusually long (up to a 50 μm), thin (~100-300 nm wide), F-actin-poor EC cell projections that we term 'nanopodia'. Immunostaining of nanopodia at both the light and electron microsopic levels localized TM4SF1 in a regularly spaced, banded pattern, forming TM4FS1-enriched domains. Live cell imaging of GFP-transduced HUVEC demonstrated that EC project nanopodia as they migrate and interact with neighboring cells. When TM4SF1 mRNA levels in EC were increased from the normal ~90 mRNA copies/cell to ~400 copies/cell through adenoviral transduction, EC projected more and longer nanopodia from the entire cell circumference but were unable to polarize or migrate effectively. When fibroblasts, which normally express TM4SF1 at ~5 copies/cell, were transduced to express TM4SF1 at EC-like levels, they formed typical TM4SF1-banded nanopodia, and broadened, EC-like lamellipodia. Mass-spectrometry demonstrated that TM4SF1 interacted with myosin-10 and β-actin, proteins involved in filopodia formation and cell migration. In summary, TM4SF1, like genuine tetraspanins, serves as a molecular organizer that interacts with membrane and cytoskeleton-associated proteins and uniquely initiates the formation of nanopodia and facilitates cell polarization and migration. PMID:21626280

  17. Pathways Regulating Spheroid Formation of Human Follicular Thyroid Cancer Cells under Simulated Microgravity Conditions: A Genetic Approach

    Directory of Open Access Journals (Sweden)

    Stefan Riwaldt

    2016-04-01

    Full Text Available Microgravity induces three-dimensional (3D growth in numerous cell types. Despite substantial efforts to clarify the underlying mechanisms for spheroid formation, the precise molecular pathways are still not known. The principal aim of this paper is to compare static 1g-control cells with spheroid forming (MCS and spheroid non-forming (AD thyroid cancer cells cultured in the same flask under simulated microgravity conditions. We investigated the morphology and gene expression patterns in human follicular thyroid cancer cells (UCLA RO82-W-1 cell line after a 24 h-exposure on the Random Positioning Machine (RPM and focused on 3D growth signaling processes. After 24 h, spheroid formation was observed in RPM-cultures together with alterations in the F-actin cytoskeleton. qPCR indicated more changes in gene expression in MCS than in AD cells. Of the 24 genes analyzed VEGFA, VEGFD, MSN, and MMP3 were upregulated in MCS compared to 1g-controls, whereas ACTB, ACTA2, KRT8, TUBB, EZR, RDX, PRKCA, CAV1, MMP9, PAI1, CTGF, MCP1 were downregulated. A pathway analysis revealed that the upregulated genes code for proteins, which promote 3D growth (angiogenesis and prevent excessive accumulation of extracellular proteins, while genes coding for structural proteins are downregulated. Pathways regulating the strength/rigidity of cytoskeletal proteins, the amount of extracellular proteins, and 3D growth may be involved in MCS formation.

  18. Factors Leading to the Formation of a Resistive Thin Film at the Bottom of Aluminum Electrolysis Cells

    Science.gov (United States)

    Coulombe, Marc-André; Soucy, Gervais; Rivoaland, Loig; Davies, Lynne

    2016-04-01

    Studies on sludge formation in aluminum electrolysis cells are rare and typically do not distinguish the deposits at the center of the cell from those composing the ledge toe because low voltage lost is expected at the center of the cell. However, high amount of sludge in the center leads to the formation of a thin film in an intermediate zone between the ledge toe and this center thick sludge accumulation. Looking at sludge deposits through composition mapping and microstructure analysis coming from four aluminum cells of two different aluminum reduction technologies, major factors leading to a thin resistive film were identified. This includes the formation of a suspension on the top of the thick deposit at the center of the cell, its displacement through magnetohydrodynamic induced movement by the metal pad, and the growth and thickening of a carbide sublayer making the thin film even more resistive. Correlation between thickening of the thin film and cathode voltage drop increase was observed. The postmortem analysis performed on six laboratory experiments was found useful to support different observations made on the industrial cells at lower cost.

  19. Effect of Cell Cycle Inhibitor Olomoucine on Astroglial Proliferation and Scar Formation after Focal Cerebral Infarction in Rats

    Institute of Scientific and Technical Information of China (English)

    MANG Gui-bin; TIAN Dai-shi; XU Yun-lan; XIE Min-jie; WANG Ping; DU Yi-xing; WANG Wei

    2011-01-01

    Background: Astrocytes become reactive following many types of CNS injuries.Excessive astrogliosis is detrimental and contributes to neuronal damage. We sought to determine whether inhibition of cell cycle could decrease the proliferation of astroglial cells and therefore reduce excessive gliosis and glial scar formation after focal ischemia. Methods: Cerebral infarctionmodel was induced by photothrombosis method. Rats were examined using MRI, and lesion volumes were estimated on day 3 post-infarction. The expression of glial fibrillary acidic protein(GFAP) and proliferating cell nuclear antigen(PCNA) was observed by immunofluorescence staining. Protein levels for GFAP, PCNA, Cyclin A and Cyclin B1 were determined by Western blot analysis from the ischemic and sham animals sacrificed at 3,7,30 days after operation. Results:Cell cycle inhibitor olomoucine significantly suppressed GFAP and PCNA expression and reduced lesion volume after cerebral ischemia. In parallel studies, we found dense astroglial scar in boundary zone of vehicle-treated rats at 7 and 30 days. Olomoucine can markedly attenuate astroglial scar formation. Western blot analysis showed increased protein levels of GFAP, PCNA,Cyclin A and Cyclin B1 after ischemia, which was reduced by olomoucine treatment. Conclusion:Our results suggested that astroglial activation, proliferation and subsequently astroglial scar formation could be partially inhibited by regulation of cell cycle. Cell cycle modulation thereby pro-vides a potential promising strategy to treat cerebral ischemia.

  20. Cell-Based High-Throughput Screening Identifies Rifapentine as an Inhibitor of Amyloid and Biofilm Formation in Escherichia coli.

    Science.gov (United States)

    Maher, Marie C; Lim, Ji Youn; Gunawan, Cheston; Cegelski, Lynette

    2015-10-01

    Escherichia coli assemble functional amyloid fibers termed curli that contribute to bacterial adhesion, biofilm formation, and host pathogenesis. We developed a cell-based high-throughput screen to identify inhibitors of curli-mediated adhesion in the laboratory strain MC4100 and curli-associated biofilm formation in the uropathogenic E. coli clinical isolate UTI89. Inhibitors of biofilm formation can operate through many mechanisms, and such inhibitors could hold therapeutic value in preventing and treating urinary tract infections. The curli-specific screen allows the identification of compounds that inhibit either curli expression, curli biogenesis, or adhesion by normally produced curli. In screening the NIH Clinical Collection of 446 compounds, we identified rifapentine as a potent inhibitor in both of these screens. Rifapentine is an antibiotic used to treat tuberculosis that targets RNA polymerase, but prevents curli-dependent adhesion and biofilm formation in E. coli at concentrations below those that affect viability. Rifapentine inhibits curli production and prevents biofilm formation on plastic, on agar, and at the air-liquid interface by inhibiting curli gene transcription. Comparisons with a cephalosporin antibiotic further revealed that curli production is not affected by standard antibiotic treatment and cell killing pressure. Thus, we reveal a new role independent of killing activity for rifapentine as an inhibitor of curli and curli-mediated biofilm formation. PMID:27623312

  1. Rapid oriented fibril formation of fish scale collagen facilitates early osteoblastic differentiation of human mesenchymal stem cells.

    Science.gov (United States)

    Matsumoto, Rena; Uemura, Toshimasa; Xu, Zhefeng; Yamaguchi, Isamu; Ikoma, Toshiyuki; Tanaka, Junzo

    2015-08-01

    We studied the effect of fibril formation of fish scale collagen on the osteoblastic differentiation of human mesenchymal stem cells (hMSCs). We found that hMSCs adhered easily to tilapia scale collagen, which remarkably accelerated the early stage of osteoblastic differentiation in hMSCs during in vitro cell culture. Osteoblastic markers such as ALP activity, osteopontin, and bone morphogenetic protein 2 were markedly upregulated when the hMSCs were cultured on a tilapia collagen surface, especially in the early osteoblastic differentiation stage. We hypothesized that this phenomenon occurs due to specific fibril formation of tilapia collagen. Thus, we examined the time course of collagen fibril formation using high-speed atomic force microscopy. Moreover, to elucidate the effect of the orientation of fibril formation on the differentiation of hMSCs, we measured ALP activity of hMSCs cultured on two types of tilapia scale collagen membranes with different degrees of fibril formation. The ALP activity in hMSCs cultured on a fibrous collagen membrane was significantly higher than on a non-fibrous collagen membrane even before adding osteoblastic differentiation medium. These results showed that the degree of the fibril formation of tilapia collagen was essential for the osteoblastic differentiation of hMSCs. PMID:25546439

  2. Culture supernatants of breast cancer cell line MDA-MB-231 treated with parthenolide inhibit the proliferation, migration, and lumen formation capacity of human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    LI Cai-juan; GUO Su-fen; SHI Tie-mei

    2012-01-01

    Background Parthenolide has been tested for anti-tumor activities,such as anti-proliferation and pro-apoptosis in recent studies.However,little is known about its role in the process of tumor angiogenesis.This study aims to investigate the effects and potential mechanisms of parthenolide on the proliferation,migration and lumen formation capacity of human umbilical vein endothelial cells.Methods Different concentrations of parthenolide were applied to the human breast cancer cell line MDA-MB-231 cells.After 24-hour incubation,the culture supematants were harvested and used to treat human umbilical vein endothelial cells for 24 hours.Then an inverted fluorescence phase contrast microscope was used to evaluate the human umbilical vein endothelial cells.The secretion of vascular endothelial growth factor (VEGF),interleukin (IL)-8 and matrix metalloproteinases (MMP)-9 in the culture supernatant of the MDA-MB-231 cells was then measured with enzyme-linked immunosorbent assay (ELISA) assays.Results Suppression of proliferation,migration,and the lumen formation capacity of human umbilical vein endothelial cells was observed in the presence of the culture supernatants from the breast cancer cell line treated with different concentrations of parthenolide.Parthenolide decreased the levels of the angiogenic factors MMP-9,VEGF,and IL-8secreted by the MDA-MB-231 cells.Conclusions Parthenolide may suppress angiogenesis through decreasing angiogenic factors secreted by breast cancer cells to interfere with the proliferation,migration and lumen-like structure formation of endothelial cells,thereby inhibiting tumor growth.It is a promising potential anti-angiogenic drug.

  3. Progressive developmental restriction, acquisition of left-right identity and cell growth behavior during lobe formation in mouse liver development.

    Science.gov (United States)

    Weiss, Mary C; Le Garrec, Jean-Francois; Coqueran, Sabrina; Strick-Marchand, Helene; Buckingham, Margaret

    2016-04-01

    To identify cell-based decisions implicated in morphogenesis of the mammalian liver, we performed clonal analysis of hepatocytes/hepatoblasts in mouse liver development, using a knock-in allele of Hnf4a/laacZ This transgene randomly undergoes a low frequency of recombination that generates a functional lacZ gene that produces β-galactosidase in tissues in which Hnf4a is expressed. Two types of β-galactosidase-positive clones were found. Most have undergone three to eight cell divisions and result from independent events (Luria-Delbrück fluctuation test); we calculate that they arose between E8.5 and E13.5. A second class was mega-clones derived from early endoderm progenitors, generating many descendants. Some originated from multi-potential founder cells, with labeled cells in the liver, pancreas and/or intestine. A few mega-clones populate only one side of the liver, indicating hepatic cell chirality. The patterns of labeled cells indicate cohesive and often oriented growth, notably in broad radial stripes, potentially implicated in the formation of liver lobes. This retrospective clonal analysis gives novel insights into clonal origins, cell behavior of progenitors and distinct properties of endoderm cells that underlie the formation and morphogenesis of the liver. PMID:26893346

  4. Progressive developmental restriction, acquisition of left-right identity and cell growth behavior during lobe formation in mouse liver development.

    Science.gov (United States)

    Weiss, Mary C; Le Garrec, Jean-Francois; Coqueran, Sabrina; Strick-Marchand, Helene; Buckingham, Margaret

    2016-04-01

    To identify cell-based decisions implicated in morphogenesis of the mammalian liver, we performed clonal analysis of hepatocytes/hepatoblasts in mouse liver development, using a knock-in allele of Hnf4a/laacZ This transgene randomly undergoes a low frequency of recombination that generates a functional lacZ gene that produces β-galactosidase in tissues in which Hnf4a is expressed. Two types of β-galactosidase-positive clones were found. Most have undergone three to eight cell divisions and result from independent events (Luria-Delbrück fluctuation test); we calculate that they arose between E8.5 and E13.5. A second class was mega-clones derived from early endoderm progenitors, generating many descendants. Some originated from multi-potential founder cells, with labeled cells in the liver, pancreas and/or intestine. A few mega-clones populate only one side of the liver, indicating hepatic cell chirality. The patterns of labeled cells indicate cohesive and often oriented growth, notably in broad radial stripes, potentially implicated in the formation of liver lobes. This retrospective clonal analysis gives novel insights into clonal origins, cell behavior of progenitors and distinct properties of endoderm cells that underlie the formation and morphogenesis of the liver.

  5. BACTERIAL BIOFILM FORMATION VERSUS MAMMALIAN CELL GROWTH ON TITANIUM-BASED MONO- AND BI-FUNCTIONAL COATINGS

    NARCIS (Netherlands)

    Subbiahdoss, Guruprakash; Pidhatika, Bidhari; Coullerez, Geraldine; Charnley, Mirren; Kuijer, Roel; van der Mei, Henny C.; Textor, Marcus; Busscher, Henk J.

    2010-01-01

    Biomaterials-associated-infections (BAI) are serious complications in modern medicine. Although non-adhesive coatings, like polymer-brush coatings, have been shown to prevent bacterial adhesion, they do not support cell growth. Bi-functional coatings are supposed to prevent biofilm formation while s

  6. Lentiviral vector-mediated siRNA knockdown of SR-PSOX inhibits foam cell formation in vitro

    Institute of Scientific and Technical Information of China (English)

    Li ZHANG; Hou-jia LIU; Tie-jun LI; Yang YANG; Xian-ling GUO; Meng-chao WU; Yao-cheng RUI; Li-xin WEI

    2008-01-01

    Aim: To investigate the expression of scavenger receptor that binds phosphatidylserine and oxidized lipoprotein (SR-PSOX)/CXC chemokine ligand 16 (CXCL16) in the human monocyte-derived cell line THP-1, and the effect of lentiviral vectors for the stable delivery of SR-PSOX/CXCL16 short hairpin RNA on foam cell formation. Methods: A lentiviral expression vector containing enhanced green fluorescence protein (GFP) and SR-PSOX small interfering RNA (siRNA) (Lenti-SR-PSOXsi), or the control siRNA (Lenti-NC) gene was constructed. A human monocyte-derived cell line THP-1 was transfected with a different multiplicity of infection (MOI) of Lenti-SR-PSOXsi or Lenti-NC, and cultured to obtain stably-transfected THP-1KD and THP-1NC cells. After incubation with oxidatively-modified, low-density lipoprotein (Ox-LDL), the expression of SR-PSOX/CXCL16 mRNA was determined by real-time PCR. The expression of the SR-PSOX/CXCL16 protein was detected by flow cytometry analysis. The effect of Lenti-SR-PSOXsi on foam cell formation was assessed by Oil red O-stain analysis. Results: Ox-LDL increased the expres-sion of SR-PSOX/CXCL 16 mRNA in a time- and dose-dependent manner in THP-1 cells. Four days after transfection with Lenti-SR-PSOXsi (MOI: 100), the percent-age of GFP expression cells was over 89.3%. The expression of the SR-PSOX/ CXCL 16 mRNA and protein in THP- 1KD cells significantly decreased compared with the parent cells, even the THP-1KD cells stimulated with 40 mg/L Ox-LDL. Ox-LDL uptake experiments in THP-1- and THP- 1KD-derived macrophages indi-cated that SR-PSOX/CXCL16 deficiency decreased the development of macroph-age-derived foam cell formation. Conclusion: The above data showed that SR-PSOX siRNA delivered by using lentiviral vectors in THP-1 cells was a powerful tool for studying the effect of SR-PSOX, and decreased the expression of the SR-PSOX gene by inhibiting macrophage-derived foam cell formation.

  7. A Fuzzy Programming approach for formation of Virtual Cells under dynamic and uncertain conditions

    Directory of Open Access Journals (Sweden)

    R.Jayachitra,

    2010-06-01

    Full Text Available Inspired by principles and advantages of the group technology (GT philosophy, part family formation for a virtual Cellular Manufacturing System (VCMS using Fuzzy logic is designed for dynamic and uncertain conditions. In real manufacturing systems, the input parameters such as part demand and the capacity are fuzzy in nature. In such cases, the fluctuations in part demand and the availability of manufacturing facilities in each period can be regarded as fuzzy. In a dynamic environment, the planning horizon can be divided into smaller time periods where each period and/or each part has different product mix and demand. A mathematical model for virtual cellular manufacturing system as binary-integer programming is proposed to minimize the total costs consisting of fixed machine costs, variable costs of all machines and the logical group movement costs. To verify the behavior of the proposed model, a comprehensive example is solved by a branch- and-bound (B&B method with the LINGO 12.0 software and the virtual cells(VC are formed by defuzzification using maximizing decision level λ (lambda-cut and the computational results are reported and compared with simulated annealing algorithm and rank order clustering algorithm .

  8. Toosendanin interferes with pore formation of botulinum toxin type A in PC12 cell membrane

    Institute of Scientific and Technical Information of China (English)

    Mu-feng LI; Yu-liang SHI

    2006-01-01

    Aim: Botulinum neurotoxins (BoNT) abort the process of neurotransmitter release at presynaptic motor nerve terminals, causing muscle paralysis. The ability of botulinum toxin to produce its effect is dependent on the ability of the light chain to cleave the SNARE proteins associated with transmitter release. Translocation of the light chain protease through the heavy chain-formed channel is a pivotal step in the intoxication process. Toosendanin (TSN), a triterpenoid derivative extracted from a Chinese traditional medicine, has been demonstrated to be an effective cure for experimental botulism. This study was designed to explore the antibotulismic mechanisms of toosendanin. Methods: The inside-out singlechannel recording patch-clamp technique was used to record the BoNT/A-induced currents in the presence and absence of TSN. Results: Channel formation was delayed and the sizes of the channels were reduced in the TSN-treated PC12cell membrane. Conclusion: The antibotulismic effect of TSN might occur via interference with toxin translocation.

  9. Highly oriented polycrystalline Cu2O film formation using RF magnetron sputtering deposition for solar cells

    Science.gov (United States)

    Noda, S.; Shima, H.; Akinaga, H.

    2014-02-01

    Room temperature sputtering deposition and re-crystallization of the deposited thin films by rapid thermal annealing have been evaluating in detail as a formation method of Cu2O active layer for solar cells, which minimize thermal budget in fabrication processes. Single phase polycrystalline Cu2O films were obtained by a magnetron rf sputtering deposition and its crystallinity and electrical characteristics were controlled by the annealing. Hall mobility was improved up to 17 cm2V-1s-1 by the annealing at 600°C for 30s. Since this value was smaller than 47 cm2V-1s-1 of the film deposited under thermal equilibrium state using pulsed laser deposition at 600°C, some contrivances were necessary to compensate the deficiency. It was understood that the sputter-deposited Cu2O films on (111)-oriented Pt films were strongly oriented to (111) face also by the self-assembly and the crystallinity was improved by the annealing preserving its orientation. The sputter-deposited film quality was expected to become equivalent to the pulsed laser deposition film from the results of X-ray diffractometry and photoluminescence.

  10. Oxidatively generated damage to the guanine moiety of DNA: Mechanistic aspects and formation in cells

    International Nuclear Information System (INIS)

    Nuclear DNA and other molecules in living systems are continuously exposed to endogenously generated oxygen species. Such species range from the unreactive superoxide radical (O2.-)-the precursor of hydrogen peroxide (H2O2)-to the highly reactive hydroxyl radical (OH.). Exogenous chemical and physical agents, such as ionizing radiation and the UVA component of solar light, can also oxidatively damage both the bases and the 2-deoxyribose moieties of cellular DNA. Over the last two decades, researchers have made major progress in understanding the oxidation degradation pathways of DNA that are most likely to occur from either oxidative metabolism or exposure to various exogenous agents. In the first part of this Account, we describe the mechanistic features of one-electron oxidation reactions of the guanine base in isolated DNA and related model compounds. These reactions illustrate the complexity of the various degradation pathways involved. Then, we briefly survey the analytical methods that can detect low amounts of oxidized bases and nucleosides in cells as they are formed. Recent data on the formation of oxidized guanine residues in cellular DNA following exposure to UVA light, ionizing radiation, and high-intensity UV pulses are also provided. We discuss these chemical reactions in the context of (OH.) radical, singlet oxygen, and two-quantum photoionization processes. (authors)

  11. Modelling plastic deformation in BCC metals: Dynamic recovery and cell formation effects

    Energy Technology Data Exchange (ETDEWEB)

    Galindo-Nava, E.I. [Department of Materials Science and Metallurgy, Pembroke Street, CB2 3QZ, University of Cambridge, Cambridge (United Kingdom); Department of Materials Science and Engineering, Mekelweg 2 2628 CD, Delft University of Technology, Delft (Netherlands); Rivera-Diaz-del-Castillo, P.E.J., E-mail: pejr2@cam.ac.uk [Department of Materials Science and Metallurgy, Pembroke Street, CB2 3QZ, University of Cambridge, Cambridge (United Kingdom)

    2012-12-15

    A recently developed model for describing plasticity in FCC metals (E.I., Galindo-Nava, P.E.J., Rivera-Diaz-del-Castillo, Mater. Sci. Eng. A 543 (2012) 110-116; E.I. Galindo-Nava, P.E.J. Rivera-Diaz-del-Castillo, Acta Mater. 60 (2012) 4370-4378) has now been applied to BCC. The core of the theory is the thermostatistical description of dislocation annihilation paths, which determines the dynamic recovery rate of the material. Input to this is the energy for the formation, migration and ordering of dislocation paths; the latter term corresponds to the statistical entropy which features strongly on the solution. The distinctions between FCC and BCC stem primarily from the possible directions and planes for dislocation slip and cross-slip, as well as from the presence of the kink-pair mechanism for dislocation migration in BCC, which are incorporated to the mathematical formulation of the model. The theory is unique in describing the stress-strain response for pure iron, molybdenum, tantalum, vanadium and tungsten employing physical parameters as input; the description is made for wide ranges of temperature and strain rate. Additionally, succinct equations to predict dislocation cell size variation with strain, strain rate and temperature are provided and validated for pure iron.

  12. In situ formation of graphene layers on graphite surfaces for efficient anodes of microbial fuel cells.

    Science.gov (United States)

    Tang, Jiahuan; Chen, Shanshan; Yuan, Yong; Cai, Xixi; Zhou, Shungui

    2015-09-15

    Graphene can be used to improve the performance of the anode in a microbial fuel cell (MFC) due to its good biocompatibility, high electrical conductivity and large surface area. However, the chemical production and modification of the graphene on the anode are environmentally hazardous because of the use of various harmful chemicals. This study reports a novel method based on the electrochemical exfoliation of a graphite plate (GP) for the in situ formation of graphene layers on the surface of a graphite electrode. When the resultant graphene-layer-based graphite plate electrode (GL/GP) was used as an anode in an MFC, a maximum power density of 0.67 ± 0.034 W/m(2) was achieved. This value corresponds to 1.72-, 1.56- and 1.26-times the maximum power densities of the original GP, exfoliated-graphene-modified GP (EG/GP) and chemically-reduced-graphene-modified GP (rGO/GP) anodes, respectively. Electrochemical measurements revealed that the high performance of the GL/GP anode was attributable to its macroporous structure, improved electron transfer and high electrochemical capacitance. The results demonstrated that the proposed method is a facile and environmentally friendly synthesis technique for the fabrication of high-performance graphene-based electrodes for use in microbial energy harvesting. PMID:25950933

  13. Fullerene mixing effect on carrier formation in bulk-hetero organic solar cell

    Science.gov (United States)

    Moritomo, Yutaka; Yasuda, Takeshi; Yonezawa, Kouhei; Sakurai, Takeaki; Takeichi, Yasuo; Suga, Hiroki; Takahashi, Yoshio; Inami, Nobuyuki; Mase, Kazuhiko; Ono, Kanta

    2015-03-01

    Organic solar cells (OSCs) with a bulk-heterojunction (BHJ) are promising energy conversion devices, because they are flexible and environmental-friendly, and can be fabricated by low-cost roll-to-roll process. Here, we systematically investigated the interrelations between photovoltaic properties and the domain morphology of the active layer in OSCs based on films of poly-(9,9-dioctylfluorene-co-bithiophene) (F8T2)/[6,6]-phenyl C71-butyric acid methyl ester (PC71BM) blend annealed at various temperatures (Tan). The scanning transmission X-ray microscopy (STXM) revealed that fullerene mixing (ΦFullerene) in the polymer matrix decreases with increase in Tan while the domain size (L) is nearly independent of Tan. The TEM-S mapping image suggests that the polymer matrix consist of polymer clusters of several nm and fullerene. We found that the charge formation efficiency (ΦCF), internal quantum efficiency (ΦIQ), and power conversion efficiency (PCE) are dominantly determined by ΦFullerene. We interpreted these observations in terms of the polymer clusters within the polymer matrix.

  14. Suppression of Hydroxycinnamate Network Formation in Cell Walls of Rice Shoots Grown under Microgravity Conditions in Space.

    Directory of Open Access Journals (Sweden)

    Kazuyuki Wakabayashi

    Full Text Available Network structures created by hydroxycinnamate cross-links within the cell wall architecture of gramineous plants make the cell wall resistant to the gravitational force of the earth. In this study, the effects of microgravity on the formation of cell wall-bound hydroxycinnamates were examined using etiolated rice shoots simultaneously grown under artificial 1 g and microgravity conditions in the Cell Biology Experiment Facility on the International Space Station. Measurement of the mechanical properties of cell walls showed that shoot cell walls became stiff during the growth period and that microgravity suppressed this stiffening. Amounts of cell wall polysaccharides, cell wall-bound phenolic acids, and lignin in rice shoots increased as the shoot grew. Microgravity did not influence changes in the amounts of cell wall polysaccharides or phenolic acid monomers such as ferulic acid (FA and p-coumaric acid, but it suppressed increases in diferulic acid (DFA isomers and lignin. Activities of the enzymes phenylalanine ammonia-lyase (PAL and cell wall-bound peroxidase (CW-PRX in shoots also increased as the shoot grew. PAL activity in microgravity-grown shoots was almost comparable to that in artificial 1 g-grown shoots, while CW-PRX activity increased less in microgravity-grown shoots than in artificial 1 g-grown shoots. Furthermore, the increases in expression levels of some class III peroxidase genes were reduced under microgravity conditions. These results suggest that a microgravity environment modifies the expression levels of certain class III peroxidase genes in rice shoots, that the resultant reduction of CW-PRX activity may be involved in suppressing DFA formation and lignin polymerization, and that this suppression may cause a decrease in cross-linkages within the cell wall architecture. The reduction in intra-network structures may contribute to keeping the cell wall loose under microgravity conditions.

  15. Formation of Nano scale Bio imprints of Muscle Cells Using UV-Cured Spin-Coated Polymers

    International Nuclear Information System (INIS)

    We report a nano scale replication method suitable for biological specimens that has potential in single cell studies and in formation of 3D biocompatible scaffolds. Earlier studies using a heat-curable polydimethylsiloxane (PDMS) or a UV-curable elastomer introduced Bio imprint replication to facilitate cell imaging. However, the replicating conditions for thermal polymerization are known to cause cell dehydration during curing. In this study, a UV-cured methacrylate copolymer was developed for use in creating replicas of living cells and was tested on rat muscle cells. Bio imprints of muscle cells were formed by spin coating under UV irradiation. The polymer replicas were then separated from the muscle cells and were analyzed under an Atomic Force Microscope (AFM), in tapping mode, because it has low tip-sample forces and thus will not destroy the fine structures of the imprint. The new polymer is biocompatible with higher replication resolution and has a faster curing process than other types of silicon-based organic polymers such as PDMS. High resolution images of the muscle cell imprints showed the micro-and nano structures of the muscle cells, including cellular fibers and structures within the cell membranes. The AFM is able to image features at nano scale resolution with the potential for recognizing abnormalities on cell membranes at early stages of disease progression.

  16. Gastric LTi cells promote lymphoid follicle formation but are limited by IRAK-M and do not alter microbial growth.

    Science.gov (United States)

    Shiu, J; Piazuelo, M B; Ding, H; Czinn, S J; Drakes, M L; Banerjee, A; Basappa, N; Kobayashi, K S; Fricke, W F; Blanchard, T G

    2015-09-01

    Lymphoid tissue inducer (LTi) cells are activated by accessory cell IL-23, and promote lymphoid tissue genesis and antibacterial peptide production by the mucosal epithelium. We investigated the role of LTi cells in the gastric mucosa in the context of microbial infection. Mice deficient in IRAK-M, a negative regulator of TLR signaling, were investigated for increased LTi cell activity, and antibody mediated LTi cell depletion was used to analyze LTi cell dependent antimicrobial activity. H. pylori infected IRAK-M deficient mice developed increased gastric IL-17 and lymphoid follicles compared to wild type mice. LTi cells were present in naive and infected mice, with increased numbers in IRAK-M deficient mice by two weeks. Helicobacter and Candida infection of LTi cell depleted rag1(-/-) mice demonstrated LTi-dependent increases in calprotectin but not RegIII proteins. However, pathogen and commensal microbiota populations remained unchanged in the presence or absence of LTi cell function. These data demonstrate LTi cells are present in the stomach and promote lymphoid follicle formation in response to infection, but are limited by IRAK-M expression. Additionally, LTi cell mediated antimicrobial peptide production at the gastric epithelium is less efficacious at protecting against microbial pathogens than has been reported for other tissues.

  17. CD4+ T Cell Help Guides Formation of CD103+ Lung-Resident Memory CD8+ T Cells during Influenza Viral Infection

    Science.gov (United States)

    Laidlaw, Brian J.; Zhang, Nianzhi; Marshall, Heather D.; Staron, Mathew M.; Guan, Tianxia; Hu, Yinghong; Cauley, Linda S.; Craft, Joe; Kaech, Susan M.

    2014-01-01

    SUMMARY Tissue-resident memory T (Trm) cells provide enhanced protection against infection at mucosal sites. Here we found that CD4+ T cells are important for the formation of functional lung-resident CD8+ T cells after influenza virus infection. In the absence of CD4+ T cells, CD8+ T cells displayed reduced expression of CD103 (Itgae), were mislocalized away from airway epithelia, and demonstrated an impaired ability to recruit CD8+ T cells to the lung air-ways upon heterosubtypic challenge. CD4+ T cell-derived interferon-γ was necessary for generating lung-resident CD103+ CD8+ Trm CD8 T cells. Furthermore, expression of the transcription factor T-bet was increased in “unhelped” lung Trm cells, and a reduction in T-bet rescued CD103 expression in the absence of CD4+ T cell help. Thus, CD4+ T cell-dependent signals are important to limit expression of T-bet and allow for the development of CD103+ CD8+ Trm cells in the lung airways following respiratory infection. PMID:25308332

  18. A channel connecting the mother cell and forespore during bacterial endospore formation

    OpenAIRE

    Meisner, Jeffrey; Xin WANG; Serrano, Monica; Henriques, Adriano O.; Moran, Charles P.

    2008-01-01

    At an early stage during Bacillus subtilis endospore development the bacterium divides asymmetrically to produce two daughter cells. The smaller cell (forespore) differentiates into the endospore, while the larger cell (mother cell) becomes a terminally differentiated cell that nurtures the developing forespore. During development the mother cell engulfs the forespore to produce a protoplast, surrounded by two bilayer membranes, which separate it from the cytoplasm of the mother cell. The act...

  19. Isothermal microcalorimetry as a tool to study solid-electrolyte interphase formation in lithium-ion cells.

    Science.gov (United States)

    Hall, David S; Glazier, Stephen L; Dahn, J R

    2016-04-28

    Isothermal microcalorimetry can be used in conjunction with electrochemical measurements to study solid-electrolyte interphase (SEI) formation reactions as they occur in a Li-ion cell. The heat flow was measured in wound cells that contained no electrolyte additives and in cells prepared with four additives that are known to produce an SEI at the negative electrode surface: vinylene carbonate (VC), fluoroethylene carbonate (FEC), pyridine boron trifluoride (PBF), and prop-1-ene-1,3-sultone (PES). For VC, two distinct features in the differential capacity (dQ/dV vs. Q) plot that align with overlapping peaks in the heat flow plot do not agree with a simple one-electron reduction followed by anionic polymerization. For FEC, three distinct differential capacity and calorimetric peaks are observed. Heat flow measurements at multiple PBF concentrations show that PBF reduction does not significantly affect the reduction of EC at higher cell voltage. The total heat flow during SEI formation in PBF- and PES-containing cells match the calculated energies in recently published reaction pathways. It is concluded that IMC may be used to study the underlying chemistry of SEI formation, especially when paired with computational studies. PMID:27056253

  20. Failure-to-thrive syndrome associated with tumor formation by Madin-Darby canine kidney cells in newborn nude mice.

    Science.gov (United States)

    Brinster, Lauren R; Omeir, Romelda L; Foseh, Gideon S; Macauley, Juliete N; Snoy, Philip J; Beren, Joel J; Teferedegne, Belete; Peden, Keith; Lewis, Andrew M

    2013-08-01

    Tumors that formed in newborn nude mice that were inoculated with 10(7) Madin-Darby canine kidney (MDCK) cells were associated with a failure-to-thrive (FTT) syndrome consisting of growth retardation, lethargy, weakness, and dehydration. Scoliosis developed in 41% of affected pups. Pups were symptomatic by week 2; severely affected pups became moribund and required euthanasia within 3 to 4 wk. Mice with FTT were classified into categories of mild, moderate, and severe disease by comparing their weight with that of age-matched normal nude mice. The MDCK-induced tumors were adenocarcinomas that invaded adjacent muscle, connective tissue, and bone; 6 of the 26 pups examined had lung metastases. The induction of FTT did not correlate with cell-line aggressiveness as estimated by histopathology or the efficiency of tumor formation (tumor-forming dose 50% endpoint range = 10(2.8) to 10(7.5)); however, tumor invasion of the paravertebral muscles likely contributed to the scoliosis noted. In contrast to the effect of MDCK cells, tumor formation observed in newborn mice inoculated with highly tumorigenic, human-tumor-derived cell lines was not associated with FTT development. We suggest that tumor formation and FTT are characteristics of these MDCK cell inocula and that FTT represents a new syndrome that may be similar to the cachexia that develops in humans with cancer or other diseases. PMID:24209967

  1. 2', 3'-Cyclic nucleotide 3'-phosphodiesterase cells derived from transplanted marrow stromal cells and host tissue contribute to perineurial compartment formation in injured rat spinal cord.

    Science.gov (United States)

    Cao, Qiong; Ding, Peng; Lu, Jia; Dheen, S Thameem; Moochhala, Shabbir; Ling, Eng-Ang

    2007-01-01

    Transdifferentiation of transplanted marrow stromal cells (MSCs) and reactive changes of glial cells in a completely transected rat spinal cord were examined. Marrow stromal cells exhibited 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) at the plasma membrane and this has allowed their identification after transplantation by immunoelectron microscopy. In the control rats, the lesion site showed activated microglia/neural macrophages and some elongated cells, whose cytoplasm was immunoreactive for CNP. Cells designated as CNP1 and apparently host-derived expressed CXCR4. In experimental rats receiving MSCs transplantation, CNP1 cells were increased noticeably. This was coupled with the occurrence of a different subset of CNP cells whose plasma membrane was CNP-immunoreactive and expressed CXCR4. These cells, designated as CNP2, enclosed both myelinated and unmyelinated neurites thus assuming a spatial configuration resembling that of Schwann cells. A remarkable feature was the extensive ramifications of CNP1 cells with long filopodia processes delineating the CNP2 cells and their associated neurites, forming many perineurial-like compartments. Present results have shown that CNP2 cells considered to be MSCs-derived can transform into cells resembling Schwann cells based on their spatial relation with the regenerating nerve fibers, whereas the CNP1 glial cells participate in formation of perineurial compartments, probably serving as conduits to guide the nerve fiber growth. The chemotactic migration of CNP cells either derived from host tissue or MSCs bearing CXCR4 may be attracted by stromal derived factor-1alpha (SDF-1alpha) produced locally. The coordinated cellular interaction between transplanted MSCs and local glial cells may promote the growth of nerve fibers through the lesion site. PMID:17061258

  2. The Ca2+ Antagonizing Effect of Chinese Cobra Venom Factor on Formation of Macrophage-derived Foam Cells

    Institute of Scientific and Technical Information of China (English)

    谭健苗; 杨向东; 姜志胜; 李亮

    2007-01-01

    Purpose CCVF was isolated from Chinese cobra (Naja naja) venom, its Ca2+ antagonizing effect on formation of macrophage-derived foam cells was explored in these studies. Methods Foam cell models were induced with C57BL/6J mouse peritoneal macrophages incubated in 10mg/L oxidized low density lipoprotein (OLDL), and their intracellular Ca2+ levels influenced both slowly and transiently by CCVF were determined with the technique of Ca2+ fluorescent indicator. Results The intracellular Ca2+ level with the macrophages incubated in 10mg/L OLDL and 10mg/L CCVF was 40.2% of the macrophages incubated in 10mg/L OLDL (P<0.05); While the transient influence of CCVF on the intracellular Ca2+ levels were not significant. Conclusion CCVF exerted a long-lasting antagonizing role on the enhancement of intracellular Ca2+ levels, thus inhibited the formation of macrophage-derived foam cell.

  3. Bleb formation is induced by alkaline but not acidic pH in estrogen receptor silenced breast cancer cells.

    Science.gov (United States)

    Khajah, Maitham A; Mathew, Princy M; Alam-Eldin, Nada S; Luqmani, Yunus A

    2015-04-01

    De novo and acquired resistance to endocrine-based therapies in breast cancer occurs in parallel with epithelial to mesenchymal transition (EMT), which is associated with enhanced proliferative and metastatic potential, and poor clinical outcome. We have established several endocrine insensitive breast cancer lines by shRNA-induced depletion of estrogen receptor (ER) by transfection of MCF7 cells. All of these exhibit EMT. We have previously reported that brief exposure of specifically ER- breast cancer cells, to extracellular alkaline pH, results in cell rounding and segregation, and leads to enhanced invasive potential. In this study we describe more detailed morphological changes and compare these with cell exposure to acidic pH. Morphological changes and localization of various molecules critical for cell adhesion and motility, associated with pH effects, were assessed by live cell microscopy, electron microscopy, and immunofluorescence. Exposure of either ER- or ER+ breast cancer cells to extracellular acidic pH did not induce significant changes in morphological appearance. Conversely, brief exposure of specifically ER silenced cells, to alkaline pH, resulted in cell contractolation and formation of bleb-like actin-rich structures which were evenly distributed on the outer membrane. Integrin α2, FAK, and JAM-1 were found in the cytoplasm streaming into the newly formed blebs. These blebs appear to be related to cell polarity and movement. Pre-treatment with cytochalasin-D or inhibitors of Rho or MLCK prevented both contractolation and bleb formation. Our data suggest that the effect of pH on the microenvironment of endocrine resistant breast cancer cells needs to be more extensively investigated. Alkaline, rather than acidic pH, appears to induce dramatic morphological changes, and enhances their invasive capabilities, through re-organization of cortical actin. PMID:25672508

  4. Osteocytes, not Osteoblasts or Lining Cells, are the Main Source of the RANKL Required for Osteoclast Formation in Remodeling Bone.

    Directory of Open Access Journals (Sweden)

    Jinhu Xiong

    Full Text Available The cytokine receptor activator of nuclear factor kappa B ligand (RANKL, encoded by the Tnfsf11 gene, is essential for osteoclastogenesis and previous studies have shown that deletion of the Tnfsf11 gene using a Dmp1-Cre transgene reduces osteoclast formation in cancellous bone by more than 70%. However, the Dmp1-Cre transgene used in those studies leads to recombination in osteocytes, osteoblasts, and lining cells making it unclear whether one or more of these cell types produce the RANKL required for osteoclast formation in cancellous bone. Because osteoblasts, osteocytes, and lining cells have distinct locations and functions, distinguishing which of these cell types are sources of RANKL is essential for understanding the orchestration of bone remodeling. To distinguish between these possibilities, we have now created transgenic mice expressing the Cre recombinase under the control of regulatory elements of the Sost gene, which is expressed in osteocytes but not osteoblasts or lining cells in murine bone. Activity of the Sost-Cre transgene in osteocytes, but not osteoblast or lining cells, was confirmed by crossing Sost-Cre transgenic mice with tdTomato and R26R Cre-reporter mice, which express tdTomato fluorescent protein or LacZ, respectively, only in cells expressing the Cre recombinase or their descendants. Deletion of the Tnfsf11 gene in Sost-Cre mice led to a threefold decrease in osteoclast number in cancellous bone and increased cancellous bone mass, mimicking the skeletal phenotype of mice in which the Tnfsf11 gene was deleted using the Dmp1-Cre transgene. These results demonstrate that osteocytes, not osteoblasts or lining cells, are the main source of the RANKL required for osteoclast formation in remodeling cancellous bone.

  5. IDENTIFYING GENES CONTROLLING FERULATE CROSS-LINKING FORMATION IN GRASS CELL WALLS

    Energy Technology Data Exchange (ETDEWEB)

    de O Buanafina, Marcia Maria

    2013-10-16

    formation or genes encoding transcription factors that control feruloylation. So it will require further investigations to confirm if we have a mutation on the ferulloyltransferase gene(s). We have also identified severe phenotypes which showed a significant change in the level of cell wall ferulates and sugars and have not survived. As this genotype did not reach flowering stage there was no seed production and so further analysis could not be done. 3. Candidate Gene Approach: Because of the likely long time expected to generate and identify candidate with mutation(s) on the feruloyltransferase gene, from our screening, we have in addition taken a bioinformatics approach in order to try to identify candidates gene(s) involved in feruloylation. Homologues of the rice feruloyl transferase genes belonging to Pfam PF02458 family were identified in Brachypodium distachyon by blasting EST sequences of putative rice arabinoxylan feruloyl transferase genes against Brachypodium and homologous sequences identified were tested for their expression level in Brachypodium. Sequences of the two Brachypodium genes, which showed highest expression and similarity to rice sequences, were used to design primers for construction of RNAi and over-expression vectors. These were transformed into Brachypodium using Agrobacterium transformation and plants generated have been analyzed for levels of cell wall ferulates and diferulates over generations T0 to T2 or T3. Our data shows a significant reduction if ferulates monomers and dimers from plants generated from RNAi::BdAT2 over 2-3 generations indicating that this gene might be a positive candidate for feruloylation in Brachypodium. However when BdAT2 was up regulated there was not much increase in the level of ferulates as would be expected. This lack of effect on the level of cell wall ferulates could be due to the CaMV::35S promoter used to drive the expression of the putative BdAT2 gene. We have shown previously that Aspergillus FAEA

  6. The concentration of intracellular nickel in Haemophilus influenzae is linked to its surface properties and cell-cell aggregation and biofilm formation.

    Science.gov (United States)

    Ng, Jiaqi; Kidd, Stephen P

    2013-04-01

    Of the known proteins which use nickel as a co-factor, Haemophilus influenzae contains only urease and glyoxalase I (gloA). We have recently reported that this pathogen harbours a unique nickel uptake system (nikKLMQO-nimR). Unusually, the disruption of the nickel uptake system (nikQ or nimR mutants) resulted in cells that aggregated and formed an increased biofilm compared to the wild type cells. Using a gloA mutant strain and urease-specific inhibitor we showed that this phenotype is not due to the loss-of-function of these enzymes. By generating H. influenzae "resting cells" which are enzymatically inactive but maintain their structural integrity we have shown that the cell aggregation in the nikQ/nimR mutants is not due to the loss of enzymatic function. The nikQ mutant was unable to accumulate nickel but the addition of excess nickel did restore intracellular nickel levels and this resulted in the nikQ mutant returning to the wild type "free-living" phenotype; cells with no aggregation and no biofilm formation. We used a range of techniques which showed that the nikQ mutant possesses changes to its cell surface properties. The mutant was more negatively charged than wild type cells as well as being more hydrophobic. Analysis of the outer membrane constituents showed that there were molecular differences. Although the nikQ mutant appears to grow the same as its wild type cell we have shown that there is a change in the "lifestyle" of these nickel limited cells and this induces changes to the surface of the cell to promote cell-cell aggregation and biofilm formation. PMID:23499478

  7. Formation of post-confluence structure in human parotid gland acinar cells on PLGA through regulation of E-cadherin.

    Science.gov (United States)

    Chan, Yen-Hui; Huang, Tsung-Wei; Chou, Ya-Shuan; Hsu, Sheng-Hao; Su, Wei-Fang; Lou, Pei-Jen; Young, Tai-Horng

    2012-01-01

    As a potential solution for patients to retrieve their lost salivary gland functions, tissue engineering of an auto-secretory device is profoundly needed. Under serum-free environment, primary human parotid gland acinar (PGAC) cells can be obtained. After reaching confluence, PGAC cells spontaneously form three-dimension (3D) cell aggregations, termed post-confluence structure (PCS), and change their behaviors. Poly (lactic-co-glycolic acid) (PLGA) has been widely used in the field of biomedical applications because of its biodegradable properties for desired functions. Nonetheless, the role of PLGA in facilitating PGAC cells to form PCS has seldom been explored to recover epithelial characteristics. In this study, PGAC cells were found to have a greater tendency to form PCS on PLGA than on tissue culture polystyrene (TCPS). By tracing cell migration paths and modulating E-cadherin activity with specific inhibitor or antibody, we demonstrated that the static force of homophilic interaction on surfaces of individual cells, but not the dynamics of cell migration, played a more important role in PCS formation. Thus, PLGA was successfully confirmed to support PGAC cells to form more PCS through the effects on enhancing E-cadherin expression, which is associated with FAK/ILK/Snail expression in PGAC cells. This result indicates that selective appropriate biomaterials may be potentially useful in generating 3D PCS on two-dimension (2D) substrate without fabricating a complex 3D scaffold.

  8. Rac function is crucial for cell migration but is not required for spreading and focal adhesion formation

    DEFF Research Database (Denmark)

    Steffen, Anika; Ladwein, Markus; Dimchev, Georgi A;

    2013-01-01

    Cell migration is commonly accompanied by protrusion of membrane ruffles and lamellipodia. In two-dimensional migration, protrusion of these thin sheets of cytoplasm is considered relevant to both exploration of new space and initiation of nascent adhesion to the substratum. Lamellipodium formation...... restored by expression of either Rac subfamily member, but not by Cdc42 or RhoG. Cells deficient in Rac showed strong reduction in wound closure and random cell migration and a notable loss of sensitivity to a chemotactic gradient. Despite these defects, Rac-deficient cells were able to spread, formed...... deficiency increased the mobility of different components in focal adhesions, potentially explaining how Rac - although not essential - can contribute to focal adhesion assembly. Together, our data demonstrate that Rac signaling is essential for lamellipodium protrusion and for efficient cell migration, but...

  9. Predicting the distribution of spiral waves from cell properties in a developmental-path model of Dictyostelium pattern formation.

    Directory of Open Access Journals (Sweden)

    Daniel Geberth

    2009-07-01

    Full Text Available The slime mold Dictyostelium discoideum is one of the model systems of biological pattern formation. One of the most successful answers to the challenge of establishing a spiral wave pattern in a colony of homogeneously distributed D. discoideum cells has been the suggestion of a developmental path the cells follow (Lauzeral and coworkers. This is a well-defined change in properties each cell undergoes on a longer time scale than the typical dynamics of the cell. Here we show that this concept leads to an inhomogeneous and systematic spatial distribution of spiral waves, which can be predicted from the distribution of cells on the developmental path. We propose specific experiments for checking whether such systematics are also found in data and thus, indirectly, provide evidence of a developmental path.

  10. Acemannan, an extracted product from Aloe vera, stimulates dental pulp cell proliferation, differentiation, mineralization, and dentin formation.

    Science.gov (United States)

    Jittapiromsak, Nawaporn; Sahawat, Dusida; Banlunara, Wijit; Sangvanich, Polkit; Thunyakitpisal, Pasutha

    2010-06-01

    This study investigated the effect of acemannan (Aloe vera gel polysaccharide) on dentin formation. Primary human dental pulp cells were treated with acemannan. New DNA synthesis, bone morphogenetic protein-2, alkaline phosphatase activity, dentin sialoprotein expression, and mineralization were determined by [(3)H]-thymidine incorporation, enzyme-linked immunosorbent assay, biochemical assay, western blotting, and Alizarin Red staining, respectively. Then the upper first molars of 24 male Sprague Dawley rats were intentionally exposed and capped with either acemannan or calcium hydroxide. At day 28, the teeth were histopathologically examined and evaluated for the degree of inflammation, dentin bridge formation, and pulp tissue organization. The results revealed that acemannan significantly increased pulp cell proliferation, bone morphogenetic protein-2, alkaline phosphatase activity, dentin sialoprotein expression, and mineralization, compared with the untreated group. The acemannan-treated group also exhibited a complete homogeneous calcified dentin bridge and good pulp tissue organization, whereas neither was detected in the calcium hydroxide-treated and sham groups. In the acemannan-treated group, either mild or no inflammation was found, whereas the other groups had various degrees of inflammation. The data suggest that acemannan promotes dentin formation by stimulating primary human dental pulp cell proliferation, differentiation, extracellular matrix formation, and mineralization. Acemannan also has pulpal biocompatibility and promotes soft tissue organization.

  11. Cell Yields of Vibrio succinogenes growing with formate and fumarate as sole carbon and energy sources in chemostat culture.

    Science.gov (United States)

    Mell, H; Bronder, M; Kröger, A

    1982-05-01

    Vibrio succinogenes which gains all the ATP by anaerobic electron transport phosphorylation, was grown in continuous culture on a defined medium with formate and fumarate as sole energy sources. The growth yield at infinite dilution rate (Ymax) was obtained by extrapolation from the growth yields measured at various dilution rates. With formate as the growth limiting substrate, Ymax was found as 14 g dry cells/mol formate. Under these conditions growth was limited by the rate of energy supply, because formate is used only as a catabolic substrate (Bronder et al. 1982). The YmaxATP calculated from the ATP requirement for cell synthesis was 18 g dry cells/mol ATP. This gives an ATP/2e ratio of 0.8. The ATP/2e ratio in vitro had been measured as 1 (Kröger and Winkler 1981). It is concluded that growing V. succinogenes gain at least 80% the stoichiometrically possible amount of ATP, when growth is limited by energy supply. PMID:7103661

  12. Silencing heme oxygenase-1 gene expression in retinal pigment epithelial cells inhibits proliferation, migration and tube formation of cocultured endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Wenjie [Ophthalmology Hospital, The First Affiliated Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001 (China); Zhang, Xiaomei, E-mail: zhangxm667@163.com [Ophthalmology Hospital, The First Affiliated Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001 (China); Lu, Hong [Ophthalmology Hospital, The First Affiliated Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001 (China); Matsukura, Makoto [Laboratory of Clinical Pharmacology and Therapeutics, Faculty of Pharmaceutical Sciences, Sojo University, 4-22-1 Ikeda, Kumamoto 860-0082 (Japan); Zhao, Jien; Shinohara, Makoto [Ashikita Institution for Developmental Disabilities, 2813 Oaza Ashikita, Ashikita-machi, Ashikita, Kumamoto 869-5461 (Japan)

    2013-05-10

    Highlights: •HO-1 is highly induced in RPE cells by hypoxia. •Inhibition of HO-1 activity and knockdown of HO-1 expression inhibit VEGF expression in RPE cells under hypoxia. •Knockdown of HO-1 in RPE cells inhibits angiogenesis of endothelial cells in vitro. -- Abstract: Heme oxygenase-1 (HO-1) plays an important role in the vasculature and in the angiogenesis of tumors, wounds and other environments. Retinal pigment epithelial (RPE) cells and choroidal endothelial cells (CECs) are the main cells involved in choroidal neovascularization (CNV), a process in which hypoxia plays an important role. Our aim was to evaluate the role of human RPE-cell HO-1 in the angiogenic activities of cocultured endothelial cells under hypoxia. Small interfering RNA (siRNA) for HO-1 was transfected into human RPE cell line ARPE-19, and zinc protoporphyrin (ZnPP) was used to inhibit HO-1 activity. Knockdown of HO-1 expression and inhibition of HO-1 activity resulted in potent reduction of the expression of vascular endothelial growth factor (VEGF) under hypoxia. Furthermore, knockdown of HO-1 suppressed the proliferation, migration and tube formation of cocultured endothelial cells. These findings indicated that HO-1 might have an angiogenic effect in CNV through modulation of VEGF expression and might be a potential target for treating CNV.

  13. Migration of microchimeric fetal cells into maternal circulation before placenta formation

    OpenAIRE

    Sunami, Rei; Komuro, Mayuko; Tagaya, Hikaru; Hirata, Shuji

    2010-01-01

    Fetal cell microchimerism is defined as the persistence of pluripotent fetal cells in the maternal body long after delivery. The exact process by which fetal cells cross the placental barrier and enter maternal circulation is still being investigated. We reported that fetal cells persist only in the maternal bone marrow and may give rise to subpopulations with the ability to differentiate into the tissue-specific mature cells within injured maternal organs. Moreover, most of the fetal cells e...

  14. Multifunctional Merkel cells: their roles in electromagnetic reception, finger-print formation, Reiki, epigenetic inheritance and hair form.

    Science.gov (United States)

    Irmak, M Kemal

    2010-08-01

    Merkel cells are located in glabrous and hairy skin and in some mucosa. They are characterized by dense-core secretory granules and cytoskeletal filaments. They are attached to neighboring keratinocytes by desmosomes and contain melanosomes similar to keratinocytes. They are excitable cells in close contact with sensory nerve endings but their function is still unclear. In this review, following roles are attributed for the first time to the Merkel cells: (1) melanosomes in Merkel cells may be involved in mammalian magnetoreception. In this model melanosome as a biological magnetite is connected by cytoskeletal filaments to mechanically gated ion channels embedded in the Merkel cell membrane. The movement of melanosome with the changing electromagnetic field may open ion channels directly producing a receptor potential that can be transmitted to brain via sensory neurons. (2) Merkel cells may be involved in finger-print formation: Merkel cells in glabrous skin are located at the base of the epidermal ridges the type of which defines the finger-print pattern. Finger-print formation starts at the 10th week of pregnancy after the arrival of Merkel cells. Keratinocyte proliferation and the buckling process observed in the basal layer of epidermis resulting in the epidermal ridges may be controlled and formed by Merkel cells. (3) Brain-Merkel cell connection is bi-directional and Merkel cells not only absorb but also radiate the electromagnetic frequencies. Hence, efferent aspects of the palmar and plantar Merkel nerve endings may form the basis of the biofield modalities such as Reiki, therapeutic touch and telekinesis. (4) Adaptive geographic variations such as skin color, craniofacial morphology and hair form result from interactions between environmental factors and epigenetic inheritance system. While environmental factors produce modifications in the body, they simultaneously induce epigenetic modifications in the oocytes and in this way adaptive changes could be

  15. Multifunctional Merkel cells: their roles in electromagnetic reception, finger-print formation, Reiki, epigenetic inheritance and hair form.

    Science.gov (United States)

    Irmak, M Kemal

    2010-08-01

    Merkel cells are located in glabrous and hairy skin and in some mucosa. They are characterized by dense-core secretory granules and cytoskeletal filaments. They are attached to neighboring keratinocytes by desmosomes and contain melanosomes similar to keratinocytes. They are excitable cells in close contact with sensory nerve endings but their function is still unclear. In this review, following roles are attributed for the first time to the Merkel cells: (1) melanosomes in Merkel cells may be involved in mammalian magnetoreception. In this model melanosome as a biological magnetite is connected by cytoskeletal filaments to mechanically gated ion channels embedded in the Merkel cell membrane. The movement of melanosome with the changing electromagnetic field may open ion channels directly producing a receptor potential that can be transmitted to brain via sensory neurons. (2) Merkel cells may be involved in finger-print formation: Merkel cells in glabrous skin are located at the base of the epidermal ridges the type of which defines the finger-print pattern. Finger-print formation starts at the 10th week of pregnancy after the arrival of Merkel cells. Keratinocyte proliferation and the buckling process observed in the basal layer of epidermis resulting in the epidermal ridges may be controlled and formed by Merkel cells. (3) Brain-Merkel cell connection is bi-directional and Merkel cells not only absorb but also radiate the electromagnetic frequencies. Hence, efferent aspects of the palmar and plantar Merkel nerve endings may form the basis of the biofield modalities such as Reiki, therapeutic touch and telekinesis. (4) Adaptive geographic variations such as skin color, craniofacial morphology and hair form result from interactions between environmental factors and epigenetic inheritance system. While environmental factors produce modifications in the body, they simultaneously induce epigenetic modifications in the oocytes and in this way adaptive changes could be

  16. Measurement of ethanol formation in single living cells of Chlamydomonas reinhardtii using synchrotron Fourier Transform Infrared spectromicroscopy

    Science.gov (United States)

    Goff, Kira L.; Quaroni, Luca; Pedersen, Tor; Wilson, Kenneth E.

    2010-02-01

    We demonstrate the capability of Fourier-Transform Infra-Red (FITR) spectroscopy to detect metabolite formation by the unicellular algae Chlamydomonas reinhardtii in solution. We show that using a synchrotron source in the microscopy configuration provides a sufficient s/n ratio to detect small molecular species accumulating at a single cell, allowing an increased sensitivity relative to measurements of bulk cultures. The formation of small molecular species, including ethanol and at least one carbonyl containing compound, can be detected with a time resolution of the order of one minute.

  17. Measurement of ethanol formation in single living cells of Chlamydomonas reinhardtii using synchrotron Fourier Transform Infrared spectromicroscopy

    International Nuclear Information System (INIS)

    We demonstrate the capability of Fourier-Transform Infra-Red (FITR) spectroscopy to detect metabolite formation by the unicellular algae Chlamydomonas reinhardtii in solution. We show that using a synchrotron source in the microscopy configuration provides a sufficient s/n ratio to detect small molecular species accumulating at a single cell, allowing an increased sensitivity relative to measurements of bulk cultures. The formation of small molecular species, including ethanol and at least one carbonyl containing compound, can be detected with a time resolution of the order of one minute.

  18. Electromagnetic particle-in-cell (PIC) method for modeling the formation of metal surface structures induced by femtosecond laser radiation

    Energy Technology Data Exchange (ETDEWEB)

    Djouder, M. [Laboratoire de Physique et Chimie Quantique, Université Mouloud Mammeri de Tizi-ouzou, BP 17 RP, 15000 Tizi-Ouzou (Algeria); Lamrous, O., E-mail: omarlamrous@mail.ummto.dz [Laboratoire de Physique et Chimie Quantique, Université Mouloud Mammeri de Tizi-ouzou, BP 17 RP, 15000 Tizi-Ouzou (Algeria); Mitiche, M.D. [Laboratoire de Physique et Chimie Quantique, Université Mouloud Mammeri de Tizi-ouzou, BP 17 RP, 15000 Tizi-Ouzou (Algeria); Itina, T.E. [Laboratoire Hubert Curien, UMR CNRS 5516/Université Jean Monnet, 18 rue de Professeur Benoît Lauras, 42000 Saint-Etienne (France); Zemirli, M. [Laboratoire de Physique et Chimie Quantique, Université Mouloud Mammeri de Tizi-ouzou, BP 17 RP, 15000 Tizi-Ouzou (Algeria)

    2013-09-01

    The particle in cell (PIC) method coupled to the finite-difference time-domain (FDTD) method is used to model the formation of laser induced periodic surface structures (LIPSS) at the early stage of femtosecond laser irradiation of smooth metal surface. The theoretical results were analyzed and compared with experimental data taken from the literature. It was shown that the optical properties of the target are not homogeneous and the ejection of electrons is such that ripples in the electron density were obtained. The Coulomb explosion mechanism was proposed to explain the ripples formation under the considered conditions.

  19. The Cell Factory Aspergillus Enters the Big Data Era: Opportunities and Challenges for Optimising Product Formation.

    Science.gov (United States)

    Meyer, Vera; Fiedler, Markus; Nitsche, Benjamin; King, Rudibert

    2015-01-01

    Living with limits. Getting more from less. Producing commodities and high-value products from renewable resources including waste. What is the driving force and quintessence of bioeconomy outlines the lifestyle and product portfolio of Aspergillus, a saprophytic genus, to which some of the top-performing microbial cell factories belong: Aspergillus niger, Aspergillus oryzae and Aspergillus terreus. What makes them so interesting for exploitation in biotechnology and how can they help us to address key challenges of the twenty-first century? How can these strains become trimmed for better growth on second-generation feedstocks and how can we enlarge their product portfolio by genetic and metabolic engineering to get more from less? On the other hand, what makes it so challenging to deduce biological meaning from the wealth of Aspergillus -omics data? And which hurdles hinder us to model and engineer industrial strains for higher productivity and better rheological performance under industrial cultivation conditions? In this review, we will address these issues by highlighting most recent findings from the Aspergillus research with a focus on fungal growth, physiology, morphology and product formation. Indeed, the last years brought us many surprising insights into model and industrial strains. They clearly told us that similar is not the same: there are different ways to make a hypha, there are more protein secretion routes than anticipated and there are different molecular and physical mechanisms which control polar growth and the development of hyphal networks. We will discuss new conceptual frameworks derived from these insights and the future scientific advances necessary to create value from Aspergillus Big Data. PMID:25616499

  20. The Cell Factory Aspergillus Enters the Big Data Era: Opportunities and Challenges for Optimising Product Formation.

    Science.gov (United States)

    Meyer, Vera; Fiedler, Markus; Nitsche, Benjamin; King, Rudibert

    2015-01-01

    Living with limits. Getting more from less. Producing commodities and high-value products from renewable resources including waste. What is the driving force and quintessence of bioeconomy outlines the lifestyle and product portfolio of Aspergillus, a saprophytic genus, to which some of the top-performing microbial cell factories belong: Aspergillus niger, Aspergillus oryzae and Aspergillus terreus. What makes them so interesting for exploitation in biotechnology and how can they help us to address key challenges of the twenty-first century? How can these strains become trimmed for better growth on second-generation feedstocks and how can we enlarge their product portfolio by genetic and metabolic engineering to get more from less? On the other hand, what makes it so challenging to deduce biological meaning from the wealth of Aspergillus -omics data? And which hurdles hinder us to model and engineer industrial strains for higher productivity and better rheological performance under industrial cultivation conditions? In this review, we will address these issues by highlighting most recent findings from the Aspergillus research with a focus on fungal growth, physiology, morphology and product formation. Indeed, the last years brought us many surprising insights into model and industrial strains. They clearly told us that similar is not the same: there are different ways to make a hypha, there are more protein secretion routes than anticipated and there are different molecular and physical mechanisms which control polar growth and the development of hyphal networks. We will discuss new conceptual frameworks derived from these insights and the future scientific advances necessary to create value from Aspergillus Big Data.

  1. Metformin inhibits the proliferation, metastasis, and cancer stem-like sphere formation in osteosarcoma MG63 cells in vitro.

    Science.gov (United States)

    Chen, Xu; Hu, Chuanzhen; Zhang, Weibin; Shen, Yuhui; Wang, Jun; Hu, Fangqiong; Yu, Pei

    2015-12-01

    Metformin is an oral drug that has been widely used to treat type 2 diabetes mellitus. Interestingly, accumulated evidence indicate that metformin may reduce the risk of cancer in patients with type 2 diabetes and inhibit tumor cell growth and survival in numerous malignancies, including osteosarcoma (OS) cells. In the present study, we aimed to investigate the effects of metformin on the proliferation, migration, invasion, and sphere formation in OS MG63 cells in vitro. Metformin suppressed OS MG63 cell proliferation in a dose- and time-dependent manner and markedly blocked anti-metastatic potentials, migration, and invasion, by downregulating matrix metalloproteinase 2 (MMP2) and MMP9. Besides, we established OS cancer stem-like cell (CSC) model with sarcosphere formation assay and demonstrated that metformin posed damage on CSCs in OS by inhibiting sphere formation and by inducing their stemness loss. The stemness of CSCs in OS such as self-renewal and differentiation potentials was both impaired with a significant decrease of Oct-4 and Nanog activation. Consistent with this, the positive rates of CD90, CD133, and stage-specific embryonic antigen-4 (SSEA-4) were all observed with reductions in response to metformin exposure. In addition, Western blot showed that metformin activated AMPKα at Tyr172, followed by a downregulated phosphorylation of mammalian target of rapamycin (mTOR)/S6 and feedback activation of p-AKT Ser(473) in both OS MG63 cells and CSCs. This indicates that AMPK/mTOR/S6 signaling pathway might be involved in the growth inhibition of both OS MG63 cells and CSCs. These results suggest that metformin, a potential anti-neoplastic agent, might make it a novel therapeutic choice for the treatment of OS in the future.

  2. Effect of Magnesium and Osteoblast Cell Presence on Hydroxyapatite Formation on (Ti,Mg)N Thin Film Coatings

    Science.gov (United States)

    Onder, Sakip; Calikoglu-Koyuncu, Ayse Ceren; Torun Kose, Gamze; Kazmanli, Kursat; Kok, Fatma Nese; Urgen, Mustafa

    2016-07-01

    TiN and (Ti,Mg)N thin film coatings were deposited on Ti substrates by an arc-physical vapor deposition technique. The effect of cell presence on hydroxyapatite (HA) formation was investigated using surfaces with four different Mg contents (0, 8.1, 11.31, and 28.49 at.%). Accelerated corrosion above 10 at.% Mg had a negative effect on the performance in terms of both cell proliferation and mineralization. In the absence of cells, Mg-free TiN coatings and low-Mg (8.1 at.%)-doped (Ti,Mg)N surfaces led to an early HA deposition (after 7 days and 14 days, respectively) in cell culture medium (DMEM), but the crystallinity was low. More crystalline HA structures were obtained in the presence of the cells. HA deposits with an ideal Ca/P ratio were obtained at least a week earlier, at day 14, in TiN and low-Mg (8.1 at.%)-doped (Ti,Mg)N compared with that of high-Mg-containing surfaces (>10 at.%). A thicker mineralized matrix was formed on low-Mg (8.1 at.%)-doped (Ti,Mg)N relative to that of the TiN sample. Low-Mg doping (coatings resulted in better cell proliferation and thicker mineralized matrix formation, so it could be a promising alternative for hard tissue applications.

  3. Stem cell-dependent formation of a functional anterior regeneration pole in planarians requires Zic and Forkhead transcription factors.

    Science.gov (United States)

    Vogg, Matthias C; Owlarn, Suthira; Pérez Rico, Yuvia A; Xie, Jianlei; Suzuki, Yoko; Gentile, Luca; Wu, Wei; Bartscherer, Kerstin

    2014-06-15

    Planarians can regenerate their head within days. This process depends on the direction of adult stem cells to wound sites and the orchestration of their progenitors to commit to appropriate lineages and to arrange into patterned tissues. We identified a zinc finger transcription factor, Smed-ZicA, as a downstream target of Smed-FoxD, a Forkhead transcription factor required for head regeneration. Smed-zicA and Smed-FoxD are co-expressed with the Wnt inhibitor notum and the Activin inhibitor follistatin in a cluster of cells at the anterior-most tip of the regenerating head - the anterior regeneration pole - and in surrounding stem cell progeny. Depletion of Smed-zicA and Smed-FoxD by RNAi abolishes notum and follistatin expression at the pole and inhibits head formation downstream of initial polarity decisions. We suggest a model in which ZicA and FoxD transcription factors synergize to control the formation of Notum- and Follistatin-producing anterior pole cells. Pole formation might constitute an early step in regeneration, resulting in a signaling center that orchestrates cellular events in the growing tissue. PMID:24704339

  4. Influence of culture media on biofilm formation by Candida species and response of sessile cells to antifungals and oxidative stress.

    Science.gov (United States)

    Serrano-Fujarte, Isela; López-Romero, Everardo; Reyna-López, Georgina Elena; Martínez-Gámez, Ma Alejandrina; Vega-González, Arturo; Cuéllar-Cruz, Mayra

    2015-01-01

    The aims of the study were to evaluate the influence of culture media on biofilm formation by C. albicans, C. glabrata, C. krusei, and C. parapsilosis and to investigate the responses of sessile cells to antifungals and reactive oxygen species (ROS) as compared to planktonic cells. For biofilm formation, the Candida species were grown at different periods of time in YP or YNB media supplemented or not with 0.2 or 2% glucose. Sessile and planktonic cells were exposed to increasing concentrations of antifungals, H2O2, menadione or silver nanoparticles (AgNPs). Biofilms were observed by scanning electron microscopy (SEM) and quantified by the XTT assay. C. albicans formed biofilms preferentially in YPD containing 2% glucose (YPD/2%), C. glabrata in glucose-free YNB or supplemented with 0.2% glucose (YNB/0.2%), while C. krusei and C. parapsilosis preferred YP, YPD/0.2%, and YPD/2%. Interestingly, only C. albicans produced an exopolymeric matrix. This is the first report dealing with the in vitro effect of the culture medium and glucose on the formation of biofilms in four Candida species as well as the resistance of sessile cells to antifungals, AgNPs, and ROS. Our results suggest that candidiasis in vivo is a multifactorial and complex process where the nutritional conditions, the human immune system, and the adaptability of the pathogen should be considered altogether to provide an effective treatment of the patient.

  5. Thrombospondin type I domain containing 7A (THSD7A) mediates endothelial cell migration and tube formation.

    Science.gov (United States)

    Wang, Chieh-Huei; Su, Pei-Tsu; Du, Xiao-Yan; Kuo, Meng-Wei; Lin, Chia-Yi; Yang, Chung-Chi; Chan, Hau-Shien; Chang, Shing-Jyh; Kuo, Calvin; Seo, Kyunga; Leung, Lawrence L; Chuang, Yung-Jen

    2010-03-01

    Angiogenesis is a highly organized process controlled by a series of molecular events. While much effort has been devoted to identifying angiogenic factors and their reciprocal receptors, far less information is available on the molecular mechanisms underlying directed endothelial cell migration. To search for novel proteins that participate in this process, we used the serial analysis of gene expression (SAGE) transcript profiling approach to identify genes that are selectively expressed in endothelial cells (ECs). Two EC SAGE libraries were constructed from human umbilical vein and artery ECs to enable data-mining against other non-ECs. A novel endothelial protein, Thrombospondin Type I Domain Containing 7A (THSD7A), with preferential expression in placenta vasculature and in human umbilical vein endothelial cells (HUVECs) was identified and targeted for further characterization. Overexpression of a THSD7A carboxyl-terminal fragment in HUVECs inhibited cell migration and disrupted tube formation, while suppression of THSD7A expression enhanced HUVEC migration and tube formation. Immunohistological analysis revealed that THSD7A was expressed at the leading edge of migrating HUVECs, and it co-localized with alpha(V)beta(3) integrin and paxillin. This distribution was dispersed from focal adhesions after disruption of the actin cytoskeleton, suggesting the involvement of THSD7A in cytoskeletal organization. Our results show that THSD7A is a novel placenta endothelial protein that mediates EC migration and tube formation, and they highlight its potential as a new target for anti-angiogenic therapy.

  6. Stem cell-dependent formation of a functional anterior regeneration pole in planarians requires Zic and Forkhead transcription factors.

    Science.gov (United States)

    Vogg, Matthias C; Owlarn, Suthira; Pérez Rico, Yuvia A; Xie, Jianlei; Suzuki, Yoko; Gentile, Luca; Wu, Wei; Bartscherer, Kerstin

    2014-06-15

    Planarians can regenerate their head within days. This process depends on the direction of adult stem cells to wound sites and the orchestration of their progenitors to commit to appropriate lineages and to arrange into patterned tissues. We identified a zinc finger transcription factor, Smed-ZicA, as a downstream target of Smed-FoxD, a Forkhead transcription factor required for head regeneration. Smed-zicA and Smed-FoxD are co-expressed with the Wnt inhibitor notum and the Activin inhibitor follistatin in a cluster of cells at the anterior-most tip of the regenerating head - the anterior regeneration pole - and in surrounding stem cell progeny. Depletion of Smed-zicA and Smed-FoxD by RNAi abolishes notum and follistatin expression at the pole and inhibits head formation downstream of initial polarity decisions. We suggest a model in which ZicA and FoxD transcription factors synergize to control the formation of Notum- and Follistatin-producing anterior pole cells. Pole formation might constitute an early step in regeneration, resulting in a signaling center that orchestrates cellular events in the growing tissue.

  7. Ceramide Kinase Contributes to Proliferation but not to Prostaglandin E2 Formation in Renal Mesangial Cells and Fibroblasts

    Directory of Open Access Journals (Sweden)

    Oleksandr Pastukhov

    2014-06-01

    Full Text Available Background/Aims: Ceramide kinase (CerK catalyzes the generation of the sphingolipid ceramide-1-phosphate (C1P which regulates various cellular functions including cell growth and death, and inflammation. Here, we used a novel catalytic inhibitor of CerK, NVP-231, and CerK knockout cells to investigate the contribution of CerK to proliferation and inflammation in renal mesangial cells and fibroblasts. Methods: Cells were treated with NVP-231 and [3H]-thymidine incorporation into DNA, [3H]-arachidonic acid release, prostaglandin E2 (PGE2 synthesis, cell cycle distribution, and apoptosis were determined. Results: Treatment of rat mesangial cells and mouse renal fibroblasts with NVP-231 decreased DNA synthesis, but not of agonist-stimulated arachidonic acid release or PGE2 synthesis. Similarly, proliferation but not arachidonic acid release or PGE2 synthesis was reduced in CERK knockout renal fibroblasts. The anti-proliferative effect of NVP-231 on mesangial cells was due to M phase arrest as determined using the mitosis markers phospho-histone H3, cdc2 and polo-like kinase-1, and induction of apoptosis. Moreover, loss of CerK sensitized cells towards stress-induced apoptosis. Conclusions: Our data demonstrate that CerK induces proliferation but not PGE2 formation of renal mesangial cells and fibroblasts, and suggest that targeted CerK inhibition has potential for treating mesangioproliferative kidney diseases.

  8. Inducible formation of breast cancer stem cells and their dynamic equilibrium with non-stem cancer cells via IL6 secretion.

    Science.gov (United States)

    Iliopoulos, Dimitrios; Hirsch, Heather A; Wang, Guannan; Struhl, Kevin

    2011-01-25

    Tumors are often heterogeneous, being composed of multiple cell types with different phenotypic and molecular properties. Cancer stem-like cells (CSCs) are a highly tumorigenic cell type found in developmentally diverse tumors or cancer cell lines, and they are often resistant to standard chemotherapeutic drugs. The origins of CSCs and their relationships to nonstem cancer cells (NSCCs) are poorly understood. In an inducible breast oncogenesis model, CSCs are generated from nontransformed cells at a specific time during the transformation process, but CSC formation is not required for transformation. MicroRNA profiles indicate that CSCs and NSCCs are related, but different cell types arising from a common nontransformed population. Interestingly, medium from the transformed population stimulates NSCCs to become CSCs, and conversion of NSCCs to CSCs occurs in mouse xenografts. Furthermore, IL6 is sufficient to convert NSCCs to CSCs in genetically different breast cell lines, human breast tumors, and a prostate cell line. Thus, breast and prostate CSCs and NSCCs do not represent distinct epigenetic states, and these CSCs do not behave as or arise from classic stem cells. Instead, tumor heterogeneity involves a dynamic equilibrium between CSCs and NSCCs mediated by IL6 and activation of the inflammatory feedback loop required for oncogenesis. This dynamic equilibrium provides an additional rationale for combining conventional chemotherapy with metformin, which selectively inhibits CSCs. PMID:21220315

  9. Influence of culture heterogeneity in cell surface charge on adhesion and biofilm formation by Enterococcus faecalis

    NARCIS (Netherlands)

    van Merode, Annet E.J.; van der Mei, HC; Busscher, HJ; Krom, BP

    2006-01-01

    Biofilm formation is an increasing problem in medicine, due to the intrinsic resistance of microorganisms in the biofilm mode of growth against the host immune system and antimicrobial therapy. Adhesion is an important step in biofilm formation, influenced, among other factors, by the surface hydrop

  10. Three-dimensional-engineered matrix to study cancer stem cells and tumorsphere formation: effect of matrix modulus.

    Science.gov (United States)

    Yang, Xiaoming; Sarvestani, Samaneh K; Moeinzadeh, Seyedsina; He, Xuezhong; Jabbari, Esmaiel

    2013-03-01

    Maintenance of cancer stem cells (CSCs) is regulated by the tumor microenvironment. Synthetic hydrogels provide the flexibility to design three-dimensional (3D) matrices to isolate and study individual factors in the tumor microenvironment. The objective of this work was to investigate the effect of matrix modulus on tumorsphere formation by breast cancer cells and maintenance of CSCs in an inert microenvironment without the interference of other factors. In that regard, 4T1 mouse breast cancer cells were encapsulated in inert polyethylene glycol diacrylate hydrogels and the effect of matrix modulus on tumorsphere formation and expression of CSC markers was investigated. The gel modulus had a strong effect on tumorsphere formation and the effect was bimodal. Tumorsphere formation and expression of CSC markers peaked after 8 days of culture. At day 8, as the matrix modulus was increased from 2.5 kPa to 5.3, 26.1, and 47.1 kPa, the average tumorsphere size changed from 37±6 μm to 57±6, 20±4, and 12±2 μm, respectively; cell number density in the gel changed from 0.8±0.1×10⁵ cells/mL to 1.7±0.2×10⁵, 0.4±0.1×10⁵, and 0.2±0.1×10⁵ cells/mL after initial encapsulation of 0.14×10⁵ cells/mL; and the expression of CD44 breast CSC marker changed from 17±4-fold to 38±9-, 3±1-, and 2±1-fold increase compared with the initial level. Similar results were obtained with MCF7 human breast carcinoma cells. Mouse 4T1 and human MCF7 cells encapsulated in the gel with 5.3 kPa modulus formed the largest tumorspheres and highest density of tumorspheres, and had highest expression of breast CSC markers CD44 and ABCG2. The inert polyethylene glycol hydrogel can be used as a model-engineered 3D matrix to study the role of individual factors in the tumor microenvironment on tumorigenesis and maintenance of CSCs without the interference of other factors. PMID:23013450

  11. Amniotic fluid stem cell-based models to study the effects of gene mutations and toxicants on male germ cell formation

    Institute of Scientific and Technical Information of China (English)

    Claudia Gundacker; Helmut Dolznig; Mario Mikula; Margit Rosner; Oliver Brandau; Markus Hengstschl(a)ger

    2012-01-01

    Male infertility is a major public health issue predominantly caused by defects in germ cell development.In the past,studies on the genetic regulation of spermatogenesis as well as on negative environmental impacts have been hampered by the fact that human germ cell development is intractable to direct analysis in vivo.Compared with model organisms including mice,there are fundamental differences in the molecular processes of human germ cell development.Therefore,an in vitro model mimicking human sperm formation would be an extremely valuable research tool.In the recent past,both human embryonic stem (ES) cells and induced pluripotent stem (iPS) cells have been reported to harbour the potential to differentiate into primordial germ cells and gametes.We here discuss the pessibility to use human amniotic fluid stem (AFS) cells as a biological model.Since their discovery in 2003,AFS cells have been characterized to differentiate into cells of all three germ layers,to be genomically stable,to have a high proliferative potential and to be non-tumourigenic.In addition,AFS cells are not subject of ethical concerns.In contrast to iPS cells,AFSs cells do not need ectopic induction of pluripotency,which is often associated with only imperfectly cleared epigenetic memory of the source cells.Since AFS cells can be derived from amniocentesis with disease-causing mutations and can be transfected with high efficiency,they could be used in probing gene functions for spermatogenesis and in screening for male reproductive toxicity.

  12. Apposition of iroquois expressing and non-expressing cells leads to cell sorting and fold formation in the Drosophila imaginal wing disc

    Directory of Open Access Journals (Sweden)

    González-Pérez Esther

    2007-09-01

    Full Text Available Abstract Background The organization of the different tissues of an animal requires mechanisms that regulate cell-cell adhesion to promote and maintain the physical separation of adjacent cell populations. In the Drosophila imaginal wing disc the iroquois homeobox genes are expressed in the notum anlage and contribute to the specification of notum identity. These genes are not expressed in the adjacent wing hinge territory. These territories are separated by an approximately straight boundary that in the mature disc is associated with an epithelial fold. The mechanism by which these two cell populations are kept separate is unclear. Results Here we show that the Iro-C genes participate in keeping the notum and wing cell populations separate. Indeed, within the notum anlage, cells not expressing Iro-C tend to join together and sort out from their Iro-C expressing neighbours. We also show that apposition of Iro-C expressing and non-expressing cells induces invagination and apico-basal shortening of the Iro-C- cells. This effect probably underlies formation of the fold that separates the notum and wing hinge territories. In addition, cells overexpressing a member of the Iro-C contact one another and become organized in a network of thin strings that surrounds and isolates large groups of non-overexpressing cells. The strings appear to exert a pulling force along their longitudinal axis. Conclusion Apposition of cells expressing and non-expressing the Iro-C, as it occurs in the notum-wing hinge border of the Drosophila wing disc, influences cell behaviour. It leads to cell sorting, and cellular invagination and apical-basal shortening. These effects probably account for keeping the prospective notum and wing hinge cell populations separate and underlie epithelial fold formation. Cells that overexpress a member of the Iro-C and that confront non-expressing cells establish contacts between themselves and become organized in a network of thin strings

  13. Efficacy of a small cell-binding peptide coated hydroxyapatite substitute on bone formation and implant fixation in sheep

    DEFF Research Database (Denmark)

    Ding, Ming; Andreasen, Christina Møller; Dencker, Mads L.;

    2015-01-01

    Cylindrical critical size defects were created at the distal femoral condyles bilaterally of eight female adult sheep. Titanium implants with 2-mm concentric gaps were inserted and the gaps were filled with one of the four materials: allograft; a synthetic 15-amino acid cell-binding peptide coated...... hydroxyapatite (ABM/P-15); hydroxyapatite + βtricalciumphosphate+ Poly-Lactic-Acid (HA/βTCP-PDLLA); or ABM/P-15+HA/βTCP-PDLLA. After nine weeks, bone-implant blocks were harvested and sectioned for micro-CT scanning, push-out test, and histomorphometry. Significant bone formation and implant fixation could...... formation in concentric gap, and its enhancements on bone formation and implant fixation were at least as good as allograft. It is suggested that ABM/P-15 might be a good alternative biomaterial for bone implant fixation in this well-validated critical-size defect gap model in sheep. Nevertheless, future...

  14. Triplet formation in fullerene multi-adduct blends for organic solar cells and its influence on device performance

    Energy Technology Data Exchange (ETDEWEB)

    Dyer-Smith, Clare [Department of Physics, Imperial College London, Blackett Laboratory, Prince Consort Road, London SW7 2AZ (United Kingdom); Department of Chemistry, Imperial College London, South Kensington Campus, London, SW7 2AZ (United Kingdom); Grantham Institute for Climate Change, Imperial College London, South Kensington Campus, London, SW7 2AZ (United Kingdom); Reynolds, Luke X. [Department of Chemistry, Imperial College London, South Kensington Campus, London, SW7 2AZ (United Kingdom); Grantham Institute for Climate Change, Imperial College London, South Kensington Campus, London, SW7 2AZ (United Kingdom); Bruno, Annalisa; Haque, Saif A. [Department of Chemistry, Imperial College London, South Kensington Campus, London, SW7 2AZ (United Kingdom); Bradley, Donal D.C.; Nelson, Jenny [Department of Physics, Imperial College London, Blackett Laboratory, Prince Consort Road, London SW7 2AZ (United Kingdom)

    2010-08-23

    In organic solar cells, high open circuit voltages may be obtained by choosing materials with a high offset between the donor highest occupied molecular orbital (HOMO) and acceptor lowest unoccupied molecular orbital (LUMO). However, increasing this energy offset can also lead to photophysical processes that compete with charge separation. In this paper the formation of triplet states is addressed in blends of polyfluorene polymers with a series of PCBM multi-adducts. Specifically, it is demonstrated that the formation of such triplets occurs when the offset energy between donor ionization potential and acceptor electron affinity is {proportional_to}1.6 eV or greater. Spectroscopic measurements support a mechanism of resonance energy transfer for triplet formation, influenced by the energy levels of the materials, but also demonstrate that the competition between processes at the donor-acceptor interface is strongly influenced by morphology. (Abstract Copyright [2010], Wiley Periodicals, Inc.)

  15. Abiotic factors in colony formation: effects of nutrition and light on extracellular polysaccharide production and cell aggregates of Microcystis aeruginosa

    Science.gov (United States)

    Yang, Zhen; Kong, Fanxiang

    2013-07-01

    Colony morphology is important for Microcystis to sustain a competitive advantage in eutrophic lakes. The mechanism of colony formation in Microcystis is currently unclear. Extracellular polysaccharide (EPS) has been reported to play an important role in cell aggregate formation of some phytoplankton. Microcystis aeruginosa was cultivated under varied abiotic conditions, including different nutrient, light, and temperature conditions, to investigate their effects on EPS production and morphological change. The results show that nutrient concentration and light intensity have great effects on EPS productionin M. aeruginosa. There was a considerable increase in EPS production after M. aeruginosa was cultivated in adjusted culture conditions similar to those present in the field (28.9 mg C/L, 1.98 mg N/L, 0.65 mg P/L, light intensity: 100 μmol/(m2 · s)). These results indicate that abiotic factors might be one of the triggers for colony formation in Microcystis.

  16. HIV gp120 induces mucus formation in human bronchial epithelial cells through CXCR4/α7-nicotinic acetylcholine receptors.

    Directory of Open Access Journals (Sweden)

    Sravanthi Gundavarapu

    Full Text Available Lung diseases such as chronic obstructive pulmonary disease (COPD, asthma, and lung infections are major causes of morbidity and mortality among HIV-infected patients even in the era of antiretroviral therapy (ART. Many of these diseases are strongly associated with smoking and smoking is more common among HIV-infected than uninfected people; however, HIV is an independent risk factor for chronic bronchitis, COPD, and asthma. The mechanism by which HIV promotes these diseases is unclear. Excessive airway mucus formation is a characteristic of these diseases and contributes to airway obstruction and lung infections. HIV gp120 plays a critical role in several HIV-related pathologies and we investigated whether HIV gp120 promoted airway mucus formation in normal human bronchial epithelial (NHBE cells. We found that NHBE cells expressed the HIV-coreceptor CXCR4 but not CCR5 and produced mucus in response to CXCR4-tropic gp120. The gp120-induced mucus formation was blocked by the inhibitors of CXCR4, α7-nicotinic acetylcholine receptor (α7-nAChR, and γ-aminobutyric acid (GABAAR but not the antagonists of CCR5 and epithelial growth factor receptor (EGFR. These results identify two distinct pathways (α7-nAChR-GABAAR and EGFR for airway mucus formation and demonstrate for the first time that HIV-gp120 induces and regulates mucus formation in the airway epithelial cells through the CXCR4-α7-nAChR-GABAAR pathway. Interestingly, lung sections from HIV ± ART and simian immunodeficiency virus (SIV ± ART have significantly more mucus and gp120-immunoreactivity than control lung sections from humans and macaques, respectively. Thus, even after ART, lungs from HIV-infected patients contain significant amounts of gp120 and mucus that may contribute to the higher incidence of obstructive pulmonary diseases in this population.

  17. Osteoprotegerin and osteoprotegerin ligand expression during human marrow stromal cell differentiation and their effect on osteoclast formation

    Institute of Scientific and Technical Information of China (English)

    YANG Lin; HAI Yong; ZHOU Jun-lin

    2011-01-01

    ackground Osteoprotegerin (OPG) and osteoprotegerin ligand (OPGL) play an important role in human bone metabolism. The aim of this research was to detect the expression of OPG and OPGL during human marrow stromal cells (hMSC) differentiation into osteoblasts (OB), and to observe their effect on osteoclasts (OC) formation in vitro to investigate bone metabolism mechanisms.Methods hMSCs were obtained from human bone marrow specimens using gradient centrifugation method, before being purified and incubated with differentiation medium to develop along the human osteoblasts (hOB) pathway. Morphology observation, biochemical detection and cell staining were performed during hMSC differentiation. OPG and OPGL mRNA levels were detected by reverse transcription-polymerase chain reaction. OPG and OPGL protein expression were determined by Western blotting. We further obtained OC progenitor cells from mice bone marrow and co-cultured with differentiating MSCs. We assessed the effect of OPG and OPGL on OC formation by identifying tartrate resistant acid phosphatase (TRAP) positive multinuclear cells.Results Optimal hMSC survival and purification were observed, along with stable biochemical indexes. Alkaline phosphatase secretion increased significantly and mineralization nodules appeared in the process of cell differentiation. OPG mRNA and protein level increased significantly, while OPGL mRNA and protein level decreased. Average levels of OPG mRNA and protein were about 2.5-fold higher than the control, while OPGL mRNA and protein levels were reduced by about one-half. In the group co-culturing with undifferentiated MSC or added OPGL, we found TRAP positive and multinuclear OC formation. However, OC formation was absent in the group co-culturing with differentiated MSC or added OPG.Conclusions During hMSC differentiation into hOB, OPG secretion increased rapidly and OPGL production decreased significantly. The OPG/OPGL ratio was also increased, while OC formation was

  18. Comparison of the colony formation and crystal violet cell proliferation assays to determine cellular radiosensitivity in a repair-deficient MCF10A cell line

    Energy Technology Data Exchange (ETDEWEB)

    Vandersickel, Veerle [Department of Basic Medical Sciences, Ghent University, Campus Heymans, De Pintelaan 185 (6B3), 9000 Gent (Belgium); Slabbert, Jacobus [NRF iThemba LABS (Laboratory for Accelerated Based Sciences), PO box 722, 7129 Somerset West (South Africa); Thierens, Hubert [Department of Basic Medical Sciences, Ghent University, Campus Heymans, De Pintelaan 185 (6B3), 9000 Gent (Belgium); Vral, Anne, E-mail: anne.Vral@UGent.b [Department of Basic Medical Sciences, Ghent University, Campus Heymans, De Pintelaan 185 (6B3), 9000 Gent (Belgium)

    2011-01-15

    Colony formation as measured by the in vitro clonogenic assay is a very important endpoint to determine cellular radiosensitivity and tumor response to radiotherapy. In the framework of assessing in vitro cellular radiosensitivity, proliferation assays could represent an attractive alternative to the clonogenic assay for cell lines that do not form proper colonies. In the present study, we compared cellular radiosensitivity measurements obtained by the crystal violet (CV) cell proliferation assay and the standard colony formation assay in repair-deficient and-proficient human MCF10A cell lines. Compared to the clonogenic assay, the CV cell proliferation assay yielded higher surviving fractions for the same radiation dose. This is reflected in larger mean inactivation dose values - a parameter that reflects the area under the survival curve. However, as the dose modifying factors obtained by both assays are comparable, the CV cell proliferation assay can be used to compare the in vitro cellular radiosensitivity of cell lines that lack the ability to form well-defined colonies.

  19. Ectopic osteogenic tissue formation by MC3T3-E1 cell-laden chitosan/hydroxyapatite composite scaffold.

    Science.gov (United States)

    Koç, Aysel; Elçin, Ayşe Eser; Elçin, Yaşar Murat

    2016-09-01

    This study evaluates the suitability of a macroporous three-dimensional chitosan/hydroxyapatite (CS/HA) composite as a bone tissue engineering scaffold using MC3T3-E1 cells. The CS/HA scaffold was produced by freeze-drying, and characterized by means of SEM and FTIR. In vitro findings demonstrated that CS/HA supported attachment and proliferation of cells, and stimulated extracellular matrix (ECM) production. Tissue biocompatibility and osteogenic capacity of the cell-laden constructs were evaluated in an ectopic Wistar rat model. In vivo results showed that the MC3T3-E1 cell-laden CS/HA was essentially histocompatible, promoted neovascularization and calcified matrix formation, and secreted osteoblast-specific protein. We conclude that the composite scaffold evaluated has potential for applications in bone regeneration. PMID:25968048

  20. Mutant KRas-Induced Mitochondrial Oxidative Stress in Acinar Cells Upregulates EGFR Signaling to Drive Formation of Pancreatic Precancerous Lesions

    Directory of Open Access Journals (Sweden)

    Geou-Yarh Liou

    2016-03-01

    Full Text Available The development of pancreatic cancer requires the acquisition of oncogenic KRas mutations and upregulation of growth factor signaling, but the relationship between these is not well established. Here, we show that mutant KRas alters mitochondrial metabolism in pancreatic acinar cells, resulting in increased generation of mitochondrial reactive oxygen species (mROS. Mitochondrial ROS then drives the dedifferentiation of acinar cells to a duct-like progenitor phenotype and progression to PanIN. This is mediated via the ROS-receptive kinase protein kinase D1 and the transcription factors NF-κB1 and NF-κB2, which upregulate expression of the epidermal growth factor, its ligands, and their sheddase ADAM17. In vivo, interception of KRas-mediated generation of mROS reduced the formation of pre-neoplastic lesions. Hence, our data provide insight into how oncogenic KRas interacts with growth factor signaling to induce the formation of pancreatic cancer.

  1. Regulatory T cells with reduced repressor capacities are extensively amplified in pulmonary sarcoid lesions and sustain granuloma formation.

    Science.gov (United States)

    Rappl, Gunter; Pabst, Stefan; Riemann, Dagmar; Schmidt, Annette; Wickenhauser, Claudia; Schütte, Wolfgang; Hombach, Andreas A; Seliger, Barbara; Grohé, Christian; Abken, Hinrich

    2011-07-01

    Sarcoidosis can evolve into a chronic disease with persistent granulomas accompanied by progressive fibrosis. While an unlimited inflammatory response suggests an impaired immune control in sarcoid lesions, it stands in contrast to the massive infiltration with CD4(+)CD25(high)FoxP3(+) regulatory T cells. We here revealed that those Treg cells in affected lung lesions were mainly derived from activated natural Treg cells with GARP (LRRC32)-positive phenotype but exhibited reduced repressor capacities despite high IL-10 and TGF-beta 1 levels. The repressive capacity of blood Treg cells, in contrast, was not impaired compared to age-matched healthy donors. Treg derived cells in granuloma lesions have undergone extensive rounds of amplifications indicated by shortened telomeres compared to blood Treg cells of the same patient. Lesional Treg derived cells moreover secreted pro-inflammatory cytokines including IL-4 which sustains granuloma formation through fibroblast amplification and the activation of mast cells, the latter indicated by the expression of membrane-bound oncostatin M. PMID:21482483

  2. TM4SF1: A tetraspanin-like protein necessary for nanopodia formation and endothelial cell migration

    OpenAIRE

    Zukauskas, Andrew; Merley, Anne; Li, Dan; Ang, Lay-Hong; Sciuto, Tracey E.; Salman, Samantha; Dvorak, Ann M.; Dvorak, Harold F.; Jaminet, Shou-Ching S.

    2011-01-01

    Transmembrane-4-L-six-family-1 (TM4SF1) is a tetraspanin-like membrane protein that is highly and selectively expressed by cultured endothelial cells (EC) and, in vivo, by EC lining angiogenic tumor blood vessels. TM4SF1 is necessary for the formation of unusually long (up to a 50 µm), thin (~ 100–300 nm wide), F-actin-poor EC cell projections that we term ‘nanopodia’. Immunostaining of nanopodia at both the light and electron microsopic levels localized TM4SF1 in a regularly spaced, banded p...

  3. The Protrusive Phase and Full Development of Integrin-Dependent Adhesions in Colon Epithelial Cells Require FAK- and ERKMediated Actin Spike Formation: Deregulation in Cancer Cells

    Directory of Open Access Journals (Sweden)

    Valerie G. Brunton

    2001-01-01

    Full Text Available Integrins play an important role in tumour progression by influencing cellular responses and matrix-dependent adhesion. However, the regulation of matrix-dependent adhesion assembly in epithelial cells is poorly understood. We have investigated the integrin and signalling requirements of cell-matrix adhesion assembly in colon carcinoma cells after plating on fibronectin. Adhesion assembly in these, and in the adenoma cells from which they were derived, was largely dependent on αvβ6 integrin and required phosphorylation of FAK on tyrosine-397. The rate of fibronectin-induced adhesion assembly and the expression of both αvβ6 integrin and FAK were increased during the adenoma-to-carcinoma transition. The matrix-dependent adhesion assembly process, particularly the final stages of complex protrusion that is required for optimal cell spreading, required the activity of extracellular signal-regulated kinase (ERK. Furthermore, phosphorylated ERK was targeted to newly forming cell-matrix adhesions in the carcinoma cells but not the adenoma cells, and inhibition of FAK-tyrosine-397 phosphorylation or MEK suppressed the appearance of phosphorylated ERK at peripheral sites. In addition, inhibition of MEK-ERK activation blocked the formation of peripheral actin microspikes that were necessary for the protrusive phase of cell-matrix adhesion assembly. Thus, MEK-ERK-dependent peripheral actin re-organization is required for the full development of integrin-induced adhesions and this pathway is stimulated in an in vitro model of colon cancer progression.

  4. Coke formation and performance of an intermediate-temperature solid oxide fuel cell operating on dimethyl ether fuel

    Energy Technology Data Exchange (ETDEWEB)

    Su, Chao; Ran, Ran; Wang, Wei; Shao, Zongping [State Key Laboratory of Materials-Oriented Chemical Engineering, College of Chemistry and Chemical Engineering, Nanjing University of Technology, No. 5 Xin Mofan Road, Nanjing 210009 (China)

    2011-02-15

    Dimethyl ether (DME) as a fuel of SOFCs is investigated with great attention paid to coke formation over the Ni-YSZ anode. DME is easily decomposed to CH{sub 4}, CO and H{sub 2} at temperatures above 700 C, with total conversion occurring at 850 C over the Ni-YSZ catalyst. These data suggest that the DME electro-oxidation likely proceeds via an indirect pathway. O{sub 2}-TPO analysis, laser Raman spectroscopy and SEM-EDX characterizations demonstrate coke formation over Ni-YSZ, which is obvious and become more prevalent at higher temperatures. The introduction of CO{sub 2} in the fuel gas decreases the CH{sub 4} selectivity and effectively suppresses coke formation above 700 C. The suppression effect is increasingly apparent at higher temperatures. At 850 C, the anode still maintains geometric integrity after exposure to DME-CO{sub 2} (1:1, volume ratio) under OCV condition. With DME or DME-CO{sub 2}, the fuel cell power output is comparable to results obtained by operating with 3% water humidified hydrogen. No obvious cell degradation from the anode is observed when operating with DME-CO{sub 2}, while it is obvious with DME. The introduction of CO{sub 2} may be a good choice to suppress the coke formation when operating on DME; however, the proper selection of operation temperature is of significant importance. (author)

  5. Cyclosporine A decreases the fluconazole minimum inhibitory concentration of Candida albicans clinical isolates but not biofilm formation and cell growth.

    Science.gov (United States)

    Wibawa, T; Nurrokhman; Baly, I; Daeli, P R; Kartasasmita, G; Wijayanti, N

    2015-03-01

    Among the genus Candida, Candida albicans is the most abundant species in humans. One of the virulent factors of C. albicans is its ability to develop biofilm. Biofilm forming microbes are characterized by decreasing of its susceptibility to antibiotics and antifungal. The fungicidal effect of fluconazole may be enhanced by cyclosporine A in laboratory engineered C. albicans strains. The aim of this work is to analyze the synergistic effect of cyclosporine A with fluconazole in C. albicans clinical isolates and the effect of cycolsporine A alone in the biofilm formation. Six fluconazole resistant and six sensitive C. albicans clinical isolates were analyzed for its minimum inhibitory concentration (MICs), biofilm formation, and cell growths. A semi-quantitative XTT [2,3-bis(2-methoxy-4-nitro-5- sulfo-phenyl)-2H-tetrazolium-5-carboxanilide] reduction assay was conducted to measure the biofilm formation. Cyclosporine A has synergistic effect with fluconazole that was shown by decreasing MICs of both fluconazole resistant and sensitive C. albicans clinical isolates. However, cyclosporine A alone did not influence the biofilm formation and cell growth of both fluconazole resistant and sensitive C. albicans clinical isolates. These results indicated that cyclosporine A might be a promising candidate of adjuvant therapy for fluconazole against both fluconazole resistant and sensitive C. albicans clinical isolates.

  6. Anhydride-functional silane immobilized onto titanium surfaces induces osteoblast cell differentiation and reduces bacterial adhesion and biofilm formation.

    Science.gov (United States)

    Godoy-Gallardo, Maria; Guillem-Marti, Jordi; Sevilla, Pablo; Manero, José M; Gil, Francisco J; Rodriguez, Daniel

    2016-02-01

    Bacterial infection in dental implants along with osseointegration failure usually leads to loss of the device. Bioactive molecules with antibacterial properties can be attached to titanium surfaces with anchoring molecules such as silanes, preventing biofilm formation and improving osseointegration. Properties of silanes as molecular binders have been thoroughly studied, but research on the biological effects of these coatings is scarce. The aim of the present study was to determine the in vitro cell response and antibacterial effects of triethoxysilypropyl succinic anhydride (TESPSA) silane anchored on titanium surfaces. X-ray photoelectron spectroscopy confirmed a successful silanization. The silanized surfaces showed no cytotoxic effects. Gene expression analyses of Sarcoma Osteogenic (SaOS-2) osteoblast-like cells cultured on TESPSA silanized surfaces reported a remarkable increase of biochemical markers related to induction of osteoblastic cell differentiation. A manifest decrease of bacterial adhesion and biofilm formation at early stages was observed on treated substrates, while favoring cell adhesion and spreading in bacteria-cell co-cultures. Surfaces treated with TESPSA could enhance a biological sealing on implant surfaces against bacteria colonization of underlying tissues. Furthermore, it can be an effective anchoring platform of biomolecules on titanium surfaces with improved osteoblastic differentiation and antibacterial properties.

  7. Slow turning lateral vessel bioreactor improves embryoid body formation and cardiogenic differentiation of mouse embryonic stem cells.

    Science.gov (United States)

    Rungarunlert, Sasitorn; Klincumhom, Nuttha; Tharasanit, Theerawat; Techakumphu, Mongkol; Pirity, Melinda K; Dinnyes, Andras

    2013-10-01

    Embryonic stem cells (ESCs) have the ability to form aggregates, which are called embryoid bodies (EBs). EBs mimic early embryonic development and are commonly produced for cardiomyogenesis. Here, we describe a method of EB formation in hydrodynamic conditions using a slow-turning lateral vessel (STLV) bioreactor and the subsequent differentiation of EBs into cardiomyocytes. EBs formed in the STLV were compared with conventional techniques, such as hanging drop (HD) or static suspension cell culture (SSC), for homogeneity of EB size, shape, proliferation, apoptosis, and in vitro cardiac differentiation. After 3 days of culture, a four-fold improvement in the yield of EB formation/mL, a six-fold enhancement in total yield of EB/mL, and a nearly 10-fold reduction of cells that failed to incorporate into EBs were achieved in STLV versus SSC. During cardiac differentiation, a 1.5- to 4.2-fold increase in the area of cardiac troponin T (cTnT) per single EB in STLV versus SSC and HD was achieved. These results demonstrate that the STLV method improves the quality and quantity of ES cells to form EBs and enhances the efficiency of cardiac differentiation. We have demonstrated that the mechanical method of cell differentiation creates different microenvironments for the cells and thus influences their lineage commitments, even when genetic origin and the culture medium are the same. Ascorbic acid (ASC) improved further cardiac commitment in differentiation assays. Hence, this culture system is suitable for the production of large numbers of cells for clinical cell replacement therapies and industrial drug testing applications. PMID:24020697

  8. A Genome-wide RNAi Screen for Microtubule Bundle Formation and Lysosome Motility Regulation in Drosophila S2 Cells

    OpenAIRE

    Amber L. Jolly; Chi-Hao Luan; Brendon E. Dusel; Sara F. Dunne; Michael Winding; Vishrut J. Dixit; Chloe Robins; Jennifer L. Saluk; David J. Logan; Anne E. Carpenter; Manu Sharma; Deborah Dean; Andrew R. Cohen; Vladimir I. Gelfand

    2016-01-01

    Long-distance intracellular transport of organelles, mRNA, and proteins (“cargo”) occurs along the microtubule cytoskeleton by the action of kinesin and dynein motor proteins, but the vast network of factors involved in regulating intracellular cargo transport are still unknown. We capitalize on the Drosophila melanogaster S2 model cell system to monitor lysosome transport along microtubule bundles, which require enzymatically active kinesin-1 motor protein for their formation. We use an auto...

  9. A genome-wide RNAi screen for microtubule bundle formation and lysosome motility regulation in Drosophila S2 cells

    OpenAIRE

    Jolly, Amber L.; Luan, Chi-Hao; Dusel, Brendon E.; Dunne, Sara Fernandez; Winding, Michael; Dixit, Vishrut J.; Robins, Chloe; Saluk, Jennifer L.; Logan, David J.; Carpenter, Anne E.; Sharma, Manu; Dean, Deborah; Cohen, Andrew R.; Gelfand, Vladimir I.

    2016-01-01

    Long-distance intracellular transport of organelles, mRNA, and proteins (“cargo”) occurs along the microtubule cytoskeleton by the action of kinesin and dynein motor proteins; the vast network of factors involved in regulating intracellular cargo transport are still unknown. We capitalize on the Drosophila melanogaster S2 model cell system to monitor lysosome transport along microtubule bundles, which require enzymatically active kinesin-1 motor protein for their formation. We use an automate...

  10. The Drosophila cell adhesion molecule Klingon is required for long-term memory formation and is regulated by Notch

    OpenAIRE

    Matsuno, Motomi; Horiuchi, Junjiro; Tully, Tim; Saitoe, Minoru

    2008-01-01

    The ruslan (rus) mutant was previously identified in a behavioral screen for mutants defective in long-lasting memory, which consists of two consolidated memory types, anesthesia-resistant memory, and protein synthesis-dependent long-term memory (LTM). We demonstrate here that rus is a new allele of klingon (klg), which encodes a homophilic cell adhesion molecule. Klg is acutely required for LTM but not anesthesia-resistant memory formation, and Klg expression increases upon LTM induction. LT...

  11. Mutations in E coli cistrons affecting adhesion to human cells do not abolish Pap pili fiber formation.

    OpenAIRE

    Norgren, M; Normark, S; Lark, D; O'Hanley, P; Schoolnik, G; Falkow, S; Svanborg-Edén, C; Båga, M; Uhlin, B E

    1984-01-01

    A chromosomal DNA fragment which mediates Pap (pili associated with pyelonephritis) pili formation, mannose-resistant hemagglutination ( MRHA ) and binding to uroepithelial cells has been isolated from the uropathogenic Escherichia coli clinical isolate J96 , and genetically studied. Analysis of polypeptides expressed by the Pap DNA led to detection of a number of polypeptides ranging in mol. wt. from 13 000 to 81 000 daltons. The gene order and transcriptional orientation for four of the cor...

  12. HIV-1 efficient entry in inner foreskin is mediated by elevated CCL5/RANTES that recruits T cells and fuels conjugate formation with Langerhans cells.

    Directory of Open Access Journals (Sweden)

    Zhicheng Zhou

    2011-06-01

    Full Text Available Male circumcision reduces acquisition of HIV-1 by 60%. Hence, the foreskin is an HIV-1 entry portal during sexual transmission. We recently reported that efficient HIV-1 transmission occurs following 1 h of polari